Results of the JRS-I LRA0401 and LRB0402 Japan Rhabdomyosarcoma Study Group trials for low-risk embryonal rhabdomyosarcoma.

BACKGROUND: Failure-free survival (FFS) rates of low-risk patients with rhabdomyosarcoma improved in Intergroup Rhabdomyosarcoma Study IV after the escalation of cyclophosphamide total dose to 26.4 g/m2. However, this dose may increase the risk of adverse events, including infertility, in some patients. The JRS-I LRA0401 and LRB0402 protocols aimed to reduce the cyclophosphamide dose to 9.6 g/m2 and 17.6 g/m2, respectively, without decreasing the FFS rates. METHODS: Subgroup-A patients received eight cycles (24 weeks) of vincristine, actinomycin D, and 1.2 g/m2/cycle cyclophosphamide. Subgroup-B patients received eight cycles (24 weeks) of vincristine, actinomycin D, and 2.2 g/m2/cycle cyclophosphamide, followed by six cycles (24 weeks) of vincristine and actinomycin D. Group II/III patients in both subgroups received radiotherapy. RESULTS: In subgroup A (n = 12), the 3-year FFS rate was 83% (95% confidence interval [CI], 48-96), and the 3-year overall survival (OS) rate was 100%. Only one isolated local recurrence was observed (8.3%). There were no unexpected grade-4 toxicities and no deaths. In subgroup B (n = 16), the 3-year FFS and OS rates were 88% (95% CI, 59-97) and 94% (95% CI, 63-99), respectively. There were no unexpected grade 4 toxicities and no deaths. CONCLUSIONS: Shorter duration therapy using vincristine, actinomycin D, and lower dose cyclophosphamide with or without radiotherapy for patients with low-risk subgroup A rhabdomyosarcoma (JRS-I LRA0401 protocol) and moderate reduction of cyclophosphamide dose for patients with low-risk subgroup B rhabdomyosarcoma (JRS-I LRB0402 protocol) did not compromise FFS.

Production of recombinant humanized monoclonal anti-human neutrophil antigen (HNA) antibodies with potential applicability as standard antibodies.

BACKGROUND: Antibodies against human neutrophil antigen (HNA) are involved in the pathogenesis of neonatal alloimmune neutropenia, autoimmune neutropenia, and transfusion-related acute lung injury. The present methods for anti-HNA antibody identification strongly depend on the presence of standard antisera with known allo/isospecificities. Here, we aimed to produce recombinant humanized antibodies to HNA from available mouse monoclonal antibodies (MoAbs). STUDY DESIGN AND METHODS: RNAs were extracted from available hybridoma cells producing mouse anti-HNA antibodies recognizing HNA-1a (TAG-1), -1b (TAG-2), -2 (TAG-4), and FcγRIIIb, and the cDNA was synthesized. Recombinant fragments consisting of the variable regions of the H and L chains of the mouse MoAb ligated to the constant region of human IgG were incorporated into an expression vector and transfected into CHO cells. Antibody specificity of the selected humanized monoclonal antibodies was confirmed, and tested by the participants of the ISBT Granulocyte Immunobiology Working Party (GIWP) workshop 2020. RESULTS: GIFT results confirmed the specific reactivity of TAGH-1 to -4, except for a cross-reactivity of TAGH-2 with HNA-1a/a neutrophils, only in flow-cytometry. MAIGA results showed clear specificity of all humanized antibodies, but the selection of the appropriate capture monoclonal antibody was essential for the test. The results of the ISBT GIWP showed high concordance among the labs. CONCLUSIONS: These are the first humanized monoclonal antibodies to HNA-1 and HNA-2 antigens produced and they will be important standard reagents for laboratories testing for neutrophil antibodies. We plan to have these humanized MoAbs available through WHO.

Pharmacological Difference Between Platelet Aggregations in Cardioembolic Stroke Patients with Direct Oral Anticoagulants: A Pilot Study.

Background Selecting the appropriate direct oral anticoagulants (DOACs) for embolic ischemic stroke patients, especially on concurrent antiplatelet therapy, is important. However, a limited number of studies have reported on the pharmacological differences in platelet aggregation of each DOAC. We aimed to evaluate the antiplatelet effects of selected DOACs, by comparing dabigatran (a direct oral thrombin inhibitor) and factor Xa (FXa) inhibitors (apixaban and rivaroxaban) in patients who had suffered a cardioembolic stroke. Methods We retrospectively evaluated 12 patients diagnosed with a cardioembolic stroke who took any DOAC without an antiplatelet drug and underwent platelet aggregation tests within 60 days from the onset of symptoms. The platelet aggregation tests were analyzed by both light transmission aggregometry and VerifyNow®. Results Six patients (50%) took dabigatran, while the other six (50%) took an FXa inhibitor (n = 4 for apixaban and n = 2 for rivaroxaban). From the light transmission aggregometry analysis, it was found that the maximal extent of aggregation for adenosine diphosphate (ADP) was significantly higher with dabigatran than with FXa inhibitors, and the ED50 value of ADP on platelet aggregation was significantly lower with dabigatran than with FXa inhibitors. Moreover, the VerifyNow® analyses revealed that P2Y12 reaction units were significantly higher with dabigatran than with FXa inhibitors. Conclusions Dabigatran had little impact on platelet aggregation compared to FXa inhibitors in patients who had suffered a cardioembolic stroke with atrial fibrillation, and who took DOACs for secondary prevention within 60 days from the onset.

Anti-human neutrophil antigen-1a, -1b, and -2 antibodies in neonates and children with immune neutropenias analyzed by extracted granulocyte antigen immunofluorescence assay.

BACKGROUND: Anti-human neutrophil antigen (HNA) antibodies have been implicated in the development of neonatal alloimmune neutropenia (NAN) and autoimmune neutropenia (AIN). There are many conventional assay methods that detect anti-HNA antibodies. However, a method to measure multiple samples and detect several anti-HNA antibodies simultaneously is needed. STUDY DESIGN AND METHODS: We developed a new method, the extracted granulocyte antigen immunofluorescence assay (EGIFA), to analyze anti-HNA-1a, -1b, and -2 antibodies in sera. The results obtained by EGIFA were evaluated in comparison with those from several standard assay methods. Anti-HNA antibodies in serum samples from nine familial cases with suspected NAN (n = 19) and children with suspected AIN (n = 88) were also measured by EGIFA. RESULTS: The evaluation of nine serum samples with anti-HNA antibodies suggested that EGIFA demonstrated equivalent specificity and superior sensitivity to monoclonal antibody-specific immobilization of granulocyte antigens and had comparable sensitivity to the granulocyte indirect immunofluorescence test. EGIFA successfully detected anti-HNA-1a or -1b antibodies in seven of nine familial cases with suspected NAN. EGIFA detected anti-HNA antibodies in 40.9% of children with suspected AIN. Among them, isolated anti-HNA-1a or -1b antibody was detected in 4.5 or 12.5% of children, respectively, and anti-HNA-2 antibody was identified in 3.4% of children. The 30.8% (16 of 52) of children negative for anti-HNA antibody by EGIFA were positive for anti-HLA antibody. CONCLUSION: EGIFA facilitated the measurement of anti-HNA-1a, -1b, and/or -2 antibodies in sera. The prompt measurement of anti-HNA antibodies will improve the diagnosis and clinical management of patients with suspected NAN or AIN.

Inflammation of colon adenoma in the setting of type 1 autoimmune pancreatitis.

Although immunoglobulin G4-related diseases (IgG4-RD) have been found to affect many organs, little is known about their effects on the colonic mucosae. Pathological examination of colon adenomas has shown inflammatory cell infiltration into the stroma. We therefore assessed the clinicopathological characteristics of colon adenomas in patients with type 1 autoimmune pancreatitis (AIP-1), a representative IgG4-RD. Both colon adenomas from patients with (IgG4 adenomas) and without (Non-IgG4 adenomas) IgG4-RD were characterized by moderate to severe lymphoplasmacytic and eosinophilic inflammation without fibrosis or phlebitis. The ratio of IgG4-positive to IgG-positive plasma cells (IgG4/IgG ratio) and the numbers of IgG4-positive plasma cells were significantly higher in IgG4 adenomas than in Non-IgG4 adenomas. IgG4-positive plasma cells tended to be distributed diffusely in lower areas of the mucosae in IgG4 adenomas. We were unable to confirm whether IgG4 adenomas constituted an IgG4-RD. However, IgG4 adenomas in the setting of IgG4-RD may provide useful pathological information, supplementing a diagnosis of IgG4-RD outside the colon, or may facilitate examination for IgG4-RD, especially AIP-1. IgG4 adenomas warrant further investigation.

Intravenous Infusion of Autologous Mesenchymal Stem Cells Expanded in Auto Serum for Chronic Spinal Cord Injury Patients: A Case Series.

Objective: The safety, feasibility, and potential functional improvement following the intravenous infusion of mesenchymal stem cells (MSCs) were investigated in patients with chronic severe spinal cord injury (SCI). Methods: The intravenous infusion of autologous MSCs cultured in auto-serum under Good Manufacturing Practices (GMP) was administered to seven patients with chronic SCI (ranging from 1.3 years to 27 years after the onset of SCI). In addition to evaluating feasibility and safety, neurological function was evaluated using the American Spinal Injury Association Impairment Scale (AIS), International Standards for Neurological Classification of Spinal Cord Injury (ISCSCI-92), and Spinal Cord Independence Measure III (SCIM-III). Results: No serious adverse events occurred. Neither CNS tumors, abnormal cell growth, nor neurological deterioration occurred in any patients. While this initial case series was not blinded, significant functional improvements and increased quality of life (QOL) were observed at 90 and 180 days post-MSC infusion compared to pre-infusion status. One patient who had an AIS grade C improved to grade D within six months after MSC infusion. Conclusions: This case series suggests that the intravenous infusion of autologous MSCs is a safe and feasible therapeutic approach for chronic SCI patients. Furthermore, our data showed significant functional improvements and better QOL after MSC infusion in patients with chronic SCI. A blind large-scale study will be necessary to fully evaluate this possibility.

Clinical significance of human neutrophil antigen-1 antibodies in children with neutropenia.

Primary autoimmune neutropenia in young children is characterized by chronic neutropenia and positivity for antibodies against human neutrophil antigens (HNAs). This study analyzed the clinical characteristics of 402 children with neutropenia to identify differences between those with and without HNA-1 antibodies (HNA1abs). HNAabs in sera were detected by granulocyte immunofluorescence testing using flow cytometry. Relative fluorescence intensity (RFI) values were used to divide patients into positive (PG, n = 302), borderline (BG, n = 34), and negative (NG, n = 66) groups. The antibodies reacted to HNA-1a alone (59%), HNA-1b alone (1%), and HNA-1a/1b (40%). The PG had a significantly lower absolute neutrophil count before definitive diagnosis and a 1.6- to 2-times greater risk of hospitalization during neutropenia than the other groups. The median duration of neutropenia was longest in the PG at 25 months, followed by 20 months in the BG and 14 months in the NG. This large-scale cohort characterizes clinically distinct groups using the RFI value for HNA1abs in young children with neutropenia. Detection of HNA1abs may aid in understanding the clinical characteristics of children with neutropenia.

Intravenous administration of auto serum-expanded autologous mesenchymal stem cells in stroke.

Transplantation of human mesenchymal stem cells has been shown to reduce infarct size and improve functional outcome in animal models of stroke. Here, we report a study designed to assess feasibility and safety of transplantation of autologous human mesenchymal stem cells expanded in autologous human serum in stroke patients. We report an unblinded study on 12 patients with ischaemic grey matter, white matter and mixed lesions, in contrast to a prior study on autologous mesenchymal stem cells expanded in foetal calf serum that focused on grey matter lesions. Cells cultured in human serum expanded more rapidly than in foetal calf serum, reducing cell preparation time and risk of transmissible disorders such as bovine spongiform encephalomyelitis. Autologous mesenchymal stem cells were delivered intravenously 36-133 days post-stroke. All patients had magnetic resonance angiography to identify vascular lesions, and magnetic resonance imaging prior to cell infusion and at intervals up to 1 year after. Magnetic resonance perfusion-imaging and 3D-tractography were carried out in some patients. Neurological status was scored using the National Institutes of Health Stroke Scale and modified Rankin scores. We did not observe any central nervous system tumours, abnormal cell growths or neurological deterioration, and there was no evidence for venous thromboembolism, systemic malignancy or systemic infection in any of the patients following stem cell infusion. The median daily rate of National Institutes of Health Stroke Scale change was 0.36 during the first week post-infusion, compared with a median daily rate of change of 0.04 from the first day of testing to immediately before infusion. Daily rates of change in National Institutes of Health Stroke Scale scores during longer post-infusion intervals that more closely matched the interval between initial scoring and cell infusion also showed an increase following cell infusion. Mean lesion volume as assessed by magnetic resonance imaging was reduced by >20% at 1 week post-cell infusion. While we would emphasize that the current study was unblinded, did not assess overall function or relative functional importance of different types of deficits, and does not exclude placebo effects or a contribution of recovery as a result of the natural history of stroke, our observations provide evidence supporting the feasibility and safety of delivery of a relatively large dose of autologous mesenchymal human stem cells, cultured in autologous human serum, into human subjects with stroke and support the need for additional blinded, placebo-controlled studies on autologous mesenchymal human stem cell infusion in stroke.

Intravenous Infusion of Autoserum-Expanded Autologous Mesenchymal Stem Cells in Patients With Chronic Brain Injury: Protocol for a Phase 2 Trial.

BACKGROUND: Brain injuries resulting from motor vehicle accidents and falls, as well as hypoxic insults and other conditions, are one of the leading causes of disability and death in the world. Current treatments are limited but include continuous rehabilitation, especially for chronic brain injury. Recent studies have demonstrated that the intravenous infusion of mesenchymal stem cells (MSCs) has therapeutic efficacy for several neurological diseases, including stroke and spinal cord injury. OBJECTIVE: The objective of our investigator-initiated clinical trial is to assess the safety and potential efficacy of the intravenous infusion of autoserum-expanded autologous MSCs for patients with chronic brain injury. METHODS: The (phase 2) trial will be a single-arm, open-label trial with the primary objective of confirming the safety and efficacy of autoserum-expanded autologous MSCs (STR-01; produced under good manufacturing practices) when administered to patients with chronic brain injury. The estimated number of enrolled participants is 6 to 20 patients with a modified Rankin Scale grade of 3 to 5. The assessment of safety and the proportion of cases in which the modified Rankin Scale grade improves by 1 point or more at 180 days after the injection of STR-01 will be performed after MSC infusion. RESULTS: We received approval for our clinical trial from the Japanese Pharmaceuticals and Medical Devices Agency on December 12, 2017. The trial will be completed on June 11, 2023. The registration term is 5 years. The recruitment of the patients for this trial started on April 20, 2018, at Sapporo Medical University Hospital in Japan. CONCLUSIONS: Our phase 2 study will aim to address the safety and efficacy of the intravenous infusion of MSCs for patients with chronic brain injury. The use of STR-01 has been performed for patients with cerebral infarction and spinal cord injury, providing encouraging results. The potential therapeutic efficacy of the systemic administration of autoserum-expanded autologous MSCs for chronic brain injury should be evaluated, given its safety and promising results for stroke and spinal cord injury. TRIAL REGISTRATION: Japan Medical Association Center for Clinical Trials JMA-IIA00333; https://tinyurl.com/nzkdfnbc. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/37898.

Repeated infusion of mesenchymal stem cells maintain the condition to inhibit deteriorated motor function, leading to an extended lifespan in the SOD1G93A rat model of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative fatal disorder in which motor neurons within the brain and spinal cord degenerate. A single infusion of mesenchymal stem cells (MSCs) delays disease progression by protecting motor neurons and restoring the blood-spinal cord barrier in the SOD1G93A transgenic ALS rat model. However, the therapeutic effect of a single infusion of MSCs is transient and does not block disease progression. In this study, we demonstrated that repeated administration of MSCs (weekly, four times) increased the survival period, protected motor functions, and reduced deterioration of locomotor activity compared to a single infusion and vehicle infusion, after which rats displayed progressive deterioration of hind limb function. We also compared the days until gait ability was lost in rats and found that the repeated-infused group maintained gait ability compared to the single-infusion and vehicle-infusion groups. These results suggest that repeated administration of MSCs may prevent the deterioration of motor function and extend the lifespan in ALS.

Intravenous infusion of mesenchymal stem cells delays disease progression in the SOD1G93A transgenic amyotrophic lateral sclerosis rat model.

ALS is a devastating neurodegenerative disease with few curative strategies. Both sporadic and familial ALS display common clinical features that show progressive paralysis. The pathogenesis remains unclear, but disruption of the blood-spinal cord barrier (BSCB) may contribute to the degeneration of motor neurons. Thus, restoration of the disrupted BSCB and neuroprotection for degenerating motor neurons could be therapeutic targets. We tested the hypothesis that an intravenous infusion of MSCs would delay disease progression through the preservation of BSCB function and increased expression of a neurotrophic factor, neurturin, in SOD1G93A ALS rats. When the open-field locomotor function was under 16 on the Basso, Beattie, and Bresnahan (BBB) scoring scale, the rats were randomized into two groups; one received an intravenous infusion of MSCs, while the other received vehicle alone. Locomotor function was recorded using BBB scoring and rotarod testing. Histological analyses, quantitative reverse transcription-polymerase chain reaction (qRT-PCR), were performed. The MSC group exhibited reduced deterioration of locomotor activity compared to the vehicle group, which displayed progressive deterioration of hind limb function. We observed the protection of motor neuron loss and preservation of microvasculature using Evans blue leakage and immunohistochemical analyses in the MSC group. Confocal microscopy revealed infused green fluorescent protein+ (GFP+) MSCs in the spinal cord, and the GFP gene was detected by nested PCR. Neurturin expression levels were significantly higher in the MSC group. Thus, restoration of the BSCB and the protection of motor neurons might be contributing mechanisms to delay disease progression in SOD1G93A ALS rats.

Intravenous infusion of auto serum-expanded autologous mesenchymal stem cells in spinal cord injury patients: 13 case series.

BACKGROUND: Although spinal cord injury (SCI) is a major cause of disability, current therapeutic options remain limited. Recent progress in cellular therapy with mesenchymal stem cells (MSCs) has provided improved function in animal models of SCI. We investigated the safety and feasibility of intravenous infusion of MSCs for SCI patients and assessed functional status after MSC infusion. METHODS: In this phase 2 study of intravenous infusion of autologous MSCs cultured in auto-serum, a single infusion of MSCs under Good Manufacturing Practice (GMP) production was delivered in 13 SCI patients. In addition to assessing feasibility and safety, neurological function was assessed using the American Spinal Injury Association Impairment Scale (ASIA), International Standards for Neurological and Functional Classification of Spinal Cord (ISCSCI-92). Ability of daily living was assessed using Spinal Cord Independence Measure (SCIM-III). The study protocol was based on advice provided by the Pharmaceuticals and Medical Devices Agency in Japan. The trial was registered with the Japan Medical Association (JMA-IIA00154). RESULTS: No serious adverse events were associated with MSC injection. There was neurologic improvement based on ASIA grade in 12 of the 13 patients at six months post-MSC infusion. Five of six patients classified as ASIA A prior to MSC infusion improved to ASIA B (3/6) or ASIA C (2/6), two ASIA B patients improved to ASIA C (1/2) or ASIA D (1/2), five ASIA C patients improved and reached a functional status of ASIA D (5/5). Notably, improvement from ASIA C to ASIA D was observed one day following MSC infusion for all five patients. Assessment of both ISCSCI-92, SCIM-III also demonstrated functional improvements at six months after MSC infusion, compared to the scores prior to MSC infusion in all patients. CONCLUSION: While we emphasize that this study was unblinded, and does not exclude placebo effects or a contribution of endogenous recovery or observer bias, our observations provide evidence supporting the feasibility, safety and functional improvements of infused MSCs into patients with SCI.

Topical insulin-like growth factor 1 treatment using gelatin hydrogels for glucocorticoid-resistant sudden sensorineural hearing loss: a prospective clinical trial.

BACKGROUND: Sudden sensorineural hearing loss (SSHL) is a common condition in which patients lose the hearing in one ear within 3 days. Systemic glucocorticoid treatments have been used as standard therapy for SSHL; however, about 20% of patients do not respond. We tested the safety and efficacy of topical insulin-like growth factor 1 (IGF1) application using gelatin hydrogels as a treatment for SSHL. METHODS: Patients with SSHL that showed no recovery to systemic glucocorticoid administration were recruited. We applied gelatin hydrogels, impregnated with recombinant human IGF1, into the middle ear. The primary outcome measure was the proportion of patients showing hearing improvement 12 weeks after the test treatment. The secondary outcome measures were the proportion of patients showing improvement at 24 weeks and the incidence of adverse events. The null hypothesis was that 33% of patients would show hearing improvement, as was reported for a historical control after hyperbaric oxygen therapy. RESULTS: In total, 25 patients received the test treatment at a median of 23 days (range 15-32) after the onset of SSHL, between 2007 and 2009. At 12 weeks after the test treatment, 48% (95% CI 28% to 69%; P = 0.086) of patients showed hearing improvement, and the proportion increased to 56% (95% CI 35% to 76%; P = 0.015) at 24 weeks. No serious adverse events were observed. CONCLUSIONS: Topical IGF1 application using gelatin hydrogels is well tolerated and may be efficacious for hearing recovery in patients with SSHL that is resistant to systemic glucocorticoids.

Intramuscular transplantation of G-CSF-mobilized CD34(+) cells in patients with critical limb ischemia: a phase I/IIa, multicenter, single-blinded, dose-escalation clinical trial.

A number of preclinical studies have indicated the therapeutic potential of endothelial progenitor cells for vascular regeneration in ischemic diseases. A phase I/IIa clinical trial of transplantation of autologous CD34(+) cells, the endothelial and hematopoietic progenitor-enriched fraction, was performed in no-option patients with atherosclerotic peripheral artery disease or Buerger's disease with critical limb ischemia (CLI). CD34(+) cells were isolated from the G-CSF-mobilized apheresis product using a magnetic cell sorting system. CD34(+) cells (10(5)/kg, n = 6; 5 x 10(5)/kg, n = 8; or 10(6)/kg, n = 3) were injected i.m. into the leg with more severe ischemia. The Efficacy Score, representing changes in the toe brachial pressure index (TBPI), Wong-Baker FACES pain rating scale, and total walking distance 12 weeks after cell transplantation, the primary endpoint, was positive, indicating improvement in limb ischemia in all patients, although no significant dose-response relationship was observed. During the 12-week observation after cell therapy, the Wong-Baker FACES pain rating scale, TBPI, transcutaneous partial oxygen pressure, total or pain-free walking distance, and ulcer size serially improved in all patients. No death or major amputation occurred, and severe adverse events were rare, although mild to moderate events relating to G-CSF and leukapheresis were frequent during the 12-week follow-up. In conclusion, the outcomes of this prospective clinical study indicate the safety and feasibility of CD34(+) cell therapy in patients with CLI. Favorable trends in efficacy parameters encourage a randomized and controlled trial in the future.

Prevention of neointimal hyperplasia induced by an endovascular stent via intravenous infusion of mesenchymal stem cells.

OBJECTIVE: In-stent restenosis after percutaneous transluminal angioplasty and stenting (PTAS) due to neointimal hyperplasia is a potential cause of clinical complications, including repeated revascularization and ischemic events. Neointimal hyperplasia induced by an inflammatory response to the stent strut may be a possible mechanism of in-stent restenosis. Intravenous infusion of bone marrow-derived mesenchymal stem cells (MSCs) has been reported to show therapeutic efficacy for cerebral stroke, presumably by an antiinflammatory effect. This study aimed to determine whether MSCs can reduce or prevent neointimal hyperplasia induced by an endovascular stent. METHODS: In this study, two types of bare metal stents were deployed using a porcine (mini-pig) model. One stent was implanted in the common carotid artery (CCA), which is considered quite similar to the human CCA, and the other was inserted in the superficial cervical artery (SCA), which is similar in size to the human middle cerebral artery. Angiographic images, intravascular ultrasound (IVUS) imaging, and microscopic images were used for analysis. RESULTS: Angiographic images and IVUS studies revealed that intravenous infusion of MSCs immediately after deployment of stents prevented in-stent stenosis of the CCA and SCA. Histological analysis also confirmed that inflammatory responses around the stent struts were reduced in both the stented CCA and SCA in the mini-pig. CONCLUSIONS: Intravenous infusion of MSCs inhibited the inflammatory reaction to an implanted stent strut, and prevented progressive neointimal hyperplasia in the stented CCA and SCA in a porcine model. Thus, MSC treatment could attenuate the recurrence of cerebral ischemic events after stenting.

Elevated brain derived neurotrophic factor levels in plasma reflect in vivo functional viability of infused mesenchymal stem cells for stroke in rats.

BACKGROUND: Intravenous infusion of mesenchymal stem cells (MSCs) derived from adult bone marrow elicits functional recovery in rat stroke models and clinical studies in patients are ongoing. Brain derived neurotrophic factor (BDNF) is a neurotrophic factor produced by MSCs and may contribute to their therapeutic efficacy. The purpose of the current study was to determine if BDNF is elevated in infarcted brain and in which compartment of blood (plasma or serum) after intravenous MSC infusion in a middle cerebral artery occlusion (MCAO) model in the rat. METHODS: In rats, a permanent middle cerebral artery occlusion (MCAO) was induced by intraluminal vascular occlusion with a microfilament and MSCs were intravenously administered 6 h after right MCAO induction. Enzyme-linked immunosorbent assay (ELISA) analysis of brain, serum and plasma BDNF were performed after the MSC infusion following the MCAO induction. Lesion volume was assessed using magnetic resonance imaging. Functional outcome was assessed using the Limb Placement Test. RESULTS: Infused MSCs reduced lesion volume and elicited functional improvement compared to the vehicle infused group. ELISA analysis of the MSC treated group revealed an increase BDNF levels in the infarcted hemisphere of the brain and plasma, but not in serum. The MSC group showed a greater increase in BDNF levels than sham control. In the MSC group, the expression of increased plasma BDNF levels correlated with increased brain BDNF levels. CONCLUSIONS: These results support the hypothesis that BDNF levels in plasma, but not serum, may be more appropriate to detect circulating BDNF in vivo following MSC infusion in a cerebral infarction rat model of ischemic stroke. Further, plasma BDNF might reflect in vivo functional viability of infused MSCs after stroke.

Long-term clinical outcome after intramuscular implantation of bone marrow mononuclear cells (Therapeutic Angiogenesis by Cell Transplantation [TACT] trial) in patients with chronic limb ischemia.

BACKGROUND: Angiogenic cell therapy by intramuscular injection of autologous bone marrow mononuclear cells was first attempted in patients with peripheral artery disease (PAD) with critical limb ischemia, and the feasibility was shown by a randomized controlled Therapeutic Angiogenesis by Cell Transplantation (TACT) study. METHODS AND RESULTS: The present study was designed to assess the 3-year safety and clinical outcomes of this angiogenic cell therapy by investigating the mortality and leg amputation-free interval as primary end points. The median follow-up time for surviving patients was 25.3 months (range, 0.8-69.0 months), and 3-year overall survival rates were 80% (95% CI 68-91) in patients with atherosclerotic peripheral arterial disease (11 died in 74 patients) and 100% (no death) in 41 patients with thromboangiitis obliterans (TAO; Buerger's disease). Three-year amputation-free rate was 60% (95% CI 46-74) in PAD and 91% (95% CI 82-100) in patients with TAO. The multivariate analysis revealed that the severity of rest pain and repeated experience of bypass surgery were the prognostic factors negatively affecting amputation-free interval. The significant improvement in the leg pain scale, ulcer size, and pain-free walking distance was maintained during at least 2 years after the therapy, although the ankle brachial index and transcutaneous oxygen pressure value did not significantly change. CONCLUSIONS: The angiogenic cell therapy using bone marrow mononuclear cells can induce a long-term improvement in limb ischemia, leading to extension of amputation-free interval. The safety and efficacy are not inferior to the conventional revascularization therapies.

Intravenous infusion of mesenchymal stem cells for protection against brainstem infarction in a persistent basilar artery occlusion model in the adult rat.

OBJECTIVE: Morbidity and mortality in patients with posterior circulation stroke remains an issue despite advances in acute stroke therapies. The intravenous infusion of mesenchymal stem cells (MSCs) elicits therapeutic efficacy in experimental supratentorial stroke models. However, since there are few reliable animal models of ischemia in the posterior circulation, the therapeutic approach with intravenous MSC infusion has not been tested. The objective of this study was to test the hypothesis that intravenously infused MSCs provide functional recovery in a newly developed model of brainstem infarction in rats. METHODS: Basilar artery (BA) occlusion (BAO) was established in rats by selectively ligating 4 points of the proximal BA with 10-0 nylon monofilament suture. The intravenous infusion of MSCs was performed 1 day after BAO induction. MRI and histological examinations were performed to assess ischemic lesion volume, while multiple behavioral tests were performed to evaluate functional recovery. RESULTS: The MSC-treated group exhibited a greater reduction in ischemic lesion volume, while behavioral testing indicated that the MSC-infused group had greater improvement than the vehicle group 28 days after the MSC infusion. Accumulated infused MSCs were observed in the ischemic brainstem lesion. CONCLUSIONS: Infused MSCs may provide neuroprotection to facilitate functional outcomes and reduce ischemic lesion volume as evaluated in a newly developed rat model of persistent BAO.

Intravenous infusion of mesenchymal stem cells promotes functional recovery in a rat model of chronic cerebral infarction.

OBJECTIVE: Intravenous infusion of mesenchymal stem cells (MSCs) derived from adult bone marrow improves behavioral function in rat models of cerebral infarction. Although clinical studies are ongoing, most studies have focused on the acute or subacute phase of stroke. In the present study, MSCs derived from bone marrow of rats were intravenously infused 8 weeks after the induction of a middle cerebral artery occlusion (MCAO) to investigate whether delayed systemic injection of MSCs improves functional outcome in the chronic phase of stroke in rats. METHODS: Eight weeks after induction of the MCAO, the rats were randomized and intravenously infused with either MSCs or vehicle. Ischemic volume and behavioral performance were examined. Blood-brain barrier (BBB) integrity was assessed by quantifying the leakage of Evans blue into the brain parenchyma after intravenous infusion. Immunohistochemical analysis was also performed to evaluate the stability of the BBB. RESULTS: Motor recovery was better in the MSC-treated group than in the vehicle-treated group, with rapid improvement (evident at 1 week post-infusion). In MSC-treated rats, reduced BBB leakage and increased microvasculature/repair and neovascularization were observed. CONCLUSIONS: These results indicate that the systemic infusion of MSCs results in functional improvement, which is associated with structural changes in the chronic phase of cerebral infarction, including in the stabilization of the BBB.

Preservation of interhemispheric cortical connections through corpus callosum following intravenous infusion of mesenchymal stem cells in a rat model of cerebral infarction.

Systemic administration of mesenchymal stem cells (MSCs) following cerebral infarction exerts functional improvements. Previous research has suggested potential therapeutic mechanisms that promote neuroprotection and synaptogenesis. These include secretion of neurotrophic factors, remodeling of neural circuits, restoration of the blood brain barrier, reduction of inflammatory infiltration and demyelination, and elevation of trophic factors. In addition to these mechanisms, we hypothesized that restored interhemispheric bilateral motor cortex connectivity might be an additional mechanism of functional recovery. In the present study, we have shown, with both MRI diffusion tensor imaging (DTI) and neuroanatomical tracing techniques using an adeno-associated virus (AAV) expressing GFP, that there was anatomical restoration of cortical interhemispheric connections through the corpus callosum after intravenous infusion of MSCs in a rat middle cerebral artery occlusion (MCAO) stroke model. Moreover, the degree of connectivity was greater in the MSC-treated group than in the vehicle-infused group. In accordance, both the thickness of corpus callosum and synaptic puncta in the contralateral (non-infarcted) motor cortex connected to the corpus callosum were greater in the MSC-treated group than in the vehicle group. Together, these results suggest that distinct preservation of interhemispheric cortical connections through corpus callosum was promoted by intravenous infusion of MSCs. This anatomical preservation of the motor cortex in the contralateral hemisphere may contribute to functional improvements following MSC therapy for cerebral stroke.