Baifeng Yu

Promoter polymorphism of the erythropoietin gene in severe diabetic eye and kidney complications

Significant morbidity and mortality among patients with diabetes mellitus result largely from a greatly increased incidence of microvascular complications. Proliferative diabetic retinopathy (PDR) and end stage renal disease (ESRD) are two of the most common and severe microvascular complications of diabetes. A high concordance exists in the development of PDR and ESRD in diabetic patients, as well as strong familial aggregation of these complications, suggesting a common underlying genetic mechanism. However, the precise gene(s) and genetic variant(s) involved remain largely unknown. Erythropoietin (EPO) is a potent angiogenic factor observed in the diabetic human and mouse eye. By a combination of case–control association and functional studies, we demonstrate that the T allele of SNP rs1617640 in the promoter of the EPO gene is significantly associated with PDR and ESRD in three European-American cohorts [Utah: P = 1.91 × 10−3; Genetics of Kidneys in Diabetes (GoKinD) Study: P = 2.66 × 10−8; and Boston: P = 2.1 × 10−2]. The EPO concentration in human vitreous body was 7.5-fold higher in normal subjects with the TT risk genotype than in those with the GG genotype. Computational analysis suggests that the risk allele (T) of rs1617640 creates a matrix match with the EVI1/MEL1 or AP1 binding site, accounting for an observed 25-fold enhancement of luciferase reporter expression as compared with the G allele. These results suggest that rs1617640 in the EPO promoter is significantly associated with PDR and ESRD. This study identifies a disease risk-associated gene and potential pathway mediating severe diabetic microvascular complications.

Genetic association of LOXL1 gene variants and exfoliation glaucoma in a Utah cohort.

Exfoliation glaucoma (XFG) is the commonest identifiable cause of secondary open-angle glaucoma worldwide, characterized by the deposition of fibrillar proteins in the anterior segment of the eye. We investigated LOXL1 gene variants previously identified to confer susceptibility to exfoliation glaucoma (XFG) in a Utah Caucasian cohort. After a standard eye examination protocol we genotyped SNPs rs2165241 and rs3825942 in 62 XFG or XFS patients and 170 normal controls. Genotype frequency distribution, odds ratios (ORs), and population attributable risks were calculated for the risk alleles. The SNP rs2165241 was significantly associated with XFG and XFS (p=4.13x10-9 for an additive model, ORhet=4.42 (2.30-8.50), ORhom=34.19 (4.48-261.00); T allele: 83.1% in cases versus 52.4% in controls). Significant association was also found for rs3825942: (p=1.89x10-6). Our findings confirm genetic association of LOXL1 with XFG and XFS and implicate a potential role of cross linking of elastin in the pathogenesis of XFG. This information will potentially guide glaucoma monitoring efforts by targeting individuals whose genetic profiles put them at higher risk for XFG.

Intrauterine Growth Restriction Influences Vascular Remodeling and Stiffening in the Weanling Rat More than Sex or Diet

Intrauterine growth restriction (IUGR) increases the incidence of adult cardiovascular disease (CVD). The sex-specific developmental mechanisms for IUGR-induced and Western high-fat diet (HFD) modification of CVD remain poorly understood. We hypothesized a maternal HFD in the Sprague-Dawley rat would augment IUGR-induced CVD in the offspring through decreased cardiac function and increased extracellular matrix (ECM) remodeling and stiffness in a sex-specific manner. HFD or regular diet (Reg) was given from 5 wk before mating through postnatal day (PND) 21. IUGR was induced by uterine artery ligation at embryonic day 19.5 (term = 21.5 days). At PND 21, echocardiographic assessments were made and carotid arteries tested for vascular compliance using pressure myography. Arterial samples were quantified for ECM constituents or fixed for histologic evaluation. The insult of IUGR (IUGR + Reg and IUGR + HFD) led to increased mechanical stiffness in both sexes (P < 0.05). The combination of IUGR + HFD increased diastolic blood pressure 47% in males (M) and 35% in females (F) compared with the Con + Reg (P < 0.05). ECM remodeling in IUGR + HFD caused fewer (M = -29%, F = -24%) but thicker elastin bands (M = 18%, F = 18%) and increased total collagen (M = 49%, F = 34%) compared with Con + Reg arteries. Remodeling in IUGR + HFD males increased medial collagen and soluble collagen (P < 0.05). Remodeling in IUGR + HFD females increased adventitial collagen and wall thickness (P < 0.05) and decreased matrix metalloproteinase 2 (MMP-2), advanced glycosylation end products (AGE), and receptor AGE (RAGE; P < 0.05). In summary, both IUGR + Reg and IUGR + HFD remodel ECM in PND 21 rats. While IUGR + HFD increases blood pressure, IUGR but not HFD increases vascular stiffness suggesting a specific mechanism of vascular remodeling that can be targeted to limit future disease. NEW & NOTEWORTHY We report intrauterine growth restriction (IUGR) increases vascular stiffening in both male and female rats through increased collagen content and altered elastin structure more than a high-fat diet (HFD) alone. Our study shows the importance of stiffness supporting the hypothesis that there are physiologic differences and potential windows for early intervention targeting vascular remodeling mechanisms.

Combination of intrauterine growth restriction and a high-fat diet impairs cholesterol elimination in rats

Background:Intrauterine growth restriction (IUGR) increases the risk of adult-onset hypercholesterolemia. High-fat diet (HFD) consumption potentiates IUGR-induced increased cholesterol. Cholesterol is converted to bile acids by Cyp7a1 in preparation for excretion. We hypothesized that IUGR rats fed a HFD will have increased cholesterol, decreased Cyp7a1 protein levels, and decreased bile acids compared to control rats fed a HFD.Methods:At day 21, IUGR and control pups were placed on one of three diets: a regular chow or one of two HFDs containing 1% or 2% cholesterol. Cholesterol levels and hepatic Cyp7a1 protein levels were quantified a postnatal week 28.Results:Both HFDs increased serum cholesterol levels in control rats, and HFD fed IUGR rats had further increased serum cholesterol up to 35-fold. Both HFDs increased hepatic cholesterol levels, and IUGR further increased hepatic cholesterol levels up to fivefold. IUGR decreased hepatic Cyp7a1 protein up to 75%, and hepatic bile acids up to 54%.Conclusion:IUGR increased cholesterol and bile acids and decreased Cyp7a1 protein in rats fed a HFD without changing food intake. These findings suggest that IUGR increases the vulnerability of HFD fed rats to hypercholesterolemia via decreased cholesterol conversion to bile acids.

IUGR disrupts the PPARγ-Setd8-H4K20me1 and Wnt signaling pathways in the juvenile rat hippocampus

Intrauterine growth restriction (IUGR) programs neurodevelopmental impairment and long‐term neurological morbidities. Neurological morbidities in IUGR infants are correlated with changes hippocampal volume. We previously demonstrated that IUGR alters hippocampal cellular composition in both neonatal and juvenile rat pups in association with altered hippocampal gene expression and epigenetic determinants. PPARγ signaling is important for neurodevelopment as well as epigenetic integrity in the brain via the PPARγ‐Setd8‐H4K20me1 axis and Wnt signaling. We hypothesized that IUGR would decrease expression of PPARγ, Setd8, and H4K20me1 in juvenile rat hippocampus. We further hypothesized that reduced PPARγ‐Setd8‐H4K20me1 would be associated with reduced Wnt signaling genes Wnt3a and β‐catenin, and wnt target gene Axin2. To test our hypothesis we used a rat model of uteroplacental insufficiency‐induced IUGR. We demonstrated that PPARγ localizes to oligodendrocytes, neurons and astrocytes within the juvenile rat hippocampus. We also demonstrated that IUGR reduces levels of PPARγ, Setd8 and H4K20me1 in male and female juvenile rat hippocampus in conjunction with reduced Wnt signaling components in only male rats. We speculate that reduced PPARγ and Wnt signaling may contribute to altered hippocampal cellular composition which, in turn, may contribute to impaired neurodevelopment and subsequent neurocognitive impairment in IUGR offspring.

Maternal tobacco smoke increased visceral adiposity and serum corticosterone levels in adult male rat offspring

Background:Maternal tobacco smoke (MTS) predisposes human and rat offspring to visceral obesity in early adulthood. Glucocorticoid excess also causes visceral obesity. We hypothesized that in utero MTS would increase visceral adiposity and alter the glucocorticoid pathway in young adult rats.Methods:We developed a novel model of in utero MTS exposure in pregnant rats by exposing them to cigarette smoke from E11.5 to term. Neonatal rats were cross-fostered to control dams and weaned to standard rat chow through young adulthood (postnatal day 60).Results:We demonstrated increased visceral adiposity (193%)*, increased visceral adipose 11-β hydroxysteroid dehydrogenase 1 mRNA (204%)*, increased serum corticosterone (147%)*, and no change in glucocorticoid receptor protein in adult male MTS rat offspring. Female rats exposed to MTS in utero demonstrated no change in visceral or subcutaneous adiposity, decreased serum corticosterone (60%)*, and decreased adipose glucocorticoid receptor protein (66%)*. *P < 0.05.Conclusion:We conclude that in utero MTS exposure increased visceral adiposity and altered in the glucocorticoid pathway in a sex-specific manner. We speculate that in utero MTS exposure programs adipose dysfunction in adult male rat offspring via alteration in the glucocorticoid pathway.

IUGR prevents IGF-1 upregulation in juvenile male mice by perturbing postnatal IGF-1 chromatin remodeling

Background:Intrauterine growth restriction (IUGR) offspring with rapid catch-up growth are at increased risk for early obesity especially in males. Persistent insulin-like growth factor-1 (IGF-1) reduction is an important risk factor. Using a mouse model of maternal hypertension-induced IUGR, we examined IGF-1 levels, promoter DNA methylation, and histone H3 covalent modifications at birth (D1). We additionally investigated whether prenatal perturbations could reset at preadolescence (D21).Methods:IUGR was induced via maternal thromboxane A2-analog infusion in mice.Results:IUGR uniformly decreased D1 IGF-1 mRNA and protein levels with reduced promoter 1 (P1) transcription and increased P1 DNA methylation. IUGR males also had increased H3K4ac at exon 5 and 3′ distal UTR. At D21, IUGR males continued to have decreased IGF-1 levels, originating from both P1 and P2 with reduced 1A variant. IUGR males also had decreased activation mark of H3K4me3 at P1 compared with sham males. In contrast, D21 IUGR females normalized their IGF-1 levels, in association with an increased activation mark of H3K4me3 at P1 compared with sham females.Conclusion:IUGR uniformly affected D1 hepatic IGF-1 epigenetic modifications in both sexes. However, at preadolescence, IUGR males are unable to correct for the prenatal reduction possibly due to a more perturbed IGF-1 chromatin structure.

Protective effect of maternal uteroplacental insufficiency on oxygen-induced retinopathy in offspring: removing bias of premature birth

To address the hypothesis that maternal uteroplacental insufficiency (UPI) increases severity of retinopathy of prematurity, we developed a composite rat model of UPI and oxygen-fluctuations and removed premature birth as a confounding factor. Timed-pregnant Sprague-Dawley dams underwent bilateral uterine artery ligation or anesthesia (control) at e19.5. Full-term pups developed in room air (RA) or an oxygen-induced retinopathy (OIR) model. Isolectin-stained retinal flat-mounts were analyzed for percent of areas of avascular/total retina (AVA) and of intravitreal neovascular/total retina (IVNV). Pup weights and serum and mRNA of liver and kidney VEGF, IGF-1, and erythropoietin (EPO) were determined. Multivariable mixed effects linear regressions and Pearson correlations were performed using STATA14. Postnatal growth restriction occurred in pups in UPI/RA, but not in UPI/OIR. Weight gain was similar between UPI/OIR and control/OIR pups. AVA was reduced and a trend toward reduced IVNV was seen in UPI/OIR compared to control/OIR. No difference in birth weights of UPI/OIR vs. control/OIR pups occurred. Serum and renal IGF-1 and EPO were significantly increased in UPI/OIR compared to control/OIR pups. In the absence of prematurity, UPI increased angiogenic factors in association with reduced OIR severity, suggesting that ischemia from UPI could yield protective angiogenic effects by offspring.

Intrauterine growth restriction combined with a maternal high‐fat diet increases hepatic cholesterol and low‐density lipoprotein receptor activity in rats

Intrauterine growth restriction (IUGR) and maternal consumption of a high‐saturated‐fat diet (HFD) increase the risk of hypercholesterolemia, a leading cause of morbidity and mortality. Many pregnant women eat a HFD, thus exposing the fetus to a HFD in utero. The cumulative effect of in utero exposure to IUGR and a HFD on offspring cholesterol levels remains unknown. Furthermore, little is known about the mechanism through which IUGR and maternal HFD consumption increase cholesterol. We hypothesize that IUGR combined with a maternal HFD would increase offspring serum and hepatic cholesterol accumulation via alteration in levels of key proteins involved in cholesterol metabolism. To test our hypothesis we used a rat model of surgically induced IUGR and fed the dams a regular diet or a HFD. HFD‐fed dams consumed the same kilocalories as regular diet‐fed dams, with no difference between surgical intervention groups. In the offspring, IUGR combined with a maternal HFD increased hepatic cholesterol levels, low‐density lipoprotein (LDL) receptor protein levels, and Ldlr activity in female rat offspring at birth and both sexes at postnatal day 14 relative to non‐IUGR offspring both from regular diet‐ and HFD‐fed dams. These findings suggest that IUGR combined with a maternal HFD increases hepatic cholesterol accumulation via increased LDL cholesterol uptake into the liver with resulting persistent increases in hepatic cholesterol accumulation.

Prenatal Exposure to a Maternal High Fat Diet Increases Hepatic Cholesterol Accumulation in Intrauterine Growth Restricted Rats in Part Through MicroRNA-122 Inhibition of Cyp7a1

Intrauterine growth restriction (IUGR) and consumption of a high saturated fat diet (HFD) increase the risk of hypercholesterolemia, a leading cause of morbidity and mortality. The mechanism through which the cumulative impact of IUGR and in utero exposure to a maternal HFD increase cholesterol levels remains unknown. Cholesterol 7α hydroxylase (Cyp7a1) initiates catabolism of cholesterol to bile acids for elimination from the body, and is regulated by microRNA-122 (miR-122). We hypothesized that IUGR rats exposed to a maternal HFD would have increased cholesterol and decreased Cyp7a1 protein levels in juvenile rats, findings which would be normalized by administration of a miR-122 inhibitor. To test our hypothesis we used a rat model of surgically induced IUGR and fed the dams a regular diet or a HFD from prior to conception through lactation. At the time of weaning, IUGR female rats exposed to a maternal HFD had increased hepatic cholesterol, decreased hepatic Cyp7a1 protein and hepatic bile acids, and increased hepatic miR-122 compared to non-IUGR rats exposed to the same HFD. In vivo inhibition of miR-122 increased hepatic Cyp7a1 protein and decreased hepatic cholesterol. Our findings suggest that IUGR combined with a maternal HFD decreased cholesterol catabolism to bile acids, in part, via miR-122 inhibition of Cyp7a1.