PPARγ lipodystrophy mutants reveal intermolecular interactions required for enhancer activation.

Peroxisome proliferator-activated receptor γ (PPARγ) is the master regulator of adipocyte differentiation, and mutations that interfere with its function cause lipodystrophy. PPARγ is a highly modular protein, and structural studies indicate that PPARγ domains engage in several intra- and inter-molecular interactions. How these interactions modulate PPARγ's ability to activate target genes in a cellular context is currently poorly understood. Here we take advantage of two previously uncharacterized lipodystrophy mutations, R212Q and E379K, that are predicted to interfere with the interaction of the hinge of PPARγ with DNA and with the interaction of PPARγ ligand binding domain (LBD) with the DNA-binding domain (DBD) of the retinoid X receptor, respectively. Using biochemical and genome-wide approaches we show that these mutations impair PPARγ function on an overlapping subset of target enhancers. The hinge region-DNA interaction appears mostly important for binding and remodelling of target enhancers in inaccessible chromatin, whereas the PPARγ-LBD:RXR-DBD interface stabilizes the PPARγ:RXR:DNA ternary complex. Our data demonstrate how in-depth analyses of lipodystrophy mutants can unravel molecular mechanisms of PPARγ function.

Smartphone Application-Assisted Home Blood Pressure Monitoring Compared With Office and Ambulatory Blood Pressure Monitoring in Patients With Hypertension: the AMUSE-BP Study.

BACKGROUND: The development of automated, smartphone application (app)-assisted home blood pressure monitoring (HBPM) allows for standardized measurement of blood pressure (BP) at home. The aim of this study was to evaluate the (diagnostic) agreement between app-assisted HBPM, automated office BP (OBP), and the reference standard 24-hour ambulatory BP monitoring (ABPM). METHODS: In this open randomized 5-way cross-over study, patients diagnosed with hypertension were randomized to one of 10 clusters, each containing 5 BP measurement methods (ABPM, HBPM, attended OBP, unattended OBP, and unattended 30-minute BP) in different order. RESULTS: In total, 113 patients were included. The average 24-hour ABPM was 126±11/73±8 mm Hg compared with 141±14/82±10 mm Hg with app-assisted HBPM, 134±13/80±9 mm Hg with unattended 30-minute BP, 137±16/81±11 mm Hg with attended OBP, and 135±15/81±10 mm Hg with unattended OBP monitoring. Diagnostic agreement between app-assisted HBPM and 24-hour ABPM for diagnosing sustained (OBP >140/90 mm Hg and ABPM ≥130/80 mm Hg or HBPM ≥135/85 mm Hg), white-coat (OBP ≥140/90 mm Hg and ABPM <130/80 mm Hg or HBPM <135/85 mm Hg), and masked hypertension (OBP <140/90 mm Hg and ABPM ≥130/80 mm Hg or HBPM ≥135/85 mm Hg) was fair-to-moderate (κ statistics ranging from 0.34 to 0.40). App-assisted HBPM had high sensitivities (78%-91%) and negative predictive values (90%-97%) for diagnosing sustained and masked hypertension. CONCLUSIONS: This study showed a considerable (diagnostic) disagreement between app-assisted HBPM and ABPM. App-assisted HBPM had high sensitivity in the diagnosis of sustained and masked hypertension and may therefore be used as complementary to, but not a replacement of, ABPM.

Anti-Xa Levels in Morbidly Obese Patients Using Apixaban or Rivaroxaban, Before and After Bariatric Surgery.

BACKGROUND: Despite limited evidence about the efficacy and safety of anticoagulation in patients post bariatric surgery, both vitamin K antagonists (VKA) and direct-acting oral anticoagulants (DOACs) are commonly prescribed. AIM: To evaluate plasma anti-Xa levels of DOACs in morbidly obese (MO) patients before and after a Roux-en-Y gastric bypass (RYGB) procedure. PATIENTS AND METHODS: Retrospective, cross-sectional, and longitudinal study of anti-Xa activity of apixaban or rivaroxaban in MO patients (N = 41). RESULTS: Preoperative analysis of plasma anti-Xa levels were within the normal range in patients using apixaban (n = 29; body mass index [BMI] 44.5 ± 5.1 kg/m2) as well as those using rivaroxaban (n = 12; BMI 42.6 ± 5.9 kg/m2). Postoperative anti-Xa levels of apixaban were all within the therapeutic range, whereas anti-Xa levels of rivaroxaban were subtherapeutic in nine out of 14 (64%) patients. Perioperative longitudinal follow-up in patients using apixaban (n = 18) showed no significant change in anti-Xa levels after RYGB. CONCLUSION: Plasma anti-Xa levels of apixaban in MO patients remained within the therapeutic range up to a body weight of 144 kg. In patients using rivaroxaban, no statistically significant relation between anti-Xa levels and bodyweight was found. After RYGB, plasma anti-Xa levels of apixaban were unaffected, whereas plasma anti-Xa levels of rivaroxaban tended to become subtherapeutic.

Assessment of Imaging Modalities Against Liver Biopsy in Nonalcoholic Fatty Liver Disease: The Amsterdam NAFLD-NASH Cohort.

BACKGROUND: Noninvasive diagnostic methods are urgently required in disease stratification and monitoring in nonalcoholic fatty liver disease (NAFLD). Multiparametric magnetic resonance imaging (MRI) is a promising technique to assess hepatic steatosis, inflammation, and fibrosis, potentially enabling noninvasive identification of individuals with active and advanced stages of NAFLD. PURPOSE: To examine the diagnostic performance of multiparametric MRI for the assessment of disease severity along the NAFLD disease spectrum with comparison to histological scores. STUDY TYPE: Prospective, cohort. POPULATION: Thirty-seven patients with NAFLD. FIELD STRENGTH/SEQUENCE: Multiparametric MRI at 3.0 T consisted of magnetic resonance (MR) spectroscopy (MRS) with multi-echo stimulated-echo acquisition mode, magnitude-based and three-point Dixon using a two-dimensional multi-echo gradient echo, MR elastography (MRE) using a generalized multishot gradient-recalled echo sequence and intravoxel incoherent motion (IVIM) using a multislice diffusion weighted single-shot echo-planar sequence. ASSESSMENT: Histological steatosis grades were compared to proton density fat fraction measured by MRS (PDFFMRS ), magnitude-based MRI (PDFFMRI-M ), and three-point Dixon (PDFFDixon ), as well as FibroScan® controlled attenuation parameter (CAP). Fibrosis and disease activity were compared to IVIM and MRE. FibroScan® liver stiffness measurements were compared to fibrosis levels. Diagnostic performance of all imaging parameters was determined for distinction between simple steatosis and nonalcoholic steatohepatitis (NASH). STATISTICAL TESTS: Spearman's rank test, Kruskal-Wallis test, Dunn's post-hoc test with Holm-Bonferroni P-value adjustment, receiver operating characteristic curve analysis. A P-value <0.05 was considered statistically significant. RESULTS: Histological steatosis grade correlated significantly with PDFFMRS (rs  = 0.66, P < 0.001), PDFFMRI-M (rs  = 0.68, P < 0.001), and PDFFDixon (rs  = 0.67, P < 0.001), whereas no correlation was found with CAP. MRE and IVIM diffusion and perfusion significantly correlated with disease activity (rs  = 0.55, P < 0.001, rs  = -0.40, P = 0.016, rs  = -0.37, P = 0.027, respectively) and fibrosis (rs  = 0.55, P < 0.001, rs  = -0.46, P = 0.0051; rs  = -0.53, P < 0.001, respectively). MRE and IVIM diffusion had the highest area-under-the-curve for distinction between simple steatosis and NASH (0.79 and 0.73, respectively). DATA CONCLUSION: Multiparametric MRI is a promising method for noninvasive, accurate, and sensitive distinction between simple hepatic steatosis and NASH, as well as for the assessment of steatosis and fibrosis severity. LEVEL OF EVIDENCE: 2 TECHNICAL EFFICACY: 2.

Arterial Wall Inflammation and Increased Hematopoietic Activity in Patients With Primary Aldosteronism.

CONTEXT: Primary aldosteronism (PA) confers an increased risk of cardiovascular disease (CVD), independent of blood pressure. Animal models have shown that aldosterone accelerates atherosclerosis through proinflammatory changes in innate immune cells; human data are scarce. OBJECTIVE: The objective of this article is to explore whether patients with PA have increased arterial wall inflammation, systemic inflammation, and reprogramming of monocytes. DESIGN: A cross-sectional cohort study compared vascular inflammation on 2'-deoxy-2'-(18F)fluoro-D-glucose; (18F-FDG) positron emission tomography-computed tomography, systemic inflammation, and monocyte phenotypes and transcriptome between PA patients and controls. SETTING: This study took place at Radboudumc and Rijnstate Hospital, the Netherlands. PATIENTS: Fifteen patients with PA and 15 age-, sex-, and blood pressure-matched controls with essential hypertension (EHT) participated. MAIN OUTCOME MEASURES AND RESULTS: PA patients displayed a higher arterial 18F-FDG uptake in the descending and abdominal aorta (P < .01, P < .05) and carotid and iliac arteries (both P < .01). In addition, bone marrow uptake was higher in PA patients (P < .05). Although PA patients had a higher monocyte-to-lymphocyte ratio (P < .05), systemic inflammatory markers, cytokine production capacity, and transcriptome of circulating monocytes did not differ. Monocyte-derived macrophages from PA patients expressed more TNFA; monocyte-derived macrophages of healthy donors cultured in PA serum displayed increased interleukin-6 and tumor necrosis factor-α production. CONCLUSIONS: Because increased arterial wall inflammation is associated with accelerated atherogenesis and unstable plaques, this might importantly contribute to the increased CVD risk in PA patients. We did not observe inflammatory reprogramming of circulating monocytes. However, subtle inflammatory changes are present in the peripheral blood cell composition and monocyte transcriptome of PA patients, and in their monocyte-derived macrophages. Most likely, arterial inflammation in PA requires interaction between various cell types.

Plasma levels of the cardiovascular protective endogenous nucleoside adenosine are reduced in patients with primary aldosteronism without affecting ischaemia-reperfusion injury: A prospective case-control study.

BACKGROUND: Patients with primary aldosteronism (PA) experience more cardiovascular events compared to patients with essential hypertension (EHT), independent from blood pressure levels. In animals, mineralocorticoid receptor antagonists limit ischaemia-reperfusion (IR) injury by increasing extracellular adenosine formation and adenosine receptor stimulation. Adenosine is an endogenous compound with profound cardiovascular protective effects. Firstly, we hypothesized that patients with PA have lower circulating adenosine levels which might contribute to the observed increased cardiovascular risk. Secondly, we hypothesized that by this mechanism, patients with PA are more susceptible to IR compared to patients with EHT. DESIGN: In our prospective study in 20 patients with PA and 20 patients with EHT, circulating adenosine was measured using a pharmacological blocker solution that halts adenosine metabolism after blood drawing. Brachial artery flow-mediated dilation (FMD) before and after forearm IR was used as a well-established method to study IR injury. RESULTS: Patients with PA had a 33% lower adenosine level compared to patients with EHT (15.3 [13.3-20.4] vs 22.7 [19.4-36.8] nmol/L, respectively, P < .01). The reduction in FMD after IR, however, did not differ between patients with PA and patients with EHT (-1.0 ± 2.9% vs -1.6 ± 1.6%, respectively, P = .52). CONCLUSIONS: As adenosine receptor stimulation induces various powerful protective cardiovascular effects, its lower concentration in patients with PA might be an important novel mechanism that contributes to their increased cardiovascular risk. We suggest that modulation of the adenosine metabolism is an exciting novel pharmacological opportunity to limit cardiovascular risk in patients with PA that needs further exploration.

Natural helix 9 mutants of PPARγ differently affect its transcriptional activity.

OBJECTIVE: The nuclear receptor PPARγ is the master regulator of adipocyte differentiation, distribution, and function. In addition, PPARγ induces terminal differentiation of several epithelial cell lineages, including colon epithelia. Loss-of-function mutations in PPARG result in familial partial lipodystrophy subtype 3 (FPDL3), a rare condition characterized by aberrant adipose tissue distribution and severe metabolic complications, including diabetes. Mutations in PPARG have also been reported in sporadic colorectal cancers, but the significance of these mutations is unclear. Studying these natural PPARG mutations provides valuable insights into structure-function relationships in the PPARγ protein. We functionally characterized a novel FPLD3-associated PPARγ L451P mutation in helix 9 of the ligand binding domain (LBD). Interestingly, substitution of the adjacent amino acid K450 was previously reported in a human colon carcinoma cell line. METHODS: We performed a detailed side-by-side functional comparison of these two PPARγ mutants. RESULTS: PPARγ L451P shows multiple intermolecular defects, including impaired cofactor binding and reduced RXRα heterodimerisation and subsequent DNA binding, but not in DBD-LBD interdomain communication. The K450Q mutant displays none of these functional defects. Other colon cancer-associated PPARγ mutants displayed diverse phenotypes, ranging from complete loss of activity to wildtype activity. CONCLUSIONS: Amino acid changes in helix 9 can differently affect LBD integrity and function. In addition, FPLD3-associated PPARγ mutations consistently cause intra- and/or intermolecular defects; colon cancer-associated PPARγ mutations on the other hand may play a role in colon cancer onset and progression, but this is not due to their effects on the most well-studied functional characteristics of PPARγ.

A Single Complex Agpat2 Allele in a Patient With Partial Lipodystrophy.

Genetic lipodystrophies are a group of rare syndromes associated with major metabolic complications - including severe insulin resistance, type 2 diabetes mellitus, and hypertriglyceridemia - which are classified according to the distribution of adipose tissue. Lipodystrophies can be present at birth or develop during life and can range from local to partial and general. With at least 18 different genes implicated so far, definite diagnosis can be challenging due to clinical and genetic heterogeneity. In an adult female patient with clinical and metabolic features of partial lipodystrophy we identified via whole genome sequencing (WGS) a single complex AGPAT2 allele [V67M;V167A], functionally equivalent to heterozygosity. AGPAT2 encodes for an acyltransferase implicated in the biosynthesis of triacylglycerol and glycerophospholipids. So far homozygous and compound heterozygous mutations in AGPAT2 have only been associated with generalized lipodystrophy. A SNP risk score analysis indicated that the index patient is not predisposed to lipodystrophy based on her genetic background. The partial phenotype in our patient is therefore more likely associated to the genetic variants in AGPAT2. To test whether the resulting double-mutant AGPAT2 protein is functional we analyzed its in vitro enzymatic activity via mass spectrometry. The resulting AGPAT2 double mutant is enzymatically inactive. Our data support the view that the current classification of lipodystrophies as strictly local, partial or generalized may have to be re-evaluated and viewed more as a continuum, both in terms of clinical presentation and underlying genetic causes. Better molecular understanding of lipodystrophies may lead to new therapies to treat adipose tissue dysfunction in common and rare diseases.

An oral mixed fat load is followed by a modest anti-inflammatory adipocytokine response in overweight patients with metabolic syndrome.

We investigated the postprandial changes in plasma levels of adipocytokines in overweight patients with metabolic syndrome after an oral fat load. After an oral fat load and during a prolonged fast, blood was drawn at 0, 2, 3, 4 and 8 h for measurement of adiponectin, adipsin, cathepsin S, chemerin, hepatic growth factor, interferon-γ-inducible protein-10, leptin, macrophage chemoattractant protein-1, macrophage migration inhibitory factor, nerve growth factor, retinol binding protein-4, resistin, serum amyloid A1, tissue inhibitor of metalloproteinase-1 and thrombopoietin using a microbead-based Luminex assay. Area under the curves (AUC) were calculated and compared. Plasma adiponectin levels were higher after an oral fat load compared to fasting at t = 2 h (950 ± 513 vs. -1,881 ± 713 ng/ml) while the plasma levels for adipsin (-9 ± 5 vs. 16 ± 5 ng/ml), chemerin (-122 ± 35 vs. 13 ± 21 ng/ml), SAA-1 (-391 ± 213 vs. 522 ± 173 ng/ml) and TPO (-335 ± 144 vs. 622 ± 216 ng/ml) were lower after an oral fat load compared to fasting. The baseline corrected AUC for IP-10 was higher after fat load compared to fasting (median -116 pg h/ml; IQR -270 to 10 vs. -21 pg h/ml; IQR -136 to 418 (p = 0.047). In conclusion, in overweight male subjects with the metabolic syndrome, an oral fat load is accompanied with a modest anti-inflammatory response of adipose tissue-derived adipocytokines.

The relation between body iron stores and adipose tissue function in patients with manifest vascular disease.

BACKGROUND: We investigated whether plasma ferritin levels through the pro-inflammatory effects of free iron are associated with adipose tissue dysfunction in a relevant population of patients with manifest vascular disease who would potentially benefit the most from further aetiological insights. MATERIALS AND METHODS: In a cohort of 355 patients with vascular diseases, the association between plasma ferritin and adiponectin levels was quantified using linear regression analysis. Interleukin-6 and adiponectin levels were measured in medium from pre-adipocytes and adipocytes after incubation with increasing concentrations of Fe(III)-citrate and after co-incubation with iron chelators or radical scavengers. RESULTS: Increasing ferritin plasma concentrations were not related to plasma adiponectin levels in patients without (ß -0·13; 95% CI -0·30 to 0·04) or with the metabolic syndrome (ß -0·04; 95% CI -0·17 to 0·10). Similar results were found in patients who developed a new cardiovascular event in the follow-up period. In vitro, incubation with increasing concentrations of Fe(III)-citrate-induced inflammation in pre-adipocyte cultures as witnessed by increased IL-6 secretion at 30 µm Fe(III)-citrate vs. control (500 ± 98 pg/mL vs. 194 ± 31 pg/mL, P = 0·03). Co-incubation of pre-adipocytes with iron chelators or radical scavengers prevented this inflammatory response. Incubation of adipocytes with 30 µm Fe(III)-citrate did not influence adiponectin secretion compared with control. CONCLUSIONS: In patients with vascular disease, there is no association between plasma ferritin and adiponectin levels. In vitro, free iron induces an inflammatory response in pre-adipocytes, but not in adipocytes. This response was blocked by co-incubation with iron chelators or radical scavengers. Adiponectin secretion by adipocytes was not influenced by free iron.

Hepatocyte growth factor and interferon-γ inducible protein-10 are related to visceral adiposity.

BACKGROUND: Increased production of chemokines by adipose tissue and defective adipose tissue oxygenation as a result of obesity may induce leucocyte infiltration and subsequent systemic inflammation. OBJECTIVES: 1-To determine the relation between the amount of visceral and subcutaneous adipose tissue and the chemokine interferon-γ-inducible protein 10 (IP-10) and angiogenic factor hepatocyte growth factor (HGF). 2-To determine the relation between the metabolic syndrome and IP-10 as well as HGF. METHODS: Patients originated from the Secondary Manifestations of ARTerial disease (SMART) cohort. In this study, a cohort of 1251 patients with manifest vascular disease was included. Subcutaneous and visceral adipose tissue thickness (SAT and VAT respectively) were measured ultrasonographically. IP-10 and HGF concentrations were measured with Luminex multiplex immuno assay in addition to fasting metabolic parameters. Linear regression analyses with adjustments for age, gender, smoking, estimated glomerular filtration rate, type 2 diabetes mellitus and medication use were applied to quantify the relations between adiposity or metabolic syndrome and IP-10 and HGF concentrations. RESULTS: VAT was significantly associated with (log)IP-10 and (log)HGF, reflected by significant higher ß-values in VAT quartile 4 compared with VAT quartile 1 (reference): ß0.155 (95%CI:0.073-0.237) for IP-10 and ß0.147 (95%CI:0.076-0.218) for HGF. Per standard deviation increase in VAT, (log)IP-10 levels increased with 0.057 pg/mL (95%CI:0.027-0.087) and (log)HGF increased with 0.051 pg/mL (95%CI:0.025-0.077). Effect estimates were not affected by including body mass index(BMI) in the model. In contrast, SAT was not associated with IP-10 and HGF. Furthermore, the presence of the metabolic syndrome was associated with IP-10 and HGF. CONCLUSIONS: Visceral adipose tissue but not subcutaneous adipose tissue is significantly associated with circulating levels of IP-10 and HGF, irrespective of BMI.

Plasma triglyceride levels increase the risk for recurrent vascular events independent of LDL-cholesterol or nonHDL-cholesterol.

BACKGROUND: Plasma triglyceride (TG) levels are known to confer an increased risk of vascular disease in healthy populations, but data in high-risk patients are scarce. In this study we evaluated the risk on recurrent vascular events conferred by increased plasma TG levels in patients with various clinical manifestations of vascular disease. METHODS: Prospective cohort study of 5731 patients with clinically manifest vascular disease. RESULTS: First new vascular events (myocardial infarction, ischemic stroke, vascular death) occurred in 782 subjects during a median follow-up of 4.9 years (interquartile range 2.5-8.1 years). Patients in the highest plasma TG quintile (>2.24 mmol/L) had a higher risk for recurrent vascular events (HR 1.45; 95%CI 1.13-1.86) compared with the lowest plasma TG quintile (<0.97 mmol/L) after adjustments for age, gender, body mass index, smoking, lipid-lowering medication and low-density lipoprotein-cholesterol. The increased risk associated with increasing plasma TG levels was irrespective of the presence of type 2 diabetes (T2DM), but only present in patients without the metabolic syndrome. Furthermore, the increased risk was particularly present in patients with coronary artery disease (CAD) (HR 1.45; 95%CI 1.02-2.08) and was not modified by other lipid levels (p-value for interaction >0.05). Plasma TG still contributed to vascular risk when other lipid levels were at target level. CONCLUSIONS: Higher plasma TG levels are associated with increased risk for recurrent vascular events, in particular in CAD patients. This increased risk is independent of the presence of T2DM and the use of lipid-lowering medication and is not modified by other lipid levels.

Obesity and dyslipidemia.

Dyslipidemia associated with obesity and the metabolic syndrome is one of the central features contributing to the increased CV risk in these patients. In view of the pandemic of the metabolic syndrome, it is imperative to fully understand the mechanisms leading to the metabolic lipid phenotype before embarking upon optimal treatment strategies. The traditional concept that insulin resistance causes increased FFA flux via increased TG hydrolysis in adipose tissue is still of a central theme in the general hypothesis. The combination of increased hepatic VLDL secretion with impaired LPL-mediated TG clearance explains the hypertriglyceridemia phenotype of the metabolic syndrome. Hence, central IR may be an important factor contributing to peripheral hypertriglyceridemia. Recently recognized regulatory systems include the profound impact of the hypothalamus on TG secretion and glucose control. In addition, dysfunctional (or inflamed) intra abdominal adipose tissue has emerged as a potent regulator of dyslipidemia and IR. It will be a challenge to design novel treatment modalities that target "dysfunctional" fat or central IR to attempt to prevent the epidemic of CV disease secondary to the metabolic syndrome.

Obesity and dyslipidemia.

The alarming and still increasing prevalence of obesity and associated cardiovascular risk raises much concern. The increase in cardiovascular risk depends to a significant extent on the changes in lipid profiles as observed in obesity. These changes are decreased high-density lipoprotein cholesterol and increased triglyceride levels. Much effort has already been expended into the elucidation of the mechanisms behind these obesity-associated lipid changes. Insulin resistance certainly plays a central role and, in addition, both hormonal and neurologic pathways have recently been found to play an important role. This article focuses on the mechanisms involved in the development of the proatherogenic lipid changes associated with obesity.

Inherited lipodystrophies and the metabolic syndrome.

Lipodystrophies represent a heterogeneous group of diseases characterized by an abnormal subcutaneous fat distribution, the extent of which can vary from localized, to partial, to generalized lipoatrophy. Whereas partial and generalized lipodystrophies are each associated with metabolic abnormalities, the localized form is not. These metabolic changes include insulin resistance with type 2 diabetes, acanthosis nigricans, dyslipidaemia predominantly consisting of hypertriglyceridaemia (associated with the onset of pancreatitis) and depressed HDL cholesterol, liver steatosis and hypertension. Affected women are often hirsute and this can be associated with the presence of polycystic ovarian syndrome (PCOS). Most of these clinical features are present to some extent in patients with the common metabolic syndrome. As the prevalence of metabolic syndrome far outweighs that of lipodystrophy, the diagnosis of this rare disorder may often be overlooked with the affected patient diagnosed as merely being 'yet' another case of metabolic syndrome. In this article, we draw attention to the importance of recognizing patients with lipodystrophy who present with metabolic abnormalities, as both the diagnostic as well as the therapeutic approach of these patients differ profoundly from patients with the metabolic syndrome.

Familial partial lipodystrophy phenotype resulting from a single-base mutation in deoxyribonucleic acid-binding domain of peroxisome proliferator-activated receptor-gamma.

CONTEXT: Familial partial lipodystrophy (FPLD) results from coding sequence mutations either in LMNA, encoding nuclear lamin A/C, or in PPARG, encoding peroxisome proliferator-activated receptor-gamma (PPARgamma). The LMNA form is called FPLD2 (MIM 151660) and the PPARG form is called FPLD3 (MIM 604367). OBJECTIVE: Our objective was to investigate whether the clinical phenotype of this proband is due to mutation(s) in PPARgamma. DESIGN: This is a case report. Patient and Setting: A 31-yr-old female with the clinical phenotype of FPLD3, i.e. lipodystrophy and early childhood diabetes with extreme insulin resistance and hypertriglyceridemia leading to recurrent pancreatitis, was assessed at an academic medical center. RESULTS: The proband was heterozygous for a novel C-->T mutation in the PPARG gene that led to the substitution of arginine 194 in PPARgamma2 isoform, a conserved residue located in the zinc finger structure involved in DNA binding, by tryptophan (R194W). The mutation was absent from the genomes of 100 healthy Caucasians. In vitro analysis of the mutated protein showed that R194W (and R166W in PPARgamma1 isoform) could not bind to DNA and had no transcriptional activity. Furthermore, R194W had no dominant-negative activity. CONCLUSIONS: The R194W mutation in PPARG disrupts its DNA binding activity and through haploinsufficiency leads to clinical manifestation of FPLD3 and the associated metabolic disturbances.

Lipase H, a new member of the triglyceride lipase family synthesized by the intestine.

We report here the molecular cloning of a novel member of the triglyceride lipase family, a 2.4-kb cDNA encoding human lipase H (LIPH) and the mouse ortholog (Liph). The human LIPH cDNA encodes a 451-amino-acid protein with a lipase domain. Mouse Liph shows 85% amino acid identity and 75% nucleotide identity to human LIPH. Human LIPH exhibits 47% identity with phosphatidylserine-specific phospholipase A1 (PS-PLA1) and 46% identity with endothelial lipase (LIPG) and lipoprotein lipase (LPL). LIPH is localized on human chromosome 3q27-q28. Northern blot analysis revealed specific expression of LIPH mRNA in intestine, lung, and pancreas. Lipase H protein was also detected in human intestine. Lipase H is a secreted protein with an apparent molecular weight of 63 kDa. Although several lipid substrates were tested, the lipid substrate of LIPG was not identified. Like the other members of this gene family, LIPH may be involved in lipid and energy metabolism.

The apolipoprotein L gene cluster has emerged recently in evolution and is expressed in human vascular tissue.

We previously isolated APOL3 (CG12-1) cDNA and now describe the isolation of APOL1 and APOL2 cDNA from an activated endothelial cell cDNA library and show their endothelialspecific expression in human vascular tissue. APOL1-APOL4 are clustered on human chromosome 22q13.1, as a result of tandem gene duplication, and were detected only in primates (humans and African green monkeys) and not in dogs, pigs, or rodents, showing that this gene cluster has arisen recently in evolution. The specific tissue distribution and gene organization suggest that these genes have diverged rapidly after duplication. This has resulted in the emergence of an additional signal peptide encoding exon that ensures secretion of the plasma high-density lipoprotein-associated APOL1. Our results show that the APOL1-APOL4 cluster might contribute to the substantial differences in the lipid metabolism of humans and mice, as dictated by the variable expression of genes involved in this process.