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Articles by L Zhang
Total Records ( 38 ) for L Zhang
  L Zhang , R Sheng and Z. Qin
 

It has long been believed that the lysosome is an important digestive organelle. There is increasing evidence that the lysosome is also involved in pathogenesis of a variety of neurodegenerative diseases, including Alzheimer's disease, Parkinson's disease, Huntington's disease, and amyotrophic lateral sclerosis. Abnormal protein degradation and deposition induced by lysosomal dysfunction may be the primary contributor to age-related neurodegeneration. In this review, the possible relationship between lysosome and various neurodegenerative diseases is described.

  L Zhang , Q Liu , Y Zhou and Y. Zhang
 

The male reproductive tracts in different species are characterized by similar patterns of male-dependent overexpression of carboxylesterases. This phenomenon indicates male sex-associated functions of these enzymes for spermatogenesis, sperm maturation, and sperm use. Recently, a novel epididymis-specific gene named Ces7 was cloned and characterized, which belongs to the carboxylesterase family. To study the functions of CES7 in sperm maturation and storage, CES7 recombinant protein was expressed in baculovirus system. The recombinant protein had carboxylesterase activity hydrolyzing cholesterol ester and choline ester. CES7 as carboxylesterase might be involved in ester hydrolysis, sperm maturation, and storage in male reproductive tract.

  L Zhang , Z Hu , C Zhu , Q Liu , Y Zhou and Y. Zhang
 

Carboxylesterases (CEs) represent a multigene family of serine-dependent enzymes. Male-dependent CEs are over-expressed in the male reproductive tract of different animal species (bivalve mollusks, fruit-flies, and mammals). Here, a novel rat epididymis-specific gene named Ces7 was cloned and characterized. It was a novel member of CE family, which was mainly expressed and secreted to the lumens of the corpus and cauda epididymis. CES7 protein was highly glycosylated as other mammalian CEs. Furthermore, Ces7 increased with age growth until sex maturation and then maintained at high level. CES7 might be one of the major CEs in male reproductive tract and contribute to the sperm fertilization.

  Y Min , W Xu , D Liu , S Shen , Y Lu , L Zhang and H. Wang
 

Dendritic cells (DCs) are important for the initiation of the adaptive immune response against Mycobacterium tuberculosis. Autophagy is an innate and adaptive defense mechanism and important for the control of M. tuberculosis. However, the role of autophagy in the adaptive immune response against M. tuberculosis remains to be determined. In the present study, we studied the effects of autophagy on the maturation of DCs infected with Bacillus Calmette–Guérin (BCG). The phenotype and function of the DCs were assessed by measuring the expression of CD86 and HLA-DR and the secretion of IL-10 and IL-6. Autophagy was evaluated by the change in LC3II, a molecular marker for autophagy. Following stimulation of autophagy, DCs that were matured in the presence of BCG showed enhanced expression of CD86 and HLA-DR and increased IL-6 production. The expression of LC3II was increased after the stimulation of autophagy. These results demonstrated that autophagy might result in the increased maturation of BCG-infected DCs, suggesting that autophagy could contribute to an enhanced adaptive immune response against M. tuberculosis.

  L Zhang , X Jia , X Peng , Q Ou , Z Zhang , C Qiu , Y Yao , F Shen , H Yang , F Ma , J Wang and Z. Yuan
 

This paper presents an liquid chromatography (LC)/mass spectrometry (MS)-based metabonomic platform that combined the discovery of differential metabolites through principal component analysis (PCA) with the verification by selective multiple reaction monitoring (MRM). These methods were applied to analyze plasma samples from liver disease patients and healthy donors. LC–MS raw data (about 1000 compounds), from the plasma of liver failure patients (n = 26) and healthy controls (n = 16), were analyzed through the PCA method and a pattern recognition profile that had significant difference between liver failure patients and healthy controls (P < 0.05) was established. The profile was verified in 165 clinical subjects. The specificity and sensitivity of this model in predicting liver failure were 94.3 and 100.0%, respectively. The differential ions with m/z of 414.5, 432.0, 520.5, and 775.0 were verified to be consistent with the results from PCA by MRM mode in 40 clinical samples, and were proved not to be caused by the medicines taken by patients through rat model experiments. The compound with m/z of 520.5 was identified to be 1-Linoleoylglycerophosphocholine or 1-Linoleoylphosphatidylcholine through exact mass measurements performed using Ion Trap–Time-of-Flight MS and METLIN Metabolite Database search. In all, it was the first time to integrate metabonomic study and MRM relative quantification of differential peaks in a large number of clinical samples. Thereafter, a rat model was used to exclude drug effects on the abundance of differential ion peaks. 1-Linoleoylglycerophosphocholine or 1-Linoleoylphosphatidylcholine, a potential biomarker, was identified. The LC/MS-based metabonomic platform could be a powerful tool for the metabonomic screening of plasma biomarkers.

  L Ji , F Fu , L Zhang , W Liu , X Cai , Q Zheng , H Zhang and F. Gao
 

It is well known that insulin possesses a cardioprotective effect and that insulin resistance is closely related to cardiovascular diseases. Peroxynitrite (ONOO) formation may trigger oxidative/nitrative stress and represent a major cytotoxic effect in heart diseases. This study was designed to investigate whether insulin attenuates ONOO generation and oxidative/nitrative stress in acute myocardial ischemia/reperfusion (MI/R). Adult male rats were subjected to 30 min of myocardial ischemia and 3 h of reperfusion. Rats randomly received vehicle, insulin, or insulin plus wortmannin. Arterial blood pressure and left ventricular pressure were monitored throughout the experiment. Insulin significantly improved cardiac functions and reduced myocardial infarction, apoptotic cell death, and blood creatine kinase/lactate dehydrogenase levels following MI/R. Myocardial ONOO formation was significantly attenuated after insulin treatment. Moreover, insulin resulted in a significant increase in Akt and endothelial nitric oxide (NO) synthase (eNOS) phosphorylation, NO production, and antioxidant capacity in ischemic/reperfused myocardial tissue. On the other hand, insulin markedly reduced MI/R-induced inducible NOS (iNOS) and gp91phox expression in cardiac tissue. Inhibition of insulin signaling with wortmannin not only blocked the cardioprotection of insulin but also markedly attenuated insulin-induced antioxidative/antinitrative effect. Furthermore, the suppression on ONOO formation by either insulin or an ONOO scavenger uric acid reduced myocardial infarct size in rats subjected to MI/R. We concluded that insulin exerts a cardioprotective effect against MI/R injury by blocking ONOO formation. Increased physiological NO production (via eNOS phosphorylation) and superoxide anion reduction contribute to the antioxidative/antinitrative effect of insulin, which can be reversed by inhibiting phosphatidylinositol 3'-kinase. These results provide important novel information on the mechanisms of cardiovascular actions of insulin.

  L Zhang , G. D Eslick , H. H. X Xia , C Wu , N Phung and N. J. Talley
 

Background: Helicobacter pylori (H. pylori) is a cause of chronic gastritis and maybe responsible for functional dyspepsia in a subset of patients. Many risk factors, such as alcohol consumption and smoking, may contribute to the colonization and infection of H. pylori in humans. However, studies on the relationship between H. pylori infection and drinking or smoking have produced conflicting results. Objective: The aim of this study was to examine whether consumption of alcohol or smoking is associated with active H. pylori infection in functional dyspepsia patients. Methods: H. pylori infection was confirmed by CLOtest and histology on at least two biopsies. Active chronic gastritis was diagnosed using the updated Sydney system. In addition to gender and age, information on drinking and smoking habits was collected using a standard questionnaire. Functional dyspepsia was diagnosed according to the Rome II diagnostic criteria. Results: H. pylori infection was positive in 27.3% of the 139 functional dyspepsia patients. Both age and gender were not significantly associated with H. pylori infection. A multiple logistic model found that alcohol consumption (OR = 9.05, 95% CI: 1.05–77.98) and pathology (active gastritis) (OR = 595.39, 95% CI: 81.43–4353.33) were associated with H. pylori infection. Active gastritis was associated with alcohol consumption (OR = 2.89, 95% CI: 1.03–8.02), smoking (OR = 2.72, 95% CI: 1.22–6.05) and age (OR = 1.03, 95% CI: 1.01–1.06). Conclusions: In patients with functional dyspepsia, there is no significant association between active H. pylori infection and smoking. However, alcohol consumption appears to be associated with H. pylori infection.

  J Fatima , T Schnelldorfer , J Barton , C. M Wood , H. J Wiste , T. C Smyrk , L Zhang , M. G Sarr , D. M Nagorney and M. B. Farnell
 

Objective  To assess the effect of R0 resection margin status and R0 en bloc resection in pancreatoduodenectomy outcomes.

Design  Retrospective medical record review.

Setting  Mayo Clinic, Rochester, Minnesota.

Patients  Patients who underwent pancreatoduodenectomy for pancreatic adenocarcinoma at our institution between January 1, 1981, and December 31, 2007, were identified and their medical records were reviewed.

Main Outcome Measure  Median survival times.

Results  A total of 617 patients underwent pancreatoduodenectomy. Median survival times after R0 en bloc resection (n = 411), R0 non–en bloc resection (n = 57), R1 resection (n = 127), and R2 resection (n = 22) were 19, 18, 15, and 10 months, respectively (P < .001). A positive resection margin was associated with death (P = .01). No difference in survival time was found between patients undergoing R0 en bloc and R0 resections after reexcision of an initial positive margin (hazard ratio, 1.19; 95% confidence interval, 0.87-1.64; P = .28).

Conclusions  R0 resection remains an important prognostic factor. Achieving R0 status by initial en bloc resection or reexcision results in similar long-term survival.

  M Zhang , L Zhang , J Zou , C Yao , H Xiao , Q Liu , J Wang , D Wang , C Wang and Z. Guo
 

Motivation: According to current consistency metrics such as percentage of overlapping genes (POG), lists of differentially expressed genes (DEGs) detected from different microarray studies for a complex disease are often highly inconsistent. This irreproducibility problem also exists in other high-throughput post-genomic areas such as proteomics and metabolism. A complex disease is often characterized with many coordinated molecular changes, which should be considered when evaluating the reproducibility of discovery lists from different studies.

Results: We proposed metrics percentage of overlapping genes-related (POGR) and normalized POGR (nPOGR) to evaluate the consistency between two DEG lists for a complex disease, considering correlated molecular changes rather than only counting gene overlaps between the lists. Based on microarray datasets of three diseases, we showed that though the POG scores for DEG lists from different studies for each disease are extremely low, the POGR and nPOGR scores can be rather high, suggesting that the apparently inconsistent DEG lists may be highly reproducible in the sense that they are actually significantly correlated. Observing different discovery results for a disease by the POGR and nPOGR scores will obviously reduce the uncertainty of the microarray studies. The proposed metrics could also be applicable in many other high-throughput post-genomic areas.

  A Di Stasi , B De Angelis , C. M Rooney , L Zhang , A Mahendravada , A. E Foster , H. E Heslop , M. K Brenner , G Dotti and B. Savoldo
 

For the adoptive transfer of tumor-directed T lymphocytes to prove effective, there will probably need to be a match between the chemokines the tumor produces and the chemokine receptors the effector T cells express. The Reed-Stemberg cells of Hodgkin lymphoma (HL) predominantly produce thymus- and activation-regulated chemokine/CC chemokine ligand 17 (TARC/CCL17) and macrophage-derived chemokine (MDC/CCL22), which preferentially attract type 2 T helper (Th2) cells and regulatory T cells (Tregs) that express the TARC/MDC-specific chemokine receptor CCR4, thus generating an immunosuppressed tumor environment. By contrast, effector CD8+ T cells lack CCR4, are nonresponsive to these chemokines and are rarely detected at the tumor site. We now show that forced expression of CCR4 by effector T cells enhances their migration to HL cells. Furthermore, T lymphocytes expressing both CCR4 and a chimeric antigen receptor directed to the HL associated antigen CD30 sustain their cytotoxic function and cytokine secretion in vitro, and produce enhanced tumor control when infused intravenously in mice engrafted with human HL. This approach may be of value in patients affected by HL.

  L Zhang , S. F Messner , J Liu and Y. A. Zhuo
 

Western research has investigated individual correlates of fear of crime with a primary focus on people's vulnerability. This vulnerability model examines the possible effects on fear of indicators of people's physical vulnerability (e.g. age and gender) and social vulnerability (e.g. income and education). As is well documented in the research on China, guanxi is a unique aspect of social capital in Chinese society. The present study argues that guanxi in the immediate neighbourhood is an important indicator of the social vulnerability of individuals in urban China. We accordingly hypothesize that residents who have strong neighbourhood guanxi are less likely to be fearful of crime. This hypothesis is assessed with data collected from a recent survey in the city of Tianjin, China. The results of multilevel analysis show that guanxi in the neighbourhood is a significant predictor of fear of crime in contemporary urban China when other important factors are controlled.

  T Holopainen , H Huang , C Chen , K. E Kim , L Zhang , F Zhou , W Han , C Li , J Yu , J Wu , G. Y Koh , K Alitalo and Y. He
 

The angiopoietin-1 (Ang1)/Tie2 signaling pathway is known to play an important role in the regulation of vascular maturation and maintenance of vessel integrity. In this study, we have investigated the effect of systemic Tie2 activation or inhibition on tumor growth and metastasis. We found that treatment with Ang1 delivered via an adenoviral vector promoted s.c. implanted tumor metastasis to the lungs. Ang1 treatment did not significantly increase vascular density in the tumors but induced enlargement of blood vessels in both the tumor and normal tissues, which increased tumor cell dissemination into the blood circulation. Ang1 also enhanced the formation of metastatic foci in the lungs when tumor cells were injected into the circulation via the tail vein. The effect of Ang1 on metastasis was validated by a simultaneous treatment with a soluble form of Tie2 (sTie2), which led to the suppression of Ang1-induced increase of tumor metastasis. Furthermore, using a highly metastatic tumor model, we confirmed that systemic treatment with sTie2 suppressed tumor metastasis to the lungs and lymph nodes, whereas tumor-associated angiogenesis and lymphangiogenesis were not significantly affected. This suggests that the Ang1/Tie2 signals contribute to tumor progression by increasing vascular entry and exit of tumor cells to facilitate tumor dissemination and establishment of metastases. [Cancer Res 2009;69(11):4656–64]

  W Qiu , E. B Carson Walter , S. F Kuan , L Zhang and J. Yu
 

Defective apoptosis contributes to tumorigenesis, although the critical molecular targets remain to be fully characterized. PUMA, a BH3-only protein essential for p53-dependent apoptosis, has been shown to suppress lymphomagenesis. In this study, we investigated the role of PUMA in intestinal tumorigenesis using two animal models. In the azoxymethane (AOM)/dextran sulfate sodium salt model, PUMA deficiency increased the multiplicity and size of colon tumors but reduced the frequency of β-catenin hotspot mutations. The absence of PUMA led to a significantly elevated incidence of precursor lesions induced by AOM. AOM was found to induce p53-dependent PUMA expression and PUMA-dependent apoptosis in the colonic crypts and stem cell compartment. Furthermore, PUMA deficiency significantly enhanced the formation of spontaneous macroadenomas and microadenomas in the distal small intestine and colon of APCMin/+ mice. These results show an essential role of PUMA-mediated apoptosis in suppressing intestinal tumorigenesis in mice. [Cancer Res 2009;69(12):4999–5006]

  L Zhang , T Deng , X Li , H Liu , H Zhou , J Ma , M Wu , M Zhou , S Shen , Z Niu , W Zhang , L Shi , B Xiang , J Lu , L Wang , D Li , H Tang and G. Li
 

microRNAs (miRNAs) are small non-coding RNAs and have been implicated in the pathology of various diseases, including cancer. Here we report that the miRNA profiles have been changed after knockdown of one of the most important oncogene c-MYC or re-expression of a candidate tumor suppressor gene SPLUNC1 in nasopharyngeal carcinoma (NPC) cells. Both c-MYC knockdown and SPLUNC1 re-expression can down-regulate microRNA-141 (miR-141). miR-141 is up-regulated in NPC specimens in comparison with normal nasopharyngeal epithelium. Inhibition of miR-141 could affect cell cycle, apoptosis, cell growth, migration and invasion in NPC cells. We found that BRD3, UBAP1 and PTEN are potential targets of miR-141, which had been confirmed following luciferase reporter assays and western blotting. BRD3 and UBAP1 are both involved in NPC carcinogenesis as confirmed through our previous studies and PTEN is a crucial tumor suppressor in many tumor types. BRD3 is involved in the regulation of the Rb/E2F pathway. Inhibition of miR-141 could affect some important molecules in the Rb/E2F, JNK2 and AKT pathways. It is well known that carcinogenesis of NPC is involved in the networks of genetic and epigenetic alteration events. We propose that miR-141- and tumor-related genes c-MYC, SPLUNC1, BRD3, UBAP1 and PTEN may constitute a gene–miRNA network to contribute to NPC development.

  L Ding , L Dong , X Chen , L Zhang , X Xu , A Ferro and B. Xu
 

Background— Left ventricular (LV) remodeling is associated with the development of heart failure after myocardial infarction. Here we investigated whether integrin-linked kinase (ILK) may regulate LV remodeling and function after myocardial infarction.

Methods and Results— Adenoviral vector expressing ILK (n=25) or empty adeno-null (n=25) was injected into rat peri-infarct myocardium after left anterior descending coronary artery ligation. ILK expression was confirmed by Western blotting and immunofluorescence. Echocardiographic and hemodynamic analyses demonstrated relatively preserved cardiac function in adeno-ILK animals. ILK treatment was associated with reduced infarct scar size, increased scar thinning ratio, and preserved LV diameter, wall thickness, cardiomyocyte size, and myofilament density. Enhanced angiogenesis and reduced fibrosis were observed in the adeno-ILK group, along with reduced apoptosis as demonstrated by terminal deoxynucleotidyl transferase–mediated dUTP nick-end labeling analysis. Moreover, increased cardiomyocyte proliferation was found in adeno-ILK animals, as measured by proliferating cell nuclear antigen, Ki-67, and phosphohistone-H3 staining. At long-term follow-up, most indices of cardiac function and hemodynamics showed no difference between adeno-ILK and control animals by 9 weeks, although LV end-systolic diameter and infarct scar size were reduced in the adeno-ILK group at this time point. Additionally, ILK overexpression was found to exert a rescue effect on remodeling when administered in a delayed fashion 1 week after coronary artery ligation.

Conclusions— ILK gene therapy improves cardiac remodeling and function in rats after myocardial infarction and is associated with increased angiogenesis, reduced apoptosis, and increased cardiomyocyte proliferation. This may represent a new approach to the treatment of postinfarct remodeling and subsequent heart failure.

  X. P Huang , Z Sun , Y Miyagi , H McDonald Kinkaid , L Zhang , R. D Weisel and R. K. Li
  Background—

Cardiac cell therapy for older patients who experience a myocardial infarction may require highly regenerative cells from young, healthy (allogeneic) donors. Bone marrow mesenchymal stem cells (MSCs) are currently under clinical investigation because they can induce cardiac repair and may also be immunoprivileged (suitable for allogeneic applications). However, it is unclear whether allogeneic MSCs retain their immunoprivilege or functional efficacy late after myocardial implantation. We evaluated the effects of MSC differentiation on the immune characteristics of cells in vitro and in vivo and monitored cardiac function for 6 months after post–myocardial infarction MSC therapy.

Methods and Results—

In the in vitro experiments, inducing MSCs to acquire myogenic, endothelial, or smooth muscle characteristics (via 5-azacytidine or cytokine treatment) increased major histocompatibility complex-Ia and -II (immunogenic) expression and reduced major histocompatibility complex-Ib (immunosuppressive) expression, in association with increased cytotoxicity in coculture with allogeneic leukocytes. In the in vivo experiments, we implanted allogeneic or syngeneic MSCs into infarcted rat myocardia. We measured cell differentiation and survival (immunohistochemistry, real-time polymerase chain reaction) and cardiac function (echocardiography, pressure-volume catheter) for 6 months. MSCs (versus media) significantly improved ventricular function for at least 3 months after implantation. Allogeneic (but not syngeneic) cells were eliminated from the heart by 5 weeks after implantation, and their functional benefits were lost within 5 months.

Conclusions—

The long-term ability of allogeneic MSCs to preserve function in the infarcted heart is limited by a biphasic immune response whereby they transition from an immunoprivileged to an immunogenic state after differentiation, which is associated with an alteration in major histocompatibility complex–immune antigen profile.

  S. M Schumacher , D. P McEwen , L Zhang , K. L Arendt , K. M Van Genderen and J. R. Martens
 

Conventional antiarrhythmic drugs target the ion permeability of channels, but increasing evidence suggests that functional ion channel density can also be modified pharmacologically. Kv1.5 mediates the ultrarapid potassium current (IKur) that controls atrial action potential duration. Given the atrial-specific expression of Kv1.5 and its alterations in human atrial fibrillation, significant effort has been made to identify novel channel blockers. In this study, treatment of HL-1 atrial myocytes expressing Kv1.5-GFP with the class I antiarrhythmic agent quinidine resulted in a dose- and temperature-dependent internalization of Kv1.5, concomitant with channel block. This quinidine-induced channel internalization was confirmed in acutely dissociated neonatal myocytes. Channel internalization was subunit-dependent, activity-independent, stereospecific, and blocked by pharmacological disruption of the endocytic machinery. Pore block and channel internalization partially overlap in the structural requirements for drug binding. Surprisingly, quinidine-induced endocytosis was calcium-dependent and therefore unrecognized by previous biophysical studies focused on isolating channel–drug interactions. Importantly, whereas acute quinidine-induced internalization was reversible, chronic treatment led to channel degradation. Together, these data reveal a novel mechanism of antiarrhythmic drug action and highlight the possibility for new agents that selectively modulate the stability of channel protein in the membrane as an approach for treating cardiac arrhythmias.

  J Cheng , Y Wang , Y Ma , B. T. y Chan , M Yang , A Liang , L Zhang , H Li and J. Du
  Rationale:

Mechanical stress plays an important role in proliferation of venous smooth muscle cells (SMCs) in neointima, a process of formation that contributes to failure of vein grafts. However, it is unknown what intracellular growth signal leads to proliferation of venous SMCs.

Objective:

The objective of this study is to identify mechanisms of mechanical stretch on neointima formation.

Methods and Results:

By a microarray analysis, we found that mechanical cyclic stretch (15% elongation) stimulated the transcription of SGK-1 (serum-, glucocorticoid-regulated kinase-1). Mechanical stretch–induced SGK-1 mRNA expression was blocked by actinomycin D. The mechanism for the SGK-1 expression involved MEK1 but not p38 or JNK signaling pathway. SGK-1 activation in response to stretch is blocked by insulin-like growth factor (IGF)-1 receptor inhibitor and mammalian target of rapamycin complex (mTORC)2 inhibitor (Ku-0063794) but not mTORC1 inhibitor (rapamycin). Mechanical stretch–induced bromodeoxyuridine incorporation was reduced by 83.5% in venous SMCs isolated from SGK-1 knockout mice. In contrast, inhibition of Akt, another downstream signal of PI3K resulted in only partial inhibition of mechanical stretch–induced proliferation of venous SMCs. Mechanical stretch also induced phosphorylation and nuclear exportation of p27kip1, whereas knockout of SGK-1 attenuated this effect of mechanical stretch on p27kip1. In vivo, we found that placement of a vein graft into artery increased SGK-1 expression. Knockout of SGK-1 effectively prevented neointima formation in vein graft. There is significant lower level of p27kip1 located in the nucleus of neointima cells in SGK-1 knockout mice compared with that of wild-type vein graft. In addition, we also found that wire injury of artery or growth factors in vitro increased expression of SGK-1.

Conclusions:

These results suggest that SGK-1 is an injury-responsive kinase that could mediate mechanical stretch–induced proliferation of vascular cells in vein graft, leading to neointima formation.

  H Ding , Y Xu , X Wang , Q Wang , L Zhang , Y Tu , J Yan , W Wang , R Hui , C. Y Wang and D. W. Wang
 

Background— Recent studies on genome-wide association have identified common variants on chromosome 9p21 associated with coronary artery disease (CAD). Given that ischemic stroke and CAD share several aspects of etiology and pathogenesis, we investigated the association of variants on chromosome 9p21 with ischemic stroke and CAD in the Chinese Han population by capturing the majority of diversity in this locus using haplotype-tagging single-nucleotide polymorphisms.

Methods and Results— We performed a shared control-cases study using 15 tagging single-nucleotide polymorphisms and 2 previously reported susceptibility single-nucleotide polymorphisms spanning 58 kb of the chromosome of 9p21 in a set of 558 patients with ischemic stroke, 510 patients with CAD, and 557 unaffected participants (controls) in the Chinese Han population. The association analyses were performed at both SNP and haplotype levels. We further verified our findings in an independent cohort of 442 ischemic stroke cases and 502 control subjects. In the first study, rs2383206, rs1004638, and rs10757278 in block 3 were significantly associated with CAD but not with ischemic stroke independent of traditional cardiovascular risk factors in additive model (P=0.002 to 0.0001, q=0.026 to 0.004). Analysis from all blocks revealed that haplotype profiles of block 3 on 9p21 were significantly different between shared control and cases of CAD (P=1.3x10–10, q=1.2x10–9) and ischemic stroke (P=1.7x10–6, q=7.7x10–6). In the expanded second case-control study, block 3 on 9p21 remained associated with ischemic stroke (P=2.6x10–4, q=6.3x10–4).

Conclusions— Our results suggest for the first time that 9p21 is a shared susceptibility locus, strongly for CAD and weakly for ischemic stroke, in a Chinese Han population.

  K Zhang , L Zhang , F Rao , B Brar , J. L Rodriguez Flores , L Taupenot and D. T. O'Connor
 

Background— Tyrosine hydroxylase (TH) is the rate-limiting enzyme in catecholamine biosynthesis. Common genetic variation at the human TH promoter predicts alterations in autonomic activity and blood pressure, but how such variation influences human traits and, specifically, whether such variation affects transcription are not yet known.

Methods and Results— Pairwise linkage disequilibrium across the TH locus indicated that common promoter variants (C-824T, G-801C, A-581G, and G-494A) were located in a single 5' linkage disequilibrium block in white, black, Hispanic, and Asian populations. Polymorphisms C-824T and A-581G were located in highly conserved regions and were predicted to disrupt known transcriptional control motifs myocyte enhancer factor-2 (MEF2), sex-determining region Y (SRY), and forkhead box D1 (FOXD1) at C-824T and G/C-rich binding factors specificity protein 1 (SP1), activating enhancer-binding protein 2 (AP2)], early growth response protein 1 (EGR1) at A-581G. At C-824T and A-581G, promoter and luciferase reporter plasmids indicated differential allele strength (T>C at C-824T; G>A at A-581G) under both basal circumstances and secretory stimulation. C-824T and A-581G displayed the most pronounced effects on both transcription in cella and catecholamine secretion in vivo. We further probed the functional significance of C-824T and A-581G by cotransfection of trans-activating factors in cella; MEF2, SRY, and FOXD1 differentially activated C-824T, whereas the G/C-rich binding factors SP1, AP2, and EGR1 differentially activated A-581G. At C-824T, factor MEF2 acted in a directionally coordinate fashion (at T>C) to explain the in vivo trait associations, whereas at A-581G, factors SP1, AP2, and EGR1 displayed similar differential actions (at G>A). Finally, chromatin immunoprecipitation demonstrated that the endogenous factors bound to the motifs in cella.

Conclusion— We conclude that common genetic variants in the proximal TH promoter, especially at C-824T and A-581G, are functional in cella and alter transcription so as to explain promoter marker-on-trait associations in vivo. MEF2, FOXD1, and SRY contribute to functional differences in C-824T expression, whereas SP1, AP2, and EGR1 mediate those of A-581G. The SRY effect on TH transcription suggests a mechanism whereby male and female sex may differ in sympathetic activity and hence blood pressure. These results point to new strategies for diagnostic and therapeutic intervention into disorders of human autonomic function and their cardiovascular consequences.

  K. J McKernan , H. E Peckham , G. L Costa , S. F McLaughlin , Y Fu , E. F Tsung , C. R Clouser , C Duncan , J. K Ichikawa , C. C Lee , Z Zhang , S. S Ranade , E. T Dimalanta , F. C Hyland , T. D Sokolsky , L Zhang , A Sheridan , H Fu , C. L Hendrickson , B Li , L Kotler , J. R Stuart , J. A Malek , J. M Manning , A. A Antipova , D. S Perez , M. P Moore , K. C Hayashibara , M. R Lyons , R. E Beaudoin , B. E Coleman , M. W Laptewicz , A. E Sannicandro , M. D Rhodes , R. K Gottimukkala , S Yang , V Bafna , A Bashir , A MacBride , C Alkan , J. M Kidd , E. E Eichler , M. G Reese , F. M De La Vega and A. P. Blanchard
 

We describe the genome sequencing of an anonymous individual of African origin using a novel ligation-based sequencing assay that enables a unique form of error correction that improves the raw accuracy of the aligned reads to >99.9%, allowing us to accurately call SNPs with as few as two reads per allele. We collected several billion mate-paired reads yielding ~18x haploid coverage of aligned sequence and close to 300x clone coverage. Over 98% of the reference genome is covered with at least one uniquely placed read, and 99.65% is spanned by at least one uniquely placed mate-paired clone. We identify over 3.8 million SNPs, 19% of which are novel. Mate-paired data are used to physically resolve haplotype phases of nearly two-thirds of the genotypes obtained and produce phased segments of up to 215 kb. We detect 226,529 intra-read indels, 5590 indels between mate-paired reads, 91 inversions, and four gene fusions. We use a novel approach for detecting indels between mate-paired reads that are smaller than the standard deviation of the insert size of the library and discover deletions in common with those detected with our intra-read approach. Dozens of mutations previously described in OMIM and hundreds of nonsynonymous single-nucleotide and structural variants in genes previously implicated in disease are identified in this individual. There is more genetic variation in the human genome still to be uncovered, and we provide guidance for future surveys in populations and cancer biopsies.

  S Maegawa , G Hinkal , H. S Kim , L Shen , L Zhang , J Zhang , N Zhang , S Liang , L. A Donehower and J. P. J. Issa
 

Aberrant methylation of promoter CpG islands in cancer is associated with silencing of tumor-suppressor genes, and age-dependent hypermethylation in normal appearing mucosa may be a risk factor for human colon cancer. It is not known whether this age-related DNA methylation phenomenon is specific to human tissues. We performed comprehensive DNA methylation profiling of promoter regions in aging mouse intestine using methylated CpG island amplification in combination with microarray analysis. By comparing C57BL/6 mice at 3-mo-old versus 35-mo-old for 3627 detectable autosomal genes, we found 774 (21%) that showed increased methylation and 466 (13%) that showed decreased methylation. We used pyrosequencing to quantitatively validate the microarray data and confirmed linear age-related methylation changes for all 12 genomic regions examined. We then examined 11 changed genomic loci for age-related methylation in other tissues. Of these, three of 11 showed similar changes in lung, seven of 11 changed in liver, and six of 11 changed in spleen, though to a lower degree than the changes seen in colon. There was partial conservation between age-related hypermethylation in human and mouse intestines, and Polycomb targets in embryonic stem cells were enriched among the hypermethylated genes. Our findings demonstrate a surprisingly high rate of hyper- and hypomethylation as a function of age in normal mouse small intestine tissues and a strong tissue-specificity to the process. We conclude that epigenetic deregulation is a common feature of aging in mammals.

  R. C DeKelver , V. M Choi , E. A Moehle , D. E Paschon , D Hockemeyer , S. H Meijsing , Y Sancak , X Cui , E. J Steine , J. C Miller , P Tam , V. V Bartsevich , X Meng , I Rupniewski , S. M Gopalan , H. C Sun , K. J Pitz , J. M Rock , L Zhang , G. D Davis , E. J Rebar , I. M Cheeseman , K. R Yamamoto , D. M Sabatini , R Jaenisch , P. D Gregory and F. D. Urnov
 

Isogenic settings are routine in model organisms, yet remain elusive for genetic experiments on human cells. We describe the use of designed zinc finger nucleases (ZFNs) for efficient transgenesis without drug selection into the PPP1R12C gene, a "safe harbor" locus known as AAVS1. ZFNs enable targeted transgenesis at a frequency of up to 15% following transient transfection of both transformed and primary human cells, including fibroblasts and hES cells. When added to this locus, transgenes such as expression cassettes for shRNAs, small-molecule-responsive cDNA expression cassettes, and reporter constructs, exhibit consistent expression and sustained function over 50 cell generations. By avoiding random integration and drug selection, this method allows bona fide isogenic settings for high-throughput functional genomics, proteomics, and regulatory DNA analysis in essentially any transformed human cell type and in primary cells.

  L Zhang , K Lau , J Cheng , H Yu , Y Li , G Sugiarto , S Huang , L Ding , V Thon , P. G Wang and X. Chen
 

Lewis x (Lex) and sialyl Lewis x (SLex)-containing glycans play important roles in numerous physiological and pathological processes. The key enzyme for the final step formation of these Lewis antigens is 1-3-fucosyltransferase. Here we report molecular cloning and functional expression of a novel Helicobacter hepaticus 1-3-fucosyltransferase (HhFT1) which shows activity towards both non-sialylated and sialylated Type II oligosaccharide acceptor substrates. It is a promising catalyst for enzymatic and chemoenzymatic synthesis of Lex, sialyl Lex and their derivatives. Unlike all other 1-3/4-fucosyltransferases characterized so far which belong to Carbohydrate Active Enzyme (CAZy, http://www.cazy.org/) glycosyltransferase family GT10, the HhFT1 shares protein sequence homology with 1-2-fucosyltransferases and belongs to CAZy glycosyltransferase family GT11. The HhFT1 is thus the first 1-3-fucosyltransferase identified in the GT11 family.

  L Zhang , J Chen , Y Wang , F Ren , W Yu and L. Cheng
  BACKGROUND

Levonorgestrel (LNG), as a dedicated emergency contraception (EC) product, has been available over-the-counter in China for 10 years. Until now, only a small number of deliveries after LNG–EC failure have been documented.

METHODS

This study was a prospective comparative cohort study. A group of 332 pregnant women who had used LNG–EC during the conception cycle was recruited, and matched to a group of 332 pregnant women without the exposure to LNG. Congenital malformations, perinatal complications and delivery circumstances were investigated in this study.

RESULTS

There were 31 pregnant women in the study group and 28 in the comparison group miscarried within 14 weeks of gestation. In the study and comparison groups, four malformations were found in each group. In the study group, both birthweight (3416 versus 3345 g, P = 0.040) and the sex ratio of birth (boys/girls, 1.14 versus 0.90, P = 0.153) were higher than in the comparison group. There were no statistically significant differences in the incidence of miscarriage or malformation or in the neonatal outcome between the two groups.

CONCLUSIONS

There was no association between the use of LNG–EC pills and the risk of major congenital malformations, pregnancy complications or any other adverse pregnancy outcomes in our study.

  D Frenkel , A. S Pachori , L Zhang , A Dembinsky Vaknin , D Farfara , S Petrovic Stojkovic , V. J Dzau and H. L. Weiner
 

Myocardial ischemia with subsequent reperfusion (MI/R) can lead to significant myocardial damage. Ischemia initiates inflammation at the blood–microvascular endothelial cell interface and contributes significantly to both acute injury and repair of the damaged tissue. We have found that MI/R injury in mice is associated with a cellular immune response to troponin. Myocardial cells exclusively synthesize troponin and release the troponin into the bloodstream following injury. Mucosally administered proteins induce T cells that secrete anti-inflammatory cytokines such as IL-10 and transforming growth factor β at the anatomical site where the protein localizes. We found that nasal administration of the three subunits of troponin (C, I and T isoforms), given prior to or 1 h following MI/R, decreased infarct size by 40% measured 24 h later. At 1.5 months following MI/R, there was a 50% reduction in infarct size and improvement in cardiac function as measured by echocardiography. Protection was associated with a reduction of cellular immunity to troponin. Immunohistochemistry demonstrated increased IL-10 and reduced IFN- in the area surrounding the ischemic infarct following nasal troponin. Adoptive transfer of CD4+ T cells to mice from nasally troponin-treated mice 1 h after the MI/R decreased infarct size by 72%, whereas CD4+ T cells from IL-10–/– mice or nasally BSA-treated mice had no effect. Our results demonstrate that IL-10-secreting CD4+ T cells induced by nasal troponin reduce injury following MI/R. Modulation of cardiac inflammation by nasal troponin provides a novel treatment to decrease myocardial damage and enhance recovery after myocardial ischemia.

  C Zhou , S Ren , S Zhou , L Zhang , C Su , Z Zhang , Q Deng and J. Zhang
  Objective

The purpose of the study was to investigate whether single-nucleotide polymorphisms of deoxyribonucleic acid repair gene excision repair cross-complementing group 1 at codon 118 and X-ray repair cross-complementing group 3 at codon 241 affected clinical outcomes in advanced non-small cell lung cancer patients receiving first-line platinum-based chemotherapy.

Methods

A total of 130 patients treated with platinum-based doublets were examined for genotyping of excision repair cross-complementing group 1 118 and X-ray repair cross-complementing group 3 241 in peripheral blood lymphocytes with the method of the TaqMan assay plus the real-time polymerase chain reaction method. Multivariate logistic or Cox's regression analyses were used to adjust for possible confounding variables.

Results

There were no differences in clinical characteristics among the different single-nucleotide polymorphisms. Overall response rate in the 130 patients was 20% with 85.4% of disease control rate. Followed up to 31 March 2008, there were 47 patients still alive. Overall survival was 15 months. No relationship was found between excision repair cross-complementing group 1 or X-ray repair cross-complementing group 3 single-nucleotide polymorphisms and tumor response to platinum-based chemotherapy. A significant correlation was found between excision repair cross-complementing group 1 118C/T single-nucleotide polymorphisms and survival (P = 0.003). In the multivariate model, the survival was highly related with excision repair cross-complementing group 1 118 C/T or T/T genotypes and tumor response to chemotherapy.

Conclusions

Overall survival was significantly improved in the patients with excision repair cross-complementing group 1 118 T/T or C/T treated by platinum-based chemotherapy.

  S Wang , L Zhang and M. Matz
 

Mining for microsatellites (also called simple sequence repeats [SSRs]) in public sequence databases of a common Indo-Pacific coral Acropora millepora identified 191 SSRs from 10 258 expressed sequence tag (EST) and 618 SSRs from 14 625 whole-genome shotgun (WGS) sequences. In contrast to other animals, trinucleotide repeats, rather than dinucleotide repeats, are dominant in the WGS-SSRs, and AAT is the most frequent trinucleotide motif in EST-SSRs. We successfully developed 40 polymorphic markers from EST-SSRs and WGS-SSRs. Both EST- and WGS-SSRs show high levels of polymorphism within corals from the same reef patch. Interestingly, markers WGS079 and WGS227 revealed SSR duplications in a few individuals, suggesting recent duplication events. Genotypic linkage disequilibrium was identified in 5 pairs of SSR markers, which will be invaluable for high-resolution studies of genetic admixture in natural populations of A. millepora. Transferability analysis showed that 25 of these markers can be successfully amplified in one of the most ubiquitous Indo-Pacific corals Acropora hyacinthus. The marker collection reported here is the largest ever developed for any reef-building coral. It holds great potential for addressing coral reef connectivity across the Indo-Pacific with an unprecedented precision, especially taking into account the cross-species transferability of a substantial number of markers.

  L Zhang , L. P Renaud and M. Kolaj
 

Burst firing mediated by a low-threshold spike (LTS) is the hallmark of many thalamic neurons. However, postburst afterhyperpolarizations (AHPs) are relatively uncommon in thalamus. We now report data from patch-clamp recordings in rat brain slice preparations that reveal an LTS-induced slow AHP (sAHP) in thalamic paraventricular (PVT) and other midline neurons, but not in ventrobasal or reticular thalamic neurons. The LTS-induced sAHP lasts 8.9 ± 0.4 s and has a novel pharmacology, with resistance to tetrodotoxin and cadmium and reduction by Ni2+ or nominally zero extracellular calcium concentration, which also attenuate both the LTS and sAHP. The sAHP is inhibited by 10 mM intracellular EGTA or by equimolar replacement of extracellular Ca2+ with Sr2+, consistent with select activation of LVA T-type Ca2+ channels and subsequent Ca2+ influx. In control media, the sAHP reverses near EK+, shifting to –78 mV in 10.1 mM [K+]o and is reduced by Ba2+ or tetraethylammonium. Although these data are consistent with opening of Ca2+-activated K+ channels, this sAHP lacks sensitivity to specific Ca2+-activated K+ channel blockers apamin, iberiotoxin, charybdotoxin, and UCL-2077. The LTS-induced sAHP is suppressed by a β-adrenoceptor agonist isoproterenol, a serotonin 5-HT7 receptor agonist 5-CT, a neuropeptide orexin-A, and by stimulation of the cAMP/protein kinase A pathway with 8-Br-cAMP and forskolin. The data suggest that PVT and certain midline thalamic neurons possess an LTS-induced sAHP that is pharmacologically distinct and may be important for information transfer in thalamic–limbic circuitry during states of attentiveness and motivation.

  X Fang , X Li , B Stanton , Y Hong , L Zhang , G Zhao , J Zhao , X Lin and D. Lin
 

Objective To assess the relationship between parental HIV/AIDS and psychosocial adjustment of children in rural central China. Methods Participants included 296 double AIDS orphans (children who had lost both their parents to AIDS), 459 single orphans (children who had lost one parent to AIDS), 466 vulnerable children who lived with HIV-infected parents, and 404 comparison children who did not experience HIV/AIDS-related illness and death in their families. The measures included depressive symptoms, loneliness, self-esteem, future expectations, hopefulness about the future, and perceived control over the future. Results AIDS orphans and vulnerable children consistently demonstrated poorer psychosocial adjustment than comparison children in the same community. The level of psychosocial adjustment was similar between single orphans and double orphans, but differed by care arrangement among double orphans. Conclusion The findings underscore the urgency and importance of culturally and developmentally appropriate intervention efforts targeting psychosocial problems among children affected by AIDS and call for more exploration of risk and resilience factors, both individual and contextual, affecting the psychosocial wellbeing of these children.

  G. S Song , H. L Zhai , Y. G Peng , L Zhang , G Wei , X. Y Chen , Y. G Xiao , L Wang , Y. J Chen , B Wu , B Chen , Y Zhang , H Chen , X. J Feng , W. K Gong , Y Liu , Z. J Yin , F Wang , G. Z Liu , H. L Xu , X. L Wei , X. L Zhao , P. B. F Ouwerkerk , T Hankemeier , T Reijmers , R. v. d Heijden , C. M Lu , M Wang , J. v. d Greef and Z. Zhu
 

Heterosis is a biological phenomenon whereby the offspring from two parents show improved and superior performance than either inbred parental lines. Hybrid rice is one of the most successful apotheoses in crops utilizing heterosis. Transcriptional profiling of F1 super-hybrid rice Liangyou-2186 and its parents by serial analysis of gene expression (SAGE) revealed 1183 differentially expressed genes (DGs), among which DGs were found significantly enriched in pathways such as photosynthesis and carbon-fixation, and most of the key genes involved in the carbon-fixation pathway exhibited up-regulated expression in F1 hybrid rice. Moreover, increased catabolic activity of corresponding enzymes and photosynthetic efficiency were also detected, which combined to indicate that carbon fixation is enhanced in F1 hybrid, and might probably be associated with the yield vigor and heterosis in super-hybrid rice. By correlating DGs with yield-related quantitative trait loci (QTL), a potential relationship between differential gene expression and phenotypic changes was also found. In addition, a regulatory network involving circadian-rhythms and light signaling pathways was also found, as previously reported in Arabidopsis, which suggest that such a network might also be related with heterosis in hybrid rice. Altogether, the present study provides another view for understanding the molecular mechanism underlying heterosis in rice.

  Z Ji , L Zhang , W Guo , C. M McHale and M. T. Smith
 

Both occupational exposure to the leukemogen benzene and in vitro exposure to its metabolite hydroquinone (HQ) lead to the induction of numerical and structural chromosome changes. Several studies have shown that HQ can form DNA adducts, disrupt microtubule assembly and inhibit DNA topoisomerase II (topo II) activity. As these are potential mechanisms underlying endoreduplication (END), a phenomenon that involves DNA amplification without corresponding cell division, we hypothesized that HQ could cause END. We measured END in the human lymphoblastoid cell line, TK6, treated with HQ (0–20 µM) and etoposide (0–0.2 µM) for 48 h. Etoposide was used as a positive control as it is a topo II poison and established human leukemogen that has previously been shown to induce END in Chinese hamster ovary cells. Both HQ and etoposide significantly induced END in a dose-dependent manner (Ptrend < 0.0001 and Ptrend = 0.0003, respectively). Since END may underlie the acquisition of high chromosome numbers by tumour cells, it may play a role in inducing genomic instability and subsequent carcinogenesis from HQ and etoposide. In order to further explore the cytogenetic effects of HQ and etoposide, we also examined specific structural changes. HQ did not induce translocations of chromosome 11 [t(11;?)] but significantly induced translocations of chromosome 21 [t(21;?)] and structural chromosome aberrations (SCA) (Ptrend = 0.0415 and Ptrend < 0.0001, respectively). Etoposide potently induced all these structural changes (Ptrend < 0.0001). The lack of an effect of HQ on t(11;?) and the reduced ability of HQ to induce t(21;?) and SCA, compared with etoposide, further suggests that HQ acts primarily as a topo II catalytic inhibitor rather than as a topo II poison in intact human cells.

  Y Xu , H Lei , H Dong , L Zhang , Q Qin , J Gao , Y Zou and X. Yan
 

Previous studies found that the forkhead transcription factor 2 (FOXL2) gene mutations are responsible for both types of blepharophimosis–ptosis–epicanthus inversus syndrome (BPES) but have not established any systematic statistic model for the complex and even contradictory results about genotype–phenotype correlations between them. This study is aimed to find possible mutations of FOXL2 gene in a Chinese family with type II BPES by using DNA sequencing and to further clarify genotype–phenotype correlations between FOXL2 mutations and BPES by using a systematic statistical method, namely Multifactor Dimensionality Reduction (MDR). A novel mutation (g.933_965dup) which could result in an expansion of the polyalanine (polyAla) tract was detected in all patients of this family. MDR analysis for intragenic mutations of FOXL2 gene reported in previous BPES studies indicated that the mutations which led to much stronger disturbance of amino acid sequence were responsible for more type I BPES, while other kinds of mutation were responsible for more type II BPES. In conclusion, the present study found a novel FOXL2 gene mutation in a Chinese BPES family and a new general genotype–phenotype correlation tendency between FOXL2 intragenic mutations and BPES, both of which expanded the knowledge about FOXL2 gene and BPES.

  G. M Polzin , W Wu , X Yan , J. M McCraw , S Abdul Salaam , A. D Tavakoli , L Zhang , D. L Ashley and C. H. Watson
  Introduction:

Standardized machine smoking measurements are poor predictors of exposure. We have refined a method using the solanesol deposited in discarded cigarette butts as a marker for estimating deliveries of mainstream smoke constituents. Developing a fast and accurate method for measuring solanesol in cigarette filters to assess tobacco smoke intake could provide a way to assess how people smoke under natural conditions. We have developed and validated a new, lower-cost, high-throughput method to measure the solanesol content in discarded cigarette filter butts and correlated these measurements with mainstream smoke deliveries of nicotine and tobacco-specific nitrosamines (TSNAs).

Methods:

Cigarettes were machine smoked under a variety of conditions to cover a wide range of nicotine deliveries and solanesol levels in the spent cigarette filter. Following machine smoking, a 1-cm portion of filter material, measured from the mouth end, was removed from the cigarette butts for analysis. Although an isotopically labeled solanesol analog is currently not commercially available, we achieved excellent quantitative results using a structurally similar compound, geranylgeraniol, as an internal standard (IS). After spiking with IS and solvent extracted, solanesol extracts were then analyzed using liquid chromatography coupled with a single-quadrupole mass analyzer. Analysis was carried out using manual preparation as well as a high-throughput 48-well format using automated liquid handlers.

Results:

Recoveries of solanesol from cigarette butts exceeded 95% with excellent precision and exhibited excellent linearity for both preparation methods. In addition, we show that the mouth-level exposure for both nicotine and TSNAs may be estimated by their relation to the solanesol retained in the cigarette filter.

Discussion:

We believe that this method provides excellent versatility and throughput for the estimation of mouth-level exposure to a wide range of toxins in cigarette smoke under naturalistic conditions. In addition, this method allows a far more accurate measure of exposure both from a single cigarette as well as from daily smoking.

  L Zhang , Q Guo and W. Zhuo
 

A new device has been developed for the measurement of the 212Pb particle size distribution indoors. This device consists of two wire screens and a back-up filter with a diameter of 2.0 cm. The sampling flow rate is typically 3.0 l min–1. After 3-h sampling time and 6-h waiting time, a CR-39 detector is used for the registration of the alpha particles from the 212Pb, deposited on the wire screens and the filter, respectively. It appears clear from field measurements that there are no appreciable differences among the particle size distributions from different dwellings within the same location and under the same climate conditions. However, the 212Pb particle size distributions from the countryside dwellings have different results from those of the city dwellings.

  L Zhang and P. J. Gallagher
 

Mind bomb 1 (Mib1) is a multidomain E3 ligase that directs ubiquitination of the Notch ligands Delta and Jagged to promote their endocytosis. Here we examine Notch-independent functions of Mib1 and find that its activities are linked to the initiation of the extrinsic cell death pathway. Expression of Mib1 induces a spontaneous, caspase-dependent cell death. Consistent with this, depletion of endogenous Mib1 decreases tumor-necrosis factor (TNF)-induced cell death. Mib1 was found to bind to cellular Fas-associated death domain (FADD)-like IL-1b converting enzyme (FLICE)-like inhibitory proteins (cFLIP-L and cFLIP-S), whereas only cFLIP-s can inhibit Mib1-induced cell death. The interaction between Mib1 and cFLIP decreases the association of caspase-8 with cFLIP, which activates caspase-8 and induces cell death. Collectively, these results suggest that in addition to a central role in Notch signaling, Mib1 has an important role in regulating the extrinsic cell death pathway.

  L Zhang , H Tu and Y. L. Li
 

As an endogenous physiologically active peptide, angiotensin (ANG) II plays an important role in the maintenance of blood pressure. In the arterial baroreceptor reflex (a pivotal regulator of blood pressure), aortic baroreceptor (AB) neurons located in the nodose ganglia (NG) are a primary afferent limb of the baroreflex. Hyperpolarization-activated currents (Ih) in the AB neurons contribute to the excitability of the AB neurons. Therefore, the present study was to measure the modulating effect of ANG II on the Ih in the primary AB neurons isolated from rats. Data from immunofluorescent and Western blot analyses showed that protein of AT1 and AT2 receptors was expressed in the nodose neurons. In the whole cell patch-clamp recording, ANG II concentration dependently enhanced the Ih density in the AB neurons (100 nM ANG II-induced 53.8 ± 3.8% increase for A-type AB neurons and 30.4 ± 7.7% increase for C-type AB neurons at test pulse –140 mV, P < 0.05). ANG II also decreased membrane excitability in the AB neurons. AT1 receptor antagonist (1 µM losartan) but not AT2 receptor antagonist (1 µM PD-123,319) totally abolished the effect of ANG II on the Ih and neuronal excitability. In addition, NADPH oxidase inhibitor (100 µM apocynin) and superoxide scavenger (1 mM tempol) also significantly blunted the ANG II-induced increase of the Ih and decrease of the membrane excitability in the AB neurons. Furthermore, losartan, apocynin, or tempol significantly attenuated the superoxide overproduction in the NG tissues induced by ANG II. These results suggest that ANG II-NADPH oxidase-superoxide signaling can activate the Ih and subsequently decrease the membrane excitability of rat AB neurons.

  J Das , G Ren , L Zhang , A. I Roberts , X Zhao , A. L.M Bothwell , L Van Kaer , Y Shi and G. Das
 

Interleukin (IL)-17–producing T helper (Th17) cells play a critical role in the pathophysiology of several autoimmune disorders. The differentiation of Th17 cells requires the simultaneous presence of an unusual combination of cytokines: IL-6, a proinflammatory cytokine, and transforming growth factor (TGF) β, an antiinflammatory cytokine. However, the molecular mechanisms by which TGF-β exerts its effects on Th17 cell differentiation remain elusive. We report that TGF-β does not directly promote Th17 cell differentiation but instead acts indirectly by blocking expression of the transcription factors signal transducer and activator of transcription (STAT) 4 and GATA-3, thus preventing Th1 and Th2 cell differentiation. In contrast, TGF-β had no effect on the expression of retinoic acid receptor–related orphan nuclear receptor t, a Th17-specific transcription factor. Interestingly, in Stat-6–/–T-bet–/– mice, which are unable to generate Th1 and Th2 cells, IL-6 alone was sufficient to induce robust differentiation of Th17 cells, whereas TGF-β had no effect, suggesting that TGF-β is dispensable for Th17 cell development. Consequently, BALB/c Stat-6–/–T-bet–/– mice, but not wild-type BALB/c mice, were highly susceptible to the development of experimental autoimmune encephalomyelitis, which could be blocked by anti–IL-17 antibodies but not by anti–TGF-β antibodies. Collectively, these data provide evidence that TGF-β is not directly required for the molecular orchestration of Th17 cell differentiation.

 
 
 
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