Human whole genome sequencing in South Africa.

The advent and evolution of next generation sequencing has considerably impacted genomic research. Until recently, South African researchers were unable to access affordable platforms capable of human whole genome sequencing locally and DNA samples had to be exported. Here we report the whole genome sequences of the first six human DNA samples sequenced and analysed at the South African Medical Research Council's Genomics Centre. We demonstrate that the data obtained is of high quality, with an average sequencing depth of 36.41, and that the output is comparable to data generated internationally on a similar platform. The Genomics Centre creates an environment where African researchers are able to access world class facilities, increasing local capacity to sequence whole genomes as well as store and analyse the data.

Programming With Varying Dietary Fat Content Alters Cardiac Insulin Receptor, Glut4 and FoxO1 Immunoreactivity in Neonatal Rats, Whereas High Fat Programming Alters Cebpa Gene Expression in Neonatal Female Rats.

Fetal programming refers to an intrauterine stimulus or insult that shapes growth, development and health outcomes. Dependent on the quality and quantity, dietary fats can be beneficial or detrimental for the growth of the fetus and can alter insulin signaling by regulating the expression of key factors. The effects of varying dietary fat content on the expression profiles of factors in the neonatal female and male rat heart were investigated and analyzed in control (10% fat), 20F (20% fat), 30F (30% fat) and 40F (40% fat which was a high fat diet used to induce high fat programming) neonatal rats. The whole neonatal heart was immunostained for insulin receptor, glucose transporter 4 (Glut4) and forkhead box protein 1 (FoxO1), followed by image analysis. The expression of 84 genes, commonly associated with the insulin signaling pathway, were then examined in 40F female and 40F male offspring. Maintenance on diets, varying in fat content during fetal life, altered the expression of cardiac factors, with changes induced from 20% fat in female neonates, but from 30% fat in male neonates. Further, CCAAT/enhancer-binding protein alpha (Cebpa) was upregulated in 40F female neonates. There was, however, differential expression of several insulin signaling genes in 40F (high fat programmed) offspring, with some tending to significance but most differences were in fold changes (≥1.5 fold). The increased immunoreactivity for insulin receptor, Glut4 and FoxO1 in 20F female and 30F male neonatal rats may reflect a compensatory response to programming to maintain cardiac physiology. Cebpa was upregulated in female offspring maintained on a high fat diet, with fold increases in other insulin signaling genes viz. Aebp1, Cfd (adipsin), Adra1d, Prkcg, Igfbp, Retn (resistin) and Ucp1. In female offspring maintained on a high fat diet, increased Cebpa gene expression (concomitant with fold increases in other insulin signaling genes) may reflect cardiac stress and an adaptative response to cardiac inflammation, stress and/or injury, after high fat programming. Diet and the sex are determinants of cardiac physiology and pathophysiology, reflecting divergent mechanisms that are sex-specific.

The Combination Effect of Aspalathin and Phenylpyruvic Acid-2-O-ß-D-glucoside from Rooibos against Hyperglycemia-Induced Cardiac Damage: An In Vitro Study.

Recent evidence shows that rooibos compounds, aspalathin and phenylpyruvic acid-2-O-ß-D-glucoside (PPAG), can independently protect cardiomyocytes from hyperglycemia-related reactive oxygen species (ROS). While aspalathin shows more potency by enhancing intracellular antioxidant defenses, PPAG acts more as an anti-apoptotic agent. Thus, to further understand the protective capabilities of these compounds against hyperglycemia-induced cardiac damage, their combinatory effect was investigated and compared to metformin. An in vitro model of H9c2 cardiomyocytes exposed to chronic glucose concentrations was employed to study the impact of such compounds on hyperglycemia-induced damage. Here, high glucose exposure impaired myocardial substrate utilization by abnormally enhancing free fatty acid oxidation while concomitantly suppressing glucose oxidation. This was paralleled by altered expression of genes involved in energy metabolism including acetyl-CoA carboxylase (ACC), 5' AMP-activated protein kinase (AMPK), and peroxisome proliferator-activated receptor-alpha (PPARα). The combination treatment improved myocardial substrate metabolism, maintained mitochondrial membrane potential, and attenuated various markers for oxidative stress including nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and glutathione content. It also showed a much-improved effect by ameliorating DNA damage when compared to metformin. The current study demonstrates that rooibos compounds offer unique cardioprotective properties against hyperglycemia-induced and potentially against diabetes-induced cardiac damage. These data also support further exploration of rooibos compounds to better assess the cardioprotective effects of different bioactive compound combinations.

Aspalathin-Enriched Green Rooibos Extract Reduces Hepatic Insulin Resistance by Modulating PI3K/AKT and AMPK Pathways.

We previously demonstrated that an aspalathin-enriched green rooibos extract (GRE) reversed palmitate-induced insulin resistance in C2C12 skeletal muscle and 3T3-L1 fat cells by modulating key effectors of insulin signalling such as phosphatidylinositol-4,5-bisphosphate 3-kinase/protein kinase B (PI3K/AKT) and AMP-activated protein kinase (AMPK). However, the effect of GRE on hepatic insulin resistance is unknown. The effects of GRE on lipid-induced hepatic insulin resistance using palmitate-exposed C3A liver cells and obese insulin resistant (OBIR) rats were explored. GRE attenuated the palmitate-induced impairment of glucose and lipid metabolism in treated C3A cells and improved insulin sensitivity in OBIR rats. Mechanistically, GRE treatment significantly increased PI3K/AKT and AMPK phosphorylation while concurrently enhancing glucose transporter 2 expression. These findings were further supported by marked stimulation of genes involved in glucose metabolism, such as insulin receptor (Insr) and insulin receptor substrate 1 and 2 (Irs1 and Irs2), as well as those involved in lipid metabolism, including Forkhead box protein O1 (FOXO1) and carnitine palmitoyl transferase 1 (CPT1) following GRE treatment. GRE showed a strong potential to ameliorate hepatic insulin resistance by improving insulin sensitivity through the regulation of PI3K/AKT, FOXO1 and AMPK-mediated pathways.

Exchange transfusions for extreme hypertriglyceridemia in a 7-week-old infant with multi-organ failure.

Severe hypertriglyceridemia is the third most common cause of acute pancreatitis and is strongly associated with an increased risk of cardiovascular disease. In infants, the most common cause of severe hypertriglyceridemia is lipoprotein lipase deficiency. We describe a 7-week-old infant with severe hypertriglyceridemia, who presented with frequent gastrointestinal bleeding, respiratory distress, a decreased level of consciousness and lipemia retinalis. Triglycerides were reduced from 734 to 2 mmol/L (64,956-177 mg/dL), by exchange transfusions. The infant made a remarkable recovery with no sequelae. This case highlights atypical, protean presentations and a potential treatment when established therapies are unavailable.

Aspalathin Protects the Heart against Hyperglycemia-Induced Oxidative Damage by Up-Regulating Nrf2 Expression.

Aspalathin (ASP) can protect H9c2 cardiomyocytes against high glucose (HG)-induced shifts in myocardial substrate preference, oxidative stress, and apoptosis. The protective mechanism of ASP remains unknown. However, as one of possible, it is well known that phytochemical flavonoids reduce oxidative stress via nuclear factor (erythroid-derived 2)-like 2 (Nrf2) activation resulting in up-regulation of antioxidant genes and enzymes. Therefore, we hypothesized that ASP protects the myocardium against HG- and hyperglycemia-induced oxidative damage by up-regulating Nrf2 expression in H9c2 cardiomyocytes and diabetic (db/db) mice, respectively. Using an oxidative stress RT² Profiler PCR array, ASP at a dose of 1 µM was demonstrated to protect H9c2 cardiomyocytes against HG-induced oxidative stress, but silencing of Nrf2 abolished this protective response of ASP and exacerbated cardiomyocyte apoptosis. Db/db mice and their non-diabetic (db/+) littermate controls were subsequently treated daily for six weeks with either a low (13 mg/kg) or high (130 mg/kg) ASP dose. Compared to nondiabetic mice the db/db mice presented increased cardiac remodeling and enlarged left ventricular wall that occurred concomitant to enhanced oxidative stress. Daily treatment of mice with ASP at a dose of 130 mg/kg for six weeks was more effective at reversing complications than both a low dose ASP or metformin, eliciting enhanced expression of Nrf2 and its downstream antioxidant genes. These results indicate that ASP maintains cellular homeostasis and protects the myocardium against hyperglycemia-induced oxidative stress through activation of Nrf2 and its downstream target genes.

Expression of UCP2 in Wistar rats varies according to age and the severity of obesity

Obesity, a complex metabolic disorder, is characterized by mitochondrial dysfunction and oxidative stress. Increased expression of uncoupling protein 2 (UCP2) during obesity is an adaptive response to suppress the production of reactive oxygen species. The aims of this study were to compare the expression of UCP2 in diet-induced obese Wistar rats that differed according to age and their severity of obesity, and to compare UCP2 expression in the liver and muscle of these rats. UCP2 messenger RNA and protein expression was increased 4.6-fold (p < 0.0001) and 3.0-fold (p < 0.05), respectively, in the liver of the older and heavier rats. In contrast, UCP2 expression was decreased twofold (p < 0.005) in the muscle of these rats, while UCP3 messenger RNA (mRNA) was increased twofold (p < 0.01). Peroxisome proliferator-activated receptor alpha (PPARα) was similarly increased (3.0-fold, p < 0.05) in the liver of the older and more severe obese rats. Total protein content was increased (2.3-fold, p < 0.0001), while 5' adenosine monophosphate-activated protein kinase (AMPK) activity was decreased (1.3-fold, p = 0.05) in the liver of the older, heavier rats. No difference in total protein content and AMPK expression was observed in the muscle of these rats. This study showed that the expression of UCP2 varies according to age and the severity of obesity and supports the widely held notion that increased UCP2 expression is an adaptive response to increased fatty acid β-oxidation and reactive oxygen species production that occurs during obesity. An understanding of metabolic adaptation is imperative to gain insight into the underlying causes of disease, thus facilitating intervention strategies to combat disease progression (AU)

No disponible

Aspalathin, a dihydrochalcone C-glucoside, protects H9c2 cardiomyocytes against high glucose induced shifts in substrate preference and apoptosis.

SCOPE: Energy deprivation in the myocardium is associated with impaired heart function. This study aims to investigate if aspalathin (ASP) can ameliorate hyperglycemic-induced shift in substrate preference and protect the myocardium against cell apoptosis. METHODS AND RESULTS: H9c2 cells were exposed to, either normal (5.5 mM) or high (33 mM) glucose concentrations for 48 h. Thereafter, cells exposed to 33 mM glucose were treated with metformin (1 µM) or ASP (1 µM), as well as a combination of metformin and ASP for 6 h. In vitro studies revealed that ASP improved glucose metabolism by decreasing fatty acid uptake and subsequent ß-oxidation through the decreased expression of adenosine monophosphate-activated protein kinase threonine 172 (pAMPK (Thr172)) and carnitine palmitoyltransferase 1 (CPT1), while increasing acetyl-CoA carboxylase (ACC) and glucose transporter 4 (GLUT4) expression. ASP inhibited high glucose induced loss of membrane potential in H9c2 cells as observed by an increase in 5' ,6,6'-tetrachloro-1,1',3,3' -tetraethylbenzimidazolyl-carbocyanine iodide (JC-1) ratio (orange\red fluorescence) and decreased apoptosis by reducing intracellular reactive oxygen species and DNA nick formation, while increasing glutathione, superoxide dismutase, uncoupling protein 2 (UCP2), and Bcl-2\Bax ratio. CONCLUSION: Our study provides evidence that ASP increases glucose oxidation and modulates fatty acid utilization producing a favorable substrate shift in H9c2 cardiomyocytes exposed to high glucose. Such a favorable shift will be of importance in the protection of cardiomyocytes in the diabetic heart.

Expression of UCP2 in Wistar rats varies according to age and the severity of obesity.

Obesity, a complex metabolic disorder, is characterized by mitochondrial dysfunction and oxidative stress. Increased expression of uncoupling protein 2 (UCP2) during obesity is an adaptive response to suppress the production of reactive oxygen species. The aims of this study were to compare the expression of UCP2 in diet-induced obese Wistar rats that differed according to age and their severity of obesity, and to compare UCP2 expression in the liver and muscle of these rats. UCP2 messenger RNA and protein expression was increased 4.6-fold (p < 0.0001) and 3.0-fold (p < 0.05), respectively, in the liver of the older and heavier rats. In contrast, UCP2 expression was decreased twofold (p < 0.005) in the muscle of these rats, while UCP3 messenger RNA (mRNA) was increased twofold (p < 0.01). Peroxisome proliferator-activated receptor alpha (PPARα) was similarly increased (3.0-fold, p < 0.05) in the liver of the older and more severe obese rats. Total protein content was increased (2.3-fold, p < 0.0001), while 5' adenosine monophosphate-activated protein kinase (AMPK) activity was decreased (1.3-fold, p = 0.05) in the liver of the older, heavier rats. No difference in total protein content and AMPK expression was observed in the muscle of these rats. This study showed that the expression of UCP2 varies according to age and the severity of obesity and supports the widely held notion that increased UCP2 expression is an adaptive response to increased fatty acid ß-oxidation and reactive oxygen species production that occurs during obesity. An understanding of metabolic adaptation is imperative to gain insight into the underlying causes of disease, thus facilitating intervention strategies to combat disease progression.

Z-2-(ß-D-glucopyranosyloxy)-3-phenylpropenoic acid, an α-hydroxy acid from rooibos (Aspalathus linearis) with hypoglycemic activity.

SCOPE: The rare enolic phenylpyruvic acid-2-O-glucoside, (PPAG:Z-2-(ß-D-glucopyranosyloxy)-3-phenylpropenoic acid), is one of the major constituents of fermented rooibos infusions. 3-Phenylpyruvic acid (2-oxo-3-phenylpropanoic acid), without the sugar moiety and with a keto form instead of an enolic arrangement, has been shown to enhance insulin release and glucose uptake in muscle cells. The purpose of this study was to assess if PPAG has similar activity on glucose metabolism. METHODS AND RESULTS: Preliminary in vitro studies confirmed that PPAG, isolated from rooibos, enhanced glucose uptake. A dose-response study in Chang cells showed that PPAG enhanced glucose uptake in the concentration range 1.0-31.6 µM (EC50 = 3.6 µM). In obese insulin-resistant rats, oral administration of PPAG lowered fasting glucose concentrations and improved oral glucose tolerance values; messenger RNA expression of glucokinase, glucose transporter 1 and 2, insulin receptor, peroxisome proliferator-activated receptor alpha, and suppressor of cytokine signaling 3, were increased in the liver. This suggests that the liver is mainly responsible for PPAG bioactivity. CONCLUSION: This study describes for the first time that PPAG increases in vitro glucose uptake and improves glucose tolerance in an obese insulin-resistant rat model, suggesting that it has potential as a new class of antidiabetic therapeutics that would contribute to the antidiabetic effect of rooibos.