Oxidised LDL modulate adipogenesis in 3T3‐L1 preadipocytes by affecting the balance between cell proliferation and differentiation

Abstract

The effects of oxidised LDL (oxLDL) on cell proliferation, apoptosis and hormone-induced differentiation have been evaluated for the first time in 3T3-L1 preadipocytes. Unlike control cells, oxLDL-treated preadipocytes showed a high proliferation rate, a low apoptosis level, and an impaired differentiation process with an increased preadipocyte factor-1 (Pref-1) mRNA expression at late times. By silencing Pref-1 mRNA or inhibiting its expression with an increased dexamethasone concentration, differentiation occurred as usual, which demonstrates the key role of Pref-1 overexpression. The results suggest a specific action of oxLDL on the adipogenesis inhibitor Pref-1, as indicated also by its reappearance in mature adipocytes treated with oxLDL. The inhibitory effects of oxLDL on differentiation required oxLDL uptake by CD36, and were associated with lipoprotein lipids. These results point to oxLDL as a modulator of adipose tissue mass and as possible link between obesity and its clinical complications.

Oxidised LDL up‐regulate CD36 expression by the Nrf2 pathway in 3T3‐L1 preadipocytes

“…Total RNA was isolated from 3T3-L1 cells with the TRIZOL reagent (Invitrogen-Life technologies, USA) as described elsewhere [6].…”

“…At confluence (day 0) the cells were induced to differentiate by differentiation mixture (DMIX) as described elsewhere [6]. In a set of experiments the cells were treated with dexamethasone to evaluate its role on Nrf2 expression.…”

“…LDL (1.019-1.063 g/ml) were isolated from pooled fresh plasma of healthy volunteers as described elsewhere [6]. Protein content was measured by LowryÕs method [20].…”

“…Cells were centrifuged at 18 000 · g for 10 min at 4°C. Supernatants, assessed for protein concentration [20], were used for Western Blot analysis as previously described [6]. Immunoblotting was carried out using antibodies that recognise Nrf2, PPARc, CD36, PKC (Santa Cruz, Biotechnology Inc., CA, USA), phospho-PKC, PKCd, or phospho-PKCd (Cell Signalling Technology Inc., MA, USA).…”

“…Several lines of evidence have demonstrated that oxLDL play a crucial role in the pathogenesis of atherosclerosis [3] and modulate growth, transcription factor and cytokine expression, influencing cell activities [4,5]. We have recently demonstrated that oxLDL are taken up through a specific, saturable process mediated by CD36, affecting the adipose tissue homeostasis by inhibiting the differentiation of preadipocytes [6]. CD36, an ubiquitous membrane glycoprotein, recognizes a variety of ligands in addition to modified lipoproteins [7,8] and serves as a fatty acid (FA) transporter in muscle and adipocytes [9], representing a tract of the mature adipocyte phenotype [10].…”