Computing cell state discriminates the aberrant hematopoiesis and activated microenvironment in Myelodysplastic syndrome (MDS) through a single cell genomic study.

BACKGROUND: Myelodysplastic syndrome (MDS) is a complicated hematopoietic malignancy characterized by bone marrow (BM) dysplasia with symptoms like anemia, neutropenia, or thrombocytopenia. MDS exhibits considerable heterogeneity in prognosis, with approximately 30% of patients progressing to acute myeloid leukemia (AML). Single cell RNA-sequencing (scRNA-seq) is a new and powerful technique to profile disease landscapes. However, the current available scRNA-seq datasets for MDS are only focused on CD34+ hematopoietic progenitor cells. We argue that using entire BM cell for MDS studies probably will be more informative for understanding the pathophysiology of MDS. METHODS: Five MDS patients and four healthy donors were enrolled in the study. Unsorted cells from BM aspiration were collected for scRNA-seq analysis to profile overall alteration in hematopoiesis. RESULTS: Standard scRNA-seq analysis of unsorted BM cells successfully profiles deficient hematopoiesis in all five MDS patients, with three classified as high-risk and two as low-risk. While no significant increase in mutation burden was observed, high-risk MDS patients exhibited T-cell activation and abnormal myelogenesis at the stages between hematopoietic stem and progenitor cells (HSPC) and granulocyte-macrophage progenitors (GMP). Transcriptional factor analysis on the aberrant myelogenesis suggests that the epigenetic regulator chromatin structural protein-encoding gene HMGA1 is highly activated in the high-risk MDS group and moderately activated in the low-risk MDS group. Perturbation of HMGA1 by CellOracle simulated deficient hematopoiesis in mouse Lineage-negative (Lin-) BM cells. Projecting MDS and AML cells on a BM cell reference by our newly developed MarcoPolo pipeline intuitively visualizes a connection for myeloid leukemia development and abnormalities of hematopoietic hierarchy, indicating that it is technically feasible to integrate all diseased bone marrow cells on a common reference map even when the size of the cohort reaches to 1,000 patients or more. CONCLUSION: Through scRNA-seq analysis on unsorted cells from BM aspiration samples of MDS patients, this study systematically profiled the development abnormalities in hematopoiesis, heterogeneity of risk, and T-cell microenvironment at the single cell level.

ATP-P2X7R-mediated microglia senescence aggravates retinal ganglion cell injury in chronic ocular hypertension.

BACKGROUND: Dysfunction of microglia during aging affects normal neuronal function and results in the occurrence of neurodegenerative diseases. Retinal microglial senescence attributes to retinal ganglion cell (RGC) death in glaucoma. This study aims to examine the role of ATP-P2X7R in the mediation of microglia senescence and glaucoma progression. METHODS: Forty-eight participants were enrolled, including 24 patients with primary open-angle glaucoma (POAG) and age-related cataract (ARC) and 24 patients with ARC only. We used ARC as the inclusion criteria because of the availability of aqueous humor (AH) before phacoemulsification. AH was collected and the adenosine triphosphate (ATP) concentration was measured by ATP Assay Kit. The chronic ocular hypertension (COH) mouse model was established by microbead occlusion. Microglia were ablated by feeding PLX5622 orally. Mouse bone marrow cells (BMCs) were prepared and infused into mice through the tail vein for the restoration of microglia function. Western blotting, qPCR and ELISA were performed to analyze protein and mRNA expression in the ocular tissue, respectively. Microglial phenotype and RGC survival were assessed by immunofluorescence. The mitochondrial membrane potential was measured using a JC-1 assay kit by flow cytometry. RESULTS: ATP concentrations in the AH were increased in older adults and patients with POAG. The expression of P2X7R was upregulated in the retinal tissues of mice with glaucoma, and functional enrichment analysis showed that P2X7R was closely related to cell aging. Through in vivo and in vitro approaches, we showed that pathological activation of ATP-P2X7R induced accelerated microglial senescence through impairing PTEN-induced kinase 1 (PINK1)-mediated mitophagy, which led to RGC damage. Additionally, we found that replacement of senescent microglia in COH model of old mice with BMCs from young mice reversed RGC damage. CONCLUSION: ATP-P2X7R induces microglia senescence by inhibiting PINK1-mediated mitophagy pathway. Specific inhibition of ATP-P2X7R may be a fundamental approach for targeted therapy of RGC injury in microglial aging-related glaucoma.

Improved YOLOv7 Algorithm for Detecting Bone Marrow Cells.

The detection and classification of bone marrow (BM) cells is a critical cornerstone for hematology diagnosis. However, the low accuracy caused by few BM-cell data samples, subtle difference between classes, and small target size, pathologists still need to perform thousands of manual identifications daily. To address the above issues, we propose an improved BM-cell-detection algorithm in this paper, called YOLOv7-CTA. Firstly, to enhance the model's sensitivity to fine-grained features, we design a new module called CoTLAN in the backbone network to enable the model to perform long-term modeling between target feature information. Then, in order to cooperate with the CoTLAN module to pay more attention to the features in the area to be detected, we integrate the coordinate attention (CoordAtt) module between the CoTLAN modules to improve the model's attention to small target features. Finally, we cluster the target boxes of the BM cell dataset based on K-means++ to generate more suitable anchor boxes, which accelerates the convergence of the improved model. In addition, in order to solve the imbalance between positive and negative samples in BM-cell pictures, we use the Focal loss function to replace the multi-class cross entropy. Experimental results demonstrate that the best mean average precision (mAP) of the proposed model reaches 88.6%, which is an improvement of 12.9%, 8.3%, and 6.7% compared with that of the Faster R-CNN model, YOLOv5l model, and YOLOv7 model, respectively. This verifies the effectiveness and superiority of the YOLOv7-CTA model in BM-cell-detection tasks.

Accumulation of Dystrophin-Positive Muscle Fibers and Improvement of Neuromuscular Junctions in mdx Mouse Muscles after Bone Marrow Transplantation under Different Conditions.

Duchenne muscular dystrophy (DMD) is a severe muscular disorder caused by mutations in the dystrophin gene. It leads to respiratory and cardiac failure and premature death at a young age. Although recent studies have greatly deepened the understanding of the primary and secondary pathogenetic mechanisms of DMD, an effective treatment remains elusive. In recent decades, stem cells have emerged as a novel therapeutic product for a variety of diseases. In this study, we investigated nonmyeloablative bone marrow cell (BMC) transplantation as a method of cell therapy for DMD in an mdx mouse model. By using BMC transplantation from GFP-positive mice, we confirmed that BMCs participate in the muscle restoration of mdx mice. We analyzed both syngeneic and allogeneic BMC transplantation under different conditions. Our data indicated that 3 Gy X-ray irradiation with subsequent BMC transplantation improved dystrophin synthesis and the structure of striated muscle fibers (SMFs) in mdx mice as well as decreasing the death rate of SMFs. In addition, we observed the normalization of neuromuscular junctions (NMJs) in mdx mice after nonmyeloablative BMC transplantation. In conclusion, we demonstrated that nonmyeloablative BMC transplantation could be considered a method for DMD treatment.

[Radioprotective effects of gallic acid on bone marrow cells in mice].

OBJECTIVE: To investigate the radioprotective effect of gallic acid(GA) on mouse bone marrow cells. METHODS: Healthy male ICR mice were randomly divided into saline control group, GA control group, X-ray irradiation group and GA protection group, with 10 mice in each group. X-ray irradiation group and normal saline control group were given 0.01 mL/g normal saline gavage, GA control group and GA protection group were given 200 mg/kg GA(20 mg/mL) gavage once a day for 14 consecutive days. On the 15 th day, 4 X-ray irradiation groups and 4 GA protection groups were given one-time X-ray irradiation to the whole body of the mice, and the absorbed doses were 1.0, 2.0, 3.0 and 4.0 Gy, respectively. The saline control group and the GA control group were not irradiated. After irradiation, detected the whole blood catalase(CAT), superoxide dismutase(SOD), malondialdehyde(MDA) and micronucleus frequency of polychromatic erythrocyte in bone marrow(MN-PCE), and use flow cytometry to detect bone marrow cell cycle, early apoptosis rate and late apoptosis rate. RESULTS: The CAT activities in the serum of mice in the 1.0, 2.0, 3.0 and 4.0 Gy GA protection groups were 2.13, 1.74, 1.49 and 1.15 U/mL, respectively, which were significantly increased compared with the corresponding X-ray irradiation group(P<0.01). SOD activities were 184.69, 156.92, 139.17 and 107.15 U/mL, which were significantly increased compared with the corresponding X-ray irradiation group(P<0.01). The contents of MDA were 3.92, 4.20, 6.32 and 9.31 nmol/mL, which were significantly lower than those of the corresponding X-ray irradiation group(P<0.05, P<0.01). The bone marrow MN-PCE rate of the mice in the 1.0, 2.0, 3.0 and 4.0 Gy GA protection groups were 4.35‰, 8.00‰, 12.90‰ and 3.80‰, respectively, which were significantly lower than those in the corresponding X-ray irradiation group(P<0.01). The proportions of G_0/G_1 phase cells of bone marrow cells in the 1.0, 2.0, 3.0 and 4.0 Gy GA protection group were 81.00%, 86.28%, 92.04% and 93.15%, respectively, which were significantly reduced compared with the corresponding X-ray irradiation group(P<0.01). The proportions of G_2/M phase cells were 4.51%, 3.05%, 2.35% and 1.81%, which were significantly increased compared with the corresponding X-ray irradiation group(P<0.05, P<0.01). The proportions of S phase cells were 15.32%, 11.36%, 5.96% and 4.92%, which were significantly increased compared with the corresponding X-ray irradiation group(P<0.01). The early apoptosis rate of bone marrow cells of mice in the 1.0, 2.0, 3.0 and 4.0 Gy GA protection groups were 3.32%, 8.96%, 12.11% and 2.26%, respectively, which were significantly reduced compared with the corresponding X-ray irradiation group(P<0.01). The late apoptosis rates of bone marrow cells were 7.21%, 11.73%, 17.11% and 19.36%, which were significantly reduced compared with the corresponding X-ray irradiation group(P<0.05, P<0.01). CONCLUSION: GA can reduce the oxidative damage, DNA damage and bone marrow cell cycle arrest of the bone marrow cells of radiation-damaged mice, inhibit the apoptosis of bone marrow cells, and have radioprotective effects on the bone marrow cells of mice.

Influence of estradiol treatment on bone marrow cell differentiation in collagenase-induced arthritis.

OBJECTIVE AND DESIGN: Estrogen is one of the important regulators of the balance between bone formation and bone resorption that can modulate the levels and activity of certain growth factors and cytokines. In this study, we investigated the effect of 17ß-estradiol (ED) on bone marrow (BM) cell differentiation in vivo and ex vivo in a mouse model of collagenase-induced osteoarthritis (CIOA). SUBJECT: ICR (CD-2) female mice were used in present experiments (total number = 75) and bone marrow cells were used for in vitro studies. TREATMENT: Mice were orally fed under different schemes with 17ß-estradiol at a dose of 2 µg or 4 µg for 30 days. METHODS: The effect of estradiol was estimated by histopathological, flow cytometry, and ELISA assays. Statistical differences were determined by one-way ANOVA. RESULTS: Estradiol treatment ameliorated cartilage destruction and osteophyte formation if started from day 0 of CIOA induction, attended with a decrease of uterine and ovarian weights. Long time treatment lowered the percentage of megakaryocyte/platelet (CD62P+) populations and osteoclast (RANK+) populations in BM. Cells obtained from estradiol-treated CIOA mice showed inhibited capacity to differentiate into RANK+ and mesenchymal cells under osteoclastogenic conditions in vitro. Estrogen decreased serum IL-6 levels. CONCLUSION: Results indicate a potential protective role for estrogen against the development of OA.

Fabrication and Biological Analysis of Highly Porous PEEK Bionanocomposites Incorporated with Carbon and Hydroxyapatite Nanoparticles for Biological Applications.

Bone regeneration for replacing and repairing damaged and defective bones in the human body has attracted much attention over the last decade. In this research, highly porous polyetheretherketone (PEEK)/hydroxyapatite (HA) bionanocomposite scaffolds reinforced with carbon fiber (CF) and carbon nanotubes (CNTs) were fabricated, and their structural, mechanical, and biological properties were studied in detail. Salt porogen (200-500 µm size) leaching methods were adapted to produce porous PEEK structures with controlled pore size and distribution, facilitating greater cellular infiltration and biological integration of PEEK composites within patient tissue. In biological tests, nanocomposites proved to be non-toxic and have very good cell viability. In addition, bone marrow cell growth was observed, and PEEK/HA biocomposites with carbon particles showed increased cell attachment over the neat PEEK/HA composites. In cell viability tests, bionanocomposites with 0.5 wt% CNTs established good attachment of cells on disks compared to neat PEEK/HA biocomposites. A similar performance was seen in culture tests of bone marrow cells (osteoblasts and osteoclasts). The 0.5 wt% CF for osteoblasts and 1 wt% CNTs for osteoclasts showed higher cell attachment. The addition of carbon-based nanomaterials into PEEK/HA has been identified as an effective approach to improve cell attachment as well as mechanical and biological properties. With confirmed cell attachment and sustained viability and proliferation of the fabricated PEEK/HA/CNTs, CF bionanocomposites were confirmed to possess excellent biocompatibility and will have potential uses in bone scaffolding and other biomedical applications.

[Efficacy of treatment of lower-limb critical ischaemia by methods of indirect revascularization]. / Éffektivnost' lecheniia kriticheskoi ishemii nizhnikh konechnostei metodami nepriamoi revaskuliarizatsii.

AIM: The study was aimed at comparing the efficacy of implantation of autologous bone marrow cells with that of revascularizing osteotrephination in treatment of lower limb chronic critical ischaemia in patients with a poor distal vascular bed. PATIENTS AND METHOD: We analysed the results of comprehensive examination and treatment of a total of 60 patients presenting with lower limb chronic critical ischaemia due to atherosclerotic lesions of the femoropopliteal-tibial segment. According to the technology of treatment, the patients were divided into two statistically homogenous groups of 30 people each. Group One patients underwent standard revascularizing osteotrephination and Group Two patients in accordance with the original technique received intramuscular implantation of 40 ml of autologous bone marrow cells, with this volume distributed in 2-ml injections to 20 points of the muscles of the crus and femur along the internal and external surface. RESULTS: The use of the original technique of treatment made it possible to achieve the clinical status in the form of moderate or minimal improvement 6 months after bone marrow cells implantation in 29 (96.7%) patients and after 12 months in 28 (93.3%) patients, whereas after revascularizing osteotrephination in 25 (83.3%) and 20 (66.7%) patients, respectively. In the remote period after 12 months, the limb was saved in 28 (93.3%) and 26 (86.7%) patients in Group Two and Group One, respectively. The patients of the second group as compared with those of the first group after 12 months demonstrated a statistically significant increase in the physical health component by 19.8% and the mental health component by 9.8%. CONCLUSION: Implantation of autologous bone-marrow cells in chronic critical limb ischaemia is pathogenetically substantiated and makes it possible to optimize the results of treatment of patients.

Evaluation of the effectiveness of infusion of bone marrow derived cell in patients with heart failure: A network meta-analysis of randomized clinical trials and cohort studies.

Background: The aim of this study was to investigate the effectiveness of bone marrow-derived cells (BMC) technology in patients with heart failure and compare it with alternative therapies, including drug therapy, cardiac resynchronization therapy pacemaker (CRT-P), cardiac resynchronization therapy defibrillator (CRT-D). Methods: A systematic review study was conducted to identify all clinical studies published by 2017. Using keywords such as "Heart Failure, BMC, Drug Therapy, CRT-D, CRT-P" and combinations of the mentioned words, we searched electronic databases, including Scopus, Cochrane Library, and PubMed. The quality of the selected studies was assessed using the Cochrane Collaboration's tool and the Newcastle-Ottawa. The primary and secondary end-points were left ventricular ejection fraction (LVEF) (%), failure cases (Number), left ventricular end-systolic volume (LVES) (ml), and left ventricular end-diastolic volume (LVED) (ml). Random-effects network meta-analyses were used to conduct a systematic comparison. Statistical analysis was done using STATA. Results: This network meta-analysis covered a total of 57 final studies and 6694 patients. The Comparative effectiveness of BMC versus CRT-D, Drug, and CRT-P methods indicated the statistically significant superiority of BMC over CRT-P (6.607, 95% CI: 2.92, 10.29) in LVEF index and overall CRT-P (-13.946, 95% CI: -18.59, -9.29) and drug therapy (-4.176, 95% CI: -8.02, -.33) in LVES index. In addition, in terms of LVED index, the BMC had statistically significant differences with CRT-P (-10.187, 95% CI: -18.85, -1.52). BMC was also dominant to all methods in failure cases as a final outcome and the difference was statistically significant i.e. BMC vs CRT-D: 0.529 (0.45, 0.62) and BMC vs Drug: 0.516 (0.44, 0.60). In none of the outcomes, the other methods were statistically more efficacious than BMC. The BMC method was superior or similar to the other methods in all outcomes. Conclusion: The results of this study showed that the BMC method, in general, and especially in terms of failure cases index, had a higher level of clinical effectiveness. However, due to the lack of data asymmetry, insufficient data and head-to-head studies, BMC in this meta-analysis might be considered as an alternative to existing treatments for heart failure.

The combination of Radix Astragali and Radix Angelicae Sinensis attenuates the IFN-γ-induced immune destruction of hematopoiesis in bone marrow cells.

BACKGROUND: Radix Astragali and Radix Angelicae Sinensis are two herbs that compose Danggui Buxue Tang (an herbal formula for treatment of anemia diseases). In this study, we explored the molecular mechanism and effective targets to immune destruction of bone marrow (BM) cells treated with Radix Astragali, Radix Angelicae Sinensis or a combination of two agents. The potential synergic advantages of two herbs should also be explored. METHODS: The constituents of Radix Astragali and Radix Angelicae Sinensis were analyzed by high performance liquid chromatography-electrospray ionization/mass spectrometer system BM cells were separated from limbs of BALB/c mice, and immune destruction was induced with IFN-γ. The percentages of hematopoietic stem cells (HSCs) and CD3+ T cells were detected by flow cytometry. The distribution of T-bet and changes in the combination of SAP and SLAM in BM cells were observed by immunofluorescence. Western blotting was used to assay the expression of key molecules of the eIF2 signaling pathway in BM cells. RESULTS: Seven constituents of Radix Astragali and six constituents of Radix Angelicae Sinensis were identified. The percentages of HSCs increased significantly after treatment with Radix Angelicae Sinensis, especially at high concentrations. The percentages of CD3+ T cells were significantly decreased after Radix Astragali and Radix Angelicae Sinensis treatment. However, the synergistic function of two-herb combinations was superior to that of the individual herbs alone. The distribution of T-bet in BM cells was decreased significantly after Radix Angelicae Sinensis treatment. The number of SLAM/SAP double-stained cells was increased significantly after Radix Astragali treatment at low concentrations. The phosphorylation levels of eIF2α were also reduced after Radix Astragali and Radix Angelicae Sinensis treatment. CONCLUSIONS: Radix Astragali and Radix Angelicae Sinensis could intervene in the immunologic balance of T lymphocytes, inhibit the apoptosis of BM cells induced by immune attack, restore the balance of the T cell immune response network and recover the hematopoietic function of HSCs. The synergistic effects of Radix Astragali and Radix Angelicae Sinensis were superior to those of each herb alone.

Bone marrow-derived mesenchymal stem cells propagate immunosuppressive/anti-inflammatory macrophages in cell-to-cell contact-independent and -dependent manners under hypoxic culture.

Immunosuppressive/anti-inflammatory macrophage (Mφ), M2-Mφ that expressed the typical M2-Mφs marker, CD206, and anti-inflammatory cytokine, interleukin (IL)-10, is beneficial and expected tool for the cytotherapy against inflammatory diseases. Here, we demonstrated that bone marrow-derived lineage-positive (Lin+) blood cells proliferated and differentiated into M2-Mφs by cooperation with the bone marrow-derived mesenchymal stem cells (MSCs) under hypoxic condition: MSCs not only promoted proliferation of undifferentiated M2-Mφs, pre-M2-Mφs, in the Lin+ fraction via a proliferative effect of the MSCs-secreted macrophage colony-stimulating factor, but also promoted M2-Mφ polarization of the pre-M2-Mφs through cell-to-cell contact with the pre-M2-Mφs. Intriguingly, an inhibitor for intercellular adhesion molecule (ICAM)-1 receptor/lymphocyte function-associated antigen (LFA)-1, Rwj50271, partially suppressed expression of CD206 in the Lin+ blood cells but an inhibitor for VCAM-1 receptor/VLA-4, BIO5192, did not, suggesting that the cell-to-cell adhesion through LFA-1 on pre-M2-Mφs and ICAM-1 on MSCs was supposed to promoted the M2-Mφ polarization. Thus, the co-culture system consisting of bone marrow-derived Lin+ blood cells and MSCs under hypoxic condition was a beneficial supplier of a number of M2-Mφs, which could be clinically applicable to inflammatory diseases.

Lyophilized bone marrow cell extract functionally restores irradiation-injured salivary glands.

OBJECTIVE: Bone marrow cell extract (BMCE) was previously reported to restore salivary gland hypofunction caused by irradiation injury. Proteins were shown to be the main active factors in BMCE. However, BMCE therapy requires multiple injections and protein denaturation is a concern during BMCE storage. This study aimed to preserve, by lyophilization (freeze-drying), the bioactive factors in BMCE. METHODS: We developed a method to freeze-dry BMCE and then to analyze its ingredients and functions in vivo. Freeze-dried (FD) BMCE, freshly prepared BMCE (positive control), or saline (vehicle control) was injected into the tail vein of mice that had received irradiation to damage their salivary glands. RESULTS: Results demonstrated that the presence of angiogenesis-related factors and cytokines in FD-BMCE remained comparable to those found in fresh BMCE. Both fresh and FD-BMCE restored comparably saliva secretion, increased cell proliferation, upregulated regenerative/repair genes, protected salivary acinar cells, parasympathetic nerves, and blood vessels from irradiation-damaged salivary glands. CONCLUSION: Lyophilization of BMCE maintained its bioactivity and therapeutic effect on irradiation-injured salivary glands. The advantages of freeze-drying BMCE are its storage and transport at ambient temperature.

Intracoronary autologous bone marrow cell transfer after myocardial infarction: the BOOST-2 randomised placebo-controlled clinical trial.

AIMS: Intracoronary infusion of autologous nucleated bone marrow cells (BMCs) enhanced the recovery of left ventricular ejection fraction (LVEF) after ST-segment elevation myocardial infarction (STEMI) in the randomised-controlled, open-label BOOST trial. We reassessed the therapeutic potential of nucleated BMCs in the randomised placebo-controlled, double-blind BOOST-2 trial conducted in 10 centres in Germany and Norway. METHODS AND RESULTS: Using a multiple arm design, we investigated the dose-response relationship and explored whether γ-irradiation which eliminates the clonogenic potential of stem and progenitor cells has an impact on BMC efficacy. Between 9 March 2006 and 16 July 2013, 153 patients with large STEMI were randomly assigned to receive a single intracoronary infusion of placebo (control group), high-dose (hi)BMCs, low-dose (lo)BMCs, irradiated hiBMCs, or irradiated loBMCs 8.1 ± 2.6 days after percutaneous coronary intervention (PCI) in addition to guideline-recommended medical treatment. Change in LVEF from baseline (before cell infusion) to 6 months as determined by MRI was the primary endpoint. The trial is registered at Current Controlled Trials (ISRCTN17457407). Baseline LVEF was 45.0 ± 8.5% in the overall population. At 6 months, LVEF had increased by 3.3 percentage points in the control group and 4.3 percentage points in the hiBMC group. The estimated treatment effect was 1.0 percentage points (95% confidence interval, -2.6 to 4.7; P = 0.57). The treatment effect of loBMCs was 0.5 percentage points (-3.0 to 4.1; P = 0.76). Likewise, irradiated BMCs did not have significant treatment effects. BMC transfer was safe and not associated with adverse clinical events. CONCLUSION: The BOOST-2 trial does not support the use of nucleated BMCs in patients with STEMI and moderately reduced LVEF treated according to current standards of early PCI and drug therapy.

Comparative analysis of cat bone marrow and adipose tissue cell cultures.

Cell culture transplantation is very promising in the treatment of various diseases. Cells obtained from a number of sources have been analysed to provide a basis for further studies in the area of regenerative medicine. The objective of the study was to compare morphological and phenotypic changes in cat adipose tissue and bone marrow cell cultures from the first to fifth passages. Adipose tissue and bone marrow were used to obtain cell cultures (coming from 3 cats) using standard methods with own modification. Phenotype changes were monitored by CD-marker identification and CD pan-keratin. The cytogenetic analysis was performed on 50 metaphase plates of cell cultures from the first to fifth passage. Cytogenetic assays showed that the adipose tissue cell culture (ATCC) at all passages was more stable than the bone marrow cell culture (BMCC).

Bone marrow transplantation improves proximal tubule dysfunction in a mouse model of Dent disease.

Dent disease is a rare X-linked tubulopathy caused by mutations in the endosomal chloride-proton exchanger (ClC-5) resulting in defective receptor-mediated endocytosis and severe proximal tubule dysfunction. Bone marrow transplantation has recently been shown to preserve kidney function in cystinosis, a lysosomal storage disease causing proximal tubule dysfunction. Here we test the effects of bone marrow transplantation in Clcn5Y/- mice, a faithful model for Dent disease. Transplantation of wild-type bone marrow in Clcn5Y/- mice significantly improved proximal tubule dysfunction, with decreased low-molecular-weight proteinuria, glycosuria, calciuria, and polyuria four months after transplantation, compared to Clcn5Y/- mice transplanted with ClC-5 knockout bone marrow. Bone marrow-derived cells engrafted in the interstitium, surrounding proximal tubule cells, which showed a rescue of the apical expression of ClC-5 and megalin receptors. The improvement of proximal tubule dysfunction correlated with Clcn5 gene expression in kidneys of mice transplanted with wild-type bone marrow cells. Coculture of Clcn5Y/- proximal tubule cells with bone marrow-derived cells confirmed rescue of ClC-5 and megalin, resulting in improved endocytosis. Nanotubular extensions between the engrafted bone marrow-derived cells and proximal tubule cells were observed in vivo and in vitro. No rescue was found when the formation of the tunneling nanotubes was prevented by actin depolymerization or when cells were physically separated by transwell inserts. Thus, bone marrow transplantation may rescue the epithelial phenotype due to an inherited endosomal defect. Direct contacts between bone marrow-derived cells and diseased tubular cells play a key role in the rescue mechanism.

Safety and efficacy of percutaneous intramyocardial bone marrow cell injection for chronic myocardial ischemia: Long-term results.

BACKGROUND: Intramyocardial injection of bone marrow cells (BMC) in refractory angina patients with chronic myocardial ischemia has shown to be safe and improve clinical status during short-term follow-up. However, scarce data are available on long-term (>12 months) safety and efficacy. Therefore, the occurrence of clinical events and the long-term clinical effects of intramyocardial BMC injection were evaluated in patients with chronic myocardial ischemia up to 10 years after treatment. METHODS AND RESULTS: Patients (n = 100, age 64 ± 9 years, male 88%) with chronic myocardial ischemia who underwent intramyocardial BMC injection between 2004 and 2010 were evaluated. During yearly outpatient clinic visits, the occurrence of clinical events was documented. In addition, clinical status was assessed according to the Canadian Cardiovascular Society (CCS) score and quality of life was measured using the Seattle Angina Questionnaire. These parameters were evaluated at baseline and during the first year, followed by cross-sectional long-term follow-up which was performed in 2011 and 2014. No adverse events considered related to the procedure occurred during 10 years of follow-up. Observed annual mortality rate and annual myocardial infarction rate were 3.8% and 1.9% per year, respectively. When compared to baseline, CCS class and quality of life remained significantly better during 5-year follow-up after BMC treatment (both P < 0.05). CONCLUSIONS: The present long-term follow-up study shows that intramyocardial BMC injection in patients with chronic myocardial ischemia is safe and improves both angina complaints and quality of life up to 5 years after BMC treatment.

Combining Growth Factor and Bone Marrow Cell Therapy Induces Bleeding and Alters Immune Response After Stroke in Mice.

BACKGROUND AND PURPOSE: Bone marrow cell (BMC)-based therapies, either the transplantation of exogenous cells or stimulation of endogenous cells by growth factors like the granulocyte colony-stimulating factor (G-CSF), are considered a promising means of treating stroke. In contrast to large preclinical evidence, however, a recent clinical stroke trial on G-CSF was neutral. We, therefore, aimed to investigate possible synergistic effects of co-administration of G-CSF and BMCs after experimental stroke in mice to enhance the efficacy compared with single treatments. METHODS: We used an animal model for experimental stroke as paradigm to study possible synergistic effects of co-administration of G-CSF and BMCs on the functional outcome and the pathophysiological mechanism. RESULTS: G-CSF treatment alone led to an improved functional outcome, a reduced infarct volume, increased blood vessel stabilization, and decreased overall inflammation. Surprisingly, the combination of G-CSF and BMCs abrogated G-CSFs' beneficial effects and resulted in increased hemorrhagic infarct transformation, altered blood-brain barrier, excessive astrogliosis, and altered immune cell polarization. These increased rates of infarct bleeding were mainly mediated by elevated matrix metalloproteinase-9-mediated blood-brain barrier breakdown in G-CSF- and BMCs-treated animals combined with an increased number of dilated and thus likely more fragile vessels in the subacute phase after cerebral ischemia. CONCLUSIONS: Our results provide new insights into both BMC-based therapies and immune cell biology and help to understand potential adverse and unexpected side effects.

Effects of Arbutin on Radiation-Induced Micronuclei in Mice Bone Marrow Cells and Its Definite Dose Reduction Factor.

BACKGROUND: Interactions of free radicals from ionizing radiation with DNA can induce DNA damage and lead to mutagenesis and carsinogenesis. With respect to radiation damage to human, it is important to protect humans from side effects induced by ionizing radiation. In the present study, the effects of arbutin were investigated by using the micronucleus test for anti-clastogenic activity, to calculate the ratio of polychromatic erythrocyte to polychromatic erythrocyte plus normochromatic erythrocyte (PCE/PCE+NCE) in order to show cell proliferation activity. METHODS: Arbutin (50, 100, and 200 mg/kg) was intraperitoneally (ip)administered to NMRI mice two hours before gamma radiation at 2 and 4 gray (Gy). The frequency of micronuclei in 1000 PCEs (MnPCEs) and the ratio of PCE/PCE+NCE were calculated for each sample. Data were statistically evaluated using one-way ANOVA, Tukey HSD test, and t-test. RESULTS: The findings indicated that gamma radiation at 2 and 4 Gy extremely increased the frequencies of MnPCE (P<0.001) while reducing PCE/PCE+NCE (P<0.001) compared to the control group. All three doses of arbutin before irradiation significantly reduced the frequencies of MnPCEs and increased the ratio of PCE/PCE+NCE in mice bone marrow compared to the non-drug-treated irradiated control (P<0.001). All three doses of arbutin had no toxicity effect on bone marrow cells. The calculated dose reduction factor (DRF) showed DRF=1.93 for 2Gy and DRF=2.22 for 4 Gy. CONCLUSION: Our results demonstrated that arbutin gives significant protection to rat bone against the clastogenic and cytotoxic effects of gamma irradiation.

Enhanced differentiation of intraepithelial lymphocytes in the intestine of polymeric immunoglobulin receptor-deficient mice.

To clarify the effect of secretory IgA (sIgA) deficiency on gut homeostasis, we examined intraepithelial lymphocytes (IELs) in the small intestine (SI) of polymeric immunoglobulin receptor-deficient (pIgR(-/-) ) mice. The pIgR(-/-) mice exhibited the accumulation of CD8αß(+) T-cell receptor (TCR)-αß(+) IELs (CD8αß(+) αß-IELs) after weaning, but no increase of CD8αß(+) γδ-IELs was detected in pIgR(-/-) TCR-ß(-/-) mice compared with pIgR(+/+) TCR-ß(-/-) mice. When 5-bromo-2'-deoxyuridine (BrdU) was given for 14 days, the proportion of BrdU-labelled cells in SI-IELs was not different between pIgR(+/+) mice and pIgR(-/-) mice. However, the proportion of BrdU-labelled CD8αß(+) -IELs became higher in pIgR(-/-) mice than pIgR(+/+) mice 10 days after discontinuing BrdU-labelling. Intravenously transferred splenic T cells migrated into the intraepithelial compartments of pIgR(+/+) TCR-ß(-/-) mice and pIgR(-/-) TCR-ß(-/-) mice to a similar extent. In contrast, in the case of injection of immature bone marrow cells, CD8αß(+) αß-IELs increased much more in the SI of pIgR(-/-) TCR-ß(-/-) mice than pIgR(+/+) TCR-ß(-/-) mice 8 weeks after the transfer. αß-IELs from pIgR(-/-) mice could produce more interferon-γ and interleukin-17 than those of pIgR(+/+) mice, and intestinal permeability tended to increase in the SI of pIgR(-/-) mice with aging. Taken together, these results indicate that activated CD8αß(+) αß-IELs preferentially accumulate in pIgR(-/-) mice through the enhanced differentiation of immature haematopoietic precursor cells, which may subsequently result in the disruption of epithelial integrity.

Characterization, Quantification, and Determination of the Toxicity of Iron Oxide Nanoparticles to the Bone Marrow Cells.

Iron oxide nanoparticles (IONPs) have been used to develop iron supplements for improving the bioavailability of iron in patients with iron deficiency, which is one of the most serious nutritional deficiencies in the world. Accurate information about the characteristics, concentration, and cytotoxicity of IONPs to the developmental and reproductive cells enables safe use of IONPs in the supplement industry. The objective of this study was to analyze the physicochemical properties and cytotoxicity of IONPs in bone marrow cells. We prepared three different types of iron samples (surface-modified iron oxide nanoparticles (SMNPs), IONPs, and iron citrate) and analyzed their physicochemical properties such as particle size distribution, zeta potential, and morphology. In addition, we examined the cytotoxicity of the IONPs in various kinds of bone marrow cells. We analyzed particle size distribution, zeta potential, iron levels, and subcellular localization of the iron samples in bone marrow cells. Our results showed that the iron samples were not cytotoxic to the bone marrow cells and did not affect the expression of cell surface markers and lipopolysaccharide (LPS)-induced the secretion of cytokines by murine bone marrow-derived dendritic cells (BMDCs). Our results may be used to investigate the interactions between nanoparticles and cells and tissues and the developmental toxicity of nanoparticles.