Anti-proliferative activity of bioactive sphingolipids from Leishmania donovani against human cervical cancer cells (HeLa)

Leishmanial total membrane lipids and sphingolipid(s) play an immunomodulatory role in the synovial fluid mononuclear cells of patients with rheumatoid arthritis. Additionally, they exhibit an anti-proliferative effect on various malignant cells, such as B16F10, as well as several acute myeloid leukaemia (AML) cells, including those referenced in K562, U937, and HL-60. Based on our previous experiment, we here represent the anti-proliferative nature of Leishmanial Sphingolipid(s) on Human cervical malignant cell HeLa. Sphingolipid fractions were isolated from attenuated Leishmania donovani by the Bligh and Dyer method, then separated into three fractions by Thin layer Chromatographic method (TLC). Three fractions were named LSL–1, LSL–2, and LSL–3. HeLa cells were treated with three fractions separately. Data were collected following the treatment with sphingolipids, applying a dose- and time-dependent approach. Trypan blue exclusion and MTT assay were performed to study the antiproliferative pattern of HeLa cells. It has been observed that LSL–1 demonstrated greater bioactivity and showed potential for anti-proliferation. We have analysed the morphological characteristics and DNA breakdown of cells after treatment with LSL–1 (60 µg/ml) using phase contrast microscopy and fluorescence microscopy. Sphingolipid (LSL–1) treated cells exhibited morphological changes, including cell shrinkage and membrane blebbing due to the externalisation of phosphatidylserine in the outer leaflet of the membrane. DNA fragmentation was verified by colorimetric assay. Analysis of morphological changes using phage contrast and fluorescence microscopy has shown that an apoptotic body-like structure is formed due to cell shrinkage and the externalisation of phosphatidylserine from the cell membrane. Additionally, DNA fragmentation was observed through Acridine orange/Ethidium Bromide (AO/EtBr) staining. The ELISA assay technique demonstrated histones containing fragmented DNA in a dose-dependent manner. Preliminary investigations have found that Leishmanial sphingolipids possess potent anti-proliferative activity against human cervical cancer cells.