Differential expression of Lp-PLA2 in obesity and type 2 diabetes and the influence of lipids

Aims/hypothesis Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a circulatory macrophage-derived factor that increases with obesity and leads to a higher risk of cardiovascular disease (CVD). Despite this, its role in adipose tissue and the adipocyte is unknown. Therefore, the aims of this study were to clarify the expression of Lp-PLA2 in relation to different adipose tissue depots and type 2 diabetes, and ascertain whether markers of obesity and type 2 diabetes correlate with circulating Lp-PLA2. A final aim was to evaluate the effect of cholesterol on cellular Lp-PLA2 in an in vitro adipocyte model. Methods Analysis of anthropometric and biochemical variables from a cohort of lean (age 44.4 ± 6.2 years; BMI 22.15 ± 1.8 kg/m2, n = 23), overweight (age 45.4 ± 12.3 years; BMI 26.99 ± 1.5 kg/m2, n = 24), obese (age 49.0 ± 9.1 years; BMI 33.74 ± 3.3 kg/m2, n = 32) and type 2 diabetic women (age 53.0 ± 6.13 years; BMI 35.08 ± 8.6 kg/m2, n = 35), as part of an ethically approved study. Gene and protein expression of PLA2 and its isoforms were assessed in adipose tissue samples, with serum analysis undertaken to assess circulating Lp-PLA2 and its association with cardiometabolic risk markers. A human adipocyte cell model, Chub-S7, was used to address the intracellular change in Lp-PLA2 in adipocytes. Results Lp-PLA2 and calcium-independent PLA2 (iPLA2) isoforms were altered by adiposity, as shown by microarray analysis (p < 0.05). Type 2 diabetes status was also observed to significantly alter gene and protein levels of Lp-PLA2 in abdominal subcutaneous (AbdSc) (p < 0.01), but not omental, adipose tissue. Furthermore, multivariate stepwise regression analysis of circulating Lp-PLA2 and metabolic markers revealed that the greatest predictor of Lp-PLA2 in non-diabetic individuals was LDL-cholesterol (p = 0.004). Additionally, in people with type 2 diabetes, oxidised LDL (oxLDL), triacylglycerols and HDL-cholesterol appeared important predictors, accounting for 59.7% of the variance (p < 0.001). Subsequent in vitro studies determined human adipocytes to be a source of Lp-PLA2, as confirmed by mRNA expression, protein levels and immunochemistry. Further in vitro experiments revealed that treatment with LDL-cholesterol or oxLDL resulted in significant upregulation of Lp-PLA2, while inhibition of Lp-PLA2 reduced oxLDL production by 19.8% (p < 0.05). Conclusions/interpretation Our study suggests adipose tissue and adipocytes are active sources of Lp-PLA2, with differential regulation by fat depot and metabolic state. Moreover, levels of circulating Lp-PLA2 appear to be influenced by unfavourable lipid profiles in type 2 diabetes, which may occur in part through regulation of LDL-cholesterol and oxLDL metabolism in adipocytes

Molecular mechanisms of adipose tissue mitochondrial (mal)adaptation in obesit

Obesity is one of the most serious public health challenges of the 21st century, associated with a wide range of debilitating and life-threatening conditions. The continuing increase in its prevalence underscores the urgent need for novel therapeutic targets, leads and strategies in the treatment of obesity. A metabolic imbalance of nutrient signal input results in adipose tissue dysfunction, and at the centre of this challenging environment lie mitochondria. While there have been studies into the functional changes of mitochondria in obesity, the underlying cellular and molecular mechanisms underlying this phenomenon are mostly unknown, especially in human adipose tissue. As such, this thesis set out to elucidate the impact of obesity on mitochondrial form and function in human adipocytes and adipose tissue. Firstly, gene expression analysis of a number of mitochondrial proteins was carried out in a large cohort of women with di↵ering levels of adiposity (lean, overweight and obese). These findings identified that the onset of obesity is accompanied by alterations of mitochondrial gene expression, worsening with weight gain, contributing to a more indepth understanding of adipose tissue mitochondria. This was revealed by alterations in genes encoding key markers of mitochondrial functions, including oxidative phosphorylation (1.2-fold # COX4I1 ), mitochondria dynamics (1.23-fold " FIS1 and 1.21-fold # MFN2)), and oxidative stress (1.29-fold " SOD2), in response to weight gain. Give the outcomes of these investigations, the molecular basis of mitochondrial maladaptation was explored by investigating the impact of endoplasmic reticulum (ER) stress on mitochondrial function. Based upon these studies, ER stress was identified as a possible attributer to mitochondrial dysfunction in obesity, as demonstrated by altered respiratory function which corresponded with diminished mitochondrial efficiency (1.43-fold #), impaired mitochondrial membrane potential (1.32-fold #) and a fragmented mitochondrial network. Subsequently, GPR120 agonism was probed as a promising therapeutic avenue to relieve the stress of mitochondria during prolonged overfeeding, though these studies did not conclusively determine this receptor to be a potential future target for treating mitochondrial function in white adipocytes. Finally, SGBS spheroids were identified as an exciting new 3D model for future studies into metabolic disease in human adipocytes. Collectively the findings presented in this thesis highlight the close relationship between obesity and mitochondrial dysfunction, which is in part precipitated by ER stress

An Overview of PARP Inhibitors for the Treatment of Breast Cancer

Loss-of-function mutations in BRCA1 and BRCA2 are detected in at least 5% of unselected patients with breast cancer (BC). These BC susceptibility genes encode proteins critical for DNA homologous recombination repair (HRR). This review provides an update on oral poly(ADP-ribose) polymerase (PARP) inhibitors for the treatment of BC. Olaparib and talazoparib are PARP inhibitors approved as monotherapies for deleterious/suspected deleterious germline BRCA-mutated, HER2-negative BC. Olaparib is approved in the USA for metastatic BC and in Europe for locally advanced/metastatic BC. Talazoparib is approved for locally advanced/metastatic BC in the USA and Europe. In phase 3 trials, olaparib and talazoparib monotherapies demonstrated significant progression-free survival benefits compared with chemotherapy. Common toxicities were effectively managed by supportive treatment and dose interruptions/reductions. Veliparib combined with platinum-based chemotherapy has also shown promise for locally advanced/metastatic BC in a phase 3 trial. Differences in efficacy and safety across PARP inhibitors (olaparib, talazoparib, veliparib, niraparib, rucaparib) may relate to differences in potency of PARP trapping on DNA and cytotoxic specificity. PARP inhibitors are being investigated in early BC, in novel combinations, and in patients without germline BRCA mutations, including those with somatic BRCA mutations and other HRR gene mutations. Ongoing phase 2/3 studies include PARP inhibitors combined with immune checkpoint inhibitors for the treatment of triple-negative BC. Wider access to testing for BRCA and other mutations, and to genetic counseling, are required to identify patients who could benefit from PARP inhibitor therapy. The advent of PARP inhibitors has potential benefits for BC treatment beyond the locally advanced/metastatic setting

A simplified and defined serum-free medium for cultivating fat across species

Cultivated meat is a promising technology with the potential to mitigate the ethical and environmental issues associated with traditional meat. Fat plays a key role in the meat flavor; therefore, development of suitable adipogenic protocols for livestock is essential. The traditional adipogenic cocktail containing IBMX, dexamethasone, insulin and rosiglitazone is not food-compatible. Here, we demonstrate that of the four inducers only insulin and rosiglitazone are necessary in both serum-free (DMAD) and serum-containing media, with DMAD outperforming FBS. Two glucocorticoid receptor activators, progesterone and hydrocortisone, found in DMAD and FBS, affect differentiation homogeneity, without playing an essential role in activating adipogenic genes. Importantly, this protocol leads to mature adipocytes in 3D culture. This was demonstrated in both media types and in four species: ruminant and monogastric. We therefore propose a simplified one-step adipogenic protocol which, given the replacement of rosiglitazone by a food-compatible PPARγ agonist, is suitable for making cultivated fat

A simplified and defined serum-free medium for cultivating fat across species

Summary: Cultivated meat is a promising technology with the potential to mitigate the ethical and environmental issues associated with traditional meat. Fat plays a key role in the meat flavor; therefore, development of suitable adipogenic protocols for livestock is essential. The traditional adipogenic cocktail containing IBMX, dexamethasone, insulin and rosiglitazone is not food-compatible. Here, we demonstrate that of the four inducers only insulin and rosiglitazone are necessary in both serum-free (DMAD) and serum-containing media, with DMAD outperforming FBS. Two glucocorticoid receptor activators, progesterone and hydrocortisone, found in DMAD and FBS, affect differentiation homogeneity, without playing an essential role in activating adipogenic genes. Importantly, this protocol leads to mature adipocytes in 3D culture. This was demonstrated in both media types and in four species: ruminant and monogastric. We therefore propose a simplified one-step adipogenic protocol which, given the replacement of rosiglitazone by a food-compatible PPARγ agonist, is suitable for making cultivated fat

UAS-Based Hyperspectral Environmental Monitoring of Acid Mine Drainage Affected Waters

The exposure of metal sulfides to air or water, either produced naturally or due to mining activities, can result in environmentally damaging acid mine drainage (AMD). This needs to be accurately monitored and remediated. In this study, we apply high-resolution unmanned aerial system (UAS)-based hyperspectral mapping tools to provide a useful, fast, and non-invasive method for the monitoring aspect. Specifically, we propose a machine learning framework to integrate visible to near-infrared (VNIR) hyperspectral data with physicochemical field data from water and sediments, together with laboratory analyses to precisely map the extent of acid mine drainage in the Tintillo River (Spain). This river collects the drainage from the western part of the Rio Tinto massive sulfide deposit and discharges large quantities of acidic water with significant amounts of dissolved metals (Fe, Al, Cu, Zn, amongst others) into the Odiel River. At the confluence of these rivers, different geochemical and mineralogical processes occur due to the interaction of very acidic water (pH 2.5–3.0) with neutral water (pH 7.0–8.0). This complexity makes the area an ideal test site for the application of hyperspectral mapping to characterize both rivers and better evaluate contaminated water bodies with remote sensing imagery. Our approach makes use of a supervised random forest (RF) regression for the extended mapping of water properties, using the samples collected in the field as ground-truth and training data. The resulting maps successfully estimate the concentration of dissolved metals and related physicochemical properties in water, and trace associated iron species (e.g., jarosite, goethite) within sediments. These results highlight the capabilities of UAS-based hyperspectral data to monitor water bodies in mining environments, by mapping their hydrogeochemical properties, using few field samples. Hence, we have demonstrated that our workflow allows the rapid discrimination and mapping of AMD contamination in water, providing an essential basis for monitoring and subsequent remediation

Protocol for differentiation of bovine adipogenic progenitor cells embedded in alginate sheets

Summary: Here, we present a cost-effective protocol to differentiate bovine fibro-adipogenic progenitors in a thin hydrogel sheet adherent to 96-well plates. We describe steps for the embedding and culturing of cells in alginate sheets, culture maintenance, and analysis. Compared to alternative three-dimensional (3D) models such as hydrogel-based microfibers, this approach simplifies automation while retaining efficient maturation of adipocytes. Embedded cells are still subjected to a 3D environment, but the sheets can be handled and analyzed like two-dimensional cultures. : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics