RETRACTED: Deletion of the scavenger receptor Scarb1 in myeloid cells does not affect bone mass

Abstract

The scavenger receptor class B member 1 (SR-B1 or Scarb1) is a glycosylated cell surface receptor for high density lipoproteins (HDL), oxidized low density lipoproteins (OxLDL), and phosphocholine-containing oxidized phospholipids (PC-OxPLs). Scarb1 is expressed in macrophages and has been shown to have both pro- and anti-atherogenic properties. It has been reported that global deletion of Scarb1 in mice leads to either high or low bone mass and that PC-OxPLs decrease osteoblastogenesis and increase osteoclastogenesis. PC-OxPLs decrease bone mass in 6-month-old mice and are critical pathogenetic factors in the bone loss caused by high fat diet or aging. We have investigated here whether Scarb1 expression in myeloid cells affects bone mass and whether PC-OxPLs exert their anti-osteogenic effects via activation of Scarb1 in macrophages. To this end, we generated mice with deletion of Scarb1 in LysM-Cre expressing cells and found that lack of Scarb1 did not affect bone mass in vivo. These results indicate that Scarb1 expression in cells of the myeloid/osteoclast lineage does not contribute to bone homeostasis. Based on this evidence, and earlier studies of ours showing that Scarb1 expression in osteoblasts does not affect bone mass, we conclude that Scarb1 is not an important mediator of the adverse effects on PC-OxPLs in osteoclasts or osteoblasts in 6-month-old mice.

Keywords: Bone mass; Macrophages; Osteoclasts; Oxidized phospholipids; Scavenger receptor class B member 1 (SR-B1 or Scarb1).

Fig. 1.

Scarb1 gene was effectively deleted in LysM-Cre targeted cells. (A) Quantitative PCR (qPCR) of loxP-flanked genomic DNA isolated from macrophages, osteoclasts, and right kidney obtained from 6-month-old female mice. Macrophages and osteoclasts were cultured from bone marrow cells as described in the methods section [for macrophages and osteoclasts Scarb1^fl/fl (n = 2), LysM-Cre;Scarb1^fl/fl (n = 6), six replicates of pooled cultures; for kidney Scarb1^fl/fl (n = 10), LysM-Cre;Scarb1^fl/fl (n = 18)]. (B) Scarb1 expression quantified by RT-PCR in the same cells as in (A). (C) qPCR of loxP-flanked genomic DNA isolated from macrophages and osteoclasts obtained from 6-month-old male mice cultured as in (A) and genomic DNA isolated from right kidney from 6-month-old male mice; [for macrophages and osteoclasts Scarb1^fl/fl (n = 2), LysM-Cre;Scarb1^fl/fl (n = 2), six replicates of pooled cultures; for kidney Scarb1^fl/fl (n = 10), LysM-Cre;Scarb1^fl/fl (n = 15)]. (D) Scarb1 expression was quantified by RT-PCR of cells cultured as in (C). Transcripts were normalized to the reference gene MRPS2. Bar and whiskers are the mean ± standard deviation (s.d.) and p values were calculated using 2-tailed unpaired t-tests.

Fig. 2.

Deletion of Scarb1 in LysM-Cre-targeted cells does not affect body weight, fat mass, lean mass, or BMD in either sex. (A) Body weight, fat mass, lean mass, and BMD measurements at the indicated sites at 2, 4 and 6 months of age in female mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 18]. (B) Weight, fat mass, lean mass, and BMD measurements at 2, 4 and 6 months of age in male mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 15]. Data are shown as mean and standard deviation. Adjusted p values, calculated by repeated measures using two-way ANOVA are shown. BMD: Bone mineral density.

Fig. 3.

Deletion of Scarb1 using LysM-Cre does not affect trabecular bone in either sex. Micro-CT determination of trabecular bone architecture in (A) vertebral and femoral metaphyseal bone of 6-month-old female mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 18] and in (B) vertebral and femoral metaphyseal bone of 6-month-old male mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 15]. Bar and whiskers are the mean ± s.d. and p values were calculated using 2-tailed unpaired t-tests.

Fig. 4.

Deletion of Scarb1 using LysM-Cre does not affect cortical bone in female or male mice. (A, C) Micro-CT determination of cortical bone in 6-month-old female [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 18] and male mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 15]. (B, D) Femoral length in female [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 15] and male mice [Scarb1^fl/fl n = 9; LysM-Cre;Scarb1^fl/fl n = 13]. Bar and whiskers are the mean ± s.d. and p values were calculated with 2-tailed unpaired t-tests.

Fig. 5.

Deletion of Scarb1 using LysM-Cre does not affect osteoclast number in vivo. (A) Measurement of bone remodeling markers in the serum of 6-month-old female and male mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 10], P1NP, N-terminal propeptide; TRAP, Tartrate-resistant acid phosphatase. (B, C) The expression of osteoclasts marker genes TRAP (Acp5), CTSK, and Igtb3 was quantified by RT-PCR in vertebral bone (L6) of 6-month-old female and male mice [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 10]. Transcripts were normalized to the reference gene transcript MRPS2. (D) Histomorphometric measurements of vertebral trabecular bone surface (L3); [Scarb1^fl/fl n = 10; LysM-Cre;Scarb1^fl/fl n = 10]. Bar and whiskers are the mean ± s.d. and p values were calculated using 2-tailed unpaired t-tests. CTSK: Cathepsin K; Igtb3: Integrin β3; N.Oc, osteoclast number; B.Pm, bone perimeter; Oc.S, osteoclast surface; BS, bone surface.