Assessing Asian Medical Students' Readiness for Diversity: Localizing Measures of Cross-Cultural Care Competence.

Phenomenon: There is now broad acceptance that the development of cross-cultural competence (CCC) supports the delivery of appropriate care to diverse groups and is an essential component in medical education worldwide. CCC training in East Asian contexts has been constrained by the fact that "cultural diversity" training globally has focused primarily on the needs of racial and ethnic communities, to the relative neglect of other groups. The present study explores Taiwanese students' perceptions of CCC provision to identify gaps in local medical education and thus facilitate a more systematic delivery of CCC. Approach: Using an adapted and translated version of the Cross-Cultural Care Survey developed by Harvard Medical School, we collected 1567 student responses from four geographically-distributed Taiwanese medical schools between 2015 and 2017. In addition to student responses, we also collected 122 clinical teacher responses from two of the four medical schools that were surveyed to cross-examine the students' self-perceived competence. The data were analyzed with SPSS and ANOVA was employed with student data to compare the differences among different stages. The analysis focuses on CCC in 4 stages of training: general education, basic pre-clinical and integrated clinical sciences, clerkship, and internship. Findings: The findings show that students felt unprepared to deal with health disparities and the needs of diverse groups and there was no evidence of an increased sense of preparedness in the development of relevant skills in the analyses of the pre-clinical to clinical stages of the curriculum. Similarly, teachers also perceived students across the different stages of training to be unprepared in dealing with the health disparities and needs of diverse groups. However, although findings from teachers' responses parallel those from students, teachers tend to perceive students to be even more unprepared than they perceive themselves to be. The training for CCC appeared inadequate from both set of data and students perceive CCC training to be less explicitly evident in the medical program as it advances from the foundation stage to the pre-clinical stage. Insights: The study raises some crucial issues in terms of diversity and CCC training in medical education programs. The fact that increased awareness of health disparities and the needs of diverse groups fails to be aligned with a sense of preparedness and skillfulness confirms that CCC has not been explicitly and sufficiently addressed in the medical curriculum, particularly in the pre-clinical stage when the focus is on acquiring scientific and technical knowledge. This study shows how a questionnaire designed by and for an American medical institution situated in a highly diverse society can be adapted so that its findings serve as a baseline for medical education programs in Taiwan, and perhaps in other countries that are beginning to acknowledge hitherto "hidden" diversity. This study also has implications which indicate that CCC is crucial in the delivery of appropriate care by members of the medical profession to diverse patients.

Emotional and Behavioral Health Needs in Elementary School Students in an Underserved Hispanic Community.

High rates of mental health problems in adolescents have been well documented; less is known about elementary school children in disadvantaged communities. We examined emotional and behavioral health needs in 202 third and fourth graders enrolled in a charter school in a largely Hispanic community. The child-reported Revised Child Anxiety and Depression Scale-25 and Teacher's Report Form were used to evaluate mental health needs as perceived by these children and their teachers. The prevalence of teacher-reported depression and child self-reported anxiety was 7.0% and 6.67%, respectively. Living in a single parent household was found to be a specific risk factor in that those children had higher rates of emotional and behavioral problems than children living with both parents. Evidence of higher depression and anxiety identified in this sample compared to national representative data suggests the need for development of culturally sensitive early prevention and intervention in this underserved community.

Infarct size-limiting effect of epoxyeicosatrienoic acid analog EET-B is mediated by hypoxia-inducible factor-1α via downregulation of prolyl hydroxylase 3.

Epoxyeicosatrienoic acids (EETs) decrease cardiac ischemia-reperfusion injury; however, the mechanism of their protective effect remains elusive. Here, we investigated the cardioprotective action of a novel EET analog, EET-B, in reperfusion and the role of hypoxia-inducible factor (HIF)-1α in such action of EET-B. Adult male rats were subjected to 30 min of left coronary artery occlusion followed by 2 h of reperfusion. Administration of 14,15-EET (2.5 mg/kg) or EET-B (2.5 mg/kg) 5 min before reperfusion reduced infarct size expressed as a percentage of the area at risk from 64.3 ± 1.3% in control to 42.6 ± 1.9% and 46.0 ± 1.6%, respectively, and their coadministration did not provide any stronger effect. The 14,15-EET antagonist 14,15-epoxyeicosa-5( Z)-enoic acid (2.5 mg/kg) inhibited the infarct size-limiting effect of EET-B (62.5 ± 1.1%). Similarly, the HIF-1α inhibitors 2-methoxyestradiol (2.5 mg/kg) and acriflavine (2 mg/kg) completely abolished the cardioprotective effect of EET-B. In a separate set of experiments, the immunoreactivity of HIF-1α and its degrading enzyme prolyl hydroxylase domain protein 3 (PHD3) were analyzed in the ischemic areas and nonischemic septa. At the end of ischemia, the HIF-1α immunogenic signal markedly increased in the ischemic area compared with the septum (10.31 ± 0.78% vs. 0.34 ± 0.08%). After 20 min and 2 h of reperfusion, HIF-1α immunoreactivity decreased to 2.40 ± 0.48% and 1.85 ± 0.43%, respectively, in the controls. EET-B blunted the decrease of HIF-1α immunoreactivity (7.80 ± 0.69% and 6.44 ± 1.37%, respectively) and significantly reduced PHD3 immunogenic signal in ischemic tissue after reperfusion. In conclusion, EET-B provides an infarct size-limiting effect at reperfusion that is mediated by HIF-1α and downregulation of its degrading enzyme PHD3. NEW & NOTEWORTHY The present study shows that EET-B is an effective agonistic 14,15-epoxyeicosatrienoic acid analog, and its administration before reperfusion markedly reduced myocardial infarction in rats. Most importantly, we demonstrate that increased hypoxia-inducible factor-1α levels play a role in cardioprotection mediated by EET-B in reperfusion likely by mechanisms including downregulation of the hypoxia-inducible factor -1α-degrading enzyme prolyl hydroxylase domain protein 3.

Sulforaphane modulates telomerase activity via epigenetic regulation in prostate cancer cell lines.

Epidemiologic studies have revealed that diets rich in sulforaphane (SFN), an isothiocyanate present in cruciferous vegetables, are associated with a marked decrease in prostate cancer incidence. The chemo-preventive role of SFN is associated with its histone de-acetylase inhibitor activity. However, the effect of SFN on chromatin composition and dynamic folding, especially in relation to HDAC inhibitor activity, remains poorly understood. In this study, we found that SFN can inhibit the expression and activity of human telomerase reverse transcriptase (hTERT), the catalytic subunit of telomerase, in 2 prostate cancer cell lines. This decrease in gene expression is correlated with SFN-induced changes in chromatin structure and composition. The SFN-mediated changes in levels of histone post-translational modifications, more specifically acetylation of histone H3 lysine 18 and di-methylation of histone H3 lysine 4, 2 modifications linked with high risk of prostate cancer recurrence, were associated with regulatory elements within the hTERT promoter region. Chromatin condensation may also play a role in SFN-mediated hTERT repression, since expression and recruitment of MeCP2, a known chromatin compactor, were altered in SFN treated prostate cancer cells. Chromatin immuno-precipitation (ChIP) of MeCP2 showed enrichment over regions of the hTERT promoter with increased nucleosome density. These combined results strongly support a role for SFN in the mediation of epigenetic events leading to the repression of hTERT in prostate cancer cells. This ability of SFN to modify chromatin composition and structure associated with target gene expression provides a new model by which dietary phytochemicals may exert their chemoprevention activity.

Fiber connectivity between the striatum and cortical and subcortical regions is associated with temperaments in Chinese males.

The seven-factor biopsychosocial model of personality distinguished four biologically based temperaments and three psychosocially based characters. Previous studies have suggested that the four temperaments-novelty seeking (NS), reward dependence (RD), harm avoidance (HA), and persistence (P)-have their respective neurobiological correlates, especially in the striatum-connected subcortical and cortical networks. However, few studies have investigated their neurobiological basis in the form of fiber connectivity between brain regions. This study correlated temperaments with fiber connectivity between the striatum and subcortical and cortical hub regions in a sample of 50 Chinese adult males. Generally consistent with our hypotheses, results showed that: (1) NS was positively correlated with fiber connectivity from the medial and lateral orbitofrontal cortex (mOFC, lOFC) and amygdala to the striatum; (2) RD was positively correlated with fiber connectivity from the mOFC, posterior cingulate cortex/retrosplenial cortex (PCC), hippocampus, and amygdala to the striatum; (3) HA was positively linked to fiber connectivity from the dorsolateral prefrontal cortex (dlPFC) and PCC to the striatum; and (4) P was positively linked to fiber connectivity from the mOFC to the striatum. These results extended the research on the neurobiological basis of temperaments by identifying their anatomical fiber connectivity correlates within the subcortical-cortical neural networks.

Comparison of recruitment methodologies for clinical trials: Results from the time for living and caring (TLC) intervention study.

BACKGROUND: Recruiting participants for research studies is a critical yet challenging task. Community-engaged recruitment strategies have gained prominence as effective means to engage diverse populations and ensure the representativeness of study samples. This case study aims to investigate the cost and effectiveness of various recruitment methods in enhancing research participation. METHODS: A comparative approach was employed to assess the outcomes of five different recruitment strategies used in the Time for Living & Caring (TLC) research study. Data on recruitment success, participant demographics, and retention rates were collected and analyzed using descriptive statistics, including ANOVA and Chi-squares, to statistically compare the outcomes associated with 5 different recruitment methodologies. The recruitment methodologies included two community-engaged strategies (community partner referral and community-based recruiters), a clinical database, social media, and word-of-mouth referral. CONCLUSION: The meta-data used to build this methodological case study describe different recruitment methodologies that may be used for clinical trials. This data-driven evaluation provides examples and considerations for researchers when developing budgets and proposals for future clinical trials. The primary finding is that there are tradeoffs in terms of cost, time, labor, and ultimately the representativeness of the sample, based on the type of recruitment methodology chosen.

Roles of endothelial nitric oxide synthase (eNOS) and mitochondrial permeability transition pore (MPTP) in epoxyeicosatrienoic acid (EET)-induced cardioprotection against infarction in intact rat hearts.

We previously demonstrated that 11,12 and 14,15-epoxeicosatrienoic acids (EETs) produce cardioprotection against ischemia-reperfusion injury in dogs and rats. Several signaling mechanisms have been implicated in the cardioprotective actions of the EETs; however, their mechanisms remain largely elusive. Since nitric oxide (NO) plays a significant role in cardioprotection and EETs have been demonstrated to induce NO production in various tissues, we hypothesized that NO is involved in mediating the EET actions in cardioprotection. To test this hypothesis, we used an in vivo rat model of infarction in which intact rat hearts were subjected to 30-min occlusion of the left coronary artery and 2-hr reperfusion. 11,12-EET or 14,15-EET (2.5mg/kg) administered 10min prior to the occlusion reduced infarct size, expressed as a percentage of the AAR (IS/AAR), from 63.9±0.8% (control) to 45.3±1.2% and 45.5±1.7%, respectively. A nonselective nitric oxide synthase (NOS) inhibitor, L-NAME (1.0mg/kg) or a selective endothelial NOS inhibitor, L-NIO (0.30mg/kg) alone did not affect IS/AAR but they completely abolished the cardioprotective effects of the EETs. On the other hand, a selective neuronal NOS inhibitor, nNOS I (0.03mg/kg) and a selective inducible NOS inhibitor, 1400W (0.10mg/kg) did not affect IS/AAR or block the cardioprotective effects of the EETs. Administration of 11,12-EET (2.5mg/kg) to the rats also transiently increased the plasma NO concentration. 14,15-EET (10µM) induced the phosphorylation of eNOS (Ser(1177)) as well as a transient increase of NO production in rat cardiomyoblast cell line (H9c2 cells). When 11,12-EET or 14,15-EET was administered at 5min prior to reperfusion, infarct size was also reduced to 42.8±2.2% and 42.6±1.9%, respectively. Interestingly, L-NAME (1.0mg/kg) and a mitochondrial KATP channel blocker, 5-HD (10mg/kg) did not abolish while a sarcolemmal KATP channel blocker, HMR 1098 (6.0mg/kg) and a mitochondrial permeability transition pore (MPTP) opener, atractyloside (5.0mg/kg) completely abolished the cardioprotection produced by the EETs. 14,15-EET (1.5mg/kg) with an inhibitor of MPTP opening, cyclosporin A (CsA, 1.0mg/kg) produced a greater reduction of infarct size than their individual administration. Conversely, an EET antagonist 14,15-epoxyeicosa-5(Z)-enoic acid (14,15-EEZE, 2.5mg/kg) completely abolished the cardioprotective effects of CsA, suggesting a role of MPTP in mediating the EET actions. Taken together, these results suggest that the cardioprotective effects of the EETs in an acute ischemia-reperfusion model are mediated by distinct mediators depending on the time of EET administration. The cardioprotective effects of EETs administered prior to ischemia were regulated by the activation of eNOS and increased NO production, while sarcKATP channels and MPTP were involved in the beneficial effects of the EETs when administered just prior to reperfusion.

Nociceptive-induced myocardial remote conditioning is mediated by neuronal gamma protein kinase C.

Deciphering the remote conditioning molecular mechanism may provide targets to develop therapeutics that can broaden the clinical application. To further investigate this, we tested whether two protein kinase C (PKC) isozymes, the ubiquitously expressed epsilon PKC (εPKC) and the neuronal-specific gamma PKC (γPKC), mediate nociceptive-induced remote myocardial conditioning. Male Sprague-Dawley rats were used for both in vivo and ex vivo myocardial ischemia-reperfusion protocols. For the in vivo studies, using a surgical abdominal incision for comparison, applying only to the abdomen either bradykinin or the εPKC activator (ψεRACK) reduced myocardial infarct size (45 ± 1, 44 ± 2 %, respectively, vs. incision: 43 ± 2 %, and control: 63 ± 2 %, P < 0.001). Western blot showed only εPKC, and not γPKC, is highly expressed in the myocardium. However, applying a selective γPKC inhibitor (γV5-3) to the abdominal skin blocked remote protection by any of these strategies. Using an ex vivo isolated heart model without an intact nervous system, only selective εPKC activation, unlike a selective classical PKC isozyme activator (activating α, ß, ßII, and γ), reduced myocardial injury. Importantly, the classical PKC isozyme activator given to the abdomen in vivo (with an intact nervous system including γPKC) during myocardial ischemia reduced infarct size as effectively as an abdominal incision or ψεRACK (45 ± 1 vs. 45 ± 2 and 47 ± 1 %, respectively). The classical PKC activator-induced protection was also blocked by spinal cord surgical transection. These findings identified potential remote conditioning mimetics, with these strategies effective even during myocardial ischemia. A novel mechanism of nociceptive-induced remote conditioning, involving γPKC, was also identified.

Intestinal microbiota determine severity of myocardial infarction in rats.

Signals from the intestinal microbiota are important for normal host physiology; alteration of the microbiota (dysbiosis) is associated with multiple disease states. We determined the effect of antibiotic-induced intestinal dysbiosis on circulating cytokine levels and severity of ischemia/reperfusion injury in the heart. Treatment of Dahl S rats with a minimally absorbed antibiotic vancomycin, in the drinking water, decreased circulating leptin levels by 38%, resulted in smaller myocardial infarcts (27% reduction), and improved recovery of postischemic mechanical function (35%) as compared with untreated controls. Vancomycin altered the abundance of intestinal bacteria and fungi, measured by 16S and 18S ribosomal DNA quantity. Pretreatment with leptin (0.12 µg/kg i.v.) 24 h before ischemia/reperfusion abolished cardioprotection produced by vancomycin treatment. Dahl S rats fed the commercially available probiotic product Goodbelly, which contains the leptin-suppressing bacteria Lactobacillus plantarum 299v, also resulted in decreased circulating leptin levels by 41%, smaller myocardial infarcts (29% reduction), and greater recovery of postischemic mechanical function (23%). Pretreatment with leptin (0.12 µg/kg i.v.) abolished cardioprotection produced by Goodbelly. This proof-of-concept study is the first to identify a mechanistic link between changes in intestinal microbiota and myocardial infarction and demonstrates that a probiotic supplement can reduce myocardial infarct size.

Instrumental lying by parents in the US and China.

The practice of lying to one's children to encourage behavioral compliance was investigated among parents in the US (N = 114) and China (N = 85). The vast majority of parents (84% in the US and 98% in China) reported having lied to their children for this purpose. Within each country, the practice most frequently took the form of falsely threatening to leave a child alone in public if he or she refused to follow the parent. Crosscultural differences were seen: A larger proportion of the parents in China reported that they employed instrumental lie-telling to promote behavioral compliance, and a larger proportion approved of this practice, as compared to the parents in the US. This difference was not seen on measures relating to the practice of lying to promote positive feelings, or on measures relating to statements about fantasy characters such as the tooth fairy. Findings are discussed with reference to sociocultural values and certain parenting-related challenges that extend across cultures.

Eribis peptide 94 reduces infarct size in rat hearts via activation of centrally located µ opioid receptors.

Eribis peptide 94 (EP 94) is a novel enkephalin derivative that binds with high potency to µ and δ opioid receptors with less affinity for the κ opioid receptor. This compound has recently been shown to produce an acute reduction in myocardial infarct size in the anesthetized pig and rat partially via an endothelial nitric oxide synthase and KATP channel-dependent mechanism. EP 94 also was found to produce a chronic reduction in infarct size 24 hours postdrug administration via the upregulation of inducible nitric oxide synthase in rats. Despite these findings, no data have emerged in which the opioid receptor subtype responsible for cardioprotection has been identified and the site of action, heart, other peripheral organs, or the central nervous system, has not been addressed. In the current study, EP 94, was administered in 2 divided doses (0.5 µg/kg, intravenously) at 5 and 10 minutes into the ischemic period, and the opioid antagonists were administered 10 minutes before the onset of the 30-minute ischemic period. The selective antagonists used were the µ receptor antagonist CTOP (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2), the δ receptor antagonists naltrindole and BNTX (7-benzylidenenaltrexone), and the κ receptor antagonist nor-BNI (norbinaltorphimine). Surprisingly, only CTOP completely blocked the cardioprotective effect of EP 94, whereas naltrindole, BNTX, and nor-BNI had modest but nonsignificant effects. Because there is controversial evidence suggesting that µ receptors may be absent in the adult rat myocardium, it was hypothesized that the protective effect of EP 94 may be mediated by an action outside the heart, perhaps in the central nervous system. To test this hypothesis, rats were pretreated with the nonselective opioid antagonist, naloxone hydrochloride, which penetrates the blood-brain barrier or naloxone methiodide, the quaternary salt of naloxone hydrochloride, which does not penetrate the blood-brain barrier before EP 94 administration. In support of a central nervous system site of action for EP 94, naloxone hydrochloride completely blocked its cardioprotective effect, whereas naloxone methiodide had no effect. These results suggest that EP 94 reduces infarct size (expressed as a percent of the area at risk) in the rat primarily via activation of central µ opioid receptors.

Acute and chronic cardioprotection by the enkephalin analogue, Eribis peptide 94, is mediated via activation of nitric oxide synthase and adenosine triphosphate-regulated potassium channels.

BACKGROUND/AIMS: Eribis peptide 94 (EP 94) is a new enkephalin derivative which potently binds to the µ- and δ-opioid receptor. In this study, we determined the effects of EP 94 and potential mechanism(s) involved in cardioprotection of the rat heart. METHODS AND RESULTS: An acute (5 and10 min into ischemia) and a chronic (24 h prior to ischemia) EP 94 administration produced a similar 30-40% reduction in infarct size/area at risk and the effects were blocked by the K(ATP) channel antagonists, HMR 1098 and 5-HD. The cardioprotective effects were blocked by a nonselective nitric oxide synthase (NOS) inhibitor (L-NAME) following acute administration and by a selective iNOS inhibitor (1400W) following chronic administration. CONCLUSION: These results suggest that EP 94 may have potential for the treatment of ischemic heart disease via a nitric oxide (NO)-K(ATP)-mediated mechanism.

Medical students' perceptions of their preparedness to care for LGBT patients in Taiwan: Is medical education keeping up with social progress?

INTRODUCTION: Integrating training on health equity of sexual and gender minorities (SGM) in medical education has been challenging globally despite emphasis on the need for medical students to develop competence to provide adequate care for diverse patient groups. This study elicits Taiwanese medical students' perceptions of their values and preparedness to care for Lesbian, Gay, Bisexual, or Transgender (LGBT) patients using a qualitative approach that considers broader societal changes, and more focused topics such as the provision of relevant training in medical education. METHODS: Eighty-nine medical students/trainees from two southern Taiwanese medical schools (one public and one private) participated in focus groups (n = 70) and individual interviews (n = 19). Qualitative analysis was conducted using inductive thematic analysis. RESULTS: Participants (i) expressed wide social acceptance and openness toward LGBT individuals, but were unsure of ways to communicate with LGBT patients; (ii) confirmed that stigmatization and biases might be developed during their training; (iii) recognized gender stereotypes could have negative impacts on clinical reasoning; (iv) considered themselves prepared to care for LGBT patients, yet equated non-discriminatory attitudes to preparedness; (v) acknowledged a lack of relevant professional skills; (vi) implicated curriculum did not address LGBT issues systematically and explicitly. CONCLUSION: This study has identified the insufficiencies of current medical training and inadequate preparedness of medical students/trainees to provide better care for LGBT patients. It provides insights for medical educators to design and implement effective medical curriculum and training, and faculty development programs to equip medical students/trainees with self-awareness and competencies to more readily provide holistic care for SGM, in keeping up with social progress, and promote health equity for a more diverse patient population.

Bioavailability and inter-conversion of sulforaphane and erucin in human subjects consuming broccoli sprouts or broccoli supplement in a cross-over study design.

Broccoli consumption may reduce the risk of various cancers and many broccoli supplements are now available. The bioavailability and excretion of the mercapturic acid pathway metabolites isothiocyanates after human consumption of broccoli supplements has not been tested. Two important isothiocyanates from broccoli are sulforaphane and erucin. We employed a cross-over study design in which 12 subjects consumed 40 g of fresh broccoli sprouts followed by a 1 month washout period and then the same 12 subjects consumed 6 pills of a broccoli supplement. As negative controls for isothiocyanate consumption four additional subjects consumed alfalfa sprouts during the first phase and placebo pills during the second. Blood and urine samples were collected for 48h during each phase and analyzed for sulforaphane and erucin metabolites using LC-MS/MS. The bioavailability of sulforaphane and erucin is dramatically lower when subjects consume broccoli supplements compared to fresh broccoli sprouts. The peaks in plasma concentrations and urinary excretion were also delayed when subjects consumed the broccoli supplement. GSTP1 polymorphisms did not affect the metabolism or excretion of sulforaphane or erucin. Sulforaphane and erucin are able to interconvert in vivo and this interconversion is consistent within each subject but variable between subjects. This study confirms that consumption of broccoli supplements devoid of myrosinase activity does not produce equivalent plasma concentrations of the bioactive isothiocyanate metabolites compared to broccoli sprouts. This has implications for people who consume the recommended serving size (1 pill) of a broccoli supplement and believe they are getting equivalent doses of isothiocyanates.

Metabolism and tissue distribution of sulforaphane in Nrf2 knockout and wild-type mice.

PURPOSE: To determine the metabolism and tissue distribution of the dietary chemoprotective agent sulforaphane following oral administration to wild-type and Nrf2 knockout (Nrf2(-/-)) mice. METHODS: Male and female wild-type and Nrf2(-/-) mice were given sulforaphane (5 or 20 µmoles) by oral gavage; plasma, liver, kidney, small intestine, colon, lung, brain and prostate were collected at 2, 6 and 24 h (h). The five major metabolites of sulforaphane were measured in tissues by high performance liquid chromatography coupled with tandem mass spectrometry. RESULTS: Sulforaphane metabolites were detected in all tissues at 2 and 6 h post gavage, with the highest concentrations in the small intestine, prostate, kidney and lung. A dose-dependent increase in sulforaphane concentrations was observed in all tissues except prostate. At 5 µmole, Nrf2(-/-) genotype had no effect on sulforaphane metabolism. Only Nrf2(-/-) females given 20 µmoles sulforaphane for 6 h exhibited a marked increase in tissue sulforaphane metabolite concentrations. The relative abundance of each metabolite was not strikingly different between genders and genotypes. CONCLUSIONS: Sulforaphane is metabolized and reaches target tissues in wild-type and Nrf2(-/-) mice. These data provide further evidence that sulforaphane is bioavailable and may be an effective dietary chemoprevention agent for several tissue sites.

Evidence for a role of opioids in epoxyeicosatrienoic acid-induced cardioprotection in rat hearts.

We previously demonstrated that several epoxyeicosatrienoic acids (EETs) produce reductions in myocardial infarct size in rats and dogs. Since a recent study demonstrated the release of opioids in mediating the antinociceptive effect of 14,15-EET, we hypothesized that endogenous opioids may also be involved in mediating the cardioprotective effect of the EETs. To test this hypothesis, we used an in vivo rat model of infarction and a rat Langendorff model. In the infarct model, hearts were subjected to 30 min occlusion of the left coronary artery and 2 h reperfusion. Animals were treated with 11,12-EET or 14,15-EET (2.5 mg/kg) alone 15 min before occlusion or with opioid antagonists [naloxone, naltrindole, nor-binaltorphimine (nor-BNI), and d-Phe-Cys-Tyr-d-Trp-Om-Thr-Pen-Thr-NH(2) (CTOP), a nonselective, a selective delta, a selective kappa, and a selective mu receptor antagonist, respectively] 10 min before EET administration. In four separate groups, antiserum to Met- and Leu-enkephalin and dynorphin-A-(1-17) was administered 50 min before the 11,12-EET administration. Infarct size expressed as a percent of the area at risk (IS/AAR) was 63.5 + or - 1.2, 45.3 + or - 1.0, and 40.9 + or - 1.2% for control, 11,12-EET, and 14,15-EET, respectively. The protective effects of 11,12-EET were abolished by pretreatment with either naloxone (60.5 + or - 1.8%), naltrindole (60.8 + or - 1.0%), nor-BNI (62.3 + or - 2.8%), or Met-enkephalin antiserum (63.2 + or - 1.7%) but not CTOP (42.0 + or - 3.0%). In isolated heart experiments, 11,12-EET was administered to the perfusate 15 min before 20 min global ischemia followed by 45 min reperfusion in control hearts or in those pretreated with pertussis toxin (48 h). 11,12-EET increased the recovery of left ventricular developed pressure from 33 + or - 1 to 45 + or - 6% (P < 0.05) and reduced IS/AAR from 37 + or - 4 to 20 + or - 3% (P < 0.05). Both pertussis toxin and naloxone abolished these beneficial effects of 11,12-EET. Taken together, these results suggest that the major cardioprotective effects of the EETs depend on activation of a G(i/o) protein-coupled delta- and/or kappa-opioid receptor.

Acute methadone treatment reduces myocardial infarct size via the delta-opioid receptor in rats during reperfusion.

BACKGROUND: Methadone is an opioid agonist often given to manage acute and chronic pain. We sought to determine whether methadone compared with morphine dose dependently reduces myocardial infarct size (IS) and whether the mechanism is delta-opioid receptor mediated. Furthermore, we examined whether myocardial IS reduction varies with the timing of methadone administration or duration of induced ischemia. METHODS: After surgical instrumentation, we divided male Sprague-Dawley rats into 3 sets. The first set was divided into groups, which received methadone (0.03-3 mg/kg), morphine (0.03-3 mg/kg), or water (placebo) 30 min before ischemia. Some animals of the first set also received the delta-opioid antagonist naltrindole (5 mg/kg) before methadone (0.3 mg/kg), morphine (0.3 mg/kg), or placebo administration. The second set of animals was divided into groups that received methadone (0.3 mg/kg) 5 min before reperfusion or 10 s after reperfusion. These 2 sets of animals were subjected to 30 min of myocardial ischemia by left anterior descending coronary artery occlusion and then 2 h of reperfusion. The third set of animals received placebo, methadone (0.3 mg/kg), or morphine (0.3 mg/kg) 5 min before reperfusion and were subjected to 45 min of ischemia by left anterior descending coronary artery occlusion with 2 h of reperfusion. Myocardial IS was assessed by staining myocardial tissue with triphenyltetrazolium chloride and expressed as a percentage of the area at risk (mean +/- sem). RESULTS: Methadone or morphine administered before ischemia reduced myocardial IS. The greatest effect was achieved at a dose of 0.3 mg/kg (methadone, 46% +/- 1%, P < 0.001 and morphine, 47% +/- 1%, P < 0.001 versus placebo, 61% +/- 1%, respectively). Naltrindole (5 mg/kg) blocked methadone-induced (0.3 mg/kg) and morphine-induced (0.3 mg/kg) cardioprotection (naltrindole + methadone, 58% +/- 1%, P < 0.001 versus methadone; and naltrindole + morphine, 58 +/- 1%, P < 0.001 versus morphine). Methadone (0.3 mg/kg) reduced myocardial IS when given 5 min before reperfusion (46% +/- 1%, P < 0.001 versus placebo) but not 10 s after reperfusion (60% +/- 1%, P = 0.675 versus placebo). No significant myocardial IS differences were seen for placebo when comparing the 45-min ischemia group (64% +/- 1%) with the 30-min ischemia group (60% +/- 1%, P = 0.069). The longer ischemia time of 45 min abrogated methadone-induced IS reduction (64% +/- 2%, P = 0.867 versus 45-min ischemia placebo group) and morphine-induced IS reduction (65% +/- 1%, P = 0.836 versus 45-min ischemia placebo group). CONCLUSIONS: These findings demonstrate that methadone and morphine produce similar myocardial IS-sparing effects that are delta-opioid receptor mediated and that are dependent on the duration of myocardial ischemia.

Human thrombopoietin reduces myocardial infarct size, apoptosis, and stunning following ischaemia/reperfusion in rats.

AIMS: Thrombopoietin (Tpo) is known for its ability to stimulate platelet production. However, it is currently unknown whether Tpo plays a physiological function in the heart. METHODS AND RESULTS: We assessed the potential protective role of Tpo in vitro and in vivo in two rat models of myocardial ischaemia/reperfusion. Tpo receptor (c-mpl) message was detected in the heart using RT-PCR, and the Tpo receptor protein was detected using western blotting and immunohistochemistry. Tpo treatment immediately before ischaemia reduced myocardial necrosis, apoptosis, and decline in ventricular function following ischaemia/reperfusion in the rat in a concentration- and dose-dependent manner with an optimal concentration of 1.0 ng/mL in vitro and an optimal dose of 0.05 microg/kg iv in vivo. Tpo also reduced infarct size when given after the onset of ischaemia or at reperfusion. Tpo activated JAK-2 (Janus kinase-2) and p44 MAPK (mitogen-activated protein kinase) during reperfusion but not prior to ischaemia. Inhibition of JAK-2 (AG-490), p42/44 MAPK (PD98059), mitochondrial K(ATP) channels (5-HD), and sarcolemmal K(ATP) channels (HMR 1098) abolished Tpo-induced resistance to injury from myocardial ischaemia/reperfusion. AG-490, PD98059, 5-HD, and HMR1098 alone had no effect on cardioprotection. Treatment with a single dose of Tpo (0.05 or 1.0 microg/kg iv) did not result in the elevation of platelet count or haematocrit over a 16-day period. CONCLUSION: A single treatment of Tpo confers cardioprotection through JAK-2, p42/44 MAPK, and K(ATP) channels, suggesting a potential therapeutic role of Tpo in the treatment of injury resulting from myocardial ischaemia and reperfusion.

Gadolinium limits myocardial infarction in the rat: dose-response, temporal relations and mechanisms.

The lanthanide cation, gadolinium (Gd) attenuates post-ischemic myocardial stunning. This study tests the hypothesis that Gd also preconditions the myocardium against infarction following ischemia-reperfusion (IR) and explores potential mechanisms underlying Gd-induced cardioprotection. Regional myocardial infarction was induced in rats by occluding the left anterior descending artery for 30 min and reperfusing for 120 min. Rats (n=6/group) were administered intravenous Gd (1 to 100 micromol/kg) 15 min prior to ischemia. Hearts were excised after reperfusion to determine infarct size (IS) and area at risk (AAR). The ratio IS/AAR (%) was reduced by Gd in a "U"-shaped, dose-dependent manner. The minimum dose that reduced IS/AAR was 5 micromol/kg (52+/-5% vs. 64+/-4%), while the dose that reduced IS/AAR maximally was 20 micromol/kg (44+/-4%). Gd also reduced IS/AAR when given 1 min before reperfusion (47+/-3%) but not when given 10 s after reperfusion (60+/-3%). Cardioprotection was maintained if IR was delayed 24-72 h after Gd administration. Cardioprotection by Gd was abolished by inhibition of JAK-2 with AG-490, of p42/44 MAPK with PD98059 or of K(ATP) channels with glibenclamide. None of these agents given alone altered IS/AAR compared with controls. Inhibition of JAK-2 also blocked Gd-induced delayed cardioprotection. Gd may have broad potential roles in IR, as it conferred immediate cardioprotection when given prior to ischemia or prior to reperfusion and delayed cardioprotection for up to 72 h after administration. The mechanism underlying Gd-induced preconditioning appears to be multi-factorial, involving JAK-2, STAT-3 and p44 MAPK pathways, as well as K(ATP) channels.

Diabetes abolishes morphine-induced cardioprotection via multiple pathways upstream of glycogen synthase kinase-3beta.

The cardioprotective effect of opioids or glycogen synthase kinase (GSK) inhibitors given at reperfusion has not been investigated in diabetes models. Therefore, nondiabetic (NDBR) or streptozotocin-induced diabetic (DBR) rat hearts were subjected to 30 min of ischemia and 2 h of reperfusion. Groups of NDBR or DBR were administered either vehicle, morphine (0.3 mg/kg), or the GSK inhibitor SB216763 (0.6 mg/kg) 5 min before reperfusion. SB216763 (but not morphine) reduced infarct size in DBRs (44 +/- 1* and 55 +/- 2%, respectively), while both agents reduced infarct size in NDBRs versus untreated NDBRs or DBRs (44 +/- 3*, 42 +/- 3*, 60 +/- 2, and 56 +/- 2%, respectively, *P < 0.001). Morphine-induced phospho- (P-)GSK3beta was reduced 5 min after reperfusion in DBRs compared with NDBRs (0.83 +/- 0.29 and 1.94 +/- 0.12 [P < 0.05] pg/microg tissue, respectively). The GSK3beta mediators, P-Akt, P-extracellular signal-related kinase (ERK)1, and P-signal transducer and activator of transcription (STAT)3, were also significantly reduced in untreated DBR compared with NDBR rats. Morphine-induced elevations of P-Akt, P-ERK1, P-p70s6, P-janus-activated kinase-2, and P-STAT3 in NDBRs were also blunted in DBRs. H9C2 cells raised in 25 mmol/l compared with 5.56 mmol/l glucose media also demonstrated reduced morphine-induced P-GSK3beta, P-Akt, P-STAT3, and P-ERK1 after 15 min. Hence, acute GSK inhibition may provide a novel therapeutic strategy for diabetic patients during an acute myocardial infarction, whereas morphine is less effective due to signaling events that adversely affect GSK3beta.