Testosterone Exacerbates the Formation of Liver Cancer Induced by Environmental N-Nitrosamines Exposure: Potential Mechanisms and Implications for Human Health.

Background: Humans are frequently exposed to N-nitrosamines through various sources, including diet, cigarette smoking, contaminated water, the atmosphere, and endogenous nitrosation. Exposure to these carcinogens may also contribute to the gender-specific incidence of liver cancer, which is significantly higher in males than in females, possibly due to the influence of endogenous hormones such as testosterone. However, the effect of testosterone on N-nitrosamine-induced liver cancer and its underlying mechanism remains unclear. Purpose: To investigate the effect of testosterone on the development of liver cancer induced by N-nitrosamines exposure. Patients and Methods: Histopathological and immunohistochemical staining techniques were employed to analyze the expression levels and nuclear localizations of key signaling molecules, including androgen receptor (AR), ß-catenin, and HMGB1, in both tumor and non-tumor regions of liver samples obtained from human patients and mice. Results: The findings demonstrated a strong correlation between AR and ß-catenin in the nuclear region of tumor areas. AR also showed a significant correlation with HMGB1 in the cytoplasmic region of non-tumor areas in both human and mice samples. The study further analyzed the expression levels and patterns of these three proteins during the progression of liver tumors. Conclusion: This study confirms that AR has the ability to modulate the expression levels and patterns of ß-catenin and HMGB1 in vivo, thereby exacerbating the progression of liver cancer induced by environmental N-nitrosamines exposure. Importantly, the effect of testosterone on the formation of liver cancer induced by environmental N-nitrosamine exposure intensifies this progression. These findings have important implications for drug safety in clinical practice and emphasize the significance of reducing N-nitrosamines exposure through conscious choices regarding diet and lifestyle to ensure environmental safety.

ATF3-CBS signaling axis coordinates ferroptosis and tumorigenesis in colorectal cancer.

The induction of ferroptosis is promising for cancer therapy. However, the mechanisms enabling cancer cells to evade ferroptosis, particularly in low-cystine environments, remain elusive. Our study delves into the intricate regulatory mechanisms of Activating transcription factor 3 (ATF3) on Cystathionine ß-synthase (CBS) under cystine deprivation stress, conferring resistance to ferroptosis in colorectal cancer (CRC) cells. Additionally, our findings establish a positively correlation between this signaling axis and CRC progression, suggesting its potential as a therapeutic target. Mechanistically, ATF3 positively regulates CBS to resist ferroptosis under cystine deprivation stress. In contrast, the suppression of CBS sensitizes CRC cells to ferroptosis through targeting the mitochondrial tricarboxylic acid (TCA) cycle. Notably, our study highlights that the ATF3-CBS signaling axis enhances ferroptosis-based CRC cancer therapy. Collectively, the findings reveal that the ATF3-CBS signaling axis is the primary feedback pathway in ferroptosis, and blocking this axis could be a potential therapeutic approach for colorectal cancer.

Improving Classification Performance in Dendritic Neuron Models through Practical Initialization Strategies.

A dendritic neuron model (DNM) is a deep neural network model with a unique dendritic tree structure and activation function. Effective initialization of its model parameters is crucial for its learning performance. This work proposes a novel initialization method specifically designed to improve the performance of DNM in classifying high-dimensional data, notable for its simplicity, speed, and straightforward implementation. Extensive experiments on benchmark datasets show that the proposed method outperforms traditional and recent initialization methods, particularly in datasets consisting of high-dimensional data. In addition, valuable insights into the behavior of DNM during training and the impact of initialization on its learning performance are provided. This research contributes to the understanding of the initialization problem in deep learning and provides insights into the development of more effective initialization methods for other types of neural network models. The proposed initialization method can serve as a reference for future research on initialization techniques in deep learning.

Mobile phone addiction and academic burnout: the mediating role of technology conflict and the protective role of mindfulness.

With the rapid development of Internet technology, more and more college students are facing the threat of mobile phone addiction. However, the relationship and underlying mechanism between mobile phone addiction and academic burnout haven't been explored in depth. This study proves the mediating role of technology conflict and the moderating role of mindfulness in the relation between mobile phone addiction and academic burnout. 752 college students were recruited to complete the questionnaire of mobile phone addiction, technology conflict, mindfulness and academic burnout. Results showed that mobile phone addiction was significantly and positively associated with academic burnout, and this relationship could be mediated by technology conflict. Besides, the direct effect of mobile phone addiction on academic burnout and the indirect effect of technology conflict in this link were moderated by mindfulness. Both these two effects are stronger for college students with lower level of mindfulness. Our findings enrich our understanding of how and when mobile phone addiction was related to academic burnout. Educational professionals and parents should take timely measure to the academic burnout of college students suffering from mobile phone addiction, particularly for those with lower level of mindfulness.

Monocyte subtype expression patterns in septic patients with diabetes are distinct from patterns observed in obese patients.

Background: Sepsis causes a high rate of mortality and long-term morbidity, associated with an imbalance of innate immunity against infections and inflammation. Obesity and diabetes increase the risk for disease severity. Monocyte dysfunction plays a major role and justify further investigations. Objective: To investigate the distribution and inflammatory phenotypes in circulating monocyte subsets in patients manifesting with sepsis including septic shock with and without obesity and diabetes. Methods: A total of 235 blood samples were tested from critically ill adult patients registered at the intensive care unit (ICU). The cohorts were divided into non-diabetic groups with or without obesity and diabetic groups with or without obesity, suffering from sepsis or septic shock. We determined frequencies of total monocytes and of monocyte subsets in the circulation and density expression levels of functional markers, including CD14, CD16, HLA-DR, CD33, CD163, CD206, and arginase-1 by flow cytometric analysis. Results: When progressing to septic shock in non-diabetic and diabetic patients, the percentages of total monocytes among the leukocyte population and of CD33+ and CD14+ monocytes among the monocyte population were consistently down-regulated compared to non-sepsis in non-diabetic and diabetic patients, respectively. Non-diabetic sepsis patients further presented with decreased CD33 and up-regulated CD163 expression density, which was absent in diabetic patients. We subsequently addressed obesity-related changes of monocytes in non-diabetic and diabetic septic patients. Obese septic patients with diabetes were unique in displaying increased monocytic CD16 and CD163 expression. However, obese septic patients without diabetes solely presented with lower amounts of non-classical monocytes. Body mass index (BMI) dependent changes were restricted to diabetic septic patients, with a significantly higher diminution of the classical monocyte subset and concomitantly increased CD16 expression densities. Conclusion: Distribution and phenotypes of monocyte subsets were differentially modulated in critically ill patients with and without metabolic disease when progressing to sepsis or septic shock. Only diabetic septic patients displayed decline of classical monocytes and increase of CD16 expression densities. Therefore, diabetes but not obesity appears to promote the inflammatory phenotype of circulating monocytes in critically ill patients.

Associations of maternal ambient temperature exposures during pregnancy with the placental weight, volume and PFR: A birth cohort study in Guangzhou, China.

BACKGROUND: The placenta performs crucial functions to ensure normal fetal development. Experimental studies have indicated associations between exposure to elevated temperatures during pregnancy and reduction in placental weight and volume. However, epidemiological studies in humans are lacking. OBJECTIVE: To estimate the associations between prenatal exposure to ambient temperature with placental weight, volume, and the placental weight to birth weight ratio (PFR). METHODS: We conducted a prospective birth cohort study using the Prenatal Environment and Offspring Health Cohort (PEOH Cohort) beginning in 2016 in Guangzhou, China. Women in early pregnancy were recruited and followed up during their hospitalization for childbirth. An inverse distance-weighted method was employed to estimate the average temperature exposure of every 4 weeks as well as the trimester-specific average temperature exposure at the individual's residential address. A generalized linear model was applied to estimate the effects of temperature exposure during pregnancy on the placental weight, volume, and PFR. RESULTS: A total of 4051 pregnant women were enrolled. Compared with the reference temperature of 20 °C, maternal exposure to 29 °C (95th centile) during late pregnancy was associated with an average of -6.03 g (95% confidence interval [CI]: -11.28 g, -0.78 g) in placental weight, -16.15 cm3 (95% CI: -26.24 cm3, -6.07 cm3) in placental volume, and 0.26 (95% CI: 0.07, 0.45) in PFR. The peak effects of high temperatures on placental weight, volume, and PFR were found from 29 to 32 weeks (ß = -3.79 g, 95% CI: -8.39 g, 0.82 g), 37 to 40 weeks (ß = -19.34 cm3, 95% CI: -30.99 cm3, -7.69 cm3), and 25 to 28 weeks (ß = 0.35, 95% CI: 0.04, 0.66), respectively. CONCLUSIONS: Maternal exposure to elevated temperatures was associated with a decrease in placental weight and volume and an increase in PFR. The associations were stronger when exposures occurred during late pregnancy.

Associations of maternal ozone exposures during pregnancy with maternal blood pressure and risk of hypertensive disorders of pregnancy: A birth cohort study in Guangzhou, China.

Although studies have assessed the associations of maternal exposure to ozone (O3) during pregnancy with blood pressure and the risk of hypertensive disorders of pregnancy (HDP), the results were inconsistent. Furthermore, no studies have been conducted in China where the ambient O3 concentration continuedly increased. The present study aimed to estimate the effects of maternal exposure to O3 during pregnancy on the HDP risk, systolic blood pressure (SBP), diastolic blood pressure (DBP) and pulse pressure (PP). All participants of pregnant women were selected from the prospective birth cohort study on Prenatal Environments and Offspring Health conducted in Guangzhou, China. A spatiotemporal land-use-regression model was used to estimate individual monthly air pollution exposure from three months before pregnancy to childbirth date. Information on HDP, SBP, DBP and PP was obtained from maternal medical records. A Logistic regression model and a mixed linear model were used to estimate the associations of maternal exposure to O3 with the risk of HDP and blood pressure (SBP, DBP and PP), respectively. We found significant associations of maternal exposure to O3 during the third (OR = 1.31, 95%CI: 1.07, 1.60) and the second month (OR = 1.25, 95%CI: 1.02, 1.51) before pregnancy with the risk of HDP. Observed significantly positive associations of O3 exposures with SBP, DBP and PP during the two months before pregnancy and during the early pregnancy. The peak effects of O3 exposure on SBP, DBP and PP were respectively observed during the second month of pregnancy (ß = 1.07  mmHg, 95%CI: 0.84, 1.31  mmHg), the first month before pregnancy (ß = 0.40  mmHg, 95%CI: 0.21, 0.50  mmHg) and the second month of pregnancy (ß = 0.78  mmHg, 95%CI: 0.59, 0.97  mmHg). Our results suggest that maternal exposure to O3 were positively associated with blood pressure and the risk of HDP, and the period from three months before pregnancy to the first trimester might be the critical exposure window.

Association between maternal antimony exposure and risk of gestational diabetes mellitus: A birth cohort study.

BACKGROUND: A small number of epidemiological studies have suggested the association of antimony (Sb) exposure with type 2 diabetes risk. However, little is known about the relationship between Sb exposure during pregnancy and risk of gestational diabetes mellitus (GDM). OBJECTIVES: To investigate the associations of urinary Sb concentrations with GDM risk and blood glucose levels in pregnant women. METHODS: We analyzed the baseline data of 1789 pregnant women enrolled in the Birth Cohort Study on Prenatal Environments and Offspring Health (PEOH) in Guangzhou, China. Sb concentrations in urine were measured by inductively coupled plasma mass spectrometry (ICP-MS). Logistic regression and analysis of covariance were used to evaluate associations of Sb exposure with GDM risk and blood glucose levels. RESULTS: A total of 437 (24.4%) women were diagnosed with GDM. The relative risk of GDM for women in the highest quartile of creatinine-corrected Sb (CC-Sb) concentrations was 1.55 [RR (95% CI) = 1.55 (1.12, 2.15), p-trend = 0.005], compared with women in the lowest quartile. Moreover, the women in the top quartile of CC-Sb levels had a 5.2% higher 1 h blood glucose and a 4.2% higher 2 h blood glucose than those in the bottom quartile. We also found an interactive effect between maternal age and CC-Sb on the risk of GDM (p-interaction < 0.001). CONCLUSION: This study suggested significant positive associations of Sb exposure with increased GDM risk and impaired blood glucose homeostasis in pregnant women, and the Sb-GDM association might be modified by maternal age.

Effects of prenatal exposure to air particulate matter on the risk of preterm birth and roles of maternal and cord blood LINE-1 methylation: A birth cohort study in Guangzhou, China.

BACKGROUND: Epidemiological studies have found that increased risk of preterm birth (PTB) is associated with higher prenatal exposure to PM10 and PM2.5, but few studies have been conducted to assess the impacts of extremely fine particulate matter (PM1) which may have more toxic effects than other types of ambient particulate air pollution (PM). Several studies have separately investigated the associations between DNA methylation and PTB risk and PM. Maternal LINE-1 methylation level negatively correlated with prenatal exposure to PM and risk of PTB. A comprehensive picture is lacking regarding the associations between prenatal exposure to PM, LINE-1 methylation, and risk of PTB. OBJECTIVES: This study aimed to estimate the effects of exposure to ambient PM (PM10, PM2.5, and PM1) of different sizes during pregnancy on risk of PTB, identify susceptible exposure windows, and illustrate the roles of LINE-1 methylation in the associations between PM and PTB risk. METHODS: The Birth Cohort Study on Prenatal Environments and Offspring Health (PEOH) has been ongoing since 2016 in Guangzhou, China. A total of 4928 pregnant women were recruited during early pregnancy, and 4278 (86.8%) were successfully followed-up. Each individual weekly exposure to PM10 and PM2.5 from 3 months before pregnancy to childbirth was assessed using a spatiotemporal land use regression model, and the weekly PM1 exposure was estimated by employing a generalized additive model. Maternal and cord blood LINE-1 methylation levels (%5mC) were tested using bisulfite-PCR pyrosequencing. A distributed lag nonlinear model incorporated with a Cox proportional hazard model was applied to assess the effect of weekly-specific maternal PM exposure on PTB risk, and a multiple-linear regression model was employed to investigate the associations between PM exposure and LINE-1 methylation levels of maternal and cord bloods. We also assessed the associations between LINE-1 methylation levels and PTB risk by using a logistic regression model. RESULTS: The risk of PTB was positively associated with PM2.5 and PM1 concentrations during the 12th to 20th gestational weeks, and the strongest association was in the fourth quartile (Q4) versus the first quartile (Q1) and observed during the 16th gestational week (PM2.5: harzard ratio [HR] = 1.18, 95%CI: 1.04-1.35, IQR = 11.94 µg/m3. PM1: HR = 1.20, 95%CI: 1.03-1.39, IQR = 11.36 µg/m3). We observed significantly negative associations of PM10(ß = -0.51%5mC per 10 µg/m3, P = 0.014), PM2.5 (ß = -0.66%5mC per 10 µg/m3, P = 0.032) and PM1 (ß = -0.67%5mC per 10 µg/m3, P = 0.032) concentrations with cord blood LINE-1 methylation levels, and a negative association between PM1 concentration and maternal LINE-1 methylation level (ß = -0.86%5mC per 10 µg/m3, P = 0.034). CONCLUSION: Higher prenatal exposure to PM1 and PM2.5 during the 12th to 20th gestational weeks was associated with increased risk of PTB. Maternal and fetal LINE-1 methylation alternation might be an underlying mechanism of PM that increasing the risk of PTB.

The Involvement of ß-Catenin/COX-2/VEGF Axis in NMDA-Caused Retinopathy.

NMDA, a molecule that is capable of producing the loss of retinal ganglia cells (RGCs), has been widely studied; however, the detailed mechanism is not yet clarified. Previously, Wnt/ß-catenin signaling has been suggested to be involved in the NMDA-induced retinopathy. In addition, previous investigations in our group demonstrated the presence of a Wnt/ß-catenin/COX-2 axis in dorsal root ganglions (DRGs). Therefore, here in this paper, we tested whether there is an association of such axis with NMDA-induced RGC loss. Rat retinal damage models generated by intravitreal injection of NMDA were used to measure the expression levels of ß-catenin, COX-2, and VEGF in retinas, and the neuron numbers of the retinal GCL of rats were counted. Then, pharmacological tools (MK801, a NMDA receptor inhibitor; Dickkopf homolog 1, a specific inhibitor of the Wnt pathway; NS-398, a COX-2 inhibitor; and bevacizumab, IVB, a VEGF inhibitor) were introduced to evaluate the detailed roles of Wnt/ß-catenin, COX-2, and VEGF in retinopathy of rats. Results demonstrated that all three factors in sequence are positively regulated neuronal loss induced by NMDA. These observations indicated that the Wnt pathway/COX-2/VEGF axis plays a pathogenic role in retinopathy and represented novel therapeutic targets.

Returning ex-patriot Chinese to Guangdong, China, increase the risk for local transmission of Zika virus.

OBJECTIVES: Fast expansion and linkage to microcephaly and Guillain Barre syndrome have made Zika virus (ZIKV) track attention of global health authority concerns. The epidemiology, virological characteristics and genetic evolution of introduced ZIKV to Guangdong, China, were investigated. METHODS: Analyses of the epidemiological characteristics and genetic diversity of ZIKV isolates were performed. RESULTS: A total of twenty-eight confirmed ZIKV infection cases were imported into China in 2016, of which 19 were imported into Guangdong, China from Venezuela (16), the Samoa Islands (1), Suriname (1) and Guatemala (1). Serial sampling studies of the cases indicated longer shedding times of ZIKV particles from urine and saliva samples than from serum and conjunctiva swab samples. Seven ZIKV strains were successfully isolated from serum, urine and conjunctiva swab samples using cell culture and neonatal mouse injection methods. Genomic analysis indicated that all viruses belonged to the Asian lineage but had different evolutionary transmission routes with different geographic origins. The molecular clock phylogenetic analysis of the ZIKV genomes indicated independent local transmission that appeared to have been previously established in Venezuela and Samoa. Additionally, we found 7 unique non-synonymous mutations in the genomes of ZIKV that were imported to China. The mutations may indicate that ZIKV has undergone independent evolutionary history not caused by sudden adaptation to Chinese hosts. CONCLUSION: The increasing number of ex-patriot Chinese returning from ZIKV hyper-endemic areas to Guangdong combined with the presence of a variety of Aedes species indicate the potential for autochthonous transmission of ZIKV in Guangdong.

Evaluation of a real-time impedance analysis platform on fungal infection.

End-point assays of in vitro cell proliferation and death have been employed to study the mechanisms of fungal pathogenesis and have shown the responses of host cells at individual time points. A new cell analysis technology has been developed that allows for the continuous measurement and quantification of cell activities, thus enabling the dynamic assessment of electrical impedance when various pathogens are cultured in vitro. In this study, this system was evaluated to determine the response of the cell line RAW264.7 to infection by several clinically relevant fungi in vitro, including Aspergillus fumigatus, Candida albicans, and melanized and albino mutant strains of Fonsecaea monophora. The results showed that infection resulted in rounding of the host cells with a loss of contact between individual cells and a decline in the electrical impedance of all test groups. However, changes in the electrical impedance were variable. Aspergillus fumigatus caused initial increases and later significant decreases in the electrical impedance, while for C. albicans and F. monophora, the effect was reduced. The melanized strain of F. monophora caused a faster change in the electrical impedance than the albino strain. Our data proved that this system can be used as an efficient tool for monitoring cellular responses to fungal infection.

Inapparent Streptococcus agalactiae infection in adult/commercial tilapia.

We report on inapparent infections in adult/commercial tilapia in major tilapia fish farms in Guangdong. A total of 146 suspected isolates were confirmed to be S. agalactiae using an API 20 Strep system and specific PCR amplification. All isolates were identified as serotype Ia using multiplex serotyping PCR. An MLST assay showed single alleles of adhP (10), atr (2), glcK (2), glnA (1), pheS (1), sdhA (3) and tkt (2), and this profile was designated 'unique ST 7'. The analysis of virulence genes resulted in 10 clusters, of which dltr-bca-sodA-spb1-cfb-bac (62, 42.47%) was the predominant virulence gene profile. The PFGE analysis of S. agalactiae yielded 6 distinct PFGE types (A, B, C, D, F and G), of which Pattern C (103) was the predominant type, accounting for approximately 70.55% (103/146) of the total S. agalactiae strains. Therefore, unlike what has been found in juvenile tilapia, in which PFGE pattern D/F is the major prevalent pattern, we found that pattern C was the major prevalent pattern in inapparent infected adult/commercial tilapia in Guangdong, China. In conclusion, we close a gap in the current understanding of S. agalactiae epidemiology and propose that researchers should be alert for inapparent S. agalactiae infections in adult/commercial tilapia to prevent a potential threat to food safety.

Identification and characterization of Clonorchis sinensis cathepsin B proteases in the pathogenesis of clonorchiasis.

BACKGROUND: Human clonorchiasis is a prevailing food-borne disease caused by Clonorchis sinensis infection. Functional characterizations of key molecules from C. sinensis could facilitate the intervention of C. sinensis associated diseases. METHODS: In this study, immunolocalization of C. sinensis cathepsin B proteases (CsCBs) in C. sinensis worms was investigated. Four CsCBs were expressed in Pichia pastoris yeast cells. Purified yCsCBs were measured for enzymatic and hydrolase activities in the presence of various host proteins. Cell proliferation, wound-healing and transwell assays were performed to show the effect of CsCBs on human cells. RESULTS: CsCBs were localized in the excretory vesicle, oral sucker and intestinal tract of C. sinensis. Recombinant yCsCBs from yeast showed active enzymatic activity at pH 5.0-5.5 and at 37-42 °C. yCsCBs can degrade various host proteins including human serum albumin, human fibronectin, human hemoglobin and human IgG. CsCBs were detected in liver tissues of mice and cancer patients afflicted with clonorchiasis. Various bioassays collectively demonstrated that CsCBs could promote cell proliferation, migration and invasion of human cancer cells. CONCLUSION: Our results demonstrated that CsCBs can degrade various human proteins and we proved that the secreted CsCBs are involved in the pathogenesis of clonorchiasis.

Oral delivery of Bacillus subtilis spore expressing enolase of Clonorchis sinensis in rat model: induce systemic and local mucosal immune responses and has no side effect on liver function.

Caused by the consumption of raw or undercooked freshwater fish containing infective metacercariae of Clonorchis sinensis, human clonorchiasis remains a major public health problem in China. In previous study, we had expressed enolase from C. sinensis (CsENO) on the surface of Bacillus subtilis spore and the recombinant spore induced a pronounced protection in terms of reduced worm burden and eggs per gram feces, suggesting B. subtilis spore as an ideal vehicle for antigen delivery by oral treatment and CsENO as a promising vaccine candidate against clonorchiasis. In the current study, we detected CsENO-specific IgG and IgA levels both in serum and in intestinal mucus from rats orally administrated with B. subtilis spore surface expressing CsENO by ELISA. Lysozyme levels in serum and in intestinal mucus were analyzed too. In addition, IgA-secreting cells in intestine epithelium of the rats were detected by immunohistochemistry assay. The intestinal villi lengths of duodenum, jejunum, and ileum were also measured. Rats orally treated with B. subtilis spore or normal saline were used as controls. Our results showed that, compared with the control groups, oral administration of B. subtilis spore expressing CsENO induced both systemic and local mucosal immune response. The recombinant spores also enhanced non-specific immune response in rats. The spores had no side effect on liver function. Moreover, it might facilitate food utilization and digestion of the rats. Our work will pave the way to clarify the involved mechanisms of protective efficacy elicited by B. subtilis spore expressing CsENO and encourage us to carry out more assessment trails of the oral treated spore to develop vaccine against clonorchiasis.

Effects of thyroxine and donepezil on hippocampal acetylcholine content, acetylcholinesterase activity, synaptotagmin-1 and SNAP-25 expression in hypothyroid adult rats.

A growing number of studies have revealed that neurocognitive impairment, induced by adult-onset hypothyroidism, may not be fully restored by traditional hormone substitution therapies, including thyroxine (T4). The present study has investigated the effect of T4 and donepezil (DON; an acetylcholinesterase (AChE) inhibitor) treatment on the hypothyroidism-induced alterations of acetylcholine (ACh) content and AChE activity. Furthermore, we examined synaptotagmin-1 (syt-1) and SNAP-25 expression in the hippocampus of adult rats. Adding 0.05% propylthiouracil to their drinking water for five weeks induced hypothyroidism in the rat models. From the fourth week, the rats were treated with T4, DON or a combination of both. Concentration of ACh and the activity of AChE was determined colorimetrically. The results demonstrated that hypothyroidism induced a significant decrease of Ach content and AChE activity (by 17 and 34%, respectively), which were restored to control values by T4 administration. DON treatment also restored Ach to the normal level. Protein levels of syt-1 and SNAP-25 were determined by immunohistochemistry. The results demonstrated that syt-1 was expressed at significantly lower levels in hypothyroid rats, while SNAP-25 levels were notably higher compared with the controls. Two-week treatment with T4 alone failed to normalize the expression levels of these two proteins, while co-administration of T4 and DON was able to induce this effect. These data suggested that the thyroid hormone, T4, may have a direct effect on the metabolism of hippocampal ACh in adult rats, and that the DON treatment may facilitate the recovery of synaptic protein impairments induced by hypothyroidism.

Sequence analysis and molecular characterization of Clonorchis sinensis hexokinase, an unusual trimeric 50-kDa glucose-6-phosphate-sensitive allosteric enzyme.

Clonorchiasis, which is induced by the infection of Clonorchis sinensis (C. sinensis), is highly associated with cholangiocarcinoma. Because the available examination, treatment and interrupting transmission provide limited opportunities to prevent infection, it is urgent to develop integrated strategies to prevent and control clonorchiasis. Glycolytic enzymes are crucial molecules for trematode survival and have been targeted for drug development. Hexokinase of C. sinensis (CsHK), the first key regulatory enzyme of the glycolytic pathway, was characterized in this study. The calculated molecular mass (Mr) of CsHK was 50.0 kDa. The obtained recombinant CsHK (rCsHK) was a homotrimer with an Mr of approximately 164 kDa, as determined using native PAGE and gel filtration. The highest activity was obtained with 50 mM glycine-NaOH at pH 10 and 100 mM Tris-HCl at pH 8.5 and 10. The kinetics of rCsHK has a moderate thermal stability. Compared to that of the corresponding negative control, the enzymatic activity was significantly inhibited by praziquantel (PZQ) and anti-rCsHK serum. rCsHK was homotropically and allosterically activated by its substrates, including glucose, mannose, fructose, and ATP. ADP exhibited mixed allosteric effect on rCsHK with respect to ATP, while inorganic pyrophosphate (PPi) displayed net allosteric activation with various allosteric systems. Fructose behaved as a dose-dependent V activator with the substrate glucose. Glucose-6-phosphate (G6P) displayed net allosteric inhibition on rCsHK with respect to ATP or glucose with various allosteric systems in a dose-independent manner. There were differences in both mRNA and protein levels of CsHK among the life stages of adult worm, metacercaria, excysted metacercaria and egg of C. sinensis, suggesting different energy requirements during different development stages. Our study furthers the understanding of the biological functions of CsHK and supports the need to screen for small molecule inhibitors of CsHK to interfere with glycolysis in C. sinensis.

The serological diagnosis of human clonorchiasis by time-resolved fluoroimmunoassay based on GST2-specific IgG4 detection.

Due to its delayed fluorescence of a lanthanide chelate, high accuracy and low background the broad linear range, long fluorescent life-time and large Stoke's shift of europium chelates, the time-resolved fluorescence has been developed for higher sensitive immunoassay. In this article, a simple, sensitive and specific method-time-resolved fluoroimmunoassay (TRFIA) was adopted for immunoassay of clonorchiasis, and recombinant glutathione transferases 2 of Clonorchis sinensis (rCsGST2) was used as a diagnostic antigen. To evaluate this novel assay for clinical applications, 409 serum samples were investigated. The diagnostic accuracy of the antigen was evaluated by receiver-operating characteristic (ROC) analysis. The area under the ROC curve (AUC) was 0.965, 95% confidence interval (CI, 0.946, 0.985). To eliminate the random influence of ambient temperature, test parameters, photometric instruments and so on, the cut-off value was expressed as ratios between the fluorescence of sample and that of a well-defined negative control serum, and the deduced cut-off value was 9.3605. At the optimum cut-off criteria, the technique has a sensitivity of 95.80%, specificity of 93.60%. And the cross reactivity revealed that its cross reactivity with Schistosoma japonicum, round worm, hook worm, whip worm, and Toxoplasma gondii was 9.3, 8.3, 7.6, 9.8, and 5.0%, respectively. Kappa score of agreement between TRFIA and microscopic examination of stools was 0.892, P < 0.05. These combined results showed that our method is feasible and could be used for the clinical determination of clonorchiasis.