Lyplal1 is dispensable for normal fat deposition in mice

Abstract

Genome-wide association studies (GWAS) have detected association between variants in or near the Lysophospholipase-like 1 (LYPLAL1) locus and metabolic traits, including central obesity, fatty liver and waist-to-hip ratio. LYPLAL1 is also known to be upregulated in the adipose tissue of obese patients. However, the physiological role of LYPLAL1 is not understood. To investigate the function of Lyplal1 in vivo we investigated the phenotype of the Lyplal1^{tm1a(KOMP)Wtsi} homozygous mouse. Body composition was unaltered in Lyplal1 knockout mice as assessed by dual-energy X-ray absorptiometry (DEXA) scanning, both on normal chow and on a high-fat diet. Adipose tissue distribution between visceral and subcutaneous fat depots was unaltered, with no change in adipocyte cell size. The response to both insulin and glucose dosing was normal in Lyplal1^{tm1a(KOMP)Wtsi} homozygous mice, with normal fasting blood glucose concentrations. RNAseq analysis of liver, muscle and adipose tissue confirmed that Lyplal1 expression was ablated with minimal additional changes in gene expression. These results suggest that Lyplal1 is dispensable for normal mouse metabolic physiology and that despite having been maintained through evolution Lyplal1 is not an essential gene, suggesting possible functional redundancy. Further studies will be required to clarify its physiological role.

Keywords: Adipose tissue; Genome-wide association study; Lyplal1; Model organism; Mouse; Obesity.

Fig. 1.

Mice were generated with the Lyplal1^tm1a allele, resulting in knockout at both the protein and the RNA level. (A) Diagram showing the Lyplal1^tm1a allele design (figure obtained from IMPC, www.mousephenotype.org/data/genes/MGI:2385115). RNA and protein were extracted from organs from 28-week-old mice. (B-I) qPCR analysis of Lyplal1 mRNA levels in gastrocnemius (B), heart (C), liver (D), kidney (E), spleen (F), BAT (G), scWAT (H) and vWAT (I). Data are presented as means±s.d. Black triangles, male Lyplal1^+/+; white triangles, male Lyplal1^tm1a/tm1a; black circles, female Lyplal1^+/+; white circles, female Lyplal1^tm1a/tm1a. (J) Protein levels of Lyplal1 and GAPDH were determined by Western blot in liver, kidney, scWAT and vWAT lysates. Representative blots are shown. [(B) n=5 female Lyplal1^tm1a/tm1a, n=7 other groups; (C,E-G) n=3 each group; (D) n=7 Lyplal1^+/+, n=8 Lyplal1^tm1a/tm1a; (H) n=8 male Lyplal1^+/+, n=6 male Lyplal1^tm1a/tm1a, n=5 female Lyplal1^+/+, n=5 female Lyplal1^tm1a/tm1a; (I) n=5 male Lyplal1^+/+, n=3 male Lyplal1^tm1a/tm1a, n=5 female Lyplal1^+/+, n=6 female Lyplal1^tm1a/tm1a; (J) n=3 samples from each sex and genotype per tissue]. KO, knockout; scWAT, subcutaneous white adipose tissue; vWAT, visceral white adipose tissue; WT, wild type.

Fig. 2.

Lyplal1 knockout does not alter body composition in Lyplal1 knockout mice fed high-fat diet from 6 weeks of age. (A) Body weights of mice up to 28 weeks of age were not altered by genotype. (B-D) Lean mass (B), fat mass (C) and fat percentage (D) were unaltered in 24-week-old mice, measured by DEXA. (E-G) scWAT (E), vWAT (F) and BAT (G) mass were unchanged in 28-week-old mice, weighed after dissection. (H) No qualitative change in scWAT morphology in knockout mice. Representative images are shown of scWAT sections from 28-week-old mice stained with H&E. (I) No change in adipocyte CSA was determined using ImageJ analysis of scWAT sections stained with H&E, analysed using PhenStat. (J) No change in fatty liver in knockout mice. Representative images are shown of liver sections from 28-week-old mice stained with Oil Red O and Haematoxylin. Black triangles, male Lyplal1^+/+; white triangles, male Lyplal1^tm1a/tm1a; black circles, female Lyplal1^+/+; white circles, female Lyplal1^tm1a/tm1a. Data are presented as means±s.d. Mixed model analysis was performed using PhenStat. [(B-D) n=10 male Lyplal1^+/+, n=11 male Lyplal1^tm1a/tm1a, n=9 female Lyplal1^+/+, n=8 female Lyplal1^tm1a/tm1a; (E,F) n=9 male Lyplal1^+/+, n=10 male Lyplal1^tm1a/tm1a, n=9 female Lyplal1^+/+, n=7 female Lyplal1^tm1a/tm1a; (G) n=10 male Lyplal1^+/+, n=10 male Lyplal1^tm1a/tm1a, n=9 female Lyplal1^+/+, n=8 female Lyplal1^tm1a/tm1a; (H,I) n=9 male Lyplal1^+/+, n=8 male Lyplal1^tm1a/tm1a, n=7 female Lyplal1^+/+, n=8 female Lyplal1^tm1a/tm1a]. BAT, brown adipose tissue; CSA, cross-sectional area; H&E, Haemotoxylin and Eosin; KO, knockout; scWAT, subcutaneous white adipose tissue; vWAT, visceral white adipose tissue; WT, wild type.

Fig. 3.

Lyplal1 knockout does not alter glucose metabolism in mice. (A-D) Intraperitoneal insulin tolerance test (ipITT) at 18 weeks of age (A) and intraperitoneal glucose tolerance test (ipGTT) at 22 weeks of age (B) demonstrated no alteration in glucose clearance in knockout mice. (C,D) The AUC was calculated for both ipITT (C) and ipGTT (D). AUC mixed model analysis was performed using Phenstat. (E) Plasma insulin concentrations after a 4 h fast were unaltered in 28-week-old knockout mice, determined by ELISA, mixed model analysis performed using PhenStat. (F-H) Indirect calorimetry of mice at 26 weeks of age. No differences were observed in average activity count (F), RER (G) or energy expenditure (H). Black triangles, male Lyplal1^+/+; white triangles, male Lyplal1^tm1a/tm1a; black circles, female Lyplal1^+/+; white circles, female Lyplal1^tm1a/tm1a; dashed line, male Lyplal1^+/+; dotted line, male Lyplal1^tm1a/tm1a; dash-dot line, female Lyplal1^+/+; dash-dot-dot line, female Lyplal1^tm1a/tm1a. Data are presented as means±s.d., with linear regression lines as appropriate. [(A,C) n=6 male Lyplal1^+/+, n=6 male Lyplal1^tm1a/tm1a, n=6 female Lyplal1^+/+, n=5 female Lyplal1^tm1a/tm1a; (B,D) n=9 male Lyplal1^+/+, n=9 male Lyplal1^tm1a/tm1a, n=10 female Lyplal1^+/+, n=8 female Lyplal1^tm1a/tm1a; (E) n=10 male Lyplal1^+/+, n=11 male Lyplal1^tm1a/tm1a, n=9 female Lyplal1^+/+, n=8 female Lyplal1^tm1a/tm1a; (F,G) n=9 male Lyplal1^+/+, n=8 male Lyplal1^tm1a/tm1a, n=6 female Lyplal1^+/+, n=7 female Lyplal1^tm1a/tm1a; (H) n=9 male Lyplal1^+/+, n=8 male Lyplal1^tm1a/tm1a, n=6 female Lyplal1^+/+, n=6 female Lyplal1^tm1a/tm1a]. AUC, area under the curve; KO, knockout; RER, respiratory exchange ratio; WT, wild type.