SummaryThe intestinal epithelium serves as an essential barrier to the outside world and is maintained by functionally distinct populations of rapidly cycling intestinal stem cells (CBC ISCs) and slowly cycling, reserve ISCs (r-ISCs). Because disruptions in the epithelial barrier can result from pathological activation of the immune system, we sought to investigate the impact of inflammation on ISC behavior during the regenerative response. In a murine model of αCD3 antibody-induced small-intestinal inflammation, r-ISCs proved highly resistant to injury, while CBC ISCs underwent apoptosis. Moreover, r-ISCs were induced to proliferate and functionally contribute to intestinal regeneration. Further analysis revealed that the inflammatory cytokines interferon gamma and tumor necrosis factor alpha led to r-ISC activation in enteroid culture, which could be blocked by the JAK/STAT inhibitor, tofacitinib. These results highlight an important role for r-ISCs in response to acute intestinal inflammation and show that JAK/STAT-1 signaling is required for the r-ISC regenerative response.

Human airway smooth muscle cells treated with lysophosphatidic acid (LPA) and epidermal growth factor (EGF) exhibit synergistic stimulation of mitogenesis (Ediger TL and Toews ML. J Pharmacol Exp Ther 294: 1076-1082, 2000). The effects of LPA treatment of human airway smooth muscle cells on EGF receptor (EGFR) regulation have now been investigated. LPA treatment for 12-24 h resulted in a twofold increase in (125)I-EGF binding and EGFR protein levels as assessed by Western blot analysis. Competition binding assays indicated single-site binding with an affinity of 3 nM, and the affinity was not changed by LPA treatment. EGFR upregulation was blocked by cycloheximide and actinomycin D, suggesting that LPA influences transcriptional regulation of EGFR expression. Inhibitor studies revealed a prominent role for activation of mitogen-activated protein kinase and p70 ribosomal S6 kinase. Both synergism and EGFR upregulation increased with increased cell density, whereas EGFR expression in control cells decreased. The similar requirements for exposure time, LPA concentrations, and cell confluence suggest that EGFR upregulation may be one contributing factor to the synergistic stimulation of mitogenesis seen with LPA plus EGF.

Simultaneous treatment of human airway smooth muscle (HASM) cells with lysophosphatidic acid (LPA) and epidermal growth factor (EGF) leads to strikingly synergistic stimulation of mitogenesis. The purpose of this study was to explore potential sites for signal integration mediating synergism, focusing on extracellular signal-regulated kinase (ERK) and transcription factors involved in proliferation and inflammation as likely candidates.Activation of ERK was analysed by immunoblotting. Transcription factor activation was assessed using HASM cells transduced with luciferase reporter gene constructs.LPA and EGF both activated ERK but had no synergistic effect when combined. LPA and EGF both activated activator protein (AP)-1, cyclic adenosine monophosphate response element-binding protein, nuclear factor of activated T-cells and the serum response element; however, only AP-1 activation exhibited synergism. Activation of the inhibitory guanine nucleotide-binding protein and of ERK signalling pathways were required for most transcription factor responses to LPA. In contrast, nuclear factor (NF)-kB was activated by LPA but not EGF and NF-kB activation was completely blocked only when Rho was inhibited. Rapid activation of Rho was observed in response to LPA but not to EGF. Importantly, inhibition of Rho selectively blocked synergism in both AP-1 activation and mitogenesis.In summary, extracellular signal-regulated kinase activation is required for many transcription factor responses to lysophosphatidic acid and epidermal growth factor, however it is not synergistic. Activation of activator protein-1 is synergistic, and Rho activation by lysophosphatidic acid is required for synergism in both activator protein-1 activation and mitogenesis. Eur Respir J 2003; 21: 759-769.

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