In-vitro effects of protease inhibitors on BAX, BCL-2 and apoptosis in two human breast cell lines (with corrigendum)

Abstract

Currently, the treatment of choice of HIV/AIDS in South Africa is the multidrug combination regimen known as HAART (highly active antiretroviral treatment). HAART, which commonly consists of nucleoside or non-nucleoside reverse transcriptase inhibitors and protease inhibitors, has radically decreased mortality and morbidity rates among people living with HIV/AIDS. The emphasis of the original development of the antiretroviral drugs was on clinical effectiveness (reducing mortality). Presently, emphasis has shifted from the initial short- term considerations to the long-term undesirable or harmful effects induced by this treatment regimen. Whether antiretroviral compounds are oncogenic is widely speculated, which led to this investigation into the effects of protease inhibitors on the expression of key apoptotic regulatory genes, BAX and BCL-2, in two human breast cell lines, MCF-7 and MCF-10A by real-time qPCR gene expression and immunofluorescence. The anti-apoptotic effects of the protease inhibitors – LPV/r were also investigated by cell death detection ELISA and acridine orange staining. This study also evaluated the cytotoxicity of the antiretroviral drugs in normal and cancer cell lines of the breast (at clinically relevant concentrations of the drugs and at different time points, 24–96 h), employing the neutral red uptake assay. The drugs and combinations tested did not alter BAX and BCL-2 gene expression and protein expression and localisation in both cell lines. In addition, the protease inhibitors–LPV/r did not inhibit camptothecin-induced apoptosis in both cell lines. We have shown that the protease inhibitors demonstrated varying degrees of cytotoxicity in the breast cells. The resulting DNA damage associated with cytotoxicity is strongly implicated in the processes of tumour initiation.

Lopinavir and Curcumin Directly Alters BAX/BCL2 and VEGF165b mRNA Levels to Suppress Human Squamous Cervical Carcinoma Cell Growth

“…Acridine Orange permeates all cells, making the nuclei appear green. Live cells have a normal green nucleus; early apoptotic cells have bright green nucleus with condensed or fragmented chromatin; late apoptotic cells display condensed and fragmented chromatin and membrane blebbing (Cañete et al, 2001;Adefolaju et al, 2015). After AO staining, culture dishes were inverted and fixed with formaldehyde vapour for 1 min to prevent the photo-damaging effects of continuous excitation on living cells due to the photosensitizing effects of most fluorescent dyes.…”

“…The mitochondrial apoptotic pathway is regulated by pro-apoptotic genes like BAX and anti-apoptotic genes such as BCL2 (Chipuk et al, 2010). As a result, the ratio of pro-apoptotic to antiapoptotic gene-expression regulates the sensitivity of cells to death signals (Bai & Wang, 2014), hence, the upregulation of BCL2 gene expression and/or downregulation of BAX is one of the important hallmarks of cancers (Green & Beere, 1999;Adefolaju et al, 2015). The effects of the combination of lopinavir and curcumin on this important pathway would provide better insight into their anticancer potentials and underlying mechanisms.…”