Abstract
In recent years, mesenchymal stem cells (MSCs) derived from umbilical cord Wharton’s jelly (ucMSCs) have gained much attention in regenerative medicine. Multiple papers have described their high differentiation potential, as well as their important immunomodulatory effects. Umbilical cords (UCs) offer an extensive source of ucMSCs for clinical applications, with the same benefits as other MSCs used in autologous cell therapies. In addition, they have the advantage that their procurement does not involve any risk, as UCs are usually discarded after birth. Considering the importance of developing a more effective therapy for treating wounds, especially chronic venous ulcers (CVUs), we previously analyzed the therapeutic potential of a specific subpopulation derived from ucMSCs, which we named DMCs. The present work aims to complement our previously published results by extending the characterization of DMCs and analyzing graft acceptance after allogeneic transplantation in serial doses in a pre-clinical trial. First, the presence of human leukocyte antigen-G (HLA-G) molecule, a potent immunosuppressive checkpoint (IC), was assessed by immunohistochemistry (IHC) directly in UCs, then cells were cultured and analyzed by flow cytometry (FC) to label membrane-bound HLA-G isoforms. RT-PCR was performed using specific primers to detect different HLA-G isoforms and key markers in cell transplant acceptance, such as PDL-1, IDO, FoxP3 and CTLA-4. To determine the immune response after re-exposure to DMCs and in vivo graft acceptance, a multiple dose assay was performed in C3H/s strain mice. In the results by IHC and FC, the expression of HLA-G was low, but when RT-PCR was performed, HLA-G expression was found in 66% of the samples analyzed and, surprisingly, we found that 88% of this expression detected in ucMSCs corresponds to isoforms that have lost the α1 domain. So far, all HLA-G isoforms described have this domain, so this finding could be due to the detection of unknown isoforms for ucMSCs. Among the analyzed markers, we found that PDL-1, FoxP3, LAG3, TNFR2, VEGF-A, NRP-1, NCAM and CLU were strongly expressed in DMCs, while CTLA-4 and IDO were moderately expressed. In the murine preclinical trial, skin samples were taken after the last cell application and histological skin analyses revealed no signs of lymphocytic infiltration or tissue abnormalities. No cellular rejection or adverse skin reactions, such as swelling or redness, were observed. These observations demonstrate the safe use of DMCs after several consecutive applications of cell doses. In conclusion, an allogeneic transplant of DMCs could be used as a possible therapeutic alternative for treating CVUs. The expression of the different selected markers in the DMCs cell line would allow the generation of a tolerogenic environment and could play a key role in the activation of angiogenesis, cell migration and proliferation for the regeneration of the tissues involved. Finally, we want to highlight that we have successfully tested the possibility of applying several serial doses, considering the treatment of patients with severe and extensive wounds, which could require more prolonged treatment.

Keywords: MSCs, allogeneic cell transplantation, umbilical cord, HLA-G, immunocheckpoints.

Abstract

Hematopoietic stem cell transplantation (HSCT) stands as a pivotal intervention for hematologic disorders like leukemia, lymphoma, and specific immune deficiencies. However, the engraftment failure and delayed neutrophils regeneration present formidable obstacles and challenges. These results in prolonged cytopenia and heightened complication risks, ultimately elevating morbidity and mortality rates. Mesenchymal stem cells (MSCs) play indispensable roles in supporting hematopoiesis, yet their intricate interplay demands further exploration and comprehension. Previous studies have shown that the synergistic action of MSCs and activated T cells enhances the function of neutrophils. To improve the success of HSCT, our research reveals that the combined infusion of allotype-cord blood HSCs (CB-HSCs) and activated T cells from the same donor with third-party MSCs facilitates the differentiation of human CB-HSCs into neutrophils in vivo. Furthermore, in a model of respiratory infection, the triple cell therapy resulted in increased accumulation of human neutrophils, secretion of IL-6 and IL-8, bacterial clearance, and reduced mortality compared to the control group. These findings illuminate an axis between allo-CB-HSCs, MSCs, and activated T cells, offering a promising avenue to enhance engraftment and mitigate complications in clinical settings. This innovative approach holds potential to expedite neutrophils production post-transplantation and bolster resistance against infections, representing a significant advancement in HSCT outcomes.

Abstract

Treatment of liver failure using stem cells is actual and a promising area of regenerative medicine. Experimental models of acute and chronic liver failure have studied the effectiveness of stem cells for the correction and treatment of morphological and functional characteristics of the liver. Male Wistar rats(n=120) in acute liver failure(ALF) and(n=160) in chronic liver failure(CLF) weighed 400–450 g. were been used. Hematopoietic stem cells(HSCs) and mesenchymal stem cells(MSCs) were obtained using standard methods. To obtain Apoptotic HSCs(aHSCs), we incubated HSCs in Custodiol solution. MSCs were adhered to implanted cell-engineered constructs(CECs) are built from biopolymer-based, microstructured, and collagen-containing hydrogel(BMCG) or recombinant spidroin rS1/9. The ALF was created by resecting 75% of the liver parenchyma. The CLF had been provoked by long-term exposure to carbon tetrachloride. Wistar rats(n=280) randomized into 7 groups: Gr.1 was a 1control with the saline treatment of the intraperitoneal injections(n=40); Gr.2 was a 2control with the saline treatment of the hepatic parenchyma(n=40); Gr.3 was with the HSCs treatment of the intraperitoneal injections(n=40); Gr.4 of aHSCs received single intraperitoneal injections(n=40). Gr.5 was loaded with MSCs into the parenchyma livers(n=40); Gr.6 was loaded with CECs based on BMCG implanted into the parenchyma livers(n=40); Gr.7 was loaded with CECs based on recombinant spidroin rS1/9 implanted into the parenchyma of their livers(n=40). August rats(n=100) made up a donor population for HSCs and MSCs to develop grafts for animals from Gr.3-7. The study length was 90 days. It’s was shown that HSCs, aHSCs and MSCs had affect biochemical test values and morphological parameters in rats with ALF and CLF. Conclusion: We found that HSC, aHSCs and MSCs were operational and active, with regenerative potential and might represent a proper option to correct and treat ALF and CLF, as well as to maintain affected liver function in patients with liver grafting needed.

Aabstract

Exposure to a space environment characterized by the absence of mechanical loading significantly reduces both the differentiation and the regenerative potential of embryonic stem cells. In a study conducted aboard a space shuttle during the Discovery mission, murine embryonic stem cells were cultured in embryoid bodies for fifteen days under microgravity conditions. These embryoid bodies were compared with those maintained under terrestrial gravitational conditions. The results revealed that, in the absence of the usual mechanical stimulation, the embryoid bodies exhibited a marked inhibition in the expression of markers associated with terminal differentiation, while retaining elevated levels of self-renewal. This finding indicates that the cells remain in an immature state, which may be interpreted as a preservation of regenerative potential, yet simultaneously poses a significant challenge for tissue repair during long-duration spaceflights. Gene expression analysis further revealed alterations in signaling pathways essential for cellular maturation, including those related to Notch and Wnt. Additionally, a reduced consumption of glucose was observed, suggesting decreased metabolic activity within the space environment. Interestingly, upon return to terrestrial gravitational conditions, the embryoid bodies previously exposed to microgravity demonstrated an increased potential to differentiate into colonies of specialized cardiac muscle cells. These findings underscore the critical role of mechanical loading in promoting normal cellular differentiation and tissue regeneration. They also highlight the necessity of developing countermeasures that simulate mechanical stimulation to ensure proper tissue repair during extended space missions.