Comprehensive two-dimensional gas chromatography mass spectrometry-based untargeted metabolomics to clarify the dynamic variations in the volatile composition of Huangjiu of different ages.

Aging plays an important role in the formation of aroma characteristics of Huangjiu, a traditional Chinese alcoholic beverage. Comprehensive two-dimensional gas chromatography mass spectrometry (GC×GC-qMS)-based untargeted metabolomics combined with a multivariate analysis was used to investigate the dynamic variations in the aroma profile of Huangjiu during aging process and to establish the relationship between the changing volatile metabolite profiles and the age-dependent sensory attributes. A total of 144 volatile metabolites were identified by GC×GC-qMS and 63 were selected as critical metabolites based on variable importance in projection values and p-values. Based on the results of principal component analysis, orthogonal partial least-squares discriminant analysis, and hierarchical clustering analysis, the samples of six different ages were divided into three groups: 1Y and 3Y samples, 5Y and 8Y samples, and 10Y and 15Y samples. The partial least-squares analysis results further revealed the relationship between the aromas attributes and variations of these volatile compounds. The high esters, aldehydes, and lactones contents contributed to the high intensities of the sweet and ester aroma attributes of the aged Huangjiu, while the high alcohols and ethyl esters contents contributed to the alcoholic and fruity aroma attributes of the newly brewed Huangjiu. These results improve our understanding of the chemical nature of the aroma characteristics of aged Huangjiu. PRACTICAL APPLICATION: Huangjiu is often labeled with its age as a measure of quality, which influences consumers' choice. Dynamic variations in volatile compounds of Huangjiu during aging and its contribution to the aroma characteristics of Huangjiu were figured out, which will assist the industry to produce better quality aged Huangjiu for consumers.

Pyrazinamide Resistance and Mutation Patterns Among Multidrug-Resistant Mycobacterium tuberculosis from Henan Province.

PURPOSE: This study was designed to identify the phenotypic and genotypic characteristics of pyrazinamide (PZA) resistance among multidrug-resistant Mycobacterium tuberculosis (MDR-TB) from Henan and to evaluate the efficacy of pncA, rpsA, and panD mutations in predicting PZA resistance. MATERIALS AND METHODS: A total of 152 MDR strains were included in this study. The Bactec MGIT system was used to determine PZA susceptibility for all strains. The pncA, rpsA, and panD genes were sequenced to identify any mutations, and the sequences were then aligned with the sequence of standard strain H37Rv. Moreover, the correlations between PZA-resistant phenotypes and treatment outcomes were analysed. RESULTS: Of the152 strains, 105 had a PZA-resistant phenotype, and 102 harboured the pncA mutation. The PZA resistance rate was higher in the strains with resistance to all four first-line drugs and those that were pre-extensively drug-resistant (pre-XDR) and extensively drug-resistant (XDR). A total of 100 different pncA mutation patterns were identified, including 80 point mutations and 20 insertions/deletions, and 32 new pncA mutation patterns were detected. In this study, 13 strains had multiple mutations. Of the11 PZA-resistant strains without pncA mutations, two harboured the rpsA mutation, and one harboured the panD mutation. With PZA susceptibility results as the reference, single-gene pncA sequencing had sensitivity of 89.52% and specificity of 89.36%. With the combination of rpsA and panD, the sensitivity increased to 92.38%, and the specificity remained the same. No significant differences were observed in the sputum smear/culture conversion rate between PZA-resistant patients and PZA-sensitive patients. However, PZA resistance was related to the time to sputum smear/culture conversion (P = 0.018). CONCLUSION: The combination of pncA, rpsA, and panD was beneficial for the timely diagnosis of PZA resistance and could provide a laboratory basis for customizing treatment regimens for MDR-TB patients.

Role of MIRU-VNTR and spoligotyping in assessing the genetic diversity of Mycobacterium tuberculosis in Henan Province, China.

BACKGROUND: Tuberculosis remains a serious threat to human health as an infectious disease in China. Henan, a most populated province in China, has a high incidence of tuberculosis (TB). Though the genetic diversity of Mycobacterium tuberculosis (MTB) has been investigated in many regions, there have been only a few studies on the molecular characteristics and drug resistance phenotypes in Henan. This is the first study on the genetic profile of MTB from Henan. METHODS: A total of 668 MTB isolates from various areas were genotyped with spoligotyping and 26-locus MIRU-VNTR (classical 24-locus MIRU-VNTR and 2 other loci). The association between TB spoligotype signatures and drug-resistant profiles was analysed. RESULTS: Our data revealed that MTB isolates circulating in Henan had a high degree of genetic variation. The Beijing family was the most predominant genotype (83.53%,n = 558), and the typical Beijing type(ST1) was the major sublineage (81.73%,n = 546). In total,668 isolates were divided into 567 different types, forming 38 clusters (2-15 isolates per cluster), and 529 unique types by 26-locus MIRU-VNTR analysis. There was no correlation between the Beijing family and gender, age at diagnosis or treatment history, whereas the Beijing family was significantly associated with all four first-line drug resistance and multidrug-resistant phenotypes. For these samples, 15 of 26 MIRU-VNTR loci had high or moderate discriminatory power according to the Hunter-Gaston discriminatory index. A combination of the 10 most polymorphic loci had similar discriminatory power as the 26-locus set. CONCLUSION: The Beijing genotype is the most prevalent family. Ten-locus MIRU-VNTR in combination with spoligotyping can efficiently classify the molecular type of MTB in Henan Province.

CD38 is a putative functional marker for side population cells in human nasopharyngeal carcinoma cell lines.

Cancer stem cells (CSCs) are thought to be responsible for cancer progression and therapeutic resistance but identification of this subpopulation requires selective markers. Fortunately, side population (SP) cells analysis brings a novel method to CSCs study. In this study, we identified SP cells, which are demonstrated rich in CSCs, in four nasopharyngeal carcinoma (NPC) cell lines. We investigated SP cells from HK-1 NPC cell line and showed CSCs characteristics in this subpopulation. SP cells displayed greater proliferation and invasion and expressed high levels of CSCs markers than NSP cells. Furthermore, our microRNA microarray analysis of SP versus NSP cells revealed that CD38-related miRNAs were down-regulated in SP cell, but the mRNA and protein level of CD38 were highly expressed in SP cells. We further searched for molecules interacting with CD38 and identified ZAP70, which was also well expressed in SP cells at both mRNA and protein levels. Our results uncover a CD38 pathway that may regulate the proliferation and migration of SP cells from HK-1 NPC cell line.

Dysregulation of the PI3K/Akt signaling pathway affects cell cycle and apoptosis of side population cells in nasopharyngeal carcinoma.

Increasing evidence has suggested that certain types of cancer possess their own stem-like cells, and that one subset of these cells, termed the side population (SP), may have an important role in tumorigenesis and cancer therapy. However, the molecular mechanisms underlying the modulation of SP cells in nasopharyngeal carcinoma (NPC) have remained elusive. In the present study, it was hypothesized that dysregulation of the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K)/Akt signaling pathway may influence SP and non-SP (NSP) phenotype. SP cells from the HK-1 NPC cell line were identified, and cancer stem cell markers were found to be highly expressed in SP cells compared with that of NSP cells. Freshly sorted SP cells demonstrated a significant increase in the proportion of cells in G0/G1 phase, while the majority of NSP cells were in the proliferative phase. Following 48 h of culture subsequent to cell sorting, the differences in cell cycle distribution between the SP and NSP cells converged. In addition, the apoptotic ratio of NSP cells was higher than that of SP cells at 24 h following sorting, but had no significant differences 48 h following sorting. To elucidate the potential mechanism mediating the cell cycle and apoptosis in SP cells, the expression levels of key molecules in the PI3K/Akt signaling pathway were evaluated. PI3K and Akt were upregulated, while 14-3-3σ protein was downregulated in SP cells when freshly sorted (0 h). However, there was no significant difference in the expression of these molecules between SP and NSP cells following 48 h of culture. These results suggested that dysregulation of the PI3K/Akt signaling pathway may be associated with the cell cycle and apoptosis of SP cells in NPC. However, further investigation is required to elucidate the detailed mechanisms underlying these effects.

Dynamic changes and functions of macrophages and M1/M2 subpopulations during ulcerative colitis-associated carcinogenesis in an AOM/DSS mouse model.

The high risk of developing colorectal carcinoma (CRC), from ulcerative colitis (UC), is well known. Macrophages are widely distributed immune cells that have an indispensable role in UC, as well as in CRC. However, little is currently known about the dynamic changes that occur in macrophage and M1/M2 macrophage subpopulations, during UC­associated carcinogenesis. The aim of the present study was to investigate the alteration of colorectal macrophages and M1/M2 macrophage subpopulations during UC­associated carcinogenesis. Both expression level alterations and functional changes were determined during UC­associated carcinogenesis in an azoxymethane/dextran sodium sulfate­induced chemically colitis­associated carcinoma mouse model of Crj:CD­1 (ICR) mice. Notable evidence from immunohistochemistry, flow cytometry, cytokine detection, and gene expression analyses demonstrated that M2 macrophages have a critical role in CRC initiation, promotion, and metastasis. M2 macrophages are associated with unbalanced pro­inflammatory and anti­inflammatory axes and aberrant enhancement of migration/invasion­associated factors. Functional changes, similar to M2 polarized macrophages, were shown to occur in the M1 macrophages, without phenotypical changes, during the development of carcinoma and metastasis. The results of the present study suggest that M2 macrophages have a pro­tumor role during UC­associated carcinogenesis. Furthermore, similar functional changes occurred in the M1 macrophages, without polarization alterations, during carcinogenesis and metastasis.

CD38 is highly expressed and affects the PI3K/Akt signaling pathway in cervical cancer.

Cervical cancer is the second most common cancer and the fifth most deadly malignancy in females worldwide, affecting 500,000 individuals each year. It is the leading cause of cancer mortality among women in developing countries. Dysregulated activation of genes, such as CD44, SOX9 and SKP2, plays a role in cervical cancer. CD38 is known to be involved in activities typical of cell surface receptors, such as signaling for activation and proliferation events and heterotypic cell adhesion. CD38 contributes to disease progression and relapse in certain tumors, such as acute myeloid and chronic lymphocytic leukemia. To the best of our knowledge, there is currently no report on the relationship between CD38 and cervical cancer. Using qPCR, immunohistochemistry, and western blot analysis, the expression levels of CD38 were investigated and found to be upregulated in cervical cancer. CD38 was correlated with dysregulation of the phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway in cervical cancer tissues in vitro. At the same time, CD38 overexpression affected the expression of PI3K, Akt, MDM2 and p53 in vivo. The results of the present study suggested that CD38 is highly expressed in cervical carcinoma tissues and play an important role in dysregulation of the PI3K/Akt signaling pathway.

[Side population cells and progress in cancer stem cell research].

In recent years, cancer stem cells have become a hotspot for global researchers. Cancer stem cell theory deems that cells with self-renewal and differentiation potential play a key role in tumor resistance and relapse. These cells are named cancer stem cells. At present, the sorting methods include the side population cell sorting technique, screening techniques based on cell surface special markers, tumor sphere cultures, label retaining cell, ALDEFLUOR assays and so on. Side population cells are a small part of cells with the capacity of efflux DNA fluorescent dye Hoechst 33342 and present a low staining intensity in flow cytometry plot. Side population cells are rich in cancer stem cells, and its sorting method has been considered simple and effective in cancer stem cell research.

Dynamic changes of peritoneal macrophages and subpopulations during ulcerative colitis to metastasis of colorectal carcinoma in a mouse model.

OBJECTIVE AND DESIGN: Patients with ulcerative colitis have increased risk of colorectal carcinoma, but little is known about how peritoneal macrophages are involved in ulcerative colitis-associated carcinogenesis. We investigated the alteration of peritoneal macrophages and M1/M2 subpopulations during ulcerative colitis-associated carcinogenesis. MATERIALS AND METHODS: Expression and functional changes in peritoneal macrophages and M1/M2 subpopulations were investigated by histopathology, flow cytometry, immunofluorescence, cytokines expression by ELISA and QRT-PCR in an azoxymethane (AOM)- and dextran sodium sulfate (DSS)-induced chemical colitis-associated carcinoma mouse model using male Crj:CD-1 (ICR) mice. RESULTS: Striking evidence observed in histopathology, flow cytometry, cytokine detection, and gene expression analysis all revealed that inflammation-associated cytokines (IL-1ß, IL-10, IL-12, IL-6, TNF-α) and migration/invasion-associated factors (G-CSF, GM-CSF, CXCR4, VEGF, TGF-ß, ICAM-1) induced by peritoneal M2 macrophages increased significantly during the progression from inflammatory hyperplasia to carcinoma and metastasis. Similar functional changes occurred during peritoneal metastasis in M1 macrophages without changed polarization. CONCLUSIONS: These results suggested that peritoneal M2 macrophages played a critical role in ulcerative colitis-associated carcinogenesis, including unbalanced pro-inflammatory and anti-inflammatory axis and enhanced expression of migration/invasion-associated factors. Furthermore, functional changes of M1 macrophages occurred without changed polarization during carcinogenesis and metastasis.

Risk of nasopharyngeal carcinoma associated with polymorphic lactotransferrin haplotypes.

Lactotransferrin (LTF) is a component of the nonspecific immune system, having antimicrobial properties against bacteria, fungi, and several viruses. The gene coding for LTF is polymorphic, with the occurrence of several common alleles in the general population. Our previous study found that LTF inhibited nasopharyngeal carcinoma (NPC) cell proliferation in vitro and in vivo. To better understand one possible mechanism of LTF-mediated antitumor activity in NPC cells, in the present study, we investigated the distribution of LTF gene polymorphisms (rs1126477, rs1126478, rs2073495, and rs9110) in NPC and revealed whether these polymorphisms were associated with LTF gene expression. It was found that rs2073495 and rs9110 were correlated significantly with NPC. The frequency of CC genotype was higher and GG or TT genotype was lower, in NPC patients compared with that in the control group (P < 0.05, χ(2) = 8.73 and 9.33, respectively). CC genotype is the risk factor for NPC. Haplotype analyses indicated that NPC patients had lower rate of 'A-G-G-T' haplotype (constructed with rs1126477, rs1126478, rs2073495, and rs9110) compared with controls (P = 4.12 × 10(-6) < 0.001, χ(2) = 21.25). The population with 'A-G-G-T' haplotype had 0.322-fold risk to be NPC. The expression of LTF gene was high in NPC tissues and control tissues with 'A-G-G-T' haplotype compared with these without its. These findings suggested that rs2073495 and rs9110 could play important roles in NPC physiological processes.