Autologous Marrow-derived Mesenchymal Stem Cells Research Articles

Transplantation of Mesenchymal Stem Cells Derived from Old Rats Improves Healing and Biomechanical Properties of Vaginal Tissue Following Surgical Incision in Aged Rats.

Pelvic floor dysfunction encompasses a group of disorders that negatively affect the quality of women's lives. These include pelvic organ prolapse (POP), urinary incontinence, and sexual dysfunction. The greatest risk factors for prolapse are increased parity and older age, with the largest group requiring surgical intervention being post-menopausal women over 65. Prolapse recurrence rates following surgery were reported to be as high as 30%. This may be attributed to ineffective healing in the elderly. Autologous stem cell transplantation during surgery may improve surgical results. In our previous studies, we showed that the transplantation of bone marrow-derived mesenchymal stem cells (MSCs) from young donor rats improved the healing of full-thickness vaginal surgical incision in the vaginal wall of old rats, demonstrated by both histological and functional analysis. In order to translate these results into the clinical reality of autologous MSC transplantation in elderly women, we sought to study whether stem cells derived from old donor animals would provide the same effect. In this study, we demonstrate that MSC transplantation attenuated the inflammatory response, increased angiogenesis, and exhibited a time-dependent impact on MMP9 localization. Most importantly, transplantation improved the restoration of the biomechanical properties of the vagina, resulting in stronger healed vaginal tissue. These results may pave the way for further translational studies focusing on the potential clinical autologous adjuvant transplantation of MSCs for POP repair for the improvement of surgical outcomes.

Autologous bone marrow derived mesenchymal stem cells are safe for the treatment of Achilles tendinopathy

Achilles tendinopathy is a disabling condition that affects more than 50% of runners. Pre-clinical studies in a large animal model of naturally-occurring tendinopathy similar to human Achilles tendinopathy has shown benefits of autologous bone marrow-derived mesenchymal stem cell (MSC) implantation. However, MSCs are advanced therapies medicinal products (ATMPs), with strict regulatory requirements. Guided by the regulator we carried out a first in man study to assess the safety and efficacy of autologous MSC injection in human patients with non-insertional Achilles tendinopathy. Ten patients, mean age 47 with mid-portion Achilles tendon pain and swelling for more than 6 months, underwent autologous cultured cell injections (median 12.2 × 106, range 5–19 × 106 cells) into their Achilles tendon. At 24 weeks follow-up, no serious adverse reactions or important medical events were observed. MOXFQ, EQ-5D-5L, and VISA-A scores improved clinically at 12 and 24 weeks. VAS pain improved increasingly at 6, 12 and 24 weeks. MOXFQ Pain and VISA-A Scores improved > 12 points from baseline to 24 weeks in 8 patients. Maximum anteroposterior tendon thickness as measured by greyscale US decreased by mean 0.8 mm at 24 weeks. This phase IIa study demonstrated the safety of autologous MSC injection for non-insertional Achilles tendinopathy and provides proof-of-concept of the technique in patients, all of whom had previously failed conservative treatments for chronic disease and leads the way for a larger randomised controlled trial.

Intracerebral Administration of Autologous Mesenchymal Stem Cells as HSV-TK Gene Vehicle for Treatment of Glioblastoma Multiform: Safety and Feasibility Assessment.

Widespread investigation has revealed the promising ability of suicidal genes in the treatment of glioma tumors; nevertheless, promoting their effects relies on the ability to apply suitable vehicles and techniques. In this study, the safety and feasibility of using bone marrow-derived mesenchymal stem cells (MSCs) in combination with prodrug for treatment of patients with primary and secondary glioblastoma multiform (GBM) was assessed. Five GBM patients were recruited. Following gross total resection of the tumor and adjuvant radiotherapy and chemotherapy, intracerebral injection of autologous MSCs transduced with lentivirus containing herpes simplex virus thymidine kinase (HSV-TK) was performed followed by intravenous administration of ganciclovir for 2weeks. The treatment was well tolerated by all patients. Mild-to-moderate fever, headache, and cerebrospinal fluid leukocytosis were evident in three, two, and one patient, respectively. The progression-free survival (PFS) and overall survival (OS) of patients were 95.79 ± 51.40 and 128.85 ± 48.81weeks, respectively. The 1-year PFS and OS were 60% and 100%, respectively, among our patients, and two patients had more than 3years of OS and more than 2years of PFS. It seems that intracerebral administration of bone marrow MSC containing the HSV-TK gene in combination with intravenous ganciclovir would be safe and feasible in the treatment of patients with GBM.

Retinal function after intravitreal injection of autologous bone marrow-derived mesenchymal stromal cells in advanced glaucoma

To report electroretinographic (ERG) findings in advanced glaucoma treated with a single intravitreal injection of bone marrow-derived mesenchymal stem cells (MSCs). Intravitreal injection of autologous MSCs (1 × 106 cells) was performed in 2 eyes from 2 patients with open-angle glaucoma in advanced stage of optic neuropathy (ClinicalTrials.gov, NCT02330978, 01.05.2015): cup/disk ratio worse than 0.9, visual field mean deviation index lower than -15dB, visual acuity of light perception, but controlled intraocular pressure. ERG tests were recorded at baseline and week 1, 4 and 48 after injection, using DTL electrodes following the ISCEV standard: After dark adaptation, ERG was elicited using white flashes of 0.01cd.s/m2 and 3.0cd.s/m2, followed by 10-min light adaptation (30cd/m2) and stimuli of 3.0cd.s/m2 and 30Hz flicker. Patients did not show improvement on visual acuity or visual field after treatment. At baseline, ERG responses showed typical findings for advanced glaucoma, with a- and b-wave amplitude and latency within normative range, but reduced photopic negative responses. No noteworthy changes were observed on ERG responses for both cases up to 1week after treatment, but at day 15, one patient showed retinal detachment with proliferative vitreoretinopathy and was removed from the trial. The other patient kept ERG responses stable throughout study period. Although no ERG response changes were observed after MSCs injection in one case, the complication observed on the second one, along with the lack of visual function improvement, warrants further studies involving modified MSCs to treat ocular disorders, including glaucoma. ClinicalTrials.gov, NCT02330978- missed in pdf.

Tonsil-derived stem cells as a new source of adult stem cells.

Located near the oropharynx, the tonsils are the primary mucosal immune organ. Tonsil tissue is a promising alternative source for the high-yield isolation of adult stem cells, and recent studies have reported the identification and isolation of tonsil-derived stem cells (T-SCs) from waste surgical tissue following tonsillectomies in relatively young donors (i.e., under 10 years old). As such, T-SCs offer several advantages, including superior proliferation and a shorter doubling time compared to bone marrow-derived mesenchymal stem cells (MSCs). T-SCs also exhibit multi-lineage differentiation, including mesodermal, endodermal (e.g., hepatocytes and parathyroid-like cells), and even ectodermal cells (e.g., Schwann cells). To this end, numbers of researchers have evaluated the practical use of T-SCs as an alternative source of autologous or allogenic MSCs. In this review, we summarize the details of T-SC isolation and identification and provide an overview of their application in cell therapy and regenerative medicine.

Towards Cell free Therapy of Premature Ovarian Insufficiency: Human Bone Marrow Mesenchymal Stem Cells Secretome Enhances Angiogenesis in Human Ovarian Microvascular Endothelial Cells.

Primary Ovarian Insufficiency (POI) refers to an ovarian loss of function in women under the age of 40. Unfortunately, currently, there is no effective treatment available for POI-related infertility. Alternatives such as the use of egg donations are culturally and ethically unacceptable to many couples. Human Bone marrow-derived Mesenchymal Stem Cells (MSCs) are known for their ability to differentiate into other cell types, once primed by the organ microenvironment. Importantly MSCs produce a vast array of bioactive factors many of them have been shown to enhance neovascularization in various tissues. Recently, preliminary data from our ongoing clinical trial revealed encouraging preliminary data after autologous MSC engraftment into the ovaries of 2 POI patients with durable elevation in serum estrogen levels and increase in size of treated ovaries sustained up to one-year post cell therapy.In this study, we investigated the action of the mechanisms of MSCs treatment on a POI ovary. We designed an in vitro study using MSC secretome and Human Ovarian Endothelial Cells (HOVECs) to understand the molecular mechanisms by which MSC mediates their angiogenic properties and regenerative effects. Human primary HOVECs were treatment with MSC secretome and examined by FACS for the expression of angiogenesis markers such as Endoglin, Tie-2, and VEGF. The formation of vessels was evaluated by using a 3D Matrigel tubulogenesis assay.We observed that the expression of proliferation marker Ki67 was significantly increased under treatment with MSC secretome in HOVEC cells (P4). MSCs secretome treatment also induced significantly higher expression of several angiogenic markers such as VEGFR2, Tie2/Tek, VE-Cadherin, Endoglin, and VEGF compared to matched control (P4). Furthermore, MSC secretome significantly increased the number of branching points in tubulogenesis assay (P4). Our study suggests that MSC secretome likely contains bioactive factors that can enhance ovarian angiogenesis. Further characterization of these factors can lead to novel therapeutic options for women with premature ovarian insufficiency and other related causes of female infertility.

Autologous Mesenchymal Stem Cell and Islet Cotransplantation: Safety and Efficacy.

Islet engraftment after transplantation is impaired by high rates of islet/β cell death caused by cellular stressors and poor graft vascularization. We studied whether cotransplantation of ex vivo expanded autologous bone marrow‐derived mesenchymal stem cells (MSCs) with islets is safe and beneficial in chronic pancreatitis patients undergoing total pancreatectomy with islet autotransplantation. MSCs were harvested from the bone marrow of three islet autotransplantation patients and expanded at our current Good Manufacturing Practices (cGMP) facility. On the day of islet transplantation, an average dose of 20.0 ± 2.6 ×106 MSCs was infused with islets via the portal vein. Adverse events and glycemic control at baseline, 6, and 12 months after transplantation were compared with data from 101 historical control patients. No adverse events directly related to the MSC infusions were observed. MSC patients required lower amounts of insulin during the peritransplantation period (p = .02 vs. controls) and had lower 12‐month fasting blood glucose levels (p = .02 vs. controls), smaller C‐peptide declines over 6 months (p = .01 vs. controls), and better quality of life compared with controls. In conclusion, our pilot study demonstrates that autologous MSC and islet cotransplantation may be a safe and potential strategy to improve islet engraftment after transplantation. (Clinicaltrials.gov registration number: NCT02384018). stem cells translational medicine 2018;7:11–19

Autologous mesenchymal stem cells or meniscal cells: what is the best cell source for regenerative meniscus treatment in an early osteoarthritis situation?

BackgroundTreatment of meniscus tears within the avascular region represents a significant challenge, particularly in a situation of early osteoarthritis. Cell-based tissue engineering approaches have shown promising results. However, studies have not found a consensus on the appropriate autologous cell source in a clinical situation, specifically in a challenging degenerative environment. The present study sought to evaluate the appropriate cell source for autologous meniscal repair in a demanding setting of early osteoarthritis.MethodsA rabbit model was used to test autologous meniscal repair. Bone marrow and medial menisci were harvested 4 weeks prior to surgery. Bone marrow-derived mesenchymal stem cells (MSCs) and meniscal cells were isolated, expanded, and seeded onto collagen-hyaluronan scaffolds before implantation. A punch defect model was performed on the lateral meniscus and then a cell-seeded scaffold was press-fit into the defect. Following 6 or 12 weeks, gross joint morphology and OARSI grade were assessed, and menisci were harvested for macroscopic, histological, and immunohistochemical evaluation using a validated meniscus scoring system. In conjunction, human meniscal cells isolated from non-repairable bucket handle tears and human MSCs were expanded and, using the pellet culture model, assessed for their meniscus-like potential in a translational setting through collagen type I and II immunostaining, collagen type II enzyme-linked immunosorbent assay (ELISA), and gene expression analysis.ResultsAfter resections of the medial menisci, all knees showed early osteoarthritic changes (average OARSI grade 3.1). However, successful repair of meniscus punch defects was performed using either meniscal cells or MSCs. Gross joint assessment demonstrated donor site morbidity for meniscal cell treatment. Furthermore, human MSCs had significantly increased collagen type II gene expression and production compared to meniscal cells (p < 0.05).ConclusionsThe regenerative potential of the meniscus by an autologous cell-based tissue engineering approach was shown even in a challenging setting of early osteoarthritis. Autologous MSCs and meniscal cells were found to have improved meniscal healing in an animal model, thus demonstrating their feasibility in a clinical setting. However, donor site morbidity, reduced availability, and reduced chondrogenic differentiation of human meniscal cells from debris of meniscal tears favors autologous MSCs for clinical use for cell-based meniscus regeneration.

Impact of Autologous Mesenchymal Stem Cell Infusion on Neuromyelitis Optica Spectrum Disorder: A Pilot, 2-Year Observational Study.

We evaluate safety and efficacy of autologous bone marrow-derived mesenchymal stem cells (MSCs) as a potential treatment for neuromyelitis optica spectrum disorder (NMOSD). Fifteen patients with NMOSD were recruited. All patients received a single intravenous infusion of 1.0 × 10(8) autologous MSC within 3-4 generations derived from bone marrow. The primary endpoints of the study were efficacy as reflected by reduction in annualized relapse rates (ARRs) and inflammatory lesions observed by MRI. At 12 months after MSC infusion, the mean ARR was reduced (1.1 vs. 0.3, P = 0.002), and the T2 or gadolinium-enhancing T1 lesions decreased in the optic nerve and spinal cord. Disability in these patients was reduced (EDSS, 4.3 vs. 4.9, P = 0.021; visual acuity, 0.4 vs. 0.5, P = 0.007). The patients had an increase in retinal nerve fiber layer thickness, optic nerve diameters and upper cervical cord area. We did not identify any serious MSC-related adverse events. At 24 months of MSC infusion, of 15 patients, 13 patients (87%) remained relapse-free, the mean ARR decreased to 0.1; the disability of 6 patients (40%) was improved, and the mean EDSS decreased to 4.0. This pilot trial demonstrates that MSC infusion is safe, reduces the relapse frequency, and mitigates neurological disability with neural structures in the optic nerve and spinal cord recover in patients with NMOSD. The beneficial effect of MSC infusion on NMOSD was maintained, at least to some degree, throughout a 2-year observational period.

Expansion and angiogenic potential of mesenchymal stem cells from patients with critical limb ischemia

Critical limb ischemia (CLI) is a life- and limb-threatening condition affecting 1% to 10% of the population with peripheral arterial disease. Traditional revascularization options are not possible for up to 50% of CLI patients, in which case, the use of cellular therapies, such as bone marrow-derived mesenchymal stem cells (MSCs), hold great promise as an alternative revascularization therapy. However, no randomized, controlled phase 3 trials to date have demonstrated an improvement in limb salvage with cellular therapies. This may be due to poor cell quality (ie, inability to generate a sufficient number of angiogenic MSCs) or to the inadequate retention and viability of MSCs after delivery, or both. Because concerns remain about the expansion and angiogenic potential of autologous MSCs in the CLI population, the objective of this study was to examine the effect of our novel culture media supplement, pooled human platelet lysate (PL), in lieu of the standard fetal bovine serum (FBS), to improve the expansion potential of MSCs from CLI patients. We also characterized the invitro angiogenic activity of MSCs from the tibia of amputated CLI limbs compared with MSCs from healthy donors. MSCs were obtained from the tibia of four CLI patients (ISC) and four ISC patients with diabetes mellitus (ISC+DM) undergoing major amputation. Healthy MSCs were aspirated from the iliac crest of four young and healthy donors. MSCs were isolated and expanded in culture with PL or FBS. MSCs from passage 3 to 6 were used for phenotypic marker expression and for adipogenic and osteogenic differentiation and were tested for their invitro angiogenic activity on human microdermal endothelial cells. In parallel MSCs were cultured to passage 11 for population-doubling calculations. MSCs from ISC and ISC+DM patients and from healthy patients exhibited appropriate expression of cell surface markers and differentiation capacity. Population doublings were significantly greater for PL-stimulated compared with FBS-stimulated MSCs in all groups. Biologically active amounts of angiogens were identified in the secretome of all MSCs without consistent trends among groups. PL expansion did not adversely affect the angiogenic activity of MSCs compared with FBS. The ISC and ISC+DM MSCs demonstrated angiogenic effects on endothelial cells similar to those of healthy and ISC MSCs. PL promotes the rapid expansion of MSCs from CLI and healthy persons. Importantly, MSCs expanded from CLI patients demonstrate the desired angiogenic activity compared with their healthy counterparts. We conclude that autologous MSCs from CLI patients can be sufficiently expanded with PL and be expected to deliver requisite angiogenic effects invivo. We expect the improved expansion of ISC and ISC+DM with PL to be helpful in improving the successful delivery of autologous MSCs to patients with CLI.

Aortic valve conduit implantation in the descending thoracic aorta in a sheep model: The outcomes of pre-seeded scaffold

BackgroundWe evaluated the outcomes of implanting pre-seeded decellularized aortic valve conduit (AVC) with bone marrow-derived mesenchymal stem cells (MSCs) in a sheep model. MethodsEight sheep AVCs were obtained under sterile conditions and decellularized by using detergent-based methods. Decellularized AVCs were seeded with autologous bone marrow-derived MSCs in a dynamic bioreactor system. Pre-seeded AVCs were implanted in the descending thoracic aorta in a sheep model. In all sheep, a decellularized pericardial patch was also anastomosed to the proximal part in order to reduce the incidence of rupture. Pathological evaluations, echocardiography, multislice computed tomography (CT), and CT angiography were performed for the evaluation of implanted AVCs. ResultsThe longest survival period was 19 months in pre-seeded animals with complete recellularization at the long-term follow-up. Immunohistochemical staining for desmin, smooth muscle actin, and cytokeratin was significantly positive in the pre-seeded samples and reached near normal ranges. CT angiography revealed no intimal tearing after 18 months of follow-up. ConclusionPre-seeded AVCs with bone marrow-derived MSCs may have satisfactory results in postoperative cell seeding capabilities with promising functional potentiality. This modality may be beneficial and may provide a new era of biological grafts in cardiovascular surgery.

Influence of carrier cells on the clinical outcome of children with neuroblastoma treated with high dose of oncolytic adenovirus delivered in mesenchymal stem cells

We report here our clinical experience of a program of compassionate use of Celyvir – autologous marrow-derived mesenchymal stem cells (MSCs) carrying an oncolytic adenovirus – for treating children with advanced metastatic neuroblastoma. Children received weekly doses of Celyvir with no concomitant treatments. The tolerance was excellent, with very mild and self-limited viral-related symptoms. Patients could be distinguished based on their response to therapy: those who had a clinical response (either complete, partial or stabilization) and those who did not respond. We found differences between patients who responded versus those who did not when analyzing their respective MSCs, at the expression levels of adhesion molecules (CCR1, CXCR1 and CXCR4) and in migration capacities in transwell assays, and in immune-related molecules (IFNγ, HLA-DR). These results suggest interpatient differences in the homing and immune modulation capacities of the therapy administered. In addition, the pretherapy immune T cell status and the T effector response were markedly different between responders and non-responders. We conclude that multidoses of Celyvir have an excellent safety profile in children with metastatic neuroblastoma. Intrinsic patients' and MSCs' factors appear to be related to clinical outcome.

Intravitreal Implantation of Genetically Modified Autologous Bone Marrow-Derived Stem Cells for Treating Retinal Disorders.

A number of retinal degenerative diseases may be amenable to treatment with continuous intraocular delivery of therapeutic agents that cannot be delivered effectively to the retina via systemic or topical administration. Among these disorders are lysosomal storage diseases resulting from deficiencies in soluble lysosomal enzymes. Most cells, including those of the retina, are able to take up these enzymes and incorporate them in active form into their lysosomes. In theory, therefore, continuous intraocular administration of a normal form of a soluble lysosomal enzyme should be able to cure the molecular defect in the retinas of subjects lacking this enzyme. Experiments were conducted to determine whether genetically modified bone marrow-derived stem cells implanted into the vitreous could be used as -vehicles for continuous delivery of such enzymes to the retina. Bone marrow-derived mesenchymal stem cells (MSCs) from normal mice were implanted into the vitreous of mice undergoing retinal degeneration as a result of a mutation in the PPT1 gene. The implanted cells appeared to survive indefinitely in the vitreous without proliferating or invading the retina. This indicates that intravitreal implantation of MSCs is likely a safe means of long-term delivery of proteins synthesized by the implanted cells. Experiments have been initiated to test the efficacy of using genetically modified autologous MSCs to inhibit retinal degeneration in a canine model of neuronal ceroid lipofuscinosis.

Interferon-Gamma Modification of Mesenchymal Stem Cells: Implications of Autologous and Allogeneic Mesenchymal Stem Cell Therapy in Allotransplantation

Bone marrow-derived mesenchymal stem cells (MSC) have unique immunomodulatory and reparative properties beneficial for allotransplantation cellular therapy. The clinical administration of autologous or allogeneic MSC with immunosuppressive drugs is able to prevent and treat allograft rejection in kidney transplant recipients, thus supporting the immunomodulatory role of MSC. Interferon-gamma (IFN-γ) is known to enhance the immunosuppressive properties of MSC. IFN-γ preactivated MSC (MSC-γ) directly or indirectly modulates T cell responses by enhancing or inducing MSC inhibitory factors. These factors are known to downregulate T cell activation, enhance T cell negative signalling, alter T cells from a proinflammatory to an anti-inflammatory phenotype, interact with antigen-presenting cells and increase or induce regulatory cells. Highly immunosuppressive MSC-γ with increased migratory and reparative capacities may aid tissue repair, prolong allograft survival and induce allotransplant tolerance in experimental models. Nevertheless, there are contradictory in vivo observations related to allogeneic MSC-γ therapy. Many studies report that allogeneic MSC are immunogenic due to their inherent expression of major histocompatibility (MHC) molecules. Enhanced expression of MHC in allogeneic MSC-γ may increase their immunogenicity and this can negatively impact allograft survival. Therefore, strategies to reduce MSC-γ immunogenicity would facilitate "off-the-shelf" MSC therapy to efficiently inhibit alloimmune rejection and promote tissue repair in allotransplantation. In this review, we examine the potential benefits of MSC therapy in the context of allotransplantation. We also discuss the use of autologous and allogeneic MSC and the issues associated with their immunogenicity in vivo, with particular focus on the use of enhanced MSC-γ cellular therapy.

Mesenchymal Stem Cells for Regenerative Therapy: Optimization of Cell Preparation Protocols

Administration of bone marrow-derived mesenchymal stem cells (MSCs) is an innovative approach for the treatment of a range of diseases that are not curable by current therapies including heart failure. A number of clinical trials have been completed and many others are ongoing; more than 2,000 patients worldwide have been administered with culture-expanded allogeneic or autologous MSCs for the treatment of various diseases, showing feasibility and safety (and some efficacy) of this approach. However, protocols for isolation and expansion of donor MSCs vary widely between these trials, which could affect the efficacy of the therapy. It is therefore important to develop international standards of MSC production, which should be evidence-based, regulatory authority-compliant, of good medical practice grade, cost-effective, and clinically practical, so that this innovative approach becomes an established widely adopted treatment. This review article summarizes protocols to isolate and expand bone marrow-derived MSCs in 47 recent clinical trials of MSC-based therapy, which were published after 2007 onwards and provided sufficient methodological information. Identified issues and possible solutions associated with the MSC production methods, including materials and protocols for isolation and expansion, are discussed with reference to relevant experimental evidence with aim of future clinical success of MSC-based therapy.

Presence of osteoclast precursor cells during ex vivo expansion of bone marrow-derived mesenchymal stem cells for autologous use in cell therapy

Background aimsTo obtain a cell product competent for clinical use in terms of cell dose and biologic properties, bone marrow-derived mesenchymal stem cells (MSCs) must be expanded ex vivo. MethodsA retrospective analysis was performed of records of 76 autologous MSC products used in phase I or II clinical studies performed in a cohort of cardiovascular patients. In all cases, native MSCs present in patient bone marrow aspirates were separated and expanded ex vivo. ResultsThe cell products were classified in two groups (A and B), according to biologic properties and expansion time (ex vivo passages) to reach the protocol-established cell dose. In group A, the population of adherent cells obtained during the expansion period (2 ± 1 passages) was composed entirely of MSCs and met the requirements of cell number and biologic features as established in the respective clinical protocol. In group B, in addition to MSCs, we observed during expansion a high proportion of ancillary cells, characterized as osteoclast precursor cells. In this case, although the biologic properties of the resulting MSC product were not affected, the yield of MSCs was significantly lower. The expansion cycles had to be increased (3 ± 1 passages). ConclusionsThese results suggest that the presence of osteoclast precursor cells in bone marrow aspirates may impose a limit for the proper clinical use of ex vivo expanded autologous bone marrow-derived MSCs.

Investigation of the immune response to autologous, allogeneic, and xenogeneic mesenchymal stem cells after intra-articular injection in horses

Mesenchymal stem cells have demonstrated immunomodulatory capabilities as well as modest efficacy in animal models of joint injury, warranting further study as a potential treatment of joint disease. The goal of the study was to investigate the blood and synovial immune and histologic response to intra-articular injection of autologous, allogeneic, and xenogeneic bone marrow-derived mesenchymal stem cells (MSC) in horses. The study group consisted of 6 five-year-old Thoroughbred mares that had been injected previously with 15 million, genetically modified autologous, allogeneic, or xenogeneic MSC into the fetlock joints. One group of autologous cells was genetically modified to permit MSC biolocalization in the synovium. To assess response to the injection, synovial biopsies were obtained via arthroscopy 60 days after MSC injection for gross, histologic and molecular analyses. Peripheral blood mononuclear cells were isolated from each horse 120 days after MSC injection and co-cultured with a monolayer of each MSC group to permit quantification of activated CD4+ lymphocytes and cytokine release (ELISA) upon re-exposure to MSC.Arthroscopic examination revealed normal synovium with no grossly detrimental effect to the synovium or cartilage. Intra-articular MSC produced a persistent mononuclear infiltrate for at least 60 days, mostly perivascular, identified as CD3+ lymphocytes. An immune response (significant increase in CD4+ lymphocytes) was detected upon re-exposure to xenogeneic but not to allogeneic or autologous MSC. An inflammatory cytokine release from peripheral blood mononuclear cell/MSC co-cultures was present in all MSC groups but was significantly greater in the xenogeneic group. In conclusion, intra-articular injection of MSC, regardless of cell origin, incited a persistent mononuclear synovitis demonstrating a sustained biologic influence of these cells. Allogeneic cells did not elicit a detectable immune response upon re-exposure using our methods.

Inflammatory effects of autologous, genetically modified autologous, allogeneic, and xenogeneic mesenchymal stem cells after intra-articular injection in horses

To compare the clinical and inflammatory joint responses to intra-articular injection of bone marrow-derived mesenchymal stem cells (MSC) including autologous, genetically modified autologous, allogeneic, or xenogeneic cells in horses. Six five-year-old Thoroughbred mares had one fetlock joint injected with Gey's balanced salt solution as the vehicle control. Each fetlock joint of each horse was subsequently injected with 15 million MSC from the described MSC groups, and were assessed for 28 days for clinical and inflammatory parameters representing synovitis, joint swelling, and pain. There were not any significant differences between autologous and genetically modified autologous MSC for synovial fluid total nucleated cell count, total protein, interleukin (IL)-6, IL-10, fetlock circumference, oedema score, pain-free range-of-motion, and soluble gene products that were detected for at least two days. Allogeneic and xenogeneic MSC produced a greater increase in peak of inflammation at 24 hours than either autologous MSC group. Genetically engineered MSC can act as vehicles to deliver gene products to the joint; further investigation into the therapeutic potential of this cell therapy is warranted. Intra-articular MSC injection resulted in a moderate acute inflammatory joint response that was greater for allogeneic and xenogeneic MSC than autologous MSC. Clinical management of this response may minimize this effect.

Effect on Ligament Marker Expression by Direct-Contact Co-culture of Mesenchymal Stem Cells and Anterior Cruciate Ligament Cells

Ligament and tendon repair is an important topic in orthopedic tissue engineering; however, the cell source for tissue regeneration has been a controversial issue. Until now, scientists have been split between the use of primary ligament fibroblasts or marrow-derived mesenchymal stem cells (MSCs). The objective of this study was to show that a co-culture of anterior cruciate ligament (ACL) cells and MSCs has a beneficial effect on ligament regeneration that is not observed when utilizing either cell source independently. Autologous ACL cells (ACLcs) and MSCs were isolated from Yorkshire pigs, expanded in vitro, and cultured in multiwell plates in varying %ACLcs/%MSCs ratios (100/0, 75/25, 50/50, 25/75, and 0/100) for 2 and 4 weeks. Quantitative mRNA expression analysis and immunofluorescent staining for ligament markers Collagen type I (Collagen-I), Collagen type III (Collagen-III), and Tenascin-C were performed. We show that Collagen-I and Tenascin-C expression is significantly enhanced over time in 50/50 co-cultures of ACLcs and MSCs (p≤0.03), but not in other groups. In addition, Collagen-III expression was significantly greater in MSC-only cultures (p≤0.03), but the Collagen-I-to-Collagen-III ratio in 50% co-culture was closest to native ligament levels. Finally, Tenascin-C expression at 4 weeks was significantly higher (p≤0.02) in ACLcs and 50% co-culture groups compared to all others. Immunofluorescent staining results support our mRNA expression data. Overall, 50/50 co-cultures had the highest Collagen-I and Tenascin-C expression, and the highest Collagen-I-to-Collagen-III ratio. Thus, we conclude that using a 50% co-culture of ACLcs and MSCs, instead of either cell population alone, may better maintain or even enhance ligament marker expression and improve healing.

Transplantation With Autologous Mesenchymal Stem Cells After Acute Myocardial Infarction Evaluated by Magnetic Resonance Imaging

The purpose of this study was to track and investigate the effects of autologous bone marrow-derived mesenchymal stem cells (MSCs) transplantation after acute myocardial infarction in swine assessed by magnetic resonance imaging (MRI). Twenty-four Chinese mini-pigs (27±3 kg) were divided into 4 groups, including control groups (groups 1 and 3) and MSCs transplantation groups (group 2, super paramagnetic iron oxide labeled and group 4, 4',6-diamidino-2-phenylindole labeled). Super paramagnetic iron oxide-labeled and 4',6-diamidino-2-phenylindole-labeled MSCs (3.0×10⁶ cells/mL) with a volume of 10 mL were injected into the left anterior descending artery by a catheter at 1 week after acute myocardial infarction, respectively. Cell distribution, cardiac functions, and scar tissue were quantitatively assessed by MRI. The reduction of the T2* value in the myocardium, spleen, and liver in group 2 was significantly greater than that in group 1. MRI showed that function and scar size at baseline and 3 days after cell infusion were not significantly different between groups 1 and 2. Six weeks later left ventricular ejection fraction (P<0.0001), end-systolic volume (P<0.05), the number of dyskinetic segments (P<0.0001), left ventricular weight index (P<0.0001), and the infarcted size (P<0.0001) in group 4 were all improved comparing with those in group 3. The majority of MSCs entrapped by the extracardial organs were mainly in the spleen. Catheter-based delivery of autologous bone marrow-derived MSCs into infarcted myocardium is feasible and effective.