Indomethacin Combined with Ciprofloxacin Improves the Prognosis of Mice under Severe Traumatic Infection via the PI3K/Akt Pathway in Macrophages.

The prevention and treatment strategies for traumatic infection often focus on the use of antibiotics, while eschew the combined treatment of the bacteria, their toxins, and inflammatory mediators. This might be a main reason the prognosis of wound victims has not improved. Although our previous work found that the combination of indomethacin (IND) and ciprofloxacin (CIP) could promote skin wound repair and enhance the immune function, the efficacy and safety of this strategy for severe traumatic infection-mediated complications remain unknown. Additionally, there is no study on the relevant target cells and molecular mechanisms. In this study, C57BL/6 adult male mice were modeled for severe traumatic infection, and the optimal doses of IND and CIP alone were determined. After that, the efficacy and safety of IND plus CIP in traumatic infection mice were explored. Then the differentially expressed genes of activated macrophages in this process were analysed and verified by transcriptomic methods and conventional experimental techniques. The role of a candidate signalling pathway (PI3K/Akt) in regulating macrophage function and drug combination therapy was evaluated. The results showed that IND plus CIP increased the survival rate, reduced the degree of inflammatory response, and enhanced the bacteriostatic effect in mice under traumatic infection. This combined therapy did not cause significant damage to the functions of important organs (liver, kidney, heart). In addition, IND combined with CIP induced macrophages to significantly change their expression levels of several cytokines, including interleukin (IL) -1ß, IL-6, IL-10, IL-22, IL-23A, IL-17A, IL-17F, cluster of differentiation (CD) 11b and other genes/encode proteins. Further study showed that intervention with the PI3K inhibitor LY294002 modulated the secretion function of the above-mentioned macrophages and Akt activation (phosphorylation at serine 473). IND plus CIP can regulate macrophage function through the PI3K/Akt signalling pathway and improve the prognosis of severe traumatic infected mice. This may be a new therapeutic strategy for the prevention and treatment of severe traumatic infection.

The performance of VCS(volume, conductivity, light scatter) parameters in distinguishing latent tuberculosis and active tuberculosis by using machine learning algorithm.

BACKGROUND: Tuberculosis is a chronic infectious disease caused by mycobacterium tuberculosis (MTB) and is the ninth leading cause of death worldwide. It is still difficult to distinguish active TB from latent TB,but it is very important for individualized management and treatment to distinguish whether patients are active or latent tuberculosis infection. METHODS: A total of 220 subjects, including active TB patients (ATB, n = 97) and latent TB patients (LTB, n = 113), were recruited in this study .46 features about blood routine indicators and the VCS parameters (volume, conductivity, light scatter) of neutrophils(NE), monocytes(MO), and lymphocytes(LY) were collected and was constructed classification model by four machine learning algorithms(logistic regression(LR), random forest(RF), support vector machine(SVM) and k-nearest neighbor(KNN)). And the area under the precision-recall curve (AUPRC) and the area under the receiver operating characteristic curve (AUROC) to estimate of the model's predictive performance for dentifying active and latent tuberculosis infection. RESULTS: After verification,among the four classifications, LR and RF had the best performance (AUROC = 1, AUPRC = 1), followed by SVM (AUROC = 0.967, AUPRC = 0.971), KNN (AUROC = 0.943, AUPRC = 0.959) in the training set. And LR had the best performance (AUROC = 0.977, AUPRC = 0.957), followed by SVM (AUROC = 0.962, AUPRC = 0.949), RF (AUROC = 0.903, AUPRC = 0.922),KNN(AUROC = 0.883, AUPRC = 0.901) in the testing set. CONCLUSIONS: The machine learning algorithm classifier based on leukocyte VCS parameters is of great value in identifying active and latent tuberculosis infection.

Leukocyte Cell Population Data (CPD) as a Discriminating Tool between Active Tuberculosis and Lung Cancer.

BACKGROUND: Cell population data (CPD) are parameters of cell size, shape, and content that can be used in the differential diagnosis of diseases such as leukemia, bacterial or viral infection, and dengue fever. The aim of this study was to screen for CPD parameters that can be used to differentiate active pulmonary tuberculosis (APTB) from lung cancer (LC) and to assess their efficacy. METHODS: Whole blood samples from 84 APTB patients, 109 LC patients, and 95 healthy volunteers were collected from January 2019 to November 2019. All samples were tested by DxH800 blood cell analyzer using VCS (volume, conductivity, and scatter) technology to obtain CPD parameters, total leukocyte count, and leukocyte classification count. The results were tested for normal distribution, followed by one-way analysis of variance (ANOVA) and area under the ROC curve (AUC) analysis to evaluate the diagnostic efficacy of CPD parameters. RESULTS: Twenty-three CPD parameters were significantly higher in the APTB group than in the LC group, 13 CPD parameters were significantly lower than in the LC group, and 6 CPD parameters were not statistically different between the two groups. The AUCs of CPD parameters between the APTB and LC groups were analyzed, and the results showed that the AUCs of nine CPD parameters were higher than 0.91, with the AUCs of neutronphil mean conductance (NMC), lymphocyte mean conductance (LMC), and monocyte mean conductance (MMC) even reaching 0.983, 0.930, and 0.996, respectively. Meanwhile, compared with the CPD parameters, white blood cells and their conventional differential counts (WBC, NE%, LY%, MO%) did not result in higher AUCs for the two groups (0.641, 0.757, 0.659, 0.733, respectively). CONCLUSIONS: Three CPD parameters (NMC, LMC, and MMC) obtained higher AUC than other indicators, and their combined diagnosis efficacy obtained 100% sensitivity and 99.1% specificity, which may be helpful for clinical differential diagnosis of APTB and LC.

Development of a novel integrated isothermal amplification system for detection of bacteria-spiked blood samples.

Bloodstream infection (BSI) caused by bacteria is highly pathogenic and lethal, and easily develops whole-body inflammatory state. Immediate identification of disease-causing bacteria can improve patient prognosis. Traditional testing methods are not only time-consuming, but such tests are limited to laboratories. Recombinase polymerase amplification combined with lateral flow dipstick (RPA-LFD) holds great promise for rapid nucleic acid detection, but the uncapping operation after amplification easily contaminates laboratories. Therefore, the establishment of a more effective integrated isothermal amplification system has become an urgent problem to be solved. In this study, we designed and fabricated a hermetically sealed integrated isothermal amplification system. Combining with this system, a set of RPA-LFD assays for detecting S. aureus, K. peneumoniae, P. aeruginosa, and H. influenza in BSI were established and evaluated. The whole process could be completed in less than 15 min and the results can be visualized by the naked eye. The developed RPA-LFD assays displayed a good sensitivity, and no cross-reactivity was observed in seven similar bacterial genera. The results obtained with 60 clinical samples indicated that the developed RPA-LFD assays had high specifcity and sensitivity for identifying S. aureus, K. peneumoniae, P. aeruginosa, and H. influenza in BSI. In conclusion, our results showed that the developed RPA-LFD assay is an alternative to existing PCR-based methods for detection of S. aureus, K. peneumoniae, P. aeruginosa, and H. influenza in BSI in primary hospitals.

Differentiation between Thalassemia Trait and Iron Deficiency Anemia Based on Low Hemoglobin Density and Microcytic Anemia Factor.

BACKGROUND: The most common causes of microcytic hypochromic anemia are thalassemia trait (TT) and iron deficiency anemia (IDA). Clinically, the differential diagnosis of TT and IDA is crucial, but it is typically challenging. Thus, in order to differentiate between TT and IDA, we seek to develop a new discriminative index on an automatic hematology analyzer utilizing the two new RBC characteristics of low hemoglobin density (LHD) and microcytic anemia factor (MAF). METHODS: We recruited a total of 323 subjects, including 115 healthy controls, 83 TT, and 125 IDA. An automated hematology analyzer (DxH800, Beckman Coulter) was used to determine peripheral blood parameters; LHD and MAF were calculated using the parameters of MCHC, Hb, and MCV. The receiver operating characteristic (ROC) curve was used to determine the cutoff values and evaluate the diagnostic value for TT and IDA. RESULTS: LHD was significantly lower in TT than IDA, whereas MAF was higher. To distinguish between TT and IDA, a new formula based on LHD and MAF was developed, with a cutoff value of 0.5, AUC of 0.9706 (95% CI: 0.9503 - 0.9909), and specificity, sensitivity, positive predictive value, and negative predictive values were 92.91%, 91.36%, 89.16%, and 94.40%, respectively. The new formula has proven advantages over conventional indices, such as RDW-SD, MCV, MCH, etc. Conclusions: The RBC parameters LHD and MAF detected by hematology analyzer could be useful for screening for TT and IDA. Our new formula outperforms other discriminant formulas in the literature with high sensitivity and specificity, is simple, rapid, and can aid in early detection and management.

Long non-coding RNA CASC7 is a promising serum biomarker for hepatocellular carcinoma.

BACKGROUND: At present, a large number of studies have found that long non-coding RNAs (lncRNAs) can be used as biomarkers for diagnosis and monitoring prognosis of hepatocellular carcinoma (HCC). The expression of lncRNA cancer susceptibility candidate 7 (CASC7) in HCC has rarely been studied. The purpose of this study was to explore the expression of CASC7 and its correlation with clinical features, and to further analyze its diagnostic value in HCC. METHODS: Serum samples were collected from 80 patients with HCC, 80 patients with chronic hepatitis B (CHB), and 80 healthy people. The expression level of serum CASC7 was detected by droplet digital PCR. Appropriate parametric and nonparametric tests were used for data analysis. RESULTS: The results showed that the expression of CASC7 in serum of patients with HCC was significantly higher than that of patients with CHB (median: 8.8 versus 2.2 copies/µl, p < 0.001) and healthy controls (median: 8.8 versus 3.8 copies/µl, p < 0.001). High expression of serum CASC7 was significantly correlated with tumor number (p = 0.005), intrahepatic metastasis (IM) (p < 0.001), tumor size (p = 0.007) and tumor-node-metastasis (TNM) stage (p = 0.008). The area under the curve (AUC) of CASC7 to distinguish HCC patients from CHB patients and healthy controls was 0.808 (95% CI: 0.742-0.874) at the cut-off value of 7.24 copies/µl with 63.8% sensitivity and 95.2% specificity. CONCLUSIONS: This study suggested that CASC7 was significantly up-regulated in serum of patients with HCC and closely related to tumor number, IM, tumor size and TNM stage, which may serve as a promising diagnostic biomarker.

Rapid detection of EGFR mutation in CTCs based on a double spiral microfluidic chip and the real-time RPA method.

Circulating tumor cells (CTCs) are cells shed from primary or metastatic tumors and spread into the peripheral bloodstream. Mutation detection in CTCs can reveal vital genetic information about the tumors and can be used for "liquid biopsy" to indicate cancer treatment and targeted medication. However, current methods to measure the mutations in CTCs are based on PCR or DNA sequencing which are cumbersome and time-consuming and require sophisticated equipment. These largely limited their applications especially in areas with poor healthcare infrastructure. Here we report a simple, convenient, and rapid method for mutation detection in CTCs, including an example of a deletion at exon 19 (Del19) of the epidermal growth factor receptor (EGFR). CTCs in the peripheral blood of NSCLC patients were first sorted by a double spiral microfluidic chip with high sorting efficiency and purity. The sorted cells were then lysed by proteinase K, and the E19del mutation was detected via real-time recombinase polymerase amplification (RPA). Combining the advantages of microfluidic sorting and real-time RPA, an accurate mutation determination was realized within 2 h without professional operation or complex data interpretation. The method detected as few as 3 cells and 1% target variants under a strongly interfering background, thus, indicating its great potential in the non-invasive diagnosis of E19del mutation for NSCLC patients. The method can be further extended by redesigning the primers and probes to detect other deletion mutations, insertion mutations, and fusion genes. It is expected to be a universal molecular diagnostic tool for real-time assessment of relevant mutations and precise adjustments in the care of oncology patients.

Centrifugal microfluidic-based multiplex recombinase polymerase amplification assay for rapid detection of SARS-CoV-2.

The COVID-19 pandemic has spread worldwide, and rapid detection of the SARS-CoV-2 virus is crucial for infection surveillance and epidemic control. This study developed a centrifugal microfluidics-based multiplex reverse transcription recombinase polymerase amplification (RT-RPA) assay for endpoint fluorescence detection of the E, N, and ORF1ab genes of SARS-CoV-2. The microscope slide-shaped microfluidic chip could simultaneously accomplish three target genes and one reference human gene (i.e., ACTB) RT-RPA reactions in 30 min, and the sensitivity was 40 RNA copies/reaction for the E gene, 20 RNA copies/reaction for the N gene, and 10 RNA copies/reaction for the ORF1ab gene. The chip demonstrated high specificity, reproducibility, and repeatability. Chip performance was also evaluated using real clinical samples. Thus, this rapid, accurate, on-site, and multiplexed nucleic acid test microfluidic chip would significantly contribute to detecting patients with COVID-19 in low-resource settings and point-of-care testing (POCT) and, in the future, could be used to detect emerging new variants of SARS-CoV-2.

Isothermal exponential amplification reactions triggered by circular templates (cEXPAR) targeting miRNA.

BACKGROUND: Isothermal exponential amplification reaction (EXPAR) is an emerging amplification technique that is most frequently used to amplify microRNA (miRNA). However, EXPAR also exhibits non-specific background amplification in the absence of the targeted sequence, which limits the attainable assay sensitivity of EXPAR. METHODS AND RESULTS: A novel modified isothermal EXPAR based on circular amplification templates (cEXPAR) was developed in this study. The circular template consists of two same linear fragments that complement the target sequence, and these two linear fragments are separated by two nicking agent recognition sequences (NARS). Compared with the linear structure template, this circular template allows DNA or RNA fragments to be randomly paired with two repeated sequences and can be successfully amplified. This reaction system developed in this study could rapidly synthesize short oligonucleotide fragments (12-22 bp) through simultaneous nicking and displacement reactions. Highly sensitive chain reactions can be specifically triggered by as low as a single copy of target molecule, and non-specific amplification can be effectively eliminated in this optimized system. Moreover, the proposed approach applied to miRNA test can discriminate single-nucleotide variations between miRNAs. CONCLUSION: The newly developed cEXPAR assay provides a useful alternative tool for rapid, sensitive, and highly specific detection of miRNAs.

A feedback loop between NONHSAT024276 and PTBP1 inhibits tumor progression and glycolysis in HCC by increasing the PKM1/PKM2 ratio.

Hepatocellular carcinoma (HCC) is one of the most common malignancies with a hallmark of aberrant metabolism. The mechanism of long noncoding RNAs (lncRNAs) underlying the aggressive behaviors and glycolysis of HCC is poorly understood. In this study, we identified, via microarray, novel lncRNA NONHSAT024276 as a potential tumor suppressor in HCC. The downregulation of NONHSAT024276 closely correlated with larger tumor volume and higher aspartate transaminase levels. Functional experiments were performed to verify the role of NONHSAT024276 in HCC progression, and the negative effects of NONHSAT024276 expression on cell proliferation and migration were identified. Mechanistically, NONHSAT024276 directly bound to polypyrimidine tract-binding protein 1 (PTBP1), downregulating it and forming a feedback loop. Furthermore, NONHSAT024276 increased the ratio of M1 and M2 isoforms of pyruvate kinase (PKM1/PKM2) and also obstructed the PTBP1/PKM-mediated glycolysis. Finally, the rescue assays confirmed that NONHSAT024276 functioned in HCC via downregulating PTBP1 to increase the PKM1/PKM2 ratio. Hence, this study supported a model in which NONHSAT024276 downregulated PTBP1 and formed a feedback loop to increase the PKM1/PKM2 ratio to inhibit glycolysis and progression of HCC, opening new prospects for preventing or treating HCC.

Predictive Value of the Peripheral Blood Parameters for Preeclampsia.

BACKGROUND: Excessive systemic inflammation plays a vital role in pathophysiology of preeclampsia (PE). The aim is to clarify the predictive value of the peripheral blood parameters including white blood cell (WBC), neutrophils, lymphocyte, monocyte, platelet count, mean platelet volume (MPV), plateletcrit, platelet distribution width (PDW), platelet-large cell ratio (PLCR), and the ratio value for PE. METHODS: This retrospective study enrolled 170 PE patients, 123 healthy control pregnant women, and 122 non-pregnant women. When pregnant women were admitted to the hospital for delivery, peripheral complete blood cell count was detected by an automatic blood cell analyzer. Clinical signs and demographic characteristics were recorded. The receiver operating characteristic (ROC) curve was used to determine the cutoff value and analyze the predictive significances for PE. Furthermore, the risk factors of PE were tested by univariate and stratified analyses. RESULTS: This study showed that WBC, neutrophil count, neutrophil percentage, NLR, NMR, and PLR# were significantly increased in PE patients as compared with pregnant control patients (p < 0.001), whereas lymphocyte percentage, monocyte percentage, and PNR were decreased. In addition, there was no significant difference in the rest of the peripheral blood parameters between women with and without PE. The ROC curve result revealed that WBC and neutrophil count had a higher AUC value than the rest of peripheral blood variables. WBC and neutrophil count are positively correlated MAP. Moreover, the WBC and neutrophil count were indicated as independent risk factors for the development of PE. CONCLUSIONS: This study clarifies that peripheral blood parameters of WBC and neutrophil count have good applied value with high sensitivity and specificity in predicting the development of PE and are also independent risk factors for the development of PE.

Epidemiology and resistance mechanisms of tigecycline- and carbapenem-resistant Enterobacter cloacae in Southwest China: a 5-year retrospective study.

OBJECTIVES: The prevalence and molecular epidemiology of tigecycline resistance in carbapenem-resistant Enterobacter cloacae (CREC) in mainland China is unknown. We aimed to investigate the molecular characteristics and mechanisms of tigecycline-resistant CREC (TCREC) in Southwest China. METHODS: We conducted a 5-year retrospective study. TCREC isolates underwent antimicrobial susceptibility testing, PFGE and MLST. We determined the presence of genes, deficiency of outer membrane proteins (OMPs) and expression of efflux pumps using PCR, reverse transcription PCR and SDS-PAGE. RESULTS: A large proportion of CREC isolates (21.7%; 36/166) were TCREC. All isolates were resistant to ertapenem, whereas 67% remained susceptible to imipenem and meropenem. ST88 (10/36; 27.8%) was predominant and was associated with moderate resistance to tigecycline and high resistance to carbapenems, followed by ST256 (3/36; 8.3%), ST78 (2/36; 5.6%), ST557 (2/36; 5.6%) and ST102 (2/36; 5.6%). blaNDM-1 (6/36; 16.7%) was the most common carbapenemase gene, and ST88 (5/6; 83.3%) was the most common type, followed by blaIMP-8 (3/36; 8.3%). Coexistence of extended-spectrum ß-lactamase (ESBL) genes and OmpF and/or OmpC loss was found in 27/36 isolates; additionally, increased co-expression of efflux pump genes acrB and oqxA was identified in 25/36 isolates, which may together contribute to co-resistance to carbapenems and tigecycline. CONCLUSION: Most ST88 strains carried carbapenemases, especially NDM-1. Overexpression of efflux pumps contributed to tigecycline resistance. The presence of carbapenemase and/or ESBL genes and lack of OMPs, but not efflux pump overexpression, may confer carbapenem resistance. Reasonable supervision and management of the epidemic of TCREC will help to stem the transmission of isolates.

The VCS Parameters of Lymphocytes may Improve Discrimination between Bacterial Infection and Viral Infection.

BACKGROUND: Distinguishing bacterial infections from viral infections is very important for accurate and appro-priate drug treatment, alleviating diseases and avoiding side effects caused by drug abuse. The aim of this study is to assess the clinical usefulness of the lymphocyte VCS (volume, conductivity, light scatter) parameters to dis-tinguish bacterial infection from viral infection. METHODS: Peripheral blood was collected from 60 viral infection patients (VIG), 63 bacterial infection patients (BIG), and 95 healthy controls (HC). The lymphocyte VCS parameters and blood routine indicators were obtained by using a hematology analyzer with VCS technology. The critical cutoff value, sensitivity and specificity were established based on receiver operator characteristic (ROC) curve analysis. RESULTS: Mean volume of lymphocytes (MV-LY), median angle light scatter of lymphocytes (MALS-LY), upper median angle light scatter of lymphocytes (UMALS-LY), neutrophil-lymphocyte ratio (NLR) were significantly increased in the bacterial infection group compared with the viral infection group and the healthy controls. The area under curve (AUC) for mean volume of lymphocytes (MV-LY) was 0.8143 for discriminating the bacterial infection group from the viral infection group. For median angle light scatter of lymphocytes (MALS-LY), the area under curve (AUC) was 0.8116. For upper median angle light scatter of lymphocytes (UMALS-LY), the area under curve (AUC) was 0.8631. For neutrophil-lymphocyte ratio (NLR), the area under curve (AUC) was 0.8513. CONCLUSIONS: This study clarifies that mean volume of lymphocytes, median angle light scatter of lymphocytes, and upper median angle light scatter of lymphocytes have good clinical practical value in distinguishing bacterial infection from viral infection and healthy controls because of its high sensitivity and specificity.

Enhanced Separation Efficiency and Purity of Circulating Tumor Cells Based on the Combined Effects of Double Sheath Fluids and Inertial Focusing.

Circulating tumor cells (CTCs) play a crucial role in solid tumor metastasis, but obtaining high purity and viability CTCs is a challenging task due to their rarity. Although various works using spiral microchannels to isolate CTCs have been reported, the sorting purity of CTCs has not been significantly improved. Herein, we developed a novel double spiral microchannel for efficient separation and enrichment of intact and high-purity CTCs based on the combined effects of two-stage inertial focusing and particle deflection. Particle deflection relies on the second sheath to produce a deflection of the focused sample flow segment at the end of the first-stage microchannel, allowing larger particles to remain focused and entered the second-stage microchannel while smaller particles moved into the first waste channel. The deflection of the focused sample flow segment was visualized. Testing by a binary mixture of 10.4 and 16.5 µm fluorescent microspheres, it showed 16.5 µm with separation efficiency of 98% and purity of 90% under the second sheath flow rate of 700 µl min-1. In biological experiments, the average purity of spiked CTCs was 74% at a high throughput of 1.5 × 108 cells min-1, and the recovery was more than 91%. Compared to the control group, the viability of separated cells was 99%. Finally, we validated the performance of the double spiral microchannel using clinical cancer blood samples. CTCs with a concentration of 2-28 counts ml-1 were separated from all 12 patients' peripheral blood. Thus, our device could be a robust and label-free liquid biopsy platform in inertial microfluidics for successful application in clinical trials.

Cell population data in identifying active tuberculosis and community-acquired pneumonia.

OBJECTIVE: Leukocyte morphological parameters known as CPD (cell population data) is detected by hematology analyzer UniCel DxH800 with VCS technology. This study aimed to investigate the diagnostic efficacy of morphological changes in CPD parameters in distinguishing active tuberculosis from community-acquired pneumonia. METHODS: From October 2018 to February 2019, 88 patients with active tuberculosis, 78 patients with community-acquired pneumonia, and 89 healthy controls were enrolled in this study. CPD was obtained using Unicel DxH800 analyzer for all whole blood samples, one-way ANOVA (non-parametric) and area analysis under ROC curve were performed. RESULTS: The neutrophil mean conductivity (NMC), monocyte mean volume (MMV), monocyte mean conductivity (MMC), lymphocyte percentage (LY%), and monocyte percentage (MO%) were significantly higher in the active tuberculosis group than in the community-acquired pneumonia group. The white blood cell (WBC) count and neutrophil percentage (NE%) were significantly lower in the active tuberculosis group than in the community-acquired pneumonia group. The analysis of the area under the ROC curve proved that WBC count, neutrophil percentage (NE%), lymphocyte percentage (LY%), and monocyte percentage (MO%) did not achieve a higher area under the curve (AUC: 0.63, 0.71, 0.62, and 0.7, respectively). However, the AUC of NMC, MMV, and MMC in the CPD parameters was 0.951, 0.877, 0.98, respectively, and the simultaneous measurement of the three parameters was 0.99. The sensitivity and specificity were 98.5% and 91.1%, respectively. CONCLUSION: The combined diagnosis of NMC, MMV, and MMC could assist the clinical diagnosis of active tuberculosis and community-acquired pneumonia.

Evaluation of the Diagnostic Value of Peripheral Blood Parameters for Neonatal Pneumonia.

BACKGROUND: To evaluate the diagnostic value of peripheral blood parameters including white blood cell (WBC), neutrophil, lymphocyte, monocyte, platelet, mean platelet volume (MPV), platelet distribution width (PDW), mean corpuscular volume (MCV), red cell distribution width (RDW), neutrophil-to-lymphocyte ratio (NLR), monocyte-to-lymphocyte (MLR), platelet-to-lymphocyte ratio (PLR) and platelet-to-neutrophil ratio (PNR) for neonatal pneumonia. METHODS: Two hundred and six full-term neonates in our hospital from January 2018 to December 2019 were enrolled, including 73 pneumonic neonates and 133 health controls. Peripheral blood parameters were measured by an automatic blood cell analyzer. While C-reactive protein (CRP) and PCT concentrations were detected by electrochemical luminescence assay. Clinical signs, characteristic population, temperature, and chest radiograph findings were recorded. The receiver operating characteristic (ROC) curve was used to determine the cutoff values and analyze the diagnosis significances for neonatal pneumonia. RESULTS: This study showed that WBC, neutrophil, RDW, NLR, and MLR levels in the pneumonic group were higher than that of the control group, whereas lymphocyte, monocyte, platelet, and PNR levels were lower (p < 0.05). The ROC curve result showed that NLR and PNR owned higher AUC values than the rest of peripheral blood variables. At a cutoff value 2.581, NLR exhibited 63.01% sensitivity, 90.98% specificity, and 0.847 area under ROC curve (AUC). In addition, at a cutoff value 52.77, PNR showed 84.93% sensitivity, 78.95% specificity, and 0.856 AUC. CONCLUSIONS: This study clarifies that peripheral blood parameter of NLR and PNR have good applied value in diagnosis neonatal pneumonia with high sensitivity and specificity.

Screening and Functional Prediction of Key Candidate Genes in Hepatitis B Virus-Associated Hepatocellular Carcinoma.

BACKGROUND: The molecular mechanism by which hepatitis B virus (HBV) induces hepatocellular carcinoma (HCC) is still unknown. The genomic expression profile and bioinformatics methods were used to investigate the potential pathogenesis and therapeutic targets for HBV-associated HCC (HBV-HCC). METHODS: The microarray dataset GSE55092 was downloaded from the Gene Expression Omnibus (GEO) database. The data was analyzed by the bioinformatics software to find differentially expressed genes (DEGs). Gene Ontology (GO) enrichment analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, ingenuity pathway analysis (IPA), and protein-protein interaction (PPI) network analysis were then performed on DEGs. The hub genes were identified using Centiscape2.2 and Molecular Complex Detection (MCODE) in the Cytoscape software (Cytoscape_v3.7.2). The survival data of these hub genes was downloaded from the Gene Expression Profiling Interactive Analysis (GEPIA). RESULTS: A total of 2264 mRNA transcripts were differentially expressed, including 764 upregulated and 1500 downregulated in tumor tissues. GO analysis revealed that these DEGs were related to the small-molecule metabolic process, xenobiotic metabolic process, and cellular nitrogen compound metabolic process. KEGG pathway analysis revealed that metabolic pathways, complement and coagulation cascades, and chemical carcinogenesis were involved. Diseases and biofunctions showed that DEGs were mainly associated with the following diseases or biological function abnormalities: cancer, organismal injury and abnormalities, gastrointestinal disease, and hepatic system disease. The top 10 upstream regulators were predicted to be activated or inhibited by Z-score and identified 25 networks. The 10 genes with the highest degree of connectivity were defined as the hub genes. Cox regression revealed that all the 10 genes (CDC20, BUB1B, KIF11, TTK, EZH2, ZWINT, NDC80, TPX2, MELK, and KIF20A) were related to the overall survival. CONCLUSION: Our study provided a registry of genes that play important roles in regulating the development of HBV-HCC, assisting us in understanding the molecular mechanisms that underlie the carcinogenesis and progression of HCC.

Effects of Blood Storage on Cell Population Data.

BACKGROUND: Cell population data (CPD) are leukocyte morphologic parameters, measured by automated hematology analyzers with VCS technology. Many studies have demonstrated that these parameters may have clinical utility for diagnosing or screening certain pathological conditions. This study is to investigate the effects of peripheral blood stored at room temperature on the CPD values and provide useful information about storage-induced CPD changes. METHODS: Venous blood samples from healthy donors kept at room temperature (18 - 25°C) for different time intervals were analyzed using a Coulter DxH800 hematology analyzer. The CPD data collected included mean cellular volume, conductivity and multiple angles of light scatters as well as their corresponding standard deviation for neutrophils, lymphocytes, and monocytes. Peripheral blood smears at each time interval were also prepared and examined microscopically. RESULTS: Peripheral blood kept at room temperature over time significantly affects the CPD values for neutrophils, lymphocytes, and monocytes. These CPD changes are correlated with the morphologic alterations observed under light microcopy, but detected much earlier. Some changes imitate clinical pathological conditions. For example, aged neutrophils showed decreased median angle light scatters, suggesting cytoplasmic degranulation, which can be seen in the case of myelodysplastic syndrome. CONCLUSIONS: This study provides valuable information about storage-induced CPD changes that can affect potential clinical application and interpretation of these automated digital morphologic parameters.

A Novel lncRNA NONHSAT053785 Acts as an Independent Risk Factor for Intrahepatic Metastasis of Hepatocellular Carcinoma.

PURPOSE: Long noncoding RNAs (lncRNAs) in body fluids have been considered as promising novel biomarkers for tumor-related diseases. The present study aimed to investigate the expression level of lncRNA NONHSAT053785 in serum and its correlation with clinical characteristics of hepatocellular carcinoma (HCC) patients. METHODS: The droplet digital PCR (ddPCR) was used to measure the serum levels of NONHSAT053785 in 112 HCC patients, 96 chronic hepatitis B (CHB) patients, and 99 healthy controls (HC). The correlation between NONHSAT053785 and clinical characteristics was analyzed by chi-square test and Spearman correlation test. The risk factors of intrahepatic metastasis (IM) were detected by univariate and multivariate analyses. Furthermore, the diagnostic value of NONHSAT053785 in HCC and its predictive ability in IM were evaluated by the receiver operating characteristic (ROC) curves. RESULTS: The level of NONHSAT053785 was significantly increased in the serum of HCC patients and was higher in HCC patients with IM as compared to those without. Additionally, the expression level of NONHSAT053785 was significantly related to IM, Child-Pugh classification, and peripheral blood indicators such as liver metabolic enzymes and positively correlated to IM, Barcelona Clinic Liver Cancer (BCLC) staging, and some peripheral blood indicators. Furthermore, the serum NONHSAT053785 was indicated as an independent predictor for IM in the elderly, non-smoking, drinking, and tumor size ≥5 cm subjects. The area under the ROC curve (AUC) was 0.801 (P <0.0001) for diagnosis of HCC and 0.678 (P =0.0015) for predicting IM. CONCLUSION: The increase in serum NONHSAT053785 levels was related to an increased risk of IM, and hence, may serve as a novel biomarker for the diagnosis of HCC and the prediction of IM.

An ultraportable and versatile point-of-care DNA testing platform.

Point-of-care testing (POCT) has broad applications in resource-limited settings. Here, a POCT platform termed POCKET (point-of-care kit for the entire test) is demonstrated that is ultraportable and versatile for analyzing multiple types of DNA in different fields in a sample-to-answer manner. The POCKET is less than 100 g and smaller than 25 cm in length. The kit consists of an integrated chip (i-chip) and a foldable box (f-box). The i-chip integrates the sample preparation with a previously unidentified, triple signal amplification. The f-box uses a smartphone as a heater, a signal detector, and a result readout. We detected different types of DNA from clinics to environment to food to agriculture. The detection is sensitive (<103 copies/ml), specific (single-base differentiation), speedy (<2 hours), and stable (>10 weeks shelf life). This inexpensive, ultraportable POCKET platform may become a versatile sample-to-answer platform for clinical diagnostics, food safety, agricultural protection, and environmental monitoring.