Regulation of human jejunal transmucosal resistance and MLC phosphorylation by Na+-glucose cotransport

Na+-nutrient cotransport-dependent regulation of paracellular permeability has been demonstrated in rodent intestine and human intestinal epithelial c ell lines. In cell lines this regulation is associated with phosphorylation of myosin II regulatory light chain (MLC). However, the subcellular locali zation of phosphorylated MLC during this regulation has not been studied an d regulation of paracellular permeability and MLC phosphorylation has not b een studied in isolated human intestine. To evaluate these events in human jejunum, isolated mucosa was mounted in Ussing chambers, characterized elec trophysiologically, and then immunostained using anti-phosphorylated MLC an d anti-total MLC antisera. MLC phosphorylation was assessed by calculating the ratio of anti-phosphorylated MLC signal to anti-total MLC signal within defined regions. Transmucosal resistance of mucosae without active Na+-glu cose cotransport was 37 +/- 3% greater than that of mucosae with active Na-glucose cotransport within 15 min. Quantitative double-label immunofluores cence showed that the phosphorylated MLC-to-total MLC ratio increased by 45 +/- 4% within the perijunctional actomyosin ring when Na+-glucose cotransp ort was active. Thus regulation of transmucosal resistance by Na+-glucose c otransport is accompanied by increased MLC phosphorylation within the perij unctional actomyosin ring. These data support the proposed critical role of the perijunctional cytoskeleton in physiological regulation of human small intestinal paracellular permeability.