Abstract: FR-PO621
TRAP Analysis of Podocyte Gene Expression in Diabetic Nephropathy
Session Information
- Diabetes Mellitus and Obesity: Basic - Experimental - II
November 03, 2017 | Location: Hall H, Morial Convention Center
Abstract Time: 10:00 AM - 10:00 AM
Category: Diabetes
- 501 Diabetes Mellitus and Obesity: Basic - Experimental
Authors
- Wang, Yinqiu, Vanderbilt University Medical Center, Nashville, Tennessee, United States
- Zhang, Ming-Zhi, Vanderbilt University Medical Center, Nashville, Tennessee, United States
- Vickers, Kasey C., Vanderbilt University Medical Center, Nashville, Tennessee, United States
- Harris, Raymond C., Vanderbilt University Medical Center, Nashville, Tennessee, United States
Background
Diabetic nephropathy (DN) is the leading cause of end-stage renal disease, and podocyte injury plays a critical role in its development. To better understand the role of podocytes in DN, we have utilized Translating Ribosome Affinity Purification (TRAP) to analyze mRNA translation in podocytes isolated from kidneys of mice with early onset of diabetes.
Methods
TRAP allows for the isolation and quantification of cell-specific active mRNA translation. We utilized a podocin-Cre transgene for podocyte-specific activation of a TRAP allele (Rosa26fsTRAP). Translated mRNAs were quantified by high-throughput rRNA-depleted total RNA sequencing. To determine responses to early diabetes, mice were made diabetic with streptozotocin and mRNA was isolated 3 weeks after onset of hyperglycemia. To date, 9 diabetics (5 female, 4 male) and 9 controls (3 female, 6 male) have been studied.
Results
With TRAP isolation, mRNA of podocin and nephrin, two podocyte markers, were enriched 50 to 70 -fold. Overall, 1255 genes were upregulated >1.5 fold and 406 genes were downregulated >1.5 fold. However, there were marked sex differences in male vs. female: Up: 1292 vs. 413; Down: 580 vs. 350; 100 genes were up and 40 genes were down in both sexes. Unlike females, in males, predominant clusters of upregulated genes were related to cell cycle, cell division and apoptosis. In both sexes, there were gene clusters related to inflammation and alterations in cell metabolism. Of interest, both sexes had increased gene clusters related to cell responses to interferons.
Conclusion
Analysis of translated mRNA from podocytes after early onset of a model of type I diabetes indicates significant differences between sexes, with evidence of more dedifferentiation and injury in males. Our ongoing studies in mice with more established DN and in models of type II diabetes should provide further insight into podocyte injury and gender responses.
Funding
- NIDDK Support