Regulatory role of nitric oxide in the reduced survival of erythrocytes in visceral leishmaniasis.
BACKGROUND
Nitric oxide (NO) plays a vital role in maintaining the survivability of circulating erythrocytes. Here we have investigated whether NO depletion associated with visceral leishmaniasis (VL) is responsible for the reduced survival of erythrocytes observed during the disease.


METHODS
Infected hamsters were treated with standard anti-leishmanial sodium stibogluconate (SAG) and NO donor isosorbide dinitrate (ISD). Erythrophagocytosis by macrophages was determined by labelling the cells with FITC followed by flow cytometry. Aggregation of band3 was estimated from band3 associated EMA fluorescence. Caspase 3 activity was measured using immunosorbent assay kit. Phosphatidylserine (PS) externalization and cell shrinkage were determined using annexin V. Aminophspholipid translocase and scramblase activities were measured following NBD-PS and NBD-PC internalization, respectively.


RESULTS
Impairment of both synthesis and uptake of NO resulted in decreased bioavailability of this signaling molecule in erythrocytes in VL. NO level was replenished after simultaneous treatment with ISD and SAG. Combination treatment decreased red cell apoptosis in infected animals by deactivating caspase 3 through s-nitrosylation. Drug treatment prevented infection-mediated ATP depletion and altered calcium homeostasis in erythrocytes. Improved metabolic environment effectively amended dysregulation of aminophospholipid translocase and scramblase, which in turn reduced cell shrinkage, and exposure of phosphatidylserine on the cell surface under the diseased condition.


CONCLUSIONS
In this study, we have identified NO depletion to be an important factor in promoting premature hemolysis with the progress of leishmanial infection. The study implicates NO to be a possible target for future drug development towards the promotion of erythrocyte survival in VL.
Chowdhury
Kaustav Dutta
KD
Indian Institute of Chemical Biology, CSIR, Kolkata-700032, India.
Sen
Gargi
G
Biswas
Tuli
T
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000378:metabolism; D000967:Antimony Sodium Gluconate / Q000494:pharmacology; D000981:Antiprotozoal Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D053148:Caspase 3 / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D006224:Cricetinae; D004912:Erythrocytes / Q000378:metabolism* / Q000469:parasitology; D006461:Hemolysis / Q000187:drug effects; D006801:Humans; D007893:Leishmania donovani; D007898:Leishmaniasis, Visceral / Q000188:drug therapy / Q000378:metabolism* / Q000469:parasitology; D008264:Macrophages / Q000378:metabolism / Q000469:parasitology; D008297:Male; D008647:Mesocricetus; D009569:Nitric Oxide / Q000378:metabolism*; D010587:Phagocytosis / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects
D001457:Anion Exchange Protein 1, Erythrocyte; D000981:Antiprotozoal Agents; D009569:Nitric Oxide; D000255:Adenosine Triphosphate; D053148:Caspase 3; D002118:Calcium; D000967:Antimony Sodium Gluconate