Recent Research Articles from UNTHSC

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5-hydroxytryptamine-mediated neurotransmission modulates spontaneous and vagal-evoked glutamate release in the nucleus of the solitary tract effect of uptake blockade.

Tue, 07/22/2014 - 4:04am
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5-hydroxytryptamine-mediated neurotransmission modulates spontaneous and vagal-evoked glutamate release in the nucleus of the solitary tract effect of uptake blockade.

J Pharmacol Exp Ther. 2014 May;349(2):288-96

Authors: Hosford PS, Mifflin SW, Ramage AG

Abstract
The effect of blockade of either 5-hydroxytryptamine (5-HT)/serotonin transporter (SERT) with citalopram or the organic cation transporter 3 (OCT3)/plasma membrane monoamine transporter (PMAT) with decynium-22 (D-22) on spontaneous and evoked release of 5-HT in the nucleus tractus solitarius (NTS) was investigated in rat brainstem slices treated with gabazine. 5-HT release was measured indirectly by changes in the frequency and amplitude of glutamatergic miniature excitatory postsynaptic currents (mEPSCs) [in the presence of tetrodotoxin (TTX)] and evoked EPSCs. Blockade of 5-HT3 receptors with granisetron reduced, whereas the 5-HT3 agonist phenylbiguanide increased, the frequency of mEPSCs. 5-HT decreased mEPSC frequency at low concentrations and increased frequency at high concentrations. This inhibition was blocked by the 5-HT1A antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-2-pyridinylcyclohexanecarboxamide (WAY-100635), which was ineffective on its own, whereas the excitation was reversed by granisetron. The addition of citalopram or D-22 caused inhibition, which was prevented by 5-HT1A blockade. Thus, in the NTS, the spontaneous release of 5-HT is able to activate 5-HT3 receptors, but not 5-HT1A receptors, as the release in their vicinity is removed by uptake. The ineffectiveness of corticosterone suggests that the low-affinity, high-capacity transporter is PMAT, not OCT3. For evoked 5-HT release, only D-22 caused an increase in the amplitude of EPSCs, with a decrease in the paired pulse ratio, and increased the number of spontaneous EPSCs after 20-Hz stimulation. Thus, for the evoked release of 5-HT, the low-affinity, high-capacity transporter PMAT, but not 5-HT transporter (5-HTT)/SERT, is important in the regulation of changes in 5-HT extracellular concentration.

PMID: 24618127 [PubMed - indexed for MEDLINE]

Establishment of human retinal microvascular endothelial cells with extended life-span.

Sat, 07/19/2014 - 4:06am
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Establishment of human retinal microvascular endothelial cells with extended life-span.

In Vivo. 2013 Nov-Dec;27(6):685-94

Authors: Kashyap MV, Ranjan AP, Shankardas J, Vishwanatha JK

Abstract
AIM: To generate and characterize a telomerase-immortalized human retinal microvascular endothelial cell (HREC) line. This cell line may be utilized as an in vitro model to study the molecular basis of several diseases of the human retina.
MATERIALS AND METHODS: Primary retinal neuronal cells were isolated and transfected with plasmid encoding full-length human telomerase reverse transcriptase (hTERT). Transfected cells were selected and characterized to determine telomerase activity, karyotype, proliferative capacity and functionality.
RESULTS: HREC-hTERT cells appear morphologically similar to primary endothelial cells and have an extended in vitro life-span. HREC-hTERT cells express the progenitor/stem cell marker nestin. They have active telomerase and a high proliferative capacity. These cells also maintain a diploid karyotype. The HREC-hTERT cells showed high colony-formation capacity and plating efficiency compared to the primary cells. These cells are capable of differentiation into neuronal and glial cell phenotypes and the differentiated cells express the astrocyte marker glial fibrillary acidic protein (GFAP) and the neuronal marker microtubule-associated protein-2 (MAP2), respectively.
CONCLUSION: The in vitro life-span of human retinal neuronal endothelial cells can be extended by ectopic expression of hTERT without altering the genetic stability and functionality of these cells. These cells will be a valuable tool to further our understanding on the role of HRECs in the human blood-retinal-barrier and in angiogenesis and neovascularization.

PMID: 24292569 [PubMed - indexed for MEDLINE]

Treatment of Acute Pulmonary Embolism: Update on Newer Pharmacologic and Interventional Strategies.

Thu, 07/17/2014 - 4:04am
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Treatment of Acute Pulmonary Embolism: Update on Newer Pharmacologic and Interventional Strategies.

Biomed Res Int. 2014;2014:410341

Authors: Pelliccia F, Schiariti M, Terzano C, Keylani AM, D'Agostino DC, Speziale G, Greco C, Gaudio C

Abstract
Acute pulmonary embolism (PE) is a common complication in hospitalized patients, spanning multiple patient populations and crossing various therapeutic disciplines. Current treatment paradigm in patients with massive PE mandates prompt risk stratification with aggressive therapeutic strategies. With the advent of endovascular technologies, various catheter-based thrombectomy and thrombolytic devices are available to treat patients with massive or submassive PE. In this paper, a variety of newer treatment strategies for PE are analyzed, with special emphasis on various interventional treatment strategies. Clinical evidence for utilizing endovascular treatment modalities, based on our institutional experience as well as a literature review, is provided.

PMID: 25025049 [PubMed - as supplied by publisher]

Validation of a Serum Screen for Alzheimer's Disease Across Assay Platforms, Species, and Tissues.

Thu, 07/17/2014 - 4:04am
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Validation of a Serum Screen for Alzheimer's Disease Across Assay Platforms, Species, and Tissues.

J Alzheimers Dis. 2014 Jul 11;

Authors: O'Bryant SE, Xiao G, Zhang F, Edwards M, German DC, Yin X, Como T, Reisch J, Huebinger RM, Graff-Radford N, Dickson D, Barber R, Hall J, O'Suilleabhain P, Grammas P

Abstract
Background: There is a significant need for rapid and cost-effective biomarkers of Alzheimer's disease (AD) for advancement of clinical practice and therapeutic trials. Objective: The aim of the current study was to cross-validate our previously published serum-based algorithm on an independent assay platform as well as validate across tissues and species. Preliminary analyses were conducted to examine the utility in distinguishing AD from non-AD neurological disease (Parkinson's disease, PD). Methods: Serum proteins from our previously published algorithm were quantified from 150 AD cases and 150 controls on the Meso Scale Discovery (MSD) platform. Serum samples were analyzed from 49 PD cases and compared to a random sample of 51 AD cases and 62 controls. Support vector machines (SVM) were used to discriminate PD versus AD versus controls. Human and AD mouse model microvessel images were quantified with HAMAMATSU imaging software. Mouse serum biomarkers were assayed via MSD. Results: Analysis of 21 serum proteins from 150 AD cases and 150 controls yielded an algorithm with sensitivity and specificity of 0.90 for correctly classifying AD. This multi-marker approach was then validated across species and tissue. Assay of the top proteins in human and AD mouse model brain microvessels correctly classified 90-100% of the samples. SVM analyses were highly accurate at distinguishing PD versus AD versus controls. Conclusions: This serum-based biomarker panel should be tested in a community-based setting to determine its utility as a first-line screen for AD and non-AD neurological diseases for primary care providers.

PMID: 25024345 [PubMed - as supplied by publisher]

In vivo Assessment of SMT19969 in the Hamster Model of Clostridium difficile Infection.

Thu, 07/17/2014 - 4:04am
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In vivo Assessment of SMT19969 in the Hamster Model of Clostridium difficile Infection.

Antimicrob Agents Chemother. 2014 Jul 14;

Authors: Weiss W, Pulse M, Vickers R

Abstract
SMT19969 (2, 2' bis(4-pyridyl)3H, 3' H 5,5 bibenzimidazole) is a novel, narrow spectrum, non-absorbable antibiotic currently in development for the treatment of Clostridium difficile infection. The comparative activity of SMT19969 and vancomycin against non-epidemic and epidemic strains of C. difficile was studied in an established hamster model. Against non-epidemic (VA11) strains, survival ranged from 80% to 95% in SMT19969 treated animals. Vancomycin exhibited 100% protection during treatment with relapse observed starting on Day 9 and 50% survival at Day 20. At 50 mg/kg, SMT19969 administered orally, qd x 5 days exhibited full protection of treated animals on dosing days and out through day 12 against epidemic strains. Vancomycin also protected during the dosing interval, but apparent relapse occurred earlier starting on day 11. SMT19969 exhibited excellent in vitro activity with an MIC of 0.25 ug/mL for all isolates. MICs for vancomycin were 2-4 fold higher at ≤0.5-1 ug/mL. All plasma samples of SMT19969 were below the limit of quantitation (25 ng/mL) at all time points, consistent with the reported lack of bioavailability of the compound. Cecal concentrations were significantly above MIC (ranging from 96μg/mL to 172μg/mL).

PMID: 25022586 [PubMed - as supplied by publisher]

Pathogenesis of Chronic Hyperglycemia: From Reductive Stress to Oxidative Stress.

Thu, 07/17/2014 - 4:04am
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Pathogenesis of Chronic Hyperglycemia: From Reductive Stress to Oxidative Stress.

J Diabetes Res. 2014;2014:137919

Authors: Yan LJ

Abstract
Chronic overnutrition creates chronic hyperglycemia that can gradually induce insulin resistance and insulin secretion impairment. These disorders, if not intervened, will eventually be followed by appearance of frank diabetes. The mechanisms of this chronic pathogenic process are complex but have been suggested to involve production of reactive oxygen species (ROS) and oxidative stress. In this review, I highlight evidence that reductive stress imposed by overflux of NADH through the mitochondrial electron transport chain is the source of oxidative stress, which is based on establishments that more NADH recycling by mitochondrial complex I leads to more electron leakage and thus more ROS production. The elevated levels of both NADH and ROS can inhibit and inactivate glyceraldehyde 3-phosphate dehydrogenase (GAPDH), respectively, resulting in blockage of the glycolytic pathway and accumulation of glycerol 3-phospate and its prior metabolites along the pathway. This accumulation then initiates all those alternative glucose metabolic pathways such as the polyol pathway and the advanced glycation pathways that otherwise are minor and insignificant under euglycemic conditions. Importantly, all these alternative pathways lead to ROS production, thus aggravating cellular oxidative stress. Therefore, reductive stress followed by oxidative stress comprises a major mechanism of hyperglycemia-induced metabolic syndrome.

PMID: 25019091 [PubMed - as supplied by publisher]

Manual therapy, exercise, and education for low back pain and pelvic pain during pregnancy.

Thu, 07/17/2014 - 4:04am
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Manual therapy, exercise, and education for low back pain and pelvic pain during pregnancy.

Am J Obstet Gynecol. 2014 Jun;210(6):592-3

Authors: Licciardone JC, Aryal S

PMID: 24607754 [PubMed - indexed for MEDLINE]

Acute improvement in hemodynamic control after osteopathic manipulative treatment in the third trimester of pregnancy.

Thu, 07/17/2014 - 4:04am
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Acute improvement in hemodynamic control after osteopathic manipulative treatment in the third trimester of pregnancy.

Complement Ther Med. 2013 Dec;21(6):618-26

Authors: Hensel KL, Pacchia CF, Smith ML

Abstract
OBJECTIVES: The physiological changes that occur during pregnancy, including increased blood volume and cardiac output, can affect hemodynamic control, most profoundly with positional changes that affect venous return to the heart. By using Osteopathic Manipulative Treatment (OMT), a body-based modality theorized to affect somatic structures related to nervous and circulatory systems, we hypothesized that OMT acutely improves both autonomic and hemodynamic control during head-up tilt and heel raise in women at 30 weeks gestation.
DESIGN: One hundred subjects were recruited at 30 weeks gestation.
SETTING: The obstetric clinics of UNTHealth in Fort Worth, TX.
INTERVENTION: Subjects were randomized into one of three treatment groups: OMT, placebo ultrasound, or time control. Ninety subjects had complete data (N=25, 31 and 34 in each group respectively).
MAIN OUTCOME MEASURES: Blood pressure and heart rate were recorded during 5 min of head-up tilt followed by 4 min of intermittent heel raising.
RESULTS: No significant differences in blood pressure, heart rate or heart rate variability were observed between groups with tilt before or after treatment (p>0.36), and heart rate variability was not different between treatment groups (p>0.55). However, blood pressure increased significantly (p=0.02) and heart rate decreased (p<0.01) during heel raise after OMT compared to placebo or time control.
CONCLUSIONS: These data suggest that OMT can acutely improve hemodynamic control during engagement of the skeletal muscle pump and this was most likely due to improvement of structural restrictions to venous return.

PMID: 24280470 [PubMed - indexed for MEDLINE]

Evaluation of a novel material, Diomics X-Swab™, for collection of DNA.

Wed, 07/16/2014 - 4:04am
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Evaluation of a novel material, Diomics X-Swab™, for collection of DNA.

Forensic Sci Int Genet. 2014 Jun 24;12C:192-198

Authors: Marshall PL, Stoljarova M, Larue BL, King JL, Budowle B

Abstract
Success of DNA typing is related to the amount of target material recovered from an evidentiary item. Generally, the more DNA that is recovered, the better the chance is of obtaining a typing result that will be robust and reliable. One method of collecting stain materials is by swabbing. Recovery of DNA from a number of commercially available swabs is not an efficient process. The X-Swab™ (Diomics Corporation, La Jolla, CA) is a unique bio-specimen collection material with highly absorptive properties and can be dissolved during certain extraction conditions. Therefore, more DNA may be collected from a substrate and be released from the swab matrix than other swabs. The ability to recover DNA from X-Swab material and success in STR typing were compared with the Copan 4N6FLOQSwab™ (Brescia, Italy), a device which utilizes a proprietary flocked-swab technology to maximize DNA collection and elution efficiency. Both types of swabs were impregnated with known amounts of DNA and body fluids and allowed to air dry. In addition, blood was placed onto glass slides, allowed to dry and collected using both types of swabs. DNA recovery was assessed by DNA quantitation and by STR typing. Results suggested that X-Swab material yielded greater DNA recovery, particularly of low quantity samples (defined as diluted neat samples), compared with the 4N6FLOQSwab. Results also indicated that X-Swab material itself enhances yield of PCR products.

PMID: 25016249 [PubMed - as supplied by publisher]

ERK5/KLF4 signaling as a common mediator of the neuroprotective effects of both nerve growth factor and hydrogen peroxide preconditioning.

Wed, 07/16/2014 - 4:04am
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ERK5/KLF4 signaling as a common mediator of the neuroprotective effects of both nerve growth factor and hydrogen peroxide preconditioning.

Age (Dordr). 2014 Aug;36(4):9685

Authors: Su C, Sun F, Cunningham RL, Rybalchenko N, Singh M

Abstract
Oxidative stress has long been implicated in the pathogenesis of various neurodegenerative disorders such as Alzheimer's disease and stroke. While high levels of oxidative stress are generally associated with cell death, a slight rise of reactive oxygen species (ROS) levels can be protective by "preconditioning" cells to develop a resistance against subsequent challenges. However, the mechanisms underlying such preconditioning (PC)-induced protection are still poorly understood. Previous studies have supported a role of ERK5 (mitogen-activated protein [MAP] kinase 5) in neuroprotection and ischemic tolerance in the hippocampus. In agreement with these findings, our data suggest that ERK5 mediates both hydrogen peroxide (H2O2)-induced PC as well as nerve growth factor (NGF)-induced neuroprotection. Activation of ERK5 partially rescued pheochromocytoma PC12 cells as well as primary hippocampal neurons from H2O2-caused death, while inhibition of ERK5 abolished NGF or PC-induced protection. These results implicate ERK5 signaling as a common downstream pathway for NGF and PC. Furthermore, both NGF and PC increased the expression of the transcription factor, KLF4, which can initiate an anti-apoptotic response in various cell types. Induction of KLF4 by NGF or PC was blocked by siERK5, suggesting that ERK5 is required in this process. siKLF4 can also attenuate NGF- or PC-induced neuroprotection. Overexpression of active MEK5 or KLF4 in H2O2-stressed cells increased Bcl-2/Bax ratio and the expression of NAIP (neuronal apoptosis inhibitory protein). Taken together, our data suggest that ERK5/KLF4 cascade is a common signaling pathway shared by at least two important mechanisms by which neurons can be protected from cell death.

PMID: 25015774 [PubMed - as supplied by publisher]

Pulmonary Arterial Hypertension in Adults: Novel Drugs and Catheter Ablation Techniques Show Promise? Systematic Review on Pharmacotherapy and Interventional Strategies.

Sun, 07/13/2014 - 4:04am
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Pulmonary Arterial Hypertension in Adults: Novel Drugs and Catheter Ablation Techniques Show Promise? Systematic Review on Pharmacotherapy and Interventional Strategies.

Biomed Res Int. 2014;2014:743868

Authors: Rosanio S, Pelliccia F, Gaudio C, Greco C, Keylani AM, D'Agostino DC

Abstract
This systematic review aims to provide an update on pharmacological and interventional strategies for the treatment of pulmonary arterial hypertension in adults. Currently US Food and Drug Administration approved drugs including prostanoids, endothelin-receptor antagonists, phosphodiesterase type-5 inhibitors, and soluble guanylate-cyclase stimulators. These agents have transformed the prognosis for pulmonary arterial hypertension patients from symptomatic improvements in exercise tolerance ten years ago to delayed disease progression today. On the other hand, percutaneous balloon atrioseptostomy by using radiofrequency perforation, cutting balloon dilatation, or insertion of butterfly stents and pulmonary artery catheter-based denervation, both associated with very low rate of major complications and death, should be considered in combination with specific drugs at an earlier stage rather than late in the progression of pulmonary arterial hypertension and before the occurrence of overt right-sided heart failure.

PMID: 25013799 [PubMed - as supplied by publisher]

The Association between Patient-Centered Attributes of Care and Patient Satisfaction.

Sun, 07/13/2014 - 4:04am
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The Association between Patient-Centered Attributes of Care and Patient Satisfaction.

Patient. 2014 Jul 11;

Authors: Tak H, Ruhnke GW, Shih YC

Abstract
BACKGROUND AND OBJECTIVE: Little is known about the attributes of care that most strongly impact satisfaction in real-world settings where patients' limited medical knowledge may restrict their ability to ascertain the true quality of care. We therefore examined the association between patient-centered attributes of physician care (thoroughness, explanation, and listening), in-office waiting time, and patient satisfaction.
METHODS: We used the Community Tracking Study Household Survey, a US nationally representative dataset (n = 71,594). Using logistic regression models, we analyzed the association between patient ratings of care attributes and patient satisfaction for the total sample and by subgroups, according to health status, physician type, and visit type.
RESULTS: Patients' perception of excellent or very good care attributes was strongly associated with being very satisfied with care received (thoroughness of care, odds ratio [OR] 2.64, 95 % confidence interval [CI] 2.31-3.02; listening, OR 2.04, 95 % CI 1.77-2.36; explanation, OR 1.63, 95 % CI 1.42-1.86), as was a waiting time of ≤10 min (OR 1.50, 95 % CI 1.39-1.63). The effect magnitude of thoroughness on satisfaction is particularly strong relative to high-quality listening and explanation among respondents in poor health, and for whom the most recent office visit was to see a generalist or for curative care.
CONCLUSIONS: Thoroughness of care was the strongest determinant of patient satisfaction, followed by physician listening and explanation. Especially with patients' improved access to current medical information, it is important for physicians to recognize that excellent communication cannot serve as a substitute for high-quality, thorough care.

PMID: 25011683 [PubMed - as supplied by publisher]

Total Cholesterol and Neuropsychiatric Symptoms in Alzheimer's Disease: The Impact of Total Cholesterol Level and Gender.

Sun, 07/13/2014 - 4:04am
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Total Cholesterol and Neuropsychiatric Symptoms in Alzheimer's Disease: The Impact of Total Cholesterol Level and Gender.

Dement Geriatr Cogn Disord. 2014 Jul 4;38(5-6):300-309

Authors: Hall JR, Wiechmann AR, Johnson LA, Edwards M, Barber RC, Cunningham R, Singh M, O'Bryant SE

Abstract
Background: Neuropsychiatric symptoms (NPS) in Alzheimer's disease (AD) are a major factor in nursing home placement and a primary cause of stress for caregivers. Elevated cholesterol has been linked to psychiatric disorders and has been shown to be a risk factor for AD and to impact disease progression. The present study investigated the relationship between cholesterol and NPS in AD. Methods: Data on cholesterol and NPS from 220 individuals (144 females, 76 males) with mild-to-moderate AD from the Texas Alzheimer's Research and Care Consortium (TARCC) cohort were analyzed. The total number of NPS and symptoms of hyperactivity, psychosis, affect and apathy were evaluated. Groups based on total cholesterol (TC; ≥200 vs. <200 mg/dl) were compared with regard to NPS. The impact of gender was also assessed. Results: Individuals with high TC had lower MMSE scores as well as significantly more NPS and more symptoms of psychosis. When stratified by gender, males with high TC had significantly more NPS than females with high TC or than males or females with low TC. Conclusion: The role of elevated cholesterol in the occurrence of NPS in AD appears to be gender and symptom specific. A cross-validation of these findings will have implications for possible treatment interventions, especially for males with high TC. © 2014 S. Karger AG, Basel.

PMID: 25011444 [PubMed - as supplied by publisher]

ANGIOTENSIN II RECEPTOR SUBTYPE 1A (AT1AR) GENE KNOCKDOWN IN THE SUBFORNICAL ORGAN (SFO) PREVENTS INCREASED DRINKING BEHAVIOR IN BILE DUCT LIGATED RATS.

Sat, 07/12/2014 - 4:05am
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ANGIOTENSIN II RECEPTOR SUBTYPE 1A (AT1AR) GENE KNOCKDOWN IN THE SUBFORNICAL ORGAN (SFO) PREVENTS INCREASED DRINKING BEHAVIOR IN BILE DUCT LIGATED RATS.

Am J Physiol Regul Integr Comp Physiol. 2014 Jul 9;

Authors: Walch JD, Nedungadi TP, Cunningham JT

Abstract
Bile duct ligation (BDL) causes congestive liver failure that initiates hemodynamic changes resulting in dilutional hyponatremia due to increased water intake and vasopressin release. This project tested the hypothesis that angiotensin signaling at the subfornical organ (SFO) augments drinking behavior in BDL rats. A genetically modified adeno-associated virus containing shRNA for angiotensin II receptor subtype 1a (AT1aR) gene was microinjected into the subfornical organ (SFO) of rats to knockdown expression. Two weeks later, BDL or sham surgery was performed. Rats were housed in metabolic chambers for measurement of fluid and food intake and urine output. The rats were euthanized 28 days after BDL surgery for analysis. A group of rats was perfused for immunohistochemistry and a second group was used for laser-capture microdissection for analysis of SFO AT1aR gene expression. BDL rats showed increased water intake that was attenuated in rats that received SFO microinjection of AT1aR shRNA. Among BDL rats treated with scrambled (control) and AT1aR shRNA, we observed an increased number of vasopressin positive cells in the supraoptic nucleus (SON) that colocalized with ΔFosB staining suggesting increased vasopressin release in both groups. These results indicate that angiotensin signalling through the SFO contributes to increased water intake, but not dilutional hyponatremia, during congestive liver failure.

PMID: 25009217 [PubMed - as supplied by publisher]

Response: Observational Study Demonstrates That OMT Is Associated With Reduced Analgesic Prescribing and Fewer Missed Work Days.

Fri, 07/11/2014 - 12:04pm

Response: Observational Study Demonstrates That OMT Is Associated With Reduced Analgesic Prescribing and Fewer Missed Work Days.

J Am Osteopath Assoc. 2014 Jul;114(7):530-1

Authors: Prinsen JK, Hensel KL, Snow RJ

PMID: 25002441 [PubMed - in process]

The OSTEOPATHIC Trial Demonstrates Significant Improvement in Patients With Chronic Low Back Pain as Manifested by Decreased Prescription Rescue Medication Use.

Fri, 07/11/2014 - 12:04pm

The OSTEOPATHIC Trial Demonstrates Significant Improvement in Patients With Chronic Low Back Pain as Manifested by Decreased Prescription Rescue Medication Use.

J Am Osteopath Assoc. 2014 Jul;114(7):528-9

Authors: Licciardone JC

PMID: 25002440 [PubMed - in process]

A high volume extraction and purification method for recovering DNA from human bone.

Wed, 07/09/2014 - 12:05pm
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A high volume extraction and purification method for recovering DNA from human bone.

Forensic Sci Int Genet. 2014 Jun 18;12C:155-160

Authors: Marshall PL, Stoljarova M, Schmedes SE, King JL, Budowle B

Abstract
DNA recovery, purity and overall extraction efficiency of a protocol employing a novel silica-based column, Hi-Flow(®) (Generon Ltd., Maidenhead, UK), were compared with that of a standard organic DNA extraction methodology. The quantities of DNA recovered by each method were compared by real-time PCR and quality of DNA by STR typing using the PowerPlex(®) ESI 17 Pro System (Promega Corporation, Madison, WI) on DNA from 10 human bone samples. Overall, the Hi-Flow method recovered comparable quantities of DNA ranging from 0.8ng±1 to 900ng±159 of DNA compared with the organic method ranging from 0.5ng±0.9 to 855ng±156 of DNA. Complete profiles (17/17 loci tested) were obtained for at least one of three replicates for 3/10 samples using the Hi-Flow method and from 2/10 samples with the organic method. All remaining bone samples yielded partial profiles for all replicates with both methods. Compared with a standard organic DNA isolation method, the results indicated that the Hi-Flow method provided equal or improved recovery and quality of DNA without the harmful effects of organic extraction. Moreover, larger extraction volumes (up to 20mL) can be employed with the Hi-Flow method which enabled more bone sample to be extracted at one time.

PMID: 24997320 [PubMed - as supplied by publisher]

Hypertrophic Cardiomyopathy Associated Lys104Glu Mutation in the Myosin Regulatory Light Chain Causes Diastolic Disturbance in Mice.

Wed, 07/09/2014 - 12:05pm
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Hypertrophic Cardiomyopathy Associated Lys104Glu Mutation in the Myosin Regulatory Light Chain Causes Diastolic Disturbance in Mice.

J Mol Cell Cardiol. 2014 Jun 30;

Authors: Huang W, Liang J, Kazmierczak K, Muthu P, Duggal D, Farman GP, Sorensen L, Pozios I, Abraham TP, Moore JR, Borejdo J, Szczesna-Cordary D

Abstract
We have examined, for the first time, the effects of the familial hypertrophic cardiomyopathy (HCM)- associated Lys104Glu mutation in the myosin regulatory light chain (RLC). Transgenic mice expressing the Lys104Glu substitution (Tg-MUT) were generated and the results compared to Tg-WT (wild-type human ventricular RLC) mice. Echocardiography with pulse wave Doppler in 6month-old Tg-MUT showed early signs of diastolic disturbance with significantly reduced E/A transmitral velocities ratio. Invasive hemodynamics in 6month-old Tg-MUT mice also demonstrated a borderline significant prolonged isovolumic relaxation time (Tau) and a tendency for slower rate of pressure decline, suggesting alterations in diastolic function in Tg-MUT. Six month-old mutant animals had no LV hypertrophy; however, at >13months they displayed significant hypertrophy and fibrosis. In skinned papillary muscles from 5-6 month-old mice a mutation induced reduction in maximal tension and slower muscle relaxation rates were observed. Mutated cross-bridges showed increased rates of binding to the thin filaments and a faster rate of the power stroke. In addition, ~2-fold lower level of RLC phosphorylation was observed in the mutant compared to Tg-WT. In line with the higher mitochondrial content seen in Tg-MUT hearts, the MUT-myosin ATPase activity was significantly higher than WT-myosin, indicating increased energy consumption. In the in vitro motility assay, MUT-myosin produced higher actin sliding velocity under zero load, but the velocity drastically decreased with applied load in the MUT vs. WT myosin. Our results suggest that diastolic disturbance (impaired muscle relaxation, lower E/A) and inefficiency of energy use (reduced contractile force and faster ATP consumption) may underlie the Lys104Glu-mediated HCM phenotype.

PMID: 24992035 [PubMed - as supplied by publisher]

Cancer-associated Isocitrate Dehydrogenase 1 (IDH1) R132H Mutation and D-2-hydroxyglutarate Stimulate Glutamine Metabolism under Hypoxia.

Wed, 07/09/2014 - 12:05pm
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Cancer-associated Isocitrate Dehydrogenase 1 (IDH1) R132H Mutation and D-2-hydroxyglutarate Stimulate Glutamine Metabolism under Hypoxia.

J Biol Chem. 2014 Jul 1;

Authors: Reitman ZJ, Duncan CG, Poteet E, Winters A, Yan LJ, Gooden DM, Spasojevic I, Boros LG, Yang SH, Yan H

Abstract
Mutations in the cytosolic NADP+-dependent isocitrate dehydrogenase (IDH1) occur in several types of cancer, and altered cellular metabolism associated with IDH1 mutations presents unique therapeutic opportunities. By altering IDH1, these mutations target a critical step in reductive glutamine metabolism, the metabolic pathway that converts glutamine ultimately to acetyl-CoA for biosynthetic processes. While IDH1-mutated cells are sensitive to therapies that target glutamine metabolism, the effect of IDH1 mutations on reductive glutamine metabolism remains poorly understood. To explore this issue, we investigated the effect of a knock-in, single-codon IDH1-R132H mutation on the metabolism of the HCT116 colorectal adenocarcinoma cell line. Here we report the R132H-isobolome by using targeted 13C isotopomer tracer fate analysis to trace the metabolic fate of glucose and glutamine in this system. We show that introduction of the R132H mutation into IDH1 upregulates the contribution of glutamine to lipogenesis in hypoxia, but not in normoxia. Treatment of cells with a D-2-hydroxyglutarate (D-2HG) ester recapitulated these changes, indicating that the alterations observed in the knocked-in cells were mediated by D-2HG produced by the IDH1 mutant. These studies provide a dynamic mechanistic basis for metabolic alterations observed in IDH1-mutated tumors and uncover potential therapeutic targets in IDH1-mutated cancers.

PMID: 24986863 [PubMed - as supplied by publisher]

The role of 5-HT2A, 5-HT 2C and mGlu2 receptors in the behavioral effects of tryptamine hallucinogens N,N-dimethyltryptamine and N,N-diisopropyltryptamine in rats and mice.

Wed, 07/09/2014 - 12:05pm
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The role of 5-HT2A, 5-HT 2C and mGlu2 receptors in the behavioral effects of tryptamine hallucinogens N,N-dimethyltryptamine and N,N-diisopropyltryptamine in rats and mice.

Psychopharmacology (Berl). 2014 Jul 3;

Authors: Carbonaro TM, Eshleman AJ, Forster MJ, Cheng K, Rice KC, Gatch MB

Abstract
RATIONALE: Serotonin 5-HT2A and 5-HT2C receptors are thought to be the primary pharmacological mechanisms for serotonin-mediated hallucinogenic drugs, but recently there has been interest in metabotropic glutamate (mGluR2) receptors as contributors to the mechanism of hallucinogens.
OBJECTIVE: The present study assesses the role of these 5-HT and glutamate receptors as molecular targets for two tryptamine hallucinogens, N,N-dimethyltryptamine (DMT) and N,N-diisopropyltryptamine (DiPT).
METHODS: Drug discrimination, head twitch, and radioligand binding assays were used. A 5-HT2AR inverse agonist (MDL100907), 5-HT2CR antagonist (SB242084), and mGluR2/3 agonist (LY379268) were tested for their ability to attenuate the discriminative stimulus effects of DMT and DiPT; an mGluR2/3 antagonist (LY341495) was tested for potentiation. MDL100907 was used to attenuate head twitches induced by DMT and DiPT. Radioligand binding studies and inosital-1-phosphate (IP-1) accumulation were performed at the 5-HT2CR for DiPT.
RESULTS: MDL100907 fully blocked the discriminative stimulus effects of DMT, but only partially blocked DiPT. SB242084 partially attenuated the discriminative stimulus effects of DiPT, but produced minimal attenuation of DMT's effects. LY379268 produced potent, but only partial blockade of the discriminative stimulus effects of DMT. LY341495 facilitated DMT- and DiPT-like effects. Both compounds elicited head twitches (DiPT>DMT) which were blocked by MDL1000907. DiPT was a low-potency full agonist at 5-HT2CR in vitro.
CONCLUSIONS: The 5-HT2AR likely plays a major role in mediating the effects of both compounds. 5-HT2C and mGluR2 receptors likely modulate the discriminative stimulus effects of both compounds to some degree.

PMID: 24985890 [PubMed - as supplied by publisher]