Development and clinical evaluation of a rapid antibody lateral flow assay for the diagnosis of SARS-CoV-2 infection.

BACKGROUND: The novel coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which has quickly spread worldwide since its outbreak in December 2019. One of the primary measures for controlling the spread of SARS-CoV-2 infection is an accurate assay for its diagnosis. SARS-CoV-2 real-time PCR kits suffer from some limitations, including false-negative results in the clinic. Therefore, there is an urgent need for the development of a rapid antibody test kit for COVID-19 diagnosis. METHODS: The nuclear capsid protein (N) and spike protein 1 (S1) fragments of SARS-CoV-2 were expressed in Escherichia coli, and rapid antibody-based tests for the diagnosis of SARS-CoV-2 infection were developed. To evaluate their clinical applications, the serum from COVID-19 patients, suspected COVID-19 patients, recovering COVID-19 patients, patients with general fever or pulmonary infection, doctors and nurses who worked at the fever clinic, and health professionals was analyzed by the rapid antibody test kits. The serum from patients infected with Mycoplasma pneumoniae and patients with respiratory tract infection was further analyzed to test its cross-reactivity with other respiratory pathogens. RESULTS: A 47 kDa N protein and 67 kDa S1 fragment of SARS-CoV-2 were successfully expressed, purified, and renatured. The rapid antibody test with recombinant N protein showed higher positive rate than the rapid IgM antibody test with recombinant S1 protein. Clinical evaluation showed that the rapid antibody test kit with recombinant N protein had 88.56 % analytical sensitivity and 97.42 % specificity for COVID-19 patients, 53.48 % positive rate for suspected COVID-19 patients, 57.14 % positive rate for recovering COVID-19 patients, and 0.5-0.8 % cross-reactivity with other respiratory pathogens. The analytical sensitivity of the kit did not significantly differ in COVID-19 patients with different disease courses (p < 0.01). CONCLUSIONS: The rapid antibody test kit with recombinant N protein has high specificity and analytical sensitivity, and can be used for the diagnosis of SARS-CoV-2 infection combined with RT-PCR.

The intestinal microbial community dissimilarity in hepatitis B virus-related liver cirrhosis patients with and without at alcohol consumption.

BACKGROUND: Chronic hepatitis B virus (HBV) infection-reduced liver functions are associated with intestinal microbial community dissimilarity. This study aimed to investigate the microbial community dissimilarity in patients with different grades of HBV-related liver cirrhosis. RESULTS: Serum endotoxin was increased with Child-Pugh (CP) class (A, B, and C). Veillonellaceae and Lachnospiraceae families were reduced in patients compared with controls. Megamonas and Veillonella genus was reduced and increased in patients compared with controls, respectively, especially in CPB and CPC groups. Correlation analysis showed that endotoxin content was significantly correlated with alcohol consumption (95% CI 0.100, 0.493), CP class (95% CI 0.289, 0.687) and Lachnospiraceae family level (95% CI - 0.539, - 0.122). Firmicutes/Bacteroidetes ratio was correlated with the level of Lachnospiraceae family (95% CI 0.013, 0.481), Veillonellaceae family (95% CI 0.284, 0.696), Megamonas genus (95% CI 0.101, 0.518) and Veillonella genus (95% CI 0.134, 0.545). All aforementioned bacteria were independent risk or protective factors for hepatitis. Alcohol consumption changed microbial community. CONCLUSIONS: Our study demonstrated that elevated Firmicutes/Bacteroidetes ratio, reduced Megamonas genus level and increased Veillonella genus level were indicators for HBV-related liver cirrhosis. Alcohol-related pathogenesis was associated with the changed microbial community.

A Novel Approach of Urinary Monohydroxyphenyl Metabolites Assay in Cancer Screening.

BACKGROUND: A noninvasive, fast, highly sensitive and simple test is needed for cancer screening in addition to the detection of biomarkers in blood. Recently, the patent (CN102565055A) for the Urinary Monohydroxyphenyl Metabolites Assay (UMM-A) was authorized, and the effectiveness of clinical application has yet to be studied further. METHODS: A retrospective study was conducted consisting of 432 cancer patients, 28 benign tumor patients, 117 non-cancerous diseases patients, and 120 healthy donors to analyze the levels of monohydroxyphenyl metabolites in the urine sample. A logistic regression model was used to study the possible confounding factors affecting the diagnostic performance and to test the probability of a case to be positive for UMM-A. RESULTS: Compared with healthy donors, non-cancerous disease, and benign tumor subjects, the positive rate and MM level of UMM-A in cancer patients have significantly increased. After the 246 retreated cancer patients were excluded, and 186 untreated cancer patients were included, with the same specificity to 77.0%, the sensitivity improved from 66.7 to 89.8%, the negative predictive value improved from 58.6 to 91.4%. CONCLUSIONS: The present study has provided important information on the diagnostic characteristics of UMM-A for untreated cancer and its potential application in cancer screening.

Association of cytotoxic T-lymphocyte antigen 4 gene with immune thrombocytopenia in Chinese Han children.

OBJECTIVES: To investigate the association of cytotoxic T lymphocyte-associated antigen 4 (CTLA4) with immune thrombocytopenia (ITP). METHODS: A case-control association analysis of 277 Chinese Han children was performed. The tagging variants rs11571315 and rs3087243 in the CTLA4 gene were detected using polymerase chain reaction-restriction fragment length polymorphism method. The expression quantitative trait loci (eQTL) analysis and quantitative real-time polymerase chain reaction were performed to determine the relationship of CTLA4 with ITP. RESULTS: Neither SNP was significantly different between case and control groups in either the genotypic or allelic distribution. The eQTL analysis results indicated that in the spleen, the rs3087243 was significant with the expression of CTLA4. The rs11571315 has similar results. Interestingly, the transcript level of CTLA4 was found to significantly decrease in patients with ITP. DISCUSSION: The autoimmune and gene etiology is implicated in the pathogen of ITP. The CTLA4 is important for negative regulation of T-cell activation, and CTLA-4 gene has been identified as a risk factor for some autoimmune diseases. However, association studies of ITP and CTLA4 gene have obtained conflicting results. This is the first study to systematically investigate the association of CTLA4 with ITP in Chinese Han children. CONCLUSIONS: The CTLA4 gene is suggested to correlate with ITP through its abnormal expression level instead of gene site mutation.

Predictive value of the neutrophil-to-lymphocyte ratio and hemoglobin insystemic lupus erythematosus.

The aim of the present study was to evaluate the association of the neutrophil-to-lymphocyte ratio (NLR) and hemoglobin levels with disease activity in patients with systemic lupus erythematosus (SLE) and to explore their clinical significance in predicting SLE. The present study included 212 patients with SLE and 201 healthy controls. All the clinical characteristics were collected from their medical records. The results revealed that the NLR was elevated and the hemoglobin level was markedly decreased in the patients with SLE compared with the healthy controls. NLR was positively correlated with the SLE Disease Activity Index 2000 (SLEDAI-2K), erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP), whereas it was not correlated with C3 or C4. The hemoglobin level was negatively correlated with SLEDAI-2K, ESR and CRP and positively correlated with C3 and C4. In addition, NLR [EXP(B)=1.986; 95% confidence interval (CI), 1.432-2.753; P=0.001] and hemoglobin [EXP(B)=0.947; 95% CI, 0.929-0.965; P=0.001] were independent predictive factors of SLE. The optimal NLR cut-off value for predicting SLE was 2.075, with 71.14% sensitivity and 69.57% specificity, whereas the optimal hemoglobin cut-off value was 131.5 mg/l, with 75.79% sensitivity and 77.98% specificity. In addition, high NLR together with low hemoglobin levels and high NLR or low hemoglobin levels had increased positive predictive values (86.05 and 66.95, respectively). High NLR with low hemoglobin levels and high NLR or low hemoglobin levels also had higher sensitivity (64.91 and 92.40, respectively) and specificity (64.91 and 18.95, respectively), compared with high NLR alone or low hemoglobin alone. In conclusion, NLR and hemoglobin may reflect SLE disease activity and may be used as markers for predicting the outcome of SLE.

The role of aryl hydrocarbon receptor in bone remodeling.

Bone remodeling is a persistent process for maintaining skeletal system homeostasis, and it depends on the dynamic equilibrium between bone-forming osteoblasts and bone-resorbing osteoclasts. Aryl hydrocarbon receptor (Ahr), a ligand-activated transcription factor, plays a pivotal role in regulating skeletal system. In order to better understand the role of Ahr in bone remodeling, we focused on bone remodeling characteristic, and the effects of Ahr on bone formation and differentiation, which suggest that Ahr is a critical control factor in the process of bone remodeling. Moreover, we discussed the impacts of Ahr on several signaling pathways related to bone remodeling, hoping to provide a theoretical basis to improve bone remodeling.

Higher serum level of myoglobin could predict more severity and poor outcome for patients with sepsis.

BACKGROUND: There have been sporadic case reports published focusing on myoglobin and sepsis. However, there are no systematic studies evaluating the correlation between myoglobin level and sepsis. This study investigated the correlation between the serum myoglobin level and the severity of septic patients. Next, we assessed the predictive value of the serum myoglobin level for the prognosis of septic patients. METHODS: Seventy septic patients were included and subdivided into the following 3 groups: sepsis group, severe sepsis group, and septic shock group. We collected blood samples at 0, 6, 12, 18, and 24hours after admission. The serum levels of myoglobin, C-reactive protein, and procalcitonin were analyzed. We also evaluated the levels of malondialdehyde, which is a biomarker for oxidative stress. RESULTS: The data indicate that the myoglobin level increased gradually within 24hours after admission. The median myoglobin levels of the sepsis, severe sepsis, and septic shock groups were 635.7, 903.6, and 1094.8µg/L, respectively (P<.05). The elevated myoglobin level was positively correlated with Sequential Organ Failure Assessment score, C-reactive protein, and procalcitonin level in septic patients. The increased myoglobin level was also associated with the mortality of septic patients. The Kaplan-Meier survival curves indicated that patients with high myoglobin levels had an elevated mortality rate. Moreover, an elevated myoglobin level indicated more oxidative stress. CONCLUSIONS: The myoglobin level can be detected in the early stage of sepsis and may serve as a potential biomarker for evaluating sepsis severity and further prognosis.

Detection of microRNA-200b may predict the inhibitory effect of gefitinib on non-small cell lung cancer and its potential mechanism.

The present study aimed to investigate the association and underlying mechanisms between microRNA-200b level and the inhibitory effect of gefitinib on non-small cell lung cancer. In total, 100 patients (43 males and 57 females; median age, 63 years) with advanced non-small cell lung cancer (NSCLC) were selected. All patients were administered with gefitinib orally (250 mg/day) and the effect of gefitinib was evaluated according to the Response Evaluation Criteria in Solid Tumors guidelines. Tumor tissue and plasma samples were collected prior to and subsequent to therapy. The microRNA-200b levels in tissues and plasma were determined by quantitative polymerase chain reaction (PCR). A549 cells were cultured in vitro and transfected with microRNA-200b mimic. Using Cell Counting Kit-8 assay, the proliferation inhibition detected was induced by 0.1 µM gefitinib in transfected or non-transfected A549 cells. Cell apoptosis and cell cycle progression were analyzed by flow cytometry and the migration of cells was observed by Transwell assay. In addition, mRNA and protein levels of insulin-like growth factor 1 receptor (IGF-1R), protein kinase B (AKT) and extracellular signal-related kinase (ERK), together with the phosphorylation of AKT and ERK in A549 cells, were determined by quantitative PCR and western blot analysis, respectively. The microRNA-200b levels in gefitinib-insensitive patients were decreased compared with gefitinib-sensitive patients. Transfection with microRNA-200b mimic increased the gefitinib induced proliferation inhibition, apoptosis and cell cycle arrest in A549 cells. Also, transfection with microRNA-200b mimic increased the migration inhibitory effect of gefitinib on A549 cells. Decreased IGF-1R expression together with reduced phosphorylation of AKT and ERK were observed following transfection of A549 cells with the microRNA 200b mimic. In conclusion, detection of microRNA-200b may predict the inhibitory effect of gefitinib on NSCLC. Upregulation of microRNA-200b led to the elevated sensitivity of glioma cells to gefitinib, and this effect may be explained as microRNA-200b being able to inhibit the expression of IGF-1R, thereby reducing the activation of downstream phosphoinositide 3-kinase/AKT and mitogen-activated protein kinase signaling pathways.

Clinical evaluation of circulating microRNA-25 level change in sepsis and its potential relationship with oxidative stress.

OBJECTIVE: To investigated the circulating microRNA expression profile in sepsis and its clinical evaluation. METHODS: 70 patients with sepsis and 30 patients with SIRS were selected and their blood samples were collected. Using liquid bead array with 3 statistical analysis approaches analyzed the circulating microRNA expression profiles, for confirming the data of liquid bead array, qRT-PCR was performed. The prognostic value of the changed microRNA in sepsis was determined and compared with CRP and PCT by analyzing the receiver operating characteristic (ROC) curves. To reveal whether the selected microRNAs could predict the outcome of patients, 28 d survival rate were calculated using Kaplan-Meier curves. Furthermore, the level of malondialdehyde (MDA), activity of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in plasma were detected and the relationship with the changed microRNA was determined. RESULTS: By integrating data from liquid bead array, we ultimately identified 6 microRNAs that were consistently changed in both of 3 statistical analysis approaches, however, only the change of microRNA-25 was significant according to the qPCR's result. The area under ROC curve showed that the clinical accuracy of microRNA-25 for sepsis diagnosis was better than CRP and PCT (AUG=0.806, 0.676 and 0.726, P<0.05).The decrease in level of microRNA-25 was correlated with the severity of sepsis, SOFA score, CRP and PCT level, meanwhile, microRNA-25 level can be used for predicting the prognosis of patients, the patients with microRNA-25 level ≤0.492 had a lower 28 d survival rate. Moreover, Decreased microRNA-25 level was related to the level of oxidative stress indicators in sepsis patients. CONCLUSIONS: MicroRNA-25 can be used as a biomarker for the diagnosis and assessment of sepsis. Meanwhile, microRNA-25 level may be associated with oxidative stress in patients with sepsis, and it is expected to become a target for anti-oxidation therapy.