Canagliflozin alleviates high glucose-induced peritoneal fibrosis via HIF-1α inhibition.

The cardioprotective effects of sodium-glucose cotransporter type 2 (SGLT2) inhibitors have been demonstrated in many studies. However, their benefits for end-stage kidney disease patients, particularly those on peritoneal dialysis, remain unclear. SGLT2 inhibition has shown peritoneal protective effects in some studies, but the mechanisms are still unknown. Herein, we investigated the peritoneal protective mechanisms of Canagliflozin in vitro by simulating hypoxia with CoCl2 in human peritoneal mesothelial cells (HPMCs) and rats by intraperitoneal injection of 4.25% peritoneal dialysate simulating chronic high glucose exposure. CoCl2 hypoxic intervention significantly increased HIF-1α abundance in HPMCs, activated TGF-ß/p-Smad3 signaling, and promoted the production of fibrotic proteins (Fibronectin, COL1A2, and α-SMA). Meanwhile, Canagliflozin significantly improved the hypoxia of HPMCs, decreased HIF-1α abundance, inhibited TGF-ß/p-Smad3 signaling, and decreased the expression of fibrotic proteins. Five-week intraperitoneal injection of 4.25% peritoneal dialysate remarkably increased peritoneal HIF-1α/TGF-ß/p-Smad3 signaling and promoted peritoneal fibrosis and peritoneal thickening. At the same time, Canagliflozin significantly inhibited the HIF-1α/TGF-ß/p-Smad3 signaling, prevented peritoneal fibrosis and peritoneal thickening, and improved peritoneal transportation and ultrafiltration. High glucose peritoneal dialysate increased the expression of peritoneal GLUT1, GLUT3 and SGLT2, all of which were inhibited by Canagliflozin. In conclusion, we showed that Canagliflozin could improve peritoneal fibrosis and function by ameliorating peritoneal hypoxia and inhibiting the HIF-1α/TGF-ß/p-Smad3 signaling pathway, providing theoretical support for the clinical use of SGLT2 inhibitors in patients on peritoneal dialysis.

[Study on effect of gypenosides on insulin sensitivity of rats with diabetes mellitus via regulating NF-κB signaling pathway].

This study focused on the ameliorative effects of gypenosides(GPS) on insulin sensitivity and inflammatory factors in rats with type 2 diabetes mellitus(T2 DM) and explored their possible molecular mechanisms. After the successful establishment of T2 DM model, diabetic rats were randomly divided into four groups, including model group, GPS groups(200, 100 mg·kg~(-1)) and metformin group(100 mg·kg~(-1)), with healthy rats serving as the control. After 6-week intragastric administration, fasting blood glucose(FBG) and oral glucose tolerance were examined. The levels of insulin, C-peptide, tumor necrosis factor-α(TNF-α), interleukin-1ß(IL-1ß), interleukin-6(IL-6) and C-reactive protein(CRP) in serum were examined. Then the homeostasis model assessment of insulin resistance(HOMA-IR) and insulin sensitivity index(ISI) were calculated. The protein expression levels of phosphorylated insulin receptor substrate-1(p-IRS-1) and phosphorylated protein kinase B(p-Akt) in skeletal muscle were measured by Western blot, as well as those of phosphorylated inhibitor of nuclear factor-κB(NF-κB) kinase ß(p-IKKß), phosphorylated alpha inhibitor of NF-κB(p-IκBα) and phosphorylated p65 subunit of NF-κB(p-p65) in adipose tissue. The relative expression levels of glucose transporter 4(GLUT4) mRNA in skeletal muscle and NF-κB mRNA in adipose tissue were measured by qRT-PCR, and the morphological changes of pancreatic tissue were observed. Compared with the model group, the GPS groups witnessed significant decrease in FBG, marked amelioration of impaired oral glucose tolerance and significant increase in ISI. Further, the high-dose GPS group saw significantly reduced HOMA-IR, TNF-α, IL-1ß and CRP, significantly increased expression levels of p-IRS-1(Tyr), p-Akt and GLUT4, and markedly inhibited p-IRS-1(Ser), p-IKKß, p-IκBα, p-p65 and NF-κB. The concentration of CRP and the expression levels of p-IRS-1(Ser), p-IKKß, p-IκBα and NF-κB were remarkably reduced in the low-dose GPS group. However, GPS was found less effective in the regulation of serum insulin, C-peptide and IL-6 levels and the alleviation of pancreatic islet injury. The results indicated that GPS can reduce FBG and improve insulin sensitivity in diabetic rats possibly by regulating the NF-κB signaling pathway, inhibiting inflammation, and thereby regulating the expression of key proteins in the insulin signaling pathway.

[Pollution Characteristics and Source Apportionment of PM2.5 in Heating and Non-heating Periods in Shenyang].

To study the atmospheric PM2.5 pollution characteristics and sources in heating and non-heating periods in Shenyang, 113 groups of effective PM2.5 samples were collected from January 29, 2015 to January 26, 2016, and the water-soluble ions, carbon constituents and elements in PM2.5 were tested. The results indicated that the average PM2.5 mass concentration in Shenyang during the sampling period was 66 µg·m-3. Among the sampled PM2.5 concentrations, 31.0% exceeded the daily value of the secondary standard limit of the Chinese National Ambient Air Quality Standard (75 µg·m-3). The average concentration and over-standard rate of PM2.5 in the heating period (90 µg·m-3, 68.6%) was higher than that of the non-heating period (51 µg·m-3, 31.4%). The concentrations of the 21 elements (except Mg, Ti, Ca, Fe, and Si), water-soluble ions (except Ca2+), OC, and EC were all higher in the heating period than in the non-heating period. The ratio of[NO3-]/[SO42-]showed that the influence of moving source increased obviously in the non-heating period, and fixed source was still the main contributor in the heating period. The water-soluble ions were the result of the interaction of fixed source and moving source. The NOR and SOR analyses showed that the secondary conversion of NOx was weak, and the secondary conversion of SO2 was obvious, especially in the non-heating period. The enrichment factor showed that the elements with high EF value mainly came from coal burning, traffic pollution, and industrial emissions. The reconstructed PM2.5 masses were highly correlated with the measured ones. The main constituents of PM2.5 in both heating and non-heating seasons were organic matter (28.0%, 23.1%), mineral dust (14.5%, 26.0%), and sulfate (15.1%, 19.9%), and PM2.5 was mainly affected by the secondary particles, combustion sources and dust sources.

[Textual research on Zhuru Tang].

Zhuru Tang( from the Effective Prescriptions for Universal Relief) listed in the Catalogue of Ancient Classical Formulas( The First Batch) by the State Administration of Traditional Chinese Medicine,is usually used to treat stomach fever and vomiting. The first step of the research and development of the classic formula compound preparations is to follow the principle of the ancient method,comb through the literature of all dynasties,and then investigate the historical evolution of the prescription,the evolution of formula significance and the clinical application. Based on this principle,we searched the Chinese Medical Classics Database and relevant literature materials to conduct textual research on the history,evolution of formula significance,clinical application,decocting method,as well as the basis and processing of traditional Chinese medicine from the perspectives of " recipe" and " medicine",in order to provide reference for the development and research of Zhuru Tang.

Cardamonin Regulates miR-21 Expression and Suppresses Angiogenesis Induced by Vascular Endothelial Growth Factor.

Cardamonin has promising potential in cancer prevention and therapy by interacting with proteins and modifying the expressions and activities, including factors of cell survival, proliferation, and angiogenesis. In our precious study, we have demonstrated that cardamonin suppressed vascular endothelial growth factor- (VEGF-) induced angiogenesis as evaluated in the mouse aortic ring assay. It is also known that microRNAs (miRNAs) play important roles in angiogenesis. Herein, we hypothesized whether antiangiogenesis effect of cardamonin in human umbilical vein endothelial cells (HUVECs) triggered by VEGF was associated with miRNAs. We found that cardamonin reduced the miR-21 expression induced by VEGF in HUVECs. Treatment with miR-21 mimics abolished the effects of cardamonin on VEGF-induced cell proliferation, migration, and angiogenesis in HUVECs. However, treatment with miR-21 inhibitors presented the opposite effects, indicating the vital role of miR-21 in this process. Our study provides a new insight of the preliminary mechanism of anti-VEGF-induced angiogenesis by cardamonin in HUVECs.

Flavonoids from the leaves of Carya cathayensis Sarg. inhibit vascular endothelial growth factor-induced angiogenesis.

The total flavonoids (TFs) were isolated from the leaves of Carya cathayensis Sarg. (LCC), a well-known Chinese medicinal herb commercially cultivated in Tianmu Mountain district, a cross area of Zhejiang and Anhui provinces in China. Five flavonoids, i.e. cardamonin, pinostrobin chalcone (PC), wogonin, chrysin, and pinocembrin were the main components of the TFs. The TFs and these pure compounds suppressed vascular endothelial growth factor (VEGF)-induced angiogenesis as detected in the mouse aortic ring assay, and cardamonin showed the best effect among them. To further elucidate the mechanisms for suppressing angiogenesis of these flavonoids, assays of VEGF-induced proliferation and migration in human umbilical vein endothelial cells (HUVECs) were performed. The TFs, cardamonin, pinocembrin, and chrysin obviously suppressed both VEGF-induced HUVEC proliferation and migration. However, PC and wogonin not only slightly inhibited VEGF-induced proliferation but also remarkably suppressed those of migration in HUVECs. Our further study showed that cardamonin decreased the phosphorylation of ERK and AKT induced by VEGF with a dose-dependent manner in HUVECs. Our findings indicate that the TFs and these pure flavonoids may become potential preventive and/or therapeutic agents against angiogenesis-related diseases.

[Effect of IL-17 on collagen I/III expression in cardiac fibroblasts isolated from BALB/c mice].

AIM: To investigate the effect of IL-17 on the expression of collagen I/III in cardiac fibroblasts and analyze its molecular mechanism. METHODS: Cardiac fibroblasts were isolated from 7-14-day-old BALB/c mice and cultured in DMEM with 10% fetal bovine serum (FBS). The cells were collected after IL-17 treatment for 0, 24, 48, 72 h. IL-17 receptors on cardiac fibroblasts were detected by PCR; the collagen I/III expression was analyzed by immunofluorescence; the PKCß, Erk1/2, NF-κB phosphorylation were investigated by Western blotting. RESULTS: IL-17RA/C was expressed on cardiac fibroblasts; after 24 h of IL-17 stimulation, the collagen I/III expression obviously increased; Western blotting showed that PKCß, Erk1/2 and NF-κB were phosphorylated on 30, 45, 45 min, respectively. CONCLUSION: IL-17 could induce the expression of collagen I/III in cardiac fibroblasts, which might be related with PKCß-ERK1/2-NF-κB phosphorylation.

[Release of HMGB1 by LPS-treated cardiac fibroblasts and its contribution to the production of collagen type I and III].

AIM: To investigate whether cardiac fibroblasts (CFs) treated by LPS can actively secrete high-mobility group box protein 1 (HMGB1) and to analyze the correlation between HMGB1 releasing and the accumulation of collagen type I , III . METHODS: CFs were isolated from the heart of 7-14-day-old BALB/c mice and cultured in DMEM with 10% fetal bovine serum (FBS). We collected the CFs and cell supernatants after treated by LPS for 0, 6, 12, 24, 36, 48 h, respectively. The mRNA and protein expression levels of HMGB1, collagen 1a1 (col1a1) and collagen 3a1 (col3a1) in CFs after LPS stimulation were detected by RT-PCR and Western blotting, respectively. The intracellular localization of HMGB1 in treated CFs was investigated by immunofluorescence. RESULTS: After 0-6 h of LPS stimulation, the mRNA levels of HMGB1, col1a1, col3a1 had no significant changes; but increased obviously at 12, 24, 36, 48 h. HMGB1 was found in the cell supernatant by Western blotting after 24 h LPS stimulation, and its expression decreased following the first rise in CFs. Meanwhile, immunofluorescence showed HMGB1 translocation from nucleus to cytoplasm. The levels of col1a1 and col3a1 were up-regulated in CFs after stimulation. CONCLUSION: LPS can induce HMGB1 translocation from nucleus to cytoplasm and across cellular membrane to the outside of CFs at a time-dependent manner. Col1a1 and Col3a1, which are closely associated with myocardial fibrosis, were obviously up-regulated by LPS stimulation, which indicates that actively released HMGB1 might contribute to myocardial fibrosis following the endotoxin induced-sepsis.

[Construction of eukaryotic co-expression vector of Porphyromonas gingivalis outer membrane protein ragB and glucocorticoid-induced tumor necrosis factor receptor ligand (GITRL) and its immunogenic analysis].

AIM: To construct eukaryotic co-expression vector of Porphyromonas gingivalis outer membrane protein ragB and mouse glucocorticoid-induced tumor necrosis factor receptor ligand (mGITRL) and to analyze its immunogenicity in vivo. METHODS: The ragB gene was obtained from pMD18-T-ragB, and then cloned into the eukaryotic expression vector pIRES and pIRES-mGITRL, respectively. The eukaryotic expression vectors: pIRES-ragB and pIRES-ragB-mGITRL were identified by double enzyme digestion and DNA sequencing, then transfected into COS7 cells by Lipofectamine(TM);2000. The expressions of ragB or mGITRL in COS7 cells were detected by Western blotting. The mice were immunized with the recombinant pIRES-ragB-mGITRL plasmid. The serum antibody level was determined by ELISA. RESULTS: pIRES-ragB and pIRES-ragB-mGITRL plasmids were successfully constructed. Western blotting showed that the targeted gene was over-expressed in COS7 cells and skeletal muscle cells, respectively. The high titers of antibodies against RagB were detected in mouse serum. CONCLUSION: The construction of pIRES-ragB-mGITRL co-expression vector provides the experimental basis for Porphyromonas gingivalis vaccine research, prevention and treatment of periodontitis.

[Rapid propagation of Bletilla striata by synthetic seeds technology].

OBJECTIVE: To establish a new manufacturing method for Bletilla striata synthetic seeds, and provided a new way for rapid propagation of B. striata, the correlated influential factors were studied. METHOD: The synthetic seeds were manufactured by taking seeds of B. striata as materials which were beforehand germinated in 1/2 MS medium for 10 days, and the influential factors such as artificial endosperm components, episperm substances, storage conditions and germination groundmass impact on the germination rate and seedling rate of the synthetic seeds were evaluated. RESULT: Compound 4.0% sodium alginate + 0.2 mol x L(-1) CaCl2 + 0.4 mg x L(-1) penicillin + 0.3% carbendazim powder + 0.2% sodium benzoate served as the best episperm substances while MS + 1.0 mg x L(-1) NAA + 2.0 mg x L(-1) KT as the best endosperm components, in which, high germination rate and seedling rate were obtained. The synthetic seeds storing in the 4 degrees C for a long time was able to have still high vitality. CONCLUSION: The B. striata synthetic seeds manufacturing system was established successfully, while efforts should be taken to improve the sowing technique of the synthetic seeds in non-sterile conditions.