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Within an advanced Langevin-hydrodynamics framework coupled to a hybrid fragmentation-coalescence hadronization model, we study heavy flavor quenching and flow in relativistic heavy-ion collisions. We investigate how the initial heavy quark spectrum, the in-medium energy loss and hadronization mechanisms of heavy quarks, the evolution profile of the pre-equilibrium stage, the medium flow, and the temperature dependence of heavy quark diffusion coefficients influence the suppression and elliptic flow of heavy mesons at the RHIC and the LHC. Our results show that the different modeling of initial conditions, pre-equilibrium evolution, and in-medium interactions can individually yield uncertainties of approximately 10-40% in D meson suppression and flow at a low transverse momentum. We also find that proper combinations of collisional versus radiative energy loss, coalescence versus fragmentation in hadronization, and the inclusion of medium flow are the most important factors for describing the suppression and elliptic flow of heavy mesons.

We develop for charmed hadron production in relativistic heavy-ion collisions a comprehensive coalescence model that includes an extensive set of s and p-wave hadronic states as well as the strict energy-momentum conservation, which ensures the boost invariance of the coalescence probability and the thermal limit of the produced hadron spectrum. By combining our hadronization scheme with an advanced Langevin-hydrodynamics model that incorporates both elastic and inelastic energy loss of heavy quarks inside the dynamical quark-gluon plasma, we obtain a successful description of the pT-integrated and differential Λc/D0 and Ds/D0 ratios measured at RHIC and the LHC. We find that including the effect of radial flow of the medium is essential for describing the enhanced Λc/D0 ratio observed in relativistic heavy-ion collisions. We also find that the puzzling larger Λc/D0 ratio observed in Au+Au collisions at RHIC than in Pb+Pb collisions at the LHC is due to the interplay between the effects of the QGP radial flow and the charm quark transverse momentum spectrum at hadronization. Our study further suggests that charmed hadrons have larger sizes in medium than in vacuum.

Contents Protein disulphide isomerase family A3 (PDIA3) has an activity of thioredoxin, widely expressed in multiple tissues and involved in multiple cellular processes. It was recently found in human and mouse sperm cells and could affect sperm-egg fusion. Therefore, the present investigation aims to identify PDIA3 mRNA and protein in rat testis and sperm cells. Rat PDIA3 cDNA was cloned by RT-PCR. The cRNA riboprobe was transcribed from PDIA3 cDNA and was used to display PDIA3 mRNA location in rat testes by in situ hybridization. PDIA3 protein distribution was also observed in testis and sperm cells by immunohistochemistry and immunocytochemistry, respectively. The rat PDIA3 transcript and protein were localized in the cells from spermatocytes to the spermatozoa phases of rat testes, mostly in the pachytene spermatocytes. PDIA3 protein was also observed on the intact sperm membrane including the tail. The rat PDIA3 gene is transcribed and translated through the whole spermatogenesis process, and the PDIA3 protein is spread all over the sperm cell membrane. The results provide some primary information about PDIA3 in testis and sperm for further study on PDIA3 function in rat spermatogenesis and sperm-egg fusion. [ABSTRACT FROM AUTHOR]

Objective: Melanoma, with its high degree of malignancy, stands as one of the most dangerous skin cancers and remains the primary cause of death from skin cancer. With studies demonstrating the potential of traditional Chinese medicine to intervene and treat melanoma, we turned our attention to celastrol. Celastrol is a triterpene compound extracted from the traditional Chinese medicine derived from Tripterygium wilfordii. Previous studies have shown that celastrol exerts inhibitory effects on various malignant tumors, including melanoma. Hence, our goal was to clarify the impact of celastrol on cell viability, apoptosis, and cell cycle progression by elucidating its effects on the PI3K/AKT/mTOR pathway., Methods: CCK-8 and wound healing assays were used to determine the effect of celastrol on the viability and migration of B16-F10 cells. Changes in cell apoptosis, cell cycle, reactive oxygen species (ROS), and mitochondrial membrane potential were detected by flow cytometry. PI3K/AKT/mTOR pathway proteins and HIF-α mRNA expression in B16-F10 cells were detected by western blotting and qPCR. Moreover, the addition of a PI3K activator demonstrated that celastrol could inhibit the function of B16-F10 cells via the PI3K/AKT/mTOR pathway., Results: Celastrol inhibited the viability and migration of B16-F10 cells. Through the inhibition of the PI3K/AKT/mTOR pathway down-regulates the expression of HIF-α mRNA, thereby causing an increase of ROS in cells and a decrease in the mitochondrial membrane potential to promote cell apoptosis and cell cycle arrest. The inhibitory effect of celastrol on B16-F10 cells was further demonstrated by co-culturing with a PI3K activator., Conclusion: Celastrol inhibits the function of B16-F10 cells by inhibiting the PI3K/AKT/mTOR cellular pathway and regulating the expression of downstream HIF-α mRNA., (© 2024. The Author(s).)

This study investigated the effects of extrusion on the physical properties of glutinous rice and addressed the challenges associated with its hardened texture and reduced taste in glutinous rice products by adding extruded glutinous rice to assess their anti-retrogradation effect compared with different improvers. Glutinous rice flour with different gelatinization degrees was obtained by changing the initial moisture content of glutinous rice grains before extrusion, and their physicochemical properties and the effect of adding them to rice products were analyzed. Results showed that with the increase in moisture content, the viscosity, water absorption index of extruded glutinous rice flour, and product viscosity increased, while the gelatinization degree, water solubility index, and product elasticity decreased, and the hardness of the rice products showed a trend of first decreasing and then increasing. Twenty percent moisture content of glutinous rice products showed the best properties mentioned above. The effects of adding different improvers on retrogradation degree, quality characteristics, microstructure, and moisture migration of glutinous rice products were analyzed by texture profile analysis, sensory evaluation, scanning electron microscopy, and low-field nuclear magnetic resonance. It was found that soybean polysaccharides, xanthan gum, and extruded glutinous rice flour had better anti-retrogradation effects, while colloid and soybean polysaccharides provided a tighter and more three-dimensional internal structure to the rice products. Our study showed that extruded glutinous rice flour had good anti-retrogradation properties and little effect on flavor and taste, but it would increase the roughness and viscosity of the products, which had advantages and disadvantages compared with other improvers., (© 2023 Institute of Food Technologists.)

Objective: This study prospectively investigates the association between immunoglobulin G (IgG) N-glycan traits and ischemic stroke (IS) risk., Methods: A nested case-control study was conducted in the China suboptimal health cohort study, which recruited 4,313 individuals in 2013-2014. Cases were identified as patients diagnosed with IS, and controls were 1:1 matched by age and sex with cases. IgG N-glycans in baseline plasma samples were analyzed., Results: A total of 99 IS cases and 99 controls were included, and 24 directly measured glycan peaks (GPs) were separated from IgG N-glycans. In directly measured GPs, GP4, GP9, GP21, GP22, GP23, and GP24 were associated with the risk of IS in men after adjusting for age, waist and hip circumference, obesity, diabetes, hypertension, and dyslipidemia. Derived glycan traits representing decreased galactosylation and sialylation were associated with IS in men (FBG2S2/(FBG2 + FBG2S1 + FBG2S2): odds ratio ( OR ) = 0.92, 95% confidence interval ( CI ): 0.87-0.97; G1 n : OR = 0.74, 95% CI : 0.63-0.87; G0 n : OR = 1.12, 95% CI : 1.03-1.22). However, these associations were not found among women., Conclusion: This study validated that altered IgG N-glycan traits were associated with incident IS in men, suggesting that sex discrepancies might exist in these associations., (Copyright © 2023 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.)

The risk of dementia increases in patients with cognitive impairment. However, it is not clear what factors contribute to the onset of dementia in those with cognitive impairment. In this prospective cohort study, we will investigate the every-five-year incidence of cognitive impairment and prognostic factors for cognitive impairment. The Jidong cognitive impairment cohort was established from April 2012 to August 2015, during which we recruited 5854 healthy participants (55.1% male) older than 45 years (mean, 57 years). Participants received a health examination in the Staff Hospital, Jidong Oilfield Branch, China National Petroleum Corporation. Baseline data and blood samples were collected. Cognitive impairment was evaluated using the Mini-Mental State Examination, and was defined as a Mini-Mental State Examination score of less than 24. Dementia was assessed using the criteria of Diagnostic and Statistical Manual of Mental Disorders (Fourth edition), the International Working Group criteria, and the Mini-Mental State Examination score. The follow-up will continue until December 2024, during which a prognostic model will be constructed. The primary outcome is the presence/absence of dementia and the secondary outcome is quality of life. Baseline screening results showed the following: (1) Cognitive impairment was apparent in 320 participants (5.5%). These participants will be excluded from the Jidong cohort study, and the remaining participants will be followed up. (2) Of the 320 participants with cognitive impairment, there was a significantly higher prevalence of illiteracy than other education levels (35.9%, P < 0.05). Age, arterial hypertension, alcohol consumption, and passive smoking differed significantly between the cognitive impairment and healthy groups (P < 0.05). Multivariate logistic regression models showed that age (odds ratio [OR] = 1.059, 95% confidence interval [CI]: 1.044-1.074) and arterial hypertension (OR = 1.665, 95% CI: 1.143-2.427) were risk factors for mild cognitive impairment. With the increase of educational level (illiteracy, primary school, junior high school, high school, university, and above), cognitive impairment gradually decreased (OR < 1, P < 0.05). (3) This cohort study has initially screened for several risk factors for cognitive impairment at baseline, and subsequent prospective data will further describe, validate, and evaluate the effects of these risk factors on cognitive impairment and dementia. These results can provide clinical evidence for the early prevention of cognitive impairment and dementia. The study was approved by the Ethics Committee of Kailuan General Hospital of Tangshan City and the Medical Ethics Committee, Staff Hospital, Jidong Oilfield Branch, China National Petroleum Corporation on July 12, 2013 (approval No. 2013 YILUNZI 1)., Competing Interests: None

Gene structure and expression regulation mechanism are the research hotspots and focus of modern life sciences. The lac operon is a cluster of genes through which Escherichia coli catabolizes lactose. It was first proposed by F. Jacob and J. Monod, who were also awarded the Nobel Prize in Physiology or Medicine in 1965 for their contributions. Thereafter, the lac operon has become the classic teaching case of the gene regulation mechanism in microbiology, genetics and molecular biology, and been highly valued by teachers and students alike. Although the conclusion is easy to follow and memorize, its rich connotation and esoteric reasoning has rendered it difficult to understand, neither is it easy for teachers to fully exploit the advantages of this teaching case. Therefore it is necessary to have an in-depth understanding of the genetic structure and working principle of the lac operon, especially the scientific background and thinking process through which scientists revealed these mysteries. In this paper, the historical discovery and analysis process of the E. coli lac operon was reviewed by following their footprints, listening to their analysis of experimental results. Based on the DNA sequences, the reasons for several unusual phenomena of lac operon expression were also discussed to exemplify the teaching value of the classic cases in genetics and molecular biology.

The classroom is the main venue for undergraduate teaching. It is worth pondering how to cultivate undergraduate's research ability in classroom teaching. Here we introduce the practices and experiences in teaching reform in genetics for training the research quality of undergraduate students from six aspects: (1) constructing the framework for curriculum framework systematicaly, (2) using the teaching content to reflect research progress, (3) explaining knowledge points with research activities, (4) explaining the scientific principles and experiments with PPT animation, (5) improving English reading ability through bilingual teaching, and (6) testing students' analysing ability through examination. These reforms stimulate undergraduate students' enthusiasm for learning, cultivate their ability to find, analyze and solve scientific problems, and improve their English reading and literature reviewing capacity, which lay a foundation for them to enter the field of scientific research.

Human noroviruses are the most important viral pathogens causing epidemic acute gastroenteritis, in which the GII.4 viruses have been predominant worldwide for the past decades. During 2014-2015 winter season, a new GII.17 variant emerged as the predominant virus in China surpassing the GII.4 virus in causing significantly increased acute gastroenteritis outbreaks. Genome sequences of the new GII.17 variant was determined and compared with other GII.17 noroviruses, revealing residue substitutions at specific locations, including the histo-blood group antigen-binding site and the putative antigenic epitopes. Further study of GII.17 outbreaks focusing on host susceptibility showed that the new GII.17 variant infected secretor individuals of A, B, O and Lewis types. Accordingly, the P particles of the new GII.17 variant bound secretor saliva samples of A, B, O and Lewis types with significantly higher binding signals than those of the P particles of the previous GII.17 variants. In addition, human sera collected from the outbreaks exhibited stronger blockade against the binding of the new GII.17 P particles to saliva samples than those against the binding between the P particles of previous GII.17 variants and saliva samples. Taken together, our data strongly suggested that the new GII.17 variant gained new histo-blood group antigen-binding ability and antigenic features, which may contribute to its predominance in causing human norovirus epidemics.

Reduction of protein phosphatase-2A (PP2A) activity is a common clinical feature of Alzheimer's disease and vascular dementia. In this study, we observed that chronic brain hypoperfusion induced by bilateral common carotid artery occlusion of rats led to PP2A inactivation based on the increase in tyrosine-307 phosphorylation and leucine-309 demethylation of PP2AC and the depression in PP2ABα. Knockdown of miR-195 using overexpression of its antisense molecule oligonucleotide (pre-AMO-miR-195) delivered by a lentivirus (lenti-pre-AMO-miR-195) increased tyrosine-307 phosphorylation and decreased both PP2ABα expression and leucine-309 methylation; these effects were prevented by the overexpression of miR-195 using lenti-pre-miR-195 and controlled by an increase in methylesterase (PME-1) and a decrease in leucine carboxyl methyltransferase-1. In vitro studies demonstrated that miR-195 regulated PME-1 expression by binding to the Ppme1 gene 3'-untranslated region (3'UTR) domain. Masking the miR-195 binding sites in the amyloid precursor protein (APP) and β-site APP cleaving enzyme 1 genes prevented miR-195-induced leucine carboxyl methyltransferase-1 elevation. We concluded that the miR-195 downregulation in chronic brain hypoperfusion involved PP2A inactivity, which was mediated by the post-transcriptional regulation PME-1, APP, and β-site APP cleaving enzyme 1 expression., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Twenty-four-month-old male C57BL/6 mice with low serum testosterone levels were used as a late-onset hypogonadism (LOH) animal model for examining the effects of velvet antler polypeptide (VAP) on sexual function and testosterone synthesis. These mice received VAP for 5 consecutive weeks by daily gavage at doses of 100, 200, or 300 mg kg-1 body weight per day (n = 10 mice per dose). Control animals (n = 10) received the same weight-based volume of vehicle. Sexual behavior and testosterone levels in serum and interstitial tissue of testis were measured after the last administration of VAP. Furthermore, to investigate the mechanisms of how VAP affects sexual behavior and testosterone synthesis in vivo, the expression of steroidogenic acute regulatory protein (StAR), cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc), and 3β-hydroxysteroid dehydrogenase (3β-HSD) in Leydig cells was also measured by immunofluorescence staining and quantitative real-time PCR. As a result, VAP produced a significant improvement in the sexual function of these aging male mice. Serum testosterone level and intratesticular testosterone (ITT) concentration also increased in the VAP-treated groups. The expression of StAR, P450scc, and 3β-HSD was also found to be enhanced in the VAP-treated groups compared with the control group. Our results suggested that VAP was effective in improving sexual function in aging male mice. The effect of velvet antler on sexual function was due to the increased expression of several rate-limiting enzymes of testosterone synthesis (StAR, P450scc, and 3β-HSD) and the following promotion of testosterone synthesis in vivo.

The JAK-STAT3 signaling pathway is one of the critical pathways regulating cell proliferation, differentiation, and apoptosis. Myocardin is regarded as a key mediator for the change of smooth muscle phenotypes. However, the relationship between STAT3 and myocardin in the vascular smooth muscle cell (VSMC) phenotypic switch has not been investigated. The goal of this study was to investigate the molecular mechanism by which STAT3 affects the myocardin-regulated VSMC phenotypic switch. Data presented in this study demonstrated that STAT3 was rapidly up-regulated after stimulation with VEGF. Inhibition of the STAT3 activation process impaired VSMC proliferation and enhanced the expression of VSMC contractile genes by increasing serum-response factor binding to the CArG-containing regions of VSMC-specific contractile genes. In contrast, the interaction between serum-response factor and its co-activator myocardin was reduced by overexpression of STAT3. In addition, treated VEGF inhibited the transcription activity of myocardin, and overexpression of STAT3 inhibited myocardin-induced up-regulation of VSMC contractile phenotype-specific genes. Although myocardin and STAT3 are negatively correlated, interestingly, both of them can enhance the expression of VEGF, suggesting a feedback loop to regulate the VSMC phenotypic switch. Taken together, these results indicate that the JAK-STAT3 signaling pathway plays a key role in controlling the phenotypic switch of VSMCs through the interactions between STAT3 and myocardin by various coordinated gene regulation pathways and feedback loops., (© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.)

Breast cancer is the leading cause of cancer death in women worldwide which is closely related to metastasis. But the exact molecular mechanism of metastasis is still not fully understood. We now report that both MRTF-A and STAT3 play important roles in migration of MDA-MB-231 breast cancer cells. Moreover, MRTF-A and STAT3 synergistically increased MDA-MB-231 cell migration by promoting the expression of migration markers urokinase-type plasminogen activator (uPA) and osteopontin (OPN) and inhibiting the expression of breast cancer metastasis suppressor 1 (BRMS1). Luciferase reporter assays demonstrated that MRTF-A and STAT3 do not affect transcription of the BRMS1 promoter. Instead, we identified a newly molecular mechanism by which MRTF-A and STAT3 synergistically controlled MDA-MB-231 cell migration by recruiting DNMT1 to hypermethylate the promoter of BRMS1 and thus affect the expression of BRMS1. Interestingly, physical interaction between MRTF-A and STAT3 synergistically promotes the transactivity of DNMT1 by binding to the GAS element within the DNMT1 promoter. Our data thus provide important and novel insights into the roles of MRTF-A and STAT3 in regulating MDA-MB-231 cell migration., (© 2015 International Union of Biochemistry and Molecular Biology.)

In Mendellian genetics, the dominance and recessiveness are used to describe the functional relationship between two alleles of one gene in a heterozygote. The allele which constitutes a phenotypical character over the other is named dominant and the one functionally masked is called recessive. The definitions thereby led to the creation of Mendel's laws on segregation and independent assortment and subsequent classic genetics. The discrimination of dominance and recessiveness originally is a requirement for Mendel's logical reasoning, but now it should be explained by cellular and molecular principles in the modern genetics. To answer the question raised by students of how the dominance and recessiveness are controlled, we reviewed the recent articles and tried to summarize the cellular and molecular basis of dominant and recessive inheritance. Clearly, understanding the essences of dominant and recessive inheritance requires us to know the dissimilarity of the alleles and their products (RNA and/or proteins), and the way of their function in cells. The alleles spatio-temporally play different roles on offering cells, tissues or organs with discernible phenotypes, namely dominant or recessive. Here, we discuss the changes of allele dominance and recessiveness at the cellular and molecular levels based on the variation of gene structure, gene regulation, function and types of gene products, in order to make students understand gene mutation and function more comprehensively and concretely.

Objective: The aim of this study was to facilitate the systematic discovery of diagnostic biomarkers of diabetic nephropathy (DN)., Methods: 3 publicly available independent cohorts were got from Gene Expression Omnibus database. Gene expression array were used to screen for genome-wide relative significance (GWRS) and genome-wide global significance (GWGS). The most significant up- and down-regulated top 100 gene signatures were identified using a fold change based model. Then the protein-protein interaction (PPI) network was constructed, while the hub genes in this PPI network were identified by centrality analysis. Modules detection was performed to explore the functions of the modules. Meanwhile, gene enrichment analysis was performed to illuminate the biological pathways and processes associated with DN., Results: The most significant up- and down-regulated top 100 gene signatures were identified and a PPI network was established. Several hub genes (VEGFA, IL8, MYC, CD14, ALB) were discovered. Several functional modules were revealed. Biological pathways including cytokine-cytokine receptor interaction and p53 signaling pathway, and processes including inflammatory response, response to wounding and enzyme linked receptor protein signaling pathway were identified., Conclusion: Our study displayed underlying biomarkers including biological pathways and several hub genes of DN., (© Georg Thieme Verlag KG Stuttgart · New York.)

Myocardin is well known to play a key role in the development of cardiomyocyte hypertrophy. But the exact molecular mechanism regulating myocardin stability and transactivity to affect cardiomyocyte hypertrophy has not been studied clearly. We now report that NF-κB (p65) can inhibit myocardin-induced cardiomyocyte hypertrophy. Then we explore the molecular mechanism of this response. First, we show that p65 can functionally repress myocardin transcriptional activity and also reduce the protein expression of myocardin. Second, the function of myocardin can be regulated by epigenetic modifications. Myocardin sumoylation is known to transactivate cardiac genes, but whether p65 can inhibit SUMO modification of myocardin is still not clear. Our data show that p65 weakens myocardin transcriptional activity through attenuating SUMO modification of myocardin by SUMO1/PIAS1, thereby impairing myocardin-mediated cardiomyocyte hypertrophy. Furthermore, the expression of myocardin can be regulated by several microRNAs, which play important roles in the development and function of the heart and muscle. We next investigated potential role of miR-1 in cardiac hypotrophy. Our results show that p65 can upregulate the level of miR-1 and miR-1 can decrease protein expression of myocardin in cardiac myocytes. Notably, miR-1 expression is also controlled by myocardin, leading to a feedback loop. These data thus provide important and novel insights into the function that p65 inhibits myocardin-mediated cardiomyocyte hypertrophy by downregulating the expression and SUMO modification of myocardin and enhancing the expression of miR-1., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Breast cancer is the leading cause of cancer death in women worldwide which is closely related to metastasis. But the exact molecular mechanism on metastasis is still not fully understood; we now report that both MRTF-A and STAT3 play important role in breast cancer migration of MDA-MB-231 cells. Moreover, MRTF-A and STAT3 synergistically increased MDA-MB-231 cell migration by promoting the expression of migration markers Myl-9 and Cyr-61. Importantly, we identified a detailed molecular mechanism of MDA-MB-231 cell migration controlled via physical interaction between MRTF-A and STAT3, which synergistically promote the transactivity of the migration marker Myl-9 and Cyr-61 by CArG box binding. Interestingly, the two signaling pathways RhoA-MRTF-A and JAK-STAT3 across talk to regulate MDA-MB-231 cell migration. Our data thus provide important and novel insights into the roles of MRTF-A and STAT3 in regulating MDA-MB-231 cell migration., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Breast cancer is the leading cause of cancer death in women worldwide. It is well known that oncogene activation and anti-oncogene inactivation affect the development and progression of breast cancer, but the role of oncogene activation and anti-oncogene inactivation in breast cancer is still not fully understood. We now report that maspin acts as a tumor suppressor gene to induce MCF-7 cell apoptosis. In addition, maspin promoter hypermethylation and histone hypoacetylation lead to silencing of maspin gene expression in MCF-7 cells. Moreover, DNA methyltransferase (DNMT) inhibitor 5-aza-2'-deoxycytidine (5-aza-dc) and/or the histone deacetylase (HDAC) inhibitor Trichostatin A (TSA) strongly up-regulated the expression of maspin in MCF-7 cells. Notably, myocardin can promote the re-expression of maspin in MCF-7 cells. Luciferase assay shows that myocardin activates the transcription of maspin promoter by CArG box. More importantly, 5-aza-dc/TSA and myocardin synergetically enhance re-expression of maspin and augment maspin-mediated apoptosis in MCF-7 cells. Thus, these data reveal the new insight that myocardin meditates apoptosis in breast cancer through affecting maspin re-expression and epigenetic modification to regulate the development of breast cancer, thereby raising the possibility of its use in breast cancer therapy., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Human chorionic gonadotropin (hCG) is a glycoprotein produced by placental trophoblasts. Previous studies indicated that hCG could be responsible for the pregnancy-induced protection against breast cancer in women. It is reported that hCG decreases proliferation and invasion of breast cancer MCF-7 cells. Our research also demonstrates that hCG can reduce the proliferation of MCF-7 cells by downregulating the expression of proliferation markers, proliferating cell nuclear antigen (PCNA), and proliferation-related Ki-67 antigen (Ki-67). Interestingly, we find here that hCG elevates the state of cellular differentiation, as characterized by the upregulation of differentiation markers, β-casein, cytokeratin-18 (CK-18), and E-cadherin. Inhibition of hCG secretion or luteinizing hormone/hCG receptors (LH/hCGRs) synthesis can weaken the effect of hCG on the induction of cell differentiation. Furthermore, hCG can suppress the expression of estrogen receptor alpha. hCG activated receptor-mediated cyclic adenosine monophosphate/protein kinase A signaling pathway. These findings indicated that a protective effect of hCG against breast cancer may be associated with its growth inhibitory and differentiation induction function in breast cancer cells., (© 2014 International Union of Biochemistry and Molecular Biology.)

Objective: To explore the application of multiple displacement amplification (MDA) combined with short tandem repeats (STRs) in preimplantation genetic diagnosis (PGD)., Methods: MDA was applied to amplify the whole genome of a single cell and to retrieve and assemble the highly heterogeneous STR loci among human population. Haplotype analytic system was established with aiming at diagnosis of the single gene diseases by selecting the STR loci located within the pathogenic genes or on both bounding sides of the pathogenic genes. At the same time, allele specific amplification, PCR-reverse dot-blotting hybridization methods and gene sequencing methods were employed for direct detection of the pathogenic genes. The STR loci located at related chromosomes were selected to carry out allele number analysis on the basis of chromosome number and structural abnormality., Results: In the study, 12 PGD systems were set up including 6 different monogenic diseases (spinal muscular atrophy, Duchenne muscular dystrophy, X-linked chronic granulomatous disease, osteopetrosis, achondroplasia, X-linked severe combined immunodeficiency), Robertsonian translocations, α-thalassemia combined with Robertsonian translocation, α- and β-double thalassemia, β-thalassemia with HLA typing and DMD with HLA typing. Then 44 PGD cycles were performed for 35 couples with different kinds of inherited diseases, which resulted in 20 healthy liveborns (12 singletons and 4 twins) and 5 ongoing pregnancies. The clinical pregnancy rate was 47.7% (21/44) per PGD cycle. The overall diagnostic rate was 94.6% (367/388). The MDA failed in 3.6% (14/388) single blastomeres. The amplification rate of the subsequent PCR was 97.1% and the average allele drop out (ADO) rate was 12.6% (range: 0-47.5%)., Conclusion: The application of MDA combined with STRs provided a generic PGD approach for different genetic disorders, especially for simultaneous diagnosis of two or more hereditary statuses. The method could greatly shorten the time of developing PGD system of new diseases, which broadens the indications of PGD.

The aim of the present study was to analyse the antiproliferative effects and mechanisms of action of protein kinase inhibitors (PKIs) in human glioblastoma multiforme (GBM) cells with different epidermal growth factor receptor (EGFR) and phosphatase and tensin homologue (PTEN) status. The GBM cell models were established by transfection of plasmids carrying wild-type EGFR, mutated EGFRvIII or PTEN and clonal selection in U87MG cells. Phosphatidylinositol 3-kinase (PI3-K)/AKT pathway-focused gene profiles were examined by real-time polymerase chain reaction-based assays, protein expression was evaluated by western blotting and the antiproliferative effects of PKI treatment were determined by the 3-(4,5-dimethyl-2 thiazoyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay in GBM cells. The cell model with intact PTEN and low EGFR levels was the most sensitive to treatment with the EGFR inhibitor erlotinib, whereas the model with EGFRvIII was the most resistant to treatment with the mitogen-activated protein kinase kinase inhibitor U0126. The dual PI3-K and mammalian target of rapamycin (mTOR) inhibitor PI103 had the most potent antiproliferative effects against all GBM cells tested. Following simultaneous stimulation of AKT and extracellular signal-regulated kinase, rapamycin concentrations > 0.5 nmol/L failed to exhibit a further growth inhibitory effect. Concurrent inhibition of mTOR and ribosomal protein s6 activity may underlie the inhibition of GBM proliferation by PKI. In conclusion, overexpression of EGFR or EGFRvIII, accompanied by a loss of PTEN, contributed to the activation of multiple intracellular signalling pathways in GBM cells. Rigorous examination of biomarkers in tumour tissues before and after treatment may be necessary to determine the efficacy of PKI therapy in patients with GBM., (© 2012 The Authors Clinical and Experimental Pharmacology and Physiology © 2012 Wiley Publishing Asia Pty Ltd.)

Acute myocardial infarction (AMI) is a leading cause of death worldwide. Most cases of AMI result from coronary atherosclerosis (AS). The pathogenic mechanisms underlying AS lesions and AMI are incompletely understood. Calcium-sensing receptors (CaSR) belong to a family of G-protein-coupled receptors. We previously discovered that CaSR was expressed in the heart tissue of adult rats. CaSR may contribute to AMI in AS. We initially established a rat model of AS by injection of vitamin D(3) and feeding with a high-fat diet. Isoproterenol (ISO) was used to induce AMI. The MB isoenzyme of creatine kinase (CK-MB), lactate dehydrogenase (LDH), cardiac troponin T (cTnT), tetrazolium chloride staining, and cardiac function parameters were selected as indicators of myocardial damage or necrosis. Cardiac apoptosis was analyzed by transferase dUTP nick-end labeling (TUNEL) assay. Expression of CaSR, Bcl-2, Bax, caspase-3, p-ERK1/2, p-JNK, and p-p38 were determined by Western blot analysis. Compared with the control group, levels of cTnT, CK-MB, and LDH; number of TUNEL-positive cells; and expression of CaSR, Bax, caspase-3, p-ERK1/2, p-JNK and p-p38, were significantly increased, whereas cardiac function and expression of Bcl-2 were decreased markedly in isoproterenol (ISO)-treated group (C/ISO) and AS groups. These changes were significant in the AS/ISO group than in the C/ISO group or AS group. The upregulation of CaSR during AS formation renders hypersensitivity to AMI. Activation of the pro-apoptotic mitochondria pathway and JNK-p38 MAPK pathway triggered by increased expression of CaSR may be one of molecular mechanisms underlying AMI in AS.

Matrix metalloproteinase-2 (MMP-2) is constitutively expressed in vascular smooth muscle cells (VSMCs) and up-regulated in atherosclerotic lesion by various stimuli, such as oxidized low-density lipoprotein (oxLDL). Calcium-sensing receptor (CaSR) is also expressed in VSMCs, but it remains unclear whether CaSR is associated with overproduction of MMP-2 in VSMCs. In this study, the expression of MMP-2 was detected by real-time PCR and Western blot analysis, and the gelatinolytic activity of MMP-2 was measured using gelatin zymography. Our results showed that oxLDL enhanced MMP-2 expression and activity in rat aortic VSMCs in a time- and dose-dependent manner. In addition, CaSR expression was up-regulated by oxLDL. Manipulating CaSR function in these cells by NPS2390 (an antagonist of CaSR) or GdCl(3) (an agonist of CaSR) affected the oxLDL-induced MMP-2 production. In VSMCs, oxLDL stimulated the rapid activation of phosphatidylinositol 3-kinase (PI3K)/Akt signal pathway, as determined by Western blot analysis. Phosphorylation of Akt and MMP-2 production stimulated by oxLDL were attenuated by LY294002 (a specific inhibitor of PI3K). Activation of Akt was suppressed by NPS2390 but enhanced by GdCl(3). In contrast, oxLDL had no stimulatory effect on the phosphorylation of JNK, and pretreatment with SP600125 (an inhibitor of JNK) produced no significant effect on oxLDL-induced MMP-2 production. These results suggest that CaSR mediates oxLDL-induced MMP-2 production in VSMCs via PI3K/Akt signal pathway.

1. Calcium-sensing receptors (CaSR) exist in a variety of tissues. In 2010, we first identified its functional expression in Buffalo rat liver (BRL) cells and demonstrated that the activation of CaSR was involved in an increased intracellular calcium through the Gq subunit-phospholipase C-inositol triphosphate pathway. However, its role and related mechanism in hepatic ischaemia/reperfusion (I/R) injury is still unclear. 2. Therefore, in the present study, BRL cells were incubated in ischaemia-mimetic solution for 4 h, then reincubated in the normal culture medium for 10 h to establish a simulated I/R model. We assayed the apoptotic ratio of BRL cells by flow cytometry and Hoechst 33342 staining; analyzed the expression of CaSR, cytochrome c (Cyt-c), caspase-3, Bcl-2, Bax, extracellular signal-regulated protein kinase (ERK), and p38 by Western blotting; and measured the concentration of intracellular calcium by laser-scanning confocal microscopy. 3. The results showed that simulated I/R increased the expression of CaSR and induced apoptosis in BRL cells. GdCl(3), a specific activator of CaSR, further increased CaSR expression, intracellular calcium, and apoptosis in BRL cells during I/R. The activation of CaSR downregulated Bcl-2 expression, upregulated Cyt-c, caspase-3, and Bax expressions, and promoted p38 and ERK-1/2 phosphorylation. 4. In conclusion, increased CaSR expression plays a vital role in apoptosis induced by I/R injury, in which its mechanism is related with calcium overload and the activation of the mitochondrial and mitogen-activated protein kinase apoptotic pathways. The regulation of CaSR activity might serve as a novel pharmacological target to prevent and treat liver disease., (© 2011 The Authors. Clinical and Experimental Pharmacology and Physiology © 2011 Blackwell Publishing Asia Pty Ltd.)

Activation of the calcium-sensing receptor (CaSR) leads to an increase of intracellular calcium concentration and alteration of cellular activities. High level of intracellular calcium is involved in hypoxia-induced proliferation of pulmonary arterial smooth muscle cells (PASMCs). However, whether the CaSR is expressed in PAMSCs and is related to the hypoxia-induced proliferation of PASMCs is unclear. In this study, the expression and distribution of CaSRs were detected by RT-PCR, western blotting and immunofluorescence; the intracellular concentration of free calcium ([Ca(2+) ](i) ) was determined by confocal laser scanning microscopy; cell proliferation was tested using an MTT and BrdU incorporation assay; cell cycle analysis was carried out using a flow cytometric assay; and the expression of proliferating cell nuclear antigen (PCNA), extracellular signal-regulated protein kinase 1,2 (ERK1,2) and AKT were analysed by western blotting. We observed that both CaSR mRNA and protein were expressed in rat PASMCs. Lowering of oxygen from 21% to 2.5% led to increased [Ca(2+) ](i) and CaSR expression. This condition of hypoxia also stimulated PASMCs proliferation accompanying with increased phosphorylation of ERK1,2 and AKT. GdCl(3) (an agonist of CaSR) or NPS2390 (an antagonist of CaSR) amplified or weakened the effect of hypoxia, respectively. PD98059 (a MEK1 inhibitor) or LY294002 (a PI3K inhibitors) decreased the up-regulation of PCNA expression and the increase of the cell proliferation index induced by hypoxia and GdCl(3) in PASMCs. Our results suggest that CaSR is expressed in rat PASMCs, and that CaSR activation through MEK1/ERK1,2 and PI3 kinase pathways is involved in hypoxia-induced proliferation of PASMCs., (© 2010 The Authors. Basic & Clinical Pharmacology & Toxicology © 2010 Nordic Pharmacological Society.)

We cloned the cDNA and genomic DNA encoding for Izumo1 of cashmere goat (Capra hircus) and sheep (Ovis aries). Analysis of 4.6 kb Izumo1 genomic sequences in sheep and goat revealed a canonical open reading frame (ORF) of 963 bp spliced by eight exons. Sheep and goat Izumo1 genes share >99% identity at both DNA and protein levels and are also highly homologous to the orthologues in cattle, mouse, rat and human. Extensive cloning and analysis of Izumo1 cDNA revealed three (del 69, del 182 and del 217) and two (del 69 and ins 30) alternative splicing isoforms in goat and sheep, respectively. All of the isoforms are derived from splicing at typical GT-AG sites leading to partial or complete truncation of the immunoglobulin (Ig)-like domain. Bioinformatics analysis showed that caprine and ovine Izumo1 proteins share similar structure with their murine orthologue. There are a signal peptide at the N-terminus (1-22 aa), a transmembrane domain at the C-terminus (302-319 aa), and an extracellular Ig-like region in the middle (161-252 aa) with a putative N-linked glycosylation site (N(205)-N-S). Alignment of Izumo1 protein sequences among 15 mammalian species displayed several highly conserved regions, including LDC and YRC motifs with cysteine residues for potential disulfide bridge formation, CPNKCG motif upstream of the Ig-like domain, GLTDYSFYRVW motif upstream of the putative N-linked glycosylation site, and a number of scattered cysteine residues. These distinctive features are very informative to pinpoint the important gene motifs and functions. The C-terminal regions, however, are more variable across species. Izumo1 cDNA sequences of goat, sheep, and cow were found to be largely homologous, and the molecular phylogenetic analysis is consistent with their morphological taxonomy. This implies the Izumo1 gene evolves from the same ancestor, and the mechanism of sperm-egg fusion in mammals may be under the same principle in which Izumo1 plays an important role.

Background: The extracellular calcium-sensing receptor (CaSR) belongs to family C of the G protein coupled receptors. Whether the CaSR is expressed in the pulmonary artery (PA) is unknown., Methods: The expression and distribution of CaSR were detected by RT-PCR, Western blotting and immunofluorescence. PA tension was detected by the pulmonary arterial ring technique, and the intracellular calcium concentration ([Ca2+]i) was detected by a laser-scanning confocal microscope., Results: The expressions of CaSR mRNA and protein were found in both rat pulmonary artery smooth muscle cells (PASMCs) and PAs. Increased levels of [Ca2+]o (extracellular calcium concentration) or Gd3+ (an agonist of CaSR) induced an increase of [Ca2+]i and PAs constriction in a concentration-dependent manner. In addition, the above-mentioned effects of Ca2+ and Gd3+ were inhibited by U73122 (specific inhibitor of PLC), 2-APB (specific antagonist of IP3 receptor), and thapsigargin (blocker of sarcoplasmic reticulum calcium ATPase)., Conclusions: CaSR is expressed in rat PASMCs, and is involved in regulation of PA tension by increasing [Ca2+]i through G-PLC-IP3 pathway.

CD9 is a glycoprotein of the transmembrane 4 superfamily (TM4SF) and is involved in various cellular processes. Some CD9 cDNA have been cloned in mammals and certain fish genera in recent years, but goat and sheep counterparts of cattle, human and mouse have not been identified. To facilitate the studies, we cloned the cDNA encoding for CD9 of cashmere goat (Capra hircus) and sheep (Ovis aries), and expressed sheep CD9 in Escherichia coli cells. Structural analysis indicated for both goat and sheep that a 1123 bp cDNA spanned an open reading frame of 681 bp which predicted a protein of 226 amino acids with a typical TM4SF structure, including four highly conserved transmembrane domains, two extracellular domains and a CCG motif, which is a hallmark of the TM4SF. The predicted amino acid sequences were highly homologous to those of cattle, mouse and human CD9. Molecular phylogenetic analysis based on CD9 cDNA sequences indicated that goat and sheep CD9 were closely related to CD9 of cattle, which is in agreement with their morphological taxonomy.

A comparison of the Vicia faba SCE with the human lymphocyte SCE test was made with regard to their capacity to detect mutagens. Twelve chemical agents that can induce sister-chromatid exchanges (SCEs) in both Vicia faba root tip cells and human lymphocytes were used. The results indicate that the plant SCE technique is a sensitive test for screening various chemical agents including food preservatives, pesticides and essences which may have mutagenic activity. A new simplified procedure was introduced for differential staining of sister chromatids in Vicia faba.