Susceptibility gene identification and risk evaluation model construction by transcriptome-wide association analysis for salt sensitivity of blood pressure.

BACKGROUND: Salt sensitivity of blood pressure (SSBP) is an intermediate phenotype of hypertension and is a predictor of long-term cardiovascular events and death. However, the genetic structures of SSBP are uncertain, and it is difficult to precisely diagnose SSBP in population. So, we aimed to identify genes related to susceptibility to the SSBP, construct a risk evaluation model, and explore the potential functions of these genes. METHODS AND RESULTS: A genome-wide association study of the systemic epidemiology of salt sensitivity (EpiSS) cohort was performed to obtain summary statistics for SSBP. Then, we conducted a transcriptome-wide association study (TWAS) of 12 tissues using FUSION software to predict the genes associated with SSBP and verified the genes with an mRNA microarray. The potential roles of the genes were explored. Risk evaluation models of SSBP were constructed based on the serial P value thresholds of polygenetic risk scores (PRSs), polygenic transcriptome risk scores (PTRSs) and their combinations of the identified genes and genetic variants from the TWAS. The TWAS revealed that 2605 genes were significantly associated with SSBP. Among these genes, 69 were differentially expressed according to the microarray analysis. The functional analysis showed that the genes identified in the TWAS were enriched in metabolic process pathways. The PRSs were correlated with PTRSs in the heart atrial appendage, adrenal gland, EBV-transformed lymphocytes, pituitary, artery coronary, artery tibial and whole blood. Multiple logistic regression models revealed that a PRS of P < 0.05 had the best predictive ability compared with other PRSs and PTRSs. The combinations of PRSs and PTRSs did not significantly increase the prediction accuracy of SSBP in the training and validation datasets. CONCLUSIONS: Several known and novel susceptibility genes for SSBP were identified via multitissue TWAS analysis. The risk evaluation model constructed with the PRS of susceptibility genes showed better diagnostic performance than the transcript levels, which could be applied to screen for SSBP high-risk individuals.

Dietary Selenium Insufficiency Induces Cardiac Inflammatory Injury in Chicks.

BACKGROUND: Laying hens undergo intensive metabolism and are vulnerable to cardiac insults. Previous research demonstrated overt heart disorders of broiler chickens induced by dietary Se deficiency. OBJECTIVES: This study aimed to reveal effects and mechanism of dietary Se insufficiency on cardiac injuries of egg-type chicks in their early life. METHODS: White Leghorn chicks (0-d-old, female) were fed a corn-soy, Se-insufficient basal diet (BD, 0.05 mg Se/kg; n = 11) or the BD supplemented with 0.3 mg Se/kg (as sodium selenite; n = 8) for 35 d. Cardiac tissues were collected at the end of study for histology and to determine its relationship with heart Se contents, selenoprotein expression profiles, antioxidant and inflammatory status, and the Toll-like receptor 4/extracellular signal-regulated kinases/p38 map kinase/c-Jun N-terminal kinase (TLR4/ERK/P38/JNK) pathway. RESULTS: Compared with those fed 0.35 mg Se/kg, chicks fed BD had significantly lower body weights and average daily gain, and 28% lower heart Se, and developed cardiac mononuclear inflammatory cell infiltration, along with elevated (P < 0.05) serum concentrations of creatine kinase, aldolase, and interleukin-1 (IL-1). The BD decreased (P < 0.05) body weight and heart glutathione contents and expression of selenoproteins but increased (P < 0.05) heart concentrations of malondialdehyde and reactive oxygen species. These changes were associated with increased (P < 0.05) mRNA and/or protein concentrations of cyclooxygenases, lipoxygenase-12, cytokines (IL-1ß), nuclear factor (NF) κB subunit, chemokines, and receptors (CCL20, CXCR1, and CXCLI2) and increased (P < 0.1) TLR4/ERK /P38/JNK in the heart of Se-insufficient chicks. CONCLUSIONS: Dietary Se insufficiency induces infiltration of mononuclear inflammatory cells in the heart of egg-type chicks. This cardiac injury was mediated by decreased functional expressions of selenoproteins, which resulted in apparent elevated oxidative stress and subsequent activations of the TLR4 pathway and NF κB.

Determination of cyclic adenosine phosphate and protein content in dormant chlamydospore and nondormant chlamydospore of Arthrobotrys flagrans.

Arthrobotrys flagrans, a nematode-eating fungus, is an effective component of animal parasitic nematode biocontrol agents. In the dried formulation, the majority of spores are in an endogenous dormant state. This study focuses on dormant chlamydospore and nondormant chlamydospore of A. flagrans to investigate the differences in cyclic adenosine monophosphate (cAMP) and protein content between the two types of spores. cAMP and soluble proteins were extracted from the nondormant chlamydospore and dormant chlamydospore of two isolates of A. flagrans. The cAMP Direct Immunoassay Kit and Bradford protein concentration assay kit (Coomassie brilliant blue method) were used to detect the cAMP and protein content in two types of spores. Results showed that the content of cAMP in dormant spores of both isolates was significantly higher than that in nondormant spores (p < 0.05). The protein content of dormant spores in DH055 bacteria was significantly higher than that of nondormant spores (p < 0.05). In addition, the protein content of dormant spores of the SDH035 strain was slightly higher than that of nondormant spores, but the difference was not significant (p > 0.05). The results obtained in this study provide evidence for the biochemical mechanism of chlamydospore dormancy or the germination of the nematophagous fungus A. flagrans.

Potential millennial-scale avian declines by humans in southern China.

Mounting observational records demonstrate human-caused faunal decline in recent decades, while accumulating archaeological evidence suggests an early biodiversity impact of human activities during the Holocene. A fundamental question arises concerning whether modern wildlife population declines began during early human disturbance. Here, we performed a population genomic analysis of six common forest birds in East Asia to address this question. For five of them, demographic history inference based on 25-33 genomes of each species revealed dramatic population declines by 4- to 48-fold over millennia (e.g. 2000-5000 thousand years ago). Nevertheless, summary statistics detected nonsignificant correlations between these population size trajectories and Holocene temperature variations, and ecological niche models explicitly predicted extensive range persistence during the Holocene, implying limited demographic consequence of Holocene climate change. Further analyses suggest high negative correlations between the reconstructed population declines and human disturbance intensities and indicate a potential driver of human activities. These findings provide a deep-time and large-scale insight into the recently recognized avifaunal decline and support an early origin hypothesis of human effects on biodiversity. Overall, our study sheds light on the current biodiversity crisis in the context of long-term human-environment interactions and offers a multi-evidential framework for quantitatively assessing the ecological consequences of human disturbance.

Evaluation of Mycobacterium tuberculosis specific antigen-stimulated CD27-CD38+IFN-γ+CD4+ T cells for discrimination of active tuberculosis.

BACKGROUND: Active tuberculosis (ATB) originates from primary Mycobacterium tuberculosis (MTB) infection or reactivation of latent tuberculosis. Besides bacteriological examination, MTB-reactive immunocytes detection can be an alternative testing for discrimination of active tuberculosis. The purpose of this study is to investigate the accuracy of peripheral blood CD27-CD38+IFN-γ+CD4+T cells in ATB diagnosis. METHODS: This prospective diagnostic accuracy study was conducted at Shanghai Pulmonary Hospital between January 2019 and December 2021. Patients with ATB, non-tuberculosis mycobacterium infection (NTM), latent tuberculosis infection (LTBI), other respiratory diseases (OD), and healthy individuals (HC) were enrolled. The accuracy of CD27-CD38+IFN-γ+CD4+/CD4+ and other phenotypic markers for ATB diagnosis was assessed. RESULTS: A total of 376 patients (237 ATB, 38 LTBI, 8 NTM, 50 OD, and 43 HC) were enrolled. The ratios of CD4+IFN-γ+CD27- and CD4+IFN-γ+CD27-CD38+ profiles in CD4+IFN-γ+ cells and the ratios of CD4+IFN-γ+CD38+, CD4+IFN-γ+CD27-, and CD4+IFN-γ+CD38+CD27- profiles in CD4+ cells in the ATB group were significantly higher than in the other groups. The area under the curve (AUC) of CD27-CD38+IFN-γ+CD4+/CD4+ for the diagnosis of ATB was the highest, with a value of 0.890. With the optimal cutoff value of 1.34 × 10-4, the sensitivity and specificity of CD27-CD38+IFN-γ+CD4+/CD4+ for ATB diagnosis was 0.869 and 0.849, respectively. CONCLUSION: CD27-CD38+IFN-γ+CD4+/CD4+ might be a potential biomarker for active tuberculosis diagnosis.

Factors predicting clinical and endoscopic remission with placebo therapy in East Asian patients with ulcerative colitis: a systematic review and meta-analysis.

BACKGROUND: The placebo effect in randomized controlled trials (RCTs) for East Asian patients with ulcerative colitis (UC) has been poorly characterized. This systematic review and meta-analysis aimed to determine the placebo rates of clinical and endoscopic remission in East Asian patients with UC and to identify factors that influenced placebo outcomes. METHODS: A comprehensive search was performed to identify all eligible RCTs. The placebo rates of clinical and endoscopic remission were pooled by random effects. Univariate meta-regression was performed to evaluate the influence of different factors on these placebo rates. RESULTS: Fourteen induction and seven maintenance RCTs were included, comprising 1781 East Asian patients with UC. The pooled placebo rates of clinical and endoscopic remission for induction trials were 13% (95% confidence interval [CI] 10-16%) and 26% (95% CI 21-32%), respectively. Corresponding values for maintenance trials were 15% (95% CI 10-23%) and 24% (95% CI 19-29%). Heterogeneity existed among the studies. On univariate meta-regression, the route of drug administration, outcome definition, disease severity, disease duration, distal disease and concomitant corticosteroids influenced the outcomes of placebo arms. CONCLUSIONS: The placebo rates of clinical and endoscopic remission in RCTs for East Asian patients with UC range from 13 to 26%, and are influenced by specific study characteristics. These results can inform the design and interpretation of future clinical trials in this region.

Au(III)-induced extracellular electron transfer by Burkholderia contaminans ZCC for the bio-recovery of gold nanoparticles.

The biorecovery of gold (Au) by microbial reduction has received increasing attention, however, the biomolecules involved and the mechanisms by which they operate to produce Au nanoparticles have been not resolved. Here we report that Burkholderia contaminans ZCC is capable of reduction of Au(III) to Au nanoparticles on the cell surface. Exposure of B. contaminans ZCC to Au(III) led to significant changes in the functional group of cell proteins, with approximately 11.1% of the (C-C/C-H) bonds being converted to CO (8.1%) and C-OH (3.0%) bonds and 29.4% of the CO bonds being converted to (C-OH/C-O-C/P-O-C) bonds, respectively. In response to Au(III), B. contaminans ZCC also displayed the ability of extracellular electron transfer (EET) via membrane proteins and could produce reduced riboflavin as verified by electrochemical and liquid chromatography-mass spectrometric results, but did not do so without Au(III) being present. Addition of exogenous reduced riboflavin to the medium suggested that B. contaminans ZCC could utilize indirect EET via riboflavin to enhance the rate of reduction of Au(III). Transcriptional analysis of the riboflavin genes (ribBDEFH) supported the view of the importance of riboflavin in the reduction of Au(III) and its importance in the biorecovery of gold.

GDF11 knockdown downregulates SMURF1 to inhibit breast cancer progression by activation of p53 and inactivation of ERα signaling.

Breast cancer (BC) is a prevalent neoplasm that occurs in women all over the world. Growth and differentiation factor 11 (GDF11) plays an essential role in cancer progression. This study focused on investigating the biological role and underlying mechanisms of GDF11 in BC. We detected the expression of GDF11 in 27 patients with BC and BC cell lines. Kaplan-Meier plotter was employed to analyze the relationship between GDF11 expression and overall survival (OS) of BC patients. The proliferative, migratory, invasive, and apoptotic abilities of T47D cells were examined. Correlation analysis of GDF11 with Smad ubiquitination regulatory factor 1 (SMURF1) was conducted. The association between GDF11 and the p53 pathway was analyzed by western blot and PFT-α (a p53 inhibitor)-mediated rescue assays. A brief analysis of the role of estrogen receptor alpha (ERα) signaling in BC progression was performed. The results showed that GDF11 was increased in BC tissues and cell lines, and the high expression of GDF11 was associated with the poor OS of BC patients. GDF11 knockdown inhibited the proliferation, migration, and invasion of T47D cells, but promoted cell apoptosis. Meanwhile, the GDF11 knockdown reduced the SMURF1 expression and invoked the p53 pathway activation. SMURF1 overexpression and PFT-α partially blocked the effects of GDF11 knockdown. In addition, GDF11 knockdown and SMURF1 silencing inhibited the activation of the ERα signaling pathway. In summary, GDF11 was involved in the progression of BC by regulating SMURF1-mediated p53 and ERα pathways, opening up a new way for BC treatment.

Population genomic, climatic and anthropogenic evidence suggest the role of human forces in endangerment of green peafowl (Pavo muticus).

Both anthropogenic impacts and historical climate change could contribute to population decline and species extinction, but their relative importance is still unclear. Emerging approaches based on genomic, climatic and anthropogenic data provide a promising analytical framework to address this question. This study applied such an integrative approach to examine potential drivers for the endangerment of the green peafowl (Pavo muticus). Several demographic reconstructions based on population genomes congruently retrieved a drastic population declination since the mid-Holocene. Furthermore, a comparison between historical and modern genomes suggested genetic diversity decrease during the last 50 years. However, climate-based ecological niche models predicted stationary general range during these periods and imply the little impact of climate change. Further analyses suggested that human disturbance intensities were negatively correlated with the green peafowl's effective population sizes and significantly associated with its survival status (extirpation or persistence). Archaeological and historical records corroborate the critical role of humans, leaving the footprint of low genomic diversity and high inbreeding in the survival populations. This study sheds light on the potential deep-time effects of human disturbance on species endangerment and offers a multi-evidential approach in examining underlying forces for population declines.

Pancreatic outflow tract reconstruction after pancreaticoduodenectomy: a meta-analysis of randomized controlled trials.

BACKGROUND: To evaluate the outcomes of pancreaticogastrostomy and pancreaticojejunostomy after pancreatoduodenectomy with the help of a meta-analysis. METHODS: Randomized controlled trials comparing pancreaticogastrostomy and pancreaticojejunostomy were searched electronically using PubMed, The Cochrane Library, and EMBASE. Fixed and random-effects were used to measure pooled estimates. Research indicators included pancreatic fistula, delayed gastric emptying, postoperative hemorrhage, intraperitoneal fluid collection, wound infection, overall postoperative complications, reoperation, and mortality. RESULTS: Overall, 10 randomized controlled trials were included in this meta-analysis, with a total of 1629 patients. The overall incidences of pancreatic fistula and intra-abdominal collections were lower in the pancreaticogastrostomy group than in the pancreaticojejunostomy group (OR=0.73, 95% CI 0.55~0.96, p=0.02; OR=0.59, 95% CI 0.37~0.96, p=0.02, respectively). The incidence of B/C grade pancreatic fistula in the pancreaticogastrostomy group was lower than that in the pancreaticojejunostomy group, but no significant difference was observed (OR=0.61, 95%CI 0.34~1.09, p=0.09). Postoperative hemorrhage was more frequent in the pancreaticogastrostomy group than in the pancreaticojejunostomy group (OR=1.52; 95% CI 1.08~2.14, p=0.02). No significant differences in terms of delayed gastric emptying, wound infection, reoperation, overall postoperative complications, mortality, exocrine function, and hospital readmission were observed between groups. CONCLUSION: This meta-analysis suggests that pancreaticogastrostomy reduces the incidence of postoperative pancreatic fistula and intraperitoneal fluid collection but increases the risk of postoperative hemorrhage compared with pancreaticojejunostomy.

Novel Models Predict Postsurgical Recurrence and Overall Survival for Patients with Hepatitis B Virus-Related Solitary Hepatocellular Carcinoma ≤10 cm and Without Portal Venous Tumor Thrombus.

BACKGROUND: The predictive model of postsurgical recurrence for solitary early hepatocellular carcinoma (SE-HCC) is not well established. The aim of this study was to develop a novel model for prediction of postsurgical recurrence and survival for patients with hepatitis B virus (HBV)-related SE-HCC ≤10 cm. PATIENTS AND METHODS: Data from 1,081 patients with HBV-related SE-HCC ≤10 cm who underwent curative liver resection from 2003 to 2016 in our center were collected retrospectively and randomly divided into the derivation cohort (n = 811) and the internal validation cohort (n = 270). Eight hundred twenty-three patients selected from another four tertiary hospitals served as the external validation cohort. Postsurgical recurrence-free survival (RFS) and overall survival (OS) predictive nomograms were generated. The discriminatory accuracies of the nomograms were compared with six conventional hepatocellular carcinoma (HCC) staging systems. RESULTS: Tumor size, differentiation, microscopic vascular invasion, preoperative α-fetoprotein, neutrophil-to-lymphocyte ratio, albumin-to-bilirubin ratio, and blood transfusion were identified as the risk factors associated with RFS and OS. RFS and OS predictive nomograms based on these seven variables were generated. The C-index was 0.83 (95% confidence interval [CI], 0.79-0.87) for the RFS-nomogram and 0.87 (95% CI, 0.83-0.91) for the OS-nomogram. Calibration curves showed good agreement between actual observation and nomogram prediction. Both C-indices of the two nomograms were substantially higher than those of the six conventional HCC staging systems (0.54-0.74 for RFS; 0.58-0.76 for OS) and those of HCC nomograms reported in literature. CONCLUSION: The novel nomograms were shown to be accurate at predicting postoperative recurrence and OS for patients with HBV-related SE-HCC ≤10 cm after curative liver resection. IMPLICATIONS FOR PRACTICE: This multicenter study proposed recurrence or mortality predictive nomograms for patients with hepatitis B virus-related solitary early hepatocellular carcinoma ≤10 cm after curative liver resection. A close postsurgical surveillance protocol and adjuvant therapy should be considered for patients at high risk of recurrence.

Secondary contact after allopatric divergence explains avian speciation and high species diversity in the Himalayan-Hengduan Mountains.

The geographical context of speciation is important for understanding speciation and community assembly. However, the predominant mode of speciation in the Himalayan-Hengduan Mountains (HHMs), a global biodiversity hotspot, remains unknown. Here, we examined the role of geography in speciation using four pairs of sister or closely related avian species that currently co-occur in the HHMs. While multilocus network analyses based on nine to eleven genes revealed deep splits between these species, several allelic networks based on individual loci suggested phylogenetic paraphyly implying a recent history of divergence. Following extensive sampling in the contact zones of these species pairs, the coalescence-based approximate Bayesian computation approach supported no gene flow during their divergence and was consistent with an allopatric speciation model. We further estimated the divergence times of the four species pairs during the middle and late Pleistocene, which were characterized by increased amplitudes of glacial variability. We found a positive relationship between their divergence times and current sympatry levels, supporting a scenario of secondary contact following allopatric speciation. The Pleistocene glacial-interglacial cycles may have led to the initial geographic population isolation; ecological divergence or mate choice might further accelerate their differentiation during secondary contact, facilitating their speciation and species accumulation in the mountainous landscape. Our findings reveal the critical role of geographic isolation in speciation in the HHMs and shed light on how this biodiversity hotspot aggregates numerous species.

Strict allopatric speciation of sky island Pyrrhula erythaca species complex.

Increasing evidence of post-divergence gene flow between taxa is shifting our understanding on the mode of speciation. A fundamental question arises concerning the circumstances under which strict allopatric speciation still holds true. Sky island populations might undergo reduced gene flow by niche conservatism to highland habitats and follow divergence in an allopatric manner. In this study, we tested this hypothesis in the sky island Grey-headed Bullfinch (Pyrrhula erythaca) species complex via statistical analyses of both genetic and ecological data. Results of coalescent-based analysis of multiple nuclear loci suggested that P. e. owstoni likely colonized Taiwan island during the severe mid-Pleistocene glacial climate followed by strictly allopatric divergence from P. e. erythaca distributed in Himalayas-Hengduan mountains and central North China. Results of ecological niche modeling suggested that their speciation may be attributed to the niche conservatism of these birds and the lack of a suitable ecological corridor during subsequent milder glacial episodes. In addition, we delimited the traditionally defined P. erythaca into two full species, P. erythaca in the Asian mainland and P. owstoni on the island of Taiwan, based on both genetic and behavioural evidences. These results suggest that ecology can have a dynamic role in allowing highland populations to expand their ranges and isolated by habitat barriers to diversify in a strictly allopatric manner.

Supplemental Nicotinamide Dose-Dependently Regulates Body Phosphorus Excretion via Altering Type II Sodium-Phosphate Co-Transporter Expressions in Laying Hens.

BACKGROUND: Dietary supplemental nicotinamide is used to treat hyperphosphatemia in humans. However, the mechanisms of its impact on body phosphorus homeostasis remain unclear. OBJECTIVE: This study was to determine effects and molecular mechanisms of 3 dietary nicotinamide concentrations on body phosphorus homeostasis in laying hens. METHODS: Hy-Line Brown layers (total = 21; 40 wk old; body weight: 1,876 ± 24 g) were individually housed (n = 7) and fed a corn-soybean meal-based diet supplemented with nicotinamide at 20 (N20), 140 (N140), and 1000 (N1000) mg/kg for 21 d. Serum phosphorus and fibroblast growth factor 23 (FGF23) concentrations, phosphorus and calcium excretion, and mRNA and/or protein of type II sodium-phosphate co-transporters (NPt2a, NPt2ab) and FGF23 and FGF23 receptors were measured in the intestines, calvaria, kidney, and liver. RESULTS: Hens in the N1000 group had a 16% lower serum phosphorus concentration and 22% greater phosphorus excretion than those in the N20 or N140 group (P ≤ 0.05). Compared with hens in the N20 group, hens in the N140 and N1000 groups, which did not differ, had 15-21% lower serum FGF23 concentrations, 19-22% greater calcium excretion, 43-56% lower ileum NPT2b protein production, and 1.5- to 1.6-fold greater kidney NPT2a protein production, respectively (all differences at P ≤ 0.05). CONCLUSIONS: Supplementing high concentrations of nicotinamide in diets for laying hens led to accelerated phosphorus and calcium excretions and decreased serum phosphorus and FGF23 concentrations, which were associated with downregulated intestinal NPt2b protein production. Our findings exclude kidney NPt2a protein production as a primary mechanism for the nicotinamide-induced body phosphorus loss.

Accessory ovary may be a treatment for infertility: Case report and review of current literatures.

Accessory ovary is a type of ovarian dysplasia, which is often defined as an ovarian tissue placed near and directly connected to the normal ovary or one of ovarian ligaments. It is often asymptomatic, mostly is found or diagnosed at laparotomy, laparoscopy or autopsy. Accessory ovary is often excised during surgery due to its potential malignant behavior. We report a case of endometriosis cyst occurred simultaneously in right side of orthotopic and accessory ovaries, together with torsion 180° of accessory ovarian cyst. Considering that the patient had not given birth and the large size of cysts, exploratory laparotomy was performed. During laparotomy, both site of ovarian cyst have been removed with orthotopic and accessory ovarian tissues preserved. After surgery, a large number of antral follicles were found both in right side of orthotopic and accessory ovaries by ultrasonography. Accessory ovary is considered to have physiological function, and can be preserved as a fertility protection measure for women who have fertility requirements. At present, the definition of ectopic ovary, accessory ovary and supernumerary ovary are very vague and rarely discussed separately. So, we proposed a new way to clarify the concepts of ectopic ovary, accessory ovary and supernumerary ovary. Moreover, we advocated that they should be discussed separately in terms of definition and management measures.

Semi-Synthesis of C-Ring Cyclopropyl Analogues of Fraxinellone and Their Insecticidal Activity Against Mythimna separata Walker.

Fraxinellone (1) is a naturally occurring degraded limonoid isolated from Meliaceae and Rutaceae plants. As a potential natural-product-based insecticidal agent, fraxinellone has been structurally modified to improve its activity. Furan ring of fraxinellone is critical in exhibiting its insecticidal activity, but with few modifications. Herein, C-ring-modified cyclopropyl analogues were semi-synthesized by Rh(II)-catalyzed cyclopropanation. The structures of the target compounds were well characterized by NMR and HRMS. The precise three-dimensional structural information of 3a was established by X-ray crystallography. Their insecticidal activity was evaluated against Mythimna separata Walker by a leaf-dipping method. Compound 3c exhibited stronger insecticidal activity than 1 and toosendanin against M. separata with teratogenic symptoms during the different periods, implying that cyclopropanation of the furan ring could strengthen the insecticidal activity of fraxinellone.

The niches of nuthatches affect their lineage evolution differently across latitude.

Ecological niche evolution can promote or hinder the differentiation of taxa and determine their distribution. Niche-mediated evolution may differ among climatic regimes, and thus, species that occur across a wide latitudinal range offer a chance to test these heterogeneous evolutionary processes. In this study, we examine (a) how many lineages have evolved across the continent-wide range of the Eurasian nuthatch (Sitta europaea), (b) whether the lineages' niches are significantly divergent or conserved and (c) how their niche evolution explains their geographic distribution. Phylogenetic reconstruction and ecological niche models (ENMs) showed that the Eurasian nuthatch contained six parapatric lineages that diverged within 2 Myr and did not share identical climatic niches. However, the niche discrepancy between these distinct lineages was relatively conserved compared with the environmental differences between their ranges and thus was unlikely to drive lineage divergence. The ENMs of southern lineages tended to cross-predict with their neighbouring lineages whereas those of northern lineages generally matched with their abutting ranges. The coalescence-based analyses revealed more stable populations for the southern lineages than the northern ones during the last glaciation cycle. In contrast to the overlapping ENMs, the smaller parapatric distribution suggests that the southern lineages might have experienced competitive exclusion to prevent them from becoming sympatric. On the other hand, the northern lineages have expanded their ranges and their current abutting distribution might have resulted from lineages adapting to different climatic conditions in allopatry. This study suggests that niche evolution may affect lineage distribution in different ways across latitude.

Hypersensitive assessment of aryl hydrocarbon receptor transcriptional activity using a novel truncated cyp1a promoter in zebrafish.

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is a persistent organic pollutant (POP), an unintentional byproduct of various industrial processes, and a human carcinogen. The expression of the cytochrome P450 1A (cyp1a) gene is upregulated in the presence of TCDD through activating the aryl hydrocarbon receptor pathway in a dose-dependent manner. Several essential response elements, including the 8 potential xenobiotic response elements in the cyp1a promoter region, have been identified to be the main functional parts for the response to TCDD. Thus, we aimed to develop a convenient and sensitive biomonitoring tool to examine the level of POPs in the environment and evaluate its potential human health risks by TCDD. Here, we established a transgenic zebrafish model with a red fluorescent reporter gene ( mCherry) using the truncated cyp1a promoter. Under exposure to TCDD, the expression pattern of mCherry in the reporter zebrafish mirrored that of endogenous cyp1a mRNA, and the primary target tissues for TCDD were the brain vessels, liver, gut, cloaca, and skin. Our results indicated that exposure of the embryos to TCDD at concentrations as low as 0.005 nM for 48 h, which did not elicit morphologic abnormalities in the embryos, markedly increased mCherry expression. In addition, the reporter embryos responded to other POPs, and primary liver cell culture of zebrafish revealed that Cyp1a protein was mainly expressed in the cytoplasm of liver cells. Furthermore, our transgenic fish embryos demonstrated that TCDD exposure can regulate the expression levels of several tumor-related factors, including epidermal growth factor, TNF-α, C-myc, proliferating cell nuclear antigen, TGF-ß, serine/threonine kinase (Akt), and phosphorylated Akt, suggesting that our transgenic fish can be used as a sensitive model to evaluate the carcinogenicity induced by TCDD exposure.-Luo, J.-J., Su, D.-S., Xie, S.-L., Liu, Y., Liu, P., Yang, X.-J., Pei D.-S. Hypersensitive assessment of aryl hydrocarbon receptor transcriptional activity using a novel truncated cyp1a promoter in zebrafish.

CRISPR/Cas9-based genome engineering of zebrafish using a seamless integration strategy.

Numerous feasible methods for inserting large fragments of exogenous DNA sequences into the zebrafish genome have been developed, as has genome editing technology using programmable nucleases. However, the coding sequences of targeted endogenous genes are disrupted, and the expression patterns of inserted exogenous genes cannot completely recapitulate those of endogenous genes. Here we describe the establishment of a novel strategy for endogenous promoter-driven and microhomology-mediated end-joining-dependent integration of a donor vector using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated (Cas) 9. We successfully integrated mCherry into the final coding sequence of targeted genes to generate seamless transgenic zebrafish lines with high efficiency. This novel seamless transgenesis technique not only maintained the integrity of the endogenous gene but also did not disrupt the function of targeted gene. Therefore, our microhomology-mediated end-joining-mediated transgenesis strategy may have broader applications in gene therapy. Moreover, this novel seamless gene-editing strategy in zebrafish provides a valuable new transgenesis technique, which was driven by endogenous promoters and in vivo animal reporter modes for translational medicine. It is expected to be a standard gene-editing technique in the field of zebrafish, leading to some important breakthroughs for studies in early embryogenesis.-Luo, J.-J., Bian, W.-P., Liu, Y., Huang, H.-Y., Yin, Q., Yang, X.-J., Pei, D.-S. CRISPR/Cas9-based genome engineering of zebrafish using a seamless integration strategy.

An in vitro investigation of telocytes-educated macrophages: morphology, heterocellular junctions, apoptosis and invasion analysis.

BACKGROUND: Telocytes (TCs), a recently discovered novel type of interstitial cells, were also found in a wide variety of human and mammalian reproductive organs/tissues, including uterus, oviduct and placenta. Previously, we demonstrated that TCs-conditioned media was capable of activating peritoneal macrophages (pMACs) through paracrine effects. This study investigates the hypothesis that direct interaction of TCs with pMACs will also play a significant role in immunoregulation of pMACs. METHODS: TCs and pMACs were derived from the uterus and intraperitoneal cavity of female BALB/c mice, respectively. TCs were identified by immunofluorescence and then co-cultured directly with pMACs for 24 h without added cytokines, to observe the in vitro biological behavior of pMACs. We used histochemical staining to study morphology and mitochondrial metabolism of pMACs, scanning electron microscopy to study heterocellular junctions, flow cytometry to investigate mitochondrial membrane potential (ΔΨm) and apoptosis, and transwell chambers to study invasion ability. Student-t test was used accordingly. RESULTS: Presently, TCs with typical structure and immunophenotype of double CD-34-positive/vimentin-positive were successfully isolated. pMACs co-cultured with TCs showed obviously morphological activation, with enhanced energy metabolism (P < 0.05). Meanwhile, direct physical cell-to-cell interaction promoted the development of heterocellular junctions between TCs and pMACs. Furthermore, TCs treatment markedly reduced the depletion of ΔΨm in co-cultured pMACs (all P < 0.05), and inhibited their apoptosis (P < 0.05). Functionally, pMACs co-cultured with TCs showed enhanced invasion ability (P < 0.05). CONCLUSIONS: Direct physical cell-to-cell interaction promoted the development of heterocellular junctions between TCs and pMACs, presumably responsible for the observed novel efficient way of pMACs activation via mitochondrial signaling pathway. TCs-educated pMACs might be a promising way to restore the defective immunosurveillance in endometriosis (EMs), led to the enhanced treatment efficacy of EMs in a simple and clinically feasible fashion.