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Lentiviral vectors are one of the most commonly used viral delivery systems for gene therapy. Vesicular stomatitis virus-G envelope glycoprotein (VSV G)-pseudotyped lentiviral vectors have been widely used in clinical studies for treatment of virus infections and genetic deficient diseases. However, the efficiency of lentiviral vector transduction has been long recognized as a limiting factor in clinical gene therapy application, especially in transducing hematopoietic stem cells. MARCH8 (membrane-associated RING-CH 8), an E3 ubiquitin ligase, has been reported to target and downregulate VSV G. Results in this study show that MARCH8 induces ubiquitination and lysosome degradation of VSV G, and knockout of MARCH8 in virus-producing cells increases lentiviral vector transduction by elevating the level of VSV G protein. We then engineered VSV G mutant that has the lysine residues in the cytoplasmic domain substituted for arginine, and showed that this G mutant resists degradation by MARCH8, and allows the enhancement of transduction efficiency of lentiviral vector particles than the parental VSV G protein. This engineered VSV G mutant thus further advances the lentiviral vector system as a powerful tool in gene therapy.

Objective Most of the studies on the herb Chuanxiong Rhizoma (CR) have focused on the l - arginine-nitric oxide (NO) pathway, but the nitrate-nitrite-NO (NO3−–NO2−–NO) pathway was rarely investigated. Therefore, the aim of this study was to evaluate the effects and mechanisms of action of CR in coronary artery disease (CAD). Methods The NO3−, NO2− and NO levels were examined in the NO3−–NO2−–NO pathway. High-performance ion chromatography was used to quantify NO3− and NO2− levels. Then, NO was quantified using a multifunctional enzyme marker with a fluorescent probe. The tension of aortic rings was measured using a multi myograph system. Results High content of NO3− and low content of NO2− was found in CR, and which could potently convert NO3− to NO2− in the presence of endogenous reductase enzyme. Incubating human coronary artery endothelial cells (HCAECs) with CR-containing serum showed that CR significantly decreased the NO3− content and increased the levels of NO2− and NO in the cells under hypoxic conditions. In addition, CR significantly relaxed isolated aortic rings when the l - arginine –NO pathway was blocked. The optimal concentration of CR for relaxation was 200 mg/mL. Conclusion CR supplements large amounts of NO in cells and vessels to achieve relaxation via the NO3−–NO2−–NO pathway, thereby making up for the deficiency caused by the lack of NO after the l - arginine-NO pathway is suppressed. This study also supports the potential use of a traditional Chinese herb for future drug development.

CRISPR-Cas provides bacteria and archaea with immunity against invading phages and foreign plasmid DNA and has been successfully adapted for gene editing in a variety of species. The class 2 type VI CRISPR-Cas effector Cas13a targets and cleaves RNA, providing protection against RNA phages. Here we report the repurposing of CRISPR-Cas13a to inhibit human immunodeficiency virus type 1 (HIV-1) infection through targeting HIV-1 RNA and diminishing viral gene expression. We observed strong inhibition of HIV-1 infection by CRISPR-Cas13a in human cells. We showed that CRISPR-Cas13a not only diminishes the level of newly synthesized viral RNA, either from the transfected plasmid DNA or from the viral DNA, which is integrated into cellular DNA, but it also targets and destroys the viral RNA that enters cells within viral capsid, leading to strong inhibition of HIV-1 infection. Together, our results suggest that CRISPR-Cas13a provides a potential novel tool to treat viral diseases in humans., Graphical Abstract, The class 2 type VI CRISPR-Cas effector Cas13a targets and cleaves RNA, providing protection against RNA phages. Yin et al. demonstrate that CRISPR-Cas13a targets and cleaves the HIV RNA genome and mRNA, leading to strong inhibition of HIV-1 infection.

Interferon-regulated myxovirus resistance protein B (MxB) is an interferon-induced GTPase belonging to the dynamin superfamily. It inhibits infection with a wide range of different viruses, including HIV-1, by impairing viral DNA entry into the nucleus. Unlike the related antiviral GTPase MxA, MxB possesses an N-terminal region that contains a nuclear localization signal and is crucial for inhibiting HIV-1. Because MxB previously has been shown to reside in both the nuclear envelope and the cytoplasm, here we used bioinformatics and biochemical approaches to identify a nuclear export signal (NES) responsible for MxB's cytoplasmic location. Using the online computational tool LocNES (Locating Nuclear Export Signals or NESs), we identified five putative NES candidates in MxB and investigated whether their deletion caused nuclear localization of MxB. Our results revealed that none of the five deletion variants relocates to the nucleus, suggesting that these five predicted NES sequences do not confer NES activity. Interestingly, deletion of one sequence, encompassing amino acids 505-527, abrogated the anti-HIV-1 activity of MxB. Further mutation experiments disclosed that amino acids 515-519, and Pro-515 in particular, regulate MxB oligomerization and its binding to HIV-1 capsid, thereby playing an important role in MxB-mediated restriction of HIV-1 infection. In summary, our results indicate that none of the five predicted NES sequences in MxB appears to be required for its nuclear export. Our findings also reveal several residues in MxB, including Pro-515, critical for its oligomerization and anti-HIV-1 function.

Introduction. Carbapenem-resistant Enterobacteriaceae (CRE) have been responsible for nosocomial outbreaks worldwide and have become endemic in several countries. Hypothesis/Gap Statement. To better understand the epidemiological trends and characteristics of CRE in the Henan province. Aim. We assessed the molecular epidemiological characteristics of 305 CRE strains isolated from patients in 19 secondary or tertiary hospitals in ten areas of the Henan province in China. Methodology. A total of 305 CRE isolates were subjected to multiple tests, including in vitro antimicrobial susceptibility testing, PCR for carbapenemase genes bla KPC, bla NDM, bla IMP, bla VIM, bla OXA-48-like. Tigecycline-resistant genes ramR, oqxR, acrR, tetA, rpsJ, tetX, tetM, tetL were analysed in five tigecycline non-susceptible carbapenem-resistant Klebsiella pneumoniae isolates (TNSCRKP). Additionally, multilocus sequence typing (MLST) was performed for carbapenem-resistant K. pneumoniae (CRKP). Results. The most common CRE species were K. pneumoniae (234, 77 %), Escherichia coli (36, 12 %) and Enterobacter cloacae (13, 4 %). All strains exhibited multi-drug resistance. Overall, 97 % (295/305) and 97 % (297/305) of the isolates were susceptible to polymyxin B and tigecycline, respectively. A total of 89 % (271/305) of the CRE isolates were carbapenemase gene-positive, including 70 % bla KPC, 13 % bla NDM, 6 % bla IMP, and 1 % combined bla KPC/bla NDM genes. K. pneumoniae carbapenemase (KPC) was the predominant carbapenemase in K. pneumoniae (87 %), whereas NDM and IMP were frequent in E. coli (53 %) and E. cloacae (69 %), respectively. Mutations in the ramR, tetA, and rpsJ genes were detected in five TNSCRKP. Moreover, 15 unique sequence types were detected, with ST11 (74 %), ST15 (9 %) and ST2237 (5 %) being dominant among K. pneumoniae strains. Conclusion. A high proportion of CRE strains were carbapenemase-positive, and five carbapenem-resistant K. pneumonia isolates were tigecycline non-susceptible, indicating a need for the ongoing surveillance of CRE and effective measures for the prevention of CRE infections.

Initial malarial infection mostly causes symptomatic illness in humans. Infection that is not fatal induces complete protection from severe illness and death, and thus complete protection from severe illness or death is granted with sufficient exposure. However, malaria parasite immunity necessitates constant exposure. Therefore, it is important to evaluate lowered immunity and recurrent susceptibility to symptomatic disease in lower transmission areas. We aimed to investigate selection pressure based on transmission levels, antimalarial drug use, and environmental factors. We whole genome sequenced (WGS) P. falciparum clinical samples from Chinese hosts working in Ghana and compared the results with the WGS data of isolates from native Ghanaians downloaded from pf3k. The P. falciparum samples were generally clustered according to their geographic origin, and Chinese imported samples showed a clear African origin with a slightly different distribution from the native Ghanaian samples. Moreover, samples collected from two host populations showed evidence of differences in the intensity of selection. Compared with native Ghanaian samples, the China-imported isolates exhibited a higher proportion of monoclonal infections, and many genes associated with RBC invasion and immune evasion were found to be under less selection pressure. There was no significant difference in the selection of drug-resistance genes due to a similar artemisinin-based combination therapy medication profile. Local selection of malarial parasites is considered to be a result of differences in the host immunity or disparity in the transmission opportunities of the host. In China, most P. falciparum infections were imported from Africa, and under these circumstances, distinct local selective pressures may be caused by varying acquired immunity and transmission intensity. This study revealed the impact of host switching on the immune system, and it may provide a better understanding of the mechanisms that enable clinical immunity to malaria.

To investigate the particle size distribution of particulate matter and the concentration of specific perfluorinated compounds in indoor dust samples from several locations. Then, we used cell-based assays to investigate the effect of perfluorinated compounds on human corneal epithelial (HCEpiC), endothelial cells (HCEC) and retinal pigment epithelial cells (RPE). Indoor dust samples were collected at five different locations and PM50–10, PM10–2.5, and PM2.5–1 were fractionized. The presence and levels of 8:2 fluorotelomer alcohol, 10:2 fluorotelomer alcohol, and perfluorooctanoic acid were detected by gas chromatography–mass spectrometry. The effect of perfluorooctanoic acid on the activation of reactive oxygen species, transepithelial resistance as well as the expression of interleukin (IL)-6 and IL-8 were determined. The basolateral media of human corneal epithelial or human corneal endothelial cells were used to treat human corneal endothelial or retinal pigment epithelial cells, respectively to indicate the potential of ocular surface inflammation may result in retinal inflammation. Among perfluorinated compounds, only perfluorooctanoic acid was detected in all indoor dust samples. Perfluorooctanoic acid had the highest concentration among all perfluorinated compounds in the samples. Exposure to perfluorooctanoic acid impaired tight junction sealing and increased the levels of reactive oxygen species in human corneal epithelial cells. In human corneal epithelial cells, secretion of IL-6 and IL-8 in both apical and basolateral media was promoted significantly by perfluorooctanoic acid treatment. Stimulation with the basolateral media from perfluorooctanoic acid-treated human corneal epithelial cells induced inflammation in human corneal endothelial cells. The treatment of retinal pigment epithelial cells with the basolateral media from stimulated human corneal endothelial cells also elicited the secretion of proinflammatory cytokines. The results indicate that perfluorooctanoic acid exposure impaired the tight junction of corneal cells and caused inflammatory reactions in the retina. Exposure of the cornea to perfluorooctanoic acid contained in particulate matter might induce oxidative stress and inflammation in the retina and represent a risk factor for age-related macular degeneration.

Abnormal expression of ADAM29 has been frequently reported in several cancers, however, its role in clear cell renal cell carcinoma (ccRCC) has not evaluated in detail. Herein, we attempt to determine the biological role and the action mechanism of ADAM29 in ccRCC. Bioinformatics analysis based on the ccRCC RNA-Seq dataset from TCGA database revealed that ADAM29 was up-expressed in ccRCC tissues by comparison with normal tissues. And a significant increase of ADAM29 expression was also observed in 3 ccRCC cell lines (UT33A, Caki-1, and786-O) in comparison with normal cell line. Besides, high level of ADAM29 was found to be connected with the poor prognosis and could be considered as an independent prognosticator for patients with ccRCC. Furthermore, functional experiments in vitro demonstrated that ADAM29 promoted the growth, invasion and migration of ccRCC cells. Moreover, Western blot assays indicated that ADAM29 was positively correlated with the level of proliferation-related proteins Cyclin D1 and PCNA and motion-related proteins MMP9 and Snail. Our data indicate that ADAM29 acts as an oncogene that increases tumour cells proliferation, invasion and migration partly by regulating the expression of Cyclin D1/PCNA/MMP9/Snail, suggesting that ADAM29 may become a prognosticator and therapeutic candidate for ccRCC.

Cyclic GMP-AMP synthase (cGAS) is a major sensor of cytosolic DNA from invading pathogens and damaged cellular organelles. Activation of cGAS promotes liquid-like phase separation and formation of membraneless cytoplasmic structures. Here, we found that cGAS bound G3BP1, a double-stranded nucleic acid helicase involved in the formation of stress granules. Loss of G3BP1 blocked subcellular cGAS condensation and suppressed the interferon response to intracellular DNA and DNA virus particles in cells. Furthermore, an RNA-dependent association with PKR promoted G3BP1 foci formation and cGAS-dependent interferon responses. Together, these results indicate that PKR promotes the formation of G3BP1-dependent, membraneless cytoplasmic structures necessary for the DNA-sensing function of cGAS in human cells. These data suggest that there is a previously unappreciated link between nucleic acid sensing pathways, which requires the formation of specialized subcellular structures.

The main purpose of this study was to examine the longitudinal mediation effects of multiple gaming motives from online game involvement to problematic Internet use (PIU). In terms of mediators, the present study included three specific gaming motivations - escapism, advancement, and socializing. A sample of 389 gamers participated in the study from freshman to junior college years. Gaming involvement was collected at the freshman year, motives at sophomore, and PIU at junior. Initially, the three gaming motives were examined separately for their longitudinal mediation effects in a series of single-mediator models. Next, the three motives were simultaneously included in a multiple-mediator model to compare the relative levels of their mediation effects from gaming involvement to PIU across a period of two years. In the multiple-mediator model, the most compelling results showed that escapism and advancement were positively associated with PIU, yet socializing was not; socializing failed to significantly predict later PIU. Also, the direct effect of gaming involvement on PIU was weak and failed to reach significant level, contradicting to the findings of previous cross-sectional studies. Many game players experience great pleasure from immersing themselves in a virtual world, and often to the extent of developing addictive symptoms. This phenomenon is referred to as “pull” effect which is considered comparable to the motive of “pursuing in-game advancement” in the present study. Conversely, frustrations or other negative experiences in real life might also bring about a “push” effect that tugs players into gaming. The motive of escapism from the real-life in this study resembles the so-called “push” effect. The authors suggest when gamers demonstrated social motive during game time, such as participating in a gamer community and collaborating with the others despite being socially withdrawn in real life, it would be less reasonably to be regarded as PIU. During game time, if gamers experience a temporary sense of well-being, it might help to vent their real-life frustration. However, an excessive use of this emotion-focused coping strategy might still increase the risk of PIU.

Multi-glycoside of Tripterygium wilfordii Hook. f. (GTW) has been proven to be clinically effective in relieving microinflammation in patients with early diabetic nephropathy (DN). However, the therapeutic mechanisms involved in vivo remain unclear. In the process of early DN, microinflammation and activation of p38 mitogen‑activated protein kinase (MAPK) and canonical nuclear factor (NF)-κB signaling pathways are the important mechanisms by which hyperglycemia contributes to glomerulosclerosis (GS). Therefore, this study aimed to examine the ameliorative effects of GTW on GS, and then to clarify its anti‑microinflammatory mechanisms by inhibiting p38 MAPK and NF-κB signaling activities in the kidney. All rats were divided into 4 groups: the sham group, the sham + GTW group, the vehicle group and the GTW group. The suitable dose of GTW and vehicle were daily administered for 8 weeks after the induction of DN by unilateral nephrectomy combined with intraperitoneal injections of streptozotocin (STZ). The general status of the rats, biochemical parameters, renal histological changes and macrophages in glomeruli, as well as expression of the key proteins in the p38 MAPK and canonical NF-κB signaling pathways and inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1β and transforming growth factor (TGF)-β1 in the kidney were examined, respectively. The results revealed that, GTW improved the general cond-ition and biochemical parameters of the rats, but did not lower blood glucose; GTW attenuated GS and suppressed glomerular microinflammation including the infiltration of ED1+ cells in glomeruli and the protein overexpression of TNF-α, IL-1β and TGF-β1 in the kidney; GTW inhibited the protein overexpression of key signaling molecules of p38 MAPK and canonical NF-κB pathways in the kidney including phosphorylated p38 MAPK, phosphorylated inhibitor protein IκB and NF-κB (p65). On the whole, we expounded that GTW, as a natural regulator in vivo, alleviates GS without affecting hyperglycemia, by exerting anti-microinflammatory effects, including reducing macrophage infiltration in glomeruli, suppressing TNF-α, IL-1β and TGF-β1 overexpression in the kidney and inhibiting p38 MAPK and NF-κB signaling activities.

The advent of nanomedicine requires novel delivery vehicles to actively target their site of action. Here, we demonstrate the development of lung-targeting drug-loaded liposomes and their efficacy, specificity and safety. Our study focuses on glucocorticoids methylprednisolone (MPS), a commonly used drug to treat lung injuries. The steroidal molecule was loaded into functionalized nano-sterically stabilized unilamellar liposomes (NSSLs). Targeting functionality was performed through conjugation of surfactant protein A (SPANb) nanobodies to form MPS–NSSLs–SPANb. MPS–NSSLs–SPANb exhibited good size distribution, morphology, and encapsulation efficiency. Animal experiments demonstrated the high specificity of MPS–NSSLs–SPANb to the lung. Treatment with MPS–NSSLs–SPANb reduced the levels of TNF-α, IL-8, and TGF-β1 in rat bronchoalveolar lavage fluid and the expression of NK-κB in the lung tissues, thereby alleviating lung injuries and increasing rat survival. The nanobody functionalized nanoparticles demonstrate superior performance to treat lung injury when compared to that of antibody functionalized systems.

Background For a long time, the Christie-Atkinson-Munch-Peterson (CAMP) test has been a standard test for the identification of Streptococcus agalactiae, and a positive result for S.agalactiae has been considered sensitive enough. Methods To confirm whether a positive CAMP test is a requirement for the identification of S.agalactiae, five suspected CAMP-negative S.agalactiae isolates from two hospitals, confirmed as Gram-positive and catalase-negative streptococci, were verified by the CAMP test in three batches of plates from two manufacturers and identified by the Phoenix system, MALDI-TOF MS, the PCR assay and the 16S rDNA gene sequencing. Results All five suspected strains were S.agalactiae, four of which were CAMP-negative and one of which was not S.agalactiae by the PCR assay. Conclusions A positive CAMP test was lacking sensitivity for the identification of S.agalactiae, and the question of whether the cfb gene is worthy of targeting should be further studied.

Objectives In recent years, Klebsiella pneumoniae has emerged as a leading cause of nosocomial infection owing to the rising prevalence of multidrug-resistant strains, particularly carbapenem-resistant isolates. In this study, the complete genome sequence of carbapenem-resistant K. pneumoniae SWU01 was determined. Methods Antimicrobial susceptibilities and hypermucoviscous phenotype were determined by the disk diffusion method and positive string test, respectively. Multilocus sequence typing (MLST) was performed using the K. pneumoniae MLST database, and capsular serotype was analysed using the BIGSdb-Kp database with the nucleotide sequence of the variable region (CD1-VR2-CD2) of wzc . The complete genome sequence was obtained via the PacBio RS II platform, and antimicrobial resistance genes were identified using ResFinder 2.1. Results SWU01 was resistant to all antibiotics tested except polymyxin B and minocycline. This strain showed a hypermucoviscous phenotype with serotype K47 belonging to the ST11 clone. The complete genome consists of a 5 536 506-bp circular chromosome and a 162 552-bp plasmid, with a G+C content of 57.4%. A total of 5537 protein-coding sequences, 85 tRNAs, 25 rRNAs, 12 non-coding RNA genes and 157 pseudogenes were identified in the genome. Thirteen acquired antibiotic resistance genes were detected (eight in the chromosome and five in the plasmid). Conclusions Here we present the first whole-genome sequence of a carbapenem-resistant hypermucoviscous K. pneumoniae isolate SWU01 with capsular serotype K47 belonging to ST11 from a patient in China, which may serve as a reference sequence for further understanding of the pathogenesis and multidrug resistance mechanisms of this species.

MOV10 has emerged as an important host antiviral factor. MOV10 not only inhibits various viruses, including human immunodeficiency virus type 1, hepatitis C virus and vesicular stomatitis virus, but also restricts the activity of retroelements long interspersed nucleotide element-1, Alu, SVA and intracisternal A particles. Here, we report that MOV10 suppresses influenza A virus infection through interacting with viral nucleoprotein (NP), sequestering viral RNP in the cytoplasm and causing the degradation of viral vRNA. The antiviral activity of MOV10 depends on the integrity of P-bodies. We also found that the antiviral activity of MOV10 is partially countered by viral NS1 protein that interferes with the interaction of MOV10 with viral NP and causes MOV10 degradation through the lysosomal pathway. Moreover, NS1-defective influenza A virus is more susceptible to MOV10 restriction. Our data not only expand the antiviral spectrum of MOV10 but also reveal the NS1 protein as the first viral antagonist of MOV10.

Freezing of gait (FOG) is a common and debilitating symptom in Parkinson’s disease (PD). The current study investigated alterations of resting-state spontaneous brain activity in PD patients with FOG. A total of 29 patients with FOG, 28 patients without FOG and 31 controls were included. All subjects underwent resting-state functional MRI, and the amplitude of low-frequency fluctuation (ALFF) was calculated to measure the spontaneous brain activity. Between-group differences and correlations with FOG severity (both subjective and objective measures) were analyzed. Compared to those without FOG, patients with FOG showed increased ALFF in right anterior cingulate cortex (ACC) and left inferior parietal lobule (IPL), as well as decreased ALFF in right superior frontal gyrus (SFG), bilateral cerebellum and left thalamus. Correlation analyses demonstrated that ALFF within the right SFG, right ACC and bilateral pallidum were positively correlated with FOG; while ALFF within the thalamus, putamen, cerebellum and sensorimotor regions were negatively correlated. Our results indicate that FOG is associated with dysfunction within frontal-parietal regions, along with increased inhibitory outputs from basal ganglia. Additionally, altered activity of cerebellum implicates its role in the pathophysiology of FOG. These findings provide further insight into the underlying neural mechanisms of FOG in PD patients.

BST-2 (tetherin, CD317, and HM1.24) is induced by interferon and restricts virus release by tethering the enveloped viruses to the cell surface. The effect of BST-2 on influenza A virus (IAV) infection has been inconclusive. In the present study, we report that BST-2 diminishes the production of IAV virus-like particles (VLPs) that are generated by viral neuraminidase and hemagglutinin proteins to a much greater degree than it inhibits the production of wild-type IAV particles. This relatively weaker inhibition of IAV is associated with reduction in BST-2 levels, which is caused by the M2 protein that interacts with BST-2 and leads to down-regulation of cell surface BST-2 via the proteasomal pathway. Similarly to the viral antagonist Vpu, M2 also rescues the production of human immunodeficiency virus-1 VLPs and IAV VLPs in the presence of BST-2. Replication of wild-type and the M2-deleted viruses were both inhibited by BST-2, with the M2-deleted IAV being more restricted. These data reveal one mechanism that IAV employs to counter restriction by BST-2.

Enterovirus 71 (EV71) infection causes hand-foot-and-mouth disease that leads to cardiopulmonary complications and death in young children. There is thus an urgent need to find new treatments to control EV71 infection. In this study, we report potent inhibition of EV71 by a polyene antibiotic Amphotericin B. Amphotericin B profoundly diminished the expression of EV71 RNA and viral proteins in the RD cells and the HEK293 cells. As a result, EV71 production was inhibited by Amphotericin B with an EC50 (50% effective concentration) of 1.75 μM in RD cells and 0.32 μM in 293 cells. In addition to EV71, EV68 was also strongly inhibited by Amphotericin B. Results of mechanistic studies revealed that Amphotericin B targeted the early stage of EV71 infection through impairing the attachment and internalization of EV71 by host cells. As an effective anti-fungi drug, Amphotericin B thus holds the promise of formulating a novel therapeutic to treat EV71 infection.

Nanobody (Nb) is a promising vector for targeted drug delivery. This study aims to identify an Nb that can specifically target the lung by binding human pulmonary surfactant protein A (SP-A). Human lung frozen tissue sections were used for 3 rounds of biospanning of our previously constructed Nb library for rat SP-A to establish a sub-library of Nb, which specifically bound human lung tissues. Phage-ELISA was performed to screen the sub-library to identify Nb4, which specifically bound human SP-A. The binding affinity Kd of Nb4 to recombinant human SP-A was 7.48 × 10−7 M. Nb4 (19 kDa) was stable at 30 °C–37 °C and pH 7.0–7.6 and specifically bound the SP-A in human lung tissue homogenates, human lung A549 cells, and human lung tissues, whereas didn’t react with human liver L-02 cells, kidney 293T cells, and human tissues from organs other than the lung. Nb4 accumulated in the lung of nude mice 5 minutes after a tail vein injection of Nb4 and was excreted 3 hours. Short-term exposure (one month) to Nb4 didn’t cause apparent liver and kidney toxicity in rats, whereas 3-month exposure resulted in mild liver and kidney injuries. Nb4 may be a promising vector to specifically deliver drugs to the lung.

Podocyte injury is closely related to proteinuria in the progress of diabetic nephropathy(DN). The pathological characters of podocyte injury mainly refer to the change of podocyte form and function, including foot process effacement, reduction of podocyte number and density, podocyte apoptosis, podocyte epithelial-mesenchymal transdifferentiation(EMT)and podocyte hypertrophy. These pathological damages are controlled by multiple signaling pathways in the kidney, such as mammalian target of rapamycin(mTOR)/autophagy pathway, transforming growth factor(TGF)-β1 pathway and Notch pathway. For podocyte injuries induced by high glucose or in murine models of DN, some Chinese herbal medicine(CHM)extracts, such as multiglycoside of Tripterygium wilfordii(GTW), triptolide(TP), astragaloside IV(AS-IV), astragalus polysaccharide(APS)and Panax notoginseng saponins(PNS), have the protective effects in vivo or in vitro. The preliminary studies in China showed that GTW improves podocyte injury in the DN model rats probably through regulating the activity of mTORC1 signaling pathway in the kidney. Therefore, it is the developmental direction for the further study to clarify the interventional effects of CHM based on podocyte injury-related signaling pathway in DN.

Enhancers frequently contain multiple binding sites for the same transcription factor. These homotypic binding sites often exhibit synergy, whereby the transcriptional output from two or more binding sites is greater than the sum of the contributions of the individual binding sites alone. Although this phenomenon is frequently observed, the mechanistic basis for homotypic binding site synergy is poorly understood. Here, we identify a bona fide cardiac-specific Prkaa2 enhancer that is synergistically activated by homotypic MEF2 binding sites. We show that two MEF2 sites in the enhancer function cooperatively due to bridging of the MEF2C-bound sites by the SAP domain-containing co-activator protein myocardin, and we show that paired sites buffer the enhancer from integration site-dependent effects on transcription in vivo. Paired MEF2 sites are prevalent in cardiac enhancers, suggesting that this might be a common mechanism underlying synergy in the control of cardiac gene expression in vivo.

The renoprotective effect of inhibitors of renin-angiotensin system (RAS) has been identified through placebo-controlled trials. However, the effect of calcium-channel blockers (CCBs) on renal system is still controversial. Our current meta-analysis includes available evidences to compare the effect of dihydropyridine CCBs and ACEIs or ARBs on renal outcomes and mortality. We also further investigate whether CCBs can be used in combination with inhibitors of RAS to improve the prognosis of patients with chronic kidney disease (CKD).Electronic databases were searched up to July 2012, for clinical randomized controlled trials, assessing the effect of dihydropyridine CCBs on the incidence of end-stage renal disease (ESRD) and all-cause mortality in contrast to ACEIs or ARBs. Eight clinical trials were included containing 25,647 participants. ESRD showed significantly higher frequency with CCBs therapy compared with ACEIs or ARBs therapy, though blood pressure was decreased similarly in both groups in every trial (OR, 1.25; 95% CI, 1.05-1.48; p = 0.01). In contrast, there was no significant difference in the incidence of all-cause mortality between these two groups, though ACEIs or ARBs exhibited better renoprotective effect compared to CCBs (OR, 0.96; 95% CI, 0.89-1.03; p = 0.24).CCBs did not increase all-cause mortality incidence in patients with CKD though they displayed weaker renoprotective, compared to ACEIs or ARBs therapy. Our results suggest the combination of a CCB and an ACEI or ARB should be a preferable antihypertensive therapy in patients with CKD, considering their higher effect in decreasing blood pressure and fewer adverse metabolic problems caused.

CD97 belongs to the adhesion GPCR family characterized by a long ECD linked to the 7TM via a GPCR proteolytic site (GPS) and plays important roles in modulating cell migration and invasion. CD97 (EGF1-5) is a splicing variant of CD97 that recognizes a specific ligand chondroitin sulfate on cell membranes and the extracellular matrix. The aim of this study was to elucidate the extracellular molecular basis of the CD97 EGF1-5 isoform in protein expression, auto-proteolysis and cell adhesion, including epidermal growth factor (EGF)-like domain, GPCR autoproteolysis-inducing (GAIN) domain, as well as GPS mutagenesis and N-glycosylation. Both wild-type (WT) CD97-ECD and its truncated, GPS mutated, PNGase F-deglycosylated, and N-glycosylation site mutated forms were expressed and purified. The auto-proteolysis of the proteins was analyzed with Western blotting and SDS-PAGE. Small angle X-ray scattering (SAXS) and molecular modeling were used to determine a structural profile of the properly expressed receptor. Potential N-glycosylation sites were identified using MS and were modulated with PNGase F digestion and glyco-site mutations. A flow cytometry-based HeLa cell attachment assay was used for all aforementioned CD97 variants to elucidate the molecular basis of CD97-HeLa interactions. A unique concentration-dependent GPS auto-proteolysis was observed in CD97 EGF1-5 isoform with the highest concentration (4 mg/mL) per sample was self-cleaved much faster than the lower concentration (0.1 mg/mL), supporting an intermolecular mechanism of auto-proteolysis that is distinct to the reported intramolecular mechanism for other CD97 isoforms. N-glycosylation affected the auto-proteolysis of CD97 EGF1-5 isoform in a similar way as the other previously reported CD97 isoforms. SAXS data for WT and deglycosylated CD97ECD revealed a spatula-like shape with GAIN and EGF domains constituting the body and handle, respectively. Structural modeling indicated a potential interaction between the GAIN and EGF5 domains accounting for the absence of expression of the GAIN domain itself, although EGF5-GAIN was expressed similarly in the wild-type protein. For HeLa cell adhesion, the GAIN-truncated forms showed dramatically reduced binding affinity. The PNGase F-deglycosylated and GPS mutated forms also exhibited reduced HeLa attachment compared with WT CD97. However, neither N-glycosylation mutagenesis nor auto-proteolysis inhibition caused by N-glycosylation mutagenesis affected CD97-HeLa cell interactions. A comparison of the HeLa binding affinities of PNGase F-digested, GPS-mutated and N-glycosylation-mutated CD97 samples revealed diverse findings, suggesting that the functions of CD97 ECD were complex, and various technologies for function validation should be utilized to avoid single-approach bias when investigating N-glycosylation and auto-proteolysis of CD97. A unique mechanism of concentration-dependent auto-proteolysis of the CD97 EGF1-5 isoform was characterized, suggesting an intermolecular mechanism that is distinct from that of other previously reported CD97 isoforms. The EGF5 and GAIN domains are likely associated with each other as CD97 expression and SAXS data revealed a potential interaction between the two domains. Finally, the GAIN and EGF domains are also important for CD97-HeLa adhesion, whereas N-glycosylation of the CD97 GAIN domain and GPS auto-proteolysis are not required for HeLa cell attachment.

Astilbin, a major bioactive compound extracted from Rhizoma smilacis glabrae (RSG), has been reported to possess immunosuppressive properties. Our study first evaluated the effect of astilbin on experimental autoimmune myasthenia gravis (EAMG) in Lewis rats. The results showed that astilbin could attenuate the severity of EAMG by decreasing antigen-specific autoantibodies with up-regulation of regulatory T cells and down-regulation of Th17 cells. In addition to, astilbin also reduced the efficiency of the antigen presenting cells on which the expression of MHC class II decreased. These results suggest that astilbin might be a candidate drug for immunoregulation of EAMG, and provide us new treatment ideas for human myasthenia gravis (MG).

Drug-induced liver injury (DILI) is often undiagnosed or misdiagnosed clinically because of diagnostic difficulties caused by lack of laboratory-specific serological markers. In this study, we comprehensively assessed the clinical characteristics, laboratory indices, hepatotoxic drugs, risk factors and outcomes concerning DILI, and explored the similarities in mechanisms between Chinese and Western drug-induced DILI. Patients with a first diagnosis of DILI and a Roussel Uclaf Causality Assessment Method (RUCAM) score >3 points were enrolled for systematic retrospective study. Their clinical characteristics, clinical classification, risk factors, laboratory indices, hepatotoxic drugs and outcomes were analyzed. Cholestatic patients had the highest alkaline phosphatase (ALP) and prothrombin time activity (PTA) levels (P

The developing heart contains an inner tube of specialized endothelium known as endocardium, which performs multiple essential functions. In spite of the essential role of the endocardium in heart development and function, the transcriptional pathways that regulate its development remain largely undefined. GATA4 is a zinc finger transcription factor that is expressed in multiple cardiovascular lineages and is required for endocardial cushion development and embryonic viability, but the transcriptional pathways upstream of Gata4 in the endocardium and its derivatives in the endocardial cushions are unknown. Here, we describe a distal enhancer from the mouse Gata4 gene that is briefly active in multiple cardiac lineages early in cardiac development but restricts to the endocardium where it remains active through cardiogenesis. The activity of this Gata4 cardiac enhancer in transgenic embryos and in cultured aortic endothelial cells is dependent on four ETS sites. To identify which ETS transcription factors might be involved in Gata4 regulation via the ETS sites in the enhancer, we determined the expression profile of 24 distinct ETS factors in embryonic mouse hearts. Among multiple ETS transcripts present, ETS1, FLI1, ETV1, ETV5, ERG, and ETV6 were the most abundant in the early embryonic heart. We found that ETS1, FLI1, and ERG were strongly expressed in the heart at embryonic day 8.5 and that ETS1 and ERG bound to the endogenous Gata4 enhancer in cultured endothelial cells. Thus, these studies define the ETS expression profile in the early embryonic heart and identify an ETS-dependent enhancer from the Gata4 locus.

The novel Influenza A (H1N1) virus is attacking the world in 2009. Among others, campuses in China, particularly most university/college campuses for bachelor students, are at-risk areas where many susceptible youngsters live. They most likely interact with each other quite often in dormitories, classrooms and refectories. We model the pandemic influenza A (H1N1) transmission through campus contacts and then forecast the effectiveness of interventions, based on a previously presented Complex Agent Network model for simulating infectious diseases [1]. Our results suggest that pandemic influenza A (H1N1) on campus will die out even with no intervention taken; the most effective intervention is still quarantining confirmed cases as early as possible and, in addition, vaccinating susceptible people can further decrease the maximum daily number of the infected. This study can support quantitative experimentation and prediction of infectious diseases within predefined areas, and assessment of intervention strategies. (C) 2010 Published by Elsevier Ltd.

International audience; We report an unexpected role for protease signaling in neural tube closure and the formation of the central nervous system. Mouse embryos lacking protease-activated receptors 1 and 2 showed defective hind-brain and posterior neuropore closure and developed exencephaly and spina bifida, important human congenital anomalies. Par1 and Par2 were expressed in surface ectoderm, and Par2 was expressed selectively along the line of closure. Ablation of G i/z and Rac1 function in these Par2-expressing cells disrupted neural tube closure, further implicating G protein-coupled receptors and identifying a likely effector pathway. Cluster analysis of protease and Par2 expression patterns revealed a group of membrane-tethered proteases often coexpressed with Par2. Among these, matriptase activated Par2 with picomolar potency, and hepsin and prostasin activated matriptase. Together, our results suggest a role for protease-activated receptor signaling in neural tube closure and identify a local protease network that may trigger Par2 signaling and monitor and regulate epithelial integrity in this context.

The cells of the second heart field (SHF) contribute to the outflow tract and right ventricle, as well as to parts of the left ventricle and atria. Isl1, a member of the LIM-homeodomain transcription factor family, is expressed early in this cardiac progenitor population and functions near the top of a transcriptional pathway essential for heart development. Isl1 is required for the survival and migration of SHF-derived cells into the early developing heart at the inflow and outflow poles. Despite this important role for Isl1 in early heart formation, the transcriptional regulation of Isl1 has remained largely undefined. Therefore, to identify transcription factors that regulate Isl1 expression in vivo, we screened the conserved noncoding sequences from the mouse Isl1 locus for enhancer activity in transgenic mouse embryos. Here, we report the identification of an enhancer from the mouse Isl1 gene that is sufficient to direct expression to the SHF and its derivatives. The Isl1 SHF enhancer contains three consensus Forkhead transcription factor binding sites that are efficiently and specifically bound by Forkhead transcription factors. Importantly, the activity of the enhancer is dependent on these three Forkhead binding sites in transgenic mouse embryos. Thus, these studies demonstrate that Isl1 is a direct transcriptional target of Forkhead transcription factors in the SHF and establish a transcriptional pathway upstream of Isl1 in the SHF.

Background The transmission through contacts among MSM (men who have sex with men) is one of the dominating contributors to HIV prevalence in industrialized countries. In Amsterdam, the capital of the Netherlands, the MSM risk group has been traced for decades. This has motivated studies which provide detailed information about MSM's risk behavior statistically, psychologically and sociologically. Despite the era of potent antiretroviral therapy, the incidence of HIV among MSM increases. In the long term the contradictory effects of risk behavior and effective therapy are still poorly understood. Methods Using a previously presented Complex Agent Network model, we describe steady and casual partnerships to predict the HIV spreading among MSM. Behavior-related parameters and values, inferred from studies on Amsterdam MSM, are fed into the model; we validate the model using historical yearly incidence data. Subsequently, we study scenarios to assess the contradictory effects of risk behavior and effective therapy, by varying corresponding values of parameters. Finally, we conduct quantitative analysis based on the resulting incidence data. Results The simulated incidence reproduces the ACS historical incidence well and helps to predict the HIV epidemic among MSM in Amsterdam. Our results show that in the long run the positive influence of effective therapy can be outweighed by an increase in risk behavior of at least 30% for MSM. Conclusion We recommend, based on the model predictions, that lowering risk behavior is the prominent control mechanism of HIV incidence even in the presence of effective therapy.

Simulating the evolution of the Human Immunodeficiency Virus (HIV) epidemic requires a detailed description of the population network, especially for small populations in which individuals can be represented in detail and accuracy. In this paper, we introduce the concept of a Complex Agent Network(CAN) to model the HIV epidemics by combining agent-based modelling and complex networks, in which agents represent individuals that have sexual interactions. The applicability of CANs is demonstrated by constructing and executing a detailed HIV epidemic model for men who have sex with men (MSM) in Amsterdam, including a distinction between steady and casual relationships. We focus on MSM contacts because they play an important role in HIV epidemics and have been tracked in Amsterdam for a long time. Our experiments show good correspondence between the historical data of the Amsterdam cohort and the simulation results., Comment: 21 pages, 4 figures, Mathematics and Computers in Simulation, added references

The SAM domain and HD domain containing protein 1 (SAMHD1) inhibits retroviruses, DNA viruses and long interspersed element 1 (LINE-1). Given that in dividing cells, SAMHD1 loses its antiviral function yet still potently restricts LINE-1, we propose that, instead of blocking viral DNA synthesis by virtue of its dNTP triphosphohydrolase activity, SAMHD1 may exploit a different mechanism to control LINE-1. Here, we report a new activity of SAMHD1 in promoting cellular stress granule assembly, which correlates with increased phosphorylation of eIF2α and diminished eIF4A/eIF4G interaction. This function of SAMHD1 enhances sequestration of LINE-1 RNP in stress granules and consequent blockade to LINE-1 retrotransposition. In support of this new mechanism of action, depletion of stress granule marker proteins G3BP1 or TIA1 abrogates stress granule formation and overcomes SAMHD1 inhibition of LINE-1. Together, these data reveal a new mechanism for SAMHD1 to control LINE-1 by activating cellular stress granule pathway., Author Summary Long interspersed element 1 (LINE-1 or L1) comprises 17% of human genome, and has played a major role in shaping the evolution of human genome. Approximately 100 copies of LINE-1 are still active in an average individual genome. Movement of these LINE-1 sequences to new loci in the genome has the potential of causing sporadic cases of disease. Among the multi-layered mechanisms by which the host controls LINE-1 activity is a group of host restriction factors including APOBEC3 proteins. SAMHD1 was known for the association of its mutations with the Aicardi-Goutieres syndrome (AGS), a congenital autoimmune disease. SAMHD1 was recently reported as a host restriction factor that inhibits a number of retroviruses and DNA viruses including human immunodeficiency virus type 1 (HIV-1) and herpes simplex virus 1 (HSV-1). Here, we demonstrate that SAMHD1 inhibits LINE-1 retrotransposition through promoting the sequestration of LINE-1 RNP within the cytoplasmic stress granules. SAMHD1 promotes the formation of large stress granules by inducing phosphorylation of eIF2α and disrupting eIF4A/eIF4G interaction. This is the first report describing the role of SAMHD1 in modulating the formation of stress granules. We envision that this function of SAMHD1 not only contributes to the inhibition of LINE-1, but also the restriction of various viruses.

Background Pandemic influenza represents a major threat to global health. Vaccination is the most economic and effective strategy to control influenza pandemic. Conventional vaccine approach, despite being effective, has a number of major deficiencies including limited range of protection, total dependence on embryonated eggs for production, and time consuming for vaccine production. There is an urgent need to develop novel vaccine strategies to overcome these deficiencies. Methodology/Principal Findings The major objective of this work was to develop a novel vaccine strategy combining recombinant haemagglutinin (HA) protein and a master cell (MC) activator C48/80 for intranasal immunization. We demonstrated in BALB/c mice that MC activator C48/80 had strong adjuvant activity when co-administered with recombinant HA protein intranasally. Vaccination with C48/80 significantly increased the serum IgG and mucosal surface IgA antibody responses against HA protein. Such increases correlated with stronger and durable neutralizing antibody activities, offering protection to vaccinated animals from disease progression after challenge with lethal dose of A/California/04/2009 live virus. Furthermore, protected animals demonstrated significant reduction in lung virus titers, minimal structural alteration in lung tissues as well as higher and balanced production of Th1 and Th2 cytokines in the stimulated splenocytes when compared to those without C48/80. Conclusions/Significance The present study demonstrates that the novel vaccine approach of combining recombinant HA and mucosal adjuvant C48/80 is safe and effective in eliciting protective immunity in mice. Future studies on the mechanism of action of C48/80 and potential combination with other vaccine strategies such as prime and boost approach may help to induce even more potent and broad immune responses against viruses from various clades.

With the exponential growth in the world population and the constant increase in human mobility, the possible impact of outbreaks of epidemics on cities is increasing, especially in high-density urban areas such as public transportation and transfer points. The volume and proximity of people in these areas can lead to an observed dramatic increase in the transmission of airborne viruses and related pathogens. Due to the critical role these areas play in transmission, it is vital that we have a comprehensive understanding of the ‘transmission highways’ in these areas to predict or prevent the spreading of infectious diseases in general. The principled approach of this paper is to combine and utilize as much information as possible from relevant sources and to integrate these data in a simulated environment that allows for scenario testing and decision support. In this paper, we describe a novel approach to study the spread of airborne diseases in cities by combining traffic information with geo-spatial data, infection dynamics and spreading characteristics. The system is currently being used in an attempt to understand the outbreak of influenza in densely populated cities in China.

During mammalian spermatogenesis, the diploid spermatogonia mature into haploid spermatozoa through a highly controlled process of mitosis, meiosis and post-meiotic morphological remodeling (spermiogenesis). Despite important progress made in this area, the molecular mechanisms underpinning this transformation are poorly understood. Our analysis of the expression and function of the putative serine–threonine kinase Fused (Fu) provides critical insight into key steps in spermatogenesis. In this report, we demonstrate that conditional inactivation of Fu in male germ cells results in infertility due to diminished sperm count, abnormal head shaping, decapitation and motility defects of the sperm. Interestingly, mutant flagellar axonemes are intact but exhibit altered periaxonemal structures that affect motility. These data suggest that Fu plays a central role in shaping the sperm head and controlling the organization of the periaxonemal structures in the flagellum. We show that Fu localizes to multiple tubulin-containing or microtubule-organizing structures, including the manchette and the acrosome–acroplaxome complex that are involved in spermatid head shaping. In addition, Fu interacts with the outer dense fiber protein Odf1, a major component of the periaxonemal structures in the sperm flagellum, and Kif27, which is detected in the manchette. We propose that disrupted Fu function in these structures underlies the head and flagellar defects in Fu -deficient sperm. Since a majority of human male infertility syndromes stem from reduced sperm motility and structural defects, uncovering Fu׳s role in spermiogenesis provides new insight into the causes of sterility and the biology of reproduction.