The mechanism for differential effect of nelfinavir and indinavir on collagen metabolism in human skin fibroblasts.

The mechanism for differential effects of human immune deficiency virus protease inhibitors (HIVPIs), nelfinavir (NEL) and indinavir (IND) on collagen metabolism disturbances was studied in human skin fibroblasts. It has been considered that HIVPIs-dependent deregulation of collagen biosynthesis involves prolidase (an enzyme providing proline for collagen biosynthesis), glutamine (Gln) (a substrate for proline biosynthesis), nuclear factor-κB (NF-κB) (a transcription factor that inhibit expression of type I collagen genes), ß1 integrin receptor and Akt signalling. It was found that NEL impaired collagen biosynthesis and the process was more pronounced in the presence of Gln, while IND stimulated collagen biosynthesis. NEL-dependent inhibition of collagen biosynthesis was accompanied by massive intracellular accumulation of type I collagen, while IND slightly induced this process. This effect of NEL was reversed by ascorbic acid but not N-acetylcysteine. The mechanism for the NEL-dependent defect in collagen metabolism was found at the level of prolidase activity, ß1 integrin signalling and NF-κB. NEL inhibited expression of ß1 integrin receptor, Akt and ERK1/2 and increased expression of p65 NF-κB. However, inhibitors of p65 NF-κB did not prevent NEL-dependent inhibition of collagen biosynthesis suggesting that this transcription factor is not involved in studied mechanism. Using PI3K inhibitor wortmannin that prevent phosphorylation of Akt revealed that NEL-dependent inhibition of Akt results in inhibition of collagen biosynthesis. The data suggest that differential effect of NEL and IND on collagen metabolism involves NEL-dependent down-regulation of Akt signalling and proline availability for collagen biosynthesis.

Acetylenic derivative of betulin induces apoptosis in endometrial adenocarcinoma cell line.

Since betulin (Bet) and its acetylenic derivative, 28-O-propynoylbetulin (proBet) were shown to induce apoptosis in several cancer cell lines, we studied the mechanism of this process in human endometrial adenocarcinoma cells (EA). Previous studies suggested that this group of compounds affect prolidase activity (proline releasing enzyme from imidodipeptides) and collagen biosynthesis (proline utilizing process) providing substrate (proline) for proline oxidase (POX) dependent apoptosis. Here we provide evidence that Bet and proBet exhibit prolidase-inducing activity in EA cell line. However, in contrast to Bet, proBet inhibited collagen biosynthesis, increased intracellular proline concentration and induced apoptosis in EA cells, as detected by caspase-3, and -9 expressions and annexin V staining. Although POX expression was not affected by both compounds, the process of apoptosis was accompanied by increase in cytoplasmic level of proline. The mechanism for proBet-induced prolidase activity was found at the level of ß1 integrin signaling. The inhibition of collagen biosynthesis was due to up-regulation of NF-κB p65, an inhibitor of collagen type I gene transcription. Although Bet and proBet induced expression of pro-apoptotic p53 in EA cells, the effect of proBet on the processes was much stronger. In contrast to proBet, Bet strongly induced expression of pro-survival factors, HIF-1α and VEGF. The data suggest that massive production of proline by proBet-dependent activation of prolidase and inhibition of proline utilization for collagen biosynthesis may represent mechanism for POX-dependent apoptosis in EA cells.