Genetic difference between two Schistosoma japonicum isolates with contrasting cercarial shedding patterns revealed by whole genome sequencing.

Schistosoma japonicum is one of the major infectious agents of human schistosomiasis, mainly endemic in China and the Philippines. We have previously reported the finding of two schistosome isolates, each with a different cercarial emergence pattern adapted to their different hosts. However, there are currently no whole-genome sequencing studies to investigate the underlining genetics of the adaptive traits. We sampled schistosomes in 2013 and 2020 from a hilly area Shitai (ST) and a marshland area Hexian (HX) of Anhui, China. Ten to 15 male or female adult worms from each site/year were sent for whole genome sequencing. Genetics were analyzed, and selection signals along genomes were detected. Gene enrichment analysis was performed for the genome regions under selection. The results revealed considerable genetic differentiation between the two isolates. The genome "windows" affected by natural selection were fewer in ST (64 windows containing 78 genes) than in HX (318 windows containing 276 genes). Twelve significantly enriched genes were identified in ST, but none in HX. These genes were mainly related to specific DNA binding and intercellular signaling transduction. Some functional region changes identified along the genome of the hilly schistosome may be related to its unique late afternoon cercarial emergence.

Title: Différence génétique entre deux isolats de Schistosoma japonicum présentant des schémas contrastés d'émergence de cercaires, révélés par séquençage du génome entier. Abstract: Schistosoma japonicum est l'un des principaux agents infectieux de la schistosomiase humaine, principalement endémique en Chine et aux Philippines. Nous avons précédemment rapporté la découverte de deux isolats de schistosomes, chacun présentant un schéma différent d'émergence de cercaires, adapté à leurs différents hôtes. Cependant, il n'existe actuellement aucune étude de séquençage du génome entier pour comprendre la génétique sous-jacente aux traits adaptatifs. Nous avons échantillonné des schistosomes en 2013 et 2020 dans une zone de collines de Shitai (ST) et une zone marécageuse de Hexian (HX) de l'Anhui, en Chine. Dix à 15 vers adultes mâles ou femelles de chaque site/année ont été envoyés pour séquençage du génome entier. La génétique a été analysée et des signaux de sélection le long des génomes ont été détectés. Une analyse d'enrichissement génétique a été réalisée pour les régions du génome sélectionnées. Les résultats ont révélé une différenciation génétique considérable entre deux isolats. Les « fenêtres ¼ du génome affectées par la sélection naturelle étaient moins nombreuses dans ST (64 fenêtres contenant 78 gènes) que dans HX (318 fenêtres contenant 276 gènes). Douze gènes significativement enrichis ont été identifiés dans ST mais aucun dans HX. Ces gènes étaient principalement liés à la liaison spécifique à l'ADN et à la transduction de la signalisation intercellulaire. Certains changements de régions fonctionnelles identifiés le long du génome du schistosome des collines peuvent être liés à son émergence cercarienne exceptionnelle en fin d'après-midi.

Trends in innovative pediatric drug development in China based on clinical trial registration data.

In China, the focus of drug research and development has gradually shifted from generic to innovative drugs. Using the Chinese Clinical Trials Registry and Information Transparency Platform, we retrospectively analyzed clinical trials of innovative pediatric drugs conducted in mainland China over the last decade. The goal of this work was to better understand the characteristics of and historical changes in innovative pediatric drug research and development (R&D) in China and to provide effective data support for policy makers and other stakeholders. This study included 198 innovative pediatric drug clinical trials. The data showed that, although some progress has been made in the R&D of innovative pediatric drugs in China, many factors limiting this progress still exist, such as concentrated R&D areas, inadequate pediatric participants, and unbalanced source distributions. The level of innovative pediatric drug R&D in China currently lags behind the global level and has not kept pace with anti-neoplastic drug R&D in China. To promote the innovative development of pediatric drugs in China, the Chinese government must develop an R&D supervision framework, improve the motivation and innovation capabilities of pharmaceutical companies, and optimize the source distribution between regions.

0.05% atropine on control of myopia progression in Chinese school children: a randomized 3-year clinical trial.

AIM: To evaluate the effect of 0.05% atropine on the control of myopia for 2y (phase I) and on spherical equivalent refraction (SER) progression for 1y (phase II) after its withdrawal in Chinese myopic children. METHODS: Totally 142 children with myopia were randomly assigned to the 0.05% atropine group or to the placebo group. In phase I, children received 1 treatment for each eye daily. In phase II, the patients received no treatment. Axial length (AL), SER, intraocular pressure (IOP) and atropine-related side effects were assessed at 6 months' intervals. RESULTS: During phase I, the mean change of SER was -0.46±0.30 D in the atropine group, compared to -1.72±1.12 D in the placebo group (P<0.001). The mean change of AL in the atropine group (0.26±0.30 mm) was significantly shorter than that in the placebo group (0.76±0.62 mm, P=0.002). In addition, in phase II (12mo after the withdrawal of atropine), there was no significant difference in AL change from the atropine group, when compared with that from the placebo group (0.31±0.25 mm vs 0.28±0.26 mm, P>0.05). Furthermore, the change in SER from the atropine group was 0.50±0.41 D, which was significantly lower than 0.72±0.60 D from placebo group, (P<0.05). Finally, there were no statistically significant differences in IOP between the treatment and control groups at any stages (all P>0.05). CONCLUSION: The use of 0.05% atropine for two consecutive years may effectively control elongation of AL and thus progression of myopia, without significant SER progression 1y after atropine withdrawal. Therefore, treatment with 0.05% atropine daily for 2y is effective and safe.

Population Genetics of Oncomelania hupensis Snails from New-Emerging Snail Habitats in a Currently Schistosoma japonicum Non-Endemic Area.

Schistosomiasis is still one of the most significant neglected tropical diseases worldwide, and China is endemic for Schistosoma japonicum. With its great achievement in schistosomiasis control, the government of China has set the goal to eliminate the parasitic disease at the country level by 2030. However, one major challenge is the remaining huge areas of habitats for the intermediate host Oncomelania hupensis. This is further exacerbated by an increasing number of new emerging snail habitats reported each year. Therefore, population genetics on snails in such areas will be useful in evaluation of snail control effect and/or dispersal. We then sampled snails from new emerging habitats in Taicang of Jiangsu, China, a currently S. japonicum non-endemic area from 2014 to 2017, and performed population genetic analyses based on nine microsatellites. Results showed that all snail populations had low genetic diversity, and most genetic variations originated from within snail populations. The estimated effective population size for the 2015 population was infinitive. All snails could be separated into two clusters, and further DIYABC analysis revealed that both the 2016 and the 2017 populations may derive from the 2015, indicating that the 2017 population must have been missed in the field survey performed in 2016. These findings may have implications in development of more practical guidelines for snail monitoring and control.

The prevalence of liver abnormalities in humans due to Schistosoma japonicum by ultrasonography in China: a meta-analysis.

BACKGROUND: Schistosoma japonicum was once one of the most severe parasitic diseases in China. After 70 years of national schistosomiasis control programmes, the prevalence and associated morbidity of the infection have been reduced to a much lower level. However, due to the low sensitivity of the current detection approaches, many minor infections in humans could not be identified and ultimately develop chronic injuries with liver abnormalities, a specific 'network' echogenic pattern under ultrasonography. Therefore, as more people take part in physical examinations, we performed this meta-analysis to estimate the overall prevalence of schistosomiasis-associated liver abnormalities in China. METHODS: The publications were searched systematically across five electronic databases. All eligible studies were assessed with quality evaluation forms. Heterogeneity of studies was determined using the I2 and Q tests. A random effects or fixed effects model was employed based on heterogeneity results. The pooled prevalence and its 95% confidence intervals were calculated with the Freeman-Tukey double arcsine transformation. All analyses were conducted using R with the "meta" package. The protocol registration number was CRD42021232982. RESULTS: A total of 19 relevant articles, including 21 studies, were included. The average score of study quality was 6.4 (total score 7), indicating high quality of all included studies. A total of 268, 247 persons were included, and 43, 917 persons were diagnosed with schistosomiasis liver abnormalities by ultrasonography. High degrees of heterogeneity existed among all studies or within subgroups. The overall pooled prevalence was 18.64% (95% CI: 11.88-26.50%). The estimate significantly increased over time and varied among provinces, with the highest in Shanghai and the lowest in Sichuan. The estimate in people aged 60 years or older was significantly higher than that in people of all ages. No significant difference was seen when based on study areas (urban or rural areas) or gender. CONCLUSION: The long-term burden of schistosomiasis in China remains large, as nearly one-fifth of the examined persons were diagnosed with schistosomiasis liver abnormalities. The pooled prevalence was associated with regions or age groups. Such may have a high reference value in the exact calculation of the disease burden and can be helpful for policy makers in prioritizing public health.

Meta-analysis of variable-temperature PCR technique performance for diagnosising Schistosoma japonicum infections in humans in endemic areas.

BACKGROUND: As China is moving onto schistosomiasis elimination/eradication, diagnostic methods with both high sensitivity and specificity for Schistosoma japonicum infections in humans are urgently needed. Microscopic identification of eggs in stool is proven to have poor sensitivity in low endemic regions, and antibody tests are unable to distinguish between current and previous infections. Polymerase chain reaction (PCR) technologies for the detection of parasite DNA have been theoretically assumed to show high diagnostic sensitivity and specificity. However, the reported performance of PCR for detecting S. japonicum infection varied greatly among studies. Therefore, we performed a meta-analysis to evaluate the overall diagnostic performance of variable-temperature PCR technologies, based on stool or blood, for detecting S. japonicum infections in humans from endemic areas. METHODS: We searched literatures in eight electronic databases, published up to 20 January 2021. The heterogeneity and publication bias of included studies were assessed statistically. The risk of bias and applicability of each eligible study were assessed using the Quality Assessment of Diagnostic Accuracy Studies 2 tool (QUADAS-2). The bivariate mixed-effects model was applied to obtain the summary estimates of diagnostic performance. The hierarchical summary receiver operating characteristic (HSROC) curve was applied to visually display the results. Subgroup analyses and multivariate regression were performed to explore the source of heterogeneity. This research was performed following the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines and was registered prospectively in PROSPERO (CRD42021233165). RESULTS: A total of 2791 papers were retrieved. After assessing for duplications and eligilibity a total of thirteen publications were retained for inclusion. These included eligible data from 4268 participants across sixteen studies. High heterogeneity existed among studies, but no publication bias was found. The pooled analyses of PCR data from all included studies resulted in a sensitivity of 0.91 (95% CI: 0.83 to 0.96), specificity of 0.85 (95% CI: 0.65 to 0.94), positive likelihood ratio of 5.90 (95% CI: 2.40 to 14.60), negative likelihood ratio of 0.10 (95% CI: 0.05 to 0.20) and a diagnostics odds ratio of 58 (95% CI: 19 to 179). Case-control studies showed significantly better performances for PCR diagnostics than cross-sectional studies. This was further evidenced by multivariate analyses. The four types of PCR approaches identified (conventional PCR, qPCR, Droplet digital PCR and nested PCR) differed significantly, with nested PCRs showing the best performance. CONCLUSIONS: Variable-temperature PCR has a satisfactory performance for diagnosing S. japonicum infections in humans in endemic areas. More high quality studies on S. japonicum diagnostic techniques, especially in low endemic areas and for the detection of dual-sex and single-sex infections are required. These will likely need to optimise a nested PCR alongside a highly sensitive gene target. They will contribute to successfully monitoring endemic areas as they move towards the WHO 2030 targets, as well as ultimately helping areas to achieve these goals.

Genetic diversity and structure of Oncomelania hupensis hupensis in two eco-epidemiological settings as revealed by the mitochondrial COX1 gene sequences.

BACKGROUND: Oncomelania hupensis hupensis is the only intermediate host of Schistosoma japonicum, the causative agent of schistosomiasis in China and is therefore of significant medical and veterinary health importance. Although tremendous progress has been achieved, there remains an understudied area of approximately 2.06 billion m2 of potential snail habitats. This area could be further increased by annual flooding. Therefore, an understanding of population genetics of snails in these areas may be useful for future monitoring and control activities. METHODS AND RESULTS: We sampled snails from Hexian (HX), Zongyang (ZY) and Shitai (ST) in Anhui (schistosomiasis transmission control), and from Hengtang (HT), Taicang (TC), Dongsan (DS) and Xisan (XS) in Jiangsu (schistosomiasis transmission interrupted), downstream of Anhui. ST, DS and XS are classified as hilly and mountainous areas, and HX, ZY, TC and HT as lake and marshland areas. The mitochondrial cytochrome c oxidase subunit I gene were sequenced. Out of 115 snails analyzed, 29 haplotypes were identified. We observed 56 (8.72%) polymorphic sites consisting of 51 transitions, four transversions and one multiple mutational change. The overall haplotype and nucleotide diversity were 0.899 and 0.01569, respectively. Snail populations in Anhui had higher genetic diversity than in Jiangsu. 73.32% of total variation was distributed among sites and 26.68% within sites. Snails were significantly separated according to eco-epidemiological settings in both network and phylogenetic analyses. CONCLUSION: Our results could provide important guidance towards assessing coevolutionary interactions of snails with S. japonicum, as well as for future molluscan host monitoring and control activities.

[Effect of massage on depressive behavior of chronic stress rats and its mechanism].

Objective: To investigate the effects and mechanisms of massage on the depressive behavior of rats with chronic stress. Methods: The rats were subjected to chronic unpredictable mild stress for 21 days and then treated with massage for 14 days. They were divided into the following groups: blank control group, model group, massage group and fluoxetine group, with 10 rats in each group. The important acupoints of bladder meridian were massaged for 10 minutes every day (with an interval of 2 minutes, 2 times in total). Body weight, open field test, sucrose intake test and water maze test were used to evaluate the behavioral changes. The expressions of Erk/p-ERK (Extracellular signal-related kinases/Phosphorylation extracellular signal-related kinases) and BDNF in prefrontal cortex were detected by Western Blot. Results: The body weight, open field sucrose intake test and water maze data of the model group were significantly lower than those of the Control Group (P＜0.01), and the contents of p-ERK and BDNF protein in prefrontal cortex were decreased significantly (P＜0.01). The body weight, open field test, sucrose intake test and water maze test data of rats in massage group and fluoxetine group were significantly higher than those in model group(P＜0.01). The contents of p-ERK and BDNF in frontal cortex were increased significantly (P＜0.05, P＜0.01), especially in fluoxetine group (P＜0.01). Conclusion: Massage may increase the phosphorylation level of ERK protein in hippocampus and prefrontal cortex, activate ERK signaling pathway, promote the expression of BDNF, and improve the depression behavior of chronic stress rats.

In vivo praziquantel efficacy of Schistosoma japonicum over time: A systematic review and meta-analysis.

Praziquantel (PZQ), the only choice of chemotherapy for schistosomiasis recommended by World Health Organization (WHO), has been widely used over 40 years. The long-term, and rapid expansion of, PZQ use for disease control across a large populations continues to raise concern regarding the potential for emergence and establishment of drug resistance. Recent research has also proposed that the long survival and low sensitivity of unpaired worms, derived from either incomplete treatment cure rates or single-sex schistosome infections within final hosts, could exacerbate the risk of PZQ resistance (PZQ-R) emerging. With the aim of assessing whether PZQ efficacy amongst S. japonicum may have changed over time in China, we performed a unique systematic review and meta-analyses on datasets which evaluated the efficacy of PZQ via laboratory assays of field S. japonicum isolates on experimental mice over time. Relevant published literatures from four electronic bibliographic databases and lists of article references were searched. Two indexes, d, a measure used in meta-analyses for worm burden difference between two groups, and r, a traditional measure for worm reduction percentage after treatment but without considering sample size were calculated for each study. A total of 25 papers including 127 experimental studies with eligible data on 2230 mice were retrieved. The pooled d (D) was 3.91 (3.56-4.25) and pooled r (R) was 54.52% (52.55%-56.52%). D significantly increased over time, whereas R non-significantly decreased; both estimates were significantly associated with the total drug dose. Such findings suggested no evidence of PZQ-R emergence S. japonicum to date. However, we consider the potential role of parasite origins, PZQ dosage, and single versus mixed gender infections of the results published to date, and the avenues now needed for further research.

Extended survival and reproductive potential of single-sex male and female Schistosoma japonicum within definitive hosts.

Schistosomiasis is caused by dioecious helminths of the genus Schistosoma. Recent work indicated that unpaired female and male schistosomes can survive within their definitive host for at least 1 year, although the viability or fertility of these worms after subsequent pairing remained untested. We performed two experiments on laboratory mice, one with female Schistosoma japonicum exposure first and male schistosomes second and another vice versa. After surviving as single-sex unpaired forms for up to 1 year, 58.5% of male and 70% of female schistosomes were able to mate and produce viable eggs. This highlights an additional biological challenge in achieving elimination of schistosomiasis.

The Predictive Value of MAP2K1/2 Mutations on Efficiency of Immunotherapy in Melanoma.

Background: MAP2K1/2 genes are mutated in approximately 8% of melanoma patients; however, the impact of MAP2K1/2 gene alterations on the efficiency of immunotherapy has not been clarified. This study focused on the correlation between MAP2K1/2 gene mutations and the treatment response. Methods: Six metastatic melanoma clinical cohorts treated with immune checkpoint inhibitors [anti-cytotoxic T lymphocyte antigen-4 (CTLA-4) or anti-programmed cell death-1 (PD-1)] were recruited in this study. RNA expression profiling results from each of these six cohorts and the Cancer Genome Atlas (TCGA) melanoma cohort were analysed to explore the mechanism related to immune activation. Results: Compared to patients with wild-type MAP2K1/2, those with MAP2K1/2 mutations in an independent anti-CTLA-4-treated cohort had higher objective response rates, longer progression-free survival, and longer overall survival (OS). These findings were further validated in a pooled anti-CTLA-4-treated cohort in terms of the OS. However, there was no correlation between MAP2K1/2 mutations and OS in the anti-PD-1-treated cohort. Subgroup Cox regression analysis suggested that patients with MAP2K1/2 mutations received fewer benefits from anti-PD-1 monotherapy than from anti-CTLA-4 treatment. Furthermore, transcriptome profiling analysis revealed that melanoma tumours with MAP2K mutation was enriched in CD8+ T cells, B cells, and neutrophil cells, also expressed high levels of CD33 and IL10, implying a potential mechanism underlying the benefit of melanoma patients with MAP2K1/2 mutations from anti-CTLA-4 treatment. Conclusions: MAP2K1/2 mutations were identified as an independent predictive factor for anti-CTLA-4 therapy in melanoma patients. Anti-CTLA-4 treatment might be more effective than anti-PD-1 therapy for patients with MAP2K1/2-mutated melanoma.

Identification of a novel CHN1 p.(Phe213Val) variant in a large Han Chinese family with congenital Duane retraction syndrome.

Duane retraction syndrome (DRS) is a neuromuscular dysfunction of the eyes. Although many causative genes of DRS have been identified in Europe and the United States, few reports have been published in regard to Chinese DRS. The aim of the present study was to explore the genetic defect of DRS in a Chinese family. Exome sequencing was used to identify the disease-causing gene for the two affected family members. Ophthalmic and physical examinations, as well as genetic screenings for variants in chimerin 1 (CHN1), were performed for all family members. Functional analyses of a CHN1 variant in 293T cells included a Rac-GTP activation assay, α2-chimaerin translocation assay, and co-immunoprecipitation assay. Genetic analysis revealed a NM_001822.7: c.637T > G variant in the CHN1 gene, which resulted in the substitution of a highly conserved C1 domain with valine at codon 213 (NP_001813.1: p.(Phe213Val)) (ClinVar Accession Number: SCV001335305). In-silico analysis revealed that the p.(Phe213Val) substitution affected the protein stability and connections among the amino acids of CHN1 in terms of its tertiary protein structure. Functional studies indicated that the p.(Phe213Val) substitution reduced Rac-GTP activity and enhanced membrane translocation in response to phorbol-myristoyl acetate (PMA). Together with previous studies, our present findings demonstrate that CHN1 may be an important causative gene for different ethnicities with DRS.

Can Combined Screening of Ultrasound and Elastography Improve Breast Cancer Identification Compared with MRI in Women with Dense Breasts-a Multicenter Prospective Study.

Objectives: To assess the performance of elastography (ES) and ultrasound (US) in predicting the malignancy of breast lesions and to compare their combined diagnostic value with that of magnetic resonance imaging (MRI). Materials and Methods: The study prospectively enrolled 242 female patients with dense breasts treated in 35 heath care facilities in China between November 2018 and October 2019. Based on conventional US and elastography, radiologists classified the degree of suspicion of breast lesions according to the US Breast Imaging Reporting and Data System (BI-RADS) criteria. The diagnostic value was compared between US BI-RADS and MRI BI-RADS, with pathological results used as the reference standard. Results: The results demonstrated that irregular tumor shape, a nonparallel growth orientation, indistinct margins, angular contours, microcalcifications, color Doppler flow and ES score on US imaging were significantly related to breast cancer in dense breasts (P=0.001; P=0.001; P=0.008; P<0.001; P=0.019; P=0.008; P=0.002, respectively). The sensitivity, specificity, PPV, NPV, accuracy and AUC of US BI-RADS category were 94.7%, 90.7%, 95.8%, 88.0%, 93.4% and 0.93 (95%CI, 0.88-0.97), respectively, while those of MRI BI-RADS category were 98.2%, 57.5%, 84.3%, 83.3%, 86.0% and 0.78 (95%CI, 0.71-0.85), respectively. MRI BI-RADS showed a significantly higher sensitivity than US BI-RADS (98.2% vs 94.7%, P=0.043), whereas US BI-RADS showed significantly higher specificity (90.7% vs 57.5%, P<0.001). US BI-RADS showed better diagnostic efficiency in differentiating nodules in dense breasts than MRI BI-RADS (AUC 0.93 vs 0.78, P<0.001). Conclusion: By combining the use of ES and conventional US, US BI-RADS had better diagnostic efficiency in differentiating nodules in dense breasts than MRI. For the diagnosis of malignant tumors in patients with dense breasts, MRI and US BI-RADS can be used as supplemental diagnostic tools to detect lesions, with US BI-RADS considered the preferred adjunctive resource.

Target Identification of Active Constituents of Shen Qi Wan to Treat Kidney Yang Deficiency Using Computational Target Fishing and Network Pharmacology.

Background: Kidney yang deficiency syndrome (KYDS) is one of the most common syndromes treated with traditional Chinese medicine (TCM) among elderly patients. Shen Qi Wan (SQW) has been effectively used in treating various diseases associated with KYDS for hundreds of years. However, due to the complex composition of SQW, the mechanism of action remains unknown. Purpose: To identify the mechanism of the SQW in the treatment of KYDS and determine the molecular targets of SQW. Methods: The potential targets of active ingredients in SQW were predicted using PharmMapper. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were carried out using the Molecule Annotation System (MAS3.0). The protein-protein interaction (PPI) network of these potential targets and "components-targets-pathways" interaction networks were constructed using Cytoscape. We also established a KYDS rat model induced by adenine to investigate the therapeutic effects of SQW. Body weight, rectal temperature, holding power, water intake, urinary output, blood urea nitrogen (BUN), serum creatinine (Scr), adrenocorticotrophic hormone (ACTH), cortisol (CORT), urine total protein (U-TP), and 17-hydroxy-corticosteroid (17-OHCS) were measured. Additionally, the mRNA expression levels of candidates were detected by qPCR. Results: KYDS-caused changes in body weight, rectal temperature, holding power, water intake, urinary output, BUN, Scr, ACTH, CORT, U-TP, and 17-OHCS were corrected to the baseline values after SQW treatment. We selected the top 10 targets of each component and obtained 79 potential targets, which were mainly enriched in the proteolysis, protein binding, transferase activity, T cell receptor signaling pathway, and focal adhesion. SRC, MAPK14, HRAS, HSP90AA1, F2, LCK, CDK2, and MMP9 were identified as targets of SQW in the treatment of KYDS. The administration of SQW significantly suppressed the expression of SRC, HSP90AA1, LCK, and CDK2 and markedly increased the expression of MAPK14, MMP9, and F2. However, HRAS levels remained unchanged. Conclusion: These findings demonstrated that SQW corrected hypothalamic-pituitary-target gland axis disorder in rats caused by KYDS. SRC, MAPK14, HRAS, HSP90AA1, F2, LCK, CDK2, and MMP9 were determined to the therapeutic target for the further investigation of SQW to ameliorate KYDS.

Intracerebroventricular injection of Aß1-42 combined with two-vessel occlusion accelerate Alzheimer's disease development in rats.

Numerous experimental studies and clinical observations suggest that cerebral ischemia may contribute to the pathogenesis of Alzheimer's disease (AD). Two-vessel occlusion caused cerebral ischemia model is often used in the study of vascular dementia (VaD). But how cerebral ischemia works on AD rat model which induced by intracerebroventricular injection of Aß1-42 remains unclear. In the following study, we investigated the characteristics of rat model caused by intracerebroventricular injection of Aß1-42 or two-vessel occlusion (2-VO) only and by both of the two operations. The animal cognitive functions were accessed by the Morris water maze. Regional cerebral blood flow was detected by Laser Doppler Blood Flowmeter. HE&Nissl staining, Congo red staining and immunohistochemistry were used to observe the status of neuronal loss, Aß deposition and the phosphorylated tau expression in hippocampus, respectively. We also measured the contents of AchE and ChAT in serum and hippocampus by Enzyme Linked Immunosorbent Assay. The MWM results showed that rats of Aß1-42+2-VO group had a disorder in cognitive functions, at an early stage of one week after modeling, comparing with rats of sham group. The regional cerebral blood flow (rCBF) was significantly reduced in Aß1-42+2-VO and 2-VO group one week after modeling, and still maintained low perfusion levels four weeks after modeling. HE and Nissl staining showed that Aß1-42+2-VO rats' hippocampal CA1 neurons were in disorder, degeneration and necrosis, severe neuronal loss from the first week to the fourth week, while this phenomenon only appeared in the fourth week after modeling in rats of Aß1-42 group and 2-VO group. Congo red staining showed that Aß1-42 + 2-VO group rats' hippocampus CA1 had amyloid deposits from the first week to the fourth week, Aß1-42 group were not find amyloid deposition significantly until four weeks after modeling, however, 2-VO group had no significant amyloid deposition all the time. Notably, IHC showed that, two weeks after modeling, the p-tau positive total area and integrated optical density of hippocampal CA1 region were significantly increased in Aß1-42 + 2-VO group rats, while 2-VO group and Aß1-42 group rats had no significantly changes all the time. We also found that the content of AchE was increased both in serum and hippocampus of Aß1-42 + 2-VO group rats, and ChAT was decreased. However, there was no significantly change in cortex of content of AchE: acetylcholinesterase (AchE) and choline acetylase (ChAT) all three groups. Together, our study suggest that intracerebroventricular injection of Aß1-42 combined with two-vessel occlusion may accelerate Alzheimer's disease development in rats. Also, this may serve as a less-time consuming new model to study the Alzheimer's disease and especially AD accompanied by cerebral ischemia.

[Effect of Zhenwu Tang on regulating of "AVP-V2R-AQP2" pathway in NRK-52E cells].

This study was aimed to investigate the effect and mechanism of Zhenwu Tang on AVP-V2R-AQP2 pathway in NRK-52E cells in vitro. Forty eight male SD rats were randomly divided into eight groups with 6 animals in each group. Distilled water or 22.68 g·kg⁻¹·d⁻¹ Zhenwu Tang(calculated by raw drug dosage meter) was given by gavage. Blood samples were collected by cardiac puncture， and the medicated serum was centrifuged from the blood by 3 000 r·min⁻¹. NRK-52E cells were treated with different medicated serum or dDAVP. The condition of cell proliferation was detected by RTCA. The distribution of V2R and AQP2 in cells were detected by immunofluorescence. The expression of V2R， PKA and AQP2 were detected by Western blot and AQP2 mRNA level was detected by real-time PCR. Results showed that the level of AQP2 mRNA(P<0.01) and protein expression of V2R， PKA and AQP2(P<0.05， P<0.01， P<0.05) of Z7d group which was treated with Zhenwu Tang medicated serum for 24 h were significantly higher than that of normal rat serum group. And the expression level of V2R， p-AQP2 and AQP2(P<0.01， P<0.05， P<0.01) of Z7d+dDAVP group were significantly increased comparing to normal rat serum group. The results indicate that the applying of Zhenwu Tang medicated serum could increase the expression level of V2R， PKA and AQP2 which exist in AVP-V2R-AQP2 pathway in NRK-52E， and there is synergistic effect between Zhenwu Tang medicated serum and dDAVP. So the pathway of AVP-V2R-AQP2 may be one of the mechanism for which Zhenwu Tang regulate balance of water transportation.

Analyzing the role of soil and rice cadmium pollution on human renal dysfunction by correlation and path analysis.

The aim of this study was to investigate the role of soil and rice pollution on human renal dysfunction. The participants were 97 inhabitants (46 men and 51 women) who are aged 50 to 60 years old and have been living in Xiaogan (Hubei, China) from birth. We collected samples of soil, rice, and urinary correspondingly. Urinary N-acetyl-ß-D-glucosaminidase (NAG) and ß-2-microglobulin (ß2MG) were used as indicators of renal dysfunction, and urinary cadmium (U-Cd) was used as indicator of total internal cadmium exposure. We made a hypothesis that soil cadmium concentration (S-Cd) and rice cadmium concentration (R-Cd) could be used as indicators of environmental cadmium exposure. Correlation and path analysis were used to estimate the relationships among the levels of rice cadmium (R-Cd), soil cadmium (S-Cd), urinary cadmium (U-Cd), and renal damage indicators (NAG and ß2MG). Our results showed that there was positive significant relationship between S-Cd (R-Cd, U-Cd), and U-NAG (U-ß2MG). The standard multiple regression describing the relationship between S-Cd (R-Cd, U-Cd) and U-NAG was Y1 = 1.26X1-6.53X2 + 9.32, where Y is U-NAG, X1 is U-Cd, X2 is S-Cd. The equation of U-ß2MG was Y = 49.32X1 + 3085.99X2 + 143.42, where Y is U-ß2MG, X1 is U-Cd, X2 is R-Cd. It is obvious that the effect of S-Cd and R-Cd on NAG or U-ß2MG cannot be ignored. Through our study, we can find that the effects of S-Cd on renal health even as significant as R-Cd. To protect people from the damage of cadmium pollution, it is vital to monitor the situation of soil and rice cadmium pollution.

Estimation of the benchmark dose of urinary cadmium as the reference level for renal dysfunction: a large sample study in five cadmium polluted areas in China.

BACKGROUND: Itai-itai disease primarily results from cadmium (Cd) exposure and is known as one of the four major pollution diseases in Japan. Cd pollution is more serious in several areas of China than in Japan. However, there is still a lack of information regarding the threshold level of Cd exposure for the adverse health effects in the general Chinese population. This study aims to evaluate the reference value of urinary Cd (UCd) for renal dysfunction in a Chinese population as the benchmark dose lower confidence limit (BMDL) based on a large sample survey. METHODS: A total of 6103 participants who lived in five Cd polluted areas of China participated in this study. We analyzed UCd levels as a biomarker of exposure and urinary ß2-microglobulin (Uß2-MG) levels as a renal tubular effect biomarker. The BMD studies were performed using BMD software. The benchmark response (BMR) was defined as a 10% additional risk above the background. RESULTS: There was a positive correlation between the UCd levels and the prevalence of Uß2-MG. The BMD of UCd for Uß2-MG was estimated for each province. The findings showed that the BMD levels were related to the participants' geographic region, which may be partially due to the large differences in Cd exposure level, ethnic group, lifestyle and diet of the sample population in these study areas. The reference level of UCd for the renal effects was further evaluated by combining the five sets of data from all 6103 subjects. The overall BMDLs of UCd for Uß2-MG with an excess risk of 10% were 2.00 µg/g creatinine (µg/g cr) in males and 1.69 µg/g cr in females, which were significantly lower than the World Health Organization (WHO) threshold level of 5 µg/g cr for Cd-related renal effects. CONCLUSIONS: The selection of the sample population and geographic region affected the BMDL evaluation. Based on the findings of this survey of a large sample population, the UCd BMDLs for Uß2-MG in males with BMRs at 10% were 2.00 µg/g cr. The BMD was slightly lower in females, which indicated that females may be relatively more sensitive to Cd exposure than males.

Benchmark Dose Estimation for Cadmium-Induced Renal Effects Based on a Large Sample Population from Five Chinese Provinces.

A survey involving 6103 participants from five Chinese provinces was conducted to evaluate the threshold value of urinary cadmium (UCd) for renal dysfunction as benchmark dose low (BMDL). The urinary N-acetyl-ß-D-glucosaminidase (UNAG) was chosen as an effect biomarker. The UCd BMDLs for UNAG ranged from 2.18 µg/g creatinine (cr) to 4.26 µg/g cr in the populations of different provinces. The selection of the sample population and area affect the evaluation of the BMDL. The reference level of UCd for renal effects was further evaluated based on the data of all 6103 subjects. With benchmark responses (BMR) of 10%/5%, the overall UCd BMDLs for males in the total population were 3.73/2.08 µg/g cr. The BMD was slightly lower in females, thereby indicating that females may be relatively more sensitive to Cd exposure than are males.

Body burden of cadmium and its related factors: a large-scale survey in China.

A survey of more than 6000 participants from four distinct non-polluted and polluted regions in China was conducted to evaluate the body burden of cadmium (Cd) on the Chinese populations using urinary Cd (UCd) as a biomarker. The findings revealed that the UCd level was 1.24 µg/g creatinine (µg/g cr) for the sample population from non-polluted Shanghai, and the UCd levels exceeded 5 µg/g cr, which is the health-based exposure limit set by the World Health Organization (WHO), in 1.1% of people. The mean UCd levels in moderately polluted (Hubei and Liaoning) and highly polluted areas (Guizhou) were 4.69 µg/g cr, 3.62 µg/g cr and 6.08 µg/g cr, respectively, and these levels were 2.9 to 4.9 times the levels observed in Shanghai. Notably, the UCd levels exceeded the recently updated human biomonitoring II values (i.e., intervention or "action level") in 44.8%-87.9% of people from these areas compared to only 5.1%-21.4% of people in Shanghai. The corresponding prevalence of elevated UCd levels (>WHO threshold, 5 µg/g cr) was also significantly higher (30.7% to 63.8% vs. 1.1%), which indicates that elevated Cd-induced health risks to residents in these areas. Age and region were significant determinants for UCd levels in a population, whereas gender did not significantly influence UCd.