Bone Marrow Mononuclear Cells Group Research Articles

#1948 Transcriptome analysis of the effect of roxadustat on osteoclasts

Abstract Background and Aims In recent years, hypoxia-inducing factor prolyl hydroxylase inhibitors (HIF-PHIs) have been widely used in the treatment of renal anemia, bringing a revolutionary breakthrough in the treatment of anemia in CKD patients. However, HIF-PHIs acts on a variety of cells and regulates the expression of multiple target genes. Some studies have reported that HIF-PHIs plays an important role in regulating bone metabolism and promoting the differentiation and calcification of osteoblasts, but its influence on osteoclasts remains controversial. This study intends to explore the effect of roxadustat, a representative drug of HIF-PHIs, on osteoclasts. Method Bone marrow mononuclear cells (BMMCs) from 6-8 weeks old male C57BL/6 wild mice were extracted and induced to differentiate into osteoclasts using macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). Meanwhile, lipopolysaccharide (LPS) was used to simulate the inflammatory environment in CKD patients. According to the experimental group, different concentrations of roxadustat (0 μM, 2.5 μM, 5 μM, 10μM) were added simultaneously. When osteoclast induction reached day 6, mRNA was extracted for transcriptome sequencing and bioinformatics analysis. Results As shown in Table 1, different concentrations of roxadustat were given to primary bone marrow monocytes under both conditions, and there were 3 biological replicates for each sample. The reproducibility scatter plot showed that the data in each group had good repeatability (Fig. 1), and the heat map for basic difference analysis of each sample was shown in Fig. 2. The trend analysis results of the two groups were shown in Fig. 3. BMMCs group and LPS stimulated group showed two upward trends, respectively. KEGG enrichment analysis of these two upward trends showed that HIF-1, MAPK, PI3K-Akt, NF-κB and osteoclast differentiation pathways were significantly enriched in both groups. It is suggested that roxadustat can promote osteoclast differentiation and increase inflammatory response with the increase of roxadustat concentration. The enrichment analysis of the two groups showed that there was no significant enrichment pathway. Conclusion We explored the differentiation of osteoclasts under normal conditions and osteoclast differentiation under simulated inflammation in CKD patients respectively. Transcriptomic analysis results showed that HIF-1, MAPK, PI3K-Akt, NF-κB and osteoclast differentiation pathways were significantly enriched with the increase of roxadustat concentration. It is suggested that the use of roxadustat may promote bone destruction and increase the risk of inflammatory response.

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.

Autologous bone marrow mononuclear cells to treat severe traumatic brain injury in children.

Autologous bone marrow mononuclear cells (BMMNCs) infused after severe traumatic brain injury have shown promise for treating the injury. We evaluated their impact in children, particularly their hypothesized ability to preserve the blood-brain barrier and diminish neuroinflammation, leading to structural CNS preservation with improved outcomes. We performed a randomized, double-blind, placebo-sham-controlled Bayesian dose-escalation clinical trial at two children's hospitals in Houston, TX and Phoenix, AZ, USA (NCT01851083). Patients 5-17 years of age with severe traumatic brain injury (Glasgow Coma Scale score ≤ 8) were randomized to BMMNC or placebo (3:2). Bone marrow harvest, cell isolation and infusion were completed by 48 h post-injury. A Bayesian continuous reassessment method was used with cohorts of size 3 in the BMMNC group to choose the safest between two doses. Primary end points were quantitative brain volumes using MRI and microstructural integrity of the corpus callosum (diffusivity and oedema measurements) at 6 months and 12 months. Long-term functional outcomes and ventilator days, intracranial pressure monitoring days, intensive care unit days and therapeutic intensity measures were compared between groups. Forty-seven patients were randomized, with 37 completing 1-year follow-up (23 BMMNC, 14 placebo). BMMNC treatment was associated with an almost 3-day (23%) reduction in ventilator days, 1-day (16%) reduction in intracranial pressure monitoring days and 3-day (14%) reduction in intensive care unit (ICU) days. White matter volume at 1 year in the BMMNC group was significantly preserved compared to placebo [decrease of 19 891 versus 40 491, respectively; mean difference of -20 600, 95% confidence interval (CI): -35 868 to -5332; P = 0.01], and the number of corpus callosum streamlines was reduced more in placebo than BMMNC, supporting evidence of preserved corpus callosum connectivity in the treated groups (-431 streamlines placebo versus -37 streamlines BMMNC; mean difference of -394, 95% CI: -803 to 15; P = 0.055), but this did not reach statistical significance due to high variability. We conclude that autologous BMMNC infusion in children within 48 h after severe traumatic brain injury is safe and feasible. Our data show that BMMNC infusion led to: (i) shorter intensive care duration and decreased ICU intensity; (ii) white matter structural preservation; and (iii) enhanced corpus callosum connectivity and improved microstructural metrics.

Intra-articular bone marrow mononuclear cell therapy improves lameness from naturally occurring equine osteoarthritis.

Osteoarthritis (OA) can be debilitating and is related to impaired resolution of synovial inflammation. Current treatments offer temporary relief of clinical signs, but have potentially deleterious side effects. Bone marrow mononuclear cells (BMNC) are a rich source of macrophage progenitors that have the ability to reduce OA symptoms in people and inflammation in experimentally-induced synovitis in horses. The objective of this study was to evaluate the ability of intra-articular BMNC therapy to improve clinical signs of naturally occurring equine OA. Horses presenting with clinical and radiographic evidence of moderate OA in a single joint were randomly assigned to 1 of 3 treatment groups: saline (negative control), triamcinolone (positive control), or BMNC (treatment group). Lameness was evaluated subjectively and objectively, joint circumference measured, and synovial fluid collected for cytology and growth factor/cytokine quantification at 0, 7, and 21 days post-injection. Data were analyzed using General Estimating Equations with significance set at p < 0.05. There were no adverse effects noted in any treatment group. There was a significant increase in synovial fluid total nucleated cell count in the BMNC-treated group on day 7 (median 440; range 20-1920 cells/uL) compared to day 0. Mononuclear cells were the predominant cell type across treatments at all time points. Joint circumference decreased significantly in the BMNC-treated group from days 7 to 21 and was significantly lower at day 21 in the BMNC-treated group compared to the saline-treated group. Median objective lameness improved significantly in the BMNC group between days 7 and 21. GM-CSF, IL-1ra, IGF-1, and TNF-α were below detectable limits and IL-6, IL-1β, FGF-2 were detectable in a limited number of synovial fluid samples. Inconsistent and limited differences were detected over time and between treatment groups for synovial fluid PGE2, SDF-1, MCP-1 and IL-10. Decreased lameness and joint circumference, coupled with a lack of adverse effects following BMNC treatment, support a larger clinical trial using BMNC therapy to treat OA in horses.

Safety and efficacy of intra-arterial bone marrow mononuclear cell transplantation in patients with acute ischaemic stroke in Spain (IBIS trial): a phase 2, randomised, open-label, standard-of-care controlled, multicentre trial

Pilot clinical trials have shown the safety of intra-arterial bone marrow mononuclear cells (BMMNCs) in stroke. However, the efficacy of different doses of intra-arterial BMMNCs in patients with acute stroke has not been tested in a randomised clinical trial. We aimed to show safety and efficacy of two different doses of autologous intra-arterial BMMNC transplantation in patients with acute stroke. The IBIS trial was a multicentre phase 2, randomised, controlled, investigator-initiated, assessor-blinded, clinical trial, in four stroke centres in Spain. We included patients (aged 18-80 years) with a non-lacunar, middle cerebral artery ischaemic stroke within 1-7 days from stroke onset and with a National Institutes of Health Stroke Scale score of 6-20. We randomly assigned patients (2:1:1) with a computer-generated randomisation sequence to standard of care (control group) or intra-arterial injection of autologous BMMNCs at one of two different doses (2 × 106 BMMNCs/kg or 5 × 106 BMMNCs/kg). The primary efficacy outcome was the proportion of patients with modified Rankin Scale scores of 0-2 at 180 days in the intention-to-treat population, comparing each BMMNC dose group and the pooled BMMNC group versus the control group. The primary safety endpoint was the proportion of serious adverse events. This trial was registered at ClinicalTrials.gov, NCT02178657 and is completed. Between April 1, 2015, and May 20, 2021, we assessed 114 patients for eligibility. We randomly assigned 77 (68%) patients: 38 (49%) to the control group, 20 (26%) to the low-dose BMMNC group, and 19 (25%) the high-dose BMMNC group. The mean age of participants was 62·4 years (SD 12·7), 46 (60%) were men, 31 (40%) were women, all were White, and 63 (82%) received thrombectomy. The median NIHSS score before randomisation was 12 (IQR 9-15), with intra-arterial BMMNC injection done a median of 6 days (4-7) after stroke onset. The primary efficacy outcome occurred in 14 (39%) patients in the control group versus ten (50%) in the low-dose group (adjusted odds ratio 2·08 [95% CI 0·55-7·85]; p=0·28), eight (44%) in the high-dose group (1·89 [0·52-6·96]; p=0·33), and 18 (47%) in the pooled BMMNC group (2·22 [0·72-6·85]; p=0·16). We found no differences in the proportion of patients who had adverse events or dose-related events, but two patients had a groin haematoma after cell injection in the low-dose BMMNC group. Intra-arterial BMMNCs were safe in patients with acute ischaemic stroke, but we found no significant improvement at 180 days on the mRS. Further clinical trials are warranted to investigate whether improvements might be possible at different timepoints. Instituto de Salud Carlos III co-funded by the European Regional Development Fund/European Social Fund, Mutua Madrileña, and the Regional Ministry of Health of Andalusia.

Biological, Functional and Genetic Characterization of Bone Marrow Mesenchymal Stromal Cells from Adult Ph+ Acute Lymphoblastic Leukemia

Objectives: To clarify the biological characteristics of bone marrow mesenchymal stromal cells (MSCs) in Ph+ acute lymphoblastic leukemia (Ph+ ALL) patients in different disease states, and compare protective effects of MSCs in different disease states on leukemia cells and explore the possible mechanisms. Methods: Bone marrow specimens from Ph+ ALL patients in various disease status (newly diagnosis, complete remission, relapse/ refractory) and healthy donors were collected. Bone marrow mononuclear cells (BMNCs) were isolated and cultured for primary MSCs. (1) CCK-8 method was used to compare proliferation ability of MSCs in vitro. (2) IC50 concentrations of dasatinib (Dasa), imatinib (Ima) and dexamethasone (Dex) in SUP-B15 were determined. SUP-B15 were co-cultured with HS-5 (a human stromal cell line) and MSCs respectively, then the apoptosis ratio of SUP-B15 cells after drug treatment was detected. (3) Transcriptome sequence (RNA-seq) of MSCs was performed, and differentially-expressed genes (DEGs) and signal pathway enrichment analysis among MSCs and BMNCs groups were performed. (5) qRT-PCR was used to detect the expression levels of candidate genes in MSCs. Results: (1) The proliferation ability of DR-MSCs in vitro was stronger than ALL-MSCs (P<0.01, Figure 1A). Proliferation ability of MSCs from refractory and relapsed group (RR-MSCs) were stronger than that from complete remission group (CR-MSCs) and newly diagnosed group (ND-MSCs)(P<0.05, P<0.05, Figure 1A). There was no significant difference between newly diagnosed-complete remission (ND-CR) paired MSCs (P>0.05). (2) The ratios of apoptotic SUP-B15 cells were significant decreased when co-cultured with HS-5 with and without the presence of Dasa, Ima, and Dex (P＜0.05, P＜0.0001, P＜0.001, P＜0.01, Figure 1B). (3) Ratios of apoptotic SUP-B15 cells were significantly reduced when co-cultured with different MSCs under the treatment of blank control, Dasa, Ima, and Dex. The abilities to prevent SUP-B15 cell apoptosis of DR-MSCs and ND-MSCs was stronger than that of CR-MSCs (P<0.001, P<0.05, P<0.01, P<0.0001, P<0.05, P<0.05, P<0.05, P<0.001) (Figure 1B). (4) The RNA-seq results showed that the gene expression thermogram among DR-MSCs, ND-MCSs and CR-MCSs were quite different (Figure 1C-D). Expression of BMP4 in ND-MSCs was significantly up-regulated (FC=3.311, P. adj=0.024) comparing with DR-MSCs. RR-MSCs expressed higher level of BMP4 than CR-MSCs (P＜0.05). The expression level of BMP6 in CR-MSCs was lower than that in ND-MSCs (P<0.05). DEGs were mainly enriched in BMP/SMADs signaling pathway by GO enrichment analysis (P. adj＜0.0001). (5) Comparing with DR-MSCs, DEGs in ND-MSCs were enriched in cytokine receptors and immune gene-related pathways by Reactome enrichment analysis (P. adj=0.023). (6) TIDE scores of CR-MSCs and RR-MSCs were higher than those of ND-MSCs (P=0.045 and P =0.06, respectively). Conclusion: Compared with DR-MSCs, Ph+ ALL-MSCs had a weaker proliferative capacity in vitro. MSCs derived from different disease status have protective effects on the apoptosis of SUP-B15 induced by drugs, the effects of DR-MSCs and ND-MSCs were stronger than that of CR-MSCs. The protective effect may be achieved through the activity of BMP/SMADs signaling pathway. Figure Legend: A. In Vitro proliferation of MSCs;B.The effect of MSCs cells on apoptosis of SUP-B15;C.Differential gene expression thermogram of DR-MSCs and ND-MCSs;D.Differential gene expression thermogram of ND-CR compared samples. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Local intramuscular transplantation of autologous bone marrow mononuclear cells for critical lower limb ischaemia.

Peripheral arterial disease is a major health problem, and in about 1% to 2% of patients, the disease progresses to critical limb ischaemia (CLI), also known as critical limb-threatening ischaemia. In a substantial number of individuals with CLI, no effective treatment options other than amputation are available, with around a quarter of these patients requiring a major amputation during the following year. This is the second update of a review first published in 2011. To evaluate the benefits and harms of local intramuscular transplantation of autologous adult bone marrow mononuclear cells (BMMNCs) as a treatment for CLI. We used standard, extensive Cochrane search methods. The latest search date was 8 November 2021. We included all randomised controlled trials (RCTs) of CLI in which participants were randomly allocated to intramuscular administration of autologous adult BMMNCs or control (either no intervention, conventional conservative therapy, or placebo). We used standard Cochrane methods. Our primary outcomes of interest were all-cause mortality, pain, and amputation. Our secondary outcomes were angiographic analysis, ankle-brachial index (ABI), pain-free walking distance, side effects and complications. We assessed the certainty of the evidence using the GRADE approach. We included four RCTs involving a total of 176 participants with a clinical diagnosis of CLI. Participants were randomised to receive either intramuscular cell implantation of BMMNCs or control. The control arms varied between studies, and included conventional therapy, diluted autologous peripheral blood, and saline. There was no clear evidence of an effect on mortality related to the administration of BMMNCs compared to control (risk ratio (RR) 1.00, 95% confidence interval (CI) 0.15 to 6.63; 3 studies, 123 participants; very low-certainty evidence). All trials assessed changes in pain severity, but the trials used different forms of pain assessment tools, so we were unable to pool data. Three studies individually reported that no differences in pain reduction were observed between the BMMNC and control groups. One study reported that reduction in rest pain was greater in the BMMNC group compared to the control group (very low-certainty evidence). All four trials reported the rate of amputation at the end of the study period. We are uncertain if amputations were reduced in the BMMNC group compared to the control group, as a possible small effect (RR 0.52, 95% CI 0.27 to 0.99; 4 studies, 176 participants; very low-certainty evidence) was lost after undertaking sensitivity analysis (RR 0.52, 95% CI 0.19 to 1.39; 2 studies, 89 participants). None of the included studies reported any angiographic analysis. Ankle-brachial index was reported differently by each study, so we were not able to pool the data. Three studies reported no changes between groups, and one study reported greater improvement in ABI (as haemodynamic improvement) in the BMMNC group compared to the control group (very low-certainty evidence). One study reported pain-free walking distance, finding no clear difference between BMMNC and control groups (low-certainty evidence). We pooled the data for side effects reported during the follow-up, and this did not show any clear difference between BMMNC and control groups (RR 2.13, 95% CI 0.50 to 8.97; 4 studies, 176 participants; very low-certainty evidence). We downgraded the certainty of the evidence due to the concerns about risk of bias, imprecision, and inconsistency. We identified a small number of studies that met our inclusion criteria, and these differed in the controls they used and how they measured important outcomes. Limited data from these trials provide very low- to low-certainty evidence, and we are unable to draw conclusions to support the use of local intramuscular transplantation of BMMNC for improving clinical outcomes in people with CLI. Evidence from larger RCTs is needed in order to provide adequate statistical power to assess the role of this procedure.

Regeneration potential of bone marrow derived mesenchymal stem cells and platelet rich plasma (PRP) on irradiation-induced damage of submandibular salivary gland in albino rats

Radiation-induced damage to salivary glands (SG) is a consequence of radiotherapy for head and neck cancers. Recovery of the irradiated SG has been studied using various regenerative approaches. This study aims to compare the regenerative potentials of platelet-rich plasma (PRP) and bone marrow mononuclear cells (BMMCs) on irradiated rat submandibular salivary glands (SMD). 32 healthy male albino rats were irradiated with a single dose of 6 Gy then classified into four groups. Group A received no treatment while the other 3 groups were injected 24 h post-radiation with a single dose of either; BMMCs (Group B), PRP (Group C), or BMMCs suspended in PRP (Group D). SMD regeneration was assessed in terms of histological changes and TGF- β1 gene expression. The results showed that compared to the untreated group all groups showed successful regeneration with group D showing the best results. A statistically significant increase in the surface area of acini and TGF- β1 gene expression was observed in group D, followed by group C, then B. Our results prove that using PRP and BMMCs could be promising in decreasing irradiation side effects on SG. Moreover, combining PRP and BMMCs gives better effects compared to each therapy alone.

Effect of Freshly Isolated Bone Marrow Mononuclear Cells and Cultured Bone Marrow Stromal Cells in Graft Cell Repopulation and Tendon-Bone Healing after Allograft Anterior Cruciate Ligament Reconstruction.

Graft cell repopulation and tendon-bone tunnel healing are important after allograft anterior cruciate ligament reconstruction (ACLR). Freshly isolated bone marrow mononuclear cells (BMMNCs) have the advantage of short isolation time during surgery and may enhance tissue regeneration. Thus, we hypothesized that the effect of intra-articular BMMNCs in post-allograft ACLR treatment is comparable to that of cultured bone marrow stromal cells (BMSCs). A rabbit model of hamstring allograft ACLR was used in this study. Animals were randomly assigned to the BMMNC, BMSC, and control groups. Fresh BMMNCs isolated from the iliac crest during surgery and cultured BMSCs at passage four were used in this study. A total of 1 × 107 BMMNCs or BMSCs in 100 µL phosphate-buffered saline were injected into the knee joint immediately after ACLR. The control group was not injected with cells. At two and six weeks post operation, we assessed graft cell repopulation with histological and cell tracking staining (PKH26), and tendon-bone healing with histological micro-computed tomography and immunohistochemical analyses for collagen I and monocyte chemoattractant protein-1 (MCP1). At two weeks post operation, there was no significant difference in the total cell population within the allograft among the three groups. However, the control group showed significantly higher cell population within the allograft than that of BM cell groups at six weeks. Histological examination of proximal tibia revealed that the intra-articular delivered cells infiltrated into the tendon-bone interface. Compared to the control group, the BM cell groups showed broader gaps with interfacial fibrocartilage healing, similar collagen I level, and higher MCP1 expression in the early stage. Micro-CT did not reveal any significant difference among the three groups. BMMNCs and BMSCs had comparable effects on cell repopulation and interfacial allograft-bone healing. Intra-articular BM cells delivery had limited benefits on graft cell repopulation and caused higher inflammation than that in the control group in the early stage, with fibrocartilage formation in the tendon-bone interface after allograft ACLR.

Neuroprotective Effects of Bone Marrow Mononuclear Cells Therapy in a Rat Model of Ischemia Stroke

Objective: Bone marrow mononuclear cells (BMMCs) are considered a potential approach to promote the recovery of stroke-induced neurological deficit. However, the exact mechanism of BMMCs in nerve function recovery is still unclear. Methods: Adult Sprague-Dawley (SD) rat models of cerebral ischemia-reperfusion injury was established by using thread method. BMMCs were transplanted into rat models. Neurological deficits were evaluated by Longa score scale. Immunohistochemistry assay were employed to examine the expression of GFAP and Nogo-A around the ischemic foci in the right frontal lobe. Caspase-3 activity was examined by Western Blot. Results: Rats in BMMCs group had lessened neurological deficits and cleaved Caspase-3 expression on day 21 after reper-fusion, as well as higher expression of GFAP [(37.62±2.45) vs. (27.62±1.69) and (38.00±1.85) vs. (27.25±1.83), P &lt; 0.05] and lower expression of Nogo-A [(28.88±2.64) vs. (32.50±1.60) and (23.87±2.36) vs. (32.00±1.85), P &lt; 0.05] on day 14 and 21 after reperfusion. Meanwhile, the expression of Nogo-A on day 21 was lower than that on day 14 after reperfusion [(23.87±2.36) vs. (28.88±2.64), P &lt; 0.05] in BMMCs group. Conclusion: These findings suggested that BMMCs treatment could improve the functional recovery of neurological deficits in rats with MCAO, which was probably related to enhanced expression of GFAP and reduced Nogo-A expression and Caspase-3 activity in the ischemic brain tissues.

Determination of the effective dose of bone marrow mononuclear cell therapy for bone healing in vivo

IntroductionCell-based therapy by bone marrow mononuclear cells (BMC) in a large-sized bone defect has already shown improved vascularization and new bone formation. First clinical trials are already being conducted. BMC were isolated from bone marrow aspirate and given back to patients in combination with a scaffold within some hours. However, the optimal concentration of BMC has not yet been determined for bone healing. With this study, we want to determine the optimal dosage of the BMC in the bone defect to support bone healing.Material and methodsScaffolds with increasing BMC concentrations were inserted into a 5 mm femoral defect, cell concentrations of 2 × 106 BMC/mL, 1 × 107 BMC/mL and 2 × 107 BMC/mL were used. Based on the initial cell number used to colonize the scaffolds, the groups are designated 1 × 106, 5 × 106 and 1 × 107 group. Bone healing was assessed biomechanically, radiologically (µCT), and histologically after 8 weeks healing time.ResultsImproved bone healing parameters were noted in the 1 × 106 and 5 × 106 BMC groups. A significantly higher BMD was observed in the 1 × 106 BMC group compared to the other groups. Histologically, a significantly increased bone growth in the defect area was observed in group 5 × 106 BMC. This finding could be supported radiologically.ConclusionIt was shown that the effective dose of BMC for bone defect healing ranges from 2 × 106 BMC/mL to 1 × 107 BMC/mL. This concentration range seems to be the therapeutic window for BMC-supported therapy of large bone defects. However, further studies are necessary to clarify the exact BMC-dose dependent mechanisms of bone defect healing and to determine the therapeutically effective range more precisely.

Comparing the efficacy of core decompression with autologous bone marrow mononuclear cell grafting with impacted bone grafting in treating bilateral femoral head necrosis

Objective To compare the efficacy of core decompression with autologous bone marrow mononuclear cell grafting with impacted bone grafting in treating bilateral femoral head necrosis. Methods From January 2011 to January 2017, a total of 44 patients with bilateral femoral head necrosis (88 hips) were admitted to the Department of Orthopaedics, China-Japan Friendship Hospital, including 35 males and 9 females with aged 34.9±7.2 years old (ranged from 22-48 years). Core decompression with autologous bone marrow mononuclear cell and impacted bone grafting were conducted to each hip joint for every patient. All patients were followed up for every 3 months at the first year postoperatively and for every 6 months thereafter. The following clinical measurement were recorded, Harris hip score (HHS), visual analogue score (VAS), the anterior-posterior and frog lateral radiographs, and CT. The 5-year survival rate of the hip was calculated with the endpoint event being defined as a need for total hip arthroplasty or other surgical intervention, or a HHS less than 70. Results The postoperative follow-up duration was 50.5±34.2 months in the impacted bone grafting group and 54.0±33.1 months in the core decompression with autologous bone marrow mononuclear cells group. Fifteen hips in the impacted bone grafting group and 13 hips in the core decompression with autologous bone marrow mononuclear cell group failed during the follow-up. The 5-year cumulative survival rates of the hips in two groups were 64.7% and 72.1%, respectively [HR=1.178, 95%CI(0.561, 2.477)]. In the impacted bone grafting group, the 5-year survival rates of the hip joints at the ARCO IIIB+IIIC and IIIA stages were 42.9% and 74.2%, respectively [HR=3.258, 95%CI(1.172, 9.059)]. In the core decompression with autologous bone marrow mononuclear cell group, the 5-year survival rates of hips at the ARCO stage I, II and IIIA stages were 50.0%, 75.3%, and 71.4%, respectively (χ2=0.757, P=0.685). Age, gender, BMI, preoperative HHS and etiology did not affect the effects of core decompression with autologous bone marrow mononuclear cell grafting or impacted bone grafting (P>0.05). The preoperative VAS of the impacted bone grafting group and the core decompression with autologous bone marrow mononuclear cell grafting group were 4.80±0.62 and 3.27±1.17, respectively (t=8.625, P<0.001). At the last follow-up, the VAS was reduced to 2.84±1.95 and 2.25±2.08, respectively (t=2.712, P=0.01; t=7.087, P<0.001) with significant difference in postoperative VAS between the two groups (t=2.489, P=0.017). The preoperative HHS of the impacted bone grafting group and the core decompression with autologous bone marrow mononuclear cell grafting group were 77.02±5.03 and 82.57±5.71, respectively (t=7.822, P<0.001). At the last follow-up, the HHS increased to 81.57±12.81 and 83.55±12.87, respectively. The difference between the preoperative and postoperative HHS was statistically significant in the impacted bone grafting group (t=2.389, P=0.021) but not in the core decompression with autologous bone marrow mononuclear cell grafting group (t=0.451, P=0.654). There was no significant difference in postoperative HHS between the two groups (t=1.353, P=0.183). Conclusion Both impacted bone grafting and core decompression with autologous bone marrow mononuclear cell grafting are safe and effective methods in treating femoral head necrosis. The ARCO stage is a risk factor affecting the prognosis of hips after impacted bone grafting, which has no effect on the mid-term survival of hips after core decompression with autologous bone marrow mononuclear cell grafting. Key words: Femur head necrosis; Bone transplantatio; Transplantation, autologous; Bone marrow transplantation

Immune Tolerance Induced by Decitabine Combined With Multiple Allogeneic Bone Marrow Mononuclear Cell Infusion

ObjectiveThe aim of our study was to investigate the strategy of immune tolerance induced by the demethylating drug decitabine (DAC) combined with multiple allogeneic bone marrow mononuclear cell (BMMNC) infusions. MethodsAn animal model with C57BL/6 mice as recipients and a BALB/c mouse as the donor was established. C57BL/6 mice were randomly divided into the following groups: DAC, BMMNC, DAC+BMMNC, and the control groups. Mice in the DAC and DAC+BMMNC groups were given DAC (5 μg/kg) on days 1, 2, 7, 8, 13, and 14. On experimental days 3, 9, and 15, BMMNCs (5 × 106/mouse/time) from BALB/c mice were infused into mice of the BMMNC and DAC+BMMNC groups. One-way mixed lymphocyte reaction (MLR), interleukin-2 reverse test, and in vitro adoptive transfer experiments were performed. The ratio of regulatory T cells in splenocytes, chimera detection, and median survival time of skin grafts were recorded. ResultsMLR showed that the stimulation index of the DAC+BMMNC group was significantly lower compared with the other 3 groups (P < .01), but still showed a strong proliferative response to the third-party unrelated donor KM mice, with no significant difference compared with the other 3 groups. The MLR low reactivity of the DAC+BMMNC group could be reversed by exogenous interleukin-2. After adding an equal amount of splenocytes from a C57BL/6 mouse in the DAC+BMMNC group, MLR of control group significantly decreased (P < .05). Flow cytometry showed the ratio of regulatory T cells in splenocytes from DAC and DAC+BMMNC groups was significantly higher than that from the BMMNC and control groups (P < .05). Chimeric rate of the DAC+BMMNC (60%) group was significantly higher than that of the BMMNC group (30%). Moreover, survival time of skin grafts in the DAC+BMMNC group was significantly longer than that in the other 3 groups (P < .001). No death or graft-vs-host disease was observed. ConclusionDAC combined with multiple allogeneic BMMNC infusion could successfully induce specific immune tolerance in mice, which may provide some new strategies to improve immune tolerance after organ transplant.

Bone Marrow Mononuclear Cells Transfer for Patients after ST-Elevated Myocardial Infarction: A Meta-Analysis of Randomized Control Trials.

PurposeResults on the clinical utility of cell therapy for ST-elevated myocardial infarction (STEMI) are controversial. This study sought to analyze the efficacy of treatment with intracoronary bone marrow mononuclear cells (BMMC) on left ventricular (LV) function and remodeling and LV diastolic and systolic function in patients with STEMI.Materials and MethodsLiterature search of PubMed and EMBASE databases between 2004 and 2017 was performed for randomized controlled trials in STEMI patients who underwent successful percutaneous coronary intervention and received intracoronary BMMC therapy. The defined end points were left ventricular ejection fraction (LVEF), left ventricular end-diastolic volume (LVEDV), and left ventricular end-systolic volume (LVESV). Also, sensitivity analysis and several subgroup analyses based on follow-up duration, timing of injection, doses of cells, and imaging modalities were conducted to strengthen the statistic power of the study.ResultsA total of 22 trials with 1360 patients were available for the current meta-analysis. The pooled statistics showed a significant improvement in LVEF {2.58 [95% confidence interval (CI), 1.32, 3.84]; p<0.001}, LVEDV [−3.73, (95% CI, −6.94, −0.52), p=0.02], and LVESV [−4.67, (95% CI, −7.07, −2.28), p<0.001] in the BMMC group, compared with the control group. However, in sensitivity analysis, a significant reduction in LVEDV disappeared, while the outcomes of LVEF and LVESV remained unchanged. The same results were presented in the subgroup analysis adjusting for imaging modalities and timing of cells injection.ConclusionBMMC transplantation in patients with STEMI was found to lead to improvement in LVEF, LVEDV, and LVESV parameters, indicating that cell therapy has a potential beneficial effect on LV remodeling and function.

The bone marrow mononuclear cells reduce the oxidative stress of cerebral infarction through PI3K/AKT/NRF2 signaling pathway.

To evaluate the mechanism of bone marrow mononuclear cells (BMMNCs) in reducing the oxidative stress after cerebral infarction through PI3K/AKT/NRF2 signaling pathway. 96 healthy SD rats, which were 6-8-week old, weighting about 250-280 g, were selected for the study. The middle cerebral artery occlusion model (MCAO) was established in SD rats using the suture method. The rats were randomly divided into sham operation group, model group, BMMNCs group and PI3K inhibitor group. 24 rats in each group were selected. 200 μl phosphate-buffered saline (PBS) solution was injected into the caudal vein of the rats in the model group, 200 μl PBS solution containing 5×106 BMMNCs that obtained by gradient centrifugation was injected into the rats in the BMMNCs group, meanwhile, in the PI3K inhibitor group, LY294002 (10 mmol/L/kg) was injected into the lateral ventricle of the brain. After the 3d, 7d and 14d, the modified neurological severity scores (mNSS) were used to evaluate the neurological function. The volume of cerebral infarction was assessed by TTC staining, the VEGF, BDNF, TNF-α, IL-1β, malondialdehyde (MDA), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) levels were detected by ELISA. The mNSS and the volume of cerebral infarction of the model group were significantly higher than those of the sham operation group (p<0.05), while the mNSS and the volume of cerebral infarction of the BMMNCs group were lower than those of the model group, higher than those of the sham operation group (p<0.05). The VEGF, BDNF, TNF-α, IL-1β, MDA, SOD and GSH-Px levels of the model group were significantly higher than those of the sham operation group (p<0.05). BMMNCs can reduce the oxidative stress, apoptosis, and inflammatory reaction through PI3K/AKT/NRF2 signaling pathway, thus promoting the secretion of nerve and vascular cytokines, improving the neurological function and reducing the infarct scope.

Efficacy, safety and influencing factors of intra-calf muscular injection of bone marrow mononuclear cells in the treatment of type 2 diabetes mellitus-induced lower extremity vascular disease.

The efficacy, safety and impact of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) associated with the intra-calf muscular injection of bone marrow mononuclear cells (BMMCs) in the treatment of type 2 diabetes mellitus (T2DM)-induced lower extremity vascular disease (LEVD) were evaluated. Patients with T2DM-LEVD were randomly divided into a control group and BMMCs group to assess the efficacy and safety of the treatment; serum VEGF and bFGF levels were detected. The BMMCs group was divided into superior genicular artery (SGA) and inferior genicular artery (IGA) subgroups as well as low-dose and high-dose subgroups for the comparison of efficacy indices. The BMMCs group exhibited significantly improved indices (P<0.05) compared with the control group and no fatalities or cancer occurred. There were no significant changes in serum VEGF and bFGF levels (P>0.05). The claudication distance in the IGA subgroup was significantly greater that in the SGA subgroup (P<0.05); the low-dose subgroup and the high-dose subgroup did not demonstrate any significant differences in each index (P>0.05). BMMC treatment for T2DM-LEVD was found to be safe and effective and had no significant impact on serum VEGF and bFGF levels in the short term; However, the degree of LEVD may affect its efficacy.

Chitosan nerve conduits seeded with autologous bone marrow mononuclear cells for 30\u2009mm goat peroneal nerve defect

In the current research, to find if the combination of chitosan nerve conduits seeded with autologous bone marrow mononuclear cells (BM-MNCs) can be used to bridge 30 mm long peroneal nerve defects in goats, 15 animals were separated into BM-MNC group (n = 5), vehicle group (n = 5), and autologous nerve graft group (n = 5). 12 months after the surgery, animals were evaluated by behavioral observation, magnetic resonance imaging tests, histomorphological and electrophysiological analysis. Results revealed that animals in BM-MNC group and autologous nerve graft group achieved fine functional recovery; magnetic resonance imaging tests and histomorphometry analysis showed that the nerve defect was bridged by myelinated nerve axons in those animals. No significant difference was found between the two groups concerning myelinated axon density, axon diameter, myelin sheath thickness and peroneal nerve action potential. Animals in vehicle group failed to achieve significant functional recovery. The results indicated that chitosan nerve conduits seeded with autologous bone marrow mononuclear cells have strong potential in bridging long peripheral nerve defects and could be applied in future clinical trials.

Comparative analysis of curative effect of bone marrow mesenchymal stem cell and bone marrow mononuclear cell transplantation for spastic cerebral palsy

BackgroundBone marrow mesenchymal stem cells (BMMSCs) and bone marrow mononuclear cells (BMMNCs) are both used to treat spastic cerebral palsy. However, the differences in therapeutic effect remain unknown.MethodsA total of 105 patients with spastic cerebral palsy were enrolled and randomly assigned to three groups: the BMMSC group, the BMMNC group and the control group. Patients in both transplantation groups received four intrathecal cell injections. Patients in the control group received Bobath therapy. The gross motor function measure (GMFM) and the fine motor function measure (FMFM) were used to evaluate the therapeutic efficacy before transplantation and 3, 6, and 12 months after transplantation.ResultsThree months after cell transplantation, scores in the A dimension of GMFM and the A and C dimensions of FMFM scores in the BMMSC group are all higher than those of the BMMNC and the control groups (P < 0.05). Six months after cell transplantation, scores in the A, B dimensions of GMFM and the A, B, C, D, and E dimensions of FMFM scores in the BMMSC group are higher than those of the BMMNC and the control groups (P < 0.05). Twelve months after cell transplantation, scores in the A, B, and C dimensions of GMFM and the A, B, C, D, and E dimensions of FMFM scores in the BMMSC group are all higher than those of the BMMNC and the control groups (P < 0.05). No obvious adverse effects were investigated during follow-up.ConclusionsBMMSC transplantation for the treatment of cerebral palsy is safe and feasible, and can improve gross motor and fine motor function significantly. In addition, compared with BMMNC, the motor function of children improved significantly in terms of gross motor and fine motor functions.

Neuroprotection by intravenous transplantation of bone marrow mononuclear cells from 5-fluorouracil pre-treated rats in a model of ischemic stroke

Our previous studies showed that bone marrow mononuclear cells (BMMNCs) from 5-fluorouracil (5-FU) pre-treated rats (named BMRMNCs) had a better therapeutic efficacy in ischemia/reperfusion rats as compared to BMMNCs from untreated rats. This study was undertaken to further explore the potential mechanisms underlying the neuroprotective effects of BMRMNCs in the same model. Rats were intravenously pre-treated with 5-FU, and BMRMNCs were collected 7 days later and subjected to flow cytometry for detection of CD34, CD45 and CD90. Middle cerebral artery occlusion (MCAO) was induced in rats, and BMMNCs and BMRMNCs were independently transplanted via the tail vein at 24 h after MCAO. NISSL staining was performed 14 days after cell transplantation and the viable cells in the hippocampus were counted. Stromal cell-derived factor 1 (SDF-1) mRNA expression was detected in the penumbra at 7 and 14 days after treatment. The contents of pro-inflammatory cytokines and growth factors as well as microvessel density (MVD) were determined at 14 days. Results showed more BMRMNCs were positive for CD34, CD45 and CD90. After transplantation, more viable cells were observed in the hippocampus of BMRMNCs treated rats. In addition, BMRMNCs transplantation significantly increased MVD, reduced pro-inflammatory cytokines and raised growth factors in the penumbra. However, the SDF-1 mRNA expression was comparable between BMRMNCs group and BMMNCs group. Our results indicate that BMRMNCs are likely to more effectively improve the local microenvironment to increase viable cells and elevate angiogenesis, exerting neuroprotective effects on cerebral ischemia in rats.

Bone Marrow Mononuclear Cell Transplantation Restores Inflammatory Balance of Cytokines after ST Segment Elevation Myocardial Infarction

BackgroundAcute myocardial infarction (AMI) launches an inflammatory response and a repair process to compensate cardiac function. During this process, the balance between proinflammatory and anti-inflammatory cytokines is important for optimal cardiac repair. Stem cell transplantation after AMI improves tissue repair and increases the ventricular ejection fraction. Here, we studied in detail the acute effect of bone marrow mononuclear cell (BMMNC) transplantation on proinflammatory and anti-inflammatory cytokines in patients with ST segment elevation myocardial infarction (STEMI).MethodsPatients with STEMI treated with thrombolysis followed by percutaneous coronary intervention (PCI) were randomly assigned to receive either BMMNC or saline as an intracoronary injection. Cardiac function was evaluated by left ventricle angiogram during the PCI and again after 6 months. The concentrations of 27 cytokines were measured from plasma samples up to 4 days after the PCI and the intracoronary injection.ResultsTwenty-six patients (control group, n = 12; BMMNC group, n = 14) from the previously reported FINCELL study (n = 80) were included to this study. At day 2, the change in the proinflammatory cytokines correlated with the change in the anti-inflammatory cytokines in both groups (Kendall’s tau, control 0.6; BMMNC 0.7). At day 4, the correlation had completely disappeared in the control group but was preserved in the BMMNC group (Kendall’s tau, control 0.3; BMMNC 0.7).ConclusionsBMMNC transplantation is associated with preserved balance between pro- and anti-inflammatory cytokines after STEMI in PCI-treated patients. This may partly explain the favorable effect of stem cell transplantation after AMI.