ObjectiveTo investigate activation of phosphatidylinositol 3 hydroxykinase (PI3K)/AKT pathway and sphingosine 1-phosphate receptor 2 (S1PR2) in peripheral blood mononuclear cells (PBMCs) of acute obstructive cholangitis (AOC) rats and their effects on systemic inflammation in rats.Methods① In vitro experiment: The isolated PBMCs from the rats were divided into 4 groups: a control group, LY294002 treatment group, lipopolysaccharide (LPS) treatment group, and LPS+LY294002 treatment group. The levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the supernatant were detected and the phosphorylation levels of PI3K and AKT and protein level of S1PR2 in the PBMCs were detected. ② In vivo experiment: The rats were randomly divided into four groups: a control group, LY294002 treatment group, AOC model group, and AOC+LY294002 treatment group. The survival rate of rats was recorded, the liver function (ALT, AST, and TBIL), TNF-α, and IL-6 levels in the serum were detected. The phosphorylation levels of PI3K and AKT and protein level of S1PR2 in the PBMCs of the rats were detected. Results① The results of in vitro experiment: The levels of TNF-α and IL-6 in the LPS+LY294002 treatment group were significantly lower than those in the LPS treatment group (P<0.050). The phosphorylation levels of PI3K and AKT and protein level of S1PR2 in the LPS+LY294002 treatment group were significantly lower than those in the LPS treatment group (P<0.050). ② The results of in vivo experiment: The survival rate of rats in the AOC+LY294002 treatment group was higher than those in the AOC group. The serum levels of ALT, AST, TBIL, TNF-α, and IL-6 in the AOC+LY294002 treatment group were significantly lower than those in the AOC model group (P<0.050). The phosphorylation levels of PI3K and AKT and protein level of S1PR2 in the AOC+LY294002 treatment group were significantly lower than those in the AOC model group (P<0.050).ConclusionInhibition of activation of PI3K/AKT pathway in PBMCs can inhibit expression of S1PR2, then alleviate systemic inflammatory response induced by AOC in rats.
ObjectiveTo assess the long-term effectiveness and safety of autologous bone marrow mononuclear cells (BM-MNC) transplantation in the treatment of critical diabetic lower arteriosclerosis obliterans (ASO). MethodsBetween January 2007 and January 2010, 61 patients with critical diabetic lower ASO were treated with standard medical therapies in 29 cases (control group) or with standard medical therapies and autologous BM-MNC transplantation in 32 cases (treatment group). There was no significant difference in gender, age, disease duration, Fontatine stage, glucose (GLU), triglyceride (TG), total cholesterol (CHOL), low-density lipoprotein-cholesterol (LDL-C), hemoglobin A1c (HbA1c), systolic blood pressure (SBP), and diastolic blood pressure (DBP) between 2 groups (P>0.05). The endpoints were overall survival (OS) and amputation-free survival (AFS). The risk indexes for ASO were observed and compared between 2 groups before and after treatments. ResultsThe patients were followed up 2-36 months, and no malignant tumor occurred. The OS rate, OS time, AFS rate, and AFS time were 82.76% (24/29), (32.31±9.08) months, 37.50% (9/24), and (21.28±13.35) months in the control group and were 78.13% (25/32), (32.47±6.96) months, 68.00% (17/25), and (28.38±9.48) months in the treatment group;all indexes showed no significant differences (P>0.05). OS rate, OS time, AFS rate, and AFS time showed no significant differences between 2 groups at the other time (P>0.05) except AFS time at 1 year, which was significantly short in the control group than the treatment group (t=2.806, P=0.007). At the endpoint of follow-up, the indexes of GLU, TG, CHOL, LDL-C, HbA1c, SBP, and DBP showed no significant differences between before and after treatments and between 2 groups (P>0.05) in 49 survival patients (24 in control group and 25 in treatment group). ConclusionAutologous BM-MNC transplantation is safe and effective in the treatment of critical diabetic lower ASO, which can significantly improve AFS rate and prolong AFS time with no risks.
Abstract: Objective To observe the changes in morphology, structure, and ventricular function of infarct heart after bone marrow mononuclear cells (BMMNC) implantation. Methods Twenty-four dogs were divided into four groups with random number table, acute myocardial infarction (AM I) control group , AM I-BMMNC group , old myocardial infarct ion (OMI) control group and OM I-BMMNC group , 6 dogs each group. Autologous BMMNC were injected into infarct and peri-infarct myocardium fo r transplantation in AM I-BMMNC group and OM I-BMMNC group. The same volume of no-cells phosphate buffered solution (PBS) was injected into the myocardium in AM Icontrol group and OM I-control group. Before and at six weeks of cell t ransplantation, ult rasonic cardiography (UCG) were performed to observe the change of heart morphology and function, then the heart was harvested for morphological and histological study. Results U CG showed that left ventricular end diastolic dimension (LV EDD) , left ventricular end diastolic volume (LVEDV ) , the thickness of left ventricular postwall (LVPW ) in AM I-BMMNC group were significantly less than those in AM I-control group (32. 5±5. 1mm vs. 36. 6±3. 4mm , 46. 7±12. 1m l vs. 57. 5±10. 1m l, 6. 2±0. 6mm vs. 6. 9±0. 9mm; P lt; 0. 05). LVEDD, LVEDV , LVPW in OM I-BMMNC group were significantly less than those in OM I-control group (32. 8±4. 2 mm vs. 36. 8±4. 4mm , 48. 2±12. 9m l vs. 60.6±16.5m l, 7. 0±0. 4mm vs. 7. 3±0. 5mm; P lt; 0. 05). The value of eject fraction (EF) in OM I-BMMNC group were significantly higher than that in OM I-control group (53. 3% ±10. 3% vs. 44. 7%±10. 1% ). Compared with their control group in morphological measurement, the increase of infarct region thickness (7. 0 ± 1. 9mm vs. 5. 0 ±2.0mm , 6.0±0. 6mm vs. 4. 0±0. 5mm; P lt; 0. 05) and the reduction of infarct region length (25. 5±5. 2mm vs. 32. 1±612mm , 33. 6±5. 5mm vs. 39. 0±3. 2mm , P lt; 0. 05) were observed after transplantation in AM I-BMMNC group and OM I-BMMNC group, no ventricular aneurysm was found in AM I-BMMNC group, and the ratio between long axis and minor axis circumference of left ventricle increased in OM I-BMMNC group (0. 581±0. 013 vs. 0. 566±0.015; P lt; 0. 05). Both in AM I-BMMNC group and OM I-BMMNC group, fluorescence expressed in transplantation region was observed, the morphology of most nuclei with fluorescencew as irregular, and the differentiated cardiocyte with fluorescence was not found in myocardium after transplantation. The histological examination showed more neovascularization after transp lantation both in AMI and in OM I, and significant lymphocyte infiltration in AM I-BMMNC group. Conclusion BMMNC implantation into infarct myocardium both in AMI and OMI have a beneficial effect, which can attenuate deleterious ventricular remodeling in morphology and st ructure, and improve neovascularization in histology, and improve the heart function.
Objective To investigate the effect of tumor associated glycoprotein-72 （TAG-72） redirected T lymphocytes on breast cancer cells. Methods Peripheral blood mononuclear cells （PBMCs）were isolated from healthy volunteers. The recombinant vector anti-TAG-72-scFv-CD3ζ-pcDNA 3.0 were transfected into PBMCs by lipofectamineTM2000 （transfection group）， PBMCs transfected with plasmid pcDNA 3.0 as control group. MCF-7 and Bcap37 cells were cocultivated with PBMCs of transfection group and control group， respectively， and antitumor response of G1 block was observed. Results G1 block rate of MCF-7 cells in transfection group was （82.3±6.9）%， which was significantly higher than that in control group 〔（43.4±3.9）%， P＜0.05〕. G1 block rate of Bcap37 cells in transfection group was （51.3±4.7）%， and not differed from that in control group 〔（45.6±2.5）%， P＞0.05〕. Conclusion TAG-72 redirected T lymphocytes can inhibit the cell proliferation of TAG-72 positive breast cancer cells， and it may provide valuable tools for the cellular immunotherapy.
Objective To study the expression of NLRP3 inflammasome and its downstream inflammatory factors in patients with chronic obstructive pulmonary disease (COPD) and healthy controls, and to reveal the effect and significance of NLRP3 inflammasome in the pathogenesis of COPD. Methods Forty patients with acute exacerbation COPD (AECOPD) who were hospitalized from November 2016 to May 2017 were recruited in the AECOPD group, and recruited in the stable COPD group when they entered the stable stage. Forty healthy individuals were recruited in the control group. General information and peripheral blood were collected from each subject. The levels of NLRP3 mRNA and caspase-1 mRNA in peripheral blood mononuclear cells were measured by real-time PCR. The levels of IL-18 and IL-1β were measured by enzyme-linked immunosorbent assay. Results The levels of NLRP3 mRNA, IL-18 and IL-1β in the AECOPD patients were significantly higher than those in the stable COPD group [2.11±0.77, 12.79 (7.10, 43.13) pg/ml, 17.02 (8.36, 52.21) pg/ml vs. 1.60±0.44, 10.66 (6.32, 18.59) pg/ml, 13.34 (7.07, 16.89) pg/ml, all P<0.05] . The levels of NLRP3 mRNA, IL-18 and IL-1β in the AECOPD patients were significantly higher than those in the control group [2.11±0.77, 12.79 (7.10, 43.13) pg/ml, 17.02 (8.36, 52.21) pg/mlvs. 1.00±0.49, 6.29 (4.73, 7.93) pg/ml, 5.93 (4.81, 9.67) pg/ml, all P<0.05]. The levels of NLRP3 mRNA, IL-18 and IL-1β were significantly higher in the stable COPD group than the control group [1.60±0.44, 10.66 (6.32, 18.59) pg/ml, 13.34 (7.07, 16.89) pg/mlvs. (1.00±0.49, 6.29 (4.73, 7.93) pg/ml, 5.93 (4.81, 9.67) pg/ml, all P<0.05]. Correlation analysis showed that the plasma IL-18 level was positive correlated with leukocyte count and neutrophil percentage in the AECOPD group (r=0.372, P<0.05;r=0.386, P<0.05). The expression of NLRP3 mRNA in the AECOPD group and stable COPD group were positively correlated with the CAT score (r=0.387, P<0.05;r=0.399, P<0.05) . Conclusion NLRP3 inflammasome is involved in the inflammatory response in COPD patients.
The bone marrow mononuclear cell（BMMNC） subset comprises mesenchymal stem cells, hematopoietic stem cells, endothelial progenitor cells. These cells can differentiate into cardiomyocytes, vascular endothelial cells and smooth muscle cells, and they can also release a wide array of cytokines that exert their effects on surrounding cells, including inducing neovascularization, preventing apoptosis of home cells and homing of endogenous systemic repairing cells. Many trials have been developed to evaluate the effect of bone marrow mononuclear cell transplantation in treating ischemia heart diseases in this country and others. Several routes have been used to deliver these cells to human myocardium or to the coronary circulation in these trials, such as intracoronary injection, intravenous infusion, direct injection into the ventricular wall, or transepicardial/transendocardial infusions，and the cells are constructed into fragmented cell sheets to improve cell retention, or some cytokines are used to enhance therapeutic effect. Although the results of the recent clinical trials in this area are rather conflicting, these therapeutic approaches seem to be promising forthe treatment of ischemic heart disease. In this review, many aspects of bone marrow mononuclear cell transplantation in myocardial infarction are summarized such as the mechanism, delivery routes, retaining of cells, homing, survival and future development, etc.
Objective To investigate the efficacy of autologous bone marrow mononuclear cells transplantation in treating lower l imb thromboangiitis obl iterans (TAO). Methods From January 2005 to November 2008, 25 patients (27 l imbs) with lower l imb TAO were treated. There were 24 males (26 l imbs) and 1 female (1 l imb), aging 16-44 years (33 years on average). Fifteen left l imbs and 12 right l imbs were involved. The median duration of disease was 2 years (from 3 months to9 years). Intermittent claudication was observed in 5 cases (5 l imbs), 16 patients (17 l imbs) had symptom of rest pain, 4 patients (5 l imbs) suffered ulcer on the distal l imbs. The results of visual analogue scale (VAS), maximum walking distance (MWD), ankle/brachial index (ABI), and transcutaneous oxygen pressure (TcPO2) before operation were (7.16 ± 1.12) points, (0.098 ± 0.043) km, 0.20 ± 0.09, and (11.78 ± 3.46) mm Hg (1 mm Hg=0.133 kPa), respectively. A total of 300 mL bone-marrow blood was extracted from the il iac bone. And then the mononuclear cells were isolated from the bone-marrow blood. All patients received cell transplantation only one time. The amount of transplantation bone marrow mononuclear cells was (1.82-29.46) × 109 (mean 13.33 × 109). Results All patients were followed up for 1 years. After 4 weeks of implantation, the results of VAS, MWD, ABI, and TcPO2 were (2.39 ± 0.51) points, (0.783 ± 0.176) km, 0.28 ± 0.16, (21.33 ± 6.57) mm Hg, respectively, showing significant difference compared with preoperative results (P lt; 0.05). The VAS, MWD, ABI, and TcPO2 increased to (2.44 ± 0.67) points, (1.199 ± 0.304) km, 0.37 ± 0.09, (27.90 ± 5.23) mm Hg after 1 year of implantation, showing significant differences compared with preoperative results (P lt; 0.05). One ulcer healed well and the improvement was obtained in other 3 cases after 4 weeks of implantation (80%). Four ulcers healed well after 1 year of implantation (80%). After 1 year of implantation, angiography revealed 37.04% affected limbs had a satisfactory neovascularization. The angiographic levels were grade 0 in 5 cases, grade 1 in 12 cases, grade 2 in 4 cases, and grade 3 in 6 cases. Conclusion Autologous bone marrow mononuclear cells transplantation could be a simple, safe, effective method to treat TAO.
【摘要】 目的 了解乙型肝炎病毒（HBV）X基因在HBV相关肝病肝移植受体术后外周单个核细胞（PBMC）和骨髓CD34+细胞内的整合情况及其对乙肝疫苗接种的影响。 方法 采集1999年6月-2005年11月25例HBV相关肝病肝移植受体的外周静脉血及其中23例的骨髓血，以密度梯度离心结合单克隆免疫磁珠分离法获取外周血单个核细胞及骨髓CD34+细胞后，提取细胞DNA。因HBV X基因的整合频率最高，设计HBV X基因的特异引物，进行HBV-Alu-PCR，终产物进行电泳并回收、连接载体、筛选扩增后测序，检测有无X基因整合。 结果 经PCR后电泳及测序分析，25例HBV相关肝病肝移植受体术后的PBMC内未检测出HBV X基因的整合，其中采集到骨髓标本的23例CD34+细胞中亦未检测到HBV X基因的整合。 结论 肝移植术后受体体内HBV微生态的剧烈改变，使HBV整合的基本条件丧失，在此情况下，外周免疫细胞及骨髓造血干/祖细胞不是发生HBV整合的适宜场所，乙肝疫苗接种效果与HBV X基因整合关系不明确。【Abstract】 Objective To investigate whether hepatitis B virus HBV X gene integrates in peripheral blood mononuclear cell (PBMC) and bone marrow CD34+ cells from HBV related liver disease recipients after liver transplantation. Methods Between June 1999 and November 2005, PBMC were obtained from 25 HBV related liver disease recipients after liver transplantation and bone marrow CD34+ cells obtained from 23 cases among them. The cellular DNA was extracted by DNA isolation and purification kit following the manufacture’s instructions. Specific primers to HBV X gene and to human Alu repeats were used to amplify the virus integration through a 3-round hemi-nest PCR. The PCR final product was judged by 1.2% agarose electrophoresis, ligated to T vector, proliferated in E. coil 5α and sequenced. Results According to agarose electrophoresis and sequencing analysis, there were no HBV X gene integration in PBMC and bone marrow CD34+ cells from HBV related liver transplant recipients after surgery. Conclusions Because of the radical change of HBV microecological environment in HBV related liver transplant recipients after operation, the fundamental condition of HBV integration has been lost, which led PBMC and bone marrow CD34+ cells not suit to HBV X gene integrate to human genome. And the impact of HBV X gene integration on HBV vaccination is still undefined.
Objective To investigate the effectiveness of autologous bone marrow mononuclear cells transplantation on lower l imb chronic venous ulcer. Methods Between May 2009 and September 2010, 17 patients with lower l imb chronic venous ulcer were treated with autologous bone marrow mononuclear cells transplantation (transplantation group) and 10patients treated without cells transplantation served as control group. In the transplantation group, there were 9 males and 8 females with age of (33.3 ± 6.1) years, including 11 cases of simple great saphenous vein varicosity and 6 cases of chronic venous insufficiency; the area of ulcer was (4.39 ± 2.46) cm2; and the duration of ulcer ranged from 3 months to 6 years. In the control group, there were 4 males and 6 females with age of (39.2 ± 10.3) years, including 7 cases of simple great saphenous vein varicosity and 3 cases of chronic venous insufficiency; and the area of ulcer was (5.51 ± 2.63) cm2; and the duration of ulcer ranged from 3 months to 2 years. All patients in both groups were classified as C6 according to Cl inical Etiology Anatomy Pathophysiology (CEAP) classification. No signficant difference was found in the general data between 2 groups (P gt; 0.05). The heal ing process of ulcer was observed. The granulation tissue was harvested for HE staining before operation and at 3 days after operation in the transplantation group. The microvessel density (MVD) and vascular endothel ial growth factor (VEGF) expression of ulcer granulation tissue were observed. Results In the transplantation group, ulcer heal ing was accelerated; complete heal ing was observed in 15 cases, partial heal ing in 1 case, and no heal ing in 1 case with the median heal ing time of 22 days. However, in the control group, the heal ing process was slower; complete heal ing of ulcer was observed in 7 cases and no heal ing in 3 cases with the median heal ing time of 57.5 days. There was significant difference in the heal ing time between 2 groups (Z=0.001 4, P=0.002 7). HE staining showed a great number of microvessels in the granulation tissue in the transplantation group. The immunohistochemical staining showed that MVD was significantly increased (t=3.120, P=0.008) after cell ransplantation (32.1 ± 12.8) when compared with that before transplantation (22.1 ± 6.7). The VEGF expressionafter transplantation (8.05% ± 5.10%) was increased sl ightly when compared with that before transplantation (6.13% ±4.20%), but the difference was not significant (t=1.150, P=0.268). Conclusion Autologous bone marrow mononuclear cellstransplantation can stimulate granulation tissue growth and improve ulcer heal ing.
【摘要】 目的 检测B细胞成熟抗原（BCMA）mRNA在系统性红斑狼疮（SLE）患者外周血单个核细胞（PBMC）的表达水平，探讨BCMA在SLE发病中的意义。 方法 纳入2006年1-11月收治的36例SLE患者，同期17例健康志愿者作为对照组，采用半定量RT-PCR法检测外周血单个核细胞中BCMA mRNA的表达，并与SLE疾病活动指数（SLEDAI）进行相关性分析。 结果 SLE患者组BCMA mRNA表达水平（0.598±0.230）均明显高于正常对照组（0.411±0.309）（Plt;0.05）。SLE患者BCMA mRNA表达水平与SLEDAI评分无相关性（P=0.590）。 结论 SLE患者BCMA mRNA表达水平的增高，可能在SLE的发病机制中具有一定的作用。【Abstract】 Objective To detect the mRNA expression of B-cell maturation antigen (BCMA) in peripheral blood mononuclear cells (PBMC) in patients with systemic lupus erythematosus (SLE), and explore the role of BCMA in the pathogenesis of SLE. Methods From January 2006 to November 2006 the expression of BCMA mRNA in PBMC of 36 patients with SLE and 17 normal controls were measured by half-quantitative RT-PCR. The linear correlation between the expression of BCMA mRNA and SLE disease activity index (SLEDAI) was assessed. Results The level of BCMA mRNA (0.598±0.230) in PBMC significantly increased in SLE patients compared with that in the normal controls (0.411±0.309) (Plt;0.05). The expression of BCMA mRNA in SLE patients showed no correlation with SLEDAI score (P=0.590). Conclusion The results suggest that the expression of BCMA mRNA might play an important role in the pathogenesis of SLE.