The effect of pH on the Ca2+-Mg2+-dependent ATPase of sarcoplasmic reticulum (SR) was investigated with a rapid mixing quench-flow apparatus capable of measuring phosphorylation and dephosphorylation at times as rapid as 4 msec. The rates of formation and decomposition of the phosphorylated intermediate (E approximately P) of the Ca2+-Mg2+-ATPase were studied in the pH range between 7.6 and 6.0. At pH 6.8, the rates of formation of the phosphorylated intermediate of the Ca2+-Mg2+-ATPase of sarcoplasmic reticulum are the same (t1/2 = 10 msec) for cardiac and skeletal sarcoplasmic reticulum preloaded with calcium, but decrease as the pH is lowered. The effect of acid pH (6.0) is more pronounced for cardiac sarcoplasmic reticulum (t 1/2 = 47 msec) than for skeletal sarcoplasmic reticulum (t 1/2 = 17 msec), in agreement with studies showing that acidosis has a more pronounced effect on cardiac muscle than on skeletal muscle. In addition, a decrease in pH results in a decrease in the rate of the E approximately P decomposition step (the slowest step in the SR reaction sequence). The E approximately P decomposition half-lives were observed to be 97 and 77 msec, respectively for cardiac and skeletal SR at pH 6.8. At pH 6.0, the half-lives were increased to 136 and 178 msec for cardiac and skeletal SR, respectively.

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To clarify the involvement of perilipin, a lipid-droplet-surface protein associated with adipocytes and steroidogenic cells, in the differentiation of sebocytes, we investigated the expression of perilipin in sebaceous glands in vivo and in vitro. Perilipin was expressed in sebaceous glands of the hamster auricle in vivo and was localized at the surface of intracellular lipid droplets in differentiated hamster sebocytes in vitro. Western blot analysis showed that perilipin with a molecular weight of approximately 57 kDa, which was identical to that in differentiated mouse 3T3-L1 adipocytes, was detected in cultured sebocytes, indicating that sebaceous glands expressed perilipin A. In addition, the production of perilipin A in cultured sebocytes was transcriptionally augmented by sebocytic-lipogenesis stimulators, insulin, and 5alpha-dihydrotestosterone, whereas it was decreased by a suppressor of sebocytic differentiation, epidermal growth factor. Furthermore, hamster sebocytes were found to express peroxisome proliferation-activating receptor alpha and gamma1, the activation of which by WY14643 and troglitazone, respectively, caused the transcriptional augmentation of perilipin A expression along with an increase in levels of triacylglycerols in lipid droplets in sebocytes. Therefore, these results provide novel evidence that the expression of perilipin A increases on the surface of intracellular lipid droplets augmented along with the differentiation of hamster sebocytes.

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