Supportive role of the Na+ transporter CmHKT1;1 from Cucumis melo in transgenic Arabidopsis salt tolerance through improved K+/Na+ balance.

KEY MESSAGE: CmHKT1;1 selectively exports Na+ from plant cells. Upon NaCl stress, its expression increased in a salt-tolerant melon cultivar. Overexpression of CmHKT1;1 increased transgenic Arabidopsis salt tolerance through improved K+/Na+ balance. High-affinity K+ transporters (HKTs) are thought to be involved in reducing Na+ in plant shoots under salt stress and modulating salt tolerance, but their function in a moderately salt-tolerant species of melon (Cucumis melo L.) remains unclear. In this study, a Na+ transporter gene, CmHKT1;1 (GenBank accession number: MK986658), was isolated from melons based on genome data. The transcript of CmHKT1;1 was relatively more abundant in roots than in stems or leaves from melon seedlings. The tobacco transient expression system showed that CmHKT1;1 was plasma-membrane localized. Upon salt stress, CmHKT1;1 expression was more strongly upregulated in a salt-tolerant melon cultivar, 'Bingxuecui' (BXC) compared with a salt-sensitive cultivar, 'Yulu' (YL). Electrophysiological evidence demonstrated that CmHKT1;1 only transported Na+, rather than K+, when expressed in Xenopus laevis oocytes. Overexpression of CmHKT1;1 increased salt sensitivity in Saccharomyces cerevisiae and salt tolerance in Arabidopsis thaliana. Under NaCl treatments, transgenic Arabidopsis plants accumulated significantly lower concentrations of Na+ in shoots than wild type plants and showed a better K+/Na+ balance, leading to better Fv/Fm, root length, biomass, and enhanced plant growth. The CmHKT1;1 gene may serve as a useful candidate for improving crop salt tolerance.

Artemisinin enhances the anti-tumor immune response in 4T1 breast cancer cells in vitro and in vivo.

BACKGROUND: Breast cancer is a prominent cause of death among women worldwide. Recent studies have demonstrated that artemisinin (ART) displays anti-tumor activity. Using a mouse breast cancer model, we investigated the effects of ART in vitro and in vivo to determine how it influences the anti-tumor immune response. METHODS: We measured the proliferation and apoptosis of 4T1 cells in vitro after ART treatment by MTT assay and FACS. To examine the effects of ART in vivo, tumor volumes and survival rates were measured in 4T1 tumor-bearing mice. FACS was used to determine the frequencies of Tregs, MDSCs, CD4+ IFN-γ+ T cells, and CTLs in the tumors and spleens of the mice. mRNA levels of the transcription factors T-bet and FOXP3 and cytokines IFN-γ, TNF-α, TGF-ß, and IL-10 were also determined by real-time RT-PCR. ELISA was used to measure TGF-ß protein levels in the cell culture supernatants. RESULTS: ART supplementation significantly increased 4T1 cell apoptosis and decreased TGF-ß levels in vitro. ART also impeded tumor growth in 4T1 TB mice and extended their survival. MDSC and Treg frequencies significantly decreased in the 4T1 TB mice after ART treatment while CD4+ IFN-γ+ T cells and CTLs significantly increased. ART significantly increased T-bet, IFN-γ, and TNF-α mRNA levels within the tumor and significantly decreased TGF-ß mRNA levels. CONCLUSION: ART supplementation hindered 4T1 tumor growth in vivo by promoting T cell activation and quelling immunosuppression from Tregs and MDSCs in the tumor.

Clinical and genetic characteristics of Chinese pediatric patients with chronic granulomatous disease.

BACKGROUND: Chronic granulomatous disease (CGD) is a rare disease in China, and very little large-scale studies have been conducted to date. We aimed to investigate the clinical and genetic features of CGD in Chinese pediatric patients. METHODS: Pediatric patients with CGD from Beijing Children's Hospital, Capital Medical University, China, were enrolled from January 2006 to December 2016. RESULTS: A total of 159 pediatric patients with CGD were enrolled. The median age of clinical onset was 1.4 months, and 73% (116/159) had clinical onset symptoms before the 1 year of age. The most common site of invasion was the lungs. The lymph nodes, liver, and skin were more frequently invaded in X-linked (XL) CGD patients than in autosomal recessive (AR) CGD patients (P < 0.05). Approximately 64% (92/144) of the pediatric patients suffered from abnormal response to BCG vaccination. The most frequent pathogens were Aspergillus and Mycobacterium tuberculosis. Gene analysis indicated that 132 cases (89%, 132/147) harbored CYBB pathogenic variants, 7 (5%, 7/147) carried CYBA pathogenic variants, 4 (3%, 4/147) had NCF1 pathogenic variants, and 4 (3%, 4/147) had NCF2 pathogenic variants. The overall mortality rate in this study was 43%, particularly the patients were males, with CYBB mutant and did not receive HSCT treatment. CONCLUSIONS: Chronic granulomatous disease is a rare disease affecting Chinese children; however, it is often diagnosed at a later age, and thus, the mortality rate is relatively high. The prevalence and the severity of disease in XL-CGD are higher than AR-CGD.

Comprehensive bioinformatics analysis identifies several potential diagnostic markers and potential roles of cyclin family members in lung adenocarcinoma.

PURPOSE: The aim of this study was to identify critical genes in lung cancer progression. METHODS: We downloaded and reanalyzed gene expression profiles from different public data-sets using comprehensive bioinformatics analysis. Differentially expressed genes (DEGs) were identified in lung adenocarcinoma tissues compared with adjacent nonmalignant lung tissues. The overlapping DEGs identified from different datasets were used for functional and pathway enrichment analyses and protein-protein interaction (PPI) analysis. Moreover, transcription factors (TFs) and miRNAs that regulated the overlapping DEGs were predicted, followed by a TF-miRNA-target network construction. Furthermore, survival analysis of genes was performed. Several genes were further validated by quantitative real-time PCR (qRT-PCR). RESULTS: A total of 647 overlapping upregulated genes and 979 overlapping downregulated genes were identified. The overlapping upregulated genes and downregulated genes were involved in different functions, such as cell cycle, p53 signaling pathway, immune response, and cell adhesion molecules (CAMs). Several genes belonging to the cyclin family, including CCNB1, CCNB2, and CCNA2, were hubs of the PPI network and TF-miRNA-target network. Additionally, genes, including NPAS2, GNG7, CHIA, and SLC2A1, were predicted to be prognosis-related DEGs. Gene expression profiles determined by bioinformatics analysis and qRT-PCR were highly comparable. CONCLUSION: CCNB1, CCNB2, CCNA2, NPAS2, GNG7, CHIA, and SLC2A1 are promising targets for the clinical diagnosis and therapy of lung adenocarcinoma.

[Characteristic of nuclear antigen 1 gene and latent membrane protein 1 gene of Epstein-Barr virus in primary EBV infection in children in Beijing area in 2005-2010].

OBJECTIVE: To analyze the characteristic of nuclear antigen 1 gene and latent membrane protein 1 gene of Epstein-Barr virus in primary EBV infection in children in Beijing area in 2005-2012. METHODS: Polymerase chain reaction (PCR) was used to amplify the EBNA-3C, EBNA1 and LMP1 genes. The amplified products were sequenced directly and the sequences were analyzed by BioEdit 7. 0. 9 and MEGA 4. 0. 2. RESULTS: Type A EBV was detected in 98% samples. Nucleotide sequence analysis of the carboxy-terminal region of EBNA1 showed that Vvvl was deteted in 98% samples. DNA sequence analysis of LMP1 C-terminus indicated that China 1 was 90% in this study. There were no significant differences in the frequency of Vvv1 and China 1 between the IM and HLH samples (P = 1.00). Linkage analysis of EBV types, EBNA1 and LMP1 variants indicated that 90% of EBV type A was associated with EBNA1-Vvv1 variant and LMP1-China 1 variant in 40 cases. Full length of LMP1 gene was successfully amplified in 35 cases. Four Chinese groups (CG1-4) were identified. The percentage of CG1-CG4 were 85%, 6%, 6% and 3%, respectively. CONCLUSION: EBV type A is predominant in primary EBV infection in children in Beijing Area. EBNA1-Vvv1 and LMP1-China 1 variants were predominant genotypes in this area. There is a high linkage between EBNA1-Vvv1 variant and LMP1-China 1 variant. Four Chinese groups (CG1-4) were identified according to the full length of LMP1 gene and CG1 was the most prevalent.

Epidemiologic and clinical characteristics of infectious mononucleosis associated with Epstein-Barr virus infection in children in Beijing, China.

BACKGROUND: infectious mononucleosis (IM) is a self-limited disease, but a few cases may have severe complications. This retrospective study was to explore the epidemiologic and clinical characteristics of IM associated with Epstein-Barr virus infection (EBV-IM) in children. METHODS: hospitalized patients with EBV-IM were enrolled during January 2005 to October 2008 in Beijing Children's Hospital Affi liated to Capital Medical University. All patients were divided into four groups: <1 year (group I), 1 to 3 years (group II), 3 to 6 years (group III), and ≥ 6 years (group IV). The epidemiology and clinical characteristics were compared among the four groups. RESULTS: totally 418 patients were enrolled, with 245 boys and 173 girls. Fever, lymphadenopathy and pharyngitis were three main manifestations of the patients. The incidences of hepatomegaly, splenomegaly and rash were higher in the patients aged below 6 years, and with age increment the incidences lowered. In contrast, the patients aged <1 year had the lowest incidence of tonsillopharyngitis. The total white blood cell count was higher in the infantile group than in the other groups (P=0.038). The infantile group had significantly lower levels of serum alanine aminotransferase and aspartate aminotransferase than the older groups (P=0.007 and P=0.012 respectively). The percentage of CD4(+) T cell subset decreased and the percentage of CD8(+) T cell subset increased with age increment. CONCLUSIONS: the incidence of EBV-IM peaked in children at age of 4 to 6 years in Northern China. Most of the patients had the classic triad of fever, lymphadenopathy and pharyngitis. Clinical symptoms, signs, laboratory findings and complications of patients varied with ages.

[Incidence and relationship of human cytomegalovirus infection and human herpesvirus 6 infection in pediatric patients after hemopoietic stem cell transplantation].

OBJECTIVE: To study the incidence of human cytomegalovirus (CMV) and human herpesvirus 6 (HHV-6) infection in pediatric patients with hemopoietic stem cell transplantation (HSCT), and to explore the relationship between CMV and HHV-6 infection in pediatric patients with HSCT. METHODS: Pediatric patients with HSCT in hemotology center of Beijing Children's Hospital were enrolled into this study from June 2007 to October 2009. Peripheral blood were collected every week after HSCT, and Fluorescent quantitation PCR and conventional PCR were used to detect CMV DNA load in serum and HHV-6 DNA in peripheral blood respectively. Genetic typing was conducted on HHV-6. RESULTS: Fifty two pediatric patients with HSCT were enrolled into this study, and six hundreds and thirty six specimens were collected totally. CMV DNA was detected in fifty two specimens from twenty cases. The median time was 56 days after HSCT. The incidence of CMV infection was 38.5% (20/52) in all HSCT patients and 47.6% (20/42) in allogene HSCT patients. The incidence of late CMV infection was 22.2% (6/27) in allogene HSCT. Three patients died of CMV infection,and two died of CMV interstitial pneumonia. HHV-6 DNA was detected in thirty three specimens from fourteen cases. The median time was 23 days after HSCT. The incidence of HHV-6 infection was 26.9% (14/52)in all HSCT patients and 31% (13/42) in allogene HSCT patients. The genotype of HHV6 was all type B. HHV-6 DNA was positive in six of twenty cases with CMV infection. The incidence of co-infection was 30% (6/20). CONCLUSIONS: There was a substantial incidence of CMV and HHV6 infection after HSCT. The relationship between earlier HHV6 infection and later CMV infection in pediatric patients with HSCT need further study.

[Comparative proteomic analysis of human large cell lung cancer cell line with high and low metastasis potentials].

OBJECTIVE: A comparative proteomic analysis on two human large cell lung cancer strains with high metastasis potential (L9981) and low metastasis potential (NL9980) were conducted. METHODS: The total proteins of the two cell lines were separated by immobilized pH gradient (IPG )-based two-dimensional gel electrophoresis. The differential expression proteins of the two cell lines were analyzed using image analysis software. Thirteen differential expressed proteins were further identified using in-gel digestion with trypsin, of which peptide extracts were prepared for MALDI-TOF MS/MS analysis by Daltonics AutoFlex TOF-TOF LIFT Mass Spectrometer (Bruker). Protein identifications were searched in the NCBInr protein database using the Mascot search engine. RESULTS: Protein image analysis indicated that 8 protein spots were observed only in L9981, not in NL9980; 8 protein spots were observed only in NL9980, not in L9981; 19 protein spots were detected in both L9981 and NL9980, of which 9 showed significantly higher volumes in L9981 than in NL9980 and 10 showed significantly higher volumes in NL9980 than in L9981. MS and biological informatics study found that the expressions of heat shock 70 kD protein 9B precursor, MTHSP75 and glutathione synthetas incerased in L9981 cells. However, a variant of P47 protein, immunoglobulin heavy chain variable region, enolasel, heat shock protein and eukaryotic translation initiation fact 3 were down-regulated in L9981 cells. Pyruvate kinase (PK) was only expressed in L9981 cells while WD-40 repeat protein was only expressed in NL9980 cells. CONCLUSIONS: The metastatic lung cancer cell lines display different protein profiles compared to the non metastatic lung cancer cell lines. The identified proteins are likely to be associated with tumor metastasis, 4hich could serve as a basis for searching potential prognosis markers of lung cancer and elucidating the mechanisms of the metastasis of lung cancer.