rottlerin and Body-Weight

In mammals, gonadotropins stimulate oocyte maturation via the epidermal growth factor (EGF) network, and the protein kinase C (PKC) signaling pathway mediates this process. Tumor necrosis factor-α converting enzyme (TACE) is an important protein responding to PKC activation. However, the detailed signaling cascade between PKC and TACE in follicle-stimulating hormone (FSH)-induced oocyte maturation in vitro remains unclear. In this study, we found that rottlerin (mallotoxin, MTX), the inhibitor of PKC δ and θ, blocked FSH-induced maturation of mouse cumulus-oocyte complexes (COCs) in vitro. We further clarified the relationship between two molecules downstream of PKC δ and θ and TACE in COCs: nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) and its products, reactive oxygen species (ROS). We proved that the respective inhibitors of NOX, ROS and TACE could block FSH-stimulated oocyte maturation dose-dependently, but these inhibitory effects could be reversed partially by amphiregulin (Areg), an EGF family member. Notably, inhibition of PKC δ and θ prevented FSH-induced translocation of two cytosolic components of NOX, p47phox and p67phox, to the plasma membrane in cumulus cells. Moreover, FSH-induced TACE activity in cumulus cells was decreased markedly by inhibition of NOX and ROS. In conclusion, PKC δ and θ possibly mediate FSH-induced meiotic resumption in mouse COCs via NOX-ROS-TACE signaling pathway.

Topics: Acetophenones; ADAM Proteins; ADAM17 Protein; Amphiregulin; Animals; Benzopyrans; Body Weight; Cells, Cultured; Female; Follicle Stimulating Hormone; Isoenzymes; Meiosis; Mice; Microscopy, Fluorescence; NADH, NADPH Oxidoreductases; NADPH Oxidase 1; Oocytes; Phagocytosis; Protein Kinase C; Protein Kinase C-delta; Protein Kinase C-theta; Reactive Oxygen Species; Signal Transduction

Protein kinase C (PKC)-delta, an upstream regulator of the Akt survival pathway, contributes to cellular dysfunction in the pathogenesis of diabetes. Herein, we examined the role of PKC-delta in neuronal apoptosis through Akt in the retinas of diabetic rats.. We used retinas from 24- and 35-week-old male Otsuka Long-Evans Tokushima fatty (OLETF) diabetic and Long-Evans Tokushima Otsuka (LETO) nondiabetic rats. To assess whether PKC-delta affects Akt signaling and cell death in OLETF rat retinas, we examined 1) PKC-delta activity and apoptosis; 2) protein levels of phosphatidylinositol 3-kinase (PI 3-kinase) p85, heat shock protein 90 (HSP90), and protein phosphatase 2A (PP2A); 3) Akt phosphorylation; and 4) Akt binding to HSP90 or PP2A in LETO and OLETF retinas in the presence or absence of rottlerin, a highly specific PKC-delta inhibitor, or small interfering RNAs (siRNAs) for PKC-delta and HSP90.. In OLETF retinas from 35-week-old rats, ganglion cell death, PKC-delta and PP2A activity, and Akt-PP2A binding were significantly increased and Akt phosphorylation and Akt-HSP90 binding were decreased compared with retinas from 24-week-old OLETF and LETO rats. Rottlerin and PKC-delta siRNA abrogated these effects in OLETF retinas from 35-week-old rats. HSP90 siRNA significantly increased ganglion cell death and Akt-PP2A complexes and markedly decreased HSP90-Akt binding and Akt phosphorylation in LETO retinas from 35-week-old rats compared with those from nontreated LETO rats.. PKC-delta activation contributes to neuro-retinal apoptosis in diabetic rats by inhibiting Akt-mediated signaling pathways.

Topics: Acetophenones; Animals; Apoptosis; Benzopyrans; Body Weight; Diabetes Mellitus, Type 2; HSP90 Heat-Shock Proteins; Immunoblotting; Immunohistochemistry; Immunoprecipitation; In Situ Nick-End Labeling; Male; Neurons; Pentobarbital; Phosphatidylinositol 3-Kinases; Propoxycaine; Protein Binding; Protein Kinase C-delta; Protein Phosphatase 2; Proto-Oncogene Proteins c-akt; Rats; Rats, Inbred OLETF; Retina; Retinal Ganglion Cells; RNA, Small Interfering; Signal Transduction

We investigated the relationship between phosphorylation of alphaB-crystallin (alphaBC) and retinal apoptosis in type 2 diabetes. The retinas of male Otsuka Long-Evans Tokushima fatty (OLETF) rats at 24 and 35 weeks were used as an animal model for type 2 diabetes and sex- and age-matched Long-Evans Tokushima Otsuka (LETO) rats were used as controls. In the retinas of 35-week OLETF rats, the interaction between alphaBC and protein kinase C delta (PKC delta) among the PKC isozymes, alphaBC phosphorylation at Ser45 (S45p-alphaBC), TUNEL-positive apoptotic ganglion cells, several apoptotic signs, and co-localization of S45p-alphaBC and TUNEL significantly increased as compared with other groups while the alphaBC-Bax interaction greatly decreased. These changes were abolished by rottlerin treatment, a highly specific PKC delta inhibitor. These results suggest that PKC delta is involved in regulation of anti-apoptotic function of alphaBC in the retina of type 2 diabetes.

Topics: Acetophenones; Age Factors; alpha-Crystallin B Chain; Animals; Apoptosis; bcl-2-Associated X Protein; Benzopyrans; Blood Glucose; Body Weight; Diabetes Mellitus, Type 2; Disease Models, Animal; Enzyme Inhibitors; Female; Gene Expression Regulation; Immunoprecipitation; In Situ Nick-End Labeling; Male; Protein Kinase C-delta; Rats; Rats, Inbred OLETF; Rats, Long-Evans; Retina; Time Factors