Local Production of Estradiol by Growth Plate Chondrocytes and its Gender-Specific Membrane Mediated Effects

摘要

Growth plate chondrocytes from both male and female rats have the nuclear receptors for 17beta-estradiol (E_2). However, cells from female rat costochondral growth plates respond to E_2 differently than cells from male rats. E_2directly affects the fluidity of chondrocyte membranes derived from female, but not male, rats. In addition, E_2 activates PKC in a nongenomic manner in female cells, and chelerylhrine, a specific inhibitor of PKC, inhibits Independent alkaline phosphatase activity in these cells, indicating that PKC is involved in the signal transduction mechanism. ERct and ERbeta have been found in plasma membranes of E_2-sensitive cells, suggesting that E_2 mediates its effects through membrane receptor-mediated mechanisms in addition to nuclear receptor-mediated pathways. This paper reviews studies that examine this hypothesis. Fourth passage resting zone (RC) and growth zone (GC) chondrocytes from female and male rat costochondral cartilage were treated with 10~(-10) to 10~(-7) M E_2 in the presence of inhibitors and activators of enzymes involved in membrane-mediated signal transduction. To examine the role of classical estrogen receptors, cultures were also treated with the estrogen agonist diethylstilbesterol (DES) and the antagonist ICI 182780. To verify that the effects of E_2 on PKC were due to a membrane-mediated response, we examined the effects of E_2 that had been conjugated to bovine serum albumin (E_2-BSA) to prevent its uptake into the ceil. To determine if the membrane receptor might function in an autocrine/paracrine manner, we examined whether costochondral chondrocytes themselves produce E_2 by examining aromatase expression and activity as a function of cell maturation state and gender. For these experiments, RC and GC cells from male rats were also used. E_2 and E2-BSA elicited comparable effects. Both forms of E_2 stimulated PKC, but only in female cells. The phosphatidylinositol-specific phospholipase C (PLC) inhibitor U73122 blocked the increase in PKC activity. Inhibition of PC-PLC, PLD, and PLA_2 had no effect; also, activation of PLA_2 was without effect. ICI 182780 did not block the stimulatory effect of either E_2 or E_2-BSA and DES also did not alter their effects on PKC. The G-protein inhibitor GDPbetaS-inhibited E_2-BSA stimulated PKC in both RC and GC cells. However, the G-protein activator GTPgammaS increased PKC in E_2-BSA treated GC cells only. Aromatase activity was present in male and female cells and was highest in female RC chondrocytes compared to male RC cells and male and femaie GC cells. Female RC cells also produced the highest levels of E2. Moreover, regulation of E_2 production by I a,25-(OH)_2D_3 was gender-dependent and was found in female cells.The results of these studies show that the gender-specific effects of E_2 involve membrane-associated mechanisms that are independent of ERalpha/ERbeta. The rapid nongenomic effect of E_2 and E_2-BSA on PKC is dependent on G-protein coupled PI-PLC. The membrane ERs may function in autocrine/paracrine regulation since RC and GC cells produce E_2 in a gender- and cell maturation-dependent and regulated manner.