Investigation of M3 muscarinic acetylcholine receptor expression in placentas of smoking women

This study examined M3 muscarinic acetylcholine receptor expression in placentas of women who smoked during pregnancy and explored the affected signaling pathways via network and functional annotation analyses.Placenta samples from 45 control subjects and 45 individuals who smoked during pregnancy were fixed in zinc-formalin and em-bedded in paraffin.Demographic data were recorded.Sections from the paraffin blocks were stained using Hematoxylin-Eosin and subjected to M3 muscarinic acetylcholine receptor immunostaining.Protein interaction networks were constructed with Cytoscape and analyzed with MCODE for module detection, while Enrichr was used for functional annotation.Histopathological analysis revealed significant degeneration in chorionic villi, increased fibrin deposition, a rise in syncytial knots, and vascular alterations, indicating that smoking adversely affects placental structure.In placental components, M3 muscarinic acetylcholine receptor immunoreactivity was mostly absent in the trophoblastic layer, syncytial knots, villous stroma, chorionic capillaries, and fibrin-rich areas.In the control group, moderate M3 muscarinic acetylcholine receptor expression was noted in connective tissue cells, whereas the trop-hoblastic layer and vascular structures displayed little reactivity.Conversely, placental sections from the smoking group showed a pronounced reduction in M3 muscarinic acetylcholine receptor expression, with a negative immunoreaction in key areas.Module and reactome pathway analysis indicated that Module 2, enriched in nicotinic acetylcholine receptor signaling, may mediate smoking-induced placental dysfunction.Additionally, Modules 1 and 3 were linked to GPCR and neurotransmitter pathways, respectively (p<0.05).The diminished M3 muscarinic acetylcholine receptor expression appears to disrupt placental function via altered nicotinic receptor signaling, potentially affecting vascular tone and nutrient exchange.