Toxicity and metabolism of 3-bromopyruvate in Caenorhabditis elegans / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

In this study, we aimed to evaluate the toxic effects, changes in life span, and expression of various metabolism-related genes in Caenorhabditis elegans, using RNA interference (RNAi) and mutant strains, after 3-bromopyruvate (3-BrPA) treatment. C. elegans was treated with various concentrations of 3-BrPA on nematode growth medium (NGM) plates, and their survival was monitored every 24 h. The expression of genes related to metabolism was measured by the real-time fluorescent quantitative polymerase chain reaction (qPCR). Nematode survival in the presence of 3-BrPA was also studied after silencing three hexokinase (HK) genes. The average life span of C. elegans cultured on NGM with 3-BrPA was shortened to 5.7 d compared with 7.7 d in the control group. hxk-1, hxk-2, and hxk-3 were overexpressed after the treatment with 3-BrPA. After successfully interfering hxk-1, hxk-2, and hxk-3, the 50% lethal concentration (LC50) of all mutant nematodes decreased with 3-BrPA treatment for 24 h compared with that of the control. All the cyp35 genes tested were overexpressed, except cyp-35B3. The induction of cyp-35A1 expression was most obvious. The LC50 values of the mutant strains cyp-35A1, cyp-35A2, cyp-35A4, cyp-35B3, and cyp-35C1 were lower than that of the control. Thus, the toxicity of 3-BrPA is closely related to its effect on hexokinase metabolism in nematodes, and the cyp-35 family plays a key role in the metabolism of 3-BrPA.

Blood pressure variability and its clinical significance in patients with acute cerebral infarction combined with H type hypertension / 中国医学装备

Objective:To investigate the blood pressure variability and its clinical significance in patients with acute cerebral infarction who combined with H type hypertension.Methods: 95 patients with acute cerebral infarction who combined with H type hypertension were enrolled in the perspective study. According to the homocysteine(Hcy) level of patients, they were divided into observation group ( Hcy ≥10mmol/L, 51cases) and control group (Hcy<10mmol/L, 44cases). The blood pressure variability and main clinical features of these patients in the two groups in 24h were observed. At the same time, the correlation between blood pressure variability and main clinical feature in observation group were analyzed.Results: The 24h systolic pressure variability, 24h diastolic pressure variability, carotid intima-media thickness and national institute of health stroke scale (NIHSS) of observation group were 14.57±4.62, 18.57±5.38, 13.39±4.85 mm and 1.27±0.17, respectively. While they were 12.48±3.78, 16.12±5.74, 11.34±4.32 mm and 1.09±0.13 in the control group, respectively. And the differences of them between the two groups were statistical significance (t=2.389,t=2.146,t=2.160,t=5.725,P<0.05). Besides, there was obvious positive correlation between 24h systolic pressure variability and NIHSS of patients with acute cerebral infarction (r=0.254,P<0.05), and there was also obvious positive correlation between 24h systolic pressure variability and thickness of carotid intima-media (r=0.256,P<0.05).Conclusion: Increased blood pressure variability in patients with acute hypertensive cerebral infarction combined with H type hypertension may be related to poor prognosis of patients and vascular intima thickness.

Consistency and clinicopathologic significance of BRAFV600E protein expression and genic mutation in papillary thyroid carcinoma / 临床与实验病理学杂志

Purpose To investigate the consistency and clinicopathologic correlation of BRAFV600E protein expression and gene mutation in papillary thyroid carcinoma. Methods BRAFV600E protein expression and genn mutation was detected respectively by immunohistochemistry of SP and real time-PCR, then the consistency between the both methods was analyzed by Kappa-test, the correlation between BRAFV600E and clinicopatho-logic parameters was analyzed by Chi-square test in papillary thyroid carcinoma. Results The gene mutation and protein expression rates of BRAFV600E were 89.3％ and 88.3％, respec-tively, the differences were not significant, the concordance rate of the both methods was 97.0％, Kappa value was 0.847, the consistence was higher, meanwhile the mutation rates between age ＜45 and ≥45 were respectively 96.8％ and 85.9％, there were significant differences, the positive rates of the both detec-tion methods were higher in thyroid capsule invaded group than non-invaded group, the differences were significant. Conclusion The both methods have higher consistency, the immunohisto-chemistry can be used as an initial screening tool for detecting gene mutation, the gene mutation of BRAFV600E is significantly associated with age and capsule invasion, the relationship is not found between BRAFV600E mutation and the other clinicopatholog-ic parameters.

MiR-200a and miR-200b target PTEN to regulate the endometrial cancer cell growth in vitro

Objective To study whether miR-200a and miR-200b target PTEN gene expression to regulate the endometrial cancer cell growth in vitro. Methods Endometrial cancer cells ECC-1 were cultured and transfected with the miR-200a and miR-200b mimics and inhibitors as well as the negative control mimics and inhibitors, and then the cell proliferation activity as well as the expression of PTEN and downstream genes in cells was determined; after transfection of miR-200a and miR-200b mimics as well as PTEN-3′UTR luciferase report gene plasmids, the fluorescence activity of luciferase reporter gene was determined. Results 12 h, 24 h and 48 h after transfection, the cell proliferation activity of miR-200a mimics group and miR-200b mimics group were significantly higher than those of NC mimics group while the cell proliferation activity of miR-200a inhibitor group and miR-200b inhibitor group were significantly lower than those of NC inhibitor group; 48 h after transfection, PTEN expression in cells and PTEN-3′UTR luciferase reporter gene fluorescence activity of miR-200a mimics group and miR-200b mimics group were significantly lower than those of NC mimics group while p-PI3K and p-Akt expression were significantly higher than those of NC mimics group; PTEN expression in cells and PTEN-3′UTR luciferase reporter gene fluorescence activity of miR-200a inhibitor group and miR-200b inhibitor group were significantly higher than those of NC inhibitor group while p-PI3K and p-Akt expression were significantly lower than those of NC inhibitor group. Conclusion miR-200a and miR-200b can promote the endometrial cancer cell growth in vitro by targeted inhibition of PTEN gene expression.

Research progress on liver stem cells / 第二军医大学学报

Liver stem cells are primitive cells with self-renewing capacity and bidirectional differentiation potential to differentiate into mature hepatocytes or biliary epithelial cells. Liver stem cells not only play roles in the homeostasis maintenance, injury repair and regeneration of liver, but also have huge potential applications in cell therapy of liver diseases, construction of bioartificial liver and liver-oriented gene therapy. In this review, we summarized the major types of liver stem cells, as well as their origins and molecular markers.

MiR-200a and miR-200b target PTEN to regulate the endometrial cancer cell growth in vitro

OBJECTIVE@#To study whether miR-200a and miR-200b target PTEN gene expression to regulate the endometrial cancer cell growth in vitro.@*METHODS@#Endometrial cancer cells ECC-1 were cultured and transfected with the miR-200a and miR-200b mimics and inhibitors as well as the negative control mimics and inhibitors, and then the cell proliferation activity as well as the expression of PTEN and downstream genes in cells was determined; after transfection of miR-200a and miR-200b mimics as well as PTEN-3'UTR luciferase report gene plasmids, the fluorescence activity of luciferase reporter gene was determined.@*RESULTS@#12 h, 24 h and 48 h after transfection, the cell proliferation activity of miR-200a mimics group and miR-200b mimics group were significantly higher than those of NC mimics group while the cell proliferation activity of miR-200a inhibitor group and miR-200b inhibitor group were significantly lower than those of NC inhibitor group; 48 h after transfection, PTEN expression in cells and PTEN-3'UTR luciferase reporter gene fluorescence activity of miR-200a mimics group and miR-200b mimics group were significantly lower than those of NC mimics group while p-PI3K and p-Akt expression were significantly higher than those of NC mimics group; PTEN expression in cells and PTEN-3'UTR luciferase reporter gene fluorescence activity of miR-200a inhibitor group and miR-200b inhibitor group were significantly higher than those of NC inhibitor group while p-PI3K and p-Akt expression were significantly lower than those of NC inhibitor group.@*CONCLUSION@#miR-200a and miR-200b can promote the endometrial cancer cell growth in vitro by targeted inhibition of PTEN gene expression.

Impact in response control, attention and hyperactivity behavior on children with obstructive sleep apnea hypopnea syndrome by integrated visual and auditory continuous performance test / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To study the impact in response control, attention and hyperactivity behavior on children with obstructive sleep apnea hypopnea syndrome (OSAHS) by the integrated visual and auditory continuous performance test (IVA-CPT).</p><p><b>METHODS</b>Fifty-one children aged between 5 and 12 years were diagnosed as OSAHS by polysomnography (PSG), and received adenotonsillectomy and adenoidectomy or only adenoidectomy. Then received IVA-CPT and PSG before surgery, 3 months after surgery and 6 months after surgery (named as first, second and third time point). These children were divided into two groups according to the disease course (group A: course of disease < 5 years; group B: course of disease ≥ 5 years). The SPSS 19.0 was used for statistical analysis.</p><p><b>RESULTS</b>By balanced test, there were no differences in gender, body mass index (BMI) and disease severity among the two groups before surgery (P > 0.05). The numbers of children with abnormal psychological behavior at three time points were 32 (62.7%), 25 (49.0%) and 8 (15.7%). The abnormal rate did not show statistical difference between the first and second time point (χ(2) = 1.49, P = 0.163), but did show statistical difference between the second and third time point (χ(2) = 12.95, P < 0.001). Repetitive measurement and analysis of variance showed that there were statistical differences in means of FRCQ, FAQ and HYP between three time points in two groups (F were 342.15, 263.12, 380.57, P < 0.001), and all the means improved with time. It also showed that there were statistical differences in means of FRCQ, FAQ and HYP between two groups at every time point (F were 167.05, 126.47, 117.683, P < 0.001). FRCQ and HYP all showed interation effect between two groups (P < 0.001). Means of apnea hypopnea index (AHI) and lowest arterial (LaSO2) were compared between three time points in two groups and all showed statistical differences (F were 99.057, 70.742, P < 0.001). Means of AHI and LaSO2 were compared between two groups at every time point. AHI and LaSO2 did not show statistical difference (P > 0.05). AHI and LaSO2 did not exist interation effect of disease course and time.</p><p><b>CONCLUSIONS</b>OSAHS obviously affect the children's response control, attention and hyperactivity behavior, but can recover gradually after adenotonsillectomy and adenoidectomy or only adenoidectomy. Therefore, Children with OSAHS should receive treatment as early as possible so as to reduce the influence on psychology.</p>

Study on the relativity among the attachment type, alexitymia and defense style of depression patients / 中华行为医学与脑科学杂志

ObjectiveTo evaluate the attachment type,the level of alexitymia and the defense style of depression patients and their relativity.Methods 32 depression patients were assessed with the adult attachment scale (AAS-1996 Revised Edition),toronto alexithymia scale (TAS) and defense style questionnaire( DSQ),and 80 healthy people were involved as controls.Results The depression patients showed significantly different attachment type from the control group( x2 =47.6,P ＜ 0.01 ).Compare with the control group,the depression patients showed higher scores in immature defense style( (5.47 ± 1.13 ) vs (3.79 ± 1.09 ) ) and neurotic defense style( (4.93 ±0.56) vs (4.20 ± 1.00),(P＜0.01) ).Compared with controls,the depression patients showed higher scores in TAS total score( (76.88 ± 8.94) vs (65.70 ± 7.98 ) ) factor Ⅰ ( ( 3.34 ± 0.77 ) vs ( 2.41 ±0.78)) and factor Ⅱ ((3.64 ±0.51) vs (2.65 ±0.55),P＜0.01),and lower scores in factor Ⅲ((2.51 ±0.78) vs (3.14 ± 1.03 ),P ＜ 0.01 ).The depression patients' adult attachment was significantly correlative with their scores of alexithymia.The depression patients' adult attachment was also significantly correlative with their scores of defense style.ConclusionThe attachment of depression patients may lead to depression via their defense style.Depression leads to alexitymia.Depression patients with different features of attachment show different level of alexitymia.

Lovastatin changes activities of lactate dehydrogenase A and B genes in rat myocardial cells / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Lactate dehydrogenase (LDH) is a crucial regulator of energy metabolism in many organs including the heart. Lovastatin is widely used in prevention and treatment of coronary heart disease and is a drug with substantial metabolic influences. Our study aimed to determine the activities of the lactate dehydrogenase A and B (LDHA and LDHB) genes following lovastatin treatment.</p><p><b>METHODS</b>The rat myocardial cell line H9c2(2-1) in culture was exposed to 100 nmol/L lovastatin for 24 hours or for five days. The functions of the LDHA and LDHB genes were examined at the transcriptional (mRNA) level with quantitative real-time polymerase chain reaction (Q-RT-PCR), and at the translational (protein) level with immunoblotting.</p><p><b>RESULTS</b>When compared with control levels, the LDHA mRNA went up by (151.65 ± 16.72)% (P = 0.0132) after 24 hours and by (175.28 ± 56.54)% (P = 0.0366) after five days of lovastatin treatment. Although 24 hours of lovastatin treatment had no significant effects on LDHB mRNA levels, when the treatment was extended to five days, LDHB mRNA levels were significantly down-regulated to (63.65 ± 15.21)% of control levels (P = 0.0117). After 24 hours of treatment with lovastatin, there were no significant changes in protein levels of either LDHA or LDHB. When treatment time was extended to five days, the protein levels of LDHA were up-regulated by (148.65 ± 11.81)% (P = 0.00969), while the protein levels of LDHB were down-regulated to (64.91 ± 5.47)% of control levels (P = 0.0192).</p><p><b>CONCLUSIONS</b>Lovastatin affects gene activities of LDHA and LDHB differently, which may reveal novel pharmacological effects of lovastatin.</p>

Relationship between serum ferritin levels and susceptibility to attention deficit hyperactivity disorder in children: a Meta analysis / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the possible relationship between serum ferritin levels and susceptibility to attention deficit hyperactivity disorder (ADHD) in children.</p><p><b>METHODS</b>The papers relating to the relationship between serum ferritin levels and susceptibility to childhood ADHD were searched in the Database CBM, CNKI, VIP and PubMed. The Meta-analysis software RevMan 5.0 was used for the heterogeneity test and for the pooled OR calculation. Sensitivity and publication bias analysis were performed.</p><p><b>RESULTS</b>Five control studies were included for the Meta analysis, including 258 cases of ADHD and 138 control cases. There was heterogeneity in the studies on the relationship between serum ferritin levels and susceptibility to childhood ADHD (P=0.003). So the studies were analyzed using the random-effect model. The pooled OR of serum ferritin levels and susceptibility to childhood ADHD was -23.09 (95%CI:-33.06-13.13; P<0.00001). The funnel plots did not indicate the existence of publication bias.</p><p><b>CONCLUSIONS</b>The results from present Meta analysis can prove that serum ferritin levels are associated with susceptibility to childhood ADHD.</p>

Influence of drug treatment on glucocorticoid receptor levels in patients with coronary heart disease / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Glucocorticoid signaling exerts major roles in inflammation, metabolism and depression, which are three crucial factors accompanying or underlying coronary heart disease. Although accumulating evidence indicates the influence of glucocorticoids on the pathology and treatment of coronary heart disease, there is still a dearth of pharmaceutical mechanisms for this relationship. This study aimed to investigate the influence of drug treatment on glucocorticoid receptor levels in coronary heart disease.</p><p><b>METHODS</b>Eighty hospitalized patients (average age (59.0 +/- 7.5) years, 46 male and 34 female) with coronary heart disease were categorized into four groups with 20 members in each according to one of the four drugs they were treated with. The four drugs were: nitrated derivative isosorbide dinitrate, the beta-adrenergic receptor blocker metoprolol, the calcium antagonist nifedipine, and the HMG-CoA reductase inhibitor lovastatin. Glucocorticoid receptor protein levels of peripheral blood lymphocytes were tested using immunoblotting analysis before and after one month of treatment.</p><p><b>RESULTS</b>Immunoblotting analysis showed increased glucocorticoid receptor levels after treatment with metoprolol and nifedipine. There were no statistically significant changes of glucocorticoid receptor levels after treatment with isosorbide dinitrate or lovastatin, although there were trends of up-regulation of glucocorticoid receptor expression after both treatments.</p><p><b>CONCLUSIONS</b>Both the beta-blocker and the calcium blocker can increase glucocorticoid receptor levels after chronic administration. This effect suggests a mechanism for their anti-inflammatory and other therapeutic roles for coronary heart disease and comorbid disorders.</p>

Higher concentration of CO₂ and 37°C stabilize the less virulent opaque cell of Candida albicans / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>Candida albicans (C. albicans) strains can spontaneously switch at a very low frequency from white to opaque phase. The ability to switch reversibly between white and opaque phenotype and contributes to the pathogenicity of C. albicans. White and opaque switching can be induced by various environmental signals. Previous study showed that opaque cells switch en masse to white when transferred in vitro to 37°C, the temperature of their animal host. The objective of the present study was to determine the effect of different concentration of carbon dioxide and temperature on white-opaque switching, and to determine the different anti-candida killing activity of white and opaque form by human monocyte-macrophage cell line THP-1.</p><p><b>METHODS</b>White-opaque switching and opaque-white switching were assayed. Modified Lee's medium supplemented with phloxine B was used to detect white and opaque forms of C. albicans under 0.03% CO2 at 25°C, 0.03% CO2 at 37°C and 5% CO2 at 37°C. Growth curve of C. albicans was monitored using OD value at 630 nm simultaneously. White and opaque forms of C. albicans and THP-1 cells were cocultured at ratio of 1:10. Colony serial dilutions were used to assay for intracellular candidacidal activity. MTT assay was used to measure the extracellular candidacidal activity.</p><p><b>RESULTS</b>Phenotype switching was successfully induced in vitro in all three strains of C. albicans. When evaluating white to opaque switching, opaque colony proportion of all colonies was 0.572 ± 0.087, 0.920 ± 0.030 and 0.985 ± 0.026 exposure of white cells to 0.03% CO2 at 25°C, 0.03% CO2 at 37°C and 5% CO2 at 37°C. When evaluating opaque to white switching, opaque colony proportion of all colonies was 0.600 ± 0.114, 0.983 ± 0.003 and 0.998 ± 0.003 exposure of white cells to 0.03% CO2 at 25°C, 0.03% CO2 at 37°C and 5% CO2 at 37°C. No significant difference of white or opaque form growth rate was found among three conditions (P > 0.05). THP-1 mediated extracellular anti-candida activity in white form was (79.80 ± 3.71)% and (56.28 ± 19.12)% at different dilution ratio, which were significantly lower than that in opaque form (100%, P < 0.01). THP-1 mediated intracellular anti-candida activity in white form ((62.98 ± 5.02)%) was significantly lower than that in opaque form ((87.07 ± 1.80)%, P < 0.01).</p><p><b>CONCLUSIONS</b>Our results showed that opaque form is more vulnerable and less virulent than that in white form. It suggested that higher concentration of CO2 and 37°C in host niches stabilize the less virulent opaque cell of C. albicans, which might have implications for pathogenesis, commensalism and mating.</p>

Insoluble beta-glucan from the cell wall of Candida albicans induces immune responses of human THP-1 monocytes through Dectin-1 / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>beta-glucan is the major structure component of Candida albicans (C. albicans) cell wall. It has been demonstrated that Dectin-1 as the principal C-type lectin pattern-recognition receptor (PRR) can recognize fungal beta-glucan and induce immune responses. In this study, we sought to clarify whether insoluble beta-glucan from the cell wall of C. albicans (CaIG) could induce immune responses in human THP-1 monocytes (a human acute monocytic leukemia cell line) and to determine the underlying mechanisms.</p><p><b>METHODS</b>Human THP-1 monocytes were challenged with CaIG in vitro. The mRNA expression of Dectin-1, Toll-like receptors (TLR2), proinflammatory cytokine (TNF-alpha) and chemokine (IL-8) was assayed by real-time reverse transcription polymerase chain reaction (RT-PCR). The secretion of TNF-a and IL-8 were measured by enzyme-linked immunosorbent assay (ELISA). H(2)O(2) release was determined by microplate fluorescent assay. Western blotting was used to analyze IkappaB-a phosphorylation and degradation.</p><p><b>RESULTS</b>Exposure of THP-1 monocytes to CaIG led to increased gene expression and secretion of TNF-alpha and IL-8. CaIG induced H(2)O(2) release in a time-dependent manner. CaIG hydrolyzed with zymolyase failed to induce gene expression and secretion of TNF-alpha, IL-8 and H(2)O(2) release. CaIG up-regulated the mRNA of Dectin-1, whereas the mRNA level of TLR2 was not altered. THP-1 monocytes challenged with CaIG resulted in the activation of NF-kappaB in a time-dependent manner. Dectin-1 inhibitor laminarin blocked the CaIG-induced production of TNF-alpha and H(2)O(2) in THP-1 monocytes, but no such effect was observed in pretreatment with anti-TLR2 neutralizing antibody and the LPS inhibitor (polymyxin B).</p><p><b>CONCLUSION</b>CaIG may play a role in activation of immune responses in human THP-1 cells through Dectin-1, not TLR2.</p>

Clinical analysis of primary percutaneous coronary intervention in patients with acute myocardial infarction / 中国基层医药

Objective To report the clinical effect of primary percutaneous coronary intervention(PCI)in patients with acute myocardial infarction(AMI).Methods A retrospective study was accomplished on the clinical data of 13 AMI patients who underwent PCI from March 2004 to April 2006.Results The infarct-related artery (IRA)was successfully recanalized by primary PCI for 12 AMI patients,without major complications occurred in these cases during hospitalization.Conclusion Primary PCI should be firstly chosen for treatment of AMI in the hospitals which could carry out PCI.

The relationship between postoperative complication and pathological features of 324 patients with neuronal intestinal malformations / 中华外科杂志

<p><b>OBJECTIVE</b>To investigate the relationship between the morphological features of different types of neuronal intestinal malformations (NIM) and their postoperative complications.</p><p><b>METHODS</b>The data of morphological and clinical features of 324 cases with NIM were analyzed retrospectively.</p><p><b>RESULTS</b>In all 324 patients, 210 cases were Hirschsprung's disease (HD), 38 intestinal neuronal dysplasia (IND), 45 mixed HD/IND, 8 hypoganglionosis, 22 combined HD/hypoganglionosis and 1 immaturity of ganglion cells. The percentages of normal neuron in bowel of different NIM were 88.1%, 24.4%, 18.4%, 4/8, 27.7% and 0/1 in HD, HD/IND, IND, hypoganglionosis, HD/hypoganglionosis and immaturity of ganglion cells respectively. There were totally 46 cases complicated with recurrent postoperative enterocolitis (EC). Incidence of recurrent postoperative EC in HD patients was 6.7% while in IND/HD and IND patients was 35.6% and 28.9%, respectively. Incidences of EC in cases with the residual IND margins and with the normal margins were 38.2% and 8.7%, respectively. Incidence of EC in cases with transanal endorectal pull-through procedure and with transabdominal procedure was 18.0% and 8.3%, respectively. Nine cases underwent another procedure because of severe persistent constipation or EC after operation, including 4 cases HD/IND, 1 case IND, 3 cases HD and 1 case HD/hypoganglionosis.</p><p><b>CONCLUSIONS</b>Neuron distribution is inconsistent with pathology of NIM. Postoperative EC are rare in the patients only with isolated HD. Furthermore, margins with residual IND and transanal endorectal pull-through procedure are risk factors to recurrent EC. However, the extension of excision about IND is uncertain and need further study.</p>

The role of subtypes of voltage-gated K+ channels in pulmonary vasoconstriction induced by 15-hydroeicosatetraenoic acid / 药学学报

<p><b>AIM</b>To observe the effect of subtypes of Kv channels in rat pulmonary artery smooth muscle cells (PASMCs) on the process of pulmonary vasoconstriction induced by 15-HETE.</p><p><b>METHODS</b>In the present study, ring of rabbit PA with specific Kv channel blockers were employed to functionally identify certain channel subtypes that took part in the process of 15-HETE induced pulmonary vasoconstriction; RT-PCR and Western blotting analysis were also used to measure the expression of subtypes of Kv in PASMCs exposed to 15-HETE,chronic hypoxia.</p><p><b>RESULTS</b>Blocking of Kv1. 1, Kv1. 2, Kv1. 3 and Kv1. 6 channels did not affect 15-HETE induced vasoconstriction in normoxic rats; 15-HETE did not affect expression of Kv1. 1 and Kv1. 2 channels; 15-HETE significantly downregulated the expression of mRNA and protein of Kv1. 5 and Kv2. 1 in rat PASMCs.</p><p><b>CONCLUSION</b>The results suggested that hypoxia may block Kv1. 5 and Kv2. 1 channels via 15-HETE mediated mechanism, leading to decrease numbers of functional Kv1. 5 and Kv2. 1 channels in PASMCs, leading to PA vasoconstriction.</p>

ERK1/2 signaling pathway is involved in 15-hydroxyeicosatetraenoic acid-induced hypoxic pulmonary vasoconstriction / 生理学报

Hypoxia-induced 15-hydroxyeicosatetraenoic acid (15-HETE) is an essential mediator to constrict pulmonary arteries (PA). The signaling pathway involved in 15-HETE-induced PA vasoconstriction remains obscure. The aim of the present study was to test the hypothesis that hypoxic PA constriction induced by 15-HETE was possibly regulated by the extracellular signal-regulated kinase-1/2 (ERK1/2) pathway. PA ring tension measurement, Western blot and immunocytochemistry were used in the study to determine the possible role of ERK1/2 in 15-HETE-induced PA vasoconstriction. The organ bath for PA rings tension study was employed. Adult male Wistar rats were raised in hypoxic environment with fractional inspired oxygen (FIO2, 0.12) for 9 d. PA 1~1.5 mm in diameter were dissected and cut into 3 mm long rings for tension study. ERK1/2 up-stream kinase (MEK) inhibitor PD98059, which blocks the activation of ERK1/2, was used. The results showed that pretreatment of PD98059 significantly blunted 15-HETE-induced PA vasoconstrictions in the rings from hypoxic rat. Moreover, in endothelium-denuded rings, PD98059 also significantly attenuated 15-HETE-induced vasoconstriction. Phosphorylation of ERK1/2 in pulmonary arterial smooth muscle cells (PASMCs) of rat was enhanced evidently when stimulated by 15-HETE. Thus, the data suggest that ERK1/2 signaling pathway is involved in 15-HETE-induced hypoxic pulmonary vasoconstriction.

15-hydroxyeicosatetraenoic acid depressed endothelial nitric oxide synthase activity in pulmonary artery / 生理学报

15-hydroxyeicosatetraenoic acid (15-HETE) plays an important role in hypoxia-induced pulmonary vasoconstriction. Release of nitric oxide (NO) is apparently decreased and activity of endothelial nitric oxide synthase (eNOS) is impaired in chronic hypoxia. However, little is known whether 15-HETE contributes to eNOS/NO pathway in the constriction induced by 15-HETE. We examined the response of rat pulmonary artery (PA) rings to 15-HETE, the production of NO, total eNOS expression and the phosphorylation of eNOS in bovine pulmonary artery endothelial cells (BPAECs) stimulated by 15-HETE. Rat PA rings were divided into three groups: endothelium intact group, endothelium denuded group, and nitro-L-arginine methyl ester (L-NAME, 0.1 mmol/L, an inhibitor of eNOS) group. Constrictions to 15-HETE were significantly enhanced in endothelium denuded group and L-NAME group (both P< 0.05 vs endothelium intact group, n= 9); BPAECs were incubated in different conditions to test nitrite production by Greiss method. Nitrite production was significantly reduced by 1 mumol/L 15-HETE (P<0.05), and increased by the lipoxygenase inhibitors, 10 mumol/L cinnamyl 3,4- dihydroxy-[alpha] -cyanocinnamate (CDC, P< 0.05) and 0.1 mmol/L nordihydroguiairetic acid (NDGA, P< 0.01 ); Western blot analysis of extracts from BPAECs incubated with 15-HETE in different time was carried out to test total eNOS expression, and the expression was changed unobviously. Immunoprecipitation (IP) and Western blot analysis of cell extracts from BPAECs treated with 2 mumol/L 15-HETE in different length of time were accomplished, using phospo-eNOS-threonine 495 (Thr495, an inhibitory site) antibody for IP, and eNOS or 15-lipoxygenase (15-LO) antibodies for Western blot. 15-HETE depressed eNOS activity by increasing the levels of phospho-eNOS-Thr 495. The data suggest that eNOS/NO pathway is involved in PA constrictions induced by 15-HETE and that 15-HETE depresses eNOS activity by phosphorylation in Thr495 site. The protein interaction between phospho-eNOS (Thr495) and 15-LO is discovered for the first time.