Identification of Rab7 as an autophagy marker: potential therapeutic approaches and the effect of Qi Teng Xiao Zhuo granule in chronic glomerulonephritis.

CONTEXT: Qi Teng Xiao Zhuo granule (QTXZG) is a traditional Chinese medicine (TCM) used for therapeutic effects on chronic glomerulonephritis (CGN). However, the underlying mechanism remains unclear. OBJECTIVE: To investigate the molecular mechanism of QTXZG on CGN by proteomics. MATERIALS AND METHODS: The CGN model was induced in Sprague-Dawley rats by injecting adriamycin (3.5 mg/kg, Day 1; 3.0 mg/kg, Day 14) twice through the tail vein. Urine samples were collected on the 21st day; and the rats divided randomly into control, adriamycin, QTXZG administration groups. Rats in the QTXZG group received QTXZG (10.8 g/kg); control and adriamycin groups were given physiological saline once per day for 30 days. Proteomics was applied to identify the candidate proteins combined with autophagy database and verified by immunofluorescence (IF) and western blots (WB). RESULTS: 278 differentially expressed proteins (DEPs) were identified based on proteomics and Rab7 was screened as an autophagy protein biomarker. In vitro cell experiments, we found that QTXZG (20%, IC50 = 23.47%) could decrease the expression of NLRP3, Caspase-1, IL-18, IL-1ß, while increasing the expression of Pink1, Parkin, Rab7, Podocalyxin. The cell apoptosis rate increased from 6.68 ± 0.07 to 11.03 ± 0.36%. Overexpression of Rab7 resulted in an increase in autophagy relevant protein expression. DISCUSSION AND CONCLUSION: TCM CGN-regulating herbs (QTXZG) can exert therapeutic effects by affecting the Rab7/Pink1/Parkin pathway to promote mitochondrial autophagy. New breakthroughs in targeted Rab7 may eventually enable such applications.

Metabolomic Analysis of Stephania tetrandra-Astragalus membranaceus Herbal Pair-Improving Nephrotic Syndrome Identifies Activation of IL-13/STAT6 Signaling Pathway.

The Stephania tetrandra−Astragalus membranaceus herbal pair (FH) is a classic herbal pair widely used in the treatment of nephrotic syndrome (NS). The effects of Stephania tetrandra (FJ) and Astragalus membranaceus (HQ) on NS have been reported, but the mechanism of their combination on the improvement of NS are still unclear. The NS model was established by injecting adriamycin into the tail vein. FH intervention reduced the levels of serum triglyceride, total cholesterol, interleukin-6 (IL-6), blood urea nitrogen (BUN), urinary protein, and the gene expression levels of aquaporin 2 (AQP2) and arginine vasopressin (AVP) in NS rats. In addition, FH improved kidney injury in NS rats by inhibiting the expression of interleukin 13 (IL-13), phospho-signal transducers, and activators of transcription 6 (p-STAT6), Bax, cleaved-caspase3, while promoting the expression of Bcl-2. By comprehensive comparison of multiple indexes, the effects of FH on lipid metabolism, glomerular filtration rate, and inflammation were superior to that of FJ and HQ. Metabonomic studies showed that, compared with FJ and HQ, FH intervention significantly regulated tricarboxylic acid (TCA) cycle, cysteine and methionine metabolism, and alanine, aspartic acid and glutamic acid metabolism. Pearson correlation analysis showed that succinic acid and L-aspartic acid were negatively correlated with urinary protein, cystatin C (Cys C) and BUN (p < 0.05). In summary, FH could reduce renal injury and improve NS through inhibiting the IL-13/STAT6 signal pathway, regulating endogenous metabolic pathways, such as TCA cycle, and inhibiting the expression of AQP2 and AVP genes. This study provides a comprehensive strategy to reveal the mechanism of FH on the treatment of NS, and also provides a reasonable way to clarify the compatibility of traditional Chinese medicine.

Network pharmacology-based identification of miRNA expression of Astragalus membranaceus in the treatment of diabetic nephropathy.

ABSTRACT: Diabetic nephropathy (DN) is a common microvascular complication of diabetic patients, along with hypertension, hyperlipemia, proteinuria, edema, and other clinical manifestations. Astragalus membranaceus (AM) is a traditional Chinese medicine and has shown significant clinical efficacy against DN. However, the overall molecular mechanism of this therapeutic effect has not been entirely elucidated. Using network pharmacology, we aimed to identify the key active ingredients and potential pharmacological mechanisms of AM in treating DN and provide scientific evidence of its clinical efficacy.The active ingredients of AM were obtained from the traditional Chinese medicine systems pharmacology database, and the potential targets of AM were identified using the therapeutic target database. DN-related target genes were acquired from the Gene Expression Omnibus microarray dataset GSE1009 and 3 widely used databases-DisGeNET, GeneCards, and Comparative Toxicogenomics Database. The DN-AM common target protein interaction network was established by using the STRING database. Active ingredients candidate targets proteins networks were constructed using Cytoscape software for visualization. Additionally, gene ontology (GO) and Kyoto encyclopedia of genes and genomes pathway analyses were performed using the Database for Annotation, Visualization, and Integrated Discovery database. Target-regulating microRNAs (miRNAs) of these hub genes were obtained from the therapeutic target database, which could then be used for further identification of AM-regulated key miRNAs.A total of 17 active ingredients and 214 target proteins were screened from AM. 61 candidate co-expressed genes with therapeutic effects against DN were obtained and considered as potential therapeutic targets. GO and Kyoto encyclopedia of genes and genomes enrichment analysis showed that these genes were mainly involved in inflammatory response, angiogenesis, oxidative stress reaction, HIF signaling pathway, tumor necrosis factor signaling pathway, and VEGF signaling pathway. In all, 636 differentially expressed genes were identified between the DN patients and control group by using microarray data, GSE1009. Lastly, VEGFA, epidermal growth factor receptor, STAT1, and GJA1 were screened as hub genes. The relationships between miRNAs and hub genes were constructed, which showed that miR-302-3p, miR-372-3p, miR-373-3p, and miR-520-3p were regulated by VEGFA and epidermal growth factor receptor. Meanwhile, VEGFA also influenced miR-15-5p, miR-16-5p, miR-17-5p, miR-20-5p, miR-93-5p, miR-106-5p, miR-195-5p, miR-424-5p, miR-497-5p, and miR-519-3p. In addition, miR-1-3p and miR-206 were regulated by VEGFA and GJA1, and miR-23-3p was regulated by STAT1 and GJA1.To our knowledge, this study revealed for the first time the characteristic multiple components, multiple targets, and multiple pathways of AM that seem to be the underlying mechanisms of action of AM in the treatment of DN with respect to miRNAs.Private information from individuals will not be published. This systematic review also does not involve endangering participant rights. Ethical approval will not be required. The results may be published in a peer-reviewed journal or disseminated at relevant conferences.

[Identification of chemical constituents in Huangdi Anxiao Capsules by UPLC-Q-TOF-MS~E combined with UNIFI software].

In order to clarify the main chemical constituents of Huangdi Anxiao Capsules, an ultra-high performance liquid coupled with quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS~E) combined with Waters UNIFI software were successfully used to rapidly identify the chemical constituents in Huangdi Anxiao Capsules. The mass spectrometry data of chemical constituents from Huangdi Anxiao Capsules were collected by UPLC-Q-TOF-MS~E, and their structures were identified by the results of UNIFI software according to relative retention time of reference standards, MS feature fragments and literature data of each compound. A total of 100 compounds in Huangdi Anxiao Capsules were identified, including 25 compounds from Pueraria Lobate Radix, 22 compounds from Coptis Rhizoma, 6 compounds from Ophiopogonis Radix, 14 compounds from Eriobotryae Folium, 22 compounds from Rehmanniae Radix, and 15 compounds from Notoginseng Radix et Rhizoma. Among them, 3 compounds were common components. These 100 compounds included flavonoids, alkaloids, saponins and organic acids. This study systematically analyzed the chemical composition of Huangdi Anxiao Capsules, so as to provide evidences for defining the chemical material basis of Huangdi Anxiao Capsules.

Uncovering the Mechanism of Astragalus membranaceus in the Treatment of Diabetic Nephropathy Based on Network Pharmacology.

BACKGROUND: Diabetic nephropathy (DN), characterized by hyperglycemia, hypertension, proteinuria, and edema, is a unique microvascular complication of diabetes. Traditional Chinese medicine (TCM) Astragalus membranaceus (AM) has been widely used for DN in China while the pharmacological mechanisms are still unclear. This work is aimed at undertaking a network pharmacology analysis to reveal the mechanism of the effects of AM in DN. Materials and Methods. In this study, chemical constituents of AM were obtained via Traditional Chinese Medicine Systems Pharmacology Database (TCMSP), and the potential targets of AM were identified using the Therapeutic Target Database (TTD). DisGeNET and GeneCards databases were used to collect DN-related target genes. DN-AM common target protein interaction network was established by using the STRING database. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were carried out to further explore the DN mechanism and therapeutic effect of AM. The network diagrams of the active component-action target and protein-protein interaction (PPI) networks were constructed using Cytoscape software. RESULTS: A total of 16 active ingredients contained and 78 putative identified target genes were screened from AM, of which 42 overlapped with the targets of DN and were considered potential therapeutic targets. The analysis of the network results showed that the AM activity of component quercetin, formononetin, calycosin, 7-O-methylisomucronulatol, and quercetin have a good binding activity with top ten screened targets, such as VEGFA, TNF, IL-6, MAPK, CCL3, NOS3, PTGS2, IL-1ß, JUN, and EGFR. GO and KEGG analyses revealed that these targets were associated with inflammatory response, angiogenesis, oxidative stress reaction, rheumatoid arthritis, and other biological process. CONCLUSIONS: This study demonstrated the multicomponent, multitarget, and multichannel characteristics of AM, which provided a novel approach for further research of the mechanism of AM in the treatment of DN.

Effect of the traditional Chinese medicine Qi Teng Xiao Zhuo granules on chronic glomerulonephritis rats studied by using long noncoding RNAs expression profiling.

AIM OF THE STUDY: Chronic glomerulonephritis (CGN) is the most common form of primary glomerular disease. Qi Teng Xiao Zhuo granules have been proposed as a prescription of traditional Chinese medicine (TCM) for treatment of CGN, however,the comprehensive molecular mechanism underlying this therapeutic effectremains unclear to date. Our study aimed to evaluate and analyze the possible roles and molecular mechanisms of Qi Teng Xiao Zhuo granule-mediated treatment of CGN induced by adriamycin in rats. MATERIALS AND METHODS: RNA-sequencing and real-time polymerase chain reaction (RT-PCR) were applied to identify specifically expressed long noncoding RNAs (lncRNAs) in glomerular tissues of rats from the control group, adriamycin-induced group, and Qi Teng Xiao Zhuo granules group (n = 3). Differentially expressed lncRNAs and mRNAs (messengerRNAs) were screened out among the 3 groups. Gene ontology (GO) and pathway enrichment analyses were performed to analyze the biological functions and pathways for mRNAs. LncRNA-mRNA co-expression network was constructed to analyse for the genes. The protein-protein interaction (PPI) network was visualized. RESULTS: A total of 473 significantly up and down-regulated lncRNAs, 753 up and down-regulated mRNAs were identified. Additionally, it is worth noting that TOP2a (topoisomerase (DNA) II alpha), with the highest connectivity degree in PPI network, was enriched in variouskinds of pathways. Coding-non-coding gene co-expression networks (CNC network) were drawn based on the correlation analysis between lncRNAs and mRNAs. Ten lncRNAs, NONRATT009275.2, NONRATT025409.2, NONRATT025419.2, MSTRG.7681.1, ENSRNOT00000084373, NONRATT000512.2, NONRATT006734.2, ENSRNOT00000084386, NONRATT021738.2, ENSRNOT00000084080, were selected to analyse the relationship between LncRNAs and Qi Teng Xiao Zhuo granules via the CNC network (Coding-non-coding gene co-expression networks) and GO analysis. Real-time PCR results confirmed that the six lncRNAs were specifically expressed in the Qi Teng Xiao Zhuo granules rats. CONCLUSIONS: The ten lncRNAs might play important roles in the Qi Teng Xiao Zhuo granules treatment of CGN. Key genes, such as Ptprc (protein tyrosine phosphatase, receptor type, C), TOP2a, Fos (FBJ osteosarcoma oncogene), Myc (myelocytomatosis oncogene), etc, may be crucial biomarkers for Qi Teng Xiao Zhuo granules.

Effects of Qi Teng Xiao Zhuo granules on circRNA expression profiles in rats with chronic glomerulonephritis.

Objectives: To screen and study circular RNA (circRNA) expression profiles in QTXZG-mediated treatment of chronic glomerulonephritis (CGN) induced by adriamycin in rats and to research the possible roles and molecular mechanisms of QTXZG. Materials and methods: Next-generation RNA sequencing was used to identify circRNA expression profiles in CGN after QTXZG treatment compared with a CGN model group and a control group. Bioinformatics analysis was performed to predict potential target miRNAs and mRNAs. GO and pathway analyses for potential target mRNAs were used to explore the potential roles of differentially expressed (DE) circRNAs. Results: We identified 31 and 21 significantly DE circRNAs between the model group vs the control group and the model group vs the QTXZG group, respectively. Four circRNAs that resulted from the establishment of the CGN model were reversed following treatment with QTXZG. Further analysis revealed that these four circRNAs may play important roles in the development of CGN. Conclusions: This study elucidated the comprehensive expression profile of circRNAs in CGN rats after QTXZG treatment for the first time. Analysis of the circRNA-miRNA-mRNA-ceRNA network to determine potential function provided a comprehensive understanding of circRNAs that may be involved in the development of CGN. The current study indicated that therapeutic effects of QTXZG on CGN may be due to regulation of circRNA expression.

[Effect of Huangdi Anxiao Capsules on zebrafish vascular lesions induced by high glucose and high fat].

Zebrafish of different strains with 5 dpf (5 days post-fertilization) were selected and fed with 0.2% high-fat diet for 8 h and 3% glucose solution for 16 halternatively during the day and night for 4 consecutive days. The zebrafish model was established and randomly divided into model group, Huangdi Anxiao Capsules (260 mg·L⁻¹) group and pioglitazone (32 mg·L⁻¹) group. The drug treatment groups were given the water-soluble drugs, with a volume of 25 mL, and incubated in a 28 °C incubator for 4 days. To detect the exposure to the corresponding drugs, the normal control group was set up. Thirty zebrafish were included in each group. The effect of Huangdi Anxiao Capsules on vascular wall thickness, fluorescence intensity of islet beta cells, fluorescence intensity of macrophages, and blood flow velocity of zebrafish were detected. The expressions of vascular endothelial growth factor (vegfaa) and angiotensin converting enzyme (ACE) were detected by RT-PCR. The results showed that compared with the model group, Huangdi Anxiao Capsules can significantly reduce the thickness of the blood vessel wall, increase the fluorescence intensity of islet ß cells and macrophages, increase the blood flow velocity in vivo, and decrease the ACE and vegfaa expressions in zebrafish. It is suggested that Huangdi Anxiao Capsules may alleviate zebrafish vascular lesions by regulating the expressions of ACE and vegfaa.

[Determination and pharmacokinetics of main components for Psoralea corylifolia-Myristica fragrants drug pair by using UPLC-MS/MS].

To conduct multiple-reaction monitoring(MRM) quantitative analysis with ultra-high performance liquid chromatography coupled with mass spectrometry method(UPLC-MS/MS), determine the concentrations of psoralen, isopsoralen, bakuchiol and dehydrodiisoeugenol in plasma under positive iron mode with chloramghenicol as internal standard, and investigate the pharmacokinetics process of the main components before and after oral administration of drug pair Psoralea corylifolia -Myristica fragrants. Thirty-six SD rats were randomly divided into three group(A, B, C) and received P. corylifolia extract, P. corylifolia-M. fragrants extract, and M. fragrants extract respectively by intragastric administration. The plasma samples were collected at different time points. In the plasma samples, psoralen, isopsoralen, bakuchiol and dehydrodiisoeugenol showed good linear relationship within concentration rages of 0.098 125 to 39.25, 0.084 37 to 33.75, 0.046 875 to 18.75, and 0.11 to 2.2 mg•L⁻¹ respectively. The precision and stability results showed that the determination method of plasma concentration for such compositions was stable and reliable. The pharmacokinetic parameters obtained by DAS 2.0 showed varying differences before and after compatibility. According to the experimental results, the compatibility of P. corylifolia and M. fragrants can significantly impact the pharmacokinetic process of main components, expand their distribution and accelerate their metabolism and elimination in vivo.

The effects of Qi Teng Xiao Zhuo granules, traditional Chinese medicine, on the expression of genes in chronic glomerulonephritis rats.

BACKGROUND AND AIM: Chronic glomerulonephritis (CGN) is a primary glomerular disease that is related to immune-mediated inflammatory diseases. Qi Teng Xiao Zhuo granules have been proposed as a prescription of traditional Chinese medicine for treatment of CGN, but the comprehensive molecular mechanism underlying this therapeutic effect is not clear to date. The aim of this study was to evaluate and analyze the possible roles and molecular mechanisms of Qi Teng Xiao Zhuo granule-mediated treatment of CGN induced by adriamycin in rats. METHODS: For gene expression analysis, four samples of glomerular tissue from rats in the Qi Teng Xiao Zhuo granule group and four samples each from the adriamycin treated and control groups were hybridized with Agilent Rat 4×44K whole genome microarrays. KEGG and Gene Ontology (GO) analyses and LIMMA, String and Cytoscape software were used to analyze the functional microarray data and screen differentially expressed genes. Hub genes were identified using Pathway Studio software. Real-time PCR was performed to verify the selected genes. RESULTS: Microarray gene expression analysis showed that Pnoc, Cacfd1, Fos, Igll1, Lcn2, and Syk were among the most downregulated genes in the Qi Teng Xiao Zhuo granule group compared with the adriamycin treated group, whereas Cyp2c7, Hsd3b6, Acsm5, and Ugt2b15 were significantly upregulated. Functional analysis demonstrated that metabolism of xenobiotics by cytochrome P450, the B cell receptor signaling pathway, and cytokine-cytokine receptor interaction pathways were significantly downregulated in the Qi Teng Xiao Zhuo granule group and that GO terms related to positive regulation of immune response, immune response-activating signal transduction, cell differentiation, cell cycle, proliferation, and adhesion were significantly affected. Fos and Syk were considered to be potential hub genes. CONCLUSIONS: In the adriamycin-induced CGN rat model, comprehensive molecular mechanisms were involved with complex gene expression alterations containing many altered pathways and GO terms. However, how Qi Teng Xiao Zhuo granules regulate these events warrants further investigation.

Comparison of fresh, dried and stir-frying gingers in decoction with blood stasis syndrome in rats based on a GC-TOF/MS metabolomics approach.

In China, ginger (Zingiberofficinale Rosc.) and its processed products, such as dried ginger and stir-frying ginger are commonly applied in traditional Chinese medicine (TCM). The paper presents the research on the effects of fresh ginger, dried ginger and stir-frying ginger extracts in blood stasis syndrome. First, a blood stasis syndrome rats model was established and then the hemorheological and blood coagulation activities were analyzed. Third, a sensitive, simple, and valid gas chromatography combined with time-of-flight mass spectrometry (GC-TOF/MS) method was established to compare the metabolic fingerprint coupled with multivariate analysis. The total 27 metabolites (16 in serum and 11 in urine) were identified and contributed to the blood stasis progress. These metabolites mainly involve six metabolism pathways in different impact-value. The altered efficacy index and metabolites can be regulated to normal levels by fresh ginger (FG), dried ginger (DG) and stir-frying ginger (SG). FG is the most effective as shown by the efficacy index, similarity analysis and peak intensity. The result presented here shows that metabolomics equipped with efficacy index makes it possible to study the blood stasis syndrome and to compare the effect and metabolites in fresh, dried and stir-frying gingers. The metabolomics approach can be recommended to study the pharmacological effect and mechanism of herbal drugs.

Panax notoginseng saponins ameliorates experimental hepatic fibrosis and hepatic stellate cell proliferation by inhibiting the Jak2/ Stat3 pathways.

OBJECTIVE: To investigate the inhibitory effect of Panax notoginseng saponins (PNS) on liver fibrosis and explore the underlying mechanisms. METHODS: Carbon tetrachloride (CCl4)-treated rats and hepatic stellate cells (HSCs) were used. The effect of PNS on CCl4-induced liver fibrosis was studied with histochemical and biochemical analysis. Transforming growth factor (TGF)-ß1, α-smooth muscle actin (α-SMA), and collagen I mRNA expression were determined by reverse transcripwhile, the protein expression levels of α-SMA, collagen I, phosphorylation-Janus activated kinase signal transducer (p-Jak2)/Jak2, and phosphorylation-activator of transcription (p-Stat)3/Stat3 were determined by immunohistochemistry and/or immunoblotting. RESULTS: PNS treatment significantly improved the liver function of rats as indicated by decreased serum enzymatic activities of alanine aminotransferase and aspartate aminotransferase. Histopathological results indicated that PNS alleviated liver damage and reduced the formation of fibrous septa. Moreover, PNS significantly decreased liver hydroxyproline and significantly attenuated expressions of collagen I, α-SMA, TGF-ß1, p-Jak2 / Jak2, and p-Stat3/Stat3 in the rat liver fibrosis model and HSCs. CONCLUSION: PNS can relieve liver fibrosis by modulating Jak2/Stat3 signaling transduction pathway, which may be one of its mechanisms to suppress hepatic fibrosis.

Paridis Rhizoma Sapoinins attenuates liver fibrosis in rats by regulating the expression of RASAL1/ERK1/2 signal pathway.

ETHNO-PHARMACOLOGICAL RELEVANCE: Paridis Rhizoma is a Chinese medicinal herb that has been used in liver disease treatment for thousands of years. Our previous studies found that Paridis Rhizoma saponins (PRS) are the critical components of Paridis Rhizoma which has good liver protection effect. However, the anti-hepatic fibrosis effect and the mechanism of PRS have seldom been reported. AIM OF THE STUDY: To investigate the potential of PRS in the treatment of experimental liver fibrosis and the underlying mechanism. MATERIALS AND METHODS: The chemical feature fingerprint of PRS was analyzed by UPLC-PDA. A total of 40 Male Sprague-Dawley (SD) rats were randomly divided into the control group, the model group, the PRS high dose group (PRS H) and the PRS low dose group (PRS L) with 10 rats in each group. The model, PRS H and L groups as liver fibrosis models were established with carbon tetrachloride (CCl4) method. PRS H and L groups were adopted PRS (300 and 150mg/kgd-1) treatment since the twelfth week of modeling till the sixteenth week. Pathological changes in hepatic tissue were examined using hematoxylin and eosin (H&E) and MASSON trichrome staining. Immunohistochemical analysis was performed to determine the protein expression of the RASAL1. RT-PCR and western blotting were used to detect the expression of ERK1/2 mRNA and protein. RESULTS: Four saponins in PRS were identified from 19 detected chromatographic peaks on UPLC-PDA by comparing to the standard compounds. PRS can improve the degeneration and necrosis of hepatic tissue, reduce the extent of its fibrous hyperplasia according to H&E and MASSON staining detection. As was detected in PRS H and L groups, PRS down-regulated p-ERK1/2 mRNA and RASAL1 protein, and up-regulated the level of p-ERK1/2 mRNA and RASAL1 protein. CONCLUSION: These results demonstrated that PRS can attenuate CCl4-induced liver fibrosis through the regulation of RAS/ERK1/2 signal pathway.

[Up-regulations of Bax and caspase-3 and down-regulation of Bcl-2 after Xinfeng capsule treatment in adjuvant-induced arthritis rats].

OBJECTIVE: To observe the effects of Xinfeng capsule (XFC) treatment on the expressions of Bcl-2, Bax and caspase-3 in the synovium tissues of the adjuvant-induced arthritis (AA) rats and explore the possible mechanisms. METHODS: Spague-Dawley (SD) rats were randomly divided into normal group, model group, XFC (1.5, 3.0, 6.0 g/kg) groups and 40 mg/kg tripterygium glycosides tablet (TGT) group, with 10 rats in each group. Arthritis rat was induced by intracutaneous innoculation of 0.1 mL Freund's complete adjuvant (FCA) in the left paw. XFC at different doses and TGT, diluted in water, were administrated via gavage, once a day from the 19th day to the 48th day after AA induction. The normal and model rats were orally administered the same volume of saline solution. Then, ankle-joint samples were taken to examine the degree of AA by HE staining. Bcl-2, Bax and caspase-3 mRNA expressions were determined by reverse transcriptase-polymerase chain reaction (RT-PCR). Meanwhile, the protein expression levels of Bcl-2, Bax and caspase-3 were detected by immunohistochemistry and Western blotting. RESULTS: Compared with the model group, XFC treatment improved histopathological changes, such as synovial hyperplaisia, pannus formation, and so on. Furthermore, XFC (3.0 and 6.0 g/kg) groups not only obviously attenuated mRNA and protein expressions of Bcl-2, but also obviously up-regulated the expressions of Bax and caspase-3 both at protein and mRNA levels. CONCLUSION: XFC has a protective effect on AA rats, which might be partly associated with its regulatory role in decreasing the expression of Bcl-2 and promoting the expressions of Bax and caspase-3.

[Simultaneous Determination of Seven Chemical Markers and Preliminary Identification of Chemical Constituents in Daodi Psoraleae Fructus-Myristicae Semen Chinese Drug Pair].

OBJECTIVE: To study the simultaneous determination method of daodi Psoraleae Fructus-Myristicae Semen Chinese drug pair for the seven ingredients, and Psoraleae Fructus-Myristicae Semen Chinese drug pair on the chemical composition of initial ownership and identification. METHODS: UPLC BEH C18 column (2.1 mm x 100 mm, 1.7 µm) was used in the determination. The flow rate was kept at 0.25 mL/min, and 2 µL of standard and sample solution were injected in each run. The mobile phase was consisted of acetonitrile and water using a gradient elution. The UPLC/Q-TOF MS condition: Waters HSS T3 (100 mm x 2.1 mm,1.7 µm); capillary voltage 3.0 kV (positive ion mode) and 2.5 kV (negative ion mode); Mass spectrometric detection was carried out on a Waters Xevo G2 Q/ TOF mass spectrometer equipped with an ESI source operating in both positive and negative ion modes. The parameters of the mass spectrometer under the ESI mode were as follows: ion source temperature 110 °C, cone gas flow 50 L/h, desolvation gas temperature 450 °C, desolvation gas flow 800 L/h. RESULTS: The seven chemical markers in the selected linear range had good linearity. The recoveries were in the range of 95.07%-98.16% and RSDs were between 1.23%-1.97%. CONCLUSION: It is suitable for the quality control and further studies of the herb in vivo of daodi Psoraleae Fructus-Myristicae Semen Chinese drug pair.

[Protective effects and mechanism of congguiyishen capsules on diabetic nephropathy rats].

OBJECTIVE: To study the curative and protective effects of Congguiyishen Capsules on the diabetic nephropathy (DN) model rats. METHODS: Established the DN model rats by intraperitoneal injection of urea bacteria element (Streptozotocin, STZ). The rats were divided into six groups including normal control group, model group, positive control group, high-dosage group, medium-dosage group and low-dosage group. After oral administration for 4 weeks, determined the 24 h urinary protein, Cr, kidney mass/body mass, FBG, Ang II, AT1R, AGTRAP and CTGF in the kidney. Observed the pathological damage of kidney tissue with Masson staining. RESULTS: After treatment, Cr, kidney mass index, 24 h urine protein, FBG and Ang II were decreased signicantly (P < 0.05). And the treatment also alleviated the pathological damage of kidney tissue. CONCLUSION: Congguiyishen Capsules have protective effect for DN model rats. The mechanism may be related to the suppression of inflammatory response and down-regulating the expression of AT1R, AGTRAP and CTGF.

[Optimization of processing technology for xanthii fructus by UPLC fingerprint technique and contents of toxicity ingredient].

The experiment's aim was to optimize the processing technology of Xanthii Fructus which through comparing the difference of UPLC fingerprint and contents of toxicity ingredient in water extract of 16 batches of processed sample. The determination condition of UPLC chromatographic and contents of toxicity ingredient were as follows. UPLC chromatographic: ACQUITY BEH C18 column (2.1 mm x 100 mm, 1.7 microm) eluted with the mobile phases of acetonitrile and 0.1% phosphoric acidwater in gradient mode, the flow rate was 0.25 mL x min(-1) and the detection wavelength was set at 327 nm. Contents of toxicity ingredient: Agilent TC-C18 column (4.6 mm x 250 mm, 5 microm), mobile phase was methanol-0.01 mol x L(-1) sodium dihydrogen phosphate (35: 65), flow rate was 1.0 mL x min(-1), and detection wavelength was 203 nm. The chromatographic fingerprints 16 batches of samples were analyzed in using the similarity evaluation system of chromatographic, fingerprint of traditional Chinese medicine, SPSS16.0 and SIMCA13.0 software, respectively. The similarity degrees of the 16 batches samples were more than 0.97, all the samples were classified into four categories, and the PCA showed that the peak area of chlorogenic acid, 3,5-dicaffeoylquinic acid and caffeic acid were significantly effect index in fingerprint of processed Xanthii Fructus sample. The outcome of determination showed that the toxicity ingredient contents of all samples reduced significantly after processing. This method can be used in optimizing the processing technology of Xanthii Fructus.

[Changes of CD4 + CD25 + Foxp3 + Regulatory T Cells in Different Stages of COPD and Intervention of Qibai Pingfei Capsules].

OBJECTIVE: To observe the changes in proportion of CD4 + CD25 + Foxp3 + regulatory T cells (Treg) in peripheral blood of different stages of CPOD and intervention of Qibai Pingfei Capsules. METHODS: The rats were randomly divided into three groups,including normal group, COPD group and Qibai Pingfei Capsules(2. 88 g/kg)group. At the end of 7,14,21 and 28 days,eight rats were sacrificed in each group. CD4 + CD25 + Foxp3 + Treg cells in peripheral blood were measured by flow cytometry method. RESULTS: Compared with normal group, the percentages of CD4 + % in peripheral blood were not significantly different at the end of 7, 14, 21 and 28 days. However, CD4 + CD25 + % and CD4 + CD25 + Foxp3 +/CD4 + were significantly increased and CD4 + CD25 + Foxp3 + Treg% were significantly decreased at the end of 14,21 and 28 days. Compared with model group, CD4 + CD25 + %, CD4 + CD25 + Foxp3 +/CD4 + were significantly decreased, CD4 + CD25 + Foxp3 + Treg % were significantly increased at different stages of CODP. CONCLUSION: Immune disorders may exist in COPD, and Treg cells may be involved in the process of COPD. Meanwhile,the protective effect in COPD rats of Qibai Pingfei Capsules may be associated with improving the percentage of suppressive CD4 + CD25 + Foxp3 + Tregs.

[Dynamic study on curative effect of Shuganjianpifang against hepatic fibrosis induced by CCl4 in rats].

OBJECTIVE: To observe the dynamic effects of Shuganjianpifang (SGJPF) in preventing rat hepatic fibrosis induced by CCl4. METHODS: The model rat was induced by subcutaneous injection of 50% CCl4 solution in olive oil (0.1 mL/100 g), twice a week for 12 weeks. The treatment groups were given by lavage with SGJPF and Colchicine respectively once a day for 16 weeks. After 4, 8, 12 and 16 weeks, six rats were randomly sacrificed in each group. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), and glutathione (GSH) in serum as well as superoxide dismutase (SOD) and malondialdehyde (MDA) in hapatic tissues were examined by colorimetric methods dynamically. Meanwhile, the histopathologic changes of hepatic tissues were observed by the HE stained method. RESULTS: Compared with normal group, the levels of ALT and AST in serum and MDA in hepatic tissues were significantly elevated, the levels of SOD in hepatic tissues and GSH in serum were significantly decreased in model group after 4, 8, 12 and 16 weeks. Compared with model group, the levels of ALT and AST in serum and MDA in hepatic tissues were significantly reduced, the levels of SOD in hepatic tissues and GSH in serum were increased significantly after 8, 12 and 16 weeks in SGJPF group and Colchicine group. Histopathologic examination results also showed that SGJPF reduced the degree of liver fibrosis after 4, 8, 12 and 16 weeks. CONCLUSION: Shuganjianpifang can obviously inhibit the formation of liver fibrosis at different phases in rats.

[Effect of shuganjianpifang on the expression of BCL-2 and BAX in rats livers with hepatic fibrosis].

OBJECTIVE: To observe the effect of shuganjianpifang on BCL-2, BAX expressions in livers of hepatic fibrosis rats and its mechanism. METHODS: Sixty SD rats were randomly divided into six groups. Rat liver fibrosis was induced by CCl4 twice a week for 12 weeks. Shuganjianpifang was used daily via lavage at 7th week for 6 weeks. The contents of ALT, AST, T-BiL and Alb in serum were measured. Liver samples were taken to examine the degree of liver fibrosis by HE staining. The experessions of BCL-2 and BAX were detected by immunohisto chemistry. The expression of BCL-2, BAX mRNA was detected by RT-PCR technology. RESULTS: As compared with the fibrotic model group, shuganjianpifang significantly reduced histopathological change, such as steatosis, deposition, decreased the contents of ALT, AST and T-BiL, up-regulated the expression of Alb. Meanwhile shuganjianpifang could effectively inhibit the expression of BAX, significantly enhanced the expression BCL-2 in liver fibrosis rats. CONCLUSION: Shuganjianpifang can resist hepatic fibrosis possibly by up-regulating BCL-2 expression and down-regulating BAX expression.