Interaction Between Photoperiod and Sex on Hepatic Lipid Homeostasis in Rats Fed an Obesogenic Diet

Abstract

Seasonal changes in day length as well as sex are critical factors influencing metabolic processes. However, their combined impact on liver lipid metabolism under obesogenic conditions remains unclear. This study aims to investigate the interaction between sex and photoperiod on liver lipid homeostasis in obese Fischer 344 (F344) rats, with a special emphasis on the underlying molecular mechanisms. To this aim, male and female rats were fed with a cafeteria diet (CAF) for 11 weeks and exposed to either long (L18, 18 h of light/day) or short (L6, 6 h of light/day) photoperiods for the last 8 weeks. Liver histological analysis, hepatic damage markers, plasmatic and hepatic lipid profiles as well as hepatic gene expression related to lipid metabolism were performed. Results indicate that males exposed to the L18 photoperiod developed hepatic macro-steatosis and exacerbated hepatic lipid accumulation, leading to increased liver weight and total hepatic lipids compared to their L6 photoperiod counterparts. At the molecular level, males under the L18 photoperiod, showed downregulation of the lipid transporter Fabp4 and upregulation of β-oxidation related genes. On the other hand, female rats exhibited a healthier lipid profile compared to males when exposed to both L6 and L18 photoperiods, and a lower degree of steatosis. Gene analysis revealed that the lipid transporter Mtp expression was significantly higher in females under the L18 photoperiod. In conclusion, photoperiod and sex influence hepatic lipid metabolism in rats fed an obesogenic diet, highlighting the importance of considering these factors in understanding and addressing metabolic disorders and reinforcing the concept of precision medicine.

Keywords: fatty liver disease; hepatic lipid metabolism; hepatic lipid transport; metabolic syndrome; obesogenic diet; photoperiod.

FIGURE 1

Experimental design. Twenty male and 20 female F344 rats were fed for 11 weeks with CAF diet and exposed to L18 and L6 photoperiods for the last 8 weeks of the experiment. CAF, cafeteria; F344, Fischer 344; L6, short photoperiod of 6 h of light per day; L12, standard photoperiod of 12 h of light per day; L18, long photoperiod of 18 h of light per day.

FIGURE 2

Representative histological sections after eosin and hematoxylin stain of liver from male and female F344 rats exposed for 8 weeks to L18 or L6 photoperiods and fed with a CAF diet for 11 weeks. Pictures were taken under the magnification of 20×. CAF, cafeteria; F344, Fischer 344; L6, short photoperiod of 6 h of light per day; L18, long photoperiod of 18 h of light per day.

FIGURE 3

Effect of sex and photoperiod on hepatic damage markers in plasma. ALT (A), AST (B), AST/ALT ratio (C) and bilirubin (D) in male and female F344 rats fed with CAF diet for 11 weeks and exposed to L18 or L6 photoperiods for the last 8 weeks. Data are expressed as minimum to maximum values, median and interquartile range (n = 10). P, photoperiod effect; S, sex effect; S × P, interaction between sex and photoperiod assessed using two‐way ANOVA; *Indicates significant differences (*p < 0.05, **p < 0.01) between groups using two‐way ANOVA followed by Tukey post hoc test. ALT, alanine aminotransferase; AST, aspartate aminotransferase; CAF, cafeteria; F, females; F344, Fischer 344; L6, short photoperiod of 6 h of light per day; L18, long photoperiod of 18 h of light per day; M, males.

FIGURE 4

Effect of sex and photoperiod on hepatic lipid parameters. Liver weight (A), total hepatic lipid content (B), hepatic total cholesterol (C), hepatic triglycerides (D), hepatic phospholipids (E) in male and female F344 rats fed with CAF diet for 11 weeks and exposed to L18 or L6 photoperiods for the last 8 weeks. P, photoperiod effect; S, sex effect; S × P, interaction between sex and photoperiod assessed using two‐way ANOVA. *Indicates significant differences (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001) and “a” indicates a trend (p < 0.1) between groups using two‐way ANOVA followed by Tukey post hoc test. CAF, cafeteria; F, females; F344, Fischer 344; H‐PL, hepatic phospholipids; H‐TAG, hepatic triglycerides; H‐TC, hepatic total cholesterol; L6, short photoperiod of 6 h of light per day; L18, long photoperiod of 18 h of light per day; M, males.

FIGURE 5

Effect of sex and photoperiods on the relative gene expression of lipogenesis‐related genes in the liver. Fasn (A), Acaca (B), Srebp‐1c (C), and Dgat (D) relative hepatic mRNA levels in male and female F344 rats fed with CAF diet for 11 weeks and exposed to L18 or L6 photoperiods for the last 8 weeks. Data are expressed as minimum to maximum values, median and interquartile range (n = 10). P, photoperiod effect; S, sex effect assessed using two‐way ANOVA for Srebp‐1C, and two‐way non‐parametric ANOVA with ART transformation for Fasn, Acaca and Dgat. *Indicates significant differences (*p < 0.05 and ** p < 0.01) between groups using Tukey as post hoc test for Srebp‐1C, and post hoc analysis with Tukey adjustment using emmeans package in R for Fasn, Acaca and Dgat. CAF, cafeteria; F344, Fischer 344; M, males; F, females; L18, long photoperiod of 18 h of light per day; L6, short photoperiod of 6 h of light per day; Fasn, Fatty acid synthase; Acaca, acetyl‐CoA carboxylase alpha; Srebp‐1c, sterol regulatory element‐binding protein 1C; Dgat, diacylglycerol O‐acyltransferase 1.

FIGURE 6

Effect of sex and photoperiods on the relative gene expression of β‐oxidation related genes in the liver. Cpt‐1α (A) and Ppar‐α (B) relative hepatic mRNA levels in male and female F344 rats fed with a CAF diet for 11 weeks and exposed to L18 or L6 photoperiods for the last 8 weeks. Data are expressed as minimum to maximum values, median and interquartile range (n = 10). P, photoperiod effect; S, sex effect; S × P, interaction between sex and photoperiod assessed using two‐way ANOVA. *Indicates significant differences (*p < 0.05, **p < 0.01) and “a” indicates a trend (p < 0.1) between groups using two‐way ANOVA followed by Tukey post hoc test. CAF, cafeteria; Cpt‐1α, carnitine palmitoyltransferase I alpha; F, females; F344, Fischer 344; L6, short photoperiod of 6 h of light per day; L18, long photoperiod of 18 h of light per day; M, males; Ppar‐α, peroxisome proliferator‐activated receptor alpha.

FIGURE 7

Effect of sex and photoperiods on the relative gene and protein expression of lipid transport‐related genes in the liver. Relative hepatic mRNA of Mtp (A) and Fabp4 (B) and relative protein expression levels of MTP (C) and FABP4 (D) in male and female F344 rats fed with CAF diet for 11 weeks and exposed to L18 or L6 for the last 8 weeks. Data are expressed as minimum to maximum values, median and interquartile range (n = 10 for relative hepatic mRNA and n = 6 for relative protein expression levels). P, photoperiod effect; S, sex effect; S × P, interaction between sex and photoperiod assessed using two‐way ANOVA for Fabp4, MTP and FABP4, and two‐way non‐parametric ANOVA with ART transformation for Mtp. *Indicates significant differences (*p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001) and “a” indicates a trend (p < 0.1) between groups using two‐way ANOVA followed by Tukey post hoc test for Fabp4, MTP and FABP4, and post hoc analysis with Tukey adjustment using emmeans package in R for Mtp. CAF, cafeteria; F, females; F344, Fischer 344; Fabp4, fatty acid binding protein 4; M, males; L6, short photoperiod of 6 h of light per day; L18, long photoperiod of 18 h of light per day; Mtp, microsomal triglyceride transfer protein.

FIGURE 8

Effect of sex and photoperiods on the relative gene expression of genes involved in hepatic cholesterol metabolism. Scarb1 (A), Abca1 (B), and Hmg‐CoA reductase (C) relative hepatic mRNA levels in male and female F344 rats fed with CAF diet for 11 weeks and exposed to L18 or L6 for the last 8 weeks. Data are expressed as minimum to maximum values, median and interquartile range (n = 10). S × P, interaction between sex and photoperiod assessed using two‐way ANOVA for Abca1 and Hmg‐CoA reductase, and two‐way non‐parametric ANOVA with ART transformation for Scarb1. Abca1, ATP‐binding cassette transporter 1; CAF, cafeteria; F, females; F344, Fischer 344; Hmg‐CoA reductase, 3‐hydroxy‐3‐methylglutaryl‐CoA reductase; L6, short photoperiod of 6 h of light per day; L18, long photoperiod of 18 h of light per day; M, males; Scarb1, scavenger receptor class B member 1.

FIGURE 9

Summary of the main metabolic pathways implicated in the management of the hepatic lipids in male and female F344 rats fed with CAF diet for 11 weeks and exposed to L18 or L6 for the last 8 weeks. Acaca, acetyl‐CoA carboxylase alpha; CAF, cafeteria; Cpt‐1α, carnitine palmitoyltransferase I alpha; Dgat, diacylglycerol O‐acyltransferase 1; F344, Fischer 344; Fabp4, fatty acid binding protein 4; Fasn, fatty acid synthase; HDL‐C, high‐density lipoprotein cholesterol; H‐TAG, hepatic triglycerides; H‐TC, hepatic total cholesterol; L18, long photoperiod of 18 h of light per day; L6, short photoperiod of 6 h of light per day; Mtp, microsomal triglyceride transfer protein; Ppar‐α, peroxisome proliferator‐activated receptor alpha; TAG, triglycerides; TC, total cholesterol; VLDL, very‐low‐density lipoprotein.