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Regenerative medicine aims to repair and regenerate damaged cells, tissues, and organs in order to restore function. Regeneration can be obtained either by cell replacement or by stimulating the body's own repair mechanisms. Importantly, a favorable environment is required before any regenerative signal can stimulate resident stem/stromal cells, and regeneration is possible only after the resolution of injury-induced inflammation. An exacerbated immune response is often present in cases of degenerative, inflammatory-based diseases. Here we discuss how amniotic membrane cells, and their derivatives, can contribute to the resolution of many diseases with altered immune response by acting on different inflammatory mediators.

We previously demonstrated that mesenchymal cells from human amniotic membrane (hAMTCs) inhibit the generation and maturation of monocyte-derived dendritic cells (DCs) in vitro. Considering the crucial role of DCs in the immune response and that epithelial cells of the human amniotic membrane (hAECs) share some of the immunoregulatory properties of hAMTCs, we investigated whether hAECs also modulate monocyte-derived DCs. We compared hAECs with hAMTCs in a cell-to-cell contact setting and their secreted factors in modulating DC differentiation and function. First, we demonstrated that primary and expanded hAMTCs strongly inhibited the differentiation of DCs and induced a shift toward M2-like macrophages. This was observed when hAMTCs were cultured in contact (hAMTC-DC cont ) or in Transwells (hAMTC-DC tw ) with monocytes and even when medium conditioned by hAMTCs was used instead of hAMTCs. hAECs also prevented DC development, but to a lesser extent than hAMTCs. hAECs were more effective when cultured in contact with monocytes (hAEC-DC cont ) rather than in Transwells (hAEC-DC tw ). The modulatory capacity of hAECs changed during passaging unlike the hAMSCs. The ability to stimulate CD4 + and CD8 + T-cell proliferation was almost completely abolished by hAMTC-DC cont , whereas hAMTC-DC tw and hAEC-DC cont displayed only a reduced ability to stimulate CD8 + T cells. Furthermore, monocytes cocultured with hAMTCs and hAECs showed some similarities, but also differences in cytokine/chemokine secretion. Similarities were observed in the inhibition of IL-12p70 and TNF-α and the increase in IL-10 in supernatants taken from monocyte-DCs cocultured with hAMTCs and hAECs in contact and Transwell settings. The inflammatory factors IL-8, CXCL9, and MIP-1α were significantly lower in hAMTC-DC cont , hAMTC-DC tw , and hAEC-DC cont conditions. In contrast, only hAMTCs (in both contact and Transwell conditions) were able to significantly increase IL-1β and CCL2. Altogether, we demonstrated that hAMTCs and hAECs affect DC differentiation, but that hAMTCs exerted a stronger inhibitory effect, abolished T-cell proliferation, and also induced more changes in cytokine/ chemokine production.

Biliary fibrosis and resultant cirrhosis are among the most common outcomes of chronic liver diseases. Currently, liver transplantation remains the only effective treatment. In seeking alternative therapeutic approaches, we focused on the potential use of the human amniotic membrane (AM). Indeed, AM has gained increasing importance for its antiscarring, anti-inflammatory, and wound-healing properties, as well as for the multipotent differentiation ability and immunomodulatory features of AM-derived cells. Intriguingly, we have recently demonstrated that placenta-derived cells reduce lung fibrosis in bleomycin-treated mice, and that AM patches reduce postischemic cardiac injury in rats. Hence, we have now investigated the effects of human AM on biliary fibrosis induced in rats through the bile duct ligation (BDL) procedure. A fragment of human AM was applied onto the liver surface after BDL and the effects on fibrosis establishment and progression were evaluated at different time points in comparison with fibrosis progression in control BDL rats. The degree of liver fibrosis was first assessed by the semiquantitative Knodell scoring system and, thereafter, by digital image morphometric analysis to quantify the area occupied by ductular reaction, activated myofibroblasts, and collagen deposition. We demonstrated a significant reduction in the severity of BDL-induced fibrosis in AM-treated rats. Indeed, while fibrosis progressed rapidly in control BDL rats, leading to cirrhosis within 6 weeks, AM-treated rats showed confined fibrosis at the portal/periportal area with no signs of cirrhosis, and a reduction in collagen deposition to about 50% of levels observed in control BDL rats. In addition, the AM was able to significantly slow the gradual progression of the ductular reaction and reduce, at all time points, the area occupied by activated myofibroblasts. These findings suggest that human AM, when applied as a patch onto the liver surface, might inhibit fibrosis progression in BDL-injured livers, and could protect against hepatic damage associated with fibrotic degeneration.

The amniotic membrane has long been applied for wound healing and treatment of ophthalmological disorders, even though the mechanisms underlying its actions remain to be clarified. Recently, cells derived from fetal membranes of human term placenta have raised strong interest in regenerative medicine for their stem cell potential and immunomodulatory features. Our study aimed to investigate the possible utility of amniotic membrane to limit postischemic cardiac injury. A fragment of human amniotic membrane was applied onto the left ventricle of rats that had undergone ischemia through left anterior descending coronary artery ligation. Echocardiographic assessment of morphological and functional cardiac parameters was then performed over a 3-month period. We demonstrated that application of an amniotic membrane fragment onto ischemic rat hearts could significantly reduce postischemic cardiac dysfunction. The amniotic membrane-treated rats showed higher preservation of cardiac dimensions and improved cardiac contractile function in terms of higher left ventricle ejection fraction, fractional shortening, and wall thickening. These improvements were apparent by day 7 after application of the amniotic membrane, persisted for at least 2 months, and occurred independently of cardiac injury severity. No engraftment of amniotic cells was detected into host cardiac tissues. Our results suggest that use of amniotic membrane may constitute a convenient vehicle for supplying cells that produce cardioprotective soluble factors, and reinforce the notion that this tissue constitutes a cell source with clinical potential that has yet to be completely revealed.

Cells derived from the amniotic membranes of human term placenta have drawn much interest for their characteristics of multipotency and low immunogenicity, supporting a variety of possible clinical applications in the field of cell transplantation and regenerative medicine. We have previously shown that cells derived from the mesenchymal region of human amnion (AMTC) can strongly inhibit T-lymphocyte proliferation. In this study, we demonstrate that AMTC can block differentiation and maturation of monocytes into dendritic cells (DC), preventing the expression of the DC marker CD1a and reducing the expression of HLA-DR, CD80, and CD83. The monocyte maturation block resulted in impaired allostimulatory ability of these cells on allogeneic T cells. In attempting to define the mechanisms responsible for these findings, we have observed that the presence of AMTC in differentiating DC cultures results in the arrest of the cells to the G 0 phase and abolishes the production of inflammatory cytokines such as TNF-α, CXCL10, CXCL9, and CCL5. Finally, we also demonstrate that the monocytic cells present in the amniotic mesenchymal region fail to differentiate toward the DC lineage. Taken together, our data suggest that the mechanisms by which AMTC exert immumodulatory effects do not only relate directly to T cells, but also include inhibition of the generation and maturation of antigen-presenting cells. In this context, AMTC represent a very attractive source of multipotent allogeneic cells that promise to be remarkably valuable for cell transplantation approaches, not only due to their low immunogenicity, but also because of the added potential of modulating immune responses, which could be fundamental both for controlling graft rejection after transplantation and also for controlling diseases characterized by inflammatory processes.

Fetal membranes (amnion and chorion) have recently raised significant attention as potential sources of stem cells. We have recently demonstrated that cells derived from human term placenta show stem cell phenotype, high plasticity, and display low immunogenicity both in vitro and in vivo. Moreover, placenta-derived cells, after xenotransplantation, are able to engraft in solid organs including the lung. On these bases, we studied the effects of fetal membrane-derived cells on a mouse model of bleomycin-induced lung fibrosis. Fetal membrane-derived cells were infused 15 min after intratracheal bleomycin instillation. Different delivery routes were used: intraperitoneal or intratracheal for both xenogeneic and allogeneic cells, and intravenous for allogeneic cells. The effects of the transplanted cells on bleomycin-induced inflammatory and fibrotic processes were then scored and compared between transplanted and control animals at different time points. By PCR and immunohistochemistry analyses, we demonstrated the presence of transplanted cells 3, 7, 9, and 14 days after transplantation. Concomitantly, we observed a clear decrease in neutrophil infiltration and a significant reduction in the severity of bleomycin-induced lung fibrosis in mice treated with placenta-derived cells, irrespective of the source (allogeneic or xenogeneic) or delivery route. Our findings constitute further evidence in support of the hypothesis that placenta-derived cells could be useful for clinical application, and warrant further studies toward the use of these cells for the repair of tissue damage associated with inflammatory and fibrotic degeneration.