The integrity of the endothelial barrier is a determinant of the prognosis of lipopolysaccharide (LPS)-induced acute lung injury (ALI). In this study, we investigated whether and how Sirtuin 1 (SIRT1) maintained the vascular integrity during ALI. An experimental model of ALI was established in mice through intratracheal administration of LPS (10 mg/kg). LPS stimulation significantly increased the pulmonary permeability and decreased the expression of SIRT1 and tight junction proteins (TJs), including occludin, claudin-5, tight junction protein 1 and tight junction protein 2. Morphological studies showed that LPS induced obvious lung injury with inflammatory cell infiltration in the interstitial and alveolar space, hemorrhage, edema, and the thickened alveolar wall compared to the control mice. Intratracheal administration of the selective SIRT1 activator SRT1720 (6.25 mg/kg) significantly attenuated LPS-induced lung injury, lung hyper-permeability and increased TJs expression, whereas intratracheal administration of the selective SIRT1 inhibitor EX527 (6.25 mg/kg) aggravated LPS-induced ALI. Similar protective effects of SIRT1 on pulmonary cellular permeability were observed in primary human pulmonary microvascular endothelial cells treated with LPS (2 mg/mL) in vitro. We further demonstrated that the RhoA/ROCK signaling pathway was activated in SIRT1 regulation of tight junction permeability. The RhoA/ROCK inhibitor Y-27632 (10 μM) increased the expression of TJs and reversed LPS- or EX527-induced hyper-permeability. In conclusion, SIRT1 ameliorates LPS-induced lung injury via decreasing endothelial tight junction permeability, possibly via RhoA/ROCK signaling pathway. This finding may contribute to the development of new therapeutic approaches for lung injury.
Acid-sensing ion channels (ASICs) are present in neurons and may contribute to chemoreception. Among six subunits of ASICs, ASIC1 is mainly expressed in the central nervous system. Recently, multiple sites in the brain including the lateral hypothalamus (LH) have been found to be sensitive to extracellular acidification. Since LH contains orexin neurons and innervates the medulla respiratory center, we hypothesize that ASIC1 is expressed on the orexin neuron and contributes to acid-induced increase in respiratory drive. To test this hypothesis, we used double immunofluorescence to determine whether ASIC1 is expressed on orexin neurons in the LH, and assessed integrated phrenic nerve discharge (iPND) in intact rats in response to acidification of the LH. We found that ASIC1 was co-localized with orexinA in the LH. Microinjection of acidified artificial cerebrospinal fluid increased the amplitude of iPND by 70% (pH 7.4 v.s. pH 6.5∶1.05±0.12 v.s. 1.70±0.10, n = 6, P<0.001) and increased the respiratory drive (peak amplitude of iPND/inspiratory time, PA/Ti) by 40% (1.10±0.23 v.s. 1.50±0.38, P<0.05). This stimulatory effect was abolished by blocking ASIC1 with a nonselective inhibitor (amiloride 10 mM), a selective inhibitor (PcTX1, 10 nM) or by damaging orexin neurons in the LH. Current results support our hypothesis that the orexin neuron in the LH can exert an excitation on respiration via ASIC1 during local acidosis. Since central acidification is involved in breathing dysfunction in a variety of pulmonary diseases, understanding its underlying mechanism may improve patient management.
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