Structural insights into biased G protein-coupled receptor signaling revealed by fluorescence spectroscopy

G protein-coupled receptors (GPCRs) are seven-transmembrane proteins that mediate most cellular responses to hormones and neurotransmitters, representing the largest group of therapeutic targets. Recent studies show that some GPCRs signal through both G protein and arrestin pathways in a ligand-specific manner. Ligands that direct signaling through a specific pathway are known as biased ligands. The arginine-vasopressin type 2 receptor (V2R), a prototypical peptide-activated GPCR, is an ideal model system to investigate the structural basis of biased signaling. Although the native hormone arginine-vasopressin leads to activation of both the stimulatory G protein (Gs) for the adenylyl cyclase and arrestin pathways, synthetic ligands exhibit highly biased signaling through either Gs alone or arrestin alone. We used purified V2R stabilized in neutral amphipols and developed fluorescence-based assays to investigate the structural basis of biased signaling for the V2R. Our studies demonstrate that the Gs-biased agonist stabilizes a conformation that is distinct from that stabilized by the arrestin-biased agonists. This study provides unique insights into the structural mechanisms of GPCR activation by biased ligands that may be relevant to the design of pathway-biased drugs.     

Understanding Transitions in Care from Hospital to Homeless Shelter: a Mixed-Methods, Community-Based Participatory Approach

BACKGROUND

Coordinated transitions from hospital to shelter for homeless patients may improve outcomes, yet patient-centered data to guide interventions are lacking.

OBJECTIVES

To understand patients?? experiences of transitions from hospital to a homeless shelter, and determine aspects of these experiences associated with perceived quality of these transitions.

DESIGNS

Mixed methods with a community-based participatory research approach, in partnership with personnel and clients from a homeless shelter.

PARTICIPANTS

Ninety-eight homeless individuals at a shelter who reported at least one acute care visit to an area hospital in the last year.

APPROACH

Using semi-structured interviews, we collected quantitative and qualitative data about transitions in care from the hospital to the shelter. We analyzed qualitative data using the constant comparative method to determine patients?? perspectives on the discharge experience, and we analyzed quantitative data using frequency analysis to determine factors associated with poor outcomes from patients?? perspective.

KEY RESULTS

Using qualitative analysis, we found homeless participants with a recent acute care visit perceived an overall lack of coordination between the hospital and shelter at the time of discharge. They also described how expectations of suboptimal coordination exacerbate delays in seeking care, and made three recommendations for improvement: 1) Hospital providers should consider housing a health concern; 2) Hospital and shelter providers should communicate during discharge planning; 3) Discharge planning should include safe transportation. In quantitative analysis of recent hospital experiences, 44?% of participants reported that housing status was assessed and 42?% reported that transportation was discussed. Twenty-seven percent reported discharge occurred after dark; 11?% reported staying on the streets with no shelter on the first night after discharge.

CONCLUSIONS

Homeless patients in our community perceived suboptimal coordination in transitions of care from the hospital to the shelter. These patients recommended improved assessment of housing status, communication between hospital and shelter providers, and arrangement of safe transportation to improve discharge safety and avoid discharge to the streets without shelter.     

Myelin specific cells infiltrate MCAO lesions and exacerbate stroke severity

Although inflammatory responses increase stroke severity, the role of immune cells specific for central nervous system (CNS) antigens remains controversial. Disruption of the blood–brain barrier (BBB) during stroke allows CNS antigens to leak into the peripheral circulation and enhances access of circulating leukocytes to the brain, including those specific for CNS antigens such as myelin oligodendrocyte glycoprotein (MOG) that can induce experimental autoimmune encephalomyelitis (EAE). We here demonstrate for the first time that myelin reactive splenocytes specific for MOG transferred into severe combined immunodeficient (SCID) mice can migrate into the infarct hemisphere of recipients subjected to 60 min middle cerebral artery occlusion (MCAO) and 96 h reperfusion; moreover these cells exacerbate infarct volume and worsen neurological deficits compared to animals transferred with na?ve splenocytes. These findings indicate that autoimmunity in the CNS can exert detrimental injury on brain cells and worsen the damage from ischemic stroke.     

Contribution of GPR30 for 1,25 dihydroxyvitamin D3 protection in EAE

Previous studies have demonstrated that vitamin D3-mediated protection in EAE occurs only in females and is dependent on the presence of diestrus levels of 17β-estradiol (E2). To evaluate the role of estrogen receptors in vitamin D3 treatment of EAE, we compared disease severity, CNS histopathology and immunological responses in vehicle and calcitrol (1,25 dihydroxyvitamin D₃) treated WT C57BL/6 mice vs. GPR30 membrane estrogen receptor (MER) knockout mice with MOG-35-55 peptide-induced EAE. Our results demonstrated that vitamin D₃-mediated prevention of clinical signs, CNS cellular lesions and demyelination observed in WT mice was abrogated in GPR30-KO mice with EAE. Regulatory effects of vitamin D₃ treatment that were MER dependent included increased levels of IL-10 and IL-6 secreted by MOG peptide-reactive splenocytes and increased expression of CCL5, CCR1 & CCR3 in spleen tissue. These results demonstrate for the first time that the MER is a key contributor to the E2-dependent effects of vitamin D₃-mediated protection in EAE.     

Sexual activity and function in women with and without pelvic floor disorders

Introduction and hypothesis

We describe differences in sexual activity and function in women with and without pelvic floor disorders (PFDs).

Methods

Heterosexual women ≥40 years of age who presented to either urogynecology or general gynecology clinics at 11 clinical sites were recruited. Women were asked if they were sexually active with a male partner. Validated questionnaires and Pelvic Organ Prolapse Quantification (POP-Q) examinations assessed urinary incontinence (UI), fecal incontinence (FI), and/or pelvic organ prolapse (POP). Sexual activity and function was measured by the Female Sexual Function Index (FSFI). Student’s t test was used to assess continuous variables; categorical variables were assessed with Fisher’s exact test and logistic regression. Univariate and multivariate analyses were used to assess the impact of pelvic floor disorders (PFDs) on FSFI total and domain scores.

Results

Five hundred and five women met eligibility requirements and gave consent for participation. Women with and without PFDs did not differ in race, body mass index (BMI), comorbid medical conditions, or hormone use. Women with PFDs were slightly older than women without PFDs (55.6?+?10.8 vs. 51.6?+?8.3 years, P <0.001); all analyses were controlled for age. Women with PFDs were as likely to be sexually active as women without PFDs (61.6 vs. 75.5 %, P?=?0.09). There was no difference in total FSFI scores between cohorts (23.2?+?8.5 vs. 24.4?+?9.2, P?=?0.23) or FSFI domain scores (all P?=?NS).

Conclusion

Rates of sexual activity and function are not different between women with and without PFDs.     

Platelet reactivity evaluated with the VASP assay following ticagrelor loading dose in acute coronary syndrome patients undergoing percutaneous coronary intervention

Background

The level of platelet reactivity (PR) inhibition obtained after P2Y12-ADP receptor antagonist loading dose (LD) is associated with the ischemic and bleeding risk following percutaneous coronary intervention (PCI) in acute coronary syndromes (ACS).

Objective

We aimed to evaluate the level of PR inhibition achieved by a 180 mg LD of ticagrelor and the rate of high on-treatment platelet reactivity (HTPR) in ACS patients undergoing PCI.

Methods

We performed a multicentre prospective observational study enrolling ACS patients undergoing PCI. Patients were included if they were admitted for ST-elevation myocardial infarction or non ST-elevation ACS. To assess PR, a VASP index was measured at least 6 and within 24 hours following a 180 mg LD of ticagrelor. HTPR was defined as a VASP index ≥ 50%.

Results

One hundred and fifteen patients were included: 31.3% of STEMI, 49.6% of NSTEMI and 19.1% of unstable angina. Following ticagrelor LD the mean VASP index was 17 ± 14%. However the response to ticagrelor was not uniform with a small inter-individual variability: inter quartile range: 7.6–22.8% and a rate of HTPR of 3.5%. A high number of patients, 65.6%, had a VASP index < 16%. None of the baseline characteristics of the study population was associated with PR. In addition, PR was similar in STEMI, NSTEMI and unstable angina (p = 0.9).

Conclusion

In ACS patients the level of PR inhibition achieved by a 180 mg loading dose of ticagrelor is not uniform and the rate of HTPR is 3.5%. A high proportion of patients exhibited a VASP index < 16%.     

Emotionally Focused Therapy: A Culturally Sensitive Approach for African American Heterosexual Couples

ABSTRACT

This article discusses unique considerations for a culturally sensitive emotionally focused therapy (EFT) for African American couples. EFT is an empirically supported treatment that significantly reduces relationship distress, yet efficacy with diverse populations remains unknown. African Americans’ chronic exposure to racism and discrimination creates a cumulative racial stress that negatively impacts their couple relationships. We propose a racially sensitive intervention for African American couples that addresses five considerations most salient for successful EFT treatment: the impact of racial stress, client trust of the therapist, gender-role expectations, and racial–gender views of emotionality and vulnerability. A case study illustrates this approach.     

Early psychotherapy process and cluster B and C personality pathology: similarities and differences in interactions with symptomatic and interpersonal distress

Abstract

In a prior study (Kolden & Klein, 1996), the authors found that the relationships between global personality pathology and early psychotherapy change processes (as defined by the Generic Model of Psychotherapy) were moderated by the extent of the patient's acute symptomatic and interpersonal distress. In the current study, the authors reanalyzed the same data to examine similarities and differences between personality disorder Clusters B (dramatic, emotional, or erratic) and C (anxious or fearful) in therapy process. In general, we found that more distressed patients reported greater defensiveness. There were no significant interactions between symptomatic distress and personality pathology in the prediction of any of the process variables. However, interpersonal distress moderated relationships between Clusters B and C and some therapy processes. Patients high in Cluster B felt more open and involved in the session when they were less distressed by their interpersonal problems at the start of therapy. In contrast, openness and insight were impeded among patients high in Cluster C when they were less distressed interpersonally. Therapists generally used more direct interventions and exploration of past experiences when working with patients higher in Cluster C pathology. However, therapists used direct interventions more specifically when patients with more severe Cluster B pathology were also higher in interpersonal distress. The discussion considers implications for the facilitation of productive early therapy process in patients suffering from Cluster B or C personality pathology.