The evaluation of modified cell infusion method to reduce febrile non-hemolytic transfusion reaction in CD19 chimeric antigen receptor T cell threapy / 中华内科杂志

Objective@#To retrospectively analyze the efficacy and safety of modified cell infusion method in reducing the incidence of febrile non-hemolytic transfusion reaction (FNHTR).@*Methods@#A total of 69 patients were enrolled in the clinical trial of CD19 chimeric antigen receptor T (CAR-T) cell treatment from February 2017 to October 2018. Study group received the modified cell infusion method, that 1×106 CAR-T cells were re-suspended in 2 mg human serum albumin with total volume of 20 ml and injected intravenously. The control group was intravenously administrated with CAR-T cell in 100 ml normal saline. The incidence of FNHTR, cytokine releasing syndrome (CRS) grade, cytokine level and efficacy were compared.@*Results@#(1)The incidence of FNHTR in the study group was 21.1%, significantly lower than that in the control group (71%)(P=0.000). (2)There was no statistical difference in cell proliferation between the study group and the control group on day 4, 7, 14 and 21 after CAR-T cell infusion (P=10.223, 3.254, 5.551, 7.605). (3)There was no statistical difference in CRS grading between the study group and the control group (P=0.767). There was no statistical difference in the levels of interleukin 2 receptor (IL-2R), IL-6, tumor necrosis factor (TNF)-α between the two groups. (4)The C-reaction protein (CRP) level of the study group was lower than that of the control group on day 4 and 7 (P=0.026, 0.007). (5)There was no statistical difference of response rates in acute lymphocytic leukemia (ALL) and non-Hodgkin lymphoma (NHL) patients between the two groups (PALL=0.842; PNHL=0.866).@*Conclusion@#The modified cell infusion method in CD19 CAR-T cell treatment reduces the incidence of treatment-related FNHTR. It does not affect the proliferation of CAR-T cells in vivo, the grading of CRS and the response rates.

Immunophenotypic characteristics of CD19 chimeric antigen receptor( CAR)-transfected Nalm-6 cells / 中华微生物学和免疫学杂志

Objective To investigate the immunophenotypic characteristics of potential leukemia cells transfected with CD19 antigen receptor( CAR) during CAR-T cell preparation. Methods Morphological chan-ges in CD19 CAR-transfected cells were observed under inverted microscope. The transfection rate and immuno-phenotype of transfected Nalm-6 cells were analyzed by flow cytometry. Secretion of cytokines in the culture sys-tem was detected by chemiluminescence. Results The transfection rate of Nalm-6 cells by CD19 CAR was (46. 50±3. 78) ％ and that of KG1a cells was (15. 70±1. 22) ％. CD19 CAR-transfected Nalm-6 cells prolifer-ated more rapidly than Nalm-6 cells ( P values on 0 d, 4 d, 7 d and 12 d were 6. 339, 3. 447, 0. 012 and 0. 009). In the culture of CD19 CAR-transfected Nalm-6 cells, cell aggregation and adhesion were observed and they gradually gathered into a group. The rate of CD19 expression was only 1. 19％ in the CD19 CAR-transfect-ed Nalm-6 cell culture system with the transfection rate of (46. 50±3. 78) ％. After increasing the proportion of Nalm-6 cells in the culture system, CD19 expression was gradually increased, while the expression of CD22 re-mained stable. CD19 expressed by Nalm-6 cells cultured in the supernatant of CD19 CAR-transfected Nalm-6 cell culture system was decreased gradually. The levels of IL-10 and TNF-αsecreted by CD19 CAR-transfected Nalm-6 cells were higher than those by Nalm-6 cells. Conclusions Results of the immunophenotypic analysis of CD19 CAR-transfected leukemia cells suggested that CD22 CAR-T cell therapy could be used as a rescue or combination therapy for CD19 CAR transfection into leukemia cells.

The evaluation of modified cell infusion method to reduce febrile non?hemolytic transfusion reaction in CD19 chimeric antigen receptor T cell threapy / 中华内科杂志

To retrospectively analyze the efficacy and safety of modified cell infusion method in reducing the incidence of febrile non?hemolytic transfusion reaction (FNHTR). Methods A total of 69 patients were enrolled in the clinical trial of CD19 chimeric antigen receptor T (CAR?T) cell treatment from February 2017 to October 2018. Study group received the modified cell infusion method, that 1×106 CAR?T cells were re?suspended in 2 mg human serum albumin with total volume of 20 ml and injected intravenously. The control group was intravenously administrated with CAR?T cell in 100 ml normal saline. The incidence of FNHTR, cytokine releasing syndrome (CRS) grade, cytokine level and efficacy were compared. Results (1)The incidence of FNHTR in the study group was 21.1%, significantly lower than that in the control group (71%)(P=0.000). (2)There was no statistical difference in cell proliferation between the study group and the control group on day 4, 7, 14 and 21 after CAR?T cell infusion (P=10.223, 3.254, 5.551, 7.605). (3)There was no statistical difference in CRS grading between the study group and the control group (P=0.767). There was no statistical difference in the levels of interleukin 2 receptor (IL?2R), IL?6, tumor necrosis factor (TNF)?α between the two groups. (4)The C?reaction protein (CRP) level of the study group was lower than that of the control group on day 4 and 7 (P=0.026, 0.007). (5)There was no statistical difference of response rates in acute lymphocytic leukemia (ALL) and non?Hodgkin lymphoma (NHL) patients between the two groups (PALL=0.842; PNHL=0.866). Conclusion The modified cell infusion method in CD19 CAR?T cell treatment reduces the incidence of treatment?related FNHTR. It does not affect the proliferation of CAR?T cells in vivo, the grading of CRS and the response rates.

Long-term effects of ultrasound-guided microwave ablation in the treatment of small renal cell carcinoma / 南方医科大学学报

<p><b>OBJECTIVE</b>To evaluate the long-term efficacy of microwave ablation in the treatment of small renal cell carcinoma (RCC).</p><p><b>METHODS</b>We retrospectively analyzed 140 cases of small cell renal carcinoma (151 lesions with a mean diameter of 2.8±0.8 cm) treated between April, 2006 and October, 2015 with ultrasound-guided microwave ablation with cooled-shaft needle antenna. One microwave ablation antenna was used for tumors less than 2 cm in diameter and 2 antennas were used for larger tumors. The patients received enhanced ultrasound and CT/MRI examinations at 1, 3, and 6 months after the operation and every 6 months thereafter. The overall survival, disease-free survival, and local tumor progression rate of the patients were evaluated.</p><p><b>RESULTS</b>The response rate of treatment (complete ablation at one month on enhanced images) was 100% in these patients. The local tumor progression rates at 1, 3, and 5 years were 0.9%, 2.0%, and 7.1%, respectively, and the 1-, 3-, and 5-year distant metastasis rates were 1.6%, 2.5%, and 7.9%, respectively. The overall survival rates of the patients at 1, 3, and 5 years were 98.4%, 94.8%, 89.5%, respectively, with disease-free survival rates of 98.4%, 93.0%, and 83.1%, respectively. No major complications occurred in these cases, and multivariate analysis showed that the tumor number (P=0.015) and tumor growth patterns (P=0.049) were independent risk factors that adversely affected the long-term outcome after surgery.</p><p><b>CONCLUSION</b>Our data show that microwave ablation is a safe and effective modality for treatment of renal cell carcinoma.</p>

Maternal vitamin D level and its relevant factors in the second and third trimesters of pregnancy in a hospital in Nanjing / 中华临床营养杂志

Objective To analyze the serum level of 25-hydroxy vitamin D [25-(OH) D] in pregnant women in Nanjing and its relationship with age,gestational week,pre-pregnancy body mass index (BMI) and season,so as to provide some guidance on clinical application of vitamin D measurement and supplementation.Methods 880 pregnant women (in 21-36 gestational weeks) regularly visiting antenatal clinic in Obstetrics and Gynecology Hospital Affiliated to Nanjing Medical University were selected from January to December 2015.Their serum 25-(OH)D concentrations were measured by enzyme-linked immunosorbent assay (ELISA).Results The mean serum 25-(OH) D concentration in these pregnant women was (42.03 ± 19.22) nmol/L and sufficient in only 42 (4.77％).Vitamin D levels showed seasonal variation (P =0.0164),with the levels in summer [(44.61 ±23.57) nmol/L] and autumn [(43.43 ± 19.31) nmol/L] being higher than spring [(39.68 ± 16.91) nmol/L] and winter [(39.65 ± 13.36) nmol/L].There was no statistically significant difference in serum vitamin D level among different gestational weeks (P =0.929 4).There was statistically significant difference in serum vitamin D level among different age groups (P =0.038 3).Vitamin D level was not associated with pre-pregnancy BMI.Conclusions Vitamin D deficiency is highly prevalent among pregnant women in Nanjing.We suggest vitamin D testing be carried out more actively.

Establishment of an mouse model of iron-overload and its impact on bone marrow hematopoiesis / 中国医学科学院学报

<p><b>OBJECTIVE</b>To establish a mouse model of iron overload by intraperitoneal injection of iron dextran and investigate the impact of iron overload on bone marrow hematopoiesis.</p><p><b>METHODS</b>A total of 40 C57BL/6 mice were divided into control group, low-dose iron group (12.5 mg/ml), middle-dose iron group (25 mg/ml), and high-dose iron group (50 mg/ml). The control group received normal saline (0.2 ml), and the rest were injected with intraperitoneal iron dextran every three days for six weeks. Iron overload was confirmed by observing the bone marrow, hepatic, and splenic iron deposits and the bone marrow labile iron pool. In addition, peripheral blood and bone marrow mononuclear cells were counted and the hematopoietic function was assessed.</p><p><b>RESULTS</b>Iron deposits in bone marrow, liver, and spleen were markedly increased in the mouse models. Bone marrow iron was deposited mostly within the matrix with no significant difference in expression of labile iron pool.Compared with control group, the ability of hematopoietic colony-forming in three interventional groups were decreased significantly (P<0.05). Bone marrow mononuclear cells counts showed no significant difference. The amounts of peripheral blood cells (white blood cells, red blood cells, platelets, and hemoglobin) in different iron groups showed no significant difference among these groups;although the platelets were decreased slightly in low-dose iron group [(780.7±39.60)×10(9)/L], middle dose iron group [(676.2±21.43)×10(9)/L], and high-dose iron group [(587.3±19.67)×10(9)/L] when compared with the control group [(926.0±28.23)×10(9)/L], there was no significant difference(P>0.05).</p><p><b>CONCLUSIONS</b>The iron-overloaded mouse model was successfully established by intraperitoneal administration of iron dextran. Iron overload can damage the hepatic, splenic, and bone marrow hematopoietic function, although no significant difference was observed in peripheral blood count.</p>

Effect and mechanism of iron-catalyzed oxidative stress on mesenchymal stem cells / 中国医学科学院学报

<p><b>OBJECTIVE</b>To explore effect of iron overload on the proliferation and apoptosis of mesenchymal stem cell(MSCs) and the possible mechanism.</p><p><b>METHODS</b>Iron overload model of MSCs was established by adding ferric ammonium citrae (FAC) into the culture medium at different concentrations (100, 200, 400 Μmol/L) and incubated for different lengths of time (12, 24, 48 h). The levels of labile iron pool (LIP) and reactive oxygen species (ROS) were measured to confirm oxidative stress state in the model. Changes in cell proliferation and apoptosis after iron overload were measured through population double time(DT)and annexin V-PI assay. Finally, the expressions of phosphorylated p38 mitogen activated protein kinase (P-p38MAPK), p38MAPK, protein kinase B (AKT), and p53 were determined through Western blot analysis to investigate which ROS-mediated signaling pathway was involved in this process.</p><p><b>RESULTS</b>The LIP level of MSCs was significantly increased by FAC treatment at 400 Μmol/L (mean fluorescence intensity 482.49±20.96 vs. 303.88±23.37, P<0.05). The level of intracellular ROS was positively correlated with the concentration of FAC and reached a peak level when cultured with 400 Μmol/L FAC (P<0.05).After treatment with 400 Μmol/L FAC at different time points (12 h, 24 h, and 48 h), the DT of MSCs was (1.47± 0.11) d, (1.80±0.13) d, and (2.04±0.14) d, respectively, which was signifcantly longer than that of the control, which was(1.20±0.05)d (P<0.05).The apoptosis rate was also significantly higher in iron overload group[(3.51±1.17)% vs.(0.66±0.62)%, P<0.05]with consequent increase in the expressions of P-p38MAPK, p38MAPK, and p53 proteins in iron overload group, while no significant difference was found in the expression of AKT.</p><p><b>CONCLUSION</b>Iron overload can inhibit the proliferation of MSCs and induce their apoptosis through the generation of ROS, which is probably due to the stimulation of p38MAPK- p53 signaling pathway.</p>

Effects of Recombinant Human Fibronectin Fragment Combined with Anti-CD3 Monoclonal Antibody on the Proliferation and Cytotoxicity of Cytokine-induced Killer Cells from Acute Leukemia / 天津医药

Objective To investigate the effects of recombinant human fibronectin fragment (RetroNectin) combined with anti-CD3 monoclonal antibody (CD3Ab) on the proliferation and cytotoxicity of cytokine-induced killer cells (CIK) from acute leukemia (AL). Methods Mononuclear cells (MNCs) were isolated from peripheral blood of complete remission AL pa-tients. The MNCs were cultured in vitro by precoating with RetroNectin (RN group), CD3Ab (CD3Ab group), RetroNectin com-bined with CD3Ab (RN+CD3Ab group) and traditional method (control group) to generate CIK. The changes of growth rate, characterization, cytotoxicity and apoptosis of CIK were determined between groups. Results The amplification of CIK was higher in experimental group than that of control group, and the amplification of CIK was higher in group RN+CD3Ab than that of in group RN and group CD3Ab (P＜0.05). The expression of CD25 positive cells was higher in group RN and group RN+CD3Ab than that of group CD3Ab and control group (P＜0.05).The percentage of G1 stage cells was lower in group RN and group RN+CD3Ab than that of group CD3Ab and control group. The percentage of S stage cells was higher in group RN and group RN+CD3Ab than that of group CD3Ab and control group (P＜0.05). The cytotoxicity was higher in group RN and group RN+CD3Ab than that of group CD3Ab and control group (P＜0.05) at the E/T scope 40∶1.The percentage of apoptotic cells was lower in group RN and group RN+CD3Ab than that of group CD3Ab and control group (P ＜ 0.05). Conclusion These in vitro studies suggest that a higher activity of immune cells could be obtained by CIK cells cultured by precoating Ret-roNectin and CD3Ab.

Mortality trend on chronic obstructive pulmonary disease in Liaoning province, 1984-2010 / 中华流行病学杂志

Objective To evaluate the mortality trend or chromc obstructive pulmonary disease (COPD) among residents in Liaoning province during the period of 1984-2010.Methods The cut-points were ascertained by Monte Carlo Permutation test in COPD mortality trend lines of Poisson regression with Joinpoint Regression Program.The annual percent changes (APC) before and after the cut-points and the average annual percent change(AAPC) of COPD mortality were examined during the period.Results Significant declining trends on COPD mortality among the urban population during 1984-2010 and that of rural population during 1999-2009 were found.The standardized urban COPD mortality rate by Chinese population declined from 243.93 per 100thousand in 1984 to 33.13 per 100 thousand in 2010.The urban 26 years AAPC was -5.8％.While the mortality in the rural population decreased from 251.33 per 100 thousand in 1999 to 102.25 per 100 thousand in 2009 in the same population.The rural 10 years' AAPC was-6.8％.The total trend of COPD mortality reduction was mainly resulted from the fast decline of bronchitis mortality.The AAPC of COPD mortality of the urban population was-9.0％ and greater than that of the rural population (-6.8％) from 1999 to 2009.The urban population had a lower COPD mortality than that of the rural population.In urban area,males had a higher COPD mortality than females,however,in the rural area,males had a lower COPD mortality than the females.Conclusion The COPD mortality among the residents of Liaoning province declined significantly from 1984 to 2010.Further studies are needed to confirm the viewpoint of WHO that the prevalence of COPD would have a continuous increasing trend in China.

In vitro effect of iron overload on bone marrow cell function by inducing the reactive oxygen species / 中华血液学杂志

<p><b>OBJECTIVE</b>To investigate the in vitro effect of iron overload on the generation of reactive oxygen species (ROS) and of bone marrow (BM) cell function.</p><p><b>METHODS</b>BM mononuclear cells (BMMNCs) were cultured with ferric citrate (FAC) at different concentrations and for different time to create iron overload and confirmed by the detection of cellular labile iron pool (LIP). The changes of ROS, apoptosis, hematopoietic colony formation (CFU-E, BFU-E, CFU-GM and CFU-mix) and the percentage of the CD34 + cells percentage were analyzed. The differences of these index were tested after the iron overload treated with deferasirox (DFO) or antioxidants (N-acetyl-L-cysteine, NAC).</p><p><b>RESULTS</b>1) When BMMNCs were cultured with FAC, the LIP was found to increase in a time and concentration dependent manner. The intracellular LIP reached maximum at 400 micromol/L of FAC for 24 hours. 2) The ROS of total cells, leukocytes and erythrocytes increased to 1.77, 1.75 and 2.12 fold respectively compared with that of normal control when cells were cultured at 400 micromol/L of FAC for 24 hours . DFO and NAC could reduce the ROS efficiently (P<0.05). 3) The apoptotic rates of the FAC treated cells [(24.80 +/- 2.99)%] increased significantly compared with that of normal control [(8.90 +/- 0.96)%]. The capacity of hematopoietic colony formation in FAC treated cells decreased markedly compared with that of normal control (P<0.05). The percentage of CD34+ cells of FAC treated cells [(0.39 +/- 0.07)%] also decreased significantly compared with that of normal control [(0.91 +/- 0.12)%]. And these changes could be recovered by addition of NAC or DFO.</p><p><b>CONCLUSION</b>Iron overload can affect the hematopoiesis by inducing the generation of ROS and this damage could be corrected by removing the excess iron and ROS of the BM cells. These findings might improve the treatment of dyshematopoiesis in patients with iron overload.</p>

Decomposition of life expectancy at birth by age and causes of death among residents of Liaoning province,1975-2005 / 中华流行病学杂志

Objective To analyze the impact of mortality by age and causes of death on life expectancy at birth among residents of Liaoning province.Methods The study included mortality data of urban and rural residents in two periods (1973-1975 and 2004-2005).Both Abridged Life Table and Arriaga method were used to calculate and to decompose life expectancy changes by age and causes of death.Results From 1975-2005,the life expectancy increased by 4.68 years in urban residents and 4.91 for rural residents with a higher increment among females than males.Most part of the increase (76.27% and 82.81% for urban and rural male,58.76% and 62.13% for urban and rural female) in life expectancy within the last 30 years could be explained by the decrease of mortality in the populations at age 0-4 and 55-74.Diseases related to respiratory system and infectious disease were contributing the most to the gap in life expectancy between the two periods.Mortality of heart disease was a negative contributor to the changes in life expectancy among both rural and urban residents while the mortalities of cerebro-vascular diseases and malignant tumors were the negative contributors for rural residents.Conclusion The increase of life expectancy in the last 30 years was mainly resulted from the decrease of mortality on both respiratory and infectious diseases.Control of chronic diseases is the key point to increase the life expectancy among the residents of Liaoning province.

Establishment of iron overloaded bone marrow model in vitro and its impact on hematopoiesis / 中国实验血液学杂志

This study was to establish an iron overload bone marrow (BM) model by co-culturing the mononuclear cells from BM with iron, and investigate its hematopoiesis changes. The iron overload model was set up by adding different concentration of ferric citrate (FAC) into the mononuclear cells from BM and culturing for different time, and the model was confirmed by detecting labile iron pool (LIP). Then the apoptosis of hematopoietic cells, ability of hematopoietic colony forming (CFU-E, BFU-E, CFU-GM and CFU-mix) and percentage of the CD34(+) cells of the BM cells all were determined. The changes of these indexes were tested after the iron-overloaded BM was treated with deferasirox (DFO). The results showed that after BM cells were cultured with FAC at different concentrations for different time, the LIP increased in time-and concentration-dependent manners. The intracellular LIP reached maximum level when cultured at 400 µmol/L of FAC for 24 hours. The detection of BM cell hematopoietic function found that the apoptotic rate of the FAC-treated cells (24.8 ± 2.99%) increased significantly, as compared with normal control (8.9 ± 0.96%)(p < 0.01). The ability of hematopoietic colony forming in FAC-treated cells decreased markedly, as compared with normal control (p < 0.05). The percentage of CD34(+) cells of FAC-treated cells (0.39 ± 0.07%) also decreased significantly, as compared with normal control (0.91 ± 0.12%)(p < 0.01). And these changes could be alleviated by adding DFO. It is concluded that the iron-overloaded model has been set by adding iron into the mononuclear cells from BM in vitro, and the hematopoietic function of iron-overloaded BM is deficient. These changes can be alleviated by removing the excess iron from the BM cells through treating with DFO. These findings would be helpful to further study the mechanism of iron-overload on the hematopoiesis of BM and also useful to find the way to treat iron-overload patients with hematopoietic disorders.

Clinical significance of leukemia stem/progenitor cell related gene expression in acute leukemia / 中国实验血液学杂志

This study was aimed to detect the expression of leukemia stem/progenitor cell (LSPC) related genes (ABCB1, BMI-1, HOXB4) in the patients with acute leukemia, and to explore its clinical significance in acute leukemia. Bone marrow samples were collected from de novo acute leukemia patients (41 cases), patients with complete remission (CR, 16 cases) and the patients with non-malignant hematologic diseases (10 cases) respectively. And the expressions of ABCB1, BMI-1, HOXB4 genes were detected by comparative real-time quantitative PCR (RQ-PCR) with SYBR Green assay. The results showed that the expressions of ABCB1, BMI-1, HOXB4 were not detected in the patients with non-malignant hematologic diseases, but were higher (relative expressive level: 4.26 ± 2.26, 3.72 ± 1.91, 3.74 ± 2.38) in de novo acute leukemia patients and lower (relative expressive level: 2.14 ± 1.47, 2.07 ± 0.99, 1.47 ± 0.89) in the acute leukemia patients with CR (p < 0.05). The expressions of LSPC related genes were lower (relative expressive level: 1.77 ± 1.29, 2.09 ± 1.26, 1.78 ± 1.49) in the patients acquired CR/partial remission (PR) than those in the patients not acquired CR/PR (relative expressive level: 7.23 ± 1.78, 3.96 ± 0.92, 4.48 ± 2.57) (p < 0.01). Univariate analysis revealed that there were more cases with the expression of LSPC immunophenotype (CD34(+)CD38(-)CD96(+) and CD34(+)CD38(-)CD123(+)) and more hyperleukocytosis cases in patients with any higher expression of LSPC related gene (p < 0.05). Analysis of multiple parameters discovered larger significance (p < 0.01). It is concluded that there is a good relationship between LSPC related genes (ABCB1, BMI-1, HOXB4) and LSPC immunophenotype. The expression of LSPC-related genes is higher in de novo acute leukemia patients, and lower in patients acquired CR/PR. The patients with higher expressed LSPC-related genes display worse response to chemotherapy, lower CR/PR rate and higher leukocytosis, the analysis of multiple parameters may be a good method for assessing the therapeutic efficacy/prognosis of acute leukemia.