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Background: Platelet transfusion is considered the standard treatment for preventing or controlling severe haemorrhage in Glanzmann thrombasthenia (GT). However, platelet transfusion can have detrimental effects, including the production of anti-GPIIb/IIIa isoantibodies or anti-HLA antibodies (Ab) and platelet transfusion refractoriness. Recombinant activated factor VII (rFVIIa) has been proposed as an alternative treatment to platelet transfusion., Methods: We analyzed data from 77 case reports including 100 subjects to investigate the effectiveness and safety of rFVIIa in combination with platelets or antifibrinolytics for preventing or treating non-surgical bleeds, as well as surgical and obstetrical procedures in GT., Results: The dosage of rFVIIa was consistent with previous recommendations (90 µg/kg per infusion). In subjects without Ab/refractoriness (n = 56), rFVIIa was effective in managing 93% of non-surgical bleeds (n = 42), 91% of minor (n = 11) and 92% of major (n = 26) surgical procedures and 89% of obstetrical procedures (n = 9). In subjects with Ab/refractoriness (n = 44), rFVIIa was effective in managing 90% of non-surgical bleeds (n = 39), and 75% of minor (n = 12) and 100% of major (n = 17) surgical procedures. The use of rFVIIa was safe, with 4 (2.7%) serious adverse events associated with rFVIIa., Conclusion: Although the use of rFVIIa is currently restricted to subjects with Ab/refractoriness or when platelets are not available, our findings suggest expanding the indications for rFVIIa to encompass GT without Ab/refractoriness. Frontline use of rFVIIa may be proposed when clinically possible to mitigate the risks associated with platelet transfusion., (© 2024 The Author(s). Haemophilia published by John Wiley & Sons Ltd.)

Dementia is frequent in nursing home (NH), affecting between 40 and 50 % of the residents and is associated in 90 % of the cases with behavioral and psychological symptoms (BPSD) which are detrimental to caregivers' working life and perceived health. We compared perceived health and working life between NHs that used or did not use nighttime personalized non-pharmacological therapies (PNPT) to address BPSD. This anonymous survey included 96 caregivers from 25 NHs which used (n = 69) or not (n = 27) nighttime PNPT. Perceived health, nursing and medical care, quality of care and internal motivation were significantly higher, stress symptoms and workload were significantly lower and there was a tendency to enhanced satisfaction with work in caregivers from NHs which used nighttime PNPT. Our present data open a new window on the benefits of PNPT., Competing Interests: Declaration of competing interest None., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective: This study described older patients receiving hospitalisation-at-home (HaH) services and identified factors associated with 30-day hospital readmission., Design: 3-year retrospective study in 2017-2019 in France., Participants: 75 108 patients aged 75 years and older who were discharged from hospital medical wards (internal medicine and geriatric units) and admitted to HaH., Primary Outcome Measure: 30-day hospital readmission., Results: The mean age of patients was 83.4 years (SD 5.7), 52.3% were male and 88.4% lived in a private household. Patients were primarily discharged from the internal medicine unit (85.3%). The top four areas of care in the HaH were palliative care, complex dressing, intravenous therapy and complex nursing care. Overall, 23.5% of patients died during their HaH stay and 27.8% were readmitted to the hospital at 30 days. In the multivariate model, male (OR 1.19, 95% CI 1.16 to 1.23), supportive cancer HaH care (OR 1.78, 95% CI 1.51 to 2.11) and very high intensity care during the previous in-person hospitalisation (OR 1.45, 95% CI 1.34 to 1.57) increased the risk of hospital readmission at 30 days. Older age (OR 0.97, 95% CI 0.97 to 0.98), living in a nursing home (OR 0.51, 95% CI 0.48 to 0.54), postsurgery HaH care (OR 0.49, 95% CI 0.41 to 0.58) and having been previously hospitalised in a geriatric unit (OR 0.81, 95% CI 0.77 to 0.85) decreased the risk of hospital readmission at 30 days., Conclusions: HaH provides complex care to very old patients, which is associated with high mortality. Several factors are associated with rehospitalisation within 30 days that could be avoided with better integration of different services with higher geriatric skills., Trial Registration Number: CNIL:2228861., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2023. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Hypophosphatemia is a recognized side effect of treatment of iron deficiency anemias with injectable iron. We analyzed 35 clinical trials that used ferric carboxymaltose (FCM) or iron sucrose (IS). Hypophosphatemia prevalence ranged from 0 to 91.7%. FCM-induced a significant (P<0.001) greater hypophosphatemia prevalence and phosphatemia decrease than IS (52.0% [95% CI: 42.2-61.8%] vs. 7.7% [95% CI: -2.8 to 18.2%] and -1.12mmol/L [95% CI: -1.36 to -0.89mmol/L] vs. -0.13mmol/L [95% CI: -0.59 to 0.32mmol/L]). FCM-induced hypophosphatemia was dose-dependent. The nadir of hypophosphatemia was reached in almost all studies after 7 and 14days. Hypophosphatemia persisted at the end of the study in 53.8% of the reported studies that used FCM and lasted up to 6months. FCM-induced an increase in intact circulating fibroblast growth factor 23 and in renal phosphorus excretion while serum 1-25 dihydroxyvitamin D was decreased. Risk factors for hypophosphatemia after FCM therapy were low basal circulating phosphate or ferritin, low body weight, high glomerular filtration rate, serum parathyroid hormone or hemoglobin and age, whereas renal insufficiency was associated with a lower risk. In conclusion, hypophosphatemia is common after treatment with injectable iron, FCM being associated with a higher risk than IS and with disorders of phosphocalcium metabolism. Monitoring of blood phosphate and 1-25 dihydroxyvitamin D could be considered during FCM therapy., (Copyright © 2023 Académie Nationale de Pharmacie. Published by Elsevier Masson SAS. All rights reserved.)

Purpose: 1) To test the hypothesis of the existence of a perinatal vitamin A (VA) programming of VA metabolism and to better understand the intestinal regulation of VA metabolism., Methods: Offspring from rats reared on a control (C) or a VA-deficient (D) diet from 6 weeks before mating until offspring weaning, i.e., 7 weeks after mating, were themselves reared on a C or D diet for 19 weeks, resulting in the following groups: C-C (parents fed C-offspring fed C), D-C, C-D and D-D. VA concentrations were measured in plasma and liver. β-Carotene bioavailability and its intestinal conversion rate to VA, as well as vitamin D and E bioavailability, were assessed after gavages with these vitamins. Expression of genes involved in VA metabolism and transport was measured in intestine and liver., Results: C-D and D-D had no detectable retinyl esters in their liver. Retinolemia, hepatic retinol concentrations and postprandial plasma retinol response to β-carotene gavage were higher in D-C than in C-C. Intestinal expression of Isx was abolished in C-D and D-D and this was concomitant with a higher expression of Bco1, Scarb1, Cd36 and Lrat in males receiving a D diet as compared to those receiving a C diet. β-Carotene, vitamin D and E bio-availabilities were lower in offspring receiving a D diet as compared to those receiving a C diet., Conclusion: A VA-deficient diet during the perinatal period modifies the metabolism of this vitamin in the offspring. Isx-mediated regulation of Bco1 and Scarb1 expression exists only in males severely deficient in this vitamin. Severe VA deficiency impairs β-carotene and vitamin D and E bioavailability., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany.)

Objectives: The geriatric population, often polymedicated, is exposed to the risk of adverse drug events. Medication reconciliation (MR), which is an interactive and pluriprofessional process, helps ensure continuity of care. The objective of this study was to analyze and to define relevant prioritization criteria for MR in older patients in order to avoid a maximum of medication errors., Methods: A clinical audit of MR at the transition points of patient admission and discharge was conducted prospectively for 10 months. Patients were selected on the basis of a prioritization procedure already established in our structure, that is the presence of at least one of the three following criteria: originating from an hospital department, severe renal failure and prescription of at-risk drugs., Results: The cohort of patients reconciled at admission included 136 patients. A total of 63 unintentional discrepancies (UDs) were identified, the majority of which (76.2%) involved drug omissions. Three criteria were identified as independent predictors of UDs risk on the entry prescription compared to the optimized drug assessment: rheumatological history, originating from an hospital department and hyponatremia. Hyponatremia was found in the present study to be the most relevant criterion that significantly increased the risk of having an UD on the patient's prescription, particularly a risk of treatment omission at admission., Conclusion: This study will allow to improve the prioritization criteria on the healthcare establishment's procedure and to implement MR in geriatric day hospitalization in order to strengthen the city-hospital link., (Copyright © 2021 Académie Nationale de Pharmacie. Published by Elsevier Masson SAS. All rights reserved.)

Modifying the galenic of dry oral forms (DOF) to be administered to patients with swallowing or behavioral disorders is frequent in long term care homes. The objective was to investigate the practice of modifying DOF galenic by nurses in home settings (NHS). A 14-question electronic survey was distributed to 1977 NHS. Almost 3/4 of respondents reported crushing tablets or opening capsules, with 37% on a daily basis. Approximately 22% did not inquire about the feasibility of modifying galenic DOF beforehand. Nearly 75% of NHS were aware about the risk of ineffectiveness after crushing tablets or opening capsules. However, only 14% mentioned the risk of overdosing associated with this practice. More than 60% of NHS never wore protective equipment when changing dosage form. Changing the dosage form of DOF is a widespread practice in home settings. Our present work urges the need to improve collaboration between prescribing physicians, nurses and pharmacists., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2021 Elsevier Inc. All rights reserved.)

We compared the efficacy of tailored non pharmacological therapies (NPT) on specific nocturnal behavioral and psychological symptoms of dementia (BPSD). This retrospective 1-year study included 84 older dependent patients institutionalized in 7 long-term care home. Dedicated assistants, who were taught by experts how to use NPT, were asked to record the occurrence of each BPSD episode, to choose a given NPT on the basis of their knowledge of the patient and the type of BPSD and to estimate its efficacy. Wandering was the most prevalent BPSD followed by agitation/aggression and screaming. The most used therapy was cognitive stimulation, followed by multisensory stimulation, reminiscence and Montessori-based. Regarding wandering, multisensory stimulation was found to be the most efficacious NPT significantly different from Montessori-based, cognitive stimulation or reminiscence. With regards to agitation/aggression or screaming, Montessori-based was found to be the most efficacious NPT significantly different from multisensory stimulation, reminiscence and cognitive stimulation., Competing Interests: Declaration of Competing Interest None., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Scope: The effect of vitamin A deficiency on vitamin A and lipid postprandial metabolism in young rats is addressed, considering the effect of sex., Methods and Results: Sprague-Dawley rats are fed either 400 UI.kg -1 vitamin A diet (vitamin A-deficient (VAD) diet) or 2300 UI.kg -1 vitamin A (control diet), before being mated. Mothers receive the same VAD or control diet during gestation and lactation. Offspring receive the same diet than mothers until 8 weeks of age. VAD diet-fed female and male offspring display a severe vitamin A deficiency with no body weight or glucose tolerance defects. Fasting plasma triglyceride concentrations are decreased in VAD diet-fed animals compared to controls (p < 0.05). Retinyl ester postprandial responses after vitamin A gavage, expressed as area under the curves, are not different in VAD diet-fed and control animals, although retinyl ester postprandial peak is significantly delayed (p < 0.05) in VAD diet-fed rats. Lipids also accumulate in the distal part of the intestine after gavage and [1- 13 C]-oleate postprandial response is decreased in VAD diet-fed males., Conclusion: Vitamin A deficiency modulates both vitamin A absorption rate and lipid postprandial metabolism, which can partly explain the altered fasting lipid status observed in VAD diet-fed offspring., (© 2021 Wiley-VCH GmbH.)

Scope: To study the effect of variation in dietary vitamin A (VA) content on its hepatic and intestinal metabolism., Methods and Results: Adult female and male rats are fed with diets containing 400, 2300, or 9858 IU kg -1 VA for 31-33 weeks. VA concentrations are measured in plasma and liver. Bioavailability and intestinal conversion efficiency of β-carotene to VA are assessed by measuring postprandial plasma β-carotene and retinyl palmitate concentrations after force-feeding rats with β-carotene. Expression of genes involved in VA metabolism, together with concentrations of RBP4, BCO1, and SR-BI proteins, are measured in the intestine and liver of female rats. Plasma retinol concentrations are lower and hepatic free retinol concentrations are higher in females than in males. There is no effect of dietary VA content on β-carotene bioavailability and its conversion efficiency, but bioavailability is higher and conversion efficiency is lower in females than in males. The expression of most genes exhibited a U-shaped dose response curve depending on VA intake., Conclusions: β-Carotene bioavailability and conversion efficiency to VA are affected by the sex of rats. Results of gene expression suggest a hormetic regulation of VA metabolism in female rats., (© 2021 Wiley-VCH GmbH.)

Mastocytosis is an orphan disease associated with many systemic symptoms, chronic handicap, and potentially marked social consequences despite improved therapies. In this study, the authors aimed to measure the effect of 2 hypnosis sessions on mastocytosis symptoms in a clinical setting. Questionnaires (pain, flushes, energy, digestive symptoms, quality of life, perceived symptom severity, and global impression of change) were completed pre- and posthypnosis intervention. Data from 20 patients were analyzed (mean age: 53.3 years, 75% female). Compared to baseline assessment, patients exhibited a significant improvement immediately after the first and second hypnosis sessions with regard to the number of days with abdominal pain, abdominal pain intensity and fatigue ( p = .03 and p = .005; p = .05 and p = .02; p = .034, and p = .039, respectively). Perceived severity of symptoms was significantly improved throughout the study ( p = .0075). Long-term improvement in global impression of change was observed in half the responders (8/16). Patients with mastocytosis had an improvement in disabling symptoms with the impact of hypnotic intervention persisting at 1 month. Several patients experienced long-term improvement.

Aims: Short-term hospitalization of community-dwelling older dependent people in a geriatric acute care unit is sometimes needed to treat an acute health problem. Serious loss of independence can lead to difficulties in maintaining home care and is followed, at hospital discharge, to institutionalization in a long-term care home. We investigated the variables, particularly those related to the paramedical staff at home, predicting a return home or an institutionalization at hospital discharge., Methods: Retrospective observational study of 398 sixty and more year-old patients, living at home, having a natural caregiver, and hospitalized in an acute care unit of the State Geriatric Center., Results: 289 (72.6%) patients returned home, 101 (25.3%) were admitted in a long-term care home, and 8 (2%) died. Independent predictors of institutionalization were length of stay in the acute care unit [adjusted OR (AOR) = 1.102, P < 0.001], disruptive behavioral and psychological symptoms of dementia (BPSD, AOR = 1.827, P = 0.039), caregiver burden (AOR = 1.976, P = 0.038), moderately severe-to-severe cognitive impairment (AOR = 2.121, P = 0.011), and living alone with a close or a remote caregiver (AOR = 2.620 and 4.446, P = 0.003 and 0.001, respectively). In-home physiotherapy was independently associated (AOR = 0.393, P = 0.002) with a lower risk of institutionalization., Conclusion: In-home physiotherapy should be recommended to community-dwelling older dependent people, especially if they are living alone and/or if they present disruptive BPSD and/or moderately severe-to-severe cognitive impairment.

Purpose: Older patients with colon cancer (CC) are vulnerable to chemotherapy toxicity and death. Establishing simple scores specific for patients with CC to predict severe chemotoxicity or early death is needed to select the best treatment strategy., Subjects, Materials, and Methods: This prospective multicenter study included patients aged ≥70 years with CC receiving adjuvant or first-line metastatic chemotherapy. Frailty markers (nutrition, physical activity, energy, mobility, strength), comprehensive geriatric assessment (functional status, comorbidities, falls, nutrition, cognition, and depression), and usual laboratory parameters were collected. Logistic or Cox regression was used to examine at 500 days the association between frailty markers, comprehensive geriatric assessment, laboratory parameters, and grade 3-4 toxicity or death., Results: A total of 97 patients (median age, 79.0 years) received adjuvant (37.1%) or metastatic (62.9%) chemotherapy. During the first 500 days, grade 3-4 toxicity occurred in 49.5%, and 30% died. The predictive model for grade 3-4 toxicity combined (polychemotherapy × 3) + (hypoalbuminemia <32 g/L × 2) + (abnormal grip strength × 1.5) + C-reactive protein >11 mg/L + Eastern Cooperative Oncology Group performance status (ECOG-PS), cutoff score >3. The predictive model for death combined (metastasis × 5) + (age × 2) + alkaline phosphatase >100 IU/mL + sex (female) + abnormal grip strength + ECOG-PS, cutoff score >6. For chemotoxicity prediction, sensitivity was 81.6% and specificity 71.4%. For death prediction, sensitivity was 89.7% and specificity was 83.6%., Conclusion: These simple and efficient "ColonPrediscores" will help to better identify older patients with CC with increased risk of chemotherapy-related toxicity and/or death., Implications for Practice: The two scores assessed in this study, called "ColonPrediscores", offer a major advantage in that they do not need a previous complete geriatric assessment, which makes them an easy-to-use tool in oncologic settings., (© AlphaMed Press 2019.)

Background: Flow cytometry essentially focuses on surface-expressed proteins, with few protocols being devoted to intracellular components. We evaluated a two-step procedure using new formaldehyde-free permeabilization and staining reagents that allow the staining of platelets and red blood cells (RBCs) from whole blood., Methods: Citrated blood was treated with the new staining protocol (NSP) or control reagent (phosphate-buffered solution bovine serum albumin) and stained with antibodies against surface or intracellular markers. The effects of the NSP on cell integrity, morphology, and content were evaluated., Results: The NSP slightly reduced the cell count (~20%) and changed the RBC morphology with a 42% mean diameter reduction. Conversely, the NSP did not affect platelet discoid morphology and led to a minor size decrease (11%). These morphological changes neither impelled a gating strategy modification nor interfered with the discrimination among populations based on surface markers. The NSP provided intracellular access to all the tested antigens: CD62P, FXIII, and CD63 in platelets and glycated and fetal hemoglobin (HbA1c and HbF) and nucleic acid in RBCs. The NSP gave excellent intra-assay precision with minimal impact on cell morphology and fluorescence labelling over time (up to 24 h)., Conclusions: With the ability to detect surface and intracellular antigens through a rapid preparation protocol without washing steps or toxic formaldehyde treatment, this NSP designed for research offers a marked improvement in the analysis of platelets and RBCs isolated directly from whole blood. Consequently, the NSP opens new avenues to investigate platelet degranulation and erythrocyte subpopulations. © 2019 International Clinical Cytometry Society., (© 2019 International Clinical Cytometry Society.)

Antiplatelet therapy through inhibition of the adenosine diphosphate (ADP)/P2Y12 pathway is commonly used in the treatment of acute coronary syndrome (ACS). Although efficient in preventing platelet activation and thrombus formation, it increases the risk of bleeding complications. In patients with ACS receiving platelet aggregation inhibitors, that is, P2Y12 blockers (n = 923), we investigated the relationship between plasma and platelet-associated CD40L levels and bleeding events (n = 71). Treatment with P2Y12 inhibitors in patients with ACS did not affect plasma-soluble CD40L levels, but decreased platelet CD40L surface expression (pCD40L) and platelet-released CD40L (rCD40L) levels in response to stimulation as compared to healthy controls. In vitro inhibition of the ADP pathway in healthy control platelets reduced both pCD40L and rCD40L levels. In a multivariable analysis, the reduced pCD40L level observed in ACS patients was significantly associated with the risk of bleeding occurrence (adjusted odds ratio = 0.15; 95% confidence interval = 0.034-0.67). P2Y12 inhibitor-treated (ticagrelor) mice exhibited a 2.5-fold increase in tail bleeding duration compared with controls. A significant reduction in bleeding duration was observed on CD40L +/+ but not CD40L -/- platelet infusion. In addition, CD40L blockade in P2Y12 inhibitor-treated blood samples from a healthy human reduced thrombus growth over immobilized collagen under arterial flow. In conclusion, measurement of pCD40L may offer a novel approach to assessing bleeding risk in patients with ACS who are being treated with P2Y12 inhibitors., (© 2019 The Authors. Research and Practice in Thrombosis and Haemostasis published by Wiley Periodicals, Inc on behalf of International Society on Thrombosis and Haemostasis.)

Increased platelet activity occurs in type 2 diabetes mellitus (T2DM) and such platelet dysregulation likely originates from altered megakaryopoiesis. We initiated identification of dysregulated pathways in megakaryocytes in the setting of T2DM. We evaluated through transcriptomic analysis, differential gene expressions in megakaryocytes from leptin receptor-deficient mice (db/db), exhibiting features of human T2DM, and control mice (db/+). Functional gene analysis revealed an upregulation of transcripts related to calcium signaling, coagulation cascade and platelet receptors in diabetic mouse megakaryocytes. We also evidenced an upregulation (7- to 9.7-fold) of genes encoding stefin A (StfA), the human ortholog of Cystatin A (CSTA), inhibitor of cathepsin B, H and L. StfA/CSTA was present in megakaryocytes and platelets and its expression increased during obesity and diabetes in rats and humans. StfA/CSTA was primarily localized at platelet membranes and granules and was released upon agonist stimulation and clot formation through a metalloprotease-dependent mechanism. StfA/CSTA did not affect platelet aggregation, but reduced platelet accumulation on immobilized collagen from flowing whole blood (1200 s -1 ). In-vivo, upon laser-induced vascular injury, platelet recruitment and thrombus formation were markedly reduced in StfA1-overexpressing mice without affecting bleeding time. The presence of CA-074Me, a cathepsin B specific inhibitor significantly reduced thrombus formation in-vitro and in-vivo in human and mouse, respectively. Our study identifies StfA/CSTA as a key contributor of platelet-dependent thrombus formation in both rodents and humans.

Objectives: To determine whether environmental rearrangements of the long-term care nursing home can affect disruptive behavioral and psychological symptoms of dementia (BPSD) in residents with dementia., Design: Prospective 6-month study., Setting: The study was conducted before (phase 1) and after (phase 2) environmental rearrangements [skylike ceiling tiles in part of the shared premises, progressive decrease of the illuminance at night together with soothing streaming music, reinforcement of the illuminance during the day, walls painted in light beige, oversized clocks in corridors, and night team clothes color (dark blue) different from that of the day team (sky blue)]., Participants: All of the patients (n = 19) of the protected unit were included in the study. They were aged 65 years or older and had an estimated life expectancy above 3 months., Measures: Number and duration of disruptive BPSD were systematically collected and analyzed over 24 hours or during late hours (6:00-12:00 pm) during each 3-month period., Results: There was no significant change in the patients' dependency, risk of fall, cognitive or depression indexes, or treatment between phase 1 and 2. Agitation/aggression and screaming were observed mainly outside the late hours as opposed to wandering episodes that were noticed essentially within the late hours. The number of patients showing wandering was significantly lower over 24 hours during phase 2. The number of agitation/physical aggression, wandering, and screaming and the mean duration of wandering episodes were significantly (P = .039, .002, .025, and .026 respectively) decreased over 24 hours following environmental rearrangements. Similarly, a significant reduction in the number and mean duration of wandering was noticed during the late hours (P = .031 and .007, respectively)., Conclusions: Our study demonstrates that BPSD prevalence can be reduced following plain environmental rearrangements aimed at improving spatial and temporal orientation., (Copyright © 2018 AMDA – The Society for Post-Acute and Long-Term Care Medicine. Published by Elsevier Inc. All rights reserved.)

Background: Precision medicine and prediction of therapeutic response requires monitoring potential biomarkers before and after treatment. Liquid biopsies provide noninvasive prognostic markers such as circulating tumor DNA and RNA. Circulating tumor RNA (ctRNA) in blood is also used to identify mutations in genes of interest, but additionally, provides information about relative expression levels of important genes. In this study, we analyzed PD-L1 expression in ctRNA isolated from various cancer types. Tumors inhibit antitumor response by modulating the immune checkpoint proteins programmed death ligand 1 (PD-L1) and its cognate receptor PD1. The expression of these genes has been implicated in evasion of immune response and resistance to targeted therapies., Methods: Blood samples were collected from gastric (GC), colorectal (CRC), lung (NSCLC), breast (BC), prostate cancer (PC) patients, and a healthy control group. ctRNA was purified from fractionated plasma, and following reverse transcription, levels of PD-L1 expression were analyzed using qPCR., Results: PD-L1 expression was detected in the plasma ctRNA of all cancer types at varying frequencies but no PD-L1 mRNA was detected in cancer-free individuals. The frequencies of PD-L1 expression were significantly different among the various cancer types but the median relative PD-L1 expression values were not significantly different. In 12 cases where plasma and tumor tissue were available from the same patients, there was a high degree of concordance between expression of PD-L1 protein in tumor tissues and PD-L1 gene expression in plasma, and both methods were equally predictive of response to nivolumab., Conclusions: PD-L1 mRNA can be detected and quantitated in ctRNA of cancer patients. These results pave the way for further studies aimed at determining whether monitoring the levels of PD-L1 mRNA in blood can identify patients who are most likely to benefit from the conventional treatment., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Chronic heart failure (CHF) is a major public health matter. Mainly affecting the elderly, it is responsible for a high rate of hospitalization due to the frequency of acute heart failure (ADHF). This represents a disabling pathology for the patient and very costly for the health care system. Our study is designed to assess a connected and portable bioelectrical impedance analysis (BIA) that could reduce these hospitalizations by preventing early ADHF., Methods: This prospective study included patients hospitalized in cardiology for ADHF. Patients achieved 3 self-measurements using the BIA during their hospitalization and answered a questionnaire evaluating the acceptability of this self-measurement. The results of these measures were compared with the clinical, biological and echocardiographic criteria of patients at the same time., Results: Twenty-three patients were included, the self-measurement during the overall duration of the hospitalization was conducted autonomously by more than 80% of the patients. The acceptability (90%) for the use of the portable BIA was excellent. Some correlations were statistically significant, such as the total water difference to the weight difference (p=0.001). There were common trends between the variation of impedance analysis measures and other evaluation criteria., Conclusion: The feasibility and acceptability of a self-measurement of bioelectrical impedance analysis by the patient in AHF opens up major prospects in the management of monitoring patients in CHF. The interest of this tool is the prevention of ADHF leading to hospitalization or re-hospitalizations now requires to be presented by new studies.

Insulin receptor (IR) plays a key role in the control of glucose homeostasis; however, the regulation of its cellular expression remains poorly understood. Here we show that the amount of biologically active IR is regulated by the cleavage of its ectodomain, by the β-site amyloid precursor protein cleaving enzyme 1 (BACE1), in a glucose concentration-dependent manner. In vivo studies demonstrate that BACE1 regulates the amount of IR and insulin signaling in the liver. During diabetes, BACE1-dependent cleavage of IR is increased and the amount of IR in the liver is reduced, whereas infusion of a BACE1 inhibitor partially restores liver IR. We suggest the potential use of BACE1 inhibitors to enhance insulin signaling during diabetes. Additionally, we show that plasma levels of cleaved IR reflect IR isoform A expression levels in liver tumors, which prompts us to propose that the measurement of circulating cleaved IR may assist hepatic cancer detection and management.

Introduction: The non-adherence to substitutive treatment by L-thyroxine is the main cause of the discordance between high thyrotropin values and high doses of the drug., Observation: In a 36-year-old patient with post-surgery hypothyroidism, thyrotropin values ranged between 100 and 400 mUI/L, although daily replacement therapy included 300 μg of L-thyroxine and 75 μg of L-triiodothyronine. The oral loading test with L-thyroxine was normal and thyrotropin serum level returned to normal values under weekly oral administration of 1000 μg L-thyroxine., Conclusion: The strategy of non-adherence treatment in hypothyroidism is well defined with oral testing of L-thyroxine, followed by oral or parenteral weekly administration of the drug. The L-thyroxine oral test is the gold standard for diagnosis after eliminating of the other conventional causes: drug interactions or digestive malabsorption. L-thyroxine treatment should be discussed on a case-by-case basis, either daily under surveillance or once weekly oral or parenteral high dose., (Copyright © 2017 Société Nationale Française de Médecine Interne (SNFMI). Published by Elsevier SAS. All rights reserved.)

Purpose: Since intrathoracic goiters (IG), either cervico-mediastinal goiters (CMGs) or mediastinal nodules (MNs), can lead to sternotomies and/or evitable reoperations, their detection is mandatory before thyroid surgery. A systematic screening by CT scan or MRI is not conceivable because of their expensiveness. We tested if conventional chest radiography (CCR) could remain a good screening tool for IG before thyroid surgery., Methods: In this retrospective study (2554 patients), CCR usefulness was evaluated in relation with patients' complaints, clinical examination, neck US, and anatomical and surgical findings., Results: CMGs (n = 67) and MNs (n = 42) were symptomatic in 10 and 5 patients, respectively. Clinical examination or neck US suspected their existence in 25 and 13 and 45 and 17 patients, respectively. Among the 50 IG detected by CCR (42 CMGs and 8 MNs), 4 CMGs and 2 MNs were missed by clinical examination or neck US. CCR failed to detect IG in 59 patients (54%): 25 CMGs (37%) and 34 MNs (80%). Twenty-eight IG (9 CMGs and 19 MNs) were discovered during surgery. CCR resulted in false positive in 88 out of 2445 patients (3.5%). CCR potentially avoided reoperation in two patients (a maximum saving of 6160 €, whereas the total cost of CCR was 54,895 €)., Conclusions: CCR should not be used routinely for the preoperative detection of IG. Surgeons should preferably use clinical examination or neck US and directly perform CT scan when a mediastinal extension is suspected.

Objective: To describe glycemic control in nursing home residents with diabetes and to evaluate the relevance of HbA1c in the detection of hypoglycemia risk., Design and Methods: Diabetes treatment, geriatric assessment, blood capillary glucose (n= 24,682), and HbA1c were collected from medical charts of 236 southern France nursing home residents during a 4-month period. Glycemic control was divided into four categories: tight, fair, and moderate or severe chronic hyperglycemia using the High Blood Glucose Index or the analysis of blood glucose frequency distribution. Hypoglycemia episodes were identified by medical or biological records., Results: Glucose control was tight in 59.3 % and fair in 19.1 % of the residents. Chronic exposure to hyperglycemia was observed in 21.6 % of the residents (severe in 9.7 % and moderate in 11.9 %). Hypoglycemia was noticed in 42/236 (17.8%), in all categories of glycemic control. Relative hypoglycemia risk was significantly (P = 0.0095) higher in residents with moderate chronic hyperglycemia compared with those with tight control. The majority of residents with hypoglycemia (39/42) or chronic hyperglycemia (45/51) were insulin-treated. The relative risk of hypoglycemia was not significantly associated with HbA1c values., Conclusion: Hypoglycemia risk in nursing home residents is observed in all categories of glycemic control. In tight control, the potency of antidiabetic treatment should be reduced. In chronic hyperglycemia, diet and treatment should be reevaluated in order to reduce glucose variability. HbA1c is not sufficient for hypoglycemia risk detection; capillary blood glucose monitoring is warranted for nursing homes residents with diabetes., Competing Interests: The authors declare that there is no conflict of interest.

Background: Alterations in the nutritional perinatal environment, such as intrauterine growth retardation with subsequent postnatal catch-up growth, program cardiovascular disease in adulthood, possibly through alterations in matrix metalloproteinase (MMP)-2 and -9. However, experimental evidences demonstrating that changes in the nutritional perinatal environment can program MMP-2 and -9 with subsequent alterations of vessel wall are lacking., Aim: The current study evaluated whether immediate postnatal overfeeding is able to alter vascular morphological indexes and circulating and/or vascular MMP2-2 and -9 status., Methods: Aortic morphology (wall thickness and percentage of incomplete elastin lamellae) and circulating and aortic MMP-2 and -9 activity (measured by gelatin zymography) and aortic MMP-2 and -9 mRNA (measured by reverse transcription polymerase chain reaction (RT-PCR)) were studied in adult male rats overfed (OF) or normofed (NF) during the immediate postnatal period., Results: Postnatal overfeeding induced early onset obesity. Adult OF rats presented with increased blood pressure and circulating MMP-2 and -9 activity. In the thoracic aorta, postnatal overfeeding increased wall thickness and decreased elastin integrity (as demonstrated by an increased percentage of incomplete elastin lamellae). OF rats showed enhanced aortic MMP-2 activity and MMP-9 mRNA levels. Circulating and aortic MMP-2 activity correlated positively with the percentage of incomplete elastin lamellae and aortic wall thickness, respectively., Conclusion: Our data demonstrate for the first time that immediate postnatal nutritional programming induces increases in circulating and aortic MMP-2 activity with parallel aortic wall alterations, such as decreased elastin integrity and enhanced thickening, showing that this experimental model is suitable for the study of perinatal nutritional programming of vascular functions., (© American Journal of Hypertension, Ltd 2015. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Immediate postnatal overfeeding in rats, obtained by reducing the litter size, results in early-onset obesity. Such experimental paradigm programs overweight, insulin resistance, dyslipidemia, increased adipose glucocorticoid metabolism [up-regulation of glucocorticoid receptor (GR) and 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1)], and overexpression of proinflammatory cytokines in mesenteric adipose tissue (MAT) in adulthood. We studied the effects of pioglitazone, a PPARγ agonist, treatment on the above-mentioned overfeeding-induced alterations. Nine-month-old rats normofed or overfed during the immediate postnatal period were given pioglitazone (3 mg/kg/day) for 6 weeks. Pioglitazone stimulated weight gain and induced a redistribution of adipose tissue toward epididymal location with enhanced plasma adiponectin. Treatment normalized postnatal overfeeding-induced metabolic alterations (increased fasting insulinemia and free fatty acids) and mesenteric overexpression of GR, 11β-HSD11, CD 68, and proinflammatory cytokines mRNAs, including plasminogen-activator inhibitor type 1. Mesenteric GR mRNA levels correlated positively with mesenteric proinflammatory cytokines mRNA concentrations. In vitro incubation of MAT obtained from overfed rats demonstrated that pioglitazone induced a down-regulation of GR gene expression and normalized glucocorticoid-induced stimulation of 11β-HSD1 and plasminogen-activator inhibitor type 1 mRNAs. Our data show for the first time that the metabolic, endocrine, and inflammatory alterations induced by early-onset postnatal obesity can be reversed by pioglitazone at the adulthood. They demonstrate that pioglitazone, in addition to its well-established effect on adipose tissue redistribution and adiponectin secretion, reverses programing-induced adipose GR, 11β-HSD1, and proinflammatory cytokines overexpression, possibly through a GR-dependent mechanism.

Objectives: To compare the feasibility, safety, and outcome of parathyroidectomy in the management of primary hyperparathyroidism (PHPT) in individuals aged 75 and older with that of those younger than 50., Design: Retrospective chart review., Setting: Department of Endocrine Surgery, Hospital Paul Desbief (Marseille, France)., Participants: Individuals who underwent surgery from June 2005 to February 2013 (N = 526) had a clinical examination and laboratory and imaging assessments to diagnose PHPT., Measurements: The clinical and biochemical characteristics and surgery outcomes of individuals younger than 50 (n = 80) were compared with the characteristics and outcomes of those aged 75 and older (n = 89)., Results: Most of the participants did not have any specific signs of PHPT, and the diagnosis of PHPT was established in some participants during routine clinical and laboratory examination. Nephrolithiasis and osteitis fibrosa cystica were observed only in the younger group. Urinary calcium decreased with age. Nine participants aged 75 and older did not undergo surgery (four declined, five had medical contraindications). Conventional surgery through transverse cervicotomy was used in the majority of participants. Cure rate was excellent (158/160), with few and reversible minor complications. The coexistence of thyroid lesions was significantly higher in the older (47.5%) than in the younger group (32.3%). Nodules and multinodular goiters were removed in the majority of participants during the parathyroidectomy procedure., Conclusion: With the exception of a few cases with severe associated comorbidities, parathyroidectomy is safe and curative and should be considered as first-line choice for older adult with PHPT., (© 2014, Copyright the Authors Journal compilation © 2014, The American Geriatrics Society.)

Objective: High endogenous thrombin potential (ETP) is associated with venous and arterial thrombosis. Better knowledge of environmental influences on ETP may help to prevent thrombosis., Methods and Results: Weaning rats exhibited high ETP values that decreased in low-fat diet and remained elevated on high-fat diet. In adult rats, high-fat diet-induced ETP increase was independent of coagulation factors, obesity, and insulin resistance and negatively associated with polyunsaturated fatty acid levels. Switching from high-fat diet to low-fat diet reversed the procoagulant phenotype with a slower kinetic than the normalization of hyperinsulinemia. In humans, ETP was independent of body weight whereas it was negatively associated with nutritional markers such as the percentage of energy provided by proteins, the protein:fat ratio, circulating phenolic compounds, and omega-3 polyunsaturated fatty acid. A recommended 3-month healthy diet with reduced energy density, including lipids, decreased ETP (-21%; P<0.0001). Changes in ETP were not associated with body weight, insulin sensitivity, or coagulation factor variations, but correlated negatively with plasma docosahexaenoic acid, a nutritional status sensitive fatty acid, and compounds reflecting vegetable intake., Conclusions: Diet plays a pivotal role in regulating ETP, independently of obesity and insulin resistance. Global nutritional recommendations could be useful in primary prevention of venous thrombosis.

Background: Early life nutritional environment plays an important role in the development of visceral adipose tissue and interacts with nutritional regulations in adulthood, leading to metabolic dysregulations., Aim: We hypothesized that the renin-angiotensin system may play a role in the programming-induced development of visceral adipose tissue., Material and Methods: We studied, using a model of programming of overweight and glucose intolerance, obtained by post-natal overfeeding with consecutive highfat diet, the status of plasma renin activity and mesenteric adipose renin-angiotensin system, including the recently identified (pro)renin receptor, in adult rats., Results: Post-natal overfeeding or high-fat feeding lead to overweight with increased visceral fat mass and adipocytes surface. When both paradigms were associated, adipocytes surface showed a disproportionate increase. A strong immunoreactivity for (pro)renin receptor was found in stromal cells. Plasma renin activity increased in programmed animals whereas (pro)renin receptor expressing cells density was stimulated by high-fat diet. There was a positive, linear relationship between plasma renin activity and (pro)renin receptor expressing cells density and adipocytes surface., Conclusions: Our experiments demonstrate that association of post-natal overfeeding and high-fat diet increased plasma renin activity and adipose (pro)renin receptor expression. Such phenomenon could explain, at least in part, the associated disproportionate adipocyte hypertrophy and its accompanying increased glucose intolerance.

In severe obesity, white adipose tissue (WAT) inflammation and macrophage infiltration are thought to contribute to WAT and whole-body insulin resistance. Specific players involved in triggering and maintaining inflammation (i.e. those regulating adipokine release and WAT macrophage recruitment, retention, or function) remain to be fully elaborated, and the degree to which moderate obesity promotes WAT inflammation remains to be clarified further. Therefore, we characterized adiposity and metabolic phenotypes in adult male C57BL/6J mice fed differing levels of dietary fat (10, 45, and 60% of energy) for 12 wk, concurrent with determinations of WAT inflammation markers and mRNA expression of leukocyte-derived integrins (CD11b, CD11c, CD11d) involved in macrophage extravasation and tissue macrophage homing/retention. As expected, a lard-based, very high-fat diet (60% energy) significantly increased adiposity and glucose intolerance compared with 10% fat-fed controls, coincident with higher retroperitoneal (RP) WAT transcript levels for proinflammatory factors and macrophage markers, including TNFα and CD68 mRNA, which were ~3- and ~15-fold of control levels, respectively (P < 0.001). Mice fed the 45% fat diet had more moderate obesity, less glucose intolerance, and lower WAT macrophage/inflammatory marker mRNA abundances compared with 60% fat-fed mice; TNFα and CD68 mRNA levels were ~2- and ~5-fold of control levels (P < 0.01). Relative WAT expression of CD11d was massively induced by obesity to an extent greater than any other inflammatory marker (to >300-fold of controls in the 45 and 60% fat groups) (P < 0.0001) and this induction was WAT specific. Because we found that CD11d expression also increased in RP-WAT of Zucker obese rats and in the subcutaneous WAT of obese adult women, this appears to be a common feature of obesity. Observed correlations of WAT macrophage transcript marker abundances with body weight in lean to modestly obese mice raises an interesting possibility that the activities of at least some WAT macrophages are closely linked to the normal adipose remodeling that is a requisite for changes in WAT energy storage capacity.

Aims/hypothesis: Obesity is associated with adipose tissue inflammation. The CD40 molecule, TNF receptor superfamily member 5 (CD40)/CD40 ligand (CD40L) pathway plays a role in the onset and maintenance of the inflammatory reaction, but has not been studied in human adipose tissue. Our aim was to examine CD40 expression by human adipocytes and its participation in adipose tissue inflammation., Methods: CD40 expression was investigated in human whole adipose tissue and during adipocyte differentiation by real-time PCR, Western blot and immunohistochemistry. The CD40/CD40L pathway was studied using recombinant CD40L (rCD40L) in adipocyte culture and neutralising antibodies in lymphocyte/adipocyte co-culture., Results: CD40 mRNA levels in subcutaneous adipose tissue were higher in the adipocyte than in the stromal-vascular fraction. CD40 expression was upregulated during adipocyte differentiation. Addition of rCD40L to adipocytes induced mitogen activated protein kinase (MAPK) activation, stimulated inflammatory adipocytokine production, and decreased insulin-induced glucose transport in parallel with a downregulation of IRS1 and GLUT4 (also known as SCL2A4). rCD40L decreased the expression of lipogenic genes and increased lipolysis. CD40 mRNA levels were significantly higher in subcutaneous adipose tissue than in visceral adipose tissue of obese patients and were positively correlated with BMI, and with IL6 and leptin mRNA levels. Lymphocyte/adipocyte co-culture led to an upregulation of proinflammatory adipocytokines and a downregulation of leptin and adiponectin. Physical separation of the two cell types attenuated these effects, suggesting the involvement of a cell-cell contact. Blocking the CD40/CD40L interaction with neutralising antibodies reduced IL-6 secretion from adipocytes., Conclusions/interpretation: Adipocyte CD40 may contribute to obesity-related inflammation and insulin resistance. T lymphocytes regulate adipocytokine production through both the release of soluble factor(s) and heterotypic contact with adipocytes involving CD40.

Tumor suppressor candidate 5 (TUSC5) is a gene expressed abundantly in white adipose tissue (WAT), brown adipose tissue (BAT), and peripheral afferent neurons. Strong adipocyte expression and increased expression following peroxisome proliferator activated receptor gamma (PPARgamma) agonist treatment of 3T3-L1 adipocytes suggested a role for Tusc5 in fat cell proliferation and/or metabolism. However, the regulation of Tusc5 in WAT and its potential association with obesity phenotypes remain unclear. We tested the hypothesis that the TUSC5 gene is a bona fide PPARgamma target and evaluated whether its WAT expression or single-nucleotide polymorphisms (SNPs) in the TUSC5 coding region are associated with human obesity. Induction of Tusc5 mRNA levels in 3T3-L1 adipocytes by troglitazone and GW1929 followed a dose-response consistent with these agents' binding affinities for PPARgamma. Chromatin immunoprecipitation (ChIP) experiments confirmed that PPARgamma protein binds a approximately -1.1 kb promotor sequence of murine TUSC5 transiently during 3T3-L1 adipogenesis, concurrent with histone H3 acetylation. No change in Tusc5 mRNA or protein levels was evident in type 2 diabetic patients treated with pioglitazone. Tusc5 expression was not induced appreciably in liver preparations overexpressing PPARs, suggesting that tissue-specific factors regulate PPARgamma responsiveness of the TUSC5 gene. Finally, we observed no differences in Tusc5 WAT expression or prevalence of coding region SNPs in lean versus obese human subjects. These studies firmly establish the murine TUSC5 gene locus as a PPARgamma target, but the significance of Tusc5 in obesity phenotypes or in the pharmacologic actions of PPARgamma agonists in humans remains equivocal.

Epicardial fat is a relatively neglected component of the heart and could be an important risk factor of cardiac disease. The objective of our study was to assess the relationship between epicardial adipose tissue (EAT) extent, fat distribution, and coronaropathy in a group of adult victims of accidental or suspicious sudden death. In 56 cadavers, we performed 34 measurements of EAT from five computerized photographs of the heart (anterior and posterior faces, and three ventricle transversal slices) and analyzed their relationship with anthropometric markers of adiposity (BMI, waist and leg circumference, thickness of abdominal and thigh subcutaneous adipose tissue (SAT)), with the presence and staging of coronary artery disease (CAD), and with markers of myocardial hypertrophy. Simple linear regressions showed that EAT measurements are highly intercorrelated (r from 0.4 to 0.6, P < 0.001), and correlate with age, waist circumference, and heart weight, and to a lesser extent, with BMI, abdominal SAT thickness, and leg SAT thickness. Multiple regression showed that age, waist circumference, and heart weight significantly and independently correlate with EAT (P < 0.0001). No other anthropometric measurement was found independently correlated with EAT. The EAT/myocardium ratios correlated positively with age and waist circumference. Anterior and posterior areas of EAT were found significantly increased in patients with CAD and correlated positively with CAD staging (P = 0.0034, r = 0.38). Anterior EAT surface was found positively associated with CAD (P = 0.01), independently of age and other adiposity measurements. Prospective studies are needed to assess the risk of occurrence/progression of CAD that relate to EAT excess.

Recently, we characterized tumor suppressor candidate 5 (Tusc5) as an adipocyte-neuron PPARgamma target gene. Our objective herein was to identify additional genes that display distinctly high expression in fat and neurons, because such a pattern could signal previously uncharacterized functional pathways shared in these disparate tissues. gamma-Synuclein, a marker of peripheral and select central nervous system neurons, was strongly expressed in white adipose tissue (WAT) and peripheral nervous system ganglia using bioinformatics and quantitative PCR approaches. Gamma-synuclein expression was determined during adipogenesis and in subcutaneous (SC) and visceral adipose tissue (VAT) from obese and nonobese humans. Gamma-synuclein mRNA increased from trace levels in preadipocytes to high levels in mature 3T3-L1 adipocytes and decreased approximately 50% following treatment with the PPARgamma agonist GW1929 (P < 0.01). Because gamma-synuclein limits growth arrest and is implicated in cancer progression in nonadipocytes, we suspected that expression would be increased in situations where WAT plasticity/adipocyte turnover are engaged. Consistent with this postulate, human WAT gamma-synuclein mRNA levels consistently increased in obesity and were higher in SC than in VAT; i.e. they increased approximately 1.7-fold in obese Pima Indian adipocytes (P = 0.003) and approximately 2-fold in SC and VAT of other obese cohorts relative to nonobese subjects. Expression correlated with leptin transcript levels in human SC and VAT (r = 0.887; P < 0.0001; n = 44). Gamma-synuclein protein was observed in rodent and human WAT but not in negative control liver. These results are consistent with the hypothesis that gamma-synuclein plays an important role in adipocyte physiology.

Objective: Alterations of the perinatal environment, which lead to increased prevalence of the metabolic syndrome in adulthood, program an upregulation of systemic and/or adipose tissue glucocorticoid metabolism (11 beta-hydroxysteroid dehydrogenase type 1 [11 beta-HSD-1]-induced corticosterone reactivation). We hypothesized that postnatal programming could modulate high-fat diet-induced adipose tissue dysregulation in adulthood., Research Design and Methods: We compared the effects of chronic (since weaning) high- or low-fat diet in postnatally normofed (control) or overfed (programmed) rats., Results: Postnatal programming accentuated high-fat diet-induced overweight, insulin resistance, glucose intolerance, and decrease in circulating and epididymal adipose tissue adiponectin. Neither manipulation altered liver function. Postnatal programming or high-fat diet increased systemic corticosterone production, which was not further modified when both manipulations were associated. Postnatal programming suppressed high-fat diet-induced decrease in mesenteric adipose tissue (MAT) glucocorticoid sensitivity and triggered high-fat diet-induced increase in MAT glucocorticoid exposure, subsequent to enhanced MAT 11 beta-HSD-1 gene expression. MAT tumor necrosis factor (TNF)-alpha, TNF-receptor 1, interleukin (IL)-6, resistin, and plasminogen activator inhibitor-1 mRNAs were not changed by high-fat feeding in control rats and showed a large increase in programmed animals, with this effect further enhanced by high-fat diet for TNF-alpha and IL-6., Conclusions: Our data show for the first time that postnatal manipulation programs high-fat diet-induced upregulation of MAT glucocorticoid exposure, sensitivity, and inflammatory status and therefore reveal the pivotal role of the environment during the perinatal period on the development of diet-induced adipose tissue dysregulation in adulthood. They also urge the need for clinical trials with specific 11 beta-HSD-1 inhibitors.

Neuroendocrine (NE) differentiation in prostate cancer (CaP) has been reported to be an early marker associated with the development of androgen independence. The mechanisms by which CaP acquires NE properties are poorly understood. In this study, a putative role of adrenomedullin (AM) in the NE differentiation was investigated. The expression of AM and AM receptors (calcitonin receptor-like receptor (CRLR)/receptor activity modifying protein-2 and -3 (RAMP2 and RAMP3) was evaluated after experimental manipulation of androgen status. Levels of AM mRNA and immunoreactive AM (ir-AM) increased four- to sevenfold in androgen-sensitive LNCaP cells after androgen withdrawal in vitro and in LNCaP xenografts in animals after castration. Treatment of LNCaP cells with androgen analogue (dihydrotestosterone; 10(-9) M) prevented the increase in AM mRNA and ir-AM levels. Interestingly, the expression of CRLR, RAMP2 and RAMP3 is not regulated by androgen status. We demonstrate that in the presence of serum, AM is able to induce an NE phenotype in LNCaP cells via CRLR/RAMP2 and RAMP3, which includes extension of neuritic processes and expression of the neuron-specific enolase (NSE), producing cGMP in a dose-dependent manner, which is mediated by a pertussis toxin-sensitive GTP-binding protein. 8-Bromo-cGMP mimicked the effects of AM on cell differentiation. We demonstrate that AM induces a G-kinase Ialpha translocation to the nucleus. The protein kinase G inhibitor KT-5823 inhibited the neurite outgrowth induced by both AM and 8-bromo-cGMP. In noncastrated animals, administration of AM enhanced expression of NSE and chromogranin A in LNCaP xenografts with a significant increase of NSE levels in serum and no changes in tumor growth. In castrated animals, intraperitoneal injection of AM resulted in a 240+/-18% (P<0.001) increase in tumor volume 36 days after treatment, indicating that the nature of effect of AM in CaP depends on the presence or absence of endogenous androgen. Together, these results demonstrate that AM may function as a mediator of NE-like differentiation in culture as well as in vivo and indicate that its production may be important for tumor resurgence following androgen ablation.

Adipose tissue synthesizes factors involved in the body's homeostasis. Thus, measurements of messenger ribonucleic acid (mRNA) concentrations are important to study the involvement of adipose tissue in various physiological and pathophysiological conditions, in particular in obesity. Because adipose tissue is highly heterogeneous, containing both a stromal and an adipocyte compartment, each one having different cellular composition and functional capacities, in situ hybridization is a powerful tool to analyze the discrete expression of the mRNAs coding for the various factors synthesized within this tissue. Presented here is a detailed protocol for in situ hybridization of mRNAs in adipose tissue using 35S-labeled single-stranded probes with sufficient details for the readers unfamiliar with histologic techniques. Included are details of tissue sectioning and preparation, probe synthesis, hybridization reaction, and macro- and microscopic signal detection.

Epicardial white adipose tissue (eWAT) is in close contact with coronary vessels and therefore could alter coronary homeostasis. Adrenomedullin (AM) is a potent vasodilatator and antioxidative peptide which has been shown to play a cytoprotective role in experimental models of acute myocardial infarction. We studied, using immunohistochemistry and qRT-PCR, the expression of AM and its receptors calcitonin receptor-like receptor (CRLR), and receptor activity-modifying protein (RAMP)2 and -3 in paired biopsies of subcutaneous WAT (sWAT) and eWAT obtained from patients with coronary artery disease (CAD) or without CAD (NCAD). In eWAT obtained from NCAD or CAD patients, immunoreactivity for AM, CRLR, and RAMP2 and -3 was detected in blood vessel walls and isolated stromal cells close to adipocytes. Some of the AM positive stromal cells colocalized CD68 immunoreactivity. eWAT from CAD patients showed increased AM immunoreactivity and AM gene expression. CRLR mRNA levels were comparable in sWAT of both groups and decreased by 40-50% in eWAT, irrespectively of the coronary status. RAMP2 mRNA concentrations did not change while RAMP3 mRNA levels increased in sWAT from CAD patients. There was a positive linear relationship between eWAT 11beta-hydroxysteroid dehydrogenase type 1 mRNA (11beta-HSD-1, the enzyme that converts inactive to active glucocorticoids) and AM mRNA. In conclusion, we demonstrate that AM and its receptors are expressed in eWAT. Our data suggest that eWAT AM, which could originate from macrophages, is related to 11beta-HSD-1 expression. AM synthesis, which is increased in eWAT during chronic CAD in humans, can play a cardioprotective role.

Significant intra-individual variation in the sequences of the ribosomal RNA (rRNA) genes is highly unusual in animal genomes; however, two classes of both 18S and 28S rRNA gene sequences have been detected in chaetognaths, a small phylum of marine invertebrates. One species, Spadella cephaloptera Busch, 1851, is well-suited to the methods of in situ analysis of gene expression, since it is totally transparent. To test our hypothesis of a possible functional division of the two classes of genes, we carried out in situ hybridization. Our results indicated that 28S class II genes are expressed intensively in the oocytes of chaetognaths. In contrast, hybridization using an heterologous probe of 28S class I genes revealed only a single and relatively weak signal in a distinct area of intestinal cells. Our results suggest that the S. cephaloptera genome contains at least three different types of rRNA 28S genes; however, those which are expressed during housekeeping conditions could not be detected in our experiments.

It has been hypothesized that sleep apnea syndrome (SAS) increases hypothalamic-pituitary-adrenal axis activity and, through increased cortisol levels, participates in the pathophysiology of metabolic and cardiovascular complications. We compared the circadian profiles of cortisol in obese men with [obSAS+; apnea-hypopnea index (AHI) >or= 20/h] and without SAS (obSAS-; AHI

Chaetognaths constitute a small marine phylum of approximately 120 species. Two classes of both 18S and 28S rRNA gene sequences have been evidenced in this phylum, even though significant intraindividual variation in the sequences of rRNA genes is unusual in animal genomes. These observations led to the hypothesis that this unusual genetic characteristic could play one or more physiological role(s). Using in situ hybridization on the frontal sections of the chaetognath Spadella cephaloptera, we found that the 18S Class I genes are expressed in the whole body, with a strong expression throughout the gut epithelium, whereas the expression of the 18S Class II genes is restricted to the oocytes. Our results could suggest that the paralog products of the 18S Class I genes are probably the "housekeeping" 18S rRNAs, whereas those of class II would only be essential in specific tissues. These results provide support for the idea that each type of 18S paralog is important for specific cellular functions and is under the control of selective factors.

Adipose tissue synthesizes all components of the renin-angiotensin system. The renin receptor (RenR) is able, on renin binding, to increase its efficiency to generate angiotensin I from angiotensinogen. We demonstrate that RenR is specifically synthesized in the stromal portion of human adipose tissue in both isolated interadipocyte stromal cells and in stromal areas. RenR is expressed at the periphery of cells, strongly suggesting a membranal localization. RenR protein expression in primary cultures of human stromal cells decreased significantly during differentiation, whereas RenR mRNA levels did not change, demonstrating that RenR was expressed in both preadipocyte and nonpreadipocyte cells, and was regulated at a posttranscriptional level. Double-labeling immunohistochemistry of human adipose tissue sections revealed that RenR was colocalized with renin, whereas incubation of 3T3-L1, a preadipocyte cell line, with renin stimulated the phosphorylation state of the intracellular signaling pathway ERK 1/2, and short exposure of human adipose stromal cells in primary culture to renin was followed by a long-lasting dose-dependent increase of angiotensin I generation, indicating that adipose RenR is functional. We show, using a large set of human adipose tissue biopsies, that RenR expression was increased in visceral compared with subcutaneous adipose tissue of lean and obese patients. Taken together with our finding that RenR was colocalized with plasminogen activator inhibitor type 1, the main inhibitor of the fibrinolytic system in visceral adipose tissue, the above-mentioned data suggest that RenR plays a role in obesity-induced visceral adipose tissue accumulation and its accompanying cardiovascular complications.

Glucocorticoids hypersensitivity may be involved in the development of abdominal obesity and insulin resistance. Eight normal weight and eight obese women received on two occasions a 3-h intravenous infusion of saline or hydrocortisone (HC) (1.5 microg x kg(-1) x min(-1)). Plasma cortisol, insulin, and glucose levels were measured every 30 min from time(-30) (min) (time(-30)) to time(240). Free fatty acids, adiponectin, and plasminogen activator inhibitor-1 (PAI-1) levels were measured at time(-30), time(180), and time(240). At time(240), subjects underwent an insulin tolerance test to obtain an index of insulin sensitivity (K(ITT)). Mean(30-240) cortisol level was similar in control and obese women after saline (74 +/- 16 vs. 75 +/- 20 microg/l) and HC (235 +/- 17 vs. 245 +/- 47 microg/l). The effect of HC on mean(180-240) insulin, mean(180-240) insulin resistance obtained by homeostasis model assessment (HOMA-IR), and K(ITT) was significant in obese (11.4 +/- 2.0 vs. 8.2 +/- 1.3 mU/l, P < 0.05; 2.37 +/- 0.5 vs. 1.64 +/- 0.3, P < 0.05; 2.81 +/- 0.9 vs. 3.32 +/- 1.02%/min, P < 0.05) but not in control women (3.9 +/- 0.6 vs. 2.8 +/- 0.5 mU/l; 0.78 +/- 0.1 vs. 0.49 +/- 0.1; 4.36 +/- 1.1 vs. 4.37 +/- 1.2%/min). In the whole population, the quantity of visceral fat, estimated by computerized tomography scan, was correlated with the increment of plasma insulin and HOMA-IR during HC infusion [Delta mean(30-240) insulin (r = 0.61, P < 0.05), Delta mean(30-240) HOMA-IR (r = 0.66, P < 0.01)]. The increase of PAI-1 between time(180) and time(240) after HC was higher in obese women (+25%) than in controls (+12%) (P < 0.05), whereas no differential effect between groups was observed for free fatty acids or adiponectin. A moderate hypercortisolism, equivalent to that induced by a mild stress, has more pronounced consequences on insulin sensitivity in abdominally obese women than in controls. These deleterious effects are correlated with the amount of visceral fat.

Objective: Adrenomedullin (AM), a potent vasodilatator and antioxidative peptide, was shown recently to be expressed by adipose tissue. The aim of our study was to investigate the precise localization of AM within human adipose tissue, and to examine AM regulation in obesity., Design: Subcutaneous (SC) and omental (OM) adipose tissues from 9 lean and 13 obese women were profiled for AM expression changes. Preadipocytes from human adipose tissue were isolated and differentiated under defined adipogenic conditions., Methods: AM expression was analyzed by immunohistochemistry, in situ hybridization and quantitative RT-PCR., Results: A strong AM expression was observed in vessel walls, stromal cell clusters and isolated stromal cells, some of them being CD 68 positive, whereas mature adipocytes were not labeled. Calcitonin receptor-like receptor and receptor activity-modifying proteins (RAMP) 2 and RAMP 3 were expressed in vessel walls. In vitro, preadipocytes of early differentiation stages spontaneously secreted AM. No difference in AM localization was found between SC and OM adipose tissue. AM levels in SC tissue did not differ between lean and obese subjects. By contrast, AM levels in OM tissue were significantly higher in obese as compared with lean women. Moreover, we found a positive relationship between OM AM and tumor necrosis factor alpha mRNA levels and AM-immunoreactive area in OM tissue followed the features of the metabolic syndrome., Conclusion: Stromal cells from human adipose tissue, including macrophages, produce AM. Its synthesis increased in the OM territory during obesity and paralleled the features of the metabolic syndrome. Therefore, AM should be considered as a new member of the adipokine family.

Intrauterine growth retardation (IUGR) is associated with increased prevalence, at the adult age, of central obesity, the metabolic syndrome, and its complications (type 2 diabetes and coronary heart disease). Programming of the corticotropic function is one of the mechanisms underlying the above-mentioned phenomenon. An increased passage of active glucocorticoids from the mother to the fetus can act, at the central nervous system level, to program an enhanced response to stress and, at the peripheral level, in adipose tissue to induce an increased local glucocorticoid exposure and sensitivity. In addition to an improvement of the health of pregnant women, early diagnosis of metabolic and hormonal disturbances is important in children with IUGR, in order to prevent a compensatory catch-up growth and its subsequent obesity, and to set up a therapeutic intervention against the deleterious consequences of hypercorticism.

Objective: Data from rodents provide evidence for a causal role of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1) in the development of obesity and its complications. In humans, 11beta-HSD-1 is increased in subcutaneous adipose tissue (SAT) of obese patients, and higher adipose 11beta-HSD-1 was associated with features of the metabolic syndrome. To date, there is no evidence for an increased expression of 11beta-HSD-1 in human visceral adipose tissue (VAT), although VAT is the major predictor for insulin resistance and the metabolic syndrome., Research Methods and Procedures: 11beta-HSD-1 and hexose-6-phosphate dehydrogenase (the enzyme responsible for the synthesis of nicotinamide adenine dinucleotide phosphate, the cofactor required for 11beta-HSD-1 oxoreductase activity) mRNA levels were measured using real-time quantitative reverse transcriptase-polymerase chain reaction in abdominal SAT and VAT biopsies obtained from 10 normal-weight and 12 obese women. Adiponectin mRNA was used as an internal control., Results: 11beta-HSD-1 mRNA concentrations were significantly increased in both SAT and VAT of obese patients (720% and 450% of controls, respectively; p < 0.05) and correlated with hexose-6-phosphate dehydrogenase mRNA levels. The level of VAT 11beta-HSD-1 mRNA correlated with anthropometric parameters: BMI (r = 0.41, p = 0.05), waist circumference (r = 0.44, p = 0.04), abdominal sagittal diameter (r = 0.51, p = 0.02), and percentage fat (r = 0.51, p = 0.02)., Discussion: Our results demonstrate for the first time that 11beta-HSD-1 mRNA expression is increased in VAT from obese patients. They strengthen the importance of 11beta-HSD-1 in human obesity and its associated complications and suggest the need of clinical studies with specific 11beta-HSD-1 inhibitors.

Background: Plasminogen activator inhibitor type 1 (PAI-1) is the main inhibitor of the fibrinolytic system and contributes to an increased risk of atherothrombosis in insulin-resistant obese patients. In adipose tissue, we have shown that PAI-1 is synthesized mainly in the visceral stromal compartment and is positively regulated by glucocorticoids. We have demonstrated that adipose tissue expression of 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD-1), an enzyme that catalyzes the conversion of inactive cortisone to active cortisol, is exaggerated in obese patients., Objectives: We hypothesized that increased action of 11beta-HSD-1 in adipose tissue of obese subjects may contribute to PAI-1 overproduction., Patients and Methods: Using in situ hybridization, we studied the expression of the mRNAs coding for PAI-1 and 11beta-HSD-1 in the stromal compartment of visceral adipose tissue obtained from obese women. The regulation of PAI-1 secretion from in vitro incubated tissue explants was also investigated., Results: Regression analysis showed a significant positive linear relationship between PAI-1 and 11beta-HSD-1 mRNAs expression. In vitro incubation of adipose tissue explants demonstrated that cortisone stimulated PAI-1 gene expression and secretion, and that these effects were inhibited by co-incubation with the 11beta-HSD inhibitor, glycyrrhetinic acid., Conclusions: Our data demonstrate that 11beta-HSD-1-driven cortisone reactivation regulates adipose PAI-1 synthesis and secretion. They suggest that the increased PAI-1 synthesis and secretion observed in obese patients can be also related, at least in part, to an increased local conversion of cortisone to cortisol. Therefore, local cortisol metabolism in adipose tissue may be involved in increasing the risk of cardiovascular disease in obese subjects.