CN117462533A - Application of genistein in preparation of Sjogren syndrome medicine - Google Patents

Application of genistein in preparation of Sjogren syndrome medicine Download PDF

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CN117462533A
CN117462533A CN202311387331.0A CN202311387331A CN117462533A CN 117462533 A CN117462533 A CN 117462533A CN 202311387331 A CN202311387331 A CN 202311387331A CN 117462533 A CN117462533 A CN 117462533A
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pharmaceutically acceptable
genistein
mice
syndrome
acceptable salt
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李江
毛天娇
陈桂萍
吴晓丹
吴丽红
王丽京
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Stomatological Hospital of Guangzhou Medical University
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Abstract

The invention belongs to the technical field of medicines, discloses application of genistein in preparation of a Sjogren syndrome medicine, and particularly discloses application of genistein, pharmaceutically acceptable salt or pharmaceutically acceptable derivatives thereof in preparation of a medicine for preventing or treating Sjogren syndrome. The application of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof in the aspect of preventing or treating the Sjogren syndrome is provided for the first time, and experiments prove that the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof has remarkable treatment effect on the Sjogren syndrome xerostomia, has no obvious toxic or side effect on other organs, and has good application prospect.

Description

Application of genistein in preparation of Sjogren syndrome medicine
Technical Field
The invention belongs to the technical field of medicines, and particularly relates to application of genistein in preparation of a Sjogren syndrome medicine.
Background
Saliva is an important component of the oral environment, is a general name of mixed liquid secreted by three pairs of large salivary glands (parotid gland, submandibular gland and sublingual gland) and a plurality of small salivary glands (labial gland, buccal gland, palatine gland and sublingual gland) in the oral cavity, and is one of important exocrine liquid of human bodies. Saliva is a digestive juice which acts first and has the functions of protecting teeth and protecting mucous membrane from immune cells, and is closely related to the functions of chewing, swallowing, taste and the like of the oral cavity. Saliva thus has an important role in the whole body.
Primary sjogren's syndrome (Primary)syndome, pSS) is an autoimmune disease mediated by the interaction of hormones, immune systems, inflammatory mediators and various environmental factors, mainly involving exocrine glands, often originating from salivary glands and/or lacrimal glands, resulting in swelling of salivary glands and/or lacrimal glands, and impaired salivary and/or lacrimal secretion. Histological features are salivary gland lymphocyte infiltration, interstitial fibrous hyperplasia, acinar atrophy, reduced expression of aquaporin 5 (AQP 5) protein, and abnormal distribution. Clinical symptoms are manifested as dry mouth and/or dry eyes. And is accompanied by various autoimmune disorders, such as positive serum anti-SSA (anti-Ro) antibodies and anti-SSB (anti-La) antibodies, and severe cases with lymphomas, with high mortality. The disease occurs worldwide and is frequently generated in women with 40-60 years old (the ratio of men to women is 1:9-1:20), the disease incidence is obviously increased under the global aging trend, the exact disease incidence and the disease incidence in China are not clear, but the number of cases reported in domestic literature is obviously increased.
At present, the clinical treatment of pSS is mainly divided into symptomatic treatment of light diseases, such as supplementing artificial saliva and tears, the curative effect is poor and has dependence, and the immunosuppressant is used for more serious patients, the curative effect is poor and has great side effects, for example, adverse reactions such as sweating, flushing, headache, nausea and the like can occur after long-term administration of hydroxychloroquine sulfate. Thus, finding a safe and effective treatment regimen is now an urgent challenge.
Genistein (GEN) is a natural isoflavone mainly existing in leguminous plants, has a molecular structure similar to that of 17 beta-estradiol in human body, and has a weak estrogenic effect. Genistein with molecular formula of C 15 H 10 O 5 CAS number 446-72-0. The pharmacological actions are as follows: has estrogenic and antiestrogenic properties, can inhibit the activity of tyrosine protein kinase (PTK) and topoisomerase II, and has anticancer, antioxidant, apoptosis inducing, and angiogenesis inhibiting effects.
Disclosure of Invention
The object of the first aspect of the present invention is to provide the use of genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives for the preparation of a medicament for the prevention or treatment of sjogren's syndrome.
The object of the second aspect of the present invention is to provide the use of genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives for the preparation of a medicament for the treatment of salivary dysfunction or for the inhibition of tissue swelling.
The object of a third aspect of the present invention is to provide the use of genistein, a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable derivative thereof for the preparation of a Treg/Th17 balance modulator, an AQP5 enhancer, an Occludin enhancer, an SSA inhibitor and/or an SSB inhibitor.
The object of the fourth aspect of the invention is to provide a product.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
in a first aspect, the present invention provides the use of genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives for the preparation of a medicament for the prevention or treatment of sjogren's syndrome.
Preferably, the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative can be used for preventing or treating sjogren's syndrome by regulating Treg/Th17 cell balance of organism.
Preferably, the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative is used for preventing or treating sjogren's syndrome by up-regulating expression of salivary gland tissues AQP5 and Occludin.
Preferably, the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative achieves the purpose of preventing or treating sjogren's syndrome by inhibiting SSA and SSB expression.
Preferably, the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative achieves the purpose of preventing or treating sjogren's syndrome by improving salivation disorder.
Preferably, the content of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof is 1-500 mu M; further 1 to 100. Mu.M.
Preferably, the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative is used as the only active ingredient of the medicament.
Preferably, the medicament is in the form of cream, spray, drop, tablet, sugar-coated tablet, enteric-coated tablet, capsule, hard capsule, soft capsule, oral liquid, buccal agent, granule, pill, powder, pellet, powder or injection.
Preferably, the medicament further comprises pharmaceutically acceptable excipients.
Preferably, the pharmaceutically acceptable auxiliary materials comprise at least one of a filler, a disintegrant, a diluent, a lubricant, a binder, a wetting agent, a flavoring agent, a suspending agent, a solvent, a sustained release agent, an emulsifying agent, an absorption enhancer, a surfactant or a preservative.
Preferably, the filler includes, but is not limited to, at least one of starch, sucrose, lactose, mannitol, sorbitol, xylitol, microcrystalline cellulose, or glucose; the binder includes, but is not limited to, at least one of cellulose derivatives, alginate, starch, water, dextrin, gelatin, hydroxypropyl cellulose, methyl cellulose, or polyvinylpyrrolidone; the diluents include, but are not limited to, at least one of lactose, sucrose, mannitol, corn starch, potato starch, calcium phosphate, calcium citrate, and crystalline cellulose; the disintegrating agent includes, but is not limited to, at least one of corn starch, potato starch, microcrystalline cellulose, sodium carboxymethyl starch, cross-linked polyvinylpyrrolidone, low-substituted hydroxypropyl cellulose, sodium carboxymethyl starch, carboxymethyl cellulose, cross-linked sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, and alginic acid; the lubricant includes, but is not limited to, at least one of stearic acid, polyethylene glycol, calcium carbonate, sodium bicarbonate, micro silica gel, talc, anhydrous silica gel, and magnesium stearate: the suspending agent comprises at least one of silica gel micropowder, beeswax, cellulose, sodium carboxymethylcellulose and solid polyethylene glycol: the wetting agent includes, but is not limited to, at least one of glycerol, tween-80, oxyhydrogenated castor oil, sodium dodecyl sulfate, and lecithin: the solvent includes, but is not limited to, at least one of ethanol, liquid polyethylene glycol, isopropanol, tween-80, glycerol, propylene glycol, and vegetable oils including, but not limited to, at least one of soybean oil, castor oil, peanut oil, blend oil: the surfactant includes, but is not limited to, at least one of sodium dodecyl benzene sulfonate, stearic acid, polyoxyethylene-polyoxypropylene copolymer, sorbitan fatty acid, and polysorbate (tween): the flavoring agent comprises at least one of aspartame, sucralose, essence, steviosin, acesulfame potassium, citric acid and saccharin sodium; the preservative includes, but is not limited to, at least one of methylparaben or propylparaben.
Preferably, the auxiliary materials comprise one or a mixture of more of starch, sorbitol, maltose, glucose, sucrose, lactose, silicon derivatives, cellulose and its derivatives, gelatin, glycerol, talcum powder, calcium carbonate, calcium bicarbonate, distilled water and cyclodextrin.
In a second aspect, the present invention provides the use of genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives for the preparation of a medicament for the treatment of salivary dysfunction or for the inhibition of tissue swelling.
Preferably, the tissue comprises at least one of the submandibular gland, thymus and spleen.
Preferably, the content of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof is 1-500 mu M; further 1 to 100. Mu.M.
Preferably, the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative is used as the only active ingredient of the medicament.
Preferably, the medicament is in the form of cream, spray, drop, tablet, sugar-coated tablet, enteric-coated tablet, capsule, hard capsule, soft capsule, oral liquid, buccal agent, granule, pill, powder, pellet, powder or injection.
Preferably, the medicament further comprises pharmaceutically acceptable excipients.
Preferably, the pharmaceutically acceptable auxiliary materials comprise at least one of a filler, a disintegrant, a diluent, a lubricant, a binder, a wetting agent, a flavoring agent, a suspending agent, a solvent, a sustained release agent, an emulsifying agent, an absorption enhancer, a surfactant or a preservative.
Preferably, the auxiliary materials comprise one or a mixture of more of starch, sorbitol, maltose, glucose, sucrose, lactose, silicon derivatives, cellulose and its derivatives, gelatin, glycerol, talcum powder, calcium carbonate, calcium bicarbonate, distilled water and cyclodextrin.
In a third aspect, the present invention provides the use of genistein, a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable derivative thereof for the preparation of a Treg/Th17 balance modulator, an AQP5 enhancer, an Occludin enhancer, an SSA inhibitor and/or an SSB inhibitor.
Preferably, the content of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof is 1-500 mu M; further 1 to 100. Mu.M.
In a fourth aspect, the present invention provides a product comprising genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives.
Preferably, the content of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof is 1-500 mu M; further 1 to 100. Mu.M.
Preferably, the product has at least one of the functions (b 1) to (b 3);
(b1) Upregulating salivary gland tissue AQP5 and Occludin expression;
(b2) Regulating body Treg/Th17 cell balance;
(b3) Inhibit SSA and SSB expression.
Preferably, the product comprises a food and/or pharmaceutical product.
The beneficial effects of the invention are as follows:
the application of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof in the aspect of preventing or treating the Sjogren syndrome is provided for the first time, and experiments prove that the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof has remarkable treatment effect on the Sjogren syndrome xerostomia, has no obvious toxic or side effect on other organs, and has good application prospect. In addition, the genistein has early research results, has definite pharmacokinetics, and can realize new use of old drugs, thereby shortening the clinical test time and reducing the research cost, providing a new breakthrough for the treatment of clinical sjogren's syndrome and providing candidate drugs for the treatment of sjogren's syndrome.
Drawings
FIG. 1 shows in vitro experiments that genistein up-regulates the expression of human salivary gland epithelial cells (HSG cells) AQP5 and Occludin and down-regulates the expression of human salivary gland epithelial cells (HSG cells) SSA and SSB.
FIG. 2 is the effect of genistein on water intake in NOD model mice.
FIG. 3 is the effect of genistein on saliva flow rate in NOD model mice.
FIG. 4 is the effect of genistein on inflammatory infiltration of mandibular gland tissue in NOD model mice; wherein A is the result of immunohistochemical staining, scale is 1000 μm and 50 μm, B is the statistical result of immunohistochemical staining, in the figure, p is less than 0.005.
FIG. 5 shows the results of HE staining of genistein with respect to toxic and side effects on other organs, on a scale of 50. Mu.m.
FIG. 6 is the effect of genistein on expression of AQP5 in submandibular gland tissue; wherein A is the result of immunohistochemical staining, scale is 50 μm, B is the statistical result graph of the immunohistochemical staining, and p is less than 0.005 in the graph.
FIG. 7 is the effect of genistein on the number of spleen Treg cells of NOD mice.
FIG. 8 is the effect of genistein on the number of Th17 cells in the spleen of NOD mice.
FIG. 9 is a graph showing the effect of genistein on enlargement of the mandibular gland, spleen and thymus in NOD mice; wherein, A is the mandibular gland, spleen and thymus of NOD mice, B is the result statistical graph, in the graph, p is less than 0.05, p is less than 0.005.
FIG. 10 shows the effect of genistein on the expression of SSA and SSB in the serum of NOD mice.
FIG. 11 is the effect of genistein on the expression of AQP5 in the mandibular gland tissue of NOD mice.
FIG. 12 is a KEGG and Go enrichment analysis of genistein group and model group mice; wherein A is the KEGG pathway that upregulates differential gene enrichment; b is KEGG channel for down-regulating differential gene enrichment, C is Go enrichment analysis result graph.
Detailed Description
The invention will now be described in detail with reference to specific examples, without limiting the scope of the invention.
The materials, reagents and the like used in this example are commercially available materials and reagents unless otherwise specified.
Example 1 in vitro experiments
Studies have shown that 50ng/mL IFN-gamma acts on salivary gland epithelial cells, and can be used to construct the disease of Sjogren's syndrome, salivary gland dermatitis, which is used to simulate pSS in vitro. This example uses 50ng/mL IFN-gamma action on treatment of human mandibular gland cells (HSG cells) to construct an in vitro pSS model. The HSG cells used in the experiment are laboratory long-term liquid nitrogen preservation cell lines containing 5% CO at 37 DEG C 2 Culturing in a cell culture incubator. The culture was performed using 1640 medium containing 10% fetal bovine serum (FBS, sigma), and when the cells were grown to 85% saturation, the passaging was digested with 0.25% trypsin containing EDTA, passaged 1 time for 2 days, and HSG cells after passaging 3 times were used for the subsequent experiments.
Model preparation and grouping treatment:
normal control group: will beHSG cells were cultured in 6-well plates with a cell number of 10 6 2mL of 1640 medium containing 1% fetal bovine serum was added to each well, and 2. Mu.L of Phosphate Buffered Saline (PBS) was added as a control, and the wells were treated for 48 hours;
IFN-gamma group: similar to the normal control group, the only difference was that PBS was replaced with IFN- γ at a concentration of 50ng/mL, and 1% fetal bovine serum was treated at an IFN- γ concentration of 100 ng/. Mu.L in 1640 medium for 48 hours
IFN-γ+GNE 10 -8 M groups: the only difference is that the PBS was replaced with IFN-gamma and genistein to give IFN-gamma concentration of 50ng/mL and GNE concentration of 10 in 1640 medium of 1% fetal bovine serum -8 M；
IFN-γ+GNE 10 -7 M groups: the only difference is that the PBS was replaced with IFN-gamma and genistein to give IFN-gamma concentration of 50ng/mL and GNE concentration of 10 in 1640 medium of 1% fetal bovine serum -7 M；
IFN-γ+GNE 10 -6 M groups: the only difference is that the PBS was replaced with IFN-gamma and genistein to give IFN-gamma concentration of 50ng/mL and GNE concentration of 10 in 1640 medium of 1% fetal bovine serum -6 M；
After cells were collected 48 hours after each of the above groups were treated, protein was lysed and collected with RIPA, BCA quantification was performed, and finally Western Blot experiments were performed with a primary antibody of AQP5 (santa, 1:500), occidin (proteontech, 1:2000), TRIM21 (SSA, proteontech, 1:2000), SSB (proteontech, 1:2000), and beta-actin (proteontech, 1:5000), respectively. The secondary antibody was then incubated and developed.
The result of Western Blot experiment on protein in HSG cells treated by different treatment groups shows that genistein can obviously up-regulate the expression of AQP5 and Occludin protein, down-regulate the expression of SSA and SSB protein, and has certain concentration dependence, and the effect is 10 -8 M to 10 -6 M was evident as the genistein concentration increased (FIG. 1).
Example 2 in vivo experiments
10 NOD female mice with 7 weeks of age and weight of 20-22 g under SPF feeding environment are selected and randomly divided into 2 groups: pSS Model group (Model) and 50mg/kg genistein group (50 mg/kg GEN), while 5 ICR female mice of 7 weeks of age and 22-24 g of body weight were selected as Normal group (Normal). After 1 week of adaptive feeding, the week water intake was measured at 8 weeks of age and the administration was started at 8 weeks of age, each group was given by gavage, the normal group and the pSS model group were given blank solvent (0.5% sodium carboxymethyl cellulose solution) by gavage, and the genistein group was given 50mg/kg of genistein by gavage at 200. Mu.L/day. Daily administration was continued for 8 weeks. The weight, water content and the like of each group of mice are observed and detected every day and recorded.
(1) Influence of genistein on NOD mice Water intake and salivation
After the mice are adaptively fed for 1 week, the water intake of each group of mice is measured continuously and weekly at the age of 8 weeks to 18 weeks, and the result is shown in figure 2, the water intake of the normal group of mice does not obviously increase with age, and the mice have obvious statistical significance (P < 0.01) compared with the pSS model group, and the water intake of the pSS model group of mice obviously increases with age; the water intake of the genistein group mice became stable after 10 weeks of age (i.e. 2 weeks of administration), even the water intake decreased with age, and the water intake recovered to approximately normal group mice water intake level, which showed significant statistical significance (P < 0.01) compared with the pSS model group, indicating that genistein significantly improved the dry mouth symptoms of NOD mice. The saliva flow rate of the mice was measured once every other week, i.e., the saliva flow rate of each group of mice was measured at the ages of 8, 10, 12, 14, and 16 weeks, and the mice were first anesthetized, then pilocarpine stimulation (0.1 mg/kg) was administered, and the saliva flow rate of the mice was collected by a capillary pipette within 10 minutes after the stimulation, and the weighing calculation was performed. The results show that: the saliva flow rate of the mice in the normal group does not obviously decrease with age, and has statistical significance (P is less than 0.01) compared with that of the mice in the pSS model group, and the saliva flow rate of the mice in the pSS model group obviously decreases with age and rapidly decreases from 10 weeks of age; while the genistein group mice at 8 weeks of age had no obvious difference from the model group mice, and the saliva flow rate was improved at 10 weeks of age (2 weeks of administration), but had no statistical difference, and the saliva flow rate became stable after 2 weeks of administration, had no obvious decrease with age, and had a remarkable statistical significance (P < 0.01) (FIG. 3), indicating that genistein can significantly improve the saliva secretion disorder of mice.
(2) NOD mouse histopathological experiments
pSS originates from salivary glands, and the submandibular gland histopathology can observe lymphocyte infiltration foci, normal acinar structure destruction and reduced expression of AQP5, and the expression and distribution of AQP5 proteins in each group are compared by HE staining, comparing lymphocyte infiltration in each group of mice, and immunohistochemical staining. Meanwhile, in order to observe the systemic toxic and side effects of genistein, HE staining is used for observing the influence of the genistein on the heart, liver, spleen, lung, kidney and pancreas of mice, and the specific operation is as follows:
when the mice are 18 weeks old (namely 10 weeks after administration), the mice are sacrificed by a carbon dioxide asphyxiation method, eyeballs are sampled, left and right mandibular gland tissues of the mice in the anterior cervical region are completely stripped and weighed together, one side of the mice is frozen in liquid nitrogen for standby, and the other side of the mice is embedded with tissues. Anterior cervical mandibular gland tissue was fixed by 4% paraformaldehyde infusion for 48h, embedded in conventional dehydrated paraffin, serial sections 4 μm thick, and then HE stained and immunohistochemical stained, respectively.
The pathological staining of the HE slice shows that the tissue acinus of the mandibular gland of the normal mice has complete structure and no obvious inflammatory infiltration, has obvious statistical significance (P is less than 0.01) compared with the model group, and obviously increases and increases lymphocyte infiltration foci in the tissue of the mandibular gland of the model group; the genistein-dosed group had little lymphocyte infiltration range and small infiltration area, and had a remarkable inflammatory improvement effect compared with the model group, with a remarkable statistical difference (P < 0.01) (FIG. 4). The genistein is shown to be capable of obviously improving infiltration of submandibular gland tissue lymphocytes of NOD mice. Further observations of heart, liver, spleen, lung, kidney and pancreas of the normal, model and genistein-dosed groups revealed no significant differences between the above tissues, indicating that genistein had no significant side effects on the systemic viscera (fig. 5).
AQP5 is a member of the water secretion protein family, and is mainly distributed on the apical membrane of salivary gland acinar cells and is responsible for salivary secretion. By immunohistochemical staining, it was found that expression of AQP5 was significantly increased in submandibular gland tissues in the normal and genistein-administered groups compared to the model group, and was localized to the apical membrane of acinar cells (P < 0.05) (fig. 6).
(3) Effects of genistein on Treg cells and Th17 cells in spleen
The core immune mechanism of pSS pathogenesis is the overactivation of helper T cells 17 (Th 17), the suppression of regulatory T cells (tregs), and the imbalance of tregs/Th 17 in the body.
On the day of dissection, mouse spleen tissue is taken for grinding, erythrocytes are lysed, and the single cell suspension is prepared by centrifugal resuspension, 1X 10 is taken 6 And (3) resuspension of mononuclear cells, living and dead cell staining, resuspension after washing, adding a CD4 antibody, uniformly mixing, incubating for 30min at room temperature in a dark place, washing after reaction, rupture of membranes, incubating a Foxp3 antibody at room temperature in a dark place, and detecting the proportion of Treg cells in spleen tissues of each group by a machine after washing and resuspension. Likewise, 1X 10 groups are taken 6 The mononuclear cells are resuspended, stimulated by PMA/Ionomycin mixcure ionomycin and placed in a 37 ℃ incubator for 5 hours, living and dead staining are carried out on the cells, after washing, the mononuclear cells are resuspended, CD4 antibodies are added, after uniform mixing, the mononuclear cells are incubated for 30 minutes at room temperature and in a dark place, after reaction, washing is carried out, membrane rupture is carried out, IL17A antibodies are incubated at room temperature and in a dark place, and after washing and resuspension, the spleen tissues of each group are detected in a flow cytometer for Th17 cell proportion.
The results are shown in FIG. 7, where the proportion of Treg cells in the spleen was increased in the normal and genistein-dosed groups compared to the model group, with statistical significance (P < 0.05) and no significant differences between the normal and genistein-dosed groups. Meanwhile, the proportion of Th17 cells in spleens of mice in the normal group and the genistein-administered group was significantly reduced by P < 0.01) compared with the model group, while there was no significant difference between the normal group and the genistein-administered group (FIG. 8).
(4) Influence of genistein on organ index
Spleen, the largest peripheral immune organ of the body, plays an important role in the immune defenses of the body and is also the main colonisation site for B lymphocytes. Thymus is the central immune organ of the organism, and is the main place for T cell differentiation, development and maturation. Spleen and thymus enlargement are widely recognized as manifestations of activation of immune system functions. pSS patients also exhibited symptoms of salivary gland enlargement, so this example further photographed and weighed spleen, thymus, and submandibular glands of each group of mice, and calculated organ index (organ index=organ (g)/mouse body weight (g)).
As shown in fig. 9, the spleen, thymus and mandibular gland of the model group showed significant swelling, consistent with the characteristics of autoimmune disease, whereas the spleen, thymus and mandibular gland of the normal group and genistein-administered group did not showed significant swelling (a in fig. 9); by counting the organ indexes of the mice in each group, the result is shown as B in fig. 9, compared with a model group, the organ indexes of the mandibular gland, spleen and thymus in a normal group are obviously reduced, the statistical significance (P < 0.05) is achieved, the swelling of the mandibular gland, thymus and spleen in a genistein administration group is obviously reduced, the organ indexes of the mandibular gland and spleen are statistically significant (P < 0.05), and the genistein can effectively reduce the swelling of the mandibular gland, spleen and thymus, and has better treatment effect.
(5) Effects of genistein on AQP5 expression in mouse mandibular gland tissue and on SSA and SSB expression in mouse serum
At present, an increase in the expression of SSA and SSB autoantibodies has been detected in serum of pSS patients, and therefore, in this example, the content of SSA antibodies and SSB antibodies in each group of mouse serum was detected by taking mouse eyeball serum and performing ELISA detection.
The results showed that the serum concentrations of SSA and SSB were significantly reduced in the genistein-dosed mice compared to the model group, and were statistically significant (fig. 10).
The total RNA in the frozen mandibular gland tissues of each group was further extracted by Trizol, and the mRNA expression of AQP5 in the tissues was detected by a real-time fluorescent quantitative PCR (qPCR) method, and the results are shown in FIG. 11, in which the mRNA expression level of AQP5 in the mandibular gland tissues of mice of the normal group and the genistein-administered group was significantly higher than that of mice of the model group (P < 0.01).
(6) KEGG and GO enrichment analysis of genistein and model groups
On the day of dissection, the mandibular gland tissue on one side is taken for stripping, surrounding fat, fascia, lymph nodes and other tissues are removed, and the mandibular gland tissue is immediately placed into a liquid nitrogen tank for preservation, and then is transferred to-80 ℃ for preservation, and is used for subsequent experiments such as RNA sequencing and the like. Transcriptome sequencing was performed on model and genistein-dosed groups, respectively, and GO and KEGG enrichment analysis was performed.
The results were as follows: as shown in fig. 12 a, the up-regulated differential genes were mainly enriched in the model group: protein digestion and absorption pathways, salivary signal pathways, cAMP signal pathways, etc., which indicate that genistein can significantly up-regulate salivary gland secretion function. FIG. 12B is a KEGG pathway down-regulating differential gene enrichment, which shows that differential genes are enriched in FOXO signaling pathway, mTOR signaling pathway, metabolic signaling pathway, etc., indicating that genistein may regulate apoptosis and autophagy via these signaling pathways, repairing inflammatory damaged cells. FIG. 12C is a GO functional enrichment analysis.
The embodiments of the present invention have been described in detail with reference to the accompanying drawings, but the present invention is not limited to the above embodiments, and various changes can be made within the knowledge of one of ordinary skill in the art without departing from the spirit of the present invention. Furthermore, embodiments of the invention and features of the embodiments may be combined with each other without conflict.

Claims (10)

1. Use of genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives for the preparation of a medicament for preventing or treating sjogren's syndrome.
2. The use according to claim 1, wherein the genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives are used for the prevention or treatment of sjogren's syndrome by modulating the Treg/Th17 cell balance of the body.
3. The use according to claim 1, wherein said genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives are used for the prevention or treatment of sjogren's syndrome by up-regulating salivary gland tissue AQP5 and Occludin expression.
4. The use according to claim 3, wherein the genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives achieve the purpose of preventing or treating sjogren's syndrome by inhibiting SSA and SSB expression.
5. The use according to claim 1, wherein the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative is for the purpose of preventing or treating sjogren's syndrome by improving salivation disorders.
6. The use according to any one of claims 1 to 5, wherein the genistein, its pharmaceutically acceptable salt or its pharmaceutically acceptable derivative is present in an amount of 1 to 500 μm.
7. Use of genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives for the preparation of a medicament for the treatment of salivary dysfunction or for the inhibition of tissue swelling.
8. Use of genistein, a pharmaceutically acceptable salt or a pharmaceutically acceptable derivative thereof for the preparation of a Treg/Th17 balance modulator, an AQP5 enhancer, an Occludin enhancer, an SSA inhibitor and/or an SSB inhibitor.
9. The use according to claim 7 or 8, characterized in that the genistein, its pharmaceutically acceptable salts or its pharmaceutically acceptable derivatives are contained in an amount of 1 to 500 μm.
10. A product comprising genistein, a pharmaceutically acceptable salt thereof or a pharmaceutically acceptable derivative thereof; preferably, the content of the genistein, the pharmaceutically acceptable salt or the pharmaceutically acceptable derivative thereof is 1-500 mu M.
CN202311387331.0A 2023-10-24 2023-10-24 Application of genistein in preparation of Sjogren syndrome medicine Pending CN117462533A (en)

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