AIM To examine the antitumor activity of clostridium difficile toxin A. METHODS Highly purified toxin A from clostridium difficile was obtained by bovine thyroglobulin affinity chromatography followed by ion exchange chromatography steps on Q sepharose. The antitumor activity of toxin A of clostridium difficile on TPC 1 cell line was studied with Vero cell line as the normal cell line. The estimating ways used in this study were trypan blue exclusion test, MTT calorimetric assay, membrane damage test using 3H Uridine and observation by optical, fluorescence microscopes. RESULTS Exposed to toxin A, the cell growth inhibition, apoptosis index, non adherent cells and membrane damage in TPC 1 cell line were much more great than that in vero cell line, and the effect was dependent upon the concentration and treating time. CONCLUSION The antitumor activity of toxin A on TPC 1 cells was much higher than that on vero cell line. The data are of potential importance for the development of toxin A and the exploration of antitumor drugs.

Objective To investigate the association of leiomyoma with death receptors(DR) and the mechanisms underlying the development of leiomyoma.Methods The semi-quantitative RT-PCR was employed to examine the mRNA expression of apoptosis-associated death receptors in the Leiomyoma tissue samples collected from 20 clinical cases.Results It was found that the mRNA expression of DR4 in leiomyoma tissue was dramatically decreased as compared to that of normal tissue controls (P<0.001),but no difference was found between leiomyoma and normal tissue controls in terms of DcR1,DcR2,DR5,TNFR1A,TNFR1B and Fas mRNA expressions (P>0.05).Conclusion It is highly possible that DR4 expression might be involved in the development of leiomyoma.More efforts will be required to elucidate the exact mechanisms underlying the development of leiomyoma.

Objective To investigate the correlation between methylation status and death-associated protein kinase 1 (DAPK1) in SiHa and Hela cell line of cervical carcinoma and intervention of DNA methyltransferase inhibitor 5-azacytidine on the expression of DAPK1 and the proliferation of the cells. Methods DAPK1 methylation status was analyzed using methylation-specific PCR methods. The expression of mRNA and protein of DAPK1 were analyzed by RT-PCR and SABC methods after the treatment with 5-azacytidine. MTT assay was used to observe the changes of proliferation activity of the cells after 5-azacytidine treatment. Results DAPK1 genes were methylated and did not express in SiHa cells in the cervical carcinoma. Its expression could be restored by 5-azacytidine. MTT assay showed 5-azacytidine could weaken the proliferation of cancerous cells. Conclusion DAPK1 methylation plays an important role in the carcinogenesis of cervical cells and can reexpress after the treatment with 5-azacytidine which also restored its inhibitory function on carcinoma.

ObjectiveTo verify the homology hypothesis of depression and insomnia.Methods All-night physiological signals of electroencephalogram (EEG), electrosculogram (EOG) and electromyogram (EEG) of 30 depression cases and 30 insomnia cases were recorded and analyzed with polysomnogram.ResultsThe rapid eye movement (REM) sleep of depression patients was more active, and there was a significant difference in all REM indexes compared with insomnia cases (P<0.01). Characteristics of insomnia cases were poor sleep continuity, easy wake after sleep-onset, and depression of slow wave sleep and REM sleep. There was no slow wave sleep in some insomnia cases.Conclusion The sleep patterns of depression and insomnia are different, REM sleep is hyperactive in depression; non-REM sleep is disturbance in insomnia.

Objective To develop a simplified bisulfite genomie sequencing(BGS)method for DNA methylation marker scanning.Methods According to modified BGS protocol,the desalt DNA treated with bisulfite were directly used for bisulfite-PCR(BSP)without alkali treatmenL Complement of the bisulfite modification Wag accomplished by a prolonged pre-denaturation stage.After BSP,a second round PCR was performed with a pair of GC tagged primers to adjust the GC content of the amplieon for direct sequencing.To assess this improved protocol,promotor methylation of TNF-α gene in 3T3-L1 cell and androgen receptor(AR)gene in Hela cell was investigated.The real time BSP for Alu was also used to compare the sensitivity of the modified assay with traditional assay.Results Both the hypermethylated TNF-α promotor and hypomethylated AR promotor were successfully sequenced by improved BGS method,and the results were consistent with that of the traditional assay.The conversion rate reached 100%,while the conversion specificity was higher than 93.75%.The sensitivity of improved BGS method inereaged significantly(t=2.978 2,P＜0.05)and showed good reproducibility.Condusion The improved BGS method is simple and sensitive,facilitating more ambitious genomic methyhtion mapping studies.

In order to develop monoclonal antibodies (McAbs) against the gp90 protein of reticuloendotheliosis virus (REV), the His-tagged gp90 protein of REV was used to immunize BALB/c mice. Hybridomas were generated by fusing mouse myeloma cells SP2/0 with the splenocytes from the immunized mice. After screening and 3 rounds of cloning process, 3 hybridomas (3G5-B8, 3G5-A10 and 1G12) that stably secreted McAbs against the REV-gp90 were obtained. The isotypes of the McAbs were determined to be IgG1, IgG1 and IgG2b. The McAbs specifically bound to gp90 in REV-infected DF-1 cells, as demonstrated by Western blotting and indirect immunofluorescence assay. The recognition regions on gp90 that were recognized by 3G5-B8/3G5-A10 and 1G12 were located between amino acids 200 to 245 and 230 to 235, respectively, as demonstrated by Western blotting analysis. These McAbs will be useful in the diagnosis and pathogenesis study of REV.
Animals ; Antibodies, Monoclonal ; biosynthesis ; Antibody Specificity ; Blotting, Western ; Epitope Mapping ; Hybridomas ; Immunoglobulin G ; biosynthesis ; Mice ; Mice, Inbred BALB C ; Reticuloendotheliosis virus ; immunology ; Viral Envelope Proteins ; immunology

Infectious bursal disease virus (IBDV) VP4 plays an important role in immunosuppression of host. In order to develop monoclonal antibodies (McAbs) against VP4, we vaccinated BALB/c mice with His-VP4, screened and subcloned positive clones. We established 4 hybridoma cell lines that stably secreted McAbs against VP4 and named these cell lines 3B3, 3H11, 4C8 and 4G6, respectively. We tested the dissociation constant (Kd) of these McAbs, and found that their K(d)s were 4.61 x 10(-11), 1.71 x 10(-10), 4.26 x 10(-11), 5.02 x 10(-11), respectively. The isotypes of these McAbs were determined to be IgG1, IgG1, IgG2b and IgG1. These McAbs specifically bound to VP4 in IBDV infected DF-1 cells as demonstrated by Western blotting analysis and fluorescence antibody assay. These McAbs would help to detect IBDV infection and to analyze the biological activities of IBDV VP4.
Animals ; Antibodies, Monoclonal ; biosynthesis ; Blotting, Western ; Cell Line ; Fluorescent Antibody Technique ; Hybridomas ; Infectious bursal disease virus ; Mice ; Mice, Inbred BALB C ; Viral Structural Proteins ; immunology

This study was performed to investigate the chemical constituents in the twigs and leaves of Harrisonia perforate. Six compounds were isolated from the 95% EtOH extract of the twigs and leaves of Harrisonia perforate by silica gel, ODS, Sephadex LH-20 column chromatographies and preparative HPLC. On the basis of chemical properties and spectra data, these compounds were identified as harriperfin E (1), kihadanin A (2), kihadanin B (3), 6α-acetoxyobacunol acetate (4), gardaubryone C (5), and β-sitosterol methyl ether (6), respectively. Compound 1 is a new chromone, and compounds 2-6 are isolated from this plant for the first time.

Objective To investigate the risk factors of delayed gastric emptying (DGE) after pancreaticoduodenectomy,in order to provide a theoretical basis for prevention and treatment of this complication.Methods The term DGE was searched in Pubmed,Medline,EMBASE,Cochrane Library,CNKI,Wanfang,and published literatures were collected to determine the risk factors of DGE after pancreaticoduodenectomy.The Review Manager 5.3 software was used in the analysis.Results A total of 52 articles were included.The results of Meta-analysis showed that age and preoperative bilirubin levels did not significantly influence the incidence of DGE.Preoperative cholangitis (OR =3.39,95％ CI 1.97 ～ 5.82),hypoalbuminemia (OR =2.53,95％ CI 1.59 ～4.02),and intraoperative blood loss of more than 1 L (OR =1.98,95％ CI 1.18 ～ 3.33) significantly increased the incidence of DGE.Pyloric resection (RR =2.06,95％ CI 1.05 ～4.05),antecolic reconstruction (RR =0.74,95％ CI 0.56 ～ 0.99) and Braun enteroenterostomy (OR =0.36,95％ CI 0.17 ～0.77) significantly decreased the risk of DGE.When compared with Roux-enY enteroenterostomy,Billroth Ⅱ enteroenterostomy reduced the incidence of clinically relevant DGE (RR =0.30,95 ％ CI 0.11 ～ 0.79).Postoperative pancreatic fistula (OR =3.84,95 ％ CI 2.71 ～ 5.44) and intraabdominal infection/abscess (OR =3.95,95％ CI 2.87 ～ 5.43) were significantly associated with a high incidence of DGE.Conclusions Hypoalbuminemia,cholangitis,large blood loss,and postoperative abdominal complications were the risk factors of DGE.Pyloric resection,antecolic reconstruction,Billroth Ⅱ enteroenterostomy,and Braun enteroenterostomy significantly reduced the incidence of DGE.Subgroup analysis showed that differences on DGE definition in studies might be an important cause for the heterogeneity in the results of the different studies.

Abstract: Objective To analyze the epidemiological characteristics of viral hepatitis E in Fujian Province from 2012-2021, and to provide scientific evidence for the prevention and control of hepatitis E in the future. Methods Descriptive epidemiological method was used to analyze hepatitis E cases in Fujian Province from 2012 to 2021. Results From 2012 to 2021, a total of 8 877 cases of hepatitis E were reported in Fujian Province from 2012-2021. The overall incidence rate showed a decreasing trend (χ2trend =458.14, P<0.001), with the lowest incidence rate of 1.32/100 000 in 2020 and an annual average incidence rate of 2.29/100 000 per year. The incidence was higher in winter and spring, with the months of March and April having the highest number of reported cases (2 146, 24.17%) and the fewest cases were reported in September (571, 6.43%). The difference in reported incidence rates between cities was statistically significant (χ2=1 877.75, P<0.01). The comprehensive experimental zone of Pingtan had the highest average reported incidence rate of 6.03/100 000, while Zhangzhou had the lowest at 0.94/100 000. The number of male cases was higher than the number of female cases, with a male to female ratio of 2.04∶1. The disease was most prevalent among middle-aged and elderly individuals, with the age group of 40-<65 years having the highest number of reported cases, accounting for 57.44% (5 099/8 877) of all cases. The age group of 50-<55 years had the highest reported incidence, with the number of reported cases increasing with age below 50 years, but decreasing with age over 50 years. As for occupational distribution, peasants had the highest proportion of the disease, accounting for 34.49% (3 062 cases) of the total cases. Conclusions The reported incidence rate of hepatitis E in Fujian showed a downward from 2012 to 2021. Due to the impact of COVID-19, incidence of the lowest was 2020, but it did not reflect the true situation of the disease, which may have affected trend of hepatitis E. In order to control and reduce the incidence of hepatitis E, efforts should be made to increase publicity and education on health knowledge and vaccination among key areas and populations, strengthen monitoring and diagnostic capability, and implement comprehensive prevention and control measures.