The present study emphasizes the impact of preconceptional paternal calorie restriction (CR) on various metabolic and physiological parameters in high-fat-fed male Wistar rats and their F1 offspring. The findings shed light on the intricate interplay between paternal diet, metabolic health, and offspring outcomes, highlighting potential mechanisms underlying transgenerational metabolic programming.
Under the influence of the HF diet, the male parents demonstrated increased body weights, gonadal fat weights and reduced testosterone levels which likely suggest their compromised male fertility [43, 44, 45]. It was also observed that the abnormal testis & sperm morphologies in the HF group led to reduced spermatogenesis and reproduction capability which is in accordance with the assessment of male fertility [38]. Studies have reported that paternal undernutrition reduces male fertility which in turn is responsible for metabolic disorders in the offspring [13, 24, 25]. In the present study, a noteworthy observation emerged in the group subjected to a 50% CR (HFCR-I) within the diet-induced obese Wistar rats. Instead of rectifying the detrimental effects induced by the high-fat diet, this level of CR resulted in a state akin to undernutrition. Consequently, this undernourished condition had adverse repercussions on male fertility and impacted the health of their offspring. Whereas, implementing a 40% CR in diet-induced obese Wistar rats (HFCR-II) nearly reinstated the effects induced by a high-fat diet. Current findings also noticed altered DNMT gene expression in the testes of parents that may show epigenetic influence on their offspring metabolism. The offspring of HF and HFCR-I have shown obesogenic tendencies with increased inflammation leading to impaired glucose homeostasis by exhibiting different pathophysiology.
A study has found that moderate CR of obese male mice leads to the upregulation of SIRT1 which has beneficial effects on reproductive health [46]. In line, another study also demonstrated that SIRT1 knock-out mice has resulted in altered number & morphology of spermatozoa, causing reduced fertility [47]. Similar trends were noticed in the present study, where the SIRT1 downregulation in HF male parents is compromising their fertility. Moreover, in HFCR-I, despite SIRT1 is upregulated, their fertility was compromised due to nutrient deprivation caused by severe CR. These findings likely suggest that HF and HFCR-I may have different molecular mechanisms impacting male fertility. Further in both the cases, it was proven to have reduced mating frequency. The HFCR-I did not alter the litter size but did alter the male-to-female sex ratio which is commensurate with a study [48]. This shows that HFCR-I males differentially produce sperms bearing X and Y chromosomes, hence the birth of male offspring is favored. Alternatively, low protein diet semen could create an environment in the uterus that is more favorable to female preimplantation embryos, leading to the loss of male embryos in a selective manner [49].
SIRT1, a histone deacetylase, causes the silencing of genes by recruiting DNMTs, and it was also reported that SIRT1 downregulation leads to the downregulation of DNMT1, DNMT3a, and DNMT3b in the breast cancer cells of mice [50]. SIRT1 is also known to inhibit the other histone deacetylase HDAC1 [51]. In the present study, irrespective of decreased and increased SIRT1 expression in HF and HFCR-I respectively; there was a downregulation of DNMT1, DNMT3a, DNMT3b and HDAC1 which could have modulated the methylation/imprinting of genes in the offspring affecting their health. Thus, this establishes the relation between the paternal CR to their DNMT’s differential expression affecting the metabolism in their offspring.
While there is ample research available on how maternal nutritional interventions affect metabolic pathways in offspring, the effects of paternal nutritional strategies remain relatively underreported [15]. In rats, maternal CR downregulates the AMPK pathway in offspring, which subsequently causes various metabolic disorders, such as obesity, CVD, and steatohepatitis. Dysregulation of the underlying molecular mechanism has been linked to hyperacetylation of PGC-1α and reduced SIRT1 expression and function [52, 53]. Similarly, in the present study, the male and female offspring born to HF and HFCR-I have shown the downregulation of AMPK/SIRT1/adiponectin pathways that led to the dysregulation of inflammation, lipid and glucose metabolism in them which clearly demonstrates the importance of paternal diet. This also indicates the effects of paternal CR are mostly similar to the maternal CR. In HFCR-II offspring the gene expression was mostly comparable to the controls. Hence, moderate CR can be considered as an effective weight loss regime in obese individuals without causing much effect on their future generations.
Earlier mice studies reported that the birthweights of male offspring are reduced under a paternal high-fat diet [54], and it was also reported that paternal malnutrition led to a decreased birth weight of female offspring [23]. Our results are in accordance with these studies in which low birthweights of male and female offspring of HF and HFCR-I showed that these offspring may be prone to metabolic disorders at an early age. It was reported earlier in mice that fathers’ high-fat diet exposure leads to increased body weight gain in offspring [55], and paternal malnutrition leads to weight gain in offspring at an early age [23, 25]. Additionally, in the present study, male and female offspring of HF and HFCR-I gained more weight after weaning, which goes along with their energy intake. This shows the pattern of premature births or children born small catching up weight at an early age. Leptin resistance with increased FER leads to obesity [56]. Further, the increased plasma leptin in the HF and HFCR-I offspring shows that they are prone to leptin resistance irrespective of their FER differences, indicating that they exhibit different pathophysiology of obesity. In mice, the father’s high-fat diet led to increased fat mass in the offspring [55], and the paternal low-protein diet in rats was also associated with increased fat mass and organ weights in the offspring [24, 57]. The present findings are also in line with those of previous studies in which the total body fat percentage increased with increased adiposity index in HF and HFCR-I offspring, demonstrating that they exhibit imbalanced body fat accumulation.
High-fat diet exposure in fathers leads to increased serum lipid and lipid synthesis in offspring [19, 58, 59]. In mice, a paternal low-protein diet has also shown increased total cholesterol and triglycerides in the offspring [57] and increased lipogenesis gene expression in male offspring [60]. The present study revealed greater plasma lipid levels in HF and HFCR-I offspring than in HFCR-II offspring, suggesting that a paternal high-fat diet and CR lead to obesogenic effects in offspring, which was further confirmed by their increased mRNA expression of the lipid synthesis genes FAS and SCD1 and decreased expression of the lipid β-oxidation genes CPT1 and ACOX2. The expression difference was greater in females than in males, demonstrating more adverse sex-specific effects of paternal diet. Interestingly, our results showed that a paternal high-fat diet and 50% CR increased liver weight and steatosis in the offspring, and these effects were severe in female offspring. The increased total cholesterol and triglycerides and the deposition of fat droplets in hepatocytes resulted in fatty liver conditions and dyslipidemia in the HF and HFCR-I offspring. These results illustrate that a father’s obesity can be inherited to their offspring, while paternal CR prompts the body to adapt to an energy-conserving mode in anticipation of limited resources. This adaptation may have led to reprogramming and imprinting in the sperm, potentially influencing the metabolic health of subsequent generations.
Multiple studies on paternal high-fat diets have shown increased inflammation in offspring [20, 58, 61 62]. A paternal low-protein diet has increased the TNF-α levels in the offspring [25]. The levels of the proinflammatory cytokines IL-6, IL-1β, TNF-α and MCP-1 in the circulation are greater in the HF and HFCR-I groups than in the HFCR-II group, resulting in increased inflammation, which is further supported by increased leptin and decreased TAC and catalase activity. In Wistar rats fed high-caloric diets, hypoadiponectinemia and hyperleptinemia restrict energy expenditure and glucose consumption, leading to reduced glycolysis and fatty acid metabolism [2]. A preconceptional father’s high-fat diet alters insulin sensitivity and glucose metabolism [58, 59]. The paternal low-protein diet has also led to impaired blood glucose levels in both male and female offspring [24, 25]. The elevated levels of fasting insulin and glucose, along with the delayed clearance of glucose upon oral glucose challenge in offspring from the HF and HFCR-I corroborate their β-cell dysfunction and dysregulated glucose homeostasis. This finding was further supported by the increased AUC, HOMA-IR and HOMA-β with decreased and increased expression of glycolytic and gluconeogenesis enzymes respectively. These findings demonstrated that the HF and HFCR-I offspring groups exhibited increased inflammation and adiposity, leading to insulin resistance.
Hence this study illustrates that the modulation of AMPK/SIRT1 pathways in the offspring alters inflammation, lipid & glucose metabolism developing obesity and its associated co-morbidities at an early age.