A spectral analysis approach was developed for detailed study of time-resolved, dynamic changes in vascular smooth muscle cell (VSMC) elasticity and adhesion to identify differences in VSMC from young and aged monkeys. Atomic force microscopy (AFM) was used to measure Young's modulus of elasticity and adhesion as assessed by fibronectin (FN) or anti-beta 1 integrin interaction with the VSMC surface. Measurements demonstrated that VSMC cells from old versus young monkeys had elevated elasticity (21.6 kPa vs 3.5 kPa or a 612% elevation in elastic modulus) and adhesion (86 pN vs 43 pN or a 200% increase in unbinding force). Spectral analysis identified three major frequency components in the temporal oscillation patterns for elasticity (ranging from 1.7×10-3 to 1.9×10-2 Hz in old and 8.4×10-4 to 1.5×10-2 in young) and showed that the amplitude of oscillation was larger (p<0.05) in old than in young at all frequencies. It was also observed that patterns of oscillation in the adhesion data were similar to the elasticity waveforms. Cell stiffness was reduced and the oscillations were inhibited by treatment with cytochalasin D, ML7 or blebbistatin indicating involvement of actin-myosin driven processes. In conclusion, these data demonstrate the efficacy of time-resolved analysis of AFM cell elasticity and adhesion measurements and that it provides a uniquely sensitive method to detect real-time functional differences in biomechanical and adhesive properties of cells. The oscillatory behavior suggests mechanisms governing elasticity and adhesion are coupled and affected differentially during aging which may link these events to changes in vascular stiffness.
Fibronectin; integrins; vascular smooth muscle cell contractile function; Young's modulus; cytoskeleton; mechanotransduction; extracellular matrix adhesion; force measurement; atomic force microscopy
AIM: To confirm the hypothesis that polymorphisms of the uncoupling protein 3 (UCP3) gene are associated with the occurrence of nonalcoholic fatty liver disease (NAFLD).
METHODS: A total of 250 NAFLD patients (147 males and 103 females) and 200 healthy individuals who served as controls (control, 109 males and 91 females), aged between 6 and 16 years were enrolled in this study. The four non-synonymous single nucleotide polymorphisms (SNPs) in the UCP3 gene polymorphisms of rs1726745, rs3781907, rs11235972 and rs1800849, were genotyped using MassArray. Body mass index (BMI), waist and hip circumference, blood pressure (BP), fasting blood glucose (FBG), insulin and lipid profiles were measured and B-ultrasound examination was performed in all subjects.
RESULTS: NAFLD patients showed risk factors for metabolic syndrome: elevated BMI, waist-to-hip ratio, BP, FBG, homeostasis model assessment-estimated insulin resistance, total triglyceride, total cholesterol and low-density lipoprotein-cholesterol, while decreased high-density lipoprotein-cholesterol level compared with the control group. The GG genotype distributions of rs11235972 in the NAFLD group differed significantly from that in the control group. We found that waist circumference between CC (58.76 ± 6.45 cm) and CT+TT (57.00 ± 5.59 cm), and hip circumference between CC (71.28 ± 7.84 cm) and CT+TT genotypes (69.06 ± 7.75 cm) were significantly different with and without rs1800849 variation (P < 0.05).
CONCLUSION: A higher prevalence of rs11235972 GG genotype was observed in the NAFLD group compared with the control group. No differences were observed for the other SNPs. However, there was a significant difference in body height in addition to waist and hip circumference between the CC (mutant type group) and CT+TT group with and without rs1800849 variation.
Nonalcoholic fatty liver disease; Uncoupling protein 3; Single nucleotide polymorphisms
Persistent inflammation is associated with a shift in spinal GABAA signaling from inhibition to excitation such that GABAA-receptor activation contributes to inflammatory hyperalgesia. We tested the hypothesis that the primary afferent is the site of the persistent inflammation-induced shift in GABAA signaling which is due to a Na+-K+-Cl−-co-transporter (NKCC1)-dependent depolarization of the GABAA current equilibrium potential (EGABA). Acutely dissociated retrogradely labeled cutaneous dorsal root ganglion (DRG) neurons from naïve and inflamed (3 days after a subcutaneous injection of complete Freund’s adjuvant) adult male rats were studied with Ca2+ imaging, western blot and gramicidin perforated patch recording. GABA evoked a Ca2+ transient in a subpopulation of small- to medium-diameter capsaicin-sensitive cutaneous neurons. Inflammation was associated with a significant increase in the magnitude of GABA-induced depolarization as well as the percentage of neurons in which GABA evoked a Ca2+ transient. There was no detectable change in NKCC1 protein or phosphoprotein at the whole ganglia level. Furthermore, the increase in excitatory response was comparable in both HEPES- and HCO3−-buffered solutions, but was only associated with a depolarization of EGABA in HCO3−-based solution. In contrast, under both recording conditions, the excitatory response was associated with an increase in GABAA current density, a decrease in low threshold K+ current density, and resting membrane potential depolarization. Our results suggest that increasing K+ conductance in afferents innervating a site of persistent inflammation may have greater efficacy in the inhibition of inflammatory hyperalgesia than attempting to drive a hyperpolarizing shift in EGABA.
Inflammatory pain; nociceptor sensitization; neuroplasticity
PIMT (also known as PIPMT/NCOA6IP/Tgs1), first isolated as a transcription coactivator PRIP (NCOA6)-interacting 96-kDa protein with RNA-binding property, possesses RNA methyltransferase activity. As a transcription coactivator binding protein, PIMT enhances the nuclear receptor transcriptional activity and its methyltransferase property is involved in the formation of the 2,2,7-trimethylguanosine cap of non-coding small RNAs, but the in vivo functions of this gene have not been fully explored. To elucidate the biological functions, we used gene targeting to generate mice with a disrupted PIMT/Tgs1 gene. Disruption of PIMT gene results in early embryonic lethality due to impairment of development around the blastocyst and uterine implantation stages. We show that PIMT is expressed in all cells of the E3.5 day blastocyst in the mouse. PIMT null mutation abolished PIMT expression in all cells of the blastocyst and caused a reduction in the expression of Oct4 and Nanog transcription factor proteins in the E3.5 blastocyst resulting in the near failure to form inner cell mass (ICM). With conditional deletion of PIMT gene, mouse embryonic fibroblasts (MEFs) exhibit defective wound healing in the scratch assay and a reduction in cell proliferation due to decreased G0/G1 transition and G2/M phase cell cycle arrest. We conclude that PIMT/NCOA6IP, which is expressed in all cells of the 3.5 day stage blastocyst, is indispensable for early embryonic development.
PIMT/ NCOA6IP/Tgs1; Embryonic lethality; Apoptosis; Blastocyst; G2/M phase arrest; Mouse Embryonic Fibroblasts; Defective wound healing
Novel Danshensu derivatives (3–8) were designed and synthesized to improve bioactivity based on the strategy of ‘medicinal chemical hybridization’. Our previous studies indicated that these compounds exhibited noticeable cardioprotective activities. Here, we investigate whether these novel Danshensu derivatives exert neuroprotective activities. An in vitro study revealed that these compounds could increase cell viability and reduce LDH (lactate dehydrogenase) leakage. Moreover, Danshensu-cysteine derivative compounds 6 and 8 could significantly inhibit lipid peroxidation of cell membrane and regulate the expression of apoptosis-related protein (Bcl-2, Bax and caspase 3). An in vivo study demonstrated that treatment with compound 6 at 30 mg/kg markedly decreased the infarct volume of MCAO (middle cerebral artery occlusion) insulted rat brain. Furthermore, treatment with compound 6 showed the antioxidant capacity by increasing the activity of SOD (superoxide dismutase) and GPx (glutathione peroxidase) and decreasing the level of MDA (malondialdehyde) and the ROS (reactive oxygen species) production significantly. These results suggested that these novel conjugates exert significant neuroprotective effects as anti-ischaemia agents and those with high potential merit further investigation.
anti-apoptosis; anti-oxidative stress; Danshensu derivatives; ischemic stroke; middle cerebral artery occlusion (MCAO); neuroprotective effect; DHE, dihydroethidium; DIPEA, diisopropylethylamine; DMEM, Dulbecco’s modified Eagle’s medium; EDC, 1-(3-dimethylaminopropyl)-3-ehylcarbodiimide hydrochloride; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GPx, glutathione peroxidase; HOBt, 1-hydroxybenzotriazole; LDH, lactate dehydrogenase; MCAO, middle cerebral artery occlusion; MDA, malondialdehyde; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide; NAC, N-acetyl-L-cysteine; NBS, neuronal bovine serum; ROS, reactive oxygen species; SOD, superoxide dismutase; TTC, 2,3,5-triphenyltetrazolium chloride
The endoplasmic reticulum (ER) plays a critical role in ensuring proper folding of newly synthesized proteins. Aberrant ER stress is reported to play a causal role in cardiovascular diseases. However, the effects of ER stress on vascular smooth muscle contractility and blood pressure remain unknown. The aim of this study was to investigate whether aberrant ER stress causes abnormal vasoconstriction and consequent high blood pressure in mice.
Methods and Results
ER stress markers, vascular smooth muscle contractility, and blood pressure were monitored in mice. Incubation of isolated aortic rings with tunicamycin or MG132, 2 structurally unrelated ER stress inducers, significantly increased both phenylephrine-induced vasoconstriction and the phosphorylation of myosin light chain (Thr18/Ser19), both of which were abrogated by pretreatment with chemical chaperones or 5-Aminoimidazole-4-carboxamide ribonucleotide and metformin, 2 potent activators for the AMP-activated protein kinase. Consistently, administration of tauroursodeoxycholic acid or 4-phenyl butyric acid, 2 structurally unrelated chemical chaperones, in AMP-activated protein kinase-α2 knockout mice lowered blood pressure and abolished abnormal vasoconstrictor response of AMP-activated protein kinase-α2 knockout mice to phenylephrine. Consistently, tunicamycin (0.01 μ/g per day) infusion markedly increased both systolic and diastolic blood pressure, both of which were ablated by coadministration of 4-phenyl butyric acid. Furthermore, 4-phenyl butyric acid or tauroursodeoxycholic acid, which suppressed angiotensin II infusion–induced ER stress markers in vivo, markedly lowered blood pressure in angiotensin II–infused mice in vivo.
We conclude that ER stress increases vascular smooth muscle contractility resulting in high blood pressure, and AMP-activated protein kinase activation mitigates high blood pressure through the suppression of ER stress in vivo.
AMPK; ER stress; hypertension; vascular smooth muscle
Isolated metastases to the spleen from gastric carcinoma is very rare. Only a few cases have been reported in the literature. We herein present a case of isolated splenic metastases in a 62-year-old man, occurring 12 mo after total gastrectomy for gastric carcinoma. The patient underwent a laparoscopic exploration, during which two lesions were found at the upper pole of the spleen, without involvement of other organs. A laparoscopic splenectomy was performed. Histological examination confirmed that the splenic tumor was a poorly differentiated adenocarcinoma similar to the primary gastric lesion. The postoperative course was uneventful and the patient has been well for 9 mo, with no tumor recurrence. The clinical data of 18 cases of isolated splenic metastases from gastric carcinoma treated by splenectomy were summarized after a literature review. To our knowledge, this is the first reported case of isolated splenic metastases undergoing laparoscopic splenectomy.
Metastasis; Splenic neoplasms; Stomach neoplasms; Laparoscopy; Splenectomy
Piebaldism is a rare autosomal dominant disorder of melanocyte development, which is mostly caused by KIT gene. The key characteristics of piebaldism include localized poliosis, congenital leukoderma, and other variable manifestations. The previous study has illustrated that the homogeneous MC1R (a gene which is associated with the hair color) variant (p.I120T) coordinating with KIT mutation may lead to auburn hair color and piebaldism. In this study, we have investigated a Chinese family with piebaldism and auburn hair color; the mutation screening of KIT and MC1R genes identified that only a splicing mutation (c. 2484+1G>A) of KIT gene cosegregated with the auburn hair color and piebaldism. The data of this study and others suggests that the KIT mutation may causes of the auburn hair color in the piebaldism patients.
Heart failure (HF) is one of the most serious diseases worldwide. S-propargyl-cysteine (SPRC), a novel modulator of endogenous hydrogen sulfide, is proved to be able to protect against acute myocardial ischemia. In order to produce more stable and sustainable hydrogen sulfide, we used controlled release formulation of SPRC (CR-SPRC) to elucidate possible cardioprotective effects on HF rats and investigate involved mechanisms on apoptosis and oxidation.
Left coronary artery was occluded to induce HF model of rat. The survival rats were randomly divided into 7 groups after 24 hours and treated with drugs for 6 weeks. Echocardiographic indexes were recorded to determine cardiac function. TTC staining was performed to determine infarct size. Plasmatic level of hydrogen sulfide was detected by modified sulfide electrode. Activity of enzyme and expression of protein were determined by colorimetry and Western blot, respectively.
The cardioprotective effects of CR-SPRC on HF rats were confirmed by significant reduction of infarct size and improvement of cardiac function, with better effects compared to normal SPRC. CR-SPRC modulated antioxidant defenses by preserving levels of GSH, CAT and SOD and reducing CK leakage. In addition, CR-SPRC elevated ratio of Bcl-2/Bax and inhibited activity of caspases to protect against myocardial apoptosis. The cardioprotective effects of CR-SPRC were mediated by hydrogen sulfide.
All experiment data indicated cardioprotective effects of CR-SPRC on HF rats. More importantly, CR-SPRC exerted better effects than normal SPRC in all respects, providing a new perspective on hydrogen sulfide-mediated drug therapy.
Recent studies have demonstrated that microalga has been widely regarded as one of the most promising raw materials of biofuels. However, lack of an economical, efficient and convenient method to harvest microalgae is a bottleneck to boost their full-scale application. Many methods of harvesting microalgae, including mechanical, electrical, biological and chemical based, have been studied to overcome this hurdle.
A new flocculation method induced by decreasing pH value of growth medium was developed for harvesting freshwater microalgae. The flocculation efficiencies were as high as 90% for Chlorococcum nivale, Chlorococcum ellipsoideum and Scenedesmus sp. with high biomass concentrations (>1g/L). The optimum flocculation efficiency was achieved at pH 4.0. The flocculation mechanism could be that the carboxylate ions of organic matters adhering on microalgal cells accepted protons when pH decreases and the negative charges were neutralized, resulting in disruption of the dispersing stability of cells and subsequent flocculation of cells. A linear correlation between biomass concentration and acid dosage was observed. Furthermore, viability of flocculated cells was determined by Evans Blue assay and few cells were found to be damaged with pH decrease. After neutralizing pH and adding nutrients to the flocculated medium, microalgae were proved to maintain a similar growth yield in the flocculated medium comparing with that in the fresh medium. The recycling of medium could contribute to the economical production from algae to biodiesel.
The study provided an economical, efficient and convenient method to harvest fresh microalgae. Advantages include capability of treating high cell biomass concentrations (>1g/L), excellent flocculation efficiencies (≥ 90%), operational simplicity, low cost and recycling of medium. It has shown the potential to overcome the hurdle of harvesting microalgae to promote full-scale application to biofuels from microalgae.
Microalgae; Harvest; Flocculation; pH decrease; Recycle
Significance: Hydrogen sulfide (H2S) has traditionally been considered a toxic environmental pollutant. In the late 1990s, the presumed solely harmful role of H2S has been challenged because H2S may also be involved in the maintenance and preservation of cardiovascular homeostasis. Recent Advances: The production of endogenous H2S has been attributed to three key enzymes, cystathionine γ-lyase (CSE), cystathionine β-synthase, and 3-mercaptopyruvate sulfurtransferase. The recognition of H2S as the third gaseous signaling molecule has stimulated research on a multitude of pathophysiologic events in the cardiovascular system. In particular, important roles in cardiovascular disorder processes are ascribed to the CSE/H2S pathway, such as atherosclerosis, myocardial infarction, hypertension, and shock. Critical Issues: Many biological activities and molecular mechanisms of H2S in the cardiovascular system have been demonstrated in studies using different tools, such as the genetic overexpression of CSE, the direct administration of H2S donors, or the use of H2S-releasing pro-drugs. Unfortunately, the role of the CSE/H2S pathway in cardiovascular disease remains controversial in numerous areas, and many questions regarding the gaseous molecule still remain unanswered. Future Directions: Advances in basic research indicate that the CSE/H2S pathway may provide potential therapeutic targets for treating cardiovascular disorders. But the molecular targets of H2S still need to be identified. Antioxid. Redox Signal. 17, 106–118.
Mitotic recombination is an effective tool for generating mutant clones in somatic tissues. Due to difficulties associated with detecting and quantifying mutant clones in mice, this technique is limited to analysis of growth related phenotypes induced by loss function of tumor suppressor genes. Here, we used the polymorphic CD45.1/CD45.2 alleles on chromosome 1 as pan-hematopoietic markers to track mosaic clones generated through mitotic recombination in developing T cells. We show that lineage specific mitotic recombination can be induced and reliably detected as CD45.1 or CD45.2 homozygous clones from the CD45.1/CD45.2 heterozygous background. We have applied this system in the analysis of a lethal mutation in the Dhx9 gene. Mosaic analysis revealed a stage specific role for Dhx9 during T cell maturation. Thus, the experimental system described in this study offers a practical means for mosaic analysis of germline mutations in the hematopoietic system.
Mitotic recombination; CD45.1/CD45.2; T lymphocyte; Dhx9; Cre/lox
Cyclooxygenase-2 (COX-2) expression is associated with many aspects of physiological and pathological conditions, including pancreatic β-cell dysfunction. Prostaglandin E2 (PGE2) production, as a consequence of COX-2 gene induction, has been reported to impair β-cell function. The molecular mechanisms involved in the regulation of COX-2 gene expression are not fully understood. We previously demonstrated that transcription factor Elk-1 significantly upregulated COX-2 gene promoter activity. In this report, we used pancreatic β-cell line (INS-1) to explore the relationships between Elk-1 and COX-2. We first investigated the effects of Elk-1 on COX-2 transcriptional regulation and expression in INS-1 cells. We thus undertook to study the binding of Elk-1 to its putative binding sites in the COX-2 promoter. We also analysed glucose-stimulated insulin secretion (GSIS) in INS-1 cells that overexpressed Elk-1. Our results demonstrate that Elk-1 efficiently upregulates COX-2 expression at least partly through directly binding to the −82/−69 region of COX-2 promoter. Overexpression of Elk-1 inhibits GSIS in INS-1 cells. These findings will be helpful for better understanding the transcriptional regulation of COX-2 in pancreatic β-cell. Moreover, Elk-1, the transcriptional regulator of COX-2 expression, will be a potential target for the prevention of β-cell dysfunction mediated by PGE2.
Apigenin (4’,5,7-trihydroxyflavone) was recently shown effective in inhibiting several cancers. The aim of this study was to investigate the effect and mechanism of apigenin in the human bladder cancer cell line T24 for the first time.
T24 cells were treated with varying concentrations and time of apigenin. Cell viability was evaluated by MTT assay. Cell motility and invasiveness were assayed by Matrigel migration and invasion assay. Flow cytometry and western blot analysis were used to detect cell apoptosis, cell cycle and signaling pathway.
The results demonstrated that apigenin suppressed proliferation and inhibited the migration and invasion potential of T24 bladder cancer cells in a dose- and time-dependent manner, which was associated with induced G2/M Phase cell cycle arrest and apoptosis. The mechanism of action is like to involve PI3K/Akt pathway and Bcl-2 family proteins. Apigenin increased caspase-3 activity and PARP cleavage, indicating that apigenin induced apoptosis in a caspase-dependent way.
These findings suggest that apigenin may be an effective way for treating human bladder cancer.
Apigenin; Bladder cancer; Cell cycle arrest; Apoptosis; Invasion
To evaluate the impact of the Janus kinase 2 single nucleotide polymorphisms (SNPs) on gastric cancer risk.
In this hospital-based, case–control study, the genotypes were identified by polymerase chain reaction–restriction fragment length polymorphism protocols in 661 individuals (359 gastric cancer patients and 302 age and sex matched cancer-free controls).
Both the frequency of A allele in rs2230724 and G allele in rs1887427 were more frequent in patients with gastric cancer (P = 0.013 and 0.001, respectively). Compared with the common genotype, subjects with the (AG+AA) genotypes of rs2230724 and the (AG+GG) genotypes of rs1887427 had a 59% and 98% increased risk of developing gastric cancer, respectively (P = 0.010, adjusted OR = 1.59, 95% CI = 1.12–2.27; P<0.001, adjusted OR = 1.98, 95% CI = 1.39–2.81, respectively). Further stratified analysis showed that the association between the risk of gastric cancer and the rare genotypes of rs2230724 were more profound in the subgroups of elder individuals (>56 years), males, nonsmokers and urban subjects, while the association between the risk and the rare genotypes of rs1887427 persisted in subgroups of younger individuals (≤56 years), males, nonsmokers and both of rural and urban subjects.
The JAK2 gene rs2230724 and rs1887427 polymorphisms are associated with an increased risk of gastric cancer in a Chinese Han population.
We compared the severity of white matter T2-hyperintensities (WMH) in the frontal lobe and occipital lobe using a visual MRI score in 102 patients with lobar intracerebral hemorrhage (ICH) diagnosed with possible or probable cerebral amyloid angiopathy (CAA), 99 patients with hypertension-related deep ICH, and 159 normal elderly subjects from a population-based cohort. The frontal-occipital (FO) gradient was used to describe the difference in the severity of WMH between the frontal lobe and occipital lobe. A higher proportion of subjects with obvious occipital dominant WMH (FO gradient ≤−2) was found among patients with lobar ICH than among healthy elderly subjects (FO gradient ≤−2: 13.7 vs. 5.7%, p = 0.03). Subjects with obvious occipital dominant WMH were more likely to have more WMH (p = 0.0006) and a significantly higher prevalence of the apolipoprotein E ε4 allele (45.8% vs. 19.4%, p = 0.04) than those who had obvious frontal dominant WMH. This finding is consistent with the relative predilection of CAA for posterior brain regions, and suggests that white matter lesions may preferentially occur in areas of greatest vascular pathology.
Intercerebral hemorrhage; White matter hyperintensity; Cerebral amyloid angiopathy
TRAM-34, a clotrimazole analog characterized as a potent and selective inhibitor of intermediate-conductance, calcium-activated K+ (IKCa) channels, has been used extensively in vitro and in vivo to study the biological roles of these channels. The major advantage of TRAM-34 over clotrimazole is the reported lack of inhibition of the former drug on cytochrome P450 (CYP) activity. CYPs, a large family of heme-containing oxidases, play essential roles in endogenous signaling and metabolic pathways, as well as in xenobiotic metabolism. However, previously published work has only characterized the effects of TRAM-34 on a single CYP isoform. To test the hypothesis that TRAM-34 may inhibit some CYP isoforms, the effects of this compound were presently studied on the activities of four rat and five human CYP isoforms. TRAM-34 inhibited recombinant rat CYP2B1, CYP2C6 and CYP2C11 and human CYP2B6, CYP2C19 and CYP3A4 with IC50 values ranging from 0.9 µM to 12.6 µM, but had no inhibitory effects (up to 80 µM) on recombinant rat CYP1A2, human CYP1A2, or human CYP19A1. TRAM-34 also had both stimulatory and inhibitory effects on human CYP3A4 activity, depending on the substrate used. These results show that low micromolar concentrations of TRAM-34 can inhibit several rat and human CYP isoforms, and suggest caution in the use of high concentrations of this drug as a selective IKCa channel blocker. In addition, in vivo use of TRAM-34 could lead to CYP-related drug-drug interactions.
Intracytoplasmic sperm injection (ICSI) is commonly used to solve male infertility problems. Previous studies showed that early environmental exposure of an embryo may influence postnatal development. To detect whether ICSI operations affect the reproductive health of a male or his offspring, we established assisted reproductive technologies (ART) conceived mouse models, and analyzed gene expression profiles in the testes of both ICSI and naturally conceived (NC) newborn F1 mice using micro-array analysis. Among the differentially expressed genes, we focused on the expression of eight male reproduction-related genes. Quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR) was used to analyze the expression of these genes in the testes of both adult and old F1 generation mice and adult F2 generation mice. Our results showed that down-regulated and somatic cell-expressed genes in newborn mice retained their differential expression patterns in adult and old F1 generation individuals, implying the persistence and fetal origin of the alteration in the expression of these genes. The intergenerational transmission of differential gene expression was observed, but most changes tended to be reduced in adult F2 generations. Controlled ovarian hyperstimulation (COH) and in vitro fertilization (IVF) mice models were added to explore the precise factors contributing to the differences in ICSI offspring. The data demonstrated that superovulation, in vitro culture, and mechanical stimulation involved in ICSI had a cumulative effect on the differential expression of these male reproductive genes.
Intracytoplasmic sperm injection; Testis; Intergenerational transmission
Longevity variability is a common feature of aging in mammals, but the mechanisms responsible for this remain largely unknown. Using microarray datasets coupled with Prediction analysis of microarrays (PAM), we identified a set of 252 cardiac transcripts predictive of relative lifespan in Wistar and Fisher 344 rats. PAM “tests” of rat heart transcriptomes from a third longer lived Fisher × Norway Brown rat strain validated the predictive value of this gene subset. The expression patterns of these genes were highly conserved, and corresponding promoter regions were employed to identify common cis–elements and trans-activating factors implicated in their control. Specifically, four transcription factors (Max, Ets2, Erg, and Msx2) present in heart displayed longevity-dependent, strain-independent changes in abundance, but only ETS2 had an expression profile that directly correlated with the relative lifespan gene set. In heart, ETS2 was prevalent in CMs and showed a high degree of myocyte-to-myocyte variability predominantly in adult rat hearts prior to the exponential increase in the rate of mortality. Exclusively in this group, elevated ETS2 significantly overlapped with TUNEL staining in heart myocytes. In response to sympathetic stimuli, ETS2 is also up-regulated, and functionally, adenovirus mediated over-expression of ETS2 promotes AIF-mediated, caspase-independent programmed necrosis exclusively in CMs that can be fully inhibited by the PARP-1 inhibitor DPQ. We conclude that variations in ETS2 abundance in hearts of adult rodents and the associated loss of CMs, contribute at least partially, to the longevity variability observed during normal aging of rats through activation of programmed necrosis.
Aging; Mortality; Rat; microarrays; PAM; transcription factors
The asymmetric unit of the title Pb-based coordination polymer, [Pb2(C24H16N2O8)(H2O)2]n, consists of one PbII cation, half of a 4,4′-(1,4-phenylene)bis(2,6-dimethylpyridine-3,5-dicarboxylate (L
4−) ligand and one coordinating water molecule. The centers of the benzene ring of the ligand and the four-membered Pb/O/Pb/O ring are located on centers of inversion. The PbII ion is coordinated in form of a distorted polyhedron by seven O atoms from four separate L
4− ligands and by one water O atom. The PbO7 polyhedra share O atoms, forming infinite zigzag [PbO4(H2O)]n chains along  that are bridged by L
4− ligands, forming a two-dimensional coordination network parallel to (001). O—H⋯O hydrogen bonds involving the water molecule are observed.
Hyperhomocysteinemia is a risk factor of atherogenesis. Soluble epoxide hydrolase (sEH) is a major enzyme hydrolyzing epoxyeicosatrienoic acids and attenuates their cardiovascular protective effects. Whether homocysteine (Hcy) regulates sEH and the underlying mechanism remains elusive.
To elucidate the mechanism by which Hcy regulates sEH expression and endothelial activation in vitro and in vivo.
Methods and Results
Hcy treatment in cultured human endothelial cells dose- and time-dependently upregulated sEH mRNA and protein. Hcy increased the expression of adhesion molecules, which was markedly reversed by inhibiting sEH activity. Hcy-induced sEH upregulation is associated with activation of activating transcription factor 6 (ATF6). Bioinformatics analysis revealed a putative ATF6-binding motif in the promoter region of the sEH gene, which was found being a methylation site. Site-directed mutagenesis and chromatin immunoprecipitation assays demonstrated that Hcy treatment or ATF6 overexpression promoted ATF6 binding to the promoter of sEH and increased its activity. Result of methylation-specific PCR revealed that the ATF6 binding site on the sEH promoter was partially methylated and was demethylated with Hcy. SiRNA knockdown of ATF6α and/or SP1 blocked, and ATF6 overexpression and DNA methyltransferase inhibitor mimicked, the effect of homocysteine on sEH upregulation. In vivo, immunofluorescence assay revealed elevated expression of sEH and adhesion molecules in the aortic intima of mice with mild hyperhomocysteinemia, which was attenuated by sEH deletion or inhibition.
ATF6 activation and DNA demethylation may coordinately contribute to Hcy-induced sEH expression and endothelial activation. Inhibition of sEH may be a therapeutic approach for treating Hcy-induced cardiovascular diseases.
sEH; homocysteine; ATF6; demethylation; endothelial cells
The XRCC1 polymorphisms have been implicated in bladder cancer risk, but individually published studies show inconsistent results. The aim of our study was to clarify the effects of XRCC1 variants on bladder cancer risk.
A systematic literature search up to September 13, 2012 was carried out in PubMed, EMBASE and Wanfang databases, and the references of retrieved articles were screened. Crude odds ratios with 95% confidence intervals were used to assess the associations between XRCC1 Arg194Trp and Arg399Gln polymorphisms and bladder cancer risk. Heterogeneity and publication bias were also evaluated.
A total of 14 and 18 studies were eligible for meta-analyses of Arg194Trp and Arg399Gln, respectively. Regrouping was adopted in accordance with the most probable appropriate genetic models. No obvious heterogeneity between studies was found. For overall bladder cancer, the pooled odds ratios for Arg194Trp and Arg399Gln were 1.69 (95% confidence interval: 1.25 to 2.28; P = 0.001) and 1.10 (95% confidence interval: 1.03 to 1.19; P = 0.008), respectively. After excluding the studies that were not in Hardy–Weinberg equilibrium, the estimated pooled odds ratio still did not change at all.
The meta-analysis results suggest that XRCC1 Arg194Trp and Arg399Gln polymorphisms may be associated with elevated bladder cancer risk.
XRCC1; Polymorphism; Bladder cancer; Meta-analysis
Diabetes is associated with increased risk of cancer at several sites, but its association with risk of bladder cancer is still controversial. We examined this association by conducting a systematic review and meta-analysis of cohort studies.
Studies were identified by searching PubMed, EMBASE, Scopus, Web of Science, Cochrane register, and Chinese National Knowledge Infrastructure (CNKI) databases through April 29, 2012. Summary relative risks (SRRs) with their corresponding 95% confidence intervals (CIs) were calculated using a random-effects model.
A total of fifteen cohort studies were included in this meta-analysis. Analysis of all studies showed that diabetes was associated with a borderline statistically significant increased risk of bladder cancer (RR 1.11, 95% CI 1.00–1.23; p<0.001 for heterogeneity; I2 = 84%). When restricting the analysis to studies that had adjusted for cigarette smoking (n = 6) or more than three confounders (n = 7), the RRs were 1.32 (95% CI 1.18–1.49) and 1.20 (95% CI 1.02–1.42), respectively. There was no significant publication bias (p = 0.62 for Egger’s regression asymmetry test).
Our findings support that diabetes was associated with an increased risk of bladder cancer. More future studies are warranted to get a better understanding of the association and to provide convincing evidence for clinical practice in bladder cancer prevention.
Genome-wide association studies (GWAS) have identified approximately three dozen single nucleotide polymorphisms (SNPs) consistently associated with prostate cancer (PCa) risk. Despite the reproducibility of these associations, the molecular mechanism for most of these SNPs has not been well elaborated as most lie within non-coding regions of the genome. Androgens play a key role in prostate carcinogenesis. Recently, using ChIP-on-chip technology, 22,447 androgen receptor (AR) binding sites have been mapped throughout the genome, greatly expanding the genomic regions potentially involved in androgen-mediated activity.
To test the hypothesis that sequence variants in AR binding sites are associated with PCa risk, we performed a systematic evaluation among two existing PCa GWAS cohorts; the Johns Hopkins Hospital and the Cancer Genetic Markers of Susceptibility (CGEMS) study population. We demonstrate that regions containing AR binding sites are significantly enriched for PCa risk-associated SNPs, i.e. more than expected by chance alone. In addition, compared with the entire genome, these newly observed risk-associated SNPs in these regions are significantly more likely to overlap with established PCa risk-associated SNPs from previous GWAS. These results are consistent with our previous finding from a bioinformatics analysis that one-third of the 33 known PCa risk-associated SNPs discovered by GWAS are located in regions of the genome containing AR binding sites.
The results to date provide novel statistical evidence suggesting an androgen-mediated mechanism by which some PCa associated SNPs act to influence PCa risk. However, these results are hypothesis generating and ultimately warrant testing through in-depth molecular analyses.
AR; prostate cancer; GWAS; pathway association study
The aim of the present study was to investigate the effects of Pseudomonas aeruginosa-mannose-sensitive hemagglutinin (PA-MSHA) on inhibiting the proliferation of bladder cancer cell lines and to further define its functional mechanisms. T24 and 5637 cells were treated with PA-MSHA at various concentrations and times. Cell proliferation was analyzed using Cell Counting Kit-8 (CCK-8) assays. The cell cycle distribution and apoptosis induced by PA-MSHA were measured by flow cytometry with propidium iodide (PI) and annexin V-fluorescein isothiocyanate (FITC) staining. Western blotting was used to evaluate the expression levels of the apoptosis-related molecules and PI3K-AKT-mTOR signaling pathway proteins. A time- and concentration-dependent cytotoxic effect of PA-MSHA was observed in the T24 and 5637 cells. Flow cytometry with PI and annexin V-FITC staining showed that the various concentrations of PA-MSHA were all able to induce the apoptosis and G0-G1 cell cycle arrest of the bladder cancer cells. Cleaved caspase-8 and -9 and Fas protein expression levels were markedly associated with an increase in the apoptosis of the bladder cancer cells. The cells stimulated with PA-MSHA also exhibited a downregulation of PI3K-AKT-mTOR signaling. PA-MSHA inhibits proliferation and induces apoptosis in the T24 and 5637 bladder cancer cell lines by modulating caspase family proteins and affecting the cell cycle regulation machinery. The PI3K-AKT-mTOR signaling pathway may be important in the direct anticancer cytotoxic effect of PA-MSHA.
apoptosis; bladder cancer; caspase; Pseudomonas aeruginosa-mannose-sensitive hemagglutinin vaccine