WO2014112663A1 - Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient - Google Patents

Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient Download PDF

Info

Publication number
WO2014112663A1
WO2014112663A1 PCT/KR2013/000357 KR2013000357W WO2014112663A1 WO 2014112663 A1 WO2014112663 A1 WO 2014112663A1 KR 2013000357 W KR2013000357 W KR 2013000357W WO 2014112663 A1 WO2014112663 A1 WO 2014112663A1
Authority
WO
WIPO (PCT)
Prior art keywords
chronic pancreatitis
pharmaceutical composition
extract
persimmon
present
Prior art date
Application number
PCT/KR2013/000357
Other languages
French (fr)
Korean (ko)
Inventor
황성연
박성주
Original Assignee
주식회사 한국전통의학연구소
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 주식회사 한국전통의학연구소 filed Critical 주식회사 한국전통의학연구소
Priority to PCT/KR2013/000357 priority Critical patent/WO2014112663A1/en
Publication of WO2014112663A1 publication Critical patent/WO2014112663A1/en

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/84—Valerianaceae (Valerian family), e.g. valerian
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A61P1/18—Drugs for disorders of the alimentary tract or the digestive system for pancreatic disorders, e.g. pancreatic enzymes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]

Definitions

  • the present invention relates to a novel use of persimmon scent, and more particularly to a pharmaceutical composition for the prevention and treatment of chronic pancreatitis, and a food composition for the prevention and improvement of chronic pancreatitis comprising the extract as an active ingredient.
  • CP chronic pancreatitis
  • the syndromes, such as abdominal pain, steatorrhea, and weight loss are exacerbated by psychosocial problems, which can lead to debilitating rectal discharge, drug addiction, and loss of health care resources (Apte, MV et al. (2010)). J. Gastroenterol. Hepatol. 25 , 1816-1826).
  • CP is mainly associated with alcohol abuse (Lankisch, PG et al. (1993) Digestion 54 , 148-155).
  • Alcoholic acute pancreatitis (AP) rarely occurs after one binge drinking, so the majority of patients are men who consume 150 g of alcohol daily on average for 10-15 years after the start of the example.
  • Persimmon Scent (Nardostachys jatamansi, NJ) has been used extensively as a tonic, stimulant and anticonvulsant in several Asian countries, as well as to treat epilepsy, pathological excitement, palpitations and cramps (Bagchi, A. , Et al., Planta Med., 57, 9697 (1991)).
  • the rooted water of the incense incense has been used for mental disorders, insomnia, blood disorders and circulatory disorders (Uniyal, MR., Et al., J. Res. Indian Med.
  • the present inventors have previously reported that sweet persimmon has a protective effect on acute pancreatitis, and pneumonia (Bae, GS et al. (2010) Pancreas 39 , 520-529; Bae, GS et al. (2011) J. Nat. Med. 65 , 63 -72).
  • NJ has been used in mental disorders, obscurity, blood diseases, and the circulatory system (Uniyal. MR and Issar, RK (1969) J. Res. Indian Med. 4 , 83).
  • NJ Nardostachys jatamansi belonging to the Valerianaceae is not known for its ability to improve alcoholic chronic pancreatitis (ACP).
  • the present inventors while studying the other effects of the sensational fragrance, using an alcohol model repeatedly added to the AP effect of NJ on the ACP, morphological and histological changes of the pancreas, in vivo and in vitro ( The present invention was completed by confirming collagen accumulation and PSC activation in vitro ) to confirm that the extract of Persimmon can be used as a pharmaceutical composition for the prevention and treatment of chronic pancreatitis.
  • An object of the present invention is to provide a pharmaceutical composition for the prevention and treatment of chronic pancreatitis (CP) and novel pancreatitis (NJ) containing a novel use, specifically, the extract of N. persimmon (NJ) as an active ingredient It is.
  • CP chronic pancreatitis
  • NJ novel pancreatitis
  • Another object of the present invention to provide a food composition for preventing and improving chronic pancreatitis comprising the extract persimmon extract as an active ingredient.
  • the present invention provides a pharmaceutical composition for the prevention and treatment of chronic pancreatitis comprising the extract of Nardostachys jatamansi NJ as an active ingredient.
  • the present invention provides a food composition for preventing and improving chronic pancreatitis, including the extract of Nardostachys jatamansi NJ as an active ingredient.
  • the present invention provides a pharmaceutical composition for the prevention and treatment of chronic pancreatitis, comprising as an active ingredient extract of Persimmon (Nardostachys jatamansi NJ).
  • the chronic pancreatitis is preferably alcoholic chronic pancreatitis, and the composition further comprises a pharmaceutically acceptable carrier.
  • the extract of the persimmon extract is water or organic solvent extract of the root of the persimmon flavor, wherein the persimmon extract is 1 to the root of the persimmon fragrance More preferably, 20 times of water is added, extracted at 80 to 150 ° C. for 1 to 24 hours, and then filtered.
  • the extract of persimmon flavor of the present invention can be obtained by extracting the root of the persimmon incense with water or an organic solvent, the lower alcohol, acetone, chloroform, methylene chloride, ether, ethyl acetate, hexane and the like can be exemplified.
  • Lower alcohols include methanol, ethanol, propanol and butanol, with ethanol being most preferred.
  • the dried persimmon root dry matter or powder 1 at a temperature of 80-150 ° C., preferably 90-100 ° C. It may be extracted for 24 hours, preferably 2 to 6 hours, more preferably 2 hours and then filtered to prepare a hydrothermal extract of the persimmon root.
  • the organic solvent extract of the persimmon fragrance root may be prepared by adding 1 to 5 times, preferably 3 times the organic solvent to the dried persimmon root dry matter or powder, extracting at room temperature, and then filtrating the filtrate obtained under reduced pressure.
  • the organic solvent may be an ethanol or methanol extract.
  • the extraction process may be repeated two or more times as necessary, and the extract obtained after filtration may be freeze-dried or reduced-pressure drying to form a powder.
  • the pharmaceutical composition of the present invention may further include other pharmaceutically acceptable herbal medicines or extracts thereof to enhance the pharmacological effect.
  • an extract of the herbal medicine may be prepared according to the extraction method, and then added to the pharmaceutical composition, or the extract obtained by extracting by the above method after mixing the saenghyang root and the herbal medicine may be included in the composition.
  • 'pharmaceutically acceptable refers to a physiologically acceptable and, when administered to a human, usually does not cause an allergic or similar reaction.
  • the herbal medicine that may be added to the composition of the present invention may be any pharmaceutically acceptable herbal medicine, for example, Angelicae tenuissimae Radix, Gastrodiae Rhizoma, Bapleuri Radix, Angelica gigantis Radix, Persicae Semen, Cinnamomi Ramulus, Rhei Rhizoma, Licorice (Glycyrrhizae Radix), Cnidii Rhizoma, Dermis (Aurantii nobilis Pericarpium), Taxa (Alismatis Rhizomaidi) ), Golden (Scutellariae Radix), Horyng (Hoelen), Peony (Paeoniae Radix), Bleach (Atractylodis Rhizoma alba), Peach (Phellodendri Cortex), Gardenia (Gardeniae Fructus), Pinelliae Tuber, Uncaria Ramuluset Uncus , Ponciri Fructus, Ginseng, Gingseng,
  • compositions according to the invention may further contain one or more pharmaceutically acceptable carriers, excipients or diluents.
  • Pharmaceutically acceptable carriers may further include, for example, carriers for oral administration or carriers for parenteral administration.
  • Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid and the like.
  • carriers for parenteral administration may include water, suitable oils, saline, aqueous glucose and glycols, and the like, and may further include stabilizers and preservatives.
  • Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid.
  • Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol.
  • Other pharmaceutically acceptable carriers may be referred to those described in the following documents (Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, PA, 1995).
  • the pharmaceutical composition for preventing or treating chronic pancreatitis of the present invention can be administered to any mammal, including humans.
  • it can be administered orally or parenterally.
  • Parenteral administration methods include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intradural, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, topical, sublingual or rectal administration.
  • the pharmaceutical composition of the present invention may be administered transdermally.
  • 'transdermal administration' refers to the administration of the pharmaceutical composition of the present invention to cells or skin to deliver the active ingredient contained in the pharmaceutical composition for the prevention or treatment of chronic pancreatitis into the skin.
  • the pharmaceutical composition of the present invention may be prepared in an injectable formulation and administered by lightly pricking the skin with a 30 gauge thin injection needle, or directly applying to the skin.
  • composition of the present invention may be formulated into a preparation for oral or parenteral administration according to the route of administration as described above.
  • compositions of the present invention may be formulated using methods known in the art as powders, granules, tablets, pills, dragees, capsules, solutions, gels, syrups, slurries, suspensions and the like.
  • oral formulations can be obtained by tablets or dragees by combining the active ingredients with solid excipients, milling them, adding suitable auxiliaries and then processing them into granule mixtures.
  • excipients examples include sugars including lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol and maltitol and starch, cellulose, including starch, corn starch, wheat starch, rice starch and potato starch, etc. Fillers such as celluloses, gelatin, polyvinylpyrrolidone, and the like, including methyl cellulose, sodium carboxymethylcellulose, hydroxypropylmethyl-cellulose, and the like. In addition, crosslinked polyvinylpyrrolidone, agar, alginic acid or sodium alginate and the like may optionally be added as a disintegrant. Furthermore, the pharmaceutical composition of the present invention may further include an anticoagulant, a lubricant, a humectant, a perfume, an emulsifier, a preservative, and the like.
  • Formulations for parenteral administration may be formulated by methods known in the art in the form of injections, creams, lotions, external ointments, oils, humectants, gels, aerosols and nasal inhalants. These formulations are described in Remington's Pharmaceutical Science, 15th Edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blaug, Seymour, a prescription generally known in all pharmaceutical chemistries.
  • the total effective amount of the extract of Persimmon Fragrance, the active ingredient of the composition of the present invention may be administered to a patient in a single dose, and may be administered in a fractionated treatment protocol administered for a long time in multiple doses. May be administered.
  • the pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the extent of the disease.
  • the preferred total dose of the sweet persimmon of the present invention may be about 100 ⁇ g to 5 mg, most preferably 500 ⁇ g to 1 mg per kg of patient body weight per day.
  • the dose of the extract is effective for taking into consideration the various factors such as the age, weight, health status, sex, severity of the disease, diet and excretion rate, as well as the route and frequency of treatment of the pharmaceutical composition Since the amount is determined, one of ordinary skill in the art will be able to determine the appropriate effective dosage according to the specific use of the sensational aroma as a prophylactic or therapeutic agent for chronic pancreatitis.
  • the pharmaceutical composition according to the present invention is not particularly limited to its formulation, route of administration and method of administration as long as the effect of the present invention is shown.
  • the present invention may be provided in the form of a food composition for the prevention and improvement of chronic pancreatitis comprising the extract of Nardostachys jatamansi NJ as an active ingredient.
  • the food composition of the present invention includes all forms such as functional foods, nutritional supplements, health foods and food additives. Food compositions of this type can be prepared in various forms according to conventional methods known in the art.
  • the persimmon extract of the present invention may be prepared by drinking tea, juice, and drink, or granulated, encapsulated, and powdered.
  • it can be prepared in the form of a composition by mixing with the extract of the sense of flavor of the present invention and known substances or active ingredients known to have the effect of preventing and improving chronic pancreatitis.
  • functional foods include beverages (including alcoholic beverages), fruits and processed foods (e.g. canned fruit, canned foods, jams, marmalade, etc.), fish, meat and processed foods (e.g. ham, sausage cornebipe) Etc.), breads and noodles (e.g. udon, soba noodles, ramen, spaghetti, macaroni, etc.), fruit juices, various drinks, cookies, syrups, dairy products (e.g. butter, cheese, etc.), edible vegetable oils, margarine, vegetable protein, It can be prepared by adding the extract of persimmon flavor of the present invention to retort foods, frozen foods, various seasonings (eg, miso, soy sauce, sauce, etc.).
  • fruits and processed foods e.g. canned fruit, canned foods, jams, marmalade, etc.
  • fish e.g. ham, sausage cornebipe
  • breads and noodles e.g. udon, soba noodles, ramen, spaghetti, macaroni, etc.
  • the persimmon aroma of the present invention in the form of food additives can be prepared in the form of powder or concentrate.
  • Pancreatic fibrosis has long been considered a late stage with little or no hope for improvement. With a more advanced understanding of the physiological mechanisms underlying chronic pancreatitis (CP) and the characteristics of astrocytic cells, the major working cells in this process, a more focused attention to the possibility of fibrosis regression was considered the only means. .
  • ACP alcoholic chronic pancreatitis
  • pancreatic acinar cells pancreatic acinar cells, collagen accumulation and pancreatic appearance
  • PSC Persimmon Fragrance Extract
  • AP is characterized by pain, edema, bleeding, saturation of acinar cells, necrosis, inflammation, and increased serum amylase and lipase; This feature is confused with chronic disease forms due to fibrosis, inflammation, collagen accumulation and reduced exocrine and endocrine function.
  • AP and CP differ in pathogenic causes, repetitive AP can gradually lead to CP (Ohashi, S. et al. (2006) Am. J. Physiol. Gastrointest. Liver Physiol. 290 , G772-781).
  • repeated administration of the cerulein model was limited in treatment time, severity, and similarity with humans.
  • mice were sufficient to induce ACP with a challenge given for 3 weeks, which was demonstrated by fibrosis, inflammation, and acinar cell destruction in the pancreas (see FIG. 1). Daily and free NJ consumption inhibited morphological damage to ACP (see FIG. 1). Amylase-positive cells are abundant in the pancreas, suggesting that exocrine function is functioning properly.
  • ACP The main question in the disease-physiology of ACP is whether the disease state can be reversed. Many clinicians often recommend patients to suppress alcohol, but this has little effect on patients. Prior research on how to improve the ACP By discontinue their alcohol intake did not find a substantial improvement (Apte, MV, etc. (2010) J. Gastroenterol Hepatol 25, 1816-1826;.. Gullo, L. , etc. (1988) Gastroenterology 95 , 1063-1068). In contrast to humans, rat AP and ACP models generally improve disease after lack of stimulation (Lugea, A. et al. (2006) Gastroenterology 131 , 885-899; Von Stamm, A. et al. (2010) Gut 60 , 238-246).
  • mice are not sufficient to demonstrate the mechanism of human CP.
  • regulation of PSC is a key factor (Masamune, A. et al. (2009) Clin. Gastroenterol. Hepatol. 7 , S48-54).
  • many studies have tried to modulate PSC by inactivating or killing PSC factors (Schwer, CI et al. (2008) J. Pharmacol. Exp. Ther. 327 , 863-871; Li, L. et al. (2011) Eur. J. Clin. Invest. 41 , 151-158; Madro, A. et al. (2008) J.
  • the present invention constituted as described above provides a novel use of persimmon aroma, chronic pancreatitis prevention and treatment composition comprising the persimmon extract as an active ingredient.
  • the composition comprising the extract of the present invention can be effectively used for the prevention and treatment of chronic pancreatitis because it effectively inhibits chronic pancreatitis caused by cerulein and ethanol.
  • Figure 1 shows the NJ effect on the morphology and tissue of the pancreas after ACP
  • A is a representative pancreatic external morphology
  • B is a representative H & E stained fragment of the pancreas
  • C is inflammation, edema, fibrosis, And histological fragments of the pancreas were recorded from 0 (normal) to 3 (severe) for the whole.
  • the photo shows a representative image of one experimental test with six mice per group. The results were similar in three additional experiments. First magnification ( ⁇ 200). * P ⁇ 0.05, the control group was treated with physiological saline.
  • Figure 2 shows the effect of NJ on acinar cell death during ACP, and the presence of acinar cells was detected by successive staining of pancreatic tissue fragments with amylase.
  • the photo shows a representative image of one experimental test with six mice per group. The results were similar in 3 additional experiments.
  • Figure 3 shows the effect of PSC activation and collagen accumulation during ACP, pancreatic tissue sections were sequentially stained with (A) ⁇ -SMA and (C) collagen to detect activated PSCs.
  • the photo shows a representative image of one experimental test with six mice per group. First magnification ( ⁇ 400).
  • (B) measured pancreatic mRNA levels of ⁇ -SMA, fibronectin and TGF- ⁇ by quantitative RT-PCR. The data represents the mean SE SE of 6 mice in each group. The results were similar in three additional experiments. * P ⁇ 0.05, control group treated with saline solution
  • Figure 4 shows the effect of NJ on PSC activation in isolated PSC, PSC was isolated from rat pancreas and culture activated for 3 weeks. On day 21, NJ was treated for 24 hours. Then, (A) desmin and (B) ⁇ -SMA were treated successfully to detect PSC isolation and activation. First magnification ( ⁇ 400). (C) detected Western blot expression of ⁇ -SMA. (D) quantified cytokine mRNA expression by PSC by real-time RT-PCR. The photograph shows a representative image of one experiment. The results were similar in three additional experiments. * P ⁇ 0.05, the control group was treated with physiological saline.
  • Cerulean Tris-HCl, NaCl, collagen, DAPI, hematoxylin, eosin, xylene, ethanol, cerulein and Triton X-100 are Sigma-Aldrich (St. Louis, MO, USA).
  • ⁇ -smooth muscle actin (SMA), collagen, and desmine were purchased from Abcam (UK).
  • Amylase and GAPDH were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
  • mice All animal experiments were performed according to a protocol approved by Wonkwang University Animal Care Committee. Experimental animals purchased 15-20 g body weight 6-8 week old C57BL / 6 mouse females (15-20 g body weight) (Orient Bio). All experimental animals were bred for 7 days in a breeding room with a room temperature of 23 ⁇ 2 °C and a contrast cycle of 12 hours. The experimental animals were fed with standard feed and water as ad libitum for 7 days to adapt to the environment, and then randomly divided into control and experimental groups. Each mouse was fasted for 18 hours before inducing alcoholic chronic pancreatitis (ACP).
  • ACP alcoholic chronic pancreatitis
  • Dried persimmon roots were purchased from General Herbs (Omni Herb, Seoul, Korea). Identification of the herb was confirmed in the Korean drug test laboratory. Voucher specimens (NO; Oh / wh / nj-43) were deposited in Wonkwang University Oriental Medicine Plant Botanical Laboratory.
  • the distilled root was put in distilled water of about 10 times and boiled for about 2 hours, and then the extract was filtered and lyophilized at -70 ° C to powder.
  • the powder was extracted with distilled water, filtered and stored at 4 ° C.
  • the powder was dissolved in physiological saline and used at the required concentration.
  • Immobilized pancreatic tissue was embedded in paraffin, cut into 4 ⁇ m sections and stained with haematoxyl-eosin for standard histological examination. Immunohistochemical staining with amylase, collagen or ⁇ -SMA was performed using a DAB immunohistohemical kit (DAKO, Cytomation, Denmark). Incubation of PSC with anti- ⁇ -SMA, or desmin, followed by incubation with Alexa Fluors 568 goat-anti rabbit IgG (1: 2,000, Invitrogen, Carlsbad, Calif., USA) for 1 hour at room temperature, DAPI, ⁇ - Immunofluorescence detection of SMA or desmine was performed. The slides were mounted with mounting medium equipped with DAPI (Vector Laboratories, Burlingame, CA, USA).
  • mRNA messenger RNA
  • Target cytokine transcription was analyzed by RT-PCR in mouse pancreatic tissue and PSC.
  • Total RNA was isolated from pancreas and PSC of mice using TriZol (Invitrogen, Carlsbad, CA, USA) and reverse transcription was performed using SuperScript II RT (Invitrogen, Carlsbad, CA, USA).
  • TaqMan quantitative RT-PCR was performed using the ABI stepone plue system according to the manufacturer's instructions (Invitrogen, Carlsbad, CA, USA).
  • HGPRT hypoxanthine-guanine phosphoribosyltransferase
  • the mixed interface was harvested, and the cells were washed and then in DMEM containing 10% fetal bovine serum (FBS), 4 mM glutamine, and antibiotics (penicillin 100 U / ml, streptomycin 100 mg / ml). Resuspend. After reaching confluency, cells were harvested and replated at the same seed density. All experiments were performed using culture-harvested cells (2-4 generations). PSCs were incubated in serum-deficient media 24 hours prior to the addition of experimental reagents.
  • FBS fetal bovine serum
  • 4 mM glutamine fetal bovine serum
  • antibiotics penicillin 100 U / ml, streptomycin 100 mg / ml
  • the lysate was boiled in the same buffer (62.5 mM Tris-HCl, pH 6.8, 2% sodium dodecyl sulfate (SDS), 20% glycerol, and 10% 2-mercaptoethanol). Then, proteins in cell lysate were separated by 10% SDS-PAGE and transferred to nitrocellulose membrane. After transfer of the protein, the membrane was blocked with 5% skim milk of PBST (PBS-Tween-20) for 2 hours at room temperature, and then reacted with ⁇ -SMA and antibody overnight.
  • PBST PBS-Tween-20
  • each blot is reacted with peroxidase-linked secondary antibody for 1 hour and the antibody-specific protein using an improved chemiluminescence detection system (Amersham, Piscataway, NJ) according to the manufacturer's recommended protocol was visualized.
  • pancreatic stromal edema pancreatic stromal edema, inflammation, infiltration, and pancreatic fibrosis were demonstrated (FIG. 1).
  • ACP lost its characteristic morphology and had edema characteristics rather than the normal pancreas.
  • NJ treatment at 5 or 10 mg / ml significantly reduced external morphological edema of the pancreas (FIG. 1).
  • FIG. 1B in histological examination, NJ treatment did not show significant protection against ACP challenge.
  • FIG. 1B In order to further investigate the exact effect of NJ on ACP, we measured the survival of pancreatic acinar cells via amylase release.
  • ACP challenged mice demonstrated disruption of pancreatic acinar cells, as indicated by reduced amylase.
  • NJ treatment significantly inhibited acinar cell death and destruction (FIG. 2).
  • PSCs express intermediate filament proteins such as desmin (13).
  • PSCs express intermediate fiber proteins such as desmin (Omary, MB et al. (2004) N. Engl. J. Med. 351 , 2087-2100). As reported previously, isolated PSCs had a desmin positive response, indicating that the isolated cells were PSCs (FIG. 4A). Upon 3-week incubation of PSCs, the PSCs are activated and express ⁇ -SMA (FIG. 4B). However, NJ treatment inhibited the expression of ⁇ -SMA in PSCs (FIGS. 4B and 4C). NJ treatment also inhibited mRNA expression of fibrosis-related genes such as ⁇ -SMA, fibronectin, and TGF- ⁇ in PSCs.
  • the present invention provides a composition for the prevention and treatment of chronic pancreatitis, which comprises the extract of Persimmon as an active ingredient as a novel use of persimmon. Since the composition of the present invention effectively inhibits chronic pancreatitis caused by cerulein, it can be effectively used for the prevention and treatment of chronic pancreatitis.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • General Health & Medical Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Medicinal Chemistry (AREA)
  • Animal Behavior & Ethology (AREA)
  • Chemical & Material Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Engineering & Computer Science (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Alternative & Traditional Medicine (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Pain & Pain Management (AREA)
  • Rheumatology (AREA)
  • Biotechnology (AREA)
  • Botany (AREA)
  • Medical Informatics (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Epidemiology (AREA)
  • Medicines Containing Plant Substances (AREA)

Abstract

The present invention relates to a novel use of Nardostachys jatamansi (NJ), more specifically relates to a pharmaceutical composition for preventing and treating chronic pancreatitis which includes an extract of Nardostachys jatamansi as an active ingredient or a food composition for preventing or improving chronic pancreatitis. The composition of the present invention effectively suppresses chronic pancreatitis induced by caerulein and ethanol, and thus can be effectively used in preventing and treating chronic pancreatitis.

Description

감송향 추출물을 활성성분으로 포함하는 만성 췌장염의 예방 및 치료용 약학적 조성물Pharmaceutical composition for the prevention and treatment of chronic pancreatitis comprising the extract of persimmon
본 발명은 감송향의 신규한 용도에 관한 것으로, 보다 상세하게는 감송향 추출물을 활성성분으로 포함하는 만성 췌장염의 예방 및 치료용 약학적 조성물 및 만성 췌장염 예방 및 개선용 식품 조성물에 관한 것이다.The present invention relates to a novel use of persimmon scent, and more particularly to a pharmaceutical composition for the prevention and treatment of chronic pancreatitis, and a food composition for the prevention and improvement of chronic pancreatitis comprising the extract as an active ingredient.
심각한 섬유증(Prominent fibrosis), 염증, 선위축(glandular atrophy), 및 유관변화는 만성 췌장염(chronic pancreatitis ; CP)의 주요한 조직학적 특징이다(Apte, M.V. 등 (2011) Antioxid. Redox Signal. 15, 2711-2722). 복통, 지방 변증(steatorrhea), 및 체중 감소와 같은 그 증후군은 심리사회적 문제에 의해 악화되므로, 직장 퇴출, 마약 중독, 및 의료 서비스 자원의 손실을 쇠약하게 할 수 있다(Apte, M.V. 등 (2010) J. Gastroenterol. Hepatol. 25,1816-1826). CP는 주로 알콜 남용과 연관된다(Lankisch, P.G. 등 (1993) Digestion 54, 148-155). 알콜성 급성 췌장염(acute pancreatitis ; AP)은 한 번의 폭음이후에 거의 발생하지 않으므로, 환자의 대부분은 예시적 시작일 이후로 10-15년 동안 평균적으로 매일 150 g의 알콜을 소비하는 남성이다.Prominent fibrosis, inflammation, glandular atrophy, and ductal change are major histological features of chronic pancreatitis (CP) (Apte, MV et al. (2011) Antioxid. Redox Signal. 15 , 2711 -2722). The syndromes, such as abdominal pain, steatorrhea, and weight loss are exacerbated by psychosocial problems, which can lead to debilitating rectal discharge, drug addiction, and loss of health care resources (Apte, MV et al. (2010)). J. Gastroenterol. Hepatol. 25 , 1816-1826). CP is mainly associated with alcohol abuse (Lankisch, PG et al. (1993) Digestion 54 , 148-155). Alcoholic acute pancreatitis (AP) rarely occurs after one binge drinking, so the majority of patients are men who consume 150 g of alcohol daily on average for 10-15 years after the start of the example.
섬유 조직의 비정상적 축적이 CP, 및 췌장암과 같은 췌장의 2가지 주요 질환의 특징적 조직학적 특징이다(Kloppel, G. 등 (2004) Virchows Arch. 445, 1-8). 췌장의 만성적 손상의 주요한 대상은 췌장 성상세포(pancreatic stellate cell ; PSC)이다(Apte, M.V. 등 (1998) Gut 43, 128-133). 배양 PSC를 이용함으로써 연구자들은 많이 필요로 되는 시험관내(in vitro) 도구를 제공받아서 건강 및 질병 상태에서 PSC 생물학을 검사하였다. PSC 비활성화 또는 근절은 CP의 널리 공지된 치료법이다(Haber, P.S. 등 (1999) Am. J. Pathol. 155, 1087-1095).Abnormal accumulation of fibrous tissue is a characteristic histological feature of two major diseases of the pancreas, such as CP, and pancreatic cancer (Kloppel, G. et al. (2004) Virchows Arch. 445 , 1-8). The major target of chronic damage to the pancreas is pancreatic stellate cells (PSCs) (Apte, MV et al. (1998) Gut 43 , 128-133). By using cultured PSCs, researchers were given a much needed in vitro tool to test PSC biology in health and disease conditions. PSC inactivation or eradication is a well known treatment of CP (Haber, PS et al. (1999) Am. J. Pathol. 155 , 1087-1095).
한편, 감송향 (Nardostachys jatamansi, NJ)은 몇몇 아시아 국가들에서 강장제, 자극제 및 항경련제로 광범위하게 사용되었을 뿐 아니라, 간질, 병적 흥분, 심계 항진 및 경련을 치료하는데도 사용되어 왔다 (Bagchi, A., 등, Planta Med., 57, 9697 (1991)). 감송향의 뿌리를 달인 물은 정신장애, 불면증, 혈액장애 및 순환계 장애에 사용되었으며 (Uniyal, MR., 등, J. Res. Indian Med. 4, 83 (1969)), 감송향의 주요한 치료 성분은 향신경 활성 및 중추 신경계 (CNS) 효과를 증진시키는 나도시논 (nardosinone)이다 (Li, p. 등, Journal of Pharmacology Science, 93, 122-125 (2003)). 그러나, 감송향이 세룰레인 유도 만성 췌장염에 미치는 효과는 아직 알려진 바 없다.On the other hand, Persimmon Scent (Nardostachys jatamansi, NJ) has been used extensively as a tonic, stimulant and anticonvulsant in several Asian countries, as well as to treat epilepsy, pathological excitement, palpitations and cramps (Bagchi, A. , Et al., Planta Med., 57, 9697 (1991)). The rooted water of the incense incense has been used for mental disorders, insomnia, blood disorders and circulatory disorders (Uniyal, MR., Et al., J. Res. Indian Med. 4, 83 (1969)), the main therapeutic component of the incense incense Is nardosinone that enhances neuronal activity and central nervous system (CNS) effects (Li, p. Et al., Journal of Pharmacology Science, 93, 122-125 (2003)). However, the effect of the extract on cerulein-induced chronic pancreatitis is still unknown.
본 발명자들은 앞서서 감송향이 급성 췌장염, 및 폐혈증에 보호 효과를 가짐을 보고하였다(Bae, G.S. 등 (2010) Pancreas 39, 520-529 ; Bae, G.S. 등 (2011) J. Nat. Med. 65, 63-72). 본 발명자들은 NJ(감송향) 효과의 측정에서 달인 뿌리를 사용하였다. NJ는 정신장애, 불명증, 혈액 질환, 및 순환계에 사용되어 왔다(Uniyal. M.R. 및 Issar, R.K. (1969) J. Res. Indian Med. 4, 83). The present inventors have previously reported that sweet persimmon has a protective effect on acute pancreatitis, and pneumonia (Bae, GS et al. (2010) Pancreas 39 , 520-529; Bae, GS et al. (2011) J. Nat. Med. 65 , 63 -72). We used decoction roots in the determination of the NJ (Samson) effect. NJ has been used in mental disorders, obscurity, blood diseases, and the circulatory system (Uniyal. MR and Issar, RK (1969) J. Res. Indian Med. 4 , 83).
그러나, 마타리과(Valerianaceae)에 속하는 감송향 (Nardostachys jatamansi, NJ)은 알콜성 만성 췌장염(alcoholic chronic pancreatitis ; ACP)을 개선하는 능력에 대해서는 공지되어 있지 않다.However, Nardostachys jatamansi (NJ) belonging to the Valerianaceae is not known for its ability to improve alcoholic chronic pancreatitis (ACP).
이에 본 발명자들은 감송향이 갖는 다른 효과를 연구하던 중, 반복적으로 AP에 추가되는 알콜 모델을 이용하여 ACP에 NJ의 효과, 췌장의 형태학적 및 조직학적 변화, 생체내(in vivo) 및 시험관내(in vitro)의 콜라겐 축적 및 PSC 활성화 측정하여 감송향 추출물이 만성 췌장염의 예방 및 치료용 약학적 조성물로 사용될 수 있음을 확인함으로써 본 발명을 완성하였다.The present inventors, while studying the other effects of the sensational fragrance, using an alcohol model repeatedly added to the AP effect of NJ on the ACP, morphological and histological changes of the pancreas, in vivo and in vitro ( The present invention was completed by confirming collagen accumulation and PSC activation in vitro ) to confirm that the extract of Persimmon can be used as a pharmaceutical composition for the prevention and treatment of chronic pancreatitis.
본 발명의 목적은 감송향의 신규한 용도, 구체적으로 감송향(NJ) 추출물을 활성성분으로 포함하는 만성 췌장염(CP)의 예방 및 치료용 약학적 조성물 및 만성 췌장염 예방 및 개선용 식품 조성물을 제공하는데 있다.SUMMARY OF THE INVENTION An object of the present invention is to provide a pharmaceutical composition for the prevention and treatment of chronic pancreatitis (CP) and novel pancreatitis (NJ) containing a novel use, specifically, the extract of N. persimmon (NJ) as an active ingredient It is.
본 발명의 또 다른 목적은 감송향 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 개선용 식품 조성물을 제공한다.Another object of the present invention to provide a food composition for preventing and improving chronic pancreatitis comprising the extract persimmon extract as an active ingredient.
상기 목적을 달성하기 위하여, 본 발명은 감송향 (Nardostachys jatamansi NJ) 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 치료용 약학적 조성물을 제공한다.In order to achieve the above object, the present invention provides a pharmaceutical composition for the prevention and treatment of chronic pancreatitis comprising the extract of Nardostachys jatamansi NJ as an active ingredient.
또한, 본 발명은 감송향 (Nardostachys jatamansi NJ) 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 개선용 식품 조성물을 제공한다.In another aspect, the present invention provides a food composition for preventing and improving chronic pancreatitis, including the extract of Nardostachys jatamansi NJ as an active ingredient.
이하, 본 발명을 상세히 설명한다.Hereinafter, the present invention will be described in detail.
본 발명은 감송향 (Nardostachys jatamansi NJ) 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 치료용 약학적 조성물을 제공한다. The present invention provides a pharmaceutical composition for the prevention and treatment of chronic pancreatitis, comprising as an active ingredient extract of Persimmon (Nardostachys jatamansi NJ).
본 발명의 만성 췌장염 예방 및 치료용 약학적 조성물에 있어서, 상기 만성 췌장염은 알콜성 만성 췌장염인 것이 바람직하고, 상기 조성물은 약학적으로 허용가능한 담체를 추가로 포함하는 것이 바람직하다. In the pharmaceutical composition for preventing and treating chronic pancreatitis of the present invention, the chronic pancreatitis is preferably alcoholic chronic pancreatitis, and the composition further comprises a pharmaceutically acceptable carrier.
또한, 본 발명의 만성 췌장염 예방 및 치료용 약학적 조성물에 있어서, 상기 감송향 추출물이 감송향의 뿌리를 물 또는 유기용매로 추출한 것이 바람직하고, 이때 상기 감송향 추출물이 감송향의 뿌리에 1 내지 20 배의 물을 가하고 80 내지 150℃에서 1 내지 24 시간 동안 추출한 후 여과하여 얻어진 것이 보다 바람직하다.In addition, in the pharmaceutical composition for preventing and treating chronic pancreatitis of the present invention, it is preferable that the extract of the persimmon extract is water or organic solvent extract of the root of the persimmon flavor, wherein the persimmon extract is 1 to the root of the persimmon fragrance More preferably, 20 times of water is added, extracted at 80 to 150 ° C. for 1 to 24 hours, and then filtered.
본 발명자들은 C57black/6 마우스에 세룰레인-유도 급성 췌장염(cerulein-induced acute pancreatitis)의 배경에 대하여 3주 동안 주기적으로 에탄올을 주입하였다. 3주 동안의 처치 후에, 조직학적 검사를 위해 췌장을 수확하였다. NJ 처리는 췌장 포상세포(pancreatic acinar cell)의 생존을 증진시켰고 (아밀라제 수준 테스트에 의해 확인) 콜라겐 축적 및 췌장 성상세포(pancreatic stellate cell ; PSC)의 활성을 감소시켰다. 추가하여, NJ 처리는 상기 활성을 감소시켰지만, PSC의 사멸을 감소시키지는 않았다. 결론적으로, 본 발명의 데이터는 NJ가 PSC 활성의 억제를 통해서 ACP를 약화시킴을 나타낸다.We injected C57black / 6 mice with ethanol periodically for 3 weeks against the background of cerulein-induced acute pancreatitis. After three weeks of treatment, the pancreas was harvested for histological examination. NJ treatment enhanced the survival of pancreatic acinar cells (confirmed by amylase level testing) and reduced collagen accumulation and pancreatic stellate cell (PSC) activity. In addition, NJ treatment reduced the activity, but did not reduce the killing of PSCs. In conclusion, the data of the present invention indicate that NJ attenuates ACP through inhibition of PSC activity.
본 발명의 감송향 추출물은 감송향의 뿌리를 물 또는 유기용매로 추출하여 얻을 수 있는데, 유기용매로는 저급알콜, 아세톤, 클로로포름, 메틸렌클로라이드, 에테르, 에틸아세테이트, 헥산 등을 예시할 수 있다. 저급알콜로는 메탄올, 에탄올, 프로판올 및 부탄올을 예시할 수 있으며, 에탄올이 가장 바람직하다.The extract of persimmon flavor of the present invention can be obtained by extracting the root of the persimmon incense with water or an organic solvent, the lower alcohol, acetone, chloroform, methylene chloride, ether, ethyl acetate, hexane and the like can be exemplified. Lower alcohols include methanol, ethanol, propanol and butanol, with ethanol being most preferred.
구체적으로, 감송향 뿌리 건조물 또는 분말에 1 내지 20 배, 바람직하게는 5 내지 15 배, 더욱 바람직하게는 10배의 물을 첨가하고 80 내지 150℃, 바람직하게는 90 내지 100℃의 온도에서 1 내지 24시간, 바람직하게는 2 내지 6시간, 더욱 바람직하게는 2시간 동안 추출한 후 여과하여 감송향 뿌리의 열수 추출물을 제조할 수 있다. 감송향 뿌리의 유기용매 추출물은 감송향 뿌리 건조물 또는 분말에 1 내지 5 배, 바람직하게는 3 배의 유기용매를 첨가하고 실온에서 추출한 후 여과하여 얻어진 여액을 감압농축하여 제조할 수 있다. 상기 유기용매는 에탄올 또는 메탄올 추출물일 수 있다. 상기 추출방법들에서 추출공정은 필요에 따라 2회 이상 반복하여 실시할 수 있으며, 여과 후 얻어진 추출물을 동결 건조 또는 감압건조시켜 분말 형태로 만들 수도 있다.Specifically, 1-20 times, preferably 5-15 times, more preferably 10 times water is added to the dried persimmon root dry matter or powder and 1 at a temperature of 80-150 ° C., preferably 90-100 ° C. It may be extracted for 24 hours, preferably 2 to 6 hours, more preferably 2 hours and then filtered to prepare a hydrothermal extract of the persimmon root. The organic solvent extract of the persimmon fragrance root may be prepared by adding 1 to 5 times, preferably 3 times the organic solvent to the dried persimmon root dry matter or powder, extracting at room temperature, and then filtrating the filtrate obtained under reduced pressure. The organic solvent may be an ethanol or methanol extract. In the above extraction methods, the extraction process may be repeated two or more times as necessary, and the extract obtained after filtration may be freeze-dried or reduced-pressure drying to form a powder.
또한, 본 발명의 약학 조성물은 약리효과를 증진시키기 위해 약학적으로 허용가능한 다른 생약재 또는 그의 추출물을 추가로 포함할 수 있다. 이 경우, 상기 추출방법에 따라 생약재의 추출물을 제조한 후 약학 조성물에 가하거나 감송향 뿌리와 생약재를 혼합한 후 상기 방법으로 추출하여 얻어진 추출물을 조성물에 포함시킬 수도 있다. 상기에서 '약학적으로 허용가능한'이란 생리학적으로 허용되고 인간에게 투여될 때, 통상적으로 알레르기반응 또는 이와 유사한 반응을 일으키지 않는 것을 말한다.In addition, the pharmaceutical composition of the present invention may further include other pharmaceutically acceptable herbal medicines or extracts thereof to enhance the pharmacological effect. In this case, an extract of the herbal medicine may be prepared according to the extraction method, and then added to the pharmaceutical composition, or the extract obtained by extracting by the above method after mixing the saenghyang root and the herbal medicine may be included in the composition. As used herein, 'pharmaceutically acceptable' refers to a physiologically acceptable and, when administered to a human, usually does not cause an allergic or similar reaction.
본 발명의 조성물에 추가될 수 있는 생약재는 약학적으로 허용되는 임의의 생약재일 수 있으며, 예를 들면, 고본 (Angelicae tenuissimae Radix), 천마 (Gastrodiae Rhizoma), 시호 (Bapleuri Radix), 당귀 (Angelicae gigantis Radix), 도인 (Persicae Semen), 계지 (Cinnamomi Ramulus), 대황 (Rhei Rhizoma), 감초(Glycyrrhizae Radix), 천궁 (Cnidii Rhizoma), 진피 (Aurantii nobilis Pericarpium), 택사 (Alismatis Rhizoma), 황련 (Coptidis Rhizoma), 황금 (Scutellariae Radix), 복령 (Hoelen), 작약 (Paeoniae Radix), 백출 (Atractylodis Rhizoma alba), 황백 (Phellodendri Cortex), 치자 (Gardeniae Fructus), 반하 (Pinelliae Tuber), 조구등 (Uncaria Ramuluset Uncus), 지실 (Ponciri Fructus), 인삼 (Gingseng), 맥문동 (Liriopis Tuber), 원지 (Polygalae Radix), 석창포 (Acori graminei Rhizoma), 창출 (Atractylodis Rhizoma alba), 감국 (Chrysanthemi Flos), 방풍 (Ledebouriellae Radix), 생강 (Zingiberis Rhizoma crudus), 망초 (Natrii sulfas), 대조 (Zizyphi Fructus), 단삼 (Salviae Radix), 목단피 (Mautan Radicis Cortex), 지황 (Rehmanniae Radix), 박하 (Menthae Herba), 산약 (Dioscoreae Rhizoma), 저령 (Polyporus), 하수오 (Polygonimultiflori Radix), 구자 (Allii tuberosi Semen), 결명자 (Cassiae Semen), 구기자 (Lycii Fructus), 독활 (Araliae cordatae Radix), 두충 (Eucommiae Cortex), 백화사설초 (Hedyotis Herba), 삼백초 (Saururus Herba), 인진 (Artemisiaecapillaris Herba), 지모 (Anemarrhenae Rhizoma), 홍화 (Carthami Flos), 황기 (Astragali Radix), 석송자 (Lycopodium), 은행잎 (Ginkgonis Folium), 황정 (Polygonati Rhizoma), 연자육 (Nelumbinis Semen), 용골 (Fossilia ossis Mastodi), 지골피 (Lycii radicis Cortex), 우슬 (Achyranthis Radix), 숙지황 (Rehmanniae Radix preparata), 흑임자 (Perillae Semen), 백자인 (Thujae Semen), 맥아 (Hordei Fructus germinatus), 토사자 (Cuscutae Semen), 파극천 (Morindae Radix), 해송 (Pini koraiensis Radix) 등을 단독으로 또는 배합하여 사용할 수 있다.The herbal medicine that may be added to the composition of the present invention may be any pharmaceutically acceptable herbal medicine, for example, Angelicae tenuissimae Radix, Gastrodiae Rhizoma, Bapleuri Radix, Angelica gigantis Radix, Persicae Semen, Cinnamomi Ramulus, Rhei Rhizoma, Licorice (Glycyrrhizae Radix), Cnidii Rhizoma, Dermis (Aurantii nobilis Pericarpium), Taxa (Alismatis Rhizomaidi) ), Golden (Scutellariae Radix), Horyng (Hoelen), Peony (Paeoniae Radix), Bleach (Atractylodis Rhizoma alba), Peach (Phellodendri Cortex), Gardenia (Gardeniae Fructus), Pinelliae Tuber, Uncaria Ramuluset Uncus , Ponciri Fructus, Ginseng, Gingseng, Liriopis Tuber, Polygalae Radix, Acori graminei Rhizoma, Creation (Atractylodis Rhizoma alba), Chrysanthemi Flos, Windproof (Ledebouriellae Radix) Ginger (Zingiberis Rhizoma crudus), forget-me-not (Natr) ii sulfas, control (Zizyphi Fructus), Salviae Radix, Bark (Mautan Radicis Cortex), Rehanniae Radix, Peppermint (Menthae Herba), Herbal Medicine (Dioscoreae Rhizoma), Heraldic Acid (Polyporus), Sio (Polygonimultiflori) ), Goji (Allii tuberosi Semen), deficiency (Cassiae Semen), goji (Lycii Fructus), venom (Araliae cordatae Radix), tofu (Eucommiae Cortex), Hyedotis Herba, Sarurus Herba, Injin (Artemisiaecapillaris) Herba, Anemarrhenae Rhizoma, Safflower (Carthami Flos), Astragali Radix, Lycopodium, Ginkgois Folium, Ginkgois Folium, Polygonati Rhizoma, Neilbinis Semen, Keel (Fossilia ossis Masto) Lycii radicis Cortex, Achyranthis Radix, Rehmanniae Radix preparata, Perillae Semen, Thujae Semen, Maltese (Hordei Fructus germinatus), Cucumber Secudae (Morindae) ), Sea bream (Pini koraiensis Radix), etc. Or by a combination it may be used.
또한, 본 발명에 따른 약학적 조성물은 하나 이상의 약학적으로 허용되는 담체, 부형제 또는 희석제를 추가로 함유할 수 있다.In addition, the pharmaceutical compositions according to the invention may further contain one or more pharmaceutically acceptable carriers, excipients or diluents.
약학적으로 허용되는 담체로는 예컨대, 경구 투여용 담체 또는 비경구 투여용 담체를 추가로 포함할 수 있다.Pharmaceutically acceptable carriers may further include, for example, carriers for oral administration or carriers for parenteral administration.
경구 투여용 담체는 락토스, 전분, 셀룰로스 유도체, 마그네슘 스테아레이트, 스테아르산 등을 포함할 수 있다.Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid and the like.
또한, 비경구 투여용 담체는 물, 적합한 오일, 식염수, 수성 글루코스 및 글리콜 등을 포함할 수 있으며, 안정화제 및 보존제를 추가로 포함할 수 있다. 적합한 안정화제로는 아황산수소나트륨, 아황산나트륨 또는 아스코르브산과 같은 항산화제가 있다. 적합한 보존제로는 벤즈알코늄 클로라이드, 메틸- 또는 프로필-파라벤 및 클로로부탄올이 있다. 그 밖의 약학적으로 허용되는 담체로는 다음의 문헌에 기재되어 있는 것을 참고로 할 수 있다(Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, PA, 1995).In addition, carriers for parenteral administration may include water, suitable oils, saline, aqueous glucose and glycols, and the like, and may further include stabilizers and preservatives. Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol. Other pharmaceutically acceptable carriers may be referred to those described in the following documents (Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, PA, 1995).
본 발명의 만성 췌장염의 예방 또는 치료용 약학적 조성물은 인간을 비롯한 포유동물에 어떠한 방법으로도 투여할 수 있다. 예를 들면, 경구 또는 비경구적으로 투여할 수 있다. 비경구적인 투여방법으로는 이에 한정되지는 않으나, 정맥내, 근육내, 동맥내, 골수내, 경막내, 심장내, 경피, 피하, 복강내, 비강내, 장관, 국소, 설하 또는 직장내 투여일 수 있다. 바람직하게는 본 발명의 약학적 조성물은 경피 투여될 수 있다. 상기에서 '경피 투여'란 본 발명의 약학적 조성물을 세포 또는 피부에 투여하여 만성 췌장염의 예방 또는 치료용 약학적 조성물에 함유된 활성성분이 피부 내로 전달되도록 하는 것을 말한다. 예컨대, 본 발명의 약학적 조성물을 주사형 제형으로 제조하여 이를 30 게이지의 가는 주사 바늘로 피부를 가볍게 단자 (prick)하는 방법, 또는 피부에 직접적으로 도포하는 방법으로 투여될 수 있다.The pharmaceutical composition for preventing or treating chronic pancreatitis of the present invention can be administered to any mammal, including humans. For example, it can be administered orally or parenterally. Parenteral administration methods include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intradural, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, intestinal, topical, sublingual or rectal administration. Can be. Preferably the pharmaceutical composition of the present invention may be administered transdermally. In the above, 'transdermal administration' refers to the administration of the pharmaceutical composition of the present invention to cells or skin to deliver the active ingredient contained in the pharmaceutical composition for the prevention or treatment of chronic pancreatitis into the skin. For example, the pharmaceutical composition of the present invention may be prepared in an injectable formulation and administered by lightly pricking the skin with a 30 gauge thin injection needle, or directly applying to the skin.
본 발명의 약학적 조성물은 상술한 바와 같은 투여 경로에 따라 경구 투여용 또는 비경구 투여용 제제로 제형화할 수 있다.The pharmaceutical composition of the present invention may be formulated into a preparation for oral or parenteral administration according to the route of administration as described above.
경구 투여용 제제의 경우에 본 발명의 조성물은 분말, 과립, 정제, 환제, 당의정제, 캡슐제, 액제, 겔제, 시럽제, 슬러리제, 현탁액 등으로 당업계에 공지된 방법을 이용하여 제형화될 수 있다. 예를 들어, 경구용 제제는 활성성분을 고체 부형제와 배합한 다음 이를 분쇄하고 적합한 보조제를 첨가한 후 과립 혼합물로 가공함으로써 정제 또는 당의정제를 수득할 수 있다. 적합한 부형제의 예로는 락토즈, 덱스트로즈, 수크로즈, 솔비톨, 만니톨, 자일리톨, 에리스리톨 및 말티톨 등을 포함하는 당류와 옥수수 전분, 밀 전분, 쌀 전분 및 감자 전분 등을 포함하는 전분류, 셀룰로즈, 메틸 셀룰로즈, 나트륨 카르복시메틸셀룰로오즈 및 하이드록시프로필메틸-셀룰로즈 등을 포함하는 셀룰로즈류, 젤라틴, 폴리비닐피롤리돈 등과 같은 충전제가 포함될 수 있다. 또한, 경우에 따라 가교결합 폴리비닐피롤리돈, 한천, 알긴산 또는 나트륨 알기네이트 등을 붕해제로 첨가할 수 있다. 나아가, 본 발명의 약학적 조성물은 항응집제, 윤활제, 습윤제, 향료, 유화제 및 방부제 등을 추가로 포함할 수 있다.In the case of preparations for oral administration, the compositions of the present invention may be formulated using methods known in the art as powders, granules, tablets, pills, dragees, capsules, solutions, gels, syrups, slurries, suspensions and the like. Can be. For example, oral formulations can be obtained by tablets or dragees by combining the active ingredients with solid excipients, milling them, adding suitable auxiliaries and then processing them into granule mixtures. Examples of suitable excipients include sugars including lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol and maltitol and starch, cellulose, including starch, corn starch, wheat starch, rice starch and potato starch, etc. Fillers such as celluloses, gelatin, polyvinylpyrrolidone, and the like, including methyl cellulose, sodium carboxymethylcellulose, hydroxypropylmethyl-cellulose, and the like. In addition, crosslinked polyvinylpyrrolidone, agar, alginic acid or sodium alginate and the like may optionally be added as a disintegrant. Furthermore, the pharmaceutical composition of the present invention may further include an anticoagulant, a lubricant, a humectant, a perfume, an emulsifier, a preservative, and the like.
비경구 투여용 제제의 경우에는 주사제, 크림제, 로션제, 외용연고제, 오일제, 보습제, 겔제, 에어로졸 및 비강흡입제의 형태로 당업계에 공지된 방법으로 제형화할 수 있다. 이들 제형은 모든 제약 화학에 일반적으로 공지된 처방서인 문헌(Remington's Pharmaceutical Science, 15th Edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blaug, Seymour)에 기재되어 있다.Formulations for parenteral administration may be formulated by methods known in the art in the form of injections, creams, lotions, external ointments, oils, humectants, gels, aerosols and nasal inhalants. These formulations are described in Remington's Pharmaceutical Science, 15th Edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blaug, Seymour, a prescription generally known in all pharmaceutical chemistries.
본 발명의 조성물의 활성성분인 감송향 추출물의 총 유효량은 단일 투여량 (single dose)으로 환자에게 투여될 수 있으며, 다중 투여량 (multiple dose)으로 장기간 투여되는 분할 치료 방법 (fractionated treatment protocol)에 의해 투여될 수 있다. 본 발명의 약학적 조성물은 질환의 정도에 따라 유효성분의 함량을 달리할 수 있다. 바람직하게 본 발명의 감송향의 바람직한 전체 용량은 1일당 환자 체중 1 ㎏ 당 약 100 ㎍ 내지 5 ㎎, 가장 바람직하게는 500 ㎍ 내지 1 ㎎일 수 있다. 그러나, 상기 감송향 추출물의 용량은 약학적 조성물의 투여경로 및 치료 횟수뿐만 아니라 환자의 연령, 체중, 건강 상태, 성별, 질환의 중증도, 식이 및 배설율 등 다양한 요인들을 고려하여 환자에 대한 유효 투여량이 결정되는 것이므로, 이러한 점을 고려할 때 당 분야의 통상적인 지식을 가진 자라면 상기 감송향을 만성 췌장염의 예방 또는 치료제로서의 특정한 용도에 따른 적절한 유효 투여량을 결정할 수 있을 것이다. 본 발명에 따른 약학적 조성물은 본 발명의 효과를 보이는 한 그 제형, 투여 경로 및 투여 방법에 특별히 제한되지 아니한다.The total effective amount of the extract of Persimmon Fragrance, the active ingredient of the composition of the present invention, may be administered to a patient in a single dose, and may be administered in a fractionated treatment protocol administered for a long time in multiple doses. May be administered. The pharmaceutical composition of the present invention may vary the content of the active ingredient depending on the extent of the disease. Preferably the preferred total dose of the sweet persimmon of the present invention may be about 100 μg to 5 mg, most preferably 500 μg to 1 mg per kg of patient body weight per day. However, the dose of the extract is effective for taking into consideration the various factors such as the age, weight, health status, sex, severity of the disease, diet and excretion rate, as well as the route and frequency of treatment of the pharmaceutical composition Since the amount is determined, one of ordinary skill in the art will be able to determine the appropriate effective dosage according to the specific use of the sensational aroma as a prophylactic or therapeutic agent for chronic pancreatitis. The pharmaceutical composition according to the present invention is not particularly limited to its formulation, route of administration and method of administration as long as the effect of the present invention is shown.
또한, 본 발명은 감송향 (Nardostachys jatamansi NJ) 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 개선용 식품 조성물의 형태로 제공될 수 있다. 본 발명의 식품 조성물은 기능성 식품 (functional food), 영양 보조제 (nutritional supplement), 건강식품 (health food) 및 식품 첨가제 (food additives) 등의 모든 형태를 포함한다. 상기 유형의 식품 조성물은 당업계에 공지된 통상적인 방법에 따라 다양한 형태로 제조할 수 있다.In addition, the present invention may be provided in the form of a food composition for the prevention and improvement of chronic pancreatitis comprising the extract of Nardostachys jatamansi NJ as an active ingredient. The food composition of the present invention includes all forms such as functional foods, nutritional supplements, health foods and food additives. Food compositions of this type can be prepared in various forms according to conventional methods known in the art.
예를 들면, 건강식품으로는 본 발명의 감송향 추출물 자체를 차, 쥬스 및 드링크의 형태로 제조하여 음용하도록 하거나, 과립화, 캡슐화 및 분말화하여 섭취할 수 있다. 또한, 본 발명의 감송향 추출물과 만성 췌장염의 예방 및 개선효과가 있다고 알려진 공지의 물질 또는 활성성분과 함께 혼합하여 조성물의 형태로 제조할 수 있다.For example, as a health food, the persimmon extract of the present invention may be prepared by drinking tea, juice, and drink, or granulated, encapsulated, and powdered. In addition, it can be prepared in the form of a composition by mixing with the extract of the sense of flavor of the present invention and known substances or active ingredients known to have the effect of preventing and improving chronic pancreatitis.
또한, 기능성 식품으로는 음료 (알콜성 음료 포함), 과실 및 그의 가공식품 (예: 과일통조림, 병조림, 잼, 마아말레이드 등), 어류, 육류 및 그 가공식품 (예: 햄, 소시지콘비이프 등), 빵류 및 면류 (예: 우동, 메밀국수, 라면, 스파게티, 마카로니 등), 과즙, 각종 드링크, 쿠키, 엿, 유제품 (예: 버터, 치즈 등), 식용식물유지, 마아가린, 식물성 단백질, 레토르트 식품, 냉동식품, 각종 조미료 (예: 된장, 간장, 소스 등) 등에 본 발명의 감송향 추출물을 첨가하여 제조할 수 있다.In addition, functional foods include beverages (including alcoholic beverages), fruits and processed foods (e.g. canned fruit, canned foods, jams, marmalade, etc.), fish, meat and processed foods (e.g. ham, sausage cornebipe) Etc.), breads and noodles (e.g. udon, soba noodles, ramen, spaghetti, macaroni, etc.), fruit juices, various drinks, cookies, syrups, dairy products (e.g. butter, cheese, etc.), edible vegetable oils, margarine, vegetable protein, It can be prepared by adding the extract of persimmon flavor of the present invention to retort foods, frozen foods, various seasonings (eg, miso, soy sauce, sauce, etc.).
또한, 본 발명의 감송향을 식품 첨가제의 형태로 사용하기 위해서는 분말 또는 농축액 형태로 제조하여 사용할 수 있다.In addition, in order to use the persimmon aroma of the present invention in the form of food additives can be prepared in the form of powder or concentrate.
참고로, 본 발명에서 언급한 뉴클레오티드 및 단백질 작업에는 다음의 문헌을 참조할 수 있다 (Maniatis et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.(1982); Sambrook et al., Molecular Cloning: A Laboratory Manual, 2d Ed., Cold Spring Harbor Laboratory Press(1989); Deutscher, M., Guide to Protein Purification Methods Enzymology, vol. 182.)For reference, reference may be made to nucleotide and protein operations mentioned in the present invention (Maniatis et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY (1982); Sambrook et. al., Molecular Cloning: A Laboratory Manual, 2d Ed., Cold Spring Harbor Laboratory Press (1989); Deutscher, M., Guide to Protein Purification Methods Enzymology, vol. 182.)
췌장의 섬유증은 오랫동안 거의 또는 전혀 개선에 대한 희망이 없는 말기 단계인 것으로 간주되어 왔다. 만성 췌장염(CP)에 깔려있는 생리학적 기전과 이러한 과정에서 주요한 작용 세포인 성상세포의 특징에 대한 보다 진전된 이해를 통해서 섬유증이 퇴행되는 가능성에 대하여 보다 집중적인 주의를 갖는 것이 유일한 수단으로 간주되었다. 본 발명에서, 최근에 보고된 알콜성 만성 췌장염(ACP) 모델 (Charrier, A.L. and Brigstock, D.R. (2010) Lab. Invest. 90, 1179-1188)을 사용하여, 췌장 포상세포, 콜라겐 축적 및 췌장 성상세포(PSC) 활성화를 억제시킴에 의하여, 감송향 추출물(NJ)이 ACP의 진행을 감소시킴을 입증하였다.Pancreatic fibrosis has long been considered a late stage with little or no hope for improvement. With a more advanced understanding of the physiological mechanisms underlying chronic pancreatitis (CP) and the characteristics of astrocytic cells, the major working cells in this process, a more focused attention to the possibility of fibrosis regression was considered the only means. . In the present invention, using the recently reported alcoholic chronic pancreatitis (ACP) model (Charrier, AL and Brigstock, DR (2010) Lab. Invest. 90 , 1179-1188), pancreatic acinar cells, collagen accumulation and pancreatic appearance By inhibiting cellular (PSC) activation, it was demonstrated that the Persimmon Fragrance Extract (NJ) reduced the progression of ACP.
AP는 통증, 부종, 출혈, 포상 세포의 액포화, 괴사, 염증, 및 증가된 혈청 아밀라제 및 리파제에 의해 특징된다; 이러한 특징은 섬유증, 염증, 콜라겐 축적 및 감소된 외분비 및 내분비 기능에 의한 만성적 질환 형태와 혼동된다. 비록, AP와 CP가 병원성 원인이 다르지만, 반복적 AP는 점차적으로 CP로의 진행을 가져올 수 있다(Ohashi, S. 등 (2006) Am. J. Physiol. Gastrointest. Liver Physiol. 290, G772-781). 그러나, 세루레인 모델의 반복적 투여는 처리 시간, 심각성, 및 인간과의 유사성에서 한정적이었다. 그러므로, 만성적인 과도한 알콜 소비가 만성 췌장염으로 진행하는데 주요한 위험 인자이기 때문에, 본 발명자들은 반복적 AP 모델에 알콜 소비를 첨가함으로써 ACP를 유도하였다(Oruc, N. and Whitcomb, D.C. (2004) Gastroenterol. Clin. North Am. 33, 733-750 ; Apte, M.V. and Wilson, J.S. (2003) Best Pract. Res. Clin. Gastroenterol. 17, 593-612 ; Schenker, S. and Montalvo, R. (1998) Recent Dev. Alcohol 14, 41-65 ; Whitcomb, D.C., Ulrich, C.D. (1999) Baillieres Best Pract. Res. Clin. Gastroenterol. 13, 253-263). 마우스에 3주 동안 주어진 도전으로 ACP를 유도하는데 충분하였고, 상기 ACP는 췌장에서 섬유증, 염증 및 포상세포 파괴에 의해 입증되었다 (도 1 참조). 매일 및 자유 NJ의 소비는 ACP에 대한 형태학적 손상을 억제하였다(도 1 참조). 아밀라제 양성 세포가 췌장에 풍부하므로, 외분비 기능이 적절하게 작용하고 있음을 암시한다.AP is characterized by pain, edema, bleeding, saturation of acinar cells, necrosis, inflammation, and increased serum amylase and lipase; This feature is confused with chronic disease forms due to fibrosis, inflammation, collagen accumulation and reduced exocrine and endocrine function. Although AP and CP differ in pathogenic causes, repetitive AP can gradually lead to CP (Ohashi, S. et al. (2006) Am. J. Physiol. Gastrointest. Liver Physiol. 290 , G772-781). However, repeated administration of the cerulein model was limited in treatment time, severity, and similarity with humans. Therefore, since chronic excessive alcohol consumption is a major risk factor for progression to chronic pancreatitis, we induced ACP by adding alcohol consumption to the repeated AP model (Oruc, N. and Whitcomb, DC (2004) Gastroenterol. Clin North Am. 33 , 733-750; Apte, MV and Wilson, JS (2003) Best Pract.Res. Clin.Gastroenterol. 17 , 593-612; Schenker, S. and Montalvo, R. (1998) Recent Dev. Alcohol 14 , 41-65; Whitcomb, DC, Ulrich, CD (1999) Baillieres Best Pract. Res. Clin. Gastroenterol. 13 , 253-263). Mice were sufficient to induce ACP with a challenge given for 3 weeks, which was demonstrated by fibrosis, inflammation, and acinar cell destruction in the pancreas (see FIG. 1). Daily and free NJ consumption inhibited morphological damage to ACP (see FIG. 1). Amylase-positive cells are abundant in the pancreas, suggesting that exocrine function is functioning properly.
반복적 AP를 위하여, 췌장은 염증 및 괴사에 도달하고, 이것은 결국 사이토카인 및 기타 자극인자의 방출을 가져왔다(Omary, M. 등 (2007) J. Clin. Invest. 117, 50-59 ; Apte, M.V. 등 (1999) Gut 44, 534-541 ; Apte, M.V. 등 (2006) J. Gastroenterol. Hepatol. 21, S97-S101). TGF-β와 같은 프로-섬유조직 발생 매개체에 의해 활성화 될 때, PSC는 α-SMA를 발현하는 근섬유아세포와 같은 세포로 전환되고, 피브로넥틴 및 콜라겐이 섬유증 부위 근처에 축적된다(Schneider, E. 등 (2001) Am. J. Physiol. Cell Physiol. 281, C532-543). 본 발명자들은 ACP 마우스는 콜라겐 축적, α-SMA, 피브로넥틴 및 TGF-β의 상당한 증가가 이루어짐을 탐지하였다. 또한, 활성화된 PSC (데스민으로 확인)는 증가된 α-SMA, 피브로넥틴 및 TGF-β를 나타내었다. 그러나, 섬유증 증상은 α-SMA, 피브로넥틴 및 TGF-β 및 콜라겐 축적의 감소에 의해 나타낸 바와 같이, NJ 처리는 섬유증 반응을 상당히 약화시켰다. 이러한 결과는 TGF-β 생성 억제를 통한 PSC 비활성화와 감소된 섬유증이 연관련됨을 암시한다.For repetitive AP, the pancreas reaches inflammation and necrosis, which eventually led to the release of cytokines and other stimulators (Omary, M. et al. (2007) J. Clin. Invest. 117 , 50-59; Apte, (1999) Gut 44 , 534-541; Apte, MV et al. (2006) J. Gastroenterol. Hepatol. 21 , S97-S101). When activated by pro-fibroblast generating media such as TGF-β, PSCs are converted to cells such as myofibroblasts expressing α-SMA, and fibronectin and collagen accumulate near the fibrotic site (Schneider, E. et al. (2001) Am. J. Physiol. Cell Physiol. 281 , C532-543). We detected that ACP mice had a significant increase in collagen accumulation, α-SMA, fibronectin and TGF-β. In addition, activated PSCs (identified with desmine) showed increased α-SMA, fibronectin and TGF-β. However, NJ treatment significantly attenuated the fibrosis response, as fibrosis symptoms were indicated by a decrease in α-SMA, fibronectin and TGF-β and collagen accumulation. These results suggest that PSC inactivation through inhibition of TGF-β production is associated with decreased fibrosis.
ACP의 병-생리학에서 주요한 의문은 질환 상태가 역전될 수 있는지이다. 많은 임상의들은 흔히 환자에게 알콜을 억제할 것을 권고하지만, 이것은 환자에게 거의 영향을 주지 못한다. 알콜 섭취를 중단함에 의해 ACP를 호전시키는 지에 대한 앞서의 연구들은 실질적인 개선을 찾지 못했다 (Apte, M.V. 등 (2010) J. Gastroenterol. Hepatol. 25,1816-1826 ; Gullo, L. 등 (1988) Gastroenterology 95, 1063-1068). 인간과는 대비되게, 쥐의 AP 및 ACP 모델은 일반적으로 자극의 결여이후에 질환이 개선된다 (Lugea, A. 등 (2006) Gastroenterology 131, 885-899 ; Vonlaufen, A. 등 (2010) Gut 60, 238-246). 그러므로, 쥐의 실험 모델은 인간 CP의 기전을 입증하는데 충분하지 않다. 그러나, 본 발명자들은 인간 CP를 치료하는 임상 자원을 제공하기 위하여 실험적 모델의 회복 기전에 초점을 맞추었다. ACP의 퇴행에서, PSC의 조절이 핵심 인자이다(Masamune, A. 등 (2009) Clin. Gastroenterol. Hepatol. 7, S48-54). 정말로, 많은 연구들이 PSC 인자를 비활성 또는 사멸시킴에 의해 PSC를 조절하고자 노력하였다 (Schwer, C.I. 등 (2008) J. Pharmacol. Exp. Ther. 327, 863-871 ; Li, L. 등 (2011) Eur. J. Clin. Invest. 41, 151-158 ; Madro, A. 등 (2008) J. Physiol. Pharmacol. 59, 239-249 ; Rickmann, M. 등 (2007) Gastroenterology 132, 2518-2132 ; Long, D. 등 (2011) J. Surg. Res. 176, 248-259 ; Aoki, H. 등 (2007) Am. J. Physiol. Cell Physiol. 292, C259-268). 본 발명의 ACP 모델에서, NJ 처리는 췌장에서 α-SMA, 피브로넥틴, 및 콜라겐 발현의 감소에 의해 나타낸 바와 같이, PSC의 비활성화를 가져왔다. 본 발명자들 분리된 쥐 PSC를 사용하여, PSC 활성화에 NJ의 직접적 효과를 조사하였다. NJ 처리는 PSC 사멸을 유도하지 않았다 (데이터 미제시). 도 4에 제시한 바와 같이, 감소된 섬유증 마커에 의해 나타낸 바와 같이, NJ는 충분히 활성화된 PSC를 역전시켰다. 이러한 결과는 섬유증에 대한 NJ의 작용 기전이 PSC를 사멸시키는 것이 아니라 PSC를 비활성화시킬 수 있음을 암시한다.The main question in the disease-physiology of ACP is whether the disease state can be reversed. Many clinicians often recommend patients to suppress alcohol, but this has little effect on patients. Prior research on how to improve the ACP By discontinue their alcohol intake did not find a substantial improvement (Apte, MV, etc. (2010) J. Gastroenterol Hepatol 25, 1816-1826;.. Gullo, L. , etc. (1988) Gastroenterology 95 , 1063-1068). In contrast to humans, rat AP and ACP models generally improve disease after lack of stimulation (Lugea, A. et al. (2006) Gastroenterology 131 , 885-899; Vonlaufen, A. et al. (2010) Gut 60 , 238-246). Therefore, experimental models of mice are not sufficient to demonstrate the mechanism of human CP. However, we focused on the recovery mechanism of the experimental model to provide clinical resources for treating human CP. In the degeneration of ACP, regulation of PSC is a key factor (Masamune, A. et al. (2009) Clin. Gastroenterol. Hepatol. 7 , S48-54). Indeed, many studies have tried to modulate PSC by inactivating or killing PSC factors (Schwer, CI et al. (2008) J. Pharmacol. Exp. Ther. 327 , 863-871; Li, L. et al. (2011) Eur. J. Clin. Invest. 41 , 151-158; Madro, A. et al. (2008) J. Physiol.Pharmacol . 59 , 239-249; Rickmann, M. et al. (2007) Gastroenterology 132 , 2518-2132; Long , D. et al. (2011) J. Surg. Res. 176 , 248-259; Aoki, H. et al. (2007) Am. J. Physiol. Cell Physiol. 292 , C259-268). In the ACP model of the present invention, NJ treatment resulted in inactivation of PSCs, as indicated by a decrease in α-SMA, fibronectin, and collagen expression in the pancreas. We isolated murine PSCs to investigate the direct effect of NJ on PSC activation. NJ treatment did not induce PSC killing (data not shown). As shown in FIG. 4, as shown by the reduced fibrosis marker, NJ reversed fully activated PSCs. These results suggest that the mechanism of action of NJ on fibrosis may inactivate PSC rather than kill PSC.
본 발명자들은 NJ의 투여가 마우스에서 췌장 섬유증의 진행을 방지함을 입증하였다. 염증 및 콜라겐 축적의 감소가 ACP에 NJ의 가능성을 암시한다. 결론적으로, 본 발명의 결과는 NJ가 PSC의 비활성화를 통해 ACP에 대한 항-섬유증 효과를 가짐을 나타낸다.We demonstrated that administration of NJ prevents the progression of pancreatic fibrosis in mice. Inflammation and reduced collagen accumulation suggest the possibility of NJ in ACP. In conclusion, the results of the present invention show that NJ has an anti-fibrotic effect on ACP through inactivation of PSC.
상기와 같이 구성되는 본 발명은 감송향의 신규한 용도로서, 감송향 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 치료용 조성물을 제공한다. 본 발명의 감송향 추출물을 포함하는 조성물은 세룰레인 및 에탄올에 의해서 유발되는 만성 췌장염을 효과적으로 억제하므로 만성 췌장염의 예방 및 치료를 위하여 효과적으로 사용될 수 있다.The present invention constituted as described above provides a novel use of persimmon aroma, chronic pancreatitis prevention and treatment composition comprising the persimmon extract as an active ingredient. The composition comprising the extract of the present invention, can be effectively used for the prevention and treatment of chronic pancreatitis because it effectively inhibits chronic pancreatitis caused by cerulein and ethanol.
도 1은 ACP 이후에 췌장의 형태 및 조직에 대한 NJ 효과를 나타낸 것으로, (A)는 대표적 췌장 외부 형태, (B)는 췌장의 대표적 H&E 염색된 단편, (C)는 염증, 부종, 섬유증, 및 전체에 대하여 0(정상)에서 3(심각)까지 췌장의 조직학적 단편을 기록하였다. 상기 사진은 그룹당 6마리의 마우스를 1 실험 테스트의 대표적 이미지를 나타낸다. 상기 결과는 3번의 추가적 실험에서 유사하였다. 최초의 확대(ㅧ200). *P ＜ 0.05, 대조군은 생리식염수 처리.Figure 1 shows the NJ effect on the morphology and tissue of the pancreas after ACP, (A) is a representative pancreatic external morphology, (B) is a representative H & E stained fragment of the pancreas, (C) is inflammation, edema, fibrosis, And histological fragments of the pancreas were recorded from 0 (normal) to 3 (severe) for the whole. The photo shows a representative image of one experimental test with six mice per group. The results were similar in three additional experiments. First magnification (ㅧ 200). * P <0.05, the control group was treated with physiological saline.
도 2는 ACP 동안 포상세포(acinar cell) 사멸에 NJ 효과를 나타낸 것으로, 아밀라제로 췌장 조직 단편을 연속적으로 염색하여 포상세포의 존재를 탐지하였다. 상기 사진은 그룹당 6마리의 마우스를 1실험 테스트의 대표적 이미지를 나타낸다. 상기 결과는 3 추가적인 실험에서 유사하였다. Figure 2 shows the effect of NJ on acinar cell death during ACP, and the presence of acinar cells was detected by successive staining of pancreatic tissue fragments with amylase. The photo shows a representative image of one experimental test with six mice per group. The results were similar in 3 additional experiments.
도 3은 ACP 동안 PSC 활성화 및 콜라겐 축적의 효과를 나타낸 것으로, 췌장 조직 절편을 (A) α-SMA 및 (C) 콜라겐으로 연속적으로 염색하여 활성화된 PSC를 탐지하였다. 상기 사진은 그룹당 6마리의 마우스를 1실험 테스트의 대표적 이미지를 나타낸다. 최초의 확대 (ㅧ400). (B)는 α-SMA, 피브로넥틴(fibronectin) 및 TGF-β의 췌장 mRNA 수준을 정량 RT-PCR로 측정하였다. 데이터는 각 그룹에서 6마리 마우스의 평균 ㅁ SE를 나타낸다. 상기 결과는 3번의 추가적 실험에서 유사하였다. *P < 0.05, 대조군은 생리식염수 처리Figure 3 shows the effect of PSC activation and collagen accumulation during ACP, pancreatic tissue sections were sequentially stained with (A) α-SMA and (C) collagen to detect activated PSCs. The photo shows a representative image of one experimental test with six mice per group. First magnification (ㅧ 400). (B) measured pancreatic mRNA levels of α-SMA, fibronectin and TGF-β by quantitative RT-PCR. The data represents the mean SE SE of 6 mice in each group. The results were similar in three additional experiments. * P <0.05, control group treated with saline solution
도 4는 분리된 PSC에서 PSC 활성화에 NJ의 효과를 나타낸 것으로, PSC를 쥐 췌장으로부터 분리하고, 3주 동안 배양 활성화시켰다. 21일째에, 24시간 동안 NJ를 처리하였다. 그리고 나서, (A) 데스민(desmin) 및 (B) α-SMA를 처리하여 성공적으로 PSC 분리 및 활성화를 탐지하였다. 최초의 확대 (ㅧ400). (C)는 α-SMA의 발현을 웨스턴 블럿으로 탐지하였다. (D)는 사이토카인의 mRNA 발현을 실시간 RT-PCR에 의해 PSC로 정량화하였다. 상기 사진은 1 실험의 대표적 이미지를 나타낸다. 상기 결과는 3번의 추가적 실험에서 유사하였다. *P ＜ 0.05, 대조군은 생리식염수 처리.Figure 4 shows the effect of NJ on PSC activation in isolated PSC, PSC was isolated from rat pancreas and culture activated for 3 weeks. On day 21, NJ was treated for 24 hours. Then, (A) desmin and (B) α-SMA were treated successfully to detect PSC isolation and activation. First magnification (ㅧ 400). (C) detected Western blot expression of α-SMA. (D) quantified cytokine mRNA expression by PSC by real-time RT-PCR. The photograph shows a representative image of one experiment. The results were similar in three additional experiments. * P <0.05, the control group was treated with physiological saline.
이하, 실시예를 통하여 본 발명을 보다 상세히 설명하고자 한다. 이들 실시예는 오로지 본 발명을 보다 구체적으로 설명하기 위한 것으로, 본 발명의 요지에 따라 본 발명의 범위가 이들 실시예에 의해 제한되지 않는다는 것은 당업계에 통상의 지식을 가진 자에게 있어서 자명할 것이다.Hereinafter, the present invention will be described in more detail with reference to Examples. These examples are only for illustrating the present invention in more detail, it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples in accordance with the gist of the present invention. .
<참조예> 통계 분석Reference Example Statistical Analysis
모든 데이터는 평균ㅁ통계오차 (SE)로 표시하였다. 변화의 유의성을 시간과 투여량 파라미터를 갖는, 2-방식 ANOVA를 사용하여 검정하였다. 실험군간의 차이는 변량분석(analysis of variance)을 사용하여 검정하였다. P<0.05인 값은 통계학적으로 유의한 것으로 간주하였다.All data are expressed as mean square error (SE). The significance of the change was assayed using a two-way ANOVA with time and dose parameters. Differences between experimental groups were tested using an analysis of variance. Values with P <0.05 were considered statistically significant.
<실시예 1> 실험 재료 및 동물Example 1 Experimental Materials and Animals
<1-1> 화합물 및 시약의 준비<1-1> Preparation of Compounds and Reagents
세룰레인 Tris-HCl, NaCl, 콜라겐, DAPI, 헤마토자일린(hematoxylin), 에오신(eosin), 자일렌(xylene), 에탄올, 세룰레인(cerulein) 및 트리톤 X-100은 Sigma-Aldrich(St. Louis, MO, USA)로부터 구입하였다. α-평활근 액틴(smooth muscle actin ; SMA), 콜라겐, 및 데스민은 Abcam (UK)로부터 구입하였다. 아밀라제 및 GAPDH는 Santa Cruz Biotechnology (Santa Cruz, CA, USA)로부터 구입하였다.Cerulean Tris-HCl, NaCl, collagen, DAPI, hematoxylin, eosin, xylene, ethanol, cerulein and Triton X-100 are Sigma-Aldrich (St. Louis, MO, USA). α-smooth muscle actin (SMA), collagen, and desmine were purchased from Abcam (UK). Amylase and GAPDH were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).
<1-2> 실험 동물의 준비<1-2> Preparation of experimental animals
모든 동물 실험은 원광대학교 동물 관리 위원회에 의해 승인된 프로토콜에 따라 수행되었다. 실험동물은 15-20 g 체중의 생후 6 내지 8주된 C57BL/6 마우스 암컷 (체중 15-20 g)을 구입(오리엔트 바이오)하였다. 모든 실험동물은 7일 동안 실내온도 23ㅁ2℃, 명암주기 12시간의 사육실에서 사육하였다. 실험동물은 표준 사료와 물을 완전 자유급식 (ad libitum)으로 7일간 공급하여 환경에 적응시킨 후, 임의로 대조군과 실험군으로 나누었다. 각 마우스들은 알콜성 만성 췌장염(ACP)을 유도하기 전에 18시간 동안 절식시켰다.All animal experiments were performed according to a protocol approved by Wonkwang University Animal Care Committee. Experimental animals purchased 15-20 g body weight 6-8 week old C57BL / 6 mouse females (15-20 g body weight) (Orient Bio). All experimental animals were bred for 7 days in a breeding room with a room temperature of 23 ㅁ 2 ℃ and a contrast cycle of 12 hours. The experimental animals were fed with standard feed and water as ad libitum for 7 days to adapt to the environment, and then randomly divided into control and experimental groups. Each mouse was fasted for 18 hours before inducing alcoholic chronic pancreatitis (ACP).
<실시예 2> 감송향 추출물의 제조Example 2 Preparation of Persimmon Extract
말린 감송향 뿌리를 일반적인 구입처(Omni Herb, Seoul, Korea)에서 구입하였다. 상기 약초에 대한 동정은 한국약물시험연구소(Korean drug test laboratory)에서 확인하였다. 확증 표본(voucher specimens)(NO; Oh/wh/nj-43)들을 원광대 한의학과 식물표본실에 기탁하였다.Dried persimmon roots were purchased from General Herbs (Omni Herb, Seoul, Korea). Identification of the herb was confirmed in the Korean drug test laboratory. Voucher specimens (NO; Oh / wh / nj-43) were deposited in Wonkwang University Oriental Medicine Plant Botanical Laboratory.
상기 감소향 뿌리에 약 10 배의 증류수에 넣고 약 2시간 끓인 다음, 추출물을 여과하고 -70℃에서 동결 건조하여 분말화하였다. 상기 분말을 증류수로 추출한 후 여과하여 4℃에 보관하였다. 상기 분말은 생리식염수에 녹여 필요한 농도로 사용하였다.The distilled root was put in distilled water of about 10 times and boiled for about 2 hours, and then the extract was filtered and lyophilized at -70 ° C to powder. The powder was extracted with distilled water, filtered and stored at 4 ° C. The powder was dissolved in physiological saline and used at the required concentration.
<실시예 3> 알콜성 만성 췌장염의 유도Example 3 Induction of Alcoholic Chronic Pancreatitis
본 발명자들은 Charrier 및 Brigstock이 보고한 빠르고 효율적 모델을 사용하였다(Charrier, A.L. and Brigstock, D.R. (2010) Lab. Invest. 90, 1179-1188). 요약하면, 6-8주령의 C57Bl/6 마우스의 복강에 하루에 한 번씩 주당 6번에 걸쳐 3주 동안 에탄올(3.2 g/㎏; 33.3% 에탄올: 67.7% 물 용액으로 투여)을 투여하였다. 각주의 특정일에, 50 ㎍/㎏의 세룰레인(cerulein)을 상기 1-2에서 준비한 마우스의 복강에 6시간 동안 매시간 투여하였다. 마우스를 사육실에 3번 옮겨서 저지방 다이어트 자유급식(ad libitum)으로 공급하였다. 마지막 에탄올 처리 하루 뒤에 마우스를 희생시키고 췌장을 절취하여 4% 파라포르알데히드 (pH 7.2-7.4)에 고정하거나 RNA 및 단백질 추출에 사용하였다.We used a fast and efficient model reported by Charrier and Brigstock (Charrier, AL and Brigstock, DR (2010) Lab. Invest. 90 , 1179-1188). In summary, ethanol (3.2 g / kg; 33.3% ethanol: administered with 67.7% water solution) was administered to the abdominal cavity of 6-8 week old C57Bl / 6 mice once a day for 3 weeks over 6 times per week. On specific days of each week, 50 μg / kg of cerulein was administered to the abdominal cavity of the mice prepared in 1-2 above for 6 hours every hour. The mice were transferred to the nursery three times and fed with a low fat diet ( ad libitum ). Mice were sacrificed one day after the last ethanol treatment and the pancreas was excised and fixed in 4% paraformaldehyde (pH 7.2-7.4) or used for RNA and protein extraction.
<실험예>Experimental Example
<1> 조직학적 검사 <1> histological examination
고정된 췌장 조직을 파라핀에 묻고, 4 ㎛ 절편으로 절단하여 표준 조직학적 검사를 위하여 해마토자일린-에오신으로 염색하였다. 아밀라제, 콜라겐 또는 α-SMA에 의한 면역조직학적 염색을 DAB immunohistohemical kit (DAKO, Cytomation, Denmark)을 사용하여 수행하였다. 항-α-SMA, 또는 데스민과 함께 PSC의 배양, 이어서 실온에서 1시간 동안 Alexa Fluors 568 goat-anti rabbit IgG (1：2,000, Invitrogen, Carlsbad, CA, USA)으로 배양으로, DAPI, α-SMA 또는 데스민의 면역형광학적 탐지를 수행하였다. DAPI(Vector Laboratories, Burlingame, CA, USA)가 구비된 mounting Medium으로 슬라이드를 마우팅시켰다.Immobilized pancreatic tissue was embedded in paraffin, cut into 4 μm sections and stained with haematoxyl-eosin for standard histological examination. Immunohistochemical staining with amylase, collagen or α-SMA was performed using a DAB immunohistohemical kit (DAKO, Cytomation, Denmark). Incubation of PSC with anti-α-SMA, or desmin, followed by incubation with Alexa Fluors 568 goat-anti rabbit IgG (1: 2,000, Invitrogen, Carlsbad, Calif., USA) for 1 hour at room temperature, DAPI, α- Immunofluorescence detection of SMA or desmine was performed. The slides were mounted with mounting medium equipped with DAPI (Vector Laboratories, Burlingame, CA, USA).
<2> 메신저 RNA (mRNA) 발현<2> messenger RNA (mRNA) expression
마우스 췌장 조직 및 PSC에서 RT-PCR에 의하여 타겟 사이토카인의 전사를 분석하였다. TriZol (Invitrogen, Carlsbad, CA, USA)을 사용하여 마우스의 췌장 및 PSC으로부터 전체 RNA를 분리하고 SuperScript II RT (Invitrogen, Carlsbad, CA, USA)를 사용하여 역전사를 수행하였다. 제조자 (Invitrogen, Carlsbad, CA, USA)의 지시에 따라 ABI stepone plue system을 사용한 TaqMan 정량 RT-PCR을 수행하였다. 각 시료에 있어서, 관심 유전자의 발현을 위하여 3번의 반독 테스트 반응과 역전사가 결여된 대조군 반응을 분석하였고, 그 결과를 "하우스키핑(housekeeping)" HGPRT(hypoxanthine-guanine phosphoribosyltransferase) mRNA의 발현으로 표준화시켰다. 임의의 발현 유닛은 관심 유전자의 발현량을 리보좀 단백질 HPRT mRNA 발현량으로 나누어 계산하였다. 다중 실시간 TaqMan PCR을 위한 정방향, 역방향, 및 프로브 올리고뉴클레오티드 프라이머를 ABI (Invitrogen, Carlsbad, CA, USA)로부터 구입하였다.Target cytokine transcription was analyzed by RT-PCR in mouse pancreatic tissue and PSC. Total RNA was isolated from pancreas and PSC of mice using TriZol (Invitrogen, Carlsbad, CA, USA) and reverse transcription was performed using SuperScript II RT (Invitrogen, Carlsbad, CA, USA). TaqMan quantitative RT-PCR was performed using the ABI stepone plue system according to the manufacturer's instructions (Invitrogen, Carlsbad, CA, USA). For each sample, three anti-toxin test reactions and a control reaction lacking reverse transcription were analyzed for expression of the gene of interest, and the results were normalized to the expression of "housekeeping" hypoxanthine-guanine phosphoribosyltransferase (HGPRT) mRNA. . Any expression unit was calculated by dividing the expression level of the gene of interest by the expression level of ribosomal protein HPRT mRNA. Forward, reverse, and probe oligonucleotide primers for multiple real-time TaqMan PCR were purchased from ABI (Invitrogen, Carlsbad, CA, USA).
<3> PSC 분리<3> PSC separation
Shinji 등 (Shinji, T. 등 (2002) Acta. Med. Okayama 56, 211-218)에 설명된 절차에 따라서, 250 내지 300 g의 체중을 갖는 수컷 Wistar rat (Orient Bio, Sungnam, KyungKiDo, South Korea)의 췌장 조직으로부터 쥐 PSC를 제작하였다. 요약하면, Hank의 완충 염용액에서 0.03% 콜라겐나제 P를 갖고 췌장을 분해시켰다. 그 결과되는 세포 현탁액을 20분 동안 1,400 g에서 3.2% 이오헥솔(iohexol) 구배로 원심분리시켰다. 성상세포(Stellate cell)를 상기 이오헥솔 용액 및 수용성 완충액의 접촉면 바로 위의 혼계면(fuzzy band)내로 분리시켰다. 상기 혼계면을 수확하고, 세포를 세척한 후 10% 우태아혈청(fetal bovine serum ; FBS), 4 mM 글루타민, 및 항생제(페니실린 100 U/㎖, 스트렙토마이신 100 ㎎/㎖)를 함유한 DMEM에 재현탁시켰다. 컨플루언신(confluency)에 도달한 후에, 세포를 수확하고 동일한 씨앗 밀도로 다시 플레이팅시켰다. 모든 실험은 배양-수확된 세포 (2-4 세대)를 사용하여 수행하였다. 실험 시약의 첨가 24시간 전에 혈청-결여 배지에서 PSC를 배양하였다.Shinji etc Shinji, T. et al. (2002)Acta. Med. Okayama                 56, 211-218), were constructed from pancreatic tissue from male Wistar rats (Orient Bio, Sungnam, KyungKiDo, South Korea) weighing 250-300 g. In summary, the pancreas was digested with 0.03% collagenase P in Hank's buffered saline solution. The resulting cell suspension was centrifuged at 3.2% iohexol gradient at 1,400 g for 20 minutes. Stellate cells were separated into fuzzy bands just above the contact surfaces of the iohexol solution and aqueous buffer. The mixed interface was harvested, and the cells were washed and then in DMEM containing 10% fetal bovine serum (FBS), 4 mM glutamine, and antibiotics (penicillin 100 U / ml, streptomycin 100 mg / ml). Resuspend. After reaching confluency, cells were harvested and replated at the same seed density. All experiments were performed using culture-harvested cells (2-4 generations). PSCs were incubated in serum-deficient media 24 hours prior to the addition of experimental reagents.
<4> 웨스턴 블럿<4> western blot
PSC를 수확하고 나서, 그 용해액을 동일한 완충액 (62.5 mM Tris-HCl, pH 6.8, 2% sodium dodecyl sulfate (SDS), 20% glycerol, and 10% 2-mercaptoethanol)에서 끓였다. 그리고 나서, 세포 용해액내 단백질을 10% SDS-PAGE에 의해 분리하고 니트로셀룰로스 멤브레인에 이전시켰다. 단백질의 이전 이후에, 상기 멤브레인을 실온에서 2시간 동안 PBST(PBS-Tween-20)의 5% 탈지유로 차단하고 나서, 하룻밤 동안 α-SMA와 항체를 반응시켰다. 3번의 세척 이후에, 각 블럿을 1시간 동안 퍼옥시다제-연결된 2차 항체와 반응시키고 제조자의 권고 프로토콜에 따라서 향상된 화학발광 탐지 시스템 (Amersham, Piscataway, NJ)을 사용하여 상기 항체-특이적 단백질을 시각화하였다. After harvesting PSC, the lysate was boiled in the same buffer (62.5 mM Tris-HCl, pH 6.8, 2% sodium dodecyl sulfate (SDS), 20% glycerol, and 10% 2-mercaptoethanol). Then, proteins in cell lysate were separated by 10% SDS-PAGE and transferred to nitrocellulose membrane. After transfer of the protein, the membrane was blocked with 5% skim milk of PBST (PBS-Tween-20) for 2 hours at room temperature, and then reacted with α-SMA and antibody overnight. After 3 washes, each blot is reacted with peroxidase-linked secondary antibody for 1 hour and the antibody-specific protein using an improved chemiluminescence detection system (Amersham, Piscataway, NJ) according to the manufacturer's recommended protocol Was visualized.
<결과 1> ACP 동안 췌장의 형태학 및 조직학적 조사를 통한 NJ의 효과<Result 1> Effect of NJ on Morphological and Histological Examination of Pancreas During ACP
본 발명자들은 ACP에 대한 췌장의 형태학 및 조직학적 특성을 조사하였다. 본 발명에서, 3주 동안의 도전 이후에, 췌장 간질 부종, 염증, 침윤, 및 췌장 섬유증이 입증되었다 (도 1). ACP는 특징적 형태를 상실하였고, 정상적 췌장보다는 부종성 특성을 가져 왔다. 5 또는 10 ㎎/㎖의 NJ 처리는 췌장의 외부 형태학적 부종을 상당히 감소시켰다 (도 1). 그러나, 조직학적 검사에서, NJ 처리는 ACP 도전에 대한 상당한 보호를 나타내지 못했다 (도 1B). ACP에 NJ의 정확한 효과를 추가적으로 조사하기 위하여, 본 발명자들은 아밀라제 방출을 통해 췌장 포상세포의 생존을 측정하였다. ACP 도전된 생쥐는 감소된 아밀라제에 의해 나타낸 바와 같이, 췌장 포상세포의 파괴를 입증하였다. 그러나, NJ 처리는 포상세포 사멸 및 파괴를 상당히 억제하였다 (도 2).We investigated the morphological and histological properties of the pancreas for ACP. In the present invention, after 3 weeks of challenge, pancreatic stromal edema, inflammation, infiltration, and pancreatic fibrosis were demonstrated (FIG. 1). ACP lost its characteristic morphology and had edema characteristics rather than the normal pancreas. NJ treatment at 5 or 10 mg / ml significantly reduced external morphological edema of the pancreas (FIG. 1). However, in histological examination, NJ treatment did not show significant protection against ACP challenge (FIG. 1B). In order to further investigate the exact effect of NJ on ACP, we measured the survival of pancreatic acinar cells via amylase release. ACP challenged mice demonstrated disruption of pancreatic acinar cells, as indicated by reduced amylase. However, NJ treatment significantly inhibited acinar cell death and destruction (FIG. 2).
<결과 2> 섬유증에 NJ의 효과<Result 2> Effect of NJ on Fibrosis
PSC를 자극하고 활성화시켰을 때, α-SMA 발현, 콜라겐 축적, 및 사이토카인 생성이 증가된다 (Masamune, A. 등 (2009) Clin. Gastroenterol. Hepatol. 7, S48-54). 그러므로, ACP 동안 활성화 마커, α-SMA와 췌장의 면역염색에 의한 모델에서, in vivo PSC 활성화를 측정하였다. 그러나, NJ 처리된 ACP 마우스는 췌장에서 감소된 α-SMA 발현을 나타내었다 (도 3A). Acta2, Fn1, Tgfβ 유전자에 대한 mRNA 수준의 실시간 RT-PCR에 의한 측정은 α-SMA, 피브로넥틴, 및 종양성장인자(tumor growth factor ; TGF-β)의 유전자 발현 수준이 ACP-도전된 마우스에서 상당히 증가되었음을 나타내었다. mRNA 발현의 증가는 ACP 동안 NJ 처리에 의해 감소된었다(도 3B). 다음으로, ACP 동안 섬유증에 NJ의 효과를 입증하기 위하여, 본 발명자들은 췌장 콜라겐 축적의 조직학적 평가를 수행하였다. ACP는 췌장에서 콜라겐 발현을 특징적으로 증가시켰다. 그러나, 콜라겐 생성은 NJ 처리된 마우스에서 유의하게 감소되지 않았다 (도 3C).When PSCs are stimulated and activated, α-SMA expression, collagen accumulation, and cytokine production are increased (Masamune, A. et al. (2009) Clin. Gastroenterol. Hepatol. 7, S48-54). Therefore, in vivo PSC activation was measured in a model by activation marker, α-SMA and pancreas immunostaining during ACP. However, NJ treated ACP mice showed reduced α-SMA expression in the pancreas (FIG. 3A). Real-time RT-PCR measurement of mRNA levels for Acta2, Fn1, and Tgfβ genes indicated that gene expression levels of α-SMA, fibronectin, and tumor growth factor (TGF-β) were significantly higher in ACP-challenged mice. Increased. The increase in mRNA expression was reduced by NJ treatment during ACP (FIG. 3B). Next, to demonstrate the effect of NJ on fibrosis during ACP, we performed histological evaluation of pancreatic collagen accumulation. ACP markedly increased collagen expression in the pancreas. However, collagen production was not significantly reduced in NJ treated mice (FIG. 3C).
<결과 3> PSC 활성화에 NJ의 효과<Result 3> Effect of NJ on PSC Activation
ACP에 NJ의 역할을 밝히기 위하여, 본 발명자들은 쥐 췌장으로부터 PSC를 분리하였다. PSCs express intermediate filament proteins such as desmin (13).To elucidate the role of NJ in ACP, we isolated PSC from rat pancreas. PSCs express intermediate filament proteins such as desmin (13).
PSC는 데스민과 같은 중간 섬유 단백질을 발현한다 (Omary, M.B. 등 (2004) N. Engl. J. Med. 351, 2087-2100). 앞서 보고한 바와 같이, 분리된 PSC는 데스민 양성 반응을 가지고, 이것은 분리된 세포가 PSC임을 나타낸다 (도 4A). PSC의 3주 배양시에, 상기 PSC는 활성화되고, α-SMA를 발현한다 (도 4B). 그러나, NJ 처리는 PSC에서 α-SMA의 발현을 억제하였다 (도 4B 및 4C). NJ 처리는 또한 PSC에서 α-SMA, 피브로넥틴, 및 TGF-β와 같은 섬유증-관련 유전자의 mRNA 발현을 억제하였다.PSCs express intermediate fiber proteins such as desmin (Omary, MB et al. (2004) N. Engl. J. Med. 351 , 2087-2100). As reported previously, isolated PSCs had a desmin positive response, indicating that the isolated cells were PSCs (FIG. 4A). Upon 3-week incubation of PSCs, the PSCs are activated and express α-SMA (FIG. 4B). However, NJ treatment inhibited the expression of α-SMA in PSCs (FIGS. 4B and 4C). NJ treatment also inhibited mRNA expression of fibrosis-related genes such as α-SMA, fibronectin, and TGF-β in PSCs.
이상 살펴본 바와 같이, 본 발명은 감송향의 신규한 용도로서, 감송향 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 치료용 조성물을 제공한다. 본 발명의 조성물은 세룰레인에 의해서 유발되는 만성 췌장염을 효과적으로 억제하므로 만성 췌장염의 예방 및 치료를 위하여 효과적으로 사용될 수 있다.As described above, the present invention provides a composition for the prevention and treatment of chronic pancreatitis, which comprises the extract of Persimmon as an active ingredient as a novel use of persimmon. Since the composition of the present invention effectively inhibits chronic pancreatitis caused by cerulein, it can be effectively used for the prevention and treatment of chronic pancreatitis.
이상, 본 발명의 바람직한 실시예를 들어 상세하게 설명하였으나, 본 발명은 상기 실시예에 한정되는 것은 아니며, 본 발명의 기술적 사상의 범위내에서 당 분야에서 통상의 지식을 가진 자에 의하여 여러 가지 변형이 가능하다.As mentioned above, although preferred embodiment of this invention was described in detail, this invention is not limited to the said embodiment, A various deformation | transformation by a person of ordinary skill in the art within the scope of the technical idea of this invention is carried out. This is possible.

Claims (7)

  1. 감송향 (Nardostachys jatamansi ; NJ) 추출물을 활성성분으로 포함하는 만성 췌장염(AP) 예방 및 치료용 약학적 조성물.A pharmaceutical composition for the prevention and treatment of chronic pancreatitis (AP) comprising the extract of Nardostachys jatamansi (NJ) as an active ingredient.
  2. 제 1항에 있어서, 상기 만성 췌장염은 알콜성 만성 췌장염(ACP)인 것을 특징으로 하는 약학적 조성물.The pharmaceutical composition of claim 1, wherein the chronic pancreatitis is alcoholic chronic pancreatitis (ACP).
  3. 제 1항에 있어서, 상기 조성물은 약학적으로 허용가능한 담체를 추가로 포함하는 것을 특징으로 하는 약학적 조성물.The pharmaceutical composition of claim 1, wherein the composition further comprises a pharmaceutically acceptable carrier.
  4. 제 1항에 있어서, 상기 감송향 추출물이 감송향의 뿌리를 물 또는 유기용매로 추출한 것임을 특징으로 하는 약학적 조성물.The pharmaceutical composition according to claim 1, wherein the extract of Persimmon scent is extracted from the root of the Persimmon scent with water or an organic solvent.
  5. 제 4항에 있어서, 상기 감송향 추출물이 감송향의 뿌리에 1 내지 20 배의 물을 가하고 80 내지 150℃에서 1 내지 24 시간 동안 추출한 후 여과하여 얻어진 것임을 특징으로 하는 약학적 조성물.The pharmaceutical composition according to claim 4, wherein the extract of persimmon is obtained by adding 1 to 20 times water to the root of persimmon and extracting at 80 to 150 ° C. for 1 to 24 hours.
  6. 제 4항에 있어서, 상기 유기용매는 에탄올 또는 메탄올인 것을 특징으로 하는 약학적 조성물.The pharmaceutical composition of claim 4, wherein the organic solvent is ethanol or methanol.
  7. 제 1항 내지 제 6항 중 어느 한 항의 감송향 (Nardostachys jatamansi NJ) 추출물을 활성성분으로 포함하는 만성 췌장염 예방 및 개선용 식품 조성물.A food composition for preventing and improving chronic pancreatitis, comprising the extract of Persimmon (Nardostachys jatamansi NJ) according to any one of claims 1 to 6 as an active ingredient.
PCT/KR2013/000357 2013-01-17 2013-01-17 Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient WO2014112663A1 (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
PCT/KR2013/000357 WO2014112663A1 (en) 2013-01-17 2013-01-17 Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
PCT/KR2013/000357 WO2014112663A1 (en) 2013-01-17 2013-01-17 Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient

Publications (1)

Publication Number Publication Date
WO2014112663A1 true WO2014112663A1 (en) 2014-07-24

Family

ID=51209748

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/KR2013/000357 WO2014112663A1 (en) 2013-01-17 2013-01-17 Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient

Country Status (1)

Country Link
WO (1) WO2014112663A1 (en)

Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1994018993A1 (en) * 1993-02-23 1994-09-01 Pharmakon Usa, Inc. Therapeutic herbal composition
US20040265335A1 (en) * 2003-05-19 2004-12-30 Amazon Biotech Inc. Herbal composition and method of treating HIV infection
US20070122495A1 (en) * 2005-11-28 2007-05-31 Sahajanand Biotech Pvt. Ltd. Herbal composition to improve psychological functions as an anxiolytic, tranquilizer, and non-narcotic sedative, as well as other physiological functions
KR20100095838A (en) * 2009-02-23 2010-09-01 주식회사한국전통의학연구소 Composition for preventing and treating acute pancreatitis comprising nardostachys jatamansi extract as an active ingredient
KR20110098500A (en) * 2010-02-26 2011-09-01 전북대학교산학협력단 Composition comprising nardostachys jatamansi extract for preventing and treating insulin dependent diabetes mellitus, food comprising thereof
KR20120111109A (en) * 2011-03-31 2012-10-10 주식회사한국전통의학연구소 Composition for treatment of pancreatic cancer and beauty expenses composition comprising extract of nardostachyos rhizoma

Patent Citations (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1994018993A1 (en) * 1993-02-23 1994-09-01 Pharmakon Usa, Inc. Therapeutic herbal composition
US20040265335A1 (en) * 2003-05-19 2004-12-30 Amazon Biotech Inc. Herbal composition and method of treating HIV infection
US20070122495A1 (en) * 2005-11-28 2007-05-31 Sahajanand Biotech Pvt. Ltd. Herbal composition to improve psychological functions as an anxiolytic, tranquilizer, and non-narcotic sedative, as well as other physiological functions
KR20100095838A (en) * 2009-02-23 2010-09-01 주식회사한국전통의학연구소 Composition for preventing and treating acute pancreatitis comprising nardostachys jatamansi extract as an active ingredient
KR20110098500A (en) * 2010-02-26 2011-09-01 전북대학교산학협력단 Composition comprising nardostachys jatamansi extract for preventing and treating insulin dependent diabetes mellitus, food comprising thereof
KR20120111109A (en) * 2011-03-31 2012-10-10 주식회사한국전통의학연구소 Composition for treatment of pancreatic cancer and beauty expenses composition comprising extract of nardostachyos rhizoma

Similar Documents

Publication Publication Date Title
KR101023780B1 (en) Composition for preventing and treating acute pancreatitis comprising Nardostachys jatamansi extract as an active ingredient
WO2012134169A2 (en) Composition for lung cancer treatment and functional food containing gleditsiae semen extract
KR101624704B1 (en) Pharmaceutical composition and food composition for prevention, treatment or improvement of hair loss or benign prostatic hyperplasia
KR20090094619A (en) Compositions for enhancing immune, or preventing or treating bacterial enteritis or Coccidiosis
KR20120122407A (en) Composition for Treatment of Renal Cell Carcinoma and Beauty Expenses Composition Comprising Extract of Dendrobii Herba
KR101500582B1 (en) Composition for preventing and treating chronic pancreatitis comprising Nardostachys jatamansi extract as an active ingredient
WO2014112663A1 (en) Pharmaceutical composition for preventing and treating chronic pancreatitis which includes extract of nardostachys jatamansi as active ingredient
KR20190136786A (en) Pharmaceutical composition for prevention or treatment of colon cancer
KR20230046054A (en) Composition for the prevention or treatment of obesity comprising Scutellariae Radi and Coptidis Rhizoma extract as an active ingredient
KR101752484B1 (en) Composition for Treatment of Renal Cell Carcinoma and Beauty Expenses Composition Comprising Extract of Pharbitis Semen
WO2012134250A2 (en) Composition for renal cancer treatment and composition for cosmetic containing nardostachyos rhizoma extract
KR20120021283A (en) Composition comprising extract of myristicae semen for treatment of prostate cancer and functional food comprising this
WO2013147340A1 (en) Composition and dietary supplement for treating lung cancer, containing frankincense extract
KR101385658B1 (en) Composition Comprising Extract of Alpiniae Semen for Treatment of Prostate Cancer and Functional Food Comprising This
WO2012134167A2 (en) Composition for lung cancer treatment and functional food containing eriobotriae folium extract
WO2012134251A2 (en) Composition for pancreatic cancer treatment and composition for cosmetics containing nardostachyos rhizoma extract
KR20150118049A (en) Composition for preventing and treating periodontal disease comprising Nardostachys jatamansi extract as an active ingredient
KR20160104824A (en) Pharmaceutical composition for prevention or treatment of Parkinson&#39;s disease comprising herbal extract or fraction thereof
WO2014069692A1 (en) Composition for treating renal cancer and functional health food containing inula britannica var. chinensis extract
KR20240047864A (en) Anti-obesity composition comprising leek and gondree extract as active ingredients and method for manufacturing the same
KR20210157522A (en) Composition for preventing, ameliorating or treating diabetic nepropathy comprising bangpungtongseongsan extract as effective component
WO2015088064A1 (en) Pharmaceutical composition containing desoxo-narchinol a and 8α-hydroxy pinoresinol as active ingredient for preventing and treating sepsis
WO2014069686A1 (en) Composition for treating renal cancer and functional health food containing trichosanthis radix extract
KR20120021281A (en) Composition comprising extract of alpiniae officinari rhizoma for treatment of prostate cancer and functional food comprising this
WO2012134163A2 (en) Composition for lung cancer treatment and composition for cosmetics containing nardostachyos rhizoma extract

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 13871614

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 13871614

Country of ref document: EP

Kind code of ref document: A1