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1.  Myocardial Repolarization Dispersion and Autonomic Nerve Activity in a Canine Experimental Acute Myocardial Infarction Model 
Evidence from a canine experimental acute myocardial infarction (MI) model shows that until the seventh week after MI the relationship between stellate ganglionic nerve and vagal nerve activities (SGNA/VNA) progressively increases.
We evaluated how autonomic nervous system activity influences temporal myocardial repolarization dispersion at this period.
We analyzed autonomic nerve activity as well as QT and RR variability from recordings previously obtained in 9 dogs. From a total 48 short-term electrocardiographic segments, 24 recorded before and 24 seven weeks after experimentally-induced MI, we obtained three indices of temporal myocardial repolarization dispersion: QTe (from q wave T to wave end), QTp (from q wave to T wave peak) and Te (from T wave peak to T wave end) variability index (QTeVI, QTpVI, TeVI). We also performed a heart rate variability power spectral analysis on the same segments.
After MI, all the QT variables increased QTeVI (median [interquartile range]) (from - 1.76[0.82] to −1.32[0.68]), QTeVI (from −1.90[1.01] to −1.45[0.78]) and TeVI (from −0.72[0.67] to −0.22[1.00]), whereas all RR spectral indexes decreased (p<0.001 for all). Distinct circadian rhythms in QTeVI (p<0.05,) QTpVI (p<0.001) and TeVI (p<0.05) appeared after MI with circadian variations resembling that of SGNA/VNA. The morning QTpVI and TeVI acrophases approached the SGNA/VNA acrophase. Conversely, the evening QTeVI acrophase coincided with another SGNA/VNA peak. After MI, regression analysis detected a positive relationship between SGNA/VNA and TeVI (R2: 0.077; β: 0.278; p< 0.001).
Temporal myocardial repolarization dispersion shows a circadian variation after MI reaching its peak at a time when sympathetic is highest and vagal activity lowest.
PMCID: PMC4078249  PMID: 24120873
2.  Role of TRPV1 Channels in Ischemia/Reperfusion-Induced Acute Kidney Injury 
PLoS ONE  2014;9(10):e109842.
Transient receptor potential vanilloid 1 (TRPV1) -positive sensory nerves are widely distributed in the kidney, suggesting that TRPV1-mediated action may participate in the regulation of renal function under pathophysiological conditions. Stimulation of TRPV1 channels protects against ischemia/reperfusion (I/R)-induced acute kidney injury (AKI). However, it is unknown whether inhibition of these channels is detrimental in AKI or not. We tested the role of TRPV1 channels in I/R-induced AKI by modulating these channels with capsaicin (TRPV1 agonist), capsazepine (TRPV1 antagonist) and using Trpv1−/− mice.
Methods and Results
Anesthetized C57BL/6 mice were subjected to 25 min of renal ischemia and 24 hrs of reperfusion. Mice were pretreated with capsaicin (0.3 mg/kg body weight) or capsazepine (50 mg/kg body weight). Capsaicin ameliorated the outcome of AKI, as measured by serum creatinine levels, tubular damage,neutrophil gelatinase-associated lipocalin (NGAL) abundance and Ly-6B.2 positive polymorphonuclear inflammatory cells in injured kidneys. Neither capsazepine nor deficiency of TRPV1 did deteriorate renal function or histology after AKI. Measurements of endovanilloids in kidney tissue indicate that 20-hydroxyeicosatetraeonic acid (20-HETE) or epoxyeicosatrienoic acids (EETs) are unlikely involved in the beneficial effects of capsaicin on I/R-induced AKI.
Activation of TRPV1 channels ameliorates I/R-induced AKI, but inhibition of these channels does not affect the outcome of AKI. Our results may have clinical implications for long-term safety of renal denervation to treat resistant hypertension in man, with respect to the function of primary sensory nerves in the response of the kidney to ischemic stimuli.
PMCID: PMC4201466  PMID: 25330307
3.  Lymphocyte Oxidative Stress/Genotoxic Effects Are Related to Serum IgG and IgA Levels in Coke Oven Workers 
The Scientific World Journal  2014;2014:801346.
We investigated oxidative stress/genotoxic effects levels, immunoglobulin levels, polycyclic aromatic hydrocarbons (PAHs) levels exposed in 126 coke oven workers and in 78 control subjects, and evaluated the association between oxidative stress/genotoxic effects levels and immunoglobulin levels. Significant differences were observed in biomarkers, including 1-hydroxypyrene levels, employment time, percentages of alcohol drinkers, MDA, 8-OHdG levels, CTL levels and CTM, MN, CA frequency, and IgG, IgA levels between the control and exposed groups. Slightly higher 1-OHP levels in smoking users were observed. For the dose-response relationship of IgG, IgA, IgM, and IgE by 1-OHP, each one percentage increase in urinary 1-OHP generates a 0.109%, 0.472%, 0.051%, and 0.067% decrease in control group and generates a 0.312%, 0.538%, 0.062%, and 0.071% decrease in exposed group, respectively. Except for age, alcohol and smoking status, IgM, and IgE, a significant correlation in urinary 1-OHP and other biomarkers in the total population was observed. Additionally, a significant negative correlation in genotoxic/oxidative damage biomarkers of MDA, 8-OH-dG, CTL levels, and immunoglobins of IgG and IgA levels, especially in coke oven workers, was found. These data suggest that oxidative stress/DNA damage induced by PAHs may play a role in toxic responses for PAHs in immunological functions.
PMCID: PMC4129152  PMID: 25136686
4.  Low-Level Vagus Nerve Stimulation Upregulates Small Conductance Calcium Activated Potassium Channels in the Stellate Ganglion 
Small conductance calcium activated potassium (SK) channels are responsible for afterhyperpolarization that suppresses nerve discharges.
To test the hypotheses that low-level vagus nerve stimulation (LL-VNS) leads to the upregulation of SK2 proteins in the LSG.
Six dogs (Group 1) underwent 1-wk LL-VNS of the left cervical vagus nerve. Five normal dogs (Group 2) were used as control. SK2 protein levels were examined by western blotting. The ratio between SK2 and glyceraldehydes-3-phosphate-dehydrogenase (GAPDH) levels was used as an arbitrary unit (AU).
We found higher SK2 expression in Group 1 (0.124 ± 0.049 AU) than Group 2 (0.085 ± 0.031 AU, P < 0.05). Immunostaining showed that the density of nerve structures stained with SK2 antibody was also higher in Group 1 (11,546 ± 7,271 μm2/mm2) than in Group 2 (5,321 ± 3,164 μm2/mm2, P < 0.05). There were significantly more ganglion cells without immunoreactivity to TH in Group 1 (11.4 ± 2.3%) than Group 2 (4.9 ± 0.7%; P < 0.05). The TH-negative ganglion cells mostly stained positive for choline acetyltransferase (ChAT) (95.9 ± 2.8% in Group 1 and 86.1 ± 4.4% in Group 2, P = 0.10). Immunofluorescence confocal microscopy revealed a significant decrease in the SK2 staining in the cytosol but an increase in the SK2 staining on the membrane of the ganglion cells in Group 1 compared to Group 2.
Left LL-VNS results in the upregulation of SK2 proteins, increased SK2 protein expression in the cell membrane and the increased TH-negative (mostly ChAT-positive) ganglion cells in the LSG. These changes may underlie the antiarrhythmic efficacy of LL-VNS in ambulatory dogs.
PMCID: PMC3671581  PMID: 23357541
Autonomic nervous system; Vagus nerve stimulation; Stellate ganglion; Small conductance calcium activated potassium channel; Western blot
5.  Association between NOD2/CARD15 gene polymorphisms and Crohn's disease in Chinese Zhuang patients 
AIM: To assess the relationship between the P268S, JW1 and N852S polymorphisms and Crohn’s disease (CD) susceptibility in Zhuang patients in Guangxi, China.
METHODS: Intestinal tissues from 102 Zhuang [48 CD and 54 ulcerative colitis (UC)] and 100 Han (50 CD and 50 UC) unrelated patients with inflammatory bowel disease and 72 Zhuang and 78 Han unrelated healthy individuals were collected in the Guangxi Zhuang Autonomous Region from January 2009 to March 2013. Genomic DNA was extracted using the phenol chloroform method. The P268S, JW1 and N852S polymorphisms were amplified using polymerase chain reaction (PCR), detected by restriction fragment length polymorphism (RFLP), and verified by gene sequencing.
RESULTS: Heterozygous mutation of P268S in the NOD2/CARD15 gene was detected in 10 CD cases (six Zhuang and four Han), two Han UC cases, and one Zhuang healthy control, and P268S was strongly associated with the Chinese Zhuang and Han CD populations (P = 0.016 and 0.022, respectively). No homozygous mutant P268S was detected in any of the groups. No significant difference was found in P268S genotype and allele frequencies between UC and control groups (P > 0.05). Patients with CD who carried P268S were likely to be ≤ 40 years of age (P = 0.040), but were not significantly different with regard to race, lesion site, complications, and other clinical features (P > 0.05). Neither JW1 nor N852S polymorphisms of the NOD2/CARD15 gene were found in any of the subjects (P > 0.05).
CONCLUSION: P268S polymorphism may be associated with CD susceptibility in the Zhuang population in the Guangxi Zhuang Autonomous Region, China. In contrast, JW1 and N852S polymorphisms may not be related to CD susceptibility in these patients.
PMCID: PMC4000511  PMID: 24782627
Crohn’s disease; NOD2/CARD15; Single nucleotide polymorphisms
6.  Electrosprayed core–shell solid dispersions of acyclovir fabricated using an epoxy-coated concentric spray head 
A novel structural solid dispersion (SD) taking the form of core–shell microparticles for poorly water-soluble drugs is reported for the first time. Using polyvinylpyrrolidone (PVP) as a hydrophilic polymer matrix, the SDs were fabricated using coaxial electrospraying (characterized by an epoxy-coated concentric spray head), although the core fluids were unprocessable using one-fluid electrospraying. Through manipulating the flow rates of the core drug-loaded solutions, two types of core–shell microparticles with tunable drug contents were prepared. They had average diameters of 1.36±0.67 and 1.74±0.58 μm, and were essentially a combination of nanocomposites with the active ingredient acyclovir (ACY) distributed in the inner core, and the sweeter sucralose and transmembrane enhancer sodium dodecyl sulfate localized in the outer shell. Differential scanning calorimetry and X-ray diffraction results demonstrated that ACY, sodium dodecyl sulfate, and sucralose were well distributed in the PVP matrix in an amorphous state because of favorable second-order interactions. In vitro dissolution and permeation studies showed that the core–shell microparticle SDs rapidly freed ACY within 1 minute and promoted nearly eightfold increases in permeation rate across the sublingual mucosa compared with raw ACY powders.
PMCID: PMC3998863  PMID: 24790437
core–shell microparticle; solid dispersion; coaxial electrospraying; poorly water-soluble drug; epoxy-coated spray head
7.  Electroanatomic Remodeling of the Left Stellate Ganglion After Myocardial Infarction 
The purpose of this study was to evaluate the changes of left stellate ganglionic nerve activity (SGNA) and left thoracic vagal nerve activity (VNA) after acute myocardial infarction (MI).
Whether MI results in remodeling of extracardiac nerve activity remains unclear.
We implanted radiotransmitters to record the SGNA, VNA, and electrocardiogram in 9 ambulatory dogs. After baseline monitoring, MI was created by 1-h balloon occlusion of the coronary arteries. The dogs were then continuously monitored for 2 months. Both stellate ganglia were stained for growth-associated protein 43 and synaptophysin. The stellate ganglia from 5 normal dogs were used as control.
MI increased 24-h integrated SGNA from 7.44 ± 7.19 Ln(Vs)/day at baseline to 8.09 ± 7.75 Ln(Vs)/day after the MI (p < 0.05). The 24-h integrated VNA before and after the MI was 5.29 ± 5.04 Ln(Vs)/day and 5.58 ± 5.15 Ln(Vs)/day, respectively (p < 0.05). A significant 24-h circadian variation was noted for the SGNA (p < 0.05) but not the VNA. The SGNA/VNA ratio also showed significant circadian variation. The nerve densities from the left SG were 63,218 ± 34,719 μm2/mm2 and 20,623 ± 4,926 μm2/mm2 for growth-associated protein 43 (p < 0.05) and were 32,116 ± 8,190 μm2/mm2 and 16,326 ± 4,679 μm2/mm2 for synaptophysin (p < 0.05) in MI and control groups, respectively. The right SG also showed increased nerve density after MI (p < 0.05).
MI results in persistent increase in the synaptic density of bilateral stellate ganglia and is associated with increased SGNA and VNA. There is a circadian variation of the SGNA/VNA ratio. These data indicate significant remodeling of the extracardiac autonomic nerve activity and structures after MI.
PMCID: PMC3975658  PMID: 22381432
acute myocardial infarction; autonomic nervous system; nerve recordings
8.  Sympathetic nerve fibers and ganglia in canine cervical vagus nerves: Localization and quantitation 
Cervical vagal nerve (CVN) stimulation may improve left ventricular ejection fraction in patients with heart failure.
To test the hypothesis that sympathetic structures are present in the CVN and to describe the location and quantitate these sympathetic components of the CVN.
We performed immunohistochemical studies of the CVN from 11 normal dogs and simultaneously recorded stellate ganglion nerve activity, left thoracic vagal nerve activity, and subcutaneous electrocardiogram in 2 additional dogs.
A total of 28 individual nerve bundles were present in the CVNs of the first 11 dogs, with an average of 1.87 ± 1.06 per dog. All CVNs contain tyrosine hydroxylase-positive (sympathetic) nerves, with a total cross-sectional area of 0.97 ± 0.38 mm2. The sympathetic nerves were nonmyelinated, typically located at the periphery of the nerve bundles and occupied 0.03%–2.80% of the CVN cross-sectional area. Cholineacetyltransferase-positive nerve fibers occupied 12.90%–42.86% of the CVN cross-sectional areas. Ten of 11 CVNs showed tyrosine hydroxylase and cholineacetyltransferase colocalization. In 2 dogs with nerve recordings, we documented heart rate acceleration during spontaneous vagal nerve activity in the absence of stellate ganglion nerve activity.
Sympathetic nerve fibers are invariably present in the CVNs of normal dogs and occupy in average up to 2.8% of the cross-sectional area. Because sympathetic nerve fibers are present in the periphery of the CVNs, they may be susceptible to activation by electrical stimulation. Spontaneous activation of the sympathetic component of the vagal nerve may accelerate the heart rate.
PMCID: PMC3758134  PMID: 23246597
Cervical vagus nerves; Sympathetic nerves; Ganglion cells; Heart failure; Vagal nerve stimulation
9.  Small Molecule Inhibitors Target the Tissue Transglutaminase and Fibronectin Interaction 
PLoS ONE  2014;9(2):e89285.
Tissue transglutaminase (TG2) mediates protein crosslinking through generation of ε−(γ-glutamyl) lysine isopeptide bonds and promotes cell adhesion through interaction with fibronectin (FN) and integrins. Cell adhesion to the peritoneal matrix regulated by TG2 facilitates ovarian cancer dissemination. Therefore, disruption of the TG2-FN complex by small molecules may inhibit cell adhesion and metastasis. A novel high throughput screening (HTS) assay based on AlphaLISA™ technology was developed to measure the formation of a complex between His-TG2 and the biotinylated FN fragment that binds TG2 and to discover small molecules that inhibit this protein-protein interaction. Several hits were identified from 10,000 compounds screened. The top candidates selected based on >70% inhibition of the TG2/FN complex formation were confirmed by using ELISA and bioassays measuring cell adhesion, migration, invasion, and proliferation. In conclusion, the AlphaLISA bead format assay measuring the TG2-FN interaction is robust and suitable for HTS of small molecules. One compound identified from the screen (TG53) potently inhibited ovarian cancer cell adhesion to FN, cell migration, and invasion and could be further developed as a potential inhibitor for ovarian cancer dissemination.
PMCID: PMC3930694  PMID: 24586660
10.  Genome-wide association study in Han Chinese identifies four new susceptibility loci for coronary artery disease 
Nature genetics  2012;44(8):890-894.
We performed a meta-analysis of 2 genome-wide association studies of coronary artery disease comprising 1,515 cases with coronary artery disease and 5,019 controls, followed by de novo replication studies in 15,460 cases and 11,472 controls, all of Chinese Han descent. We successfully identified four new loci for coronary artery disease reaching genome-wide significance (P < 5 × 10−8), which mapped in or near TTC32-WDR35, GUCY1A3, C6orf10-BTNL2 and ATP2B1. We also replicated four loci previously identified in European populations (PHACTR1, TCF21, CDKN2A/B and C12orf51). These findings provide new insights into biological pathways for the susceptibility of coronary artery disease in Chinese Han population.
PMCID: PMC3927410  PMID: 22751097
11.  Differential expression and regulation of prohibitin during curcumin-induced apoptosis of immortalized human epidermal HaCaT cells 
Prohibitin (PHB), also known as inhibin, is important in cell proliferation, differentiation and apoptosis. This protein localizes to the inner membrane of mitochondria, where it acts as a chaperone protein, and is also found in the nucleus, where it negatively regulates transcription. The tumor-suppressive role of PHB in cell proliferation appears to be contradictory. In this study, we investigated the existence, localization and alterations in the expression of PHB in the whole cell and nuclear matrix and analyzed its co-localization with the expression products of related genes. The western blot analysis results revealed that PHB exists in the composition of nuclear matrix proteins and that the expression level of PHB is significantly increased in the whole cell and markedly decreased in the nuclear matrix after curcumin (1,7-bis(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione) treatment. The laser confocal scanning microscope results demonstrated the co-localization of PHB with p53, c-Myc, Bax, and Fas in HaCaT cells, and this co-localization region was transferred as a result of curcumin treatment. In addition, the results of the GST pull-down assay demonstrated the direct interaction of PHB with p53, c-Myc and Bax but not Fas in vitro. Results of the present study confirmed that the expression and distribution of PHB, which is a nuclear matrix protein, affect the apoptosis of HaCaT cells and its co-localization with specific gene products connected with cell apoptosis.
PMCID: PMC3926502  PMID: 24402549
prohibitin; nuclear matrix; curcumin; cell apoptosis; differential expression
12.  Prolonged Combination Therapy is More Effective than Monotherapy in Management of Chronic Hepatitis B Patients With Sustained Virological Response: An Experience From a ‘Real-World’ Clinical Setting 
Little is known about the duration of combination therapy for patients with chronic hepatitis B (CHB) and suboptimal response to nucleos(t)ide analogues(NAs) monotherapy.
This study aimed to assess whether monotherapy could be used for treatment of CHB patients, who poorly responded to Adefovir Dipivoxil (ADV) but obtained good responses after at least 12-month lamivudine (LAM) or telbivudine (LdT) add-on therapy.
Patients and Methods:
Forty-five patients were enrolled, and the baseline time-point was determined according to enrollment data. Twenty-six patients chose to continue combination therapy (LAM+ADV or LdT+ADV, Group A) and 19 patients switched to single-drug maintenance therapy (LAM or LdT or ADV, Group B).
There were no significant differences between two groups in baseline characteristics (P > 0.05). At 12th month, sustained virological response rate was greater in group A compared to group B (96.2% vs. 47.4%, P < 0.001), and the rates of NAs-associated resistance were 0% in group A and 15.8% in group B. Alanine aminotransferase normalization rate was also significantly higher in group A compared with group B (92.3% vs. 36.8%, P < 0.001). Among hepatitis positive patients with Be antigen (HBeAg)-, 40% (4/10) in group A and 9.1% (1/11) in group B achieved HBeAg seroconversion at the 12th month. Of patients in group B with positive-HBeAg before the previous combination therapy and detectable HBV DNA at 6 months of previous combination therapy were associated with high risks of viral relapse after switching to single-drug maintenance therapy.
Prematurely switching to single-drug maintenance therapy would be resulted in viral relapse, and prolonged combination therapy was effective to maintain sustained responses for patients with initial suboptimal response to ADV.
PMCID: PMC3955527  PMID: 24693412
Hepatitis B, Chronic; Combined Modality Therapy; Hepatitis Be Antigens
13.  A Highly Selective and Potent PTP-MEG2 Inhibitor with Therapeutic Potential for Type 2 Diabetes 
Journal of the American Chemical Society  2012;134(43):18116-18124.
Protein tyrosine phosphatases (PTPs) constitute a large family of signaling enzymes that control the cellular levels of protein tyrosine phosphorylation. A detailed understanding of PTP functions in normal physiology and in pathogenic conditions has been hampered by the absence of PTP-specific, cell-permeable small molecule agents. We present a stepwise focused library approach that transforms a weak and general nonhydrolyzable pTyr mimetic (F2Pmp, phosphonodifluoromethyl phenylalanine) into a highly potent and selective inhibitor of PTP-MEG2, an antagonist of hepatic insulin signaling. The crystal structures of the PTP-MEG2-inhibitor complexes provide direct evidence that potent and selective PTP inhibitors can be obtained by introducing molecular diversity into the F2Pmp scaffold to engage both the active site and unique nearby peripheral binding pockets. Importantly, the PTP-MEG2 inhibitor possesses highly efficacious cellular activity and is capable of augmenting insulin signaling and improving insulin sensitivity and glucose homeostasis in diet-induced obese mice. The results indicate that F2Pmp can be converted into highly potent and selective PTP inhibitory agents with excellent in vivo efficacy. Given the general nature of the approach, this strategy should be applicable to other members of the PTP superfamily.
PMCID: PMC3505080  PMID: 23075115
14.  Multiple Genomic Recombination Events in the Evolution of Saffold Cardiovirus 
PLoS ONE  2013;8(9):e74947.
Saffold cardiovirus (SAFV) is a new human cardiovirus with 11 identified genotypes. Little is known about the natural history and pathogenicity of SAFVs.
Methodology/Principal Findings
We sequenced the genome of five SAFV-1 strains which were identified from fecal samples taken from children with viral diarrhea in Beijing, China between March 2006 and November 2007, and analyzed the phylogenetic and phylodynamic properties of SAFVs using the genome sequences of every known SAFV genotypes. We identified multiple recombination events in our SAFV-1 strains, specifically recombination between SAFV-2, -3, -4, -9, -10 and the prototype SAFV-1 strain in the VP4 region and recombination between SAFV-4, -6, -8, -10, -11 and prototype SAFV-1 in the VP1/2A region. Notably, recombination in the structural gene VP4 is a rare event in Cardiovirus. The ratio of nonsynonymous substitutions to synonymous substitutions indicates a purifying selection of the SAFV genome. Phylogenetic and molecular clock analysis indicates the existence of at least two subclades of SAFV-1 with different origins. Subclade 1 includes two strains isolated from Pakistan, whereas subclade 2 includes the prototype strain and strains isolated in China, Pakistan, and Afghanistan. The most recent common ancestor of all SAFV genotypes dates to the 1710s, and SAFV-1, -2, and -3 to the 1940s, 1950s, and 1960s, respectively. No obvious relationship between variation and pathogenicity exists in the critical domains of the CD and EF loops of viral capsid proteins or the multi-functional proteins L based on animo acid sequence identity comparison between SAFV genotypes.
Our findings suggest that intertypic recombination plays an important role in the diversity of SAFVs, highlighting the diversity of the five strains with the previously described SAFV-1 strains.
PMCID: PMC3781130  PMID: 24086404
15.  Sang-qi Granula Reduces Blood Pressure and Myocardial Fibrosis by Suppressing Inflammatory Responses Associated with the Peroxisome Proliferator-Activated Receptors and Nuclear Factor κB Protein in Spontaneously Hypertensive Rats 
Aim. Sang-qi Granula (SQ) is a compound prepared from Chinese herbs and is currently used for treatment of hypertension in China. Given its protective effects on cardial function in decreasing blood pressure, we investigated the mechanism of protective effects of SQ on myocardium. Methods. 16 male normal Wistar-Kyoto rats and 16 spontaneous hypertension rats (SHR) were employed without medical treatment. 16 SHR were employed with SQ treatment. Rats in each group were sacrificed at two time points (8-week treatment and 16-week treatment). Blood pressure (BP), and heart weight/body weight (HW/BW) were measured. The expression of myeloperoxidase (MCP-1), ICAM-1, TNF-α, and CD68-positive cells was assessed. The interstitial collagen volume fraction (CVF), perivascular collagen volume area (PVCA), and the expression of TGF-β, Smad-3, PPARα, γ, and NF-κB (P65 and P50) were observed. Results. SQ significantly inhibited the elevation of the blood pressure and HW/BW of SHR. Next, SQ prevented myocardial fibrosis. Finally, a proinflammatory mediator associated with NF-κB (TNF-α, ICAM-1, MCP-1, CD68), TGF-β, and Smad-3 related to collagen deposition, which is upregulated in SHR group, was significantly suppressed by SQ. Expression of NF-κB was decreased in SHQ+SQ group compared to PPARα, and γ expression was increased by SQ. Conclusion. Treatment with SQ ameliorates cardial fibrosis induced by hypertension by attenuating the upregulation of ICAM-1, TNF-α, MCP-1, TGF-β, Smad-3, P65, and P50 expression and improving PPARα and PPARγ expression level. The results suggest that SQ may be an option for preventing cardial fibrosis through PPAR signalling pathway.
PMCID: PMC3793543  PMID: 24171042
16.  Oncolytic vesicular stomatitis virus quantitatively and qualitatively improves primary CD8+ T-cell responses to anticancer vaccines 
Oncoimmunology  2013;2(8):e26013.
The ability of heterologous prime-boost vaccination to elicit robust CD8+ T cell responses has been well documented. In contrast, relatively little is known about how this immunotherapeutic strategy impacts the functional qualities of expanded T cells in the course of effector and memory responses. Using vesicular stomatitis virus (VSV) as a boosting vector in mice, we demonstrate that a massive secondary expansion of CD8+ T cells can be achieved shortly after priming with recombinant adenoviral vectors. Importantly, VSV-boosted CD8+ T cells were more potent than those primed by adenoviruses only, as measured by cytokine production, granzyme B expression, and functional avidity. Upon adoptive transfer, equivalent numbers of VSV-expanded CD8+ T cells were more effective (on a per-cell basis) in mediating antitumor and antiviral immunity than T cells only primed with adenoviruses. Furthermore, VSV boosting accelerated the progression of expanded CD8+ T lymphocytes to a central memory phenotype, thereby altering the effector memory profile typically associated with adenoviral vaccination. Finally, the functional superiority of VSV-expanded T cells remained evident 100 d after boosting, suggesting that VSV-driven immunological responses are of sufficient duration for therapeutic applications. Our data strongly support the choice of VSV as a boosting vector in prime-boost vaccination strategies, enabling a rapid amplification of CD8+ T cells and improving the quality of expanded T cells during both early and late immunological responses.
PMCID: PMC3782525  PMID: 24083086
adenovirus; CD8+ T cells; prime-boost; vaccination; vesicular stomatitis virus
17.  Genetic analysis of microsomal epoxide hydrolase gene and its association with lung cancer risk 
The human microsomal epoxide hydrolase (EH) gene contains polymorphic alleles, which may be linked to increased risk for tobacco-related lung cancer. The purpose of this study is to screen new polymorphisms and determine whether these polymorphisms can be used to predict individual susceptibility to lung cancer. The PCR-single strand conformation polymorphism (SSCP) analysis was used to screen for polymorphisms in the coding region of the EH gene. Eleven polymorphisms, including previously reported polymorphisms, were identified and the prevalence of these variants was assessed in at least 50 healthy Caucasians and African Americans. Among the eleven polymorphisms, the prevalence of the amino acid-changing EH polymorphisms in codons 43, 113, and 139 was examined in 182 Caucasian incident cases with primary lung cancer, as well as in 365 frequency-matched controls to examine the role of EH polymorphisms in lung cancer risk. A significant increase in lung cancer risk was observed for predicted high EH activity genotypes (OR = 2.3, 95% CI = 1.2–4.3) as compared to low EH activity genotypes. This association was more pronounced among patients with lung adenocarcinoma (OR = 4.7, 95% CI = 1.7–13.1). These results suggest that the EH polymorphism plays an important role in lung cancer risk and is linked to tobacco smoke exposure.
PMCID: PMC3715303  PMID: 15901990
Epoxide hydrolase; lung cancer; genetic polymorphism; metabolism
18.  Polymorphisms for microsomal epoxide hydrolase and genetic susceptibility to COPD 
Although smoking is the major causal factor in the development of chronic obstructive pulmonary disease (COPD), only 10–20% of chronic heavy cigarette smokers develop symptomatic COPD, which suggests the presence of genetic susceptibility. The human microsomal epoxide hydrolase (EH) is a metabolizing enzyme which involves the process of numerous reactive epoxide intermediates and contains polymorphic alleles which are associated with altered EH activity and may be linked to increased risk for COPD. To determine whether the EH polymorphisms contributed to increased risk for COPD, prevalence of the EH codons 113 and 139 polymorphisms were compared between COPD patients and controls using a PCR-RFLP analysis using genomic DNA isolated from 131 COPD patients and 262 individually matched controls by age (± 5 years) among Caucasians with 1:2 ratio. Significantly increased risk for COPD was observed for subjects with the EH113His/His genotypes (OR =2.4, 95% CI=1.1–5.1). These results were consistent with the fact that a significant trend towards increased risk was observed with predicted less protective EH codon 113 genotypes (p = 0.03, trend test). A similar association was not observed for EH codon139 polymorphism. As expected, a significant correlation between smoking dose and severity of COPD was observed (p<0.001). These results suggest that EH codon 113 polymorphism may modify risk for COPD.
PMCID: PMC3705731  PMID: 15702235
chronic obstructive pulmonary disease; epoxide hydrolase; genetic polymorphism; genetic susceptibility
19.  Polymorphisms in the promoter region of neutrophil elastase gene and lung cancer risk 
The neutrophil elastase (NE) gene encodes a powerful serine protease that is involved in the process of normal tissue turnover, natural host defense or tissue damage in acute and chronic inflammatory disorders. Furthermore, NE was suggested as one of the determinant factors of individual susceptibility to lung cancer resulting from imbalance between α1-antitrypsin (AT) and NE. To determine whether NE plays a role in risk for lung cancer, we screened polymorphisms in the promoter region of the NE gene and assessed the role of the NE polymorphisms in the risk for lung cancer. We confirmed three previously identified polymorphisms which are located at −903, −741, and extra 52 bp STS relative to the transcription initiation site. In addition, two new polymorphisms at −832 (G/T) and −789 (C/T) were identified. Their rare allelic frequencies of new polymorphism are 0.02 and 0.01, respectively, among Caucasians. The prevalence of the NE −903 (T/T) and (T/G) genotypes were 0.88 and 0.12 in controls as compared to 0.96 and 0.04 in lung cancer patients using genomic DNA isolated from 113 Caucasian lung cancer cases and 131 controls. A significant increase in lung cancer risk was observed for expected high NE activity genotypes (OR = 3.2, 95% CI = 1.02–10.3) as compared to low NE activity genotypes. These results were consistent with previous in vitro functional analysis, which reported an approximately two-fold increase enzyme expression with the −903T/−741G allele as compared to the −903G/−741A variant. These results confirm that the NE promoter region polymorphisms may influence in risk for lung cancer.
PMCID: PMC3698610  PMID: 15892999
Neutrophil elastase; Lung cancer; Genetic polymorphism; Cancer susceptibility
20.  Neural Control of Ventricular Rate in Ambulatory Dogs with Pacing Induced Sustained Atrial Fibrillation 
We hypothesize that inferior vena cava-inferior atrial ganglionated plexus nerve activity (IVC-IAGPNA) is responsible for the ventricular rate (VR) control during atrial fibrillation (AF) in ambulatory dogs.
Methods and Results
We recorded bilateral cervical vagal nerve activity (VNA) and IVC-IAGPNA during baseline sinus rhythm and during pacing-induced sustained AF in 6 ambulatory dogs. Integrated nerve activities and average VR were measured every 10-s over 24-hour periods. LVNA was associated with VR reduction during AF in 5 dogs (from 211 bpm, 95% confidence interval [CI], 186 to 233 to 178 bpm [95% CI, 145 to 210], p<0.001) and RVNA in 1 dog (208 bpm [95% CI, 197 to 223] to 181 bpm [95% CI, 163 to 200], p<0.01). There were good correlations between IVC-IAGPNA and LVNA in the former 5 dogs, and between IVC-IAGPNA and RVNA in the latter dog. IVC-IAGPNA was associated with VR reduction in all dogs studied. RVNA was associated with baseline sinus rate reduction from 105 bpm (95% CI, 95 to 116) to 77 bpm (95% CI, 64 to 91, p<0.01) in 4 dogs while LVNA was associated with sinus rate reduction from 111 bpm (95% CI, 90 to 1250) to 81 bpm (95% CI, 67 to 103, p<0.01) in 2 dogs.
IVC-IAGPNA is invariably associated with VR reduction during AF. In comparison, right or left VNA was associated with VR reduction only when it co-activates with the IVC-IAGPNA. The vagus nerve that controls VR during AF may be different than that controls sinus rhythm.
PMCID: PMC3405983  PMID: 22586260
atrial fibrillation; atrioventricular node; ECG; nervous system, autonomic; ventricular rate
21.  Acute Myocardial Infarction Induces Bilateral Stellate Ganglia Neural Remodeling in Rabbits 
Cardiovascular Pathology  2011;21(3):143-148.
Myocardial infarction (MI) results in cardiac nerve sprouting in the myocardium. Whether or not similar neural remodeling occurs in the stellate ganglia (SG) is unknown. We aimed to test the hypothesis that MI induces bilateral SG nerve sprouting.
Acute MI was created by coronary artery ligation in rabbits (n=12). Serum nerve growth factor (NGF) level was measured by enzyme-linked immunosorbent assay (ELISA). The hearts and bilateral SGs were harvested for immunohistochemistry after 1 week in 6 rabbits, and after 1 month in 6 rabbits. Immunostaining for tyrosine hydroxylase (TH), growth-associated protein 43 (GAP43), cholineacetyltransferase (ChAT) and synaptophysin (SYN) was performed to determine the magnitude of nerve sprouting. Tissues from 6 normal rabbits were used as controls. Nerve density was determined by computerized morphometry.
MI results in increased serum NGF levels at 1 week (1519.8±632.2 ng/ml) that persists to 1 month (1361.2±176.3 ng/ml) as compared to controls (89.6±34.9 ng/ml), (p=.0002, and , p=.0001, respectively). Immunostaining demonstrated nerve sprouting and hyperinnervation in both SGs after MI. The nerve densities (µm2/ganglion cell) in SG 1 week after MI, 1 month after MI and in control groups, respectively, were: GAP43, 278±96, 225±39 and 149±57 (p=.01); SYN, 244±152, 268±115 and 102±60 (p=.02); TH, 233±71, 180±50 and 135±68 (p=.047); ChAT, 244±100, 208±46 and 130±41 µm2/cell (p=.01).
MI increases serum NGF levels and induces nerve sprouting and hyperinnervation in bilateral SGs for at least 1 month after MI. The hyperinnervation includes both postganglionic adrenergic axons and preganglionic cholinergic axons in the SG.
PMCID: PMC3267867  PMID: 22001051
Myocardial Infarction; Ventricular Arrhythmia; Autonomic Nervous System; Stellate Ganglion; Nerve Sprouting; Sudden Cardiac Death
22.  Regulation of Alveolar Epithelial Na+ Channels by ERK1/2 in Chlorine-Breathing Mice 
The mechanisms by which the exposure of mice to Cl2 decreases vectorial Na+ transport and fluid clearance across their distal lung spaces have not been elucidated. We examined the biophysical, biochemical, and physiological changes of rodent lung epithelial Na+ channels (ENaCs) after exposure to Cl2, and identified the mechanisms involved. We measured amiloride-sensitive short-circuit currents (Iamil) across isolated alveolar Type II (ATII) cell monolayers and ENaC single-channel properties by patching ATII and ATI cells in situ. α-ENaC, γ-ENaC, total and phosphorylated extracellular signal-related kinase (ERK)1/2, and advanced products of lipid peroxidation in ATII cells were measured by Western blot analysis. Concentrations of reactive intermediates were assessed by electron spin resonance (ESR). Amiloride-sensitive Na+ channels with conductances of 4.5 and 18 pS were evident in ATI and ATII cells in situ of air-breathing mice. At 1 hour and 24 hours after exposure to Cl2, the open probabilities of these two channels decreased. This effect was prevented by incubating lung slices with inhibitors of ERK1/2 or of proteasomes and lysosomes. The exposure of ATII cell monolayers to Cl2 increased concentrations of reactive intermediates, leading to ERK1/2 phosphorylation and decreased Iamil and α-ENaC concentrations at 1 hour and 24 hours after exposure. The administration of antioxidants to ATII cells before and after exposure to Cl2 decreased concentrations of reactive intermediates and ERK1/2 activation, which mitigated the decrease in Iamil and ENaC concentrations. The reactive intermediates formed during and after exposure to Cl2 activated ERK1/2 in ATII cells in vitro and in vivo, leading to decreased ENaC concentrations and activity.
PMCID: PMC3326429  PMID: 21997487
lung slices; patch clamp; radicals
23.  A new method of kidney biopsy using low dose CT-guidance with coaxial trocar and bard biopsy gun 
To explore a new method of kidney biopsy with coaxial trocar and bard biopsy gun under low dose computed tomography (CT)-guidance and evaluate its accuracy, safety, and efficacy.
Sixty patients underwent renal biopsy under CT-guidance. They were randomly divided into two groups: group I, low dose CT-guided (120 kV and 25 or 50 mAs) and group II, standard dose CT-guided (120 kV and 250 mAs). For group I, the coaxial trocar was accurately placed adjacent to the renal capsule of the lower pole, the needle core was removed, and samples were obtained with a bard biopsy gun. For group II, the coaxial trocar was not used. Total number of passes, mean biopsy diameter, mean glomeruli per specimen, mean operation time, mean scanning time, and mean radiation dose were noted. Dose-length product (DLP) was used to calculate the radiation doses. After 24 hours of the biopsy, ultrasound was repeated to identify any subcapsular hematoma.
Success rate of biopsy in group I was 100% while using low dose CT-guidance along with coaxial trocar renal. There was no statistic differences bewteen group I and II in the total number of passes, mean biopsy diameter, mean glomeruli per specimen and mean time of operation and CT scanning. The average DLP of group I was lower as compared to the value of group II (p <0.05).
Kidney biopsy using coaxial trocar and bard biopsy gun under low dose CT was an accurate, simple and safe method for diagnosis and treatment of kidney diseases. It can be used for repeat and multiple biopsies, particularly suitable for obese and renal atrophy patients in whom the kidneys are difficult to image.
PMCID: PMC3561167  PMID: 23294600
Kidney biopsy; Low dose CT scanning; Bard biopsy gun; Coaxial trocar
24.  Neural mechanisms of atrial fibrillation 
Current Opinion in Cardiology  2012;27(1):24-28.
Purpose of review
The autonomic nerve system is a potentially potent modulator of the initiation and perpetuation of atrial fibrillation (AF). This review will briefly summarize the neural mechanisms of AF.
Recent findings
Complex interactions exist between the sympathetic and parasympathetic nervous system on the atrial electrophysiologic properties. Direct autonomic recordings in canine models demonstrated simultaneous sympathovagal discharges are the most common triggers of paroxysmal atrial tachycardia and paroxysmal AF. Also, intrinsic cardiac autonomic nerve can serve as a sole triggering factor for the initiation of AF. Modulation of autonomic nervous system (ANS) by electrical stimulation has been tried as a treatment strategy clinically and experimentally. Recent studies showed that autonomic nervous system modulation can suppress the stellate ganglion nerve activity and reduce the incidence of paroxysmal atrial tachyarrhythmias in ambulatory dogs.
The autonomic nerve system influences the initiation and perpetuation of AF. Scientific advances toward a better understanding of the complex interrelationships of the various components of the ANS will hopefully lead to improvement of treatments for this common arrhythmia.
PMCID: PMC3279730  PMID: 22139702
Autonomic nerve; atrial fibrillation; vagal nerve stimulation
25.  Analysis of TLR4 and TLR2 polymorphisms in inflammatory bowel disease in a Guangxi Zhuang population 
AIM: To study the polymorphisms of toll-like receptor 4 (TLR4) gene Asp299Gly, Thr399Ile and TLR2 gene Arg753Gln, Arg677Trp and susceptibility to inflammatory bowel disease (IBD) in the Zhuang population from Guangxi, China.
METHODS: A case-control study was performed from February 2007 to October 2011 which included 146 Zhuang patients with IBD in the experimental group and 164 healthy Zhuang subjects who acted as the control group. All patients and healthy subjects were from the Guangxi Zhuang Autonomous Region of China. Genomic DNA was extracted from intestinal tissue by the phenol chloroform method. TLR4 gene Asp299Gly, Thr399Ile and TLR2 gene Arg753Gln, Arg677Trp were amplified by polymerase chain reaction (PCR), and then detected by PCR-restriction fragment length polymorphism (RFLP).
RESULTS: The TLR4 gene Asp299Gly was digested using Nco I restriction enzyme, and a single band of 249 bp was observed which showed that it was a wild type (AA). The TLR4 gene Thr399Ile was digested using Hinf Irestriction enzyme and only the wild type (CC) was detected. In addition, the TLR2 gene Arg677Trp was digested using Aci I restriction enzyme and only the wild type (CC) was detected. The TLR2 gene Arg753Gln was digested using Pst I restriction enzyme. Only the wild type (GG) as a single band of 254 bp was observed during RFLP. Overall, no heterozygous or homozygous single nucleotide polymorphism mutations were found in patients with Crohn’s disease and ulcerative colitis both in the TLR4 gene Asp299Gly, Thr399Ile and the TLR2 gene Arg677Trp, Arg753Gln in the Zhuang population from the Guangxi Zhuang Autonomous Region of China.
CONCLUSION: The TLR4 gene Asp299Gly, Thr399Ile and TLR2 gene Arg753Gln, Arg677Trp polymorphisms may not be associated with IBD in the Zhuang population from the Guangxi Zhuang Autonomous Region of China.
PMCID: PMC3520176  PMID: 23239925
Toll-like receptor 2; Toll-like receptor 4; Inflammatory bowel disease; Gene polymorphism

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