Anoikis, i.e. apoptosis induced by detachment from the extracellular matrix, is thought to be involved in the shedding of enterocytes at the tip of intestinal villi. Mechanisms controlling enterocyte survival are poorly understood. We investigated the role of E-cadherin, a key protein of cell-cell adhesion, in the control of anoikis of normal intestinal epithelial cells, by detaching murine villus epithelial cells from the underlying basement membrane while preserving cell-cell interactions. We show that upon the loss of anchorage, normal enterocytes execute a program of apoptosis within minutes, via a Bcl-2-regulated and caspase-9-dependent pathway. E-cadherin is lost early from cell-cell contacts. This process precedes the execution phase of detachment-induced apoptosis as it is only weakly modulated by Bcl-2 overexpression or caspase inhibition. E-cadherin loss, however, is efficiently prevented by lysosome and proteasome inhibitors. We also found that a blocking anti-E-cadherin antibody increases the rate of anoikis, whereas the activation of E-cadherin using E-cadherin-Fc chimera proteins reduces anoikis. In conclusion, our results stress the striking sensitivity of normal enterocytes to the loss of anchorage and the contribution of E-cadherin to the control of their survival/apoptosis balance. They open new perspectives on the key role of this protein, which is dysregulated in the intestinal epithelium in both inflammatory bowel disease and cancer.

Enterocytes of the intestinal epithelium are continually regenerated. They arise from precursor cells in crypts, migrate along villi, and finally die, 3–4 days later, when they reach the villus apex. Their death is thought to occur by anoikis, a form of apoptosis induced by cell detachment, but the mechanism of this process remains poorly understood. We have previously shown that a key event in the onset of anoikis in normal enterocytes detached from the basal lamina is the disruption of adherens junctions mediated by E-cadherin (Fouquet S, Lugo-Martinez VH, Faussat AM, Renaud F, Cardot P, Chambaz J, Pincon-Raymond M, Thenet S. J Biol Chem 279: 43061–43069, 2004). Here we have further investigated the mechanisms underlying this disassembly of the adherens junctions. We show that disruption of the junctions occurs through endocytosis of E-cadherin and that this process depends on the tyrosine-kinase activity of the epidermal growth factor receptor (EGFR). Activation of EGFR was detected in detached enterocytes before E-cadherin disappearance. Specific inhibition of EGFR by tyrphostin AG-1478 maintained E-cadherin and its cytoplasmic partners β- and α-catenin at cell-cell contacts and decreased anoikis. Finally, EGFR activation was evidenced in the intestinal epithelium in vivo, in rare individual cells, which were shown to lose their interactions with the basal lamina. We conclude that EGFR is activated as enterocytes become detached from the basal lamina, and that this mechanism contributes to the disruption of E-cadherin-dependent junctions leading to anoikis. This suggests that EGFR participates in the physiological elimination of the enterocytes.

Human adipose-derived mesenchymal stem cells (hADMSCs) are believed to be potential key factors for starting the regenerative process after tissue injury. However, an efficient method of delivering these regenerative cells to an external wound site is still lacking. Human amnion and pig skin have long been used as skin wound dressings for the treatment of burns and other skin lesions. Herein, we present the generation of two constructs using these two biomaterials as effective scaffolds for the culture of hADMSCs. It was found that hADMSCs seeded onto radiosterilized human amnion and pig skin are viable and proliferate. These cells are able to migrate over these scaffolds as demonstrated by using time-lapse microscopy. In addition, the scaffolds induce hADMSCs to secrete interleukin-10, an important negative regulator of inflammation, and interleukin-1β, a proinflammatory protein. The interplay between these two proteins has been proven to be vital for a balanced restoration of all necessary tissues. Thus, radiosterilized human amnion and pig skin are likely suitable scaffolds for delivery of hADMSCs transplants that could promote tissue regeneration in skin injuries like patients with burn injuries.

We assessed the role of plasminogen activator inhibitor‐1 (PAI‐1) and matrix metalloproteinase 9 (MMP9) in wound healing process and in the bone marrow mononuclear cells (BMMNC)‐related effects on physiological and pathological wound healing. A full thickness excision wound was created by removal of the skin on the midback of irradiated and nonirradiated animals. Angiogenesis and re‐epithelialization were markedly increased in PAI‐1−/− mice compared to wild‐type (WT) animals. We revealed high MMP activity in tissue of PAI‐1−/− animals. Of interest, the wound healing process was reduced in PAI‐1−/−:MMP9−/− animals compared to PAI‐1−/− mice, suggesting a key role of MMP9 in beneficial effect of PAI‐1 deficiency on wound closure. To unravel the role of PAI‐1 in BMMNC relative effects, mice were treated with or without local injection of BMMNC isolated from WT, PAI‐1−/−, and PAI‐1−/−: MMP9−/− animals for 14 days (10⁶ cells, n = 6 per group). In WT nonirradiated mice, transplantation of BMMNC isolated from PAI‐1−/− animals enhanced wound formation when compared with WT BMMNC. BMMNC differentiation into cells with endothelial phenotype was enhanced by PAI‐1 deficiency. These effects were abrogated in PAI‐1−/−:MMP9−/− and MMP9−/− BMMNC. In addition, using chimeric mice, we demonstrated that PAI‐1 deficiency environment increased the BMMNC‐GFP recruitment to the wound site, whereas this effect was abrogated when using PAI‐1−/−:MMP9−/− BMMNC. PAI‐1 deficiency, at least through MMP9 upregulation, enhanced wound healing and BMMNC therapeutic potential in irradiated and nonirradiated animals. Stem Cells2012;30:1436–1446

A plasmid vector carrying the immunoprotective amino-terminal fragment ofTaenia soliumparamyosin (VW2-1) was designed for genetic vaccination studies. Mice that were genetically immunized with VW2-1 and challenged by intraperitoneal inoculation ofTaenia crassicepscysticerci showed 43 to 48% reductions in the parasite burden, values which were similar to values obtained previously when the recombinant protein was used.