Modified Mesenchymal Stem Cells Research Articles

Neuroprotective effect of tanshinone IIA-modified mesenchymal stem cells in a lipopolysaccharide-induced neuroinflammation model

In this study, the neuroprotective potential of tanshinone IIA (TIIA)-modified mesenchymal stem cells (MSC) were investigated using a murine model of lipopolysaccharide (LPS)-induced neuroinflammation. The cognitive performance of the mice was assessed using the Y-maze and Morris water maze tests, while immunofluorescence and Western blot analyses were employed to evaluate the hippocampal expression of pertinent markers and inflammatory factors, respectively. The results from the behavioral experiments demonstrated discernible differences in learning and memory abilities between the model group and the control group (P < 0.05), confirming the successful induction of neuroinflammation. Both the MSC and TIIA-MSC groups exhibited enhancements in the cognitive abilities of neuroinflammatory mice, with the TIIA-MSC group demonstrating a more pronounced improvement (P < 0.01). Immunofluorescence analysis revealed significant activation of microglia in the model group, while the MSC and TIIA-MSC groups exhibited a reduction in hippocampal microglial activation, with the TIIA-MSC group displaying a more substantial decrease. A statistically significant difference in the expression levels of IL-1, IL-6, and TNF-α was observed between the model and control groups (P < 0.05), indicating that IL-1, IL-6, and TNF-α were downregulated in both the MSC and TIIA-MSC groups. Notably, the downregulatory effect was more prominent in the TIIA-MSC group (P < 0.01). Compared to MSC treatment alone, the administration of TIIA-modified MSC demonstrated a superior protective effect against lipopolysaccharide-induced neuroinflammation. These findings underscore the potential therapeutic efficacy of TIIA-modified MSC in mitigating neuroinflammatory responses.

Enhanced tumor targeting and timely viral release of mesenchymal stem cells/oncolytic virus complex due to GRP78 and inducible E1B55K expressions greatly increase the antitumor effect of systemic treatment

Systemic delivery of oncolytic viruses has been widely regarded as an impractical option for antitumor treatment. Here, we selected two target genes as leading components, and significant therapeutic effects were obtained by simultaneously reducing the expression of transforming growth factor β 1 (TGF-β1) and heat shock protein 27 (HSP27) in various cancer cell types. Downregulation of HSP27 reduced the cellular levels of tumor progression-related proteins, and the simultaneous downregulation of HSP27 and TGF-β1 increased tumor cell death beyond that observed with TGF-β1 downregulation alone. To increase the potential for systemic administration, we generated modified mesenchymal stem cells (MSCs) to act as oncolytic adenovirus factories and carriers and assessed bioavailability in tumors after MSC injection. The MSCs were modified to express 78-kDa glucose-regulated protein (GRP78) and adenovirus early-region 1B 55 kDa (E1B55K). The tightly controlled inducible system permitted selective timing of viral release from carrier MSCs within the tumor. This approach significantly improved viral production, tumor targeting, timely viral release at the tumor site, and antitumor efficacy of the oncolytic adenovirus. These combined results demonstrate that engineered MSCs can significantly enhance the antitumor effects of oncolytic viruses without adverse safety issues, which may greatly extend the clinical applicability of oncolytic adenoviruses.

Unveiling the improved targeting migration of mesenchymal stem cells with CXC chemokine receptor 3-modification using intravital NIR-II photoacoustic imaging

BackgroundTherapy with genetically modified mesenchymal stem cells (MSCs) has clinical translation promise. Optimizing the targeting migratory ability of MSCs relies on accurate imaging of the distribution and extravasation kinetics of MSCs, and the corresponding imaging results could be used to predict therapeutic outcomes and guide the optimization of the treatment program. Among the different imaging modalities, second near-infrared (NIR-II) optical-resolution photoacoustic microscopy (OR-PAM) has merits, including a fine resolution, a deep penetration, a high sensitivity, and a large signal-to-background ratio. It would be an ideal candidate for precise monitoring of MSCs, although it has not been tested for this purpose so far.ResultsPenetrating peptide-decorated conjugated polymer nanoparticles (TAT-CPNPs) with strong NIR-II absorbance were used to label chemokine-receptor genetically modified MSCs, which were subsequently evaluated under intravital NIR-II OR-PAM regarding their targeting migratory ability. Based on the upregulation of chemokine (C-X-C motif) ligand 10 in the inflamed ears of contact hypersensitivity mice, MSCs with overexpression of corresponding receptor, chemokine (C-X-C motif) receptor 3 (Cxcr3) were successfully generated (MSCCxcr3). TAT-CPNPs labeling enabled NIR-II photoacoustic imaging to discern MSCCxcr3 covered by 1.2 cm of chicken breast tissue. Longitudinal OR-PAM imaging revealed enhanced inflammation-targeting migration of MSCCxcr3 over time attributed to Cxcr3 gene modification, which was further validated by histological analysis.ConclusionsTAT-CPNPs-assisted NIR-II PA imaging is promising for monitoring distribution and extravasation kinetics of MSCs, which would greatly facilitate optimizing MSC-based therapy.Graphical

Calcitonin gene related peptide modified mesenchymal stem cells reduce restenosis after carotid balloon injury in rats

This work aimed to investigate the effects of calcitonin gene-related peptide (CGRP)-modified mesenchymal stem cells (MSCs) on vascular stenosis in carotid balloon-injured rats. The CGRP gene labeled with EGFP was transfected into bone marrow MSCs, and CGRP protein expression in MSCs was confirmed by immunofluorescence assays. A rat carotid balloon injury model was established using a surgical method. CGRP-modified MSCs were orthotopically transplanted into the injured area of the rats. At 28 days after cell transplantation, EGFP and CD31 expression was detected by immunofluorescence staining. Hematoxylin-eosin (H&E) staining was used to detect the intima/media area of the injured carotid artery stenosis site, and the expression of proliferating cell nuclear antigen (PCNA) was detected by immunohistochemistry. MSCs from rat bone marrow positively expressed CD29 and negatively expressed CD45. In vivo immunofluorescence staining showed that EGFP expression was significantly increased at the vascular injury site of rats transplanted with MSC_CGRP compared with the control group on the 28th day after cell transplantation and that the damaged vessels exhibited continuous CD31 expression. H&E staining showed that the vascular intimal proliferation area of rats transplanted with MSC_CGRP was significantly reduced compared with that of other groups. Furthermore, the immunohistochemistry results showed that PCNA expression in the endothelium of the MSC_CGRP group was lower than that of the other groups. Therefore, MSCs transfected with the CGRP gene can express the CGRP protein, and the implantation of CGRP-modified MSCs at the damaged site after carotid balloon-induced injury can reduce the neointimal area.

Mesenchymal stem cells modified by FGF21 and GLP1 ameliorate lipid metabolism while reducing blood glucose in type 2 diabetic mice

ObjectiveThe purpose of this study was to investigate the therapeutic effects of genetically modified mesenchymal stem cells (MSCs) in the treatment of type 2 diabetes mellitus (T2DM) in order to identify a new method for treating diabetes that differs from traditional medicine and to provide a new means by which to fundamentally improve or treat diabetes.MethodsMSCs derived from adipose tissue were modified to overexpress FGF21 and GLP1, which was achieved through lentiviral particle transduction. The cells were transplanted into BKS.Cg-Dock7m+/+Leprdb/Nju mice (T2DM mouse model). Injections of physiological saline (0.1 mL) and liraglutide (0.5 mg/kg) were used as negative and positive controls, respectively. ELISA or Western blotting was used for protein analysis, and quantitative real-time PCR was used for gene expression analysis.ResultsGenetic modification had no effects on the morphology, differentiation ability, or immunophenotype of MSCs. Moreover, MSC-FGF21+GLP1 cells exhibited significantly increased secretion of FGF21 and GLP1. In the T2DM mouse model, the transplantation of MSC-FGF21+GLP1 cells ameliorated the changes in blood glucose and weight, promoted the secretion of insulin, enhanced the recovery of liver structures, and improved the profiles of lipids. Moreover, FGF21 and GLP1 exerted synergistic effects in the regulation of glucolipid metabolism by controlling the expression of insulin, srebp1, and srebp2.ConclusionStem cell treatment based on MSCs modified to overexpress the FGF21 and GLP1 genes is an effective approach for the treatment of T2DM.

Preclinical Studies and Clinical Trials with Mesenchymal Stem Cell for Demyelinating Diseases: A Systematic Review.

Different studies have been performed to investigate stem cell administration as a promising tool for recovery of injured tissue in multiple sclerosis (MS), the most common demyelinating disease. In the present systematic review, the electronic databases of PubMed and ScienceDirect were searched to screen English language studies published until April 2020. The results obtained from experimental autoimmune encephalomyelitis (EAE) animals revealed that modified mesenchymal stem cells (MSCs) transplantation was associated with remyelination, inflammation suppression and oligodendrocyte precursor cells regeneration. Clinical trials indicated that 70% of the patients with MS showed disease stabilization following MSCs administration. Although MSC therapy has showed to be effective in the improvement of some patients with MS, designing larger placebo-controlled clinical trials with MSCs expressing immune- regulators or MSCs-exosomes may provide a novel viewpoint in the treatment of MS.

Efficacy and Safety of Intravenous Mesenchymal Stem Cells for Ischemic Stroke.

To test whether autologous modified mesenchymal stem cells (MSCs) improve recovery in patients with chronic major stroke. In this prospective, open-label, randomized controlled trial with blinded outcome evaluation, patients with severe middle cerebral artery territory infarct within 90 days of symptom onset were assigned, in a 2:1 ratio, to receive preconditioned autologous MSC injections (MSC group) or standard treatment alone (control group). The primary outcome was the score on the modified Rankin Scale (mRS) at 3 months. The secondary outcome was to further demonstrate motor recovery. A total of 39 and 15 patients were included in the MSC and control groups, respectively, for the final intention-to-treat analysis. Mean age of patients was 68 (range 28-83) years, and mean interval between stroke onset to randomization was 20.2 (range 5-89) days. Baseline characteristics were not different between groups. There was no significant difference between the groups in the mRS score shift at 3 months (p = 0.732). However, secondary analyses showed significant improvements in lower extremity motor function in the MSC group compared to the control group (change in the leg score of the Motricity Index, p = 0.023), which was notable among patients with low predicted recovery potential. There were no serious treatment-related adverse events. IV application of preconditioned, autologous MSCs with autologous serum was feasible and safe in patients with chronic major stroke. MSC treatment was not associated with improvements in the 3-month mRS score, but we did observe leg motor improvement in detailed functional analyses. This study provides Class III evidence that autologous MSCs do not improve 90-day outcomes in patients with chronic stroke. NCT01716481.

Overexpression of pigment epithelium-derived factor in placenta-derived mesenchymal stem cells promotes mitochondrial biogenesis in retinal cells

AbstractPigment epithelium-derived factor (PEDF) plays a role in protecting retinal pigment epithelial (RPE) cells from oxidative stress (OS), a causative factor of RPE cell death. Genetically modified mesenchymal stem cells (MSCs) can be used to treat critical and incurable retinal diseases. Here, we overexpressed PEDF in placenta-derived MSCs (PD-MSCsPEDF, PEDF+) using a nonviral gene delivery system and evaluated the characteristics of PD-MSCsPEDF and their potential regenerative effects on RPE cells damaged by H2O2-induced OS. PD-MSCsPEDF maintained their stemness, cell surface marker, and differentiation potential characteristics. Compared to naive cells, PD-MSCsPEDF promoted mitochondrial respiration by enhancing biogenesis regulators (e.g., NRF1, PPARGC1A, and TFAM) as well as antioxidant enzymes (e.g., HMOXs, SODs, and GPX1). Compared to OS-damaged RPE cells cocultured with naive cells, OS-damaged RPE cells cocultured with PD-MSCsPEDF showed PEDF upregulation and VEGF downregulation. The expression levels of antioxidant genes and RPE-specific genes, such as RPE65, RGR, and RRH, were significantly increased in RPE cells cocultured with PD-MSCsPEDF. Furthermore, OS-damaged RPE cells cocultured with PD-MSCsPEDF had dramatically enhanced mitochondrial functions, and antiapoptotic effects improved due to cell survival signaling pathways. In the H2O2-induced retinal degeneration rat model, compared to administration of the naive counterpart, intravitreal administration of PD-MSCsPEDF alleviated proinflammatory cytokines and restored retinal structure and function by increasing PEDF expression and decreasing VEGF expression. Intravitreal administration of PD-MSCsPEDF also protected retinal degeneration against OS by increasing antioxidant gene expression and regulating the mitochondrial ROS levels and biogenesis. Taken together, PEDF overexpression in PD-MSCs improved the mitochondrial activities and induced OS-damaged RPE cell regeneration by regulating the oxidative status and mitochondrial biogenesis in vitro and in vivo. These data suggest that genetic modification of PEDF in PD-MSCs might be a new cell therapy for the treatment of retinal degenerative diseases.Overexpression of pigment epithelium-derived factor (PEDF) in placenta-derived mesenchymal stem cells (PD-MSCs) improved the mitochondrial activities, and induced regeneration of oxidative stress-damaged RPE through regulating oxidative status and mitochondrial biogenesis. Therefore, genetic modification of PD-MSCs with PEDF might be a new cell therapy for treatment of retinal degenerative diseases.

Heme oxygenase-1 modified mesenchymal stem cells in the treatment of ischemia-reperfusion injury in lung transplantation

Objective To explore the protective mechanism of Heme Oxygenase-1 (HO-1) modified mesenchymal stem cells (MSC) in the treatment of lung transplantation ischemia-reperfusion injury (IRI). Methods Sixty SD rats were randomly divided into Surgical control (Ⅰ), simple MSCs (Ⅱ), empty vector-MSCs (Ⅲ), HO-1 modified MSCs (Ⅳ) were established. After establishing orthotopic left lung transplantation model in rats, group Ⅱ, Ⅲ and Ⅳ were injected with MSCs, empty vector-MSCs and HO-1-MSCs from pulmonary artery fine needles of recipient rats. Four hours later, partial pressure of oxygen (PaO2), partial pressure of carbon dioxide (PaCO2), tumor necrosis factor-alpha (TNF-α), Interleukin-10 (IL-10), wet/dry ratio (W/D), superoxide dismutase (SOD), malondialdehyde (MDA) and myeloperoxidase (MPO) were measured. Histopathological examination. Detection of HO-1-mRNA expression in transplants by reverse transcription polymerase chain reaction (RT-PCR). Multigroup mean comparison using One-way ANOVA. Results PaO2: Group Ⅰ, Ⅱ, Ⅲ and Ⅳ were (70.40±4.64), (87.53±4.01), (87.27±4.06), (96.80±647) mmHg (1 mmHg=0.133 kPa) gradually increased (F=75.578, P 0.05). PaCO2: group Ⅰ, Ⅱ, Ⅲ and Ⅳ were (32.67±3.31), (27.80±2.98), (27.60±2.41), (19.87±3.02) mmHg gradually decreased (F=48.286, P 0.05). W/D: Group Ⅰ, Ⅱ, Ⅲ and Ⅳ were (4.96±0.22), (4.04±0.28), (4.11±0.31), (3.85±0.27) gradually decreased (F=48.992, P 0.05). MDA content: In group Ⅰ, Ⅱ, Ⅲ and Ⅳ, the levels of MDA were (126.80±11.14), (114.27±16.45), (114.06±17.68), (102.40±14.21) nmol/g gradually decreased (F=6.551, P 0.05). MPO content: group Ⅰ, Ⅱ, Ⅲ and Ⅳ were (53.51±3.06), (45.07±3, 32), (44.45±3.37), (40.60±2.47) U/g decreased gradually (F=46.907, P 0.05). SOD activity: In group Ⅰ, Ⅱ, Ⅲ and Ⅳ, SOD activity increased gradually [(304.60±34.74), (329.87±19.44), (329.87±19.44), (338.73±15.77) U/g, (F=6.038, P 0.05). TNF-α: In group Ⅰ, Ⅱ, Ⅲ and Ⅳ, the levels of TNF-αwere (61.56±2.83), (48.32±4.11), (48.53±4.20), (36.77±3.79) ng/L gradually decreased (F=108.215, P 0.05). IL-10: The levels of IL-10 in group Ⅰ, Ⅱ, Ⅲ and Ⅳ were (23.43±2.37), (34.96±3.04), (35.48±3.00), (41.16±3.12) ng/L gradually increased (F=98.751, P 0.05). Under light microscopy, neutrophils infiltrated pulmonary interstitium and inflammatory exudation was seen in alveolar cavity. Inflammation in group Ⅳ was the lightest in each group. The amplification bands of HO-1 mRNA in group IV were significantly stronger than other groups. Conclusion HO-1 gene modified MSCs have synergistic anti-injury effect in the treatment of lung transplantation IRI. Key words: Heme oxygenase-1; Mesenchymal stem cells; Lung transplantation ischemia-reperfusion injury

Dynamics of host and graft after cell sheet transplantation: Basic study for the application of amyotrophic lateral sclerosis

Stem cells offer great hope for the therapy of neurological disorders. Using a human artificial chromosome (HAC), we generated modified mesenchymal stem cells (MSCs), termed HAC-MSC that express 3 growth factors and 2 marker proteins including luciferase, and previously demonstrated that intrathecal administration of HAC-MSCs extended the lifespan in a mouse model of amyotrophic lateral sclerosis (ALS). However, donor cells disappeared rapidly after transplantation. To overcome this poor survival, we transplanted the HAC-MSCs as a sheet structure which retained the extracellular matrix. We investigated, here, whether cell sheet showed a longer survival than intrathecal administration. Also, the therapeutic effects on ALS model mice were examined.In vivo imaging showed that luciferase signals increased immediately after transplantation up to 7 days, and these signals were sustained for up to 14 days. In contrast, following intrathecal administration, signals were drastically decreased by day 3. Moreover, cell sheet transplantation successfully prolonged the survival of donor HAC-MSCs. Cell sheet transplantation increased the level of p-Akt at the graft area. Pathologically, none of the donor cells differentiated into neurons, astrocytes or microglial cells. When the cell sheet was transplanted into ALS model mice, there was an encouraging trend in the delayed onset of symptoms and increased lifespan. If each group was subdivided into rapid and slow progressors based on cut-off values for respective median survival, the survival of rapid progressors differed significantly between groups (treated vs. sham-operated = 145.4 ± 1.4 vs. 139.2 ± 1.2). The effect of HAC-MSC sheet transplantation still has a temporally narrow therapeutic window.Further improvement could be achieved by optimization of the transplantation conditions, e.g. co-transplantation of HAC-MSCs with endothelial progenitor cells.

Research progress in mesenchymal stem cells modified by Heme oxygenase 1

To review the literature reports on research progress of Heme oxygenase 1 (HO-1) modified mesenchymal stem cells (MSCs). The significance, effects, and related mechanism of HO-1 modification of MSCs were summarized by consulting the related literatures and reports of HO-1 modification of MSCs. HO-1 modification of MSCs has important research value. It can effectively enhance the anti-oxidative stress and anti-apoptotic properties of MSCs in complex internal environment after transplantation into vivo. It can also effectively enhance the immune regulation function of MSCs. It can improve the anti-injury, repair, and immune regulation effect of MSCs in various disease models and research fields. The basic research of HO-1 modified MSCs has made remarkable progress, which is expected to be applied in clinical trials and provide theoretical basis and reference value for stem cell therapy.

Delivery of mesenchymal stem cells-derived extracellular vesicles with enriched miR-185 inhibits progression of OPMD

Oral leukoplakia is one of the most common oral potentially malignant disorders (OPMDs) and its malignant transformation to oral cancer is highly associated with chronic inflammation. Extracellular vesicles (EVs) or exosome-delivered microRNAs modulate inflammatory responses and alleviate irritations that predisposes to cancer. We previously reported that microRNA-185 (miR-185) was significantly decreased in the buccal tissue of patients with oral cancer. In this study, we utilized genetically modified mesenchymal stem cells (MSCs) derived EVs with high expression of miR-185 to pasted MSC-EV-miR-185 on buccal lesions in dimethylbenzanthracene (DMBA) induced OPMD model. We found that treatment with MSC-EV-miR-185 remarkably attenuated inflammation severity and significantly decreased the incidence and the number of dysplasia in the OPMD tissue. Immunohistochemistry showed significantly decreased expression of proliferation marker PCNA and angiogenic marker CD31 in the lesion treated with MSC-EV-miR-185. Furthermore, miR-185 specifically targeted Akt genes by promoting activation of the apoptotic pathway, confirmed by the increased levels of activated caspase-3 and 9. In conclusion, genetically modified MSC-derived EVs enriched with miR-185 alleviate inflammatory response, inhibit cell proliferation and angiogenesis, and induce cell apoptosis, suggesting that their potential role as a novel therapeutic option for OPMD.

Stem Cell Based Drug Delivery for Protection of Auditory Neurons in a Guinea Pig Model of Cochlear Implantation.

Background: The success of a cochlear implant (CI), which is the standard therapy for patients suffering from severe to profound sensorineural hearing loss, depends on the number and excitability of spiral ganglion neurons (SGNs). Brain-derived neurotrophic factor (BDNF) has a protective effect on SGNs but should be applied chronically to guarantee their lifelong survival. Long-term administration of BDNF could be achieved using genetically modified mesenchymal stem cells (MSCs), but these cells should be protected – by ultra-high viscous (UHV-) alginate (‘alginate-MSCs’) – from the recipient immune system and from uncontrolled migration.Methods: Brain-derived neurotrophic factor-producing MSCs were encapsulated in UHV-alginate. Four experimental groups were investigated using guinea pigs as an animal model. Three of them were systemically deafened and (unilaterally) received one of the following: (I) a CI; (II) an alginate-MSC-coated CI; (III) an injection of alginate-embedded MSCs into the scala tympani followed by CI insertion and alginate polymerization. Group IV was normal hearing, with CI insertion in both ears and a unilateral injection of alginate-MSCs. Using acoustically evoked auditory brainstem response measurements, hearing thresholds were determined before implantation and before sacrificing the animals. Electrode impedance was measured weekly. Four weeks after implantation, the animals were sacrificed and the SGN density and degree of fibrosis were evaluated.Results: The MSCs survived being implanted for 4 weeks in vivo. Neither the alginate-MSC injection nor the coating affected electrode impedance or fibrosis. CI insertion with and without previous alginate injection in normal-hearing animals resulted in increased hearing thresholds within the high-frequency range. Low-frequency hearing loss was additionally observed in the alginate-injected and implanted cochleae, but not in those treated only with a CI. In deafened animals, the alginate-MSC coating of the CI significantly prevented SGN from degeneration, but the injection of alginate-MSCs did not.Conclusion: Brain-derived neurotrophic factor-producing MSCs encapsulated in UHV-alginate prevent SGNs from degeneration in the form of coating on the CI surface, but not in the form of an injection. No increase in fibrosis or impedance was detected. Further research and development aimed at verifying long-term mechanical and biological properties of coated electrodes in vitro and in vivo, in combination with chronic electrical stimulation, is needed before the current concept can be tested in clinical trials.

11 - A phase I trial of mesenchymal stem cells transfected with a plasmid secreting interferon beta in advanced ovarian cancer

Background: Treatment of ovarian cancer with a platinum-taxane is effective but associated with significant relapse. We demonstrated genetically modified mesenchymal stem cells (MSC) preferentially engraft and secrete gene products such as interferon beta (IFNB) at tumor sites, resulting in complete responses in animal models.

Enhanced effect of human mesenchymal stem cells expressing human TNF-αR-Fc and HO-1 gene on porcine islet xenotransplantation in humanized mice.

Porcine islet xenotransplantation is considered an attractive alternative treatment for type 1 diabetes mellitus. However, it is largely limited because of initial rejection due to Instant Blood-Mediated Inflammatory Reaction (IBMIR), oxidative stress, and inflammatory responses. Recently, soluble tumor necrosis factor-ɑ receptor type I (sTNF-αR) and heme oxygenase (HO)-1 genes (HO-1/sTNF-αR) have been shown to improve the viability and functionality of porcine islets after transplantation. In this study, genetically modified mesenchymal stem cells (MSCs) expressing the HO-1/sTNF-αR genes (HO-1/sTNF-αR-MSC) were developed using an adenoviral system, and porcine islet viability and function were confirmed by in vitro tests such as GSIS, AO/PI, and the ADP/ATP ratio after coculturing with HO-1/sTNF-αR-MSCs. Subsequently, isolated porcine islets were transplanted underneath the kidney capsule of diabetic humanized mice without MSCs, with MSCs or with HO-1/sTNF-αR-MSCs. According to the results, the HO-1/sTNF-αR-MSC-treated group exhibited improved survival of porcine islets and could reverse hyperglycemia more than porcine islets not treated with MSCs or islets cotransplanted with MSCs. Moreover, the HO-1/sTNF-αR-MSC group maintained its morphological characteristics and the insulin secretion pattern of transplanted porcine islets similar to endogenous islets in immunocompetent humanized mice. Our results suggest that HO-1/sTNF-αR-MSCs are efficient tools for porcine islet xenotransplantation, and this study may provide basic information forpre-clinical animal models and future clinical trials of porcine islet xenotransplantation.

EPO modified MSCs can inhibit asthmatic airway remodeling in an animal model.

There was no effective measures can be obtained at present to reverse or prevent airway remodeling. We investigated the therapeutic effect of Erythropoietin (EPO) gene modified mesenchymal stem cells (MSCs) on asthmatic airway remodeling and the possible underlied molecular mechanisms. EPO gene was transfected into MSCs via lentivirus vector. The transfected cells (EPO-MSCs) were identified by flow cytometry and the EPO secreting function was detected by PCR and Western blot. MSCs or EPO-MSCs were administrated to albumin (OVA)-induced chronic asthmatic mouse model via tail veins. The asthmatic phenotype was analyzed. Number of cells in bronchoalveolar lavage fluid (BALF) was counted using a hemocytometer. Histological findings of airways were evaluated by microscopic examination. The concentrations of interleukin 4(IL-4), interleukin 5(IL-5), and interleukin 13(IL-13) in lung homogenate were determined by ELISA. The activation state of transforming growth factor-β 1 (TGF-β1), Transforming growth factor beta-activated kinase 1 (TAK1), and p38 Mitogen Activated Protein Kinase (p38MAPK) signaling was detected by Real-Time PCR and Western blotting. EPO-MSCs were successfully constructed. EPO-MSCs showed a more potently suppressive effect on local asthmatic airway inflammation and the level of IL-4, IL-5, and IL-13 in lung tissue than MSCs. Moreover, the numbers of goblet cells, the thicknesses of smooth muscle layer, collagen density, percentage of proliferating cell nuclear antigen positive (PCNA+ ) mesenchymal cells, and von Willebrand factor positive(vWF+ ) vessels were also significantly inhibited by EPO-MSCs. Furthermore, EPO-MSCs could downregulate the expression of TGF-β1, TAK1, and p38MAPK in lung tissue both in mRNA level and in protein level. EPO gene modified MSCs may more efficiently attenuate asthmatic airway remodeling, which maybe related with the downregulation of TGF-β1-TAK1-p38MAPK pathway activity.

T Cell-Activating Mesenchymal Stem Cells as a Biotherapeutic for HCC

The outcome for advanced stage hepatocellular carcinoma (HCC) remains poor, highlighting the need for novel therapies. Genetically modified mesenchymal stem cells (MSCs) are actively being explored as cancer therapeutics due to their inherent ability to migrate to tumor sites. We reasoned that MSCs can be genetically modified to redirect T cells to Glypican-3 (GPC3)+ HCC, and genetically modified these with viral vectors encoding a GPC3/CD3 bispecific T cell engager (GPC3-ENG), a bispecifc T cell engager specific for an irrelevant antigen (EGFRvIII), and/or costimulatory molecules (CD80 and 41BBL). Coculture of GPC3+ cells, GPC3-ENG MSCs, and T cells resulted in T cell activation, as judged by interferon γ (IFNγ) production and killing of tumor cells by T cells. Modification of GPC3-ENG MSCs with CD80 and 41BBL was required for antigen-dependent interleukin-2 (IL-2) production by T cells and resulted in faster tumor cell killing by redirected T cells. In vivo, GPC3-ENG MSCs ± costimulatory molecules had antitumor activity in the HUH7 HCC xenograft model, resulting in a survival advantage. In conclusion, MSCs genetically modified to express GPC3-ENG ± costimulatory molecules redirect T cells to GPC3+ tumor cells and have potent antitumor activity. Thus, further preclinical exploration of our modified approach to GPC3-targeted immunotherapy for HCC is warranted.

Transdifferentiation of brain-derived neurotrophic factor (BDNF)-secreting mesenchymal stem cells significantly enhance BDNF secretion and Schwann cell marker proteins

The use of genetically modified mesenchymal stem cells (MSCs) is a rapidly growing area of research targeting delivery of therapeutic factors for neuro-repair. Cells can be programmed to hypersecrete various growth/trophic factors such as brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and nerve growth factor (NGF) to promote regenerative neurite outgrowth. In addition to genetic modifications, MSCs can be subjected to transdifferentiation protocols to generate neural cell types to physically and biologically support nerve regeneration. In this study, we have taken a novel approach by combining these two unique strategies and evaluated the impact of transdifferentiating genetically modified MSCs into a Schwann cell-like phenotype. After 8 days in transdifferentiation media, approximately 30-50% of transdifferentiated BDNF-secreting cells immunolabeled for Schwann cell markers such as S100β, S100, and p75NTR. An enhancement was observed 20 days after inducing transdifferentiation with minimal decreases in expression levels. BDNF production was quantified by ELISA, and its biological activity tested via the PC12-TrkB cell assay. Importantly, the bioactivity of secreted BDNF was verified by the increased neurite outgrowth of PC12-TrkB cells. These findings demonstrate that not only is BDNF actively secreted by the transdifferentiated BDNF-MSCs, but also that it has the capacity to promote neurite sprouting and regeneration. Given the fact that BDNF production remained stable for over 20 days, we believe that these cells have the capacity to produce sustainable, effective, BDNF concentrations over prolonged time periods and should be tested within an invivo system for future experiments.

Cranial bone regeneration via BMP-2 encoding mesenchymal stem cells

Cranial bone repair and regeneration via tissue engineering principles has attracted a great deal of interest from researchers during last decade. Here, within this study, 6 mm critical-sized bone defect regeneration via genetically modified mesenchymal stem cells (MSC) were monitored up to 4 months. Cranial bone repair and new bone formations were evaluated by histological staining and real time PCR analysis in five different groups including autograft and bone morphogenetic protein-2 (BMP-2) transfected MSC groups. Results presented here indicate a proper cranial regeneration in autograft groups and a prospering regeneration for hBMP-2 encoding mesenchymal stem cells.

254. Hepatocyte Growth Factor Gene Modification Enhances the Inflammation-Alleviating Effect of Umbilical Cord Derived Mesenchymal Stem Cells in a Bronchiolitis Obliterans Model

BACKGROUNDS: Bronchiolitis obliterans is a serious, steadily, noninfectious, and fatal pulmonary complication. Currently specific and effective therapeutics for prevention or reversal of bronchiolitis obliterans is lacking. In this study, we evaluated the therapeutic effect of hepatocyte growth factor (HGF) gene modified mesenchymal stem cells (MSCs) on bronchiolitis obliterans.METHODS: A mouse model of experimental bronchiolitis obliterans was established by subcutaneously transplanting the tracheas from C57BL/6 mice into Balb/C recipients. These model mice were administered saline, Ad-HGF-modified human umbilical cord-MSCs (MSCs-HGF) or Ad-Null-modified MSCs (MSCs-Null). The distribution of the transplanted cells was detected by using real-time PCR (rt-PCR). The therapeutic effects of MSCs-Null and MSCs-HGF were evaluated by using FACS for lymphocyte immunophenotype of spleen, ELISA and rt-PCR for cytokine expression, and histopathological analysis for the transplanted trachea.RESULTS: The administrated MSCs mainly distributed in the transplanted trachea, blood, liver and lung of the recipients; HGF gene modification could enhance the engraftment of MSCs. The histopathological recovery of allografts tracheas was improved significantly after MSCs-Null and MSCs-HGF treatment and the beneficial effects were particularly observed in MSCs-HGFtreated mice. Furthermore, the allo-transplantation induced immunophenotype disorders of the spleen cells, including Treg, Th1, Th2 and Th17 cells, were attenuated in both cell treated groups. MSCs-HGF treatment reduced expression and secretion of inflammation cytokines interferon-gamma and increased expression of anti-inflammatory cytokine IL-4 and IL-10. It also decreased the expression level of the profibrosis factor transforming growth factor-beta.b.CONCLUSIONS: Treatment of bronchiolitis obliterans with HGF gene modified MSCs results in reduction of local inflammation and promotion in recovery of allografts tracheas histopathology in a mouse model. These findings might provide an effective therapeutic strategy for bronchiolitis obliterans.