Apoptosis-like cell death in Leishmania donovani treated with KalsomeTM10, a new liposomal amphotericin B.
OBJECTIVE
The present study aimed to elucidate the cell death mechanism in Leishmania donovani upon treatment with KalsomeTM10, a new liposomal amphotericin B.


RESULTS
We studied morphological alterations in promastigotes through phase contrast and scanning electron microscopy. Phosphatidylserine (PS) exposure, loss of mitochondrial membrane potential and disruption of mitochondrial integrity was determined by flow cytometry using annexinV-FITC, JC-1 and mitotraker, respectively. For analysing oxidative stress, generation of H2O2 (bioluminescence kit) and mitochondrial superoxide O2- (mitosox) were measured. DNA fragmentation was evaluated using terminal deoxyribonucleotidyl transferase mediated dUTP nick-end labelling (TUNEL) and DNA laddering assay. We found that KalsomeTM10 is more effective then Ambisome against the promastigote as well as intracellular amastigote forms. The mechanistic study showed that KalsomeTM10 induced several morphological alterations in promastigotes typical of apoptosis. KalsomeTM10 treatment showed a dose- and time-dependent exposure of PS in promastigotes. Further, study on mitochondrial pathway revealed loss of mitochondrial membrane potential as well as disruption in mitochondrial integrity with depletion of intracellular pool of ATP. KalsomeTM10 treated promastigotes showed increased ROS production, diminished GSH levels and increased caspase-like activity. DNA fragmentation and cell cycle arrest was observed in KalsomeTM10 treated promastigotes. Apoptotic DNA fragmentation was also observed in KalsomeTM10 treated intracellular amastigotes. KalsomeTM10 induced generation of ROS and nitric oxide leads to the killing of the intracellular parasites. Moreover, endocytosis is indispensable for KalsomeTM10 mediated anti-leishmanial effect in host macrophage.


CONCLUSIONS
KalsomeTM10 induces apoptotic-like cell death in L. donovani parasites to exhibit its anti-leishmanial function.
Infectious Diseases and Immunology Division, Indian Institute of Chemical Biology, Jadavpur, Kolkata, West Bengal, India.
Md
M
Shadab
Infectious Diseases and Immunology Division, Indian Institute of Chemical Biology, Jadavpur, Kolkata, West Bengal, India.
Baijayanti
B
Jha
Infectious Diseases and Immunology Division, Indian Institute of Chemical Biology, Jadavpur, Kolkata, West Bengal, India.
Mohammad
M
Asad
Infectious Diseases and Immunology Division, Indian Institute of Chemical Biology, Jadavpur, Kolkata, West Bengal, India.
Makaraju
M
Deepthi
Infectious Diseases and Immunology Division, Indian Institute of Chemical Biology, Jadavpur, Kolkata, West Bengal, India.
Mohd
M
Kamran
Infectious Diseases and Immunology Division, Indian Institute of Chemical Biology, Jadavpur, Kolkata, West Bengal, India.
Nahid
N
Ali
D000255:Adenosine Triphosphate / Q000378:metabolism; D000666:Amphotericin B / Q000494:pharmacology*; D000818:Animals; D000981:Antiprotozoal Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D020169:Caspases / Q000378:metabolism; D059447:Cell Cycle Checkpoints / Q000187:drug effects; D002462:Cell Membrane / Q000378:metabolism; D002470:Cell Survival; D053938:DNA Fragmentation / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004705:Endocytosis; D005978:Glutathione / Q000378:metabolism; D007893:Leishmania donovani / Q000187:drug effects* / Q000502:physiology*; D015227:Lipid Peroxidation / Q000187:drug effects; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000469:parasitology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism; D009569:Nitric Oxide; D010084:Oxidation-Reduction; D010718:Phosphatidylserines / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism
D000981:Antiprotozoal Agents; D010718:Phosphatidylserines; D017382:Reactive Oxygen Species; C068538:liposomal amphotericin B; D009569:Nitric Oxide; D000666:Amphotericin B; D000255:Adenosine Triphosphate; D020169:Caspases; D005978:Glutathione