Defective apoptosis in intestinal and mesenteric adipose tissue of crohn's disease patients

Background: Crohn's disease (CD) is associated with complex pathogenic pathways involving defects in apoptosis mechanisms. Recently, mesenteric adipose tissue (MAT) has been associated with CD ethiopathology, since adipose thickening is detected close to the affected intestinal area. However, the potential role of altered apoptosis in MAT of CD has not been addressed. Aims: To evaluate apoptosis in the intestinal mucosa and MAT of patients with CD. Methods: Samples of intestinal mucosa and MAT from patients with ileocecal CD and from non-inflammatory bowel diseases patients (controls) were studied. Apoptosis was assessed by TUNEL assay and correlated with the adipocytes histological morphometric analysis. The transcriptional and protein analysis of selected genes and proteins related to apoptosis were determined. Results: TUNEL assay showed fewer apoptotic cells in CD, when compared to the control groups, both in the intestinal mucosa and in MAT. In addition, the number of apoptotic cells (TUNEL) correlated significantly with the area and perimeter of the adipose cells in MAT. Transcriptomic and proteomic analysis reveal a significantly lower transcript and protein levels of Bax in the intestinal mucosa of CD, compared to the controls; low protein levels of Bax were found localized in the lamina propria and not in the epithelium of this tissue. Furthermore, higher level of Bcl-2 and low level of Caspase 3 were seen in the MAT of CD patients. Conclusion: The defective apoptosis in MAT may explain the singular morphological characteristics of this tissue in CD, which may be implicated in the pathophysiology of the disease. © 2014 Dias et al.Crohn's disease (CD) is associated with complex pathogenic pathways involving defects in apoptosis mechanisms. Recently, mesenteric adipose tissue (MAT) has been associated with CD ethiopathology, since adipose thickening is detected close to the affected intestinal area. However, the potential role of altered apoptosis in MAT of CD has not been addressed. Aims: To evaluate apoptosis in the intestinal mucosa and MAT of patients with CD. Methods: Samples of intestinal mucosa and MAT from patients with ileocecal CD and from non-inflammatory bowel diseases patients (controls) were studied. Apoptosis was assessed by TUNEL assay and correlated with the adipocytes histological morphometric analysis. The transcriptional and protein analysis of selected genes and proteins related to apoptosis were determined. Results: TUNEL assay showed fewer apoptotic cells in CD, when compared to the control groups, both in the intestinal mucosa and in MAT. In addition, the number of apoptotic cells (TUNEL) correlated significantly with the area and perimeter of the adipose cells in MAT. Transcriptomic and proteomic analysis reveal a significantly lower transcript and protein levels of Bax in the intestinal mucosa of CD, compared to the controls; low protein levels of Bax were found localized in the lamina propria and not in the epithelium of this tissue. Furthermore, higher level of Bcl-2 and low level of Caspase 3 were seen in the MAT of CD patients. Conclusion: The defective apoptosis in MAT may explain the singular morphological characteristics of this tissue in CD, which may be implicated in the pathophysiology of the disease96e9854

ER stress activation in the intestinal mucosa but not in mesenteric adipose tissue is associated with inflammation in Crohn’s disease patients

Chronic/abnormal activation of endoplasmic reticulum (ER) stress is linked to the exacerbation of the inflammatory process and has been recently linked to Crohn’s disease (CD) pathophysiology. We investigated the intestinal mucosa and the mesenteric adipose tissue (MAT) collected from CD patients with active disease (CD group) and from non-IBD patients (CTR group) to study ER stress activation and to address tissue-specific modulation in CD. The intestinal mucosa of CD patients showed an upregulation in the expression of ER stress related genes, including ATF3, DNAJC3, STC2, DDIT3, CALR, HSPA5 and HSP90B1. Results showed that EIF2AK3 gene was upregulated, along with increased protein expression of p-eIF2α and p-eIF2α/eIF2α ratio. Additionally, ERN1 gene expression was upregulated, along with an increased spliced/activated form sXBP1 protein. Despite the upregulation of ATF6 gene expression in the intestinal mucosa of CD patients, no differences were found in ATF6 protein expression. Lastly, the analysis of MAT revealed unchanged levels of ER stress markers along with no differences in the activation of UPR. However, chaperone gene expression was modulated in the MAT of CD patients. To conclude, our results address tissue-specific differences in UPR activation in CD and point the ER stress as an important pro-inflammatory mechanism in CD, specifically in the intestinal mucosa149CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP401270/2016-502-P-28707/20122016/01638-

Toll-like Receptor 4, F4/80 And Pro-inflammatory Cytokines In Intestinal And Mesenteric Fat Tissue Of Crohn's Disease.

Crohn's disease (CD) is a chronic intestinal ailment with a multifactorial etiology, whose incidence has increased during the last three decades. Recently, a role for mesenteric fat has been proposed in CD pathophysiology, since fat hypertrophy is detected nearby the affected intestinal area; however, there are few studies on this aspect. To evaluate inflammatory activity in intestinal mucosa and mesenteric fat tissue of patients with CD and controls. Ten patients with ileocecal CD and 16 patients with non-inflammatory disease (control groups) were studied. The specimens were snap-frozen and the expression of TLR-4, F4/80, IL1-β and IL-6 were determined by immunoblot of protein extracts. TLR4 RNA level were measured using RT-PCR. The t Test was applied (p<0.05). The local ethical committee approved the study. The intestinal mucosa of CD group had significantly higher protein levels of TLR-4, F4/80, IL-1β and IL-6 than the controls. The gene expression of TLR4 was lower in the intestinal mucosa of CD compared to the control group. Regard the mesenteric fat tissue, there was no statistical difference related to TLR-4, F4/80, IL-1β and IL-6 proteins expression. These findings may result from an up-regulation of macrophage activation and intracellular pathways activated by bacterial antigens, which are more important in intestinal mucosa than fat tissue in CD patients. This may represent an anomalous regulation of innate immunity and could contribute to the production of proinflammatory mediators and disease development.698-10

Maternal consumption of a high-fat diet modulates the inflammatory response in their offspring, mediated by the M1 muscarinic receptor

IntroductionHigh-fat diet (HFD) consumption is associated with various metabolic disorders and diseases. Both pre-pregnancy and maternal obesity can have long-term consequences on offspring health. Furthermore, consuming an HFD in adulthood significantly increases the risk of obesity and metabolic disorders. However, an intriguing phenomenon known as the obesity paradox suggests that obesity may confer a protective effect on mortality outcomes in sepsis. In sepsis, activation of the cholinergic anti-inflammatory pathway (CAP) can help mitigate systemic inflammation. We employed a metabolic programming model to explore the relationship between maternal HFD consumption and offspring response to sepsis.MethodsWe fed female mice either a standard diet (SC) or an HFD during the pre-pregnancy, pregnancy, and lactation periods. Subsequently, we evaluated 28-day-old male offspring. ResultsNotably, we discovered that offspring from HFD-fed dams (HFD-O) exhibited a higher survival rate compared with offspring from SC-fed dams (SC-O). Importantly, inhibition of the m1 muscarinic acetylcholine receptor (m1mAChR), involved in the CAP, in the hypothalamus abolished this protection. The expression of m1mAChR in the hypothalamus was higher in HFD-O at different ages, peaking on day 28. Treatment with an m1mAChR agonist could modulate the inflammatory response in peripheral tissues. Specifically, CAP activation was greater in the liver of HFD-O following agonist treatment. Interestingly, lipopolysaccharide (LPS) challenge failed to induce a more inflammatory state in HFD-O, in contrast to SC-O, and agonist treatment had no additional effect. Analysis of spleen immune cells revealed a distinct phenotype in HFD-O, characterized by elevated levels of CD4+ lymphocytes rather than CD8+ lymphocytes. Moreover, basal Il17 messenger RNA (mRNA) levels were lower while Il22 mRNA levels were higher in HFD-O, and we observed the same pattern after LPS challenge. DiscussionFurther examination of myeloid cells isolated from bone marrow and allowed to differentiate showed that HFD-O macrophages displayed an anti-inflammatory phenotype. Additionally, treatment with the m1mAChR agonist contributed to reducing inflammatory marker levels in both groups. In summary, our findings demonstrate that HFD-O are protected against LPS-induced sepsis, and this protection is mediated by the central m1mAChR. Moreover, the inflammatory response in the liver, spleen, and bone marrow-differentiated macrophages is diminished. However, more extensive analysis is necessary to elucidate the specific mechanisms by which m1mAChR modulates the immune response during sepsis

High-Fat Diet Induces Apoptosis of Hypothalamic Neurons

Consumption of dietary fats is amongst the most important environmental factors leading to obesity. In rodents, the consumption of fat-rich diets blunts leptin and insulin anorexigenic signaling in the hypothalamus by a mechanism dependent on the in situ activation of inflammation. Since inflammatory signal transduction can lead to the activation of apoptotic signaling pathways, we evaluated the effect of high-fat feeding on the induction of apoptosis of hypothalamic cells. Here, we show that consumption of dietary fats induce apoptosis of neurons and a reduction of synaptic inputs in the arcuate nucleus and lateral hypothalamus. This effect is dependent upon diet composition, and not on caloric intake, since pair-feeding is not sufficient to reduce the expression of apoptotic markers. The presence of an intact TLR4 receptor, protects cells from further apoptotic signals. In diet-induced inflammation of the hypothalamus, TLR4 exerts a dual function, on one side activating pro-inflammatory pathways that play a central role in the development of resistance to leptin and insulin, and on the other side restraining further damage by controlling the apoptotic activity

Hyperleptinemia Is Required for the Development of Leptin Resistance

Leptin regulates body weight by signaling to the brain the availability of energy stored as fat. This negative feedback loop becomes disrupted in most obese individuals, resulting in a state known as leptin resistance. The physiological causes of leptin resistance remain poorly understood. Here we test the hypothesis that hyperleptinemia is required for the development of leptin resistance in diet-induced obese mice. We show that mice whose plasma leptin has been clamped to lean levels develop obesity in response to a high-fat diet, and the magnitude of this obesity is indistinguishable from wild-type controls. Yet these obese animals with constant low levels of plasma leptin remain highly sensitive to exogenous leptin even after long-term exposure to a high fat diet. This shows that dietary fats alone are insufficient to block the response to leptin. The data also suggest that hyperleptinemia itself can contribute to leptin resistance by downregulating cellular response to leptin as has been shown for other hormones