Transglutaminases and Obesity in Humans: Association of F13A1 to Adipocyte Hypertrophy and Adipose Tissue Immune Response

Transglutaminases TG2 and FXIII-A have recently been linked to adipose tissue biology and obesity, however, human studies for TG family members in adipocytes have not been conducted. In this study, we investigated the association of TGM family members to acquired weight gain in a rare set of monozygotic (MZ) twins discordant for body weight, i.e., heavy–lean twin pairs. We report that F13A1 is the only TGM family member showing significantly altered, higher expression in adipose tissue of the heavier twin. Our previous work linked adipocyte F13A1 to increased weight, body fat mass, adipocyte size, and pro-inflammatory pathways. Here, we explored further the link of F13A1 to adipocyte size in the MZ twins via a previously conducted TWA study that was further mined for genes that specifically associate to hypertrophic adipocytes. We report that differential expression of F13A1 (ΔHeavy–Lean) associated with 47 genes which were linked via gene enrichment analysis to immune response, leucocyte and neutrophil activation, as well as cytokine response and signaling. Our work brings further support to the role of F13A1 in the human adipose tissue pathology, suggesting a role in the cascade that links hypertrophic adipocytes with inflammation

Plasma Membrane Factor XIIIA Transglutaminase Activity Regulates Osteoblast Matrix Secretion and Deposition by Affecting Microtubule Dynamics

Transglutaminase activity, arising potentially from transglutaminase 2 (TG2) and Factor XIIIA (FXIIIA), has been linked to osteoblast differentiation where it is required for type I collagen and fibronectin matrix deposition. In this study we have used an irreversible TG-inhibitor to ‘block –and-track’ enzyme(s) targeted during osteoblast differentiation. We show that the irreversible TG-inhibitor is highly potent in inhibiting osteoblast differentiation and mineralization and reduces secretion of both fibronectin and type I collagen and their release from the cell surface. Tracking of the dansyl probe by Western blotting and immunofluorescence microscopy demonstrated that the inhibitor targets plasma membrane-associated FXIIIA. TG2 appears not to contribute to crosslinking activity on the osteoblast surface. Inhibition of FXIIIA with NC9 resulted in defective secretory vesicle delivery to the plasma membrane which was attributable to a disorganized microtubule network and decreased microtubule association with the plasma membrane. NC9 inhibition of FXIIIA resulted in destabilization of microtubules as assessed by cellular Glu-tubulin levels. Furthermore, NC9 blocked modification of Glu-tubulin into 150 kDa high-molecular weight Glu-tubulin form which was specifically localized to the plasma membrane. FXIIIA enzyme and its crosslinking activity were colocalized with plasma membrane-associated tubulin, and thus, it appears that FXIIIA crosslinking activity is directed towards stabilizing the interaction of microtubules with the plasma membrane. Our work provides the first mechanistic cues as to how transglutaminase activity could affect protein secretion and matrix deposition in osteoblasts and suggests a novel function for plasma membrane FXIIIA in microtubule dynamics

Transglutaminases in Monocytes and Macrophages

Macrophages are key players in various inflammatory disorders and pathological conditions via phagocytosis and orchestrating immune responses. They are highly heterogeneous in terms of their phenotypes and functions by adaptation to different organs and tissue environments. Upon damage or infection, monocytes are rapidly recruited to tissues and differentiate into macrophages. Transglutaminases (TGs) are a family of structurally and functionally related enzymes with Ca2+-dependent transamidation and deamidation activity. Numerous studies have shown that TGs, particularly TG2 and Factor XIII-A, are extensively involved in monocyte- and macrophage-mediated physiological and pathological processes. In the present review, we outline the current knowledge of the role of TGs in the adhesion and extravasation of monocytes, the expression of TGs during macrophage differentiation, and the regulation of TG2 expression by various pro- and anti-inflammatory mediators in macrophages. Furthermore, we summarize the role of TGs in macrophage phagocytosis and the understanding of the mechanisms involved. Finally, we review the roles of TGs in tissue-specific macrophages, including monocytes/macrophages in vasculature, alveolar and interstitial macrophages in lung, microglia and infiltrated monocytes/macrophages in central nervous system, and osteoclasts in bone. Based on the studies in this review, we conclude that monocyte- and macrophage-derived TGs are involved in inflammatory processes in these organs. However, more in vivo studies and clinical studies during different stages of these processes are required to determine the accurate roles of TGs, their substrates, and the mechanisms-of-action

Transglutaminase-mediated oligomerization promotes osteoblast adhesive properties of osteopontin and bone sialoprotein

Tissue transglutaminase (TG2) is a widely distributed, protein-crosslinking enzyme having a prominent role in cell adhesion as a β1 integrin co-receptor for fibronectin. In bone and teeth, its substrates include the matricellular proteins osteopontin (OPN) and bone sialoprotein (BSP). The aim of this study was to examine effects of TG2-mediated crosslinking and oligomerization of OPN and BSP on osteoblast cell adhesion. We show that surfaces coated with oligomerized OPN and BSP promote MC3T3-E1/C4 osteoblastic cell adhesion significantly better than surfaces coated with the monomeric form of the proteins. Both OPN and BSP oligomer-adherent cells showed more cytoplasmic extensions than those cells grown on the monomer-coated surfaces indicative of increased cell connectivity. Our study suggests a role for TG2 in promoting the cell adhesion function of two matricellular substrate proteins prominent in bone, tooth cementum and certain tumors

Peptidic Inhibitors and a Fluorescent Probe for the Selective Inhibition and Labelling of Factor XIIIa Transglutaminase

Factor XIIIa (FXIIIa) is a transglutaminase of major therapeutic interest for the development of anticoagulants due to its essential role in the blood coagulation cascade. While numerous FXIIIa inhibitors have been reported, they failed to reach clinical evaluation due to their lack of metabolic stability and low selectivity over transglutaminase 2 (TG2). Furthermore, the chemical tools available for the study of FXIIIa activity and localization are extremely limited. To combat these shortcomings, we designed, synthesised, and evaluated a library of 21 novel FXIIIa inhibitors. Electrophilic warheads, linker lengths, and hydrophobic units were varied on small molecule and peptidic scaffolds to optimize isozyme selectivity and potency. A previously reported FXIIIa inhibitor was then adapted for the design of a probe bearing a rhodamine B moiety, producing the innovative KM93 as the first known fluorescent probe designed to selectively label active FXIIIa with high efficiency (kinact/KI = 127,300 M−1 min−1) and 6.5-fold selectivity over TG2. The probe KM93 facilitated fluorescent microscopy studies within bone marrow macrophages, labelling FXIIIa with high efficiency and selectivity in cell culture. The structure–activity trends with these novel inhibitors and probes will help in the future study of the activity, inhibition, and localization of FXIIIa

Peptidic Inhibitors and a Fluorescent Probe for the Selective Inhibition and Labelling of Factor XIIIa Transglutaminase

Factor XIIIa (FXIIIa) is a transglutaminase of major therapeutic interest for the development of anticoagulants due to its essential role in the blood coagulation cascade. While numerous FXIIIa inhibitors have been reported, they failed to reach clinical evaluation due to their lack of metabolic stability and low selectivity over transglutaminase 2 (TG2). Furthermore, the chemical tools available for the study of FXIIIa activity and localization are extremely limited. To combat these shortcomings, we designed, synthesised, and evaluated a library of 21 novel FXIIIa inhibitors. Electrophilic warheads, linker lengths, and hydrophobic units were varied on small molecule and peptidic scaffolds to optimize isozyme selectivity and potency. A previously reported FXIIIa inhibitor was then adapted for the design of a probe bearing a rhodamine B moiety, producing the innovative KM93 as the first known fluorescent probe designed to selectively label active FXIIIa with high efficiency (kinact/KI = 127,300 M−1 min−1) and 6.5-fold selectivity over TG2. The probe KM93 facilitated fluorescent microscopy studies within bone marrow macrophages, labelling FXIIIa with high efficiency and selectivity in cell culture. The structure–activity trends with these novel inhibitors and probes will help in the future study of the activity, inhibition, and localization of FXIIIa

Circulating undercarboxylated osteocalcin and gingival crevicular fluid tumour necrosis factor-α in children.

International audienceOsteocalcin, a protein secreted by osteoblasts during bone formation, is negatively associated with adult periodontal disease. Little is known about this association in children. To examine the extent to which plasma undercarboxylated osteocalcin (ucOC) is associated with gingival crevicular fluid tumour necrosis factor-alpha (GCF TNF-α) - a potential marker of gingival inflammation - in children. We used data from the Quebec Adipose and Lifestyle InvesTigation in Youth cohort, an ongoing longitudinal study on the natural history of obesity among Caucasian children with a family history of obesity in Quebec, Canada. This cross-sectional analysis from the baseline visit includes 120 children aged 8-10 years. Plasma ucOC and GCF TNF-α levels were determined by enzyme-linked immunosorbent assay. Linear regression analyses, adjusting for age, gender, family income, sexual maturity stage, daily physical activity, obesity, and fasting glucose were conducted, with TNF-α level as the dependent variable. A 1-ng/ml increase in ucOC was associated with a 0.96% decrease (95% confidence interval (CI): -1.69, -0.23) in GCF TNF-α level. A negative association between a marker of bone formation and a marker of gingival inflammation was observed as early as childhood among Caucasian children with a family history of obesity

Collagens regulating adipose tissue formation and functions

Abstract The globally increasing prevalence of obesity is associated with the development of metabolic diseases such as type 2 diabetes, dyslipidemia, and fatty liver. Excess adipose tissue (AT) often leads to its malfunction and to a systemic metabolic dysfunction because, in addition to storing lipids, AT is an active endocrine system. Adipocytes are embedded in a unique extracellular matrix (ECM), which provides structural support to the cells as well as participating in the regulation of their functions, such as proliferation and differentiation. Adipocytes have a thin pericellular layer of a specialized ECM, referred to as the basement membrane (BM), which is an important functional unit that lies between cells and tissue stroma. Collagens form a major group of proteins in the ECM, and some of them, especially the BM-associated collagens, support AT functions and participate in the regulation of adipocyte differentiation. In pathological conditions such as obesity, AT often proceeds to fibrosis, characterized by the accumulation of large collagen bundles, which disturbs the natural functions of the AT. In this review, we summarize the current knowledge on the vertebrate collagens that are important for AT development and function and include basic information on some other important ECM components, principally fibronectin, of the AT. We also briefly discuss the function of AT collagens in certain metabolic diseases in which they have been shown to play central roles