CN115400160A - Application of saponin endosperm extract in preparing medicine for treating ulcerative colitis - Google Patents
Application of saponin endosperm extract in preparing medicine for treating ulcerative colitis Download PDFInfo
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Abstract
The invention provides an application of saponin endosperm extract in preparing and treating ulcerative colitis, wherein the saponin endosperm extract is prepared by (1) pulverizing saponin endosperm, sieving, slowly adding into boiling water while stirring, mixing uniformly, decocting with slow fire, and concentrating to a certain concentration to obtain saponin endosperm medicinal liquid. (2) Pulverizing endosperm of fructus Gleditsiae Abnormalis, sieving, adding into water, and heating to obtain product A; pulverizing semen Gleditsiae Abnormalis endosperm, sieving, adding the above A, stirring, mixing, decocting with slow fire, and concentrating to obtain semen Gleditsiae endosperm medicinal liquid. The obtained fructus Gleditsiae Abnormalis endosperm extract has effects of moistening dryness, relaxing bowels, dispelling pathogenic wind, relieving swelling, invigorating spleen, invigorating qi, and eliminating dampness, and can be used for treating colitis ulcerosa. It also has certain effect in improving injury of lung, liver and kidney.
Description
Technical Field
The invention relates to a new application of saponin endosperm extract, in particular to an application of saponin endosperm extract in preparing and treating ulcerative colitis.
Background
The endosperm of Gleditsia sinensis is Gleditsia sinensis (Gleditsia sinensisLam.) or Yunnan Gleditsia pod (Gleditsia japonica var. delavayi(franch.) l.c.li) seed endosperm is prepared by boiling the seeds in water, peeling off the seed coat, collecting the endosperm, and drying; or taking fresh seeds, peeling off seed coats, taking endosperm of the seeds, and drying to obtain the product. Belongs to a high-energy, high-carbohydrate, low-protein and low-fat food. And contains vegetable dietary fiber. The record of the book of materia medica book of drawings: white meat in the stone is used as lung-treating drug. The "white meat in kernel" is the endosperm of the seed of Gleditsia sinensis Lam. At present, the chemical components and the pharmacological activity of the saponin endosperm are rarely researched, and particularly, no relevant data report is found in research on ulcerative colitis.
Ulcerative Colitis (UC) is a chronic nonspecific inflammatory disease of the colon and rectum whose etiology is not yet well understood, and the lesions are confined to the large intestinal mucosa and submucosa. Lesions are localized in the sigmoid colon and rectum and may extend to the descending colon, even the entire colon. The disease course is long and the attack is often repeated. The disease is seen at any age, but is most seen in 20-30 years old. The cause of ulcerative colitis is still unknown to date. Genetic factors may have a certain position. Psychological factors play an important role in the progression of the disease, with the primary presence of pathophysiological states such as depression or social distance being clearly improved after colectomy. Ulcerative colitis is thought to be an autoimmune disease.
The pathogenesis of inflammatory bowel disease is currently believed to be the result of interactions between foreign substances causing host responses, genes, and immune influences. The existing treatments are generally divided into drug treatment and surgical treatment. The drug therapy mainly comprises western medicines, the preparation of sulfasalazine salicylic acid is a main therapeutic medicine, such as adisa, mesalazine and the like, and the common corticosteroid is prednisone or dexamethasone, but the long-term hormone maintenance is not considered to prevent relapse at present. Hydrocortisone or dexamethasone can be used for intravenous drip in the acute attack stage, but whether hormone should be continuously used in the chronic stage is different, and long-term use is not advocated due to certain side effects of the hormone. The traditional Chinese medicine for treating diarrhea type ulcerative colitis can be used for treating diarrhea type ulcerative colitis, and has the advantages of ideal effect, small side effect and high safety.
Disclosure of Invention
In order to solve the technical problems, the invention provides application of saponin endosperm extract in preparing and treating ulcerative colitis, and solves the problem of how to extract saponin endosperm extract to treat ulcerative colitis. In order to realize the purpose, the invention is realized by the following technical scheme:
use of semen Gleditsiae extract in preparing medicine for treating colitis ulcerosa is provided.
The application of the saponin endosperm extract in preparing the medicine for treating ulcerative colitis comprises the following steps: pulverizing semen Gleditsiae endosperm, sieving, slowly adding into boiling water while stirring, mixing, decocting with slow fire, and concentrating to obtain fluid extract or soft extract.
Specifically, the application of the saponin endosperm extract in preparing the medicine for treating ulcerative colitis comprises the following steps: pulverizing semen Gleditsiae, sieving with 8-500 mesh sieve, slowly adding 2-12 times of boiling water while stirring, mixing, decocting with slow fire for 10-60min, and concentrating to obtain fluid extract or soft extract.
The application of the saponin endosperm extract in preparing the medicine for treating ulcerative colitis comprises the following steps:
a) Pulverizing endosperm of fructus Gleditsiae Abnormalis, sieving, adding into water, and heating to obtain product A;
b) Pulverizing semen Gleditsiae endosperm, sieving, adding above A, stirring, mixing, decocting with slow fire, and concentrating to obtain fluid extract or soft extract.
5. Use of an endosperm extract of saponin as claimed in claim 4 in the manufacture of a medicament for the treatment of ulcerative colitis, wherein: the preparation method of the saponin endosperm extract comprises the following steps:
a) Pulverizing semen Gleditsiae, sieving with 8-500 mesh sieve, adding 3-12 times of water, and heating for 2-10 min to obtain product A;
b) Pulverizing semen Gleditsiae Abnormalis endosperm, sieving with 8-500 mesh sieve, adding into the above A, stirring, mixing, decocting with slow fire for 10-60min, and concentrating to obtain fluid extract or soft extract.
Use of endosperm of fructus Gleditsiae Abnormalis in preparing medicine for treating colitis ulcerosa is provided.
A medicine for treating cerebral ulcerative colitis comprises saponin endosperm extract or saponin endosperm.
A medicine for treating cerebral ulcerative colitis contains extract of semen Gleditsiae (fructus Gleditsiae Abnormalis) endosperm or semen Gleditsiae (fructus Gleditsiae Abnormalis) endosperm as main active ingredient.
The saponin endosperm is used for treating ulcerative colitis, can inhibit the excessive activation of a TLR 4/NF-kB/NLRP 3 signal channel, down-regulates the expression of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 proteins and genes, reduces the release of IL-1 beta, IL-6, IL-17 and TNF-alpha proinflammatory factors, and plays a role in treating ulcerative colitis.
The invention has the beneficial effects that:
the saponin endosperm can improve the symptoms of weight loss, loose stool, hematochezia and the like of patients with ulcerative colitis, protect and repair colon pathological tissue injury of patients with ulcerative colitis, inhibit the release of related inflammatory factors, and play a role in resisting inflammation, thereby inhibiting the occurrence and development of ulcerative colitis. It also has effect in improving lung injury.
Drawings
FIG. 1 changes in body weight of mice;
figure 2 mouse DAI index (compared to normal control group: △△ to representP<0.01; comparison with model group: ** representP<0.01,. IndicatesP<0.05; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low-dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group);
FIG. 3 Colon map of mice;
fig. 4 colon length statistical graph (compared to normal control: △△ representP<0.01; comparison with model groups: ** to representP<0.01 denotesP<0.05; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group);
fig. 5 mouse spleen index change (compared to normal control group: △ to representP<0.05; comparison with model groups: ** to representP<0.01, * To representP<0.05; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group);
figure 6 effect of colon tissue score in mice (compared to normal control group: △△ to representP<0.01; andmodel group comparison: ** to representP<0.01, * To representP<0.05; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group);
FIG. 7 pathological section of colon tissue of mouse (x 100) (C normal group; M model group; Y positive drug mesalazine group; D saponin endosperm low dose group; Z saponin endosperm high dose group; G saponin endosperm high dose group);
fig. 8 effect of saponin endosperm on colonic tissue inflammatory factor content in mice (compared to normal control: △△ representP<0.01; comparison with model group: ** to representP<0.01, * To representP<0.05; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group);
FIG. 9 is a striped graph of colon TLR4, NF-kB-p 65, NLRP3 and Caspase-1 protein and endoglin of each group of mice (C normal group; M model group; Y positive drug mesalazine group; D saponin endosperm low dose group; Z saponin endosperm dose group; G saponin endosperm high dose group);
FIG. 10 histogram of TLR4, NF-. Kappa.B-p 65, NLRP3 and Caspase-1 protein grey value expression in colon tissue of mice (compared to normal controls: △△ to representP<0.01; comparison with model group: ** to representP<0.01; c, normal group; m model group; mesalazine group of Y positive drugs; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group).
FIG. 11 histogram of TLR4, NF-. Kappa.B-p 65, NLRP3 and Caspase-1 mRNA expression in colon tissue of mice (compared to normal controls: △△ to representP<0.01; comparison with model groups: ** to representP<0.01; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g saponin endosperm high dose group);
FIG. 12 pathological section of mouse lung tissue (x 100) (C Normal group; M model group; Y Positive drug mesalazine group; D Saponin endosperm Low dose group; Z Saponin endosperm high dose group; G Saponin endosperm high dose group)
Fig. 13 histogram of ALT, AST, CREA, UREA content in mouse serum (compared to normal control group: △△ to representP<0.01, △ RepresentP<0.05; comparison with model groups: ** representP<0.01, * To representP<0.05; c, normal group; m model group; a group of Y positive drugs mesalazine; d, a saponin endosperm low dose group; z gleditsia sinensis endosperm dosage group; g high-dose endosperm Gleditsia sinensis group)
In order to make the technical solutions of the present invention better understood and enable those skilled in the art to practice the present invention, the following embodiments are further described, but the present invention is not limited to the following embodiments.
Detailed Description
Example 1:
the process comprises the following steps:
pulverizing semen Gleditsiae (fructus Gleditsiae Abnormalis endosperm 100 g), sieving with 8-500 mesh sieve, slowly adding 2-12 times of boiling water while stirring, mixing, decocting with slow fire for 5-60min, and concentrating to a certain concentration to obtain semen Gleditsiae endosperm medicinal liquid.
Mixing 100g of semen Gleditsiae endosperm medicinal liquid with 80% pregelatinized starch, granulating, sieving, and making into capsule 1000.
The efficacy is as follows: moistening dryness, relaxing bowels, dispelling pathogenic wind, relieving swelling, invigorating spleen, invigorating qi, and eliminating dampness.
The using method comprises the following steps: is administered orally.
The usage and dosage are as follows: it is administered orally 2 granules at a time, 3 times daily.
Example 2:
the process comprises the following steps:
a) Pulverizing semen Gleditsiae endosperm 50g, sieving with 8-500 mesh sieve, adding 3-12 times of water, and heating for 2-10 min to obtain product A;
b) Pulverizing semen Gleditsiae endosperm 50g, sieving with 8-500 mesh sieve, adding into the above A product while stirring, mixing, decocting with slow fire for 5-60min, and concentrating to obtain semen Gleditsiae endosperm medicinal liquid.
Mixing 100g of semen Gleditsiae endosperm medicinal liquid with 80% pregelatinized starch, granulating, sieving, and making into capsule 1000.
The efficacy is as follows: moistening dryness, relaxing bowels, dispelling pathogenic wind, relieving swelling, invigorating spleen, invigorating qi, and eliminating dampness.
The using method comprises the following steps: is administered orally.
The usage and dosage are as follows: it is administered orally 2 granules at a time, 3 times daily.
The inventors carried out a number of experiments and the following were studies of the extraction method of the present invention:
1. reagent preparation and Experimental grouping
1. Preparation of medicinal solutions
1.1 preparation of the saponin endosperm liquor: pulverizing semen Gleditsiae, sieving (8-500 mesh) to obtain semen Gleditsiae endosperm powder, slowly adding 2-12 times of boiling water while stirring, decocting with slow fire for 5-60min, and concentrating to obtain semen Gleditsiae endosperm medicinal liquid.
1.2 preparation of the second liquid medicine of the saponin endosperm: heating water 3-12 times of semen Gleditsiae endosperm powder for 2-10 min, mixing with semen Gleditsiae endosperm powder under stirring, decocting with slow fire for 5-60min, and concentrating to obtain semen Gleditsiae endosperm medicinal liquid.
The rest of the reagents
Mesalamine (Losan Pharma GmbH); dextran sulfate sodium (MP corporation), 2.5% solution was prepared before use.
Experimental animal
SPF-grade kunming mice, 4 weeks old, 18 to 22 g, male, sbefu (beijing) biotechnology limited provide [ license number: SCXK (Jing) 2019-0010] which is bred in a pharmacological laboratory of the university of traditional Chinese medicine in Guizhou in cages.
Molding method
Dextran Sodium Sulfate (DSS) powder is prepared into 2.5% DSS solution by using Wahaha purified water, and the DSS solution is provided for a mouse model group, a positive medicament group and a saponin endosperm high-medium low-dose group to drink daily and freely, wherein the freshly prepared DSS solution is replaced every two days, and molding lasts for 8 days.
5. Experiment grouping
SPF grade male Kunming mice were 72 randomly divided into 6 groups of 12 mice each. The test results were respectively blank group, model group, positive drug mesalazine control group (administration dose of 0.52 g/kg), saponin endosperm low dose group (administration dose of 0.3379 g/kg), saponin endosperm middle dose group (administration dose of 0.6758 g/kg) and saponin endosperm high dose group (administration dose of 1.3515 g/kg). The gavage amount is 0.2ml/10g, and the blank group is administrated with distilled water in equal amount twice daily for 10 days.
After the last administration, the eyeballs of the mice are picked up to collect blood, the blood is collected and centrifuged at 12000 r at 4 ℃ for 10 min, and the serum is separated and stored in a refrigerator at-80 ℃. Rapidly dissecting the abdomen along the midline of the abdomen, cutting out a colon specimen (from the anus to the cecum), taking out and placing on ice, immediately measuring and recording the length of the colon specimen with a graduated scale, then flushing the intestinal cavity with physiological saline for 3 times, sucking dry with filter paper, breaking the colon along the longitudinal axis, observing and recording histological changes; then taking part of colon, and placing the part of colon in 4% paraformaldehyde for fixation for HE detection; the remaining colon was stored in a-80 ℃ freezer for future use.
2. Medicinal effect of saponin endosperm for preventing and treating ulcerative colitis
1. Effect of the endosperm of Gleditsia sinensis on the disease Activity index in mice
After administration of DSS solutions, the general condition of the mice was recorded daily and scored for Disease Activity Index (DAI). DAI = (weight loss score + stool trait score + stool blood score)/3. The specific scoring criteria are shown in table 1, and the results are shown in fig. 1 and 2.
TABLE 1 DAI index score criteria
Weight loss (%) | Stool character | Occult blood in | Scoring | |
0 | Is normal and normal | Is normal and normal | 0 | |
1-5 | Shaped soft stool | Macroscopic bloody stool (+) | 1 | |
5-10 | Semi-thin stool | ++ | 2 | |
10-20 | Thin stool | +++ | 3 | |
>20 | Severe loose stool | >+++ | 4 |
The DAI index of the model group mice is extremely remarkably increased compared with that of the normal group (P< 0.01). Compared with the model group, the positive drug mesalazine can reduce the DAI index of UC mice, but has no significant difference (PGreater than 0.05), the low dose of the saponin endosperm can remarkably reduce the DAI index of UC mice (A) (thePLess than 0.01), the saponin endosperm and high dose can obviously reduce the DAI index of UC mice (P<0.05)。
Effect of the endosperm of Gleditsia on Colon Length in mice
After the last administration, the mouse eyes were removed and blood was taken, the abdomen was rapidly dissected along the midline of the abdomen, a colon specimen was taken (from the anus up to the cecum), and the colon specimen was taken out and placed on ice, and its length was immediately measured and recorded with a graduated scale, with the results shown in table 2 and fig. 3 and 4.
Table 2 mouse colon length (' X ± s, n = 8)
Group of | Colon Length (cm) |
Normal group | 10.338±0.628 |
Model set | 8.250±0.975 △△ |
Positive group | 9.438±1.074 |
Low dose group | 9.963±0.890 * |
Middle dose group | 9.563±0.883 |
High dose group | 10.238±0.571 ** |
Note: comparison with normal control group: △△ representP<0.01; comparison with model groups: ** to representP<0.01; * RepresentP<0.05。
As shown in Table 3 and FIGS. 3 and 4, the colon length was extremely significantly shortened in the model group mice as compared with the normal group (P< 0.01). Compared with the model group, the low-dose saponin endosperm significantly inhibits the colon shortening of UC mice (P< 0.05), the endosperm of Chinese honeylocust fruit can remarkably inhibit the colon shortening of UC mice at high dose (P< 0.01), the dose and the positive drug mesalazine in the endosperm of the saponin can inhibit the colon shortening of UC mice (without significant difference), (2)P>0.05)。
Effect of the endosperm of Gleditsia sinensis on the spleen index of mice
After the mice were dissected, spleens were taken and weighed, and mouse spleen indices were calculated, and the results are shown in table 3 and fig. 5.
TABLE 3 mouse spleen index (` X. + -. S, n = 8)
Group of | Spleen index |
Normal group | 3.698±0.746 |
Model set | 5.743±1.112 △ |
Positive group | 5.238±1.900 |
Low dose group | 3.571±1.129 * |
Middle dose group | 3.510±0.614 ** |
High dose group | 3.035±0.879 ** |
Note: comparison with normal control group: △ to represent P<0.05; comparison with model groups: ** to representP<0.01; * To representP<0.05。
Spleen index was significantly increased in the model group mice compared with that in the normal group (P< 0.05). Compared with the model group, the spleen index of UC mice is greatly reduced by the middle dose and the high dose of the saponin endosperm: (P< 0.01), low-dose of saponin endosperm significantly reduces UC mouse spleen index: (P< 0.05), the positive drug mesalazine can also reduce the spleen index of UC mice, but has no significant difference (P>0.05)。
Effect of the endosperm of Gleditsia on Colon histopathological changes in mice
HE staining was performed on the colon tissue of mice and histopathological scoring was performed, and the results are shown in table 4 and fig. 6, and pathological sections of colon tissue of mice are shown in fig. 7.
TABLE 4 mouse Colon TDI index (` X. + -. S, n = 8)
Group of | TDI |
Normal group | 0.00±0.000 |
Model set | 1.266±0.493 △△ |
Positive group | 0.600±0.830 * |
Low dose group | 0.332±0.408 ** |
Middle dose group | 0.668±0.625 |
High dose group | 0.000±0.000 ** |
Note: comparison with normal control group: △△ representP<0.01; comparison with model groups: ** to representP<0.01; * To representP<0.05。
The colon tissue structure of the normal group of mice is clear and complete, the mucosal surface is coated with a single layer of columnar epithelium, the shape of columnar epithelial cells is normal, the arrangement of large intestinal glands in the inherent layer is dense, the number of goblet cells is normal, the muscle fiber structure of the muscle layer is complete and clear, no obvious pathological change is seen in other tissues, and the histological score is low; the structural integrity of colon tissue is seriously damaged in a model group, epithelial cells are necrosed, nuclei are dissolved, lamina propria cells are necrosed, cytoplasm vacuole is formed, nuclei are shrinked and disintegrated, muscle fibers of muscle layers are necrosed, inflammatory cell infiltration is seen in necrotic areas, single round and deep-stained lymphocytes and rod-shaped nuclear neutrophils are taken as main components, fibrous tissue hyperplasia is also seen, fibroblasts with oblong nuclei and fibrous cell hyperplasia with long spindle-shaped nuclei are included, and the histological score is remarkably increased compared with that of a normal group (the tissue is shown in the specification and the patent application shows that the colon tissue is not damaged by the tissueP<0.01). Compared with model group, the low dose and high dose of saponin endosperm can reduce the inflammatory cell infiltration degree of the UC mouse colon tissue, recover the arrangement of goblet cells and glands, and have lower histological score (P< 0.01); the positive medicine can improve the necrosis of epithelial cells and lamina propria cells of the colon tissue of UC mice and relieve inflammationDegree of sexual cell infiltration, low histological score (P< 0.05); the dosage in the endosperm of the Chinese honeylocust improves the infiltration degree of inflammatory cells, but the histological score is lower and has no significant difference (P>0.05)。
Effect of the endosperm of Gleditsia sinensis on the expression of inflammatory factors IL-1 beta, IL-6, IL-17 and TNF-alpha in colon tissue of mice
The contents of inflammatory factors IL-1 beta, IL-6, IL-17 and TNF-alpha in colon tissues of mice were detected by the method required by the kit, and the results are shown in Table 5 and FIG. 8.
TABLE 5 groups of mice colon IL-1 β, IL-6, IL-17, TNF- α (` X. + -. S, n = 8)
Group of | IL-1β(pg/mL) | IL-6(pg/mL) | IL-17(pg/mL) | TNF-α(pg/mL) |
Normal group | 8.298±0.610 | 29.910±1.386 | 8.895±0.624 | 68.485±7.373 |
Model set | 12.507±0.677 △△ | 38.873±5.018 △△ | 13.501±1.360 △△ | 83.720±3.650 △△ |
Positive group | 10.932±0.747 ** | 28.838±2.560 ** | 10.495±0.797 ** | 73.495±4.644 ** |
Low dose group | 9.810±0.871 ** | 31.979±5.076 ** | 11.015±1.497 ** | 75.223±5.314 * |
Medium dose group | 10.802±1.422 ** | 31.517±3.902 ** | 11.385±1.873 ** | 75.701±10.292 * |
High dose group | 9.977±0.660 ** | 31.396±5.328 ** | 11.802±0.624 ** | 73.771±8.185 ** |
Note: comparison with normal control group: △△ representP<0.01; comparison with model group: ** representP<0.01。
Compared with the normal group, the colon tissue of the model group mice has extremely obviously increased contents of IL-1 beta, IL-6, IL-17 and TNF-alpha (P< 0.01). Compared with the model group, the low-dose, the medium-dose, the high-dose and the positive drug mesalazine of the saponin endosperm can remarkably reduce the contents of IL-1 beta, IL-6 and IL-17 in the colon tissue of a UC mouse (see the description in the specification)PLess than 0.01), the high-dose saponin endosperm and the positive drug mesalazine can remarkably reduce the content of TNF-alpha in the colon tissue of a UC mouse (see the description below)PLess than 0.01), the low dose and the medium dose of the saponin endosperm can obviously reduce the content of TNF-alpha in the colon tissue of UC miceP<0.05)。
Discussing the mechanism of action of saponin in preventing and treating ulcerative colitis based on TLR 4/NF-kB/NLRP 3 signal channel
(1) Effect of Gleditsia sinensis endosperm on expression of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 proteins in colon tissue of mice
Western blotting was performed on mouse colon tissues, and the results are shown in Table 6, FIGS. 9 and 10.
TABLE 6 Gray-value expression of colon TLR4, NF-. Kappa.B-p 65, NLRP3 and Caspase-1 proteins in each group of mice (` X. + -. S, n = 5)
Group of | TLR4 | NF-κB-p65 | NLRP3 | Caspase-1 |
Normal group | 0.485±0.131 | 0.392±0.152 | 0.160±0.085 | 0.141±0.041 |
Model set | 0.976±0.299 △△ | 0.946±0.621 △△ | 0.902±0.211 △△ | 0.727±0.395 △△ |
Positive group | 0.332±0.200 ** | 0.262±0.180 ** | 0.327±0.279 ** | 0.174±0.092 ** |
Low dose group | 0.146±0.084 ** | 0.093±0.012 ** | 0.375±0.301 ** | 0.224±0.043 ** |
Middle dose group | 0.282±0.176 ** | 0.141±0.118 ** | 0.388±0.218 ** | 0.376±0.206 ** |
High dose group | 0.192±0.074 ** | 0.158±0.130 ** | 0.327±0.330 ** | 0.334±0.125 ** |
Note: comparison with normal control group: △△ to representP<0.01; comparison with model group: ** representP<0.01。
Compared with the normal group, the expression of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 protein in colon tissues of mice in the model group is greatly increased (P< 0.01). Compared with the model group, the saponin endosperm low dose group, the saponin endosperm medium dose group, the saponin endosperm high dose group and the positive drug mesalazine can remarkably reduce the expression of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 proteins in the colon tissues of UC mice ((P<0.01)。
(2) Effect of Gleditsia sinensis endosperm on expression of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 mRNA in colon tissue of mice
The results of RT-PCR detection of mouse colon tissue are shown in Table 7 and FIG. 11.
TABLE 7 mouse Colon TLR4, NF-. Kappa.B-p 65, NLRP3, and Caspase-1 mRNA expression (` X. + -. S, n = 5)
Group of | TLR4 | NF-κB-p65 | NLRP3 | Caspase-1 |
Normal group | 0.983±0.255 | 1.108±0.220 | 1.164±0.693 | 1.118±0.207 |
Model set | 2.918±0.651 △△ | 3.609±0.562 △△ | 17.197±7.938 △△ | 4.307±0.686 △△ |
Positive group | 1.391±0.419 ** | 1.321±0.235 ** | 3.394±2.824 ** | 1.153±0.107 ** |
Low dose group | 2.155±0.212 ** | 2.878±0.141 ** | 9.691±3.738 ** | 3.352±0.717 ** |
Middle dose group | 1.674±0.264 ** | 2.186±0.061 ** | 6.442±3.297 ** | 2.714±0.646 ** |
High dose group | 1.485±0.147 ** | 1.688±0.281 ** | 5.811±3.134 ** | 1.597±0.345 ** |
Note: comparison with normal control group: △△ to representP<0.01; comparison with model groups: ** representP<0.01。
Compared with the normal group, the expression of the mRNA of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 in colon tissues of the mice in the model group is greatly increased (P< 0.01). Compared with a model group, the saponin endosperm, the low dose, the medium dose, the high dose and the positive drug mesalazine greatly reduce the expression of TLR4, NF-kB-p 65, NLRP3 and Caspase-1 mRNA in the colon tissue of UC mice ((P<0.01)。
And conclusion:
the saponin endosperm can reduce the Disease Activity Index (DAI) of a DSS-induced UC mouse, inhibit the shortening of colon length, reduce spleen index and intestinal mucosa injury index, restore the structural integrity of colon tissues, improve the necrosis condition of epithelial cells, lamina propria cells and muscularis fibers, reduce inflammatory cell infiltration in necrotic areas and inhibit the proliferation of fibrous tissues, and shows that the saponin endosperm has the function of resisting ulcerative colitis, and the action mechanism of the saponin endosperm is related to the inhibition of the release of IL-1 beta, IL-6, IL-17 and TNF-alpha inflammatory factors and the inhibition of the injury of colon inflammation by inhibiting the expression of main target molecules TLR4, NF-kappa B-P65, NLRP3 and Caspase-1 proteins and genes on a TLR 4/NF-kappa B/NLRP3 signal channel.
3. Effect of Cassia nomame endosperm on histopathological changes of lung of mice
After the mouse was dissected, lung tissue was taken and observed by HE staining, and the results are shown in fig. 12.
The lung tissue pleura structure of the normal group of mice is normal, the wall is thin, no connective tissue is proliferated and thickened, all levels of bronchial structures are complete and clear, the epithelial cell morphology is normal, the alveolar epithelial cell morphology of the respiratory part of the lung is normal, and other obvious pathological changes are not seen. The bronchial structures of all levels of lung tissues are normal, the bronchial ciliated epithelium is arranged regularly, no obvious cell shedding is seen, the alveolar epithelial cells are normal, slight or mild bleeding in the alveolar cavity is caused, no obvious fibroplasia and inflammatory cell infiltration are seen in the pulmonary interstitium, and no obvious pathological changes are seen in other tissues. Compared with a model group, the low-dose saponin endosperm, the medium-dose saponin endosperm, the high-dose saponin endosperm and the positive drug mesalazine can improve the phenomenon of alveolar intracavity bleeding of UC mice.
And (4) conclusion:
the saponin endosperm can improve the alveolar hemorrhage in the lung tissue of the UC mouse, and shows that the saponin endosperm also has a prevention and treatment effect on lung injury.
4. Effect of Gleditsia sinensis endosperm on mouse liver function index
After the last administration, the eyeballs of the mice were removed and blood was collected, and blood serum was collected after blood centrifugation for biochemical blood test, and the results are shown in table 8 and fig. 13.
TABLE 8 serum ALT, AST, CREA, UREA conditions of mice (` X. + -. S, n = 5)
Group of | ALT(U/L) | AST(U/L) | UREA(mmol/L) | CREA(μmol/L) |
Normal group | 29.020±3.817 | 132.780±13.942 | 7.518±1.113 | 28.800±8.566 |
Model set | 37.580±3.544 △ | 166.500±18.120 △△ | 9.596±0.786 △△ | 33.840±12.073 |
Positive group | 37.860±4.245 | 137.300±4.287 * | 10.818±1.195 | 30.660±10.078 |
Low dose group | 31.260±7.270 | 131.860±11.160 ** | 9.206±0.476 | 29.260±11.186 |
Middle dose group | 32.620±5.282 | 130.740±28.721 ** | 8.216±1.565 | 28.200±5.335 |
High dose group | 32.100±5.250 | 140.040±21.685 * | 8.022±1.391 * | 26.880±5.858 |
Note: comparison with normal control group: △△ to representP<0.01, △ RepresentP<0.05; comparison with model groups: ** to representP<0.01; * RepresentP<0.05。
ALT significant elevation in serum of model group mice compared with normal group: (P< 0.05). Compared with a model group, the saponin endosperm can reduce the ALT content in the blood serum of UC mice at each dose, but has no significant difference (theP> 0.05); AST is extremely significantly elevated in model group mice (PLess than 0.01), the saponin endosperm low-dose group and the saponin endosperm high-dose group can remarkably reduce the AST content in the blood serum of UC mice, and the positive drug mesalazine and the saponin endosperm high-dose group can remarkably reduce the AST content in the blood serum of UC mice.
The serum of mice in the model group has extremely significant UREA increase compared with the normal group (P< 0.01). Compared with a model group, the high dose of the saponin endosperm can obviously reduce the content of the UREA in the blood serum of the UC mouse, and the low dose and the medium dose of the saponin endosperm can reduce the content of the UREA in the blood serum of the UC mouse, but no obvious difference exists (P> 0.05); serum CREA levels in model group mice were elevated, but there was no significant difference (PMore than 0.05), the low-dose group, the medium-dose group, the high-dose group and the positive drug mesalazine of the saponin endosperm can reduce the CREA content in the serum of UC mice, but no significant difference exists (P>0.05)。
And (4) conclusion: the saponin endosperm can reduce the content of AST, ALT, UREA and CREA in serum, and has certain improvement effect on liver and kidney injury.
Claims (8)
1. Use of semen Gleditsiae extract in preparing medicine for treating colitis ulcerosa is provided.
2. Use of an endosperm extract of saponin as claimed in claim 1 in the manufacture of a medicament for the treatment of ulcerative colitis, wherein: the preparation method of the saponin endosperm extract comprises the following steps: pulverizing semen Gleditsiae endosperm, sieving, slowly adding into boiling water while stirring, mixing, decocting with slow fire, and concentrating to obtain fluid extract or soft extract.
3. Use of an endosperm extract of saponin as claimed in claim 2 in the manufacture of a medicament for the treatment of ulcerative colitis, wherein: the preparation method of the saponin endosperm extract comprises the following steps: pulverizing semen Gleditsiae endosperm, sieving with 8-500 mesh sieve, slowly adding 2-12 times of boiling water while stirring, mixing, decocting with slow fire for 10-60min, and concentrating to obtain fluid extract or soft extract.
4. Use of an endosperm extract of saponin as claimed in claim 1 in the manufacture of a medicament for the treatment of ulcerative colitis, wherein: the preparation method of the saponin endosperm extract comprises the following steps:
a) Pulverizing endosperm of fructus Gleditsiae Abnormalis, sieving, adding into water, and heating to obtain product A;
b) Pulverizing semen Gleditsiae Abnormalis endosperm, sieving, adding the above A, stirring, mixing, decocting with slow fire, and concentrating to obtain fluid extract or soft extract.
5. Use of an endosperm extract of saponin as claimed in claim 4 in the manufacture of a medicament for the treatment of ulcerative colitis, wherein: the preparation method of the saponin endosperm extract comprises the following steps:
a) Pulverizing semen Gleditsiae, sieving with 8-500 mesh sieve, adding 3-12 times of water, and heating for 2-10 min to obtain product A;
b) Pulverizing semen Gleditsiae endosperm, sieving with 8-500 mesh sieve, adding into the above A product while stirring, mixing, decocting with slow fire for 10-60min, and concentrating to obtain fluid extract or soft extract.
6. Use of fructus Gleditsiae Abnormalis endosperm in preparing medicine for treating colitis ulcerosa is provided.
7. A medicine for treating cerebral ulcerative colitis is characterized in that: the medicine comprises saponin endosperm extract or saponin endosperm.
8. A medicine for treating the cerebral ulcerative colitis is characterized in that: the main active component of the medicine is saponin endosperm extract or saponin endosperm.
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