Year of Award

8-1-2020

Degree Type

Thesis

Degree Name

Master of Philosophy (MPhil)

Department

School of Chinese Medicine, Teaching and Research Division

Principal Supervisor

Yu, Zhiling

Keywords

Glomerulonephritis ; Kidney glomerulus ; Herbs ; Drugs ; Medicine, Chinese

Language

English

Abstract

Yi-Shen-Hua-Shi (YSHS) granule is a Chinese patent drug for treating chronic glomerulonephritis (CGN). It was marketed in 2009. However, up to now, there is no report about the quality and pharmacological activities of this product. In this work,we evaluated the quality and anti-CGN effects of the drug. To evaluate the quality of the granule, a qualitative and quantitative HPLC-DAD analytical method was developed. For qualitative analysis, HPLC fingerprint of ten batches of YSHS granule was established. The fingerprints were analyzed using similarity evaluation, hierarchical cluster analysis (HCA), principal components analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) based on 15 characteristic fingerprint peaks. Similarity values of 10-batche samples were all above 0.960, indicating a stable quality. Minor differences were observed among batches by HCA and PCA. For quantification analysis, contents of six constituents in the granule were simultaneously measured. To establish the chemical profile of the granule, a HPLC-Q-TOF- MS/MS method was developed. A total of 105 peaks were detected using HPLC-Q-TOF-MS/MS in the granule, of which, 99 were tentatively identified as terpenoids, flavonoids, coumarins, alkaloids, phenols and other types of compounds, and 15 were further validated with reference substances. HPLC fingerprint chromatogram establishment, quantification analysis of 6 constituents and compound identification should improve the quality control of YSHS granule. To study the pharmacological activities of the granule, we investigated its anti-CGN effects and TGFβ signaling-related mechanism of action. A CGN rat model was established by injection of cationization-bovine serum albumin (C-BSA) for five weeks. After C-BSA injection, drugs were intragastrically administered to the rats once daily for four weeks. Clinical signs were recorded daily. Urine and serum biochemical parameters were analyzed using respective kits. Protein levels were examined by Western blotting. Pathological changes of renal tissues were evaluated using HE and Masson's trichrome staining. No significant differences in body weights and clinical signs were found among normal, model and drug treatment groups. Proteinuria; albuminuria; increased urine volume; elevated creatinine, urea nitrogen, triglyceride levels and total cholesterol in serum; decreased serum total protein and albumin; as well as renal pathological damages and fibrosis were observed in CGN model rats. YSHS granule ameliorated all the abnormal behavioral and biochemical changes in the model rats. Mechanistic investigations revealed that YSHS granule down-regulated proteins levels of TGFβ1, phospho-Smad2/3 (Thr 8) and Smad4 in rat renal tissues. These findings indicate that the drug has anti-CGN effects in rats, and inhibiting TGFβ signaling contributes to the underlying mechanisms. In summary, our chemical analytical studies will help in improving the quality control of YSHS granule. Our bioactivity and mechanistic studies provide a pharmacological basis for the clinical use of the granule in treating CGN.

Comments

Principal supervisor: Professor Yu Zhiling ; Thesis submitted to the School of Chinese Medicine, Teaching and Research Division

Bibliography

Includes bibliographical references (pages 79-92)



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