CN103919839A

CN103919839A - Application of Trichosanthes kirilowii, safflower and hemlock parsley for relieving oxidative stress injury to drosophila intestinal tract

Info

Publication number: CN103919839A
Application number: CN201410155605.8A
Authority: CN
Inventors: 金丽华; 周洋
Original assignee: Northeast Forestry University
Current assignee: Northeast Forestry University
Priority date: 2014-04-18
Filing date: 2014-04-18
Publication date: 2014-07-16
Anticipated expiration: 2034-04-18
Also published as: CN103919839B

Abstract

The invention relates to application of Trichosanthes kirilowii, safflower and hemlock parsley extracts for relieving oxidative stress injury to drosophila intestinal tract. The Trichosanthes kirilowii, safflower and hemlock parsley extracts are obtained by a water extraction method, a repairing experiment of the Trichosanthes kirilowii, safflower and hemlock parsley extracts on an oxidative injury model of the drosophila intestinal tract is performed, and the result finds that the Trichosanthes kirilowii, safflower and hemlock parsley extracts have excellent activity on the aspects of resisting oxidation, scavenging free radicals and the like, and can be used to prepare antioxidative medicaments, in particular medicaments resisting oxidative injury to the intestinal tract.

Description

The application in alleviating the damage of fruit bat intestinal oxidative stress of Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong

Technical field

The present invention relates to the application aspect alleviation fruit bat intestinal oxidative damage of Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract, relate to fruit bat natural immunity technical field.

Background technology

Intestinal not only has digestive and absorptive functions, and is the immune organ of human body maximum.When pathogenic microorganism invasion intestinal, intestinal inherent immunity system is being born local defense effect.Intestinal inherent immunity system mainly comprises intestinal mucosa epithelium barrier, inherent immunity cell, inherent immunity molecule etc.Wherein function of intestinal mucosa barrier in patient is a key character of intestinal, when external environment, stress, the factor such as infection can break normal flora in intestinal mucosal barrier and the balance between host while stimulating intestinal, causes the imbalance of intestinal homeostasis.

Fruit bat is as important model organism, and signal transduction pathway and mammal that its intestinal structure, cellularity and stem cell participate in have a lot of similarities.Therefore study fruit bat intestinal immune regulatory mechanism and intestinal stem cell differentiation mechanism and there is important directive function to disclosing the pathogenesis of human intestine's relevant disease.Fruit bat adult intestinal tissue homeostasis depends on the removing of dead cell and the generation of new cell, and the intestinal stem cell (ISC) with cell division capacity plays most important effect aspect the maintaining of tissue homeostasis.An intestinal stem cell carries out asymmetric division and can produce two cells, one of them cell is for keeping the ISC of stem cell characteristic, another is of short duration nondividing enteroblast (enteroblast, EB), enteroblast can continue to be divided into absorptive-type enterocyte (enterocyte, EC) or the endocrine cell of secreting type (enteroendocrine cell, EE).Combined effect by these cells realizes maintaining of intestinal tissue and upgrades.Yet, when stem cell number changes or can cause the exhaustion of organ-system during function reduction, bring out canceration and organism aging process.

Fructus Trichosanthis (Trichosanthes kirilowii Maxim.) has clearing away heat and eliminating phlegm, relieving stuffiness of the chest by dispersing aggregation of pathogens, moisturizes the effects such as laxation, is used for the treatment of the diseases such as cough due to lung-heat, the turbid Huang of expectorant are thick, obstruction of qi in the chest and cardialgia.Main containing compositions such as volatile oil, oils, sterols, Fructus Trichosanthis ester alkali, aminoacid, have antibacterial, anticancer, improve hypoxia-bearing capability, reduce the multiple effects such as serum cholesterol.Flos Carthami (Carthamus tinctorius L.) is traditional medicine, has the effect of promoting blood circulation to restore menstrual flow, eliminating stasis to stop pain, is used for the treatment of circulation disorder.Its effective ingredient is mainly Carthamus yellow Flos Carthami red pigment.Wherein, Carthamus yellow has the hydroxy radical of removing, suppresses lipid peroxidation, and neuroprotective is avoided the effects such as damage.Rhizoma Chuanxiong (Ligusticum chuanxiong Hort.) has the effect of blood circulation promoting and blood stasis dispelling, wind-expelling pain-stopping, for diseases such as treating swelling and pain by traumatic injury, headache, rheumatic arthralgia.Main containing compositions such as volatile oil, alkaloid, phenols, polysaccharide and organic acid, Phthalide lactones, there is the effects such as analgesia, antitumor, antibacterial and external removing free radical.Yet, not yet have pair Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract for alleviating the relevant report of intestinal oxidative damage at present.

Summary of the invention

The object of the invention is to disclose the application in alleviating intestinal oxidative damage of Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract.

Above-mentioned purpose of the present invention is achieved by following scheme:

First give fruit bat feeding oxidative stress thing (Paraquat and H ₂o ₂) set up fruit bat intestinal model of oxidative, Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract are carried out to the experiments such as drosophila survival rate and intestinal stem cell and enteroblast quantitative observation, secondly to fruit bat difference feeding Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract, observe life span of drosophila melanogaster and aging fruit bat intestinal stem cell and enteroblast number change.Found that Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract have good antioxidation, removing free radical isoreactivity, can be used for preparing anti-oxidation medicine, especially treat the medicine of intestinal oxidative damage.

Accompanying drawing explanation

Fig. 1 is that Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract are to Paraquat and H ₂o ₂the impact of fruit bat survival rate after processing;

Fig. 2 is that after Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract are processed Paraquat, enterocyte immunostaining result and the intestinal of fruit bat is dry thin

The impact of born of the same parents and enteroblast quantity;

Fig. 3 is the impact on life span of drosophila melanogaster of Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract;

Fig. 4 be Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract to the enterocyte immunostaining result of the aging fruit bat of 30day and intestinal stem cell and

The impact of enteroblast quantity;

The specific embodiment

Below in conjunction with example, the invention will be further described:

Laboratory animal: wild type W ¹¹¹⁸strain Drosophila melanogaster (Drosophila melanogaster) is preserved for laboratory, and esgGal4UAS-GFP attacks professor Rong Wen by Peking University and provides.Fruit bat cultivation temperature is 25 ℃, relative humidity 60%.

The preparation of fruit bat culture medium: matched group culture medium prescription is Semen Maydis powder 14.2g, agar 1.1g, yeast powder 3.3g, Semen sojae atricolor powder 1.9g, white sugar 17g, propanoic acid 0.94mL, deionized water 200mL.In experimental group, add and the isopyknic Fructus Trichosanthis of matched group deionized water, Flos Carthami and Rhizoma Chuanxiong extracting solution, in experimental group, the concentration of Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract is 10%.

Each experiment reagent that the present embodiment relates to is commercially available analytical pure: Paraquat and H ₂o ₂deng.

The preparation of embodiment 1 Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extracting solution

Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong respectively take 20g, add 400mL deionized water to soak 12h, moderate heat boils 3h, filter, filtering residue adds 200mL deionized water to use the same method to extract 1 time again, merge 2 filtrates and be concentrated into 100mL, obtaining mass concentration is the Chinese herbal medicine water extraction liquid of 0.2g/mL (20%), puts-40 ℃ and saves backup.

Embodiment 2 Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract are processed the impact of rear fruit bat intestinal stem cell and enteroblast quantity on Paraquat

With the fruit bat cultivated in ordinary culture medium as a control group, the fruit bat of cultivating in Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong culture medium is as experimental group.Random collecting matched group and the experimental group fruit bat of 3～5d that sprouted wings respectively, female male drosophila is each 15 in every group.To transferring in the fruit bat pipe that is placed with 5 metafiltration paper after the hungry processing of each group fruit bat 2h, in filter paper, add 6mM paraquat or 1%H ₂o ₂solution (containing 5% sucrose), makes filter paper fully moistening, and every 24h changes a filter paper and records fruit bat death toll.

Result shows: paraquat processes after fruit bat 6d, matched group survival rate drops to 1.1%, and the experimental group survival rate of feeding Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong is respectively 75.5% (* * * P<0.001), 95.6% (* * * P<0.001) and 67.7% (* * P<0.01), compares with matched group and improved respectively 74.4%, 94.5% and 66.6% (Figure 1A).In addition, H ₂o ₂process after fruit bat 6d, the experimental group survival rate of feeding Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong is respectively 41.1% (* * P<0.01), 50% (* * P<0.01) and 54.4% (* * * P<0.001), is matched group (16.7%) 2.4,2.99 and 3.2 times (Figure 1B).The above results explanation, Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract can be alleviated paraquat and H ₂o ₂the oxidative stress damage of induction, has improved the survival rate of fruit bat significantly.

Embodiment 3 Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract are processed the rear enterocyte immunostaining result of fruit bat and the impact of intestinal stem cell cell and enteroblast quantity to Paraquat

The female fruit bat feeding of random collecting matched group and experimental group emergence 4-5d is containing paraquat (10mM) the solution 16h of 5% sucrose, under microscope, extract 10-12 bar fruit bat intestinal in paraformaldehyde, room temperature is 30min fixedly, the DAPI 10min that dyes, fluorescence microscopy Microscopic observation taking pictures after glycerol mounting.

Result shows: GFP positive cell (precursor in the experimental group of feeding Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong, comprising intestinal stem cell and enteroblast) quantity is significantly lower than matched group, and compare with matched group and reduced respectively by 47.1%, 38.6%, 31.3% (* * * P<0.001).The results are shown in Figure 2.

Embodiment 4 Fructus Trichosanthis, Flos Carthami and the impact of Rhizoma Chuanxiong extract on life span of drosophila melanogaster

The fruit bat of random collecting matched group and experimental group emergence 4d.Male and female are separated, 30 every group.Every 2d changes a subculture, and regularly add up fruit bat death toll every day, until all dead.

Result shows: compare with matched group, Fructus Trichosanthis extract can extend female, the male drosophila life-span; Flos Carthami or Rhizoma Chuanxiong extract only extend female life span of drosophila melanogaster and not remarkable on male drosophila life-span impact, the results are shown in Figure 3.

The impact on the enterocyte immunostaining result of the aging fruit bat of 30day and intestinal stem cell and enteroblast quantity of embodiment 5 Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract

The female fruit bat of random collecting matched group and experimental group emergence 4d is cultivated after 30d, extracts 10-12 bar fruit bat intestinal under microscope in paraformaldehyde, and room temperature is 30min fixedly, the DAPI 10min that dyes, fluorescence microscopy Microscopic observation taking pictures after glycerol mounting.

Result shows: GFP positive cell (precursor in the experimental group of feeding Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong, comprising intestinal stem cell and enteroblast) quantity is significantly lower than matched group, and compare with matched group and reduced respectively by 30.2%, 28.6%, 41.9% (* * * P<0.001).The results are shown in Figure 4.

In sum, by the analysis to the life-span of the survival rate of fruit bat, fruit bat, intestinal stem cell and enteroblast number change, we think that Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract can alleviate fruit bat intestinal oxidative damage.

Claims

1. Fructus Trichosanthis, Flos Carthami and the Rhizoma Chuanxiong extract application in alleviating fruit bat intestinal oxidative damage, is characterized in that, described Chinese herbal medicine extract is water extract.

2. application according to claim 1, is characterized in that, gives the fruit bat feeding paraquat(6mM of emergence 3-5d) and H ₂o ₂(1%), Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract can improve drosophila survival rate.

3. application according to claim 1, is characterized in that, gives the female fruit bat feeding paraquat(10mM of emergence 4-5d) solution 16h, Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract can reduce intestinal stem cell and enteroblast quantity.

4. application according to claim 1, is characterized in that, distinguishes feeding Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract to female, the male drosophila of emergence 4d, and Fructus Trichosanthis extract can extend female, the male drosophila life-span; Flos Carthami and Rhizoma Chuanxiong extract only extend female life span of drosophila melanogaster.

5. application according to claim 1, is characterized in that, the female fruit bat of emergence 4d is cultivated after 30d, and Fructus Trichosanthis, Flos Carthami and Rhizoma Chuanxiong extract can reduce old and feeble fruit bat intestinal stem cell and enteroblast quantity.