Abdul B. Abou-Samra

PTH and PTHrP Signaling in Osteoblasts

The striking clinical benefit of PTH in osteoporosis began a new era of skeletal anabolic agents. Several studies have been performed, new studies are emerging out and yet controversies remain on PTH anabolic action in bone. This review focuses on the molecular aspects of PTH and PTHrP signaling in light of old players and recent advances in understanding the control of osteoblast proliferation, differentiation and function.

Elevated circulating lipasin/betatrophin in human type 2 diabetes and obesity

Lipasin (also known as C19ORF80, RIFL, ANGPTL8 and betatrophin) is a newly discovered circulating factor that regulates lipid metabolism and promotes pancreatic β-cell proliferation. Whether circulating levels of lipasin in humans are altered in a) type 2 diabetes; b) obesity and c) the postprandial state, however, is unknown. The current study aimed to compare serum lipasin levels in those who were a) non-diabetic (N = 15) or diabetic (BMI- and age-matched; N = 14); b) lean or obese (N = 53 totally) and c) fasting and 2 hours following a defined meal (N = 12). Serum lipasin levels were determined by the enzyme-linked immunosorbent assay. Lipasin levels [mean ± SEM] were increased by more than two fold (P < 0.001) in the diabetic patients (5.56 ± 0.73 ng/mL) as compared to the control subjects (2.19 ± 0.24 ng/mL). Serum lipasin levels were positively correlated with BMI (rho = 0.49, P < 0.001), and showed a 35% increase 2 hours following a defined meal (P = 0.009). Therefore, lipasin/betatrophin is nutritionally-regulated hepatokine that is increased in human type 2 diabetes and obesity.

An explanation for recent discrepancies in levels of human circulating betatrophin.

To the Editor: In addition to hyperglycaemia and hyperinsulinaemia, type 2 diabetes mellitus is often associated with hypertriglyceridaemia. Recently, we and others have identified a novel circulating factor, referred to as lipasin [1], refeeding induced in fat and liver (RIFL) [2], angiopoietin-like protein 8 (ANGPTL8) [3, 4], and betatrophin [5], which may have a dual role in mediating both triacylglycerol metabolism and glucose homeostasis. Lipasin mRNA levels in liver and fat, where it is predominantly expressed, are suppressed by fasting and highly induced by feeding and insulin resistance [1–3, 5]. Serum triacylglycerol levels are increased in lipasin overexpressing mice and decreased in lipasin knockout mice, and this likely occurs through a mechanism involving regulation of the activity of lipoprotein lipase (LPL) either directly [1] or indirectly by promoting ANGPTL3 cleavage [3]. Indeed, the knockout mice exhibit higher LPL activity [4]. The discovery of betatrophin as a potent and specific stimulator of pancreatic beta cell proliferation has drawn significant attention because it represents an intriguing therapeutic target for promoting beta cell regeneration in both type 1 and type 2 diabetes [5]. As a logical next step to understanding the roles of betatrophin in human physiology and pathology, there is a growing interest in examining circulating levels of the protein in humans [6–9]. Betatrophin levels have been found to be increased in type 1 [6] and type 2 diabetes [8, 9], and to be associated with atherogenic lipid profiles [7] and BMI [9]. However, there are notable discrepancies among results of these studies. For instance, levels of fasting betatrophin (1) ranged from 0.3 ng/ml [6] to 2.2 ng/ml [9] in lean and nondiabetic individuals; (2) were uncorrelated [6, 7] or positively correlated with BMI [9]; and (3) were either unaltered [7] or increased in type 2 diabetes [8, 9]. We propose that all these results can be correct, but they reflect different betatrophin species, because different ELISA kits, although accurate and reliable, can generate different results because of betatrophin proteolytic regulation [10]. We compared the betatrophin kits manufactured by EIAAB (Catalogue no. E1164H; Wuhan, China) and Phoenix Pharmaceuticals (Catalogue no. EK-051-55; Burlingame, CA, USA). When measuring a human recombinant betatrophin (EIAAB), both kits were accurate, and the correlation coefficient of values based on the two kits was 0.99 by linear regression (Fig. 1a). We next examined circulating betatrophin levels in sera collected by Valley Biomedical (Winchester, VA, USA) from 30 human male individuals (BMI range 17.8 to 58.1 kg/m) following overnight fasting; aliquoted samples were saved at −80°C. The study participants were given informed consent and the study protocol was approved by the Institutional Review Boards of Wayne State University. Based on the Phoenix kit, the mean serum betatrophin level in lean individuals (BMI <25 kg/m) was significantly lower than that in overweight and obese individuals (BMI ≥25), (2.36±0.25 ng/ml vs 3.63±0.29 ng/ml [mean±SEM]; Z. Fu :A. B. Abou-Samra :R. Zhang (*) Center for Molecular Medicine and Genetics, School of Medicine, Wayne State University, 540 East Canfield Street, Detroit, MI 48201, USA e-mail: rzhang@med.wayne.edu

Lipasin, thermoregulated in brown fat, is a novel but atypical member of the angiopoietin-like protein family.

Hyperlipidemia is a major contributor to cardiovascular diseases. Members of the angiopoietin-like protein family (ANGPTLs) are important determinants of blood lipid levels. Lipasin, a newly identified gene that regulates serum triglycerides, is homologous to ANGPTL3s N-terminal domain, which is sufficient and necessary for blood lipid regulation. Brown fat is critical in mediating energy homeostasis. Thermogenesis is the primary function of brown fat, in which Lipasin and some ANGPTLs are abundant; it is unknown, however, whether these genes are thermoregulated. We therefore comprehensively examined the thermoregulation of Lipasin and ANGPTLs in brown fat. Here we show that Lipasin is a novel but atypical member of the ANGPTL family because it is within the same branch as ANGPTL3 and 4 by phylogenetic analysis. The mRNA levels of Lipasin are dramatically increased in the cold environment (4 °C for 4 h) whereas those of ANGPTL4 and ANGPTL2 are suppressed. Fasting dramatically suppresses Lipasin but increases ANGPTL4. High-fat diet treatment increases Lipasin, but reduces ANGPTL2. The distinct transcriptional regulations of Lipasin, ANGPTL2 and ANGPTL4 in brown fat in response to cold exposure and nutritional stimulation suggest distinct physiological roles for ANGPTL family members in mediating thermogenesis and energy homeostasis.

A dual role of lipasin (betatrophin) in lipid metabolism and glucose homeostasis: consensus and controversy

Metabolic syndrome includes glucose intolerance and dyslipidemia, both of which are strong risk factors for developing diabetes and atherosclerotic cardiovascular diseases. Recently, multiple groups independently studied a previously uncharacterized gene, officially named C19orf80 (human) and Gm6484 (mouse), but more commonly known as RIFL, Angptl8, betatrophin and lipasin. Both exciting and conflicting results have been obtained, and significant controversy is ongoing. Accumulating evidence from genome wide association studies and mouse genetic studies convincingly shows that lipasin is involved in lipid regulation. However, the mechanism of action, the identity of transcription factors mediating its nutritional regulation, circulating levels, and relationship among lipasin, Angptl3 and Angptl4, remain elusive. Betatrophin represents a promising drug target for replenishing β-cell mass, but current results have not been conclusive regarding its potency and specificity. Here, we summarize the consensus and controversy regarding functions of lipasin/betatrophin based on currently available evidence.

Distinct roles for Mitogen Activated Protein Kinase Phosphatase-1 (MKP-1) and ERK-MAPK in PTH1R signaling during osteoblast proliferation and differentiation

Parathyroid hormone (PTH) and PTH-related protein (PTHrP) activate one single receptor (PTH1R) which mediates catabolic and anabolic actions in the bone. Activation of PTH1R modulates multiple intracellular signaling responses. We previously reported that PTH and PTHrP down-regulate pERK1/2 and cyclin D1 in differentiated osteoblasts. In this study we investigate the role of MAPK phosphatase-1 (MKP-1) in PTHrP regulation of ERK1/2 activity in relation to osteoblast proliferation, differentiation and bone formation. Here we show that PTHrP increases MKP-1 expression in differentiated osteoblastic MC3T3-E1 cells, primary cultures of differentiated bone marrow stromal cells (BMSCs) and calvarial osteoblasts. PTHrP had no effect on MKP-1 expression in proliferating osteoblastic cells. Overexpression of MKP-1 in MC-4 cells inhibited osteoblastic cell proliferation. Cell extracts from differentiated MC-4 cells treated with PTHrP inactivate/dephosphorylate pERK1/2 in vitro; immunodepletion of MKP-1 blocked the ability of the extract to dephosphorylate pERK1/2; these data indicate that MKP-1 is involved in PTHrP-induced pERK1/2 dephosphorylation in the differentiated osteoblastic cells. PTHrP regulation of MKP-1 expression is partially dependent on PKA and PKC pathways. Treatment of nude mice, bearing ectopic ossicles, with intermittent PTH for 3weeks, up-regulated MKP-1 and osteocalcin, a bone formation marker, with an increase in bone formation. These data indicate that PTH and PTHrP increase MKP-1 expression in differentiated osteoblasts; and that MKP-1 induces growth arrest of osteoblasts, via inactivating pERK1/2 and down-regulating cyclin D1; and identify MKP-1 as a possible mediator of the anabolic actions of PTH1R in mature osteoblasts.

Relationship of vitamin D and parathyroid hormone with obesity and body composition in African Americans

Background  Obesity disproportionately affects African Americans (AA) (especially women), and is linked to depressed 25‐hydroxyvitamin D (25‐OH D) and elevated parathyroid hormone (PTH). The relationship of 25‐OH D and PTH with body composition and size in AA is not well known.

Nuclear export mediated regulation of microRNAs: potential target for drug intervention.

MicroRNAs (miRNAs) are short non-coding RNAs that have been recognized to regulate the expression of uncountable number of genes. Their aberrant expression has been found to be linked to the pathology of many diseases including cancer. There is a drive to develop miRNA targeted therapeutics for different diseases especially cancer. Nevertheless, reining in these short non-coding RNAs is not as straightforward as originally thought. This is in view of the recent discoveries that miRNAs are under epigenetic regulations at multiple levels. Exportin 5 protein (XPO5) nuclear export mediated regulation of miRNAs is one such important epigenetic mechanism. XPO5 is responsible for exporting precursor miRNAs through the nuclear membrane to the cytoplasm, and is thus a critical step in miRNA biogenesis. A number of studies have shown that variations in components of the miRNA biogenesis pathways, particularly the aberrant expression of XPO5, increase the risk of developing cancer. In addition to XPO5, the Exportin 1 protein (XPO1) or chromosome region maintenance 1 (CRM1) can also carry miRNA export function. These findings are supported by pathway analyses that reveal certain miRNAs as direct interaction partners of CRM1. An in depth understanding of miRNA export mediated regulatory mechanisms is important for the successful design of clinically viable therapeutics. In this review, we describe the current knowledge on the mechanisms of miRNA nuclear transport mediated regulation and propose strategies to selectively block this important mechanism in cancer.

A lipasin/Angptl8 monoclonal antibody lowers mouse serum triglycerides involving increased postprandial activity of the cardiac lipoprotein lipase

Lipasin/Angptl8 is a feeding-induced hepatokine that regulates triglyceride (TAG) metabolism; its therapeutical potential, mechanism of action, and relation to the lipoprotein lipase (LPL), however, remain elusive. We generated five monoclonal lipasin antibodies, among which one lowered the serum TAG level when injected into mice, and the epitope was determined to be EIQVEE. Lipasin-deficient mice exhibited elevated postprandial activity of LPL in the heart and skeletal muscle, but not in white adipose tissue (WAT), suggesting that lipasin suppresses the activity of LPL specifically in cardiac and skeletal muscles. Consistently, mice injected with the effective antibody or with lipasin deficiency had increased postprandial cardiac LPL activity and lower TAG levels only in the fed state. These results suggest that lipasin acts, at least in part, in an endocrine manner. We propose the following model: feeding induces lipasin, activating the lipasin-Angptl3 pathway, which inhibits LPL in cardiac and skeletal muscles to direct circulating TAG to WAT for storage; conversely, fasting induces Angptl4, which inhibits LPL in WAT to direct circulating TAG to cardiac and skeletal muscles for oxidation. This model suggests a general mechanism by which TAG trafficking is coordinated by lipasin, Angptl3 and Angptl4 at different nutritional statuses.

Increased Interaction with Insulin Receptor Substrate-1, a Novel Abnormality in Insulin Resistance and Type 2 Diabetes

Insulin receptor substrate 1 (IRS1) is a key mediator of insulin signal transduction. Perturbations involving IRS1 complexes may lead to the development of insulin resistance and type 2 diabetes (T2D). Surprisingly little is known about the proteins that interact with IRS1 in humans under health and disease conditions. We used a proteomic approach to assess IRS1 interaction partners in skeletal muscle from lean healthy control subjects (LCs), obese insulin-resistant nondiabetic control subjects (OCs), and participants with T2D before and after insulin infusion. We identified 113 novel endogenous IRS1 interaction partners, which represents the largest IRS1 interactome in humans and provides new targets for studies of IRS1 complexes in various diseases. Furthermore, we generated the first global picture of IRS1 interaction partners in LCs, and how they differ in OCs and T2D patients. Interestingly, dozens of proteins in OCs and/or T2D patients exhibited increased associations with IRS1 compared with LCs under the basal and/or insulin-stimulated conditions, revealing multiple new dysfunctional IRS1 pathways in OCs and T2D patients. This novel abnormality, increased interaction of multiple proteins with IRS1 in obesity and T2D in humans, provides new insights into the molecular mechanism of insulin resistance and identifies new targets for T2D drug development.