Unpacking the Role of Extracellular Vesicles in Ischemic and Hemorrhagic Stroke: Pathophysiology and Therapeutic Implications.

Stroke is a leading cause of death and disability worldwide. Inflammation and microvascular dysfunction have been associated with brain injury and long-term disability after both ischemic and hemorrhagic stroke. Recent studies have suggested a potential role of extracellular vesicles (EVs) as a link underlying these pathogenic processes. EVs are cell-derived particles enveloped by a lipid bilayer, containing proteins, lipids, and nucleic acids. From a functional standpoint, EVs can facilitate intercellular communication, including across the blood-brain barrier (BBB). Recent advances in EV research have shown a preferential release of EVs from specific cell types in the context of stroke, some of which were associated with increased neuroinflammation, microvascular dysfunction, and neuronal cytotoxicity while others offered a degree of neuroprotection. However, one historic challenge in the studies of EVs in stroke is the lack of consistent definitions and methods to analyze EVs, only recently updated in the MISEV2018 guidelines. Given limitations and complexity in the treatment of stroke, particularly delivery of therapeutics across the BBB, increasing attention has been paid towards manipulating EVs as one vehicle that can permit targeted therapeutic delivery to the central nervous system. These discoveries point towards a future where a better understanding of EVs will advance our knowledge of stroke-associated mechanisms of cerebral and systemic injury and contribute to the development of novel treatments. Here, we review the role that EVs play in ischemic and hemorrhagic stroke.

Bone marrow mesenchymal stromal cells in a 3D system produce higher concentration of extracellular vesicles (EVs) with increased complexity and enhanced neuronal growth properties.

PURPOSE: Extracellular vesicles (EVs) derived from mesenchymal stromal cells (MSCs) have been demonstrated to possess great potential in preclinical models. An efficient biomanufacturing platform is necessary for scale up production for clinical therapeutic applications. The aim of this study is to investigate the potential differences in neuro-regenerative properties of MSC-derived EVs generated in 2D versus 3D culture systems. METHOD: Human bone marrow MSCs (BM-MSCs) were cultured in 2D monolayer and 3D bioreactor systems. EVs were isolated using ultracentrifugation followed by size and concentration measurements utilizing dynamic light scattering (NanoSight) and by fluorescence staining (ExoView). Mouse trigeminal ganglia (TG) neurons were isolated from BALB/c mice and cultured in the presence or absence of EVs derived from 2D or 3D culture systems. Neuronal growth and morphology were monitored over 5 days followed by immunostaining for ß3 tubulin. Confocal images were analyzed by Neurolucida software to obtain the density and length of the neurites. RESULTS: The NanoSight tracking analysis revealed a remarkable increase (24-fold change) in the concentration of EVs obtained from the 3D versus 2D culture condition. ExoView analysis showed a significantly higher concentration of CD63, CD81, and CD9 markers in the EVs derived from 3D versus 2D conditions. Furthermore, a notable shift toward a more heterogeneous phenotype was observed in the 3D-derived EVs compared to those from 2D culture systems. EVs derived from both culture conditions remarkably induced neurite growth and elongation after 5 days in culture compared to untreated control. Neurolucida analysis of the immunostaining images (ß3 tubulin) showed a significant increase in neurite length in TG neurons treated with 3D- versus 2D-derived EVs (3301.5 µm vs. 1860.5 µm, P < 0.05). Finally, Sholl analysis demonstrated a significant increase in complexity of the neuronal growth in neurons treated with 3D- versus 2D-derived EVs (P < 0.05). CONCLUSION: This study highlights considerable differences in EVs obtained from different culture microenvironments, which could have implications for their therapeutic effects and potency. The 3D culture system seems to provide a preferred environment that modulates the paracrine function of the cells and the release of a higher number of EVs with enhanced biophysical properties and functions in the context of neurite elongation and growth.

Combinatorial therapy with immunosuppressive, immunomodulatory and tear substitute eyedrops ("Triple Play") in Recalcitrant Immunological Ocular Surface Diseases.

PURPOSE: The current paradigm for therapy of recalcitrant ocular surface diseases (OSD) consists of a sequential, step-up treatment approach. A combinatorial topical therapy (anti-inflammatory/immunosuppressive [steroid] with immunomodulatory [pooled human immune globulin] and tear substitute [serum]) that simultaneously targets several immunological pathways may be more efficacious. This report evaluates if the combinatorial therapy resulted in clinical benefit in patients with recalcitrant OSD. METHODS: We performed a retrospective case study of patients receiving topical, preservative-free, compounded formulations of steroids, pooled human immune globulin, and serum tears. Outcome measures included visual acuity, ocular surface disease index (OSDI), ocular discomfort score, subjective global assessment (SGA), corneal staining, conjunctival redness, and slit lamp photographs. RESULTS: Patients consisted of one male and 11 females ranging in age from 27 to 87 years old. Pathologies included ocular graft-versus-host disease (n = 4), Sjögren's syndrome (n = 3), ocular cicatricial pemphigoid (n = 1), pemphigus vulgaris (n = 1), peripheral ulcerative keratitis (n = 1), Stevens-Johnson syndrome (n = 1), and giant papillary conjunctivitis (n = 1). All patients were "improved" or "much improved" on SGA after combinatorial therapy. There was a clinically meaningful reduction in OSDI, ocular discomfort, corneal staining, and conjunctival injection. Additionally, three patients had improvement in their visual acuity (one from 20/400 to 20/20). Adverse effects included increased intraocular pressure in two patients, presumably due to topical steroid use. CONCLUSIONS: Combinatorial therapy provides clinical benefit by reducing the symptoms and signs in recalcitrant OSD. Our study provides the rationale for performing prospective clinical trials to evaluate the efficacy of combinatorial therapy for treating recalcitrant OSD.

Complicated Neuraxial Anesthesia in a Patient With Tethered Spinal Cord.

Spinal anesthesia is a common anesthetic used in many surgical procedures. Anatomical variations and diseases make this procedure require extra considerations. Extra care should be taken into consideration in the pre-anesthesia evaluation when neuraxial anesthesia is planned. We present the case of a patient undergoing cesarean section for the arrest of labor, who had a complicated labor course due to unrecognized history of a tethered spinal cord leading to inadequate analgesia, requiring replacement of epidural analgesia and spinal anesthesia. Ultimately, the decision was made to proceed with general anesthesia due to the patient's discomfort. Exhaustive chart review after surgery showed a past MRI with evidence of a tethered spinal cord.

Systemic Immune-Inflammation Index Predicts Delayed Cerebral Vasospasm After Aneurysmal Subarachnoid Hemorrhage.

BACKGROUND: Delayed cerebral vasospasm is a feared complication of aneurysmal subarachnoid hemorrhage (SAH). OBJECTIVE: To investigate the relationship of systemic inflammation, measured using the systemic immune-inflammation (SII) index, with delayed angiographic or sonographic vasospasm. We hypothesize that early elevations in SII index serve as an independent predictor of vasospasm. METHODS: We retrospectively reviewed the medical records of 289 SAH patients for angiographic or sonographic evidence of delayed cerebral vasospasm. SII index [(neutrophils × platelets/lymphocytes)/1000] was calculated from laboratory data at admission and dichotomized based on whether or not the patient developed vasospasm. Multivariable logistic regression and receiver operating characteristic (ROC) analysis were performed to determine the ability of SII index to predict the development of vasospasm. RESULTS: A total of 246 patients were included in our study, of which 166 (67.5%) developed angiographic or sonographic evidence of cerebral vasospasm. Admission SII index was elevated for SAH in patients with vasospasm compared to those without (P < .001). In univariate logistic regression, leukocytes, neutrophils, lymphocytes, neutrophil-lymphocyte ratio (NLR), and SII index were associated with vasospasm. After adjustment for age, aneurysm location, diabetes mellitus, hyperlipidemia, and modified Fisher scale, SII index remained an independent predictor of vasospasm (odds ratio 1.386, P = .003). ROC analysis revealed that SII index accurately distinguished between patients who develop vasospasm vs those who do not (area under the curve = 0.767, P < .001). CONCLUSION: Early elevation in SII index can independently predict the development of delayed cerebral vasospasm in aneurysmal SAH.

Inflammasome Caspase-1 Activity is Elevated in Cerebrospinal Fluid After Aneurysmal Subarachnoid Hemorrhage and Predicts Functional Outcome.

BACKGROUND/OBJECTIVE: Subarachnoid hemorrhage (SAH) is a devastating neurological injury, further complicated by few available methods to objectively predict outcomes. With the recent shift in focus to neuroinflammation as a potential cause of adverse outcomes following SAH, we investigated the inflammasome-derived enzyme, caspase-1, as a potential biomarker for poor functional outcome. METHODS: SAH patients were recruited from a regional stroke referral center. Cerebrospinal fluid (CSF) samples from 18 SAH subjects were collected via an external ventricular drain and obtained as close as possible to admission (within 72 h). For control subjects, we collected CSF from 9 patients undergoing lumbar puncture with normal CSF. Caspase-1 activity was measured using commercially available luminescence assays. SAH subjects were categorized at hospital discharge into those with good outcomes (Glasgow Outcome Scale, GOS, of 4-5) and poor outcomes (GOS of 1-3). RESULTS: CSF analysis demonstrated a nearly seven-fold increase in caspase-1 activity in SAH patients compared to controls (p < 0.0001). Within the SAH group, 10 patients (55.6%) had good outcomes and 8 patients (44.4%) had poor outcomes. Mean caspase-1 activity in the poor outcome group was approximately three-times higher than the good outcome group (p = 0.001). Caspase-1 activity was significantly correlated with GOS score (r = - 0.705, p = 0.001). Receiver operating characteristic curve analysis showed that caspase-1 activity can accurately differentiate between patients with good versus poor functional outcome (area under the curve 0.944, p = 0.002). CONCLUSIONS: Inflammasome-derived caspase-1 activity is elevated in the CSF of SAH patients compared to controls and higher levels correlate with worse functional outcome.

Impact of a Formulation Containing Unusual Polyunsaturated Fatty Acids, Trace Elements, Polyphenols and Plant Sterols on Insulin Resistance and Associated Disturbances.

INTRODUCTION: To evaluate the effect of a lipid-based formulation containing unusual polyunsaturated fatty acids, trace elements, polyphenols and plant sterols on insulin resistance and its associated disturbances among adults at risk of diabetes. METHODS: This was an 8-week, three-arm, open-label randomized clinical trial. We studied individuals aged ≥ 18 years old with diabetes risk given by a body mass index ≥ 25 kg/m2 or a FinnRisc score ≥ 13/20. Participants were randomly assigned to receive: 7 ml sunflower oil (control group), 3.5 ml of the study formulation + 3.5 ml of sunflower oil (low-dose group) or 7 ml of study formulation (high-dose group). RESULTS: We randomized 25 individuals. After one withdrawal in the high-dose group, the study sample comprised nine patients in the control, nine in the low-dose and six in the high-dose groups. The insulin sensitivity increased significantly and in a dose-dependent fashion, up to 10% in the high-dose group. At week 8 the low-dose group exhibited lower glycemic excursions during the oral glucose tolerance test (OGTT), especially 1 h after the glucose challenge (32 mg/dl or 23% lower vs. control group). The incremental area under the glucose curve in the OGTT was 17.1% lower in the low-dose group vs. the control group. Waist circumference increased in the control group, remained constant in the low-dose group and decreased in the high-dose group. C-reactive protein decreased in both formulation groups, up to 50% in the high-dose group. Participants in the formulation groups exhibited increased secretion of GLP-1 and plasma irisin at week 8 vs. the control group. CONCLUSION: The formulation induced favorable changes in insulin sensitivity, glucose tolerance, abdominal obesity and inflammation. These effects and their durability will need to be assessed in larger studies. TRIAL REGISTRATION: NCT03512665. FUNDING: Team Foods Colombia.

Assessment of Prognostic Value of Left Ventricular Global Longitudinal Strain for Early Prediction of Chemotherapy-Induced Cardiotoxicity: A Systematic Review and Meta-analysis.

Importance: Echocardiographic left ventricular global longitudinal strain (GLS) detects early subclinical ventricular dysfunction and can be used in patients receiving potentially cardiotoxic chemotherapy. A meta-analysis of the prognostic value of GLS for cancer therapy-related cardiac dysfunction (CTRCD) has not been performed, to our knowledge. Objective: To explore the prognostic value of GLS for the prediction of CTRCD. Data Sources: Systematic search of the MEDLINE, Embase, Scopus, and the Cochrane Library databases from database inception to June 1, 2018. Study Selection: Cohort studies assessing the prognostic or discriminatory performance of GLS before or during chemotherapy for subsequent CTRCD. Data Extraction and Synthesis: Random-effects meta-analysis and hierarchical summary receiver operating characteristic curves (HSROCs) were used to summarize the prognostic and discriminatory performance of different GLS indices. Publication bias was assessed using the Egger test, and meta-regression was performed to assess sources of heterogeneity. Main Outcomes and Measures: The primary outcome was CTRCD, defined as a clinically significant change in left ventricular ejection fraction with or without new-onset heart failure symptoms. Results: Analysis included 21 studies comprising 1782 patients with cancer, including breast cancer, hematologic malignancies, or sarcomas, treated with anthracyclines with or without trastuzumab. The incidence of CTRCD ranged from 9.3% to 43.8% over a mean follow-up of 4.2 to 23.0 months (pooled incidence, 21.0%). For active treatment absolute GLS (9 studies), the high-risk cutoff values ranged from -21.0% to -13.8%, with worse GLS associated with a higher CTRCD risk (odds ratio, 12.27; 95% CI, 7.73-19.47; area under the HSROC, 0.86; 95% CI, 0.83-0.89). For relative changes vs a baseline value (9 studies), cutoff values ranged from 2.3% to 15.9%, with a greater decrease linked to a 16-fold higher risk of CTRCD (odds ratio, 15.82; 95% CI, 5.84-42.85; area under the HSROC, 0.86; 95% CI, 0.83-0.89). Both indices showed significant publication bias. Meta-regression identified differences in sample size and CTRCD definition but not GLS cutoff value as significant sources of interstudy heterogeneity. Conclusions and Relevance: In this meta-analysis, measurement of GLS after initiation of potentially cardiotoxic chemotherapy with anthracyclines with or without trastuzumab had good prognostic performance for subsequent CTRCD. However, risk of bias in the original studies, publication bias, and limited data on the incremental value of GLS and its optimal cutoff values highlight the need for larger prospective multicenter studies.

A comparative study of different methods for automatic identification of clopidogrel-induced bleedings in electronic health records.

Electronic health records (EHRs) linked with biobanks have been recognized as valuable data sources for pharmacogenomic studies, which require identification of patients with certain adverse drug reactions (ADRs) from a large population. Since manual chart review is costly and time-consuming, automatic methods to accurately identify patients with ADRs have been called for. In this study, we developed and compared different informatics approaches to identify ADRs from EHRs, using clopidogrel-induced bleeding as our case study. Three different types of methods were investigated: 1) rule-based methods; 2) machine learning-based methods; and 3) scoring function-based methods. Our results show that both machine learning and scoring methods are effective and the scoring method can achieve a high precision with a reasonable recall. We also analyzed the contributions of different types of features and found that the temporality information between clopidogrel and bleeding events, as well as textual evidence from physicians' assertion of the adverse events are helpful. We believe that our findings are valuable in advancing EHR-based pharmacogenomic studies.

Plasma hepatocyte growth factor is a novel marker of AL cardiac amyloidosis.

BACKGROUND: Cardiac amyloidosis is an infiltrative cardiomyopathy that is challenging to diagnose. We hypothesized that the novel biomarkers hepatocyte growth factor (HGF), galectin-3 (GAL-3), interleukin-6 (IL-6), and vascular endothelial growth factor (VEGF) would be elevated in cardiac amyloidosis and may be able to discriminate from non-cardiac systemic amyloidosis or other cardiomyopathies with similar clinical or morphologic characteristics. METHODS: Patients were selected from the Vanderbilt Main Heart Registry according to the following groups: (1) amyloid light-chain (AL) cardiac amyloidosis (n = 26); (2) transthyretin (ATTR) cardiac amyloidosis (n = 7); (3) left ventricular hypertrophy (LVH) (n = 45); (4) systolic heart failure (n = 42); and (5) non-cardiac systemic amyloidosis (n = 7). Biomarkers were measured in stored plasma samples. Biomarkers' discrimination performance in predicting AL cardiac amyloidosis (i.e., Concordance index) was reported. A survival analysis was used to explore the relationship between HGF levels and mortality among AL cardiac amyloidosis patients. RESULTS: HGF levels were markedly elevated in patients with AL cardiac amyloidosis (median = 622, interquartile range (IQR): 299-1228 pg/mL) compared with the other groups, including those with non-cardiac systemic amyloidosis (median = 134, IQR: 94-163 pg/mL, p < 0.001). HGF was not a specific marker for ATTR amyloidosis. Gal-3 was elevated in all groups with amyloidosis but could not differentiate between those with and without cardiac involvement. There was no difference in IL-6 or VEGF between those with AL cardiac amyloidosis compared to other groups (p = 0.13 and 0.057, respectively). CONCLUSIONS: HGF may be a specific marker that distinguishes AL cardiac amyloidosis from other cardiomyopathies with similar clinical or morphologic characteristics. Further studies are necessary to determine whether HGF levels predict the likelihood of survival.

Evaluation of the F2R IVS-14A/T PAR1 polymorphism with subsequent cardiovascular events and bleeding in patients who have undergone percutaneous coronary intervention.

Abnormal platelet reactivity is associated with recurrent ischemia and bleeding following percutaneous coronary intervention (PCI). Protease-activated receptor-1 (PAR1), encoded by F2R, is a high affinity thrombin receptor on platelets and the target of the antiplatelet drug vorapaxar. The intronic single nucleotide polymorphism F2R IVS-14 A/T affects PAR1 receptor density and function. We hypothesized that carriers of the T allele, who have been shown to have decreased platelet reactivity, would be at lower risk for thrombotic events, but higher risk for bleeding following PCI. Using BioVU, the Vanderbilt DNA repository linked to the electronic medical record, we studied 660 patients who underwent PCI for unstable or stable coronary artery disease. Primary outcome measures were major adverse cardiovascular events (MACE, composite of revascularization, MI, stroke, death) and bleeding (assessed by Bleeding Academic Research Consortium scale) over 24 months. The minor allele (T) frequency was 14.8 %. There were no genotypic differences in the frequency of MACE (33.7, 28.8, and 31.6 % for A/A, A/T, and T/T respectively, P = 0.50) or bleeding (15.7, 14.7, and 18.8 % for A/A, A/T, and T/T respectively, P = 0.90). In a Cox regression model, fully adjusted for age, race, sex, BMI, and smoking status, carrying a T allele was not associated with MACE (HR 1.19, 95 % CI 0.89-1.59, P = 0.23) or bleeding (HR 0.73, 95 % CI 0.37-1.4, P = 0.34). In conclusion, in our population, F2R IVS-14 PAR1 variability does not affect risk of MACE or bleeding following PCI.

Understanding the role of prostaglandin E2 in regulating human platelet activity in health and disease.

The platelet thrombus is the major pathologic entity in acute coronary syndromes, and antiplatelet agents are a mainstay of therapy. However, individual patient responsiveness to current antiplatelet drugs is variable, and all drugs carry a risk of bleeding. An understanding of the complex role of Prostaglandin E2 (PGE2) in regulating thrombosis offers opportunities for the development of novel individualized antiplatelet treatment. However, deciphering the platelet regulatory function of PGE2 has long been confounded by non-standardized experimental conditions, extrapolation of murine data to humans, and phenotypic differences in PGE2 response. This review synthesizes past and current knowledge about PGE2 effects on platelet biology, presents a rationale for standardization of experimental protocols, and provides insight into a molecular mechanism by which PGE2-activated pathways could be targeted for new personalized antiplatelet therapy to inhibit pathologic thrombosis without affecting hemostasis.

Antithrombotic therapy in patients with acute coronary syndromes: how to make the right choice?

ìAcute coronary syndrome (ACS), a spectrum of clinical scenarios including ST-segment elevation myocardial infarction, non-ST-segment elevation myocardial infarction and unstable angina, is one of the main reasons of hospital admissions and the leading cause of mortality in the developed nations. The underlying pathological substrate in ACS is a rupture of the atherosclerotic plaque leading to either complete or partial thrombosis of the infarct-related vessel. Both activation of platelets and coagulation cascade play a crucial role in thrombus formation. Multiple pharmacological agents have been developed to interfere in this process and to achieve ruptured plaque stabilization. Given the unfavorable impact of bleeding events on clinical outcomes, the development of newer drugs inhibiting platelet aggregation or thrombin formation used for the treatment of patients during acute ACS phase or for the prophylaxis of future ischemic events is targeted on the optimal balance of benefits and risks for each individual. This review provides comprehensive analysis of the current status of antiplatelet and antithrombotic therapy in patients with ACS.

Catecholamine pathway gene variation is associated with norepinephrine and epinephrine concentrations at rest and after exercise.

OBJECTIVE: To examine the hypothesis that genetic variation in enzymes and transporters associated with synthesis, storage, release, and metabolism of catecholamines contributes to the interindividual variability in plasma catecholamine concentrations at rest and after exercise. METHODS: We measured plasma norepinephrine (NE) and epinephrine concentrations at rest and after a standardized exercise protocol in 165 healthy individuals (60% White, 40% African-American) and examined 29 functional or common variants in 14 genes involved in synthesis, transport, or metabolism of catecholamines. We examined the relationship between genotypes and NE concentrations at rest and the increase after exercise (ΔNE) by multiple linear regression with adjustment for covariates [age, race, sex, BMI, fitness, and resting NE (for ΔNE)]. As a secondary outcome, we carried out similar analyses for epinephrine concentrations. RESULTS: There was large interindividual variability in resting NE (mean, 204±102 pg/ml; range, 39-616 pg/ml) and ΔNE (mean, 256±206 pg/ml; range, -97 to 953 pg/ml). Resting NE was significantly associated with variants of four genes: CYB561 (P<0.001), VMAT2 (P=0.016), CHGA (P=0.039), and PNMT (P=0.038). ΔNE after exercise was associated with three variants of PNMT (P=0.041) and COMT (P=0.033 and 0.035), and resting and exercise epinephrine concentrations were associated with two variants each. CONCLUSION: The findings of this exploratory study suggest that variation in catecholamine pathway genes contributes to the interindividual variability in plasma NE and epinephrine concentrations at rest and after exercise.

CYP2A6 genetic variation and dexmedetomidine disposition.

PURPOSE: There is a large interindividual variability in dexmedetomidine dose requirements for sedation of patients in intensive care units (ICU). Cytochrome P450 2A6 (CYP2A6) mediates an important route of dexmedetomidine metabolism, and genetic variation in CYP2A6 affects the clearance of other substrate drugs. We examined whether CYP2A6 genotypes affect dexmedetomidine disposition. METHODS: In 43 critically ill ICU patients receiving dexmedetomidine infusions adjusted to achieve the desired level of sedation, we determined a median of five plasma dexmedetomidine concentrations each. Forty subjects were genotyped for five common CYP2A6 alleles and grouped into normal (n = 33), intermediate (n = 5), and slow metabolizers (n = 2). RESULTS: Using a Bayesian hierarchical nonlinear mixture model, estimated dexmedetomidine clearance was 49.1 L/h (posterior mean; 95% credible interval 41.4-57.6 L/h). There were no significant differences in dexmedetomidine clearance among normal, intermediate, and slow CYP2A6 metabolizer groups. CONCLUSION: Genetic variation in CYP2A6 does not appear to be an important determinant of dexmedetomidine clearance in ICU patients.

The effects of high-frequency jet ventilation (HFJV) on pneumoperitoneum-induced cardiovascular changes during laparoscopic surgery.

BACKGROUND: Standard mechanical ventilation may cause adverse cardiovascular effects in addition to those already related to positive-pressure pneumoperitoneum (PP) during laparoscopic surgery. High-frequency jet ventilation (HFJV) is associated with much less airway pressure, with potentially less influence on venous return, thus potentially it may reduce those effects. The aim of this study was to evaluate the benefits of HFJV to reduce the adverse cardiovascular effects during laparoscopic cholecystectomy. METHODS: We conducted a randomized prospective trial, comparing 12 patients undergoing elective laparoscopic cholecystectomy under conventional mechanical ventilation (control) with 13 similar subjects under HFJV (study). Both groups were categorized as ASA I-II and underwent total intravenous anesthesia. Cardiac functionality was continuously evaluated by analysis of arterial pressure wave changes (Edwards Flo-Trac sensor and Vigileo monitor). RESULTS: There was no significant difference between both groups regarding age, gender, BMI, and general medical condition, as well as hemodynamic stability and blood gases throughout surgery. A significant reduction in cardiac output was noted in the control group under PP during the initiation of anti-Trendelenburg position (from 5.6 to 5.0 l/min, P = 0.049). A reciprocal change was observed regarding stroke volume. Such changes were not noticed under HFJV. Total peripheral resistance was significantly increased during PP, and heart rate was not significantly affected throughout surgery in both groups. Unexpectedly, we did not observe marked adverse hemodynamic changes in the control group during PP without position adjustment. CONCLUSIONS: The use of HFJV in upper laparoscopic surgery can impede the adverse cardiovascular changes that usually occur during induction of PP. We also suggest that the use of total intravenous anesthesia (as used in our study) may also lessen the cardiovascular impairment during PP.

Genetic variations in the α(2A)-adrenoreceptor are associated with blood pressure response to the agonist dexmedetomidine.

BACKGROUND: α(2A)-Adrenoceptors (α(2A)-ARs) have important roles in sympathetic cardiovascular regulation. Variants of ADRA2A affect gene transcription and expression and are associated with insulin release and risk for type 2 diabetes. We examined whether ADRA2A variants are also associated with cardiovascular responses to the selective α(2)-AR-agonist dexmedetomidine. METHODS AND RESULTS: Seventy-three healthy subjects participated in a placebo-controlled, single-blind study. After 3 infusions of placebo, subjects received 3 incremental infusions of dexmedetomidine (cumulative dose, 0.4 µg/kg). Primary outcomes were changes in systolic blood pressure (SBP) and plasma norepinephrine concentrations, measured as difference of the area-under-the-curve during placebo and dexmedetomidine infusions (ΔAUC). We used multiple linear regression analysis to examine the associations between 9 ADRA2A tagging variants and 5 inferred haplotypes and ΔAUC after adjustment for covariates. Homozygous carriers of rs553668 and the corresponding haplotype 4, previously associated with increased α(2A)-AR expression, had a 2.2-fold greater decrease in AUC(SBP) after dexmedetomidine (adjusted P=0.006); similarly, the maximum decrease in SBP was 24.7±8.1 mm Hg compared with 13.6±5.9 mm Hg in carriers of the wild-type allele (P=0.007). Carriers of haplotype 3, previously associated with reduced α(2A)-AR expression, had a 44% smaller decrease in AUC(SBP) (P=0.013). Haplotype information significantly improved the model predicting the decrease in SBP (P<0.001). There were similar but nonsignificant trends for diastolic blood pressure and heart rate. Genotypes were not significantly associated with norepinephrine responses. CONCLUSIONS: Common ADRA2A variants are associated with the hypotensive response to dexmedetomidine. Effects of specific variants/haplotypes in vivo are compatible with their known effects on gene expression in vitro.

Effects of variation in the human alpha2A- and alpha2C-adrenoceptor genes on cognitive tasks and pain perception.

BACKGROUND: The mechanisms underlying interindividual variability in pain perception and cognitive responses are undefined but highly heritable. alpha(2C)- and alpha(2A)-adrenergic receptors regulate noradrenergic activity and are important mediators of pain perception and analgesia. We hypothesized that common genetic variants in these genes, particularly the ADRA2C 322-325 deletion variant, affect pain perception or cognitive responses. METHODS: We studied 73 healthy subjects (37 Caucasians and 36 African-Americans) aged 25.4+/-4.6years. Pain response to a cold pressor test was measured using a 10cm visual analog scale and again on the next day, after three infusions of the selective alpha(2)-agonist dexmedetomidine. Standardized cognitive tests were administered at baseline and after each infusion. The contribution of ADRA2C deletion genotype, dexmedetomidine concentration, and other covariates to pain perception and cognitive responses was determined using multiple linear regression models. Secondary analysis examined the effects of ADRA2A and other ADRA2C variants on pain perception. RESULTS: ADRA2C Del homozygotes had higher pain scores in response to cold at baseline (6.3+/-1.8cm) and after dexmedetomidine (5.6+/-2.2cm) than insertion allele carriers (4.6+/-2.1cm [baseline] and 3.8+/-1.9cm [after dexmedetomidine]; adjusted P-values=0.019 and 0.004, respectively). Cognitive responses were unrelated to ADRA2C Ins/Del genotype. None of the other ADRA2A and ADRA2C variants was significantly related to cold pain sensitivity before dexmedetomidine; after dexmedetomidine, ADRA2A rs1800038 was marginally associated (P=0.03). CONCLUSION: The common ADRA2C del322-325 variant affected pain perception before and after dexmedetomidine but did not affect other cognitive responses, suggesting that it contributes to interindividual variability in pain perception.

GRK5 Gln41Leu polymorphism is not associated with sensitivity to beta(1)-adrenergic blockade in humans.

AIMS: A common, functionally significant polymorphism in GRK5 (Gln41Leu) encodes a gain-of-function enzyme that enhances desensitization of the beta(1)-adrenergic receptor. GRK5 Leu41 has been postulated to confer endogenous 'genetic beta-blockade' and contribute to an attenuated response to beta-blockers in black subjects. The effects of this GRK5 variant on sensitivity to a beta-blocker have not been studied in humans. We hypothesized that the GRK5 Gln41Leu variant contributes to interindividual variability in response to beta-blockade and to the ethnic difference in sensitivity between black and Caucasian individuals. MATERIALS & METHODS: We measured the heart rate at rest and during a graded incremental exercise in 154 healthy subjects (85 white and 69 black) before and after an oral administration of 25 mg atenolol. We determined the genotypes of GRK5 (Gln41Leu), beta(1)-adrenergic receptor (ADRB1 Ser49Gly and Arg389Gly) genotypes and plasma atenolol concentrations. The effects of genotype and covariates on sensitivity to atenolol, measured as the reduction in exercise-induced tachycardia, were determined using multiple regression analyses. RESULTS: The minor allele frequency of GRK5 Leu41 was 32.6% in blacks and 0% in whites. Black individuals were less sensitive to atenolol than white individuals (p < or = 0.011) but this was not explained by the GRK5 genotype. The GRK5 genotype had no effect on resting heart rate before (p = 0.61) and after adjustment for age, sex, ethnicity, atenolol concentrations, BMI and ADRB1 genotypes (p = 0.81). The decrease in heart rate after atenolol administration did not differ significantly according to the GRK5 genotype at rest or after exercise, before (all p > 0.14) and after statistical adjustment for covariates (all p > 0.17). CONCLUSION: The GRK5 Gln41Leu polymorphism does not affect sensitivity to the beta(1)-adrenergic blocker, atenolol, during acute physiological adrenergic stimulation, nor does it contribute to the ethnic differences in sensitivity to atenolol among black and Caucasian individuals.

The alpha2C-adrenoceptor deletion322-325 variant and cold-induced vasoconstriction.

OBJECTIVES: Cold-induced vasoconstriction is mediated in part by selective enhancement of local alpha(2C)-adrenoceptor (alpha(2C)-AR) activity. A common insertion-deletion variant in the alpha(2C)-AR gene (ADRA2C del322-325) results in an approximately 85% reduction of agonist-mediated function in vitro. We tested the hypothesis that individuals with the ADRA2C del322-325 variant have attenuated vasoconstriction in response to cold. METHODS: Cutaneous digital blood flow (flux) was measured by laser Doppler flowmetry in a controlled environment at room temperature and during two cycles of graduated local heat and cold exposure in 31 subjects. Temperature-response curves were analyzed to estimate the following measures: E(min) (minimal flux during cooling), and ET(50) and ET(90) (the local temperature at which flux decreased by 50 and 90%, respectively). RESULTS: We found no significant genotypic differences in E(min) (24.3 +/- 19.5, 30.0 +/- 20.5, and 21.5 +/- 25.9 AU for ins/ins, ins/del, and del/del genotypes, respectively; P = 0.48), ET(50) (25.5 +/- 6.0, 25.1 +/- 6.7, and 25.1 +/- 7.1 degrees C; P = 0.99), or ET(90) (20.5 +/- 4.7, 22.1 +/- 4.0, and 20.8 +/- 6.7 degrees C; P = 0.77) in either the first or second heating and cooling cycle (cycle 1 values presented). INTERPRETATION: The ADRA2C del322-325 variant did not affect vascular sensitivity to local cold exposure.