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Early and effective identification of severe coronavirus disease 2019 (COVID-19) may allow us to improve the outcomes of associated severe acute respiratory illness with fever and respiratory symptoms. This study analyzed plasma concentrations of heat shock protein gp96 in nonsevere (including mild and typical) and severe (including severe and critical) patients with COVID-19 to evaluate its potential as a predictive and prognostic biomarker for disease severity. Plasma gp96 levels that were positively correlated with interleukin-6 (IL-6) levels were significantly elevated in COVID-19 patients admitted to the hospital but not in non-COVID-19 patients with less severe respiratory impairment. Meanwhile, significantly higher gp96 levels were observed in severe than nonsevere patients. Moreover, the continuous decline of plasma gp96 levels predicted disease remission and recovery, whereas its persistently high levels indicated poor prognosis in COVID-19 patients during hospitalization. Finally, monocytes were identified as the major IL-6 producers under exogenous gp96 stimulation. Our results demonstrate that plasma gp96 may be a useful predictive and prognostic biomarker for disease severity and outcome of COVID-19. IMPORTANCE Early and effective identification of severe COVID-19 may allow us to improve the outcomes of associated severe acute respiratory illness with fever and respiratory symptoms. Some heat shock proteins (Hsps) are released during oxidative stress, cytotoxic injury, and viral infection and behave as danger-associated molecular patterns (DAMPs). This study analyzed plasma concentrations of Hsp gp96 in nonsevere and severe patients with COVID-19. Significantly higher plasma gp96 levels were observed in severe than those in nonsevere patients, and its persistently high levels indicated poor prognosis in COVID-19 patients. The results demonstrate that plasma gp96 may be a useful predictive and prognostic biomarker for disease severity and outcome of COVID-19.

Background and aims Hepatocellular carcinoma (HCC) is one of the most common malignancy with poor prognosis, and the mortality rate remains high. More than 70% of HCC patients have recurrence within 5 years after treatment. The purpose of this study is to evaluate the prognostic values of serum markers with retrospective data. Methods We applied real‐world data (RWD) to analyze the prognostic values of six serum markers for HCC patients after treatment, including α‐fetoprotein (AFP), α‐fetoprotein‐L3 (AFP‐L3), Golgi protein73 (GP73), alanine aminotransferase (ALT), albumin (ALB), and total bilirubin (TBil). A total of 268 cases were enrolled to analyze recurrence‐free survival (RFS), and 104 cases were used to analyze overall survival (OS). Results Our results demonstrated that patients with higher AFP and AFP‐L3 had shorter RFS (p = 0.016 and 0.004), while higher GP73, ALT, and TBil experienced longer RFS (p = 0.000, 0.020, and 0.019). Patients with high‐level GP73, ALT, TBil, and low‐level ALB had significantly higher mortality rate (p=0.035, 0.008, 0.010, and 0.005). Multivariate analysis revealed that GP73 (HR = 1.548, p = 0.001) and ALT (HR = 1.316, p = 0.046) were identified as independent prognostic factors for RFS, ALB (HR = 0.127, p = 0.007), and ALT (HR = 0.237, p = 0.01) were identified as independent prognostic factors for OS. Subgroups analysis showed that GP73 had better prognostic values than other serum markers in early‐stage HCC (p = 0.023). Conclusions Our study demonstrates that AFP, AFP‐L3, and GP73 can be used as prognostic indicators for predicting the recurrence of HCC, while liver function tests have better survival prediction values. GP73 can act as a promising prognostic marker for early‐stage HCC., Prior to nasotracheal intubation (NTI), topical nasal vasoconstrictors are used to prevent NTI‐related epistaxis (NTIRE). Our study shows that well‐lubricated nasotracheal intubation does not require pretreatment with ephedrine to reduce NTIRE.

Increasing evidence has shown that the metabolism and clearance of molecular targeted agents, such as sorafenib, plays an important role in mediating the resistance of HCC cells to these agents. Metabolism of sorafenib is performed by oxidative metabolism, which is initially mediated by CYP3A4. Thus, targeting CYP3A4 is a promising approach to enhance the sensitivity of HCC cells to chemotherapeutic agents. In the present work, we examined the association between CYP3A4 and the prognosis of HCC patients receiving sorafenib. Using the online tool miRDB, we predicted that has-microRNA-4277 (miR-4277), an online miRNA targets the 3’UTR of the transcript of cyp3a4. Furthermore, overexpression of miR-4277 in HCC cells repressed the expression of CYP3A4 and reduced the elimination of sorafenib in HCC cells. Moreover, miR-4277 enhanced the sensitivity of HCC cells to sorafenib in vitro and in vivo. Therefore, our results not only expand our understanding of CYP3A4 regulation in HCC, but also provide evidence for the use of miR-4277 as a potential therapeutic in advanced HCC.

Background Whether achieving sustained virological response (SVR) in patients with hepatitis C attains complete elimination of hepatitis C virus (HCV) is unknown, because occult HCV infection (OCI), defined as the detection of HCV-RNA in hepatocytes or peripheral blood mononuclear cells (PBMC) in absence of serum HCV-RNA, may occur. We thus investigated the prevalence and clinical relevance of OCI. Methods Subjects from three hospitals who had achieved serum HCV clearance, including 60 of Direct-acting antiviral agents (DAAs) induced SVR, 50 of pegylated interferon plus ribavirin (PR) induced SVR, and 30 of spontaneous resolution, were subjected to detect HCV-RNA in liver by robust RNAscope assay and PBMC by qPCR. Paired liver biopsies at baseline and at SVR24 were analyzed. Results OCI was detected in 16 of 140 subjects (11.4%), with 15.0% in DAA-based group, 10.0% in PR group and 6.7% in spontaneously resolved group. In DAA-based subgroups, the incidence of OCI was gradually increased in group of solely DAA(s) therapy, combining DAA and PR therapy and combining DAA and ribavirin therapy. OCI is more frequent in patients with genotype 3. No correlation between baseline viral load, interleukin-28B genotype, baseline transaminases, post-SVR transaminases and OCI were found. However, OCI was significantly linked with severity of fibrosis and active inflammation at post-SVR, even considering basal fibrosis status. In addition, both the magnitude and the frequency of fibrosis regression were lower in patients with OCI than in those without OCI. In the multivariate analysis, PR therapy was identified an independent negative prognostic factor for both hepatic inflammation (P = .022) and fibrosis regression (P = .015). Importantly, we found HCV relapse in one of the OCI patients at 48 weeks after the end of PR treatment. Conclusions HCV-RNA can persist in hepatocytes and/or PBMC in a certain of patients who achieved spontaneous or treatment-induced HCV RNA clearance from serum and associated with persistent histological abnormality. Our findings provide new insights into cure of HCV and could influence the following-up scenario after SVR.

Background Early and accurate diagnosis is the key to improving survival in cirrhotic patients with ascitic fluid infection. Aims To investigate the usefulness of adjusted Intensive Care Infection Score (ICISΔ) for diagnosis of ascites infection in cirrhotic patients. Methods Cirrhotic patients with ascites (n = 125) were enrolled, and the efficacy of ICIS and ICISΔ for predicting ascites infection was evaluated. ICISΔ was created by using the weighted variation of each ICIS parameter. Results The area under the curves (AUCs) of ICIS for the diagnosis of ascites infection were 0.90 (95% CI: 0.84–0.95), 0.85 (95% CI: 0.79–0.90), and 0.87 (95% CI: 0.81–0.93), for SBP, culture-negative SBP, and combined SBP/culture-negative SBP, respectively. ICIS was optimized and diagnostic accuracy was obviously improved. ICISΔ had high AUCs of 0.99 (95% CI: 0.93–1.00) for SBP, 0.98 (95% CI: 0.83–1.00) for culture-negative SBP, and 0.98 (95% CI: 0.94–1.00) for the combination group. The optimal cutoff was identified as ICISΔ > 2, which had >97.8% sensitivity and 100% specificity for diagnosis of both SBP and culture-negative SBP. The ICISΔ had significantly higher AUCs than PCT and CPR in both groups (P = 0.002–0.008). ICISΔ kinetics could differentiate between SBP and culture-negative SBP patients. From sterile ascites, through culture-negative SBP to SBP, three ICISΔ parameters showed an increasing trend. Conclusions ICIS and ICISΔ are simple, rapid, accurate and cost-effective methods for the diagnosis of ascites infection in cirrhotic patients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection causes a broad clinical spectrum of coronavirus disease 2019 (COVID-19). The development of COVID-19 may be the result of a complex interaction between the microbial, environmental, and host genetic components. To reveal genetic determinants of susceptibility to COVID-19 severity in the Chinese population, we performed a genome-wide association study on 885 severe or critical COVID-19 patients (cases) and 546 mild or moderate patients (controls) from two hospitals, Huoshenshan and Union hospitals at Wuhan city in China. We identified two loci on chromosome 11q23.3 and 11q14.2, which are significantly associated with the COVID-19 severity in the meta-analyses of the two cohorts (index rs1712779: odds ratio [OR] = 0.49; 95% confidence interval [CI], 0.38–0.63 for T allele; P = 1.38 × 10−8; and index rs10831496: OR = 1.66; 95% CI, 1.38–1.98 for A allele; P = 4.04 × 10−8, respectively). The results for rs1712779 were validated in other two small COVID-19 cohorts in the Asian populations (P = 0.029 and 0.031, respectively). Furthermore, we identified significant eQTL associations for REXO2, C11orf71, NNMT, and CADM1 at 11q23.3, and CTSC at 11q14.2, respectively. In conclusion, our findings highlight two loci at 11q23.3 and 11q14.2 conferring susceptibility to the severity of COVID-19, which might provide novel insights into the pathogenesis and clinical treatment of this disease.

BACKGROUND. Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory coronavirus 2 (SARS-CoV-2), has become a pandemic. This study addresses the clinical and immunopathological characteristics of severe COVID-19. METHODS. Sixty-nine patients with COVID-19 were classified into severe and nonsevere groups to analyze their clinical and laboratory characteristics. A panel of blood cytokines was quantified over time. Biopsy specimens from 2 deceased cases were obtained for immunopathological, ultrastructural, and in situ hybridization examinations. RESULTS. Circulating cytokines, including IL-8, IL-6, TNF-α, IP10, MCP1, and RANTES, were significantly elevated in patients with severe COVID-19. Dynamic IL-6 and IL-8 were associated with disease progression. SARS-CoV-2 was demonstrated to infect type II and type I pneumocytes and endothelial cells, leading to severe lung damage through cell pyroptosis and apoptosis. In severe cases, lymphopenia, neutrophilia, depletion of CD4(+) and CD8(+) T lymphocytes, and massive macrophage and neutrophil infiltrates were observed in both blood and lung tissues. CONCLUSIONS. A panel of circulating cytokines could be used to predict disease deterioration and inform clinical interventions. Severe pulmonary damage was predominantly attributed to both cytopathy caused by SARS-CoV-2 and immunopathologic damage. Strategies that prohibit pulmonary recruitment and overactivation of inflammatory cells by suppressing cytokine storm might improve the outcomes of patients with severe COVID-19.

Background: An outbreak of severe respiratory illness caused by a novel coronaviruses (2019- nCoV) in China since December 2019 This work aim to evaluate the c

In this paper, we report the development of a label-free biosensor, based on surface plasmon resonance, for real-time monitoring of captured human CD4+T-cells, and their dynamic cytokine production upon specific antigen stimulation. Microarrays of CD4+T-cells, and interferon gamma (IFN-γ) specific antibody (Ab) spots were printed side by side onto the poly(OEGMA-co-HEMA) matrix. The placement of CD4+ T-cells near the anti IFN-γ Ab-coated spots ensured a high local concentration of secreted IFN-γ for detection. We have demonstrated that this approach enables the detection of tuberculosis (TB) infection in clinical samples, with an overall sensitivity of 85.5% and an overall specificity of 97.7%.

More than 70% of hepatocellular carcinoma (HCC) cases develop as a consequence of liver cirrhosis (LC). Here we have evaluated the diagnostic potential of four serum biomarkers, and developed models for HCC diagnosis and differentiation from LC patients. Serum levels of α-fetoprotein (AFP), AFP-L3, des-γ-carboxy prothrombin (DCP), and Golgi protein 73 (GP73) were analyzed in 114 advanced HCC patients, 81 early stage HCC patients, and 152 LC patients. Multilayer perceptron (MLP) and radial basis function (RBF) neural networks were used to construct the diagnostic models. Using all stages, HCC diagnostic models had a higher sensitivity (>70%) than the individual serum biomarkers, whereas only early stage HCC diagnostic models had a higher specificity (>80%). The early stage HCC diagnostic models could not be used as HCC screening tools due to their low sensitivity (about 40%). These results suggest that a combination of the two models might be used as a screening tool to distinguish early stage HCC patients from LC patients, thus improving prevention and treatment of HCC.

BACKGROUND Differentiation of malignant from benign liver tumors remains a challenging problem. In recent years, mass spectrometry (MS) technique has emerged as a promising strategy to diagnose a wide range of malignant tumors. The purpose of this study was to establish classification models to distinguish benign and malignant liver tumors and identify the liver cancer-specific peptides by mass spectrometry. MATERIAL AND METHODS In our study, serum samples from 43 patients with malignant liver tumors and 52 patients with benign liver tumors were treated with weak cation-exchange chromatography Magnetic Beads (MB-WCX) kits and analyzed by the Matrix-Assisted Laser Desorption Time of Flight Mass Spectrometry (MALDI-TOF-MS). Then we established genetic algorithm (GA), supervised neural networks (SNN), and quick classifier (QC) models to distinguish malignant from benign liver tumors. To confirm the clinical applicability of the established models, the blinded validation test was performed in 50 clinical serum samples. Discriminatory peaks associated with malignant liver tumors were subsequently identified by a qTOF Synapt G2-S system. RESULTS A total of 27 discriminant peaks (p

Background: Whether achieving sustained virological response (SVR) in patients with hepatitis C attains complete elimination of hepatitis C virus (HCV) is unknown, because occult HCV infection (OCI), defined as the detection of HCV-RNA in hepatocytes or peripheral blood mononuclear cells (PBMC) in absence of serum HCV-RNA, may occur. We thus investigated the prevalence and clinical relevance of OCI. Methods: Subjects from three hospitals who had achieved serum HCV clearance, including 60 of Direct-acting antiviral agents (DAAs) induced SVR, 50 of pegylated interferon plus ribavirin (PR) induced SVR, and 30 of spontaneous recovery, were subjected to detect HCV-RNA in hepatocytes and PBMC. Paired liver biopsies at baseline and post-SVR were analyzed. Results: OCI was detected in 16 of 140 subjects (11.4%), with 15.0% in DAA-based group, 10.0% in PR group and 6.7% in spontaneously resolved group. OCI is more frequent in patients with genotype 3. No correlation between baseline viral load, interleukin-28B genotype, baseline transaminases, post-SVR transaminases and OCI were found. However, OCI was significantly linked with severity of fibrosis at post-SVR, even considering basal fibrosis status. In addition, both the magnitude and the frequency of fibrosis regression were lower in patients with OCI than in those without OCI. In the multivariate analysis, PR therapy was identified an independent negative prognostic factor for both hepatic inflammation (P = 0.022) and fibrosis regression (P = 0.015). Conclusions: HCV-RNA can persist in hepatocytes and/or PBMC in a certain of patients albeit achieving serum resolution of hepatitis C and associated with persistent histological abnormality. Funding Statement: This work was supported by grants from National Natural Science Foundation of China (NNSFC) (No. 81673654) (to J.Zhao); NNSFC (No. 31770186) (to Y. Wang); NNSFC (No. 81802020) (to Y. Wang); and the National S&T Major Project for Infectious Diseases (No. 2017ZX10302201001007) (to J.Zhao). Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: This study was done in accordance with the ethical principles of the Declaration of Helsinki as revised in 2013 and was approved by the Ethics Committee by each institution’s human research committee, Peking University People’s Hospital, the first Hospital of Jilin University and Chinese PLA General Hospital. Informed consent was obtained from all participants prior to enrolment. Written informed consent was received from participants prior to inclusion in the study.

Inflammatory cytokine gene polymorphisms may influence the hepatic and extrahepatic HBV-related disease. In this study, we aimed to investigate the relationship between polymorphisms of IL-17, IL-21 gene and HBV related hepatocellular carcinoma in Chinese Han population.We performed a multi-center study comprised 866 HBV-related hepatocellular carcinoma (HCC) patients and 1086 unrelated patients with a diagnosis of chronic hepatitis B (CHB) as control to evaluate the effects of IL-17 (rs4711998), IL-21 SNPs (rs12508721, rs13143866 and rs2221903) and the susceptibility of HCC. MassARRAY technology was utilized to genotype. Enzyme-linked immunosorbent assay (ELISA) was used to detect the serum IL-17 and IL-21 level. Quantitative real time polymerase chain reaction (qRT-PCR) was used to analyze the serum viral loads.In logistic regression analysis, our results showed the frequency of rs4711998 allele G in CHB group was significantly higher than that in HCC group (P = 0.042, 0.859(0.743-0.994)), and it is present only among females. Compared to HCC group, rs13143866 A allele was more likely to appear in HCC group (P = 0.015, 1.268 (1.049-1.532)). The frequency of AA also showed different between HCC group and CHB groups (P = 0.011, 3.135 (1.292-7.603)), which showed strong sex-specific relationships. ELISA showed a higher serum IL-17 and IL-21 expression in HCC patients compared to CHB patients (P all0.05). Haplotype rs12508721C/rs13143866A/rs2221903T in male HCC group was statistically higher than in male CHB group(P = 0.013) but not in females (P 0.05).We suggested rs4711998 allele A as risk factors for women to develop HBV related-HCC in Chinese Han population. rs13143866 allele A as risk factors to develop HBV related-HCC in Chinese male population. Male patients with haplotype rs12508721C/rs13143866A/rs2221903T may with 1.3-fold risk for HBV-related HCC.

Background It is difficult to diagnose ascites infection early in cirrhotic patients. The present study was to create and evaluate a new bioscore combined with PCT, sNFI and dCHC in the diagnosis of ascites infection in cirrhotic patients. Methods Two hundred and fifty-nine consecutive patients were enrolled; of which 51 patients were culture-positive spontaneous bacterial peritonitis (culture-positive SBP) and 58 patients were culture-negative SBP. The efficacy of procalcitonin(PCT), c-reactive protein (CRP), white blood cell (WBC), mean fluorescence intensity of mature neutrophils(sNFI) and difference in hemoglobin concentration between newly formed and mature red blood cells(dCHC) for diagnosing ascites infection was examined. These parameters were used to create a scoring system. The scoring system was analyzed by logistic regression analysis to determine which parameters were statistically different between ascites infection and non-ascites infection patients. Receiver operating characteristic curve (ROC) was used to analyze the diagnostic ability of bioscore for ascites infection. Results In ROC analysis, the area under the curves (AUC) for PCT was 0.852 (95% CI 0.803–0.921, P

Forty-five KPC-producing Enterobacteriaceae strains were isolated from multiple departments in a Chinese public hospital from 2014 to 2015. Genome sequencing of four representative strains, namely Proteus mirabilis GN2, Serratia marcescens GN26, Morganella morganii GN28, and Klebsiella aerogenes E20, indicated the presence of blaKPC-2-carrying IncX6 plasmids pGN2-KPC, pGN26-KPC, pGN28-KPC, and pE20-KPC in the four strains, respectively. These plasmids were genetically closely related to one another and to the only previously sequenced IncX6 plasmid, pKPC3_SZ. Each of the plasmids carried a single accessory module containing the blaKPC-2/3-carrying ΔTn6296 derivatives. The ΔTn6292 element from pGN26-KPC also contained qnrS, which was absent from all other plasmids. Overall, pKPC3_SZ-like blaKPC-carrying IncX6 plasmids were detected by PCR in 44.4% of the KPC-producing isolates, which included K. aerogenes, P. mirabilis, S. marcescens, M. morganii, Escherichia coli, and Klebsiella pneumoniae, and were obtained from six different departments of the hospital. Data presented herein provided insights into the genomic diversity and evolution of IncX6 plasmids, as well as the dissemination and epidemiology of blaKPC-carrying IncX6 plasmids among Enterobacteriaceae in a hospital setting.

Background: Direct-acting antiviral agents (DAAs) have opened a new era for treating chronic hepatitis C virus (HCV) infection. However, whether DAA induced SVR attains real cure and complete eradication of HCV RNA is unknown, because occult HCV infection (OCI), as defined the detection of HCV RNA in hepatocytes or peripheral blood mononuclear cells (PBMC) in absence of serum of HCV RNA, may occur despite SVR and the clinical significance is yet to be established. We thus investigated the prevalence and provisional outcome of OCI in patients who achieved spontaneous or therapy-induced resolution of hepatitis C. Methods: Chronic HCV subjects from three tertiary hospitals who had achieved serum viral clearance, including 60 of DAAs treatment, 50 of pegylated interferon plus ribavirin (PR) treatment, and 30 of spontaneous recovery, were subjected to detect HCV RNA in hepatocytes by in suit hybridization as well as in PBMC by PCR. Liver function testes, HCV genotypes, and paired liver biopsies of pre- and post-treatment for each patient were acquired for comparison analysis. Findings: 16 of 140 subjects (11·4%) were found to have OCI, with the highest (15·0%) rate in DAA-based treated subjects, 10·0% in PR treated subjects and 6·7% in spontaneously resolved cases. Rates of OCI were 16·7% and 11·1% among patients with PR-free regimen and PR-based regimen, respectively. The existence of intermediate negative strand of HCV genome in all PBMCs of OCI patients suggested ongoing viral replication. The occurrence of OCI is more frequent in patients with genotype 3 HCV but less in patients with genotype 1. No correlation between baseline HCV viral load, interleukin-28B genotype, baseline transaminases, post transaminases and occurrence of OCI was found. While notably, OCI patients had more advanced liver fibrosis and active inflammation at both baseline and post- treatment. In addition, fibrosis scores were significantly decreased after SVR only in patients without OCI (P

Nonspecific interactions (NSIs) and irreproducibility greatly reduce the accuracy of antigen-antibody screening, which is key to the discovery of monoclonal antibody drugs and biomarkers identification. We previously developed a solid supporting material, polymer-coated initiator integrated poly(dimethysiloxane) (iPDMS), which is able to provide near-zero background for microarray screening. Here, we applied two monoclonal antibodies (mAbs), namely, anti-FLAG and HM1, to screen an iPDMS-based peptide microarray with 2083 peptides from 62 proteins to evaluate NSIs and irreproducibility. In addition to recognizing their cognate epitopes, the two mAbs also cross-reacted with random sequences, especially when they were used at high concentrations. At 50 μg mL(-1), 295 peptides (14.2% of the peptide library) had positive reactions to anti-FLAG and only 39 peptides (1.9%) reacted positively to HM1. Virtually all cross-reactions disappeared when the [mAbs] reached 0.01 μg mL(-1). Reproducible experiments of 404 peptides at various [mAbs] showed that only specific interactions, molecular mimicry, and mimotope were reproducible between different experiments. These findings suggest that irreproducibility was at least partially caused by NSIs. We also demonstrated that repeating tests and mAb dilution could effectively avoid NSI-related irreproducibility in serological screening. This will not only largely simplify the data analysis, but will also make immunoassays more reliable for clinical research.

Dengue fever is caused by the dengue virus (DENV), and DENV1 is the prevalent epidemic serotype in south China. A new lateral flow assay (LFA) based on a near-infrared (NIR) fluorescent dye was developed to detect anti-DENV1 IgG antibodies. DyLight-800 was used as the marker conjugated to goat anti-human IgG antibodies, and recombinant dengue type 1 envelope protein was used as the capture protein on the test line. Twenty samples from patients infected with DENV1 and 160 negative controls were analyzed using this new NIR-LFA. The results of the NIR-LFA were compared with the results of Panbio Dengue IgG ELISA and the Dengue Duo IgM/IgG Cassette. Nineteen confirmed DENV1-positive samples were identified by NIR-LFA, giving 95% (19/20) sensitivity. No significant differences existed in the results when the 20 primary clinical samples were analyzed using NIR-LFA, Panbio ELISA, and the Dengue Duo Cassette. However, NIR-LFA had a lower limit of detection than IgG ELISA and Duo IgM/IgG Cassette did when analyzing a 2-fold dilution series of the 19 samples positively identified by NIR-LFA. When incorporated with an NIR POCT device, the new NIR-LFA was rapid, easy to use, and highly sensitive in detecting DENV1, and has potential for application to clinical diagnosis.

Hepatocellular carcinoma (HCC) is a highly lethal malignant tumor evolved from cirrhosis. It is quite significant to seek accurate, easy markers for early warning and diagnosis of HCC. Through prospective cohort follow-up study and mass spectrometry, we discovered and verified a serum marker valuable for early warning and diagnosis. Follow-up observation was performed on cirrhosis patients. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was adopted to detect the serums of patients, and the serum polypeptides with a potential value in early HCC warning and diagnosis were screened. Electrospray ionization quadrupole time-of-flight tandem mass spectrometry (ESI-Q-TOF-MS/MS) was exploited to identify these screened polypeptides. Moreover, the serum marker concentration was determined by ELISA to validate the clinical value of the serum marker. Among 109 cirrhosis patients followed up for two years, 29 patients (26.6%) finally progressed into HCC. MALDI-TOF MS shows that the concentration of a 3155.66Da polypeptide was significantly different between the patients that progressed into HCC and those not. Through MS/MS identification, it is confirmed that the polypeptide is inter-alpha-trypsin inhibitor heavy chain 4 (ITIH4). The serum ITIH4 concentrations in two groups were measured with ELISA and compared with Alpha-fetoprotein (AFP). Results show that serum ITIH4 and AFP concentrations were negatively correlated (r=−0.263, p=0.0006), and the ITIH4 concentration had a significant intergroup difference (p=0.000). Receiver operating characteristic (ROC) curve indicates that its predictive value (area under the curve, AUC) is 0.667, superior to AFP. For the patients progressing into HCC, serum samples were separately collected when they were recruited and diagnosed as cirrhosis. Measurement on these samples reveals that ITIH4 was declining during the progression of HCC (p=0.006). By virtue of mass spectrometry, we discovered and identified a biomarker valuable for early HCC warning and diagnosis. This marker overperforms the commonly used AFP, demonstrating a bright prospect.

Tuberculosis (TB) remains a major worldwide health problem and has caused millions of deaths in the past few years. Current diagnostic methods, such as sputum smear microscopy and sputum culture, are time-consuming and cannot prevent the rapid spreading of TB during the diagnostic period. In this connection, detecting biomarkers specific to TB at molecular level in plasma of patients will provide a rapid means for diagnosis. In this study, we first evaluated the differential expression of the long non-coding RNAs (lncRNAs) in the plasma from patients with TB (TB positive), community acquired pneumonia (CAP) and healthy individuals (CG) using lncRNA microarray scanning. It was found that there were 2116 specific lncRNAs differentially expressed in the TB positive samples (1102 up-regulated and 1014 down-regulated), which accounted for 6.96% of total lncRNAs. Twelve differentially expressed lncRNAs discovered in microarray were subsequently validated by using real-time quantitative PCR (RT-qPCR). Two lncRNAs (ENST00000354432 and ENST00000427151) were further validated with more Tuberculosis samples. These results suggested the expression level of lncRNAs and the two validated lncRNAs in plasma could be the potential molecular biomarkers for the rapid diagnosis of Tuberculosis.

Background The increase in the amount of extended spectrum beta-lactamases (ESBL)-producing gram-negative bacteria is seriously threatening human health in recent years. Therefore, it is necessary to develop a rapid and reliable method for identification of ESBLs. The purpose of this study was to establish a novel method to discriminate between ESBL-producing and non- ESBL-producing bacteria by using the matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) technique. Material/methods We detected hydrolyzed production of cefotaxime after incubation with 69 gram-negative bacteria by using MALDI-TOF-MS. Then we established genetic algorithm (GA), supervised neural networks (SNN), and quick classifier (QC) models using several peaks to identify ESBL-producing strains. To confirm the clinical applicability of the models established, a blinded validation test was performed in 34 clinical isolated strains. Results Using ClinPro Tools software, we identified 4 peaks (456 Da, 396 Da, 370 Da, and 371 Da) in mass spectra of cefotaxime solution that have high enough specificity to discriminate ESBL-producing from non- ESBL-producing strains. Recognition capability of models established were 97.5% (GA), 92.5% (SNN), and 92.5% (QC), and cross validation rates were 90.15% (GA), 97.62 (SNN), and 97.62% (QC). The accuracy rates of the blinded validation test were 82.4% (GA), 88.2% (SNN), and 82.4% (QC). Conclusions Our results demonstrate that identification of ESBLs strains by MALDI-TOF-MS has potential clinical value and could be widely used in the future as a routine test in clinical microbiology laboratories.

The early prediction values of diagnostic markers for hepatocellular carcinoma (HCC) are still unclear at present. This study evaluated the prediction value of ten serum markers in HCC. A total of 109 cases of hepatic cirrhosis patients were followed up for 36 months and the relationship between the lifetime risk of developing HCC and levels of serum markers was analyzed. 31.2 (34/109) percent of hepatic cirrhosis patients developed HCC during the study’s timeframe. Higher alpha-fetoprotein (AFP), alpha-fetoprotein-L3 (AFP-L3), alanine aminotransferase (ALT), and AFP-L3/AFP ratio levels are potential risk factors for malignization in hepatic cirrhosis patients (RR=2.99, 2.92, 2.72, and 2.34); serum Golgi protein 73 (GP73) level of hepatic cirrhosis patients decreased significantly after developing HCC (t=2.212;p=0.041). The detection of ALT, AFP, AFP-L3, and GP73 has a certain guiding significance to predict the risk of HCC in hepatic cirrhosis patients.

Golgi protein-73 (GP73) is upregulated in cancers and viral infections; however, its role in human immunodeficiency virus (HIV) and acquired immune deficiency syndrome (AIDS) remains undetermined. GP73 was evaluated as a biomarker of HIV progression and AIDS treatment efficacy. Forty-eight HIV patients (≤350 CD4 + T cells/μL) undergoing highly active antiretroviral therapy (HAART group) and 18 HIV patients expected to undergo HAART within 9 months (>350 CD4 + T cells/μL) (control group) were enrolled in a prospective, single center, cohort study from May 2009 to Jun 2012. Blood aspartate aminotransferase, alanine aminotransferase (ALT), cholesterol, triglycerides, and total bilirubin were assessed at baseline, 2 weeks, and 1, 3, 6, 9, and 12 months (HAART group) or 3 month intervals (control group). Serum HIV RNA level (viral load) was determined by reverse-transcriptase polymerase chain reaction (RT-PCR), and serum and peripheral blood mononuclear cell (PBMC) GP73 concentration were determined by chemiluminescent immunoassay kit and western blot, respectively. Significant positive and negative correlations in baseline serum GP73 concentration and HIV viral load (r = 0.39, P

Nucleolin, as a multifunctional protein, has been demonstrated to play an oncogenic role in human hepatocellular carcinoma (HCC). The aim of this study was to investigate the expression pattern of nucleolin in HCC and determine its correlation with tumor progression and prognosis. Nucleolin expression at both mRNA and protein levels in HCC and adjacent nonneoplastic tissues were respectively detected by quantitative real time polymerase chain reaction (Q-PCR), immunohistochemistry and western blotting. Nucleolin expression, at both mRNA and protein levels, was significantly higher in HCC tissues than in the adjacent nonneoplastic tissues (both P < 0.001). In addition, the elevated nucleolin expression was markedly correlated with advanced tumor stage (P = 0.001), high tumor grade (P = 0.02) and serum AFP level (P = 0.008). Moreover, HCC patients with high nucleolin expression had shorter 5-year disease-free survival and shorter 5-year overall survival than those with low expression (both P < 0.001). Furthermore, the Cox proportional hazards model showed that nucleolin expression was an independent poor prognostic factor for both 5-year disease-free survival (hazards ratio [HR] = 3.696, 95% confidence interval [CI] = 1.662-8.138, P = 0.01) and 5-year overall survival (HR = 3.872, CI = 1.681-8.392, P = 0.01) in HCC. These results showed that the markedly and consistently increasing expression of nucleolin may be associated with aggressive characteristics of HCC, and implied that nucleolin expression may serve as a promising biochemical marker for predicting the clinical outcome of patients with this malignancy. The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/13000_2014_175 .

BACKGROUND: Influenza has emerged every year but a complete profile of laboratory indices throughout the disease course remains unknown. METHODS: Clinical data was collected from 28 confirmed cases of the pandemic influenza H1N1 2009. The levels of serum iron (Fe), carbon dioxide combining power (CO2-CP), total complement hemolytic activity (CH50), C-reactive protein (CRP), and white blood cell (WBC) and differential count were analyzed. RESULTS: Major laboratory abnormalities recokled for patients upon admission were lymphopenia (96.4%), eosinopenia (50.0%), hypoferremia (92.9%), decreased levels of serum CO2-CP (60.7%), increased levels of serum CRP (84.6%) and serum CH50 (71.4%). The serum iron and CO2-CP concentration and the counts for lymphocytes, eosinophils, and basophils were significantly increased four days after sickness was noticed compared with the first three days of illness (p < 0.05). The total WBC and neutrophil counts were significantly decreased four days after onset of illness compared with the counts over the first three days (p < 0.05). The monocyte count and CRP concentration was significantly decreased 7 days after onset of illness compared with first 3 days after illness onset (p < 0.05). The serum CH50 concentrations were higher than the normal range during disease course and significantly elevated 7 days after onset of illness compared with the first 6 days after illness onset (p < 0.05). CONCLUSIONS: The serum levels of iron, CO2-CP, CH50, CRP, and WBC and differential count Were significantly varied during the whole pandemic influenza (H1N1) 2009. The development of WBC count in patients with influenza may be an effective predictor for severity of illness.

This study was designed to investigate the role of serum GP73 for diagnosing significant fibrosis in patients with chronic hepatitis B virus (HBV) infections. Two populations were enrollment. All subjects were patients with chronic HBV infections. First population included 761 patients, who received liver stiffness measurement; the second population included 633 patients, who undertaken liver biopsy, in which 472 patients with nearly normal ALT. All patients received serum GP73 test. The effect of GP73 recombinant protein to HepG2 cells and LX2 cells were observed in vitro. Results showed that serum GP73 concentration is correlated with liver stiffness (r = 0.601). The area under ROC curve is 0.76. The sensitivity and specificity of GP73 for significant fibrosis (≥F2) diagnosis were 62.81%, 80.05% respectively (cut off: 76.6 ng/ml). Serum GP73 concentration was significantly correlated with the grading of fibrosis (r = 0.32, and 0.35, in 633 and 472 patients, respectively.) GP73 had a striking performance for diagnosing S2 in patients with chronic HBV infections. In 472 patients with nearly normal ALT, the sensitivity and specificity of GP73 for S2 diagnosis were 62.5% and 80.0% respectively, where the cut-off was set at 82 ng/ml. GP73 recombinant protein may prompt LX2 cells proliferation at the concentration 10-100 ng/ml. The present results indicated that GP73 may be a marker for diagnosing significant fibrosis in patients with chronic HBV infections, and may be a new contributor to fibrogensis.

Background Our previous study identified AKT1, AKT2 and AKT3 as unfavorable prognostic factors for patients with hepatocellular carcinoma (HCC). However, limited data are available on their exact mechanisms in HCC. Since microRNAs (miRNAs) are implicated in various human cancers including HCC, we aimed to screen miRNAs targeting AKTs and investigate their underlying mechanisms in HCC by integrating bioinformatics prediction, network analysis, functional assay and clinical validation. Methods Five online programs of miRNA target prediction and RNAhybrid which calculate the minimum free energy (MFE) of the duplex miRNA:mRNA were used to screen optimized miRNA-AKT interactions. Then, miRNA-regulated protein interaction network was constructed and 5 topological features (‘Degree’, ‘Node-betweenness’, ‘Edge-betweenness’, ‘Closeness’ and ‘Modularity’) were analyzed to link candidate miRNA-AKT interactions to oncogenesis and cancer hallmarks. Further systematic experiments were performed to validate the prediction results. Results Six optimized miRNA-AKT interactions (miR-149-AKT1, miR-302d-AKT1, miR-184-AKT2, miR-708-AKT2, miR-122-AKT3 and miR-124-AKT3) were obtained by combining the miRNA target prediction and MFE calculation. Then, 103 validated targets for the 6 candidate miRNAs were collected from miRTarBase. According to the enrichment analysis on GO items and KEGG pathways, these validated targets were significantly enriched in many known oncogenic pathways for HCC. In addition, miRNA-regulated protein interaction network were divided into 5 functional modules. Importantly, AKT1 and its interaction with mTOR respectively had the highest node-betweenness and edge-betweenness, implying their bottleneck roles in the network. Further experiments confirmed that miRNA-149 directly targeted AKT1 in HCC by a miRNA luciferase reporter approach. Then, re-expression of miR-149 significantly inhibited HCC cell proliferation and tumorigenicity by regulating AKT1/mTOR pathway. Notably, miR-149 down-regulation in clinical HCC tissues was correlated with tumor aggressiveness and poor prognosis of patients. Conclusion This comprehensive analysis identified a list of miRNAs targeting AKTs and revealed their critical roles in HCC malignant progression. Especially, miR-149 may function as a tumor suppressive miRNA and play an important role in inhibiting the HCC tumorigenesis by modulating the AKT/mTOR pathway. Our clinical evidence also highlight the prognostic potential of miR-149 in HCC. The newly identified miR-149/AKT/mTOR axis might be a promising therapeutic target in the prevention and treatment of HCC. Electronic supplementary material The online version of this article (doi:10.1186/1476-4598-13-253) contains supplementary material, which is available to authorized users.