RESUMO
Cypermethrin is reported to affect astrocytes in rat brain; however, its mechanism of action is obscure. Here, we observed an increase in apoptosis in the cortical astrocytes upon treatment of rats with cypermethrin. We then characterized the mechanism governing the apoptosis. Because the epidermal growth factor receptor (EGFR) signaling regulates the survival of astrocytes, we investigated the effect of cypermethrin on EGFR activation. The astrocytes exhibited an early and irreversible attenuation in the basal EGFR phosphorylation. Supportively, molecular docking studies revealed considerable homology in the docking mode of cypermethrin and the known EGFR inhibitors, erlotinib and AG1478, to the kinase domain of EGFR. Furthermore, treatment with cypermethrin demonstrated a downregulation in the intracellular and secreted levels of heparin-binding epidermal growth factor (HB-EGF), an EGFR ligand. AG1478 reduced the synthesis of HB-EGF, suggesting the dependence of HB-EGF on EGFR activation. In addition, a neutralizing antibody against HB-EGF diminished the basal EGFR levels, indicating ligand-dependent expression of EGFR. Likewise, cypermethrin caused irreversible suppression in the basal EGFR levels, which induced apoptosis in astrocytes. The apoptosis was prevented by exogenous HB-EGF. These data imply an autocrine/paracrine mode of action of HB-EGF-EGFR in astrocyte survival. Consequently, cypermethrin induced a mitochondria-mediated apoptosis, characterized by rise in Bax/Bcl-2 ratio and cleavage of caspase-9, -3, and -7, and the effect was prevented by HB-EGF. HB-EGF activated the extracellular signal-regulated kinases and AKT pathways that protected against apoptosis. Together, these data demonstrate that cypermethrin induces astrocyte apoptosis by disrupting the autocrine/paracrine mode of HB-EGF-EGFR signaling at two levels, irreversible loss of basal EGFR and downregulation of HB-EGF.
Assuntos
Apoptose/efeitos dos fármacos , Astrócitos/efeitos dos fármacos , Comunicação Autócrina/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Receptores ErbB/efeitos dos fármacos , Comunicação Parácrina/efeitos dos fármacos , Piretrinas/toxicidade , Transdução de Sinais/efeitos dos fármacos , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Astrócitos/metabolismo , Astrócitos/patologia , Células Cultivadas , Córtex Cerebral/metabolismo , Córtex Cerebral/patologia , Relação Dose-Resposta a Droga , Receptores ErbB/química , Receptores ErbB/metabolismo , Fator de Crescimento Semelhante a EGF de Ligação à Heparina , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Ligantes , Masculino , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Modelos Moleculares , Estrutura Molecular , Fosforilação , Conformação Proteica , Inibidores de Proteínas Quinases/farmacologia , Piretrinas/química , Ratos , Ratos Wistar , Relação Estrutura-Atividade , Fatores de TempoRESUMO
Inadequate maternal intake of omega-3-fatty acids (omega3 FAs) causes adverse neurodevelopmental outcome in the progeny; however, their molecular mechanism of action is obscure. Since omega3 FAs are known to inhibit neuronal apoptosis during neuro-degeneration, we investigated their possible contribution in regulating neuronal apoptosis during brain development. Using rat model of hypothyroidism-induced neuronal apoptosis, we provide evidence for anti-apoptotic role of omega3 FAs during cerebellar development. omega3 FAs were supplemented as a mixture of docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) to pregnant and lactating rats, and primary hypothyroidism was induced by administering methimazole. The cerebella from postnatal day 16 (d16) pups were isolated, and studies on apoptosis were conducted. We observed that omega3 FA-supplementation significantly reduced DNA fragmentation and caspase-3 activation in developing cerebellum of hypothyroid pups. The protection provided by omega3 FAs was associated with their ability to prevent increases in the level of pro-apoptotic basal cell lymphoma protein-2 (Bcl-2)-associated X protein (Bax) in the cerebellum during thyroid hormone (TH) deficiency. omega3 FAs increased the levels of anti-apoptotic proteins like Bcl-2 and Bcl-extra large (Bcl-x(L)), known to be repressed in hypothyroidism. omega3 FAs also restored levels of cerebellar phospho (p)-AKT, phospho-extracellular regulated kinase (p-ERK) and phospho-c-Jun N-terminal kinase (p-JNK), which were altered by hypothyroid insults, without interfering with the expression of TH responsive gene, myelin basic protein (mbp). Taken together, these results supplement an insight into the molecular mechanism of action of omega3 FAs in developing brain that involves regulation of apoptotic signaling pathways under stress.