Search

Intrahepatic cholangiocarcinoma (ICC) is a rare but highly aggressive malignant tumor arising within the liver, with a 5-year survival rate of only 20–40% after surgery. The role of interleukin-8 (IL-8) in ICC progression remains elusive. A transcriptomic approach based on IL-8 stimulation first revealed significant upregulation of the prometastatic gene CD97 and key epithelial–mesenchymal transition (EMT) factors E-cadherin and vimentin. Immunohistochemistry of 125 ICC tissues confirmed the positive correlation between IL-8 and CD97. Multivariable Cox regression indicated that they are both independent predictors of ICC prognosis. Mechanistically, IL-8 treatment induced CD97 expression at 50 and 100 ng/ml in QBC-939 and QBE cells, respectively. Moreover, the induction of cell migration and invasion upon IL-8 treatment was attenuated by CD97 RNA interference, and the expression of EMT-associated genes was dramatically inhibited. To determine whether CXCR1 or CXCR2 are downstream effectors of IL-8, siCXCR2 was applied and shown to significantly attenuate the oncogenic effects of IL-8 by inhibiting the phosphorylation of PI3K/AKT. Finally, the induction of CD97 expression by the PI3K pathway was verified by treatment with the inhibitor LY294002. In vivo, the significant tumor growth and lung metastasis effects induced by intraperitoneal injection of IL-8 were greatly inhibited by silencing CD97 in nude mice. Collectively, the study presents a novel mechanism of the IL-8-CXCR2-PI3K/AKT axis in regulating CD97 expression, which leads to ICC metastasis mainly through EMT. The study may provide alternatives for targeting the tumor microenvironment in metastatic ICC. [ABSTRACT FROM AUTHOR]

Arbuscular mycorrhizal (AM) fungi enhance plant stress tolerance, but it is unclear whether AM fungi affect heat tolerance in cucumbers. This study aimed to analyze how an AM fungus, Diversispora versiformis, affected growth, chlorophyll, five osmolytes, and plasma membrane intrinsic proteins (PIPs) and heat shock protein 70 (Hsp70) gene expression in cucumber leaves after a short-term (80 h) heat stress. Heat treatment significantly reduced root AM fungal colonization rate (0.26 folds). Heat treatment also distinctly suppressed plant height, stem diameter, and biomass, whereas AM fungal inoculation improved these growth variables as well as the chlorophyll index, with the benefit being more obvious under heat than under no-heat stress conditions. Heat treatment triggered differential changes in osmolytes (sucrose, fructose, and betaine) of inoculated and uninoculated cucumbers, whereas inoculation with AM fungus significantly raised leaf sucrose, fructose, glucose, betaine, and proline levels when compared to non-AM fungal inoculation. Heat treatment increased the expression of two (CsPIP1;6 and CsPIP2;1) of eight CsPIPs in inoculated and uninoculated plants, whereas AM fungal inoculation up-regulated the expression of CsPIP1;6, CsPIP2;1, and CsPIP2;6 under heat stress conditions. Hsp70s expressed differently in inoculated and uninoculated plants under heat versus no-heat stress, with 6 of 11 CsHsp70s down-regulated in inoculated plants. Under heat stress conditions, AM fungus only up-regulated CsHsp70-8 expression in 11 Hsp70s, while another eight CsHsp70s were down-regulated. Heat treatment and AM fungal inoculation both increased the expression of CsHsp70-8 and CsPIP1;6. It was concluded that AM fungus-inoculated cucumbers have high levels of growth, chlorophyll, and osmolytes under heat stress and do not require high CsPIPs and CsHsp70s expression to tolerate a short-term heat treatment. [ABSTRACT FROM AUTHOR]

Preeclampsia (PE) differentially impairs female and male fetal endothelial cell function, which is associated with an increased risk of adult‐onset cardiovascular disorders in children born to mothers with PE. However, the underlying mechanisms are poorly defined. We hypothesize that dysregulation of microRNA‐29a‐3p and 29c‐3p (miR‐29a/c‐3p) in PE disturbs gene expression and cellular responses to cytokines in fetal endothelial cells in a fetal sex‐dependent manner. RT‐qPCR analysis of miR‐29a/c‐3p was performed on female and male unpassaged (P0) human umbilical vein endothelial cells (HUVECs) from normotensive (NT) pregnancies and PE. Bioinformatic analysis of an RNA‐seq dataset was performed to identify PE‐dysregulated miR‐29a/c‐3p target genes in female and male P0‐HUVECs. Gain‐ and loss‐of‐function assays were conducted to determine the effects of miR‐29a/c‐3p on endothelial monolayer integrity and proliferation in response to transforming growth factor‐β1 (TGFβ1) and tumour necrosis factor‐α (TNFα) in NT and PE HUVECs at passage 1. We observed that PE downregulated miR‐29a/c‐3p in male and female P0‐HUVECs. PE dysregulated significantly more miR‐29a/c‐3p target genes in female vs. male P0‐HUVECs. Many of these PE‐differentially dysregulated miR‐29a/c‐3p target genes are associated with critical cardiovascular diseases and endothelial function. We further demonstrated that miR‐29a/c‐3p knockdown specifically recovered the PE‐abolished TGFβ1‐induced strengthening of endothelial monolayer integrity in female HUVECs, while miR‐29a/c‐3p overexpression specifically enhanced the TNFα‐promoted cell proliferation in male PE HUVECs. In conclusion, PE downregulates miR‐29a/c‐3p expression and differentially dysregulates miR‐29a/c‐3p target genes associated with cardiovascular diseases and endothelial function in female and male fetal endothelial cells, possibly contributing to the fetal sex‐specific endothelial dysfunction observed in PE. Key points: Preeclampsia differentially impairs female and male fetal endothelial cell function in responses to cytokines. Pro‐inflammatory cytokines are elevated in maternal circulation during pregnancy in preeclampsia.MicroRNAs are critical regulators of endothelial cell function during pregnancy. We have previously reported that preeclampsia downregulated microRNA‐29a‐3p and 29c‐3p (miR‐29a/c‐3p) in primary fetal endothelial cells. However, it is unknown if PE differentially dysregulates the expression of miR‐29a/c‐3p in female and male fetal endothelial cells.We show that preeclampsia downregulates miR‐29a/c‐3p in male and female HUVECs and preeclampsia dysregulates cardiovascular disease‐ and endothelial function‐associated miR‐29a/c‐3p target genes in HUVECs in a fetal sex‐specific manner.MiR‐29a/c‐3p differentially mediate cell responses to cytokines in female and male fetal endothelial cells from preeclampsia.We have revealed fetal sex‐specific dysregulation of miR‐29a/c‐3p target genes in fetal endothelial cells from preeclampsia. This differential dysregulation may contribute to fetal sex‐specific endothelial dysfunction in offspring born to preeclamptic mothers. [ABSTRACT FROM AUTHOR]

Urological chronic pelvic pain syndrome (UCPPS) manifests as pelvic pain with frequent urination and has a 10% prevalence rate without effective therapy. Nanoceria (cerium oxide nanoparticles [CNPs]) were synthesized in this study to achieve potential long-term pain relief, using a commonly used UCPPS mouse model with cyclophosphamide-induced cystitis. Transcriptome sequencing analysis revealed that serpin family B member 2 (SerpinB2) was the most upregulated marker in mouse bladder, and SerpinB2 was downregulated with CNP pretreatment. The transcriptome sequencing analysis results agreed with quantitative polymerase chain reaction and western blot analysis results for the expression of related mRNAs and proteins. Analysis of Gene Expression Omnibus (GEO) datasets revealed that SerpinB2 was a differentially upregulated gene in human UCPPS. In vitro SerpinB2 knockdown downregulated proinflammatory chemokine expression (chemokine receptor CXCR3 and C-X-C motif chemokine ligand 10) upon treatment with 4-hydroperoxycyclophosphamide. In conclusion, CNP pretreatment may prevent the development of UCPPS, and reactive oxygen species (ROS) scavenging and SerpinB2 downregulation may modulate the immune response in UCPPS. [ABSTRACT FROM AUTHOR]

Objective: To analyze the expression of protein kinase C delta (PKCẟ, PRKCD) in Liver hepatocellular carcinoma and its clinical significance based on bioinformatics, in order to provide a new direction for the study of therapeutic targets for Liver hepatocellular carcinoma. Methods: The PRKCD gene expression data and patient clinical information data in Liver hepatocellular carcinoma tissues and adjacent tissues were downloaded from The TCGA (The Cancer Genome Atlas) and UCSC Xena databases. The mRNA expression of PRKCD in Liver hepatocellular carcinoma was analyzed, and the protein expression of PRKCD in Liver hepatocellular carcinoma was analyzed by Human Protein Atlas (HPA) databases. Furthermore, its relationship with clinicopathological features and prognosis in patients with Liver hepatocellular carcinoma was analyzed. GSEA enrichment analysis were carried out for PRKCD in Liver hepatocellular carcinoma. Finally, the causes of changes in PRKCD expression were analyzed from the perspective of genetic and epigenetics based on collated liver hepatocellular carcinoma data. Results: Both the mRNA and protein expression level of PRKCD gene in Liver hepatocellular carcinoma tissues was significantly higher than that in adjacent tissues. Its expression level was correlated with TNM stage, Histologic grade,Alpha fetoprotein (AFP) and Living status in clinicopathological features, and its expression level has certain clinical diagnostic accuracy. Kaplan-Meier analysis showed that the prognosis of patients with low PRKCD expression was significantly better than that with high PRKCD expression. Univariate and multivariate Cox regression analysis showed that PRKCD was an independent prognostic factor in patients with Liver hepatocellular carcinoma, and it was also found that the risk ratio of TNM stage III and stage IV survival curve (HR) was significantly greater than that of stage I and stage II. GSEA analysis showed that it was enriched in cell cycles and DNA replication, and was positively correlated with PRKCD. Enrichment analysis found that PRKCD is mainly involved in the chemokine signaling pathway, NOD-like receptor signaling pathway, PPAR signaling pathway, adipocytokine signaling pathway and T-cell receptor signaling pathway. Finally, through genetic and epigenetic analysis, it is found that the increase in the number of copies of genes will increase the expression level of PRKCD, and the methylation level is negatively correlated with the expression level of PRKCD. Conclusion: PRKCD gene is upregulated in Liver hepatocellular carcinoma tissues, and its overexpression level is closely related to patient poor prognosis. [ABSTRACT FROM AUTHOR]

Background: Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disease characterized by apoptotic clearance deficiency provoking autoimmune responses and leading to multiple organ damage. PPAR-δ, encoded by the PPARD gene, was induced in macrophages promoting the timely disposal of apoptotic cells. Biological studies had provided solid foundation of PPARD involvement in SLE; it is worthwhile to further explore the genetic contribution of PPARD to SLE.Methods: We performed a discovery-replication genetic association study. The discovery study was based on previous reported GWAS data. And the replication study was conducted in 1003 SLE patients and 815 healthy controls from Henan, Middle East of China. Further, we analyzed the eQTL effect to identify possible functional significance.Results: In the genetic association analysis, we observed significant association between the risk C allele of rs4713853 (p = 0.03, OR 1.167, 95% CI 1.015-1.341) and increased SLE susceptibility. Moreover, individuals with the risk C allele were associated with lower expression of PPARD and DEF6. Our clinical analysis showed that SLE patients with the risk C allele of rs4713853 were more likely to present a higher proportion of anti-Sm antibody presence (CC+CT vs. TT, 20.0% vs. 14.2%, p = 0.039) and higher level of Scr (median inter quarter range CC+CT vs. TT, 56 48-71 vs. 54 46-64 μmol/L, p = 0.002).Conclusions: In conclusion, our study identified a novel association between PPARD rs4713853 and SLE susceptibility in Chinese populations. By integrating multiple layers of analysis, we suggested that PPARD might be a main candidate in the pathogenesis of SLE. [ABSTRACT FROM AUTHOR]

Background: Gene expression patterns can be used as prognostic biomarkers in various types of cancers. We aimed to identify a gene expression pattern for individual distant metastatic risk assessment in patients with locoregionally advanced nasopharyngeal carcinoma.Methods: In this multicentre, retrospective, cohort analysis, we included 937 patients with locoregionally advanced nasopharyngeal carcinoma from three Chinese hospitals: the Sun Yat-sen University Cancer Center (Guangzhou, China), the Affiliated Hospital of Guilin Medical University (Guilin, China), and the First People's Hospital of Foshan (Foshan, China). Using microarray analysis, we profiled mRNA gene expression between 24 paired locoregionally advanced nasopharyngeal carcinoma tumours from patients at Sun Yat-sen University Cancer Center with or without distant metastasis after radical treatment. Differentially expressed genes were examined using digital expression profiling in a training cohort (Guangzhou training cohort; n=410) to build a gene classifier using a penalised regression model. We validated the prognostic accuracy of this gene classifier in an internal validation cohort (Guangzhou internal validation cohort, n=204) and two external independent cohorts (Guilin cohort, n=165; Foshan cohort, n=158). The primary endpoint was distant metastasis-free survival. Secondary endpoints were disease-free survival and overall survival.Findings: We identified 137 differentially expressed genes between metastatic and non-metastatic locoregionally advanced nasopharyngeal carcinoma tissues. A distant metastasis gene signature for locoregionally advanced nasopharyngeal carcinoma (DMGN) that consisted of 13 genes was generated to classify patients into high-risk and low-risk groups in the training cohort. Patients with high-risk scores in the training cohort had shorter distant metastasis-free survival (hazard ratio [HR] 4·93, 95% CI 2·99-8·16; p<0·0001), disease-free survival (HR 3·51, 2·43-5·07; p<0·0001), and overall survival (HR 3·22, 2·18-4·76; p<0·0001) than patients with low-risk scores. The prognostic accuracy of DMGN was validated in the internal and external cohorts. Furthermore, among patients with low-risk scores in the combined training and internal cohorts, concurrent chemotherapy improved distant metastasis-free survival compared with those patients who did not receive concurrent chemotherapy (HR 0·40, 95% CI 0·19-0·83; p=0·011), whereas patients with high-risk scores did not benefit from concurrent chemotherapy (HR 1·03, 0·71-1·50; p=0·876). This was also validated in the two external cohorts combined. We developed a nomogram based on the DMGN and other variables that predicted an individual's risk of distant metastasis, which was strengthened by adding Epstein-Barr virus DNA status.Interpretation: The DMGN is a reliable prognostic tool for distant metastasis in patients with locoregionally advanced nasopharyngeal carcinoma and might be able to predict which patients benefit from concurrent chemotherapy. It has the potential to guide treatment decisions for patients at different risk of distant metastasis.Funding: The National Natural Science Foundation of China, the National Science & Technology Pillar Program during the Twelfth Five-year Plan Period, the Natural Science Foundation of Guang Dong Province, the National Key Research and Development Program of China, the Innovation Team Development Plan of the Ministry of Education, the Health & Medical Collaborative Innovation Project of Guangzhou City, China, and the Program of Introducing Talents of Discipline to Universities. [ABSTRACT FROM AUTHOR]

Background: Talin-1 is a cytoskeletal protein that plays an important role in tumourgenesis, migration and metastasis in several malignant tumors. The aim of this study was to evaluate the expression and prognostic value of Talin-1 in nasopharyngeal carcinoma (NPC). Methods: Talin-1 mRNA and protein expression were examined in NPC cell lines and clinical nasopharyngeal tissues by quantitative RT-PCR, agarose gel electrophoresis and western blotting. The expression of Talin-1 was analyzed by immunohistochemical staining in 233 paraffin-embedded NPC specimens with clinical follow-up data and cox regression analysis was used to identify independent prognostic factors. The functional role of Talin-1 in NPC cell lines was evaluated by small interfering RNA-mediated depletion of the protein followed by the wound healing and transwell invasion assays. Results: The expression of Talin-1 was significantly upregulated in most NPC cell lines and clinical tissues at both the mRNA and protein levels. High expression of Talin-1 was significantly associated with distant metastasis (P= 0.001) and patient death (P= 0.001). In addition, high expression of Talin-1 was associated with significantly poorer overall survival (OS: HR, 2.15; 95% CI, 1.28-3.63; P= 0.003) and poorer distant metastasis-free survival (DMFS: HR, 2.39; 95% CI, 1.38-4.15; P= 0.001). Cox regression analysis indicated that high expression of Talin-1 and TNM stage were independent prognostic indicators (both P < 0.05). Stratified analysis demonstrated that high expression of Talin-1 was associated with significantly poorer survival in patients with advanced stage disease (stage III-IV, HR, 1.91; 95% CI, 1.09-3.35; P = 0.02 for OS and HR, 2.22; 95% CI, 1.24-3.99; P = 0.006 for DMFS). Furthermore, the depletion of Talin-1 suppressed the migratory and invasive ability of NPC cells in vitro. Conclusions: Our data demonstrate that high expression of Talin-1 is associated with significantly poorer OS and poorer DMFS in NPC and depletion of Talin-1 expression inhibited NPC cell migration and invasion. Talin-1 may serve as novel prognostic biomarker in NPC. [ABSTRACT FROM AUTHOR]

Cancer Targeting Gene-Viro-Therapy (CTGVT) is a promising cancer therapeutical strategy that strengthens the anti-tumour effect of oncolytic virus by expressing inserted foreign anti-tumour genes. In this work, we constructed a novel adenoviral vector controlled by the tumour-specific survivin promoter on the basis of the ZD55 vector, which is an E1B55KD gene deleted vector we previously constructed. Compared with the original ZD55 vector, this new adenoviral vector (ZD55SP/E1A) showed much better ability of replication and reporter gene expression. We then combined anti-tumour gene interleukine-24 (IL-24) with an RNA polymerase III-dependent U6 promoter driving short hairpin RNA (shRNA) that targets M-phase phosphoprotein 1 (MPHOSPH1, a newly identified oncogene) by inserting the IL-24 and the shRNA of MPHOSPH1 (shMPP1) expression cassettes into the new ZD55SP/E1A vector. Our results demonstrated excellent anti-tumour effect of ZD55SP/E1A-IL-24-shMPP1 in vitro on multiple cancer cell lines such as lung cancer, liver cancer and ovarian caner. At high multiplicity-of-infection (MOI), ZD55SP/E1A-IL-24-shMPP1 triggered post-mitotic apoptosis in cancer cells by inducing prolonged mitotic arrest; while at low MOI, senescence was induced. More importantly, ZD55SP/E1A-IL-24-shMPP1 also showed excellent anti-tumour effects in vivo on SW620 xenograft nude mice. In conclusion, our strategy of constructing an IL-24 and shMPP1 dual gene expressing oncolytic adenoviral vector, which is regulated by the survivin promoter and E1B55KD deletion, could be a promising method of cancer gene therapy. [ABSTRACT FROM AUTHOR]

In an effort to gain deeper insight into the molecular processes underlying neurodegeneration in Parkinson's disease, we performed gene expression profiling at several early time points after MPTP-injection into old (1-year) mice. We used a PCR-based gene expression profiling method, digital expression pattern display (DEPD), a method of very high sensitivity and reproducibility, which displays almost all transcripts of a tissue. To identify cell death-associated genes, we defined clusters of differentially expressed transcripts with expression behaviour that correlated with the temporal profile of cell death progression and characterized one of these cell death clusters further. We selected one of the strongest regulated genes, the serum and glucocorticoid-regulated kinase 1 (sgk1), and validated its differential expression by Northern blot analysis, semiquantitative PCR andin situhybridization. Up-regulation ofsgk1(i) coincides with the onset of dopaminergic cell death in both the 8-week acute and 1-year subacute MPTP models, (ii) spans the entire brain, (iii) is attenuated by thel-deprenyl-mediated inhibition of the MPTP conversion to its active metabolite MPP+ and (iv) is not induced by dehydration. This study demonstrated that the combination of the DEPD technology, clustering analysis and a detailed histopathology is a useful tool for elucidating molecular pathways in neurodegenerative diseases. [ABSTRACT FROM AUTHOR]

Parkin is an E3 ubiquitin ligase causally involved in the pathogenesis of autosomal recessive juvenile parkinsonism. In this paper, we analysed the formation of alternative splice products and the spatio-temporal expression pattern of parkin during pre- and postnatal mouse development. Using RT-PCR, Northern blot, in situ hybridization, Western blot analysis, and immunohistochemistry we found (i) alternative splice forms of parkin; (ii) an early and widespread expression of parkin mRNA and protein in the CNS and several organs, already at E10/12; (iii) a marked increase in expression level during midgestational development (E15–18) in the CNS, followed by a steady increase until adulthood; (iv) an ubiquitous distribution throughout CNS ontogeny. Our results show that parkin expression is correlated with cell maturation and suggests an important physiological role of parkin in neurons that is at no time limited to the dopaminergic system. [Copyright &y& Elsevier]

Background: Alternative polyadenylation (APA) is a widespread phenomenon in the posttranscriptional regulation of gene expression that generates mRNAs with alternative 3′-untranslated regions (3'UTRs). APA contributes to the pathogenesis of various diseases, including cancer. However, the potential role of APA in the development of nasopharyngeal carcinoma (NPC) remains largely unknown. Methods: A strategy of sequencing APA sites (SAPAS) based on second-generation sequencing technology was carried out to explore the global patterns of APA sites and identify genes with tandem 3'UTRs in samples from 6 NPC and 6 normal nasopharyngeal epithelial tissue (NNET). Sequencing results were then validated using quantitative RT-PCR in a larger cohort of 16 NPC and 16 NNET samples. Results: The sequencing data showed that the use of tandem APA sites was prevalent in NPC, and numerous genes with APA-switching events were discovered. In total, we identified 195 genes with significant differences in the tandem 3'UTR length between NPC and NNET; including 119 genes switching to distal poly (A) sites and 76 genes switching to proximal poly (A) sites. Several gene ontology (GO) terms were enriched in the list of genes with switched APA sites, including regulation of cell migration, macromolecule catabolic process, protein catabolic process, proteolysis, small conjugating protein ligase activity, and ubiquitin-protein ligase activity. Conclusions: APA site-switching events are prevalent in NPC. APA-mediated regulation of gene expression may play an important role in the development of NPC, and more detailed studies targeting genes with APA-switching events may contribute to the development of novel future therapeutic strategies for NPC. [ABSTRACT FROM AUTHOR]

• Whether and how soil AMF enhance heat stress tolerance of cucumber were studied. • Heat stress decreased root AMF colonization and soil hyphal length. • AMF increased cucumber biomass and leaf gas exchange under heat stress. • Most cshsp70s and cspips were up-regulated in inoculated plants under HS versus ST. • Under HS, AMF up-regulated expressions of 11 of 12 cshsp70s and 12 of 14 cspips. Cucumbers are frequently subjected to heat stress (HS) during the summer, which severely limits their growth and fruit yield and quality. Soil arbuscular mycorrhizal fungi (AMF) can enhance the abiotic stress tolerance of host plants, but it is unclear whether and how soil arbuscular mycorrhizal fungi (AMF) enhance the HS tolerance of cucumber. This work aimed to assess the effects of an arbuscular mycorrhizal fungus, Diversispora versiformis , on plant growth, leaf gas exchange, and expressions of plasma membrane intrinsic proteins (PIPs) and heat shock protein 70 (Hsp70) genes in roots of cucumber under HS (38 °C/30 °C, day/night temperature, 16 h/8 h) and suitable temperature (ST) (25 °C/18 °C, day/night temperature, 16 h/8 h) conditions. The HS treatment significantly decreased root mycorrhizal colonization and soil mycelium length by 0.26- and 0.55-fold, respectively, compared with the ST treatment. The biomass inhibition of cucumber by HS versus ST was more pronounced in uninoculated plants than inoculated plants. Inoculation with D. versiformis , however, significantly increased cucumber biomass and leaf gas exchange under both ST and HS conditions when compared to the uninoculated treatment. The HS treatment had almost no effect on or down-regulated the expression of CsHsp70s and CsPIPs genes in uninoculated plants, while most CsHsp70s and CsPIPs gene expressions were up-regulated in inoculated plants under HS versus ST conditions, suggesting the active heat resistance of mycorrhizal plants. Additionally, under ST conditions, D. versiformis up-regulated CsHsp70–11, CsPIP1;2 , and CsPIP2;6 expressions. Under HS conditions, D. versiformis increased the expression of 11 of 12 CsHsp70s by 1.17‒6.14 folds as well as 12 of 14 CsPIPs by 1.82‒144.43 folds, with CsPIP1.5 and CsPIP2;7 up-regulated up to 144.43- and 82.46-fold, respectively. It is concluded that D. versiformis -inoculated cucumbers exhibited higher heat tolerance than uninoculated plants, associated with up-regulated expression of most CsHsp70s and CsPIPs genes. [ABSTRACT FROM AUTHOR]

Abstract Mutations in a protein designated Parkin were shown to be involved in the pathogenesis of autosomal recessive juvenile parkinsonism. Nothing is known about its regional and subcellular distribution in the mouse. In order to elucidate the Parkin mRNA and protein distribution in the adult mouse, the mouse cDNA was cloned and polyclonal antisera were generated against the N-terminal part of mouse Parkin. The antibodies were shown to be specific using Western blot analysis, immunostaining of cells transfected with mouse Parkin and pre-absorption tests. The Parkin protein expression profile was studied using immunohistochemistry and Western blot analysis and was compared with that of the mRNA yielded by in situ hybridization and RT-PCR analysis. Parkin protein was widely distributed in all subdivisions of the mouse brain. Low levels were found in the telencephalon and diencephalon, while the brainstem contained a large number of cells heavily expressing Parkin. Ultrastructural analysis and double immunohistochemistry revealed that the majority of Parkin-expressing cells were neurons, while only single glial cells exhibited immunostaining. The protein was distributed nonhomogeneously throughout the entire cytoplasm. A subpopulation of Parkin-immunopositive cells displayed speckled immunodeposits in the nucleus. Dopaminergic cells of the substantia nigra pars compacta exhibited high levels of Parkin mRNA but no Parkin protein, while the striatum contained immunopositive profiles but no mRNA signals. Our data indicate that Parkin is neither restricted to a single functional system nor associated with a particular transmitter system. The speckled nuclear distribution of Parkin immunoreactivity strongly suggests a role for Parkin in gene expression. [ABSTRACT FROM AUTHOR]

HMG-CoA reductase (HMGCR) is the rate-limiting enzyme in cholesterol de novo biosynthesis and its degradation may bring therapeutic benefits for the treatment of cardiovascular disease (CVD) and nonalcoholic steatohepatitis (NASH). Before, we disclosed compound HMG499 as a potent HMGCR degrader, which could be a promising agent for treating CVD, however its side-effect of promoting cholesterol accumulation in cells should be eliminated before progression. Herein, a series of novel heterocyclic ring-fused analogs of HMG499 were synthesized and investigated for their activities of stimulating HMGCR degradation using a HMGCR (TM1-8)-GFP reporting system. Among them, the most active compound 29 (QH536) showed an EC 50 of 0.22 μΜ in promoting HMGCR degradation, which was about 2 times more potent than HMG499 (EC 50 = 0.43 μM). Interestingly, 29 was different from HMG499, it had no side-effect of inducing cholesterol accumulation in cells. Mechanistic studies disclosed that 29 could significantly decrease statin-induced accumulation of HMGCR protein via ubiquitination and degradation of HMGCR through ubiquitin-proteasome pathway and inhibit the cholesterol biosynthesis in cells. Therefore, these heterocyclic ring-fused analogs could be used as promising leads for the development of new types of agents against CVD. Furthermore, 29 also lowered cholesterol levels and suppressed TGFβ1-induced proliferation of LX-2 hepatic stellate cells in a dose-dependent manner. In particular, 29 not only decreased the NASH associated fibrotic mRNA and protein expression of α-SMA, COL1A1, TIMP1 and TGFβ1 but also suppressed cholesterol levels and inflammatory genes of TNF-α, IL-6 an IL-1β in RAW264.7 macrophage cells, indicating that 29 may bring therapeutic benefit to treat NASH. A series of heterocyclic ring-fused analogs of HMG499 were synthesized and their activities of promoting HMG-CoA reductase degradation and side-effects of inducing cholesterol accumulation in cells were evaluated. [Display omitted] • A series of novel heterocyclic ring-fused analogs of HMG499 were synthesized. • The activities of promoting HMGCR degradation of these analogs were evaluated. • Compound 29 is the most potent HMGCR degrader. • Compound 29 possessed no side-effect of inducing cholesterol accumulation. [ABSTRACT FROM AUTHOR]

Recent evidence has indicated that miRNAs play important roles in carcinogenesis. The identification of dysregulated miRNAs and the target genes they regulate might enhance our understanding of the molecular mechanisms of nasopharyngeal carcinoma (NPC). A microarray analysis was performed to identify dysregulated miRNAs in NPC tissue samples, and protein-coding genes targeted by three or more downregulated miRNAs were selected using miRWalk and used in a pathway enrichment analysis. Nineteen KEGG pathways were selected by DAVID, including the MAPK, focal adhesion, gap junction, ECM–receptor interaction, TGF-beta, and p53 signalling pathways, most of which are involved in NPC carcinogenesis and progression. MiR-143 was significantly downregulated in NPC cell lines and clinical samples. The ectopic expression of miR-143 suppressed NPC cell viability, colony formation, and anchorage-independent growth in vitro, and it inhibited xenograft tumour growth in vivo. Furthermore, KRAS was confirmed as a direct target of miR-143, and silencing KRAS expression suppressed NPC cell viability and proliferation. The miR-143/ KRAS pathway provides new insight into the molecular mechanisms that regulate the development and progression of NPC, and it provides novel therapeutic targets for NPC. [ABSTRACT FROM AUTHOR]