Superfund Research Program
PCB Effects on Uterine Muscle
Project Leader: Rita Loch-Caruso (University of Michigan)
Grant Number: P42ES004911
Funding Period: 1995 - 2006
Polychlorinated biphenyls (PCBs) are widespread environmental contaminants. Previous epidemiologic and laboratory studies indicate that some PCBs alter gestation length. Because PCBs accumulate in uterus and placenta during pregnancy, these tissues may be direct targets of PCB action. Parturition requires development of coordinated, forceful, phasic uterine contractions. This study aims to elucidate structure-activity relationships and signal transduction mechanisms for PCB-induced alteration of uterine muscle contraction. The most recent studies described here include investigations of mechanisms by which non-coplanar ortho-substituted PCB congeners induce desynchronization of uterine contractions and modify prostaglandin synthesis in the uterus and amnion.
The non-coplanar ortho-substituted PCB congener 2,2'-cholorobiphenyl (CB) was used as a model PCB that promoted desynchronization of uterine contractions, characterized by irregular contraction frequency and amplitude. Because gap junctions form intercellular channels between the uterine smooth muscle (myometrial) cells and allow the uterus to contract in a coordinated manner with increased force at term, the team investigated inhibition of myometrial gap junctions as a mechanism by which 2,2'-CB induces uterine desynchronization. In myometrial cells in culture, gap junctions were inhibited in a concentration-dependent manner by 2,2'-CB, as measured by Lucifer yellow dye transfer. This inhibition of myometrial gap junctions was accompanied by phosphorylation of MEK/ERK, suggesting activation of MAP kinase, and phosphorylation of the gap junction protein connexin43 at serine255, a MAP kinase site. Phosphorylation at serine368, a protein kinase C site, was unaffected by 2,2'-CB. Pharmacologic inhibition of MEK partially reversed 2,2'-CB-induced inhibition of myometrial gap junctions in cells in culture and also restored contraction force and synchronization of uterine strips exposed to 2,2'-CB in muscle baths. These data suggest that 2,2'-CB inhibition of myometrial gap junctions is mediated by a MAP kinase pathway, and that inhibition of myometrial gap junctions by the MAP kinase mechanism underlies 2,2'-CB-induced desynchronization of uterine contractility.
Because 2,2',4,6-CB was shown by others to significantly activate phospholipase A2 (PLA2) in non-uterine cells, this congener was used as a model PCB in investigations of PCB actions on PLA2 in pregnant rat myometrium. Multiple PLA2 isoforms (sPLA2-IIA, cPLA2, and iPLA2) were identified by immunoblotting in late gestation rat myometrial cells, and were shown to mediate PCB-induced release of arachidonic acid from membrane phospholipids. Arachidonic acid serves as the precursor to eicosanoids that modulate uterine contraction, and cultured rat myometrial cells increased generation of prostaglandins in response to 2,2',4,6-CB.
Immunoblot analysis revealed increased abundance of sPLA2-IIA and a 50 kDa iPLA2-related protein in gestation day (gd) 20 uterus compared with gd10 uterus, whereas uterine cPLA2 expression was unchanged. A significant increase in uterine contraction frequency was observed in gd20 but not gd10 uterine strips treated with 2,2',4,6-CB compared to solvent controls. Pharmacologic inhibition of iPLA2 reversed 2,2',4,6-CB-induced stimulation of contraction frequency in gd20 uterine strips, whereas sPLA2 inhibition had no significant effect. These results suggest that gestational age-dependent sensitivity to the PCB congener 2,2',4,6-CB is related to the ability of 2,2',4,6-CB to activate iPLA2 activity and, potentially, to the increased abundance of the 50 kDa iPLA2-related protein.