AMPK Deficiency Increases DNA Methylation and Aggravates Colorectal Tumorigenesis in AOM/DSS Mice.

The incidence of colorectal cancer (CRC) is closely linked to metabolic diseases. Accumulating evidence suggests the regulatory role of AMP-activated protein kinase (AMPK) in cancer metabolic reprogramming. In this study, wild-type and AMPK knockout mice were subjected to azoxymethane-induced and dextran sulfate sodium (AOM/DSS)-promoted colitis-associated CRC induction. A stable AMPK-deficient Caco-2 cell line was also established for the mechanistic studies. The data showed that AMPK deficiency accelerated CRC development, characterized by increased tumor number, tumor size, and hyperplasia in AOM/DSS-treated mice. The aggravated colorectal tumorigenesis resulting from AMPK ablation was associated with reduced α-ketoglutarate production and ten-eleven translocation hydroxylase 2 (TET2) transcription, correlated with the reduced mismatch repair protein mutL homolog 1 (MLH1) protein. Furthermore, in AMPK-deficient Caco-2 cells, the mRNA expression of mismatch repair and tumor suppressor genes, intracellular α-ketoglutarate, and the protein level of TET2 were also downregulated. AMPK deficiency also increased hypermethylation in the CpG islands of Mlh1 in both colonic tissues and Caco-2 cells. In conclusion, AMPK deficiency leads to reduced α-ketoglutarate concentration and elevates the suppressive epigenetic modifications of tumor suppressor genes in gut epithelial cells, thereby increasing the risk of colorectal tumorigenesis. Given the modifiable nature of AMPK activity, it holds promise as a prospective molecular target for the prevention and treatment of CRC.

Vitamin a supplementation during pregnancy in shaping child growth outcomes: A meta-analysis.

Abnormal fetal growth increases risks of childhood health complications. Vitamin A supplementation (VAS) is highly accessible, but literature inconsistency regarding effects of maternal VAS on fetal and childhood growth outcomes exists, deterring pregnant women from VAS during pregnancy. This meta-analysis aimed to analyze effects of vitamin A only or vitamin A + co-intervention during pregnancy in healthy mothers (MH) or with complications (MC, night blindness and HIV positive) on perinatal growth outcomes, also assess VAS dose impacts. The Cochrane Library, PubMed, ScienceDirect, Scopus, Embase and Web of Science databases were searched from inception to July 15, 2021. We covered subgroup analyses, including VAS in MH or MC within randomized controlled trial (RCT) or observational studies (OS). Fifty-five studies were included in this meta-analysis (426,098 pregnancies). Vitamin A decreased risk of preterm birth by 9% in MH-RCT (P < 0.001), by 62% in MH-OS (P = 0.029), by 10% in MC-RCT (P = 0.089); decreased LBW by 24% in MC-RCT (P = 0.032); increased neonatal weight in MC-RCT (SMD 0.96; P = 0.051). Besides, vitamin A + co-intervention decreased risks of preterm by 18% in MH-OS (P = 0.021); LBW by 25% in MH-OS (P < 0.001); by 32% in MC-RCT (P = 0.006); decreased neonatal defects by 33% in MH-OS (P = 0.064); decreased anemia by 25% in MH-OS (P = 0.0003); increased neonatal weight in MH-OS (SMD 0.51; P = 0.014); and increased neonatal length in MH-OS (SMD 1.83; P = 0.013). Meta-regression of VAS dose with individual outcomes was not significant, and no side effects were observed for VAS doses up to 4000 mcg (RAE/d). Regardless of maternal health conditions, VAS during pregnancy can safely and effectively improve fetal development and neonatal health even in mothers without VAD.

Maternal Exercise Before and During Pregnancy Facilitates Embryonic Myogenesis by Enhancing Thyroid Hormone Signaling.

Background: Maternal exercise (ME) improves fetal and offspring muscle development, but mechanisms remain to be established. Since the thyroid hormone (TH) is critical for cell differentiation during embryonic development, we hypothesized that ME elevates TH receptor (THR) signaling in embryos, which promotes embryonic myogenesis. Methods: Female mice were exercised daily on a treadmill or received a daily TH, triiodothyronine (T3) injection. Embryos (embryonic day 12.5 [E12.5]) and P19 cells were used for studying effects of TH on embryonic myogenesis. TH levels in serum and embryos after ME or T3I were analyzed. Expression of TH signaling related genes and myogenic genes was assessed. THRα binding to the promoters of myogenic genes was investigated by chromatin immunoprecipitation-qantitative polymerase chain reaction (ChIP-qPCR). A CRISPR/CAS9 plasmid was utilized to knock out THRα in P19 cells. Results: ME elevated TH levels in both maternal circulation and embryos, which were correlated with enhanced TH signaling and myogenesis. At E12.5, both myogenic determinants (Pax3, Pax7) and myogenic regulatory factors (Myf5, Myod) were upregulated in ME embryos. ME increased THRα content and elevated messenger RNA (mRNA) expression of TH transporter Slc16a2 and deiodinase Dio2. In addition, the THRα binding to the promoters of Pax3/7 was increased. In P19 embryoid bodies, T3 promoted myogenic differentiation, which was abolished by ablating THRα. Furthermore, maternal daily injection of T3 at a level matching exercised mothers promoted embryonic myogenesis. Conclusions: ME promotes TH delivery to the embryos and enhances embryonic myogenesis, which is partially mediated by enhanced TH signaling in ME embryos.

Dietary succinate supplementation to maternal mice improves fetal brown adipose tissue development and thermogenesis of female offspring.

Succinic acid widely exists in foods and is used as a food additive. Succinate not only serves as an energy substrate, but also induces protein succinylation. Histone succinylation activates gene transcription. The brown adipose tissue (BAT) is critical for prevention of obesity and metabolic dysfunction, and the fetal stage is pivotal for BAT development. Up to now, the role of maternal succinate supplementation on fetal BAT development and offspring BAT function remains unexamined. To test, female C57BL/6J mice (2-month-old) were separated into 2 groups, received with or without 0.5% succinic acid in drinking water during gestation and lactation. After weaning, female offspring were challenged with high fat diet (HFD) for 12 weeks. Newborn, female weanling, and HFD female offspring mice were analyzed. For neonatal and weaning mice, the BAT weight relative to the whole body weight was significantly increased in the succinate group. The expression of PGC-1α, a key transcription co-activator promoting mitochondrial biogenesis, was elevated in BAT of female neonatal and offspring born to succinate-fed dams. Consistently, maternal succinate supplementation enhanced thermogenesis and the expression of thermogenic genes in offspring BAT. Additionally, maternal succinate supplementation protected female offspring against HFD-induced obesity. Furthermore, in C3H10T1/2 cells, succinate supplementation promoted PGC-1α expression and brown adipogenesis. Mechanistically, succinate supplementation increased permissive histone succinylation and H3K4me3 modification in the Ppargc1a promoter, which correlated with the higher expression of Ppargc1a. In conclusion, maternal succinate supplementation during pregnancy and lactation enhanced fetal BAT development and offspring BAT thermogenesis, which prevented HFD-induced obesity and metabolism dysfunction in offspring.

Effects of Maternal Exercise During Pregnancy on Perinatal Growth and Childhood Obesity Outcomes: A Meta-analysis and Meta-regression.

BACKGROUND: Perinatal growth abnormalities program susceptibility to childhood obesity, which is further exaggerated by maternal overweight and obesity (MO) during pregnancy. Exercise is highly accessible, but reports about the benefits of maternal exercise on fetal growth and childhood obesity outcomes are inconsistent, reducing the incentives for pregnant women to participate in exercise to improve children's perinatal growth. OBJECTIVE: This systematic review and meta-analysis aims to establish evidence-based efficacy of exercise in mothers with normal weight (MNW) and MO during pregnancy in reducing the risks of perinatal growth abnormalities and childhood obesity. In addition, the impacts of exercise volume are also assessed. METHODS: The PubMed, ScienceDirect, Web of Science, and Cochrane Library databases were searched from inception to February 15, 2020. We included randomized controlled trials with exercise-only intervention or exercise with other confounders in pregnant MNW (body mass index, BMI 18.5-24.9 kg/m2) and MO (BMI ≥ 25 kg/m2), which were further subgrouped in the meta-analysis. Primary outcomes included birth weight, preterm birth, small for gestational age (SGA), large for gestational age (LGA), infant and childhood weight, and childhood obesity. A linear meta-regression analysis was also used to explore the effects of exercise volume on outcomes. RESULTS: 99 studies were included in the meta-analysis (n = 596,876), and individual study quality ranged from fair to good according to the Newcastle-Ottawa scale assessment. Exercise only interventions in MNW reduced preterm birth by 15% (26 studies, n = 76,132; odds ratio [OR] 0.85; 95% CI 0.72, 1.01; I2 = 83.3%), SGA by 17% (33 studies, n = 92,351; OR 0.83; 95% CI 0.71, 0.98; I2 = 74.5%) and LGA by 17% (29 studies, n = 84,310; OR 0.83; 95% CI 0.74, 0.95; I2 = 60.4%). Exercise only interventions in MO reduced preterm birth by 33% (2 studies, n = 3,050; OR 0.67; 95% CI 0.70, 0.96; I2 = 0%), SGA by 27% (8 studies, n = 3,909; OR 0.73; 95% CI 0.50, 1.05; I2 = 40.4%) and LGA by 55% (9 studies, n = 81,581; OR 0.45; 95% CI 0.18, 1.11; I2 = 98.3%). Exercise only interventions in MNW reduced childhood obesity by 53% (3 studies, n = 6,920; OR 0.47; 95% CI 0.36, 0.63; I2 = 77.0%). However, no significant effect was observed in outcomes from exercise confounders in either MNW or MO. In the meta-regression, the volume of exercise-only intervention in MNW was negatively associated with birth weight, greatly driven by volumes more than 810 metabolic equivalents (MET)-min per week. Other outcomes were not associated with exercise volume. CONCLUSIONS: This systematic review and meta-analysis suggests that exercise during pregnancy in both MNW and MO safely and effectively reduce the risks of preterm birth, SGA, and LGA. Furthermore, MNW exercise also reduces the risk of childhood obesity. Overall, regardless of prepregnancy BMI, maternal exercise during pregnancy provides an excellent opportunity to mitigate the high prevalence of adverse birth outcomes and childhood obesity.

Grape pomace inhibits colon carcinogenesis by suppressing cell proliferation and inducing epigenetic modifications.

Grape pomace (GP), a by-product of the wine and juice industry, is rich in bioflavonoids and dietary fibers. We hypothesized that GP has protective effects against colitis-associated colorectal cancer (CRC). Nine-week-old female mice were fed a control diet (CON) or CON with 5% grape pomace (GP) for 2 weeks, when mice were subjected to azoxymethane (AOM)/dextran sulfate sodium (DSS) induced-CRC induction. GP supplementation ameliorated the disease activity index (DAI) score, reduced tumor number, tumor size and pathological scores in AOM/DSS treated mice. Furthermore, dietary GP suppressed colonic expression of inflammatory cytokines, IL-1ß and TNF-α, and inhibited NF-κB inflammatory signaling, while increased anti-inflammatory cytokine TGF-ß mRNA expression. Colorectal inflammation is known to enhance Wnt signaling and cell proliferation. In agreement, the content of ß-catenin, a key downstream mediator of Wnt signaling, was reduced as was the expression of Cyclin D1, phosphorylation and content of p53 and PCNA level in GP-fed mice. In addition, GP reduced the expression of ALDH1, a marker of cell stemness, and increased the expression of Cdx2, a key transcription factor initiating epithelial cell differentiation, DNA methylation of the promoter region of Cdx2 gene and hypermethylation of CpG island methylator phenotype (CIMP), which commonly occurs during CRC carcinogenesis, was alleviated in the GP group. In conclusion, GP supplementation suppressed colitis-associated CRC carcinogenesis, which was associated with the suppression of inflammation and cell proliferation and the enhancement of DNA demethylation in Cdx2 and CIMP genes in the colon. These data suggest that dietary GP supplementation has preventive effects against colorectal carcinogenesis.

Dietary alpha-ketoglutarate promotes beige adipogenesis and prevents obesity in middle-aged mice.

Aging usually involves the progressive development of certain illnesses, including diabetes and obesity. Due to incapacity to form new white adipocytes, adipose expansion in aged mice primarily depends on adipocyte hypertrophy, which induces metabolic dysfunction. On the other hand, brown adipose tissue burns fatty acids, preventing ectopic lipid accumulation and metabolic diseases. However, the capacity of brown/beige adipogenesis declines inevitably during the aging process. Previously, we reported that DNA demethylation in the Prdm16 promoter is required for beige adipogenesis. DNA methylation is mediated by ten-eleven family proteins (TET) using alpha-ketoglutarate (AKG) as a cofactor. Here, we demonstrated that the circulatory AKG concentration was reduced in middle-aged mice (10-month-old) compared with young mice (2-month-old). Through AKG administration replenishing the AKG pool, aged mice were associated with the lower body weight gain and fat mass, and improved glucose tolerance after challenged with high-fat diet (HFD). These metabolic changes are accompanied by increased expression of brown adipose genes and proteins in inguinal adipose tissue. Cold-induced brown/beige adipogenesis was impeded in HFD mice, whereas AKG rescued the impairment of beige adipocyte functionality in middle-aged mice. Besides, AKG administration up-regulated Prdm16 expression, which was correlated with an increase of DNA demethylation in the Prdm16 promoter. In summary, AKG supplementation promotes beige adipogenesis and alleviates HFD-induced obesity in middle-aged mice, which is associated with enhanced DNA demethylation of the Prdm16 gene.

Retinoic acid induces white adipose tissue browning by increasing adipose vascularity and inducing beige adipogenesis of PDGFRα+ adipose progenitors.

Formation of beige adipocytes within white adipose tissue enhances energy expenditure, which is a promising strategy to reduce obesity and prevent metabolic symptoms. Vitamin A and its bioactive metabolite, retinoic acid (RA), have regulatory roles in lipid metabolism. Here we report that RA induces white adipose tissue browning via activating vascular endothelial growth factor (VEGF) signaling. RA triggered angiogenesis and elicited de novo generation of platelet-derived growth factor receptor α positive (PDGFRα+) adipose precursor cells via VEGFA/VEGFR2 signaling. In addition, RA promoted beige/brown adipocyte formation from capillary networks in vitro. Using PDGFRα tracking mice, we found that the vascular system acted as an adipogenic repository by containing PDGFRα+ progenitors which differentiated into beige adipocytes under RA or VEGF164 treatments. Conditional knockout of VEGF receptors blocked RA-stimulated white adipose tissue browning. Moreover, the VEGFA and RA activated p38MAPK to enhance the binding of RA receptor to RA response elements of the Prdm16 promoter and upregulated Prdm16 transcription. In conclusion, RA induces white adipose tissue browning by increasing adipose vascularity and promoting beige adipogenesis of PDGFRα+ adipose progenitors.

The effects of chronic ethanol consumption during early pregnancy on conceptus health and uterine function in pigs.

BACKGROUND: Maternal ethanol consumption impairs fetal health, but it is unclear if this occurs through direct actions on the conceptus or indirectly through effects on the uterus. The objective of this study was to determine if chronic ethanol consumption in swine would impair early embryonic and fetal health either through direct effects on the conceptus or indirect effects on the endometrium. METHODS: Four experiments evaluated the effects of chronic ethanol consumption during early pregnancy. Female pigs were fed either 350 ml of 95% ethanol or an isocaloric amount of dextrose at 10 to 14-hr intervals beginning on day 10 after pubertal estrus and continuing until ovariohysterectomy 11 to 35 days after mating. At the second estrus, pigs were mated to a fertile boar that did not consume alcohol. RESULTS: In experiment 1, ethanol consumption increased (p < 0.01) blood alcohol concentrations that peaked 2-3 hr after feeding. In experiment 2, ethanol was detectable in uterine flushings 2 hr after feeding on day 11 of pregnancy and was highly correlated (r = 0.989, p < 0.001) with blood alcohol concentration. In experiment 3, ethanol consumption did not affect endometrial phospholipase C activity on days 11 and 16 of pregnancy but decreased (p < 0.05) basal endometrial prostaglandin F(2alpha) production on day 16. However, ethanol consumption did not decrease the number of conceptuses on day 11 or conceptus DNA content on days 11 or 16. In experiment 4, ethanol consumption decreased (p < 0.05) fetal survival rate to 58% versus 85% in dextrose-fed controls on day 35 of pregnancy. For viable conceptuses, ethanol consumption reduced (p < 0.01) fetal weight, fetal crown-rump length, placental weight and volume of placental (chorio-allantoic + amniotic) fluid. CONCLUSION: These results indicate that chronic ethanol consumption may impair conceptus health directly or indirectly through actions upon the endometrium. Thus, the pig may be a valuable experimental model for studies on the effects of maternal alcohol consumption on conceptus development.