Dynapaenia and sarcopaenia in chronic haemodialysis patients: do muscle weakness and atrophy similarly influence poor outcome?

BACKGROUND: Sarcopaenia, defined as a decline in both muscle mass and function, has been recognized as a major determinant of poor outcome in haemodialysis (HD) patients. It is generally assumed that sarcopaenia is driven by muscle atrophy related to protein-energy wasting. However, dynapaenia, defined as weakness without atrophy, has been characterized by a different disease phenotype from sarcopaenia. The aim of this study was to compare the characteristics and prognosis of sarcopaenic and dynapaenic patients among a prospective cohort of chronic HD (CHD) patients. METHODS: Two hundred and thirty-two CHD patients were enrolled from January to July 2016 and then followed prospectively until December 2018. At inclusion, weakness and atrophy were, respectively, evaluated by maximal voluntary force (MVF) and creatinine index (CI). Sarcopaenia was defined as the association of weakness and atrophy (MVF and CI below the median) while dynapaenia was defined as weakness not related to atrophy (MVF below the median, and CI above the median). RESULTS: From a total of 187 prevalent CHD patients [65% of men, age 65.3 (49.7-82.0) years], 44 died during the follow-up period of 23.7 (12.4-34.9) months. Sarcopaenia and dynapaenia were observed in 33.7 and 16% of the patients, respectively. Compared with patients with sarcopaenia, patients with dynapaenia were younger and with a lower Charlson score. In contrast, mortality rate was similar in both groups (38 and 27%, respectively). After adjustment for age, sex, lean tissue index, serum albumin, high-sensitivity C-reactive protein (hs-CRP), haemoglobin (Hb), normalized protein catabolic rate (nPCR), dialysis vintage and Charlson score, only patients with dynapaenia were at increased risk of death [hazard ratio (HR) = 2.99, confidence interval 1.18-7.61; P = 0.02]. CONCLUSIONS: Screening for muscle functionality is highly warranted to identify patients with muscle functional impairment without muscle atrophy. In contrast to sarcopaenia, dynapaenia should appear as a phenotype induced by uraemic milieu, characterized by young patients with low Charlson score and poor prognosis outcome independently of serum albumin, hs-CRP, Hb, nPCR and dialysis vintage.

High dietary intake of palm oils compromises glucose tolerance whereas high dietary intake of olive oil compromises liver lipid metabolism and integrity.

PURPOSE: Palm (PO) and olive oils (OO) are the two most consumed and/or used oils in the world for food elaboration. These oils should not be confused with the solid palm stearin which is widely used in pastry making. Large number of studies was reported dealing with adverse/beneficial cardiovascular effects of PO and OO, whereas few studies were conducted to compare their potential effects on hepatic steatosis and liver lipid metabolism. The aim of this study was to compare the metabolic effects of high intake of POs (both crude and refined) and virgin OO on surrogate parameters of glucose tolerance, hepatic lipid metabolism and liver integrity. METHODS: Thirty-two young male Wistar rats were divided into four equal groups and fed either control diet (11% energy from fat) or three high-fat diets rich in crude or refined POs or in OO (56% energy from fat), during 12 weeks. Systemic blood and liver biochemical parameters linked to glucose and lipid metabolism as well as hepatic steatosis and liver fatty acid composition were explored. The inflammation and oxidative stress status as well as the expression of several genes/proteins were also analyzed. RESULTS: The major effects of POs intake concerned glucose metabolism and liver fatty acid composition, whereas the major effects of OO intake concerned hepatic TG accumulation, inflammation, and cytolysis. CONCLUSIONS: In conclusion, high dietary intake of PO compromises glucose tolerance whereas high dietary intake of OO compromises hepatic lipid composition and liver integrity. However, adverse hepatic effects of OO observed in this study may not be transposed to human since, (a) the rodent model could lead to different effects than those observed in humans and (b) the average normal OO amounts ingested in the population are lower than those corresponding to a high-fat diet. So, further studies are needed to determine a maximum non-invasive dietary intake of OO.

20-Week follow-up of hepatic steatosis installation and liver mitochondrial structure and activity and their interrelation in rats fed a high-fat-high-fructose diet.

The incidence of obesity and its metabolic complications are rapidly increasing and become a major public health issue. This trend is associated with an increase in the prevalence of non-alcoholic fatty liver disease (NAFLD), insulin resistance and diabetes. The sequence of events leading to NAFLD progression and mitochondrial dysfunction and their interrelation remains to be elucidated. This study aimed to explore the installation and progression of NAFLD and its association with the liver mitochondrial structure and activity changes in rats fed an obesogenic diet up to 20 weeks. Male Wistar rats were fed either a standard or high-fat-high-fructose (HFHFR) diet and killed on 4, 8, 12, 16 and 20 weeks of diet intake. Rats fed the HFHFR diet developed mildly overweight, associated with increased adipose tissue weight, hepatic steatosis, hyperglycaemia and hyperinsulinaemia after 8 weeks of HFHFR diet. Hepatic steatosis and many biochemical modifications plateaued at 8-12 weeks of HFHFR diet with slight amelioration afterwards. Interestingly, several biochemical and physiological parameters of mitochondrial function, as well as its phospholipid composition, in particular cardiolipin content, were tightly related to hepatic steatosis installation. These results showed once again the interrelation between hepatic steatosis development and mitochondrial activity alterations without being able to say whether the mitochondrial alterations preceded or followed the installation/progression of hepatic steatosis. Because both hepatic steatosis and mitochondrial alterations occurred as early as 4 weeks of diet, future studies should consider these four 1st weeks to reveal the exact interconnection between these major consequences of obesogenic diet intake.

Adaptation and psychometric properties of the spanish version of the yp-core (Young Person's Clinical Outcomes in Routine Evaluation).

INTRODUCTION: Given the increasing prevalence of mental health problems in the general population, it is indispensable to use assessment tools aimed to assess the outcome of therapeutic interventions in order to refine the process of psychological rehabilitation. METHOD: We describe the process of adaptation into Spanish and a first psychometric study of the Young Person's- Clinical Outcomes in Routine Evaluation (YP-CORE), an instrument designed to measure the outcome in terms of general distress of therapeutic interventions in young people (11-16 years). 104 adolescents participated in the clinical and 131 in the non-clinical samples. RESULTS: Analyses showed good levels of acceptability, adequate internal consistency and acceptable test-retest stability, with moderately high correlations between administrations. In addition, the instrument yielded significant correlations with all dimensions of the Youth Self Report, the highest being between both total scores. Crucially, discriminated between clinical and non-clinical samples and showed a small effect of age but a larger effect of gender, with higher scores for females. The Principal Component Analysis replicates the original structure. Cut-off scores to calculate the reliable and clinically significant change are provided. CONCLUSIONS: These results support initial use of the instrument though there are certain limitations that indicate the need for more research with larger and more representative samples, in which the psychometric properties of the instrument should be verified.

Functionalized Mesoporous Silica Nanoparticle with Antioxidants as a New Carrier That Generates Lower Oxidative Stress Impact on Cells.

Mesoporous silica nanoparticles (MSNs) were covalently coated with antioxidant molecules, namely, caffeic acid (MSN-CAF) or rutin (MSN-RUT), in order to diminish the impact of oxidative stress induced after transfection into cells, thus generating safer carriers used for either drug delivery or other applications. Two cellular models involved in the entry of NPs in the body were used for this purpose: the intestinal Caco-2 and the epidermal HaCaT cell lines. Rutin gave the best results in terms of antioxidant capacities preservation during coupling procedures, cellular toxicity alleviation, and decrease of ROS level after 24 h incubation of cells with grafted nanoparticles. These protective effects of rutin were found more pronounced in HaCaT than in Caco-2 cells, indicating some cellular specificity toward defense against oxidative stress. In order to gain more insight about the Nrf2 response, a stable transfected HaCaT cell line bearing repeats of the antioxidant response element (ARE) in front of a luciferase reporter gene was generated. In this cell line, both tBHQ and quercetin (Nrf2 agonists), but not rutin, were able to induce, in a dose-dependent fashion, the luciferase response. Interestingly, at high concentration, MSN-RUT was able to induce a strong Nrf2 protective response in HaCaT cells, accompanied by a comparable induction of HO-1 mRNA. The level of these responses was again less important in Caco-2 cells. To conclude, in keratinocyte cell line, the coupling of rutin to silica nanoparticles was beneficial in term of ROS reduction, cellular viability, and protective effects mediated through the activation of the Nrf2 antioxidant pathway.

A mitochondrial-targeted ubiquinone modulates muscle lipid profile and improves mitochondrial respiration in obesogenic diet-fed rats.

The prevalence of the metabolic syndrome components including abdominal obesity, dyslipidaemia and insulin resistance is increasing in both developed and developing countries. It is generally accepted that the development of these features is preceded by, or accompanied with, impaired mitochondrial function. The present study was designed to analyse the effects of a mitochondrial-targeted lipophilic ubiquinone (MitoQ) on muscle lipid profile modulation and mitochondrial function in obesogenic diet-fed rats. For this purpose, twenty-four young male Sprague-Dawley rats were divided into three groups and fed one of the following diets: (1) control, (2) high fat (HF) and (3) HF+MitoQ. After 8 weeks, mitochondrial function markers and lipid metabolism/profile modifications in skeletal muscle were measured. The HF diet was effective at inducing the major features of the metabolic syndrome--namely, obesity, hepatic enlargement and glucose intolerance. MitoQ intake prevented the increase in rat body weight, attenuated the increase in adipose tissue and liver weights and partially reversed glucose intolerance. At the muscle level, the HF diet induced moderate TAG accumulation associated with important modifications in the muscle phospholipid classes and in the fatty acid composition of total muscle lipid. These lipid modifications were accompanied with decrease in mitochondrial respiration. MitoQ intake corrected the lipid alterations and restored mitochondrial respiration. These results indicate that MitoQ protected obesogenic diet-fed rats from some features of the metabolic syndrome through its effects on muscle lipid metabolism and mitochondrial activity. These findings suggest that MitoQ is a promising candidate for future human trials in the metabolic syndrome prevention.

Diets Rich in Olive Oil, Palm Oil, or Lard Alter Mitochondrial Biogenesis and Mitochondrial Membrane Composition in Rat Liver.

Palm oil (crude or refined) and lard are rich in SFA, while olive oil is rich in polyunsaturated fatty acids. SFA are considered harmful to health, while polyunsaturated fatty acids are beneficial to health. The aim of this study was to determine the effect of diets rich in crude PO, refined PO, OO, or lard on the mitochondrial membrane, the activity of mitochondrial respiratory chain complexes, and mitochondrial biogenesis. This was an experimental study in male Wistar rats fed a diet containing 30% of each oil. Rats had free access to food and water. After being fed for 12 weeks, animals were sacrificed and liver mitochondria were collected. This collection was used to determine membrane potential and ROS production, membrane phospholipid and fatty acid composition, citrate synthase activity and respiratory chain complex, cardiolipin synthase protein expression, and expression of selected genes involved in mitochondrial biogenesis. We found that diets rich in olive oil, palm oil, or lard altered mitochondrial biogenesis by significantly decreasing Pgc1α gene expression and altered the fatty acid composition of rat liver mitochondrial membrane PL.

Impact of Highly Saturated versus Unsaturated Fat Intake on Carbohydrate Metabolism and Vascular Reactivity in Rat.

Palm olein (PO) and lard are considered harmful to health because of their highly saturated fatty acid content. On the contrary, olive oil (OO) with its high level of polyunsaturated fatty acids is considered healthier. This study aims to evaluate the effects of high consumption of these oils on carbohydrate metabolism and vascular function. Male Wistar rats were fed ad libitum for 12 weeks with different high fat diets (HFD) containing 30% of each oil. Systemic glycemia, insulinemia, and lipidemia were assessed by routine methods or by ELISA. GLUT4 muscular expression and hepatic and muscular Akt phosphorylation were analyzed by western blot. Vascular function was evaluated, ex vivo, on aortic rings and on the variations of isometric tensions. The results show that fasting blood glucose was increased with PO and OO diets and decreased with lard. Compared to control diet, this increase was significant only with PO diet. The area under the curve of IPGTT was increased in all HFD groups. Compared to control diet, this increase was significant only with PO. In contrast, stimulation of the pathway with insulin showed a significant decrease in Akt phosphorylation in all HFD compared to control diet. KCl and phenylephrine induced strong, dose-dependent vasoconstriction of rat aortas in all groups, but KCl EC50 values were increased with lard and OO diets. The inhibitory effect of tempol was absent in PO and lard and attenuated in OO. Vascular insulin sensitivity was decreased in all HFD groups. This decreased sensitivity of insulin was more important with PO and lard when compared to OO diet. In conclusion, the results of this study clearly show that high consumption of palm olein, olive oil, and lard can compromise glucose tolerance and thus insulin sensitivity. Furthermore, palm olein and lard have a more deleterious effect than olive oil on the contractile function of the aorta. Excessive consumption of saturated or unsaturated fatty acids is harmful to health, regardless of their vegetable or animal origin.

Skeletal Muscle Phenotype in Patients Undergoing Long-Term Hemodialysis Awaiting Kidney Transplantation.

BACKGROUND AND OBJECTIVES: Age and comorbidity-related sarcopenia represent a main cause of muscle dysfunction in patients on long-term hemodialysis. However, recent findings suggest muscle abnormalities that are not associated with sarcopenia. The aim of this study was to isolate functional and cellular muscle abnormalities independently of other major confounding factors, including malnutrition, age, comorbidity, or sedentary lifestyle, which are common in patients on maintenance hemodialysis. To overcome these confounding factors, alterations in skeletal muscle were analyzed in highly selected patients on long-term hemodialysis undergoing kidney transplantation. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: In total, 22 patients on long-term hemodialysis scheduled for kidney transplantation with few comorbidities, but with a long-term uremic milieu exposure, and 22 age, sex, and physical activity level frequency-matched control participants were recruited. We compared biochemical, functional, and molecular characteristics of the skeletal muscle using maximal voluntary force and endurance of the quadriceps, 6-minute walking test, and muscle biopsy of vastus lateralis. For statistical analysis, mean comparison and multiple regression tests were used. RESULTS: In patients on long-term hemodialysis, muscle endurance was lower, whereas maximal voluntary force was not significantly different. We observed a transition from type I (oxidative) to type II (glycolytic) muscle fibers, and an alteration of mitochondrial structure (swelling) without changes in DNA content, genome replication (peroxisome proliferator activator receptor γ coactivator-1α and mitochondrial transcription factor A), regulation of fusion (mitofusin and optic atrophy 1), or fission (dynamin-related protein 1). Notably, there were autophagosome structures containing glycogen along with mitochondrial debris, with a higher expression of light chain 3 (LC3) protein, indicating phagophore formation. This was associated with a greater conversion of LC3-I to LC3-II and the expression of Gabaralp1 and Bnip3l genes involved in mitophagy. CONCLUSIONS: In this highly selected long-term hemodialysis population, a low oxidative phenotype could be defined by a poor endurance, a fiber-type switch, and an alteration of mitochondria structure, without evidence of sarcopenia. This phenotype could be related to uremia through the activation of autophagy/mitophagy. CLINICAL TRIAL REGISTRATION NUMBERS: NCT02794142 and NCT02040363.

Peripancreatic Adipose Tissue Remodeling and Inflammation during High Fat Intake of Palm Oils or Lard in Rats.

Excessive fat consumption leads to the development of ectopic adipose tissues, affecting the organs they surround. Peripancreatic adipose tissue is implicated in glucose homeostasis regulation and can be impaired in obesity. High palm oil consumption's effects on health are still debated. We hypothesised that crude and refined palm oil high-fat feeding may have contrasting effects on peripancreatic adipocyte hypertrophy, inflammation and lipid oxidation compound production in obese rats. In Wistar rats, morphological changes, inflammation and isoprostanoid production following oxidative stress were assessed in peripancreatic adipose tissue after 12 weeks of diets enriched in crude or refined palm oil or lard (56% energy from fat in each case) versus a standard chow diet (11% energy from fat). Epididymal white and periaortic brown adipose tissues were also included in the study. A refined palm oil diet disturbed glucose homeostasis and promoted lipid deposition in periaortic locations, as well as adipocyte hypertrophy, macrophage infiltration and isoprostanoid (5-F2c-isoprostane and 7(RS)-ST-Δ8-11-dihomo-isofuran) production in peripancreatic adipose tissue. Crude palm oil induced a lower impact on adipose deposits than its refined form and lard. Our results show that the antioxidant composition of crude palm oil may have a protective effect on ectopic adipose tissues under the condition of excessive fat intake.

Effects of high-fat diets on inflammation and antioxidant status in rats : comparison between palm olein and olive oil.

Palm olein (PO) and olive oil (OO) are widely consumed in the world. PO is considered harmful to health, whereas OO is considered healthy. The aim of the study was to compare the effects of consumption of these oils on antioxidant status and inflammation in rats. This was an experimental study in male wistar rats fed a diet containing 30% of each oil. Rats had free access to food and water. After being fed for 12 weeks, animals were sacrificed and liver and aortic blood were collected. Plasma was used for the determination of interleukin-6 (IL-6) and oxidative stress parameters (Superoxide dismutase -SOD; Gluthation peroxidase - GPx; Thiobarbituric acid reactive substances - TBARS; Thiol groups and isoprostane). The inflammation and oxidative stress status as well as the expression of several genes/proteins were also analyzed in liver homogenate. No significant differences were observed between PO and OO in plasma and liver levels of the studied inflammation and oxidative stress parameters. This study showed that the consumption of PO induces an antioxidant status superimposable to that of OO.   Key words : Palm olein - Olive oil - Oxidative stress - Inflammation - High fat diet.

Impact of diets rich in olive oil, palm oil or lard on myokine expression in rats.

It has recently emerged that myokines may be an important skeletal muscle adaptive response to obesogenic diets in sedentary subjects (who do not exercise). This study aimed to assess the influence of various high fat (HF) diets rich in either crude palm oil (cPO), refined palm oil (rPO), olive oil (OO) or lard on the modulation of myokine gene expression in the gastrocnemius. Five groups of 8 rats were each fed HF or control diet for 12 weeks. Systemic parameters concerning glucose, insulin, inflammation, cholesterol, triglycerides (TG) and transaminases were assessed by routine methods or ELISA. Akt and ACC phosphorylation were analyzed by WB in the soleus. Mitochondrial density, inflammation, and the gene expression of 17 myokines and the apelin receptor (Apj) were assessed by qPCR in the gastrocnemius. We found that HF diet-fed rats were insulin resistant and Akt phosphorylation decreased in the soleus muscle, but without any change in Glut4 gene expression. Systemic (IL-6) and muscle inflammation (NFκB and IκB) were not affected by the HF diets as well as TBARS, and ASAT level was enhanced with OO diet. Soleus pACC phosphorylation and gastrocnemius mitochondrial density were not significantly altered. The gene expression of some myokines was respectively increased (myostatin and Il-15) and decreased (Fndc5 and apelin) with the HF diets, whatever the type of fat used. The gene expression of two myokines with anti-inflammatory properties, Il-10 and myonectin, was dependent on the type of fat used and was most increased respectively with cPO or both rPO and OO diets. In conclusion, high-fat diets can differentially modulate the expression of some myokines, either in a dependent manner or independently of their composition.

New stably transfected bioluminescent cells expressing FLAG epitope-tagged estrogen receptors to study their chromatin recruitment.

BACKGROUND: Biological actions of estrogens are mediated by the two specific estrogen receptors ERalpha and ERbeta. However, due to the absence of adequate cellular models, their respective transcriptional activities are still poorly understood. For instance, the evaluation of such differing properties on the transcription of responsive genes using ChIP experiments was hindered by the deficiency of cells exhibiting the same genotypic background and properties but expressing only one of the ERs. We describe here the generation of such cells, using an estrogen receptor negative HELN cell line that was derived from HeLa cells stably transfected with an ERE-driven luciferase plasmid. These HELN-Falpha and HELN-Fbeta cell lines stably express either the alpha or beta (full length) estrogen receptor tagged with the FLAG epitope. The use of antibodies directed against the FLAG epitope allowed a direct comparative evaluation of the respective actions of both ERs using ChIP. RESULTS: HELN-Falpha and HELN-Fbeta cell lines were found to express comparable levels of their corresponding tagged receptors with a Kd for estradiol binding of 0.03 and 0.27 nM respectively. The presence of a stably transfected ERE-driven luciferase plasmid in these cells allowed the direct evaluation of the transcriptional activity of both tagged receptors, using natural or synthetic estrogens. FLAG-ERalpha and FLAG-ERbeta were found to exhibit similar transcriptional activity, as indicated by a kinetic evaluation of the transcriptional activation of the luciferase gene during 10 hrs of treatment with estradiol. The validity of these model cells was further confirmed by the predictable transcriptional regulations measured upon treatments with ERalpha or ERbeta specific ligands. The similar immunoprecipitation efficiency of both tagged receptors by an anti-FLAG antibody allowed the assessment of their kinetic recruitment on the synthetic luciferase promoter (containing an estrogen response element) by ChIP assays during 8 hours. A biphasic curve was obtained for both FLAG-ERalpha and FLAG-ERbeta, with a peak occurring either at 2 hr or at 1 hr, respectively, and a second one following 4 hr of E2 stimulation in both cases. In MCF-7 cells, the recruitment of ERalpha also exhibited a biphasic behaviour; with the second peak however not so important than in the HeLa cell lines. CONCLUSION: In HELN derived cell lines, no fundamental differences between kinetics were observed during 8 hours for FLAG-ERalpha and FLAG-ERbeta, as well as for polymerase II recruitment. However, the relative importance of recruitment between 1 hr and 4 hr was found to be different in HeLa cell line expressing exogenous tagged ERalpha and in MCF-7 cell line expressing endogenous ER.

Increased expression of the HDAC9 gene is associated with antiestrogen resistance of breast cancers.

Estrogens play a pivotal role in breast cancer etiology, and endocrine therapy remains the main first line treatment for estrogen receptor-alpha (ERα)-positive breast cancer. ER are transcription factors whose activity is finely regulated by various regulatory complexes, including histone deacetylases (HDACs). Here, we investigated the role of HDAC9 in ERα signaling and response to antiestrogens in breast cancer cells. Various Michigan Cancer Foundation-7 (MCF7) breast cancer cell lines that overexpress class IIa HDAC9 or that are resistant to the partial antiestrogen 4-hydroxy-tamoxifen (OHTam) were used to study phenotypic changes in response to ER ligands by using transcriptomic and gene set enrichment analyses. Kaplan-Meier survival analyses were performed using public transcriptomic datasets from human breast cancer biopsies. In MCF7 breast cancer cells, HDAC9 decreased ERα mRNA and protein expression and inhibited its transcriptional activity. Conversely, HDAC9 mRNA was strongly overexpressed in OHTam-resistant MCF7 cells and in ERα-negative breast tumor cell lines. Moreover, HDAC9-overexpressing cells were less sensitive to OHTam antiproliferative effects compared with parental MCF7 cells. Several genes (including MUC1, SMC3 and S100P) were similarly deregulated in OHTam-resistant and in HDAC9-overexpressing MCF7 cells. Finally, HDAC9 expression was positively associated with genes upregulated in endocrine therapy-resistant breast cancers and high HDAC9 levels were associated with worse prognosis in patients treated with OHTam. These results demonstrate the complex interactions of class IIa HDAC9 with ERα signaling in breast cancer cells and its effect on the response to hormone therapy.

Physical inactivity and protein energy wasting play independent roles in muscle weakness in maintenance haemodialysis patients.

BACKGROUND: Muscle weakness is associated with increased mortality risk in chronic haemodialysis (CHD) patients. Protein energy wasting (PEW) and low physical activity could impair muscle quality and contribute to muscle weakness beyond muscle wasting in these patients. Aim of this study was to assess clinical and biological parameters involved in the reduction of muscle strength of CHD patients. METHODS: One hundred and twenty-three CHD patients (80 males, 43 females; 68,8 [57.9-78.8] y.o.) were included in this study. Maximal voluntary force (MVF) of quadriceps was assessed using a belt-stabilized hand-held dynamometer. Muscle quality was evaluated by muscle specific torque, defined as the strength per unit of muscle mass. Muscle mass was estimated using lean tissue index (LTI), skeletal muscle mass (SMM) assessed by bioelectrical impedance analysis and creatinine index (CI). Voorrips questionnaire was used to estimate physical activity. Criteria for the diagnosis of PEW were serum albumin, body mass index < 23 kg/m2, creatinine index < 18.82 mg/kg/d and low dietary protein intake estimated by nPCR < 0.80g/kg/d. RESULTS: MVF was 76.1 [58.2-111.7] N.m. and was associated with CI (ß = 5.3 [2.2-8.4], p = 0.001), LTI (ß = 2.8 [0.6-5.1], p = 0.013), Voorrips score (ß = 17.4 [2.9-31.9], p = 0.02) and serum albumin (ß = 1.9 [0.5-3.2], p = 0.006). Only serum albumin (ß = 0.09 [0.03-0.15], p = 0.003), Voorrips score (ß = 0.8 [0.2-1.5], p = 0.005) and CI (ß = 0.2 [0.1-0.3], p<0.001) remained associated with muscle specific torque. Thirty patients have dynapenia defined as impaired MVF with maintained SMM and were younger with high hs-CRP (p = 0.001), PEW criteria (p<0.001) and low Voorrips score (p = 0.001), and reduced dialysis vintage (p<0.046). CONCLUSIONS: Beyond atrophy, physical inactivity and PEW conspire to impair muscle strength and specific torque in CHD patients and could be related to muscle quality. TRIAL REGISTRATION: ClinicalTrials.gov NCT02806089.

Tamoxifen resistance and epigenetic modifications in breast cancer cell lines.

Epigenetic mechanisms play crucial roles in many processes, including neoplasia, genomic imprinting, gene silencing, differentiation, embryogenesis and X chromosome inactivation. Their relevance in human disease and therapy has grown rapidly with the recent emergence of drugs that target for example DNA methylation or histone acetylation. Epigenetic effects were also recently highlighted by the deciphering of the mechanism of action of steroid hormones and anti-hormones acting through nuclear receptors. In this review, we focus on the epigenetic effects associated with long-term treatment of breast cancer cells with the antiestrogen (AE) tamoxifen, in the context of resistance appearance. We summarize the data obtained with a model cell line developed in our laboratory supporting a role for HP1 proteins in the irreversible inactivation of gene expression by long-term treatment with AE.

Receptor-interacting protein 140 is a repressor of the androgen receptor activity.

The androgen receptor (AR) is a ligand-activated transcription factor that controls growth and survival of prostate cancer cells. In the present study, we investigated the regulation of AR activity by the receptor-interacting protein 140 (RIP140). We first showed that RIP140 could be coimmunoprecipitated with the receptor when coexpressed in 293T cells. This interaction appeared physiologically relevant because chromatin immunoprecipitation assays revealed that, under R1881 treatment, RIP140 could be recruited to the prostate-specific antigen encoding gene in LNCaP cells. In vitro glutathione S-transferase pull-down assays provided evidence that the carboxy-terminal domain of AR could interact with different regions of RIP140. By means of fluorescent proteins, we demonstrated that ligand-activated AR was not only able to translocate to the nucleus but also to relocate RIP140 from very structured nuclear foci to a diffuse pattern. Overexpression of RIP140 strongly repressed AR-dependent transactivation by preferentially targeting the ligand binding domain-dependent activity. Moreover, disruption of RIP140 expression induced AR overactivation, thus revealing RIP140 as a strong AR repressor. We analyzed its mechanism of transrepression and first demonstrated that different regions of RIP140 could mediate AR-dependent repression. We then showed that the carboxy-terminal end of RIP140 could reverse transcriptional intermediary factor 2-dependent overactivation of AR. The use of mutants of RIP140 allowed us to suggest that C-terminal binding protein played no role in RIP140-dependent inhibition of AR activity, whereas histone deacetylases partly regulated that transrepression. Finally, we provided evidence for a stimulation of RIP140 mRNA expression in LNCaP cells under androgen treatment, further emphasizing the role of RIP140 in androgen signaling.

The nuclear receptor liver receptor homolog-1 is an estrogen receptor target gene.

Liver receptor homolog-1 (LRH-1) is a nuclear receptor previously known to have distinct functions during mouse development and essential roles in cholesterol homeostasis. Recently, a new role for LRH-1 has been discovered in tumor progression, giving LRH-1 potential transforming functions. In order to identify critical factors stimulating LRH-1 expression leading to deregulated cellular proliferation, we studied its expression and its regulation in several breast cancer cell lines. We observed that LRH-1 expression was increased in estrogen receptor (ER) alpha expressing cell lines, whereas weak-to-no expression was found in nonexpressing ERalpha cell lines. In MCF7, LRH-1 expression was highly induced after treatment with 17beta-estradiol (E2). This transcriptional regulation was the result of a direct binding of the ER to the LRH-1 promoter, as demonstrated by gelshift and chromatin immunoprecipitation assays. Interestingly, siRNA-mediated inactivation of LRH-1 decreased the E2-dependent proliferation of MCF7 cells. Finally, LRH-1 protein expression was detected by immunohistochemistry in tumor cells of human mammary ductal carcinomas. Altogether, these data demonstrate that LRH-1 is transcriptionally regulated by the ER alpha and reinforce the hypothesis that LRH-1 could exert potential oncogenic effects during breast cancer formation.

Negative regulation of hormone signaling by RIP140.

Receptor interacting protein (RIP) 140 is a negative transcriptional regulator of nuclear hormone receptors which is required for the maintenance of energy homeostasis and ovulation. Despite its recruitment by agonist-liganded receptors, this protein exhibits a strong repressive activity which was initially attributed to competition with coactivator binding on nuclear receptors. However, RIP140 also exerts active repression implicating the Carboxyl-terminal binding proteins (CtBPs) and histone deacetylases (HDACs). We recently demonstrated that the Carboxyl-terminal region of the molecule contains two additional silencing domains which require post-translational modifications to be fully active. In human breast cancer cells, RIP140 expression is up-regulated at the transcriptional level by various ligands of nuclear receptors. We have recently cloned the human RIP140 gene and defined the mechanism of its regulation by estrogens. In order to better characterize the role of RIP140 in hormone signaling, we have studied its interaction with the androgen receptor and demonstrated its ability to repress transcriptional regulation by androgens. RIP140 also inhibits transactivation by estrogen receptor-related receptors (ERRalpha, beta and gamma) on natural or artificial reporter genes containing different types of response elements. Surprisingly, RIP140 positively regulates ERR transactivation when the receptors are recruited to target promoters through interaction with the Sp1 transcription factor and this effect could involve titration of histone deacetylases. Altogether, these results underline that transcriptional regulation of hormone signaling by the cofactor RIP140 involves complex mechanisms relying on multiple domains and partners.

An estrogen-responsive element-targeted histone deacetylase enzyme has an antiestrogen activity that differs from that of hydroxytamoxifen.

We showed previously that prolonged treatment of a MCF-7-derived cell line with hydroxytamoxifen (OHT) induces the irreversible silencing of some estrogen-responsive genes, whereas OHT-resistant cell growth appears simultaneously (E. Badia et al., Cancer Res., 60: 4130-4138, 2000). Based on the hypothesis that particular gene silencings could be involved in triggering the resistance phenomenon, we focused our study on the mechanism of OHT-induced silencing. More precisely, we wished to determine to what extent the recruited histone deacetylase (HDAC) activity, which is known to be involved in the repressive effect induced by antagonist ligands of nuclear receptors, could participate in various aspects of OHT effects, particularly in gene silencing. A fusion protein (HDAC-EG) of human HDAC1 fused with the estrogen receptor DNA-binding domain and the glucocorticoid receptor ligand-binding domain allowed targeting of chimeric HDAC1 activity on estrogen-responsive elements (EREs) in the presence of glucocorticoid ligands. When HDAC-EG was transiently expressed in HeLa cells together with estrogen receptor, an antiestrogen-like effect was obtained on an ERE-controlled luciferase reporter gene in the presence of agonist or antagonist glucocorticoids. In MCF-7-derived cells stably expressing HDAC-EG and an estrogen-regulated luciferase, liganded HDAC-EG again produced an antiestrogenic effect on expression of natural estrogen-regulated genes such as pS2, progesterone receptor, and cathepsin D and cell growth together with chimeric luciferase gene expression. However, a prolonged HDAC-EG-mediated antiestrogen effect did not lead to irreversible luciferase gene silencing, as OHT does. It nevertheless accelerated the OHT-driven phenomenon. The antiestrogen effect of OHT thus differs from that of an ERE-targeted HDAC1 activity that might participate in irreversible silencing but is not sufficient to trigger it.