The Effects and Regulatory Mechanism of Targeting CXC Chemokine Receptor 1/2 Combined with Ara-C on the Malignant Biological Behaviors of U937 Cells of Acute Myeloid Leukemia / 中国实验血液学杂志

OBJECTIVE@#To investigate and analyze the effect of CXC chemokine receptor 1/2 (CXCR1/2) targeting inhibitor Reparixin combined with cytarabine (Ara-C) on the malignant biological behaviors of acute myeloid leukemia cells and its effect on the expression of the CXCR family, while exploring the accompanying molecular mechanism, providing scientific basis and reference for new molecular markers and targeted therapy for AML.@*METHODS@#Acute myeloid leukemia U937 cells were treated with different concentrations of Reparixin, Ara-C alone or in combination, and the cell morphology was observed under an inverted microscope; Wright-Giemsa staining was used to detect cell morphological changes; CCK-8 method was used to detect cell proliferation; the ability of cell invasion was detected by Transwell chamber method; the ability of colony formation was detected by colony formation assay; cell apoptosis was detected by Hoechst 33258 fluorescent staining and Annexin V/PI double-staining flow cytometry; monodansylcadaverine(MDC) staining was used to detect cell autophagy; the expression of apoptosis, autophagy and related signaling pathway proteins was detected by Western blot and the expression changes of CXCR family were detected by real-time quantitative polymerase chain reaction (qRT-PCR).@*RESULTS@#Reparixin could inhibit the proliferation, invasion, migration and clone formation ability of U937 cells. Compared with the single drug group, when U937 cells were intervened by Reparixin combined with Ara-C, the malignant biological behaviors such as proliferation, invasion and colony formation were significantly decreased, and the levels of apoptosis and autophagy were significantly increased (P<0.01). After Reparixin combined with Ara-C intervenes in U937 cells, it can up-regulate the expression of the pro-apoptotic protein Bax and significantly down-regulate the expression of the anti-apoptotic protein Bcl-2, and also hydrolyze and activate Caspase-3, thereby inducing cell apoptosis. Reparixin combined with Ara-C could up-regulate the expressions of LC3Ⅱ and Beclin-1 proteins in U937 cells, and the ratio of LC3Ⅱ/LC3Ⅰ in cells was significantly up-regulated compared with single drug or control group (P<0.01). MDC result showed that the green granules of vesicles increased significantly, and a large number of broken cells were seen (P<0.01). Reparixin combined with Ara-C can significantly inhibit the phosphorylation level of PI3K, AKT and NF-κB signaling molecule, inhibit the malignant biological behavior of cells by inhibiting the activation of PI3K/AKT/NF-κB pathway, and induce programmed cell death. Ara-C intervention in U937 cells had no effect on the expression of CXCR family (P>0.05). The expression of CXCR1, CXCR2, and CXCR4 mRNA could be down-regulated by Reparixin single-agent intervention in U937 cells (P<0.05), and the expression of CXCR2 was more significantly down-regulated than the control group and other CXCRs (P<0.01). When Reparixin and Ara-C intervened in combination, the down-regulated levels of CXCR1 and CXCR2 were more significant than those in the single-drug group (P<0.01), while the relative expressions of CXCR4 and CXCR7 mRNA had no significant difference compared with the single-drug group (P>0.05).@*CONCLUSION@#Reparixin combined with Ara-C can synergistically inhibit the malignant biological behaviors of U937 cells such as proliferation, invasion, migration and clone formation, and induce autophagy and apoptosis. The mechanism may be related to affecting the proteins expression of Bcl-2 family and down-regulating the proteins expression of CXCR family, while inhibiting the PI3K/AKT/NF-κB signaling pathway.

Experimental Study on the Mechanism of Mangiferin Inhibiting Malignant Biological Characteristics of Multiple Myeloma and Exerting Anticancer Effect / 中国实验血液学杂志

OBJECTIVE@#To investigate the effect of pure Chinese herbal extract Mangiferin on the malignant biological behaviors of multiple myeloma (MM) cells, and to analyze the molecular mechanism of the anti-myeloma effect of Mangiferin, so as to provide experimental basis for MM replacement therapy.@*METHODS@#U266 and RPMI8226 of human MM cell lines were intervened with different concentrations of Mangiferin. Cell proliferation was detected by CCK-8 method. Annexin V/PI double staining flow cytometry was used to detect cell apoptosis. Western blot was used to detect the expression of apoptosis and related signaling pathway proteins, and real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of matrix metalloproteinase (MMP) and CXC chemokine receptor (CXCR) family.@*RESULTS@#Mangiferin could inhibit the proliferation activity of U266 and RPMI8226 cells and induce cells apoptosis. After Mangiferin intervened in U266, RPMI8226 cells for 48 h, the expression of Bcl-2 family pro-apoptotic protein Bax was up-regulated, while the expression of survivin and Bcl-xL proteins was down-regulated and caspase-3 was hydrolyzed and activated to promote cell apoptosis, besides, the expression of Bcl-2 protein in U266 cells was also significantly down-regulated to induce apoptosis (P<0.05). After Mangiferin intervenes in MM cells, it can not only increase the expression level of tumor suppressor p53, but also induce programmed cell death of MM cells by inhibiting the expression of anti-apoptotic molecules and down-regulating the phosphorylation levels of AKT and NF-κB. In addition, after the intervention of Mangiferin, the expressions of CXCR4, MMP2 and MMP9 in U266 cells were down-regulated (P<0.05), while there is no effect on the expressions of CXCR2, CXCR7 and MMP13 (P>0.05). However, the expressions of CXCR4, MMP9, and MMP13 in RPMI8226 cells were down-regulated (P<0.01), the expression of MMP2 was weakly affected, and the expression of CXCR2 and CXCR7 was basically not affected (P>0.05).@*CONCLUSION@#Mangiferin can inhibit the proliferation and induce apoptosis of MM cells, and its mechanism may be related to inhibiting the activation of NF-κB signaling pathway, affecting the expression of Bcl-2 family proteins, and inhibiting the expression of core members of MMP and CXCR family.

Research advances of small molecule inhibitors in the treatment of mixed lineage leukemia / 药学学报

Acute leukemia (AL) is a kind of malignant clonal disease of hematopoietic stem cells. Rearrangement of mixed lineage leukemia (MLL) gene can be observed in about 5%-10% of AL patients. Currently, AL patients with MLL-rearrangements (MLL-r) lack effective treatment and are usually associated with poor prognoses. Recent studies have shown that many epigenetic regulators are directly or indirectly involved in the occurrence and development of AL carrying MLL-r (MLL), which provides a biological basis for the use of epigenetic regulation strategies to treat MLL. In this review, we start from the epigenetic regulation mechanism of MLL, and select representative drug targets to briefly analyze the relationship between each target and MLL and summarize the development progress of their inhibitors, hoping to provide reference for the subsequent research and development of drugs for the treatment of MLL.

Effect of staging treatment of / 中国针灸

OBJECTIVE@#To compare the effect of @*METHODS@#A total of 74 patients with RIF of thin endometrium type undergoing freeze-thaw embryo transfer were randomly divided into an observation group (37 cases) and a control group (37 cases). The patients in the control group were treated with freeze-thaw embryo transfer in hormone replacement cycle, and the estradiol valerate tablets were taken orally from the fifth day of menstruation, 2 mg per day. On the basis of the control group, the observation group was additionally treated with @*RESULTS@#The clinical pregnancy rate was 37.8% (14/37) in the observation group, which was higher than 16.2% (6/37) in the control group (@*CONCLUSION@#On the basis of conventional medication,

Effects of Combined Infusion of Mesenchymal Stem Cells and Endothelial Progenitor Cells on Lung Injury after Hematopoietic Stem Cell Transplantation / 中国实验血液学杂志

OBJECTIVE@#To investigate the effects of combined infusion of mesenchymal stem cells (MSC) and endothelial progenitor cells (EPC) on lung injury after hematopoietic stem cell transplantation (HSCT).@*METHODS@#The experiment was divided into normal control group, irradiation group, bone marrow cell transplantation group (BMT group), BMT+EPC group, BMT+MSC group and BMT+EPC+MSC group. The model of HSCT was established, on the 30th day after transplantation, the mice were sacrificed. Then lung tissue was taken for testing. The mRNA expression levels of VEGF, IL-18, IL-12b were detected by RT-PCR, and protein expression level of NLRP3 was detected by Western blot. The expression of MPO and CD146 was observed by immunohistochemistry assay.@*RESULTS@#The expression level of VEGF gene in BMT+EPC+MSC group was significantly higher than that in other groups (P＜0.01). The expression level of IL-18 and IL-12b gene was the highest in BMT group and the lowest in BMT+EPC+MSC group, and the difference was statistically significant (P＜0.05). HSCT could increase the expression of NLRP3 protein, and the BMT+EPC+MSC could significantly reduce the level of NLRP3 protein in lung cells, tending to normal. Compared with normal tissues, the BMT+EPC+MSC could improve the lung tissue structure more effectively, the expression of MPO positive cells was lower, and the expression of VEGF positive cells was higher.@*CONCLUSIONS@#The combined infusion of MSC and EPC can promote capillary regeneration, alleviate inflammation and promote lung repair after HSCT, which is superior to single EPC or MSC infusion.

Effects of Different Stimulators on Proliferation of Peripheral Blood Lymphocyte Subsets / 中国实验血液学杂志

OBJECTIVE@#To investigate the effects of different stimultors (PHA, PMA and IL-2) and culture systems (PBMC and whole blood) on the proliferation of human peripheral blood lymphocyte subsets, so as to provide the experimental basis for selecting the appropriate system according to the experimental purposes.@*METHODS@#A total of 10 ml serum samples were collected from healthy volunteers (n=6). The 300 μl whole blood was directly used to detect lymphocyte subsets by flow cytometry. The 400 μl whole blood were inoculated respectively with 3 different stimulators at 37℃ and 5% CO2 for 60 h; Three different stimulators were also added to the PBMC which were isolated from 2 ml whole blood. Then the proliferation ability of lymphocyte subsets was analyzed by flow cytometry.@*RESULTS@#After the PBMC were stimulated with PHA, CD4CD8CD3 lymphocytes were the most subset; The proportion of CD3CD4 T lymphocytes and CD3CD19 B lymphocytes decreased after being stimulated by PMA (P<0.01, P<0.05); the lymphocyte subset ratio had no significant change after being stimulated by IL-2. After the whole blood system was stimulated with PHA, the CD4/CD8 T lymphoblasts were main subsets, the counts of B lymphocytes and NK cells were reduced; after being stimulated with PMA, the number of CD8CD3 T lymphoblast and CD4CD8T lymphocytes increased, the B/NK cells were not distinguished with the surface markers; after the whole blood system was stimulated with IL-2, the proportion of NK cells significantly increased (P<0.05).@*CONCLUSION@#Lymphocyte proliferation stimulated by PMA is the fastest, while the effect of IL-2 on the lymphocyte subset proportion stimulated by IL-2 is the minimal. After being stimulated by PHA the division cycles of lymphocyte are the most.

In Vitro Efficacy of Six Alternative Antibiotics against Multidrug Resistant Escherichia Coli and Klebsiella Pneumoniae from Urinary Tract Infections

<p><b>INTRODUCTION</b>Increasing resistance in Escherichia coli and Klebsiella pneumoniae to firstline antibiotics makes therapeutic options for urinary tract infections (UTIs) challenging. This study investigated the in vitro efficacies of 6 antibiotics against multidrug resistant (MDR) uropathogens.</p><p><b>MATERIALS AND METHODS</b>Minimum inhibitory concentrations to ceftibuten, cefpodoxime, fosfomycin, mecillinam, temocillin, and trimethoprim were determined against 155 MDR-isolates of E. coli and K. pneumoniae. The presence of extended-spectrum beta-lactamases (ESBL) and plasmid-borne AmpC enzymes was determined by phenotypic testing with genotyping performed by multiplex polymerase chain reaction.</p><p><b>RESULTS</b>Temocillin demonstrated highest susceptibility rates for both E. coli (95%) and K. pneumoniae (95%) when breakpoints for uncomplicated UTIs were applied; however, temocillin susceptibility was substantially lower when "systemic infection" breakpoints were used. Fosfomycin demonstrated the best in vitro efficacy of the orally available agents, with 78% and 69% of E. coli and K. pneumoniae isolates susceptible, respectively. The next most effective antibiotics were ceftibuten (45%) and mecillinam (32%). ESBL and ampC genes were present in 47 (30%) and 59 (38%) isolates.</p><p><b>CONCLUSION</b>This study demonstrated few oral therapeutic options for MDR-uropathogens, with fosfomycin demonstrating the best in vitro activity.</p>

Expression of Bim, Bax and Bak in the process of gingipain-induced osteoblast apoptosis / 中华口腔医学杂志

<p><b>OBJECTIVE</b>To establish osteoblast apoptosis model induced by gingipains, and to examine the expression of pro-apoptotic protein Bcl-2 interacting mediator (Bim), Bcl-2 associated X protein (Bax) and Bcl-2 antagonist/killer (Bak).</p><p><b>METHODS</b>Gingipain and gingipain acticity were extracted and measured. Mouse osteoblast cell line MC3T3-E1 cells were cultured in the presence of 0.453, 0.906, 1.812 U/L gingipains for 0, 16, 24 and 48 h. Apoptosis was examined by 4',6-diamidino-2-phenylindole (DAPI) staining or annexin V/propidine iodide (PI) staining.Protein expression of Bim, Bax and Bak was determined by Western blotting after osteoblasts were cultured with 1.812 U/L gingipain for 0, 4, 8, 16, 24 and 48 h. Osteoblasts were cultured with 1.812 U/L gingipain which had been inhibited with N-alpha-tosyl L-lysyl-chlorom ethylketone (TLCK). Western blotting was used to detect Bim expression and DAPI staining to measure apoptosis.</p><p><b>RESULTS</b>Arginine-specific proteinases (Rgp) activity was (18.11 ± 2.11) U/L and specific proteinases (Kgp) was (1.02 ± 0.25) U/L. Percentage of osteoblast apoptosis induced by 1.812 U/L gingipain rose to (6.31 ± 0.37)% after 16 h, and reached (11.20 ± 0.35)% at 24 h and (10.80 ± 0.46)% after 48 h with DAPI staining. Annexin V/PI staining supported the result from DAPI staining.Bim protein level increased during osteoblast apoptosis, the relative fold rose to (0.31 ± 0.03) after 4 h (about 2 fold compared to control), peaking at 24 h (0.57 ± 0.05, 3-4 fold compared to control). Proteinase inhibitor TLCK effectively blocked the activity of gingipain and inhibited up-regulation of Bim induced by gingipains from (0.58 ± 0.04) to (0.14 ± 0.03). The percentage of osteoblast apoptosis decreased from (11.20 ± 0.35)% to (4.31 ± 0.38)% in the presence of TLCK. Expression of Bax remained unchanged when cells were cultured with or without gingipains. Bak was under the detectable level in MC3T3-E1.</p><p><b>CONCLUSIONS</b>1.812 U/L gingipains induced osteoblast apoptosis. Protein expression of Bim was up-regulated during cell apoptosis and was down-regulated when gingipain inhibited with TLCK, suggesting that Bim was involved in osteoblast apoptosis induced by gingipain. Inhibition of Bim protein expression protected osteoblast from apoptosis.</p>

Kinetic parametric estimation in animal PET molecular imaging based on artificial immune network / 中华核医学杂志

Objective To develop an accurate,reliable method without the need of initialization in animal PET modeling for estimation of the tracer kinetic parameters based on the artificial immune network.Methods The hepatic and left ventricular time activity curves (TACs) were obtained by drawing ROIs of liver tissue and left ventricle on dynamic 18F-FDG PET imaging of small mice.Meanwhile,the blood TAC was analyzed by sampling the tail vein blood at different time points after injection.The artificial immune network for parametric optimization of pharmacokinetics (PKAIN) was adapted to estimate the model parameters and the metabolic rate of glucose (Ki) was calculated.Results TACs of liver,left ventricle and tail vein blood were obtained.Based on the artificial immune network,Ki in 3 mice was estimated as 0.0024,0.0417 and 0.0047,respectively.The average weighted residual sum of squares of the output model generated by PKAIN was less than 0.0745 with a maximum standard deviation of 0.0084,which indicated that the proposed PKAIN method can provide accurate and reliable parametric estimation.Conclusion The PKAIN method could provide accurate and reliable tracer kinetic modeling in animal PET imaging without the need of initialization of model parameters.

Two flavonoids from Lagopsis supina / 药学学报

<p><b>AIM</b>To study the chemical constituents of Lagopsis supina.</p><p><b>METHODS</b>Compounds were isolated by column chromatography of silica gel and Sephadex LH-20, and the structures were determined by spectral analysis.</p><p><b>RESULTS</b>Two compounds were isolated and elucidated as apigenin-7-O-(6"-(E)-p-coumaroyl)-beta-D-galactopyranoside (I) and apigenin-7-O-(3",6"-di-(E)-p-coumaroyl)-beta-D-galactopyranoside (II).</p><p><b>CONCLUSION</b>I and II are new compounds.</p>