Participation of the endoplasmic reticulum protein chaperone thio-oxidoreductase in gonadotropin-releasing hormone receptor expression at the plasma membrane

Chaperone members of the protein disulfide isomerase family can catalyze the thiol-disulfide exchange reaction with pairs of cysteines. There are 14 protein disulfide isomerase family members, but the ability to catalyze a thiol disulfide exchange reaction has not been demonstrated for all of them. Human endoplasmic reticulum protein chaperone thio-oxidoreductase (ERp18) shows partial oxidative activity as a protein disulfide isomerase. The aim of the present study was to evaluate the participation of ERp18 in gonadotropin-releasing hormone receptor (GnRHR) expression at the plasma membrane. Cos-7 cells were cultured, plated, and transfected with 25 ng (unless indicated) wild-type human GnRHR (hGnRHR) or mutant GnRHR (Cys14Ala and Cys200Ala) and pcDNA3.1 without insert (empty vector) or ERp18 cDNA (75 ng/well), pre-loaded for 18 h with 1 µCi myo-[2-3H(N)]-inositol in 0.25 mL DMEM and treated for 2 h with buserelin. We observed a decrease in maximal inositol phosphate (IP) production in response to buserelin in the cells co-transfected with hGnRHR, and a decrease from 20 to 75 ng of ERp18 compared with cells co-transfected with hGnRHR and empty vector. The decrease in maximal IP was proportional to the amount of ERp18 DNA over the range examined. Mutants (Cys14Ala and Cys200Ala) that could not form the Cys14-Cys200 bridge essential for plasma membrane routing of the hGnRHR did not modify maximal IP production when they were co-transfected with ERp18. These results suggest that ERp18 has a reduction role on disulfide bonds in wild-type hGnRHR folding.

Inhibitory effect of spironolactone on the contractions of the rabbit uterine horn and the human myometrial strip.

The spontaneous contractions of the rabbit uterine horns and the human myometrial strips were stimulated by oxytocin and buserelin acetate in isolated preparations. Spironolactone application to these models produced inhibitory effects on the contractions. It is concluded that spironolactone has inhibitory effect on the rabbit uterine horn and the human myometrial strip contractions.

Supressäo hipofisária com altas doses do agonista do hormônio liberador de gonatrofinas / Pituitary suppresion with high dosage of gonadotropin releasing hormone agonist

Neste trabalho, quatro mulheres voluntárias, ovulatórias foram submetidas a administraçäo diária do agonista do hormônio liberador de gonadotrofinas (aGnRH). O aGnRH (Buserelin) foi administrado intranasal na dose de 900 ug/dia durante 12 dias dividio em 3 doses de 300 ug, iniciando-se as 8:00hs. Amostras sanguíneas foram colhidas diariamente, imediatamente antes da primeria dose do Buserelin (8:00hs), sendo que nos dias 1, 4, 8 e 12 foram colhidos 3 amostras adicionais 60, 120 e 180 minutos após a primeira amostra. Avaliaram-se os níveis plasmáticos do hormônio luteinizante (LH), hormônio folículo estimulante (FSH e estradiol (E2). A administraçäo do aGnRH resultou inicialmente numa grande liberaçäo do LH (média de 60 mUI/ml) e do FSH (média de 20 mUI/ml) no 2§ dia, com queda progressiva a partir do 3§ dia, com estabilizaçäo a partir do 7§ dia, em valores em torno de 10 mUI/ml para ambos hormônios. A desensibilizaçäo hipofisária já foi observada a partir do quarto dia do ciclo, visto que houve um aumento do LH, após a administraçäo do aGnRH, nas amostras colhidas aos 180 minutos de apenas 79,3 por cento no quarto dia, contra 1.642 por cento nas amostras do primeiro dia. Para o FSH o incremento do 1§ dia foi de 162 por cento contra 25 por cento do 4§ dia. Os níveis de E2 apresentaram queda de seus valores a partir do 7§ dia, sendo que mostraram-se abaixo de 60pg/ml no 12§ dia, apesar dos níveis de LH e FSH se situarem em torno de 10mUI/ml. Os resultados sugerem que embora mantenha níveis razoáveis circulantes de LH e FSH mesmo no 12§ dia, a partir do 4§ dia, a hipófise já seria refratária ao GnRH endógeno e que os níveis plasmáticos de LH e FSH näo deveriam ser considerados como parâmetros para avaliaçäo do tempo necessário para a supressäo hipofisária quando do uso do aGnRH