👤 S Maxwell

Endothelial to mesenchymal transition (EndMT), the transformation of endothelial cells into a mesenchymal-like state, is regulated by various factors, including transcription factors such as activator protein 1 (AP-1). While recent studies have confirmed the role of EndMT in atherosclerosis, the involvement of AP-1 in EndMT, particularly in the context of human diabetes, remains unclear. This study aimed to elucidate the role of the AP-1 transcription factor complex in EndMT associated with atherosclerosis in diabetes, utilising both an in vivo preclinical model and an ex vivo model using patient-derived serum for translational relevance. Additionally, it sought to profile gene expression changes following AP-1 inhibition in an EndMT model under high glucose conditions. Serum from patients with and without type 2 diabetes mellitus (T2DM) was used to assess EndMT in primary human aortic endothelial cells (HAECs) in the presence and absence of the AP-1 inhibitor T-5224. EndMT was evaluated through immunofluorescent staining of these cells and of aortic sections from a murine model of diabetes-associated atherosclerosis in a preclinical early intervention study. Furthermore, HAECs were used to explore the effects of AP-1 inhibition on the transcriptional signature of EndMT. Patient-derived serum induced EndMT in HAECs, which T-5224 effectively prevented, as confirmed by immunofluorescent staining. Immunofluorescent analysis of the aortic sinus also revealed that T-5224 treatment inhibited EndMT, leading to reduced atherosclerosis in Apoe This study identifies AP-1 inhibition with T-5224 as a potential therapeutic approach for EndMT resulting in reduced atherosclerosis in diabetes. The use of human serum underscores the translational relevance of these findings. Show less

Chronic kidney disease (CKD) is a complex disorder that has become a high prevalence global health problem, with diabetes being its predominant pathophysiologic driver. Autosomal genetic variation only explains some of the predisposition to kidney disease. Variations in the mitochondrial genome (mtDNA) and nuclear-encoded mitochondrial genes (NEMG) are implicated in susceptibility to kidney disease and CKD progression, but they have not been thoroughly explored. Our aim was to investigate the association of variation in both mtDNA and NEMG with CKD (and related traits), with a particular focus on diabetes. We used the UK Biobank (UKB) and UK-ROI, an independent collection of individuals with type 1 diabetes mellitus (T1DM) patients. Fourteen mitochondrial variants were associated with estimated glomerular filtration rate (eGFR) in UKB. Mitochondrial variants and haplogroups U, H and J were associated with eGFR and serum variables. Mitochondrial haplogroup H was associated with all the serum variables regardless of the presence of diabetes. Mitochondrial haplogroup X was associated with end-stage kidney disease (ESKD) in UKB. We confirmed the influence of several known NEMG on kidney disease and function and found novel associations for SLC39A13, CFL1, ACP2 or ATP5G1 with serum variables and kidney damage, and for SLC4A1, NUP210 and MYH14 with ESKD. The G allele of TBC1D32-rs113987180 was associated with higher risk of ESKD in patients with diabetes (OR:9.879; CI We identified novel variants both in mtDNA and NEMG which may explain some of the missing heritability for CKD and kidney phenotypes. We confirmed the role of MT-ND5 and mitochondrial haplogroup H on renal disease (serum variables), and identified the MT-ND5-rs41535848G variant, along with mitochondrial haplogroup X, associated with higher risk of ESKD. Despite most of the associations were independent of diabetes, we also showed potential roles for NEMG in T1DM. Show less

A number of epidemiological and genetic studies have attempted to determine whether levels of circulating lipids are associated with risks of various cancers, including breast cancer (BC). However, it remains unclear whether a causal relationship exists between lipids and BC. If alteration of lipid levels also reduced risk of BC, this could present a target for disease prevention. This study aimed to assess a potential causal relationship between genetic variants associated with plasma lipid traits (high-density lipoprotein, HDL; low-density lipoprotein, LDL; triglycerides, TGs) with risk for BC using Mendelian randomization (MR). Data from genome-wide association studies in up to 215,551 participants from the Million Veteran Program (MVP) were used to construct genetic instruments for plasma lipid traits. The effect of these instruments on BC risk was evaluated using genetic data from the BCAC (Breast Cancer Association Consortium) based on 122,977 BC cases and 105,974 controls. Using MR, we observed that a 1-standard-deviation genetically determined increase in HDL levels is associated with an increased risk for all BCs (HDL: OR [odds ratio] = 1.08, 95% confidence interval [CI] = 1.04-1.13, P < 0.001). Multivariable MR analysis, which adjusted for the effects of LDL, TGs, body mass index (BMI), and age at menarche, corroborated this observation for HDL (OR = 1.06, 95% CI = 1.03-1.10, P = 4.9 × 10-4) and also found a relationship between LDL and BC risk (OR = 1.03, 95% CI = 1.01-1.07, P = 0.02). We did not observe a difference in these relationships when stratified by breast tumor estrogen receptor (ER) status. We repeated this analysis using genetic variants independent of the leading association at core HDL pathway genes and found that these variants were also associated with risk for BCs (OR = 1.11, 95% CI = 1.06-1.16, P = 1.5 × 10-6), including locus-specific associations at ABCA1 (ATP Binding Cassette Subfamily A Member 1), APOE-APOC1-APOC4-APOC2 (Apolipoproteins E, C1, C4, and C2), and CETP (Cholesteryl Ester Transfer Protein). In addition, we found evidence that genetic variation at the ABO locus is associated with both lipid levels and BC. Through multiple statistical approaches, we minimized and tested for the confounding effects of pleiotropy and population stratification on our analysis; however, the possible existence of residual pleiotropy and stratification remains a limitation of this study. We observed that genetically elevated plasma HDL and LDL levels appear to be associated with increased BC risk. Future studies are required to understand the mechanism underlying this putative causal relationship, with the goal of developing potential therapeutic strategies aimed at altering the cholesterol-mediated effect on BC risk. Show less

Mild islet inflammation has been suggested as a contributing factor to beta cell failure in type 2 diabetes. Macrophage levels are elevated in the islets of humans and mice with type 2 diabetes, but their effects on beta cells are not understood. Our goal was to examine the gene expression changes in islet-associated macrophages in obesity models with opposing disposition to diabetes development and to assess their potential contribution to beta cell (mal)adaptation. Islets were isolated from lean control mice, obese diabetes-prone db/db mice and obese diabetes-resistant ob/ob mice. Macrophages were sorted using flow cytometry. Islets were treated ex vivo with clodronate-containing liposomes to deplete macrophages. Gene expression was assessed by real-time RT-PCR. Macrophage levels were increased in islets from db/db mice but not in islets from ob/ob mice compared with lean control mice. Macrophages from db/db and ob/ob islets displayed distinct changes in gene expression compared with control islet macrophages, suggesting differential shifts in functional state. Macrophages from db/db islets displayed increased expression of interferon regulatory factor 5 (Irf5), IL-1 receptor antagonist (Il1rn) and mannose receptor C-type 1 (Mrc1), whereas macrophages from ob/ob islets showed elevated levels of transforming growth factor beta 1 (Tgfb1) and reduced IL-1β (Il1b). Clodronate-liposome treatment of islets depleted macrophages, as evidenced by reduced mRNA expression of Cd11b (also known as Itgam) and F4/80 (also known as Adgre1) compared with PBS-liposome-treated islets. The depletion of macrophages in db/db islets increased the expression of genes related to beta cell identity. The mRNA levels of islet-associated transcription factors (Mafa and Pdx1), glucose transporter (Glut2 [also known as Slc2a2]), ATP-sensitive K The findings of this study suggest that distinct alterations in islet macrophages of obese mice are critically important for the disruption of beta cell gene expression in diabetes. Show less

Persons with type 2 diabetes mellitus (T2DM) have an elevated risk of atherosclerosis. High-density lipoproteins (HDL) normally protect against cardiovascular disease (CVD), but this may be attenuated by serum amyloid A (SAA). In a case-control study of young females, blood samples were compared between subjects with T2DM ( Show less

Compounds targeting somatostatin-receptor-type-2 (SSTR2) are useful for small bowel neuroendocrine tumor (SBNET) and pancreatic neuroendocrine tumor (PNET) imaging and treatment. We recently characterized expression of 13 cell surface receptor genes in SBNETs and PNETs, identifying three drug targets (GIPR, OXTR, and OPRK1). This study set out to characterize expression of this gene panel in the less common neuroendocrine tumors of the stomach and duodenum (gastric and duodenal neuroendocrine tumors [GDNETs]). Primary tumors and adjacent normal tissue were collected at surgery, RNA was extracted, and expression of 13 target genes was determined by quantitative polymerase chain reaction. Expression was normalized to GAPDH and POLR2A internal control genes. Expression relative to normal tissue (ddCT) and absolute expression (dCT) were calculated. Wilcoxon tests compared median expression with false discovery rate correction for multiple comparisons. Gene expression was similar in two gastric and seven duodenal tumors, and these were analyzed together. Like SBNETs (n = 63) and PNETs (n = 51), GDNETs showed significant overexpression compared with normal tissue of BRS3, GIPR, GRM1, GPR113, OPRK1, and SSTR2 (P < 0.05 for all). Of these, SSTR2 had the highest absolute expression in GDNETs (median dCT 4.0). Absolute expression of BRS3, GRM1, GPR113, and OPRK1 was significantly lower than SSTR2 in GDNETs (P < 0.05 for all), whereas expression of GIPR was similar to SSTR2 (median 4.3, P = 0.4). As in SBNETs and PNETs, GIPR shows absolute expression close to SSTR2 but has greater overexpression relative to normal tissue (21.1 versus 3.5-fold overexpression). We conclude that GIPR could provide an improved signal-to-noise ratio for imaging versus SSTR2 and represents a promising novel therapeutic target in GDNETs. Show less

Fetal akinesia deformation sequence syndrome (FADS) is a heterogeneous disorder characterised by fetal akinesia and developmental defects including, in some case, pterygia. Multiple pterygium syndromes (MPS) are traditionally divided into prenatally lethal and non-lethal (such as Escobar) types. Previously, we and others reported that homozygous mutations in the fetal acetylcholine receptor gamma subunit (CHRNG) can cause both lethal and non-lethal MPS, demonstrating that pterygia resulted from fetal akinesia, and that mutations in the acetylcholine receptor subunits CHRNA1, CHRND, and Rapsyn (RAPSN) can also result in a MPS/FADS phenotype. We hypothesised that mutations in other acetylcholine receptor related genes may interfere with neurotransmission at the neuromuscular junction and so we analysed 14 cases of lethal MPS/FADS without CHRNG, CHRNA1, CHRNB1, CHRND, or RAPSN mutations for mutations in DOK7. A homozygous DOK7 splice site mutation, c.331+1G>T, was identified in a family with three children affected with lethal FADS. Previously DOK7 mutations have been reported to underlie a congenital myaesthenic syndrome with a characteristic "limb girdle" pattern of muscle weakness. This finding is consistent with the hypothesis that whereas incomplete loss of DOK7 function may cause congenital myasthenia, more severe loss of function can result in a lethal fetal akinesia phenotype. Show less

Congenital myasthenic syndromes are inherited disorders of neuromuscular transmission characterized by fatigable muscle weakness. Autosomal recessive acetylcholine receptor (AChR) deficiency syndromes, in which levels of this receptor at the neuromuscular junction are severely reduced, may be caused by mutations within genes encoding the AChR or the AChR-clustering protein, rapsyn. Most patients have mutations within the rapsyn coding region and are either homozygous for N88K or heteroallelic for N88K and a second mutation. In some cases the second allele carries a null mutation but in many the mutations are missense, and are located in different functional domains. Little is known about the functional effects of these mutations, but we hypothesize that they would have an effect on AChR clustering by a variety of mechanisms that might correlate with disease severity. Here we expressed RAPSN mutations A25V, N88K, R91L, L361R and K373del in TE671 cells and in rapsyn-/- myotubes to determine their pathogenic mechanisms. The A25Vmutation impaired colocalization of rapsyn with AChR and prevented agrin-induced AChR clusters in rapsyn-/- myotubes. In TE671 cells, R91L reduced the ability of rapsyn to self-associate, and K373del-rapsyn was significantly less stable than wild-type. The effects of mutations L361R and N88K were more subtle: in TE671 cells, in comparison with wild-type rapsyn, L361R-rapsyn showed reduced expression/stability, and both N88K-rapsyn and L361R-rapsyn showed significantly reduced co-localization with AChR. N88K-rapsyn and L361R-rapsyn could effectively mediate agrin-induced AChR clusters, but these were reduced in number and were less stable than with wild-type rapsyn. The disease severity of patients harbouring the compound allelic mutations was greater than that of patients with homozygous rapsyn mutation N88K, suggesting that the second mutant allele may largely determine severity. Show less