Inhibition of PKCepsilon improves glucose-stimulated insulin secretion and reduces insulin clearance.

In type 2 diabetes, pancreatic beta cells fail to secrete sufficient insulin to overcome peripheral insulin resistance. Intracellular lipid accumulation contributes to beta cell failure through poorly defined mechanisms. Here we report a role for the lipid-regulated protein kinase C isoform PKCepsilon in beta cell dysfunction. Deletion of PKCepsilon augmented insulin secretion and prevented glucose intolerance in fat-fed mice. Importantly, a PKCepsilon-inhibitory peptide improved insulin availability and glucose tolerance in db/db mice with preexisting diabetes. Functional ablation of PKCepsilon selectively enhanced insulin release ex vivo from diabetic or lipid-pretreated islets and optimized the glucose-regulated lipid partitioning that amplifies the secretory response. Independently, PKCepsilon deletion also augmented insulin availability by reducing both whole-body insulin clearance and insulin uptake by hepatocytes. Our findings implicate PKCepsilon in the etiology of beta cell dysfunction and highlight that enhancement of insulin availability, through separate effects on liver and beta cells, provides a rationale for inhibiting PKCepsilon to treat type 2 diabetes.

Synergistic cytotoxicity between tamoxifen and the plant toxin persin in human breast cancer cells is dependent on Bim expression and mediated by modulation of ceramide metabolism.

Phytochemicals have provided an abundant source of novel therapeutics for the treatment of human cancers. We have previously described a novel plant toxin, persin, derived from avocado leaves, which has unique in vivo actions in the mammary epithelium and Bim-dependent, cytotoxic effects in human breast cancer cells in vitro. Compounds structurally similar to persin, such as the polyunsaturated fatty acid, conjugated linoleic acid, can attenuate steroid hormone receptor signaling and modulate the response of breast cancer cells to antiestrogens. Here, we provide evidence that persin may have similar effects by showing its potent proapoptotic synergy with the antiestrogen 4-hydroxytamoxifen. However, although persin transcriptionally down-regulates estrogen receptor (ER) expression, unlike conjugated linoleic acid, it also shows efficacy in ER-negative breast cancer cells, both alone and in combination with 4-hydroxytamoxifen, whereas normal breast epithelial cells are unaffected, suggesting it may act via a distinct, ER-independent mechanism. These proapoptotic synergistic interactions are associated with increased de novo ceramide synthesis and are dependent on expression of the proapoptotic protein Bim. These data show that persin should be further investigated as a potential novel cancer therapeutic agent because it significantly enhances the sensitivity of breast cancer cells to the cytotoxic effects of tamoxifen, regardless of their ER status, while displaying apparent specificity for the malignant phenotype.

Increased fatty acid desaturation and enhanced expression of stearoyl coenzyme A desaturase protects pancreatic beta-cells from lipoapoptosis.

Increased availability of fatty acids causes cell death and dysfunction in beta-cell lines, isolated islets, and animal models of diabetes. From the MIN6 beta-cell line, we selected two subpools that are resistant to palmitate-induced apoptosis. Protection was not universal because palmitate-resistant cells remained sensitive to cytokine- and streptozotocin-induced apoptosis. Palmitate oxidation and incorporation into cholesterol ester (but not triglycerides) were significantly higher in palmitate-resistant cells than in control cells. Consistent with these findings, transcript profiling revealed increased expression in palmitate-resistant cells of several beta-oxidation genes as well as a 2.8-fold upregulation of stearoyl-CoA desaturase 1 (SCD1). Correspondingly, the oleate-to-palmitate ratio of palmitate-resistant cells was double that of palmitate-pretreated control cells. At least some of this additional oleate in palmitate-resistant cells was incorporated into cholesterol ester stored in the form of large cytosolic lipid bodies. However, blocking cholesterol ester formation did not render palmitate-resistant cells sensitive to palmitate-induced apoptosis. On the other hand, an inhibitor of SCD1, 10,12-conjugated linoleic acid, dose dependently overcame the resistance of palmitate-resistant cells to lipoapoptosis. Our results suggest that desaturation per se is more important in protecting beta-cells from the cytotoxic effects of palmitate than is the nature of neutral lipid storage pool thus generated.

The diverse roles of protein kinase C in pancreatic beta-cell function.

Members of the serine/threonine PKC (protein kinase C) family perform diverse functions in multiple cell types. All members of the family are activated in signalling cascades triggered by occupation of cell surface receptors, but the cPKC (conventional PKC) and nPKC (novel PKC) isoforms are also responsive to fatty acid metabolites. PKC isoforms are involved in various aspects of pancreatic beta-cell function, including cell proliferation, differentiation and death, as well as regulation of secretion in response to glucose and muscarinic receptor agonists. Recently, the nPKC isoform, PKCepsilon, has also been implicated in the loss of insulin secretory responsiveness that underpins the development of Type 2 diabetes.

An in vitro study of the effects of exposure to a GSM signal in two human cell lines: monocytic U937 and neuroblastoma SK-N-SH.

The use of mobile phones is increasing, which also increases the population's exposure to global system of mobile communications (GSM) signals. Questions of safety and possible biological effects are of concern and to date, remain largely unanswered. In order to examine possible biological effects of a GSM-like signal at a cellular level, we exposed two human cell lines (one of neuronal (SK-N-SH) and the other of monocytoid (U937) origin) to a 900 MHz RF signal, pulsed at 217 Hz, producing a specific absorption rate (SAR) of 0.2 W/kg. Putative effects were assessed by comparing radiofrequency-exposed cells to sham-exposed cells using a variety of assay techniques. For the cell line SK-N-SH, effects were specifically assessed by gene microarray, followed by real-time PCR of the genes of interest, Western blot analysis was used to measure heat shock protein levels, and flow cytometry to measure cell cycle distributions and apoptosis. Effects of radiofrequency on the cell line U937 were assessed by cell viability and cell cycle analysis. From our study of these two cell lines, we found no significant difference between sham-exposed versus radiofrequency-exposed cells in any of the assays or conditions examined.