José Ignacio Rodríguez-Hermosa

MiRNA Expression Profile of Human Subcutaneous Adipose and during Adipocyte Differentiation

Background Potential regulators of adipogenesis include microRNAs (miRNAs), small non-coding RNAs that have been recently shown related to adiposity and differentially expressed in fat depots. However, to date no study is available, to our knowledge, regarding miRNAs expression profile during human adipogenesis. Thereby, the aim of this study was to investigate whether miRNA pattern in human fat cells and subcutaneous adipose tissue is associated to obesity and co-morbidities and whether miRNA expression profile in adipocytes is linked to adipogenesis. Methodology/Principal Findings We performed a global miRNA expression microarray of 723 human and 76 viral mature miRNAs in human adipocytes during differentiation and in subcutaneous fat samples from non-obese (n = 6) and obese with (n = 9) and without (n = 13) Type-2 Diabetes Mellitus (DM-2) women. Changes in adipogenesis-related miRNAs were then validated by RT-PCR. Fifty of 799 miRNAs (6.2%) significantly differed between fat cells from lean and obese subjects. Seventy miRNAs (8.8%) were highly and significantly up or down-regulated in mature adipocytes as compared to pre-adipocytes. Otherwise, 17 of these 799 miRNAs (2.1%) were correlated with anthropometrical (BMI) and/or metabolic (fasting glucose and/or triglycerides) parameters. We identified 11 miRNAs (1.4%) significantly deregulated in subcutaneous fat from obese subjects with and without DM-2. Interestingly, most of these changes were associated with miRNAs also significantly deregulated during adipocyte differentiation. Conclusions/Significance The remarkable inverse miRNA profile revealed for human pre-adipocytes and mature adipocytes hints at a closely crosstalk between miRNAs and adipogenesis. Such candidates may represent biomarkers and therapeutic targets for obesity and obesity-related complications.

The gene expression of the main lipogenic enzymes is downregulated in visceral adipose tissue of obese subjects

Contradictory findings regarding the gene expression of the main lipogenic enzymes in human adipose tissue depots have been reported. In this cross‐sectional study, we aimed to evaluate the mRNA expression of fatty acid synthase (FAS) and acetyl‐CoA carboxilase (ACC) in omental and subcutaneous (SC) fat depots from subjects who varied widely in terms of body fat mass. FAS and ACC gene expression were evaluated by real time‐PCR in 188 samples of visceral adipose tissue which were obtained during elective surgical procedures in 119 women and 69 men. Decreased sex‐adjusted FAS (−59%) and ACC (−49%) mRNA were found in visceral adipose tissue from obese subjects, with and without diabetes mellitus type 2 (DM‐2), compared with lean subjects (both P < 0.0001). FAS mRNA was also decreased (−40%) in fat depots from overweight subjects (P < 0.05). Indeed, FAS mRNA was significantly and positively associated with ACC gene expression (r = 0.316, P < 0.0001) and negatively with BMI (r = −0.274), waist circumference (r = −0.437), systolic blood pressure (r = −0.310), serum glucose (r = −0.277), and fasting triglycerides (r = −0.226), among others (all P < 0.0001). Similar associations were observed for ACC gene expression levels. In a representative subgroup of nonobese (n = 4) and obese women (n = 6), relative FAS gene expression levels significantly correlated (r = 0.657, P = 0.034; n = 10) with FAS protein values. FAS protein levels were also inversely correlated with blood glucose (r = −0.640, P = 0.046) and fasting triglycerides (r = −0.832, P = 0.010). In conclusion, the gene expression of the main lipogenic enzymes is downregulated in visceral adipose tissue from obese subjects.

A role for adipocyte-derived lipopolysaccharide-binding protein in inflammation- and obesity-associated adipose tissue dysfunction

Aims/hypothesisCirculating lipopolysaccharide-binding protein (LBP) is an acute-phase reactant known to be increased in obesity. We hypothesised that LBP is produced by adipose tissue (AT) in association with obesity.MethodsLBP mRNA and LBP protein levels were analysed in AT from three cross-sectional (n = 210, n = 144 and n = 28) and three longitudinal (n = 8, n = 25, n = 20) human cohorts; in AT from genetically manipulated mice; in isolated adipocytes; and in human and murine cell lines. The effects of a high-fat diet and exposure to lipopolysaccharide (LPS) and peroxisome proliferator-activated receptor (PPAR)γ agonist were explored. Functional in vitro and ex vivo experiments were also performed.ResultsLBP synthesis and release was demonstrated to increase with adipocyte differentiation in human and mouse AT, isolated adipocytes and human and mouse cell lines (Simpson–Golabi–Behmel syndrome [SGBS], human multipotent adipose-derived stem [hMAD] and 3T3-L1 cells). AT LBP expression was robustly associated with inflammatory markers and increased with metabolic deterioration and insulin resistance in two independent cross-sectional human cohorts. AT LBP also increased longitudinally with weight gain and excessive fat accretion in both humans and mice, and decreased with weight loss (in two other independent cohorts), in humans with acquired lipodystrophy, and after ex vivo exposure to PPARγ agonist. Inflammatory agents such as LPS and TNF-α led to increased AT LBP expression in vivo in mice and in vitro, while this effect was prevented in Cd14-knockout mice. Functionally, LBP knockdown using short hairpin (sh)RNA or anti-LBP antibody led to increases in markers of adipogenesis and decreased adipocyte inflammation in human adipocytes.Conclusions/interpretationCollectively, these findings suggest that LBP might have an essential role in inflammation- and obesity-associated AT dysfunction.

Subcutaneous Fat Shows Higher Thyroid Hormone Receptor-α1 Gene Expression Than Omental Fat

The aims of this work were to evaluate thyroid hormone receptor‐α (TRα), TRα1, and TRα2 mRNA gene expression and TRα1:TRα2 ratio, identified as candidate factors for explaining regional differences between human adipose tissue depots. TRα, TRα1, and TRα2 mRNA levels, and the gene expressions of arginine–serine‐rich, splicing factor 2 (SF2), heterogeneous nuclear ribonucleoprotein H1 (hnRNP H1), heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1), and Spot 14 (S14) were evaluated in 76 paired adipose tissue samples obtained from a population of 38 women who varied widely in terms of obesity and body fat distribution. Gene expression for these factors was also studied in stromal‐vascular cells (SVCs) and mature adipocytes (MAs) from eight paired fat depots. TRα gene and TRα1 mRNA expression were increased 1.46‐fold (P = 0.006) and 1.80‐fold (P < 0.0001), respectively, in subcutaneous (SC) vs. visceral fat. These differences in gene expression levels were most significant in the obese group, in which the TRα1:TRα2 ratio was 2.24‐fold (P < 0.0001) higher in SC vs. visceral fat. S14 gene expression was also increased by 2.42‐fold (P < 0.0001) and correlated significantly with TRα and TRα1 gene expression and with the TRα1:TRα2 ratio. In agreement with these findings, hnRNP A1:SF2 ratio was decreased by 1.39‐fold (P = 0.001). TRα and S14 levels were 2.1‐fold (P < 0.0001) and 112.4‐fold (P < 0.0001), respectively, higher in MAs than in SVCs from both fat depots. In summary, genes for TR‐α, their upstream regulators, and downstream effectors were differentially expressed in SC vs. omental (OM) adipose tissue. Our findings suggest that TRα1 could contribute to SC adipose tissue expandability in obese subjects.

Factores de riesgo de dehiscencia aguda de la pared abdominal tras laparotomía en adultos

Resumen Introduccion La evisceracion total es la salida de visceras abdominales por una dehiscencia de todos los planos de la pared, tras una laparotomia. Cuanto mayores son los factores de riesgo mas elevada es la probabilidad de sufrir evisceracion. Pacientes y metodo Estudio retrospectivo de los pacientes con evisceracion tratados en los ultimos 9 anos. Resultados En 12.622 pacientes laparotomizados se detectaron 57 evisceraciones (45 varones y 12 mujeres; edad media 70 anos). El diagnostico fue de peritonitis en 26 casos y de oclusion intestinal en 19. La cirugia fue urgente en 48 pacientes. Fue preciso reintervenir en 12 pacientes. En todos se detectaron complicaciones postoperatorias, entre las que destacaba la infeccion de la herida y el ileo paralitico. En la clinica predomino el manchado del aposito. La analitica mostro leucocitosis, hipoproteinemia y anemia. Se repararon mediante cierre simple y/o puntos totales y en 6 casos se asociaron mallas. La morbilidad posterior fue del 77%. En 26 pacientes fue preciso aplicar cuidados intensivos. La estancia media fue de 28,5 dias. Hubo 41 curaciones (72%) y 16 muertes (28%). Tras revisar 18 parametros implicados en la evisceracion, 45 pacientes (80%) presentaban 9 o mas factores de riesgo. Conclusiones La evisceracion es una enfermedad muy grave, con una elevada morbimortalidad. Los factores de riesgo mas frecuentes, en nuestra serie, fueron: edad > 65 anos, inestabilidad hemodinamica, aumento de la presion intraabdominal, cirugia urgente, infeccion de la herida o de la pared, hipoproteinemia y anemia. Dado que estos factores de riesgo son predecibles, ante la suma de varias de estas causas deberiamos anadir medidas de refuerzo al cierre dela pared abdominal.

Type I iodothyronine 5'-deiodinase mRNA and activity is increased in adipose tissue of obese subjects.

Differentiation and metabolism of adipose tissue are modulated by thyroid hormones (THs), but relatively little is known about the metabolism of THs in this tissue. Expression of the genes for type I iodothyronine 5′-deiodinase (D1), leptin (LEP) and stearoyl-CoA desaturase 1 (SCD-1) was evaluated in omental (OM) and subcutaneous (SC) fat using a cohort of 70 humans. Activities of iodothyronine deiodinases (D1, D2 and D3) were assessed in a randomly selected subpopulation of 19 subjects. D1 expression was upregulated in both OM (P=0.011) and SC (P=0.003) fat of obese subjects. Concomitantly, OM (P=0.002) and SC (P=0.028) LEP expression were increased in obesity, associated with both D1 mRNA (r=0.315, P=0.014) and activity (r=0.647, P=0.023) and inversely related to SCD-1 (r=−0.266, P=0.034) expression in SC fat. Also D1 (but not D2 and D3) activity was increased in OM (∼fourfold, P=0.010) and SC (∼eightfold, P=0.004) fat of obese when compared with non-obese subjects and correlated in both OM (r=0.528, P=0.036) and SC (r=0.749, P=0.005) fat with body mass index. Our results document increased D1 gene expression and activity in adipose tissue of obese humans and suggest a role of 3,5,3′-triiodo-L-thyronine formed by D1 in response to leptin in the modulation of adipose tissue metabolism.

Asymptomatic bilateral adrenal pheochromocytoma in a patient with a germline V804M mutation in the RET proto-oncogene.

A diagnosis of bilateral pheochromocytoma warrants exclusion of hereditary pheochromocytoma.

Study of caveolin-1 gene expression in whole adipose tissue and its subfractions and during differentiation of human adipocytes

ContextCaveolins are 21-24 kDa integral membrane proteins that serve as scaffolds to recruit numerous signaling molecules. Specific subclasses of caveolae carry out specific functions in cell metabolism. In particular, triglycerides are synthesized at the site of fatty acid entry in one of these caveolae classes.Objective and MethodsWe studied the expression of caveolin-1 (CAV-1) gene in association with metabolic variables in 90 visceral and 55 subcutaneous adipose tissue samples from subjects with a wide range of fat mass, in the stromovascular fraction (SVC) and isolated adipocytes, and during differentiation of human adipocytes.ResultsCAV-1 gene expression was significantly decreased in visceral adipose tissue (v-CAV-1) of obese subjects. v-CAV-1 was positively associated with several lipogenic genes such as acetyl-coA carboxylase (ACACA, r = 0.34, p = 0.004) and spot-14 (r = 0.33, p = 0.004). In non-obese subjects v-CAV-1 also correlated with fatty acid synthase (FAS, r = 0.60, p < 0.0001). Subcutaneous (sc) adipose tissue (sc-CAV-1) gene expression was not associated with these lipogenic factors when obese and non-obese subjects were studied together. In obese subjects, however, sc-CAV-1 was associated with fatty acid synthase (FAS, r = 0.36, p = 0.02), sterol regulatory element binding protein-1c (SREBP-1c (r = 0.58, p < 0.0001), ACACA (r = 0.33, p = 0.03), spot-14 (r = 0.36, p = 0.02), PPAR-γ co-activator-1 (PGC-1, r = 0.88, n = 19). In these obese subjects, sc-CAV-1 was also associated with fasting triglycerides (r = -0.50, p < 0.0001).CAV-1 expression in mature adipocytes was significantly higher than in stromal vascular cells. CAV-1 gene expression in adipocytes from subcutaneous adipose tissue (but not in adipocytes from visceral adipose tissue) was significatively associated with fasting triglycerides. CAV-1 gene expression did not change significantly during differentiation of human preadipocytes from lean or obese subjects despite significant increase of FAS gene expression.ConclusionDecreased CAV-1 gene expression was simultaneously linked to increased triglycerides and decreased lipogenic gene expression among obese subjects, paralleling the observations of hypertriglyceridemia in CAV-1 knockout mice. However, the regulation of CAV-1 gene expression seems independent of the adipogenic program.

CIDEC/FSP27 and PLIN1 gene expression run in parallel to mitochondrial genes in human adipose tissue, both increasing after weight loss.

Objective:FSP27 KO mice showed enhanced expression of mitochondrial genes, increased mitochondrial activity and smaller lipid droplets. Here, we aimed to investigate lipid droplet protein (CIDEC/FSP27 and perilipinA (PLIN1)) gene expression in human adipose tissue in association with obesity, insulin resistance and mitochondrial gene expression.Design and subjects:In cohort 1, CIDEC/FSP27, PLIN1, adipogenic (FASN, ACACA, PPARG, GLUT4) and mitochondrial (PPARGC1A, PPARGC1B, TFAM, MT-CO3) gene expression were analyzed in 171 adipose tissue samples (88 visceral adipose tissue (VAT) and 83 subcutaneous adipose tissue (SAT) depots) and in a time course experiment in human subcutaneous and visceral preadipocytes using real-time PCR. In cohort 2, the effects of bariatric surgery-induced weight loss were also evaluated in six caucasian morbidly obese women. Additionally, in cohort 2 FSP27 and PLIN1 protein levels were measured using western blotting.Results:CIDEC/FSP27 (1.03±0.52 vs 0.49±0.23 relative gene expression unit (R.U.), P<0.0001) and PLIN1 (1.32±0.82 vs 0.63±0.42 R.U., P<0.0001) gene were significantly more expressed in SAT than in VAT. In VAT, CIDEC/FSP27 and PLIN1 gene expression decreased with body mass index, percent fat mass, fasting glucose, fasting insulin, HOMA and were positively associated with adipogenic (PPARG, GLUT4, FASN and ACACA) and mitochondrial biogenesis (PPARGC1A, PPARGC1B, TFAM and MT-CO3)-related genes. Mitochondrial gene expression increased during adipocyte differentiation in parallel to FSP27 and PLIN1 and other adipogenic genes. After bariatric surgery-induced weight loss, PLIN1 and CIDEC/FSP27 gene and protein expression in SAT increased significantly in parallel to adipogenic and mitochondrial genes.Conclusion:These findings suggest a positive functional interaction between CIDEC/FSP27, PLIN1 and mitochondrial biogenesis-related genes in human adipose tissue.

Study of lactoferrin gene expression in human and mouse adipose tissue, human preadipocytes and mouse 3T3-L1 fibroblasts. Association with adipogenic and inflammatory markers

Lactoferrin is considered an epithelial protein present in different gland secretions. Administration of exogenous lactoferrin is also known to modulate adipogenesis and insulin action in human adipocytes. Here, we aimed to investigate lactoferrin gene expression (real-time polymerase chain reaction) and protein (enzyme-linked immunosorbent assay) levels in human (n=143) and mice adipose tissue samples, in adipose tissue fractions and during human preadipocyte and 3T3-L1 cell line differentiation, evaluating the effects of inducers (rosiglitazone) and disruptors (inflammatory factors) of adipocyte differentiation. Lactoferrin (LTF) gene and protein were detectable at relatively high levels in whole adipose tissue and isolated adipocytes in direct association with low-density lipoprotein-related protein 1 (LRP1, its putative receptor). Obese subjects with type 2 diabetes and increased triglycerides had the lowest levels of LTF gene expression in subcutaneous adipose tissue. Specifically, LTF gene expression was significantly increased in adipocytes, mainly from lean subjects, increasing during differentiation in parallel to adipogenic genes and gene markers of lipid droplets. The induction or disruption of adipogenesis led to concomitant changes (increase and decrease, respectively) of lactoferrin levels during adipocyte differentiation also in parallel to gene markers of adipogenesis and lipid droplet development. The administration of lactoferrin led to autopotentiated increased expression of the LTF gene. The decreased lactoferrin mRNA levels in association with obesity and diabetes were replicated in mice adipose tissue. In conclusion, this is the first observation, to our knowledge, of lactoferrin gene expression in whole adipose tissue and isolated adipocytes, increasing during adipogenesis and suggesting a possible contribution in adipose tissue physiology through LRP1.