RESUMO
Leucine-rich repeat-containing G protein-coupled receptor 5 (LGR5) and B-cell-specific Moloney murine leukemia virus insertion site 1 (BMI1) are markers of fast-cycling and quiescent intestinal stem cells, respectively. To determine the functions of these proteins in large animals, we investigated their effects on the proliferation of intestinal epithelial cells from pigs. Our results indicated that LGR5 and BMI1 are highly conserved proteins and that the pig proteins have greater homology with the human proteins than do mouse proteins. Overexpression of either LGR5 or BMI1 promoted cell proliferation and WNT/ß-catenin signaling in pig intestinal epithelial cells (IPEC-J2). Moreover, the activation of WNT/ß-catenin signaling by recombinant human WNT3A protein increased cell proliferation and LGR5 and BMI1 protein levels. Conversely, inhibition of WNT/ß-catenin signaling using XAV939 reduced cell proliferation and LGR5 and BMI1 protein levels. This is the first report that LGR5 and BMI1 can increase proliferation of pig intestinal epithelial cells by activating WNT/ß-catenin signaling.
Assuntos
Proliferação de Células/fisiologia , Complexo Repressor Polycomb 1/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Via de Sinalização Wnt/fisiologia , beta Catenina/metabolismo , Animais , Proliferação de Células/efeitos dos fármacos , Proliferação de Células/genética , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Compostos Heterocíclicos com 3 Anéis/farmacologia , Humanos , Intestinos/citologia , Complexo Repressor Polycomb 1/genética , Receptores Acoplados a Proteínas G/genética , Suínos , Via de Sinalização Wnt/efeitos dos fármacos , Via de Sinalização Wnt/genética , Proteína Wnt3A/genética , Proteína Wnt3A/metabolismoRESUMO
The focal adhesion kinase (FAK) signaling pathway contributes to the cell migration and adhesion that is critical for wound healing and regeneration of damaged muscle, but its function in skeletal muscle satellite cells (SCs) is less clear. We compared the migration and adhesion of SCs derived from two species of pig (Lantang and Landrace) in vitro, and explored how FAK signaling modulates the two processes. The results showed that Lantang SCs had greater ability to migrate and adhere to fibronection (P < 0.05) than Landrace SCs. Compared to Landrace SCs, Lantang SCs expressed many more focal adhesion (FA) sites, which were indicated by the presence of p-paxillin (Tyr118), and exhibited less F-actin reorganization 24 h after seeding onto fibronectin. Levels of p-FAK (Tyr397) and p-paxillin (Tyr118) were greater (P < 0.05) in Lantang SCs than Landrace SCs after migration for 24 h. Similarly, Lantang SCs showed much higher levels of p-FAK (Tyr397), p-paxillin (Tyr118) and p-Akt (Ser473) than Landrace SCs 2 h after adhesion. Treatment with the FAK inhibitor PF-573228 (5 or 10 µmol/L) inhibited Lantang SC migration and adhesion to fibronectin (P < 0.05), decreased levels of p-paxillin (Tyr118) and p-Akt (Ser473) (P < 0.05), and suppressed the formation of FA sites on migrating SCs. Thus FAK appears to play a key role in the regulation of SC migration and adhesion necessary for muscle regeneration.
