Alleviation of acute pancreatitis-associated lung injury by inhibiting the p38 mitogen-activated protein kinase pathway in pulmonary microvascular endothelial cells

BACKGROUND Previous reports have suggested that the p38 mitogen-activated protein kinase signaling pathway is involved in the development of severe acute pancreatitis (SAP)-related acute lung injury (ALI). Inhibition of p38 by SB203580 blocked the inflammatory responses in SAP-ALI. However, the precise mechanism associated with p38 is unclear, particularly in pulmonary microvascular endothelial cell (PMVEC) injury. AIM To determine its role in the tumor necrosis factor-alpha (TNF-alpha)-induced inflammation and apoptosis of PMVECs in vitro. We then conducted in vivo experiments to confirm the effect of SB203580-mediated p38 inhibition on SAP-ALI. METHODS In vitro, PMVEC were transfected with mitogen-activated protein kinase kinase 6 (Glu), which constitutively activates p38, and then stimulated with TNF-alpha. Flow cytometry and western blotting were performed to detect the cell apoptosis and inflammatory cytokine levels, respectively. In vivo, SAP-ALI was induced by 5% sodium taurocholate and three different doses of SB203580 (2.5, 5.0 or 10.0 mg/kg) were intraperitoneally injected prior to SAP induction. SAP-ALI was assessed by performing pulmonary histopathology assays, measuring myeloperoxidase activity, conducting arterial blood gas analyses and measuring TNF-alpha, interleukin (IL)-1 beta and IL-6 levels. Lung microvascular permeability was measured by determining bronchoalveolar lavage fluid protein concentration, Evans blue extravasation and ultrastructural changes in PMVECs. The apoptotic death of pulmonary cells was confirmed by performing a terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling analysis and examining the Bcl2, Bax, Bim and cle-caspase3 levels. The proteins levels of P-p38, NF kappa B, I kappa B, P-signal transducer and activator of transcription-3, nuclear factor erythroid 2-related factor 2, HO-1 and Myd88 were detected in the lungs to further evaluate the potential mechanism underlying the protective effect of SB203580. RESULTS In vitro, mitogen-activated protein kinase (Glu) transfection resulted in higher apoptotic rates and cytokine (IL-1 beta and IL-6) levels in TNF-alpha-treated PMVECs. In vivo, SB2035080 attenuated lung histopathological injury, decreased inflammatory activity (TNF-alpha, IL-1 beta, IL-6 and myeloperoxidase) and preserved pulmonary function. Furthermore, SB203580 significantly reversed changes in the bronchoalveolar lavage fluid protein concentration, Evans blue accumulation, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive cell numbers, apoptosis-related proteins (cle-caspase3, Bim and Bax) and endothelial microstructure. Moreover, SB203580 significantly reduced the pulmonary P-p38, NF kappa B, P-signal transducer and activator of transcription-3 and Myd88 levels but increased the I kappa B and HO-1 levels. CONCLUSION p38 inhibition may protect against SAP-ALI by alleviating inflammation and the apoptotic death of PMVECs.

Automated summary

This study aimed to investigate the protective effect of p38 inhibition against SAP-ALI. In vitro experiments demonstrated that p38 activation increased apoptotic rates and cytokine levels in PMVECs. In vivo experiments showed that SB203580 could alleviate lung histopathological injury, reduce inflammatory activity, and preserve pulmonary function.