Comparison of reamer irrigator aspirator (RIA) suspension versus bone marrow aspirate concentrate (BMC) for percutaneous treatment of long bone nonunions-A preclinical canine model.

OBJECTIVE: To compare the bone healing effects of percutaneously delivered bone marrow aspirate concentrate (BMC) versus reamer irrigator aspirator (RIA) suspension in a validated preclinical canine ulnar nonunion model. We hypothesized that BMC would be superior to RIA in inducing bone formation across a nonunion site after percutaneous application. The null hypothesis was that BMC and RIA would be equivalent. METHODS: A bilateral ulnar nonunion model (n= 6; 3 matched pairs) was created. Eight weeks after segmental ulnar ostectomy, RIA from the ipsilateral femur and BMC from the proximal humerus were harvested and percutaneously administered into either the left or right ulnar defect. The same volume (3 ml) of RIA suspension and BMC were applied on each side. Eight weeks after treatment, the dogs were euthanized, and the nonunions were evaluated using radiographic, biomechanical, and histologic assessments. RESULTS: All dogs survived for the intended study duration, formed radiographic nonunions 8 weeks after segmental ulnar ostectomy, and underwent the assigned percutaneous treatment. Radiographic and macroscopic assessments of bone healing at the defect sites revealed superior bridging-callous formation in BMC-treated nonunions. Histologic analyses revealed greater amount of bony bridging and callous formation in the BMC group. Biomechanical testing of the treated nonunions did not reveal any significant differences. CONCLUSION: Bone marrow aspirate concentrate (BMC) had important advantages over Reamer Irrigator Aspirator (RIA) suspension for percutaneous augmentation of bone healing in a validated preclinical canine ulnar nonunion model based on clinically relevant radiographic and histologic measures of bone formation.

Long-term hematopoietic transfer of the anti-cancer and lifespan-extending capabilities of a genetically engineered blood system by transplantation of bone marrow mononuclear cells.

A causal relationship exists among the aging process, organ decay and disfunction, and the occurrence of various diseases including cancer. A genetically engineered mouse model, termed Klf1K74R/K74R or Klf1(K74R), carrying mutation on the well-conserved sumoylation site of the hematopoietic transcription factor KLF1/EKLF has been generated that possesses extended lifespan and healthy characteristics, including cancer resistance. We show that the healthy longevity characteristics of the Klf1(K74R) mice, as exemplified by their higher anti-cancer capability, are likely gender-, age-, and genetic background-independent. Significantly, the anti-cancer capability, in particular that against melanoma as well as hepatocellular carcinoma, and lifespan-extending property of Klf1(K74R) mice, could be transferred to wild-type mice via transplantation of their bone marrow mononuclear cells at a young age of the latter. Furthermore, NK(K74R) cells carry higher in vitro cancer cell-killing ability than wild-type NK cells. Targeted/global gene expression profiling analysis has identified changes in the expression of specific proteins, including the immune checkpoint factors PDCD and CD274, and cellular pathways in the leukocytes of the Klf1(K74R) that are in the directions of anti-cancer and/or anti-aging. This study demonstrates the feasibility of developing a transferable hematopoietic/blood system for long-term anti-cancer and, potentially, for anti-aging.

Technology Behind Cell Therapy Augmentation of Fracture Healing: Concentrated Bone Marrow Aspirate.

With an aging population, and an anticipated increase in overall fracture incidence, a sound understanding of bone healing and how technology can optimize this process is crucial. Concentrated bone marrow aspirate (cBMA) is a technology that capitalizes on skeletal stem and progenitor cells (SSPCs) to enhance the regenerative capacity of bone. This overview highlights the science behind cBMA, discusses the role of SSPCs in bone homeostasis and fracture repair, and briefly details the clinical evidence supporting the use of cBMA in fracture healing. Despite promising early clinical results, a lack of standardization in harvest and processing techniques, coupled with patient variability, presents challenges in optimizing the use of cBMA. However, cBMA remains an emerging technology that may certainly play a crucial role in the future of fracture healing augmentation.

Plasma-Based Scaffold Containing Bone-Marrow Mononuclear Cells Promotes Wound Healing in a Mouse Model of Pressure Injury.

Pressure injuries, or pressure ulcers, are a common problem that may lead to infections and major complications, besides being a social and economic burden due to the costs of treatment and hospitalization. While surgery is sometimes necessary, this also has complications such as recurrence or wound dehiscence. Among the newer methods of pressure injury treatment, advanced therapies are an interesting option. This study examines the healing properties of bone marrow mononuclear cells (BM-MNCs) embedded in a plasma-based scaffold in a mouse model. Pressure ulcers were created on the backs of mice (2 per mouse) using magnets and assigned to a group of ulcers that were left untreated (Control, n = 15), treated with plasma scaffold (Plasma, n = 15), or treated with plasma scaffold containing BM-MNC (Plasma + BM-MNC, n = 15). Each group was examined at three time points (3, 7, and 14 days) after the onset of treatment. At each time point, animals were subjected to biometric assessment, bioluminescence imaging, and tomography. Once treatment had finished, skin biopsies were processed for histological and wound healing reverse transcription polymerase chain reaction (RT-PCR) array studies. While wound closure percentages were higher in the Plasma and Plasma + BM-MNC groups, differences were not significant, and thus descriptive data are provided. In all individuals, the presence of donor cells was revealed by immunohistochemistry on posttreatment onset Days 3, 7, and 14. In the Plasma + BM-MNC group, less inflammation was observed by positron emission tomography-computed tomography (PET/CT) imaging of the mice at 7 days, and a complete morphometabolic response was produced at 14 days, in accordance with histological results. A much more pronounced inflammatory process was observed in controls than in the other two groups, and this persisted until Day 14 after treatment onset. RT-PCR array gene expression patterns were also found to vary significantly, with the greatest difference noted between both treatments at 14 days when 11 genes were differentially expressed.

Percutaneous bone marrow concentrate and platelet products versus exercise therapy for the treatment of rotator cuff tears: a randomized controlled, crossover trial with 2-year follow-up.

BACKGROUND: Surgical repair is recommended for the treatment of high-grade partial and full thickness rotator cuff tears, although evidence shows surgery is not necessarily superior to non-surgical therapy. The purpose of this study was to compare percutaneous orthobiologic treatment to a home exercise therapy program for supraspinatus tears. METHODS: In this randomized-controlled, crossover design, participants with a torn supraspinatus tendon received either 'BMC treatment', consisting of a combination of autologous bone marrow concentrate (BMC) and platelet products, or underwent a home exercise therapy program. After three months, patients randomized to exercise therapy could crossover to receive BMC treatment if not satisfied with shoulder progression. Patient-reported outcomes of Numeric Pain Scale (NPS), Disabilities of the Arm, Shoulder, and Hand, (DASH), and a modified Single Assessment Numeric Evaluation (SANE) were collected at 1, 3, 6, 12, and 24 months. Pre- and post-treatment MRI were assessed using the Snyder Classification system. RESULTS: Fifty-one patients were enrolled and randomized to the BMC treatment group (n = 34) or the exercise therapy group (n = 17). Significantly greater improvement in median ΔDASH, ΔNPS, and SANE scores were reported by the BMC treatment group compared to the exercise therapy group (-11.7 vs -3.8, P = 0.01; -2.0 vs 0.5, P = 0.004; and 50.0 vs 0.0, P < 0.001; respectively) after three months. Patient-reported outcomes continued to progress through the study's two-year follow-up period without a serious adverse event. Of patients with both pre- and post-treatment MRIs, a majority (73%) showed evidence of healing post-BMC treatment. CONCLUSIONS: Patients reported significantly greater changes in function, pain, and overall improvement following BMC treatment compared to exercise therapy for high grade partial and full thickness supraspinatus tears. TRIAL REGISTRATION: This protocol was registered with www. CLINICALTRIALS: gov (NCT01788683; 11/02/2013).

Transplantation of Donor-Recipient Chimeric Cells Restores Peripheral Blood Cell Populations and Increases Survival after Total Body Irradiation-Induced Injury in a Rat Experimental Model.

Current therapies for acute radiation syndrome (ARS) involve bone marrow transplantation (BMT), leading to graft-versus-host disease (GvHD). To address this challenge, we have developed a novel donor-recipient chimeric cell (DRCC) therapy to increase survival and prevent GvHD following total body irradiation (TBI)-induced hematopoietic injury without the need for immunosuppression. In this study, 20 Lewis rats were exposed to 7 Gy TBI to induce ARS, and we assessed the efficacy of various cellular therapies following systemic intraosseous administration. Twenty Lewis rats were randomly divided into four experimental groups (n = 5/group): saline control, allogeneic bone marrow transplantation (alloBMT), DRCC, and alloBMT + DRCC. DRCC were created by polyethylene glycol-mediated fusion of bone marrow cells from 24 ACI (RT1a) and 24 Lewis (RT11) rat donors. Fusion feasibility was confirmed by flow cytometry and confocal microscopy. The impact of different therapies on post-irradiation peripheral blood cell recovery was evaluated through complete blood count, while GvHD signs were monitored clinically and histopathologically. The chimeric state of DRCC was confirmed. Post-alloBMT mortality was 60%, whereas DRCC and alloBMT + DRCC therapies achieved 100% survival. DRCC therapy also led to the highest white blood cell counts and minimal GvHD changes in kidney and skin samples, in contrast to alloBMT treatment. In this study, transplantation of DRCC promoted the recovery of peripheral blood cell populations after TBI without the development of GVHD. This study introduces a novel and promising DRCC-based bridging therapy for treating ARS and extending survival without GvHD.

Autologous concentrated bone marrow injection for precollapse osteonecrosis of the femoral head concurrent with contralateral total hip arthroplasty: protocol for a clinical trial.

INTRODUCTION: The femoral head contralateral to the collapsed femoral head requiring total hip arthroplasty (THA) often manifests in the precollapse stage of osteonecrosis of the femoral head (ONFH). It is not yet demonstrated how autologous concentrated bone marrow injection may prevent collapse of the femoral head concurrent with contralateral THA. The primary objective is to evaluate the efficacy of autologous concentrated bone marrow injection for the contralateral, non-collapsed, femoral head in patients with bilateral ONFH, with the ipsilateral collapsed femoral head undergoing THA. METHODS AND ANALYSIS: This is a multicentre, prospective, non-randomised, historical-data controlled study. We will recruit patients with ONFH who are scheduled for THA and possess a non-collapsed contralateral femoral head. Autologous bone marrow will be collected using a point-of-care device. After concentration, the bone marrow will be injected into the non-collapsed femoral head following the completion of THA in the contralateral hip. The primary outcome is the percentage of femoral head collapse evaluated by an independent data monitoring committee using plain X-rays in two directions 2 years after autologous concentrated bone marrow injection. Postinjection safety, adverse events, pain and hip function will also be assessed. The patients will be evaluated preoperatively, and at 6 months, 1 year and 2 years postoperatively. ETHICS AND DISSEMINATION: This protocol has been approved by the Certified Committee for Regenerative Medicine of Tokyo Medical and Dental University and Japan's Ministry of Healthy, Labour and Welfare and will be performed as a class III regenerative medicine protocol, in accordance with Japan's Act on the Safety of Regenerative Medicine. The results of this study will be submitted to a peer-review journal for publication. The results of this study are expected to provide evidence to support the inclusion of autologous concentrated bone marrow injections in the non-collapsed femoral head in Japan's national insurance coverage. TRIAL REGISTRATION NUMBER: jRCTc032200229.

Comparable outcomes of allogeneic peripheral blood versus bone marrow hematopoietic stem cell transplantation from a sibling donor for pediatric patients.

Traditionally, bone marrow (BM) has been preferred as a source of stem cells (SCs) in pediatric hematopoietic SC transplantation (HSCT); however, the use of peripheral blood SCs (PBSC) has recently increased. With advancing graft-versus-host disease (GVHD) prophylaxis, whether the BM is still a better SC source than PB in sibling donor HSCT remains controversial. Here, we compared the results of BM transplantation (BMT) and PBSC transplantation (PBSCT) in pediatric patients with malignant or non-malignant diseases receiving sibling HSCT using a total of 7.5 mg/kg of anti-thymocyte globulin (ATG). We retrospectively reviewed children who received HSCT from a sibling donor between 2005 and 2020 at Seoul National University Children's Hospital. Of the 86 patients, 40 underwent BMT, and 46 underwent PBSCT. Fifty- six patients had malignant diseases, whereas thirty patients had non-malignant diseases. All conditioning regimens comprised ATG. Busulfan-based myeloablative conditioning regimens were administered to patients with malignant diseases and approximately half of those with non-malignant diseases. The remaining half of the patients with non-malignant diseases were administered cyclophosphamide-based reduced- intensity conditioning regimens. According to studies conducted at our center, all BM donors received G-CSF before harvest to achieve early engraftment. In all 86 patients (47 males and 39 females), the median age at the time of HSCT was 11.4 (range, 0.7 - 24.6) years. The median follow-up period was 57.9 (range, 0.9-228.6) months, and the corresponding values for those with BM and PBSC were 77 (range, 2.4-228.6) months and 48.7 (range, 0.9-213.2) months, respectively. Engraftment failure occurred in one patient with BM and no patient with PBSC. The cumulative incidence of acute GVHD with grades II-IV was higher in PBSC (BM 2.5%, PBSC 26.1%, p = 0.002), but there was no significant difference in those with grades III-IV acute GVHD (BM 0%, PBSC 6.5%, p = 0.3703) and extensive chronic GVHD (BM 2.5%, PBSC 11.6%, p = 0.1004). There were no significant differences in treatment-related mortality (TRM) (BM 14.2%, PBSC 6.8%, p = 0.453), 5-year event-free survival (EFS) (BM 71.5%, PBSC 76.2%, p = 0.874), and overall survival (OS) rates (BM 80.8%, PBSC 80.3%, p = 0.867) between BM and PBSC in the univariate analysis. In the multivariate analysis, which included all factors with p < 0.50 in the univariate analysis, there was no significant prognostic factor for EFS or OS. There was no significant difference in the relapse incidence between BM and PBSC among patients with malignant diseases (BM 14.2%, PBSC 6.8%, p = 0.453). Additionally, there were no significant differences in the TRM, 5-year EFS, and OS rates between malignant and non-malignant diseases nor between the busulfan-based myeloablative regimen and reduced-intensity chemotherapy using cyclophosphamide. In this study, we showed no significant differences in EFS, OS, TRM, and GVHD, except for acute GVHD grades II-IV, between BMT and PBSCT from sibling donors, using ATG (a total of 7.5 mg/kg). Therefore, PB collection, which is less invasive for donors and less labor-intensive for doctors, could also be considered an acceptable SC source for sibling donor HSCT in children.

Modulation of lipid profile by secretory phospholipase A2 group IIA: Verification with a transgenic mouse model.

We previously demonstrated a positive relation of secretory phospholipase A2 group IIA (sPLA2-IIA) with circulating high-density lipoprotein cholesterol (HDL-C) in patients with coronary artery disease, and sPLA2-IIA increased cholesterol efflux in THP-1 cells through peroxisome proliferator-activated receptor-γ (PPAR-γ)/liver X receptor α/ATP-binding cassette transporter A1 (ABCA1) signaling pathway. The aim of the present study was to examine the role of sPLA2-IIA over-expression on lipid profile in a transgenic mouse model. Fifteen apoE-/- and C57BL/7 female mice received bone marrow transplantation from transgenic SPLA2-IIA mice, and treated with specific PPAR-γ inhibitor GW9662. High fat diet was given after one week of bone marrow transplantation, and animals were sacrificed after twelve weeks. Immunohistochemical staining showed over-expression of sPLA2-IIA protein in the lung and spleen. The circulating level of HDL-C, but not that of low-density lipoprotein cholesterol (LDL-C), total cholesterol, or total triglyceride, was increased by sPLA2-IIA over-expression, and was subsequently reversed by GW9662 treatment. Over-expression of sPLA2-IIA resulted in augmented expression of cholesterol transporter ABCA1 at mRNA level in the aortas, and at protein level in macrophages, co-localized with macrophage specific antigen CD68. GW9662 exerted potent inhibitory effects on sPLA2-IIA-induced ABCA1 expression. Conclusively, we demonstrated the effects of sPLA2-IIA on circulating HDL-C level and the expression of ABCA1, possibly through regulation of PPAR-γ signaling in transgenic mouse model, that is in concert with the conditions in patients with coronary artery disease.

Systematic review and meta-analysis of the effect of bone marrow-derived cell therapies on hind limb perfusion.

Preclinical and clinical studies on the administration of bone marrow-derived cells to restore perfusion show conflicting results. We conducted a systematic review and meta-analysis on preclinical studies to assess the efficacy of bone marrow-derived cells in the hind limb ischemia model and identify possible determinants of therapeutic efficacy. In vivo animal studies were identified using a systematic search in PubMed and EMBASE on 10 January 2022. 85 studies were included for systematic review and meta-analysis. Study characteristics and outcome data on relative perfusion were extracted. The pooled mean difference was estimated using a random effects model. Risk of bias was assessed for all included studies. We found a significant increase in perfusion in the affected limb after administration of bone marrow-derived cells compared to that in the control groups. However, there was a high heterogeneity between studies, which could not be explained. There was a high degree of incomplete reporting across studies. We therefore conclude that the current quality of preclinical research is insufficient (low certainty level as per GRADE assessment) to identify specific factors that might improve human clinical trials.

Donor regulatory T cells rapidly adapt to recipient tissues to control murine acute graft-versus-host disease.

The adoptive transfer of regulatory T cells is a promising strategy to prevent graft-versus-host disease after allogeneic bone marrow transplantation. Here, we use a major histocompatibility complex-mismatched mouse model to follow the fate of in vitro expanded donor regulatory T cells upon migration to target organs. Employing comprehensive gene expression and repertoire profiling, we show that they retain their suppressive function and plasticity after transfer. Upon entering non-lymphoid tissues, donor regulatory T cells acquire organ-specific gene expression profiles resembling tissue-resident cells and activate hallmark suppressive and cytotoxic pathways, most evidently in the colon, when co-transplanted with graft-versus-host disease-inducing conventional T cells. Dominant T cell receptor clonotypes overlap between organs and across recipients and their relative abundance correlates with protection efficacy. Thus, this study reveals donor regulatory T cell selection and adaptation mechanisms in target organs and highlights protective features of Treg to guide the development of improved graft-versus-host disease prevention strategies.

Effects of CD34+ cell dose on engraftment and long-term outcomes after allogeneic bone marrow transplantation.

BACKGROUND: The number of CD34+ cells in the graft is generally associated with time to engraftment and survival in transplantation using cord blood or allogeneic peripheral blood stem cells. However, the significance of abundant CD34+ in bone marrow transplantation (BMT) remained unclear. METHODS: We retrospectively reviewed 207 consecutive adult patients who underwent their first BMT at Jichi Medical University between January 2009 and June 2021. RESULTS: The median nucleated cell count (NCC) and CD34+ cell dose were 2.17 × 108/kg (range .56-8.52) and 1.75 × 106/kg (.21-5.84), respectively. Compared with 104 patients in the low CD34+ group (below the median), 103 patients in the high CD34+ group (above the median) showed faster engraftment at day +28 in terms of neutrophil (84.6% vs. 94.2%; p =  .001), reticulocyte (51.5% vs. 79.6%; p < .001), and platelet (39.4% vs. 72.8%; p < .001). There were no significant differences in overall survival, relapse, nonrelapse mortality, acute or chronic graft-versus-host disease, or infectious complications between the two groups in univariate and multivariate analyses. Low or high NCC had no significant effect on overall survival, nonrelapse mortality, cumulative incidence of relapse and graft-versus-host disease, either. While a positive correlation was observed between NCC and the CD34+ cell dose, a high CD34+ cell dose was associated with rapid hematopoietic recovery, even in patients with NCC below the median. CONCLUSION: Measurement of CD34+ cell dose in addition to NCC was useful for predicting hematopoietic recovery, but seemed to have little influence on the long-term outcome in BMT.

COVID-19 surveillance in a bone marrow transplantation unit: experience from a Brazilian tertiary-care teaching hospital.

PURPOSE: In this work, we aimed to describe the strategy of the weekly SARS-CoV-2 RT-PCR surveillance program that was implemented in our bone marrow transplantation (BMT) unit. METHODS: Our unit performed SARS-CoV-2 RT-PCR before admission and then weekly during hospitalization even if the patient was asymptomatic. From May 2021 to May 2022, we collected data from all patients that were admitted in the BMT unit to perform transplantation. The total of SARS-CoV-2 RT-PCR performed and the positive rate were described. RESULTS: During the study period, 65 patients were admitted for HSCT. A total of 414 SARS-CoV-2 RT-PCR were performed. Two cases were detected (positivity rate, 0.48%). After the positive test, both patients were isolated outside the BMT unit. CONCLUSION: We postulate that diagnosing these patients and isolating them outside the transplantation unit may have prevented secondary symptomatic cases.

RIO-kinase 2 is essential for hematopoiesis.

Regulation of protein synthesis is a key factor in hematopoietic stem cell maintenance and differentiation. Rio-kinase 2 (RIOK2) is a ribosome biogenesis factor that has recently been described an important regulator of human blood cell development. Additionally, we have previously identified RIOK2 as a regulator of protein synthesis and a potential target for the treatment of acute myeloid leukemia (AML). However, its functional relevance in several organ systems, including normal hematopoiesis, is not well understood. Here, we investigate the consequences of RIOK2 loss on normal hematopoiesis using two different conditional knockout mouse models. Using competitive and non-competitive bone marrow transplantations, we demonstrate that RIOK2 is essential for the differentiation of hematopoietic stem and progenitor cells (HSPCs) as well as for the maintenance of fully differentiated blood cells in vivo as well as in vitro. Loss of RIOK2 leads to rapid death in full-body knockout mice as well as mice with RIOK2 loss specific to the hematopoietic system. Taken together, our results indicate that regulation of protein synthesis and ribosome biogenesis by RIOK2 is essential for the function of the hematopoietic system.

IL-6-mediated endothelial injury impairs antiviral humoral immunity after bone marrow transplantation.

Endothelial function and integrity are compromised after allogeneic bone marrow transplantation (BMT), but how this affects immune responses broadly remains unknown. Using a preclinical model of CMV reactivation after BMT, we found compromised antiviral humoral responses induced by IL-6 signaling. IL-6 signaling in T cells maintained Th1 cells, resulting in sustained IFN-γ secretion, which promoted endothelial cell (EC) injury, loss of the neonatal Fc receptor (FcRn) responsible for IgG recycling, and rapid IgG loss. T cell-specific deletion of IL-6R led to persistence of recipient-derived, CMV-specific IgG and inhibited CMV reactivation. Deletion of IFN-γ in donor T cells also eliminated EC injury and FcRn loss. In a phase III clinical trial, blockade of IL-6R with tocilizumab promoted CMV-specific IgG persistence and significantly attenuated early HCMV reactivation. In sum, IL-6 invoked IFN-γ-dependent EC injury and consequent IgG loss, leading to CMV reactivation. Hence, cytokine inhibition represents a logical strategy to prevent endothelial injury, thereby preserving humoral immunity after immunotherapy.

Acute Graft versus Host Disease in Beta Thalassemia Patients Following Allogeneic Haematopoietic Stem Cell Transplantation.

OBJECTIVE: To analyse the frequency, risk factors, and clinical symptoms of acute graft-versus-host disease (aGvHD) in patients with beta-thalassemia major after allogeneic haematopoietic stem cell transplantation (HSCT). STUDY DESIGN: Descriptive study. Place and Duration of the Study: Department of Clinical Haematology, Armed Forces Bone Marrow Transplant Centre, Rawalpindi, Pakistan, from January 2017 to December 2021. METHODOLOGY: Data were obtained from patients diagnosed with bone and tissue malignancies (BTM) who had undergone haematopoietic stem cell transplantation (HSCT) and experienced aGVHD. Patients who experienced initial graft failure and individuals who underwent subsequent bone marrow transplantation were excluded. RESULTS: Total of 117 patients diagnosed with BTM underwent fully matched HSCT, including 76 (65%) males, and 41 (35%) females. The median age of the patients undergoing transplantation was 7.34±7.32 years and the donors' median age was 7.6±9.85 years. Among the donors, 53 (45.3%) were males and 64 (54.7%) were females. Gender disparity was observed in 46 (39.3%) instances as a female donor matched with a male recipient. A total of 106 individuals underwent bone marrow harvest (BMH); with 5 (4.3%) patients receiving peripheral blood stem cells (PBSC) and 6 (5.2%) patients receiving both BMH and PBSC. Acute GvHD was observed in 50 (42.7%) patients, including 30 (60%) males and 20 (40%) females. Grade I GvHD occurred in 32 (27.3%) individuals, Grade II GvHD in 16 (13.7%) patients, and Grade III GvHD in one (0.8%) patient. It had no statistically significant association with recipient/donor age, gender disparity, the source of the graft source, the dose of stem cells, or the presence of thymoglobulin (TG). CONCLUSION: Acute GvHD was observed in high frequency in Beta-thalassemia patients receiving morrow harvesting proportional to their gender distribution. Associated factors were GvHD prophylaxis measure, mucositis and, CMV reactivation. KEY WORDS: Beta thalassemia major patients, Acute graft versus host disease, Allogeneic haematopoietic stem cell.

[Establishment and Evaluation Strategy of an in vitro Cell Model of Bone Marrow Microenvironment Injury in Mouse Acute Graft-Versus-Host Disease].

OBJECTIVE: To establish a mesenchymal stem cell（MSC）-based in vitro cell model for the evaluation of mouse bone marrow acute graft-versus-host disease (aGVHD). METHODS: Female C57BL/6N mice aged 6-8 weeks were used as bone marrow and lymphocyte donors, and female BALB/c mice aged 6-8 weeks were used as aGVHD recipients. The recipient mouse received a lethal dose (8.0 Gy,72.76 cGy/min) of total body γ irradiation, and injected with donor mouse derived bone marrow cells (1×107/mouse) in 6-8 hours post irradiation to establish a bone marrow transplantation (BMT) mouse model (n=20). In addition, the recipient mice received a lethal dose (8.0 Gy,72.76 cGy/min) of total body γ irradiation, and injected with donor mouse derived bone marrow cells (1×107/mouse) and spleen lymphocytes (2×106/mouse) in 6-8 hours post irradiation to establish a mouse aGVHD model (n=20). On the day 7 after modeling, the recipient mice were anesthetized and the blood was harvested post eyeball enucleation. The serum was collected by centrifugation. Mouse MSCs were isolated and cultured with the addition of 2%, 5%, and 10% recipient serum from BMT group or aGVHD group respectively. The colony-forming unit-fibroblast（CFU-F) experiment was performed to evaluate the potential effects of serums on the self-renewal ability of MSC. The expression of CD29 and CD105 of MSC was evaluated by immunofluorescence staining. In addition, the expression of self-renewal-related genes including Oct-4, Sox-2, and Nanog in MSC was detected by real-time fluorescence quantitative PCR（RT-qPCR). RESULTS: We successfully established an in vitro cell model that could mimic the bone marrow microenvironment damage of the mouse with aGVHD. CFU-F assay showed that, on day 7 after the culture, compared with the BMT group, MSC colony formation ability of aGVHD serum concentrations groups of 2% and 5% was significantly reduced （P < 0．05）; after the culture, at day 14, compared with the BMT group, MSC colony formation ability in different aGVHD serum concentration was significantly reduced （P < 0．05）. The immunofluorescence staining showed that, compared with the BMT group, the proportion of MSC surface molecules CD29+ and CD105+ cells was significantly dereased in the aGVHD serum concentration group (P < 0.05), the most significant difference was at a serum concentration of 10% （P < 0．001, P < 0.01）. The results of RT-qPCR detection showed that the expression of the MSC self-renewal-related genes Oct-4, Sox-2, and Nanog was decreased, the most significant difference was observed at an aGVHD serum concentration of 10% （P < 0.01,P < 0．001,P < 0．001）. CONCLUSION: By co-culturing different concentrations of mouse aGVHD serum and mouse MSC, we found that the addition of mouse aGVHD serum at different concentrations impaired the MSC self-renewal ability, which providing a new tool for the field of aGVHD bone marrow microenvironment damage.

Tratamiento de la pseudoartrosis de radio con injerto de cresta iliaca y células madre del aspirado de médula ósea. A propósito de un caso / Treatment of radial pseudarthrosis with iliac crest graft and bone marrow aspirate stem cells. A case report

La pseudoartrosis es una complicación caracterizada por la ausencia de consolidación del hueso a los 9 meses desde el inicio de la fractura, con falta de progresión radiológica los últimos 3 meses, siendo sus principales causas el exceso de movimiento en el foco de fractura y una insuficiente vascularización. A pesar de no tratarse de una complicación frecuente, los huesos del antebrazo ocupan el 4.º puesto en incidencia de presentación. El manejo anestésico de la patología quirúrgica del miembro superior se realiza generalmente en régimen ambulatorio con técnicas de anestesia regional guiadas por ecografía. Estas técnicas tienen una doble función: anestesia durante el propio acto quirúrgico con una mínima variabilidad sobre el estado basal del paciente y analgesia en el postoperatorio inmediato, permitiendo de esta manera el alta a domicilio de forma más segura y precoz. Presentamos el caso de un varón de 34 años, con desarrollo de pseudoartrosis atrófica tras fractura diafisaria de radio, en el que se realiza injerto óseo de cresta ilíaca y aspirado de células madre como estímulo de la osteogénesis.(AU)

Pseudarthrosis is a complication characterised by the absence of bone healing 9 months after the onset of the fracture, with a lack of radiological progressionin the last 3 months, and its main causes are excessive movement at the fracture site and insufficient vascularisation. Despite not being a frequent complica-tion, the bones of the forearm occupy the fourth place in incidence of presentation. The anaesthetic management of surgical pathology of the upper limb isgenerally performed on an outpatient basis with regional anaesthesia techniques guided by ultrasound. These techniques have a dual function: anaesthesiaduring the surgical act with minimal variability over the patient’s baseline condition and analgesia in the immediate postoperative period, thus allowing forsafer and earlier discharge home. We present the case of a 34-year-old male with diagnosis of atrophic pseudarthrosis following a diaphyseal fracture of theradius, in whom iliac crest bone grafting and stem cell aspiration were performed to stimulate osteogenesis.(AU)