Fatty liver is a silent manifestation of the metabolic syndrome that affects millions worldwide leading to metabolic-associated fatty liver disease (MAFLD) with both a lean and overweight phenotype. There is a high risk of cardiovascular complications and mortality associated with the disease, that is more prevalent than liver pathology-related deaths. To study changes occurring in early stages of the disease can shed a light on the extend of those associations related to inter-organ crosstalk.
We hypothesized that chronic fructose intake leads to liver steatosis and can induce liver mitochondrial stress and an alarm response in the heart. In this project, we have investigated the consequences of fructose intake in the liver and heart, with a focus on mitochondrial stress and its drivers in liver-heart crosstalk under these conditions.
Methods
Animal experiments were designed according to European guidelines (FELASA; EU animal research directive 86/609/EEC and 2010/63/ EU) and approved by the local authority of the National Animal Research Authority in Norway. Male rats (Sprague-Dawley, 250 g) were given normal chow and tap water ad libitum (CON, n=12) or a 15% fructose drink (FRU, n=12) for 16 weeks. Body composition was measured throughout the intervention (EchoMRI) as well as tail-cuff measurement of blood pressure (CODA) and tail-vein blood samples. Serum cytokines (Bioplex) were analyzed. Tissue from liver and heart was obtained postmortem following 16-weeks fructose intake. Liver triglycerides (TG) were assessed by a colorimetric assay. Cardiac and liver mitochondrial respiratory capacity was determined in tissue homogenate (O2K, Oroboros) and mitochondrial H2O2 production was determined in the heart. Histology and Reverse Transcription quantitative PCR were used to assess changes in liver and heart, including hematoxylin and eosin (H&E), PicroSirius Red (PSR), Oil Red O (ORO) and dihydroethidium (DHE) stainings.
Results
16-week fructose intake increased body weight, percentage of fat mass and induced liver steatosis. Histological analysis determined increased ORO staining of lipid droplets in liver and TG in tissue homogenate was increased. H&E staining did not reveal macrovesicular steatosis, nor was increased liver collagen deposition observed with PSR. Mean arterial pressure as well as serum levels of interleukin-6 and tumour necrosis factor-alpha were elevated in fructose (FRU) rats. The heart weights were elevated in FRU however histological analysis did not reveal changes in cardiomyocyte size by H&E staining, collagen deposition, lipid droplet and reactive oxygen species formation in the nuclei measured by DHE. Complex I-linked mitochondrial oxidative phosphorylation (CI-OXPHOS) and mitochondrial respiratory capacity was lower in liver homogenate from FRU and mRNA expression indicated metabolic changes, mitochondrial stress and elevated Fibroblast Growth Factor 21 (FGF21). CI-OXPHOS was also lower in heart homogenates from FRU along with altered mitochondrial H2O2 production.
Conclusion
Liver steatosis is associated with cardiac adaptations that may contribute to the development of heart failure. Increased FGF21 expression levels in the liver indicate potential driver of detrimental liver-heart crosstalk, a role that is of interest for further investigation.