Novel clinical biomarkers in blood and pleural effusion for diagnosing patients with tuberculosis distinguishing from malignant tumor.

Pleural effusion (PE) is a common manifestation of tuberculosis (TB) and malignant tumors but tuberculous PE (TPE) is difficult to distinguish from malignant PE (MPE), especially by noninvasive detection indicators. This study aimed to find effective detection indices in blood and PE for differentiating TB from a malignant tumor. A total of 815 patients who were diagnosed with TB or cancer in Hubei Shiyan Taihe Hospital from 2014 to 2017 were collected. Amongst them, 717 were found to have PE by thoracoscopy. Clinical characteristics, patients' blood parameters and PE indicator information were summarized for analysis. Patients with MPE had higher percentages to be bloody and negative of Rivalta test in PE than those with TPE. For clinical indicators, comparison of the specific parameters in blood showed that 18 indicators were higher in the TPE group than in the MPE group. By contrast, 12 indicators were higher in the MPE group than in the TPE group (P < .01). In addition, in PE tests, 3 parameters were higher in the TPE group, whereas other 4 parameters were higher in the MPE group (P < .01). Then, for clinical diagnosing practice, ROC analysis and principal component analysis were applied. The top 6 relevant indicators with area under curve over 0.70 were screened out as follows: hydrothorax adenosine dehydrogenase (pADA, 0.90), hydrothorax high-sensitivity C reactive protein (0.79), percentage of blood monocyte (sMONp, 0.75), blood high-sensitivity C reactive protein (sHsCRP, 0.73), erythrocyte sedimentation rate (0.71) and blood D-dimer (0.70). Moreover, logistic regression model revealed that a specific combination of 3 biomarkers, namely, pADA, sMONp and sHsCRP, could enhance the distinguishment of TB from malignant tumor with PE (area under curve = 0.944, 95% confidence interval = 0.925-0.964). The diagnostic function of the top single marker pADA in patients from different groups was analyzed and it was found to maintain high specificity and sensitivity. The 6 indicators, namely, pADA, hydrothorax high-sensitivity C reactive protein, sMONp, sHsCRP, sESR and blood D-dimer, showed significant diagnostic value for clinicians. Further, the combination of pADA, sMONp and sHsCRP has high accuracy for differential diagnosis for the first time. Most interestingly, the single marker pADA maintained high specificity and sensitivity in patients with different statuses and thus has great value for rapid and accurate diagnosis of suspected cases.

Early lung cancer diagnostic biomarker discovery by machine learning methods.

Early diagnosis has been proved to improve survival rate of lung cancer patients. The availability of blood-based screening could increase early lung cancer patient uptake. Our present study attempted to discover Chinese patients' plasma metabolites as diagnostic biomarkers for lung cancer. In this work, we use a pioneering interdisciplinary mechanism, which is firstly applied to lung cancer, to detect early lung cancer diagnostic biomarkers by combining metabolomics and machine learning methods. We collected total 110 lung cancer patients and 43 healthy individuals in our study. Levels of 61 plasma metabolites were from targeted metabolomic study using LC-MS/MS. A specific combination of six metabolic biomarkers note-worthily enabling the discrimination between stage I lung cancer patients and healthy individuals (AUC = 0.989, Sensitivity = 98.1%, Specificity = 100.0%). And the top 5 relative importance metabolic biomarkers developed by FCBF algorithm also could be potential screening biomarkers for early detection of lung cancer. Naïve Bayes is recommended as an exploitable tool for early lung tumor prediction. This research will provide strong support for the feasibility of blood-based screening, and bring a more accurate, quick and integrated application tool for early lung cancer diagnostic. The proposed interdisciplinary method could be adapted to other cancer beyond lung cancer.

Triptolide induces apoptosis through the calcium/calmodulin­dependent protein kinase kinaseß/AMP­activated protein kinase signaling pathway in non­small cell lung cancer cells.

Triptolide, a triterpene extracted from the Chinese herb Tripterygium wilfordii, has been reported to exert multiple bioactivities, including immunosuppressive, anti­inflammatory and anticancer effects. Although the anticancer effect of triptolide has attracted significant attention, the specific anticancer mechanism in non­small­cell lung cancer (NSCLC) remains unclear. The present study aimed to investigate the anticancer effect of triptolide in the H1395 NSCLC cell line and to determine its mechanism of action. The results revealed that triptolide significantly inhibited the cell viability of NSCLC cells in a dose­dependent manner, which was suggested to be through inducing apoptosis. In addition, triptolide was revealed to activate the calcium (Ca2+)/calmodulin­dependent protein kinase kinase ß (CaMKKß)/AMP­activated protein kinase (AMPK) signaling pathway by regulating the intracellular Ca2+ concentration levels, which increased the phosphorylation levels of AMPK and reduced the phosphorylation levels of AKT, ultimately leading to apoptosis. The CaMKKß blocker STO­609 and the AMPK blocker Compound C significantly inhibited the apoptosis­promoting effect of triptolide. In conclusion, the results of the present study suggested that triptolide may induce apoptosis through the CaMKKß­AMPK signaling pathway and may be a promising drug for the treatment of NSCLC.

Prevalence and molecular characterization of parechovirus A in children with acute gastroenteritis in Shenzhen, 2016-2018.

Parechovirus A (PeV-A), which causes a wide variety of diseases, is prevalent among young children. However, little is currently known about PeV-A infections in children with acute gastroenteritis in mainland China. In this study, we investigated the molecular epidemiology of acute gastroenteritis in Shenzhen, southern China, with an emphasis on PeV-A infections. A total of 1220 stool specimens from 1220 outpatient children under 5 years old with acute gastroenteritis were collected from January 2016 to December 2018. Viral RNA was detected by a real-time RT-PCR and PCR method. The PeV-A isolates were genotyped by sequencing the VP3/VP1 region. Of 1220 specimens, 148 (12.1%) were positive for PeV-A. The predominant genotype was PeV-A 1B (68.9%), followed by PeV-A 4 (12.2%), PeV-A 14 (6.1%), PeV-A 1A (5.4%), PeV-A 6 (2.7%), PeV-A 3 (2.7%) and PeV-A 5 (2.0%). It was found that 68.2% of PeV-A infections occurred in the summer and rainy months (June to September) in southern China. The majority of PeV-A-positive patients (97.3%) were younger than 24 months old. PeV-A coinfection with norovirus, rotavirus, astrovirus and adenovirus was found in thirty specimens (30/148, 20.3%), five specimens (5/148, 3.4%), five specimens (5/148, 3.4%), and two specimens (2/148, 1.4%), respectively. Coinfections with more than one other enteric virus were not observed in any of the PeV-A-positive specimens. Phylogenetic analysis revealed that the PeV-A isolates from Shenzhen were closely related to each other and to strains circulating in China, suggesting endemic circulation of PeV-A in China. The results of this study indicate that PeV-A is one of important pathogens of acute gastroenteritis in young children and that coinfection is a possible mode of PeV-A infection. PeV-A associated with acute gastroenteritis exhibited high genotypic diversity in Shenzhen, southern China.

miR­20b promotes growth of non­small cell lung cancer through a positive feedback loop of the Wnt/ß­catenin signaling pathway.

microRNAs (miRNAs or miRs) are endogenous noncoding single­stranded RNA molecules that can regulate gene expression by targeting the 3'­untranslated region and play an important role in many biological and pathological processes, such as inflammation and cancer. In this study, we found that miR­20b was significantly increased in human non­small cell lung cancer (NSCLC) cell lines and patient tissues, suggesting that it may possess a carcinogenic role in lung cancer. This miRNA promoted the proliferation, migration and invasion of NSCLC cells by targeting and downregulating the expression of adenomatous polyposis coli (APC), which is a negative regulator of the canonical Wnt signaling pathway. Wnt signaling activation may increase transcription of miR­20b. Therefore, miR­20b and canonical Wnt signaling were coupled through a feed­forward positive feedback loop, forming a biological regulatory circuit. Finally, an in vivo investigation further demonstrated that an increase in miR­20b promoted the growth of cancer cells. Overall, our findings offer evidence that miR­20b may contribute to the development of NSCLC by inhibiting APC via the canonical Wnt signaling pathway.

MicroRNA-421 confers paclitaxel resistance by binding to the KEAP1 3'UTR and predicts poor survival in non-small cell lung cancer.

MicroRNAs regulate post-transcriptional gene expression and play important roles in multiple cellular processes. In this study, we found that miR-421 suppresses kelch-like ECH-associated protein 1(KEAP1) expression by targeting its 3'-untranslated region (3'UTR). A Q-PCR assay demonstrated that miR-421 is overexpressed in non-small cell lung cancer (NSCLC), especially in A549 cells. Consistently, the level of miR-421 was higher in clinical blood samples from lung cancer patients than in those from normal healthy donors, suggesting that miR-421 is an important lung cancer biomarker. Interestingly, overexpression of miR-421 reduced the level of KEAP1 expression, which further promoted lung cancer cell migration and invasion, as well as inhibited cell apoptosis both in vivo and in vitro. Furthermore, knockdown of miR-421 expression with an antisense morpholino oligonucleotide (AMO) increased ROS levels and treatment sensitivity to paclitaxel in vitro and in vivo, indicating that high miR-421 expression may at least partly account for paclitaxel tolerance in lung cancer patients. To find the upstream regulator of miR-421, one of the candidates, ß-catenin, was knocked out via the CRISPR/Cas9 method in A549 cells. Our data showed that inhibiting ß-catenin reduced miR-421 levels in A549 cells. In addition, ß-catenin upregulation enhanced miR-421 expression, indicating that ß-catenin regulates the expression of miR-421 in lung cancer. Taken together, our findings reveal the critical role of miR-421 in paclitaxel drug resistance and its upstream and downstream regulatory mechanisms. Therefore, miR-421 may serve as a potential molecular therapeutic target in lung cancer, and AMOs may be a potential treatment strategy.

p53 sensitizes chemoresistant non-small cell lung cancer via elevation of reactive oxygen species and suppression of EGFR/PI3K/AKT signaling.

BACKGROUND: Non-small cell lung cancer (NSCLC) is the leading cause of cancer deaths primarily due to chemoresistance. Somatic mutation of TP53 (36%) and epidermal growth factor receptor (EGFR; > 30%) are major contributors to cisplatin (CDDP) resistance. Substantial evidence suggests the elevated levels of reactive oxygen species (ROS) is a key determinant in cancer. The elevated ROS can affect the cellular responses to chemotherapeutic treatments. Although the role of EGFR in PI3K/Akt signaling cascade in NSCLC is extensively studied, the molecular link between EGFR and p53 and the role of ROS in pathogenesis of NSCLC are limitedly addressed. In this study, we investigated the role of p53 in regulation of ROS production and EGFR signaling, and the chemosensitivity of NSCLC. METHODS: In multiple NSCLC cell lines with varied p53 and EGFR status, we compared and examined the protein contents involved in EGFR-Akt-P53 signaling loop (EGFR, P-EGFR, Akt, P-Akt, p53, P-p53) by Western blot. Apoptosis was determined based on nuclear morphological assessment using Hoechst 33258 staining. Cellular ROS levels were measured by dichlorofluorescin diacetate (DCFDA) staining followed by flow cytometry analysis. RESULTS: We have demonstrated for the first time that activation of p53 sensitizes chemoresistant NSCLC cells to CDDP by down-regulating EGFR signaling pathway and promoting intracellular ROS production. Likewise, blocking EGFR/PI3K/AKT signaling with PI3K inhibitor elicited a similar response. Our findings suggest that CDDP-induced apoptosis in chemosensitive NSCLC cells involves p53 activation, leading to suppressed EGFR signaling and ROS production. In contrast, in chemoresistant NSCLC, activated Akt promotes EGFR signaling by the positive feedback loop and suppresses CDDP-induced ROS production and apoptosis. CONCLUSION: Collectively, our study reveals that the interaction of the p53 and Akt feedback loops determine the fate of NSCLC cells and their CDDP sensitivity.

Clinical statistics analysis on the characteristics of pneumoconiosis of Chinese miner population.

BACKGROUND: Pneumoconiosis is one of the most common occupational diseases, which shows the progressive and irreversible pathological changes. It ultimately can induce pulmonary failure and lead to death. To date, these patients have no curative treatment option under the current standard of care, so it is especially important to delay the onset of the disease and slow down its progression. Therefore, understanding of clinical features of pneumoconiosis is particularly critical for medical intervention. METHODS: We collected the clinical data from 118 pneumoconiosis cases of miners admitted in hospital and processed the statistics analysis by using the Chi-square test and the risk assessment. RESULTS: Compared to other types of miners, gold miners are liable to cause Broncho-pulmonary co-infection with Chi-square value 18.748 and the P value <0.001. However, unexpectedly, the smoking miners displayed a better Activities of Daily Living (ADLs) compared to non-smokers, which showed 19.318 of Chi-square score and less than 0.001 of P value. And this connection was associated with the dust exposure time (P<0.05), showing the increasing risk of non-smoking miners occurred as the increasing time exposed to dust. In addition, our analysis indicated that the probability of smoking miners suffered from Broncho-pulmonary co-infection was less than non-smoking miners with Chi-square value 8.044 and P<0.01, which was also associated with the dust exposure time tendentiously, though P>0.05. Moreover, smoking history exhibited a deteriorating effect to the overall survival (OS) with 9.546 of Chi-square value and P<0.05, in accordance with smoking reducing life time. Interestingly, pneumoconiosis drugs could extend the smokers' OS, but not non-smokers'. CONCLUSIONS: Our studies suggest that the history of smoking and exposure time of dust play important roles in the development of pneumoconiosis and smoking could be a factor that determines the treatment options depending on patients' smoking history.

Relevance of EGFR gene mutation with pathological features and prognosis in patients with non-small-cell lung carcinoma.

OBJECTIVE: To study the relevance of EGFR gene mutation with pathological features and prognosis in patients with non-small-cell lung carcinoma. METHODS: A total of 297 patients from July 2009 to May 2013 were chosen as objects. EGFR gene mutation were detected with fluorescence quantitative PCR. Relevance of EGFR gene mutation with clinical and pathological features was analyzed, and the prognosis of EGFR- mutant-patients and that of EGFR- wide type-patients was compared. RESULTS: In 297 patients, 136 (45.79%) showed EGFR gene mutation. EGFR gene mutation had no significant relevance with age, gender, smoking history, family history of cancer and clinical stage (P>0.05); there was significant relevance between EGFR gene mutation and blood type, pathologic types, differentiation and diameter of cancer (P<0.05). The difference between prognosis of EGFR- mutant-patients and that of EGFR- wide type-patients was statistical significance (P<0.05). CONCLUSIONS: EGFR gene mutation has significant relevance with pathological features, the prognosis of EGFR-mutant-patients is better than that of EGFR- wide type-patients.

No association between XRCC1 gene Arg194Trp polymorphism and risk of lung cancer: evidence based on an updated cumulative meta-analysis.

X-ray repair cross-complementing group 1 (XRCC1) gene Arg194Trp polymorphism has been reported to be associated with risk of lung cancer in many published studies. Nevertheless, the research results were inconclusive and conflicting. To reach conclusive results, several meta-analysis studies were conducted by combining results from literature reports through pooling analysis. However, these previous meta-analysis studies were still not consistent. Hence, we used an updated and cumulative meta-analysis to get a more comprehensive and precise result from 25 case-control studies searching through the PubMed database up to September 1, 2013. The meta-analysis was carried out by the Comprehensive Meta-Analysis software and the odds ratio (OR) with 95 % confidence interval (CI) was used to estimate the pooled effect. The result involving 8,876 lung cancer patients and 11,210 controls revealed that XRCC1 Arg194Trp polymorphism was not associated with lung cancer risk [(OR=0.97, 95 %CI=0.92-1.03) for Trp vs. Arg; (OR=0.92, 95 % CI=0.85-0.98) for ArgTrp vs. ArgArg; (OR=1.07, 95 % CI=0.92-1.23) for TrpTrp vs. ArgArg; (OR=0.93, 95 % CI=0.87-1.00) for (TrpTrp + ArgTrp) vs. ArgArg; and (OR=1.08, 95 % CI=0.94-1.25) for TrpTrp vs. (ArgTrp + ArgArg)]. The cumulative meta-analysis showed that the results maintained the same, while the ORs with 95 % CI were more stable with the accumulation of case-control studies. The sensitivity and subgroups analyses showed that the results were robust and not affected by any single study with no publication bias. Relevant studies might not be needed for supporting these results.

Clinical study of simultaneous lung volume reduction surgery during resection of pulmonary or esophageal neoplasms.

BACKGROUND: If the emphysema lesions are not symmetrical, unilateral lung volume reduction surgery (LVRS) can be carried out on the more severe side. The aim of this research was to evaluate the feasibility and effects of LVRS performed simultaneously with resection of pulmonary and esophageal neoplasms. METHODS: Forty-five patients with pulmonary neoplasm and 37 patients with esophageal neoplasm were randomly assigned to group A or group B. In group A, LVRS was performed simultaneously on the same side as thoracotomy. In group B, only tumor resection was performed. The nonfunctional lung area was determined by preoperative chest computed tomography and lung ventilation/perfusion scan. The lung volume removed was about 20% to 30% of the lobes on one side. Preoperative and postoperative indexes including pulmonary function testing variables, arterial blood gas analysis variables, dyspnea scale, 6-minute walk distance, etc., were compared between the groups. RESULTS: There were no surgical deaths in this study. The postoperative forced vital capacity in 1 second, PaO2, PaCO2, dyspnea scale, and 6-minute walk distance were improved significantly in group A, whereas these indexes did not change or decreased slightly in group B. CONCLUSIONS: For tumor patients who have associated emphysema, simultaneous LVRS not only increases the chance of receiving surgical therapy, but also improves the postoperative quality of life of the patient. LVRS has expanded the surgical indication for tumor patients.

[Effects of recombinant adeno-virus vector carrying interleukin-5 antisense on the expression of interleukin-5 mRNA and protein in CD4+ T lymphocytes of asthmatic rats].

OBJECTIVE: To construct recombinant adeno-associated virus vector carrying antisense interleukin-5 (IL-5) gene (rAAV-asIL-5), and to explore the effects of this virus transfection on IL-5 mRNA and protein in CD(4)(+) T lymphocytes of asthmatic rats. METHODS: The eukaryotic antisense IL-5 expressing vector plasmid of recombinant adeno-associated virus (pasIL-5/rAAV) was constructed by gene recombination technique. The rAAV-asIL-5 particles were produced by co-transfection of pasIL-5/rAAV, pXX2, and pXX6 in package cell 293 through phosphate calcium deposit, and the titers of rAAV-asIL-5 were measured by Southern blot. The rAAV-asIL-5 particles were transfected into CD(4)(+) T lymphocytes obtained by gradient of Ficoll and immunomagnetic beads from the peripheral blood of asthmatic rats. Then IL-5 mRNA in T lymphocytes and IL-5 protein in supernatant of cell culture were determined with semi-quantitative RT-PCR and ELISA respectively. RESULTS: (1) The rAAV-asIL-5 was constructed and identified, and the titer of rAAV-asIL-5 was 1.3 x 10(11) virus particles/ml. (2) The relative ratio A of absorbance (IL-5/beta-actin) of rAAV group was 1.0515 +/- 0.1477, which was significantly lower than that of the control group (1.4271 +/- 0.1655) (n = 6, P < 0.01). (3) The protein level of IL-5 in supernatant of culture of rAAV group was (12.0840 +/- 1.4769) ng/L, significantly lower than that of the control group [(15.3590 +/- 1.2685) ng/L, n = 6, P < 0.01]. CONCLUSION: Construction of rAAV-asIL-5 was successful, and transfection of this virus was capable of inhibiting the expression of IL-5 mRNA and protein in CD(4)(+) T lymphocytes of asthmatic rats. The results of this study provide experimental data for further study of gene therapy for asthma.