Bone Marrow Nucleated Cells Research Articles

Transcriptome Profiling Analysis Reveals the Potential Mechanisms of Three Bioactive Ingredients of Fufang E'jiao Jiang During Chemotherapy-Induced Myelosuppression in Mice.

Although multiple bioactive components have been identified in Fufang E’jiao Jiang (FEJ), their hematopoietic effects and molecular mode of action in vivo are still not fully understood. In the current study, we analyzed the effects of martynoside, R-notoginsenoside R2 (R2), and 20S-ginsenoside Rg2 (Rg2) in a 5-fluorouracil-induced myelosuppression mouse model. Bone marrow nucleated cells (BMNCs) counts, hematopoietic progenitor cell colony-forming unit (CFU) assay, as well as flow cytometry analysis of Lin-/c-kit+/Sca-1+ hematopoietic stem cell (HSC) population were conducted, and bone marrow cells were subjected to RNA sequencing. The transcriptome data were processed based on the differentially expressed genes. The results of the analysis show that each of the three compounds stimulates BMNCs and HSC growth, as well as burst-forming unit-erythroid and colony-forming unit granulocyte-monocyte colony expansion. The most relevant transcriptional changes appeared to be involved in regulation of hematopoietic cell lineage, NF-κB and TNF-α signaling, inhibition of inflammation, and acceleration of hematopoietic cell recovery. Notably, the individual compounds shared similar but specified transcriptome profiles. Taken together, the hematopoietic effects for the three tested compounds of FEJ are confirmed in this myelosuppression mouse model. The transcriptome maps of these effects provide valuable information concerning their underlying mechanisms and provide a framework for the continued study of the complex mode of action of FEJ.

Chimerism and gene therapy - Lessons learned from non-conditioned murine bone marrow transplantation models.

Hematopoietic stem and progenitor cells (HSPCs) can be used as a vector for gene therapies. In order to predict the number of HSPCs cells necessary to achieve the target level of chimerism in an autologous setting, syngeneic male bone marrow (BM) cells were transplanted into 35 non-conditioned female BALB/c mice. The resulting chimerism was determined at 6-53weeks using qPCR, cell subpopulation sorting, and colony-forming units (CFU) analysis. After the transplantation of 125.8±2.5million nucleated BM cells, the BM of recipients contained 20.0±2.8% donor cells, representing a chimerism of 0.16±0.02% per one million transplanted nucleated BM cells. Chimerism levels in the BM, neutrophils, and Bcells were comparable, whereas in Tcells it was lower, and in CFU was approximately twice greater than in BM. By extrapolating our murine data, and data from some previous studies to a human non-conditioned autologous CD34+ HSPC transplantation setting, we conclude that approximately 44million CD34+ HSPCs would be needed to achieve 20% donor chimerism in a 70-kg human, which could serve as a starting point for the future use of HSCPs in gene and cell therapy.

Apoptosis induction in K562 human myelogenous leukaemia cells is connected to the modulation of Wnt/β-catenin signalling by BHX, a novel pyrazoline derivative.

The goal of this study was to explore the effects of BHX on human chronic myeloid leukaemia (CML) cells and to elucidate the underlying molecular mechanism. CML cell line K562 cells were treated with BHX. The effects of BHX on cell proliferation, apoptosis and cell cycle were detected. Subsequently, the caspase, ATP activity, Ca2+ , ROS and mitochondrial membrane potential (MMP) levels treated with various concentrations of BHX were analysed. The variation of relevant proteins and genes was detected. Further, toxicity of BHX on peripheral blood cells, bone marrow-nucleated cells (BMNC) and organ index were investigated on mice. Results showed that BHX suppressed K562 cell proliferation in a dose-dependent manner and induced apoptosis and G0/G1 phase arrest. BHX induced mitochondria-mediated apoptosis, which was associated with downregulation of MMP, activation of caspase-3 and caspase-9, generation of intracellular ROS and elevation of Ca2+ in K562 cells. In treated cells, ATP levels were decreased, expression of total β-catenin, phosphorylated β-catenin and β-catenin in the nucleus was decreased, and expression of cell cycle-related proteins was decreased. Further analysis revealed that BHX lowered the transcriptional level of β-catenin. Lastly, BHX treatment significantly reduced the number of white blood cells, but had no effect on BMNC and organ index. These findings provide further insight into the potential use of BHX as an anti-cancer agent against human leukaemia.

Effects of extract of Urtica dioica L. (stinging nettle) on the immune response of rats with severe malnutrition.

Background. Severe malnutrition is the main cause of immunodeficiency in children under five years old in areas of economic depression in developing countries, and it is associated with generalized atrophy of lymphoid tissues. Urtica dioica L. is a medicinal herb with multiple pharmacological functions; it acts as modulator of the immune response, stimulating the proliferation of T lymphocytes in vitro. Objective: To determine the ability of U. dioica L. to modulate the immune response of malnourished rats in the first 21 days of life. Methods: Third-degree malnutrition during lactation was induced by food competition, and treatment with U. dioica L was tested. Day-old Wistar rats were randomly assigned to three groups: a) the well-nourished group, b) the untreated malnourished group and c) the malnourished group treated with 0.2 g/mL U. dioica L. on days 14, 16, 18 and 20. On day 21, animals were euthanized, and the thymus, spleen, bone marrow and blood were obtained. Lymphocyte subpopulations were determined in the thymus and leukocyte subpopulations in blood. The total number of nucleated bone marrow cells was determined, and the proliferative capability of spleen lymphocytes from malnourished rats treated with U. dioica L. was compared to that of other groups. Results: U. dioica L. decreased CD4-CD8- lymphocytes and increased CD4+CD8+ lymphocytes in the thymus, while in the blood, it increased CD4+ cells, monocytes and total T lymphocytes. The proliferation of spleen cells was stimulated in the malnourished rats, and a decrease in total nucleated bone marrow cells was observed. An increase in the red blood cell count, hemoglobin and white blood cell count was observed in malnourished rats treated with Urtica dioica L. (p≤0.05). Conclusion: U. dioica L. had immunomodulatory effects since it promoted the differentiation of CD4-CD8- T lymphocytes towards CD4+CD8+ T lymphocytes and counteracted anemia and leucopenia associated with severe malnutrition. Key words: Severe malnutrition, immunodeficiency, immunomodulatory activity, T lymphocytes, Urtica dioica L.

Impaired Mitophagy of Nucleated Erythroid Cells Leads to Anemia in Patients with Myelodysplastic Syndromes.

Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal stem cell disorders characterized by cytopenia and dysplasia. Anemia is the most common symptom in patients with MDS. Mitophagy and mitochondrial dysfunction might be involved in the development of MDS. In this study, we investigated the change of mitophagy in erythroid precursors in MDS patients. We found that NIX-mediated mitophagy was impaired in bone marrow nucleated red blood cells (NRBC) of MDS patients, associated with an increased amount of damaged mitochondria and increased ROS level which might lead to apoptosis and ineffective erythropoiesis. The results showed that the amount of mitochondria in GlycoA+ NRBC positively correlated with the count of ring sideroblasts in bone marrow samples. Meanwhile, the level of autophagy-associated marker LC3B in GlycoA+ NRBC had a positive correlation with hemoglobin (Hb) levels, and the amount of mitochondria in GlycoA+ NRBC had a negative correlation with Hb levels in high-risk MDS patients. Our results indicated that mitophagy might involve the pathogenesis of anemia associated with MDS. Autophagy might be a novel target in treatments of MDS patients.

Osteogenesis induced by a three-dimensional bioimplant composed of demineralised bone matrix, collagen, hydroxyapatite, and bone marrow-derived cells in massive bone defects: An experimental study

with bioimplants vs. bioimplant plus bone marrow nucleated cells (BMNCs) showed a notorious osteogenic effect, but with greater osteoid synthesis and cellularity in the latter. These results suggest that combined treatment with bioimplants and BMNCs have an additive effect on massive bone defects in lambs. These experimental results could be applied to repair damaged human bone.

Multiple Autologous Bone Marrow-Derived CD271+ Mesenchymal Stem Cell Transplantation Overcomes Drug-Resistant Epilepsy in Children.

There is a need among patients suffering from drug‐resistant epilepsy (DRE) for more efficient and less toxic treatments. The objective of the present study was to assess the safety, feasibility, and potential efficacy of autologous bone marrow cell transplantation in pediatric patients with DRE. Two females and two males (11 months to 6 years) were enrolled and underwent a combined therapy consisting of autologous bone marrow nucleated cells (BMNCs) transplantation (intrathecal: 0.5 × 109; intravenous: 0.38 × 109–1.72 × 109) followed by four rounds of intrathecal bone marrow mesenchymal stem cells (BMMSCs) transplantation (18.5 × 106–40 × 106) every 3 months. The BMMSCs used were a unique population derived from CD271‐positive cells. The neurological evaluation included magnetic resonance imaging, electroencephalography (EEG), and cognitive development assessment. The characteristics of BMMSCs were evaluated. Four intravenous and 20 intrathecal transplantations into the cerebrospinal fluid were performed. There were no adverse events, and the therapy was safe and feasible over 2 years of follow‐up. The therapy resulted in neurological and cognitive improvement in all patients, including a reduction in the number of epileptic seizures (from 10 per day to 1 per week) and an absence of status epilepticus episodes (from 4 per week to 0 per week). The number of discharges on the EEG evaluation was decreased, and cognitive improvement was noted with respect to reactions to light and sound, emotions, and motor function. An analysis of the BMMSCs' characteristics revealed the expression of neurotrophic, proangiogenic, and tissue remodeling factors, and the immunomodulatory potential. Our results demonstrate the safety and feasibility of BMNCs and BMMSCs transplantations and the considerable neurological and cognitive improvement in children with DRE. stem cells translational medicine 2018;7:20–33

Improvement of icaritin on hematopoietic function in cyclophosphamide-induced myelosuppression mice

Context: Icaritin (ICT), an intestinal metabolite of prenylflavonoids from Herba Epimedii, has been known to regulate many immune processes. But there are little studies of ICT on hematopoietic function.Objective: We aimed to investigate the improvement of ICT on hematopoietic function in cyclophosphamide (CTX)-induced myelosuppression mice.Methods: Mice were given CTX (50 mg/kg) by i.p. for five days to produce bone marrow depression model. 48 h after last treated with CTX, ICT was administrated at 10 mg/kg/d by p.o. for five days. Blood routine, body weight, thymus index and spleen index were tested. The bone marrow hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs), cell cycle and apoptosis of HSCs were quantified by flow cytometry. The bone marrow nucleated cells were counted by an automated cell counter. The histology of femoral bone was examined by Haemotoxylin and Eosin (H&E) staining. Serum erythropoietin (EPO), granulocyte colony-stimulating factor (G-CSF) and thyroperoxidase (TPO) were tested by ELISA kit.Results: ICT (10 mg/kg) protected against CTX-induced myelosuppression, is evidenced by increased blood cell numbers, body weight, thymus index, spleen index and improved femoral bone morphology. ICT corrected the reduction of bone marrow HSCs and HPCs, promoted bone marrow HSCs entering the proliferative cycle phase and prevented cells proceeding to the apoptosis phase. Meanwhile, ICT increased the release of G-CSF and TPO in model mice serum.Conclusion: These results demonstrated that ICT improves myelosuppression by improving bone marrow hematopoietic microenvironment, promoting the proliferation and differentiation of HSCs, inhibiting the apoptosis of HSCs and stimulating the expression of G-CSF and TPO.

STEM/PROGENITOR CELLS ADHESION STRENGTHENING TO SYNTHETIC MATERIAL USING EXTRACELLULAR MATRIX

Background: Tissue-engineered vascular grafts of small diameter have being widely used in coronary artery bypass grafting. However, the rate of settling of these grafts with endothelial cells is insufficient. Aims:The aim of the study was to selucidate the adhesion of endothelial progenitor cells and mesenchymal stem cells to synthetic materials (polystyrol, polytetrafluoroethylene) preprocessed with extracellular matrix (gelatin, fibronectin, collagen) in vitro.Materials and methods: For the study,the endothelial progenitor cells (EPC) were isolated from the peripheral blood of patients with ischemic heart disease, and mesenchymal stem cells (MSC) — from bone marrow of Wistar rats. Commercial monoclonal antibodies for flow- cytometry were used to determine the phenotype of EPC. To isolate the early and late EPC from mononuclear cells of peripheral blood, cells were raised for 8 and 16 days on a gelatin or fibronectin based substrate. The commercial kits for enzyme linked immunoassay were applied to assess levels of cytokine production and nitric oxide by early and late EPC conditioned by gelatin or fibronectin on the 8th and 16th days of growth. To conduct the study the MSC were isolated from bone marrow of rats. To determine the attachment of adherent fraction of nucleated bone marrow cells, cell morphology, the adipogenic and osteogenic differentiation were evaluated. To assess migration of MSC in real time within 24 hours on the device Cell-IQ, we used «closure/wound healing» test. The commercial kits for enzyme linked immunoassay were applied to assess levels of cytokine production and nitric oxide by MSC; and components of the extracellular matrix (fibronectin, collagen I and type IV) ― to assess the increased adhesion of MSC to the polytetrafluoroethylene. Results: The results demonstrated that endothelial progenitor cells adhere to both gelatin and fibronectin and confirmed the influence of these extracellular matrix components on the cytokine levels produced by early and late endothelial cells. The combination of fibronectin with type I or IV collagen or the combination of thereof promotes the adhesion to polytetrafluoroethylene and colonization of the graft.Conclusions: Preprocessing of synthetic material (polystyrene, polytetrafluoroethylene) enhances adhesion and growth of EPC and MSC which can be implemented when creating tissue-engineered vascular grafts for small diameter coronary artery bypass grafting with specified conditions of settlement by the cells involved in neointima formation.

Effect of ERK1/2 Signaling Pathway Inhibitor PD98059 on the Expression of Ras, BRaf, MEK, ERK1/2 in Marrow Nucleated Red Blood Cells of CMS Patients

To investigate the effect of ERK1 / 2 signaling pathway inhibitor PD98059 on Ras, Raf, MEK, ERK1, ERK2 expression in order to explore a new way for basic research and clinical treatment of the chronic mountain sickness(CMS). Sixteen CMS patients were selected, the bone marrow was collected for isolation of monomuclear cells (MNC), the cells were sorted by using CD71 and CD235a antibody magnetic beads, then positive cells were diveded into 5 groups: blank control, DMSO and PD98059 5, 10 and 20 µmol/L, and were cultured in hypoxid condition for 72 hours. The Ras-GTP levels in supernatant was detected by ELISA, the RT-PCR was used to determine the expression of BRaf, MEK, ERK1, ERK2 mRNA in nucleated red blood cells, and the Western blot method was used to detect expression of BRaf, MEK, ERK1, ERK2 protein. PD98059 had no effect on the level of Ras-GTP in each groups. Compared with the blank control group, the expression levels of BRaf, MEK mRNA in DMSO group were not statistically significant (P values were 0.826, 0.298). Compared with the PD98059 20 mol/L group, the expression level of ERK1/2 mRNA was statistically significant (P=0.001, 0.002). Compared with the blank control group, expression levels of p-BRaf, p-MEK protein in DMSO group were not statistically significant (P=0.370, 0.351). Compared with the PD98059 20 mol/L group, the difference of p-ERK1/2 protein level in other 4 groups were statistically significant (P values were <0.001, 0.007). PD98059 can up-regulate the expressions of ERK1/2 miRNA and p-ERK1/2 protein in bone marrow nucleated red blood cells, the Ras / Raf / MEK / ERK 1/2 pathway is the main signal transduction pathway in regulating bone marrow nucleated red blood cells, suggesting that Ras/Raf /MEK /ERK 1/2 pathway may be involved in the pathogenesis of chronic mountain sickness process.

The value of bone marrow imprint in diagnosis of plasma cell myeloma and monitoring the remission after chemotherapy

Objective To investigate the value of bone marrow imprint in the diagnosis of plasma cell myeloma and the remission rate after chemotherapy. Methods Bone marrow aspiration, bone marrow imprint, and bone marrow biopsy of plasma cell myeloma with 128 patients were collected. The bone marrow biopsy was used as the standard. Bone marrow imprint was compared to bone marrow aspiration in bone marrow nucleated cells quantity, infiltration degree, and diagnostic coincidence rate. Sensitivity, specificity, positive predictive value, and Youden index were evaluated. Results The results of bone marrow imprint showed that the number of nucleated cells was better than that of bone marrow aspiration (P 0.05). Bone marrow biopsy combined with CD38 (+ ) showed the degree of infiltration of bone marrow as the standard, and the bone marrow imprint was better than that of bone marrow aspiration (P 0.05). To evaluate the compliance rate of bone marrow infiltration in bone marrow imprint: In phase I, the compliance rate of bone marrow aspiration was 95.83% (23/24) and bone marrow imprint of 91.67% (22/24), the difference was not statistically significant (P>0.05); In phase II, bone marrow imprint coincidence rate was 92.86% (65/70), which was significantly higher than that of bone marrow aspiration in 65.71% (46/70) (P<0.05); In phase III, bone marrow imprint coincidence rate was 79.41% (27/34), which was significantly higher than that of bone marrow aspiration in 44.12% (15/34) (P<0.05). The other parameters, such as sensitivity, specificity, positive predictive value, and Youden index were better than those of bone marrow aspiration. Conclusions Bone marrow imprint has characteristics that both bone marrow aspiration and bone marrow biopsy have. It has obvious advantages in diagnosis of plasma cell myeloma and monitoring the remission after chemotherapy. Key words: Bone marrow examination; Biopsy; Microtomy; Multiple myeloma/DI/DT

CRISPR/Cas9-Mediated Deletion of Foxn1 in NOD/SCID/IL2rg\u2212/\u2212 Mice Results in Severe Immunodeficiency

Immunodeficient mice engrafted with either normal or cancerous human cells are widely used in basic and translational research. In particular, NOD/SCID/IL2rg−/− mice can support the growth of various types of human cancer cells. However, the hairs of these mice interfere with the observation and imaging of engrafted tissues. Therefore, novel hairless strains exhibiting comparable immunodeficiency would be beneficial. Recently, the CRISPR/Cas9 system has been used for efficient multiplexed genome editing. In the present study, we generated a novel strain of nude NOD/SCID/IL2rg−/− (NSIN) mice by knocking out Foxn1 from NOD/SCID/IL2rg−/− (NSI) mice using the CRISPR/Cas9 system. The NSIN mice were deficient in B, T, and NK cells and not only showed impaired T cell reconstitution and thymus regeneration after allogeneic bone marrow nucleated cell transplantation but also exhibited improved capacity to graft both leukemic and solid tumor cells compared with NSI, NOG, and NDG mice. Moreover, the NSIN mice facilitated the monitoring and in vivo imaging of both leukemia and solid tumors. Therefore, our NSIN mice provide a new platform for xenograft mouse models in basic and translational research.

Telomere length and somatic mutations in correlation with response to immunosuppressive treatment in aplastic anaemia

We investigated the frequencies of cytogenetic aberrations and somatic mutations of prognostic relevance in 393 patients with aplastic anaemia (AA). Clonality was determined by G-banding/fluorescence insitu hybridization (FISH) (n=245), and targeted capture sequencing was performed for 88 haematopoiesis-related genes (n=70). The telomere length (TL) of bone marrow nucleated cells was measured at the single cell level by FISH (n=135). Eighteen (4·6%) patients showed disease progression, and monosomy 7 (50·0%) was the most predominant cytogenetic evolution at disease transformation. One third of patients (32·9%) presented at least 1 mutation; the most frequently mutated genes were NOTCH1, NF1, SCRIB, BCOR and DNMT3A. The patient group with clonal changes (30·7%) showed an adverse response to immunosuppressive treatment (IST), compared to the non-clonal group, but this finding did not show statistical significance. The TL of AA patients was significantly shorter than normal control and patients with clonal changes showed significantly shorter TLs. Patients with TL>5·9 showed a higher response rate to IST (P=0·048). In conclusion, the patients with clonal changes or TL attrition showed a poor response to IST. Shorter TL can be used not only as a biomarker, but also as a predictive marker for treatment response to IST.

Black Phosphorus Quantum Dot Induced Oxidative Stress and Toxicity in Living Cells and Mice.

Black phosphorus (BP), as an emerging successor to layered two-dimensional materials, has attracted extensive interest in cancer therapy. Toxicological studies on BP are of great importance for potential biomedical applications, yet not systemically explored. Herein, toxicity and oxidative stress of BP quantum dots (BPQDs) at cellular, tissue, and whole-body levels are evaluated by performing the systemic in vivo and in vitro experiments. In vitro investigations show that BPQDs at high concentration (200 μg/mL) exhibit significant apoptotic effects on HeLa cells. In vivo investigations indicate that oxidative stress, including lipid peroxidation, reduction of catalase activity, DNA breaks, and bone marrow nucleated cells (BMNC) damage, can be induced by BPQDs transiently but recovered gradually to healthy levels. No apparent pathological damages are observed in all organs, especially in the spleen and kidneys, during the 30-day period. This work clearly shows that BPQDs can cause acute toxicities by oxidative stress responses, but the inflammatory reactions can be recovered gradually with time for up to 30 days. Thus, BPQDs do not give rise to long-term appreciable toxicological responses.

G-CSF Indirectly Induces Apoptosis of Osteoblasts During Hematopoietic Stem Cell Mobilization.

The objective of this study was to explore the mechanism underlying osteoblast suppression in the process of hematopoietic stem cells mobilization induced by granulocyte colony‐stimulating factor (G‐CSF). The apoptosis of human and mouse osteoblasts was examined by detecting caspase 3. The levels of serum DKK1 and osteocalcin in the supernatant of co‐culture of mouse osteoblasts and mouse bone marrow nucleated cells were measured. The number of mouse osteoblasts co‐cultured with mouse bone marrow nucleated cells was measured and the osteocalcin mRNA level was also measured. The G‐CSF‐induced decrease in osteoblast function was partly due to the apoptosis of osteoblasts. There was no significant difference in the level of serum DKK1 in healthy donors before and 5 days after mobilization. The osteocalcin gene and protein expression was significantly different in co‐cultured osteoblasts with bone marrow nucleated cells treated with and without G‐CSF. Osteoblasts undergo apoptosis during mobilization and G‐CSF affects osteoblasts through bone marrow nucleated cells.

Hepatoprotective Effect of Carboxymethyl Pachyman in Fluorouracil-Treated CT26-Bearing Mice.

5-Fluorouracil (5-FU) is the chemotherapeutic agent of first choice for the treatment ofcolorectal cancer, however, treatment-related liver toxicity remains a major concern. Thereby, it is desirable to search for novel therapeutic approaches that can effectively enhance curative effects and reduce the toxic side effects of 5-FU. Carboxymethyl Pachyman (CMP) exhibits strong antitumor properties, but the antitumor and hepatoprotective effects of CMP and the molecular mechanisms behind these activities, are however poorly explored. Thereby, the purpose of the present study was to evaluate the hepatoprotective effect of CMP in 5-FU-treated CT26-bearing mice, and further explore the underlying mechanism(s) of action. Initially, a CT26 colon carcinoma xenograft mice model was established. The immune organ indexes, blood indicators, liver tissue injury, and indicators associated with inflammation, antioxidant and apoptosis were then measured. Our results showed that CMP administration increased the tumor inhibitory rates of 5-FU and, meanwhile, it reversed reduction of peripheral white blood cells (WBC) and bone marrow nucleated cells (BMNC), increase of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), and decrease of superoxide dismutase (SOD), catalase (CAT), GSH-Px and glutathione(GSH) induced by 5-FU. Moreover, CMP in combination with 5-FU alleviated severe liver injury induced by 5-FU via reducing the levels of ROS, IL-1β, and IL-6, decreasing expression of p-IκB-α, NF-κB, p-NF-κB, pp38 and Bax, and elevating levels of Nrf2, GCL, HO-1 and Bcl-2. Collectively, these outcomes suggested that CMP effectively enhanced the curative effects of 5-FU and simultaneously reduced the liver injuries induced by 5-FU in CT26-bearing mice, and the mechanism may be associated with regulation of NF-κB, Nrf2-ARE and MAPK/P38/JNK pathways.

Study on autophagy in nucleated red blood cells in patients with myelodysplastic syndromes

目的研究骨髓增生异常综合征（MDS）患者有核红细胞的自噬水平。方法54例MDS患者为实验组，对照组33例。采用透射电镜观察自噬过程，流式细胞术检测骨髓细胞中GlycoA+有核红细胞自噬相关蛋白微管相关蛋白轻链3B（LC3B）的表达水平，Real-time PCR法检测GlycoA+有核红细胞自噬相关基因AMPK、ULK1和mTOR mRNA的表达，Western blot法检测GlycoA+有核红细胞线粒体外膜蛋白TOM20的表达。结果MDS患者有核红细胞中不易观察到自噬体及自噬溶酶体。相对高危组MDS患者有核红细胞LC3B表达（0.22±0.12）显著低于对照组（0.43±0.22，P<0.001）及相对低危组（0.40±0.16，P=0.001）。相对高危组患者AMPK、ULK1 mRNA表达水平明显低于对照组[AMPK：0.26（0.60）对1.00（2.07），ULK1：0.27（3.31）对1.07（4.41），P值均<0.017]，mTOR mRNA表达水平明显高于对照组[1.82（3.74）对1.26（1.38），P<0.017]。相对高危组MDS患者有核红细胞LC3B水平与HGB呈正相关（r=0.529，P=0.009）。相对高危组MDS患者有核红细胞TOM20蛋白（9.42±4.42）表达升高。结论高危MDS患者有核红细胞自噬水平下降。

Radioprotective effect of ultra-small carbon quantum dots in mice in vivo

Objective To study the radioprotective effects of ultra-small carbon quantum dots (CQDs) in mice in vivo, and to reveal the protective mechanism, as well as to study the body immune response to CQDs and the toxicity in vivo. Methods Mice were injected with different concentrations of CQDs solution. Mice models of systemic radiation injury were constructed by high dose gamma rays radiation. The 30-days survival rate, bone marrow DNA, femoral bone marrow nucleated cells (BMNC), tissue superoxide dismutase (SOD) and malondialdehyde (MDA) were measured to evaluate the radioprotective effects of CQDs, and to investigate the possible protective mechanism. The toxicity of CQDs in vivo was studied by measuring the changes of body weight, liver index and spleen index before and after injections of CQDs in mice. Results CQDs showed obvious radioprotective effects on mice in vivo. Compared with the control group, the 30-days survival rate of irradiated mice treated with CQDs increased from 0% to 40%. CQDs could effectively reduce the hematopoietic system damages caused by radiation, and increase the level of bone marrow DNA, femur BMNC, liver and lung SOD, as well as reduce the production of MDA in liver and lung. The results of immunological reaction tests showed that CQDs had less toxicity in vivo and did not trigger the body immune response. Conclusions CQDs has tremendous application prospects in the field of radiation protection. This study can provide new ideas for the application of new nano-materials in medical field. Key words: Carbon quantum dots; Radiation protection; In vivo toxicity

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

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. 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. 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.

NIX-mediated mitochondrial autophagy in pathogenesis of myelodysplastic syndrome anemia

Objective: To investigate the change of NIX level of bone marrow nucleated red blood cells in anemia patients with myelodysplastic syndromes (MDS), to explore the significance of NIX-mediated mitochondrial autophagy in the pathogenesis of MDS anemia. Methods: A total of 54 patients with MDS diagnosed in the Department of Hematology of General Hospital, Tianjin Medical University from July 2015 to July 2016 were enrolled into the MDS group, 33 cases of immune thrombocytopenia or idiopathic leukopenia as controls.The level of NIX, the number of mitochondria, mitochondrial membrane potential, the level of reactive oxygen species (ROS) in GlycoA(+) nucleated red blood cells were measured by flow cytometry; the level of NIX mRNA was measured by PCR. Results: (1) The expression of NIX in GlycoA(+) nucleated red blood cells in high-risk MDS patients (0.61±0.24) was significantly lower than that in controls (0.79±0.16, P=0.027), and lower than that in low-risk MDS patients (0.81±0.15, P=0.011), while there was no significant difference between the controls and low-risk MDS patients. The expression of NIX mRNA in GlycoA(+) nucleated red blood cells in high-risk MDS group (0.36±0.09) was lower than that in the controls (1.44±0.41, P=0.027) and that in the low-risk group (1.02±0.22, P=0.012); there was no significant difference between the controls and the low-risk group. (2) The number of mitochondria in GlycoA(+) nucleated red blood cells in high-risk MDS patients (937.17±707.85) was significantly higher than that in the controls (513.49±372.33, P=0.019) and that in low-risk MDS patients (461.74±438.02, P=0.008); while there was no significant difference between low-risk MDS patients and the controls. (3) The level of mitochondrial membrane potential in GlycoA(+) nucleated red blood cells in high-risk MDS patients (0.33±0.18) was significantly lower than that in the controls (0.61±0.32, P=0.001) and that in low-risk MDS patients (0.61±0.34, P=0.001); with no significant difference between low-risk MDS patients and the controls. (4)The level of ROS in GlycoA(+) nucleated red blood cells in high-risk MDS patients (438.65±322.83) was significantly higher than that in the controls (242.77±136.87, P=0.006), and higher than that in low-risk MDS patients (197.40±95.07, P=0.001); no significantly different between low-risk MDS patients and the controls. (5) The number of mitochondria in GlycoA(+) nucleated red blood cell was positively correlated with the percentage of ring sideroblast (r=0.457, P=0.028) in the MDS patients.(6) The number of mitochondria in GlycoA(+) nucleated red blood cells was negatively correlated with the concentration of hemoglobin (r=-0.521, P=0.009) in high-risk MDS patients, but not correlated with the concentration of hemoglobin in low-risk MDS patients. Conclusion: NIX level is reduced in nucleated red blood cells of high-risk MDS patients, which leads to impaired mitochondrial autophagy, increased damaged mitochondria and apoptosis of nucleated red blood cells, thus related with anemia.