| Summary: | 碩士 === 國立宜蘭大學 === 生物技術研究所碩士班 === 97 === Glomerulonephritis (GN) is the major cause of end-stage renal diseases worldwide. They are considered to be consequences of injury primarily to the glomerulus. Because of noninvasiveness and simplicity of specimen collection, human urine is one of the most useful biofluids for proteomics study. Identification of urinary biomarkers may lead to the development of a simple diagnostic test for earlier disease detection, or improvement in prognosis. In the current study, we seek to discover urine biomarkers for glomerular nephritis, including poststreptococcal glomerulonephritis (PSGN), Henoch-Schoenlein purpura nephritis (HSPN) and immunoglobulin A nephropathy (IgAN) using gel-based (2-DE) and gel-free proteomics approaches. Four significantly altered proteins were identified by LC-ESI-Q-TOF-MS/MS as Tamm-Horsfall protein, precursor of Alpha1-acid glycoprotein 1 also called orosomucoid-1, thioredoxin peroxidase B, and chain B of haemoglobin. Western blot and 2-D western analyses were performed to evaluate the sensitivity and specificity of these proteins. Among the four proteins, Alpha1-acid glycoprotein 1 and thioredoxin peroxidase B were considered potential biomarkers for glomerulonephritis.
Furthermore, in order to evaluate the possible mechanism of renal tubular cell injury caused by Chinese herb nephropathy (CHN), aristolochic acid (AA) was applied to treat human renal proximal tubular epithelial cell line, HK-2. AA is a natural nephrotoxant from Aristolochiaceae plants which is widely used in traditional Chinese medicine (TCM). Previous studies have shown that AA is responsible for CHN (also called aristolochic acid nephropathy, AAN), a rapidly progressive interstitial nephropathy. In Asian countries, where TCMs are very popular, the complexity of the pharmacopoeia represents a high risk for AAN because of the frequent substitution of the botanical products by AA-containing herbs. In the present study, we applied gel-based proteomics (2-DE) to reveal the mechanism of AA-induced cytotoxicity for HK-2 cell line. Seven significantly altered proteins were identified by LC-ESI-Q-TOF-MS/MS as myosin, chaperonin, heat shock 70 kDa protein 8, tropomyosin 3, vimentin, beta actin, and calreticulin precursor. These protein are involved in cytoskeleton structure and transdifferentiation, indicating that AA may trigger cytoskeleton reorganization and cell transdifferentiation of HK-2 cell.
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