Efficacy of BMMSCs on aGVHD and Its Correlation with SerumInflammatory Cytokines in Pediatric Patients with Severe Refractory Acute Graft-Versus-Host Disease / 中国实验血液学杂志

OBJECTIVE@#To investigate the efficacy of bone marrow mesenchymal stem cells (BMMSC) on children with refractory graft-versus-host disease (GVHD) and to judge the efficacy of BMMSC by dynamically monitoring the changes of cytokines in children with GVHD before and after infusion of BMMSC, so as to provide a theoretical basis for clarifying the mechanism of BMMSC.@*METHODS@#17 children with refractory aGVHD including 7 of grade II, 6 cases of grade III and 4 cases of grade IV after allo-HSCT were enrolled. All the children with aGVHD, who received routine immunosuppressive therapy, but the state of disease not improved, were treated with immunosuppressive drugs combined with BMMSC infusion. Study endpoints included safety of BMMSC infusion, response to BMMSC, and overall response of aGVHD. The serum levels of IL-2α, IL-6, IL-10, IL-8 and TNF-α in aGVHD patients were measured by chemiluminescence before infusion of BMMSCs and Day 7, Day 14 after infusion of BMMSCs.@*RESULTS@#The cumulative median dose of BMMSCs was 5.5 (3.4-11.1) × 10/kg for average of 3.7 times, and the median time of 16.5 (4-95) days for the first infusion of MSCs. In 17 cases of refractory GVHD, 14 responded to treatment, whereas 3 patients failed. The total effective rate was 82.4% and no adverse reactions occurred. Of the 14 survived cases (82.4%), the median follow-up time was 944 (559-1245) days from the first infusion of MSCs. The levels of TNF-α in children with grade II, III and IV GVHD before treatment were 9.5±4.3 pg/ml, 16.3±10.9 pg/ml and 35.8±21.2 pg/ml respectively. The difference between grade II and IV, III and IV was statistically significant (P<0.05). Compared with the ineffective group of BMMSC infusion, the serum TNF-αlevel in the BMMSCs treatment effective group was 10.8±5.6 pg/ml vs 40.6±14.8 pg/ml (t=-3.901, P<0.05) before treatment. In the effective group of BMMSCs infusion, IL-10 20±17.4 pg/ml of day 14 was significantly higher than that 7.3±3.1 pg/ml before the treatment (t=-2.850, P<0.05), while , the serum levels of IL-2α, IL-6, IL-8, TNF-α were not statistically significantly different (P＞0.05).@*CONCLUSION@#The infusion of BMMSC is safe and effective in the treatment of refractory GVHD in children. TNF-αlevel relates with the severity of GVHD. BMMSC may play an anti-GVHD role by up regulating the level of cytokine IL-10 in vivo.

Clinical Efficacy of NOPHO-AML 2004 Regimen for Treatment of Children with Acute Myelocytic Leukemia (Non-M3) / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the efficacy and safety of NOPHO-AML 2004 chemotherapy regimen for treatment of children with acute myelocytic leukemia(non-M3).</p><p><b>METHODS</b>Thirty-three patients aged 1-13 with acute myelocytic leukemia (non-M3) were diagnosed from January 2013 to June 2017. FAB typing showed that 1 case in M0, 4 cases in M1, 12 cases in M2, 5 cases in M4, 8 cases in M5, 1 case in M6, and 2 cases in M7; Risk stratification showed that: 19 cases in standard risk, and 14 cases in high risk. All patients were treated with NOPHO-AML 2004 chemotherapy regimen. SPSS 22.0 software was used, the Kaplan-Meier survival analysis method and Cox regression model were used for statistical analysis.</p><p><b>RESULTS</b>In the first course of treatment (AIET), among 33 child patients there were 27 cases with complete remission, and 5 cases with non-remission, thus the remission rate was 81.8%. Out of the 5 child patients without remission, 4 cases reached to the complete remission after the second course (AM), and 1 case did not remission, thus the total remission rate was 96.9%.9 cases (27.3%) underwent bone marrow recurrence and the median recurrence time was 30 months after complete continuous remission. Univariate analysis showed that age and erythrocyte transfusion frequency were significant factors to affect the early treatment response; the multiple Cox regression analysis showed that: age >7, MRD positive, erythrocyte transfusion >4 times and poor response to early treatment were independent risk factors for recurrence; Allogeneic hematopoietic stem cell transplantation(HSCT) in 8 high-risk children received enhanced chemotherapy had better efficacy as compared with the chemotherapy alone. The 3-year event-free survival rate was 59.9%, and 3-year overall survival rate was 69.2%. 33 children patients experienced varying degrees of infection and myelosuppression, or drug-related gastrointestinal reactions and allergic reactions, patients were tolerable to these side reactions after active symptomatic treatment.</p><p><b>CONCLUSION</b>NOPHO-AML 2004 chemotherapy regimen has high response rate and good tolerance, early treatment response is an important factor influencing prognosis. Age and repeated red blood cell infusions are the important factors influencing the prognosis, which promote bone marrow recurrence in AML children. For the children suffered from clinical high-risk AML, the NOPHO-AML 2004 chemotherapy regimen combined with HSCT can improve the prognosis of patients.</p>

Effects of blocking co-stimulatory signals on immunotolerance rejection of sensitized recipient after hematopoietic stem cell transplantation / 中国实验血液学杂志

This research was aimed to explore the effects of blocking B7/CD28 and CD40/CD154 co-stimulatory signals on engraftment of hematopoietic stem cell in the sensitized recipient so as to provide the experimental evidence for the treatment of sensitized recipient's immune rejection after clinical allogeneic hematopoietic stem cell transplantation (HSCT). The BALB/c mice were divided into 4 groups: (1)mice sensitized on 7 day before transplant; (2)mice were sensitized on 7 day before transplant, and injected CTLA4Ig+anti-CD154 applied; (3)normal mice injected by corresponding isotype control IgG of CTLA4Ig and anti-CD154; (4)normal blank control mice. Each group had 15 mice. On day 0, mice of each group were irradiated lethally 8 Gy by linear accelerator, and the bone marrow cells of C57BL/6 labeled by fluorescence staining were injected via the tail vein. The fluorescent cell level in peripheral blood and organ tissue at different time points were detected by flow cytometry (FCM) for homing assessment. Survival rates and hematopoietic reconstitution were also monitored and recorded. The results showed that application of CTLA4Ig anti-CD154 could promote implantation of allogeneic HSC in sensitized recipients, induce the immune tolerance, prolong their survival time and accelerate the hematopoietic reconstitution within 28 days, compared with the sensitized group. It is concluded that applying CTLA4Ig and anti-CD154 can enhance the engraftment of HSCT and induce immune tolerance in the sensitized recipient after allogeneic HSCT and accelerate the hematopoietic reconstitution.

Effects of CD4(+)CD25(+) regulatory T cells on allogeneic hematopoietic stem cell transplantation in sensitized mice / 中国实验血液学杂志

The aim of this study was to investigate the effects of CD4(+)CD25(+) regulatory T cells (Treg) on allogeneic hematopoietic stem cell transplantation (HSCT) in sensitized mice so as to provide experimental evidence for clinical treatment of allogeneic HSCT rejection in sensitized recipients. The BALB/c mice were divided into 5 groups: group A - mice were sensitized with injection of splenocytes; group B - mice were sensitized with splenocytes and treated with >5×10(5) Treg on day 7 before transplantation; group C - mice were sensitized with splenocytes and treated with 5×10(5) Treg on day 13 and 7 before transplantation; group D - mice were not sensitized, but treated with equal volume of PBS as control; group E - blank control. Each group had 15 mice. On day 0 of transplantation, mice in each group were irradiated lethally with 8 Gy by linear accelerator, and the bone marrow cells of C57BL/6 labeled by fluorescence staining were intravenously injected via the tail vein. The fluorescent cells in peripheral blood and organ tissue were detected by flow cytometry on different time points for homing assessment. Survival rates and hematopoietic reconstitution were also recorded and monitored. The results showed that on 12 and 24 hours after transplantation, as compared with the sensitized group, the number of fluorescence homing cells in different tissue of the applied Treg groups increased significantly and the differences were statistically significant (P < 0.05). The mice in sensitized group and blank control group all died on the 6-13 day, whereas the median survival time of mice in applied Treg once and twice were 15 days and 16 days respectively. Comparing with sensitized group, the difference was statistically significant (P < 0.001), but there was no significant difference between these two groups applied regulatory T cell (P > 0.05). It is concluded that applying Treg can induce immune tolerance of sensitized recipient to allogeneic HSCT and inhibit immune destruction and prolong the survival time, but can not induce full immune tolerance and at last sensitized mice died of rejection of hematopoietic stem cells.

Acute leukemia child with ocular Kaposi's sarcoma after hematopoietic stem cell transplantation: a case report and literatures review / 中华血液学杂志

<p><b>OBJECTIVE</b>To summarize clinical features of eye Kaposis' sarcoma ( KS ) in leukemia child after peripheral blood stem cell transplantation (PBSCT).</p><p><b>METHODS</b>One 13 years-old child with acute lymphoblastic leukemia (ALL) and negative HIV test who developed KS restricted in right conjunctiva, cornea and sclera after successful allogeneic PBSCT was reviewed retrospectively.</p><p><b>RESULTS</b>The child suffered from T cell type ALL. He received immunosuppressive treatment after PBSCT, and had once extensive herpes zoster restricted in skin. Seven months after PBSCT, he had blurred vision with right eye and slowly neoplasm formed in cornea and conjunctiva. Pathological examination confirmed KS with changes like capillary hemangioma, atypical fusiform cell, typical immunochemistry and positive immunofluorescent result of HHV8. He received excision of lump of cornea, conjunctiva, sclera and transplantation of cornea and sclera. Antiviral therapy was given together with anti-infection, prevention of cornea rejection and biotherapy. He kept right eye and hand-move eyesight, survived without GVHD or recurrence of ALL and KS.</p><p><b>CONCLUSION</b>This was the first ocular KS case in ALL child after PBSCT, without correlation with HIV infection. Complete excision combined with biotherapy was safe and effective for single ocular lesions.</p>

In vitro amplification of CD4(+) CD25(+) regulatory T cells and identification of amplified T cell immunosuppressive function / 中国实验血液学杂志

This study was purposed to compare the effect of 3 different cell components for expanding CD4(+) CD25(+) Treg in vitro, and identify their immunosuppressive function. CD4(+) T cells, CD4(+) CD25(-)T cells and CD4(+) CD25(+)T cells were isolated from mouse splenocytes by MACS and then expanded in vitro. Phenotype of the T cell lines and expression of the FOXP3 was determined by flow cytometry. The inhibitory effect of expanded CD4(+) CD25(+) T cells on CD4(+) CD25(-)T cells was tested by MLR method. The results showed that the Treg cells from all the three groups were expanded significantly after culture for 2 weeks. In the CD4(+) T cells group, the proliferation rate was (77.8 ± 5.32) folds with a percentage of Treg cells increasing from (6.61 ± 1.00)% to (15.33 ± 1.31)%. The proliferation rate in the CD4(+) CD25(-) T cells group was (95.20 ± 7.67) folds, with the percentage of CD4(+) CD25(+) T cells raising from (0.37 ± 0.13)% to (9.84 ± 0.98)%. The proliferation rate in the CD4(+) CD25(+) T cells group was (41.20 ± 6.92) folds, the proportion of Treg cells decreased from (86.75 ± 1.25)% to (85.32 ± 1.62)%, and the expression of Foxp3 decreased from (76.92 ± 1.72)% to (75.33 ± 2.11)% during the culture, there were not significant differences in the cell purity and the expression of Foxp3, compared with pre-amplification. The inhibitory test showed that the expanded CD4(+) CD25(+) T cells could inhibit the proliferation of CD4(+) CD25(-) T cells in vitro in a cell dose-dependent manner. It is concluded that the amplification of CD4(+) CD25(+) Treg cells is successful in vitro, especially in the CD4(+) CD25(+) T cells group, the cell purity and Foxp3 gene is not obviously changes after amplification.

Separation and amplification of CD4(+)CD25(+) regulatory T cells from sensitized mice / 中国实验血液学杂志

The aim of this study was to separate and amplify CD4(+)CD25(+)Treg cells from splenocytes of sensitized mice. The percentage of CD4(+)CD25(+)Treg cells was detected by flow cytometry in sensitized and normal control mice. CD4(+)T, CD4(+)CD25(+)Treg and CD4(+)CD25(-) T cells were isolated from mouse splenocytes by MACS. CD4(+)CD25(+)Treg cells were expanded in vitro cultures in addition of CD3/CD28 MACSiBead and IL-2. The activity of cells was detected with 0.4 trypan blue staining. The purity of cells after sorting, the main surface marker and the level of Foxp3 were detected by flow cytometry. The results showed that CD4(+)CD25(+)Treg cell proportion was higher in sensitized mice than normal control mice (P < 0.05). The average purity of CD4(+)CD25(+)Treg cells was 87. The activity of these cells was more than 97, and the expression of Foxp3 in these cells was high. The amplification multiples achieved 42 times after 2 weeks in vitro. The percentage of CD4(+)CD25(+) regulatory T cells was 85.32, and the expression of Foxp3 decreased from (76.92 ± 1.72) to (75.33 ± 2.11) (P > 0.05). It is concluded that the sorting of CD4(+)CD25(+)Treg cells is isolated successfully by MACS without affecting the vitality of target cells. The amplification of CD4(+)CD25(+)Treg cells is successful in vitro. Expression of surface markers and Foxp3 gene does not obviously change after amplification, so that to establish a practical method to recover and enlarge the amount of CD4(+)CD25(+)Treg cells in good condition.

Experimental study on rejection of allogeneic donor bone marrow cells in sensitized recipients / 中华血液学杂志

<p><b>OBJECTIVE</b>To establish a murine model of sensitization, and investigate the effect and mechanism of sensitization on allogeneic donor bone marrow cells (BMCs).</p><p><b>METHODS</b>Sensitized BALB/c mice were established by transfusions of allogeneic splenocytes. The donor reactive antibodies were detected by binding and complement-dependent cytotoxicity assays. After irradiation, 1 × 10(7) BMCs of C57BL/6 donor mice were injected into non-sensitized or sensitized BALB/c recipient mice. The distribution pattern of donor BMCs in peripheral blood, spleen and bone marrow of recipient mice were analyzed at different time points (2 h, 12 h and 48 h) post transplantation. Hematopoietic recovery post transplantation was assessed, and survival was monitored. Moreover, sera and splenocytes derived from non-sensitized or sensitized recipients were incubated with allogeneic BMCs in vitro, and the cytotoxic indexes were calculated in the immune experiments.</p><p><b>RESULTS</b>The binding and complement-dependent cytotoxicity assays showed that a high level of donor reactive antibodies was presented in sensitized sera. Compared with the non-sensitized recipients, the homing assay showed significantly decreased distributions of allogeneic donor BMCs in peripheral blood, spleen and femur of sensitized recipients. Non-sensitized recipients survived long term after irradiation, while all the sensitized recipients died within 12-15 days. Fourteen days post transplantation, the white blood cells and BMCs of non-sensitized recipients were (3240 ± 300) × 10(6)/L and (396 ± 27) × 10(6)/femur, respectively; while the white blood cells and BMCs of sensitized recipients were (320 ± 80) × 10(6)/L and (6 ± 2) × 10(6)/femur, respectively; the differences were statistically significant between this two groups (P < 0.05). Seven days post transplantation, the percentage of donor cells in bone marrow of non-sensitized and sensitized recipients was (48.07 ± 4.70)% and (0.77 ± 0.11)%, respectively, and the differences were statistically significant (P < 0.05). Furthermore, the white blood cells and BMCs following transplantation decreased along with time in sensitized recipients. The immune experiments of complement-dependent cytotoxicity reaction, cytotoxic T lymphocytes reaction and antibody-dependent cellular cytotoxicity showed the cytotoxic indexes were higher in sensitized group than the non-sensitized group.</p><p><b>CONCLUSION</b>A sensitized model was established by transfusions of allogeneic spleen cells. Allogeneic donor BMCs were rejected in sensitized recipients, and its mechanism might be through immune impairment pathways.</p>

Transfusion of hematopoietic stem/progenitor cells into marrow cavity in sensitized mouse model / 中国实验血液学杂志

The study was aimed to investigate the strategy of transfusion of allogeneic hematopoietic stem/progenitor cells (HS/PC) into marrow cavity of mouse model in sensitized transplantation. A sensitized BALB/c mouse model was established by repeated transfusion of allogeneic spleen cells. The normal BALB/c mice were used as non-sensitized controls. The non-sensitized or sensitized recipients were transplanted by transfusion of allogeneic HS/PCs into bone marrow cavity. The survival rate and hematopoietic recovery were monitored. Moreover, non-sensitized and sensitized sera were obtained and incubated with allogeneic HS/PC respectively, the percentage of dead cells was calculated using complement-dependent cytotoxicity (CDC) tests. The results showed that non-sensitized recipients got long-term survival after the transfusion of HS/PC into marrow cavity, and the hematopoietic recovery increased along with time. However, among the sensitized recipients, one mouse died of anesthetic accident, the other 9 mice (9/10) died within 2 weeks after the transfusion of HS/PC in marrow cavity, and the hematopoietic recovery declined along with time. Histopathologic analysis demonstrated that the sensitized recipients died of bone marrow failure. The results of CDC tests showed that the percentage of dead cells in non-sensitized and sensitized group was 7.80 ± 1.93% and 50.80 ± 3.12%, respectively, and the differences were statistically significant (p < 0.05), indicating sensitized sera were capable of impairing allogeneic HS/PC. It is concluded that the strategy of the marrow cavity transfusion of HS/PC can not enhance engraftment of allogeneic donor cells in sensitized recipients.

Therapeutic effect of focal adhesion kinase gene silence on leukemia / 中国实验血液学杂志

This study was aimed to investigate the effects of focal adhesion kinase (FAK) gene silence on leukemia cell growth, leukemogenesis and efficacy of chemotherapy drug. Vector containing lentiviral-FAK-shRNA was constructed and transfected into BCR/ABL-BaF3 leukemic cells, the cell growth and apoptosis were detected in vitro. The effect of FAK shRNA on leukemogenesis was studied in a murine model with leukemia. The apoptosis of leukemia cells and survival of leukemic mice treated by FAK shRNA combined with drug STI571 were monitored. The results showed that FAK gene expression was knocked down by lentiviral-FAK-shRNA. FAK gene silencing inhibited leukemia cell growth in vitro. The apoptosis test results showed that the percentages of Annexin V(+) cells in vector control group and FAK shRNA group were (3.46 ± 0.56)% and (7.3 ± 0.79)%, respectively, and the difference was statistically significant (p < 0.05). The mice in vector control group died at day 21 to 27, while the mice in FAK shRNA group died between day 52 and 60, and the difference was statistically significant (p < 0.05). Moreover, FAK gene silence combined with drug STI571 could enhance the apoptosis of leukemia cells and prolong survival time of leukemic mice. It is concluded that FAK gene silence inhibits leukemogenesis and promotes efficacy of chemotherapy drug on leukemia cells, indicating FAK gene silence may be considered as a new therapeutic strategy for leukemia.

Establishment of sensitized animal models and their sensitization effects on engraftment of hematopoietic stem cells / 中国实验血液学杂志

This study was aimed to establish sensitized animal models, explore the changes of immune function in these sensitized recipients, and investigate effects of sensitization on the engraftment of hematopoietic stem cells (HSCs). Different doses of spleen cells (1x10(5), 1x10(6) and 1x10(6)x2 at intervals of 7 days) from C57BL/6 were infused into BALB/c, the immunity function of sensitized models was tested by complement-dependent cytotoxicity method, mixed lymphocyte reaction and ELISA. After irradiation with gamma-ray of 60Co in dose 8 Gy, sensitized mice were transplanted 1x10(7) C57BL/6 bone marrow cells via tail vein or intra-bone marrow, and survival rate was detected daily. The results showed that different levels of donor reactive antibody were induced in all sensitized models. Comparing with normal mice, profound proliferation of spleen cells were found in groups of injected 1x10(6) and 1x10(6), continuous injections at intervals of 7 days. Sensitized model received bone marrow cells of C57BL/6 via tail vein died on day 10 to 14 after transplantation, and sensitized model mice received bone marrow cells of 1x10(6)x2 at intervals of 7 days via intra-bone marrow also died within two weeks after transplantation. It is concluded that different sensitized mouse models are established by different doses of allogeneic spleen cells infusion, the changes of immune function in sensitized mice are correlative with sensitization. Donor HSCs are rejected in sensitized models, and the engraftment can not be improved by intra-bone marrow injection.

Experimental study on the effect of sensitized sera on engraftment of hematopoietic stem cells / 中华血液学杂志

<p><b>OBJECTIVE</b>To evaluate the effect of sensitized sera on engraftment of hematopoietic stem cells (HSCs) and its mechanism.</p><p><b>METHODS</b>Bone marrow cells (BMCs) from C57BL/6 mice were incubated with sensitized sera (group I) or normal sera (group II), and then washed and transplanted into irradiated BALB/c mice. The survival analysis and engraftment evaluation of the recipients were observed. The incubated BMCs were bound with goat anti mouse IgG for and labeled with Annexin V for apoptosis detection.</p><p><b>RESULTS</b>Seven out of ten recipient mice in group I died of bone marrow failure at day 10 after transplantation, while all of those in group II were long-term survived. Engraftment assay showed recipients blood count and BMCs progressively decreased along with time passing in group I; in addition, the chimeric percentage of donor cells progressively decreased as well. The percentage of BMCs binding with goat anti mouse IgG in group I or group II were (90.3 +/- 5.1)% and (5.2 +/- 2.4)%, respectively, and the difference was statistically significant (P < 0.01). However, no significant difference was found in the apoptosis detection between the two groups (P > 0.05).</p><p><b>CONCLUSION</b>The engraftment capacity of HSCs is significantly impaired by sensitized sera, the antibodies in sensitized sera may bind to antigens expressed on HSCs but do not induce apoptosis.</p>

Effect of the serum panel reactive antibody on proliferation and differentiation of cord blood CD34+ cells in vitro / 中华儿科杂志

<p><b>OBJECTIVE</b>The low rate of engraftment in children with beta-thalassemia has seriously restricted the popularity of the hematopoietic stem cell transplantation (HSCT). Panel reactive antibody (PRA) has been regarded as one of the important factors for the success of kidney transplantation. Poly-transfusion before transplantation is associated with the production of PRA. Also PRA is produced in the children with beta-thalassemia major who need poly-transfusion for life. PRA might be one of factors inducing the low rate of engraftment in children with beta-thalassemia. This study focused on observing the effect of PRA on the proliferation, differentiation, apoptosis and necrosis of cord blood CD34(+) cells in vitro by incubating the cord blood CD34(+) cells with serum containing PRA.</p><p><b>METHOD</b>Seven samples of cord blood were collected and the HLA typing for every sample was done. Seven sera positive for PRA and seven negative sera were selected respectively. Mononuclear cells (MNCs) were obtained by Ficoll-Hypaque density gradient centrifugation. CD34(+) cells were isolated from MNCs by positive selection using an immunomagnetic separation (CD34(+) progenitor cell isolation kit and auto-MACS). The CD34(+) cells of umbilical cord blood were incubated with the serum and complement in the following groups: A (absence of serum), B (presence of PRA positive serum), C (presence of PRA positive serum and complement), D (presence of complement), and E (presence of PRA negative serum). After incubation the samples were centrifuged and the supernatant was collected for LDH detection. At the same time the CD34(+) cells were harvested for assessing the expression of Annexin V and CD95 of the CD34(+) cells by flow cytometry and also for the detection of the DNA synthesis by (3)H-TaR incorporation. Meanwhile the cells were inoculated into the methylcellulose cultural system. The proliferation and hematopoietic potential of the CD34(+) cell of cord blood by the colony formation assay were detected on the day 10.</p><p><b>RESULT</b>The concentration of LDH in group A was (20.71 +/- 2.81) U/L, which was significantly lower than that in group B (64.28 +/- 5.12) U/L and group C (84.29 +/- 4.99) U/L. The concentration of LDH in group B was significantly lower than that in group C, while there were no significant differences in the concentration of LDH among groups A, D and E (P > 0.05). The cpm in group A was (22 629 +/- 3288), which was significantly higher than that in group B (4598 +/- 2178) and group C (1626 +/- 1192). And the cpm in group B was significantly higher than that in group C. There were no significant differences in the cpm among groups A, D and E (P > 0.05). On day 10 of culture, the total colonies, granulocyte-macrophage colony forming unit (CFU-GM), mixed colony forming unit (CFU-GEMM) and erythroid burst colony forming unit (BFU-E) in group A were significantly higher than that in group B and C. The total colonies, CFU-GM and CFU-GEMM in group B were significantly higher than those in group C. No significant differences were found in the total colonies, CFU-GM, CFU-GEMM and BFU-E among groups A, D and E (P > 0.05). There were no statistically significant differences in the CD95 and Annexin V expression among all the groups (P > 0.05).</p><p><b>CONCLUSION</b>PRA could impair the membrane, decrease the DNA synthesis, and inhibit the colony formation of CD34(+) cord blood cells, which could be strengthened by the presence of the complement at the given concentration in our study. PRA had no significant influence on the apoptosis of CD34(+) cells in vitro.</p>

Effects of sensitized sera on bone marrow transplantation in a murine model / 中国免疫学杂志

Objective:To investigate the effects of sensitized sera on bone marrow transplantation and clarify the role of antibody in the process of rejection.Methods:Two hundred microlitres sensitized sera or non-sensitized sera were injected into normal BALB/c one day before transplantation.Ten millions (1?107) bone marrow cells from C57BL/6 were transfused to the recipients after lethal irradiation.The donor-reactive antibodies in recipients before transplantation were measured by complement-dependent cytotoxicity (CDC) method.Moreover,the survival analysis and engraftment evaluation of the recipients were carried post transplantation.Results:The CDC results showed that donor-reactive antibodies existed in the recipients which had received sensitized sera transfusion.Eighty percent (80%) of the recipients received sensitized sera transfusion died of bone marrow failure about 10 days post transplantation,while the recipients received non-sensitized sera transfusion were long-term alive.Furthermore,the hematopoietic recovery and percentage of donor chimera analysis declined along with time in the sensitized sera transfusion recipients,and there were significant differences compared with those in the non-sensitized sera transfusion recipients (P

FAK gene silencing induces apoptosis of leukemic cells in vitro / 中国病理生理杂志

AIM:Targeting of focal adhesion kinase (FAK) gene,we aim to construct FAK shRNA lentiviral vector and to identify its function on the growth of leukemic cells.METHODS:FAK shRNA was chemically synthesized,and inserted into a GFP-lentiviral plasmid through molecular biology methods.After packaged and concentrated,the lentiviral-FAK-shRNA-vector was transduced into a human leukemic cell line.FAK gene expression was detected by reverse transcriptional PCR and Western blotting.Cell apoptosis was measured by Annexin V labeling.RESULTS:The results showed that FAK shRNA was successfully inserted into the lentival vector,and the infection efficiency varied from 10% to 25%.Compared to the control vector (lentival vector without FAK shRNA),FAK shRNA inhibited the expression of FAK mRNA and protein by 40% and 70%,respectively.Moreover,the results of apoptosis experiment showed that the percentages of Annexin V+ cells in control vector group and FAK shRNA group were (4.19 ? 0.36) % and (8.48 ? 0.58) % respectively,the difference was statistically significant (P