Abstract: SA-PO134

Fluorescence Lifetime Imaging Microscopy Reveals Gradual Accumulation of Collagens and NADH Lifetime Decrease in Human Kidney Glomeruli with Diabetic Disease Progression

Session Information

Category: Diabetes

  • 503 Diabetes Mellitus and Obesity: Translational

Authors

  • Dobrinskikh, Evgenia, University of Colorado, Denver, Aurora, Colorado, United States
  • Henriksen, Kammi J., The University of Chicago, Chicago, Illinois, United States
  • Levi, Moshe, University of Colorado Denver, Aurora, Colorado, United States
Background


Diabetes mellitus is a heterogeneous group of diseases which is a leading cause of renal cell and tissue damage, fibrosis and eventual renal failure. In view of recent therapies aimed at the pathogenesis of fibrosis generation and progression, sensitive and quantitative techniques for recording fibrosis becomes necessary.

Methods


We have applied Two Photon Excitation (TPE), Second Harmonic Generation (SHG) and Fluorescence Lifetime Imaging Microscopy (FLIM) for label-free imaging of kidney sections from kidney biopsies from human diabetic subjects. We have applied the phasor approach for FLIM analysis, which allows for the visual determination of collagens and other extracellular matrix components localization, and metabolic state of the kidney (free to bound NADH ratio) taking advantage of the specific autofluorescence characteristics of these molecules.

Results

In kidney biopsies obtained from diabetic humans, compared to biopsies obtained from nondiabetic subjects, we have determined that there is a strong SHG signal around the glomerulus and tubulointerstitial areas, which indicates presence of fibrosis. FLIM shows graduate increase of different types of collagens in the glomerulus and tubulointerstitial areas with the diabetes progression, which suggests different organization of extracellular matrix. NADH signal decreases and its lifetime shifts to the shorter lifetime in diabetic’s kidneys that corresponds to different metabolic state of the tissue. FLIM also might determine relative degree of the disease progression based on the ratio of NADH lifetimes in different regions in diabetic compared to nondiabetic control kidneys.

Conclusion


TPE-SHG and FLIM imaging is a sensitive technique for label-free imaging, which can show metabolic state and ECM accumulation with the disease progression of the kidney based on the autofluorescence of the ECM components and NADH.

Funding

  • NIDDK Support