BACKGROUND:Metabolic dysfunction associated steatotic liver disease (MASLD), the most common chronic liver disease globally, may originate early in life. While maternal obesity is linked to offspring MASLD, the roles of paternal obesity and mediation by childhood adiposity remain unclear. OBJECTIVES:This study evaluates prospective associations between pre-pregnancy biparental adiposity and offspring MASLD in adulthood. DESIGN:We included 1933 offspring from the UK Avon Longitudinal Study of Parents and Children (ALSPAC) to assess the associations between parental pre-pregnancy body mass index (BMI) and odds of offspring MASLD at age 24 years. MASLD was defined as hepatic steatosis on transient elastography and ≥1 cardiometabolic risk factors. We evaluated causal mediation by childhood adiposity measures. RESULTS:At age 24 years, 10.4% of offspring had MASLD. Pre-pregnancy maternal and paternal obesity were independently associated with an increased odds of offspring MASLD. Each 1 kg/m2 increase in maternal BMI increased the odds of MASLD by 10% (Odds Ratio [OR] 1.10, 95% CI 1.06 to 1.14), while each 1 kg/m2 increase in paternal BMI raised the odds by 9% (OR 1.09, 95% CI 1.04 to 1.13). Biparental overweight or obesity was associated with 3.73 times the odds of offspring MASLD (OR 3.73, 95% CI 2.43 to 5.73) compared with parents with a normal BMI, with 67% of this association mediated by cumulative excess childhood BMI, a defined area under the curve for BMI Z score >1 for ages 7-17 years. CONCLUSIONS:Excess parental adiposity pre-pregnancy was associated with a higher odds of offspring MASLD, mediated by cumulative excess childhood BMI, highlighting the potential of life course interventions to reduce the risk of MASLD in future generations.

Disclosure: H. Hinrichs: None. N. Ramavath: None. M. Young: None. F. Ramirez Victorino: None. T.M. Bigley: None. M.D. Thompson: None. Introduction/BackgroundMaternal obesogenic diet exposure (MODE) promotes worse steatotic liver disease (SLD) in offspring. One mechanism of transmission is via vertical transfer of an altered microbiome to offspring, but the mechanism for how an altered offspring microbiome increases susceptibility to SLD is not clear. A critical early life event termed the ‘weaning reaction’ is dependent on the early microbiome and when altered, results in worse pathologic inflammation. We have shown that MODE attenuates the weaning reaction promoting worse SLD in mice in an early microbiome-dependent manner. We sought to define a potential intermediate between the early microbiome changes and attenuation of weaning reaction.Materials and MethodsBeginning at four weeks of age, female mice were fed either chow (CON) or high fat, fructose, cholesterol (HFFC) diet for 6 weeks before being bred with lean males. For cross-foster studies, offspring were switched between CON and HFFC dams within 24 hours of birth. Mice were gavaged with ursodeoxycholic acid (UDCA) daily between 2 and 3 weeks of age to assess the role of UDCA supplementation in the weaning reaction and immune cell development. Tissues were collected from offspring between 2 and 4 weeks of age. Ileal expression of Tnfa and Ifng was measured. Histology was performed on liver to assess inflammation and proliferation. BAs in liver and cecal contents were measured via mass spectrometry. Flow cytometry was performed on ileum at 4 weeks of age.ResultsBA profiling on 3-week-old cecal contents and liver identified an increase in abundance of tauro- and glycine- conjugated primary BAs and a decrease in the abundance of secondary BAs (UDCA) in HFFC offspring. Cross fostering shows that shifts in BAs are dependent on the fostering dam diet. HFFC offspring exhibit increased hepatocyte proliferation and hepatic inflammation in the perinatal period that is dependent on the fostering dam diet. HFFC offspring had a reduction in CD45+ lymphocytes and CD4+Foxp3+ regulatory T cells in the ileum. UDCA supplementation reestablished the weaning reaction and increased CD45+ lymphocytes and CD4+Foxp3+ regulatory T cells in the ileum of HFFC offspring.ConclusionsThese findings identify that that MODE alters early gut immune programming events in offspring intestine with modifications in BA metabolism. Supplementing MODE offspring with secondary BAs like UDCA may be an approach to restore the weaning reaction and immune cell populations. Presentation: Monday, July 14, 2025

Maternal obesogenic diet exposure (MODE) promotes fibroinflammatory liver disease in offspring via vertical transfer of an altered microbiome. The mechanism for how an altered offspring microbiome increases susceptibility to liver disease is not clear. A critical early life event termed the 'weaning reaction' is dependent on the early microbiome and when altered, results in worse pathologic inflammation. MODE attenuates the weaning reaction promoting worse liver disease in mice in an early microbiome-dependent manner. Using our MODE model and cross-fostering approaches we assessed the effect of MODE on neonatal gut-liver axis development. MODE shifts the bile acid (BA) profile, expression of intestinal barrier genes, and establishment of gut immune cell populations in the offspring. Among the BA changes, UDCA is decreased in MODE offspring and supplementation with UDCA at 2 weeks of age reestablishes the weaning reaction and gut immune cell development. MODE offspring exhibit worse hepatic inflammation and fibrosis during dextran sodium sulfate induced colitis. These findings identify that MODE alters early gut immune programming events in offspring intestine with modifications in BA metabolism. Supplementing MODE offspring with secondary BAs may be an approach to restore the weaning reaction and immune cell populations.

Introduction: Metabolic Dysfunction Associated Steatotic Liver Disease (MASLD) is a chronic liver disease that affects up to a fourth of the adults in the US. Maternal obesogenic diet exposure (MODE) programs worse MASLD in offspring. The beneficial effects of exercise on metabolic disease are well defined. We hypothesized that offspring exercise would mitigate the developmental programming effects of MODE. Methods: To determine the effect of offspring exercise, male offspring from dams fed chow or High Fat Fructose and Cholesterol (HFFC) diet underwent voluntary exercise. The three exercise conditions include voluntary exercise on a wheel for 3 weeks before (pre exercise) feeding a MASLD-inducing diet, exercise from 3 weeks before and during MASLD-diet feeding (continuous exercise) and sedentary conditions. Body weight and liver weights were measured. Histologic analysis and qPCR was performed to assess disease severity. Results: Exercise before and during HFFC feeding decreased body weight and liver weight to body weight ratio. Preliminary results show a reduction in steatosis and inflammation in offspring with continued exercise during HFFC feeding. Both pre-exercise alone and continued exercise during HFFC feeding show a trend toward attenuating the increase in fibrosis observed in MODE offspring. Conclusion: Preliminary data support that exercise may attenuate developmentally programmed worsening of MASLD by MODE. Continual exercise appears to have a greater benefit than pre-exercise alone. Disclosure E. Cantio: None. E. Reginelli: None. H.M. Hinrichs: None. M. Young: None. M. Thompson: None. Funding National Institute of Diabetes And Digestive And Kidney Diseases of the National Institutes of Health (1R25DK113652)

The mechanisms by which maternal obesity increases the susceptibility to steatotic liver disease in offspring are incompletely understood. Models using different maternal obesogenic diets (MODEs) display phenotypic variability, likely reflecting the influence of timing and diet composition. This study compared three maternal obesogenic diets using standardized exposure times to identify differences in offspring disease progression. This study found that the severity of hepatic inflammation and fibrosis in the offspring depends on the composition of the maternal obesogenic diet. Offspring cecal microbiome composition was shifted in all MODE groups relative to control. Decreased α-diversity in some MODE offspring with shifts in abundance of multiple genera were suggestive of delayed maturation of the microbiome. The weaning reaction typically characterized by a spike in intestinal expression of Tnfa and Ifng was attenuated in MODE offspring in an early microbiome-dependent manner using cross-fostering. Cross-fostering also switched the severity of disease progression in offspring dependent on the diet of the fostering dam. These results identify maternal diet composition and timing of exposure as modifiers in mediating transmissible changes in the microbiome. These changes in the early microbiome alter a critical window during weaning that drives susceptibility to progressive liver disease in the offspring.

Abstract Disclosure: H. Hinrichs: None. S.J. Ballentine: None. M.D. Thompson: None. Background: The Developmental Origins of Health and Disease (DOHaD) hypothesis states that development of chronic disease is impacted by early life or in utero exposures. Recent studies have found that exposure to maternal obesity/obesogenic diet increases the susceptibility of offspring to non-alcoholic fatty liver disease (NAFLD). We have shown that maternal obesogenic diet exposure (MODE) in mice leads to worse NAFLD progression to fibrosis in first generation offspring with associated changes in the microbiome. Evidence indicates that some developmentally programmed phenotypes are passed transgenerationally or to the third generation of offspring. The goal of the current study was to evaluate if MODE-induced programming of worse NAFLD progression is passed transgenerationally. Methods: F0 Female mice were fed chow (CON) or high fat-fructose-cholesterol (HFFC) diet for 6 weeks and bred with lean males. F1 and F2 females were fed chow diet and mated with chow-fed males to yield an F3 generation where only the F0 female was fed HFFC diet. F3 male offspring were weaned to HFFC diet for 7 weeks to induce progressive NAFLD. Liver was collected for histopathologic analysis. NAFLD activity scoring (NAS) was performed by a pathologist blinded to group. Results: F3 HFFC offspring had decreased liver weight and liver weight/body weight ratio compared to F3 CON offspring. F3 HFFC offspring had increased hepatic free fatty acids, but no difference in hepatic triglycerides and cholesterol. Mac-2 and CD45 staining was increased in F3 HFFC offspring. Sirius red staining for collagen was also increased in F3 HFFC offspring. NAS scoring confirmed histologic finding showing an increased frequency of higher inflammation and total NAS scores in F3 HFFC offspring. Fibrosis scores were also higher in F3 HFFC offspring. Conclusions: MODE leads to transgenerational transmission of worse NAFLD progression in male offspring. This suggests a role for an epigenetic mechanism involved in developmental programming of NAFLD. Future studies will focus on the mechanisms of this transgenerational event. Presentation: Friday, June 16, 2023

Abstract Disclosure: N. Ramavath: None. H. Hinrichs: None. M. Young: None. M. Thompson: None. Background: Obesity is a complex metabolic syndrome associated with a wide range of abnormalities in liver known as non-alcoholic fatty liver diseases (NAFLD). Evidence supports that maternal obesity/obesogenic diet exposure programs increased NAFLD offspring. The liver responds to lipotoxic hepatocyte injury through hepatocyte proliferation. It is not clear whether maternal obesogenic diet exposure affects the capacity of offspring hepatocytes to proliferate. We hypothesized that early impact of maternal/ obesogenic diet exposure in early life or utero exposure enhances hepatocyte proliferation capacity in offspring. Methods: Female mice were fed chow or high fat-fructose-cholesterol (HFFC) diet for 6 weeks and bred with lean males Liver and serum samples were collected at different postnatal time periods 5, 10, 15, 20, 25, and 30 days to analyze histological and molecular markers for liver proliferation. In addition, a small group of offspring from each group were weaned to the HFFC diet for 7 weeks to analyze progression of NAFLD (non -alcoholic fatty liver diseases). Results: No significant difference between neonatal body weight and liver weight was observed; however, the liver weight/body weight was increased at day 20 in HFFC offspring compared to chow offspring. The number of ki-67 positive hepatocytes peaked in both groups at day 15. An increase in ki-67 positive hepatocytes was observed at 15, 20, and 30 days in HFFC offspring compared to chow offspring. After 7 weeks of HFFC diet feeding, the number of Ki-67 positive hepatocytes was higher in HFFC offspring in comparison with Chow offspring. Conclusion: Maternal obesogenic diet exposure affected post-natal and NAFLD-associated hepatocyte proliferation in offspring with an increase in hepatocyte proliferative capacity. This could have implications for the ability of maternal obesity exposed offspring to respond to hepatic injury and potential for development of hepatic cancer. Presentation: Thursday, June 15, 2023

Maternal obesity programs the risk for development of nonalcoholic fatty liver disease (NAFLD) in offspring. Maternal exercise is a potential intervention to prevent developmentally programmed phenotypes. We hypothesized that maternal exercise would protect from progression of NAFLD in offspring previously exposed to a maternal obesogenic diet. Female mice were fed chow (CON) or high fat, fructose, cholesterol (HFFC) and bred with lean males. A subset had an exercise wheel introduced 4 weeks after starting the diet to allow for voluntary exercise. The offspring were weaned to the HFFC diet for 7 weeks to induce NAFLD. Serum, adipose, and liver tissue were collected for metabolic, histologic, and gene expression analyses. Cecal contents were collected for 16S sequencing. Global metabolomics was performed on liver. Female mice fed the HFFC diet had increased body weight prior to adding an exercise wheel. Female mice fed the HFFC diet had an increase in exercise distance relative to CON during the preconception period. Exercise distance was similar between groups during pregnancy and lactation. CON-active and HFFC-active offspring exhibited decreased inflammation compared with offspring from sedentary dams. Fibrosis increased in offspring from HFFC-sedentary dams compared with CON-sedentary. Offspring from exercised HFFC dams exhibited less fibrosis than offspring from sedentary HFFC dams. While maternal diet significantly affected the microbiome of offspring, the effect of maternal exercise was minimal. Metabolomics analysis revealed shifts in multiple metabolites including several involved in bile acid, 1-carbon, histidine, and acylcarnitine metabolism. This study provides preclinical evidence that maternal exercise is a potential approach to prevent developmentally programmed liver disease progression in offspring.

Human and animal model data show that maternal obesity promotes nonalcoholic fatty liver disease in offspring and alters bile acid (BA) homeostasis. Here we investigated whether offspring exposed to maternal obesogenic diets exhibited greater cholestatic injury. We fed female C57Bl6 mice conventional chow (CON) or high fat/high sucrose (HF/HS) diet and then bred them with lean males. Offspring were fed 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) for 2 weeks to induce cholestasis, and a subgroup was then fed CON for an additional 10 days. Additionally, to evaluate the role of the gut microbiome, we fed antibiotic-treated mice cecal contents from CON or HF/HS offspring, followed by DDC for 2 weeks. We found that HF/HS offspring fed DDC exhibited increased fine branching of the bile duct (ductular reaction) and fibrosis but did not differ in BA pool size or intrahepatic BA profile compared to offspring of mice fed CON. We also found that after 10 days recovery, HF/HS offspring exhibited sustained ductular reaction and periportal fibrosis, while lesions in CON offspring were resolved. In addition, cecal microbiome transplant from HF/HS offspring donors worsened ductular reaction, inflammation, and fibrosis in mice fed DDC. Finally, transfer of the microbiome from HF/HS offspring replicated the cholestatic liver injury phenotype. Taken together, we conclude that maternal HF/HS diet predisposes offspring to increased cholestatic injury after DDC feeding and delays recovery after returning to CON diets. These findings highlight the impact of maternal obesogenic diet on hepatobiliary injury and repair pathways during experimental cholestasis.

Mice exposed in gestation to maternal high-fat/high-sucrose (HF/HS) diet develop altered bile acid (BA) homeostasis. We hypothesized that these reflect an altered microbiome and asked if microbiota transplanted from HF/HS offspring change hepatic BA and lipid metabolism to determine the directionality of effect. Female mice were fed HF/HS or chow (CON) for 6 wk and bred with lean males. 16S sequencing was performed to compare taxa in offspring. Cecal microbiome transplantation (CMT) was performed from HF/HS or CON offspring into antibiotic-treated mice fed chow or high fructose. BA, lipid metabolic, and gene expression analyses were performed in recipient mice. Gut microbiomes from HF/HS offspring segregated from CON offspring, with increased Firmicutes to Bacteriodetes ratios and Verrucomicrobial abundance. After CMT was performed, HF/HS-recipient mice had larger BA pools, increased intrahepatic muricholic acid, and decreased deoxycholic acid species. HF/HS-recipient mice exhibited downregulated hepatic Mrp2, increased hepatic Oatp1b2, and decreased ileal Asbt mRNA expression. HF/HS-recipient mice exhibited decreased cecal butyrate and increased hepatic expression of Il6. HF/HS-recipient mice had larger livers and increased intrahepatic triglyceride versus CON-recipient mice after fructose feeding, with increased hepatic mRNA expression of lipogenic genes including Srebf1, Fabp1, Mogat1, and Mogat2. CMT from HF/HS offspring increased BA pool and shifted the composition of the intrahepatic BA pool. CMT from HF/HS donor offspring increased fructose-induced liver triglyceride accumulation. These findings support a causal role for vertical transfer of an altered microbiome in hepatic BA and lipid metabolism in HF/HS offspring. NEW & NOTEWORTHY We utilized a mouse model of maternal obesogenic diet exposure to evaluate the effect on offspring microbiome and bile acid homeostasis. We identified shifts in the offspring microbiome associated with changes in cecal bile acid levels. Transfer of the microbiome from maternal obesogenic diet-exposed offspring to microbiome-depleted mice altered bile acid homeostasis and increased fructose-induced hepatic steatosis.