Characterization of the Major Phosphoinositide-specific Phospholipase C of Human Amnion

Abstract

Most of the phosphoinositide-specific phospholipase C activity in human amnion at term was found to be attributable to a single isoform (Mr 85,000). Phospholipase C purified from amnion catalyzed the calcium-dependent hydrolysis of both phosphatidylinositol and phosphatidylinositol 4,5-bisphosphate. The high phospholipase C activity of amnion cells isolated at 38-41 weeks of gestation declined greater than 80% during the initial 2-5 days of culture to values characteristic of amnion tissue in early gestation. Activities of phospholipase A2 and phosphatidylinositol synthase remained essentially unaltered during this period of culture. Loss of phospholipase C activity was apparently due neither to the appearance of an inhibitor nor to the loss of an activator and most likely reflected a decrease in the amount of enzyme in amnion cells. Basal production of prostaglandin E2 (PGE2) by amnion cells also declined greatly during the period of loss of phospholipase C activity. Involvement of phospholipase C in the regulation of amnion prostaglandin production was also supported by the finding that the phospholipase C inhibitor, U-73122, potently inhibited amnion cell PGE2 production. In contrast, vasopressin, which appears to stimulate prostaglandin production in amnion cells by a phospholipase C-dependent mechanism, was equipotent in stimulating PGE2 production by amnion cells on Day 2 and Day 5 of culture, even though phospholipase C activity had declined by more than 75%. Furthermore, epidermal growth factor stimulation of PGE2 production by amnion cells appeared to be largely attributable to an increase in prostaglandin H synthase activity and did not involve an increase in phospholipase C activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Intrauterine Tissue Prostaglandin Synthesis: Regulatory Mechanisms

“…DiRenzo and colleagues 41 measured the specific activity of PLC in amnion and decidua obtained after elective caesarean section and spontaneous labor at term, and found no effect of labor on enzyme activity. Bala and associates 77 confirmed the observation that the concentration of PLC in term human amnion is characteristically high; however, they found that the increased levels of the enzyme were not maintained when amnion tissue or dispersed amnion cells were cultured for 2-10 days. The loss of PLC was selec tive, because it was not accompanied by losses in the activity of other enzymes of lipid metabolism such as PLA 2 and phosphatidylinositol synthase, and was not prevented by the inclusion of serum in the cul ture medium.…”

“…This suggests that PLC activation per se does not necessarily lead to increased PGE 2 output in amnion cells. Furthermore, Bala and coworkers 77 observed that the spontaneous loss of PLC activity did not diminish the PGE 2 response of cultured amnion cells to vasopressin. A possible interpreta tion of this result is that PLC-δ, the isoform appar ently expressed in the amnion at high levels, is not involved in the agonist-induced changes of ,PLC activity and PGE 2 synthesis.…”

“…Human decidua, chorion, and amnion especially, contain high levels of PLC activity. 41,77 The inositol phospholipid-specific PLC detected in gestational tissues hydrolyzes phosphatidylinositol as well as phosphatidylinositol-4,5-bisphosphate. 16,22 The presence of Ca + + is mandatory for the enzyme reaction to take place.…”

“…Bala and colleagues 77 purified the PLC of human amnion about 1000-fold and established that a single, 85-kDa form of the enzyme was responsible for most of the activity in the crude extracts. Based on its molecular weight and enzymatic properties, amnion PLC appears to be related to the Mr 85 000 isoform of PLC (PLC-8) found in bovine brain 74,75 and rat brain.…”

Nutrients Induce an Increase in Inositol 1,4,5‐Trisphosphate in Soybean Cells: Implication for the Involvement of Phosphoinositide‐Specific Phospholipase C in DNA Synthesis

“…We have shown that replenishment of nutrients can activate PI-PLC over an extended period of time. We have used U-73122, a compound that has been extensively used in studying possible functions of PI-PLC in mammals (for example, Bala et al, 1990 [1] ; Bleasdale et al, 1990 [4] ; Hirose et al, 1999 [19] ; Powis et al, 1991 [41] ; Smith et al, 1990 b [50] ) and plants (Knight et al, 1997 [24] ; Koch et al, 1998 [25] ; Pingret et al, 1998 [40] ; StaxØn et al, 1999 [52] ). Its near-identical analogue U-73343 does not inhibit PI-PLC.…”