Snap25 attenuates neuronal injury via reducing ferroptosis in acute ischemic stroke.

Due to the limited clinical treatment options for acute ischemic stroke (AIS), there is still an urgent requirement for in-depth research on the pathogenesis of AIS and the development of efficient therapeutic approaches and agents. Literature reports reveal that ferroptosis may play an important role in the pathogenesis of AIS. However, the specific mechanism and the molecular target of action of ferroptosis in AIS injury remains unclear. In this study, we constructed AIS rat and PC12 cell models. We applied RNAi-mediated knockdown and gene overexpression technologies to investigate whether Snap25 (Synaptosome-associated protein 25 kDa) can regulate the level of AIS damage by interfering with the level of ferroptosis in AIS. The in vivo and in vitro results revealed that the level of ferroptosis significantly increased in the AIS model. Snap25 gene overexpression significantly inhibited the ferroptosis level and decreased the AIS damage and OGD/R injury level in the model group. Snap25 silencing exacerbated the ferroptosis level and aggravated OGD/R injury in PC12 cells. The overexpression and silencing of Snap25 can significantly affect the expression level of ROS, suggesting that the regulatory effect on the ROS level may be an important factor in regulating ferroptosis in AIS by Snap25. In conclusion, the findings of this study suggested that Snap25 has a protective effect against ischemia/reperfusion injury by reducing ROS levels and ferroptosis levels. This study further confirmed the involvement of ferroptosis in the process of AIS injury and explored the regulatory mechanism of Snap25 on the ferroptosis level in AIS, which could provide a promising therapeutic target for ischemic stroke treatment.

High prevalence of vitamin D deficiency in Shenzhen pregnant women.

BACKGROUND: Vitamin D deficiency is a public health problem worldwide. Vitamin D deficiency in pregnant women often leads to negative clinical consequence and has been distributed differently in certain latitudes. Here, we aimed to determine the prevalence of vitamin D deficiency in pregnant women in Shenzhen City and investigate the influencing factors. METHODS: A total of 27,166 healthy pregnant women, undergoing prenatal examinations in our hospital between July 2014 and December 2018, were enrolled in our study. Maternal characteristics, including the duration of pregnancy, age and enrollment time, were recorded. The concentrations of serum 25(OH)D in the blood samples were detected by immunochemistry assays. RESULTS: For the total study population, the median serum 25(OH)D concentration was 23.36 [17.98-29.51] ng/mL, and 34.3% and 42.4% of the participants exhibited vitamin D deficiency (serum 25(OH) D < 20 ng/mL) and insufficiency (serum 25(OH)D 21-29 ng/mL), respectively. Vitamin D deficiency decreased with gestation (37.83%, 33.8%, and 29.3% for the first trimester, second trimester and third trimester, respectively, p < .001) and decreased by age (36.03%, 35.20%, 31.86% and 29.83%, for the age groups 18-24, 25-29, 30-34 and 35-46 years, respectively, p < .001). This prevalence had conspicuous seasonality (winter vs. autumn, OR 3.69, 95% CI: 3.42-3.99, p < .001). Temperature was positively associated with women's serum 25(OH)D level (r = 0.48, p < .001). CONCLUSIONS: Overall, we demonstrated that vitamin D deficiency in pregnant women in Shenzhen was common and was affected by gestation, age and season/temperature.

The Associations of Genital Mycoplasmas with Female Infertility and Adverse Pregnancy Outcomes: a Systematic Review and Meta-analysis.

The roles of genital mycoplasmas including Mycoplasma genitalium (M. genitalium), Mycoplasma hominis (M. hominis), Ureaplasma urealyticum (U. urealyticum), and Ureaplasma parvum (U. parvum) in reproductive diseases are equivocal. To investigate whether genital mycoplasmas are risk factors of female infertility and adverse pregnancy outcomes, we performed a systematic review and meta-analysis. Electronic databases were searched for related studies. A random-effects model or fixed-effects model was employed to generate forest plots. Pooled odd ratios (ORs) with 95% confidence intervals (CIs) were applied to measure the strength of associations. Meanwhile, heterogeneity was evaluated by H statistic and I2 statistic, and publication bias was explored by funnel plots based on Egger's test and Begg's test. The search yielded 2054 relevant records, and 35 articles were ultimately included for meta-analysis. M. genitalium was a significant risk factor for female infertility (OR, 13.03 [95% CI, 3.46-48.98]) and preterm birth (PTB) (OR, 1.81 [95% CI, 1.17-2.80]), but not for spontaneous abortion (SA) (OR, 0.58 [95% CI, 0.25-1.35]). M. hominis can significantly increase the potential risk of female infertility (OR, 1.56 [95% CI, 1.02-2.38]), SA (OR, 9.14 [95% CI, 4.14-20.18]), stillbirth (OR, 3.98 [95% CI, 1.39-11.36]), and premature rupture of membranes (PROM) (OR, 1.79 [95% CI, 1.26-2.55]), but was not associated with PTB (OR, 1.29 [95% CI, 0.78-2.15]). U. urealyticum had no significant risk effect on female infertility (OR, 0.68 [95% CI, 0.42-1.11]). Coinfections of M. hominis and Ureaplasma were significantly associated with female infertility, SA, and stillbirth, but not with PROM. On the basis of current evidences, this meta-analysis supports that M. genitalium is a risk factor for female infertility and PTB; M. hominis is a potential risk factor for female infertility, SA, stillbirth, and PROM; U. urealyticum has no significant association with female infertility; and the relationship of U. parvum with female infertility and adverse pregnancy outcomes needs to be paid more attention to and remains to be further revealed.

Rapid and accurate diagnosis of Chlamydia trachomatis in the urogenital tract by a dual-gene multiplex qPCR method.

Introduction. Chlamydia trachomatis (C. trachomatis, CT) is an obligatory intracellular bacterium that causes urogenital tract infections and leads to severe reproductive consequences. Therefore, a rapid and accurate detection method with high sensitivity and specificity is an urgent requirement for the routine diagnosis of C. trachomatis infections.Aim. In this study, we aimed to develop a multiplex quantitative real-time PCR (qPCR) assay based on two target regions for accurate detection of C. trachomatis in urogenital tract infections.Methodology. Primers and probes based on the conserved regions of the cryptic plasmid and 23S rRNA gene were designed. Then, two qPCR assays were established to screen for the optimal probe and primers for each of the two target regions. Subsequently, the multiplex qPCR method was developed and optimized. For the diagnostic efficiency evaluation, 1284 urogenital specimens were tested by the newly developed multiplex qPCR method, an immunological assay and a singleplex qPCR assay widely used in hospitals.Results. The multiplex qPCR method could amplify both target regions in the range of 1.0×102-1.0×108 copies ml-1 with a strong linear relationship, and lower limits of detection (LODs) for both targets reached 2 copies PCR-1. For the multiplex qPCR method, the diagnostic sensitivity and specificity was 100.0 % (134/134) and 99.3 % (1142/1150), respectively. For the singleplex qPCR assay, the diagnostic sensitivity and specificity was 88.8 % (119/134) and 100.0 % (1150/1150), respectively. For the immunological assay, the diagnostic sensitivity and specificity was 47.0 % (63/134) and 100.0 % (1150/1150), respectively.Conclusion. In this study, a multiplex qPCR assay with high sensitivity and specificity for rapid (≤2.0 h) and accurate diagnosis of C. trachomatis was developed. The qPCR assay has the potential to be used as a routine diagnostic method in clinical microbiology laboratories.

Ratio-Based Method To Identify True Biomarkers by Normalizing Circulating ncRNA Sequencing and Quantitative PCR Data.

Recent studies have indicated that circulating noncoding RNAs (ncRNAs) such as miRNAs are stable biomarkers for the diagnosis and prognosis of human diseases. However, due to low concentrations of circulating ncRNAs in blood, data normalization in plasma/serum ncRNA experiments using next-generation sequencing and quantitative real time RT-qPCR is a challenge. We found that the current normalization methods based on synthetic external spiked-in controls or published endogenous miRNA controls are inappropriate as they are not stably expressed and therefore fail to reliably detect differentially expressed ncRNAs. Using the alternative of individual ncRNAs as biomarkers, we considered a ratio-based normalization method calculated taking the ratio of any two ncRNAs in the same sample and used the resulting ratios as biomarkers. We mathematically verified the method to be independent of spiked-in and internal controls, and more robust than existing reference control based normalization methods to identify differentially expressed ncRNAs as potential biomarkers for human diseases. Thus, the ratio-based method can solve the difficult normalization problem for circuiting ncRNA data to identify reliable biomarkers to meet real clinical practice.

Plasma small ncRNA pair panels as novel biomarkers for early-stage lung adenocarcinoma screening.

BACKGROUND: Lung cancer is a major cause of cancer-related mortality worldwide, and around two-thirds of patients have metastasis at diagnosis. Thus, detecting lung cancer at an early stage could reduce mortality. Aberrant levels of circulating small non-coding RNAs (small ncRNAs) are potential diagnostic or prognostic markers for lung cancer. We aimed to identify plasma small ncRNA pairs that could be used for early screening and detection of lung adenocarcinoma (LAC). RESULTS: A panel of seven small ncRNA pair ratios could differentiate patients with LAC or benign lung disease from high-risk controls with an area under the curve (AUC) of 100.0%, a sensitivity of 100.0% and a specificity of 100.0% at the training stage (which included 50 patients with early-stage LAC, 35 patients with benign diseases and 29 high-risk controls) and an AUC of 90.2%, a sensitivity of 91.5% and a specificity of 80.4% at the validation stage (which included 44 patients with early-stage LAC, 32 patients with benign diseases and 51 high-risk controls). The same panel could distinguish LAC from high-risk controls with an AUC of 100.0%, a sensitivity of 100.0% and a specificity of 100.0% at the training stage and an AUC of 89.5%, a sensitivity of 85.4% and a specificity of 83.3% at the validation stage. Another panel of five small ncRNA pair ratios (different from the first) was able to differentiate LAC from benign disease with an AUC of 82.0%, a sensitivity of 81.1% and a specificity of 78.1% in the training cohort and an AUC of 74.2%, a sensitivity of 70.4% and a specificity of 72.7% in the validation cohort. CONCLUSIONS: Several small ncRNA pair ratios were identified as markers capable of discerning patients with LAC from those with benign lesions or high-risk control individuals.

Rapid diagnosis of human adenovirus B, C and E in the respiratory tract using multiplex quantitative polymerase chain reaction.

Human adenovirus (HAdV) is increasingly recognized as a major cause of human respiratory tract viral infections. Its outbreaks and epidemics in various populations resulted in considerable morbidity and mortality. Therefore, a rapid and specific assay for HAdV in clinical samples is of crucial importance to diagnosing HAdV infections. The present study aimed to develop and evaluate a multiplex quantitative polymerase chain reaction (qPCR) assay for the rapid detection and accurate quantification of HAdV B, C and E. The lower limit of detection for this assay was two genomic copies per reaction, and quantitative linearity ranged from 2 to 2x106 copies per reaction of the input viral DNA. Furthermore, 3,160 throat swab samples that tested HAdV negative by the immunofluorescence assay were collected and retested using the multiplex qPCR assay. The results showed that 2,906 samples were HAdV negative and the other 254 samples were HAdV positive. The HAdV species identified included B (184 samples), C (51 samples), and E (39 samples). Among the three HAdV species, HAdV B and E were detected from 8 samples, and HAdV C and E were detected from other 12 samples. The overall results demonstrated that the sensitivity and specificity of the proposed assay were 100% (254/254) and 99.6% (2894/2906), respectively. From the perspective of routine clinical diagnosis, this assay represented a rapid (≤1.5 h) and economic strategy, and had the potential to be used for the rapid and accurate diagnosis of human respiratory infections caused by HAdV B, C and E.

Evaluation of Plasma miR-21 and miR-152 as Diagnostic Biomarkers for Common Types of Human Cancers.

Stable blood based miRNA species have allowed for the differentiation of patients with various types of cancer. Therefore, specific blood-based miRNA might be considered as a methodology which could be informative of the presence of cancer potentially from multiple distinct organ sites. Recently, miR-21 has been identified as an "oncomir" in various tumors while miR-152 as a tumor suppressor. In this study, we investigated whether circulating miR-21 and miR-152 can be used for early detection of lung cancer (LuCa), colorectal carcinoma (CRC), breast cancer (BrCa) and prostate cancer (PCa), with distinguishing cancer from various benign lesions on these organ sites. We measured the two miRNA levels by using real-time RT-PCR in plasma samples from a total of 204 cancer patients, 159 various benign lesions, and 228 normal subjects. We observed significantly elevated expression of miR-21 and miR-152 in LuCa, CRC, and BrCa when compared with normal controls. We also found upregulation of plasma miR-21 and miR-152 levels in patients with benign lesions of lung and breast, as compared to normal controls, respectively. No significant expression variation of the two miRNAs was observed in PCa or prostatic benign lesions as compared to healthy controls. Receiver operating characteristic (ROC) analyses revealed that miR-21 and/or miR-152 can discriminate LuCa, CRC and BrCa from normal controls. Our results suggest that plasma miR-21 and miR-152 may serve as non-specific noninvasive biomarkers for early screening of LuCa, CRC, and BrCa, but not PCa.

Diagnostic utility of neutrophil CD64 as a marker for early-onset sepsis in preterm neonates.

INTRODUCTION: Neutrophil CD64 has been proposed as an early marker of sepsis. This study aims to evaluate the diagnostic utility of neutrophil CD64 for identification of early-onset sepsis in preterm neonates. METHODS: The prospective study was conducted in a neonatal intensive care unit between November 2010 and June 2011. Preterm neonates in whom infection was suspected when they were <12 hours of age were enrolled. Complete blood count with differential, blood culture, neutrophil CD11b and CD64 measurement were performed. Receiver operating characteristic curve analysis was performed to evaluate the performance of neutrophil CD64 as biomarker of sepsis. RESULTS: A total of 158 preterm neonates was enrolled, 88 of whom were suspected infection. The suspected sepsis group was of lesser gestational age (P<0.001) and lower birth weight (P<0.001), compared with controls. The hematologic profiles of the suspected sepsis group were characterized by higher white blood cell count, neutrophil counts and C-reactive protein. The suspected sepsis neonates had significantly higher neutrophil CD64 expression compared with controls. Neutrophil CD64 had an area value under the curve of 0.869 with an optimal cutoff values of 1010 phycoerythrin molecules bound/cell and it had a high sensitivity (81.82%) and negative predictive value (77.4%). The level of neutrophil CD64 was independent of antibiotic therapy within 24 hours after the onset of sepsis in preterm neonates. CONCLUSIONS: Neutrophil CD64 is a highly sensitive marker for suspected early-onset sepsis in preterm neonates. Our study suggests that neutrophil CD64 may be incorporated as a valuable marker to diagnose infection.

Phenotypic and molecular characterization of multidrug resistant Klebsiella pneumoniae isolated from a university teaching hospital, China.

The multidrug-resistant rate of Klebsiella pneumoniae has risen rapidly worldwide. To better understand the multidrug resistance situation and molecular characterization of Klebsiella pneumoniae, a total of 153 Klebsiella pneumoniae isolates were collected, and drug susceptibility test was performed to detect its susceptibility patterns to 13 kinds of antibiotics. Phenotypic tests for carbapenemases ESBLs and AmpC enzyme-producing strains were performed to detect the resistance phenotype of the isolates. Then PCR amplification and sequencing analysis were performed for the drug resistance determinants. The results showed that 63 strains harbored bla CTX-M gene, and 14 strains harbored bla DHA gene. Moreover, there were 5 strains carrying bla KPC gene, among which 4 strains carried bla CTX-M, bla DHA and bla KPC genes, and these 4 strains were also resistant to imipenem. Our data indicated that drug-resistant Klebsiella pneumoniae were highly prevalent in the hospital. Thus it is warranted that surveillance of epidemiology of those resistant isolates should be a cause for concern, and appropriate drugs should be chosen.

The role of procalcitonin in the identification of invasive fungal infection-a systemic review and meta-analysis.

We aimed to summarize evidence on the accuracy of procalcitonin (PCT) test in differentiating fungal infection from other causes of infection. We searched electronic database for original researches that reported diagnostic performance of PCT alone or compare with other biomarkers to diagnose invasive fungal infection (IFI). We included 8 qualifying studies studying 474 episodes of suspected fungal infection with 155 (32.7%) probable or proven IFIs. Four studies compared IFI to bacterial sepsis, in which the pooled positive likelihood ratios and negative likelihood ratios were 4.65 (95% confidence interval [CI], 2.46-8.79) and 0.15 (95% CI, 0.05-0.41), respectively. Another 4 studies compared IFI to uninfected individuals, in which the pooled positive likelihood ratios and negative likelihood ratios were 4.01 (95% CI, 2.04-7.88) and 0.23 (0.07-0.77), respectively. The existing literature suggests good diagnostic accuracy for the PCT test for discrimination between IFIs and bacterial infection or noninfectious conditions. Given the high heterogeneity, medical decisions should be based on both PCT test results and clinical findings.

Comparative studies on interactions of baicalein, baicalin and scutellarin with lysozyme.

The interactions of baicalein, baicalin and scutellarin with lysozyme (LYSO) were studied by fluorescence and UV spectroscopy. The results showed that all the three flavones can quench the fluorescence of LYSO via static quenching with the distance between the donor and acceptor less than 7 nm. The hydroxyl at B-ring gave flavones an advantage to binding with LYSO. Electrostatic forces played a major role in stabilizing baicalein-LYSO complex and baicalin-LYSO complex, whereas hydrophobic interactions in scutellarin-LYSO. Furthermore, the presence of pantothenic acid can increase the binding constant and the number of binding sites between flavones and LYSO.