Kidney Week

Abstract: FR-PO264

Single-Cell Spatial Metabolomics Reveal That Methylthioadenosine Phosphorylase (MTAP)-Derived Adenine Production Regulates Tubular Mitochondrial Bioenergetics in Type 1 Diabetes (T1D)

Session Information

• Diabetic Kidney Disease: Basic - 1
  October 25, 2024 | Location: Exhibit Hall, Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Diabetic Kidney Disease

• 701 Diabetic Kidney Disease: Basic

Authors

• Maity, Soumya, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States

Soumya Maity, PhD

• Tamayo, Ian M., The University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States

Ian M. Tamayo, MS

• Zhang, Guanshi, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States

Guanshi Zhang, PhD

• Kroma, Alaa, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States

Alaa Kroma

• Lee, Hak Joo, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States

Hak Joo Lee, PhD

• Pyle, Laura, University of Colorado Anschutz Medical Campus School of Medicine, Aurora, Colorado, United States

Laura Pyle, PhD

• Choi, Ye Ji, University of Colorado Anschutz Medical Campus School of Medicine, Aurora, Colorado, United States

Ye Ji Choi, MPH

• Bjornstad, Petter, University of Colorado Anschutz Medical Campus School of Medicine, Aurora, Colorado, United States

Petter Bjornstad, MD

• Sharma, Kumar, The University of Texas Health Science Center at San Antonio, San Antonio, Texas, United States

Kumar Sharma, MD, FASN

Background

Type 1 diabetic patients (T1D) have an estimated 50% risk of developing diabetic kidney disease (DKD) over time. Mitochondrial dysfunction is associated with DKD progression. Our prior study identified endogenous adenine accumulation as a causative factor for DKD and methylthioadenosine phosphorylase (MTAP) as a major adenine-producing enzyme in Type 2 diabetes. In this study, we employed matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) to determine the cell-type-specific MTAP-derived adenine effect on mitochondrial bioenergetics.

Methods

Kidney biopsies from young T1D patients (n=17; 24.4±2.6 year) and healthy controls (HC) (n=7; 23.7±4.05 year) were subjected to MSI to estimate the adenine abundance in situ. Human proximal tubular (HK2) cells and rat podocytes were co-cultured in normal glucose (5.5mM) containing media on the ITO-coated slide. The cell monolayer was fixed with 4% formalin followed by MSI with 10 μm spatial resolution to evaluate cell-type specific adenine production. The pre-MALDI autofluorescence (AF) image was overlaid with the MALDI ion image to determine the metabolomic signature of a single cell. Western blot and mitochondrial oxygen consumption rate (OCR) were performed in MTAP knockdown mouse proximal tubular (MCT) cells to investigate the role of MTAP in adenine-induced renal pathology and mitochondrial bioenergetics.

Results

MSI data revealed that adenine is significantly accumulated in T1D kidneys compared to HC. Single-cell spatial metabolomics revealed substantially higher adenine production in HK2 cells compared to podocytes. Moreover, 24h 25 mM glucose (HG) treatment showed adenine overproduction in HK2 cells compared to normal glucose. MTAP protein level was significantly increased with HG, suggesting MTAP-derived adenine overproduction in diabetic conditions. Knockdown of MTAP with siRNA enhanced OCR and inhibited HG-induced fibronectin expression in MCT cells, indicating intracellular adenine abundance regulates mitochondrial bioenergetics and DKD pathology in proximal tubules.

Conclusion

Our findings demonstrate that MTAP-derived adenine overproduction may be associated with tubular mitochondrial dysfunction in diabetes, and high kidney adenine may be an early driver of kidney disease in T1D.

Funding

• NIDDK Support