Increased vascular thromboxane generation impairs dilation of skeletal muscle arterioles of obese Zucker rats with reduced oxygen tension

This study determined if altered vascular prostacyclin (PGI(2)) and/or thromboxane A(2) (TxA(2)) production with reduced Po(2) contributes to impaired hypoxic dilation of skeletal muscle resistance arterioles of obese Zucker rats (OZRs) versus lean Zucker rats (LZRs). Mechanical responses were assessed in isolated gracilis muscle arterioles following reductions in Po(2) under control conditions and following pharmacological interventions inhibiting arachidonic acid metabolism and nitric oxide synthase and alleviating elevated vascular oxidant stress. The production of arachidonic acid metabolites was assessed using pooled arteries from OZRs and LZRs in response to reduced Po(2). Hypoxic dilation, endothelium-dependent in both strains, was attenuated in OZRs versus LZRs. Nitric oxide synthase inhibition had no significant impact on hypoxic dilation in either strain. Cyclooxygenase inhibition dramatically reduced hypoxic dilation in LZRs and abolished responses in OZRs. Treatment of arterioles from OZRs with polyethylene glycol-superoxide dismutase improved hypoxic dilation, and this improvement was entirely cyclooxygenase dependent. Vascular PGI(2) production with reduced Po(2) was similar between strains, although TxA(2) production was increased in OZRs, a difference that was attenuated by treatment of vessels from OZRs with polyethylene glycol-superoxide dismutase. Both blockade of PGH(2)/TxA(2) receptors and inhibition of thromboxane synthase increased hypoxic dilation in OZR arterioles. These results suggest that a contributing mechanism underlying impaired hypoxic dilation of skeletal muscle arterioles of OZRs may be an increased vascular production of TxA(2), which competes against the vasodilator influences of PGI(2). These results also suggest that the elevated vascular oxidant stress inherent in metabolic syndrome may contribute to the increased vascular TxA(2) production and may blunt vascular sensitivity to PGI(2).     

Polarized human cholangiocytes release distinct populations of apical and basolateral small extracellular vesicles

Intercellular communication is critical for organismal homeostasis, and defects can contribute to human disease states. Polarized epithelial cells execute distinct signaling agendas via apical and basolateral surfaces to communicate with different cell types. Small extracellular vesicles (sEVs), including exosomes and small microvesicles, represent an understudied form of intercellular communication in polarized cells. Human cholangiocytes, epithelial cells lining bile ducts, were cultured as polarized epithelia in a Transwell system as a model with which to study polarized sEV communication. Characterization of isolated apically and basolaterally released EVs revealed enrichment in sEVs. However, differences in apical and basolateral sEV composition and numbers were observed. Genetic or pharmacological perturbation of cellular machinery involved in the biogenesis of intralumenal vesicles at endosomes (the source of exosomes) revealed general and domain-specific effects on sEV biogenesis/release. Additionally, analyses of signaling revealed distinct profiles of activation depending on sEV population, target cell, and the function of the endosomal sorting complex required for transport (ESCRT)-associated factor ALG-2–interacting protein X (ALIX) within the donor cells. These results support the conclusion that polarized cholangiocytes release distinct sEV pools to mediate communication via their apical and basolateral domains and suggest that defective ESCRT function may contribute to disease states through altered sEV signaling.     

Inhibition of microsomal calcium sequestration causes an impairment of initiation of protein synthesis in perfused rat liver

The present study examined the effect of 2,5-di-(tert-butyl)-hydroquinone (tBuHQ), an inhibitor of liver microsomal calcium sequestration, on initiation of protein synthesis in perfused rat liver. Perfusion of livers with a concentration of tBuHQ previously shown to completely inhibit microsomal calcium sequestration in isolated hepatocytes caused a 50% inhibition of protein synthesis. The inhibition was characterized by an increase in liver content of free ribosomal particles and a decrease in polysomes indicating that peptide-chain initiation was slowed relative to elongation. Furthermore, the inhibition was associated with a 7.5-fold increase in the proportion of the alpha-subunit of eukaryotic initiation factor 2 (eIF-2) present in the phosphorylated form and a reduction in the activity of eukaryotic initiation factor 2B (eIF-2B) to 37% of the control value. The results suggest that protein synthesis in rat liver is regulated directly by changes in intracellular calcium concentration through a mechanism involving modulation of the phosphorylation state of eIF-2 alpha.