Involvement of Synthesis and Degradation Pathways of Collagen Type IV in Human Glomerulosclerosis: Molecular Analysis by in situ Reverse Transcription and Competitive Polymerase Chain Reaction

2015 ◽  
pp. 12-16 ◽  
Author(s):  
Ciro Esposito ◽  
Aneeta Patel ◽  
Zhi Hong Liu ◽  
Gary E. Striker ◽  
Liliane J. Striker
Keyword(s):  
Polymerase Chain Reaction ◽  
Molecular Analysis ◽  
Reverse Transcription ◽  
Collagen Type Iv ◽  
Degradation Pathways ◽  
Competitive Polymerase Chain Reaction ◽  
Type Iv ◽  
Polymerase Chain ◽  
Synthesis And Degradation

scholarly journals The contribution of increased collagen synthesis to human glomerulosclerosis: a quantitative analysis of alpha 2IV collagen mRNA expression by competitive polymerase chain reaction.

1992 ◽  
Vol 176 (6) ◽  
pp. 1571-1576 ◽  
Author(s):  
E P Peten ◽  
L J Striker ◽  
M A Carome ◽  
S J Elliott ◽  
C W Yang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Collagen Synthesis ◽  
Type Iv Collagen ◽  
Chain Reaction ◽  
Collagen Mrna ◽  
Type Iv ◽  
Polymerase Chain

We previously reported that one of the main components of the sclerotic material in human glomerular diseases was type IV collagen. In this study we examined the contribution of increased synthesis to this process at the gene expression level. Sufficient material has not been available to study type IV collagen synthesis by normal or sclerotic glomeruli in humans. We took advantage of the availability of nephrectomy specimens from patients with renal carcinoma, and of the observation that approximately 50% of these patients develop varying degrees of glomerulosclerosis. We microdissected glomeruli from 10 patients and analyzed them using in situ reverse transcription coupled with polymerase chain reaction (PCR) analyses (in situ RT-PCR). alpha 2IV collagen mRNA, after reverse transcription into cDNA, was detected in all patients and appeared to be increased in those with glomerulosclerosis (n = 5). A competitive PCR assay was developed to quantitate this change. There was an average 3.7-fold increase in glomerular type IV collagen cDNA in patients with significant sclerosis. This change was not due to an increased number of glomerular cells. Thus, glomerulosclerosis in humans is associated with an elevation of glomerular type IV collagen gene expression, suggesting that increased synthesis of type IV collagen may represent one component of this process.

A Reverse Transcription-Quantitative Competitive Polymerase Chain Reaction (RT-qcPCR) Technique to Measure Cytokine Gene Expression in Domestic Mammals

1996 ◽  
Vol 33 (2) ◽  
pp. 242-248 ◽  
Author(s):  
J. B. Rottman ◽  
W. A. F. Tompkins ◽  
M. B. Tompkins
Keyword(s):  
Polymerase Chain Reaction ◽  
Reverse Transcription ◽  
Human Disease ◽  
Interleukin 2 ◽  
Basic Research ◽  
Experimental Models ◽  
Cytokine Gene Expression ◽  
Chain Reaction ◽  
Competitive Polymerase Chain Reaction ◽  
Polymerase Chain

Inbred strains of rats and mice have long been used to study basic mechanisms of human disease. Our knowledge of the rodent and human immune systems has increased in recent years, largely because of the availability of reagents and techniques specific for these species. In contrast, outbred animals, including domestic companion and food animals, have not been used routinely as experimental models for human disease, largely because reagents and assays necessary for basic research in immunology and physiology have not been available. Here, using consensus cytokine nucleic acid sequences, we adapt a previously described reverse transcription-quantitative competitive polymerase chain reaction technique to measure interleukin 2 (IL2), IL4, IL6, IL10, IL12, interferon γ, tumor necrosis factor α, and glyceraldehyde-3-phosphate dehydrogenase mRNA expression in the cow, cat, dog, horse, and pig. We demonstrate that the assay is sensitive, accurate, and reproducible.

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