The Effects of Patchouli Alcohol on Diarrhea-Predominant Irritable Bowel Syndrome are Correlated with Phenotypic Plasticity in Myenteric Neurons and the Targeted Regulation of Myosin Va.

Patchouli alcohol (PA) has been widely used for the treatment of diarrhea-predominant irritable bowel syndrome (IBS-D) in traditional Chinese medicine, and the related mechanism remains to be fully understood. Our previous study has indicated that PA significantly reduced visceral sensitivity and defecation area in IBS-D rats. In this study, we prepared an IBS-D rat model and observed the dynamic intestinal motility and colonic longitudinal muscle and myenteric plexus (LMMP) neurons, as well as their subtypes at D14, D21, and D28. After PA administration, we observed the effects on the changes in intestinal motility, colonic LMMP neurons, and LMMP Myosin Va in IBS-D rats and their co-localization with inhibitory neurotransmitter-related proteins. The results indicated that PA treatment could alleviate IBS-D symptoms, regulate the abnormal expression of LMMP neurons, increase Myosin Va expression, up-regulate co-localization levels of Myosin Va with neuronal nitric oxide synthase (nNOS), and promote co-localization levels of Myosin Va with vasoactive intestinal polypeptide (VIP). In conclusion, this study demonstrated the neuropathic alterations in the colon of chronic restraint stress-induced IBS-D rat model. PA reversed the neuropathological alteration by affecting the transport process of nNOS and VIP vesicles via Myosin Va and the function of LMMP inhibitory neurons, and these effects were related to the mechanism of enteric nervous system (ENS) remodeling.

Suo Quan Wan Protects Mouse From Early Diabetic Bladder Dysfunction by Mediating Motor Protein Myosin Va and Transporter Protein SLC17A9.

Objective: To investigate the effects of Suo Quan Wan (SQW), a traditional Chinese herbal formula, on the overactive bladder (OAB) of type 2 diabetes mellitus (T2DM) mouse models, particularly on its function of mediating the gene and protein expression levels of myosin Va and SLC17A9. Materials and Methods: After 4 weeks high-fat diet (HFD) feeding, C57BL/6J mice were injected with streptozotocin (100 mg/kg) for four times. After 3 weeks, the diabetic mice were treated with SQW for another 3 weeks. Voided stain on paper assay, fasting blood glucose (FBG) test, and oral glucose tolerance test (OGTT) were conducted. Urodynamic test, tension test [α,ß-methylene ATP, electrical-field stimulation (EFS), KCl, and carbachol] and histomorphometry were also performed. Western blot analysis and qPCR assays were used to quantify the expression levels of myosin Va and SLC17A9. Results: The diabetic mice exhibited decreased weight but increased water intake, urine production, FBG, and OGTT. No significant changes were observed after 3 weeks SQW treatment. Urodynamic test indicated that the non-voiding contraction (NVC) frequency, maximum bladder capacity (MBC), residual volume (RV), and bladder compliance (BC) were remarkably increased in the diabetic mice, whereas the voided efficiency (VE) was decreased as a feature of overactivity. Compared with the model mice, SQW treatment significantly improved urodynamic urination with decreased NVC, MBC, RV, and BC, and increased VE. Histomorphometry results showed that the bladder wall of the diabetic mice thickened, and SQW effectively attenuated the pathological alterations. The contract responses of bladder strips to all stimulators were higher in the DSM strips of diabetic mice, whereas SQW treatment markedly decreased the contraction response for all stimuli. Moreover, the protein and gene expression levels of myosin Va and SLC17A9 were up-regulated in the bladders of diabetic mice, but SQW treatment restored such alterations. Conclusion: T2DM mice exhibited the early phase of diabetic bladder dysfunction (DBD) characterized by OAB and bladder dysfunction. SQW can improve the bladder storage and micturition of DBD mice by mediating the protein and gene expression levels of myosin Va and SLC17A9 in the bladder, instead of improving the blood glucose level.

Inhibitory effects of patchouli alcohol on stress-induced diarrhea-predominant irritable bowel syndrome.

AIM: To elucidate the mechanism of patchouli alcohol (PA) in treatment of rat models of diarrhea-predominant irritable bowel syndrome (IBS-D). METHODS: We studied the effects of PA on colonic spontaneous motility using its cumulative log concentration (3 × 10-7 mol/L to 1 × 10-4 mol/L). We then determined the responses of the proximal and distal colon segments of rats to the following stimuli: (1) carbachol (1 × 10-9 mol/L to 1 × 10-5 mol/L); (2) neurotransmitter antagonists including Nω-nitro-L-arginine methyl ester hydrochloride (10 µmol/L) and (1R*, 2S*)-4-[2-Iodo-6-(methylamino)-9H-purin-9-yl]-2-(phosphonooxy)bicyclo[3.1.0]hexane-1-methanol dihydrogen phosphate ester tetraammonium salt (1 µmol/L); (3) agonist α,ß-methyleneadenosine 5'-triphosphate trisodium salt (100 µmol/L); and (4) single KCl doses (120 mmol/L). The effects of blockers against antagonist responses were also assessed by pretreatment with PA (100 µmol/L) for 1 min. Electrical-field stimulation (40 V, 2-30 Hz, 0.5 ms pulse duration, and 10 s) was performed to observe nonadrenergic, noncholinergic neurotransmitter release in IBS-D rat colon. The ATP level of Kreb's solution was also determined. RESULTS: PA exerted a concentration-dependent inhibitory effect on the spontaneous contraction of the colonic longitudinal smooth muscle, and the half maximal effective concentration (EC50) was 41.9 µmol/L. In comparison with the KCl-treated IBS-D group, the contractile response (mg contractions) in the PA + KCl-treated IBS-D group (11.87 ± 3.34) was significantly decreased in the peak tension (P < 0.01). Compared with CCh-treated IBS-D rat colon, the cholinergic contractile response of IBS-D rat colonic smooth muscle (EC50 = 0.94 µmol/L) was significantly decreased by PA (EC50 = 37.43 µmol/L) (P < 0.05). Lack of nitrergic neurotransmitter release in stress-induced IBS-D rats showed contraction effects on colonic smooth muscle. Pretreatment with PA resulted in inhibitory effect on L-NAME-induced (10 µmol/L) contraction (P < 0.05). ATP might not be the main neurotransmitter involved in inhibitory effects of PA in the colonic relaxation of stress-induced IBS-D rats. CONCLUSION: PA application may serve as a new therapeutic approach for IBS-D.

Effect of the serum of rats treated with Suo Quan pill on embryonic stem cells.

Suo Quan pill(SQP), a well-known and classical traditional Chinese medicine compound, consists of three traditional Chinese medicine: Alpinia oxyphylla Miq., Lindera aggregata (Sims) Kosterm., Dioscorea opposite. Its effect was summarized as supplementing kidney-yang and shrinkaging urination. This study evaluated the effects of the serum of rats treated with Suo Quan pill on embryonic stem cells(ES cells). Cell proliferation was detected by MTT assay. Cell cycle and apoptosis of ES cells were evaluated with flow cytometry. Nanog mRNA expression was verified by fluorescence quantitative PCR and Nanog protein in ES cells was determined by Western blot. The serum of SQP-treated rats not only promoted ES cells proliferation and Nanog expression in ES cells, but also inhibited H202 stimulated cell apoptosis. Furthermore, the serum of rats containing SQP affected the cell cycle distribution of ES cells, reducing the percentage of cells in G0/G1phase and increasing the percentage of cells in G2/M phase, increasing the proliferation index of ES cells. These results illustrate that the enhanced effect of SQP on ES cells proliferation is in part due to the increased expression of Nanog in ES cells, the accelerated cell cycle period and the inhibited apoptosis of ES cells.

[Mechanism of anti-Helicobacter pylori urease activity of patchouli alcohol].

To investigate the effect of patchouli alcohol on inhibiting Helicobater pylori urease activity, and its effect on expression levels of related genes, and lay the foundation for further research on the effect of patchouli alcohol on H. pylori colonization and infection. H. pyloriwas cultured and identified by gram staining, rapid urease test (RUT) and PCR method. Then agar dilution method was used to detect the bacterial survival after 1 h intervention by different concentrations of patchouli alcoholin the acidic (pH 5.3) and neutral (pH 7.0) conditions; berthelot method was used to detect urease activity and RT-qPCR method was used to detect the expression changes of ureA, ureB, ureE, ureH, ureI, and nixA related urease genes. The results showed that the survival rate of H. pyloriwas not significantly changed but the urease activity was obviously decreased after intervention by different concentrations of patchouli alcohol; meanwhile, the expression levels of ureA, ureB, ureE, ureH, ureI, and nixA were decreased to different degrees. Therefore, patchouli alcohol could inhibit H. pylori urease activity in both acidic and neutral conditions, and the mechanism may be related to down-regulation of urease gene expression.

Danshensu, a major water-soluble component of Salvia miltiorrhiza, enhances the radioresponse for Lewis Lung Carcinoma xenografts in mice.

The molecule 3-(3,4-dihydroxyphenyl)-2-hydroxypropanoic acid (danshensu), a herbal preparation used in traditional Chinese medicine, has been found to possess potential antitumor and anti-angiogenesis effects. The aim of the present study was to investigate the efficacy of the combination of radiation therapy (RT) with danshensu in the treatment of Lewis lung carcinoma (LLC) xenografts, whilst exploring and evaluating the mechanism involved. In total, 8-week old female C57BL/6J mice were randomly assigned into 3 groups to receive: RT, RT + cisplatin and RT + danshensu, respectively, when LLC reached 100-150 mm3. Each group was divided into 7 subgroups according to the different irradiation doses that were administered. Tumor growth curves were created and the sensitization enhancement ratios of the drugs were calculated. The experiment was then repeated, and the 4 groups of tumor-bearing mice were treated with natural saline, danshensu, RT + danshensu and RT, respectively. The mice were sacrificed on day 7, and tumor tissue and blood were collected to determine microvessel density, the expression of proangiogenic factors, and the levels of blood thromboxane B2 and 6-keto-prostaglandin-F1α. Tumor hypoxia was also detected using in vivo fluorescence imaging. With respect to LLC xenografts, treatment with danshensu + RT significantly enhanced the effects of tumor growth inhibition (P<0.05). Furthermore, tumor vasculature was remodeled and microcirculation was improved, which significantly reduced tumor hypoxia (P<0.05). The present study demonstrated that danshensu significantly enhanced the radioresponse of LLC xenografts in mice. The mechanism involved may be associated with the alleviation of tumor cell hypoxia following treatment with danshensu + RT, caused by the improvement of tumor microcirculation and the remodeling of tumor vasculature.

[Effects of Suoquan Wan and Shenqi Wan on Urethra Function and ß-AR Function of Detrusor in Natural Aged Rats].

OBJECTIVE: To investigate the effect of Suoquan Wan(SQW) and Jinkui Shenqi Wan(JKSQW) on urethra function and ß6-AR function of detrusor in natural aged rats. METHODS: young rats(3 months) and Aged rats(15 months) were chosen. Young rats were chosen as control, aged rats were randomly divided into three groups: model, JKSQW and SQW groups, 12 rats in each group. JKSQW and SQW were given the appropriate concentration once a day for six weeks. The effects of SQW and JKSQW on relaxation function of bladder detrusor was investigated, and their effects on bladder induced by ß3-AR agonist and ß3-AR agonist were further studied. Then, their effects on pressure of urethra were observed. RESULTS: Compared with the young group, detrusor compliance of natural aged model rats was increased, Emax and IA of agonist of ß3-AR including BRL37344 and ISO were decreased(P <0. 01), while PA2 of antagonist of ß3-AR were increased(P <0. 05). Compared with model group, SQW and JKSQW decreased the bladder compliance(P <0. 05), and increased Emax, IA and PD2 due to sensitivity of detrusor to agonist of ß3-AR and ß3-AR including BRL37344 and ISO(P <0. 05 or P < 0. 01) ,while decreased PA2 of antagonist of ß3-AR(P <0. 05). MUCP, MUP and FUL of aged rats were lower than those of normal rats. But SQW and JKSQW increased MUCP and MUP, and JKSQW increased FUL of aged rats(P <0. 05 or P <0. 01). CONCLUSION: SQW and JKSQW can remarkably adjust ß3-AR function on the detrusor and improve the closure ability of bladder detrusor of the natural aged rats.

[Study on efficacy and mechanism of weiyangning pills against experimental gastric ulcer].

OBJECTIVE: To study the efficacy and mechanism of Weiyangning pills against experimental gastric ulcer. METHOD: The gastric ulcer model were established by acetic acid, water-immersion stress, aspirin induction, pyloric ligation in rats, in order to observe the effect of Weiyangning pills against experimental gastric ulcer and study its effect on the content of nitric oxide (NO) and epidermal growth factor (EGF), gastric mucosal blood flow, the content of PGE2, gastric secretion, gastric acid content and the activity of pepsin. RESULT: Weiyangning pills markedly reduced index of gastric ulcers of various types, increased the content of NO, EGF, PGE2 and gastric mucosal blood flow, inhibited gastric secretion and gastric acid content, and decreased the activity of pepsin. CONCLUSION: Weiyangning pills has a significant effect against experimental gastric ulcer, which is related to the reduction of gastric mucosa damage factors (gastric acid and pepsin) and the increase in gastric mucosa's function as a barrier and its recovery effects, such as NO, EGF, PGE2 and gastric mucosal blood flow.

[The influence of suoquan capsule on the mRNA expression of CYP11B2 in deficiency of the kidney and diuresis rats].

OBJECTIVE: To explore the mechanism of nourishing kidney and reducing urine effect of suoquan capsule by discussing the adjusting action on ALD synthase. METHODS: Built the model of deficiency of the kidney and diuresis by adenine,inspectd the contents of Cort, ALD in blood and the mRNA expression of CYP11B2 with ELISA and RT-PCR technology of the mice. RESULTS: Compared with model group, Suoquan capsule could remarkedly increase the contents of Cort, ALD in blood and the mRNA expression of CYP1 1 B2 of model rats. CONCLUSIONS: Suoquan capsule can increase the contents of Cort, ALD in blood and the mRNA expression of CYP11B2 in deficiency of the kidney and diuresis rats. Promote the combining of ALD by adjusting the ALD synthase may be the mechanism of nourishing the kidney and reducing urine of Souquan capsule from ALD synthase approach.

[Impacts of the formula of Suoquanwan(SQW) on expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency].

OBJECTIVE: To observe the impacts of the formula of Suoquanwan (SQW) on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency. METHODS: The model rats were induced by adenine (250 mg/kg) for 4 weeks, then treated respectively with SQW or dDAVP. The expression of AQP-2 mRNA and AVPR-V2 mRNA in kidney of Yang-deficiency model by realtime fluorescence quantitative PCR method were investigated. RESULTS: In model rats, the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney decreased, dDAVP and SQW high dose could increased the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. The others had no influence on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. CONCLUSION: SQW can increase the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency.