Bioresorbable Bilayered Elastomer/Hydrogel Constructs with Gradual Interfaces for the Fast Actuation of Self-Rolling Tubes.

In the biomedical field, self-rolling materials provide interesting opportunities to develop medical devices suitable for drug or cell encapsulation. However, to date, a major limitation for medical applications is the use of non-biodegradable and non-biocompatible polymers that are often reported for such applications or the slow actuation witnessed with degradable systems. In this work, biodegradable self-rolling tubes that exhibit a spontaneous and rapid actuation when immersed in water are designed. Photo-crosslinkable hydrophilic and hydrophobic poly(ethylene glycol)-poly(lactide) (PEG-PLA) star-shaped copolymers are prepared and used to prepare bilayered constructs. Thanks to the discrete mechanical and swelling properties of each layer and the cohesive/gradual nature of the interface, the resulting bilayered films are able to self-roll in water in less than 30 s depending on the nature of the hydrophilic layer and on the shape of the sample. The cytocompatibility and degradability of the materials are demonstrated and confirm the potential of such self-rolling resorbable biomaterials in the field of temporary medical devices.

Embedding MSCs in Si-HPMC hydrogel decreased MSC-directed host immune response and increased the regenerative potential of macrophages.

Embedding mesenchymal stromal cells (MSCs) in biomaterial is a subject of increasing interest in the field of Regenerative Medicine. Speeding up the clinical use of MSCs is dependent on the use of non-syngeneic models in accordance with Good Manufacturing Practices (GMP) requirements and on costs. To this end, in this study, we analyzed the in vivo host immune response following local injection of silanized hydroxypropyl methylcellulose (Si-HPMC)-embedded human MSCs in a rat model developing colorectal damage induced by ionizing radiation. Plasma and lymphocytes from mesenteric lymph nodes were harvested in addition to colonic tissue. We set up tests, using flow cytometry and a live imaging system, to highlight the response to specific antibodies and measure the cytotoxicity of lymphocytes against injected MSCs. We demonstrated that Si-HPMC protects MSCs from specific antibodies production and from apoptosis by lymphocytes. We also observed that Si-HPMC does not modify innate immune response infiltrate in vivo, and that in vitro co-culture of Si-HPMC-embedded MSCs impacts macrophage inflammatory response depending on the microenvironment but, more importantly, increases the macrophage regenerative response through Wnt-family and VEGF gene expression. This study furthers our understanding of the mechanisms involved, with a view to improving the therapeutic benefits of biomaterial-assisted cell therapy by modulating the host immune response. The decrease in specific immune response against injected MSCs protected by Si-HPMC also opens up new possibilities for allogeneic clinical use.

Refining the mandibular osteoradionecrosis rat model by in vivo longitudinal µCT analysis.

Osteoradionecrosis (ORN) is one of the most feared side effects of radiotherapy following cancers of the upper aero-digestive tract and leading to severe functional defects in patients. Today, our lack of knowledge about the physiopathology restricts the development of new treatments. In this study, we refined the ORN rat model and quantitatively studied the progression of the disease. We tested the impact of radiation doses from 20 to 40 Gy, delivered with incident 4MV X-ray beams on the left mandible of the inbred Lewis Rat. We used micro-computed tomography (µCT) to obtain in vivo images for longitudinal bone imaging and ex vivo images after animal perfusion with barium sulphate contrast agent for vessel imaging. We compared quantification methods by analyzing 3D images and 2D measurements to determine the most appropriate and precise method according to the degree of damage. We defined 25 Gy as the minimum irradiation dose combined with the median molar extraction necessary to develop non-regenerative bone necrosis. µCT image analyses were correlated with clinical and histological analyses. This refined model and accurate methods for bone and vessel quantification will improve our knowledge of the progression of ORN pathology and allow us to test the efficacy of new regenerative medicine procedures.

Development of extracellular vesicle-based medicinal products: A position paper of the group "Extracellular Vesicle translatiOn to clinicaL perspectiVEs - EVOLVE France".

Extracellular vesicles (EV) are emergent therapeutic effectors that have reached clinical trial investigation. To translate EV-based therapeutic to clinic, the challenge is to demonstrate quality, safety, and efficacy, as required for any medicinal product. EV research translation into medicinal products is an exciting and challenging perspective. Recent papers, provide important guidance on regulatory aspects of pharmaceutical development, defining EVs for therapeutic applications and critical considerations for the development of potency tests. In addition, the ISEV Task Force on Regulatory Affairs and Clinical Use of EV-based Therapeutics as well as the Exosomes Committee from the ISCT are expected to contribute in an active way to the development of EV-based medicinal products by providing update on the scientific progress in EVs field, information to patients and expert resource network for regulatory bodies. The contribution of our work group "Extracellular Vesicle translatiOn to clinicaL perspectiVEs - EVOLVE France", created in 2020, can be positioned in complement to all these important initiatives. Based on complementary scientific, technical, and medical expertise, we provide EV-specific recommendations for manufacturing, quality control, analytics, non-clinical development, and clinical trials, according to current European legislation. We especially focus on early phase clinical trials concerning immediate needs in the field. The main contents of the investigational medicinal product dossier, marketing authorization applications, and critical guideline information are outlined for the transition from research to clinical development and ultimate market authorization.

Regenerative medicine for digestive fistulae therapy: Benefits, challenges and promises of stem/stromal cells and emergent perspectives via their extracellular vesicles.

Despite current management strategies, digestive fistulae remain extremely debilitating complications associated with significant morbidity and mortality, generating a need to develop innovative therapies in these indications. A number of clinical trials and experimental studies have thus investigated the potential of stem/stromal cells (SCs) or SC-derived extracellular vesicles (EVs) administration for post-surgical and Crohn's-associated fistulae. This review summarizes the physiopathology and current standards-of-care for digestive fistulae, along with relevant evidence from animal and clinical studies regarding SC or EV treatment for post-surgical digestive fistulae. Additionally, existing preclinical models of fistulizing Crohn's disease and results of SC therapy trials in this indication will be presented. The optimal formulation and administration protocol of SC therapy products for gastrointestinal fistula treatment and the challenges for a widespread use of darvadstrocel (Alofisel) in clinical practice will be discussed. Finally, the potential advantages of EV therapy and the obstacles towards their clinical translation will be introduced.

The stromal vascular fraction mitigates radiation-induced gastrointestinal syndrome in mice.

BACKGROUND: The intestine is particularly sensitive to moderate-high radiation dose and the development of gastrointestinal syndrome (GIS) leads to the rapid loss of intestinal mucosal integrity, resulting in bacterial infiltration, sepsis that comprise patient survival. There is an urgent need for effective and rapid therapeutic countermeasures. The stromal vascular fraction (SVF) derived from adipose tissue is an easily accessible source of cells with angiogenic, anti-inflammatory and regenerative properties. We studied the therapeutic impact of SVF and its action on the intestinal stem cell compartment. METHODS: Mice exposed to the abdominal radiation (18 Gy) received a single intravenous injection of stromal vascular fraction (SVF) (2.5 × 106 cells), obtained by enzymatic digestion of inguinal fat tissue, on the day of irradiation. Mortality was evaluated as well as intestinal regeneration by histological analyses and absorption function. RESULTS: The SVF treatment limited the weight loss of the mice and inhibited the intestinal permeability and mortality after abdominal irradiation. Histological analyses showed that SVF treatment stimulated the regeneration of the epithelium by promoting numerous enlarged hyperproliferative zones. SVF restored CD24+/lysozyme- and Paneth cell populations in the ISC compartment with the presence of Paneth Ki67+ cells. SVF has an anti-inflammatory effect by repressing pro-inflammatory cytokines, increasing M2 macrophages in the ileum and anti-inflammatory monocyte subtypes CD11b+Ly6clowCX3CR1high in the spleen. CONCLUSIONS: Through the pleiotropic effects that contribute to limiting radiation-induced lethality, SVF opens up attractive prospects for the treatment of emergency GIS.

BMP Antagonists Secreted by Mesenchymal Stromal Cells Improve Colonic Organoid Formation: Application for the Treatment of Radiation-induced Injury.

Radiation therapy is crucial in the therapeutic arsenal to cure cancers; however, non-neoplastic tissues around an abdominopelvic tumor can be damaged by ionizing radiation. In particular, the radio-induced death of highly proliferative stem/progenitor cells of the colonic mucosa could induce severe ulcers. The importance of sequelae for patients with gastrointestinal complications after radiotherapy and the absence of satisfactory management has opened the field to the testing of innovative treatments. The aim of this study was to use adult epithelial cells from the colon, to reduce colonic injuries in an animal model reproducing radiation damage observed in patients. We demonstrated that transplanted in vitro-amplified epithelial cells from colonic organoids (ECO) of C57/Bl6 mice expressing green fluorescent protein implant, proliferate, and differentiate in irradiated mucosa and reduce ulcer size. To improve the therapeutic benefit of ECO-based treatment with clinical translatability, we performed co-injection of ECO with mesenchymal stromal cells (MSCs), cells involved in niche function and widely used in clinical trials. We observed in vivo an improvement of the therapeutic benefit and in vitro analysis highlighted that co-culture of MSCs with ECO increases the number, proliferation, and size of colonic organoids. We also demonstrated, using gene expression analysis and siRNA inhibition, the involvement of bone morphogenetic protein antagonists in MSC-induced organoid formation. This study provides evidence of the potential of ECO to limit late radiation effects on the colon and opens perspectives on combined strategies to improve their amplification abilities and therapeutic effects.

Heparan Sulfate Mimetics: A New Way to Optimize Therapeutic Effects of Hydrogel-Embedded Mesenchymal Stromal Cells in Colonic Radiation-Induced Damage.

Clinical expression of gastrointestinal radiation toxicity on non-cancerous tissue could be very life threatening and clinicians must deal increasingly with the management of late side effects of radiotherapy. Cell therapy, in particular mesenchymal stromal cell (MSC) therapy, has shown promising results in numerous preclinical animal studies and thus has emerged as a new hope for patient refractory to current treatments. However, many stem cell clinical trials do not confer any beneficial effect suggesting a real need to accelerate research towards the successful clinical application of stem cell therapy. In this study, we propose a new concept to improve the procedure of MSC-based treatment for greater efficacy and clinical translatability. We demonstrated that heparan sulfate mimetic (HS-m) injections that restore the extracellular matrix network and enhance the biological activity of growth factors, associated with local injection of MSC protected in a hydrogel, that increase cell engraftment and cell survival, improve the therapeutic benefit of MSC treatment in two animal models relevant of the human pathology. For the first time, a decrease of the injury score in the ulcerated area was observed with this combined treatment. We also demonstrated that the combined treatment favored the epithelial regenerative process. In this study, we identified a new way, clinically applicable, to optimize stem-cell therapy and could be proposed to patients suffering from severe colonic defect after radiotherapy.

A biomaterial-assisted mesenchymal stromal cell therapy alleviates colonic radiation-induced damage.

Healthy tissues surrounding abdomino-pelvic tumours can be impaired by radiotherapy, leading to chronic gastrointestinal complications with substantial mortality. Adipose-derived Mesenchymal Stromal Cells (Ad-MSCs) represent a promising strategy to reduce intestinal lesions. However, systemic administration of Ad-MSCs results in low cell engraftment within the injured tissue. Biomaterials, able to encapsulate and withstand Ad-MSCs, can overcome these limitations. A silanized hydroxypropylmethyl cellulose (Si-HPMC) hydrogel has been designed and characterized for injectable cell delivery using the operative catheter of a colonoscope. We demonstrated that hydrogel loaded-Ad-MSCs were viable, able to secrete trophic factors and responsive to the inflammatory environment. In a rat model of radiation-induced severe colonic damage, Ad-MSC + Si-HPMC improve colonic epithelial structure and hyperpermeability compared with Ad-MSCs injected intravenously or locally. This therapeutic benefit is associated with greater engraftment of Si-HPMC-embedded Ad-MSCs in the irradiated colonic mucosa. Moreover, macrophage infiltration near the injection site was less pronounced when Ad-MSCs were embedded in the hydrogel. Si-HPMC induces modulation of chemoattractant secretion by Ad-MSCs that could contribute to the decrease in macrophage infiltrate. Si-HPMC is suitable for cell delivery by colonoscopy and induces protection of Ad-MSCs in the tissue potentiating their therapeutic effect and could be proposed to patients suffering from colon diseases.

Adipose-Derived Mesenchymal Stromal Cells Improve the Healing of Colonic Anastomoses Following High Dose of Irradiation Through Anti-Inflammatory and Angiogenic Processes.

Cancer patients treated with radiotherapy (RT) could develop severe late side effects that affect their quality of life. Long-term bowel complications after RT are mainly characterized by a transmural fibrosis that could lead to intestinal obstruction. Today, surgical resection is the only effective treatment. However, preoperative RT increases the risk of anastomotic leakage. In this study, we attempted to use mesenchymal stromal cells from adipose tissue (Ad-MSCs) to improve colonic anastomosis after high-dose irradiation. MSCs were isolated from the subcutaneous fat of rats, amplified in vitro, and characterized by flow cytometry. An animal model of late radiation side effects was induced by local irradiation of the colon. Colonic anastomosis was performed 4 wk after irradiation. It was analyzed another 4 wk later (i.e., 8 wk after irradiation). The Ad-MSC-treated group received injections several times before and after the surgical procedure. The therapeutic benefit of the Ad-MSC treatment was determined by colonoscopy and histology. The inflammatory process was investigated using Fluorine-182-Fluoro-2-Deoxy-d-Glucose Positron Emission Tomography and Computed Tomography (18F-FDG-PET/CT) imaging and macrophage infiltrate analyses. Vascular density was assessed using immunohistochemistry. Results show that Ad-MSC treatment reduces ulcer size, increases mucosal vascular density, and limits hemorrhage. We also determined that 1 Ad-MSC injection limits the inflammatory process, as evaluated through 18F-FDG-PET-CT (at 4 wk), with a greater proportion of type 2 macrophages after iterative cell injections (8 wk). In conclusion, Ad-MSC injections promote anastomotic healing in an irradiated colon through enhanced vessel formation and reduced inflammation. This study also determined parameters that could be improved in further investigations.

Bowel Radiation Injury: Complexity of the Pathophysiology and Promises of Cell and Tissue Engineering.

Ionizing radiation is effective to treat malignant pelvic cancers, but the toxicity to surrounding healthy tissue remains a substantial limitation. Early and late side effects not only limit the escalation of the radiation dose to the tumor but may also be life-threatening in some patients. Numerous preclinical studies determined specific mechanisms induced after irradiation in different compartments of the intestine. This review outlines the complexity of the pathogenesis, highlighting the roles of the epithelial barrier in the vascular network, and the inflammatory microenvironment, which together lead to chronic fibrosis. Despite the large number of pharmacological molecules available, the studies presented in this review provide encouraging proof of concept regarding the use of mesenchymal stromal cell (MSC) therapy to treat radiation-induced intestinal damage. The therapeutic efficacy of MSCs has been demonstrated in animal models and in patients, but an enormous number of cells and multiple injections are needed due to their poor engraftment capacity. Moreover, it has been observed that although MSCs have pleiotropic effects, some intestinal compartments are less restored after a high dose of irradiation. Future research should seek to optimize the efficacy of the injected cells, particularly with regard to extending their life span in the irradiated tissue. Moreover, improving the host microenvironment, combining MSCs with other specific regenerative cells, or introducing new tissue engineering strategies could be tested as methods to treat the severe side effects of pelvic radiotherapy.

Development of mandibular osteoradionecrosis in rats: Importance of dental extraction.

OBJECTIVES: To develop an animal model of mandibular osteoradionecrosis (ORN) using a high-energy radiation source (as used in human therapeutics) and to assess the role of tooth extraction on ORN development. MATERIALS AND METHODS (STUDY DESIGN): Ten animals were irradiated with a single 35- or 50-Gy dose. Three weeks later, the second left mandibular molar was extracted from three animals in each group. Nine weeks after irradiation, the animals were euthanized, with an injection of contrast agent in the bloodstream to highlight vascularization. Mandibles were harvested and studied using micro-CT, histology, tartrate-resistant acid phosphatase activity and scanning electron microscopy. RESULTS: This study demonstrates that a single 50-Gy dose associated with molar extraction is necessary for ORN development. In these conditions, absence of healing of the mucosa and bone, dental effects, fibrosis, an increase in osteoclast activity and a decrease in vascularization were observed. We also determined that molar extraction increases the impact of the cellular effects of radiation. CONCLUSION: The mandibular ORN animal model was validated after 50-Gy irradiation and molar extraction. The results of this study therefore support an animal ORN model and tissue engineering strategies will now be developed to regenerate bone for patients with head and neck cancer.

TH17 predominant T-cell responses in radiation-induced bowel disease are modulated by treatment with adipose-derived mesenchymal stromal cells.

Radiation proctitis is an insidious disease associated with substantial morbidity and mortality. It may develop following the treatment of several cancers by radiotherapy when normal colorectal tissues are present in the irradiation field. There is no unified approach for the assessment and treatment of this disease, partly due to insufficient knowledge about the mechanism involved in the development of radiation proctitis. However, unresolved inflammation is hypothesized to have an important role in late side effects. This study aimed to analyse the involvement of specific immunity in colorectal damage developing after localized irradiation, and evaluate the benefit of immunomodulatory mesenchymal stromal cells isolated from adipose tissue (Ad-MSCs) for reduction of late side effects. Our experimental model of colorectal irradiation induced severe colonic mucosal damage and fibrosis that was associated with T-cell infiltration. Immune cell activation was investigated; adoptive transfer of T cells in nude rats showed stronger colonization by T cells isolated from irradiated rats. The predominant role of T cells in late radiation-induced damage and regeneration processes was highlighted by in vivo depletion experiments. Treatments using Ad-MSCs reduced T-cell infiltration in the colon and reduced established colonic damage as measured by histological score, functional circular muscle contractibility, and collagen deposition. Here, we have demonstrated for the first time the predominance of the TH17 population compared to TH1 and TH2 in radiation-induced bowel disease, and that this is reduced after Ad-MSC treatment. Additionally, we demonstrated in vitro that IL17 acts directly on colonic smooth muscle cells to induce expression of pro-inflammatory genes that could participate in the development of radiation-induced injury. Our data demonstrate that the TH17 population is specifically induced during development of radiation-induced side effects in the colon. Moreover, Ad-MSC treatment modulates the TH17 population and reduces the extracellular matrix remodelling process induced following irradiation.

Persistent visceral allodynia in rats exposed to colorectal irradiation is reversed by mesenchymal stromal cell treatment.

Each year, millions of people worldwide are treated for primary or recurrent pelvic malignancies, involving radiotherapy in almost 50% of cases. Delayed development of visceral complications after radiotherapy is recognized in cancer survivors. Therapeutic doses of radiation may lead to the damage of healthy tissue around the tumor and abdominal pain. Because of the lack of experimental models, the underlying mechanisms of radiation-induced long-lasting visceral pain are still unknown. This makes managing radiation-induced pain difficult, and the therapeutic strategies proposed are mostly inefficient. The aim of our study was to develop an animal model of radiation-induced visceral hypersensitivity to (1) analyze some cellular and molecular mechanisms involved and (2) to test a therapeutic strategy using mesenchymal stromal cells (MSCs). Using a single 27-Grays colorectal irradiation in rats, we showed that such exposure induces a persistent visceral allodynia that is associated with an increased spinal sensitization (enhanced p-ERK neurons), colonic neuroplasticity (as increased density of substance P nerve fibers), and colonic mast cell hyperplasia and hypertrophy. Mast cell stabilization by ketotifen provided evidence of their functional involvement in radiation-induced allodynia. Finally, intravenous injection of 1.5 million MSCs, 4 weeks after irradiation, induced a time-dependent reversion of the visceral allodynia and a reduction of the number of anatomical interactions between mast cells and PGP9.5+ nerve fibers. Moreover, unlike ketotifen, MSC treatment has the key advantage to limit radiation-induced colonic ulceration. This work provides new insights into the potential use of MSCs as cellular therapy in the treatment of pelvic radiation disease.

Mesenchymal stem cell therapy stimulates endogenous host progenitor cells to improve colonic epithelial regeneration.

Patients who undergo pelvic radiotherapy may develop severe and chronic complications resulting from gastrointestinal alterations. The lack of curative treatment highlights the importance of novel and effective therapeutic strategies. We thus tested the therapeutic benefit of mesenchymal stem cells (MSC) treatment and proposed molecular mechanisms of action. MSC efficacy was tested in an experimental model of radiation-induced severe colonic ulceration histologically similar to that observed in patients. In this model, MSC from bone marrow were administered intravenously, immediately or three weeks (established lesions) after irradiation. MSC therapy reduces radiation-induced colonic ulceration and increases animal survival. MSC treatment induces therapeutic efficacy whatever the time of cell infusion. Infused-MSC engraft in the colon but also increase endogenous MSC mobilization in blood that have lasting benefits over time. In vitro analysis demonstrates that the MSC effect is mediated by paracrine mechanisms through the non-canonical WNT (Wingless integration site) pathway. In irradiated rat colons, MSC treatment increases the expression of the non-canonical WNT4 ligand by epithelial cells. The epithelial regenerative process is improved after MSC injection by stimulation of colonic epithelial cells positive for SOX9 (SRY-box containing gene 9) progenitor/stem cell markers. This study demonstrates that MSC treatment induces stimulation of endogenous host progenitor cells to improve the regenerative process and constitutes an initial approach to arguing in favor of the use of MSC to limit/reduce colorectal damage induced by radiation.

Characterization and histological localization of multipotent mesenchymal stromal cells in the human postnatal thymus.

The aim of this work was to characterize multipotent mesenchymal stromal cells (MSCs) in the postnatal human thymus and to localize these MSCs in the organ. Adherent cells isolated from thymus samples were characterized by cell-surface antigen expression. This showed that adherent cells have a MSC profile as assessed by the expression of CD73 and CD105 markers and the lack of CD45 expression. These cells are able to differentiate in vitro into adipocytes, osteoblasts, and chondrocytes and to inhibit mixed lymphocyte reaction. This indicates that isolated cells have all of the characteristics of MSC. The fibroblast colony-forming unit (CFU-F) assay was used to determine their frequency in the postnatal thymus. This frequency was 60.9 +/- 14.8 CFU-F per 1 x 10(5) freshly isolated mononuclear cells. Moreover, taking advantage of CD34 and CD105 expression, immunohistological staining allowed us to localize MSC within interlobular trabeculae in close contact with the outer cortex. Polymerase chain reaction experiments indicated that thymic MSC expressed interleukin-7 and stromal cell-derived factor-1 messenger RNA. Overall, these results confirm previous findings of the presence in the adult human thymus of multipotent MSCs with a phenotype similar to adipose-derived adult stem cells. These results also show for the first time a histological localization of MSC in an organ. This suggests a possible role of thymic MSC in intrathymic differentiation.

Immunosuppressive effects of mesenchymal stem cells: involvement of HLA-G.

INTRODUCTION: Mesenchymal stem cells (MSCs) possess unique immunomodulatory properties. They are able to suppress allogenic T-cell response and modify maturation of antigen-presenting cells. Their role in the treatment of severe graft versus host disease has been reported. The underlying molecular mechanisms of immunosuppression are currently being investigated. Histocompatibility locus antigen (HLA)-G is a nonclassical major histocompatibility complex class I antigen with strong immune-inhibitory properties. METHODS: We studied the role of HLA-G on MSC-induced immunosuppression. The expression of HLA-G on human MSCs cultured alone and in mixed lymphocytes reaction (MSC/MLR) was analyzed. RESULTS: We found that HLA-G can be detected on MSCs by real-time reverse-phase polymerase chain reaction, immunofluorescence, flow cytometry (52.4+/-3.6%), and enzyme-linked immunosorbent assay in the supernatant (38.7+/-5.2 ng/mL). HLA-G protein expression is constitutive and the level is not modified upon stimulation by allogenic lymphocytes in MSC/MLR. The functional role of HLA-G protein expressed by MSCs was analyzed using the 87G anti-HLA-G blocking antibody in a MSC/MLR. We found that blocking HLA-G molecule significantly raised lymphocyte proliferation in MSC/MLR (35.5%, P=0.01). CONCLUSION: Our findings provide evidences supporting involvement of HLA-G in the immunosuppressive properties of MSCs. These results emphasize the potential application of MSCs as a relevant therapeutic candidate in transplantation.

Identification of IL-10 and TGF-beta transcripts involved in the inhibition of T-lymphocyte proliferation during cell contact with human mesenchymal stem cells.

Mesenchymal stem cells (MSC) inhibit the response of allogeneic T lymphocytes in culture. Because the mechanisms of this effect may differ according to the existence of cell contact, we investigated the differences in gene expression of inhibitory molecules during MSC-T lymphocyte coculture when cell contact does and does not occur. Human MSC and T lymphocytes were cultured together in standard and transwell cultures. MSC gene expression was analyzed by semiquantitative real-time RT-PCR. MSC elicited a high dose-dependent inhibition of T lymphocytes in cultures with cell contact, but inhibition occurred even without cell contact. In both cases, we observed significant upregulation of IDO, LIF, and HLA-G, along with downregulation of HGF and SDF1. In cultures with cell contact, IL-10 and TGF-beta transcripts were expressed in a significantly higher level than in cultures without this contact. Furthermore, in the latter, the increased inhibition of T-cell proliferation was positively correlated with IDO gene expression and negatively correlated with SDF1 gene expression. MSC appear to induce T-cell tolerance by two distinct mechanisms. The first of these, which does not require cell contact, induces expression of the tolerogenic genes IDO, LIF, and HLA-G. The second mechanism, which is contact dependent, modulates IL-10 and TGF-beta gene expression. These two mechanisms probably play separate roles in MSC-induced tolerance in allogeneic hematopoietic stem cell transplantation.

Human mesenchymal stem cells favour healing of the cutaneous radiation syndrome in a xenogenic transplant model.

It has been suggested that human mesenchymal stem cells (hMSC) could be used to repair numerous injured tissues. We have studied the potential use of hMSC to limit radiation-induced skin lesions. Immunodeficient NOD/SCID mice were locally irradiated to the leg (30 Gy, dose rate 2.7 Gy/min) using a (60)Co source to induce a severe skin lesion. Cultured bone marrow hMSC were delivered intravenously to the mice. The irradiated skin samples were studied for the presence of the human cells, the severity of the lesions and the healing process. Macroscopic analysis and histology results showed that the lesions were evolving to a less severe degree of radiation dermatitis after hMSC transplant when compared to irradiated non-transplanted controls. Clinical scores for the studied skin parameters of treated mice were significantly improved. A faster healing was observed when compared to untreated mouse. Immunohistology and polymerase chain reaction analysis provided evidence that the human cells were found in the irradiated area. These results suggest a possible use of hMSC for the treatment of the early phase of the cutaneous radiation syndrome. A successful transplant of stem cells and subsequent reduction in radiation-induced complication may open the road to completely new strategies in cutaneous radiation syndrome therapy.

Telomere-driven genomic instability in cancer cells.

Telomeres, the ends of linear chromosomes, play a major role in the maintenance of genome integrity. Telomerase or alternative lengthening of telomeres (ALT) mechanisms exist in most cancer cells in order to stabilize telomere length by the addition of telomeric repeats. Telomere loss can be dramatically mutagenic. Chromosomes lacking one telomere remain unstable until they are capped, generating chromosomal instability, gene amplification via breakage/fusion/bridge (B/F/B) cycles and resulting in chromosome imbalances. The chronology of the occurrence of gene amplification and chromosome imbalances detected in human tumors is still unknown. All of the aberrations that occur prior to, during or after activation of a telomere maintenance mechanism promote the development of cancer.