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Recent Research Articles from UNTHSC

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Total testosterone and neuropsychiatric symptoms in elderly men with Alzheimer's disease.

Sun, 05/10/2015 - 3:29am
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Total testosterone and neuropsychiatric symptoms in elderly men with Alzheimer's disease.

Alzheimers Res Ther. 2015;7(1):24

Authors: Hall JR, Wiechmann AR, Cunningham RL, Johnson LA, Edwards M, Barber RC, Singh M, Winter S, O'Bryant SE, Texas Alzheimer’s Research and Care Consortium

Abstract
INTRODUCTION: There has been a significant increase in the use of testosterone in aging men, but little investigation into its impact on men with Alzheimer's disease (AD). The findings of the few studies that have been done are inconsistent. In the present study, we investigated the relationship between total testosterone (TT) and neuropsychiatric symptoms (NPS) in a well-characterized sample of elderly men with mild to moderate AD.
METHODS: The sample, which was drawn from the Texas Alzheimer's Research Care Consortium Longitudinal Research Cohort, included 87 men who met the criteria for mild to moderate AD. The occurrence of NPS was gathered from caregivers and/or family members with the Neuropsychiatric Inventory. TT was analyzed, and the sample was divided into a low-testosterone group (TT ≤2.5 ng/ml; n = 44) and a borderline/normal group (TT ≥2.6 ng/ml; n = 43).
RESULTS: TT was correlated with symptoms of hallucinations, delusions, agitation, irritability and motor activity. The borderline/normal group was significantly more likely to have hallucinations (odds ratio (OR) = 5.56), delusions (OR = 3.87), motor activity (OR = 3.13) and irritability (OR = 2.77) than the low-testosterone group. Health status and apolipoprotein E ε4 status were not significant factors.
CONCLUSIONS: The findings of the present study have implications for the use of testosterone replacement therapy in men with AD or the prodromal stage of the disease.

PMID: 25937840 [PubMed]

AMPA receptor desensitization is the determinant of AMPA receptor mediated excitotoxicity in purified retinal ganglion cells.

Sun, 05/10/2015 - 3:29am
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AMPA receptor desensitization is the determinant of AMPA receptor mediated excitotoxicity in purified retinal ganglion cells.

Exp Eye Res. 2015 Mar;132:136-50

Authors: Park YH, Mueller BH, McGrady NR, Ma HY, Yorio T

Abstract
The ionotropic glutamate receptors (iGLuR) have been hypothesized to play a role in neuronal pathogenesis by mediating excitotoxic death. Previous studies on iGluR in the retina have focused on two broad classes of receptors: NMDA and non-NMDA receptors including the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic receptor (AMPAR) and kainate receptor. In this study, we examined the role of receptor desensitization on the specific excitotoxic effects of AMPAR activation on primary retinal ganglion cells (RGCs). Purified rat RGCs were isolated from postnatal day 4-7 Sprague-Dawley rats. Calcium imaging was used to identify the functionality of the AMPARs and selectivity of the s-AMPA agonist. Phosphorylated CREB and ERK1/2 expression were performed following s-AMPA treatment. s-AMPA excitotoxicity was determined by JC-1 mitochondrial membrane depolarization assay, caspase 3/7 luciferase activity assay, immunoblot analysis for α-fodrin, and Live (calcein AM)/Dead (ethidium homodimer-1) assay. RGC cultures of 98% purity, lacking Iba1 and GFAP expression were used for the present studies. Isolated prenatal RGCs expressed calcium permeable AMPAR and s-AMPA (100 μM) treatment of cultured RGCs significantly increased phosphorylation of CREB but not that of ERK1/2. A prolonged (6 h) AMPAR activation in purified RGCs using s-AMPA (100 μM) did not depolarize the RGC mitochondrial membrane potential. In addition, treatment of cultured RGCs with s-AMPA, both in the presence and absence of trophic factors (BDNF and CNTF), did not increase caspase 3/7 activities or the cleavage of α-fodrin (neuronal apoptosis marker), as compared to untreated controls. Lastly, a significant increase in cell survival of RGCs was observed after s-AMPA treatment as compared to control untreated RGCs. However, preventing the desensitization of AMPAR with the treatment with either kainic acid (100 μM) or the combination of s-AMPA and cyclothiazide (50 μM) significantly reduced cell survivability. Activation of the AMPAR in RGCs does not appear to activate a signaling cascade to apoptosis, suggesting that RGCs in vitro are not susceptible to AMPA excitotoxicity as previously hypothesized. Conversely, preventing AMPAR desensitization through differential agonist activation caused AMPAR mediated excitotoxicity. Activation of the AMPAR in increasing CREB phosphorylation was dependent on the presence of calcium, which may help explain this action in increasing RGC survival.

PMID: 25643624 [PubMed - indexed for MEDLINE]

Angiotensin II type 1a receptors in subfornical organ contribute towards chronic intermittent hypoxia-associated sustained increase in mean arterial pressure.

Wed, 05/06/2015 - 3:29am
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Angiotensin II type 1a receptors in subfornical organ contribute towards chronic intermittent hypoxia-associated sustained increase in mean arterial pressure.

Am J Physiol Heart Circ Physiol. 2015 Mar 1;308(5):H435-46

Authors: Saxena A, Little JT, Nedungadi TP, Cunningham JT

Abstract
Sleep apnea is associated with hypertension. The mechanisms contributing to a sustained increase in mean arterial pressure (MAP) even during normoxic awake-state remain unknown. Rats exposed to chronic intermittent hypoxia for 7 days, a model of the hypoxemia associated with sleep apnea, exhibit sustained increases in MAP even during the normoxic dark phase. Activation of the renin-angiotensin system (RAS) has been implicated in chronic intermittent hypoxia (CIH) hypertension. Since the subfornical organ (SFO) serves as a primary target for the central actions of circulating ANG II, we tested the effects of ANG II type 1a receptor (AT1aR) knockdown in the SFO on the sustained increase in MAP in this CIH model. Adeno-associated virus carrying green fluorescent protein (GFP) and small-hairpin RNA against either AT1aR or a scrambled control sequence (SCM) was stereotaxically injected in the SFO of rats. After recovery, MAP, heart rate, respiratory rate, and activity were continuously recorded using radiotelemetry. In the normoxic groups, the recorded variables did not deviate from the baseline values. Both CIH groups exhibited significant increases in MAP during CIH exposures (P < 0.05). During the normoxic dark phase in the CIH groups, only the SCM-injected group exhibited a sustained increase in MAP (P < 0.05). The AT1aR-CIH group showed significant decreases in FosB/ΔFosB staining in the median preoptic nucleus and the paraventricular nuclei of the hypothalamus compared with the SCM-CIH group. Our data indicate that AT1aRs in the SFO are critical for the sustained elevation in MAP and increased FosB/ΔFosB expression in forebrain autonomic nuclei associated with CIH.

PMID: 25539713 [PubMed - indexed for MEDLINE]

microRNA- 124 targets Tip110 expression and regulates hematopoiesis.

Sat, 05/02/2015 - 3:32am

microRNA- 124 targets Tip110 expression and regulates hematopoiesis.

Stem Cells Dev. 2015 Apr 30;

Authors: Liu Y, Huang X, Timani K, Broxmeyer HE, He JJ

Abstract
MicroRNA (miR) regulates hematopoiesis through targeting different genes posttranscriptionally. We have recently shown that Tip110 expression is down-regulated during hematopoietic stem cell (HSC) differentiation. However, the underlying mechanisms are not known. In this study, we identified a conserved miR-124 binding site on the Tip110 3' untranslated region (3'UTR) and showed that Tip110 was down-regulated by miR-124 through its 3'UTR. We then examined the relationship among miR-124 and Tip110 expression and differentiation of human cord blood CD34+ cells. We found that miR-124 was expressed in a low level in human cord blood CD34+ cells, but it was considerably up-regulated during culturing and differentiation of these cells. Moreover, we demonstrated that miR-124 expression decreased Tip110 expression and promoted differentiation of human cord blood CD34+ cells, while miR-124 knockdown increased Tip110 expression, slowed down differentiation of human cord blood CD34+ cells, and caused an expansion of hematopoietic progenitor cells in vitro. Finally, we used mouse embryonic fibroblasts derived from Tip110 transgenic mice, performed the exon array analysis and found that Tip110 altered a number of genes in the hematopoiesis pathways. Dnmt3a as de novo methyltransferase was also significantly up regulated. That miR-124 was markedly up-regulated during human cord blood CD34+ cell differentiation could be the result of direct loss of its promoter methylation from Dnmt3a. together, our study demonstrates that miR-124 regulates Tip110 expression and differentiation of human cord blood CD34+ cells and suggests important roles of miR- 124/Tip110 in hematopoiesis.

PMID: 25928721 [PubMed - as supplied by publisher]

Treatment for preventing tuberculosis in children and adolescents: a randomized clinical trial of a 3-month, 12-dose regimen of a combination of rifapentine and isoniazid.

Sat, 05/02/2015 - 3:32am
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Treatment for preventing tuberculosis in children and adolescents: a randomized clinical trial of a 3-month, 12-dose regimen of a combination of rifapentine and isoniazid.

JAMA Pediatr. 2015 Mar;169(3):247-55

Authors: Villarino ME, Scott NA, Weis SE, Weiner M, Conde MB, Jones B, Nachman S, Oliveira R, Moro RN, Shang N, Goldberg SV, Sterling TR, International Maternal Pediatric and Adolescents AIDS Clinical Trials Group, Tuberculosis Trials Consortium

Abstract
IMPORTANCE: Three months of a once-weekly combination of rifapentine and isoniazid for treatment of latent tuberculosis infection is safe and effective for persons 12 years or older. Published data for children are limited.
OBJECTIVES: To compare treatment safety and assess noninferiority treatment effectiveness of combination therapy with rifapentine and isoniazid vs 9 months of isoniazid treatment for latent tuberculosis infection in children.
DESIGN, SETTING, AND PARTICIPANTS: A pediatric cohort nested within a randomized, open-label clinical trial conducted from June 11, 2001, through December 17, 2010, with follow-up through September 5, 2013, in 29 study sites in the United States, Canada, Brazil, Hong Kong (China), and Spain. Participants were children (aged 2-17 years) who were eligible for treatment of latent tuberculosis infection.
INTERVENTIONS: Twelve once-weekly doses of the combination drugs, given with supervision by a health care professional, for 3 months vs 270 daily doses of isoniazid, without supervision by a health care professional, for 9 months.
MAIN OUTCOMES AND MEASURES: We compared rates of treatment discontinuation because of adverse events (AEs), toxicity grades 1 to 4, and deaths from any cause. The equivalence margin for the comparison of AE-related discontinuation rates was 5%. Tuberculosis disease diagnosed within 33 months of enrollment was the main end point for testing effectiveness. The noninferiority margin was 0.75%.
RESULTS: Of 1058 children enrolled, 905 were eligible for evaluation of effectiveness. Of 471 in the combination-therapy group, 415 (88.1%) completed treatment vs 351 of 434 (80.9%) in the isoniazid-only group (P = .003). The 95% CI for the difference in rates of discontinuation attributed to an AE was -2.6 to 0.1, which was within the equivalence range. In the safety population, 3 of 539 participants (0.6%) who took the combination drugs had a grade 3 AE vs 1 of 493 (0.2%) who received isoniazid only. Neither arm had any hepatotoxicity, grade 4 AEs, or treatment-attributed death. None of the 471 in the combination-therapy group developed tuberculosis vs 3 of 434 (cumulative rate, 0.74%) in the isoniazid-only group, for a difference of -0.74% and an upper bound of the 95% CI of the difference of +0.32%, which met the noninferiority criterion.
CONCLUSIONS AND RELEVANCE: Treatment with the combination of rifapentine and isoniazid was as effective as isoniazid-only treatment for the prevention of tuberculosis in children aged 2 to 17 years. The combination-therapy group had a higher treatment completion rate than did the isoniazid-only group and was safe.
TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT00023452.

PMID: 25580725 [PubMed - indexed for MEDLINE]

Exosomes Are Unlikely Involved in Intercellular Nef Transfer.

Fri, 05/01/2015 - 3:30am

Exosomes Are Unlikely Involved in Intercellular Nef Transfer.

PLoS One. 2015;10(4):e0124436

Authors: Luo X, Fan Y, Park IW, He JJ

Abstract
HIV-1 Nef is an important pathogenic factor for HIV/AIDS pathogenesis. Several recent studies including ours have demonstrated that Nef can be transferred to neighboring cells and alters the function of these cells. However, how the intercellular Nef transfer occurs is in dispute. In the current study, we attempted to address this important issue using several complementary strategies, a panel of exosomal markers, and human CD4+ T lymphocyte cell line Jurkat and a commonly used cell line 293T. First, we showed that Nef was transferred from Nef-expressing or HIV-infected Jurkat to naïve Jurkat and other non-Jurkat cells and that the transfer required the membrane targeting function of Nef and was cell density-dependent. Then, we showed that Nef transfer was cell-cell contact-dependent, as exposure to culture supernatants or exosomes from HIV-infected Jurkat or Nef-expressing Jurkat and 293T led to little Nef detection in the target cells Jurkat. Thirdly, we demonstrated that Nef was only detected to be associated with HIV virions but not with acetylcholinesterase (AChE+) exosomes from HIV-infected Jurkat and not in the exosomes from Nef-expressing Jurkat. In comparison, when it was over-expressed in 293T, Nef was detected in detergent-insoluble AChE+/CD81low/TSG101low exosomes, but not in detergent-soluble AChE-/CD81high/TSG101high exosomes. Lastly, microscopic imaging showed no significant Nef detection in exosomal vesicle-like structures in and out 293T. Taken together, these results show that exosomes are unlikely involved in intercellular Nef transfer. In addition, this study reveals existence of two types of exosomes: AChE+/CD81low/TSG101low exosomes and AChE-/CD81high/TSG101high exosomes.

PMID: 25919665 [PubMed - as supplied by publisher]

Activation of mTOR: a culprit of Alzheimer's disease?

Fri, 05/01/2015 - 3:30am
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Activation of mTOR: a culprit of Alzheimer's disease?

Neuropsychiatr Dis Treat. 2015;11:1015-30

Authors: Cai Z, Chen G, He W, Xiao M, Yan LJ

Abstract
Alzheimer's disease (AD) is characterized by cognitive impairment in clinical presentation, and by β-amyloid (Aβ) production and the hyper-phosphorylation of tau in basic research. More highlights demonstrate that the activation of the mammalian target of rapamycin (mTOR) enhances Aβ generation and deposition by modulating amyloid precursor protein (APP) metabolism and upregulating β- and γ-secretases. mTOR, an inhibitor of autophagy, decreases Aβ clearance by scissoring autophagy function. mTOR regulates Aβ generation or Aβ clearance by regulating several key signaling pathways, including phosphoinositide 3-kinase (PI3-K)/protein kinase B (Akt), glycogen synthase kinase 3 [GSK-3], AMP-activated protein kinase (AMPK), and insulin/insulin-like growth factor 1 (IGF-1). The activation of mTOR is also a contributor to aberrant hyperphosphorylated tau. Rapamycin, the inhibitor of mTOR, may mitigate cognitive impairment and inhibit the pathologies associated with amyloid plaques and neurofibrillary tangles by promoting autophagy. Furthermore, the upstream and downstream components of mTOR signaling are involved in the pathogenesis and progression of AD. Hence, inhibiting the activation of mTOR may be an important therapeutic target for AD.

PMID: 25914534 [PubMed]

Circulating mitochondrial DNA and Toll-like receptor 9 are associated with vascular dysfunction in spontaneously hypertensive rats.

Wed, 04/29/2015 - 3:29am

Circulating mitochondrial DNA and Toll-like receptor 9 are associated with vascular dysfunction in spontaneously hypertensive rats.

Cardiovasc Res. 2015 Apr 24;

Authors: McCarthy CG, Wenceslau CF, Goulopoulou S, Ogbi S, Baban B, Sullivan JC, Matsumoto T, Webb RC

Abstract
AIMS: Immune system activation is a common feature of hypertension pathogenesis. However, the mechanisms that initiate this activation are not well understood. Innate immune system recognition and response to danger is becoming apparent in many cardiovascular diseases. Danger signals can arise from not only pathogens, but also damage-associated molecular patterns (DAMPs). Our first hypothesis was that the DAMP, mitochondrial DNA (mtDNA), which is recognized by Toll-like receptor 9 (TLR9), is elevated in the circulation of spontaneously hypertensive rats (SHR), and that the deoxyribonuclease enzymes responsible for its degradation, have decreased activity in SHR. Based on these novel SHR phenotypes, we further hypothesized that (1) treatment of SHR with an inhibitory oligodinucleotide for TLR9 (ODN2088) would lower blood pressure and that (2) treatment of normotensive rats with a TLR9-specific CpG oligonucleotide (ODN2395) would cause endothelial dysfunction and increase blood pressure.
METHODS AND RESULTS: We observed that SHR have elevated circulating mtDNA and diminished deoxyribonuclease I and II activity. Additionally, treatment of SHR with ODN2088 lowered systolic blood pressure. On the other hand, treatment of normotensive rats with ODN2395 increased systolic blood pressure and rendered their arteries less sensitive to acetylcholine-induced relaxation and more sensitive to norepinephrine-induced contraction. This dysfunctional vasoreactivity was due to increased cyclooxygenase and p38 mitogen-activated protein kinase activation, increased reactive oxygen species generation, and reduced nitric oxide bioavailability.
CONCLUSION: Circulating mtDNA and impaired deoxyribonuclease activity may lead to the activation of the innate immune system, via TLR9, and contribute to elevated arterial pressure and vascular dysfunction in SHR.

PMID: 25910936 [PubMed - as supplied by publisher]

CRH promotes S. pneumoniae growth in vitro and increases lung carriage in mice.

Sun, 04/26/2015 - 3:30am

CRH promotes S. pneumoniae growth in vitro and increases lung carriage in mice.

Front Microbiol. 2015;6:279

Authors: Ndjom CG, Jones HP

Abstract
Streptococcus pneumoniae (S. pneumoniae), a commensal across the nasal passages, is responsible for the majority of infectious pneumonia cases worldwide. Previous studies have shown that hormonal factors may be influential in regulating S. pneumoniae's transition from a non-pathogen to a pathogenic state. The current study investigated the effects of corticotropin-releasing hormone (CRH), a peptide hormone involved in stress, on the pathogenicity of S. pneumoniae. Mice were infected with CRH-treated S. pneumoniae via intranasal route, showing an increase in pulmonary bacterial burden. We also quantified S. pneumoniae's response to CRH through limited serial dilutions and growth curve analysis. We demonstrated that CRH promotes S. pneumoniae titer-dependent proliferation, as well as accelerates log-phase growth. Results also showed an increase in pneumococcal-associated virulence protein A virulence gene expression in response to CRH. These results demonstrate a role for CRH in S. pneumoniae pathogenicity, thus implicating CRH in mediating the transition of S. pneumoniae into a pathogenic state.

PMID: 25904910 [PubMed]

Flu-like and Other Systemic Drug Reactions Among Persons Receiving Weekly Rifapentine plus Isoniazid or Daily Isoniazid for Treatment of Latent Tuberculosis Infection in the PREVENT TB study.

Sun, 04/26/2015 - 3:30am

Flu-like and Other Systemic Drug Reactions Among Persons Receiving Weekly Rifapentine plus Isoniazid or Daily Isoniazid for Treatment of Latent Tuberculosis Infection in the PREVENT TB study.

Clin Infect Dis. 2015 Apr 22;

Authors: Sterling TR, Moro RN, Borisov AS, Phillips E, Shepherd G, Adkinson NF, Weis S, Ho C, Villarino ME, Tuberculosis Trials Consortium

Abstract
BACKGROUND:  Weekly rifapentine plus isoniazid for 3 months (3HP) is as effective as daily isoniazid (9H) for 9 months for latent tuberculosis infection in high-risk persons, but there have been reports of possible flu-like syndrome.
METHODS:  We identified clinically significant systemic drug reactions (SDR) and evaluated risk factors in patients who did not complete treatment in the PREVENT TB study.
RESULTS:  Among 7,552 persons who received >1 dose of study drug, 153 had a SDR: 138/3,893(3.5%) with 3HP vs. 15/3,659(0.4%) with 9H(P<0.001). In the 3HP arm, 87(63%) had flu-like syndrome and 23(17%) had cutaneous reactions; 13/3,893(0.3%) had severe reactions (6 were hypotensive) and 6 reported syncope. Symptoms occurred after a median of 3 doses, and 4 hours after the dose; median time to resolution was 24 hours. There were no deaths. In multivariate logistic regression analysis, factors independently associated with SDR included receipt of 3HP (aOR 9.4;95%CI:5.5, 16.2), white non-Hispanic race/ethnicity (aOR 3.3;95%CI:2.3, 4.7), female sex (aOR 2.0;95% CI:1.4, 2.9), age >35 years (aOR 2.0;95% CI:1.4, 2.9), and lower body mass index (BMI; P=0.009). In a separate multivariate analysis among persons who received 3HP, severe SDR were associated with white non-Hispanic race/ethnicity (aOR 5.4;95% CI:1.8, 16.3), and receipt of concomitant non-study medications (aOR 5.9;95% CI:1.3, 27.1).
CONCLUSIONS:  SDR were more common with 3HP, and mostly flu-like. Persons of white race, female sex, older age, and lower BMI were at increased risk. Severe reactions were rare and associated with 3HP, concomitant medication, and white race. The underlying mechanism is unclear.

PMID: 25904367 [PubMed - as supplied by publisher]

A weighted U-statistic for genetic association analyses of sequencing data.

Sun, 04/26/2015 - 3:30am
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A weighted U-statistic for genetic association analyses of sequencing data.

Genet Epidemiol. 2014 Dec;38(8):699-708

Authors: Wei C, Li M, He Z, Vsevolozhskaya O, Schaid DJ, Lu Q

Abstract
With advancements in next-generation sequencing technology, a massive amount of sequencing data is generated, which offers a great opportunity to comprehensively investigate the role of rare variants in the genetic etiology of complex diseases. Nevertheless, the high-dimensional sequencing data poses a great challenge for statistical analysis. The association analyses based on traditional statistical methods suffer substantial power loss because of the low frequency of genetic variants and the extremely high dimensionality of the data. We developed a Weighted U Sequencing test, referred to as WU-SEQ, for the high-dimensional association analysis of sequencing data. Based on a nonparametric U-statistic, WU-SEQ makes no assumption of the underlying disease model and phenotype distribution, and can be applied to a variety of phenotypes. Through simulation studies and an empirical study, we showed that WU-SEQ outperformed a commonly used sequence kernel association test (SKAT) method when the underlying assumptions were violated (e.g., the phenotype followed a heavy-tailed distribution). Even when the assumptions were satisfied, WU-SEQ still attained comparable performance to SKAT. Finally, we applied WU-SEQ to sequencing data from the Dallas Heart Study (DHS), and detected an association between ANGPTL 4 and very low density lipoprotein cholesterol.

PMID: 25331574 [PubMed - indexed for MEDLINE]

Stress induces p38 MAPK-mediated phosphorylation and inhibition of Drosha-dependent cell survival.

Sun, 04/26/2015 - 3:30am
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Stress induces p38 MAPK-mediated phosphorylation and inhibition of Drosha-dependent cell survival.

Mol Cell. 2015 Feb 19;57(4):721-34

Authors: Yang Q, Li W, She H, Dou J, Duong DM, Du Y, Yang SH, Seyfried NT, Fu H, Gao G, Mao Z

Abstract
MicroRNAs (miRNAs) regulate the translational potential of their mRNA targets and control many cellular processes. The key step in canonical miRNA biogenesis is the cleavage of the primary transcripts by the nuclear RNase III enzyme Drosha. Emerging evidence suggests that the miRNA biogenic cascade is tightly controlled. However, little is known whether Drosha is regulated. Here, we show that Drosha is targeted by stress. Under stress, p38 MAPK directly phosphorylates Drosha at its N terminus. This reduces its interaction with DiGeorge syndrome critical region gene 8 and promotes its nuclear export and degradation by calpain. This regulatory mechanism mediates stress-induced inhibition of Drosha function. Reduction of Drosha sensitizes cells to stress and increases death. In contrast, increase in Drosha attenuates stress-induced death. These findings reveal a critical regulatory mechanism by which stress engages p38 MAPK pathway to destabilize Drosha and inhibit Drosha-mediated cellular survival.

PMID: 25699712 [PubMed - indexed for MEDLINE]

A nanotherapy strategy significantly enhances anticryptosporidial activity of an inhibitor of bifunctional thymidylate synthase-dihydrofolate reductase from Cryptosporidium.

Fri, 04/24/2015 - 7:29am

A nanotherapy strategy significantly enhances anticryptosporidial activity of an inhibitor of bifunctional thymidylate synthase-dihydrofolate reductase from Cryptosporidium.

Bioorg Med Chem Lett. 2015 Apr 4;

Authors: Mukerjee A, Iyidogan P, Castellanos-Gonzalez A, Cisneros JA, Czyzyk D, Ranjan AP, Jorgensen WL, White AC, Vishwanatha JK, Anderson KS

Abstract
Cryptosporidiosis, a gastrointestinal disease caused by protozoans of the genus Cryptosporidium, is a common cause of diarrheal diseases and often fatal in immunocompromised individuals. Bifunctional thymidylate synthase-dihydrofolate reductase (TS-DHFR) from Cryptosporidium hominis (C. hominis) has been a molecular target for inhibitor design. C. hominis TS-DHFR inhibitors with nM potency at a biochemical level have been developed however drug delivery to achieve comparable antiparasitic activity in Cryptosporidium infected cell culture has been a major hurdle for designing effective therapies. Previous mechanistic and structural studies have identified compound 906 as a nM C. hominis TS-DHFR inhibitor in vitro, having μM antiparasitic activity in cell culture. In this work, proof of concept studies are presented using a nanotherapy approach to improve drug delivery and the antiparasitic activity of 906 in cell culture. We utilized PLGA nanoparticles that were loaded with 906 (NP-906) and conjugated with antibodies to the Cryptosporidium specific protein, CP2, on the nanoparticle surface in order to specifically target the parasite. Our results indicate that CP2 labeled NP-906 (CP2-NP-906) reduces the level of parasites by 200-fold in cell culture, while NP-906 resulted in 4.4-fold decrease. Moreover, the anticryptosporidial potency of 906 improved 15 to 78-fold confirming the utility of the antibody conjugated nanoparticles as an effective drug delivery strategy.

PMID: 25900220 [PubMed - as supplied by publisher]

Streptozotocin-induced type 1 diabetes in rodents as a model for studying mitochondrial mechanisms of diabetic β cell glucotoxicity.

Thu, 04/23/2015 - 3:29am

Streptozotocin-induced type 1 diabetes in rodents as a model for studying mitochondrial mechanisms of diabetic β cell glucotoxicity.

Diabetes Metab Syndr Obes. 2015;8:181-8

Authors: Wu J, Yan LJ

Abstract
Chronic hyperglycemia and the corresponding glucotoxicity are the main pathogenic mechanisms of diabetes and its complications. Streptozotocin (STZ)-induced diabetic animal models are useful platforms for the understanding of β cell glucotoxicity in diabetes. As diabetes induced by a single STZ injection is often referred to as type 1 diabetes that is caused by STZ's partial destruction of pancreas, one question often being asked is whether the STZ type 1 diabetes animal model is a good model for studying the mitochondrial mechanisms of β cell glucotoxicity. In this mini review, we provide evidence garnered from the literature that the STZ type 1 diabetes is indeed a suitable model for studying mitochondrial mechanisms of diabetic β cell glucotoxicity. Evidence presented includes: 1) continued β cell derangement is due to chronic hyperglycemia after STZ is completely eliminated out of the body; 2) STZ diabetes can be reversed by insulin treatment, which indicates that β cell responds to treatment and shows ability to regenerate; and 3) STZ diabetes can be ameliorated or alleviated by administration of phytochemicals. In addition, mechanisms of STZ action and fundamental gaps in understanding mitochondrial mechanisms of β cell dysfunction are also discussed.

PMID: 25897251 [PubMed]

Assessment of subunit-dependent direct gating and allosteric modulatory effects of carisoprodol at GABAA receptors.

Thu, 04/23/2015 - 3:29am

Assessment of subunit-dependent direct gating and allosteric modulatory effects of carisoprodol at GABAA receptors.

Neuropharmacology. 2015 Apr 17;

Authors: Kumar M, González LA, Dillon GH

Abstract
Carisoprodol is a widely prescribed muscle relaxant, abuse of which has grown considerably in recent years. It directly activates and allosterically modulates α1β2γ2 GABAARs, although the site(s) of action are unknown. To gain insight into the actions of carisoprodol, subunit-dependent effects of this drug were assessed. Whole-cell patch clamp recordings were obtained from HEK293 cells expressing α1β2, α1β3 or αxβzγ2 (where x = 1-6 and z = 1-3) GABAARs, and in receptors incorporating the δ subunit (modeling extrasynaptic receptors). The ability to directly gate and allosterically potentiate GABA-gated currents was observed for all configurations. Presence or absence of the γ2 subunit did not affect the ability of carisoprodol to directly gate or allosterically modulate the receptor. Presence of the β1 subunit conferred highest efficacy for direct activation relative to maximum GABA currents, while presence of the β2 subunit conferred highest efficacy for allosteric modulation of the GABA response. With regard to α subunits, carisoprodol was most efficacious at enhancing the actions of GABA in receptors incorporating the α1 subunit. The ability to directly gate the receptor was generally comparable regardless of the α subunit isoform, although receptors incorporating the α3 subunit showed significantly reduced direct gating efficacy and affinity. In extrasynaptic (α1β3δ and α4β3δ) receptors, carisoprodol had greater efficacy than GABA as a direct gating agonist. In addition, carisoprodol allosterically potentiated both EC20 and saturating GABA concentrations in these receptors. In assessing voltage-dependence, we found direct gating and inhibitory effects were insensitive to membrane voltage, whereas allosteric modulatory effects were affected by membrane voltage. Our findings demonstrate direct and allosteric effects of carisoprodol at synaptic and extrasynpatic GABAARs and that subunit isoform influences these effects.

PMID: 25896767 [PubMed - as supplied by publisher]

Risk factors for and assessment of symptomatic pseudarthrosis after lumbar pedicle subtraction osteotomy in adult spinal deformity.

Thu, 04/23/2015 - 3:29am
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Risk factors for and assessment of symptomatic pseudarthrosis after lumbar pedicle subtraction osteotomy in adult spinal deformity.

Spine (Phila Pa 1976). 2014 Jul 1;39(15):1190-5

Authors: Dickson DD, Lenke LG, Bridwell KH, Koester LA

Abstract
STUDY DESIGN: Retrospective review of prospectively collected data.
OBJECTIVE: To assess the prevalence, risk factors, and clinical outcomes for pseudarthrosis after a lumbar pedicle subtraction osteotomy (PSO).
SUMMARY OF BACKGROUND DATA: There exists no large series that examines pseudarthrosis rates of PSOs.
METHODS: Data of 171 consecutive patients with adult deformity who underwent a lumbar PSO by 2 surgeons at a single institution with a minimum 2-year follow-up were analyzed. Pseudarthrosis diagnosed through sagittal malalignment and instrumentation failure noted on radiograph was confirmed intraoperatively.
RESULTS: Eighteen (10.5%) of 171 patients developed pseudarthrosis after a PSO. Eleven of the 18 patients (6.4% of all patients, 61.1% of the 18 patients with pseudarthrosis) had pseudarthrosis at the PSO site, L3 being the most common; other locations included the lumbosacral junction (4/18), thoracolumbar junction (2/18), and upper thoracic spine (1/18). Preoperative pseudarthrosis level was a predictor of the postoperative level of pseudarthrosis (93%). Fifteen of the 18 patients (83%) had no interbody fusion directly above or below the PSO site, 16 (88%) had a history of pseudarthrosis at the time of PSO surgery and 2 of 3 patients who had prior radiation to the lumbar region developed pseudarthrosis. Most pseudarthroses occurred within the first 2 years (n = 13/18), between 2 and 5 years (n = 3/18), and more than 5 years (n = 2/18) postoperatively. Prior pseudarthrosis (P < 0.0001), pseudarthrosis at the PSO site (P < 0.0001), prior decompression in the lumbar region (P = 0.0037), prior radiation to the lumbar region (P < 0.0001), and presence of inflammatory/neurological disorders (P < 0.0036) were identified as risk factors. All 18 patients with pseudarthroses required revision surgery (posterior-only surgery, n = 12; anteroposterior surgery, n = 6) due to loss of sagittal alignment and pain. The mean pre-revision Scoliosis Research Society score was 85, post-revision score was 95 (P = 0.0166), and the mean pre-revision Oswestry Disability Index score was 42.5, post-revision score was 34.5 (P = 0.0203).
CONCLUSION: The overall prevalence of pseudarthrosis was 10.5% of which 61% occurred at the actual PSO site and Scoliosis Research Society and Oswestry Disability Index scores improved significantly after pseudarthrosis repair.
LEVEL OF EVIDENCE: 4.

PMID: 25171067 [PubMed - indexed for MEDLINE]

Detergent screening of the human voltage-gated proton channel using fluorescence-detection size-exclusion chromatography.

Thu, 04/23/2015 - 3:29am
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Detergent screening of the human voltage-gated proton channel using fluorescence-detection size-exclusion chromatography.

Protein Sci. 2014 Aug;23(8):1136-47

Authors: Agharkar A, Rzadkowolski J, McBroom M, Gonzales EB

Abstract
The human voltage-gated proton channel (Hv1) is a membrane protein consisting of four transmembrane domains and intracellular amino- and carboxy-termini. The protein is activated by membrane depolarization, similar to other voltage-sensitive proteins. However, the Hv1 proton channel lacks a traditional ion pore. The human Hv1 proton channel has been implicated in mediating sperm capacitance, stroke, and most recently as a biomarker/mediator of cancer metastasis. Recently, the three-dimensional structures for homologues of this voltage-gated proton channel were reported. However, it is not clear what artificial environment is needed to facilitate the isolation and purification of the human Hv1 proton channel for structural study. In the present study, we generated a chimeric protein that placed an enhanced green fluorescent protein (EGFP) to the amino-terminus of the human Hv1 proton channel (termed EGFP-Hv1). The chimeric protein was expressed in a baculovirus expression system using Sf9 cells and subjected to detergent screening using fluorescence-detection size-exclusion chromatography. The EGFP-Hv1 proton channel can be solubilized in the zwitterionic detergent Anzergent 3-12 and the nonionic n-dodecyl-β-d-maltoside (DDM) with little protein aggregation and a prominent monomeric protein peak at 48 h postinfection. Furthermore, we demonstrate that the chimeric protein exhibits a monomeric protein peak, which is distinguishable from protein aggregates, at the final size-exclusion chromatography purification step. Taken together, we can conclude that solubilization in DDM will provide a useable final product for further structural characterization of the full-length human Hv1 proton channel.

PMID: 24863684 [PubMed - indexed for MEDLINE]

Neonatal Variables, Altitude of Residence and Aymara Ancestry in Northern Chile.

Wed, 04/22/2015 - 3:29am

Neonatal Variables, Altitude of Residence and Aymara Ancestry in Northern Chile.

PLoS One. 2015;10(4):e0121834

Authors: Rothhammer F, Fuentes-Guajardo M, Chakraborty R, Lorenzo Bermejo J, Dittmar M

Abstract
Studies performed in the Andean plateau, one of the highest inhabited areas in the world, have reported that reduced availability of oxygen is associated to fetal growth retardation and lower birth weight, which are established predictors of morbidity and mortality during the first year of life. To test this hypothesis, perinatal variables of neonates born at the Juan Noé Hospital of Arica, Chile, were analyzed in relation to altitude of residence and Aymara ancestry of their mothers. The study population comprised the offspring of 5,295 mothers born between February 2004 and August 2010. Information included birth weight, height, head circumference, gestational age, altitude of residence and socioeconomic status, and was obtained from medical records. Mother´s ancestry was assessed based on surnames which were linked to percentages of Aymara admixture estimates relying on 40 selected ancestry informative markers. After correcting for the effect of multicollinearity among predictor variables, neonates born to mothers with an increased component of Aymara ancestry showed significantly higher birth weight and height at sea level, a marginally significant (p-value 0.06) decrease of birth weight and a significant decrease of height with altitude in comparison with the offspring of mothers with low Aymara ancestry. Since observed tendencies are suggestive of a possible genetic adaptation to hypoxia of the Chilean Aymara, we discuss briefly preliminary evidence related to fetal oxygen transport, particularly polymorphisms in the promoters of the HBG1 and HBG2 genes that are modulators of HbF synthesis, obtained in this ethnic group.

PMID: 25885573 [PubMed - as supplied by publisher]

Inhibition of triple-negative and Herceptin-resistant breast cancer cell proliferation and migration by Annexin A2 antibodies.

Sat, 04/18/2015 - 3:31am
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Inhibition of triple-negative and Herceptin-resistant breast cancer cell proliferation and migration by Annexin A2 antibodies.

Br J Cancer. 2014 Dec 9;111(12):2328-41

Authors: Chaudhary P, Thamake SI, Shetty P, Vishwanatha JK

Abstract
BACKGROUND: Annexin A2 (AnxA2), a calcium-dependent phospholipid binding protein, is abundantly present at the surface of triple-negative and Herceptin-resistant breast cancer cells. Interactions between cell-surface AnxA2 and tyrosine kinase receptors have an important role in the tumour microenvironment and act together to enhance tumour growth. The mechanism supporting this role is still unknown.
METHODS: The membrane function of AnxA2 was blocked by incubating cells with anti-AnxA2 antibodies. Western blotting, immunoprecipitation, immunofluorescence, 1-(4,5-Dimethylthiazol-2-yl)-3,5-diphenylformazan (MTT), flow cytometry, Clonogenic, and wound-healing assays were performed in this study.
RESULTS: We demonstrate that AnxA2 interacts with epidermal growth factor receptor (EGFR) at the cell surface and has an important role in cancer cell proliferation and migration by modulating EGFR functions. Blocking AnxA2 function at the cell surface by anti-AnxA2 antibody suppressed the EGF-induced EGFR tyrosine phosphorylation and internalisation by blocking its homodimerisation. Furthermore, addition of AnxA2 antibody significantly inhibited the EGFR-dependent PI3K-AKT and Raf-MEK-ERK downstream pathways under both EGF-induced and basal growth conditions, resulting in lower cell proliferation and migration.
CONCLUSIONS: These findings suggest that cell-surface AnxA2 has an important regulatory role in EGFR-mediated oncogenic processes by keeping EGFR signalling events in an activated state. Therefore, AnxA2 could potentially be used as a therapeutic target in triple-negative and Herceptin-resistant breast cancers.

PMID: 25321192 [PubMed - indexed for MEDLINE]

Diversity of piroplasms detected in blood-fed and questing ticks from several states in the United States.

Sat, 04/18/2015 - 3:31am
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Diversity of piroplasms detected in blood-fed and questing ticks from several states in the United States.

Ticks Tick Borne Dis. 2014 Jun;5(4):373-80

Authors: Shock BC, Moncayo A, Cohen S, Mitchell EA, Williamson PC, Lopez G, Garrison LE, Yabsley MJ

Abstract
Piroplasms in the genera Babesia, Theileria, and Cytauxzoon are tick-borne parasites that may be animal and human pathogens. Most piroplasms with known life cycles are transmitted by ixodid ticks; however, for many species, the vector is unknown. This study was conducted to determine the prevalence and diversity of piroplasms in ticks from several US states. Piroplasm-specific polymerase chain reaction (PCR) assays were used to test 1631 ticks from Georgia (n=486), Kentucky (n=103), Pennsylvania (n=1), Tennessee (n=626), and Texas (n=414). Ticks were either questing (n=42) or collected from animals (n=627) or humans (n=962). The 2 primary species tested were Dermacentor variabilis (n=702) and Amblyomma americanum (n=743), but Amblyomma cajennense (n=99), Amblyomma maculatum (n=16), Ixodes scapularis (n=4), I. woodi (n=1), and unidentified Amblyomma spp. nymphs (n=64) were also tested. A low prevalence of piroplasms was detected with 37 (2.3%), 35 (2.1%), and 9 (0.6%) ticks positive for Theileria spp., Babesia spp., or Cytauxzoon felis, respectively. Based on sequence analysis, at least 6 Babesia spp. were detected and 15 of the 35 (41%) Babesia-positive ticks were A. americanum, 19 (56%) were D. variabilis, and one (3%) was an I. scapularis. Nine Babesia-positive ticks were removed from humans from Kentucky (n=1), Georgia (n=2), Texas (n=5), and Pennsylvania (n=1). Three Babesia-positive ticks were questing A. americanum which represents the first report of Babesia-infected questing Amblyomma in the US. Theileria infections were only detected in A. americanum, and all sequences were similar to white-tailed deer associated Theileria spp. C. felis was only detected in D. variabilis. These data suggest that A. americanum may be a vector of Babesia spp., although experimental studies are needed to confirm vector competence. Finally, these data demonstrate a high diversity of piroplasms in both questing and partially fed ticks in the US; although, host-blood meals can be present in non-questing ticks.

PMID: 24709338 [PubMed - indexed for MEDLINE]

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