Formononetin, an Active Component of Astragalus Membranaceus, Inhibits the Pathogenesis and Progression of Esophageal Cancer Through the COX-2/Cyclin D1 Axis.

BACKGROUND: The goal was to investigate the inhibitory effect of formononetin, an active component in Astragalus membranaceus, on the pathogenesis and development of esophageal cancer and the mechanism of action. METHODS: The expression of COX-2 in cancer tissue and paracancerous tissue of patients with esophageal cancer detected early. C57BL/6 mice were used to construct a 4-nitroquinoline 1-oxide (4-NQO)-induced esophageal cancer model to verify the inhibitory effect of formononetin on the pathogenesis of esophageal cancer. Additionally, human esophageal cancer cells were treated with formononetin, and the effects on the proliferation and cell cycle of esophageal cancer cells were assessed by the CCK-8 assay and flow cytometry. Changes in the expression levels of cyclin D1 and COX-2 mRNA in cells were detected by RT-qPCR and western blot (WB) analysis. RESULTS: The expression level of COX-2 mRNA in esophageal cancer tissue was significantly higher than that in paracancerous tissue. In the mouse cancer model, the incidence of esophageal cancer in mice in the formononetin treatment group was significantly reduced at week 18 (0/15 vs. 2/15) and at week 24 (6/15 vs. 13/15) (all p < 0.05). Formononetin significantly inhibited the proliferation ability of KYSE170 and KYSE150 cells and inhibited the protein expression of COX-2 and cyclin D1 (both p < 0.05). CONCLUSIONS: Formononetin, an active component of Astragalus membranaceus, can prevent the pathogenesis and progression of esophageal cancer by reducing the expression of the inflammatory proteins COX-2 and cyclin D1.

Molecular docking and in vitro experiments verified that kaempferol induced apoptosis and inhibited human HepG2 cell proliferation by targeting BAX, CDK1, and JUN.

Hepatocellular carcinoma, as a common liver cirrhosis complication, has become the sixth most common cancer worldwide, and its increasing incidence has resulted in considerable medical and economic burdens. As a natural polyphenolic compound, kaempferol has exhibits a wide range of antitumor activities against multiple cancer targets. In this study, the Autodock software was used for molecular docking to simulate the interaction process between kaempferol and HCC targets and the PyMOL software was used for visualization. Proliferation of kaempferol HepG2 cells under the effect of kaempferol was detected using Cell Counting Kit-8 (CCK-8) assay, and the apoptosis rate of HepG2 cells was detected using flow cytometry. The expressions of proteins BAX, CDK1, and JUN protein expressions were detected by Western blot. Molecular docking found that the kaempferol ligand has 3 rotatable bonds, 6 nonpolar hydrogen atoms, and 12 aromatic carbon atoms, and can form complexes with the kaempferol targets P53, BAX, AR, CDK1, and JUN through electrostatic energy. GO (Gene Ontology) enrichment analysis suggests that kaempferol regulates the biological function of hepatocellular carcinoma cells and is related to apoptosis. Cell Counting Kit-8 assay suggested that Kaempferol can significantly inhibited HepG2 cell proliferation, and the inhibition rate increased with the increase in drug concentration and incubation time. Moreover, kaempferol can promoted HepG2 cell apoptosis in a dose-dependent manner. This compound upregulated BAX and JUN expression and downregulated CDK1 expression. Thus, Kaempferol can promote HepG2 cell apoptosis, and the regulatory mechanism may be related to the regulation of the expression levels of the apoptosis-related proteins BAX, CDK1, and JUN.

Determining novel candidate anti-hepatocellular carcinoma drugs using interaction networks and molecular docking between drug targets and natural compounds of SiNiSan.

BACKGROUND: SiNiSan (SNS) is an ancient traditional Chinese medicine (TCM) used to treat liver and spleen deficiencies. We studied the unique advantages of using SNS to treat hepatocellular carcinoma (HCC) with multiple components and targets to determine its potential mechanism of action. METHODS: The active compounds from the individual herbs in the SNS formula and their targets were mined from Traditional Chinese Medicine Systems Pharmacology Database (TCMSP). HCC-associated targets were collected from the TCGA and GEO databases and samples were collected from patients with stage III hepatocellular carcinoma. A compound-disease target network was constructed, visualized, and analyzed using Cytoscape software. We built a protein-protein interaction (PPI) network using the String database. We enriched and analyzed key targets using GSEA, GO, and KEGG in order to explore their functions. Autodock software was used to simulate the process of SNS molecules acting on HCC targets. RESULTS: A total of 113 candidate compounds were taken from SNS, and 64 of the same targets were chosen from HCC and SNS. The predominant targets genes were PTGS2, ESR1, CHEK1, CCNA2, NOS2 and AR; kaempferol and quercetin from SNS were the principal ingredients in HCC treatment. The compounds may work against HCC due to a cellular response to steroid hormones and histone phosphorylation. The P53 signaling pathway was significantly enriched in the gene set GSEA enrichment analysis and differential gene KEGG enrichment analysis. CONCLUSIONS: Our results showed that the SNS component has a large number of stage III HCC targets. Among the targets, the sex hormone receptors, the AR and ESR1 genes, are the core targets of SNS component and the most active proteins in the PPI network. In addition, quercetin, which has the most targets, can act on the main targets (BAX, CDK1, CCNB1, SERPINE1, CHEK2, and IGFBP3) of the P53 pathway to treat HCC.

Human infection with a novel reassortant Eurasian-avian lineage swine H1N1 virus in northern China.

Influenza A virus infections occur in different species, causing mild to severe respiratory symptoms that lead to a heavy disease burden. Eurasian avian-like swine influenza A(H1N1) viruses (EAS-H1N1) are predominant in pigs and occasionally infect humans. An influenza A(H1N1) virus was isolated from a boy who was suffering from fever and headache and designated as A/Tianjin-baodi/1606/2018(H1N1). Full-genome sequencing and phylogenetic analysis revealed that A/Tianjin-baodi/1606/2018(H1N1) is a novel reassortant EAS-H1N1 containing gene segments from EAS-H1N1 (HA and NA), classical swine H1N1(NS) and A(H1N1)pdm09(PB2, PB2, PA, NP and M) viruses. The isolation and analysis of A/Tianjin-baodi/1606/2018(H1) provide further evidence that EAS-H1N1 poses a threat to human health and greater attention should be paid to surveillance of influenza virus infection in pigs and humans.

Expression and clinical significance of melanoma antigen genes MAGE-A1 and MAGE-A3 in glioma tissues / 中国肿瘤生物治疗杂志

glioma; U87 cell; U251 cell; MAGE-A1; MAGE-A3; DNA methylation; histone acetylation@#To detect the expressions of melanoma antigen genes MAGE-A1 and MAGE-A3 in glioma tissues and to explore their clinical significance. Methods: Seventy-eight surgically resected glioma specimens and 15 normal brain tissue samples from donors suffered traffic accidence were collected at the Department of Neurosurgery, the Fourth Hospital of Hebei Medical University between January 2006 and January 2010, and the mRNAexpressions of MAGE-A1 and MAGE-A3 in collected tissues were detected with RT-PCR; their associations with the overall survival of patients were also analyzed.The promoter methylation status of the two genes was observed with methylation specific PCR, and the relationship between the gene expressions and promoter methylation status was analyzed. The expressions of MAGE-A1 and MAGE-A3 genes in U251 and U87 glioma cell lines were detected by RT-PCR before and after the treatment with DNA methyltransferase inhibitor 5-aza-CdR and/or histone deacetylase inhibitor trichostatin A (TSA). Results:The positive expression rates of MAGE-A1 and MAGE-A3 genes in glioma tissues were 65.34% and 38.46%, respectively; however, the two genes were not detected in 15 cases of normal brain tissues.The 5-year overall survival of patients in MAGEA1 positive expression group was shorter than that of negative expression group (P<0.05). There was significant correlation between the mRNA expressions of two genes and their promoter methylation status (all P<0.01). There was no mRNA expressions of MAGEA1 and MAGE-A3 in U87 cells untreated with 5-Aza-CdR and TSA, but a small amount of MAGE-A1 mRNA and MAGE-A3 mRNA were detected in U251 cells. TSA alone could not activate the expression of MAGE-A1 and MAGE-A3 genes. 5-Aza-CdR alone or in combination with TSA could activate the expression of both genes, and the combined effect was better than that of single administration. Conclusion: There are different degrees of MAGE-A1 and-A3 expression in glioma tissues, and the expression of MAGE-A1 is a negative prognostic factor for glioma patients. DNApromoter methylation and histone acetylation are important mechanisms of the activation of MAGE-A1 and MAGE-A3 expression.

A big pertussis outbreak in a primary school with high vaccination coverage in northern China: An evidence of the emerging of the disease in China.

BACKGROUND: A big pertussis outbreak occurred in a primary school with high vaccination coverage in northern China. An investigation was carried out in order to calculate the attack rate and identify the risk factors. METHODS: Between May 12 and July 29, an investigation was carried out in the primary school, which included 383 students and 27 teachers. Three definitions were used to distinguish the cases: confirmed, epidemiologically linked and suspected cases. A total of 232 blood samples were collected and examined by ELISA among healthy children in another primary school. RESULTS: A total of 138 suspected pertussis cases were counted, of which 116 students were confirmed. The attack rate among students was as high as 30.29%. The pertussis outbreak lasted 88 days, and had quaternary cases of transmission. Migrant children were almost four times as likely to catch the disease as local children (p = 0.005). In addition, students who had received the last dose of pertussis vaccine more than 4 years prior were three times more likely of becoming ill than those less than 4 years (p = 0.006). The average level of antibodies to pertussis was 30.99 IU/mL among healthy children. No statistically significant difference was observed between DTaP and DTwP (p = 0.843). CONCLUSIONS: This pertussis outbreak in a primary school with high vaccination coverage was an evidence of the pertussis resurgence in China. The major risk factor we identified was the waning of immunity in the years after pertussis vaccination. Booster vaccination for students should be given.

Detection of Viruses and Mycoplasma pneumoniae in Hospitalized Patients with Severe Acute Respiratory Infection in Northern China, 2015-2016.

Severe acute respiratory infection (SARI) presents a huge disease and economic burden worldwide. The present study described the frequency and types of different infectious etiologies among hospitalized patients with SARI in Tianjin, China, during 2015 and 2016. Basic information, in addition to a throat or serum sample, was collected from SARI patients. Nine viruses were detected using reverse transcription polymerase chain reaction, and Mycoplasma pneumoniae was detected using the Serodia Myco II gelatin particle agglutination test. A total of 585 specimens from 2,290 SARI cases were collected. The most common infection (19.66%, 115/585) was M. pneumoniae, followed by influenza virus A/B (6.15%, 36/585), and respiratory syncytial virus (4.96%, 29/585). Identification of viral or M. pneumoniae infections was the highest in the pediatric medicine ward (74.84%, 119/159), followed by the intensive care unit (37.04%, 80/216) and respiratory medicine ward (34.29%, 72/210). M. pneumoniae was highest (38.71%, 24/62) in the 5-14-year age group. Influenza was the main infection in January 2015 and March 2016. The correlation coefficient for the proportion of hospitalized cases of SARI and the positive detection rate within the same week was 0.25. M. pneumoniae and influenza were the leading pathogens among hospitalized SARI patients. A continued surveillance of hospitalized cases of SARI can detect emerging diseases, such as avian influenza A (H7N9) virus and other respiratory disease outbreaks.

Functional analysis of the uropathogenic Escherichia coli R049 gene.

The objective of this study was to determine the function of the novel uropathogenic Escherichia coli (UPEC) gene R049 during host infection. We infected the urinary tracts of mice with E. coli UPEC132 or the R049 deletion mutant UPEC132ΔR049.The mouse kidneys were harvested at 4 and 8h post-infection and screened for differentially expressed genes by microarray analysis. We identified 379 and 515 differentially expressed genes at 4 and 8 h post-infection, respectively. Thirty-four of these genes were associated with inflammatory and immune signaling pathways, including those related to mitogen-activated protein kinase signaling, leukocyte transendothelial migration, cytokine-cytokine receptor interaction, Toll-like receptor signaling, and apoptosis. Protein binding (GO 0005515) was the most prevalent molecular function in the Gene Ontology terms related to differentially expressed genes. In conclusion, R049 expression in UPEC132 is related to the early innate immune and inflammatory responses in UPEC-infected hosts. This work lays the foundation for further research on anti-infective immunity against UPEC.

Design and Evaluation of a Multi-Epitope Peptide of Human Metapneumovirus.

OBJECTIVES: No licensed vaccines or therapeutic agents for human metapneumovirus (hMPV) infection exist to date. We aimed to construct a multi-epitope peptide (MEP) of hMPV to show promising results for epitope-based vaccine development. METHODS: Six independent algorithms were screened to predict B-cell epitopes of hMPV, and three algorithms were used to predict cytotoxic T lymphocyte and T helper (Th) lymphocyte epitopes. Predicted epitopes were assembled in series with the spacers GPGPG and KK introduced, termed MEP. Recombinant mep genes were inserted into pET32a(+) plasmid and expressed in Escherichia coli strain BL21 (DE3). BALB/c mice were immunized with MEP with different adjuvants. Antibody titer, lymphocyte proliferation, cytotoxic T lymphocyte (CTL) activity and splenocyte cytokines were detected 2 weeks later after the last immunization. Microneutralization assay was used to detect neutralizing antibodies. RESULTS: Six B-cell epitopes, four CTL epitopes and two Th epitopes were screened to construct the mep gene. Expressed MEP induced >104 antibodies in BALB/c mice, and produced anti-MEP antibody reacting with hMPV strains specifically as detected in indirect fluorescent assay (the titer was 160). The lymphocyte proliferation index, CTL activity and splenocyte cytokines of the MEP immunization groups were higher than in the control group (p < 0.05). Both IgG1 and IgG2a antibodies could be detected in the different groups, and balanced Th1/Th2 cytokines were secreted by splenocytes in these groups. The mean neutralizing titers of the MEP+CpG ODN, MEP+Alum and MEP+Alum+ CpG ODN groups were 87 (95% CI 50-126), 93 (95% CI 67-121) and 96 (95% CI 69-147), respectively. CONCLUSION: MEP of hMPV elicited both strong humoral immunity and cell-mediated immunity in mice. The anti-MEP serum could neutralize hMPV infection in vitro. Joint use of CpG ODN and aluminum hydroxide adjuvants obtained the best immune effects. This study may contribute to hMPV epitope-based vaccine development.

An outbreak of acute respiratory disease in China caused by human adenovirus type B55 in a physical training facility.

OBJECTIVES: To investigate the cause of an acute respiratory tract infection (ARTI) outbreak. METHODS: Thirty-eight clinical samples were collected from 19 patients in an ARTI outbreak that occurred in a physical training facility in January 2013; patient demographic information was also collected. In addition, 60 influenza virus-negative samples from febrile respiratory patients were collected from the same community at the same time to determine whether these were the same infections. Multiplex PCR (multi-PCR) was used to detect the possible pathogen in these samples. All human adenovirus (HAdV)-positive samples were inoculated onto Hep-2 cells for isolation. HAdV isolates were typed by hexon gene, fiber gene, and whole genome sequencing using primers designed in-house and compared to different type/serotype HAdVs downloaded from GenBank. Phylogenetic analysis was used to determine the type of the HAdV. RESULTS: Of the 38 samples, 34 from 17 cases were HAdV-positive; two of them were co-infected, one with respiratory syncytial virus A and the other with human rhinovirus. The hexon gene open reading frame (ORF; 2841 nucleotides (nt)) and fiber gene ORF (978 nt) were obtained from four HAdV strains (TJ-2013-92, TJ-2013-94, TJ-2013-100, TJ-2013-122) from three upper respiratory infection cases and one pneumonia case. They were all completely identical. One HAdV isolate, TJ-2013-90, was selected for whole genome sequencing; 34238 nt were obtained. Phylogenetic analysis showed the whole genome of TJ-2013-90 to be clustered together with HAdV-B55/HAdV-B11a. Three of 60 influenza virus-negative specimens were HAdV-positive, but hexon and fiber gene analysis showed that they were grouped in different branches to the HAdV isolates from this outbreak. CONCLUSIONS: The cause of this ARTI outbreak was HAdV-B55. This was another outbreak caused by this re-emerging virus. Continuous surveillance of respiratory adenovirus is necessary for disease control.

[The status of pertussis infection and molecular epidemiological characteristics of pertussis in Tianjin, 2013].

OBJECTIVE: To understand the status of pertussis infection and characteristics of molecular epidemiology of pertussis in 2013 in Tianjin. METHODS: Totally, 181 suspected pertussis cases were selected and their nasopharyngeal swabs and serum were sampled at the Disease Monitoring Settings in Tianjin. Real-time PCR was used to detect Bordetella pertussis double target genes and enzyme linked immune-sorbent assay (ELISA) method was used to detect the specific pertussis toxin IgG (PT-IgG) antibody. Fimbriae 2 (FIM2) and Fimbriae 3 (FIM3) genes of pertussis was amplified by PCR for sequencing, from 30 pertussis DNA positive samples. RESULTS: The positive rate of Real-time PCR was 68.24% in 148 cases and the positive rate of PT-IgG antibody was 55.56% in 108 cases. Among 101 cases that nucleic acid were positive, the median duration of disease was 11 days. Among the PT-IgG Positive cases (60 cases), the median duration of disease was 21 days. In cases under 1 year old, the Real-time PCR testing positive rate was 84.28%. Positive rates among other age groups, the differences were statistically significant. Nucleotide homologies of FIM2 and FIM3 genes from 30 pertussis strains were 99.6%-100.0%, while it was 99% when compared to both international standard Tohama strain and Chinese vaccine strain. CONCLUSION: Detection of pertussis by Real-time PCR from clinical nasopharyngeal swab sample was quick and sensitive for the diagnosis. Bordetella pertussis epidemic strains in Tianjin area appeared close relation with both the international standards and China vaccine strains.

The expression and clinical significance of melanoma-associated antigen-A1, -A3 and -A11 in glioma.

Melanoma-associated antigens (MAGEs) were initially identified in melanoma and have since been widely studied. Melanoma-associated antigen-As (MAGE-As), a subfamily of MAGEs, are expressed in germ cells and various types of cancer, and are considered to be ideal targets for cancer immunotherapy. Glial cells and melanocytes originate from the neural ectoderm, so tumors derived from these two types of cells, i.e. gliomas and melanomas, may have common biological characteristics. However, studies on the expression of the MAGE-A family in gliomas are limited and conflicting. In the present study, the expression levels of MAGE-A1, -A3 and -A11 were detected by immunohistochemistry, and the association of their expression levels with the clinicopathological parameters, overall survival (OS) and ki-67 labeling indices of glioma patients were analyzed. The results showed that i) the expression levels of MAGE-A1, -A3 and -A11 proteins in the glioma tissues were 64.1, 51.3 and 57.7%, respectively and that no MAGE-A1, -A3 or -A11 expression was detected in the normal brain specimens; ii) the expression levels of MAGE-A1 and -A11 increased with ascending pathological grades and were positively correlated with the ki-67 labeling index; and iii) the OS of the patients in the groups with high MAGE-A1 (P=0.005) and -A11 (P=0.019) expression was statistically lower compared with the groups with low expression and no significant differences in OS were detected between the patients in the groups with high and low MAGE-A3 expression (P=0.304). Based on these results, we conclude that MAGE-A1, -A3 and -A11 may be used as ideal targets for glioma immunotherapy, and that MAGE-A1 and -A11 expression may be involved in tumor cell proliferation. These proteins may be potential indicators of a poor prognosis in glioma patients.

Epidemiology and full genome sequence analysis of H1N1pdm09 from Northeast China.

Pandemic influenza A (H1N1) 2009 virus (H1N1pdm09) was a novel tri-assortment virus that emerged in Mexico and North America in 2009 and caused the first influenza pandemic in the 21st century. This study investigated the prevalence pattern and molecular characteristics of H1N1pdm09 in three continuous years from April 2009 to March 2012 in populations of Tianjin, Northeast China. Totally, 3,068 influenza viruses (25.4 %) were detected from 12,089 respiratory specimens. Among them, 41.4 % (1,269/3,068) were H1N1pdm09 positive. 15.1 % (192/1,269) severe respiratory infection cases were H1N1pdm09 positive. H1N1pdm09 was the predominant prevalence subtype in October 2009-March 2010 (69.1 %, 930/1,346) and October 2010-March 2011 (42.1 %, 220/523). Eight isolated H1N1pdm09 viruses from severe infection/death cases in three different years were selected to sequence the whole genome through splicing the sequences following 46 PCRs. HA sequences of seven H1N1pdm09 isolates from mild infection cases were detected. Phylogenetic analysis showed that HA, NA, M, NP and NS genes of H1N1pdm09 viruses gathered together with swine influenza A (H1N1), whereas PB2 and PA genes originated from avian influenza virus, and PB1 gene originated from human seasonal influenza virus. Identity analysis indicated that all the genes were highly conserved. Compared with vaccine strain A/California/07/2009(H1N1), the maximal mutation gene was HA (0.7-2.6 %), then NA (0.6-1.7 %), last one was M (mutation rate 0-0.6 %). More site substitutions were observed in 2011 isolates than in 2009 and 2010 isolates of HA (p = 0.002), NA (p = 0.003) and PA (p = 0.001) proteins. The amino acid substitution rates were varied among eight gene segments, ranging from 7.39 × 10(-4) for PB2 to 7.40 × 10(-3) for NA. The higher d N / d S rates were observed in HA, PA and NS segments in H1N1pdm09 in Tianjin. Three HA amino acid site substitutions occurred at the HA receptor-binding sites and antigenic determinant, including S179N and K180T (located at antigenic site Sa) in A/Tianjinhedong/SWL44/2011(H1) and A/Tianjinjinnan/SWL41/2011(H1), and D239N (located at antigenic site Ca) in A/Tianjinninghe/SWL49/2009(H1). Antigenic drift may have occurred in H1N1pdm09 with time. No oseltamivir-resistance site substitution was observed at 275 and 295 sites. Amino acid residue site at 31 in M2 protein was N in all 8 isolates, which suggested that H1N1pdm09 was resistant to amantadine.