Insulin-Like Effects of Visfatin on Human Osteoblasts

Abstract

Visfatin (also known as pre-B cell colony-enhancing factor or PBEF) is a novel adipocytokine that is highly expressed in visceral fat and upregulated in obesity and type 2 diabetes mellitus. Visfatin binds to and activates the insulin receptor (IR), thereby exerting insulin-mimetic effects in various cell lines. IR has been detected in osteoblasts, which is consistent with the role of insulin as an important osteotropic hormone. This study investigated the actions of visfatin on human primary osteoblasts. The expression and tyrosine phosphorylation of IR, IR substrate-1 (IRS-1), and IRS-2 were determined by immunoprecipitation and immunoblotting. Cell proliferation was determined by measuring [(3)H]thymidine incorporation and cell number. Glucose uptake was determined by measuring 2-[(3)H]deoxyglucose incorporation. Real-time quantitative reverse-transcription polymerase chain reaction (PCR) was used for determining alkaline phosphatase (ALP), osteocalcin, and type I collagen mRNA expression. Enzyme-linked immunosorbent assay and radioimmunoassay were used for measuring ALP activity, osteocalcin secretion, and type I collagen production. We found that visfatin induced tyrosine phosphorylation of IR, IRS-1, and IRS-2. Moreover, the effects of visfatin - glucose uptake, proliferation, and type I collagen enhancement of cultured human osteoblast-like cells - bore a close resemblance to those of insulin and were inhibited by hydroxy-2-naphthalenylmethylphosphonic acid tris-acetoxymethyl ester, a specific inhibitor of IR tyrosine kinase activity. We also unexpectedly found that visfatin downregulated osteocalcin secretion from human osteoblast-like cells. These data indicate that the regulation of glucose uptake, proliferation, and type I collagen production by visfatin in human osteoblasts involves IR phosphorylation, the same signal-transduction pathway used by insulin.

The role of adipocytokines in the pathogenesis of knee joint osteoarthritis

“…Higher adipocytokine levels were found among women with larger osteophytes (>10 mm) than in those with no or smaller osteophytes (<10 mm) [70]. Visfatin has been also found to influence osteoblasts proliferation and type II collagen production [71].…”

Suppression of leukocyte infiltration and cartilage degradation by selective inhibition of pre-B cell colony-enhancing factor/visfatin/nicotinamide phosphoribosyltransferase: Apo866-mediated therapy in human fibroblasts and murine collagen-induced arthrit

“…Several groups have shown that PBEF interacts with the insulin receptor (8-10); however, this notion remains controversial (11,12), and it may be that PBEF signals through an alternative signaling route. It is also unlikely that this signaling effect is attributable to the endotoxin that may be present in PBEF preparations, since coincubation with polymixin B shows no abrogation of PBEF signaling (6,10,13).In vitro activation of signaling pathways by PBEF has been shown to regulate the expression of metalloproteinases, chemokines, and cytokines (6,7,(9)(10)(11)(12), but the in vivo contribution of this mediator of arthritis progression remains unclear. The recently developed small molecule inhibitor APO866 has been shown to act as a competitive inhibitor of PBEF/NAMPT activity by inhibiting binding of its natural substrate, nicotinamide (14,15).…”

“…Several groups have shown that PBEF interacts with the insulin receptor (8)(9)(10); however, this notion remains controversial (11,12), and it may be that PBEF signals through an alternative signaling route. It is also unlikely that this signaling effect is attributable to the endotoxin that may be present in PBEF preparations, since coincubation with polymixin B shows no abrogation of PBEF signaling (6,10,13).…”

“…In vitro activation of signaling pathways by PBEF has been shown to regulate the expression of metalloproteinases, chemokines, and cytokines (6,7,(9)(10)(11)(12), but the in vivo contribution of this mediator of arthritis progression remains unclear. The recently developed small molecule inhibitor APO866 has been shown to act as a competitive inhibitor of PBEF/NAMPT activity by inhibiting binding of its natural substrate, nicotinamide (14,15).…”

L-carnitine and taurine synergistically inhibit the proliferation and osteoblastic differentiation of vascular smooth muscle cells

“…DNA and protein synthesis assay DNA and protein synthesis were assessed using scintillation counting to measure the incorporation of [ 3 H]thymidine (7.4×10 4 Bq/mL) or [ 3 H]leucine into DNA or protein, respectively, as previously described [29][30][31] . Briefly, cells were plated at a density of 2×10 4 /well in 24-well plates, cultured in growth medium and treated with vehicle controls, 5−20 mmol/L TAU, 1−10 mmol/L LC or 20 mmol/L TAU+10 mmol/L LC for 48 h. Then, the plates were washed with PBS, and 10% trichloroacetic acid solution was added to the wells to precipitate the DNA and protein.…”