Autologous Adipose Tissue-derived Mesenchymal Stem Research Articles

Seven-Year Longitudinal Study: Clinical Evaluation of Knee Osteoarthritic Patients Treated with Mesenchymal Stem Cells.

Background/Objectives: Numerous studies have demonstrated the safety and efficacy of intraarticular stem cell injections for treating osteoarthritic knee joints, reporting symptom reduction and pain relief within a few months of treatment. Here, we report the results of a 7-year follow-up after a single intraarticular injection of 0.5-1 × 107 autologous adipose tissue-derived mesenchymal stem cells in patients with OA (Kellgren-Lawrence grade 2 to 4). Methods: Nine patients were treated, and two patients had bilateral disease. Patients were evaluated clinically and radiologically using X-ray and MRI. A comprehensive statistical analysis was undertaken to evaluate the obtained results. Results: All clinical scores and range of motion significantly improved within the first six months after injection. At the 18-month time point, a significant improvement in cartilage structure was observed on MRI while X-ray showed no changes in subchondral bone of distal femur and proximal tibia. At the 60-month time point, the clinical scores were still improved compared to baseline, except for the range of motion, which decreased almost back to the baseline level. At 84 months, the clinical scores decreased significantly toward the baseline level, but the MRI structural characteristics of cartilage still remained significantly better than those measured at baseline. Conclusions: Adipose tissue-derived stem cell therapy has substantial long-term clinical effects on patients with knee osteoarthritis.

Safety and efficacy of autologous adipose tissue-derived stem cell transplantation in aging-related low-grade inflammation patients: a single-group, open-label, phase I clinical trial

BackgroundInflamm-aging is associated with the rate of aging and is significantly related to diseases such as Alzheimer’s disease, Parkinson’s disease, atherosclerosis, heart disease, and age-related degenerative diseases such as type II diabetes and osteoporosis. This study aims to evaluate the safety and efficiency of autologous adipose tissue-derived mesenchymal stem cell (AD-MSC) transplantation in aging-related low-grade inflammation patients.MethodsThis study is a single-group, open-label, phase I clinical trial in which patients treated with 2 infusions (100 million cells i.v) of autologous AD-MSCs were initially evaluated in 12 inflamm-aging patients who concurrently had highly proinflammatory cytokines and 2 of the following 3 diseases: diabetes, dyslipidemia, and obesity. The treatment effects were evaluated based on plasma cytokines.ResultsDuring the study’s follow-up period, no adverse effects were observed in AD-MSC injection patients. Compared to baseline (D-44), the inflammatory cytokines IL-1α, IL-1β, IL-8, IL-6, and TNF-α were significantly reduced after 180 days (D180) of MSC infusion. IL-4/IL-10 at 90 days (D90) and IL-2/IL-10 at D180 increased, reversing the imbalance between proinflammatory and inflammatory ratios in the patients.ConclusionAD-MSCs represent a potential intervention to prevent age-related inflammation in patients.Trial registrationClinicalTrials.gov number is NCT05827757, first registered on 13th Oct 2020

Intrathecal delivery of adipose-derived mesenchymal stem cells in traumatic spinal cord injury: Phase I trial

Intrathecal delivery of autologous culture-expanded adipose tissue-derived mesenchymal stem cells (AD-MSC) could be utilized to treat traumatic spinal cord injury (SCI). This Phase I trial (ClinicalTrials.gov: NCT03308565) included 10 patients with American Spinal Injury Association Impairment Scale (AIS) grade A or B at the time of injury. The study’s primary outcome was the safety profile, as captured by the nature and frequency of adverse events. Secondary outcomes included changes in sensory and motor scores, imaging, cerebrospinal fluid markers, and somatosensory evoked potentials. The manufacturing and delivery of the regimen were successful for all patients. The most commonly reported adverse events were headache and musculoskeletal pain, observed in 8 patients. No serious AEs were observed. At final follow-up, seven patients demonstrated improvement in AIS grade from the time of injection. In conclusion, the study met the primary endpoint, demonstrating that AD-MSC harvesting and administration were well-tolerated in patients with traumatic SCI.

Long-Term Effects of Adipose-Derived Stem Cells for the Treatment of Bilateral Limbal Stem Cell Deficiency

Purpose To determine the safety and feasibility of human autologous adipose tissue-derived adult mesenchymal stem cells (ASCs) for ocular surface regeneration in patients with bilateral limbal stem-cell deficiency (LSCD). Methods A phase IIa clinical trial was designed (https://Clinicaltrials.gov, NCT01808378) with 8 patients, 3 of whom had aniridia, 2 meibomian glands diseases, 2 multiple surgeries and 1 chronic chemical injury. The therapeutic protocol was as follows: 6-mm of central corneal epithelium was removed, 400,000 ASCs were injected into each limboconjunctival quadrant, 400,000 ASCs were suspended over the cornea for 20 min, and finally the cornea was covered with an amniotic membrane patch. Results No adverse events were detected after a mean of 86,5 months of follow-up. One year after surgery, 6 of the 8 transplants were scored as successful, five patients had improved uncorrected visual acuity (mean of 12 letters), two patients presented epithelial defects (also present at baseline) and the mean percentage of corneal neovascularization was of 28.75% (36.98%, at baseline). Re-examination 24 months after treatment disclosed preserved efficacy in 4 patients. At the last visit (after a mean of 86,5 months of follow up) epithelial defects were absent in all patients although improvement in all of the variables was only maintained in patient 3 (meibomian glands agenesia). Conclusion ASCs are a feasible and conservative therapy for treating bilateral LSCD. The therapeutic effect differs between etiologies and diminishes over time.

Impact and Possible Mechanism(s) of Adipose Tissue-Derived Mesenchymal Stem Cells on T-Cell Proliferation in Patients With Rheumatic Disease.

Objectives: Systemic lupus erythematosus (SLE) and systemic sclerosis (SSc) are chronic wasting, incurable rheumatic diseases of autoimmune background, in which T cells play a critical pathogenic role. Autologous adipose tissue-derived mesenchymal stem cells (ASCs) may represent an alternative therapeutic option for SLE and SSc patients, but the biology of these cells is poorly understood.Methods: Herein, we evaluated the anti-proliferative impact of ASCs of healthy donors (HD/ASCs, 5 reference cell lines), SLE patients (n = 20), and SSc patients (n = 20) on T lymphocytes. To assess the direct and indirect pathway of ASCs action, peripheral blood mononuclear cells (PBMCs) and purified CD4+ T cells of HD were activated and co-cultured in cell-to-cell contact (C-C) and transwell (T-W) conditions with untreated or cytokine (TNF + IFNΥ, TI)-licensed ASCs, then analyzed by flow cytometry to rate the proliferation response of CD8+ and/or CD4+ T cells. The concentrations of kynurenines, prostaglandin E2 (PGE2), interleukin 10 (IL-10), and transforming growth factor β (TGFβ) were measured from culture supernatants. Specific inhibitors of these factors (1-MT, indomethacin, and cytokine-neutralizing antibody) were used to assess their contribution to anti-proliferative ASCs action.Results: All tested ASCs significantly decreased the number of proliferating CD4+ and CD8+ T cells, the number of division/proliferating cell (PI), and fold expansion (RI), and similarly upregulated kynurenines and PGE2, but not cytokine levels, in the co-cultures with both types of target cells. However, TI-treated SLE/ASCs and SSc/ASCs exerted a slightly weaker inhibitory effect on CD4+ T-cell replication than their respective HD/ASCs. All ASCs acted mainly via soluble factors. Their anti-proliferative effect was stronger, and kynurenine levels were higher in the T-W condition than the C-C condition. Blocking experiments indicated an involvement of kynurenine pathway in inhibiting the number of proliferating cells, PI, and RI values as well as PGE2 role in decreasing the number of proliferating cells. TGFβ did not contribute to ASCs anti-proliferative capabilities, while IL-10 seems to be involved in such activity of only SLE/ASCs.Conclusion: The results indicate that SLE/ASCs and SSc/ASCs retain their capability to restrain the expansion of allogeneic CD4+ and CD8+ T cells and act by similar mechanisms as ASCs of healthy donors and thus may have therapeutic value.

Mesenchymal stem cell therapy attenuates complement C3 deposition and improves the delicate equilibrium between angiogenic and anti-angiogenic factors in abortion-prone mice

Immunological disorders are one of the main causes of recurrent spontaneous abortions (RSA). A rapidly expanding body of evidence indicates that excessive activation of the complement system is critically involved in the development of miscarriages. In the CBA/J × DBA/2 murine model of recurrent miscarriage, exaggerated and unrestrained complement activation is reported to be the underlying cause of angiogenic factor imbalance and persistent inflammation. We have previously shown that mesenchymal stem cell (MSC) therapy can significantly reduce the abortion rate in abortion-prone mice through regulating the feto-maternal immune response. In the present study, we hypothesized that MSCs might improve the balance of angiogenic factors at the feto-maternal unit of CBA/J × DBA/2 mice by restraining complement activation and deposition.To explore this hypothesis, autologous adipose tissue-derived mesenchymal stem cells (AD-MSCs) were administered intra-peritoneally to abortion-prone mice on the 4.5th day of gestation. Control mice received PBS as vehicle. On day 13.5 of pregnancy, deposition of the complement component C3 and expression levels of Crry, CFD (adipsin), VEGF, PlGF and FLT-1 were measured at the feto-maternal interface by immunohistochemistry and real-time PCR, respectively. Decidual cells were also cultured in RPMI 1640 medium for 48 h and VEGF and sFLT-1 protein levels were quantified in supernatants using enzyme-linked immunosorbent assay (ELISA).Our results indicated that MSC therapy significantly reduced C3 deposition and adipsin transcription in the fetal-maternal interface of abortion-prone mice. Furthermore, administration of MSCs robustly upregulated the mRNA expression levels of Crry, VEGF, PlGF and FLT-1 in the placenta and decidua of CBA/J × DBA/2 mice. Consistently, the in vitro results demonstrated that decidual cells obtained from MSC-treated dams produced increased concentrations of VEGF in culture supernatants, with concomitant decreased levels of sFLT-1 protein.Here, we show for the first time that adoptive transfer of MSCs rectifies the disturbed balance of angiogenic factors observed at the feto-maternal unit of CBA/J × DBA/2 mice, in part at least, through inhibiting excessive complement activation and promoting the production of angiogenic factors. Collectively, these alterations seem to play a pivotal role in reducing the abortion rate and improving the intrauterine condition for the benefit of the fetus.

Case Report: Successful Therapy of Spontaneously Occurring Canine Degenerative Lumbosacral Stenosis Using Autologous Adipose Tissue-Derived Mesenchymal Stem Cells

The management of degenerative lumbosacral stenosis (DLSS) in dogs usually requires aggressive, costly surgical treatments that may themselves present complications, while do not fully resolve the symptoms of the disease. In this study, the dog diagnosed with severe DLSS, with hind limb paresis, was treated using a new and least invasive treatment. Cultured autologous adipose tissue-derived mesenchymal stem cells (AT-MSCs) were injected bilaterally at the level of L7-S1, in the vicinity of the external aperture of the intervertebral foramen of DLSS patient. In the previously described treatments of spontaneous intervertebral disc degeneration in dogs, intradiscal injections of MSCs did not lead to positive effects. Here, we report a marked improvement in clinical outcome measures related to the ability of a dog to walk and trot, which were expressed by a numeric rating scale based on a veterinary assessment questionnaire. The improved status persisted throughout the observed time course of 4.5 years after the AT-MSC transplantation. To the best of our knowledge, this is the first case of successful therapy, with long-term positive effect, of spontaneously occurring canine DLSS using presented treatment that, we believe, represents a contribution to current knowledge in this field and may shape both animal and human DLSS treatment options.

Autologous Adipose Tissue-Derived Mesenchymal Stem Cells Introduced by Biliary Stents or Local Immersion in Porcine Bile Duct Anastomoses.

Biliary complications (strictures and leaks) represent major limitations in living donor liver transplantation. Mesenchymal stem cells (MSCs) are a promising modality to prevent biliary complications because of immunosuppressive and angiogenic properties. Our goal was to evaluate the safety of adipose-derived MSC delivery to biliary anastomoses in a porcine model. Secondary objectives were defining the optimal method of delivery (intraluminal versus extraluminal) and to investigate MSC engraftment, angiogenesis, and fibrosis. Pigs were divided into 3 groups. Animals underwent adipose collection, MSC isolation, and expansion. Two weeks later, animals underwent bile duct transection, reanastomosis, and stent insertion. Group 1 received plastic stents wrapped in unseeded Vicryl mesh. Group 2 received stents wrapped in MSC-seeded mesh. Group 3 received unwrapped stents with the anastomosis immersed in an MSC suspension. Animals were killed 1 month after stent insertion when cholangiograms and biliary tissue were obtained. Serum was collected for liver biochemistries. Tissue was used for hematoxylin-eosin and trichrome staining and immunohistochemistry for MSC markers (CD44 and CD34) and for a marker of neoangiogenesis (CD31). There were no intraoperative complications. One pig died on postoperative day 3 due to acute cholangitis. All others recovered without complications. Cholangiography demonstrated no biliary leaks and minimal luminal narrowing. Surviving animals exhibited no symptoms, abnormal liver biochemistries, or clinically significant biliary stricturing. Group 3 showed significantly greater CD44 and CD34 staining, indicating MSC engraftment. Fibrosis was reduced at the anastomotic site in group 3 based on trichrome stain. CD31 staining of group 3 was more pronounced, supporting enhanced neoangiogenesis. In conclusion, adipose-derived MSCs were safely applied to biliary anastomoses. MSCs were locally engrafted within the bile duct and may have beneficial effects in terms of fibrosis and angiogenesis.

Intratendinous Injection of Mesenchymal Stem Cells for the Treatment of Rotator Cuff Disease: A 2-Year Follow-Up Study

To assess the mid-term safety and efficacy of an intratendinous injection of autologous adipose tissue-derived mesenchymal stem cells (AD MSCs) for rotator cuff disease at 2-year follow-up. The first part of the study consisted of 3 dose-escalation groups, with 3 patients each, for the evaluation of safety: low-dose (1.0× 107 cells), mid-dose (5.0× 107), and high-dose (1.0× 108) groups. For the second part, we planned to include 9 patients receiving the high dose for the evaluation of exploratory efficacy. Clinical outcomes were assessed according to pain, range of motion, muscle strength, functional scores, overall satisfaction and function, and presence of failure. Structural outcomes included changes in volume of tendon defects measured using magnetic resonance imaging. This study enrolled 19 patients (9 for the first part and 10 for the second part) with partial-thickness rotator cuff tears. There were no treatment-related adverse events at minimum 2-year follow-up. Intratendinous injection of AD MSCs reduced shoulder pain by approximately 90% at 1 and 2 years in the mid- and high-dose groups. The strength of the supraspinatus, infraspinatus, and teres minor significantly increased by greater than 50% at 2 years in the high-dose group. Shoulder function measured with 6 commonly used scores improved for up to 2 years in all dose groups. Structural outcomes evaluated with magnetic resonance imaging showed that the volume of bursal-sided defects in the high-dose group nearly disappeared at 1 year and did not recur at up to 2 years. No failures-defined as the performance of any kind of shoulder surgery or return of the Shoulder Pain and Disability Index score to the preinjection level-occurred during follow-up. This study showed continued safety and efficacy of an intratendinous injection of AD MSCs for the treatment of partial-thickness rotator cuff tears over a 2-year period through regeneration of tendon defects. Level III, retrospective comparative study.

Treatment of Buerger’s disease (Thromboangiitis obliterans) with autologous adipose tissue-derived mesenchymal stem cell: Report of three cases

Buerger’s disease or Thromboangiitis obliterans is an orphan vascular disease that most commonly affects nerves, small or medium-sized vessels in the upper and lower extremities, and is characterized by a non-atherosclerotic, segmental, inflammatory disorder. The etiology and the pathogenesis of the disease have not been fully elucidated. Although various interventions have been adopted recently, there is still no effective treatment for the prevention of the progression of the disease. This report presents three clinical cases that show the efficacies of autologous adipose tissue-derived mesenchymal stem cell (AdMSC) treatment in Buerger’s disease. Three male patients diagnosed with Buerger’s disease were between 46 and 55 years and had a smoking history. AdMSCs (5X106 cells/kg body weight) were injected intramuscularly into at least 38 points of the ischemic legion of the lower limb at one time. The patients were checked for safety and efficacy at one, three, and six months after AdMSC injection. No severe adverse events and no adverse drug events were observed in physical examination, vital signs, and laboratory tests for all three patients. Ulcers in the affected legs of the patients were healed completely after the treatment. Visual Analogue Scale scores and all the criteria (activities, emotional, pain, social, symptoms and total) of the King's College Hospital's Vascular Quality of Life Questionnaire (VascuQOL) of all the patients were improved from baseline to six months follow-up. Digital Infrared Thermal Imaging showed the gradual alleviation of lesions in the leg. Angiogenesis in the affected limbs was identified by CT-Angiography after AdMSC injection. The present cases show the improvement in patients with Buerger’s disease with the observation of angiogenesis after intramuscular injection of autologous AdMSCs. This suggests that autologous AdMSC can be an effective alternative treatment for Buerger’s disease.

Case Report: Exploratory treatment with multiple intravenous infusion of the autologous adipose tissue-derived mesenchymal stem cells for the treatment of Diamond-Blackfan anemia patient

Diamond-Blackfan anemia (DBA) is a rare congenital erythropoietic disorder characterized by erythroblastopenia. Conventional treatments of DBA are the administrations of corticosteroids and blood transfusions for mitigation of anemia, and bone marrow transplantation. However, there are hurdles to overcome for long-term use and the conventional treatment. Mesenchymal stem cells (MSCs) have been noted as a novel alternative cell therapy in various diseases, and adipose tissue-derived MSCs (AdMSCs) are known for their versatile efficacies and feasibility. Here, we report the potential efficacies and the safety of intravenous administration of the autologous AdMSC in a patient with DBA for the first time. The isolation and characterization of autologous AdMSCs from a girl aged 11 years, 10 months with DBA were carried out due to the mutation of ribosomal protein s24 (RPS24). AdMSCs, diluted to 1 x 108 cells in 100 ml of saline, were infused intravenously for 1 hour. Intravenous administration of AdMSCs was carried out 5 times in 2-week intervals, and the patient was checked using various assessments (vital signs, physical examination, laboratory tests, adverse events, etc) at every visit. After 3, 6 and 9 months from the first administration of AdMSCs, red blood cell (RBC) count, hemoglobin value, and hematocrit were assessed for the efficacy. There were no side effects or adverse events observed during the treatment. Although showing subnormal values, the RBC number, hemoglobin level, and hematocrit were improved 9 months after the systemic administration of AdMSCs from baseline; the RBC count (x106 /μl), hemoglobin level (g/dl) and hematocrit level (%) were increased from 1.58 to 2.38, 5.6 to 8.3, and 16.9 to 26.1, respectively. The present case reported the first AdMSC administration for DBA patient and indicates it is possible that the intravenous administration of autologous AdMSC can be a safe alternative for DBA treatment.

Systemic Infusion of Autologous Adipose Tissue-Derived Mesenchymal Stem Cells in Peritoneal Dialysis Patients: Feasibility and Safety.

Objective Using mesenchymal stem cells (MSCs) is regarded as a new therapeutic approach for improving fibrotic diseases. the aim of this study to evaluate the feasibility and safety of systemic infusion of autologous adipose tissue-derived MSCs (AD-MSCs) in peritoneal dialysis (PD) patients with expected peritoneal fibrosis. Materials and Methods This study was a prospective, open-label, non-randomized, placebo-free, phase I clinical trial. Case group consisted of nine eligible renal failure patients with more than two years of history of being on PD. Autologous AD-MSCs were obtained through lipoaspiration and expanded under good manufacturing practice conditions. Patients received 1.2 ± 0.1×106 cell/kg of AD-MSCs via cubital vein and then were followed for six months at time points of baseline, and then 3 weeks, 6 weeks, 12 weeks, 16 weeks and 24 weeks after infusion. Clinical, biochemical and peritoneal equilibration test (PET) were performed to assess the safety and probable change in peritoneal solute transport parameters. Results No serious adverse events and no catheter-related complications were found in the participants. 14 minor reported adverse events were self-limited or subsided after supportive treatment. One patient developed an episode of peritonitis and another patient experienced exit site infection, which did not appear to be related to the procedure. A significant decrease in the rate of solute transport across peritoneal membrane was detected by PET (D/P cr=0.77 vs. 0.73, P=0.02). ConclusionThis study, for the first time, showed the feasibility and safety of AD-MSCs in PD patients and the potentials for positive changes in solute transport. Further studies with larger samples, longer follow-up, and randomized blind control groups to elucidate the most effective route, frequency and dose of MSCs administration, are necessary (Registration Number: IRCT2015052415841N2).

Intratendinous Injection of Autologous Adipose Tissue-Derived Mesenchymal Stem Cells for the Treatment of Rotator Cuff Disease: A First-In-Human Trial.

Despite relatively good results of current symptomatic treatments for rotator cuff disease, there has been an unmet need for fundamental treatments to halt or reverse the progress of disease. The purpose of this study was to assess the safety and efficacy of intratendinous injection of autologous adipose tissue-derived mesenchymal stem cells (AD MSCs) in patients with rotator cuff disease. The first part of the study consists of three dose-escalation cohorts; the low- (1.0 × 107 cells), mid- (5.0 × 107 ), and high-dose (1.0 × 108 ) groups with three patients each for the evaluation of the safety and tolerability. The second part included nine patients receiving the high-dose for the evaluation of the exploratory efficacy. The primary outcomes were the safety and the shoulder pain and disability index (SPADI). Secondary outcomes included clinical, radiological, and arthroscopic evaluations. Twenty patients were enrolled in the study, and two patients were excluded. Intratendinous injection of AD MSCs was not associated with adverse events. It significantly decreased the SPADI scores by 80% and 77% in the mid- and high-dose groups, respectively. Shoulder pain was significantly alleviated by 71% in the high-dose group. Magnetic resonance imaging examination showed that volume of the bursal-side defect significantly decreased by 90% in the high-dose group. Arthroscopic examination demonstrated that volume of the articular- and bursal-side defects decreased by 83% and 90% in the mid- and high-dose groups, respectively. Intratendinous injection of autologous AD MSCs in patient with a partial-thickness rotator cuff tear did not cause adverse events, but improved shoulder function, and relieved pain through regeneration of rotator cuff tendon. Stem Cells 2018;36:1441-1450.

A Prospective, Nonrandomized, no Placebo-Controlled, Phase I/II Clinical Trial Assessing the Safety and Efficacy of Intramuscular Injection of Autologous Adipose Tissue-Derived Mesenchymal Stem Cells in Patients With Severe Buerger's Disease.

Buerger's disease is a rare and severe disease affecting the blood vessels of the limbs. Adipose tissue-derived mesenchymal stem cells (ADSCs) have the potential to cure Buerger's disease when developed as a stem cell drug. In the present study, we conducted a prospective, nonrandomized, no placebo-controlled, phase I/II clinical trial with a 2-year follow-up questionnaire survey. A total of 17 patients were intramuscularly administered autologous ADSCs at a dose of 5 million cells/kg. The incidence of adverse events (AEs), adverse drug reaction (ADR), and serious adverse events (SAEs) was monitored. No ADRs and SAEs related to stem cell treatment occurred during the 6-month follow-up. In terms of efficacy, the primary endpoint was increase in total walking distance (TWD). The secondary endpoint was improvement in rest pain, increase in pain-free walking distance (PFWD), toe-brachial pressure index (TBPI), transcutaneous oxygen pressure (TcPO2), and arterial brachial pressure index (ABPI). ADSCs demonstrated significant functional improvement results including increased TWD, PFWD, and rest pain reduction. No amputations were reported during the 6-month clinical trial period and in the follow-up questionnaire survey more than 2 years after the ADSC injection. In conclusion, intramuscular injection of ADSCs is very safe and is shown to prompt functional improvement in patients with severe Buerger's disease at a dosage of 300 million cells per 60 kg of body weight. However, the confirmatory therapeutic efficacy and angiogenesis need further study.

Phase Ib Open Clinical Trial to Assess the Safety of Autologous Mesenchymal Stem Cells for the Treatment of Nonrevascularizable Critical Lower Limb Ischemia

Background: Critical limb ischemia is a highly disabling disease, characterized by chronic pain at rest, ulceration and tissue tropism attributable to arterial occlusion. Despite significant advances in recent years in treating this disease, there are patients who, for technical reasons or because of the benefit/risk balance, have no therapeutic options other than amputation of the affected limb. Objective: This study’s objective was to assess the feasibility and safety of autologous Adipose Tissue-Derived Mesenchymal Stem Cell (AT-MSC) implantation in patients with lower limb ischemia who are not candidates for surgical or endovascular revascularization. Methods: This is a pragmatic, phase Ib, open, one-arm clinical trial, with a 1-year follow-up after cell implantation. The dose was 1 × 106 AT-MSCs/kg. AT-MSCs were diluted into a final volume of 25 mL of Ringer solution and injected as 25 aliquots of 1 mL into each injection site on the limb. Injection sites were selected below the knee at 25 different sites of the ischemic calf muscle along the tibial and peroneal arteries. Liposuction was done in the abdomen. Results: A total of 7 patients underwent treatment for 21 months. Two patients showed no serious complications with the liposuction, only pain and mild infection. No serious cell implantation-related adverse event occurred during the follow-up, although 2 patients had to undergo amputation. The ankle-brachial index and clinical assessment of the limb improved during the follow-up. Conclusion: In conclusion, AT-MSC treatment of critical limb ischemia is feasible, safe and has promising initial results for salvaging limbs in the short term.

Silicone Substrate with Collagen and Carbon Nanotubes Exposed to Pulsed Current for MSC Osteodifferentiation.

Autologous human adipose tissue-derived mesenchymal stem cells (MSCs) have the potential for clinical translation through their induction into osteoblasts for regeneration. Bone healing can be driven by biophysical stimulation using electricity for activating quiescent adult stem cells. It is hypothesized that application of electric current will enhance their osteogenic differentiation, and addition of conductive carbon nanotubes (CNTs) to the cell substrate will provide increased efficiency in current transmission. Cultured MSCs were seeded and grown onto fabricated silicone-based composites containing collagen and CNT fibers. Chemical inducers, namely, glycerol phosphate, dexamethasone, and vitamin C, were then added to the medium, and pulsatile submilliampere electrical currents (about half mA for 5 cycles at 4 mHz, twice a week) were applied for two weeks. Calcium deposition indicative of MSC differentiation and osteoblastic activity was quantified through Alizarin Red S and spectroscopy. It was found that pulsed current significantly increased osteodifferentiation on silicone-collagen films without CNTs. Under no external current, the presence of 10% (m/m) CNTs led to a significant and almost triple upregulation of calcium deposition. Both CNTs and current parameters did not appear to be synergistic. These conditions of enhanced osteoblastic activities may further be explored ultimately towards future therapeutic use of MSCs.

Isolation of autologous adipose tissue-derived mesenchymal stem cells for bone repair

IntroductionAdipose tissue represents an abundant and accessible source of adult stem cells that can differentiate into cells and tissues of mesodermal origin, including osteogenic cells. MethodsThis paper describes the procedure to obtain a 5-cm3 saline sample, containing the adipose-derived stem cells (ASCs) pellet, starting from lipoaspirate obtained from a conventional abdominal liposuction. ResultsA mean of 2.5×106 cells is isolated for each procedure; 35% (875000) of these are CD34+/CD45− cells, which express a subset of both positive (CD10, CD13, CD44, CD59, CD73, CD90, HLAABC) and negative (CD33, CD39, CD102, CD106, CD146, HLADR) cell-associated surface antigens, characterizing them as ASCs. ConclusionsThis procedure is easy, effective, economic and safe. It allows the harvesting of a significant number of ASCs that are ready for one-step bony regenerative surgical procedures.

Autologous adipose tissue‑derived mesenchymal stem cells are involved in rat liver regeneration following repeat partial hepatectomy.

Adipose tissue-derived mesenchymal stem cells (ADSCs) have been considered to be attractive and readily available adult mesenchymal stem cells, and they are becoming increasingly popular for use in regenerative cell therapy, as they are readily accessible through minimally invasive techniques. The present study investigated whether autologous ADSC transplantation promoted liver regeneration following a repeat partial hepatectomy in rats. The rats were divided into three groups as follows: 70% partial hepatectomy (PH) group; repeat PH (R-PH) group and R-PH/ADSC group, subjected to R-PH and treated with autologous ADSCs via portal vein injection. In each group, the rats were sacrificed at different time points postoperatively in order to evaluate the changes in liver function and to estimate the liver regenerative response. The expression of proliferating cell nuclear antigen (PCNA) labeling index in the liver was measured using immunohistochemistry. The expression levels of hepatocyte growth factor (HGF) mRNA were measured using reverse transcription polymerase chain reaction. The results showed that regeneration of the remaining liver following R-PH was significantly promoted by ADSC transplantation, as shown by a significant increase in liver to body weight ratio and the PCNA labeling index at 24 h post-hepatectomy. Additionally, ADSC transplantation markedly inhibited the elevation of serum levels of alanine aminotransferase, aspartate aminotransferase and total bilirubin, increased HGF content and also attenuated hepatic vacuolar degeneration 24 h postoperatively. Furthermore, the liver was found to almost fully recover from hepatocellular damage due to hepatectomy among the three groups at 168 h postoperatively. These results indicated that autologous ADSC transplantation enhanced the regenerative capacity of the remnant liver tissues in the early phase following R-PH.

Osteochondral Regeneration of the Loading-bearing Site Using a Scaffold Free Three-dimensional Construct of Adipose Tissue-derived Mesenchymal Stem Cells in Pigs

Background: Many surgical strategies for reconstruction of both bone and cartilage have ever been investigated to restore joint structure and function in the late stages of Osteoarthritis (OA). This study was designed to investigate the regeneration of articular cartilage and subchondral bone in the loading-bearing site using a three-dimensional (3D) construct of autologous adipose tissue-derived mesenchymal stem cells (AT-MSCs). Methods: A 3D construct consisting of approximately 1,920 spheroids each containing 5.0 × 104 AT-MSCs was implanted into an osteochondral defect (with a diameter of 6.8 mm and a depth of 6 mm) in the right femoral medial condyle in five adult mini-pigs. The contralateral (left femoral) defect was the control. At three and six months post-operatively, the defects were evaluated using both CT and MR imaging. The radiolucent volume (RV, mm3) of the defects was calculated based on the multiplanar reconstruction of the CT images. MR images and gross and histologic pathology features were scored using a modified-MOCART system and the ICRS system, respectively, at six months post-operatively. Results: The percentages of RVs at three and six months compared with those immediately after the surgeries were significantly decreased in the implanted defects compared with the control defects. The total scores of modified- MOCART system were also significantly increased in the implanted sites comparing to the controls. Although there were no statistical differences in the average of gross scores, the average histological scores were significantly higher in the implanted sites than in the control sites. Conclusion: This is the first report suggesting that implantation of a scaffold-free three dimensional construct of only AT-MSCs into the osteochondral defect regenerates the original cartilage and subchondral bone structures over six months post-operatively in the loading-bearing site of large animal.

Implantation of Autologous Adipose Tissue-Derived Mesenchymal Stem Cells in Foot Fat Pad in Rats.

The foot fat pad (FFP) bears body weight and may become a source of foot pain during aging. This study investigated the regenerative effects of autologous adipose tissue-derived mesenchymal stem cells (AT-MSCs) in the FFP of rats. Fat tissue was harvested from a total of 30 male Sprague-Dawley rats for isolation of AT-MSCs. The cells were cultured, adipogenic differentiation was induced for 1 week, and the AT-MSCs were labeled with fluorescent dye before injection. AT-MSCs (5 × 10(4) in 50 µL of saline) were injected into the second infradigital pad in the right hindfoot of the rat of origin. Saline only (50 µL) was injected into the corresponding fat pad in the left hind paw of each rat. Rats (n = 10) were euthanized at 1, 2, and 3 weeks, and the second infradigital fat pads were dissected for histologic examination. The fluorescence-labeled AT-MSCs were present in the foot pads throughout the 3-week experimental period. On histologic testing, the area of fat pad units (FPUs) in the fat pads that received AT-MSC injections was greater than that in the control fat pads. Although the thickness of septae was not changed by AT-MSC injections, the density of elastic fibers in the septae was increased in the fat pads with implanted AT-MSCs. In this short-term study, the implanted AT-MSCs largely survived and might have stimulated the expansion of individual FPUs and increased the density of elastic fibers in the FFP in this rat model. These data support the development of stem cell therapies for age-associated degeneration in FFP in humans.