Adipose tissue macrophage (ATM)-driven inflammation plays a key role in insulin resistance; however, factors activating ATMs are poorly understood. Using a proteomics approach, we show that markers of classical activation are absent on ATMs from obese humans, but readily detectable on airway macrophages of patients with cystic fibrosis, a disease of chronic bacterial infection. Moreover, treating macrophages with glucose, insulin, and palmitate – conditions characteristic of the metabolic syndrome – produces a ‘metabolically-activated’ phenotype distinct from classical activation. Markers of metabolic activation are expressed by pro-inflammatory ATMs in obese humans/mice and are positively correlated with adiposity. Metabolic activation is driven by independent pro- and anti-inflammatory pathways, which regulate balance between cytokine production and lipid metabolism. We identify PPARγ and p62/SQSTM1 as two key proteins that promote lipid metabolism and limit inflammation in metabolically-activated macrophages. Collectively, our data provide important mechanistic insights into pathways that drive the metabolic disease-specific phenotype of macrophages.

SummaryThe concept of glucolipotoxicity refers to the combined, deleterious effects of elevated glucose and fatty acid levels on pancreatic beta-cell function and survival. Significant progress has been made in recent years towards a better understanding of the cellular and molecular basis of glucolipotoxicity in the beta cell. The permissive effect of elevated glucose on the detrimental actions of fatty acids stems from the influence of glucose on intracellular fatty-acid metabolism, promoting the synthesis of cellular lipids. The combination of excessive levels of fatty acids and glucose therefore leads to decreased insulin secretion, impaired insulin gene expression, and beta-cell death by apoptosis, all of which probably have distinct underlying mechanisms. Recent studies from our laboratory have identified several pathways implicated in fatty-acid inhibition of insulin gene expression, including the extracellular-regulated kinase (ERK1/2) pathway; the metabolic sensor Per-Arnt-Sim kinase (PASK); and the ATF6 branch of the unfolded protein response. We have also confirmed in vivo in rats that the decrease in insulin gene expression is an early defect which precedes any detectable abnormality in insulin secretion. While the role of glucolipotoxicity in humans is still debated, the inhibitory effects of chronically elevated fatty acid levels has been clearly demonstrated in several studies, at least in individuals genetically predisposed to developing type 2 diabetes. It is therefore likely that glucolipotoxicity contributes to beta-cell failure in type 2 diabetes as well as to the decline in beta-cell function observed after the onset of the disease.

The objective of this comprehensive review is to summarize and discuss the available evidence of how adipose tissue inflammation affects insulin sensitivity and glucose tolerance. Low-grade, chronic adipose tissue inflammation is characterized by infiltration of macrophages and other immune cell populations into adipose tissue, and a shift towards more pro-inflammatory subtypes of leukocytes. The infiltration of pro-inflammatory cells in adipose tissue is associated with an increased production of key chemokines such as C-C motif chemokine ligand 2, pro-inflammatory cytokines including tumor necrosis factor α and interleukins 1β and 6, as well as reduced expression of the key insulin sensitizing adipokine, adiponectin. In both rodent models and humans, adipose tissue inflammation is consistently associated with excess fat mass and insulin resistance. In humans, associations with insulin resistance are stronger and more consistent for inflammation in visceral as opposed to subcutaneous fat. Further, genetic alterations in mouse models of obesity that reduce adipose tissue inflammation are – almost without exception - associated with improved insulin sensitivity. However, a dissociation between adipose tissue inflammation and insulin resistance can be observed in very few rodent models of obesity as well as in humans following bariatric surgery- or low-calorie diet-induced weight loss, illustrating that the etiology of insulin resistance is multifactorial. Taken together, adipose tissue inflammation is a key factor in the development of insulin resistance and type 2 diabetes in obesity, along with other factors that likely include inflammation and fat accumulation in other metabolically active tissues.

The first objective of this study was to describe the effect of on-farm heat treatment of colostrum on colostral bacteria counts and IgG concentrations. The second objective was to describe the effect of feeding heat-treated (vs. raw) colostrum on passive transfer of colostral immune and nutritional parameters in neonatal calves. Pooled batches of colostrum were mixed and divided equally: one half was fed raw whereas the other half was fed after heat treatment at 60 degrees C for 60 min using a commercial on-farm batch pasteurizer. Colostrum samples were cultured for total bacteria count and total coliform count and analyzed for total IgG concentration. Forty-nine Holstein calves were fed either raw colostrum (n = 24) or heat-treated colostrums (n = 25) within 1 to 2 h after birth. Serum samples collected from calves at 0 h (precolostrum) and 24 h (postcolostrum) were assayed for serum total protein; IgG, IgA, and IgM concentrations; peripheral total leukocyte counts; neutrophil counts; lymphocyte counts; lymphocyte phenotypes; vitamin A, vitamin E, cholesterol, and beta-carotene concentrations. Serum samples collected from 2- to 5-d-old calves were tested for immunoglobulin function via a bovine viral diarrhea virus type I serum neutralization titer and for neutrophil bacterial opsonization activity. On-farm batch heat treatment of colostrum at 60 degrees C for 60 min resulted in lower colostrum bacteria concentrations while maintaining colostral IgG concentration. Calves fed heat-treated colostrum had significantly greater serum total protein and IgG concentrations at 24 h, plus greater apparent efficiency of IgG absorption (total protein = 6.3 mg/dL; IgG = 22.3 mg/mL; apparent efficiency of absorption = 35.6%) compared with calves fed raw colostrum (TP = 5.9 mg/dL; IgG = 18.1 mg/mL; apparent efficiency of absorption = 26.1%). There was no effect of treatment on serum concentrations of IgA, IgM, vitamin A, vitamin E, cholesterol, beta-carotene or vitamin E:cholesterol ratio, or on serum bovine viral diarrhea virus type I serum neutralization titers. There was no difference between treatment groups when examining calf plasma total leukocyte counts, neutrophil counts, lymphocyte counts, or neutrophil opsonization activity. However, the latter results were considered inconclusive.

Abnormalities in lipid metabolism have been proposed as contributing factors to both defective insulin secretion from the pancreatic beta cell and peripheral insulin resistance in type 2 diabetes. Previously, we have shown that prolonged exposure of isolated rat islets of Langerhans to excessive fatty acid levels impairs insulin gene transcription. This study was designed to assess whether palmitate alters the expression and binding activity of the key regulatory factors pancreas-duodenum homeobox-1 (PDX-1), MafA, and Beta2, which respectively bind to the A3, C1, and E1 elements in the proximal region of the insulin promoter. Nuclear extracts of isolated rat islets cultured with 0.5 mM palmitate exhibited reduced binding activity to the A3 and C1 elements but not the E1 element. Palmitate did not affect the overall expression of PDX-1 but reduced its nuclear localization. In contrast, palmitate blocked the stimulation of MafA mRNA and protein expression by glucose. Combined adenovirus-mediated overexpression of PDX-1 and MafA in islets completely prevented the inhibition of insulin gene expression by palmitate. These results demonstrate that prolonged exposure of islets to palmitate inhibits insulin gene transcription by impairing nuclear localization of PDX-1 and cellular expression of MafA.The prevalence of diabetes mellitus is increasing dramatically in Western countries, in part because of the increase in obesity. Type 2 diabetes mellitus, the most frequent form of the disease, is characterized by defective insulin secretion from the pancreatic beta cells and peripheral insulin resistance. According to the lipotoxicity hypothesis, abnormalities in lipid metabolism contribute to both defects (1) and in particular to the inexorable decline of beta cell function observed during the course of the disease (2). However, the mechanisms of lipotoxicity in the beta cell remain largely unknown.In vitro, prolonged exposure to excessive concentrations of fatty acids inhibits glucose-stimulated insulin secretion (3-7) and insulin gene expression (8 -11). Previous studies in our laboratory have shown that deleterious effects of fatty acids appear mediated by distinct mechanisms; whereas inhibition of insulin secretion is observed after culture with palmitate, oleate, and other fatty acids (7), insulin gene expression is only affected by palmitate and is mediated via de novo synthesis of ceramide (11). In isolated rat islets, we have shown that palmitate markedly blunts the activation by glucose of an insulin promoter reporter construct, indicating a transcriptional mode of action (11). However, the mechanisms by which palmitate affects the insulin promoter are unknown.Both beta cell-specific expression and metabolic regulation of the insulin gene are conferred by a highly conserved region lying ϳ340 bp upstream of the transcription initiation site that constitutes the promoter/enhancer region (12-14). The main glucose-responsive elements on the insulin promoter are the highly conserved A3 (15), C1 (16), and E1 (16) sites,...

The islet-enriched MafA, PDX-1, and BETA2 activators contribute to both ␤ cell-specific and glucose-responsive insulin gene transcription. To investigate how these factors impart activation, their combined impact upon insulin enhancer-driven expression was first examined in non-␤ cell line transfection assays. Individual expression of PDX-1 and BETA2 led to little or no activation, whereas MafA alone did so modestly. MafA together with PDX-1 or BETA2 produced synergistic activation, with even higher insulin promoter activity found when all three proteins were present. Stimulation was attenuated upon compromising either MafA transactivation or DNA-binding activity. MafA interacted with endogenous PDX-1 and BETA2 in coimmunoprecipitation and in vitro GST pull-down assays, suggesting that regulation involved direct binding. Dominant-negative acting and small interfering RNAs of MafA also profoundly reduced insulin promoter activity in ␤ cell lines. In addition, MafA was induced in parallel with insulin mRNA expression in glucose-stimulated rat islets. Insulin mRNA levels were also elevated in rat islets by adenoviral-mediated expression of MafA. Collectively, these results suggest that MafA plays a key role in coordinating and controlling the level of insulin gene expression in islet ␤ cells.Insulin is selectively expressed in the pancreatic ␤ cells of the islet of Langerhans. Restricted expression is due to a unique combination of factors that stimulate through conserved enhancer region sequences located approximately between nucleotides Ϫ340 and Ϫ90 relative to the transcription start site (1-4). Detailed analysis has revealed that activation is primarily controlled by PAX6, PDX-1, MafA, and BETA2 binding to the C2 (Ϫ317 to Ϫ311 bp), A3 (Ϫ201 to Ϫ196 bp), C1 (Ϫ126 to Ϫ101 bp), and E1 (Ϫ100 to Ϫ91 bp) elements, respectively (5, 6). These distinct factors are enriched in islet cells, with BETA2 (7, 8) and PAX6 (9) present in all islet cell types, PDX-1 in ␤ and a subset of ␦ cells

The insulin gene is expressed almost exclusively in pancreatic beta-cells. Metabolic regulation of insulin gene expression enables the beta-cell to maintain adequate stores of intracellular insulin to sustain the secretory demand. Glucose is the major physiologic regulator of insulin gene expression; it coordinately controls the recruitment of transcription factors [e.g., pancreatic/duodenal homeobox-1 (PDX-1), mammalian homologue of avian MafA/L-Maf (MafA), Beta2/Neuro D (B2), the rate of transcription, and the stability of insulin mRNA. However, chronically elevated levels of glucose (glucotoxicity) and lipids (lipotoxicity) also contribute to the worsening of beta-cell function in type 2 diabetes, in part via inhibition of insulin gene expression. The mechanisms of glucotoxicity, which involve decreased binding activities of PDX-1 and MafA and increased activity of C/EBPbeta, are mediated by high-glucose-induced generation of oxidative stress. On the other hand, lipotoxicity is mediated by de novo ceramide synthesis and involves inhibition of PDX-1 nuclear translocation and MafA gene expression. Glucotoxicity and lipotoxicity have common targets, which makes their combination particularly harmful to insulin gene expression and beta-cell function in type 2 diabetes.

In comparison with UW organ preservation, exposure of pancreata to the TLM result in greater islet yields and extended preservation times.