Objective:To investigate the inhibitory effects of melanoma differentiation associated gene-7(mda-7/IL- 24)on lymphoma cell line Namalwa in vitro and in vivo.Methods:Using RT-PCR,the expression of mda-7/IL-24 was examined in 10 malignant hematopoietic cell lines,including Namalwa,Raji,K562,NB4,U937,Ramous,CEM,KG1a, HL60,J6-1,etc.The coding region of mda-7/IL-24 was cloned from LPS-treated human peripheral blood mononuclear cells(PBMC)by RT-PCR,and the eukaryotic expression vector pTarget-IL-24 was constructed.The recombinant vector, after sequenced,was transfected into Namalwa cell line via lipofectamine reagent.The stable expression transfectants were selected by G418.The expression of mda-7/IL-24 mRNA and protein was verified by RT-PCR and Western blotting.MTT assay,colony forming assay,apoptosis detection,and tumorigenesis in nude mice were used to assess the effects of mda- 7/IL-24 on tumor proliferation,growth characteristics,colony forming,apoptosis,and tumorigenesis.Results:Expression of mda-7/IL-24 mRNA was not found in any of the 10 malignant hematopoietic cell lines and the expression of mda-7/IL- 24 mRNA and protein was found in Namalwa cells transfected with recombinant plasmid pTarget-IL-24.Significant de- crease in tumor cell viability was observed in Namalwa cells stably transfected with mda-7/IL-24,compared with control cells transfected with empty plasmid pTarget(P

To produce specific monoclonal antibody (McAb) against human thrombomodulin (hTM), the full-length hTM cDNA-expressing plasmid pThr402 was transfected into CHO cells by Lipofectamine 2000 reagent. The hTM-expressing CHO cells, which was confirmed by flow cytometry and Western blot, were obtained by G418 selection. Then the McAb against hTM was prepared with classic hybridoma technique. A cell line of CHO-TM5 with high level of hTM was used to immunize female Balb/c mice 3 times at an interval of 4 weeks. On the third day after the third immunization, mice were sacrificed and spleen cells were harvested to prepare hybridoma cells with SP2/0 cells at the ratio of 10 to 1. Hybridoma cells were then cultured at 96-well plates for screening. Cellular enzyme-linked immunoabsorbent assay (CELISA) was applied twice. The first CELISA was done with polythene ELISA plate with a monolayer of CHO-TM5 cells. The positive clones from the first screen were then selected by reacting with similar screening ELISA plate but with CHO cell monolayer instead. Only clones that were positive for the first screening and negative for the second screening were kept, and called as CHO-TM5(+)CHO(-) hybridoma cells. Balb/c mice were intraperitoneally injected with the selected hybridoma cells. Ascites were collected and monoclonal antibodies were purified using FPLC, and its Ig class, subclass, and titer were then determined respectively. The specificity of the yielded McAb was identified with CELISA, flow cytometry, ABC immunohistochemistry and immunoblotting. One line of hybridoma cells with high expression of specific McAb against hTM, NH-1, was obtained. The Ig subclass of the McAb was IgG1 and the titer of ascitic McAb was 1x10(-6). Flow cytometry, CELISA and Western blot assays demonstrated that McAb NH-1 could specifically recognize hTM expressed in CHO-TM5 cells and human umbilical vascular endothelial cells. Meanwhile, the tissue specificity of antigen recognized by McAb NH-1 was identified by immunohistochemical ABC staining. NH-1 can specifically recognize the natural hTM expressed mainly in vascular endothelial cells, which will potentially be useful for investigation of the functions and clinic values of hTM.
Animals ; Antibodies, Monoclonal ; biosynthesis ; immunology ; Antibody Specificity ; CHO Cells ; Cricetulus ; Female ; Humans ; Hybridomas ; secretion ; Mice ; Mice, Inbred BALB C ; Thrombomodulin ; immunology ; Transfection

Objective To investigate the expression and variation of MIP 1β,MIP-2,and IL-12p70 in mice with bloodstream infection caused by 4 kinds of bacteria.Methods CD-1 (ICR) mouse models of bloodstream infection with Staphylococcus aureus (S.aureus),Enterococcus f aecalis (E.f aecalis),Escherichia coli (E.coli),and K lebsiella pneumoniae (K.pneumoniae) were established.After mice in each trial group and PBS control group were infected by bacteria for 0.5h,1h,3h,6h,12h,24h,and 48h,concentrations of MIP-1β,MIP-2,and IL-12p70 were detected by Luminex liquid suspension chip system.Results Concentrations of MIP-1β increased significantly 1h after bacteria was in blood,S.aureus,E.faecalis,E.coli,K.pneumoniae,and control groups were (134.5 ± 18.3),(61.5 ± 15.4),(3 354.0 ±809.0),(6 888.4 ± 1 100.2),and (28.9 ± 4.6) pg/mL respectively;the peak values of IL-12p70 were (389.3 ± 118.1),(127.6 ± 10.0),(42.2 ± 3.5),(62.8 ± 8.4),and (4.8 ± 0.3) pg/mL respectively.Concentrations of MIP-1β and MIP-2 in E.coli and K.pneumoniae groups were significantly higher than other trial groups and control group (all P＜0.01),while concentrations of IL-12p70 in S.aureus and E.faecalis groups were both significantly higher than E.coli,K.pneumoniae,and control groups (all P＜0.01).Conclusion Concentrations of MIP-1β and MIP-2 in E.coli and K.pneumoniae groups were both significantly higher than those in S.aureus and E.faecalis groups,while concentrations of IL-12p70 in S.aureus and E.faecalis groups were both significantly higher than those in E.coli and K.pneumoniae groups.The combination detection of multiple cytokines or chemokines are valuable in predicting gram-positive or gram-negative bacterial infection,and can provide basis for treatment of early infection.

OBJECTIVETo investigate cytotoxicity and genotoxicity of benzo(a)pyrene (B(a)P) by 16HBE-CYP1A1 cells which are human bronchial epithelial cell with CYP1A1 transformed.

METHODSExpression of CYP1A1 and mEH of cell models were tested by real-time quantitative polymerase chain reaction. Cells were treated with 0, 1, 5, 10 and 20 micromol/L B(a)P for 24 h. Adverse effects of B(a)P were tested by cytokinesis-block micronucleus (CBMN) cytome assays. Cytotoxicity was assessed by the nuclear division index (NDI), frequency of necrotic and apoptotic cells. Genetic damages were assessed by frequencies of CBMN, nucleoplasmic bridges (NPBs) and nuclear buds (NBUDs).

RESULTSHigh levels of CYP1A1 and mEH were found in 16HBE-CYP1A1 cells (relative mRNA content was 7.8 x 10(-4) and 0.030 respectively). In 16HBE-CYP1A1 cells, NDI were decreased in 1, 5, 10 and 20 micromol/L B(a)P treated groups, 1.92 +/- 0.04, 1.71 +/- 0.01, 1.61 +/- 0.04, and 1.41 +/- 0.01, respectively; and lower than control group (2.08 +/- 0.03). Compared with control group ((82.67 +/- 6.66)%), the binucleated cells ratios were decreased, (76.33 +/- 3.51)%, (66.33 +/- 0.58)%, (51.67 +/- 1.53)% and (39.0 +/- 1.0)% respectively.Necrotic cells ratios were (1.93 +/- 0.42)%, (2.20 +/- 0.53)%, (8.07 +/- 0.90)% and (15.27 +/- 2.80)%, respectively, higher than control group ((0.47 +/- 0.11)%). The differences were significant (F values were 899.94, 303.33, 240.87, P < 0.01). Apoptotic cells were increased at lower groups and decreased to normal at higher groups treated by B(a)P. They were (1.20 +/- 0.53)%, (2.00 +/- 0.20)%, (1.47 +/- 0.12)%, (1.20 +/- 0.00)% and (1.20 +/- 0.00)%, respectively. Analysis on biomarkers of genetic damage, the significant dose-effect relationship were observed in NPBs and NBUDs (F values were 50.23, 121.09, P < 0.01, respectively). Frequencies of NPBs were (4.67 +/- 2.89) per thousand, (7.33 +/- 1.53) per thousand, (10.67 +/- 2.08) per thousand and (11.00 +/- 1.00) per thousand respectively. Frequencies of NBUDs were (2.33 +/- 0.58) per thousand, (4.00 +/- 1.00) per thousand, (5.00 +/- 1.00) per thousand, and (7.67 +/- 1.16) per thousand respectively. However, the dose-relationship of CBMN last only to 10 micromol/L B(a)P treated groups in 16HBE-CYP1A1 cells, and frequencies of CBMN were (8.33 +/- 3.21) per thousand, (14.67 +/- 1.15) per thousand, respectively. Frequency of CBMN was (16.67 +/- 2.88) per thousand in 20 micromol/L B(a)P treated group, lower than 10 micromol/L B(a)P treated group ((17.67 +/- 2.08) per thousand). In 16HBEV control cells, the cytotoxicity was found only in higher B(a)P treated groups and frequencies of CBMN, NPBs and NBUDs were increased also. While no significant differences were observed between 5, 10, 20 micromol/L B(a)P treated groups (they were (6.37 +/- 2.08) per thousand, (9.33 +/- 1.52) per thousand, (9.33 +/- 3.21) per thousand; (4.33 +/- 1.53) per thousand, (6.00 +/- 2.65) per thousand, (5.33 +/- 1.53) per thousand and (2.33 +/- 0.58) per thousand, (3.33 +/- 1.16) per thousand, (3.67 +/- 1.16) per thousand, respectively).

CONCLUSIONSThe genetic damages were more severe after treated with activated B(a)P, which may be induced by decreased NDI, increased necrotic cells and inhibition of apoptosis.

Apoptosis ; drug effects ; Benzo(a)pyrene ; toxicity ; Cell Division ; drug effects ; Cell Line, Transformed ; DNA Damage ; Humans ; Micronuclei, Chromosome-Defective

This study was aimed to investigate the significance of detecting the antigen-receptor gene rearrangement clonality in the diagnosis of lymphoma. Paraffin-embedding and HE staining of samples from 31 patients with lymphomas were performed for morphologic observation by light microscope. Immunophenotype was analyzed by the immunohistochemistry (IHC) method. The clonality of antigen-receptor gene rearrangement was detected by BIOMED-2 Assay Kit. The results showed that among the 31 cases, 12 cases were suspected to be T-cell lymphoma, 1 case was suspected to be T-cell reactive hyperplasia, and 16 cases were suspected to be B-cell lymphoma, 2 cases were B-cell reactive hyperplasia. The detection results showed that the positivity of Ig gene rearrangement clonality was 94.44% (17/18), the positivity of TCR gene rearrangement clonality was 92.31% (12/13), the other two cases were negative. Finally, 12 cases were diagnosed to be T-cell lymphoma and 17 cases were B-cell lymphoma. The other two cases were reactive lymphoid proliferations. And the positivity rate in the 31 patients with lymphomas was 93%. It is concluded that the detection of antigen-receptor gene rearrangement clonality is a useful assistant method in the diagnosis of lymphoma.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Child ; Female ; Gene Rearrangement, T-Lymphocyte ; Humans ; Lymphoma ; diagnosis ; pathology ; Lymphoma, B-Cell ; diagnosis ; pathology ; Lymphoma, T-Cell ; diagnosis ; pathology ; Male ; Middle Aged ; Receptors, Antigen, T-Cell ; genetics ; Young Adult

BACKGROUND: Morinda officinalis oligosaccharide is the main active ingredient of morinda officinalis extract,which can promote the angiogenesis of ischemic tissue, but the mechanism is unknown. At present, there are two ways for endothelial repair:vascular endothelial cell division or differentiation from vascular endothelial progenitor cells in the peripheral blood. Here, we attempted to explain the pro-angiogenesis mechanism of morinda officinalis oligosaccharide by exploring whether there is a correlation between morinda officinalis oligosaccharide and the biological function of vascular endothelial progenitor cells, thereby providing experimental reference for new drug development. OBJECTIVE: To observe the effect of morindae officinalis oligosaccharide on the proliferation, differentiation and paracrine of vascular endothelial progenitor cells. METHODS: Vascular endothelial progenitor cells were isolated from healthy human peripheral blood, and divided into two groups: control group (without morindae officinalis oligosaccharide) and experimental group (with 0.15 g/L morindae officinalis oligosaccharide), followed by 48 hours of in vitro culture.The proliferation of vascular endothelial progenitor cells was tested by fluorescent staining;the ratio of vascular endothelial progenitor cells expressing CD31 was detected by flow cytometry; and the levels of vascular endothelial growth factor, stromal cell-derived factor 1 and interleukin 8 were analyzed by enzyme-linked immunosorbent method. RESULTS AND CONCLUSION: The percentage of vascular endothelial cells expressing CD34, CD133 or VEGFR- 2 was (84.72±4.34)%. After 48 hours of culture by 0.15 g/L morindae officinalis oligosaccharide, the proliferation rate and the positive expression of CD31 in the experimental group were significantly higher than those in the control group (P < 0.05). The levels of vascular endothelial growth factor, stromal cell-derived factor 1 and interleukin 8 in the experimental group were also higher than those in the control group (P < 0.05). To conclude, morindae officinalis oligosaccharide can promote the proliferation and differentiation of vascular endothelial progenitor cells, and meanwhile, it can stimulate the release of vascular endothelial growth factor, stromal cell-derived factor 1 and interleukin 8 from vascular endothelial progenitor cells through the paracrine pathway. Consequently, it is a potential drug for myocardial ischemic diseases.

Objective To investigate the effects of liver X receptor (LXR) agonist on adiposederived mesenchymal stem cells (AD-MSCs) implantation into infarcted hearts of mice.Method AD-MSCFluc+ which stably expressed firefly luciferase (Fluc) were isolated from β-actin-Fluc transgenic mice and characterized by flowcytometry.Male FVB mice were randomly allocated into the following four groups (n=10each):(1) sham group; (2) MI + PBSgroup; (3) MI + AD-MSCFluc+ group; (4) MI + AD-MSCFluc + LXR agonist (T0901317) group.AD-MSCFluc+ or PBS were injected intramyocardially into periinfarcted region of mice heart after permanent left anterior descending ( LAD ) artery ligation.Bioluminensence imaging (BLI)was performed for quantification of injected cells retention and survival.Cardiac function was evaluated by echocardiography.Results The AD-MSCFluc+ were positive for CD44 and CD90 by flowcytometry.BLI evidenced the firefly luciferase expression of AD-MSCFluc+ which was positively correlated with cell numbers ( r2 =0.98 ).The results of BLI in vivo revealed that LXR agonist could improve the survival of AD-MSCFluc+ at day 7,14 and 21 after transplantation compared with AD-MSCFluc+ alone group.Cardiac function was further improved in combination therapy group compared with AD-MSCFluc+ alone group (P＜0.05).Conclusions LXR agonist T0901317 can improve the retention and survival of intramyocardial injected AD-MSCFluc+ post-MI,and the combination therapy of T0901317 and AD-MSCFluc+ has a synergetic effect on improving cardiae funetion in this model.

This study was aimed at analyzing the effectiveness of vector control efforts at the 19th Asian Games in Hang-zhou,to provide a reference for future vector control at similar large-scale events.We collected and analyzed vector organism monitoring data for various venues at Asian Games(branch)villages and designated reception hotels(hotels and guesthouses)associated with the Asian Games in Hangzhou from April to October of 2023.Excel 2019 software and SPSS 20.0 software were used to organize the data.Paired t tests(for normally distributed data)or Wilcoxon signed rank tests(for data not nor-mally distributed)were performed,and count data were analyzed statistically with x2 tests.The larva mosquito path index at Asian Games venues peaked in June and then declined month by month.The first circle area of the adult mosquito human trapping index peaked in June,and the second circle area peaked in July.The mosquito indicators reached the corre-sponding control standards before the Asian Games.Moreo-ver,the differences in mosquito density before and after the Asian Games were statistically significant(all P＜0.05).Both the positivity rate and density of flies in rooms reached level A in the first and second circle areas from April to October.The proportion of indoor fly-proof facilities meeting the standards was lowest in May,then gradually increased.The corresponding control standards were reached in the first round in the fourth week of September,and in the second round in the first week of October.The difference in the density of positivity for room flies before and after the Asian Games was statistically significant(all P＜0.05).Rodent density reached the corre-sponding control standards in the second week of September.The proportion of anti-rodent facilities meeting the standards was lowest in June,and then gradually increased.The first round began to meet the corresponding control standards in the fourth week of September,and the second round began to meet the corresponding control standards in the first week of October.Ex-cept for the outdoor rat density path index,which was statistically significant before and after the Asian Games,the other indi-cators showed no statistical significance(P＞0.05).The cockroach density in the first circle area reached level A in all months,and that in the second circle area reached level B in mid-August.Vector control in the Hangzhou Asian Games achieved the ex-pected results,with no vector infringement or local vector-borne infectious diseases occurring.In future vector control for large-scale events,vector density monitoring should be performed as early as possible,and comprehensive prevention and control measures should be taken to maintain low vector density and prevent vector infringement incidents.

Objective To understand the epidemiologic characteristics of endemic typhus in Baoshan city. Methods Epidemiological data were collected and characteristics were analyzed. IgG antibody(Ab) of Rickettsia mooseri and Orientia tsutsuganushi in serum of patients were tested using both Weil-Felix and IFA method. The Rickettsia mooseri gltA gene, Rickettsia prowazekii gltA gene,Orientia tsutsugamushi 56 kDa protein gene, SFGR ompA gene, Ehrlichia sp. 16S rRNA gene and Anaplasma sp. 16S rRNA gene in spleen of mice were examined by PCR. Results Fifty- eight endemic typhus cases were found in Longyang district of Baoshan city, during July to August, 2009.Among them, 48 cases were confirmed by clinical diagnosis and 10 cases by laboratory tests. The Ab of Orientia tsutsugamushi Karp serotype was detected in 3 cases from laboratory diagnosis. The spleen samples from 85 Rattns flavipectus were tested using PCR. Of them, 3 samples for Rickettsia mooseri gltA gene showed positive (positive rate was 3.5% ), and the homology of 3 Rickettsia mooseri and Rickettsia mooseri Wilmington strain (GenBank U59714.1) was 100% through comparing gene sequence. The results of PCR for detecting Rickettsia prowazekii, Orientia tsutsugamushi, SFGR,Anaplasma sp. and Ehrlichia. sp were all negative. Conclusion The outbreak of endemic typhus was confirmed in Longyang district of Baoshan city through epidemiological data, clinical diagnosis and laboratory tests. Rickettsia mooseri DNA was detected in the dominant Raw flavipectus, suggesting that endemic typhus did exist in the local areas.

The function of circulatory system, nervous system and endocrine system is significantly changed in hypoxic environments. These changes affect the absorption, distribution, metabolism, and excretion of drugs in the body. Drug metabolizing enzymes and transporters are the main factors affecting drug metabolism; microRNA (miRNA) can act directly on drug metabolizing enzymes and transporters and can regulate their genes through hypoxia-inducible factor, inflammatory cytokines, and nuclear receptors. This article reviews the effect of hypoxia on drug metabolizing enzymes and transporters and the mechanisms by which miRNA modulates these proteins and their expression during hypoxia.