Parathyroid hormone-related protein (PTHrP) inhibits mitochondrial-dependent apoptosis through CK2.

Over the past decade, parathyroid hormone-related protein (PTHrP) has been identified as a key survival factor for cells subjected to apoptotic stimuli. Its anti-apoptotic activity has been attributed to nuclear accumulation of the intact protein, or a synthetic peptide corresponding to its nuclear targeting sequence (NTS), which promotes rapid exit of nutrient deprived cells from the cell cycle. Intracellular PTHrP also inhibited apoptosis by blocking tumor necrosis factor alpha (TNFalpha)-induced apoptosis by blocking signaling from the "death receptor" and preventing damage to the mitochondrial membrane. In both cases, the anti-apoptotic activity was significantly reduced in the presence of a nuclear deficient form of PTHrP with a (88)K/E K/E.K/I(91) mutation in the NTS. The current work was undertaken to determine the mechanism by which nuclear PTHrP blocked mitochondrial-mediated apoptosis. Using sub-cellular fractionation and functional assays we showed that pre-treatment of HEK293 cells with exogenous NTS peptide before inducing apoptosis with TNFalpha was as effective as expression of the full-length protein in inhibiting apoptosis. Inhibition of apoptosis was associated with increased expression of protein kinase casein kinase 2 (CK2) and in sustained CK2 accumulation and activity in the nuclear fraction. In primary chondrogenic cells harvested from the limb buds of PTHrP(+/-) and PTHrP(-/-) embryonic mice, there was a dose-dependent decrease in CK2 expression and activity that correlated with increased susceptibility to apoptosis. Taken together the results indicate that nuclear accumulation of PTHrP effectively inhibits mitochondrial-mediated apoptosis through regulation of the expression, activity, and sub-cellular trafficking of CK2.

Possible mechanisms underlying the mitogenic action of heptachlor in rat hepatocytes.

The worldwide use of the organochlorine pesticide heptachlor has led to widespread contamination in the environment. Like many other organochlorine pesticides, heptachlor is considered to pose a threat to human health. It has been shown that heptachlor is a tumor-promoting agent, but the mechanisms involved still remain unclear. The negative response of heptachlor in in vitro genotoxicity test suggests that this pesticide displays its carcinogenicity through epigenetic pathways. With the growing evidence that proliferation accounts for the tumor-promoting effects of many agents, the purpose of this work was to investigate the mechanisms involved in the mitogenic activity of heptachlor in quiescent rat hepatocytes and to understand the properties of this compound as a tumor promoter in the liver. Heptachlor triggered significant proliferation in quiescent rat hepatocytes. Two mechanisms were delineated to support the mitogenic effect in the hepatocyte: activation of key kinases in signaling pathways and inhibition of apoptosis. Exposure to heptachlor led to activation of protein kinase C mitogenactivated protein kinases. Moreover, these results indicate that like many tumor promoters, heptachlor strongly inhibited TGFbeta-induced apoptosis and cytochrome c release into the cytosol. The levels of the anti-apoptotic protein Bcl-2 were also increased in the presence of heptachlor. In conclusion, these results indicate that heptachlor alters basic cell function by interfering with key cellular signaling pathways.

Vitellogenin in tilapia male fishes exposed to organochlorine pesticides in Ouémé River in Republic of Benin.

In many African countries, the economy largely depends on agriculture. Pesticides are therefore likely to represent an important source of xenoestrogens in contaminated rivers and lagoons. The largely uncontrolled use of diverse pesticides led us to hypothesize that these agents, and particularly organochlorine compounds, may pose a serious problem in the Republic of Benin. To verify our hypothesis, tilapia (Sarotherodon melanotheron) from five sites in the southern part of the main Ouémé River were analyzed. Ouémé River drains the southern region of the country. Vitellogenin (Vtg) was used as an indicator of contaminated sites. This approach has its limitations, because there are a wide variety of man-made chemicals present in the aquatic environment likely to induce Vtg in male fish. Therefore, in this study this approach allows us to define potential contaminated target sites. In order to determine whether the presence of Vtg could be attributable to pesticides, organochlorine pesticides in the flesh of tilapia were also analyzed. Significant amounts of Vtg in fish from contaminated sites were detected, and were correlated with organochlorine pesticide levels in tissue. These results indicate that organochlorine pesticides are present in the Ouémé River and that these compounds can act as endocrine modulators in this ecosystem. Eating fish from contaminated rivers, such as the Ouémé River, may contribute to the accumulation of high concentrations of these pesticides in the body, leading to exposure to their negative effects.

Assessing the estrogenic potential of organochlorine pesticides in primary cultures of male rainbow trout (Oncorhynchus mykiss) hepatocytes using vitellogenin as a biomarker.

Many organochlorine pesticides are suspected of impairing natural hormonal function in organisms by mimicking endogenous estrogen. The aim of this study was to assess the estrogenic activity of the organochlorine pesticides o,p'-DDT, dieldrin, aldrin, heptachlor, mirex and DDT in rainbow trout hepatocyte cultures using vitellogenin (Vtg) as the biomarker. A wide range of pesticide concentrations (0.0001-100 microM) was evaluated. Among the pesticides tested, o,p'-DDT was the most potent inducer of Vtg. The lower potency of technical grade DDT relative to o,p'-DDT could be explained by the fact that this pesticide is a mixture of two different pesticides (18% o,p'-DDT and 77% p,p'-DDT). This suggests that o,p'-DDT is a stronger inducer of Vtg than p,p'-DDT. A simple hypothesis could be that pesticides mixed together competed for the same receptor to favor the formation of a complex with reduced activity towards EREs. If these compounds are classified according to the level of Vtg secreted, we observed the following decreasing order: 17beta-estradiol (E(2))>o,p'-DDT>dieldrin>aldrin>DDT. Non-toxic levels of these compounds competed with E(2) for binding to the estrogen receptor. Heptachlor and mirex did not induce Vtg. Since the latter compounds failed to stimulate Vtg production, the possibility that they could interfere with the estrogenic response by inhibiting E(2) action was tested. In the presence of heptachlor, Vtg production triggered by E(2) significantly decreased. The EC50 value for inhibition of ER binding by heptachlor was cytotoxic for hepatocytes in culture, and this could in part explain the lack of Vtg response observed with this compound at the concentrations tested. Our results indicate that organochlorine pesticides can act as positive or negative modulators of estrogenic function in rainbow trout.