LOC100131429) was significantly associated with log-PV  after adjusting for multiple testing. Other noteworthy SNPs that were nominally associated  with log-PV included rs9583484 (intronic SNP in COL4A2), rs10146527 (intronic SNP in NRXN3), rs9909466 (SNP near RPL32P31), and rs2241606 (synonymous SNP in SLC12A7). We found several SNPs in metabolic loci associated with PV. Further studies are needed to confirm our results and elucidate the mechanism between these genetic loci, PV, and clinical BPH.Prostate enlargement leading to clinical benign prostatic hyperplasia (BPH) is associated with metabolic dysregulation and obesity. The genetic basis of this association is unclear. Our objective was to evaluate whether single nucleotide polymorphisms (SNPs) previously associated with metabolic disorders are also associated with prostate volume (PV). Participants included 876 men referred for prostate biopsy and found to be prostate cancer free. PV was measured by transrectal ultrasound. Samples were genotyped using the Illumina Cardio-MetaboChip platform. Multivariable adjusted linear regression models were used to evaluate SNPs (additive coding) in relation to natural-log transformed (log) PV. We compared SNP-PV results from biopsy-negative men to 442 men with low-grade prostate cancer with similar levels of obesity and PV. Beta-coefficients from the discovery and replication samples were then aggregated with fixed effects inverse variance weighted meta-analysis. SNP rs11736129 (near the pseudo-gene  A highly prevalent condition in aging men, benign prostatic hyperplasia (BPH) is the non-malignant proliferation of the epithelial and stromal cells in the prostate gland . It is oThe etiology of BPH is not fully understood. In the last decade, interest has increased for metabolic syndrome and markers of metabolism as potential risk factors for BPH and LUTS –7. SeverIRX4, ITGA6, and RFX6 genes were also associated with increased BPH risk or increased BPH aggressiveness [While genetic susceptibility to metabolic dysregulation may contribute to BPH progression, few studies have evaluated genetic susceptibility to BPH. A study among twins found high concordance rates for benign prostate disease, suggesting a hereditary influence . Most stsiveness . The exaWe used the Illumina Cardio-MetaboChip to evaluate associations between PV and over 100,000 genetic polymorphisms in genes or genomic regions previously associated with anthropomorphic and metabolic traits . ConsideParticipants for this study were participants of the Nashville Men’s Health Study (NMHS), a multi-center, rapid-recruitment protocol targeting men seeking a diagnostic prostate biopsy. Briefly, among men referred for prostate biopsy to VUMC and to the Tennessee Valley Veterans Administration in Nashville between January 1, 2006 and March 30, 2011, men who were ≥40 years of age, had English proficiency sufficient for informed consent, no prior diagnosis of PC, no prior prostate surgery, and reported no current use of androgen supplementation were considered eligible for participation. Institutional review boards at Vanderbilt University Medical Center (VUMC) and at the Tennessee Valley Veterans Administration  approved all study aspects including and not limited to recruitment, data collection protocols and written informed consent procedures. All study participants provided written informed consent. Recruitment procedures were conducted prior to the prostate biopsy and transrectal ultrasound (TRUS) procedures to prevent selection bias during data collection associated with knowledge of outcome.Trained research staff collected information on demographic characteristics and lifestyle factors, and measured anthropometric traits including height, weight, hip and waist circumference, which were used to calculate body mass index (BMI) and waist-to-hip ratio (WHR). Blood samples were genotyped with the Illumina Cardio-MetaboChip , which iA total of 969 eligible men free of PC with DNA availability were randomly selected for genotyping and served as our primary research group to examine the relationship between SNPs and prostate volume. We also wanted to investigate the consistency of any signals in a separate group of 520 men with low-grade PC. These men were recruited from the same recruitment protocol as the biopsy-negative men, with identical data and bio-specimen protocols. The rationale for the comparison is that low-grade and localized PC lesions are common, and with the exception of the detection of a localized tumor, men without PC at biopsy and men with low-grade PC are similar with regard to prostate size, prostate-specific antigen (PSA) levels, BMI, and the prevalence of CVD and other obesity-related comorbidities. We intentionally did not include high-grade PC cases from the validation sample because genetic factors leading to advanced PC are likely to be distinct from those involved in prostate enlargement, and men with high-grade PC had significantly smaller prostate volume and higher PSA levels than men with either low-grade PC or men without PC at biopsy. In other words, elevated PSA levels in men diagnosed with high-grade PC are likely a consequence of the PC, whereas elevated PSA levels among men with low-grade PC are likely, or in large part, a consequence of increasing PV, similar to men with elevated PSA and without PC ,35.-6 . We further excluded 38 individuals (30 PC-free men and 2 PC cases) as they did not have data available for prostate volume. Our final analysis consisted of a discovery sample of 876 men without PC at biopsy and a validation sample of 442 men with low-grade PC, for whom data were also available for both prostate volume and genotype information. We limited our investigation to SNPs that had minor allele frequencies of 5% or more, which left us with a total of 100,559 SNPs available for primary data analysis. We used principal component analysis with EIGENSOFT to evaluate the presence of population substructure in our study sample. We derived 10 principal components after LD-pruning our dataset to include 39,980 independent markers with minor allele frequency ≥0.05. A plot representing the first two principal components for our samples along with reference populations from the International HapMap Project has been published previously [Nine hundred and forty one men without PC and 459 low-grade PC cases were successfully genotyped. Post genotype-calling quality control (QC) procedures were conducted using the software package PLINK . Individeviously .We transformed the continuous PV (ml) variable using the natural-log function to approximate the normal distribution. We compared the characteristics of men without PC and men with low-grade PC with the student’s t-test with unequal variances for comparison of group means, Mann-Whitney rank sum test for the comparison of group medians, and Pearson’s chi-squared test for categorical variables. To ensure that the associations between major risk factors for PV in our study sample are comparable to that of other studies that have evaluated PV as a dependent variable, and also to ensure that the associations are similar among men without PC and men with low-grade PC, we used linear regression to evaluate the relationship between important covariates and log-transformed PV.-7) that considers the multiple tests in our study. We evaluated genomic inflation with a quantile-quantile plot  and natural-log transformed PV in two models 1) a minimally adjusted model including age and 10 principal components; 2) a fully adjusted model including age, height, BMI and 10 principal components in men without PC and men with low-grade PC, separately. Results from both models were similar; we present results from fully adjusted models. We first analyzed men without PC and men with low-grade PC separately, and then to summarize results from these two groups we performed an inverse variance weighted fixed effect meta-analysis of the beta-coefficients from our models considering men without PC and men with low-grade PC. We present beta-coefficients and standard errors as change of log-PV per minor allele. To account for multiple-testing we chose to estimate the family-wise error rate (FWER) in our study. We first used simpleM to estimate the effective number of independent tests . Brieflyile plot .To separate the genetic vs. non-genetic contributions of height, BMI and WHR on prostate volume, we created genetic risk scores (GRS) for height, BMI, and WHR from SNPs previously identified by GIANT and CHARGE consortium analyses to be associated with BMI , WHR 4040, and hWe then used STATA to regress natural-log transformed values of prostate volume against un-weighted and weighted genetic risk scores for height, BMI and WHR in linear regression models that adjusted for age and 10 principal components of ancestry, and with or without traits of interest .Distribution of characteristics in men without PC and men with low-grade PC are presented in Non-genetic predictors of PV were similar across diagnostic groups. Age, height, BMI, and statin use were all independently and positively associated with increased prostate volume in participants without a PC diagnosis and participants with low-grade PC . WHR and-4 , and heterogeneity in the rs11736129 and PV association was low between the two clinical groups. (I2 = 11%), indicating a similar association within each group. SNP rs9583484, located in the intronic region of the COL4A2 gene was nominally associated with PV , with a stronger association among biopsy-negative men (I2 = 51%). Additionally, two SNPs near NRXN3 , a SNP near RPL32P31  and a synonymous polymorphism in the SLC12A7 exon  were nominally associated with log-PV at the p-value threshold of < 1x10-04.A total of 100,538 SNPs with minor allele frequencies of 5% or higher were available for our single variant analyses. We estimated the association between each SNP and PV in multivariable linear regression models adjusting for age, height, BMI, and 10 principal components summarizing ancestry. Analyses were conducted within each group, and then inverse variance weighted fixed effect meta-analysis was used to combine beta coefficients from the two clinical groups. There were 5 independent loci associated with PV from the meta-analysis at a p-value threshold of less than 1x10-4 . SNP rs1-4 (-5) near ZEB1 and ARHGAP12, and rs12662869  in and intron of SLC17A1/A4. In men with low-grade PC, PV was associated with SNPs in the intronic regions of PKP2, AKAP13, and SNPs near LOC100131429, AGTR1, and ANAPC1.We next identified loci associated with log-PV within each clinical group at a p-value threshold of <1x10-4 . Among mAs anthropomorphic traits such as height and measures of obesity are consistently associated with prostate enlargement, we additionally investigated the associations between GRSs of height, WHR and BMI in relation to log-PV in each group . Both unMetabolic syndrome and its components have been hypothesized to increase risk for BPH and LUTS, however it is not clear if these associations are at least in part mediated by genetic susceptibility to the metabolic syndrome, and if so, by which specific variants. Hypothesizing that genetic variants related to the metabolic syndrome may also be positively associated with BPH, we evaluated SNPs throughout the genome that have been implicated with several metabolic disorders in relation to prostate volume in men without PC and men with low-grade PC. As the first study to systematically assess this hypothesis, we report a genome-wide significant association and also provide preliminary evidence for several additional loci putatively associated with prostate volume.LOC100131429, which bears sequence similarity to the armadillo repeat containing 1 gene (ARMC1) which encodes a metal ion binding protein. While pseudo-genes lack coding potential due to the presence of various mutations such as premature stop codons and frame shifts, unprocessed pseudo-genes like the LOC100131429 may be transcribed. The SNP rs11736129 is annotated by ENCODE to lie within an enhancer histone mark, and thus may be an expression quantitative trait locus [MYLKP1 pseudo-gene and its functional counterpart, the expression of the non-coding RNA of a pseudo-gene may inhibit the expression of the functional gene by decreasing its mRNA stability [LOC100121429 pseudo-gene and the ARMC1 gene are expressed in normal human prostate tissues [ARMC1 gene was associated with childhood obesity in a Hispanic population [ARMC1 pseudo-gene and BPH are speculative and require confirmation at this point. SLC7A11 is the closest protein-coding gene near rs11736129, which is approximately 250KB from the SNP. The gene product of SLC7A11 is a component of an anionic antiporter transport system which regulates cysteine and glutamine transport. This transport system, also known as the xCT antiporter system, has been proposed as a drug intervention target for cancers such as common triple-negative breast cancer [rs11736129, the most statistically significant result in our analysis, lies approximately 13 kilo-bases downstream of the pseudo-gene tability . RNA exp tissues . The ARMpulation , howevert cancer , glioma t cancer  and panct cancer .NRXN3 associated with PV. NRXN3 gene product plays important role in cell adhesion, is expressed in the prostate tissue [NRXN3 was significantly associated with waist-circumference in a large analysis by the CHARGE consortium [COL4A2 gene, a gene providing the major structural component of basement membranes. COL4A2 is expressed in normal prostate tissues, induced by androgens, and the C-terminal portion of the COL4A2 gene is thought to inhibit angiogenesis [COL4A2 has been suggested as a biomarker for screening BPH [COL4A1 and COL4A2 may be over-expressed, and COL3A1 and COL5A2 expression may be repressed by androgen exposure [RPL32P31 pseudogene and polymorphisms in the solute carrier family of genes  involved in the transport of sodium and other inorganic compounds across the cell membrane and the sodium/potassium channel (SLC17A1).Among loci for which evidence was suggestive, we found two polymorphisms in the e tissue , and hase tissue . Interesnsortium , with BMnsortium  and clinnsortium . Similarogenesis . COL4A2 ning BPH . Pritchaexposure , suggesta priori determined metabolic genetic variants throughout the genome in relation to PV, our results are preliminary and need to be replicated by other independent studies before drawing any definitive conclusions. Assuming a minor allele frequency of 0.10, a conservative p-value threshold of , and a sample size of 1300 participants (assuming both PC-free men and low-grade PC cases together), we had approximately 83% power to detect a beta-coefficient of 0.15. The effect estimates that we observed for the most part were less than 0.15, and we were likely underpowered to detect these smaller associations. With the exception of SNP at LOC100131429 that reached genome-wide significance , the associations for all of the other SNPs reported here are not statistically significant after adjustment for multiple-comparisons.As the first study to systematically evaluate We note several limitations of this study. Firstly, our analysis is limited to the evaluation between SNPs and total prostate volume. We did not have information on transitional zonal volume; availability of this information would have provided for a more refined analysis as transitional zone has been shown to be a better predictor of BPH severity than total prostate volume . UnderstDespite these caveats, our study has several strengths. The measurement of PV and accession of covariates for all participants in the study were taken prior to prostate biopsy, therefore reducing the potential for information bias. We used PC-free men as our discovery sample and men with low-grade PC as our validation sample after first demonstrating that, aside from prostate volume, these two groups had similar body size and clinical characteristics. Obesity may have a separate association with advanced PC , and we Finally, we evaluated the compound genetic components of metabolism-related traits such as height, BMI and WHR in relation to prostate volume. Height, BMI, and WHR were positively associated with prostate volume. However, our investigation of genetic risk scores for these anthropomorphic traits did not identify a shared genetic component between body size measures and prostate enlargement, suggesting that the association between obesity or height with prostate size is a consequence of endocrine factors or other non-genetic factors.In conclusion, we identified several genetic loci related to metabolic disorders that were also associated with prostate enlargement. Replication of these loci in additional populations and subsequent functional and mechanistic studies will be needed to further understand the genetic epidemiology of BPH.S1 Fig2). Plot shows the expected p-values under the uniform distribution versus the observed p-values from study. PC = Prostate Cancer.Quantile-quantile plot for the meta-analysis p-values from fully adjusted model: adjusted for age, height, BMI and 10 principal components; BMI = body mass index (kg/m(TIFF)Click here for additional data file.


P<0.001) as compared to Non-Occl-Con pigs; however, Occl-Ex pigs exhibited partial recovery (P<0.001) intermediate to the other two groups. BK induced relaxation was reduced in LADs from Occl-Ex and Occl-Sed pigs (P<0.001), compared to Non-Occl-Con, and exercise modestly increased responses to BK (P<0.05). In addition, SNP, PE, Ang II, and ET-1 responses were not significantly different among the groups. Our results indicate that ‘simple’ occlusion of the femoral arteries induces vascular dysfunction in conduit vessels distinct from the affected hindlimbs, as evident in blunted coronary vasorelaxation responses to ADP and BK. These findings imply that PAI, even in the absence of frank atherogenic vascular disease, contributes to vascular dysfunction in the coronary arteries that could exacerbate disease outcome in patients with peripheral artery disease. Further, regular daily physical activity partially recovered the deficit observed in the coronary arteries.The present study was designed to determine whether adult swine with peripheral artery insufficiency (PAI) would exhibit vascular dysfunction in vessels distinct from the affected distal limbs, the coronary conduit arteries. Moreover, we sought to evaluate the effect of exercise training on coronary vasomotor function in PAI. Eighteen female healthy young Yucatan miniature swine were randomly assigned to either occluded exercise trained , or occluded-sedentary , or non-occluded, non-exercised control  groups. Occl-Ex pigs were progressively trained by running on a treadmill . The left descending artery (LAD) and left circumflex (LCX) coronary arteries were harvested. Vasorelaxation to adenosine diphosphate (ADP), bradykinin (BK), and sodium nitro-prusside (SNP) were assessed in LAD’s; while constrictor responses to phenylephrine (PE), angiotensin II (Ang II), and endothelin-1 (ET-1) were assessed in LCX’s. Vasorelaxation to ADP was reduced in LADs from Occl-Sed and Occl-Ex pigs ( PAD is a major cause of disability, loss of work, and lifestyle changes in the United States. It affects approximately 8 to 12 million persons in the country, and its prevalence increases with advancing age, as approximately 12 to 20% of people older than 70 years suffer from this condition.Peripheral artery disease (PAD) impedes blood flow typically to the lower extremities leading to a mismatch of oxygen delivery that becomes more evident during physical activity. The impaired perfusion observed in PAD can foster functional and structural modifications in the affected limb  which attempt to ameliorate the symptoms. This is especially true if the patients become physically active. However, PAD patients also exhibit decreased nitric oxide (NO) activity that is associated with an increased production of vasoconstrictors and sympathetic outflow,6.One half of all PAD patients older than 55 years are asymptomatic; whilst among the symptomatic, 40% experience intermittent claudication, the most common clinical manifestation of PAD, and 10% have limb ischemia at rest with increased mortality due to cardiovascular disease, especially CAD , as well as a 6.6-fold higher overall mortality than non-PAD patients. Interestingly, in patients with concomitant CAD and PAD the presence of peripheral inflammation has been postulated to contribute to coronary endothelial dysfunction, mostly due to a reduction in NO bioavailability. Indeed, many conceive that the association of PAD with CAD is simply the result of generalized atherosclerosis progressing in both arterial trees. However, an estimated 35 to 50% of all patients with PAD do not exhibit CAD or cerebral vascular disease (CVD),11. There are studies reporting a widespread incidence of normal coronary arteries among PAD patients–14. For instance, coronary angiography was found to be normal in 85% of symptomatic PAD patients . Duran et al. reported that coronary artery angiography was within normal limits in 28% of patients with PAD . Furthermore, the study by Hertzer et al. found that CAD was present in only 270 of 468 elderly patients (58%) with PAD who were living in a long-term health care facility. Nonetheless, these PAD patients in the absence of frank CAD and CVD have the same markedly increased morbidity and mortality as those PAD patients with attendant central vascular disease. It is possible that the ischemic limb muscles in PAD release substances and/or initiate neural reflex responses systemically that lead to coronary artery dysfunction and potentially greater susceptibility to CAD. It is well documented that exercise training improves walking ability and reduces cardiovascular mortality and morbidity of claudicant patients,17. Indeed, Sakamoto et al. reported that supervised exercise training reduced overall cardiovascular mortality by 52% and morbidity by 30% among PAD patients. Based on this background, the present study was designed to determine whether in an atherosclerosis free, preclinical animal model of peripheral artery insufficiency (PAI) bilateral occlusion of the femoral arteries would lead to vascular dysfunction within a vascular bed, the coronary conduit vessels, distant from the affected hindlimb vasculature. This animal model of PAI exhibits limb blood flow that is non-ischemic at rest, but is impaired during exercise,19, similar to flow patterns seen with intermittent claudication. In addition, we sought to evaluate the effect of exercise training on coronary endothelial function in this animal model of PAI. Exercise training has been shown to increase endothelial-dependent vasorelaxation of peripheral vessels in patients with intermittent claudication, and in patients with CAD. Thus, we hypothesized that a preclinical animal model of PAI would exhibit coronary dysfunction and that this would be ameliorated by exercise.PAD patients are reported to have increased incidence of coronary artery disease (CAD)–25, were used in this study. The pigs were housed in the animal care facility in rooms maintained at 20–23°C with a 12h: 12h light-dark cycle. They were fed a normal diet (Purina Laboratory 5082 Mini-Pig Breeder Chow) with ad libitum access to water. All experimental protocols were approved by the Institutional Animal Care and Use Committee of the University of Missouri.Healthy young female Yucatan miniature swine , a breed relatively resistant to atherogenesis.Animals were pre-anesthetized with ketamine (35 mg/ kg)-xylazine (2.25 mg/kg) intramuscular injection. A tracheal tube was inserted, and secured. Pigs were anesthetized by using 2% isoflurane gas with a closed circuit anesthesia machine and an auto-respirator (Ohmed 7000). Surgical areas were cleaned with warm soap water, sterilized with Betadine and alcohol. A 4cm incision was made on the pig’s neck to expose left carotid artery. A medical perfusion catheter  was inserted via the carotid artery and advanced into the thoracic aorta for blood pressure monitoring. Blood pressure, respiration rate, heart rate, ECG, and body temperature were monitored during the surgery procedure. A 4 cm longitudinal skin incision was made in each groin area above the femoral artery which was occluded with 2–0 surgical silk at 2 cm distal from the inguinal ligament. The femoral occlusion animal model employed in this experiment has been used extensively by our laboratoryAt the beginning of the study all pigs were familiarized with walking on a motorized treadmill for 5 days prior to surgery and then randomly assigned into exercise and sedentary groups. The occluded-exercise  were endurance trained on a treadmill at 0% grade, beginning one week post-surgery; whilst the occluded-sedentary  were limited to cage activity within a 2 x 4 m pen. Occl-Ex pigs underwent progressive daily training sessions 5 days/week for 12 weeks. The training protocol included a warm-up prolonged running and cools down stages, where the durations and intensity of exercise bouts were increased as the animals were capable. Briefly, warm-up and cool down stages lasted for 5 min each at 1.5 mph. The endurance stage was running at a constant speed of 3.1 mph, building up to a total time of 75 min as the animals were capable. This intensity could easily be achieved by the occluded animals for approximately 75 min/session within approximately 9 weeks after beginning training. Exercise stress tests were conducted for both occluded groups, once every week by running on the treadmill beginning at 1mph and increasing the speed by 1mph each min until the pig was unable to continue. Blood pressure and heart rate were monitored during the stress tests. In addition, non-occluded pigs, age-matched, served as a control group ; these pigs were not exercise neither trained nor given the stress test each week.At the end of the 12 weeks of exercise training, pigs were anesthetized with intramuscular ketamine (35 mg/ kg)-xylazine (2.25 mg/kg), and intravenous thiopenthal (25 mg/ kg) for deep anesthesia, and the heart was removed to achieve euthanasia and placed in iced Krebs solution. The left anterior descending (LAD) and left circumflex (LCX) coronary arteries were removed from the heart. The arteries were trimmed of connective tissue and fat in cold Krebs bicarbonate buffer solution; and cut into six LAD and two LCX vessel rings each 3–4 mm in length. Cut rings were photographed on an Olympus SZH video microscope which was connected to a Spot Insight camera . Then, the rings morphological characteristics  were measured using Image J software .in vitro studies, and mounted on myographs  by positioning two stainless steel wires in the lumen of each ring. Arterial rings were placed in a 20 mL bath of Krebs bicarbonate solution. The optimal point in the length-tension relationship (Lmax) was established through incremental (10% of passive outside diameter) increases in stretch in combination with potassium chloride (KCl: 50 mM) administration to the rings. The remainder of the study was performed at the experimentally determined Lmax of each ring. Contractile responses to KCl were performed prior to vasorelaxation studies. KCl (80 mM) was administered twice to all of the arterial rings until the increase in tension had reached a plateau (10 min). Following the KCl experiments, Krebs bicarbonate solution was replaced every 20 min until resting tension was achieved.Six LAD and two LCX rings were used, for −9–10−4 M) and bradykinin ; whilst the assessment of endothelium-independent vasorelaxation utilized increasing doses of sodium nitroprusside . The LAD rings were preconstricted with prostaglandin F2α  and allowed to achieve a plateau in tension development before the addition of vasodilators. For each pig’s heart ring one was left untreated (intact). Ring two was pretreated with nitro-L-arginine methyl ester  to inhibit the nitric oxide synthase (NOS) pathway. The third ring was pretreated with indomethacin  to inhibit cyclooxygenase (COX), the enzyme responsible for prostacyclin (PGI2) and prostaglandin production. Ring four was pretreated with a combination of L-NAME and Indo to assess the importance of NOS-and COX-independent mechanisms of relaxation. Ring five was pretreated with a combination of L-NAME, Indo, and potassium chloride (KCl) to block NOS, COX and the non-NOS, non-COX pathway, endothelium-derived hyperpolarizing factor (EDHF). Finally, the endothelium was removed from ring six; “denuded” by gentle rubbing of the luminal surface with fine-tipped forceps to study the endothelium- independent mechanisms, explicitly smooth muscle relaxation. Adequate ring denudation was tested by examining responses to maximal BK (30 mM). Ring six was considered to be properly denuded by the inability of BK to induce vasorelaxation (i.e. less than 5%) in PGF2α (30 μM) preconstricted rings. The order of agonists throughout the entire study was ADP, BK, and SNP. Following each agonist induced dose response; Krebs bicarbonate solution was replaced at 20min intervals until resting tension of all arterial rings was reached (roughly 60min), before the next protocol was initiated.It was designed to examine the relaxation responses. Endothelium-dependent, dose-dependent vasorelaxation was assessed in six LAD rings using cumulative addition of adenosine diphosphate , angiotensin II  and endothelin-1 . Ring one was left untreated; “intact”, whilst on ring two the endothelium was removed as described above; “denuded”.The vasoconstrictor properties were assessed in two LCX rings using cumulative addition of phenylephrine  and maintained at 37°C. Angiotensin II was purchased from Bachem Americas , and all other drugs and chemicals were purchased from Sigma Chemical .The Krebs–bicarbonate buffer solution contained (in mM) 131.5NaCl, 5.0KCl, 1.2NaH50) values, and maximal relaxation responses were determined via one-way ANOVA with a Kruskal-Wallis test using Graph Pad Prism version 5.0a. Means of the EC50 values are presented as the negative log of the molar concentration. The analysis of concentration-response curves was performed using the mixed procedure in SAS version 9. The statistical model used was a three-factor analysis of variance with one between subjects factor , and two within subject factors (treatment and dose). The Mixed procedure also allows us to model for heterogeneous variances across doses. Pairwise comparisons were made between groups for fixed treatment and dose levels using Least Squares Means. Nonparametric statistical methods were used to perform a series of tests on dose-response data in the cases where examination of residual plots from a three-factor analysis of variance model indicated that the assumption of normality of the error terms was suspect. Specifically, group differences at each dose-treatment combination were looked at with the Wilcoxon rank sum test. Treatment differences at each dose-group combination were also examined with the Wilcoxon signed-rank test on the differences. A false discovery rate adjustment was used for multiple tests in view of the large number of tests considered. Differences with false discovery rate-adjusted P-values of P<0.05 were considered significant.All values are means ± SE. Differences among groups regarding ring characteristics, half-maximal effective concentration (ECP<0.01). By the end of the training period, animals in the Occl-Ex group could run up to a higher intensity of 6–7 mph, as compared to only approximately 5mph for the Occl-Sed group during the weekly graded treadmill running test. In addition, Occl-Ex pigs showed a significantly lower peak blood pressure during the stress test than the Occl-Sed pigs (121±4 mmHg and 135±7 mmHg respectively (P<0.01). The peak heart rates were not significantly different between groups .All Occl-Ex pigs completed the training program. The body weights were Non-Occl-Con, 27.5±0.74 kg; Occl-Ex, 39.4±2.2 kg; and Occl-Sed, 42.9±4.21kg. Although we included age-matched controls, the Non-Occl-Con were significantly smaller than both, Occl-Ex and Occl-Sed, (P<0.05) of the rings from the Non-Occl-Con pigs.Structural and functional characteristics of rings harvested from LAD and LCX are presented in . There wP<0.001) in intact rings from both Occl-Ex and Occl-Sed pigs than the Non-Occl-Con pigs (P<0.05) by approximately a log-dose (P<0.001) in a dose-dependent manner, compared to the Occl-Sed group. BK-induced relaxation was partially inhibited by the addition of L-NAME in all three groups (P<0.001) (P<0.001), suggesting the presence of a prostanoid constrictor in the occluded vessels (P<0.001), indicating the presence of a COX-sensitive vasoconstrictor even in the absence of occlusion. With the combination of inhibitors, L-Name and Indo, BK-induced relaxation was reduced in all three groups (P<0.001), with a marked reduction in the Non-Occl-Con group such that the values were now below those of the Occl-Sed and Occl-Ex groups  ; while t vessels . HoweverP<0.001; . BK-indue groups . Finallye groups .P<0.001), and exhibited a greater EC50 (P<0.001) from the deficit of the Occl-Sed group. ADP-induced relaxation was in part inhibited by L-NAME in all groups (P<0.001) that of the Non-Occl-Con group with the relaxation curve of the Occl-Ex intermediate (P<0.05) in a dose-dependent manner (P<0.001), in a dose-dependent manner, when Indo was added to the bath (P<0.001) and the Occl-Ex group greater than the Occl-Sed group (P<0.05) in a dose-dependent manner. Consistently, the relative sensitivities to ADP, as evident in EC50  and increasing values in the Occl-Sed group (P<0.05) with dose-group interactions (P<0.001). It should be recognized that the ADP-induced relaxation remained rather robust for all three groups , indicating that pathways different from NOS and COX are important for vasorelaxation. In contrast, ADP-induced relaxation was markedly inhibited by L-NAME + Indo + KCl  remained. This implies that some non-NOS pathway contributes to the coronary dysfunction caused by occlusion of the femoral arteries. Further, the lack of differences in the SNP-induced responses among groups indicates that the responsiveness of the smooth muscle to NO is not an important factor underpinning the reductions in vessel relaxation observed with intact vessels from the occluded groups. Second, significant improvements in BK- & ADP-induced relaxation became evident with inhibition of the COX pathways with Indo  contribute to vasorelaxation in coronary arteries. Indeed, it has been reported that EDHF-mediated responses play an important role in conduit arteries for endothelium-dependent relaxation, especially the coronary vascular bed. The observation that BK-stimulated dilatory responses of the occluded groups were actually greater (2Y1 receptors (“purinoceptors”) located on vascular smooth muscle cells of porcine coronary arteries. However, the fact that the dilatory responses were similar in Non-Occl- Con, Occl-Sed, and Occl-Ex animals after denudation , that was evident with both BK and ADP, was eliminated by the inhibition of both the NOS and COX pathways with L-NAME and Indo, respectively. This implies that either or both NOS and COX pathways are altered by exercise training and contribute to the improved vasoresponsiveness by an enhanced NO-dependent vasorelaxation and/or a reduced COX-dependent vasoconstrictor effect. On one hand, the differences between the Occl-Ex and Occl-Sed groups  or in the presence of Indo (leading to an enhanced NOS-dependent vasodilatation). It is unclear why this was not observed with BK    were notion)  may actually be an advantage in the present experiment as it potentially eliminates confounding factors that often present as comorbidities in human PAD patients. Thus, while patients with intermittent claudication can exhibit endothelial dysfunction,6 and an increased inflammatory status with impairment of the coronary arteries it is difficult to assess the effects of PAI as an independent cause of any central vascular dysfunction. In contrast, in the present study we are able to evaluate distinct outcomes of ‘simple’ PAI in relatively young pigs that are free from atherosclerotic/inflammatory disease. Certainly, Yucatan miniature swine are highly resistant to frank atherogenesis. Minimal signs of atherogenesis are only evident after feeding them an exceptionally high-fat diet, representing 46% of their daily caloric intake, for an extended period of time (20–24 weeks)–24. Even then the extent of atherosclerosis in central and peripheral arteries is relatively modest characterized by a Stary Stage I-III classification,24,25, similar to that reported in studies of early human atherosclerosis. Thus, our experiment could expose consequences of PAI that are manifest, even in the absence of extensive comorbidities.Preclinical animal models of PAI, similar to that used in this experiment, have been well characterized and extensively used to investigate potential therapeutic approaches to treat patients with PAD. This appears to be exaggerated in PAD patients, as ischemia induced by exercise promotes oxidative stress and inflammation that has been linked to an attenuation of endothelium-dependent vascular smooth muscle relaxation. Accordingly, we could speculate that the decreased coronary vasorelaxation reported in this study might be the consequence of altered NO availability induced by excessive oxidative stress even with simple cage activity in all of the occluded pigs. However, our results indicate occlusion promulgates changes in signal pathways far more extensive than the NOS. On the other hand, it is apparent that chronic physical activity, similar to that experienced by our trained pigs, exerts an anti-inflammatory effect,38. Elevated biomarkers of chronic inflammation are reduced by exercise training in patients with chronic cardiovascular disease, including PAD,40. If this occurred in our Occl-Ex pigs, it could have contributed to the ameliorated dilatory dysfunction observed in the coronary vessels with exercise training. Second, there is the potential that PAI leads to a hypersympathetic state. Sympathetic nervous activity (SNA) is expected to increase in response to exercise  and is exacerbated during ischemic exercise. In PAD patients there is an exaggerated increase in blood pressure during walkingthat is related to walking speed and duration, as well as the severity of disease,34. This increased blood pressure has been attributed to an enhanced sympathetic output through the ischemic pressor response,42,4. Similar increases in mean arterial blood pressure, attributed to an observed increase in SNA, have been reported in studies using animal models of PAI. An increase in SNA during activity could have contributed to our results, as the associated increase in blood pressure driven by an enhanced SNA is known to be a cardiovascular risk factor leading to a reduced endothelial dependent vasodilatation. Interestingly, it has been reported that exercise training can normalize SNA, alleviate levels of vasoactive substances leading to vasoconstriction, and promote a reduction in systemic blood pressure. Consistent with these observations, our data showed that Occl-Ex pigs exhibited significantly lower peak blood pressure during the stress test than the Occl-Sed pigs .The most important finding of the present study is that coronary artery endothelial dysfunction was observed in other- wise healthy pigs following bilateral occlusion of the femoral arteries. Although the precipitating cause of this dysfunction is presently unclear, a couple of contributing factors can be proposed. First, it is now well recognized that active muscle, even in normal healthy humans, releases an abundance of potent cytokines, including factors potentially involved in the inflammatory process–14 representing approximately 35 to 50% of all patients with PAD,11. Further, exercise training partially ameliorated the dilatory deficit. The potential that muscle- derived cyotkines/factors and/or a hypersympathetic state and their amelioration by training contributed to these responses remains to be determined. To the extent that our findings apply to these human patients, this implies that individuals with PAD are at risk of coronary dysfunction, even in the absence of generalized atherosclerotic disease that affects central vessels.In conclusion, we report here that bilateral occlusion of the femoral arteries, used to establish a preclinical model of PAI, induces coronary endothelial dysfunction, evident as a reduced vasodilatory responsiveness. These observations are potentially most significant in view of the sizeable number of PAD patients that are free from frank CAD


Arabidopsis thaliana. SA can influence activity and/or localization of transcriptional regulators by post-translational modifications of transcription factors and co-regulators. SA-induced redox changes, mediated by thioredoxins and glutaredoxins, modify transcriptional regulators that are involved in suppression of JA-dependent genes, such as NPR1 and TGA transcription factors, which affects their localization or DNA binding activity. Furthermore, SA can mediate sequestering of JA-responsive transcription factors away from their target genes by stalling them in the cytosol or in complexes with repressor proteins in the nucleus. SA also affects JA-induced transcription by inducing degradation of transcription factors with an activating role in JA signaling, as was shown for the ERF transcription factor ORA59. Additionally, SA can induce negative regulators, among which WRKY transcription factors, that can directly or indirectly inhibit JA-responsive gene expression. Finally, at the DNA level, modification of histones by SA-dependent factors can result in repression of JA-responsive genes. These diverse and complex regulatory mechanisms affect important signaling hubs in the integration of hormone signaling networks. Some pathogens have evolved effectors that highjack hormone crosstalk mechanisms for their own good, which are described in this review as well.Transcriptional regulation is a central process in plant immunity. The induction or repression of defense genes is orchestrated by signaling networks that are directed by plant hormones of which salicylic acid (SA) and jasmonic acid (JA) are the major players. Extensive cross-communication between the hormone signaling pathways allows for fine tuning of transcriptional programs, determining resistance to invaders and trade-offs with plant development. Here, we give an overview of how SA can control transcriptional reprogramming of JA-induced genes in  The activation of inducible immune responses in the plant is tightly regulated, ensuring an effective and cost-efficient response to pathogenic microbes and herbivorous insects . RecogniArabidopsis thaliana. Consequently, this review is based primarily on research with Arabidopsis, but we are aware that other plant species may regulate the interplay between hormone signaling pathways differently. We aim to focus on general mechanisms affecting transcriptional regulation that could also apply to other plant species. Hormone-modulated regulation of disease resistance is primarily achieved through effects on gene transcription. Activation or repression of target genes is accomplished by physical interaction between trans-acting proteins, such as transcription factors, and cis-acting DNA elements. Transcription factors and co-regulators can themselves be controlled at the transcriptional level, but they are also subject to post-translational modification through reduction or oxidation, sequestration, phosphorylation, degradation, or interaction with other transcription factors or co-factors . Regulation of NPR1 monomer levels in the nucleus is also dependent on SA. NPR1 and NPR1-homologs NPR3 and NPR4 were described to be SA-receptors . Two branches are distinguished in JA-dependent signaling: (i) MYC2 is the master regulator of the MYC branch, which is co-regulated by JA and ABA, activating downstream marker genes VSP2 and LOX2  is predominantly regulated at the level of gene transcription . First, aos/dde2 . Using tpression . Furtherpression . HoweverThe activation of the immune response in plants is associated with rapid production of reactive oxygen intermediates (ROI) and increased levels of nitric oxide (NO). Redox-sensing small-molecule couples, such as reduced and oxidized glutathione, can limit damage from these redox active molecules. Moreover, these redox sensors transduce changes in ROI and NO levels into posttranslational modifications by reduction or oxidation of cysteine residues of transcriptional regulators, causing changes in transcriptional activity . Redox sS-nitrosylation of cysteine residues via NO donor S-nitrosoglutathione . SA triggers cycles of cellular reduction and oxidation, measurable for example by enhanced total glutathione levels and a higher ratio of reduced to oxidized glutathione after SA treatment   . Other tPDF1.2 expression, i.e., within 30 h after application of SA. In addition, treatment with glutathione synthesis inhibitor BSO blocked SA-mediated antagonism of PDF1.2 expression , where overexpression of OsNPR1 suppressed JA-responsive gene expression and defense against insects. However, when a mutated form of OsNPR1 was overexpressed that was constitutively present in the nucleus, herbivore resistance and expression of a JA-responsive gene were not affected . Moreover, JA-induced PDF1.2 expression is not impaired when GRX480 is overexpressed in the triple mutant tga2/tga5/tga6 background, showing that the function of this GRX in suppression of JA-responses is dependent on these TGA transcription factors  in this phenomenon. GRXs are small ubiquitous redox enzymes that use glutathione to reduce their targets . SA is kignaling . In fact pathway . Their apression . TGA tra factors .Salicylic acid could antagonize JA signaling by preventing accessibility of JA-responsive transcriptional regulators to their target genes. This could be achieved by sequestering transcription factors in inactive complexes or by degradation of positive regulators.ABI4 and ABI5. After ABA treatment, the ABA receptor ABAR interacts with WRKY40, which is then recruited to the cytosol. By this recruitment, binding of WRKY40 to ABA responsive promoters is inhibited and repression of ABA responsive genes is lifted  that can subsequently phosphorylate transcriptional regulators. Phosphorylation of transcription factors influences gene transcription by changing the binding strength to DNA, or affecting sequestration or stability . In partsis gene . It has ne  in promoters. In Arabidopsis, 10 TGAs exist of which several have been shown to interact with NPR1 . In non-induced conditions, suppression of PR1 by TGAs has also been reported . Transcriptional regulation of ORA59 by TGAs is in line with the observation that the GCC-box is enriched in the promoter elements of ACC-induced, SA-suppressed genes. How can TGA factors act as both activators and repressors in different hormone signaling pathways? Possibly, different co-factors can be recruited to TGA factors depending on both the promoter context and the hormonal context. In the case of activation of transcription by SA, TGAs have been shown to interact with transcriptional activators NPR1 and GRAS protein SLC14 . GRXs were shown to down-regulate ORA59 expression in a TGA-dependent manner, as discussed in “Role of Reduction of Transcriptional Regulators in SA Signaling” .Both  of NPR1  or was e of NPR1 . HoweverArabidopsis contain an EAR domain, which is an active repressor domain that interacts with the general co-repressor TPL . Overexpression of SA-induced WRKY70 suppressed MeJA-induced PDF1.2 expression  at the PR1 promoter, which is associated with increased transcription, was enhanced in these plants. Conversely, tri-methylation of H3K27, which is associated with transcriptional silencing, was enriched at the PDF1.2 promoter , suggesting that histone modifications were responsible for the observed increased or decreased transcription  underlying antagonistic effects of SA on JA-mediated transcriptional responses and highlighted several transcriptional regulators  as signal integrators. However, there is still much unknown about hormonal crosstalk mechanisms. The use of whole-transcriptome sequencing techniques after combinatorial hormone treatment or pathogen infection will aid in the identification and characterization of additional transcriptional regulators that can act as nodes of convergence in multiple signaling pathways . CombiniThe authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Stage IV melanoma has historically had a poor prognosis because of lack of responsiveness to traditional chemotherapeutics. The 1-year survival rate of patients diagnosed with stage IV melanoma varies between 33 and 66 %. Depending on the number, the location, and the resectability of distant metastases, common treatment options comprise surgery, systemic medical therapy, and radiotherapy.Single-agent DTIC has been the standard chemotherapy regimen for metastatic melanoma since 1970s. From a recent meta-analysis Luo et al [Patients who have limited sites of metastatic disease, a long disease-free survival and a tumor doubling time higher than 60 days, may be amenable to surgical resection and complete surgical excision of limited metastatic disease can result in prolonged relapse-free survival in carefully selected patients. The potential for complete surgical resection to benefit individual patients is suggested by the results of a prospective, phase II study from the Southwest Oncology Group. In this prospective, multicenter study, 64 of 77 carefully selected patients were able to undergo complete resection of all sites of metastatic disease. Overall, the three and four-year survival rates were 36 and 31 percent, respectively, although late relapses continued to be observed after this time.Ipilimumab is a fully human monoclonal antibody directed against cytotoxic T-lymphocyteassociated antigen-4 (CTLA-4), a negative regulator of T-cell-mediated immune responses. In Phase III trials, ipilimumab treatment significantly extended overall survival (OS) compared with control in both pretreated and treatment-naϊve patients. Approximately 10% of patients present objective responses by standard criteria, whereas 10–20% have stable disease or minor responses, that translate into a clinical benefit. In patients who initially achieve clinical benefit and then relapse months or years later, re-treatment with ipilimumab can be also advantageous.Vemurafenib stands as the first personalized treatment in melanoma based on a specific mutation, with a favorable impact on survival. The overall survival rate at 6 months was 77% and 58% at 12.As a consequence, future improvements of this targeted and personalized approaches are expected from ongoing clinical trials aiming at potentiate the activity of BRAF inhibitors through combination with other molecules. These combination therapies also aim at lowering the observed skin toxicities.


The findings were consistently observed after propensity matching analysis. In conclusion, the risk of mortality was lower for AF patients who were treated according to the antithrombotic recommendations of the 2012 ESC guidelines, guided by the CHA2DS2-VASc score. Better efforts to implement guidelines would lead to improved outcomes for patients with AF.This study compared the risk of mortality in atrial fibrillation (AF) patients treated adherent to the 2012 European Society of Cardiology (ESC) guidelines for stroke prevention and those who were not treated according to guideline recommendations. This study used the Taiwan National Health Insurance Research Database. From 1996 to 2011, 354,649 newly diagnosed AF patients were identified as the study population. Among the study cohort, 45,595 and 309,054 patients were defined as Guideline-Adherent and Non-Adherent groups, respectively. During the follow up of 1,480,280 person-years, 133,552 (37.7%) patients experienced mortality. The risk of mortality was lower among AF patients whose treatment was adherent to the guideline recommendation for stroke prevention than those whose treatment was not  with an adjusted hazard ratio of 0.62  after adjusting for age, gender, CHA Atrial fibrillation (AF) is the most common and prevalent arrhythmia in the worldwide population1357892DS2-VASc  risk score, is recommended and advocated in current guidelines to stratify individual stroke risk in patients with AF142DS2-VASc score of 0 for males and 1 for females) do not require antithrombotic therapy. AF patients with ≥1 additional stroke risk factor beyond gender should be offered OACs for effective stroke prevention.Oral anticoagulants (OACs) significantly reduce the risk of AF-related stroke11Despite the clear recommendations for stroke prevention in the guidelines, OACs are still under-used in real-world practice161719202122The protocol of the present study is similar to our previous studies2526272829Information about important co-morbid conditions of each individual was retrieved from the medical claims based on the International Classification of Diseases-Ninth Revision-Clinical Modification codes. We defined patients with a certain disease only when it was a discharge diagnosis or confirmed more than twice in an outpatient department. The diagnostic accuracies of important co-morbidities, such as hyperlipidemia, heart failure, hypertension, myocardial infarction, diabetes mellitus, and chronic obstructive pulmonary disease, have been validated previously31From January 1, 1996, to December 31, 2011, a total of 354,649 AF patients aged 20 years or older were identified from the NHIRD as the study population. AF was diagnosed using the International Classification of Diseases-Ninth Revision-Clinical Modification code (427.31) registered by the physicians responsible for treatment. To ensure diagnostic accuracy, we defined patients with AF when it was a discharge diagnosis or confirmed more than twice in the outpatient department342DS2-VASc risk score was calculated for each patient by assigning 1 point each for age between 65–74 years; history of hypertension, diabetes mellitus, recent heart failure, and/or vascular disease ; and female sex. Two points each were assigned for a history of stroke, TIA, or age 75 years or olderThe CHA2DS2-VASc score. According to the 2012 ESC AF guidelines, Guideline-adherent was defined as the following: (1) For AF males with a CHA2DS2-VASc score of 0 and AF females with a score of 1 (for gender), no anti-thrombotic drug was necessary, (2) For AF patients with an additional risk factor of the CHA2DS2-VASc scheme in addition to gender, an OAC should be prescribed. Patients who received treatment different from these recommendations were defined as “Non-adherent”. The clinical endpoint was defined as all-cause mortality.Guideline adherence was based on compliance with anti-thrombotic recommendations for AF stroke prevention according to the CHAWe performed propensity score–matched analyses for patients adherent or non-adherent to ESC guideline. We calculated propensity scores for the likelihoods of adherence to guideline compared to non-adherence by multivariate logistic regression analyses, conditional on all baseline covariates listed in t test was used for the analysis of continuous variables, and the differences between nominal variables were compared by the chi-square test. The incidence rate of all-cause mortality was calculated by dividing the number of events by person-time at risk with the 95% confidence interval (CI) estimated by using Fisher’s exact test. The risk of all-cause mortality was assessed using Cox regression analysis. Statistical significance was set at p < 0.05, and all statistical analyses were carried out with SPSS 17.0 .Data were presented as the mean ± standard deviation (SD) for continuous variables and proportions for categorical variables. An unpaired 2-tailed Student 2DS2-VASc score is shown in 2DS2-VASc score.2DS2-VASc score compared to Non-adherent patients. A total of 43.8% of patients in the ESC guideline Non-adherent group received anti-platelet agents for stroke prevention.During the follow up of 1,480,280 person-years, the primary endpoint of mortality occurred in 133,552 (37.7%) patients. The risk of mortality was lower in Guideline-adherent AF patients than in Non-adherent AF patients , . The cum2DS2-VASc score, baseline differences and the use of antiplatelet therapy  than other ethnicitiesThe guideline adherence rate (13%) we observed in the present study is lower than in other observational studies, which cumulatively report an adherence rate between 53–75%2324372 score for stroke risk stratification232DS2-VASc-based guidelines are limited. Recently, Lip et al. reported from the EORP-AF Pilot General Registryet al., the use of warfarin not only reduced the risk of stroke by 64%, it also reduced the risk of all-cause mortality by about 25%Several previous studies have investigated the impact of guideline adherence on the clinical outcomes of systemic thromboembolism (TE) and bleeding232DS2-VASc score. A common argument to withhold OAC in the elderly is the risk of major bleeding, particularly ICH. Certain patients with multiple comorbidities have a higher HAS-BLED  score, and an increased risk of major bleeding with OAC use2DS2-VASc score, reserving the HAS-BLED score to assist clinicians in identifying correctable risk factors for bleedingOlder age and multiple comorbidities likely move physicians to discard OAC use precluding compliance with guideline recommendations. Similar to other observational studies414243This study was an observational study and has several limitations. First, patients were categorized into guideline-adherent or non-adherent groups according to the baseline antithrombotic treatments the patients received. The strategies for AF stroke prevention could be changed during the follow-up. We could not ascertain the drug adherence and the reason for non-adherence during the follow-up. However, at the timing when patients experienced mortality or at the end of follow up, more than 80% of patients were still categorized as the same groups as they were at the baseline. Furthermore, since patients who did not receive antithrombotic agents at enrollment were allowed to take OACs during follow-up according to the guideline recommendation and were not excluded from the analysis, the true risk of mortality for patients in the Non-adherent group could be even higher than what we reported here. Therefore, the true difference in mortality risk between these 2 groups could potentially be even larger. Second, the baseline characteristics were different between the 2 groups, and therefore we were not able to exclude the possibility that Non-adherent patients were older and sicker, which may have led to the observed increase in mortality. We adjusted for these potential confounders using multivariate Cox regression and propensity matching analyses. Third, we were not able to calculate the HAS-BLED  score for each patient because details on the amount of alcohol intake and biochemical indices of renal/liver function were not available in this registry database. The concern of severe bleeding for patients with a high HAS-BLED score may prohibit physicians from prescribing OACs, and a further study is necessary to explore the reasons why OACs were not used. Lastly, the present study only Taiwanese patients, and the suitability of result extrapolation to other populations remains uncertain.2DS2-VASc score. Better efforts to implement guidelines would lead to improved outcomes for patients with AF.In this nationwide AF cohort, the risk of mortality was lower for patients who were treated according to antithrombotic recommendations of the 2012 ESC AF guidelines guided by the CHAHow to cite this article: Li, C.-H. et al. European Society of Cardiology Guideline-Adherent Antithrombotic Treatment and Risk of Mortality in Asian Patients with Atrial Fibrillation. Sci. Rep.6, 30734; doi: 10.1038/srep30734 (2016).


Objective: Adipose-derived stem cells secrete various growth factors that promote hair growth. This study examined the effects of adipose-derived stem cell-conditioned medium on alopecia. Methods: Adipose-derived stem cell-conditioned medium was intradermally injected in 22 patients (11 men and 11 women) with alopecia. Patients received treatment every 3 to 5 weeks for a total of 6 sessions. Hair numbers were counted using trichograms before and after treatment. A half-side comparison study was also performed in 10 patients (8 men and 2 women). Results: Hair numbers were significantly increased after treatment in both male (including those without finasteride administration) and female patients. In the half-side comparison study, the increase in hair numbers was significantly higher on the treatment side than on the placebo side. Conclusion: Treatment using adipose-derived stem cell-conditioned medium appears highly effective for alopecia and may represent a new therapy for hair regeneration. Conservative treatments for alopecia include oral administration of finasteride and topical administration of minoxidil. However, these conventional therapies are ineffective in some patients. Furthermore, finasteride administration is indicated only for androgenic alopecia and is therefore applied only in male patients. New therapies that are both more effective and able to be used in female patients are required.,4,6,7Adipose-derived stem cells are multipotent cells that have shown potential for regenerative medicine.2In this study, to obtain more objective data, we examined changes in hair numbers as measured using trichograms and evaluated the effects of adipose-derived stem cell-conditioned medium on hair regeneration.The study followed principles in the Declaration of Helsinki. Twenty-two patients  with alopecia were treated with adipose-derived stem cell-conditioned medium and examined with trichograms both before starting and after completing treatment. In the other group of 10 patients , a half-side comparison study was performed as described later. Informed consent was obtained from all patients prior to enrolment in the study.,2 of solution. A total volume of 3 to 4 mL was injected during each session of treatment. Patients received treatment every 3 to 5 weeks for a total of 6 sessions. Finasteride was also administered to 6 male patients, but no female patients, during the study.We used a commercial product containing protein solution from adipose-derived stem cell-conditioned medium , as previously described.62) centered on the tattoo was counted by 2 technicians blind to the patients, and the average number was recorded as data. Trichograms were made before starting treatment and at 1 to 3 months after the final treatment .The intersection of a line extending cranially from the lateral angle of an eye (left or right) and a line connecting both ears coronally was marked on the scalp with tattoo. A circle 2 cm in diameter centered on this mark was shaved. TrichoScope images were recorded using a PowerShot 450 digital camera . The number of hairs within a circle of 11 mm in diameter  received adipose-derived stem cell-conditioned medium treatment on the left side and placebo treatment  on the right. Treatment was again performed every 3 to 5 weeks for a total of 6 sessions. Both sides of the scalp were examined using trichograms before starting treatment and after treatment. None of the patients were administered finasteride during this study.P < .05 were considered statistically significant.Data are expressed as mean ± standard error of the mean. Data before and after treatment, or data from both sides in the half-side comparison, were compared using the Wilcoxon signed rank test. Values of Numbers of hairs in the same area of the same patient were accurately counted using trichograms before and after treatment . The numIn the half-side comparison study, the number of hairs was significantly increased after treatment on both the left (adipose-derived stem cell-conditioned medium) and right (placebo) sides . HoweverThis study used trichograms to objectively show the positive effects of adipose-derived stem cell-conditioned medium on alopecia. To obtain accurate data, shaving and marking the area for the trichogram is important. We believe that trichograms, particularly as used in this study, offer a reliable and useful method for assessing the treatment of alopecia, although the method is semi-invasive for patients.Treatment with adipose-derived stem cell-conditioned medium was effective, even in patients who did not take finasteride. We believe that this treatment may offer an alternative to finasteride administration and thus appears particularly useful for female patients. Although no significant difference in improvement of alopecia was seen between male patients treated using adipose-derived stem cell-conditioned medium with and without finasteride administration, we got the impression that the combination of adipose-derived stem cell-conditioned medium and finasteride is preferable to single treatment. The combination therapy was effective in our previous study as well.,11,Adipose-derived stem cell-conditioned medium is rich in growth factors such as vascular endothelial growth factor, hepatocyte growth factor, platelet-derived growth factor, and insulin-like growth factor 1.2In the half-side comparison study, the finding that the number of hairs increased even on the right (placebo) side was somewhat surprising. This result might suggest that the injection itself activates hair growth via tissue injury, or that adipose-derived stem cell-conditioned medium injected on one side can affect the other side, perhaps through the local circulation.Contraindications for adipose-derived stem cell-conditioned medium treatment are similar to those for general mesotherapy, including local skin disease, inflammation, infection, allergic disease, autoimmune disease, pregnancy, cancer, and current anticoagulant therapy. The most common complication is pain during and after injection, but this can be prevented or managed with supraorbital nerve block, occipital nerve block, local anesthesia, cooling, or administration of nonsteroidal anti-inflammatory drugs.In conclusion, treatment using adipose-derived stem cell-conditioned medium appears highly effective for alopecia and may represent a new avenue of therapy for hair regeneration. More details about adipose-derived stem cell-conditioned medium treatment, such as long-term (over a period of years) effects and histological changes, should be elucidated in the future.


Schistosoma haematobium, a parasite of major public health importance. Mathematical models of epidemiological trends suggest that antifecundity immunity is dependent on antibody responses to adult-worm-derived antigen.Immunity that reduces worm fecundity and, in turn, reduces morbidity is proposed for Schistosoma-specific antigen that is steadily secreted by adult worms. Fecundity was modeled against host age, infection transmission intensity, and antibody responses specific to soluble worm antigen (SWA), tegument allergen-like 1, and 28-kDa glutathione-S-transferase.For a Malian cohort  residing in high-transmission fishing villages or a moderate-transmission village, worm fecundity was assessed using the ratio of urinary egg excretion to levels of circulating anodic antigen, a Worm fecundity declined steadily until a host age of 11 years. Among children, host age and transmission were negatively associated with worm fecundity. A significant interaction term between host age and transmission indicates that antifecundity immunity develops earlier in high-transmission areas. SWA immunoglobulin G1 (IgG1) levels explained the effect of transmission on antifecundity immunity.Antifecundity immunity, which is likely to be protective against severe morbidity, develops rapidly during childhood. Antifecundity immunity is associated with SWA-IgG1, with higher infection transmission increasing this response at an earlier age, leading to earlier development of antifecundity immunity. Schistosoma cause debilitating chronic infections affecting an estimated 240 million people . Findings were log transformed prior to analysis. Dependence of worm fecundity on host age was examined for improved model fit, using piecewise regression. The fit of the piecewise regression model with the lowest residual standard error was compared with a nonpiecewise regression model by analysis of variance.For individuals aged <11 years, the dependence of worm fecundity on host antibody levels and transmission intensity , including interaction terms between host age and transmission, were explored. Models were reduced by stepwise removal of nonsignificant predictors, and the results are expressed as the geometric mean (GM) ratio.S. mansoni adults will contribute to CAA levels, reducing the apparent S. haematobium egg to adult worm ratio. For cultural reasons, the provision of stool samples was low, resulting in a number of missing data points (70 of 276). Multiple imputation by chained equations, using predictive mean matching and 25 imputations, was used to estimate these missing values . Predictors in the imputation were age, transmission intensity, sex, ln(caa + 1), ln(Sh eggs + 1), ln(SWA-IgG1), ln(SWA-IgG4), ln(SWA-IgE) and ln(SWA-IgA), ln(Sh28GST-IgG1), ln(Sh28GST-IgG3), ln(Sh28GST-IgA), detectable ShTAL1-IgG1, and IgG4 and IgE responses, with complete data sets available for all. In models of worm fecundity for individuals aged <11 years, the reported statistics are the pooled fits from models adjusted for the 25 imputations of S. mansoni infection intensity.The presence of The association between SWA-IgG1–specific responses and host age and transmission intensity were analyzed by linear regression analysis.The GM egg counts and worm burdens, stratified by age, for the participating villages are shown in Figure Supplementary Figure 2). This piecewise regression model had a significantly better fit than a nonpiecewise linear regression model . In the piecewise regression model, for individuals aged ≥11 years, there was no significant relationship between age and worm fecundity , but individuals <11 years of age had both a significantly greater y-axis intercept  and a significantly different slope of the regression line , compared with those aged ≥11 years .When worm fecundity was expressed as a ratio of egg excretion to CAA levels, a nonlinear relationship was observed between host age and worm fecundity Figure . Worm feP < .001). Lower worm fecundity occurred in children residing in the high-transmission villages . An interaction term between host age and transmission was also significant , reflecting the lower gradient of the host age/worm-fecundity slope in the higher-transmission villages. This was caused by the lower intercept on the y-axis for children in these higher-transmission villages .The association between host age and worm fecundity was explored further in individuals <11 years of age, taking into account the sex of the host, the differing transmission intensities, and the interaction between host age and transmission. There was a significant negative relationship between host age and worm fecundity , but neither the interaction term between age and transmission  nor transmission intensity itself  remained significant after the addition of SWA-IgG1 and were removed from the model. Both age  and transmission intensity  were significant predictors of SWA-IgG1. Therefore, the transmission-driven increase in SWA-IgG1 levels explains the earlier decrease in worm fecundity observed in the high transmission villages. Age remained a significant negative predictor of worm fecundity , indicating that other age-dependent mediators of antifecundity immunity exist.IgG1, IgG4, IgE, and IgA responses to SWA were individually entered into univariant linear regression models of worm fecundity for those aged <11 years. All 4 antibody responses were significantly negatively associated with worm fecundity, but IgG1 responses explained the greatest variance in worm fecundity, with the lowest residual standard error (Table P = .097), but transmission itself remained significant .When SWA-IgG1 was replaced by SWA-IgG4, the antibody isotype with the next best fit in univariant analysis of worm fecundity, the interaction term between age and transmission was no longer significant , respectively). Sh28GST-IgA was a significant negative predictor of worm fecundity in a comparable model . However, when added to a model adjusted for age and SWA-IgG1 level, the negative association between Sh28GST-IgA and worm fecundity was no longer significant .Sh28GST-IgG1 and Sh28GST-IgG3 levels were not significantly associated with worm fecundity in models adjusted for age, transmission intensity, and the interaction term between the 2 , but ShTAL1-IgG4 and ShTAL1-IgE responses were significant negative predictors of worm fecundity in comparable models . When added to a model adjusted for age and SWA-IgG1 level, the negative associations between worm fecundity and ShTAL1-IgG4 and ShTAL1-IgE responses were no longer significant  and 0.46 , respectively).ShTAL1-IgG1 responses were not significantly associated with worm fecundity in models adjusted for age, transmission intensity, and the interaction term between the 2 .S. haematobium, developing by 15 years of age, but not for S. mansoni [, for which deviations away from a linear relationship between CAA and egg count have been attributed to crowding effects reducing the fitness of the adult worms [In a previous study, antifecundity immunity was proposed to exist for  mansoni , for whilt worms . It is nChanges in host hormone levels at puberty could possibly explain the observed reduction in egg excretion from the age of 11 years in this study and from the age of 15 years reported in a previous study . HoweverS. haematobium is phylogenetically within a clade of schistosomes infecting ungulates and other primates, and it is so closely related to some members that hybridization occurs [S. mansoni infections. In S. bovis experimental infections, antifecundity immunity can be transferred via serum [S. haematobium infection challenge experiments of patas monkeys [S. bovis and the closely related S. matteei, the ability of sera to inhibit GST function is not related to antibody levels [n occurs , 34. It ia serum , and vacia serum , a findi monkeys . In the  monkeys . IgG3 re monkeys . Howevery levels , indicatS. mansoni cryptic tegument-associated protein, and the pattern of the antibody responses to ShTAL1 are similar to those of SmTAL1, with IgG4 and IgE levels increasing with age and after treatment [S. mansoni homologue, is predominantly expressed in the adult worm . ShTAL1-IgG4 and ShTAL1-IgE responses were negatively associated with worm fecundity, after adjustment for age and transmission, indicating that antigens exposed upon worm death may give rise to antibody responses that mediate antifecundity immunity. However, after adjustment for SWA-IgG1, these relationships were no longer significant.A pattern-oriented mathematical model predicts that antibody to antigens exposed upon worm death mediate antifecundity immunity . Here, wreatment . AdditioSWA is an extract from adult worms that includes numerous antigens, many of which are cryptic during the life of the worm, with some found on the surface or excreted/secreted and others shared/cross-reactive with other life stages of the parasite. Those on the surface or excreted/secreted and those shared by the eggs (because eggs die in the tissues) will be continuously exposed to the host. It was levels of SWA-IgG1 that were explanatory of the observed transmission intensity dependence of antifecundity immunity, indicating that this isotype may play a predominant role. IgG1 is raised against cryptic antigens, as indicted by a boost in levels after treatment induced worm death , but it S. haematobium antifecundity/morbidity vaccine, which would improve the quality of life for millions of individuals.To conclude, a reduction in worm fecundity was observed during childhood and developed earlier in higher-transmission villages. The IgG1 responses to worm antigen were explanatory of this transmission dependence, implicating this antibody isotype in antifecundity immunity. The specific target of this response remains to be clarified, but the results add impetus to research into candidates for a potential The Journal of Infectious Diseases online . Supplementary materials consist of data provided by the author that are published to benefit the reader. The posted materials are not copyedited. The contents of all supplementary data are the sole responsibility of the authors. Questions or messages regarding errors should be addressed to the author.Supplementary materials are available at


N = 121, 83% response) and for student pharmacists who completed both pre- and post-intervention surveys . Of those who completed both pre- and post-intervention surveys, 59% obtained genotyping. Student pharmacists demonstrated a significant increase in their knowledge of pharmacogenomic resources  and confidence in applying pharmacogenomic information to manage patients' drug therapy , particularly if the student had received genotyping. Student pharmacists understanding of the risks and benefits of using personal genome testing services significantly increased  along with agreement that personal genomics would likely play an important role in their future career , particularly among students who participated in genotyping. The educational intervention, including personal genotyping, was feasible, and positively enhanced students' reflections, and attitudes toward pharmacogenomics in a professional pharmacy program.Pharmacogenomics, once hailed as a futuristic approach to pharmacotherapy, has transitioned to clinical implementation. Although logistic and economic limitations to clinical pharmacogenomics are being superseded by external measures such as preemptive genotyping, implementation by clinicians has met resistance, partly due to a lack of education. Pharmacists, with extensive training in pharmacology and pharmacotherapy and accessibility to patients, are ideally suited to champion clinical pharmacogenomics. This study aimed to analyze the outcomes of an innovative pharmacogenomic teaching approach. Second-year student pharmacists enrolled in a required, 15-week pharmaceutical care lab course in 2015 completed educational activities including lectures and small group work focusing on practical pharmacogenomics. Reflecting the current landscape of direct-to-consumer (DTC) genomic testing, students were offered 23andMe genotyping. Students completed surveys regarding their attitudes and confidence on pharmacogenomics prior to and following the educational intervention. Paired pre- and post-intervention responses were analyzed with McNemar's test for binary comparisons and the Wilcoxon signed-rank test for Likert items. Responses between genotyped and non-genotyped students were analyzed with Fisher's exact test for binary comparisons and the Mann-Whitney U-test for Likert items. Responses were analyzed for all student pharmacists who voluntarily completed the pre-intervention survey ( The political, social, and economic landscape of personalized medicine, particularly pharmacogenomics, is in the midst of a transformation. In early 2015, plans for the landmark Precision Medicine Initiative (PMI) were established with the goal to advance biomedical precision medicine research and broaden personalized treatment. This initiative included a $215 million investment in the National Institutes of Health (NIH), the Food and Drug Administration (FDA), and the National Coordinator for Health Information Technology  personal genotyping and reduced cost for sequencing have brought pharmacogenomics to the forefront of regulatory and public attention. Along these lines, the FDA has increased regulation of personalized genome testing companies. Marketing approval for several DTC personal genome tests, classified as medical devices by the FDA, have been required to substantiate safety and efficacy claims and mitigate risk Lancet, . These rhttp://www.fda.gov/Drugs/ScienceResearch/ResearchAreas/Pharmacogenetics/ucm083378.htm, US Food and Drug Administration). Additionally, guidelines exist via groups, namely the Pharmacogenomics Knowledgebase (PharmGKB at https://www.pharmgkb.org/index.jsp) and the Clinical Pharmacogenomics Implementation Consortium (CPIC at https://cpicpgx.org/), tasked with determining the clinical impact of gene-variant-drug pairs in the medical community  at Chapel Hill. The cohort included students on both the Chapel Hill and Asheville campuses who were enrolled in a required 15-week course titled Pharmacy 404L: Pharmaceutical Care Lab during the Spring 2015 semester. One hundred forty-five students were enrolled in the course, and participation in this study was voluntary. This study was determined to be exempt from review by the UNC Institutional Review Board and was conducted in accordance to Good Clinical Practice, International Conference of Harmonization guidelines, and all applicable state and federal laws.Pharmaceutical Care Lab consisted of weekly, 1-h large group class sessions for all 145 students and weekly, 4-h small group sessions of 8–10 students led by a pharmacy post-graduate resident. On week 8 of the course, during a 1-h large group lecture, student pharmacists completed a pre-intervention survey, participated in an introductory lecture on pharmacogenomics, and received information regarding voluntary and anonymous personal genomic testing via 23andMe . The large group lecture included the following student learning objectives: define pharmacogenomics, discuss the importance of pharmacogenomics in drug therapy, examine how pharmacogenomics is used to manage drug therapy, and provide examples of pharmacogenomic-guided algorithms. On week 11 of the course, pharmacogenomic cases, and relevant clinical resources were discussed in small group sessions. Pharmacy residents facilitated case discussions and were provided with an instructor's guide. A final wrap-up large group lecture was provided on week 15 with the following student learning objectives: discuss 23andMe results, demonstrate how to obtain pertinent pharmacogenomic information and utilize online resources, and review a clinical case focusing on the use of pharmacogenomics to manage drug therapy. Overall, eight different drug-gene pairs were described using clinical cases. Student pharmacists were asked to complete the post-intervention survey immediately following this final lecture. A detailed outline of the educational intervention is presented in Supplementary Table https://www.pharmgkb.org/). The spreadsheet was designed using gene haplotype translation tables from PharmGKB to assign haplotypes to CYP2C19, CYP2C9, CYP3A5, CYP2D6, DPYD, TPMT, G6PD, IFNL3, SLCO1B1, and VKORC1. For each gene, when no changes from the reference haplotype were identified, the formula returned the reference haplotype; if haplotypes could not be determined due to missing or conflicting information, no haplotypes were reported. The limitations of the spreadsheet and the 23andMe genotyping were explained in the informational sheet along with the identified haplotypes. Hyperlinks to CPIC guideline webpages for the various drugs affected by the genes were included in the spreadsheet. Extensive office hours were offered to student pharmacists pre- and post-genotyping. Although not requested, referrals for follow-up health appointments to discuss specific concerns were available.The voluntary 23andMe personal genome test was offered to student pharmacists at a reduced price of $30.00 through funding from the UNC Center for Pharmacogenomics and Individualized Therapy (CPIT). Student pharmacists signed up for genotyping online, and kits were sent directly to their residences from 23andMe. Student pharmacists collected samples and sent them back to 23andMe, after which they received their results ~4–6 weeks later via the 23andMe website. At the time of the completion of this study, 23andMe provided information limited to ancestry and raw data from the Illumina® HumanOmniExpress-24 format chip consisting of 730,525 markers . Student pharmacists received results for 23andMe prior to the conclusion of the course. They were instructed on how to use the 23andMe website and download raw data but were also provided with demo profile information if they opted out of testing. Student pharmacists were referred to third party websites for detailed health information along with precautions and limitations of using various online resources. Student pharmacists extracted personal pharmacogenomic data from the raw 23andMe genotype file using an Excel spreadsheet developed by our lab and interpreted data using the PharmGKB and CPIC guidelines . Student pharmacist responses were linked using an alphanumeric code to preserve anonymity. Paired pre- and post-intervention responses were analyzed with McNemar's test for binary comparisons and the Wilcoxon signed-rank test for Likert items. Responses between genotyped and non-genotyped students were analyzed with Fisher's exact test for binary comparisons and the Mann-Whitney U-test for Likert items. Results were considered statistically significant if N = 121, 83% response rate). The post-intervention survey response rate was 35% (N = 51). We focused our analyses on a linked subset of student pharmacists who completed both the pre- and post-intervention surveys . This comparison subset was further delineated into the genotyped group  and non-genotyped group . Student pharmacists self-described their ethnicity as 73.6% “White or Caucasian (not Hispanic or Latino),” 19.3% “Asian,” 6.6% “Black or African American,” 3.3% “Hispanic or Latino,” and 1% “Native Hawaiian or Pacific Islander.” The majority of student pharmacists reported having obtained a bachelor degree prior to pharmacy school (68.6%) and having taken genetics (60.3%). One student pharmacist completed personal genotyping using 23andMe prior to the implementation of this study and was therefore removed from subsequent analyses comparing pre- and post-intervention groups. There were no statistically significant differences in demographics between the genotyped and non-genotyped groups or between those who completed the pre-intervention survey and students in the linked subset group who completed both the pre- and post-intervention surveys , and none had pharmacogenomic testing performed in a medical setting  of student pharmacists were interested in providing saliva samples to 23andMe for personal genome testing. However, only 69.6% (N = 23) of genotyped and 43.8% (N = 7) of non-genotyped student pharmacists originally affirmed interest in this personal genotyping opportunity prior to the intervention. There was not a significant relationship between student pharmacists' personal experiences with medications and participation in personal genotyping in our study population; a majority of student pharmacists were taking medications (67.8%) and had tried medications that were ineffective (53.7%) or had side effects (76%).In the pre-intervention survey, a majority . There were no additional statistically significant changes in pre- to post-intervention survey responses to the five questions focused on personal reflections and attitudes toward pharmacogenomics.The pre-intervention survey indicated that a majority of student pharmacists were comfortable with the use of personal pharmacogenomic information to guide clinicians in selecting the appropriate medication (73.6%) and dose of medication 72.7%) for themselves . A significant difference was upheld in agreement with this statement in both the genotyped  and non-genotyped groups . Student pharmacists would largely recommend pharmacogenomic testing to manage therapy prospectively on the pre-intervention survey (65.3%), yet no significant difference was found pre- to post-intervention. A significantly larger agreement with this statement occurred in the genotyped group when compared with the non-genotyped group on the post-intervention survey . A minority of student pharmacists were confident in applying pharmacogenomic information to manage patients' drug therapy on the pre-intervention survey (39.7%). However, a significant increase in agreement with this statement was present within the pre- to post-intervention linked subset survey group  and particularly with the student pharmacists who were genotyped . Most students agreed that pharmacogenomic information should be stored in the patient's medical record on the pre-intervention survey (85.1%). However, a decrease in agreement with this statement occurred from the pre- to post-intervention linked subset surveys . Student pharmacists largely agreed that pharmacogenomics would likely play an important role in their future careers (77.7%).Overall, student pharmacists agreed that pharmacogenomics was useful in managing drug therapy in the pre-intervention survey  should have access to the results of pharmacogenomic tests. From the pre- to post-intervention linked subset surveys, similar percentages of student pharmacists agreed that patients (98 vs. 93%), healthcare providers (100 vs. 95%), and health insurance companies (18 vs. 23%) should have access. On the pre-intervention survey, few student pharmacists agreed that life insurance companies (10.7%) and family members (0.8%) should have access.On the post-intervention survey, genotyped and non-genotyped student pharmacists were asked if they could explain the rationale for pharmacogenomic testing (82.6 vs. 62.5%), identify therapeutic areas in which pharmacogenomic testing is required (65.2 vs. 56.3%) or recommended (78.3 vs. 56.3%), discuss the risks and benefits of pharmacogenomic testing (73.9 vs. 56.3%), and interpret the results of pharmacogenomic testing from patients , particularly in the non-genotyped group . Again, a slight majority of student pharmacists indicated that they knew enough about genetics to understand personal genome results in the pre-intervention group (51.2%). Student pharmacists in the pre-intervention group largely agreed that personal genomics would likely play an important role in their future careers (61.2%). A significant increase in student pharmacist agreement occurred with this prompt between the pre- and post-intervention surveys in the linked subset group . This significant increase was upheld between the pre- and post-intervention surveys of the linked subset for the genotyped group  and between the genotyped and non-genotyped post-intervention groups . In the pre-intervention survey, few student pharmacists agreed that physicians and pharmacists have enough knowledge to help individuals interpret the results of their personal genome tests . Few student pharmacists agreed that most people could accurately interpret their personal genome test results on the pre-intervention survey (4.1%), which did not significantly change in the post-intervention survey. Few student pharmacists agreed that personal genome testing companies provide an accurate analysis and interpretation of genotype data on the pre-intervention survey (19.8%). The majority of student pharmacists in the pre-intervention subset agreed that personal genome testing companies should be regulated by the federal government .A slight majority of students agreed with understanding the risks and benefits of using personal genome testing services in the pre-intervention group  in the post-intervention survey group indicated that they had provided saliva samples to 23andMe for personal genome testing. The most common reasons for undergoing personal genotyping included satisfying general curiosity about their genetic makeup (96%), learning about their ancestry (83%), gaining access to their raw genetic data for further analysis (74%), and price (52%). Reasons for not undergoing personal genotyping in the post-intervention survey group included not being curious about their genetic makeup (35%), not wanting to learn about their ancestry (18%), not wanting access to raw genetic data that could be used to inform them of future risks (35%), and price (53%). Of the 16 non-genotyped student pharmacists in the post-intervention survey group, 35% of students responded “yes” and 41% responded “maybe” when asked if they wished that they had undergone personal genotyping as a part of Pharmaceutical Care Lab.An additional set of prompts was presented to those student pharmacists who elected to undergo personal genome testing through this course . Overall, student pharmacists were pleased with their decision to undergo personal genome testing (87%). Few experienced anxiety when deciding to undergo personal genome testing (13%) or when waiting for their results (4.3%), and none experienced anxiety after receiving personal genome testing results. Most student pharmacists agreed that the opportunity to ask a healthcare professional for help in interpreting the results is an important component to a personal genome testing offer (82.6%).The landscape of clinical pharmacogenomics is rapidly changing. Pharmacists have indicated a desire to increase pharmacogenomics-related responsibilities, including advising clinicians and patients on pharmacogenomics-guided therapeutic selection and adjustment (Owusu-Obeng et al., Several educational initiatives have been proposed for fulfilling pharmacist competencies in pharmacogenomics, ranging from more traditional approaches, such as increased didactic coursework and case studies, to more innovative learning methods, such as massive open online courses (PharmGENEd, Ma et al., Increased availability and visibility of genomics to the general public via DTC advertising may become the ultimate driver of mainstream pharmacogenomics (Chua and Kennedy, In this study, the survey instrument was adapted from the tools developed by Salari et al. and Ormond et al., both of which were administered to medical and graduate students (Ormond et al., Additional prompts were added due to the student pharmacist population surveyed in this study. Genotyping positively influenced confidence in applying pharmacogenomics information to manage patients' drug therapy and agreement that personal genomics would likely play an important role in students' future careers. Interestingly, perhaps due to knowledge of current limitations in pharmacogenomics, a significantly smaller percentage of students in the linked subset from pre- to post-intervention surveys agreed that pharmacogenomics information should be stored in the medical record.The survey included an exploratory post-intervention questionnaire set developed from Lee et al., who established a train-the-trainer approach for broad-scale dissemination of pharmacogenomics information to students (Lee et al., Although this study was conducted at a single institution with students in a specific professional curriculum, it contributes to our understanding of how educational interventions can impact student understanding of personalized genotyping in the health professions. One limitation of this survey was a low post-intervention survey response rate, which could have resulted in selection bias. Despite this low response rate, we were able to glean important information about this intervention. To protect anonymity, surveys were not linked to grades, making it difficult to determine our impact on student confidence beyond the student self-reported data collected. Also, the timing of the post-intervention survey was not ideal, as the survey was released at the end of the semester. However, it is a valuable endeavor to continue optimizing this intervention for 2016, and it is anticipated that this educational intervention and the offer for personal pharmacogenomic testing will become a staple within the curriculum. Overall, the educational intervention, including personal genotyping, within this study was feasible and positively enhanced reflections and attitudes toward pharmacogenomics in a professional capacity.AF, CSB, KS, CLB, and TW designed and implemented the educational intervention. AF, CSB, JM, and NW created and modified the survey instrument. AF and JM analyzed the data, and OS created the pharmacogenomic tool for students to analyze their data. All authors contributed to the writing and editing of this manuscript.The University of North Carolina Center for Pharmacogenomics and Individualized Therapy subsidized the cost of 23andMe kits for student pharmacists.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.