Modulation of the NAMPT/SIRT axis in HeLa cells exposed to adipose-derived mesenchymal stem cell-conditioned medium

Cervical cancer cell survival depends on coordinated metabolic and stress response pathways. Adipose tissue-derived mesenchymal stem/stormal cells (AD-MSCs) release soluble factors and extracellular vesicle cargo that can alter tumor cell behavior, but their effects on the NAMPT/sirtuin axis in cervical cancer cells are not well defined. We therefore examined how AD-MSC-conditioned medium (CM) affects HeLa cell viability, apoptosis, and expression of NAMPT, SIRT1, SIRT2, and SIRT7.

Previously isolated, characterized, and cryopreserved human AD-MSCs were thawed, expanded, and used to prepare CM. HeLa cells were exposed to 10%-100% CM for 48 h, and viability was measured by MTT assay. The 20% and 60% CM conditions were then evaluated by Annexin V/PI flow cytometry and qRT-PCR. NAMPT, SIRT1, SIRT2, and SIRT7 transcripts were quantified relative to GAPDH.

CM exposure reduced HeLa cell viability, with an estimated IC50 at 38.99%. The 20% condition increased the apoptotic cell fraction, whereas the response to 60% CM was not significant. At 20% CM, SIRT2 (p = 0.0036) expression decreased, SIRT7 expression increased (p = 0.0036), SIRT1 (p = 0.6286) and NAMPT(p = 0.3277) expression did not differ significantly from the comparison groups.

Under the conditions tested, AD-MSC-CM reduced HeLa cell viability and promoted apoptosis most clearly at 20% CM. The accompanying decrease in SIRT2 and increase in SIRT7 point to a paracrine response involving metabolic and sirtuin related signaling. Identification of the active CM components and confirmation at the protein, enzyme-activity, and pathway levels are required before a causal mechanism can be assigned.