Influence of Oridonin on the Icilling Acitivity of NK-92 MI Cells Targeting Cell THP1 and Its Mechanism / 中国实验血液学杂志

OBJECTIVE@#To investigate the influence of oridonin on the killing activity of NK-92 MI cells targeting THP1 and the related mechanism.@*METHODS@#The killing activity of NK-92 MI to THP1 before and after oridonin treatment was detected by LDH release assay; the expression of natural killer cell ligands activating receptor D (NKG2D, including MICA, MICB, ULBP1, ULBP2 and ULBP3) was detected by real-time quantitative polymerase chain reaction (qRT-PCR) and Western blot respectively; the expression of cytokine TNF-α, TNF-β and IFN-γ in the co-culture supernatant of NK-92 MI cells and THP1 cells were measured by ELISA.@*RESULTS@#The killing efficiency after oridonin treatment at different effector-target ratio (1:1, 5:1, 10:1) was all significantly up-regulated in comparison with that before oridonin treatment (P＜0.05). QRT-PCR and Western blot showed that the expressions of mRNA and protein levels of MICB, ULBP1, ULBP2 increased to varying degree (P＜0.05), but the expression levels of MICA and ULBP3 were not statistically significant between experimental group and control group (P＞0.05). ELISA results indicated that IFN-γ and TNF-β release were significantly increased after oridonin treatment (P＜0.05), however, the TNF-α release was not statistically different in comparison with control group (P＞0.05).@*CONCLUSION@#Oridonin can significantly improve killing efficiency of NK-92 MI on THP1, that might be related with up-regulation of MICB, ULBP1 and ULBP2 expression and promotion of IFN-γ and TNF-β release.

Effect of PD-L1 Expression on Activity of NK Killing AML Cell Lines and Its Mechanisms / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To expolore the effect of programmed death receptor ligand 1 (PD-L1) expression level on killing effect of different cell lines of acute myeloid leukemia (AML) and its possible mechanism.</p><p><b>METHODS</b>Peripheral blood from healthy individuals was collected routinely; NK cells were isolated using immunomagnetic beads; PD-L1 expression level was detected by flow cytometry; the killing effect of NK cells on acute myelogenous leukemia cell lines was evaluated with LDH release method and monoclonal antibody blocking experiment; the expression levels of IFN-γ and IL-2 in the supernatants from the co-cultured effector/targer cells were measured by ELISA.</p><p><b>RESULTS</b>The ratio of CD3CD56NK cells increased from （12.44±3.48）% to (71.29±5.65）%. The flow cytometry showed that KG-1a cells lowly expressed PD-L1 (8.35±4.12)%, but the THP cells a highly expressed PD-L1 (76.42±26.54)%. Meanwhile, the NK cells displayed a more efficient killing effect on KG-1a cells than that of THP1 cells (P<0.05). Moreover, PD-L1 monoclonal antibody could reinforce NK cell killing effect and, promote the secretion of IFN-γ and IL-2 in 5 acute myelogenous leukemia cell lines to varying degree.</p><p><b>CONCLUSION</b>The killing effect of NK cells on acute myelogenous leukemia cell line is inversely proportional to PD-L1 expression; blocking PD1/PD-L1 binding can significantly enhance the killing efficiency of effector-target cells, which way be related with promoting the release of IFN-γ and IL-2.</p>

Role of voltage-gated sodium channel in tumor metastasis / 国际药学研究杂志

The metastasis is the most important biological characteristic of malignant tumors. Once distant metastasis occurs in the tumor tissue, this usually means that the tumor has entered the advanced stage, so it is difficult to cure with local treatment alone, which is the main cause of death. It has been found that voltage-gated sodium channels (VGSC) are expressed not only in excitable cells but also in many metastatic cells, particularly in certain types of cancer cells and the expression of VGSC is related to cancer migration, invasion and metastasis in vivo. Therefore, this article reviews recent studies on the VGSC in tumor invasion and metastasis.

Effect of Guiqiyiyuan ointment on bystander effect in rat models of 12C6+ beam radiation damage / 解放军医学杂志

Objective To investigate the bystander effect injury to lung-heart-liver-spleen caused by 12C6+ beam radiation,and explore the prevention and treatment effect of Guiqiyiyuan ointment on the injury and its mechanism.Methods Ninety healthy male Wistar rats were randomly divided into 3 groups:NC group (normal control,normal saline 2ml/kg,n=30),SR group (simple radiation,8Gy,2ml/kg,n=30),GO group (Guiqiyiyuan ointment 11.83g/kg,radiation,8Gy,n=30).All the rats received intragastric administration for 7 days.The right side of the lung was modeled by 12C6+ beam radiation.After modeling,the rats were killed at 48h.The heart,liver and spleen were taken.The malonaldehyde (MDA),glutathione (GSH),glutathione peroxidase (GSH-Px),superoxide dismutase (SOD) contents were measured by colorimetry,DNA methylation rate was assayed by ELISA,and the expressions of Dnmt1,Dnmt3a and Dnmt3b were detected by immunohistochemistry.Results Compared with NC group,the contents of SOD,GSH and GSH-Px decreased (P＜0.01),MDA increased (P＜0.01),the level of DNA methylation decreased (P＜0.01),and the expressions of Dnmt1,Dnmt3a and Dnmt3b increased in SR group (P＜0.01).Compared with SR group,the contents of SOD,GSH and GSH-Px increased (P＜0.01),MDA decreased (P＜0.01),the level of DNA methylation increased (P＜0.01),and the expressions of Dnmt1,Dnmt3a and Dnmt3b decreased in GO group (P＜0.01).Dnmtl,Dnmt3a and Dnmt3b proteins were expressed in the cytoplasm of myocardial cells,hepatocytes and peripheral B cells of the white pulp in spleen,in all the groups.The color of NC group was light brown-brown,showing a weak positive expression.The color of SR group was brown-brown,showing a strong positive expression.The color of GO group was light brown-tan,showing a moderate positive expression.Conclusion The Guiqiyiyuan ointment can reduce the bystander effect caused by the 12C6+ beam radiation,and its mechanism is related to improving the oxidative stress reaction and the level of DNA methylation.

Effect of Silencing SET Gene on Acute Promyelocytic Leukemia NB4-R1 Cells / 中国实验血液学杂志

<p><b>OBJECTIVE</b>To investigate the effect of silencing SET gene on the biological characteristics of acute promyelocytic leukemia NB4-R1 cells.</p><p><b>METHODS</b>The expression vector of pGCSIL containing SET-shRNA were transfected into 293T cells by using other packaging plasmids. The supernatant of the 293T cells was harvested for lentivirus. The SET-shRNA lentiviral vector was transfected into acute promyelocytic leukemia NB4-R1 cells and a stably transfected cell line was established. Real-time quantitative PCR and Western blot were used to assay the silencing efficiency on SET gene and the expression of PP2A. The cell cycle distribution was tested by flow cytometry.</p><p><b>RESULTS</b>The expression of SET in experimental group statistically decreased as compared with that of the control group. The expression of PP2A was obviously raised at the level of mRNA and protein. The percentage of NB4-R1 cells in G0/G1 phase significantly increased, while the percentage of cells in S phase significantly decreased.</p><p><b>CONCLUSION</b>The silencing gene in acute promyelocytic leukemia NB4-R1 cells using SET-shRNA lentiviral vector can increase the expression of PP2A and interfere of the cell cycle in NB4-R1 cells. This study has laid a experimental base for targed therapy of patients with acute promyelocytic leukemia.</p>

A multi-center, randomized, double-blind clinical study on Bushen Huoxue in treatment of ovulatory dysfunction caused infertility / 中国中药杂志

<p><b>OBJECTIVE</b>The multi-center, randomized, double-blind, double-simulated and positive-control trial was used to verify the contribution degree of Bushen Huoxue for the treatment of ovulatory dysfunction caused infertility, which provided scientific basis for clinical treatment.</p><p><b>METHOD</b>According to diagnostic, inclusion and exclusion criteria, we observed 349 patients which were divided into the treated group (n = 177, treated with Bushen Huoxue ricipe) and control group (n = 172, treated with clomiphene). Ovulation rate, pregnancy rate, clinical effective rate of traditional Chinese medicine, endometrium and diameter of dominant follicle were observed. Serum reproductive endocrine hormones were assayed before and after treatment.</p><p><b>RESULT</b>The treated group showed ovulation rate of 69.34%, with pregnancy rate of 41.35%. The clinical effective rate of treated group and control group were 91.73% and 80.77%. There was remarkable difference in endometrium (P < 0.05) and remarkbale difference in sex hormones PRL and E₂in treated group at prior-treatment and post-treatment (P < 0.05). No adverse effects were found in the experiment. Security indicators did not show abnormal change.</p><p><b>CONCLUSION</b>The comparison between the two groups showed that the treated group was significantly different from control group in the pregnancy rate (P < 0.05), without notable difference in ovulation rate. There was significant difference in clinical effective rate between the treated group and control group. Both the two groups could contribute to the mature development and discharge of the follicles. The growth of endometrium and endometrial receptivity in the treated group were higher than control group. The treated group has regulatory effect on PRL and E₂.</p>

Establishment of the human acute promyelocytic leukemia pathological model in the SCID beige mice by using NB4 cell line / 中国实验血液学杂志

<p><b>OBJECTIVE</b>This study was to establish a stable, effective and reproducible human acute promyelocytic leukemia model in severe combined immunodeficient (SCID) mice by using NB4 cell line, and to investigate the disease course character and biological behaviors.</p><p><b>METHODS</b>Three-five-week-old SCID beige mice were divided randomly into two groups: experimental and control group. SCID mice of experimental group were transplanted by tail vein (iv) injection of 5×10(6) NB4 cells. The WBC cell count and the positive rate of promyelocytes in peripheral blood were dynamically monitored by using smears. Morphological examination and histopathological assay were employed to confirm NB4 cell infiltration in organs (liver, spleen, lung, kidney and brain). The expression level of PML-RARα fusion protein was detected by Western blot.</p><p><b>RESULTS</b>Within two weeks there was no significant difference in peripheral blood WBC count between two groups (P > 0.05), meanwhile, NB4 cells were not found. At the day 21 and 28 after inoculation, the peripheral blood white blood cell count of experimental group reached to (4.79 ± 1.13)×10(9)/L and (7.62 ± 2.24)×10(9)/L respectively, which were significantly higher than that in control group (P < 0.05); simultaneously, the positive rates of promyelocytes on smears were (2.14 ± 0.63)% and (6.6 ± 2.76)%, respectively. Morphological observation showed single or multiple tumor lumps at day 21 after inoculation; HE staining of tissue biopsies demonstrated a large number of promyelocyte in the liver, spleen, lung, kidney and brain tissue. Cell immunofluorescence results showed that the CD33 expression of bone marrow cells in mice of experimental group was strongly positive (P < 0.05). Western blot confirmed that the PML-RARα fusion protein was expressed variously in liver, kidney and brain tissue.</p><p><b>CONCLUSIONS</b>The human acute promyelocytic leukemia SCID mouse model is succesfully established by tail vein injection of NB4 cells. This model can mimic the characters of involved bone marrow and diffuse growth of cells. This model is a useful tool to explore the pathogenic mechanism and experimental treatment of human leukemia.</p>

Apoptosis of retinoic acid-resistant APL NB4-R1 cells induced by As4S4 and its molecular mechanism / 中国实验血液学杂志

<p><b>OBJECTIVE</b>This study was to investigate the apoptosis-inducing effect of As(4)S(4) on the retinoic acid-resistant acute promyelocytic leukemia (APL) NB4-R1 cells and its potential mechanisms.</p><p><b>METHODS</b>The leukemia cell line NB4-R1 was cultured in vitro and divided into control group and treatment group. The apoptosis rate and cell cycle were detected by flow cytometry. The apoptotic DNA fragments were analyzed by agarose gel electrophoresis. The changes of BCL-2, BAX and Caspase-3 were determined by Western blot.</p><p><b>RESULTS</b>After NB4-R1 cells were treated with As(4)S(4)(25 µmol/L) for 0 h, 24 h, 48 h, the percentage of early apoptotic cells was obviously raised from 0% to 24.49% and 47.41%, the percentage of late apoptotic cells were elevated from 0.08% to 14.72% and 20.70%. Compared with control group, the DNA degradation revealed a characteristic DNA ladder during agarose gel electrophoresis after treatment for 24 h. The drug significantly induced an accumulation of the S phase cell population from 31.85% of the untreated cells to 42.53% and 55.12% treated with the different time whereas the NB4-R1 cells in G0/G1 phase decreased from 57.30% to 37.56% and 28.51%. As(4)S(4) could decrease the expression of BCL-2 and increase the level of BAX. Pro-caspase-3 could be cleaved into small active fragments under the apoptotic stimulation.</p><p><b>CONCLUSION</b>As(4)S(4) can efficiently induce NB4-R1 cell apoptosis, which may be related with the down-regulation of BCL-2 and the up-regulation of BAX, as well as the activation of Caspase-3.</p>

Efficient reprogramming of human cord blood CD34(+) cells for formation of induced pluripotent stem cells with non-integrating plasmid system / 中国实验血液学杂志

This study was to establish the episomal vector reprogramming method to reprogram iPSC from human cord blood (CB) CD34(+) cells. The non-integrating plasmids of pEB-C5 and pEB-Tg were transfected into short-term cultured CB CD34(+) cells by using the nucleofector, so as to demonstrate efficient reprogramming of CB CD34(+) cells. Within 14 days of one-time transfection by two plasmids together, up to 200 iPSC-like colonies per 2 million transfected CB CD34(+) cells were generated. The results showed that the pluripotency of iPSC-derived CB CD34(+) cells was similar to that of hESC and the karyotypes of iPSC were normal. In addition, no vector integration was found in iPSC of 9th and 10th passages. Furthermore, hiPSC formed teratoma with three embryonic germ layers. It is concluded that the integration-free method to generate human iPSC from CB CD34(+) cells is reliable and can provide new ways for both research and future clinical applications.

Effect on expression of vascular smooth muscle cell calcium channel in rats exposed in low temperature / 中华劳动卫生职业病杂志

<p><b>OBJECTIVE</b>To observe the expression of vascular smooth muscle cells calcium channel α1C subunit (LTCCα1C) in rats exposed in low temperature.</p><p><b>METHODS</b>Cold-induced hypertension was established and blood pressure was measured every two weeks. The mRNA expression of L type calcium channel α1C was determined by RT-PCR.</p><p><b>RESULTS</b>The blood pressure of the rats exposed to cold environment increased. The blood pressure of experimental groups [(102.8 ± 2.25) mm Hg] began to increase from the first two weeks, compared with the control group [(89.2 ± 3.73) mm Hg], there were significant difference (P < 0.05). The blood pressure of experimental groups were (114.5 ± 4.21), (121.9 ± 3.03) mm Hg respectively at 4, 6 weeks. Compared with the control group, the expression of LTCCα1C mRNA of the cold exposure group increased significantly (P < 0.01). There was a significant correlation between the expression of LTCCα1C mRNA and the blood pressure of the rats (r = 0.86, P < 0.01).</p><p><b>CONCLUSION</b>Repeated cold exposure can establish cold-induced hypertension, and the level of vascular smooth muscle cells LTCCα1C expression increase.</p>

Study of differential proteins in lung adenocarcinoma using laser capture microdissection combined with liquid chip-mass spectrometry technology / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>In recent years the proportion of lung adenocarcinoma (adCA) which occurs in lung cancer patients has increased. Using laser capture microdissection (LCM) combined with liquid chip-mass spectrometry technology, we aimed to screen lung cancer biomarkers by studying the proteins in the tissues of adCA.</p><p><b>METHODS</b>We used LCM and magnetic bead based weak cation exchange (MB-WCX) to separate and purify the homogeneous adCA cells and normal cells from six cases of fresh adCA and matched normal lung tissues. The proteins were analyzed and identified by matrix assisted laser desorption/ionization time-of-fight mass spectrometry (MALDI-OF-MS). We screened for the best pattern using a radial basic function neural network algorithm.</p><p><b>RESULTS</b>About 2.895 × 10(6) and 1.584 × 10(6) cells were satisfactorily obtained by LCM from six cases of fresh lung adCA and matched normal lung tissues, respectively. The homogeneities of cell population were estimated to be over 95% as determined by microscopic visualization. Comparing the differentially expressed proteins between the lung adCA and the matched normal lung group, 221 and 239 protein peaks, respectively, were found in the mass-to-charge ration (M/Z) between 800 Da and 10 000 Da. According to t test, the expression of two protein peaks at 7521.5 M/Z and 5079.3 M/Z had the largest difference between tissues. They were more weakly expressed in the lung adCA compared to the matched normal group. The two protein peaks could accurately separate the lung adCA from the matched normal lung group by the sample distribution chart. A discriminatory pattern which can separate the lung adCA from the matched normal lung tissue consisting of three proteins at 3358.1 M/Z, 5079.3 M/Z and 7521.5 M/Z was established by a radial basic function neural network algorithm with a sensitivity of 100% and a specificity of 100%.</p><p><b>CONCLUSIONS</b>Differential proteins in lung adCA were screened using LCM combined with liquid chip-mass spectrometry technology, and a biomarker model was established. It is possible that this technology is going to become a powerful tool in screening and early diagnosis of lung adCA.</p>

Expression of synaptophysin in experimental diffuse brain injury / 法医学杂志

OBJECT@#To investigate the changes in the expression_level of synaptophysin following diffuse brain injury (DBI) in rats and to correlate the changes of the synaptophysin expression_level with the post injury time interval.@*METHODS@#Wister rats were used as a DBI model induced by Marmarou method. The changes of synaptophysin immunoreactivity on coronal sections of the rats sampled at different post-injury time intervals were used as a marker. The densitometry of the synaptophysin immunoreactivity was documented by imaging technique and analyzed by SPSS software.@*RESULTS@#The expression level of synaptophysin in DBI rats showed dynamic changes following DBI as well as during the repairing period.@*CONCLUSION@#The changes of synaptophysin level may be used as a marker for estimation of the post injury time interval in DBI.

Time-dependent expression of astroglial S100beta following diffuse brain injury in rats / 法医学杂志

OBJECTIVE@#To investigate the dynamics of the induction of S100beta in different parts of rat brain following the diffuse brain injury.@*METHODS@#Immunohistochemistry and auto-image analysis were to determine the expression of astroglial S100beta after diffuse brain injury in rats. Forty rats were distributed into groups according to injury time of 30min, and2,4,12,24h, and 3,6 d after diffuse brain injury, and normal rats as control.@*RESULTS@#The number of S100beta positive cells in the four areas increased significantly followed by a decrease, and then a further increase. The expression of S100beta could be detected increasing in 2h, and increased significantly in 4h, and it reached apex 12h after DBI, and decreased gradually to the level less than normal 3d, and returned to normal 7d following injury. In the postmortem injury groups, there were no significant changes in anti-S100beta immunoreactivities in four areas of brain compared to the control group.@*CONCLUSION@#The present study showed the time-dependent expression of S100beta is obvious following diffuse brain injury, and suggested S100beta be suitable as a marker for brain injury age determination.

Clinical value of esophageal mucosal iodine stain during esophagoscopy for patients without swallowing symptoms / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the clinical value of esophageal mucosal iodine stain during esophagoscopy for patients with early esophageal carcinoma or precancerous lesions without swallowing symptoms, through analyzing the correlation between endoscopic findings and pathological results of biopsy on the suspicious spots.</p><p><b>METHODS</b>For 366 patients examined by iodine stain during esophagoscopy, the position, size, shape and boundary of all visible unstained lesions were recorded and multiple biopsies were taken on the unstained spots.</p><p><b>RESULTS</b>Before iodine stain, 462 lesions had been discovered in 366 patients. However, 478 abnormal lesions stained in 341 patients were detected after iodine stain, the remaining 25 gave no abnormal findings. More than 1/3 of patients were found to have more than 2 abnormally stained lesions. 28.4% of them (104 cases) had moderate or severe dysplasia or early esophageal cancer. The sensitivity of iodine stain in this series was 89.8%.</p><p><b>CONCLUSION</b>Iodine stain is very useful in detecting occult early esophageal carcinoma and precancerous lesions. The degree of coloration and the margin of suspicious spots are closely correlated with the pathological results.</p>