Abstract: FR-PO232
APOL1 Preserves Podocyte Differentiation in High Glucose Milieu through Down-Regulation of MicroRNA193a
Session Information
- Apoptosis, Proliferation, Autophagy, Cell Senescence, Cell Transformation
November 03, 2017 | Location: Hall H, Morial Convention Center
Abstract Time: 10:00 AM - 10:00 AM
Category: Cell Biology
- 202 Apoptosis, Proliferation, Autophagy, Cell Senescence, Cell Transformation
Authors
- Mishra, Abheepsa, Feinstein Institute of Medical Research, Northwell Health, MANHASSET, New York, United States
- Ayasolla, Kamesh R., Feinstein Institute for Medical Research, Great Neck, New York, United States
- Lan, Xiqian, Feinstein Institute for Medical Research, Great Neck, New York, United States
- Kumar, Vinod, Fienstine Institute for Medical Research, New York, New York, United States
- Aslam, Rukhsana, Feinstein Institute for medical research, Glenoaks, New York, United States
- Hussain, Ali, Feinstein Institute of Medical Research, New York, New York, United States
- Marashi Shoshtari, Seyedeh Shadafarin, The Feinstein Institute for Medical Research, Manhasset, New York, United States
- Bhooplapur, Manali, Feinstein Institute for medical research, Glenoaks, New York, United States
- Chowdhary, Sheetal, Feinstein Institute of Medical Research, New York, New York, United States
- Malhotra, Ashwani, Feinstein Inst.Med research and NSLIJ, Manhasset, New York, United States
- Singhal, Pravin C., North Shore LIJ Health System, Great Neck, New York, United States
Background
Dibetic podocytopathy is characterized by significant proteinuria an indication of loss of integrity of glomerular filatration barrier. High glucose milieu has been demonstrated to promote dedifferentiation of podocytes (PDs) contributing to the loss of integrity of glomerular filtration barrier; however, the involved mechanism is far from clear. APOL1 is expressed in kidneys of certain primates including humans. APOL1 risk alleles (G1 and G2) have been reported to be podocytotoxic, however, the role of APOL1G0 (wild-type) is far from clear.We hypothesize that APOL1 facilitates preservation of the molecular integrity of podocytes in adverse milieus such as high glucose through down-regulation of microRNA (miR) 193a.
Methods
To evaluate the effect of high glucose milieu, differentiated human podocytes (DIF-PDs, after incubation for 10 days at 37°C) were incubated in media containing different concentrations of glucose (5, 15, 25, 30, and 35 mM) for 48 hours. To evaluate the role of miR193a, DIF-PDs were incubated in media containing normal glucose (5mM, NGM), high glucose (30 mM) with/without miR193a inhibitor (25 nm) for 48 hours. To establish a causal relationship, DIF-PDs were transfected with either control or APOL1/miR193a siRNA followed by incubation in either normal (5 mM, NGM) or high glucose (HGM) media for 48 hours. To confirm a relationship, DIF-PDs were transfected with either vector or APOL1 lentivirus and then incubated in media containing either normal or high glucose for 48 hours. Proteins and RNA were extracted. Protein blots were probed for APOL1, WT1, podocalyxin, nephrin and reprobed for actin. RNAs were assayed for miR193a.
Results
HGM down regulated WT1 and nephrin (2.5 fold) expressions but increased (3-fold) miR193a levels in PDs. HGM down regulated PD expression of APOL1 in a dose-dependent manner. PDs knocked down for APOL1 displayed enhanced (2.2 fold) levels of miR193a, whereas, PDs knocked down for miR193a displayed increased (2.5 fold) expression of APOL1. Over expression of APOL1 in PDs preserved podocyte molecular phenotype in HGM.
Conclusion
High glucose dedifferentiates PDs through down-regulation of APOL1; however, overexpression of APOL1 preserves PDs molecular integrity.
Funding
- NIDDK Support