Improved efficacy of mesenchymal stromal cells stably expressing CXCR4 and IL-10 in a xenogeneic graft versus host disease mouse model.

Previous clinical trials have shown that mesenchymal stromal cells (MSCs) can modulate graft versus host disease (GvHD) after allogeneic hematopoietic transplantation, although with variable efficacy. To improve the anti-GvHD effect of these cells, adipose tissue derived-human MSCs (Ad-MSCs) were transduced with a lentiviral vector conferring stable expression of CXCR4, a molecule involved in cell migration to inflamed sites, and IL-10, a cytokine with potent anti-inflammatory properties. In vitro experiments showed that the expression of these molecules in Ad-MSCs (named CXCR4-IL10-MSCs) efficiently enhanced their migration towards SDF-1α and also improved their immunomodulatory properties compared to unmodified Ad-MSCs (WT-MSCs). Moreover, using a humanized GvHD mouse model, CXCR4-IL10-MSCs showed improved therapeutic effects, which were confirmed by histopathologic analysis in the target organs. Additionally, compared to WT-MSCs, CXCR4-IL10-MSCs induced a more marked reduction in the number of pro-inflammatory Th1 and Th17 cells, a higher polarization towards an anti-inflammatory T cell profile (CD3+-IL10+ cells), and increased the number of regulatory T and B cells. Our in vitro and in vivo studies strongly suggest that CXCR4-IL10-MSCs should constitute an important new generation of MSCs for the treatment of GvHD in patients transplanted with allogeneic hematopoietic grafts.

The Current Status of Mesenchymal Stromal Cells: Controversies, Unresolved Issues and Some Promising Solutions to Improve Their Therapeutic Efficacy.

Mesenchymal stromal cells (MSCs) currently constitute the most frequently used cell type in advanced therapies with different purposes, most of which are related with inflammatory processes. Although the therapeutic efficacy of these cells has been clearly demonstrated in different disease animal models and in numerous human phase I/II clinical trials, only very few phase III trials using MSCs have demonstrated the expected potential therapeutic benefit. On the other hand, diverse controversial issues on the biology and clinical applications of MSCs, including their specific phenotype, the requirement of an inflammatory environment to induce immunosuppression, the relevance of the cell dose and their administration schedule, the cell delivery route (intravascular/systemic vs. local cell delivery), and the selected cell product (i.e., use of autologous vs. allogeneic MSCs, freshly cultured vs. frozen and thawed MSCs, MSCs vs. MSC-derived extracellular vesicles, etc.) persist. In the current review article, we have addressed these issues with special emphasis in the new approaches to improve the properties and functional capabilities of MSCs after distinct cell bioengineering strategies.

Enhanced anti-inflammatory effects of mesenchymal stromal cells mediated by the transient ectopic expression of CXCR4 and IL10.

BACKGROUND: Mesenchymal stromal cells (MSCs) constitute one of the cell types most frequently used in cell therapy. Although several studies have shown the efficacy of these cells to modulate inflammation in different animal models, the results obtained in human clinical trials have been more modest. Here, we aimed at improving the therapeutic properties of MSCs by inducing a transient expression of two molecules that could enhance two different properties of these cells. With the purpose of improving MSC migration towards inflamed sites, we induced a transient expression of the C-X-C chemokine receptor type 4 (CXCR4). Additionally, to augment the anti-inflammatory properties of MSCs, a transient expression of the anti-inflammatory cytokine, interleukin 10 (IL10), was also induced. METHODS: Human adipose tissue-derived MSCs were transfected with messenger RNAs carrying the codon-optimized versions of CXCR4 and/or IL10. mRNA-transfected MSCs were then studied, first to evaluate whether the characteristic phenotype of MSCs was modified. Additionally, in vitro and also in vivo studies in an LPS-induced inflamed pad model were conducted to evaluate the impact associated to the transient expression of CXCR4 and/or IL10 in MSCs. RESULTS: Transfection of MSCs with CXCR4 and/or IL10 mRNAs induced a transient expression of these molecules without modifying the characteristic phenotype of MSCs. In vitro studies then revealed that the ectopic expression of CXCR4 significantly enhanced the migration of MSCs towards SDF-1, while an increased immunosuppression was associated with the ectopic expression of IL10. Finally, in vivo experiments showed that the co-expression of CXCR4 and IL10 increased the homing of MSCs into inflamed pads and induced an enhanced anti-inflammatory effect, compared to wild-type MSCs. CONCLUSIONS: Our results demonstrate that the transient co-expression of CXCR4 and IL10 enhances the therapeutic potential of MSCs in a local inflammation mouse model, suggesting that these mRNA-modified cells may constitute a new step in the development of more efficient cell therapies for the treatment of inflammatory diseases.

Improved Hematopoietic Gene Therapy in a Mouse Model of Fanconi Anemia Mediated by Mesenchymal Stromal Cells.

In this study we propose a novel approach based on the use of mesenchymal stromal cells (MSCs), aiming at limiting risks of graft failure in gene therapy protocols associated with low conditioning regimens. Because the engraftment of corrected hematopoietic stem cells (HSCs) is particularly challenging in Fanconi anemia (FA), we have investigated the relevance of MSCs in an experimental model of FA gene therapy. Our results showed, first, that risks of graft failure in recipients conditioned with a moderate dose of 5 Gy and infused with limited numbers of wild-type HSCs are significantly higher in Fanca-/- recipients as compared with wild-type recipients. However, when wild-type HSC numbers inducing 30-50% of graft failures in Fanca-/- recipients were coinfused with MSCs, no graft failures were observed. Moreover, graft failures associated with the infusion of low numbers of gene-corrected Fanca-/- HSCs were also significantly overcome by MSC coinfusion. Our study shows for the first time that MSC coinfusion constitutes a simple and nontoxic approach to minimize risks of graft failure in gene therapy applications associated with low conditioning regimens and infusion of limited numbers of corrected HSCs.

Comparative analysis of the immunomodulatory capacities of human bone marrow- and adipose tissue-derived mesenchymal stromal cells from the same donor.

BACKGROUND AIMS: The immunomodulatory properties of mesenchymal stromal cells (MSCs), together with their tissue regenerative potential, make them interesting candidates for clinical application. METHODS: In the current study, we analyzed the in vitro immunomodulatory effects of MSCs derived from bone marrow (BM-MSCs) and from adipose tissue (AT-MSCs) obtained from the same donor on both innate and acquired immunity cells. BM-MSCs and AT-MSCs were expanded to fourth or fifth passage and co-cultured with T cells, monocytes or natural killer (NK) cells isolated from human peripheral blood and stimulated in vitro. The possible differing impact of MSCs obtained from distinct sources on phenotype, cell proliferation and differentiation, cytokine production and function of these immune cells was comparatively analyzed. RESULTS: BM-MSCs and AT-MSCs induced a similar decrease in NK-cell proliferation, cytokine secretion and expression of both activating receptors and cytotoxic molecules. However, only BM-MSCs significantly reduced NK-cell cytotoxic activity, although both MSC populations showed the same susceptibility to NK-cell-mediated lysis. AT-MSCs were more potent in inhibiting dendritic-cell (DC) differentiation than BM-MSC, but both MSC populations similarly reduced the ability of DCs to induce CD4(+) T-cell proliferation and cytokine production. BM-MSCs and AT-MSCs induced a similar decrease in T-cell proliferation and production of inflammatory cytokines after activation. CONCLUSIONS: AT-MSCs and BM-MSCs from the same donor had similar immunomodulatory capacity on both innate and acquired immunity cells. Thus, other variables, such as accessibility of samples or the frequency of MSCs in the tissue should be considered to select the source of MSC for cell therapy.

Mesenchymal stromal cells enhance the engraftment of hematopoietic stem cells in an autologous mouse transplantation model.

INTRODUCTION: Studies have proposed that mesenchymal stem cells (MSCs) improve the hematopoietic engraftment in allogeneic or xenogeneic transplants and this is probably due to the MSCs' immunosuppressive properties. Our study aimed to discern, for the first time, whether MSC infusion could facilitate the engraftment of hematopoietic stem cells (HSCs) in autologous transplantations models, where no immune rejection of donor HSCs is expected. METHODS: Recipient mice (CD45.2) mice, conditioned with moderate doses of radiation (5-7 Gy), were transplanted with low numbers of HSCs (CD45.1/CD45.2) either as a sole population or co-infused with increasing numbers of adipose-derived-MSCs (Ad-MSCs). The influence of Ad-MSC infusion on the short-term and long-term engraftment of donor HSCs was investigated. Additionally, homing assays and studies related with the administration route and with the Ad-MSC/HSC interaction were conducted. RESULTS: Our data show that the co-infusion of Ad-MSCs with low numbers of purified HSCs significantly improves the short-term and long-term hematopoietic reconstitution of recipients conditioned with moderate irradiation doses. This effect was Ad-MSC dose-dependent and associated with an increased homing of transplanted HSCs in recipients' bone marrow. In vivo and in vitro experiments also indicate that the Ad-MSC effects observed in this autologous transplant model are not due to paracrine effects but rather are related to Ad-MSC and HSC interactions, allowing us to propose that Ad-MSCs may act as HSC carriers, facilitating the migration and homing of the HSCs to recipient bone marrow niches. CONCLUSION: Our results demonstrate that Ad-MSCs facilitate the engraftment of purified HSCs in an autologous mouse transplantation model, opening new perspectives in the application of Ad-MSCs in autologous transplants, including HSC gene therapy.

Role of cytochrome P-450 genetic polymorphisms in oral carcinogenesis.

Oral cancer is one of the most frequent head and neck cancers, and epidemiological studies have shown that smoking is a major risk factor in this pathology. However, as not all smokers develop oral cancer, some individuals must be more susceptible to develop this disease. This individual susceptibility has been related to different genetic variants in metabolizing enzymes. The cytochrome P-450 (CYP) family of enzymes metabolizes tobacco-related carcinogens producing reactive metabolites, which could cause DNA damage. Because of their functional role in the metabolism of tobacco-related compounds, the genetic polymorphisms found in the genes that code for CYP enzymes have been suggested to modulate oral cancer risk and contribute to individual susceptibility. In this review, we analyze and update the available evidence in the literature regarding the polymorphisms of CYP genes in relation to the susceptibility of developing oral cancer.

Alcohol-dehydrogenase (ADH1B) Arg48His polymorphism in Basque Country patients with oral and laryngeal cancer: preliminary study.

Oral and laryngeal cancer has a high incidence in the Basque Country (Spain), the main risk factors in this pathology being regular consumption of tobacco and alcohol. However, since not all the individuals exposed to these risk factors develop cancer, the individual genetic susceptibility should be investigated in this population. The aim of this study was to determine the distribution of alcohol dehydrogenase-1B polymorphism (Arg48His; rs1229984) in our region and analyze its association with the risk of oral and laryngeal squamous cell carcinoma. Samples from 87 patients with oral or laryngeal cancer and 242 healthy controls were analyzed. Multivariate logistic regression analysis showed that the combined Arg/His and His/His genotypes were associated with a reduced risk of head and neck squamous cell carcinoma (odds ratio: 0.203; 95% confidence interval: 0.052-0.796). In conclusion, the histidine allele was associated with a reduced risk of oral and laryngeal cancer in the Basque Country.