Osbpl8 Deficiency in Mouse Causes an Elevation of High-Density Lipoproteins and Gender-Specific Alterations of Lipid Metabolism

Peer reviewe

Gene trap (GT) insertion site in Osbpl8.

<p>Insertion of the pGT vector in the 5^th intron of the Osbpl8 gene in chromosome 10 generates a β-geo fusion mRNA transcript through the use of the En-2 splice acceptor (SA) contained in the vector. The insertion site (nt position indicated) was identified by PCR, using primers targeting gDNA in 5^th intron of Osbpl8 gene (Fwd), and in the 1^st intron of En2 (Rev1).</p

Non-cholesterol sterols in the plasma of WT and Osbpl8KO mice (n = 6).

#<p>T-test, comparison between genotypes; *p<0.05, **p<0.01.</p

Liver tissue lipid content of the WT and Osbpl8KO mice.

*<p>T-test, comparison between genotypes, n = 6.</p

Western blot analysis of HDL apolipoproteins in WT and KO mouse plasma and high-density lipoprotein fractions.

<p>A. Chow diet-fed animals; ApoA-II, ApoC-III, and ApoE (identified on the right) in the fasting plasma (Pla) of WT and KO mice of both genders (identified at the top); Bottom panel, ApoE in the HDL peak fractions of plasma fractioned on a Suprose HR 6 10/30 column. B. Western diet-fed animals; ApoE in the plasma (Pla) and HDL fractions (HDL) of WT and KO mice of both genders (identified at the top). The plasma loading was 3 µl/lane of 1/40 dilution, and that of Superose 6 HR 10/30 HDL fractions 150 µl/lane concentrated by acetone precipitation. The analysis was carried out for pooled plasma and HDL fractions isolated from the same pools; Chow diet, 3 animals/pool; Western diet, 6 animals/pool.</p

HDL catabolism in Osbpl8KO mice.

<p>Chow-fed WT and Osbpl8KO (n = 4–6) female (A) or male (B) animals were injected with Alexa568-labeled mouse HDL (40 µg protein/animal), and the fluorescence signal (y-axis; mean ± s.e.m., % of starting value at 5 min post-injection) in their plasma as a function of time (x-axis) was measured as specified in Materials and <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0058856#s4" target="_blank">Methods</a>.</p

Western blot analysis of ORP8 in KO mouse tissues and peritoneal macrophage.

<p>Total protein specimens (20 µg protein/lane) of tissues with abundant ORP8 expression: spleen, brain, kidney, and liver (WT and Osbpl8KO animals), and of peritoneal macrophage (WT, Osbpl8+/−, and Osbpl8−/−) were analyzed by Western blotting with ORP8 and β-actin antibodies. ORP8 runs for an unknown reason as a doublet of bands with the apparent molecular masses of 97 and 101 kDa, as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0058856#pone.0058856-Yan1" target="_blank">[10]</a>.</p

Fasting plasma lipid concentrations of WT and Osbpl8KO mice before (n = 12) and after (n = 6) the diets.

#<p>TC, total cholesterol; FC, free cholesterol; TG, triglycerides; PL, choline-containing phospholipids; mmol/L.</p>##<p>T-test, comparison between genotypes; *p<0.05, **p<0.01, ***p<0.001.</p>§<p>apolipoprotein A-I; mg/ml.</p