Involvement of the cytoplasmic cysteine-238 of CD40 in its up-regulation of CD23 expression and its enhancement of TLR4-triggered responses.

CD40, a member of the tumor necrosis factor receptor superfamily, plays a key role in both adaptive and innate immunity. Engagement of CD40 with its natural trimeric ligand or with cross-linked antibodies results in disulfide-linked CD40 (dl-CD40) homodimer formation, a process mediated by the cysteine-238 residues of the cytoplasmic tail of CD40. The present study was designed to elucidate the biological relevance of cysteine-238-mediated dl-CD40 homodimers to the expression of CD23 on B cells and to investigate its possible involvement in the innate response. Our results indicate that cysteine-238-mediated dl-CD40 homodimerization is required for CD40-induced activation of PI3-kinase/Akt signaling and the subsequent CD23 expression, as inhibition of dl-CD40 homodimer formation through a point mutation-approach specifically impairs these responses. Interestingly, cysteine-238-mediated dl-CD40 homodimers are also shown to play a crucial role in Toll-like receptor 4-induced CD23 expression, further validating the importance of this system in bridging innate and adaptive immune responses. This process also necessitates the activation of the PI3-kinase/Akt cascade. Thus, our results highlight new roles for CD40 and cysteine-238-mediated CD40 homodimers in cell biology and identify a potential new target for therapeutic strategies against CD40-associated chronic inflammatory diseases.

Interaction of CD154 with different receptors and its role in bidirectional signals.

In addition to its classical receptor, CD40, it is now well established that CD154 also binds αIIbß3, α5ß1, and αMß2 integrins. Although these integrins are all members of the same family, they bind CD154 differently. The current investigation aims to analyze the interaction of CD154 with α5ß1 and αMß2 and investigate its role in bidirectional signals in various human cell lines. Results obtained herein indicate that the CD154 residues involved in the interaction with α5ß1 are N151 and Q166, whereas those involved in αMß2 binding are common to residues required for CD40, namely Y145 and R203. Soluble CD40/CD154 or αMß2/CD154 complexes do not interfere with the binding of CD154 to α5ß1-positive cells, but inhibit the binding of CD154 to CD40- or αMß2-positive cells, respectively. Ligation of CD154 on CD154-positive cells with soluble CD40, αIIbß3, α5ß1, or αMß2 stimulates intracellular signaling, including MAPK phosphorylation. Given that CD154 exists as a trimer, our data strongly suggest that CD154 may bind concomitantly to two receptors of the same or different family, and biologically activate cells expressing both receptors. The characterization of CD154/receptor interactions helps the identification of new therapeutic targets for the prevention and/or treatment of CD154-associated autoimmune and inflammatory diseases.

Vascular smooth muscle cell alterations triggered by mice adipocytes: role of high-fat diet.

AIM: Inherent mechanisms leading to vascular smooth muscle cells (VSMC) alterations in obesitylinked type 2 diabetes (T2D) situation remain to be clarified. This study evaluates the impact of supernatant of adipocytes extracted from mice fed high-fat-diets (HFD) on the proliferation and apoptosis of VSMC. METHODS: Adipocytes were extracted from visceral white fat pads of male and female C57Bl6 mice showing different stages of metabolic alterations after 20 weeks of vegetal or animal HFD feeding. These cells were stimulated or not with insulin or glucose to condition VSMC media. After 24h of stimulation with adipocyte supernatants (AdS), VSMC proliferation and sustainability were assessed in the absence and presence of AdS. CD36 and insulin receptor mRNA levels were also evaluated. RESULTS: Proliferation and viability of VSMC were significantly modulated by the nature of the AdS used and the gender of mice from which adipocytes have been extracted. The most extensive effects on VSMC were triggered by adipocytes from males fed animal HFD and females fed vegetal HFD. These effects were concurrent with increased leptin concentration and decreased adiponectin levels in AdS. In addition, adipocytes of HFD-fed mice increased caspase-3 activity and apoptosis in VSMC. Significant up-regulation of CD36 mRNA was also found in these cells. CONCLUSION: Adipocytes of HFD-fed mice induce VSMC alterations. These changes involved mouse gender, most probably correlated to the diet-induced adipocyte secretion profile. Greater sensitivity to AdS effects in VSMC raises concerns about the more frequent cardiovascular events associated with obesity in the presence of T2D, which impairs adipocyte activity.

Nature of fatty acids in high fat diets differentially delineates obesity-linked metabolic syndrome components in male and female C57BL/6J mice.

BACKGROUND: Adverse effects of high-fat diets (HFD) on metabolic homeostasis are linked to adipose tissue dysfunction. The goal of this study was to examine the effect of the HFD nature on adipose tissue activity, metabolic disturbances and glucose homeostasis alterations in male mice compared with female mice. METHODS: C57BL/6J mice were fed either a chow diet or HFD including vegetal (VD) or animal (AD) fat. Body weight, plasmatic parameters and adipose tissue mRNA expression levels of key genes were evaluated after 20 weeks of HFD feeding. RESULTS: HFD-fed mice were significantly heavier than control at the end of the protocol. Greater abdominal visceral fat accumulation was observed in mice fed with AD compared to those fed a chow diet or VD. Correlated with weight gain, leptin levels in systemic circulation were increased in HFD-fed mice in both sexes with a significant higher level in AD group compared to VD group. Circulating adiponectin levels as well as adipose tissue mRNA expression levels were significantly decreased in HFD-fed male mice. Although its plasma levels remained unchanged in females, adiponectin mRNA levels were significantly reduced in adipose tissue of both HFD-fed groups with a more marked decrease in AD group compared to VD group. Only HFD-fed male mice were diabetic with increased fasting glycaemia. On the other hand, insulin levels were only increased in AD-fed group in both sexes associated with increased resistin levels. VD did not induce any apparent metabolic alteration in females despite the increased weight gain. Peroxisome Proliferator-Activated Receptors gamma-2 (PPARγ2) and estrogen receptor alpha (ERα) mRNA expression levels in adipose tissue were decreased up to 70% in HFD-fed mice but were more markedly reduced in male mice as compared with female mice. CONCLUSIONS: The nature of dietary fat determines the extent of metabolic alterations reflected in adipocytes through modifications in the pattern of adipokines secretion and modulation of key genes mRNA expression. Compared with males, female mice demonstrate higher capacity in controlling glucose homeostasis in response to 20 weeks HFD feeding. Our data suggest gender specific interactions between the diet's fatty acid source, the adipocyte-secreted proteins and metabolic disorders.