PTH and Phospholipase A2 in the ageing process of intestinal cells

GENTILI CLAUDIA; MORELLI SUSANA; RUSSO DE BOLAND ANA

JOURNAL OF CELLULAR BIOCHEMISTRY

Wiley-Liss

Lugar: USA; Año: 2004 vol. 93 p. 312 - 326

0730-2312

In this study we analyzed, for the first time, alterations in phospholipase A2 (PLA2) activity and response toparathyroid hormone (PTH) in rat enterocytes with aging. We found that PTH, rapidly stimulate arachidonic acid (AA)release in rat duodenal cells (þ1- to 2-fold), an effect that is greatly potentiated by aging (þ4-fold). We also found thathormone-inducedAArelease in young animals is Ca2þ-dependent via cPLA2, whileAAreleased byPTHin cells from agedrats is due to the activation of cPLA2 and the Ca2þ-independent PLA2 (iPLA2). In enterocytes from 3 months old rats, PTHinduced, in a time and dose-dependent fashion, the phosphorylation of cPLA2 on serine 505, with a maximun at 10 min(þ7-fold). Basal levels of cPLA2 serine-phosphorylation were higher in old enterocytes, affecting the hormone responsewhich was greatly diminished (þ2-fold at 10 min). cPLA2 phosphorylation impairment in old animals was not related tochanges of cPLA2 protein expression and did not explain the substantial increase on PTH-induced AA release with aging,further suggesting the involvement of a different PLA2 isoform. Intracellular Ca2þ chelation (BAPTA-AM, 5 mM) suppressedthe serine phosphorylation of cPLA2 in both, young and aged rats, demonstrating that intracellular Ca2þ is required for fullactivation of cPLA2 in enterocytes stimulated with PTH. Hormone effect on cPLA2 was suppressed to a great extent by theMAP kinases ERK 1 and ERK2 inhibitor, PD 98059 (20 mM), the cAMP antagonist, Rp-cAMP, and the PKC inhibitorRo31820 both, in young and aged animals. Enterocytes exposure to PTH also resulted in phospho-cPLA2 translocationfrom cytosol to nuclei and membrane fractions, where phospholipase subtrates reside. Hormone-induced enzymetranslocation is also modified by aging where, in contrast to young animals, part of phospho-cPLA2 remained cytosolic.Collectively, these data suggest that PTH activates in duodenal cells, a Ca2þ-dependent cytosolic PLA2 and attendantAA release and that this activation requires prior stimulation of intracellular ERK1/2, PKA, and PKC. cPLA2 is themajor enzyme responsible for AA release in young enterocytes while cPLA2 and the Ca2þ-independent iPLA2, potentiatePTH-induced AA release in aged cells. Impairment of PTH activation of PLA2 isoforms upon aging may result inabnormal hormone regulation of membrane fluidity and permeability and thereby affecting intestinal cell membranefunction. J. Cell. Biochem. 93: 312–326, 2004. 2004 Wiley-Liss, Inc.