Increased expression of iNOS in the postural muscles

Increased expression of iNOS in the postural muscles CHIR-99021 purchase may act as a protective factor through HIF-1 alpha stabilization. Copyright (C) 2012 S. Karger AG, Basel”
“The medial amygdaloid nucleus (MeA) is a part of the limbic system and is involved in cardiovascular modulation. We previously reported that microinjection of noradrenaline (NA) into the MeA of unanesthetized rats caused

pressor and bradycardiac responses, which were mediated by acute vasopressin release into the systemic circulation. In the present study, we tested the possible involvement of magnocellular neurons of the paraventricular (PVN) and/or supraoptic (SON) of the hypothalamus that synthesize vasopressin in the cardiovascular pathway activated by the microinjection of NA into the MeA. Pressor

AZD4547 research buy and bradycardiac responses to the microinjection of NA (27 nmol/100 nL) into the MeA were blocked by pretreatment of either the PVN or the SON with cobalt chloride (CoCl2, 1 mM/100 nL), thus indicating that both hypothalamic nuclei mediate the cardiovascular responses evoked by microinjection of NA Into the MeA. Our results suggest that the pressor and bradycardiac response caused by the microinjection of NA into the MeA is mediated by magnocellular neurons in both the PVN and SON. (C) 2012 IBRO. Published by Elsevier Ltd. All rights reserved.”
“In the present study we investigated motor programming constraints implied by the Generalized Motor Program (GMP) view. A response precuing task was used in which participants performed aiming movements of either short or long duration to either a near or a far target position. Precues provided either no Levetiracetam advance information or partial information about extent or duration or fully specified the aiming movement. Reaction time (RT) decreased and

late Contingent Negative Variation (CNV) amplitude increased with the amount of advance information. In contrast to predictions of the GMP view, the extent precue led to faster responses and larger CNV amplitude than the duration precue. We conclude that late CNV amplitude reflects independent parameter specification processes at an abstract level at which GMP’s motor programming constraints do not apply.”
“Astrocytes contribute to neurotransmission through a variety of mechanisms ranging from synapse isolation to active signaling. Astroglial involvement in neurophysiology has been mostly investigated at the single-cell level. However, a unique feature of astrocytes is their high level of intercellular connectivity mediated by connexins, the proteins forming gap junction (GJ) channels. These astroglial GJ circuits enable the rapid intercellular exchange of ions, metabolites, and neuroactive substances.

Genetic association results at these loci were similar on univari

Genetic association results at these loci were similar on univariate and multivariate analysis. No candidate genes met the criteria for statistical significance.

Conclusions: Two single nucleotide polymorphisms, rs9810233 and

rs1920101, which are 25 kb apart, are associated with erectile dysfunction, although they do not meet the standard Acalabrutinib order genome-wide association study significance criterion of p < 5 x 10(-8). Other studies with larger sample sizes are required to determine whether ALCAM represents a novel gene in the pathogenesis of diabetes associated erectile dysfunction.”
“In this study, we analysed facial expression detection and facial mimicry behaviour in response to an emotional empathic task. We posited that a “”simulation mechanism”" may be related to emotional face detection and that this

mechanism click here could be supported by prefrontal cortical structures. To evaluate our hypothesis, we requested 16 subjects to emotionally empathise with facial stimuli and applied high frequency repeated transcranial magnetic stimulation (rTMS) to the medial prefrontal cortex (MPFC) in order to induce an increased response to the facial expressions of emotion. To determine whether emotional content affects empathic behaviour, we also varied the AZD6738 purchase stimulus valence (negative vs. positive vs. neutral faces). We found

that autonomic (facial zygomatic and corrugator electromyography (EMG) subjective response) and detection (correct responses (CRs); response times (RTs)) measures were modulated by the MPFC activity. Specifically, when prefrontal structures were activated, subjects displayed an increased performance in CRs and reduced RTs for face recognition, as well as increased emotion-specific EMG responses in comparison to sham effects and control site stimulation. In fact, the zygomatic muscle was more responsive in cases of positive emotion (happiness), whereas corrugator activity was more robust during periods of negative emotion (fear, anger, disgust). However, an overall stronger effect was observed in response to negative, and potentially aversive, faces when compared with responses to positive and neutral faces. Finally, a direct correlation was found between the psychophysiological responses and detection measures. In summary, these results suggest that a “”simulation mechanism”" underlies emotion detection in an empathic situation that includes both EMG and behavioural responses. This mechanism appears to be supported and regulated by the MPFC area. (C) 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

Materials and Methods: Eight patients with Parkinson’s disease an

Materials and Methods: Eight patients with Parkinson’s disease and detrusor overactivity refractory to anticholinergics were injected with

100 U botulinum toxin type A. Daytime and nighttime urinary frequency, and urinary incontinence episodes were selleck products recorded. Patients also completed a standardized quality of life questionnaire on incontinence and a visual analog scale on the impact of bladder problems on daily life activities, and underwent urodynamic assessment, including pressure flow studies. Clinical and urodynamic assessment was performed before, and 1, 3 and 6 months after injection.

Results: In all patients 100 U botulinum toxin type A induced decreased daytime and nighttime urinary frequency, a decreased number of

urinary incontinence episodes, increased quality of life scores and, as shown by increased maximum cystometric capacity, improved urodynamic findings. In 2 patients with Parkinson’s disease post-void residual urine volume developed.

Conclusions: Intradetrusor injection of 100 U botulinum toxin type A induced clinical and urodynamic improvement in overactive bladder symptoms that lasted at least 6 months in patients with Parkinson’s disease.”
“A 7-year-old girl with a history of recurrent urinary tract infection since the age of 3 years is known to have bilateral, moderately severe (grade III) vesicoureteral Elacridar reflux. Renal scintigraphy with technetium-99-labeled dimercaptosuccinic acid has revealed bilateral scarring in the upper poles of her kidneys, with more severe scarring on the left kidney than on the right. Despite ongoing antimicrobial prophylaxis, she has recently had another febrile urinary tract infection, which

responded most well to antibiotic treatment. Radionuclide cystography reveals persistent bilateral, moderately severe vesicoureteral reflux. The patient has no history of constipation or dysfunctional voiding. She is referred to a pediatric urologist, who discusses with the patient and her parents the various treatment options, including endoscopic correction.”
“The current research synthesis integrates the findings of 111 independent samples from 54 economically developing countries that examined the relation between economic status and subjective well-being (SWB). The average economic status-SWB effect size was strongest among low-income developing economies (r = .28) and for samples that were least educated (r = .36). The relation was weakest among high-income developing economies (r = .10) and for highly educated samples (r = .13). Controlling for numerous covariates, the partial r effect size remained significant for the least-educated samples (pr = .18).

Lyn inhibition or cholesterol depletion abrogated imatinib-induce

Lyn inhibition or cholesterol depletion abrogated imatinib-induced migration, and dual Src/Abl kinase inhibitor dasatinib induced fewer CML cells to migrate to the stroma. These findings demonstrate the novel mechanism of microenvironment-mediated resistance through lipid raft modulation, which involves compartmental changes of the multivalent CXCR4 and Lyn complex. We propose https://www.selleckchem.com/products/etomoxir-na-salt.html that pharmacological targeting of lipid rafts may eliminate bone marrow-resident

CML cells through interference with microenvironment-mediated resistance.”
“The fusiform face area, a high-level visual area, is pivotal in processing facial information. This area receives inputs from the left and right visual fields unlike the primary visual area, which only receives inputs from its contralateral visual field. Response of the fusiform face area to ipsilateral stimulation depends on the signals crossing over at the corpus callosum. We investigated the distribution of voxelwise activation to determine whether ipsilateral-dominant

voxels exist in the fusiform face area using high spatial resolution selleck chemical functional MRI at 7 T. We further examined the possible functional differences between ipsilateral-dominant and contralateral-dominant voxels. By unilateral visual field stimulation, we detected ipsilateral-dominant voxels in the right fusiform face area. Their distribution was spatially heterogeneous. We tested upright and inverted facial stimulation confined to unilateral visual fields and found that these ipsilateral-dominant voxels had a different functional nature from contralateral-dominant voxels. NeuroReport 24: 53-57 (C) 2013 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins. NeuroReport 2013, 24:53-57″
“Aim: To examine various potential factors that may predict early and overall mortality.

Design and Methods: We carried out an observational prospective study of a cohort of incident patients starting dialysis IWR-1 cost in a UK centre.

Univariate analysis of factors and co-morbidities potentially affecting survival on dialysis were analysed to potential predictors. Factors

affecting 1 year mortality were analysed using the t-test, the Mann-Whitney U-test or the chi-square test as appropriate. Mortality over the 5-year follow-up period was analysed using the Kaplan-Meier method.

Results: Ninety-four patients [predominantly Caucasian (98%)], of mean age 63 years (15.6) (56% > 65 years) with a slight male preponderance were studied. Vascular disease (39%) and sepsis (33%) accounted for most of the deaths and a significant proportion of mortality was seen in the first year (56%). Patients with early mortality were older (68 vs. 61 years, P = 0.05) with lower haemoglobin (8.4 vs. 9.4 g/dl, P = 0.01) at the start of dialysis, commenced dialysis with a lower eGFR (5.4 vs. 6.5 ml/min/1.73 m(2), P = 0.

The non-classical HMM showed

the highest rate in the clas

The non-classical HMM showed

the highest rate in the classification with values above 95% in training and 100% in test Although the higher accuracy of the HMM, the DIM showed simplicity for the RNase Ill classification with low computational cost. Such simplicity was evaluated in respect to HMM and ANN models for the functional annotation of a new bacterial RNase Ill class member, isolated and annotated by our group. (C) 2010 Elsevier Ltd. All rights reserved.”
“2-Chloronicotinic acid is receiving much attention for its effective applications as a key precursor in the synthesis of pesticides and medicines. In this study, a strain ZJB-09149 U0126 research buy converting 2-chloro-3-cyanopyridine to 2-chloronicotinic acid was newly isolated and identified as Rhodococcus erythropolis, based on its physiological and biological tests, and 16S rDNA sequence analysis. In addition, the effects of inducer, carbon source and nitrogen source were examined. Maximum activity was achieved when the above parameters were set as 8 g/l e-caprolactam, Pevonedistat 7 g/l yeast extract and 5 g/l maltose.

Moreover, the biotransformation pathway of 2-chloro-3-cyanopyridine to 2-chloronicotinic acid in strain ZJB-09149 was investigated as well. This study revealed that the nitrile hydratase (NHase) and amidase expressed in R. erythropolis ZJB-09149 are responsible for the conversion of 2-chloro-3-cyanopyridine. This is the first time to report on the biotransformation preparation of 2-chloronicotinic acid.”
“Stochastic resonance (SR) is a phenomenon occurring in nonlinear systems by which the ability to process information, for instance the detection of weak signals is statistically enhanced by a non-zero level of noise. SR effects have been observed in a great variety of systems, comprising selleck chemical electronic circuits, optical devices, chemical reactions and neurons. In this paper we report the SR phenomena occurring in the execution of an extremely simple behavioral rule inspired from bacteria chemotaxis. The phenomena are quantitatively analyzed by using Markov chain models and Monte Carlo simulations.

(C) 2011 Elsevier Ltd. All rights reserved.”
“Co-cultivation of mutant Penicillium oxalicum SAUE-3.510 and Pleurotus ostreatus MTCC 1804 was evaluated for the production of xylanase-laccase mixture under solid-state fermentation (SSF) condition. Growth compatibility between mutant P. oxalicum SAUE-3.510 and white rot fungi (P. ostreatus MTCC 1804, Trametes hirsuta MTCC 136 and Pycnoporus sp. MTCC 137) was analyzed by growing them on potato dextrose agar plate. Extracellular enzyme activities were determined spectrophotometrically. Under derived conditions, paired culturing of mutant P. oxalicum SAUE-3.510 and P. ostreatus MTCC 1804 resulted in 58% and 33% higher levels of xylanase and laccase production, respectively.

Adult male rats underwent

either an FPI or sham injury A

Adult male rats underwent

either an FPI or sham injury. Additional rats were only exposed to anesthesia. HPA regulation was evaluated by measuring the effects of dexamethasone (DEX) treatment on CORT and ACTH. Tail vein blood was collected following 30-min restraint stress, at post-injury days (PID) 1, 7 and 14, prior to (0 min) and at 30, 60, 90 and 120 min after stress onset. Results from these studies indicate that the stress response was significantly more pronounced after FPI in that CORT and ACTH restraint-induced increases were more pronounced and longer lasting compared to controls. DEX suppression of CORT and ACTH was observed in all groups, suggesting that stress hyper-responsiveness after mild FPI is not attributable to reduced sensitivity of CORT feedback regulation. The increased sensitivity to stressful events in the first two post-injury weeks after BIBW2992 a mild FPI may have a negative impact on early rehabilitative therapies. (c) 2011 IBRO.

Published by Elsevier Ltd. All rights reserved.”
“While endocannabinoid modulation of both GABAergic and glutamatergic synaptic transmission and plasticity has been extensively investigated, our understanding of the role of endocannabinoids in protecting neurons from harmful insults remains limited. 2-Arachidonoylglycerol (2-AG), the most abundant endogenous ligand and a full agonist selleck screening library for cannabinoid receptors, exhibits anti-inflammatory and neuroprotective effects via a CBI receptor RGFP966 research buy (CB1R)-mediated mechanism. However, it is still not clear whether 2-AG is also able to protect neurons from beta-amyloid (A beta)-induced neurodegeneration. Here, we demonstrate that exogenous application of 2-AG significantly protected hippocampal neurons in culture against A beta-induced neurodegeneration and apoptosis. This neuroprotective effect was blocked by SR141716 (SR-1), a selective CB1R antagonist,

but not by SR144528 (SR-2), a selective CB2R antagonist, or capsazepine (CAP), a selective transient receptor potential cation channels, subfamily V, member 1 (TRPV1) receptor antagonist. To determine whether endogenous 2-AG is capable of protecting neurons from A beta insults, hippocampal neurons in culture were treated with URB602 or JZL184, selective inhibitors of monoacylglycerol lipase (MAGL), the enzyme hydrolyzing 2-AG. MAGL inhibition that elevates endogenous levels of 2-AG also significantly reduced A beta-induced neurodegeneration and apoptosis. The 2-AG-produced neuroprotective effects appear to be mediated via CB1R-dependent suppression of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and nuclear factor-kappa B (NF-kappa B) phosphorylation and cyclooxygenase-2 (COX-2) expression. Our results suggest that elevation of endogenous 2-AG by inhibiting its hydrolysis has potential as a novel efficacious therapeutic approach for preventing, ameliorating or treating Alzheimer’s disease. (c) 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

The distributions of H3-K4 mono-, di-, and tri-methylation exhibi

The distributions of H3-K4 mono-, di-, and tri-methylation exhibited selleckchem exactly the same pattern as LSD1. LSD1 expression was induced both region and cell specifically after ischemic/perfusion, and complied with the two-peak mode of expression.

These studies revealed a tightly regulated distribution for LSD1 in the brain

of rats under ischemic insult, suggesting a critical role in neuron function. (C) 2010 Elsevier Ireland Ltd and the Japan Neuroscience Society All rights reserved.”
“The envelope of the human hepatitis B virus (HBV) contains three membrane proteins (L, M, and S). They accomplish different functions in HBV infectivity and nucleocapsid envelopment. Infectivity determinants have been assigned to the N-terminal part of the pre-S1 domain of the L protein and the antigenic loop of the S domain in the L and/or S protein. Nucleocapsid envelopment requires a C-terminal sequence within pre-S1, including

the five N-terminal amino acids of pre-S2 as part of the L protein. However, the role of the M protein and the pre-S2 domain of the L protein are not entirely understood. We addressed this question and analyzed assembly competence and infectivity of viruses that lack the M protein and, at the same click here time, carry alterations in the pre-S2 domain of L. These include deletions, in part frameshift mutations and a randomization of virtually the entire pre-S2 sequence. We found that the M protein is dispensable for HBV in vitro infectivity.

Viruses that lack the M protein and contain a mostly randomized pre-S2 sequence assemble properly and are infectious in HepaRG cells and primary human hepatocytes. While deletions of 20 amino acids in the pre-S2 domain of L protein allowed the production of infectious virions, more extended deletions interfered with assembly. This indicates that the pre-S2 domain of the L protein serves an important role for virus assembly, presumably as a spacer that supports conformational changes of L protein but does not participate as part of the M protein or as a subdomain of the L protein in virus entry.”
“The human cerebral neocortex is divided into BIBW2992 datasheet six layers consisting of specific neuronal cell types and connections To determine the distribution of cortical neurons during early development, we examined the expressions of layer-specific markers in human midterm fetal brains Layer V marker ZNF312 is expressed in most cortical areas, but not in the prospective somatosensory association area. Expression of layer IV marker ROR beta is also diminished in this region but Increased in the primary visual cortex, where expression of ZNF312 is reduced. Our results indicate that ZNF312 and other layer markers have area dependent expressions in the human fetal cortex (C) 2010 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved.”
“Bats are the second largest group of mammals on earth and act as reservoirs of many emerging viruses.

GB virus B (GBV-B), the virus phylogenetically most closely relat

GB virus B (GBV-B), the virus phylogenetically most closely related to HCV, causes hepatitis in tamarins. We have demonstrated the suitability of the tamarin as a host for GBV-B and as a surrogate nonhuman primate model for HCV infection, and we have initiated studies of GBV-B infection in a closely PRT062607 datasheet related species, the common marmoset (Callithrix jacchus). Here, we demonstrate that marmosets exhibit two phenotypes upon infection with GBV-B: the susceptible phenotype and the partially resistant phenotype. In addition, we identify changes that may correlate with adaptation of the virus to the partially resistant host. GBV-B

was serially passaged five times through 14 marmosets as one lineage and two times through 6 marmosets as a second lineage. Virus adapted to the marmosets and eventually exhibited robust infections in two separate lineages, lineages 1 and 2. A third lineage was initiated with a molecular clone, and again, susceptible and partially resistant phenotypes were observed. Three isolates were fully sequenced (from lineage 1), and 21 nucleotide changes were observed, with six amino acid changes. We speculate that the marmoset partially resistant phenotype may be due to a polymorphism in the marmoset population that affects critical virus-host interactions and that wild-type GBV-B is capable of rapidly adapting

to this altered host.”
“A mouse model of amyotrophic LY294002 lateral sclerosis-parkinsonism-dementia complex based on the consumption of cycad seed flour was used to determine whether the observed pathology of motor neuron loss begins in the distal axons or the spinal cord. Assessments of neuromuscular junction integrity and motor neurons were performed at multiple time points. Mice fed cycad pellets performed worse on the wire hang than controls. Microglial activation in cycad-fed mice was observed with motor neuron degeneration at 12 weeks, but reactive astrocyte proliferation was not observed. After 33 weeks of cycad feeding, motor neuron loss had stabilized, with no evidence of neuromuscular Selleck Bafilomycin A1 junction endplate denervation. These data suggest that neuronal pathology

begins at the soma and proceeds distally in a ‘dying forward’ pattern. NeuroReport 20:1284-1289 (C) 2009 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.”
“Viruses utilize host factors in many steps of their life cycles. Yet, little is known about host factors that contribute to the life cycle of hepatitis B virus (HBV), which replicates its genome by reverse transcription. To identify host factors that contribute to viral reverse transcription, we sought to identify cellular proteins that interact with HBV polymerase (Pol) by using affinity purification coupled with mass spectrometry. One of the HBV Pol-interacting host factors identified was DDX3 DEAD-box RNA helicase, which unwinds RNA in an ATPase-dependent manner.

The concentration of intracellular chloride in neurons is mainly

The concentration of intracellular chloride in neurons is mainly regulated by two cation-chloride cotransporters, the potassium-chloride cotransporter 2 (KCC2) and the sodium-potassium-chloride co-transporter 1 (NKCC1). In this study, we measured the reversal potential

of IPSPs (E(IPSP)) of lumbar motoneurons during the first postnatal BX-795 concentration week and we investigated the expression of KCC2 and NKCC1 in the ventral horn of the spinal cord from the embryonic day 17 to the postnatal day 20 in the rat. Our results suggest that the negative shift of E(IPSP) from above to below the resting membrane potential occurs during the first postnatal week when the expression of KCC2 increases significantly and the expression of NKCC1 decreases. KCC2 immunolabeling surrounded motoneurons, presumably in the plasma membrane and NKCC1 immunolabeling appeared outside this KCC2-labeled fine strip. Taken together, the present results indicate that maturation of chloride homeostasis is not completed at birth in the rat and that the upregulation of KCC2 plays a key role in the shift from depolarizing to hyperpolarizing IPSPs. (C) 2009 IBRO. Published by Elsevier Ltd. All rights reserved.”
“The

present study investigated a skill-level-dependent interaction between gravity and muscular force when striking piano keys. Kinetic AZD5363 in vitro analysis of the arm during the downswing motion performed by expert and novice piano players was made using an inverse dynamic technique. The corresponding activities of the elbow agonist and antagonist muscles were simultaneously recorded using electromyography (EMG). Muscular torque at the elbow joint was computed while excluding the effects of gravitational and motion-dependent interaction torques. During descending the forearm to strike the keys, the experts kept the activation of the triceps (movement agonist) muscle close to the resting level, and decreased anti-gravity activity of the biceps muscle across all loudness levels. This suggested that elbow extension

torque was produced by gravity without the contribution of agonist muscular work. For the novices, on the other hand, a distinct activity in the triceps muscle appeared during the middle of the downswing, and its amount and duration we e increased with increasing loudness. Therefore, for the novices, agonist muscular force was the GABA Receptor predominant contributor to the acceleration of elbow extension during the downswing. We concluded that a balance shift from muscular force dependency to gravity dependency for the generation of a target joint torque occurs with long-term piano training. This shift would support the notion of non-muscular force utilization for improving physiological efficiency of limb movement with respect to the effective use of gravity. (C) 2009 IBRO. Published by Elsevier Ltd. All rights reserved.”
“Objectives. Controversy exists over the optimal hospital type to which high-risk surgical patients should be referred for operative management.

Seroprevalence in humans was examined by screening sera from 30 a

Seroprevalence in humans was examined by screening sera from 30 adults and 17 young children for PVM-neutralizing activity. Sera from a single child (6%) and 40% of adults had low neutralizing activity against PVM, which could be consistent with increasing incidence of exposure following early childhood. There was no cross-reaction of human or AGM sera between RSV and PVM and no cross-protection in the mouse model. In native Western blots, human sera reacted with RSV but not PVM proteins under conditions in which AGM immune sera reacted strongly. Serum reactivity was further evaluated by flow cytometry using unfixed Vero cells infected with PVM or RSV expressing

green fluorescent protein (GFP) as a measure of viral gene expression. The reactivity of human sera against RSV-infected cells correlated with GFP expression, whereas reactivity against PVM-infected cells was low and LCZ696 in vivo S3I-201 uncorrelated with GFP expression. Thus, PVM specificity was not evident. Our results indicate that the PVM-neutralizing activity of human sera is not due to RSV- or PVM-specific antibodies but may be due to low-affinity, polyreactive natural antibodies of the IgG subclass. The absence of PVM-specific antibodies and restriction in nonhuman primates makes PVM unlikely to be a human pathogen.”
“In membrane protein biochemical

and structural studies, detergents are used to mimic membrane environment and maintain functional, stable conformation of membrane proteins in the absence of lipid bilayers. However, detergent concentration, esp. molar ratio of membrane protein to detergent

is usually unknown. Here, a gas chromatography-mass spectrometry selected ion monitoring (GC-MS-SIM) method was developed to quantify four detergents which are frequently used in membrane protein structural studies. To remove excessive detergents, a filtered centrifugation using Centricon tubes was applied. A membrane protein Ig-Beta fragment in four different detergent micelles was exemplified. Detergent concentrations in the upper and lower fraction of the Centricon tube were measured after each round of centrifugation. The results were very consistent to basic properties of detergent micelles in aqueous solvents. Therefore, coupling of GC-MS-SIM and detergent removal by Centricon Enzalutamide solubility dmso tubes, detergents concentration, esp. molar ratio of membrane protein to detergent could be controlled, which will expedite membrane protein structural and biochemical studies. (C) 2009 Elsevier Inc. All rights reserved.”
“BACKGROUND

Menorrhagia is a common problem, yet evidence to inform decisions about therapy is limited. In a pragmatic, multicenter, randomized trial, we compared the levonorgestrel-releasing intrauterine system (levonorgestrel-IUS) with usual medical treatment in women with menorrhagia who presented to their primary care providers.