Bone marrow-derived mesenchymal stem cells (MSCs) modulate immune response and can produce significant levels of transforming growth factor-β (TGF-β) and interleukin-6 (IL-6). These 2 cytokines represent the key factors that reciprocally regulate the development and polarization of naive T-cells into regulatory T-cell (Treg) population or proinflammatory T helper 17 (Th17) cells. In the present study we demonstrate that MSCs and their products effectively regulate expression of transcription factors Foxp3 and RORγt and control the development of Tregs and Th17 cells in a population of alloantigen-activated mouse spleen cells or purified CD4⁺CD25⁻ T-cells. The immunomodulatory effects of MSCs were more pronounced when these cells were stimulated to secrete TGF-β alone or TGF-β together with IL-6. Unstimulated MSCs produce TGF-β, but not IL-6, and the production of TGF-β can be further enhanced by the anti-inflammatory cytokines IL-10 or TGF-β. In the presence of proinflammatory cytokines, MSCs secrete significant levels of IL-6, in addition to a spontaneous production of TGF-β. MSCs producing TGF-β induced preferentially expression of Foxp3 and activation of Tregs, whereas MSC supernatants containing TGF-β together with IL-6 supported RORγt expression and development of Th17 cells. The effects of MSC supernatants were blocked by the inclusion of neutralization monoclonal antibody anti-TGF-β or anti-IL-6 into the culture system. The results showed that MSCs represent important players that reciprocally regulate the development and differentiation of uncommitted naive T-cells into anti-inflammatory Foxp3⁺ Tregs or proinflammatory RORγt⁺ Th17 cell population and thereby can modulate autoimmune, immunopathological, and transplantation reactions.

Limbal stem cells (LSC), which reside in the basal layer of the limbus, are thought to be responsible for corneal epithelial healing after injury. When the cornea is damaged, LSC start to proliferate, differentiate, and migrate to the site of injury. To characterize the signaling molecules ensuring communication between the cornea and LSC, we established a mouse model of mechanical corneal damage. The central cornea or limbal tissue was excised at different time intervals after injury, and the expression of genes in the explants was determined. It was observed that a number of genes for growth and differentiation factors were significantly upregulated in the cornea rapidly after injury. The ability of these factors to regulate the differentiation and proliferation of limbal cells was tested. It was found that the insulin-like growth factor-I (IGF-I), which is rapidly overexpressed after injury, enhances the expression of IGF receptor in limbal cells and induces the differentiation of LSC into cells expressing the corneal cell marker, cytokeratin K12, without any effect on limbal cell proliferation. In contrast, the epidermal growth factor (EGF) and fibroblast growth factor-β (FGF-β), which are also produced by the damaged corneal epithelium, supported limbal cell proliferation without any effect on their differentiation. Other factors did not affect limbal cell differentiation or proliferation. Thus, IGF-I was identified as the main factor stimulating the expression of IGF receptors in limbal cells and inducing the differentiation of LSC into cells expressing corneal epithelial cell markers. The proliferation of these cells was supported by EGF and FGF.

Stem cell (SC) therapy represents a promising approach to treat a wide variety of injuries, inherited diseases, or acquired SC deficiencies. One of the major problems associated with SC therapy remains the absence of a suitable matrix for SC growth and transfer. We describe here the growth and metabolic characteristics of mouse limbal stem cells (LSCs) and mesenchymal stem cells (MSCs) growing on 3D nanofiber scaffolds fabricated from polyamide 6/12 (PA6/12). The nanofibers were prepared by the original needleless electrospun Nanospider technology, which enables to create nanofibers of defined diameter, porosity, and a basis weight. Copolymer PA6/12 was selected on the basis of the stability of its nanofibers in aqueous solutions, its biocompatibility, and its superior properties as a matrix for the growth of LSCs, MSCs, and corneal epithelial and endothelial cell lines. The morphology, growth properties, and viability of cells grown on PA6/12 nanofibers were comparable with those grown on plastic. LSCs labeled with the fluorescent dye PKH26 and grown on PA6/12 nanofibers were transferred onto the damaged ocular surface, where their seeding and survival were monitored. Cotransfer of LSCs with MSCs, which have immunosuppressive properties, significantly inhibited local inflammatory reactions and supported the healing process. The results thus show that nanofibers prepared from copolymer PA6/12 represent a convenient scaffold for growth of LSCs and MSCs and transfer to treat SC deficiencies and various ocular surface injuries.

The immunosuppressive effects of UV radiation have been well documented. This suppression has been attributed to the action of the cis form of urocanic acid (UCA), a photoproduct of trans-UCA, a natural constituent of the skin. Here, we show that mouse spleen cells preincubated with cis-UCA have a diminished proliferative response to allogeneic cells in MLC and to stimulation with anti-CD3 mAb. Cells preincubated with cis-UCA also had a decreased ability to serve as APC and to stimulate the proliferation of allogeneic lymphocytes in MLC. Simultaneously, the production of IL-2 and IFN-γ by cells preincubated with cis-UCA was decreased. However, IL-10 gene expression and IL-10 protein secretion by spleen cells stimulated in the presence of cis-UCA were significantly enhanced. The principal cell population displaying the cis-UCA-induced elevated production of IL-10 was CD4+ T cells, which were shown to be a direct target of cis-UCA action. This was also supported by the observation that production of IL-10 by stimulated splenic non-T cells or by macrophages was not altered by cis-UCA. The enhanced production of IL-10 by activated CD4+ T cells may represent a novel pathway of UVB radiation-induced, cis-UCA-mediated immunosuppression. We suggest that the elevated production of IL-10 by activated CD4+ T cells may account for the suppressor T cell phenomena described in UV-irradiated recipients.