Abstract
Purpose:
Diabetic retinopathy (DR) is a vasodegenerative condition with apoptosis of endothelial cells and pericytes resulting in widespread areas of ischemia. Contrary to the belief that duration of diabetes is one of the strongest predictors for development of DR, we identified a unique cohort of patients who in spite of long-standing (>40 yrs) poorly controlled diabetes, remained free of DR. We reasoned that hematopoietic stem cells (HSCs) and vascular progenitors were unique in this group of patients. Microarray analysis of HSCs from this ‘protected cohort’ revealed upregulation of tumor suppressor protein ataxia telangiectasia mutated (ATM). We hypothesized that the loss of ATM in bone marrow HSCs could result in inadequate retinal repair and accelerated development of DR.
Methods:
We developed gender mismatched mouse chimeras in which hematopoietic tissue of wild type mice was replaced with that from ATM-/- mice, WT. ATM-/- chimera. These chimeras were sacrificed 6 months post induction of diabetes with streptozotocin and tissues were harvested for further analysis. Long-term repopulating (LTR) and short-term repopulating (STR)-HSCs were evaluated as lin-Sca1+c-kit+CD34- and lin-Sca1+c-kit+CD34+ cells respectively using flow cytometry. Retinas were processed for trypsin digestion to evaluate the degree of DR and femurs were embedded to quantify the numbers of LTR and STR-HSCs.
Results:
We observed a 50% decrease (p<0.05) in the number of LTR-HSCs in diabetic mice. This decrease in LTR-HSCs was further accelerated in diabetic WT.ATM -/- chimeras (p<0.05). The diabetic WT.ATM -/- also showed a tendency of myeloid bias and a 2-fold increase (p<0.05) in STR-HSCs was observed compared to non-diabetic WT.ATM -/- chimeras. Cell cycle analysis further revealed a profound decrease (p<0.05) in quiescent (G0 phase) LTR-HSCs in control and diabetic WT.ATM-/- chimeras. Also, a substantial portion of STR-HSCs was in either G1 or G2 phase, suggesting a complete decline of quiescent HSCs in. WT.ATM-/-chimeras. Quantification of retinas in diabetic WT.ATM-/- chimeras showed an accelerated increase in the number of acellular capillaries.
Conclusions:
In conclusion, our study suggests the critical role of ATM in protecting bone marrow HSCs from diabetic stress and highlights the importance of maintaining long-term repopulating ability of HSCs to participate in retinal vascular repair in DR.
Keywords: 499 diabetic retinopathy •
721 stem cells •
498 diabetes