Corosolic acid inhibits metastatic response of human renal cell carcinoma cells by modulating ERK/MMP2 signaling.

Corosolic acid (CA), a plant-derived pentacyclic triterpenoid, has potent anti-inflammatory, anti-metabolic, and anti-neoplastic actions against a variety of human cancers. However, the specific mechanism by which CA inhibits the progression of renal cell carcinoma (RCC) is yet unclear. We found that CA (≤8 µM) had no influence on either the growth or viability of RCC cell lines (786-O, ACHN, and Caki-1) or normal HK2 cells. However, in a dose-dependent manner, CA prevented the invasion and migration of RCC cells. Human protease array analysis showed that CA reduced MMP2 expression. At increasing concentrations of CA, the expression of MMP2 was dose-dependently reduced, as shown by western blot and RT-PCR analyses as well as immunofluorescence staining. CA also stimulated ERK1/2 phosphorylation in 786-O and Caki-1 cells. Transfection of CA-treated RCC cells with siRNA-ERK restored MMP2 protein expression and the motility and invasion capabilities of RCC cells. Molecular docking study results showed that CA and MMP2 interact strongly. These findings elucidate the mechanism by which CA prevents RCC cells from migrating and invading, and these findings indicate that CA may be a potential anti-metastatic therapy for RCC.

Nurses' knowledge of and attitude toward postoperative patient-controlled analgesia (PCA) and the associated factors.

BACKGROUND: Postoperative pain control is pivotal for surgical care; it facilitates patient recovery. Although patient-controlled analgesia (PCA) has been available for decades, inadequate pain control remains. Nurses' knowledge of and attitude toward PCA may influence the efficacy on clinic application. PURPOSE: The purpose of this study is to evaluate nurses' knowledge of and attitude toward postoperative PCA and investigate the associated factors. METHODS: This is a cross-sectional study. We enrolled registered nurses from a 2200-bed medical center in northern Taiwan within one year. The participants completed an anonymous self-reported PCA knowledge inventory and PCA attitude inventory. Data were analyzed descriptively and associated were tested using logistic regression. RESULTS: With 303 participants enrolled, we discovered that nurses had limited knowledge of and a negative attitude toward PCA. Under half of the participants know how to set up a bolus dose and lockout intervals. The majority held misconceptions regarding side effect management for opioids. The minority agree to increase the dose when a patient experienced persistent pain or suggested the use of PCA. Surprisingly, participants with a bachelor's or master's degree had lower knowledge scores than those with a junior college degree. Those with 6-10 years of work experience also are lower than those with under 5 years of experience. However, the participants with experience of using PCA for patient care had higher knowledge scores and a more positive attitude. CONCLUSIONS: Although postoperative PCA has been available for decades and education programs are routinely provided, nurses had limited knowledge of and a negative attitude toward PCA. A higher education level and longer work experience were not associated with more knowledge. The current education programs on PCA should be revised to enhance their efficacy in delivering up-to-date knowledge and situation training which may convey supportive attitude toward clinical application of PCA.

Evaluation of the awareness of Western Australian SunSmart campaigns between 2008 and 2022.

ISSUE ADDRESSED: It is unknown whether SunSmart health promotion campaigns in Western Australia are still effectively reaching their target audience of young people (under 45 years). This study examined trends over time in awareness, relevancy and believability of SunSmart advertisements and identified socio-demographic characteristics and risk factors associated with campaign awareness. METHOD: Linear regression and log-binomial modelling were undertaken using data from the annual SunSmart post-campaign evaluation surveys between 2008/2009 and 2021/2022. SunSmart campaigns were analysed and categorised into the following themes: (1) personal real-life stories; (2) daily activities/sun exposure leads to skin cancer; or (3) cartoon/animated. RESULTS: Between 2008 and 2022, there were declines in total awareness (74.2% to 20.4%), unprompted awareness (33.7% to 4.9%) and relevancy (89.5% to 54.8%) of SunSmart advertisements (representing annual percent decreases of 3.6%, 3.1% and 1.8%, respectively). However, believability remained high over time (>94% in each annual survey). Trends were inconsistent between the awareness of campaign themes and socio-demographic characteristics and risk factors. Several campaigns had greater awareness in their subsequent years, compared with the first campaign year. CONCLUSION: In more recent years, SunSmart advertisements and campaigns may not have reached their target audience. In addition to socio-demographic characteristics, particularly age, advertisement factors may also affect the awareness of specific campaigns. SO WHAT?: Given the changing advertising landscape and its rising costs, ongoing funding is pertinent to increase the reach of future SunSmart campaigns. Increasing advertisements on alternative platforms and designing campaigns which separately target adolescents and adults need to be considered.

Loss of α7nAChR enhances endothelial-to-mesenchymal transition after myocardial infarction via NF-κB activation.

The myocardial fibrosis in response to myocardial infarction (MI) is closely related to the dysbalance of endothelial-to-mesenchymal transition (EndMT). Although numerous reports indicate that α7 nicotinic acetylcholine receptor (α7nAChR) activates the cholinergic anti-inflammatory pathway (CAP) to regulate the magnitude of inflammatory responses, the role of α7nAChR in myocardial fibrosis, as well as the underlying mechanisms, have not been elucidated. In this study, we evaluated cardiac function, fibrosis, and EndMT signaling using a mouse model of MI and interleukin (IL)-1ß-induced human cardiac microvascular endothelial cells (HCMECs). In vivo, α7nAChR deletion increased cardiac dysfunction, exacerbated the cardiac inflammatory response, and NF-κB activation, and enhanced EndMT, as shown by higher expression levels of fibroblast markers (FSP-1, α-SMA, collagen I, Snail) and decreased levels of the FGFR1, glucocorticoid receptor (GR) and endothelial marker (CD31) compared to wild-type mice. In vitro, the pharmacological activation of α7nAChR with PNU282987 significantly inhibited IL-1ß-induced EndMT, as shown by a reduced transition to the fibroblast-like phenotype and the expression of fibrotic markers. Moreover, the IL-1ß-mediated activation of NF-κB pathway was suppressed by PNU282987. This anti-EndMT effect of α7nAChR was associated with regulation of Snail. Furthermore, Western blot analysis further revealed that the GR antagonist RU38486 could partially counteract the effect of PNU282987 on NF-κB expression. In conclusion, our results show that α7nAChR is involved in cardiac fibrosis by inhibiting EndMT, providing a novel approach to the treatment of MI.

Secondary structures, dynamics, and DNA binding of the homeodomain of human SIX1.

Human sine oculis homeobox homolog (SIX) 1 contains a homeodomain (HD), which is important for binding to DNA. In this study, we carried out structural studies on the HD of human SIX1 using nuclear magnetic resonance (NMR) spectroscopy. Its secondary structures and dynamics in solution were explored. HD is well-structured in solution, and our study shows that it contains three α-helices. Dynamics study indicates that the N- and C-terminal residues of HD are flexible in solution. HD of human SIX1 exhibits molecular interactions with a short double-strand DNA sequence evidenced by the 1 H-15 N-heteronuclear single quantum correlation (HSQC) and 19 F-NMR experiments. Our current study provides structural information for HD of human SIX1. Further studies indicate that this construct can be utilized to study SIX1 and DNA interactions.

Curcumin enhances p-cresyl sulfate-induced cytotoxic effects on renal tubular cells.

Indoxyl sulfate (IS) and p-cresyl sulfate (PCS), protein-bound uremic toxins, can induce oxidative stress and cause renal disease progression. However, the different cytotoxic effects on renal cells between IS and PCS are not stated. Due to uremic toxins are generally found in CKD patients, the mechanisms of uremic toxins-induced renal injury are required to study. Curcumin has anti-oxidant, anti-inflammatory and anti-apoptotic effects which may be potential used to protect against renal damage. In contrast, curcumin also exert cytotoxic effects on various cells. In addition, curcumin may reduce or enhance cytotoxicity combined with different chemicals treatments. However, whether curcumin may influence uremic toxins-induced renal injury is unclear. The goal of this study is to compare the different cytotoxic effects on renal cells between IS and PCS treatment, as well as the synergistic or antagonistic effects by combination treatments with curcumin and PCS. Our experimental result shows the PCS exerts a stronger antiproliferative effect on renal tubular cells than IS treatment. In addition, our study firstly demonstrates that curcumin enhances PCS-induced cell cytotoxicity through caspase-dependent apoptotic pathway and cell cycle alteration.

Analysis of EZH2 Genetic Variants on Triple-Negative Breast Cancer Susceptibility and Pathology.

Triple-negative breast cancer (TNBC) is the third most common female cancer in Taiwan. EZH2 plays an important role in cancer development through transcriptional repression by chromatin remodeling. However, the expression of EZH2 in breast cancer is highly correlated with tumorigenesis, and patient survival is not matched to TNBC. Furthermore, it has not been determined if specific EZH2 genetic variants are associated with breast cancer risk. In this paper, we evaluated the survival of different types of breast cancer. The results indicated that a lower expression of EZH2 led to poor survival of TNBC patients. Therefore, we aimed at studying the relationship between genetic polymorphisms of EZH2 and susceptibility to TNBC in Taiwan. Four single-nucleotide polymorphisms (SNPs) of EZH2 (rs6950683, rs2302427, rs3757441, and rs41277434) were analyzed by real-time PCR genotyping in 176 patients with TNBC and 1000 cancer-free controls. The results showed that TNBC patients under 60 years old who carried a TC or CC genotype at EZH2 rs6950683 and re3757441 had a tumor size of 20 mm or smaller (T1). Thus, this study is the first to examine the age and mutant genes associated with EZH2 SNPs in TNBC progression and development in Taiwan.

[Sleep-improving mechanism of Chaiqin Ningshen Granules in insomnia rats: based on hippocampal metabonomics].

With the ultra high performance liquid chromatography-quadruple-electrostatic field orbitrap high resolution mass spectrometry(UHPLC-Q Exactive Orbitrap-MS)-based metabonomics technology, this study aims to analyze the effect of Chaiqin Ningshen Granules(CNG) on endogenous metabolites in insomnia rats of liver depression syndrome and explore the sleep-improving mechanism of this prescription. Parachlorophenylalanine(PCPA, ip) and chronic stimulation were combined to induce insomnia of liver depression pattern in rats, and the effect of CNG on the macroscopic signs, hemorheology, and neurotransmitters in the hippocampus of insomnia rats of liver depression syndrome was observed. After the administration, rat hippocampus was collected for liquid chromatography-mass spectrometry(LC-MS) analysis of the metabolomics. Principal component analysis(PCA), partial least squares discriminant analysis(PLS-DA), and orthogonal partial least squares discriminant analysis(OPLS-DA) were employed for analyzing the metabolites in rat hippocampus and screening potential biomarkers. MetPA was used to yield the related metabolic pathways and metabolic networks. The results show that the drugs can significantly improve the mental state, liver depression, and blood stasis of rats, significantly increase the content of 5-hydroxytryptamine(5-HT) and gamma aminobutyric acid(GABA) in hippocampus(except low-dose CNG), and significantly reduce the content of glucose(Glu)(except low-dose CNG). Among them, estazolam and high-dose CNG had better effect than others. Metabolomics analysis yielded 27 potential biomarkers related to insomnia. MetPA analysis showed 4 metabolic pathways of estazolam in intervening insomnia and 3 metabolic pathways of high-dose CNG in intervening insomnia, involving purine metabolism, glycerophospholipid metabolism, histidine metabolism, and caffeine metabolism. CNG can alleviate insomnia by regulating endogenous differential metabolites and further related metabolic pathways. The result lays a basis for further elucidating the mechanism of CNG in improving sleep.

CISD1 protects against atherosclerosis by suppressing lipid accumulation and inflammation via mediating Drp1.

Atherosclerosis still remains the leading cause of morbidity and mortality worldwide, and deeper understanding of target signaling that protect from the atherosclerosis progression may provide novel therapeutic strategies. CDGSH iron-sulfur domain-containing protein 1 (CISD1) is a protein localized on the outer membrane of mitochondria, and plays key roles in regulating cell death and oxidative stress. However, its potential on atherosclerosis development and the underlying mechanisms are largely unknown. Here, in our study, we found markedly decreased CISD1 expression in lipid-laden THP1 macrophages. Notably, lentivirus (LV)-mediated CISD1 over-expression remarkably ameliorated lipid deposition in macrophages stimulated by ox-LDL. Furthermore, cellular total ROS and mitochondrial ROS generation, and impairment of mitochondrial membrane potential (MMP) were highly induced by ox-LDL in THP1 cells, while being considerably reversed upon CISD1 over-expression. Inflammatory response caused by ox-LDL was also significantly restrained in macrophages with CISD1 over-expression. Mechanistically, we found that CISD1 could interact with dynamin-related protein 1 (Drp1). Intriguingly, CISD1-improved mitochondrial dysfunction and inflammation in ox-LDL-treated macrophages were strongly abolished by Drp1 over-expression, indicating that Drp1 suppression might be necessary for CISD1 to perform its protective effects in vitro. In high fat diet (HFD)-fed apolipoprotein E-deficient (ApoE-/-) mice, tail vein injection of lentiviral vector expressing CISD1 remarkably decreased atherosclerotic lesion area, serum LDL cholesterol levels and triglyceride contents. Inflammatory response, cellular total and mitochondrial ROS production, and Drp1 expression levels in aorta tissues were also dramatically ameliorated in HFD-fed ApoE-/- mice, contributing to the inhibition of atherosclerosis in vivo. Therefore, improving CISD1 expression may be a novel therapeutic strategy for atherosclerosis treatment.

Complementary effects of pine bark extract supplementation on inattention, impulsivity, and antioxidative status in children with attention-deficit hyperactivity disorder: A double-blinded randomized placebo-controlled cross-over study.

The purpose of this study was to investigate the complementary effects of polyphenolic compounds from pine bark extract (PE) as a strong antioxidative substrate on the symptoms of inattention and impulsivity in children with attention-deficit hyperactivity disorder (ADHD). This was a randomized, double-blind, crossover, placebo-controlled study that included two experimental units (4 weeks with PE supplementation and 4 weeks with placebo supplementation) separated by a 2-week washout period. ADHD participants were supplemented with 25 mg or 50 mg PE. We recruited 20 participants (17 boys and 3 girls) with a mean age of 10.0 ± 2.1 years. PE supplementation caused a significant reduction in the inattention and hyperactivity-impulsivity items of SNAP-IV. During the period of PE supplementation, the item of commissions in the Continuous Performance Test III (CPT III) significantly decreased, which was used to evaluate the symptoms of inattention and impulsivity. In addition, the erythrocytic reduced glutathione/oxidized glutathione ratio significantly increased, and the plasma TBARs level significantly decreased after 4 weeks of PE supplementation. However, there was no significant correlation between CPT III (commission) and antioxidative status indictors. PE supplementation may have potential effects of ameliorating inattention and impulsivity, and elevating the antioxidative status in children with ADHD.

C-reactive protein derived from perivascular adipose tissue accelerates injury-induced neointimal hyperplasia.

AIM: Inflammation within the perivascular adipose tissue (PVAT) in obesity plays an important role in cardiovascular disorders. C-reactive protein (CRP) level in obesity patients is significantly increased and associated with the occurrence and progression of cardiovascular disease. We tested the hypothesis CRP derived from PVAT in obesity contributes to vascular remodeling after injury. METHODS: A high-fat diet (HFD) significantly increased CRP expression in PVAT. We transplanted thoracic aortic PVAT from wild-type (WT) or transgenic CRP-expressing (CRPTG) mice to the injured femoral artery in WT mice. RESULTS: At 4 weeks after femoral artery injury, the neointimal/media ratio was increased significantly in WT mice that received PVAT from CRPTG mice compared with that in WT mice that received WT PVAT. Transplanted CRPTG PVAT also significantly accelerated adventitial macrophage infiltration and vasa vasorum proliferation. It was revealed greater macrophage infiltration in CRPTG adipose tissue than in WT adipose tissue and CRP significantly increased the adhesion rate of monocytes through receptor Fcγ RI. Proteome profiling showed CRP over-expression promoted the expression of chemokine (C-X-C motif) ligand 7 (CXCL7) in adipose tissue, transwell assay showed CRP increased monocyte migration indirectly via the induction of CXCL7 expression in adipocytes. CONCLUSION: CRP derived from PVAT was significantly increased in HFD mice and promoted neointimal hyperplasia after vascular injury.

Chronic hepatitis B virus infection.

Chronic hepatitis B virus infection is a global public health threat that causes considerable liver-related morbidity and mortality. It is acquired at birth or later via person-to-person transmission. Vaccination effectively prevents infection and chronic hepatitis B virus carriage. In chronically infected patients, an elevated serum hepatitis B virus DNA concentration is the main risk factor for disease progression, although there are other clinical and viral parameters that influence disease outcomes. In addition to liver biochemistry, virological markers, and abdominal ultrasonography, non-invasive assessment of liver fibrosis is emerging as an important assessment modality. Long-term nucleos(t)ide-analogue therapy is safe and well tolerated, achieves potent viral suppression, and reduces the incidence of liver-related complications. However, a need to optimise management remains. Promising novel therapies are at the developmental stage. With current vaccines, therapies, and an emphasis on improving linkage to care, WHO's goal of eliminating hepatitis B virus as a global health threat by 2030 is achievable.

Expression, purification of Zika virus membrane protein-NS2B in detergent micelles for NMR studies.

The Zika virus (ZIKV) genome encodes a polyprotein that can be post-translationally processed into functional viral proteins. The viral protease is indispensable in the maturation of viral proteins. The Zika protease comprises of two components crucial for catalysis. The N-terminal region of NS3 contains the catalytic triad and approximately 40 amino acids of NS2B are essential for folding and protease activity. NS2B is a membrane protein with transmembrane domains that are critical for the localization of NS3 to the membrane. In this study, we expressed and purified full-length NS2B from ZIKV in E. coli. Purified NS2B was then reconstituted into lyso-myristoyl phosphatidylglycerol (LMPG) micelles. It was found that compared to wild type NS2B, NS2B C11S mutation in LMPG exhibited dispersed cross peaks in the 1H15N-HSQC spectrum, thereby suggesting the feasibility for structural characterization using solution NMR spectroscopy.

Opportunities and barriers for providing HIV testing through community health centers in mainland China: a nationwide cross-sectional survey.

BACKGROUND: Primary care may be an avenue to increase coverage of HIV testing but it is unclear what challenges primary healthcare professionals in low- and middle-income countries face. We describe the HIV testing practices in community health centres (CHCs) and explore the staff's attitude towards further development of HIV testing services at the primary care level in China. METHODS: We conducted a national, cross-sectional survey using a stratified random sample of CHCs in 20 cities in 2015. Questionnaires were completed by primary care doctors and nurses in CHCs, and included questions regarding their demographics, clinical experience and their views on the facilitators and barriers to offering HIV testing in their CHC. Multivariate logistic regression was conducted to examine the association between staff who would offer HIV testing and their sociodemographic characteristics. RESULTS: A total of 3580 staff from 158 CHCs participated. Despite the majority (81%) agreeing that HIV testing was an important part of healthcare, only 25% would provide HIV testing when requested by a patient. The majority (71%) were concerned about reimbursement, and half (47%) cited lack of training as a major barrier. Almost half (44%) believed that treating people belonging to high-risk populations would scare other patients away, and 6% openly expressed their dislike of people belonging to high-risk populations. Staff who would offer HIV testing were younger (adjusted odds ratio (aOR) 0.97 per year increase in age, 95% confidence interval (CI):0.97-0.98); trained as a doctor compared to a nurse (aOR 1.79, 95%CI:1.46-2.15); held a bachelor degree or above (aOR 1.34, 95%CI:1.11-1.62); and had previous HIV training (aOR 1.55, 95%CI:1.27-1.89). CONCLUSIONS: Improving HIV training of CHC staff, including addressing stigmatizing attitudes, and improving financial reimbursement may help increase HIV testing coverage in China.

High-Fat Diet Aggravates the Intestinal Barrier Injury via TLR4-RIP3 Pathway in a Rat Model of Severe Acute Pancreatitis.

OBJECTIVE: For patients with severe acute pancreatitis (SAP), a high body mass index (BMI) increases the possibility of infection derived from the intestine. In this study, we evaluate whether TAK242 can alleviate severe acute pancreatitis-associated injury of intestinal barrier in high-fat diet-fed rats. METHODS: A SAP model was established by retrograde injection of 5% sodium taurocholate into the biliary-pancreatic duct. Thirty Sprague-Dawley (SD) adult rats were randomly divided into five groups: standard rat chow (SRC) normal (SN), SRC SAP (SAP), high-fat diet normal (HN), HFD SAP (HSAP), and TLR4 inhibitor pretreatment HFD SAP (HAPT) groups. Intraperitoneal injection of 3 mg/kg TAK242 was administered 30 minutes before SAP model establishment in the HAPT group. Rats were sacrificed 12 hours after SAP modeling, followed by blood and pancreatic and distal ileum tissue collection for further analyses. Changes in the pathology responses of the rats in each group were assessed. RESULT: Analyses of serum amylase, lipase, cholesterol, triglyceride, IL-1ß, IL-6, DAO, and serum endotoxin as well as tight junction protein expression including zonula occluden-1 and occludin indicated that high-fat diet aggravated SAP-induced intestinal barrier injury via increasing inflammatory response. In addition, the level of necroptosis was significantly higher in the SAP group compared with the SN group while the HSAP group exhibited more necroptosis compared with the SAP group, indicating the important role of necroptosis in pancreatitis-associated gut injury and illustrating that high-fat diet aggravated necroptosis of the ileum. Pretreatment with TLR4 inhibitor significantly alleviated inflammatory response and reduced necroptosis and level of oxidative stress while improving intestinal barrier function. CONCLUSION: High-fat diet aggravated SAP-induced intestinal barrier injury via inflammatory reactions, necroptosis, and oxidative stress. Inhibition of TLR4 by TAK242 reduced inflammation, alleviated necroptosis, and lowered the level of oxidative stress and then protected the intestinal barrier dysfunction from SAP in high-fat diet-fed rats.

Identifying Reducing and Capping Sites of Protein-Encapsulated Gold Nanoclusters.

The reducing and capping sites along with their local structure impact photo properties of the red bovine serum albumin-capped Au nanocluster (BSA-AuNC), however, they are hard to identify. We developped a workflow and relevant techniques using mass spectrometry (MS) to identify the reducing and capping sites of BSA-AuNCs involved in their formation and fluorescence. Digestion without disulfide cleavages yielded an Au core fraction exhibiting red fluorescence and [AunSm] ion signals and a non-core fraction exhibiting neither of them. The core fraction was identified to mainly be comprised of peptides containing cysteine residues. The fluorescence and [AunSm] signals were quenched by tris(2-carboxyethyl)phosphine, confirming that disulfide groups were required for nanocluster stabilization and fluorescence. By MS sequencing, the disulfide pairs, C75-C91/C90-C101 in domain IA, C315-C360/C359-C368 in domain IIB, and C513-C558/C557-C566 in domain IIIB, were identified to be main capping sites of red AuNCs. Peptides containing oxidized cysteines (sulfinic or cysteic acid) were identified as reducing sites mainly in the non-core fraction, suggesting that disulfide cleavages by oxidization and conformational changes contributed to the subsequent growth of nanoclusters at nearby intact disulfide pairs. This is the first report on precise identification of the reducing and capping sites of BSA-AuNCs.

Secondary structure and membrane topology of dengue virus NS4A protein in micelles.

Dengue virus (DENV) non-structural (NS) 4A is a membrane protein essential for viral replication. The N-terminal region of NS4A contains several helices interacting with the cell membrane and the C-terminal region consists of three potential transmembrane regions. The secondary structure of the intact NS4A is not known as the previous structural studies were carried out on its fragments. In this study, we purified the full-length NS4A of DENV serotype 4 into dodecylphosphocholine (DPC) micelles. Solution NMR studies reveal that NS4A contains six helices in DPC micelles. The N-terminal three helices are amphipathic and interact with the membrane. The C-terminal three helices are embedded in micelles. Our results suggest that NS4A contains three transmembrane helices. Our studies provide for the first time structural information of the intact NS4A of DENV and will be useful for further understanding its role in viral replication.

Characterization of molecular interactions between Zika virus protease and peptides derived from the C-terminus of NS2B.

Zika virus (ZIKV) protease is a two-component complex in which NS3 contains the catalytic triad and NS2B cofactor region is important for protease folding and activity. A protease construct-eZiPro without the transmembrane domains of NS2B was designed. Structural study on eZiPro reveals that the Thr-Gly-Lys-Arg (TGKR) sequence at the C-terminus of NS2B binds to the active site after cleavage. The bZiPro construct only contains NS2B cofactor region and the N-terminus of NS3 without any artificial linker or protease cleavage site, giving rise to an empty pocket accessible to substrate and inhibitor binding. Herein, we demonstrate that the TGKR sequence of NS2B in eZiPro is dynamic. Peptides from NS2B with various lengths exhibit different binding affinities to bZiPro. TGKR binding to the active site in eZiPro does not affect protease binding to small-molecule compounds. Our results suggest that eZiPro will also be useful for evaluating small-molecule protease inhibitors.

Endoplasmic reticulum stress in perivascular adipose tissue promotes destabilization of atherosclerotic plaque by regulating GM-CSF paracrine.

BACKGROUND: Perivascular adipose tissue (PVAT) accelerates plaque progression and increases cardiovascular risk. We tested the hypothesis that PVAT contributed to plaque vulnerability and investigated whether endoplasmic reticulum stress (ER stress) in PVAT played an important role in vulnerable plaque. METHODS: We transplanted thoracic aortic PVAT or subcutaneous adipose tissue as a control, from donor mice to carotid arteries of recipient apolipoprotein E deficient (apoE-/-) mice after removing carotid artery collar placed for 6 weeks. Two weeks after transplantation, ER stress inhibitor 4-phenyl butyric acid (4-PBA) was locally administrated to the transplanted PVAT and then animals were euthanized after 4 weeks. Immunohistochemistry was performed to quantify plaque composition and neovascularization. Mouse angiogenesis antibody array kit was used to test the angiogenic factors produced by transplanted adipose tissue. In vitro tube formation assay, scratch wound migration assay and mouse aortic ring assay were used to assess the angiogenic capacity of supernatant of transplanted PVAT. RESULTS: Ultrastructural detection by transmission electron microscopy showed transplanted PVAT was a mixed population of white and brown adipocytes with abundant mitochondria. Transplanted PVAT increased the intraplaque macrophage infiltration, lipid core, intimal and vasa vasorum neovascularization and MMP2/9 expression in plaque while decreased smooth muscle cells and collagen in atherosclerotic plaque, which were restored by local 4-PBA-treatment. Antibody array analysis showed that 4-PBA reduced several angiogenic factors [Granulocyte Macrophage Colony Stimulating Factor (GM-CSF), MCP-1, IL-6] secreted by PVAT. Besides, conditioned medium from 4-PBA treated-PVAT inhibited tube formation and migration capacity of endothelial cells and ex vivo mouse aortic ring angiogenesis compared to conditioned medium from transplanted PVAT. mRNA expression and protein levels of GM-CSF were markedly elevated in adipocytes under ER stress which would be suppressed by 4-PBA. In addition, ER stress enhanced NF-κB binding to the promoter of the mouse GM-CSF gene in adipocytes confirmed by Chromatin immunoprecipitation analyses. CONCLUSIONS: Our findings demonstrate that ER stress in PVAT destabilizes atherosclerotic plaque, in part through increasing GM-CSF paracrine via transcription factor NF-κB.

Improving the hepatitis cascade: assessing hepatitis testing and its management in primary health care in China.

Objective: The study aimed to decentralize hepatitis testing and management services to primary care in China. Methods: A nationwide representative provider survey amongst community health centres (CHCs) using randomized stratified sampling methods was conducted between September and December 2015. One hundred and eighty CHCs and frontline primary care practitioners from 20 cities across three administrative regions of Western, Central and Eastern China were invited to participate. Results: One hundred and forty-nine clinicians-in-charge (79%), 1734 doctors and 1846 nurses participated (86%). Majority of CHCs (80%, 95% CI: 74-87) offered hepatitis B testing, but just over half (55%, 95% CI: 46-65) offered hepatitis C testing. The majority of doctors (87%) and nurses (85%) felt that there were benefits for providing hepatitis testing at CHCs. The major barriers for not offering hepatitis testing were lack of training (54%) and financial support (23%). Multivariate analysis showed that the major determinants for CHCs to offer hepatitis B and C testing were the number of nurses (AOR 1.1) and written policies for hepatitis B diagnosis (AOR 12.7-27.1), and for hepatitis B the availability of reproductive health service. Conclusions: Primary care providers in China could play a pivotal role in screening, diagnosing and treating millions of people with chronic hepatitis B and C in China.