The lipidome of posttraumatic stress disorder.

Posttraumatic stress disorder (PTSD) can develop after trauma exposure. Some studies report that women develop PTSD at twice the rate of men, despite greater trauma exposure in men. Lipids and their metabolites (lipidome) regulate a myriad of key biological processes and pathways such as membrane integrity, oxidative stress, and neuroinflammation in the brain by maintaining neuronal connectivity and homeostasis. In this study, we analyzed the lipidome of 40 individuals with PTSD and 40 trauma-exposed non-PTSD individuals. Plasma samples were analyzed for lipidomics using Quadrupole Time-of-Flight (QToF) mass spectrometry. Additionally, ~ 90 measures were collected, on sleep, mental and physical health indices. Sleep quality worsened as PTSD severity increased in both sexes. The lipidomics analysis identified a total of 348 quantifiable known lipid metabolites and 1951 lipid metabolites that are yet unknown; known metabolites were part of 13 classes of lipids. After adjusting for sleep quality, in women with PTSD, only one lipid subclass, phosphatidylethanolamine (PE) was altered, whereas, in men with PTSD, 9 out of 13 subclasses were altered compared to non-PTSD women and men, respectively. Severe PTSD was associated with 22% and 5% of altered lipid metabolites in men and women, respectively. Of the changed metabolites, only 0.5% measures (2 PEs and cholesterol) were common between women and men with PTSD. Several sphingomyelins, PEs, ceramides, and triglycerides were increased in men with severe PTSD. The triglycerides and ceramide metabolites that were most highly increased were correlated with cholesterol metabolites and systolic blood pressure in men but not always in women with PTSD. Alterations in triglycerides and ceramides are linked with cardiac health and metabolic function in humans. Thus, disturbed sleep and higher weight may have contributed to changes in the lipidome found in PTSD.

Immunological Misfiring and Sex Differences/Similarities in Early COVID-19 Studies: Missed Opportunities of Making a Real IMPACT.

COVID-19-associated intensive care unit (ICU) admissions were recognized as critical health issues that contributed to morbidity and mortality in SARS-CoV-2-infected patients. Severe symptoms in COVID-19 patients are often accompanied by cytokine release syndrome. Here, we analyzed publicly available data from the Yale IMPACT cohort to address immunological misfiring and sex differences in early COVID-19 patients. In 2020, SARS-CoV-2 was considered far more pathogenic and lethal than other circulating respiratory viruses, and the inclusion of SARS-CoV-2 negative patients in IMPACT cohorts confounds many findings. We ascertained the impact of several important biological variables such as days from symptom onset (DFSO); pre-existing risk factors, including obesity; and early COVID-19 treatments on significantly changed immunological measures in ICU-admitted COVID-19 patients that survived versus those that did not. Deceased patients had 19 unique measures that were not shared with ICU patients including increased granzyme-B-producing GzB+CD8+ T cells and interferon-γ. Male COVID-19 patients in ICU experienced many more changes in immunological and clinical measures than female ICU patients (25% vs. ~16%, respectively). A total of 13/124 measures including CCL5, CCL17, IL-18, IFNα2, Fractalkine, classical monocytes, T cells, and CD4Temra exhibited significant sex differences in female vs. male COVID-19 patients. A total of nine measures including IL-21, CCL5, and CD4Temra differed significantly between female and male healthy controls. Immunosuppressed patients experienced the most decreases in CD4Temra and CD8Tem cell numbers. None of the early COVID-19 treatments were effective in reducing levels of IL-6, a major component of the cytokine storm. Obesity (BMI >30) was the most impactful risk factor for COVID-19-related deaths and worst clinical outcomes. Our analysis highlights the contribution of biological sex, risk factors, and early treatments with respect to COVID-19-related ICU admission and progression to morbidity and mortality.

Regulation of Lung Immune Tone by the Gut-Lung Axis via Dietary Fiber, Gut Microbiota, and Short-Chain Fatty Acids.

Lung immune tone, i.e. the immune state of the lung, can vary between individuals and over a single individual's lifetime, and its basis and regulation in the context of inflammatory responses to injury is poorly understood. The gut microbiome, through the gut-lung axis, can influence lung injury outcomes but how the diet and microbiota affect lung immune tone is also unclear. We hypothesized that lung immune tone would be influenced by the presence of fiber-fermenting short-chain fatty acid (SCFA)-producing gut bacteria. To test this hypothesis, we conducted a fiber diet intervention study followed by lung injury in mice and profiled gut microbiota using 16S sequencing, metabolomics, and lung immune tone. We also studied germ-free mice to evaluate lung immune tone in the absence of microbiota and performed in vitro mechanistic studies on immune tone and metabolic programming of alveolar macrophages exposed to the SCFA propionate (C3). Mice on high-fiber diet were protected from sterile lung injury compared to mice on a fiber-free diet. This protection strongly correlated with lower lung immune tone, elevated propionate levels and enrichment of specific fecal microbiota taxa; conversely, lower levels of SCFAs and an increase in other fatty acid metabolites and bacterial taxa correlated with increased lung immune tone and increased lung injury in the fiber-free group. In vitro , C3 reduced lung alveolar macrophage immune tone (through suppression of IL-1ß and IL-18) and metabolically reprogrammed them (switching from glycolysis to oxidative phosphorylation after LPS challenge). Overall, our findings reveal that the gut-lung axis, through dietary fiber intake and enrichment of SCFA-producing gut bacteria, can regulate innate lung immune tone via IL-1ß and IL-18 pathways. These results provide a rationale for the therapeutic development of dietary interventions to preserve or enhance specific aspects of host lung immunity.

The Biological Basis of Sex Differences in Athletic Performance: Consensus Statement for the American College of Sports Medicine.

ABSTRACT: Biological sex is a primary determinant of athletic performance because of fundamental sex differences in anatomy and physiology dictated by sex chromosomes and sex hormones. Adult men are typically stronger, more powerful, and faster than women of similar age and training status. Thus, for athletic events and sports relying on endurance, muscle strength, speed, and power, males typically outperform females by 10%-30% depending on the requirements of the event. These sex differences in performance emerge with the onset of puberty and coincide with the increase in endogenous sex steroid hormones, in particular testosterone in males, which increases 30-fold by adulthood, but remains low in females. The primary goal of this consensus statement is to provide the latest scientific knowledge and mechanisms for the sex differences in athletic performance. This review highlights the differences in anatomy and physiology between males and females that are primary determinants of the sex differences in athletic performance and in response to exercise training, and the role of sex steroid hormones (particularly testosterone and estradiol). We also identify historical and nonphysiological factors that influence the sex differences in performance. Finally, we identify gaps in the knowledge of sex differences in athletic performance and the underlying mechanisms, providing substantial opportunities for high-impact studies. A major step toward closing the knowledge gap is to include more and equitable numbers of women to that of men in mechanistic studies that determine any of the sex differences in response to an acute bout of exercise, exercise training, and athletic performance.

Unraveling corticotropin-releasing factor family-orchestrated signaling and function in both sexes.

Stress responses to physical, psychological, environmental, or cellular stressors, has two arms: initiation and recovery. Corticotropin-releasing factor (CRF) is primarily responsible for regulating and/or initiating stress responses via, whereas urocortins (UCNs) are involved in the recovery response to stress via feedback inhibition. Stress is a loaded, polysemous word and is experienced in a myriad of ways. Some stressors are good for an individual, in fact essential, whereas other stressors are associated with bad outcomes. Perceived stress, like beauty, lies in the eye of the beholder, and hence the same stressor can result in individual-specific outcomes. In mammals, there are two main biological sexes with reproduction as primary function. Reproduction and nutrition can also be viewed as stressors; based on a body of work from my laboratory, we propose that the functions of all other organs have co-evolved to optimize and facilitate an individual's nutritional and reproductive functions. Hence, sex differences in physiologically relevant outcomes are innate and occur at all levels- molecular, endocrine, immune, and (patho)physiological. CRF and three UCNs are peptide hormones that mediate their physiological effects by binding to two known G protein-coupled receptors (GPCRs), CRF1 and CRF2. Expression and function of CRF family of hormones and their receptors is likely to be sexually dimorphic in all organs. In this chapter, based on the large body of work from others and my laboratory, an overview of the CRF family with special emphasis on sex-specific actions of peripherally expressed CRF2 receptor in health and disease is provided.

Impact of sex and comorbid diabetes on hospitalization outcomes in acute pancreatitis: A large United States population-based study.

Backgrounds: Data on the association between comorbid diabetes mellitus (DM) and acute pancreatitis (AP) remains limited. Utilizing a large, nationwide database, we aimed to examine the impact of comorbid diabetes mellitus on patients admitted for acute pancreatitis. Methods: This was a retrospective case-control study of adult patients with AP utilizing the National Inpatient Sample from 2015-2018, using ICD-10 codes. Hospitalization outcomes of patients admitted for AP with comorbid DM were compared to those without comorbid DM at the time of admission. The primary outcome was a mortality difference between the cohorts. Multivariable-adjusted cox proportional hazards model analysis was performed. Data was analyzed as both sex aggregated, and sex segregated. Results: 940,789 adult patients with AP were included, of which 256,330 (27.3%) had comorbid DM. Comorbid DM was associated with a 31% increased risk of inpatient mortality (aOR: 1.31; p = 0.004), a 53% increased risk of developing sepsis (aOR: 1.53; p = 0.002), increased hospital length of stay (LOS) (4.5 days vs. 3.7 days; p < 0.001), and hospital costs ($9934 vs. $8486; p < 0.001). Whites admitted for AP with comorbid DM were at a 49% increased risk of mortality as compared to Hispanics (aOR: 1.49; p < 0.0001). Different comorbidities had sex-specific risks; men admitted for AP with comorbid DM were at a 28% increased risk of mortality (aOR: 1.28; p < 0.0001) as compared to women. Men with comorbid DM plus obesity or hypertension were also at increased risk of mortality as compared to women, whereas women with comorbid DM plus renal failure were at greater risk of mortality as compared to men. Conclusions: Comorbid DM appears to be a risk factor for adverse hospitalization outcomes in patients admitted for AP with male sex and race as additional risk factors. Future prospective studies are warranted to confirm these findings to better risk stratify this patient population.

Organ-Specific Glucose Uptake: Does Sex Matter?

Glucose uptake by peripheral organs is essential for maintaining blood glucose levels within normal range. Impaired glucose uptake is a hallmark of type 2 diabetes (T2D) and metabolic syndrome and is characterized by insulin resistance. Male sex is an independent risk factor for the development of T2D. We tested whether sex and diet are independent variables for differential glucose uptake by various organs. Here, in a longitudinal study, we used 18F-fluorodeoxyglucose (FDG) and positron emission tomography (PET) to determine baseline differences in whole-body glucose uptake in young male and female mice on chow and high-fat diets. We report that sex and diet are important independent variables that account for differential glucose uptake in brown fat, skeletal muscle, liver, heart, kidney, and the stomach, but not the brain, lungs, pancreas, small intestine, or perigonadal adipose. Of the seven organs analyzed, two organs, namely brown fat, and the heart had the highest concentrations of FDG, followed by the brain, kidneys, and skeletal muscle on chow diet. Young female mice had 47% greater FDG uptake in the brown fat compared to male mice, whereas skeletal muscle FDG uptake was 49% greater in male mice. The high-fat diet inhibited FDG uptake in brown fat, skeletal muscle, and the heart, three major organs involved in uptake, whereas brain uptake was enhanced in both sexes. These foundational and groundbreaking findings suggest that mechanisms of glucose homeostasis are context- and organ-dependent and highlight the need to study sex-specific outcomes and mechanisms for diseases such as T2D, obesity, and metabolic syndrome.

Neuron-Glia-Immune Triad and Cortico-Limbic System in Pathology of Pain.

Pain is an unpleasant sensation that alerts one to the presence of obnoxious stimuli or sensations. These stimuli are transferred by sensory neurons to the dorsal root ganglia-spinal cord and finally to the brain. Glial cells in the peripheral nervous system, astrocytes in the brain, dorsal root ganglia, and immune cells all contribute to the development, maintenance, and resolution of pain. Both innate and adaptive immune responses modulate pain perception and behavior. Neutrophils, microglial, and T cell activation, essential components of the innate and adaptive immune responses, can play both excitatory and inhibitory roles and are involved in the transition from acute to chronic pain. Immune responses may also exacerbate pain perception by modulating the function of the cortical-limbic brain regions involved in behavioral and emotional responses. The link between an emotional state and pain perception is larger than what is widely acknowledged. In positive psychological states, perception of pain along with other somatic symptoms decreases, whereas in negative psychological states, these symptoms may worsen. Sex differences in mechanisms of pain perception are not well studied. In this review, we highlight what is known, controversies, and the gaps in this field.

Low-density lipoprotein receptor-related protein 1 (LRP1) is a novel receptor for apolipoprotein A4 (APOA4) in adipose tissue.

Apolipoprotein A4 (APOA4) is one of the most abundant and versatile apolipoproteins facilitating lipid transport and metabolism. APOA4 is synthesized in the small intestine, packaged onto chylomicrons, secreted into intestinal lymph and transported via circulation to several tissues, including adipose. Since its discovery nearly 4 decades ago, to date, only platelet integrin αIIbß3 has been identified as APOA4 receptor in the plasma. Using co-immunoprecipitation coupled with mass spectrometry, we probed the APOA4 interactome in mouse gonadal fat tissue, where ApoA4 gene is not transcribed but APOA4 protein is abundant. We demonstrate that lipoprotein receptor-related protein 1 (LRP1) is the cognate receptor for APOA4 in adipose tissue. LRP1 colocalized with APOA4 in adipocytes; it interacted with APOA4 under fasting condition and their interaction was enhanced during lipid feeding concomitant with increased APOA4 levels in plasma. In 3T3-L1 mature adipocytes, APOA4 promoted glucose uptake both in absence and presence of insulin in a dose-dependent manner. Knockdown of LRP1 abrogated APOA4-induced glucose uptake as well as activation of phosphatidylinositol 3 kinase (PI3K)-mediated protein kinase B (AKT). Taken together, we identified LRP1 as a novel receptor for APOA4 in promoting glucose uptake. Considering both APOA4 and LRP1 are multifunctional players in lipid and glucose metabolism, our finding opens up a door to better understand the molecular mechanisms along APOA4-LRP1 axis, whose dysregulation leads to obesity, cardiovascular disease, and diabetes.

Stress and corticotropin releasing factor (CRF) promote necrotizing enterocolitis in a formula-fed neonatal rat model.

The etiology of necrotizing enterocolitis (NEC) is not known. Alterations in gut microbiome, mucosal barrier function, immune cell activation, and blood flow are characterized events in its development, with stress as a contributing factor. The hormone corticotropin-releasing factor (CRF) is a key mediator of stress responses and influences these aforementioned processes. CRF signaling is modulated by NEC's main risk factors of prematurity and formula feeding. Using an established neonatal rat model of NEC, we tested hypotheses that: (i) increased CRF levels-as seen during stress-promote NEC in formula-fed (FF) newborn rats, and (ii) antagonism of CRF action ameliorates NEC. Newborn pups were formula-fed to initiate gut inflammation and randomized to: no stress, no stress with subcutaneous CRF administration, stress (acute hypoxia followed by cold exposure-NEC model), or stress after pretreatment with the CRF peptide antagonist Astressin. Dam-fed unstressed and stressed littermates served as controls. NEC incidence and severity in the terminal ileum were determined using a histologic scoring system. Changes in CRF, CRF receptor (CRFRs), and toll-like receptor 4 (TLR4) expression levels were determined by immunofluorescence and immunoblotting, respectively. Stress exposure in FF neonates resulted in 40.0% NEC incidence, whereas exogenous CRF administration resulted in 51.7% NEC incidence compared to 8.7% in FF non-stressed neonates (p<0.001). Astressin prevented development of NEC in FF-stressed neonates (7.7% vs. 40.0%; p = 0.003). CRF and CRFR immunoreactivity increased in the ileum of neonates with NEC compared to dam-fed controls or FF unstressed pups. Immunoblotting confirmed increased TLR4 protein levels in FF stressed (NEC model) animals vs. controls, and Astressin treatment restored TLR4 to control levels. Peripheral CRF may serve as specific pharmacologic target for the prevention and treatment of NEC.

Sex Differences in the Exocrine Pancreas and Associated Diseases.

Differences in pancreatic anatomy, size, and function exist in men and women. The anatomical differences could contribute to the increase in complications associated with pancreatic surgery in women. Although diagnostic criteria for pancreatitis are the same in men and women, major sex differences in etiology are reported. Alcohol and tobacco predominate in men, whereas idiopathic and obstructive etiologies predominate in women. Circulating levels of estrogens, progesterone, and androgens contribute significantly to overall health outcomes; premenopausal women have lower prevalence of cardiovascular and pancreatic diseases suggesting protective effects of estrogens, whereas androgens promote growth of normal and cancerous cells. Sex chromosomes and gonadal and nongonadal hormones together determine an individual's sex, which is distinct from gender or gender identity. Human pancreatic disease etiology, outcomes, and sex-specific mechanisms are largely unknown. In rodents of both sexes, glucocorticoids and estrogens from the adrenal glands influence pancreatic secretion and acinar cell zymogen granule numbers. Lack of corticotropin-releasing factor receptor 2 function, a G protein-coupled receptor whose expression is regulated by both estrogens and glucocorticoids, causes sex-specific changes in pancreatic histopathology, zymogen granule numbers, and endoplasmic reticulum ultrastructure changes in acute pancreatitis model. Here, we review existing literature on sex differences in the normal exocrine pancreas and mechanisms that operate at homeostasis and diseased states in both sexes. Finally, we review pregnancy-related pancreatic diseases and discuss the effects of sex differences on proposed treatments in pancreatic disease.

Human Placenta Buffers the Fetus from Adverse Effects of Perceived Maternal Stress.

Maternal stress during pregnancy is linked to several negative birth outcomes. The placenta, a unique pregnancy-specific organ, not only nourishes and protects the fetus but is also the major source of progesterone and estrogens. As the placenta becomes the primary source of maternal progesterone (P4) and estradiol between 6-9 weeks of gestation, and these hormones are critical for maintaining pregnancy, maternal stress may modulate levels of these steroids to impact birth outcomes. The objective was to test whether maternal perceived stress crosses the placental barrier to modulate fetal steroids, including cortisol, which is a downstream indicator of maternal hypothalamic-pituitary-adrenal (HPA) axis regulation and is associated with negative fetal outcomes. Nulliparous women, 18 years or older, with no known history of adrenal or endocrine illness were recruited during their third trimester of pregnancy at the University of California San Francisco (UCSF) Mission Bay hospital obstetrics clinics. Simultaneous measurement of 10 steroid metabolites in maternal (plasma and hair) and fetal (cord blood and placenta) samples was performed using tandem mass spectrometry along with assessment of the perceived stress score and sociodemographic status. While the maternal perceived stress score (PSS) and sociodemographic status were positively associated with each other and each with the body mass index (BMI) (r = 0.73, p = 0.0008; r = 0.48, p = 0.05; r = 0.59, p = 0.014, respectively), PSS did not correlate with maternal or fetal cortisol, cortisone levels, or fetal birth weight. Regardless of maternal PSS or BMI, fetal steroid levels remained stable and unaffected. Progesterone was the only steroid analyte quantifiable in maternal hair and correlated positively with PSS (r = 0.964, p = 0.003), whereas cord estradiol was negatively associated with PSS (r = -0.94, p = 0.017). In conclusion, hair progesterone might serve as a better marker of maternal stress than cortisol or cortisone and maternal PSS negatively impacts fetal estradiol levels. Findings have implications for improved biomarkers of stress and targets for future research to identify factors that buffer the fetus from adverse effects of maternal stress.

Considering Sex as a Biological Variable in Basic and Clinical Studies: An Endocrine Society Scientific Statement.

In May 2014, the National Institutes of Health (NIH) stated its intent to "require applicants to consider sex as a biological variable (SABV) in the design and analysis of NIH-funded research involving animals and cells." Since then, proposed research plans that include animals routinely state that both sexes/genders will be used; however, in many instances, researchers and reviewers are at a loss about the issue of sex differences. Moreover, the terms sex and gender are used interchangeably by many researchers, further complicating the issue. In addition, the sex or gender of the researcher might influence study outcomes, especially those concerning behavioral studies, in both animals and humans. The act of observation may change the outcome (the "observer effect") and any experimental manipulation, no matter how well-controlled, is subject to it. This is nowhere more applicable than in physiology and behavior. The sex of established cultured cell lines is another issue, in addition to aneuploidy; chromosomal numbers can change as cells are passaged. Additionally, culture medium contains steroids, growth hormone, and insulin that might influence expression of various genes. These issues often are not taken into account, determined, or even considered. Issues pertaining to the "sex" of cultured cells are beyond the scope of this Statement. However, we will discuss the factors that influence sex and gender in both basic research (that using animal models) and clinical research (that involving human subjects), as well as in some areas of science where sex differences are routinely studied. Sex differences in baseline physiology and associated mechanisms form the foundation for understanding sex differences in diseases pathology, treatments, and outcomes. The purpose of this Statement is to highlight lessons learned, caveats, and what to consider when evaluating data pertaining to sex differences, using 3 areas of research as examples; it is not intended to serve as a guideline for research design.

Vagal gut-brain signaling mediates amygdaloid plasticity, affect, and pain in a functional dyspepsia model.

Functional dyspepsia (FD) is associated with chronic gastrointestinal distress and with anxiety and depression. Here, we hypothesized that aberrant gastric signals, transmitted by the vagus nerve, may alter key brain regions modulating affective and pain behavior. Using a previously validated rat model of FD characterized by gastric hypersensitivity, depression-like behavior, and anxiety-like behavior, we found that vagal activity - in response to gastric distention - was increased in FD rats. The FD phenotype was associated with gastric mast cell hyperplasia and increased expression of corticotrophin-releasing factor (Crh) and decreased brain-derived neurotrophic factor genes in the central amygdala. Subdiaphragmatic vagotomy reversed these changes and restored affective behavior to that of controls. Vagotomy partially attenuated pain responses to gastric distention, which may be mediated by central reflexes in the periaqueductal gray, as determined by local injection of lidocaine. Ketotifen, a mast cell stabilizer, reduced vagal hypersensitivity, normalized affective behavior, and attenuated gastric hyperalgesia. In conclusion, vagal activity, partially driven by gastric mast cells, induces long-lasting changes in Crh signaling in the amygdala that may be responsible for enhanced pain and enhanced anxiety- and depression-like behaviors. Together, these results support a "bottom-up" pathway involving the gut-brain axis in the pathogenesis of both gastric pain and psychiatric comorbidity in FD.

Effects of stress-related peptides on chloride secretion in the mouse proximal colon.

BACKGROUND: Stress increases intestinal secretion and exacerbates symptoms of irritable bowel syndrome (IBS). Peripherally derived corticotropin-releasing factor (CRF) is known to mediate stress-induced intestinal secretion, presumably by activation of CRF1 receptors in the gut. The present study aimed to ascertain the role of CRF2 activation in intestinal secretion by three other members of CRF peptide family, urocortin (UCN) 1-3, in wild type (WT) and CRF2 knockout (Crhr2-/- ) mice. METHODS: Mucosal/submucosal preparations from proximal colon of WT and Crhr2-/- mice of both sexes were mounted in Ussing chambers for measurement of short-circuit current (Isc ) as an indicator of ion secretion. KEY RESULTS: Male mice demonstrated a significantly higher baseline Isc than female in both WT and Crhr2-/- genotypes. CRF and UCN1-3 (1 µM) caused greater increases in colonic Isc (ΔIsc ) in male than female. Colonic Isc response to the selective CRF1 agonist, stressin1, was similar in both sexes. In male mice, the selective CRF2 agonists (UCN2 and UCN3) caused significantly greater ΔIsc than CRF and stressin1. UCN2- and UCN3-evoked ΔISC was significantly reduced in preparations pretreated with the selective CRF2 antagonist antisauvagine-30 and in Crhr2-/- mice. The prosecretory effects of urocortins were due to increases in Cl- secretion and involved enteric neurons and mast cells. CONCLUSIONS AND INFERENCE: The findings revealed sex differences in baseline colonic secretion and responses to stress-related peptides. CRF2 receptors play a more prominent role in colonic secretion in male mice. The greater baseline secretion and responses to UCNs may contribute to the higher prevalence of diarrhea-predominant IBS in males.

Corticotropin-Releasing Factor Family: A Stress Hormone-Receptor System's Emerging Role in Mediating Sex-Specific Signaling.

No organ in the body is impervious to the effects of stress, and a coordinated response from all organs is essential to deal with stressors. A dysregulated stress response that fails to bring systems back to homeostasis leads to compromised function and ultimately a diseased state. The components of the corticotropin-releasing factor (CRF) family, an ancient and evolutionarily conserved stress hormone-receptor system, helps both initiate stress responses and bring systems back to homeostasis once the stressors are removed. The mammalian CRF family comprises of four known agonists, CRF and urocortins (UCN1-3), and two known G protein-coupled receptors (GPCRs), CRF1 and CRF2. Evolutionarily, precursors of CRF- and urocortin-like peptides and their receptors were involved in osmoregulation/diuretic functions, in addition to nutrient sensing. Both CRF and UCN1 peptide hormones as well as their receptors appeared after a duplication event nearly 400 million years ago. All four agonists and both CRF receptors show sex-specific changes in expression and/or function, and single nucleotide polymorphisms are associated with a plethora of human diseases. CRF receptors harbor N-terminal cleavable peptide sequences, conferring biased ligand properties. CRF receptors have the ability to heteromerize with each other as well as with other GPCRs. Taken together, CRF receptors and their agonists due to their versatile functional adaptability mediate nuanced responses and are uniquely positioned to orchestrate sex-specific signaling and function in several tissues.

Protective effects of urocortin 2 against caerulein-induced acute pancreatitis.

Because little is known about the role of corticotropin-releasing factor (CRF) agonists in regulating responses in pancreatitis, we evaluated the effects of urocortin 2 (UCN2) and stressin1 in caerulein-induced acute pancreatitis (AP) model in rats. Male rats were pretreated with UCN2 or stressin1 for 30 min followed by induction of AP with supraphysiologic doses of caerulein. Serum amylase and lipase activity, pancreatic tissue necrosis, immune cell infiltrate, nuclear factor (NF)-κB activity, trypsin levels, and intracellular Ca2+ ([Ca2+]i) were ascertained. UCN2, but not stressin1 attenuated the severity of AP in rats. UCN2, but not stressin1, reduced serum amylase and lipase activity, cell necrosis and inflammatory cell infiltration in AP. NF-κB activity in pancreatic nuclear extracts increased in AP and UCN2 treatment reduced caerulein-induced increases in NF-κB activity by 42%. UCN2 treatment prevented caerulein-induced degradation of IκB-α in the cytosolic fraction as well as increased levels of p65 subunit of NF-κB in the cytosolic fraction. Pancreatic UCN2 levels decreased in AP compared with saline. UCN2 evoked [Ca2+]i responses in primary acinar cells and abolished caerulein-evoked [Ca2+]i responses at 0.1nM, and decreased by ~50% at 1.0nM caerulein. UCN2 stimulation resulted in redistribution of a portion of F-actin from the apical to the basolateral pole. UCN2 prevented the massive redistribution of F-actin observed with supraphysiologic doses of caerulein. UCN2, but not stressin1 attenuated severity of an experimental pancreatitis model. The protective effects of UCN2, including anti-inflammatory and anti-necrotic effects involve activation of the CRF2 receptor, [Ca2+]i signaling, and inhibition of NF-κB activity.

Apolipoprotein A-IV: A Multifunctional Protein Involved in Protection against Atherosclerosis and Diabetes.

Apolipoprotein A-IV (apoA-IV) is a lipid-binding protein, which is primarily synthesized in the small intestine, packaged into chylomicrons, and secreted into intestinal lymph during fat absorption. In the circulation, apoA-IV is present on chylomicron remnants, high-density lipoproteins, and also in lipid-free form. ApoA-IV is involved in a myriad of physiological processes such as lipid absorption and metabolism, anti-atherosclerosis, platelet aggregation and thrombosis, glucose homeostasis, and food intake. ApoA-IV deficiency is associated with atherosclerosis and diabetes, which renders it as a potential therapeutic target for treatment of these diseases. While much has been learned about the physiological functions of apoA-IV using rodent models, the action of apoA-IV at the cellular and molecular levels is less understood, let alone apoA-IV-interacting partners. In this review, we will summarize the findings on the molecular function of apoA-IV and apoA-IV-interacting proteins. The information will shed light on the discovery of apoA-IV receptors and the understanding of the molecular mechanism underlying its mode of action.

Probing the Interactome of Corticotropin-Releasing Factor Receptor Heteromers Using Mass Spectrometry.

Mass spectrometry is a sensitive technique used in the field of proteomics that allows for simultaneous detection and characterization of several proteins in a sample. It is also a powerful methodology to elucidate protein-protein interactions in a sequence-dependent and unbiased manner. G protein-coupled receptors (GPCRs) seldom function in isolation and characterization of proteins present in the receptor complex (or its interactome) is critical for understanding the vast spectrum of functions these receptors perform in a context-dependent manner. Here, we describe a mass spectrometry-based method to sequence and characterize proteins present in heteromeric complexes formed by corticotropin-releasing factor (CRF) receptors that belong to class B GPCRs. CRF receptor heteromeric complexes were identified in HEK293 cells co-transfected with tagged CRF receptors 1 and 2. CRF receptors were immunoprecipitated using antibodies against the tags from transfected HEK293 cells and proteins in their interactome were identified using liquid chromatography mass spectrometry method (LC-MS/MS). Both CRF receptors were identified in this interactome. A few of the proteins identified in the CRF receptor interactome using MS were confirmed to be true interactions using traditional co-immunoprecipitation and Western blotting methods.