Prognostic Ability of Enhancer RNAs in Metastasis of Non-Small Cell Lung Cancer.

(1) Background: Non-small cell lung cancer (NSCLC) is the most common lung cancer. Enhancer RNA (eRNA) has potential utility in the diagnosis, prognosis and treatment of cancer, but the role of eRNAs in NSCLC metastasis is not clear; (2) Methods: Differentially expressed transcription factors (DETFs), enhancer RNAs (DEEs), and target genes (DETGs) between primary NSCLC and metastatic NSCLC were identified. Prognostic DEEs (PDEEs) were screened by Cox regression analyses and a predicting model for metastatic NSCLC was constructed. We identified DEE interactions with DETFs, DETGs, reverse phase protein arrays (RPPA) protein chips, immunocytes, and pathways to construct a regulation network using Pearson correlation. Finally, the mechanisms and clinical significance were explained using multi-dimensional validation unambiguously; (3) Results: A total of 255 DEEs were identified, and 24 PDEEs were selected into the multivariate Cox regression model (AUC = 0.699). Additionally, the NSCLC metastasis-specific regulation network was constructed, and six key PDEEs were defined (ANXA8L1, CASTOR2, CYP4B1, GTF2H2C, PSMF1 and TNS4); (4) Conclusions: This study focused on the exploration of the prognostic value of eRNAs in the metastasis of NSCLC. Finally, six eRNAs were identified as potential markers for the prediction of metastasis of NSCLC.

Platelets Restrict the Oxidative Burst in Phagocytes and Facilitate Primary Progressive Tuberculosis.

Rationale: Platelets are generated in the capillaries of the lung, control hemostasis, and display immunological functions. Tuberculosis primarily affects the lung, and patients show platelet changes and hemoptysis. A role of platelets in immunopathology of pulmonary tuberculosis requires careful assessment.Objectives: To identify the dynamics and interaction partners of platelets in the respiratory tissue and establish their impact on the outcome of pulmonary tuberculosis.Methods: Investigations were primarily performed in murine models of primary progressive pulmonary tuberculosis, by analysis of mouse strains with variable susceptibility to Mycobacterium tuberculosis infection using platelet depletion and delivery of antiplatelet drugs.Measurements and Main Results: Platelets were present at the site of infection and formed aggregates with different myeloid subsets during experimental tuberculosis. Such aggregates were also detected in patients with tuberculosis. Platelets were detrimental during the early phase of infection, and this effect was uncoupled from their canonical activation. Platelets left lung cell dynamics and patterns of antimycobacterial T-cell responses unchanged but hampered antimicrobial defense by restricting production of reactive oxygen species in lung-residing myeloid cells.Conclusions: Platelets are detrimental in primary progressive pulmonary tuberculosis, orchestrate lung immunity by modulating innate immune responsiveness, and may be amenable to new interventions for this deadly disease.

A deletion in the RD105 region confers resistance to multiple drugs in Mycobacterium tuberculosis.

BACKGROUND: The emergence of drug-resistant strains of Mycobacterium tuberculosis (Mtb), especially those that are multidrug resistant poses a serious threat to global tuberculosis control. However, the mechanism underlying the occurrence of drug resistance against more than one drug is poorly understood. Given that the Beijing/W strains are associated with outbreaks and multidrug resistance, they may harbor a genetic advantage and provide useful insight into the disease. One marker found in all Beijing/W Mtb strains is a deletion of RD105 region that results in a gene fusion, Rv0071/74, with a variable number (3-9 m) of VDP (V: Val, D: Asp; P: Pro) repeats (coded by gtggacccg repeat sequences) at the N-terminal. Here, we report that this variable number of VDP repeats in Rv0071/74 regulates the development of multidrug resistance. RESULTS: We collected and analyzed 1255 Beijing/W clinical strains. The results showed that the number of VDP repeats in Rv0071/74 was related to the development of multidrug resistance, and the deletion of Rv0071/74-9 m from Beijing/W clinical strain restored drug susceptibility. Rv0071/74-9 m also increased resistance to multiple drugs when transferred to different mycobacterial strains. Cell-free assays indicate that the domain carrying 4-9 VDP repeats (4-9 m) showed a variable binding affinity with peptidoglycan and Rv0071/74 cleaves peptidoglycan. Furthermore, Rv0071/74-9 m increased cell wall thickness and reduced the intracellular concentration of antibiotics. CONCLUSIONS: These findings not only identify Rv0071/74 with VDP repeats as a newly identified multidrug resistance gene but also provide a new model for the development of multiple drug resistance.

Notch4 Negatively Regulates the Inflammatory Response to Mycobacterium tuberculosis Infection by Inhibiting TAK1 Activation.

Tuberculosis, caused by Mycobacterium tuberculosis infection, remains a global threat to human health, but knowledge of the molecular mechanisms underlying the pathogenesis of tuberculosis is still limited. Although Notch4, a member of the Notch receptor family, is involved in the initiation of mammary tumors, its function in M. tuberculosis infection remains unclear. In this study, we found that Notch4-deficient mice were more resistant to M. tuberculosis infection, with a much lower bacterial burden and fewer pathological changes in the lungs. Notch4 inhibited M. tuberculosis-induced production of proinflammatory cytokines by interaction with TAK1 and inhibition of its activation. Furthermore, we found that Notch intracellular domain 4 prevented TRAF6 autoubiquitination and suppressed TRAF6-mediated TAK1 polyubiquitination. Finally, Notch inhibitors made mice more resistant to M. tuberculosis infection. These results suggest that Notch4 is a negative regulator of M. tuberculosis-induced inflammatory response, and treatment with a Notch inhibitor could serve as a new therapeutic strategy for tuberculosis.

Integrative Analysis of Proteome and Ubiquitylome Reveals Unique Features of Lysosomal and Endocytic Pathways in Gefitinib-Resistant Non-Small Cell Lung Cancer Cells.

Non-small cell lung cancer (NSCLC) patients carrying EGFR activating mutations treated with gefitinib, a tyrosine kinase inhibitor, will develop drug resistance. Ubiquitylation is one of major posttranslational modifications of proteins affecting the stability or function of proteins. However, the role of protein ubiquitylation in gefitinib resistance is poorly understood. To systematically identify the global change in protein expression and ubiquitylation during gefitinib resistance, a quantitative global proteome and ubiquitylome study in a pair of gefitinib-resistant and sensitive NSCLC cells is carried out. Altogether, changes in expression of 3773 proteins are quantified, and changes in ubiquitylation of 2893 lysine sites in 1415 proteins are measured in both cells. Interestingly, lysosomal and endocytic pathways, which are involved in autophagy regulation, are enriched with upregulated proteins or ubiquitylated proteins in gefitinib-resistant cells. In addition, HMGA2 overexpression or ALOX5 knockdown suppresses gefitinib resistance in NSCLC cells by inhibiting autophagy. Overall, these results reveal the previously unknown global ubiquitylome and proteomic features associated with gefitinib resistance, uncover the opposing roles of HMGA2 or ALOX5 in regulating gefitinib resistance and autophagy, and will help to identify new therapeutic targets in overcoming gefitinib resistance.

Platelets direct monocyte differentiation into epithelioid-like multinucleated giant foam cells with suppressive capacity upon mycobacterial stimulation.

BACKGROUND: Epithelioid, foam, and multinucleated giant cells (MNGCs) are characteristics of tuberculosis granulomas, yet the precise genesis and functions of these transformed macrophages are unclear. We evaluated the role of platelets as drivers of macrophage transformation in mycobacterial infection. METHODS: We employed flow cytometry and microscopy to assess cellular phenotype and phagocytosis. Immune assays allowed quantification of cytokines and chemokines, whereas gene microarray technology was applied to estimate global transcriptome alterations. Immunohistochemical investigations of tuberculosis granulomas substantiated our findings at the site of infection. RESULTS: Monocytes differentiated in presence of platelets (MP-Macs) acquired a foamy, epithelioid appearance and gave rise to MNGCs (MP-MNGCs). MP-Macs up-regulated activation markers, phagocytosed mycobacteria, and released abundant interleukin 10. Upon extended culture, MP-Macs shared transcriptional features with epithelioid cells and M2 macrophages and up-regulated CXCL5 transcripts. In line with this, CXCL5 concentrations were significantly increased in airways of active tuberculosis patients. The platelet-specific CD42b antigen was detected in MP-Macs, likewise in macrophages, MNGCs, and epithelioid cells within tuberculosis granulomas, along with the platelet aggregation-inducing factor PDPN. CONCLUSIONS: Platelets drive macrophage differentiation into MNGCs with characteristics of epithelioid, foam, and giant cells observed in tuberculosis granulomas. Our data define platelets as novel participants in tuberculosis pathogenesis.

Clinical efficacy of tetrandrine in artificial stone-associated silicosis: A retrospective cohort study.

Background: Outbreaks of silicosis have occurred among workers in the artificial stone (AS) industry, and there is currently no effective antifibrosis treatment for silicosis. Design: A retrospective cohort study. Methods: We retrospectively analyzed the clinical data of 89 artificial stone-associated silicosis patients treated in Shanghai Pulmonary Hospital (China). Patients who agreed to be administered tetrandrine entered the observation group and those who disagreed entered the control group. Changes in chest HRCT, pulmonary function, and clinical symptoms of patients in two groups were compared pre- and post-treatment. Results: After treatment for 3-12 months, 56.5%-65.4% of patients in the observation group showed improvements in HRCT imaging, while there was no improvement in the control group (p < 0.05). Disease progression occurred in 0%-17.4% of patients in the observation group after 3-12 months of treatment compared with 44.4%-92.0% of patients in the control group (p < 0.05). After 3 months of treatment, the forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1), and diffusing capacity of the lung for carbon monoxide (DLco) in the observation group increased by 136.7 ± 189.2 mL (p < 0.05), 124.2 ± 169.9 mL (p < 0.05), and 1.4 ± 2.3 mL/min/mmHg (p > 0.05), respectively, while those in the control group decreased (145.8 ± 356.5; 107.5 ± 272.1; 1.9 ± 3.8). After 6 months of treatment, FVC, FEV1, and DLco in the observation group increased by 207.8 ± 372.2 mL (p > 0.05), 107.8 ± 295.2 mL (p > 0.05) and 0.7 ± 6.0 mL/min/mmHg (p > 0.05), respectively, while those of the control group decreased (383.3 ± 536.7; 215.6 ± 228.9; 1.4 ± 1.7). The incidences of clinical symptoms such as cough, expectoration, dyspnea, chest tightness, and chest pain in the observation group were decreased-after treatment (all p < 0.05), while the incidences of these symptoms increased in the control group, although the change was not statistically significant (all p > 0.05). Conclusion: Tetrandrine can control and delay the progression of AS-associated silicosis fibrosis, with improved chest HRCT imaging and pulmonary function.

Clinical value of ELISA-MPT64 for the diagnosis of tuberculous pleurisy.

Tuberculous pleurisy is one of the common extrapulmonary tuberculosis diseases. However, the diagnosis of tuberculous pleurisy still lacks a useful and effective tool, mainly due to paucity of Mycobacterium tuberculosis organisms in pleural effusion. Previous studies have confirmed that the MPT64 protein is highly specific and is secreted only by M. tuberculosis (MTB) complex. Therefore, in this study, we developed ELISA based on recombinantly expressed MPT64 in combination with rabbit polyclonal antibodies. The ELISA-MPT64 method was validated using MTB strains and tested against clinical samples. Nested PCR, Löwenstein-Jensen (L-J) culture and smear microscopy were employed as the comparative tools for assessing the performance of the assay. Our results demonstrate that the newly established ELISA-MPT64 technique is a rapid and useful tool for the diagnosis of tuberculous pleurisy.

Risk factors for neonatal hyperbilirubinemia: a systematic review and meta-analysis.

Background: Hyperbilirubinemia is the most common cause of neonatal hospitalization and, although it generally has a good prognosis, a significant percentage of neonatal patients maintain a high bilirubin level, which can lead to severe complications, including lifelong disability such as growth retardation, encephalopathy, autism and hearing impairment. The study of risk factors for neonatal hyperbilirubinemia has been controversial. Therefore, we evaluated the risk factors of neonatal hyperbilirubinemia using a meta-analysis. Methods: Relevant English and Chinese studies that discussed risk factors for neonatal hyperbilirubinemia were retrieved from the PubMed, EMBASE, Medline, Central, China National Knowledge Infrastructure (CNKI), Wanfang and China Science Digital Library (CSDL). The literature took newborns as the research object, set up a control group, and observed the relationship between exposure factors and neonatal hyperbilirubinemia. The combined effect size was expressed by odds ratio (OR) and 95% confidence interval (CI). The Chi-square test was used to test heterogeneity of the studies, and if it existed, subgroup analyses were used to explore the source of heterogeneity, and the random-effects model was selected for the combined analysis. The fixed-effects model was chosen for the combined analysis if there was no heterogeneity. Publication bias was assessed using Egger's test and funnel plot. Results: Risk factors for neonatal hyperbilirubinemia were exclusive breastfeeding (BF: OR =1.74, 95% CI: 1.42, 2.12, Z=5.43, P<0.00001); glucose-6-phosphate dehydrogenase deficiency (G6PD: OR =1.62, 95% CI: 1.44, 1.81, Z=8.39, P<0.00001); maternal-fetal ABO blood group incompatibility (OR =1.64, 95% CI: 1.42, 1.89, Z=6.75, P<0.00001); and preterm birth (PTB: OR =1.31, 95% CI: 1.17, 1.47, Z=4.60, P<0.00001); there was no heterogeneity or publication bias among the studies (BF: χ2=5.34, P=0.25, I2=25%; G6PD: χ2=4.40, P=0.49, I2=0%; ABO: χ2=1.91, P=0.75, I2=0%; PTB: χ2=0.81, P=0.67, I2=0%). Conclusions: Exclusive breastfeeding, G6PD deficiency, ABO incompatibility and premature birth were confirmed as risk factors for neonatal hyperbilirubinemia. Pregnant women with risk factors should be monitored more closely and clinical intervention should be given in a timely manner.

Evaluation of Aptamer Fluorescence Microscopy in the Diagnosis of Pulmonary Tuberculosis.

Sputum smear microscopy for tuberculosis diagnosis has stood the test of time. However, due to its low sensitivity, the positive detection rate for tuberculosis in clinical specimens is not high. To improve the sensitivity of microscopic observation in Mycobacterium tuberculosis (MTB) detection, we developed the MTB-specific aptamer MA1. To further improve the binding reactivity of MA1 with MTB, we constructed a new derivative aptamer with a pocket-stem-loop-structure, MA1-39, and identified it to have high binding reactivity with the MTB reference strain. We developed an aptamer fluorescence microscopy test for MTB based on MA1-39 and evaluated its feasibility for diagnosing pulmonary tuberculosis. Among 56 tested strains, MA1-39 was proven to effectively discriminate MTB from the control strains, including 12 non-tuberculosis mycobacterial (NTM) reference strains, 6 NTM isolates, and 7 other bacteria. Next, this approach was applied to 169 clinical sputum samples from suspected tuberculosis patients and non-tuberculosis controls. Molecular tests together with both clinical and bacteriological identification were used as a protocol to evaluate the efficacy of aptamer detection. Compared with the traditional acid-fast staining light microscope, the aptamer fluorescence microscope showed a higher detection rate for MTB in clinical specimens (48.8% versus 32.6%), and the specificities of the two techniques had almost no significant difference (90.4% versus 94%). In addition, aptamer fluorescence microscopy showed the same positive predictive value (PPV) as staining (84% versus 84.9%), but a higher negative predictive value (NPV; 63% versus 57.3%). In conclusion, the newly established aptamer fluorescence microscopy approach is likely to be a feasible method for microbiological diagnosis of tuberculosis. IMPORTANCE We established an aptamer fluorescence microscopy approach for rapid detection of MTB in clinical sputum samples. The use of aptamer probes was proven to significantly increase the sensitivity of sputum smear microscopy. In resource-limited countries, microscopy is currently a fast, simple, and very common test method in many laboratories, and it will remain the primary means of microbiological diagnosis of tuberculosis in the foreseeable future. Improving detection techniques can further enhance the clinical application value of this ancient diagnostic method. Since aptamer fluorescence microscopy can provide rapid and sensitive results, it may be a feasible and useful method in resource-limited settings.

Risk Factors of Silicosis Progression: A Retrospective Cohort Study in China.

Background: Silicosis poses a threat to workers' health due to the irreversible lung lesions. Design: A retrospective cohort study. Methods: A total of 259 patients [80 worked with artificial stone (AS), 179 with non-artificial stone (non-AS)] with confirmed silicosis were included in this study. Forty-one of AS and 91 of non-AS had approximately 2 years' follow-up records [lung function tests and high-resolution computer tomography (HRCT)]. Compared with the first records, increased, densified, or newly emerging lesions in lung HRCT images were judged as progression of the disease. Cox proportional hazards models were used to determine the risk factors. Kaplan-Meier survival curve and log-rank test were used to compare prognostic factors for cumulative risk of progression. Results: In 132 patients with median follow-up of 24.0 months (IQR, 13.8, 24.9), 66 patients showed progression, in them, 36 (87.8%) were from AS group and 30 (32.9%) from non-AS group. Working experience of AS processing (hazard ratio, 5.671; 95% CI, 3.048-10.550) and complicated silicosis in CT images (hazard ratio, 2.373; 95% CI, 1.379-4.082) were the main risk factors associated with progression. Forced vital capacity decreased after 1-year (241.5 vs. 55.2 mL) and 2-year (328.1 vs. 68.8 mL) follow-up in the two groups (AS vs. non-AS). History of anti-tuberculosis medication, chest oppression and pain, ground-glass opacity, pleural abnormalities, and restrictive pulmonary dysfunction were more frequently found on HRCT images in the AS group than non-AS group. Lung functions (DLCO, %) were lower in the current/former smokers than the non-smokers (P < 0.05) in AS patients. Conclusion: Prevention and protection rules are needed to be enforced in the occupation involving AS processing; smoking may be associated with declined lung function in AS patients.

Perspective on sequence evolution of microsatellite locus (CCG)n in Rv0050 gene from Mycobacterium tuberculosis.

BACKGROUND: The mycobacterial genome is inclined to polymerase slippage and a high mutation rate in microsatellite regions due to high GC content and absence of a mismatch repair system. However, the exact molecular mechanisms underlying microsatellite variation have not been fully elucidated. Here, we investigated mutation events in the hyper-variable trinucleotide microsatellite locus MML0050 located in the Rv0050 gene of W-Beijing and non-W-Beijing Mycobacterium tuberculosis strains in order to gain insight into the genomic structure and activity of repeated regions. RESULTS: Size analysis indicated the presence of five alleles that differed in length by three base pairs. Moreover, nucleotide gains occurred more frequently than loses in this trinucleotide microsatellite. Mutation frequency was not completely related with the total length, though the relative frequency in the longest allele was remarkably higher than that in the shortest. Sequence analysis was able to detect seven alleles and revealed that point mutations enhanced the level of locus variation. Introduction of an interruptive motif correlated with the total allele length and genetic lineage, rather than the length of the longest stretch of perfect repeats. Finally, the level of locus variation was drastically different between the two genetic lineages. CONCLUSION: The Rv0050 locus encodes the bifunctional penicillin-binding protein ponA1 and is essential to mycobacterial survival. Our investigations of this particularly dynamic genomic region provide insights into the overall mode of microsatellite evolution. Specifically, replication slippage was implicated in the mutational process of this microsatellite and a sequence-based genetic analysis was necessary to determine that point mutation events acted to maintain microsatellite size integrity while providing genomic diversity.

Study on the effectiveness and safety of Xingpi Yanger granule combined with Saccharomyces boulardii for rotavirus enteritis in children: A protocol for systematic review and meta-analysis.

BACKGROUND: To systematically evaluate the effectiveness and safety of traditional Chinese medicine preparation XPYEG combined with SBI and SBI alone in the treatment of REC, and to provide the reference in drugs for the clinical treatment of children with rotavirus enteritis. METHODS: Retrieving the English databases: PubMed, Cochrane Library and Embase; Chinese databases: CNKI, CBM and WANFANG Data. Retrieving a randomized controlled trial of XPYEG and SBI in the treatment of REC. The retrieval time is from the above database until September 2020. The retrieval strategy of combining free words and subject words is adopted, and the references included in the literature are searched manually in accordance with the literature studied in this paper and not included in the above database. Two researchers screen the literature according to the literature inclusion and exclusion criteria, extract valid data and evaluate the quality of the literature, and cross-check it. Using the RevMan 5.3 software to conduct the meta-analysis on the main outcome and secondary outcome indicators of the included literature, while assessing the evidence quality of included study. RESULTS: The effectiveness and safety of XPYEG and SBI in the treatment of REC are presented through the main and secondary outcome indicators. OSF REGISTRATION NUMBER: DOI 10.17605/OSF.IO/3QSZG. CONCLUSION: This study will conclude whether the combination of XPYEG and SBI is more effective than SBI alone in the treatment of REC, and whether the medication increases the risk of adverse reactions compared with single medication. ETHICS AND DISSEMINATION: This study does not involve the specific patients, and all research data comes from publicly available professional literature, so an ethics committee is not required to conduct an ethical review and approval of the study.

Identification and evaluation of a new nucleic acid amplification test target for specific detection of Mycobacterium tuberculosis.

BACKGROUND: Accurate and early diagnosis of tuberculosis (TB) is of major importance in the management and control of TB. Because the conventional bacteriological diagnosis of TB has several limitations, nucleic acid amplification (NAA) tests have emerged as promising alternatives. A potential problem with NAA tests is that some strains lack a target, which may be the one of main reasons for the much lower and highly variable accuracy in diagnosis. A possible solution may be to use more valid and applicable targets to increase detection accuracy. METHODS: In this paper, we designed a two-step program to obtain NAA test targets. Inter-simple sequence repeats (ISSR) based on oligonucleotide (GTG)(5) were first constructed to genotype Mycobacterium strains to obtain Mycobacterium tuberculosis (MTB)-specific fragment. Second, sequence characterized amplified region (SCAR) markers were developed from these species-specific sequences to identify MTB. Some 312 Mycobacterium strains were used to evaluate the efficacy of the SCAR markers, IS6110 element [specific identification of Mycobacterium tuberculosis complex (MTC)] and 16SrRNA gene (specific identification of Mycobacterium) amplification, together with traditional bacteriology testing was used as a control. RESULTS: MTB-specific sequences located in a gene coding for Rv1508c, as a new NAA test target, were obtained using ISSR-PCR genotyping. Based on these sequences, the SCAR primer pairs MISP1 and MISP2 were designed. All 312 strains from Mycobacterium accurately produced the genus-specific 16SrRNA amplicon. 271 MTB strains and M. africanum were positive. However, all nontuberculous mycobacteria (NTM) strains and 1 MTB strain named 1143 were negative in both SCAR and IS6110 PCR amplification. M. bovis, bacille Calmette-Guérin (BCG) were IS6110-PCR positive, while SCAR-PCR was negative. Strain 1143 was defined as M. arupense with 99% identity by 16SrRNA gene sequencing identification, despite being diagnosed as MTB using traditional testing. CONCLUSIONS: SCAR markers developed with this two-step program can be used as a new NAA test target to correctly detect MTB.

The effect of preventive use of hydrolyzed protein formula milk on gastrointestinal diseases and physical development of premature infants: A protocol for systematic review and meta-analysis.

BACKGROUND: Because of the controversy in clinical nutritional support therapy of hydrolyzed protein formula milk and standard preterm infant formula (SPIF) in premature infants. In this study, the effectiveness and safety of preventive use of hydrolyzed protein formula milk in reducing gastrointestinal diseases and promoting physical development of premature infants are scientifically evaluated by systematic evaluation. To help find the suitable nutritional support for premature infants. METHODS: To search the database of Chinese and English by computer: SinoMed, CNKI, WanFang Data, VIP, PubMed, EMbase and The Cochrane Library, and to collect randomized controlled trials on the application of hydrolyzed protein formula milk in nutritional support treatment of premature infants compared with SPIF. The retrieval time limit is from the establishment of each database to September 1, 2020. Two authors independently completed the paper search, and sorting out the main outcome indicator and secondary outcome indicator in the selected literature, and the data are statistically analyzed by Review Manager software (RevManV.5.3.0) and STATA 13.0. RESULTS: This study will provide a high-quality evidence on the effects of hydrolyzed protein formula milk on gastrointestinal diseases and physical development of premature infants. CONCLUSION: At present, the clinicians are controversial about the safety and effectiveness of hydrolyzed protein formula milk and SPIF in the nutritional support therapy of premature infants. This study will compare the effectiveness and safety of these 2 nutritional support methods, and make a comprehensive analysis of the influence of hydrolyzed protein formula milk on gastrointestinal diseases and physical development of premature infants, and finally give a positive conclusion. OSF REGISTRATION NUMBER: DOI 10.17605/OSF.IO/UQD92.

Changes in Physical Meat Traits, Protein Solubility, and the Microstructure of Different Beef Muscles during Post-Mortem Aging.

This study was performed to compare the differences in pH, myofibril fragmentation index (MFI), total protein solubility (TPS), sarcoplasmic protein solubility (SPS), myofibrillar protein solubility (MPS), and the microstructure of seven beef muscles during aging. From the six beef carcasses of Xinjiang brown cattle, a total of 252 samples from semitendinosus (ST), longissimus thoracis (LT), rhomboideus (RH), gastrocnemius (GN), infraspinatus (IN), psoas major (PM), and biceps femoris (BF) muscles were collected, portioned, and assigned to six aging periods (1, 3, 7, 9, 11, and 14 day/s) and 42 samples were used per storage period. IN muscle showed the highest pH (p < 0.05) from 1 to 14 days and the lowest TPS (p < 0.01) from 9 to 14 days with respect to the other muscles. Moreover, the changes in IN were further supported by transmission electron microscopy due to the destruction of the myofibril structure. The highest value of MFI was tested in ST muscle from 7 to 14 days. The total protein solubility in PM, RH, and GN muscles were not affected (p > 0.05) as the aging period increased. The lowest TPS was found in the RH muscle on day 1, 3, and 7 and in the IN muscle on day 9, 11, and 14. The pH showed negative correlations with the MFI, TPS, and MPS (p < 0.01). The results suggest that changes in protein solubility and muscle fiber structure are related to muscle location in the carcass during aging. These results provide new insights to optimize the processing and storage of different beef muscles and enhance our understanding of the biological characteristics of Xinjiang brown cattle muscles.

[Identification of the Mycobacterium tuberculosis complex by detecting recombinant secretory protein MPT64].

OBJECTIVE: To identify the Mycobacterium tuberculosis complex by detecting the secretory protein MPT64. METHODS: The gene mpt64 was amplified by polymerase chain reaction (PCR) from the genome of Mycobacterium tuberculosis H(37)Rv strain and cloned into expression vector. Immune sera from rabbits by recombinant proteins MPT64, were used to make enzyme-labeled antibodies and coated antibodies. A double antibody sandwich enzyme-linked immunosorbent assay (ELISA) was established to detect the secretory protein MPT64 of the culture supernatants in Mycobacterium strains. Results of ELISA were compared to those of the gene mpt64 amplified by PCR. RESULTS: A recombinant vector was constructed. The minimum detectable concentration of MPT64 was 0.01 mg/L. A total of 27 reference strains and 170 clinical isolate strains were evaluated. PCR for Mycobacterium tuberculosis reference strain, Mycobacterium bovis reference strain and Mycobacterium africanum reference strain was positive, but that for other reference stains was negative, consistent with the results of ELISA. In the 170 clinical isolate stains, the positive result of PCR and ELISA was 98.2% (111/113) and 97.3% (110/113) respectively, while the specificity of PCR and ELISA was both 100%, no positive result in non-tuberculosis mycobacterium strains. CONCLUSION: Identification of the Mycobacterium tuberculosis complex by detecting secretory protein MPT64 is rapid, sensitive, and specific, which can be used routinely in clinical laboratories.

Sex Disparity in Severity of Lung Lesions in Newly Identified Tuberculosis Is Age-Associated.

Background: The age-associated characteristic of computed tomography (CT) images of tuberculosis (TB) and the reason for male bias in TB are still not clear. Methods: We compared the CT images, clinical inflammatory indices and sputum bacterial counts between 594 non-smoking men and women with newly diagnosed TB with matched large span of ages from 15 to 92 years old. Logistic regression analyses were used to identify the cavity-associated factors of men and women, separately and in combination. Results: Sputum bacterial counts, ratio of cavities, lung injury scores, and level of C reactive protein were significantly higher in men than in women with ages from 15 to 74, but not in cases older than 75. In CT images, thick walled cavity, cicatricial emphysema and parenchymal bands were present in men at ages of 15-74 more than matched women. Ratios of cases with lobular emphysema and pleural effusion were higher in men after age of 56. While ratios of cases with parenchymal bands, calcification, pleural effusion, pleural thickening, lobular emphysema and bronchovascular distortion increased with aging, those of centrilobular nodules, micronodules and tree in bud decreased with aging in men. Erythrocyte sedimentation rate (ESR) increased with aging, but no differences were found between men and women in ESR or T-SPOT TB tests. Higher complement C4 and lower body mass index in men and positive result in anti-TB antibody test in women were strongly associated with the presence of cavity. Conclusions: The sex bias in TB is age-associated. TB prevention, treatment and research should take differences of sex and age into account.

Continuous treatment with recombinant Mycobacterium tuberculosis CFP-10-ESAT-6 protein activated human monocyte while deactivated LPS-stimulated macrophage.

Influence of the recombinant culture filtered protein 10 (CFP-10) and early-secreted antigenic target 6kDa protein (ESAT-6) (r-CFP-10-ESAT-6, rCE) of Mycobacterium tuberculosis (Mtb) on human monocyte and macrophage activation was investigated using human monocyte, monocyte like THP-1 cell line and monocyte derived macrophage (MDM). rCE solely enhanced TNF-alpha release from human monocytes and THP-1 cells in a dose- and time-dependent manner. rCE enhanced expression of CD80 and CD40, it also synergized with IFN-gamma in induction of TNF-alpha production and HLA-DR expression. Pharmacological agents that selectively inhibit mitogen activated protein kinase activation markedly suppressed rCE-induced TNF- alpha release. However, continuous presence of rCE (>72h) during monocyte to macrophage differentiation inhibited macrophage response to LPS stimulation. Collectively, these data suggest that rCE might have differential influence on monocyte and macrophage activation, which might be correlated with Mtb immune evasion.