Loss of NRF-2 and PGC-1α genes leads to retinal pigment epithelium damage resembling dry age-related macular degeneration

Age-related macular degeneration (AMD) is a multi-factorial disease that is the leading cause of irreversible and severe vision loss in the developed countries. It has been suggested that the pathogenesis of dry AMD involves impaired protein degradation in retinal pigment epithelial cells (RPE). RPE cells are constantly exposed to oxidative stress that may lead to the accumulation of damaged cellular proteins, DNA and lipids and evoke tissue deterioration during the aging process. The ubiquitin-proteasome pathway and the lysosomal/autophagosomal pathway are the two major proteolytic systems in eukaryotic cells. NRF-2 (nuclear factor-erythroid 2-related factor-2) and PGC-1 alpha (peroxisome proliferator-activated receptor gamma coactivator-1 alpha) are master transcription factors in the regulation of cellular detoxification. We investigated the role of NRF-2 and PGC-1 alpha in the regulation of RPE cell structure and function by using global double knockout (dKO) mice. The NRF-2/PGC-1 alpha dKO mice exhibited significant age-dependent RPE degeneration, accumulation of the oxidative stress marker, 4-HNE (4-hydroxynonenal), the endoplasmic reticulum stress markers GRP78 (glucose-regulated protein 78) and ATF4 (activating transcription factor 4), and damaged mitochondria. Moreover, levels of protein ubiquitination and autophagy markers p62/SQSTM1 (sequestosome 1), Beclin-1 and LC3B (microtubule associated protein 1 light chain 3 beta) were significantly increased together with the Iba-1 (ionized calcium binding adaptor molecule 1) mononuclear phagocyte marker and an enlargement of RPE size. These histopathological changes of RPE were accompanied by photoreceptor dysmorphology and vision loss as revealed by electroretinography. Consequently, these novel findings suggest that the NRF-2/PGC-1 alpha dKO mouse is a valuable model for investigating the role of proteasomal and autophagy clearance in the RPE and in the development of dry AMD.Peer reviewe

PGC-1α4 Interacts with REST to Upregulate Neuronal Genes and Augment Energy Consumption in Developing Cardiomyocytes

Transcriptional coactivator PGC-1α is a main regulator of cardiac energy metabolism. In addition to canonical PGC-1α1, other PGC-1α isoforms have been found to exert specific biological functions in a variety of tissues. We investigated the expression patterns and the biological effects of the non-canonical isoforms in the heart. We used RNA sequencing data to identify the expression patterns of PGC-1α isoforms in the heart. To evaluate the biological effects of the alternative isoform expression, we generated a transgenic mouse with cardiac-specific overexpression of PGC-1α4 and analysed the cardiac phenotype with a wide spectrum of physiological and biophysical tools. Our results show that non-canonical isoforms are expressed in the heart, and that the main variant PGC-1α4 is induced by β-adrenergic signalling in adult cardiomyocytes. Cardiomyocyte specific PGC-1α4 overexpression in mice relieves the RE1-Silencing Transcription factor (REST)-mediated suppression of neuronal genes during foetal heart development. The resulting de-repression of REST target genes induces a cardiac phenotype with increased cellular energy consumption, resulting in postnatal dilated cardiomyopathy. These results propose a new concept for actions of the PGC-1α protein family where activation of the Pgc-1α gene, through its isoforms, induces a phenotype with concurrent supply and demand for cellular energy. These data highlight the biological roles of the different PGC-1α isoforms, which should be considered when future therapies are developed

The effect of diet on fatty acid composition of elongated bovine conceptuses

AbstraktiConceptus elongation is crucial for establishment of pregnancy in ruminants. During the elongation step, endometrium secreted lipids are required as a source of energy and for the remodeling and proliferation of cellular membranes (Ribeiro et al., Reprod 2016;152:R115-R126). The present study investigated the effects of diets designed to cause 15 % reduction in milk fat synthesis (milk fat depression) on fatty acid (FA) composition of elongated bovine conceptuses. The day-14 conceptuses were collected from 22 Nordic Red multiparous dairy cows during two sequential indoor housing periods. Treatments comprised a grass silage based basal diet (CO; 10 cows), basal diet supplemented with 12 g/d of rumen protected trans-10, cis-12 CLA (Lutrell Pure, BASF, Germany) (CLA; 8 cows), and grass silage based diet containing high-starch concentrate components and supplemented with 26.7 g/kg diet dry matter of sunflower oil and 13.3 g/kg diet dry matter of fish oil (MFD; 4 cows). CLA supplement was mixed in total mixed ration and administered in two equal proportions per day. Oil supplements replaced concentrate ingredients. Cows were randomly allocated to the treatments immediately after parturitions. The diets were total mixed rations with 55:45 forage:concentrate ratio on dry matter basis. In total of 45, 35 and 13 conceptuses having visible embryonic discs were used after recovery from the superovulated CO, CLA and MFD donors 130 days after parturitions, respectively. Lipids were extracted separately from each cryopreserved conceptus using a mixture of hexane and 2-propanol (3:2, vol/vol). FA were transesterified to methyl esters using methanolic sodium methoxide and analyzed with a gas chromatograph equipped with a flame ionization detector (Shingfield et al., Anim Sci 2003;77:165–179). Data were analyzed using linear mixed models with MIXED procedure in SAS 9.4. The proportions of the most abundant FA in conceptuses, cis-9 18:1 (30-32 g/100g FA, %), 16:0 (25-26 %), and 18:0 (12-14 %), did not differ between treatments (P > 0.10). In addition, total proportions of cis unsaturated FA (53-54 %) and saturated FA (43-45 %) and ratio of cis unsaturated FA to saturated FA was not different among treatments (P > 0.10). CLA had no effect on the conceptus’ FA composition compared with CO. However, MFD induced higher (P < 0.01) 22:6n-3, and lower (P < 0.05) 22:4n-6 and 22:5n-6 proportions compared with CLA and CO. The proportion of cis-12 18:1 and trans-9, cis-12 18:2, which are biohydrogenation products of 18:2n-6 found in rich amounts in sunflower oil, tended to be higher (P < 0.10) in MFD compared with other treatments. In conclusion, although CLA and MDF caused changes in lipogenesis in the mammary gland and milk fat depression, a substantial effect on the FA composition of conceptuses was not observed. However, unsaturated FA deriving from the MFD diet and metabolized in the rumen biohydrogenation processes had a specific impact on conceptus FA profiles during elongation stage.201

Obesity represses CYP2R1, the vitamin D 25‐hydroxylase, in the liver and extrahepatic tissues

Abstract Low plasma level of 25‐hydroxyvitamin D (25‐OH‐D), namely vitamin D deficiency, is associated with obesity and weight loss improves 25‐OH‐D status. However, the mechanism behind obesity‐induced vitamin D deficiency remains unclear. Here, we report that obesity suppresses the expression of cytochrome P450 (CYP) 2R1, the main vitamin D 25‐hydroxylase, in both mice and humans. In humans, weight loss induced by gastric bypass surgery increased the expression of CYP2R1 in the s.c. adipose tissue suggesting recovery after the obesity‐induced suppression. At the same time, CYP27B1, the vitamin D 1α‐hydroxylase, was repressed by the weight loss. In a mouse (C57BL/6N) model of diet‐induced obesity, the plasma 25‐OH‐D was decreased. In accordance, the CYP2R1 expression was strongly repressed in the liver. Moreover, obesity repressed the expression of CYP2R1 in several extrahepatic tissues, the kidney, brown adipose tissue, and testis, but not in the white adipose tissue. Obesity had a similar effect in both male and female mice. In mice, obesity repressed expression of the vitamin D receptor in brown adipose tissue. Obesity also upregulated the expression of the vitamin D receptor and CYP24A1 in the s.c. adipose tissue of a subset of mice; however, no effect was observed in the human s.c. adipose tissue. In summary, we show that obesity affects CYP2R1 expression both in the mouse and human tissues. We suggest that in mouse the CYP2R1 repression in the liver plays an important role in obesity‐induced vitamin D deficiency. Currently, it is unclear whether the CYP2R1 downregulation in extrahepatic tissues could contribute to the obesity‐induced low plasma 25‐OH‐D, however, this phenomenon may affect at least the local 25‐OH‐D concentrations

Fasting-Induced Transcription Factors Repress Vitamin D Bioactivation, a Mechanism for Vitamin D Deficiency in Diabetes

Low 25-hydroxyvitamin D levels correlate with the prevalence of diabetes; however, the mechanisms remain uncertain. Here, we show that nutritional deprivation-responsive mechanisms regulate vitamin D metabolism. Both fasting and diabetes suppressed hepatic cytochrome P450 (CYP) 2R1, the main vitamin D 25-hydroxylase responsible for the first bioactivation step. Overexpression of coactivator peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1 alpha), induced physiologically by fasting and pathologically in diabetes, resulted in dramatic downregulation of CYP2R1 in mouse hepatocytes in an estrogen-related receptor alpha (ERR alpha)-dependent manner. However, PGC-1 alpha knockout did not prevent fasting-induced suppression of CYP2R1 in the liver, indicating that additional factors contribute to the CYP2R1 repression. Furthermore, glucocorticoid receptor (GR) activation repressed the liver CYP2R1, suggesting GR involvement in the regulation of CYP2R1. GR antagonist mifepristone partially prevented CYP2R1 repression during fasting, suggesting that glucocorticoids and GR contribute to the CYP2R1 repression during fasting. Moreover, fasting upregulated the vitamin D catabolizing CYP24A1 in the kidney through the PGC-1 alpha-ERR alpha pathway. Our study uncovers a molecular mechanism for vitamin D deficiency in diabetes and reveals a novel negative feedback mechanism that controls crosstalk between energy homeostasis and the vitamin D pathway

Fasting-induced transcription factors repress vitamin D bioactivation, a mechanism for vitamin D deficiency in diabetes

Abstract Low 25-hydroxyvitamin D levels correlate with the prevalence of diabetes; however, the mechanisms remain uncertain. Here, we show that nutritional deprivation–responsive mechanisms regulate vitamin D metabolism. Both fasting and diabetes suppressed hepatic cytochrome P450 (CYP) 2R1, the main vitamin D 25-hydroxylase responsible for the first bioactivation step. Overexpression of coactivator peroxisome proliferator–activated receptor γ coactivator 1-α (PGC-1α), induced physiologically by fasting and pathologically in diabetes, resulted in dramatic downregulation of CYP2R1 in mouse hepatocytes in an estrogen-related receptor α (ERRα)–dependent manner. However, PGC-1α knockout did not prevent fasting-induced suppression of CYP2R1 in the liver, indicating that additional factors contribute to the CYP2R1 repression. Furthermore, glucocorticoid receptor (GR) activation repressed the liver CYP2R1, suggesting GR involvement in the regulation of CYP2R1. GR antagonist mifepristone partially prevented CYP2R1 repression during fasting, suggesting that glucocorticoids and GR contribute to the CYP2R1 repression during fasting. Moreover, fasting upregulated the vitamin D catabolizing CYP24A1 in the kidney through the PGC-1α-ERRα pathway. Our study uncovers a molecular mechanism for vitamin D deficiency in diabetes and reveals a novel negative feedback mechanism that controls crosstalk between energy homeostasis and the vitamin D pathway

Loss of NRF-2 and PGC-1α genes leads to retinal pigment epithelium damage resembling dry age-related macular degeneration

Age-related macular degeneration (AMD) is a multi-factorial disease that is the leading cause of irreversible and severe vision loss in the developed countries. It has been suggested that the pathogenesis of dry AMD involves impaired protein degradation in retinal pigment epithelial cells (RPE). RPE cells are constantly exposed to oxidative stress that may lead to the accumulation of damaged cellular proteins, DNA and lipids and evoke tissue deterioration during the aging process. The ubiquitin-proteasome pathway and the lysosomal/autophagosomal pathway are the two major proteolytic systems in eukaryotic cells. NRF-2 (nuclear factor-erythroid 2-related factor-2) and PGC-1α (peroxisome proliferator-activated receptor gamma coactivator-1 alpha) are master transcription factors in the regulation of cellular detoxification. We investigated the role of NRF-2 and PGC-1α in the regulation of RPE cell structure and function by using global double knockout (dKO) mice. The NRF-2/PGC-1α dKO mice exhibited significant age-dependent RPE degeneration, accumulation of the oxidative stress marker, 4-HNE (4-hydroxynonenal), the endoplasmic reticulum stress markers GRP78 (glucose-regulated protein 78) and ATF4 (activating transcription factor 4), and damaged mitochondria. Moreover, levels of protein ubiquitination and autophagy markers p62/SQSTM1 (sequestosome 1), Beclin-1 and LC3B (microtubule associated protein 1 light chain 3 beta) were significantly increased together with the Iba-1 (ionized calcium binding adaptor molecule 1) mononuclear phagocyte marker and an enlargement of RPE size. These histopathological changes of RPE were accompanied by photoreceptor dysmorphology and vision loss as revealed by electroretinography. Consequently, these novel findings suggest that the NRF-2/PGC-1α dKO mouse is a valuable model for investigating the role of proteasomal and autophagy clearance in the RPE and in the development of dry AMD