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Fractalkine (CX3CL1), a new factor protecting β-cells against TNFα.

Rutti S, Arous C, Schvartz D, Timper K, Sanchez JC, Dermitzakis E, Donath MY, Halban PA, Bouzakri K - Mol Metab (2014)

Bottom Line: CX3CL1 decreased human (but not rat) β-cell apoptosis.Moreover, CX3CL1 decreases basal apoptosis of human β-cells.We further demonstrate that CX3CL1 protects β-cells from the adverse effects of TNFα on their function by restoring the expression and phosphorylation of key proteins of the insulin secretion pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetic Medicine and Development, Geneva University, Geneva, Switzerland.

ABSTRACT

Objective: We have previously shown the existence of a muscle-pancreas intercommunication axis in which CX3CL1 (fractalkine), a CX3C chemokine produced by skeletal muscle cells, could be implicated. It has recently been shown that the fractalkine system modulates murine β-cell function. However, the impact of CX3CL1 on human islet cells especially regarding a protective role against cytokine-induced apoptosis remains to be investigated.

Methods: Gene expression was determined using RNA sequencing in human islets, sorted β- and non-β-cells. Glucose-stimulated insulin secretion (GSIS) and glucagon secretion from human islets was measured following 24 h exposure to 1-50 ng/ml CX3CL1. GSIS and specific protein phosphorylation were measured in rat sorted β-cells exposed to CX3CL1 for 48 h alone or in the presence of TNFα (20 ng/ml). Rat and human β-cell apoptosis (TUNEL) and rat β-cell proliferation (BrdU incorporation) were assessed after 24 h treatment with increasing concentrations of CX3CL1.

Results: Both CX3CL1 and its receptor CX3CR1 are expressed in human islets. However, CX3CL1 is more expressed in non-β-cells than in β-cells while its receptor is more expressed in β-cells. CX3CL1 decreased human (but not rat) β-cell apoptosis. CX3CL1 inhibited human islet glucagon secretion stimulated by low glucose but did not impact human islet and rat sorted β-cell GSIS. However, CX3CL1 completely prevented the adverse effect of TNFα on GSIS and on molecular mechanisms involved in insulin granule trafficking by restoring the phosphorylation (Akt, AS160, paxillin) and expression (IRS2, ICAM-1, Sorcin, PCSK1) of key proteins involved in these processes.

Conclusions: We demonstrate for the first time that human islets express and secrete CX3CL1 and CX3CL1 impacts them by decreasing glucagon secretion without affecting insulin secretion. Moreover, CX3CL1 decreases basal apoptosis of human β-cells. We further demonstrate that CX3CL1 protects β-cells from the adverse effects of TNFα on their function by restoring the expression and phosphorylation of key proteins of the insulin secretion pathway.

No MeSH data available.


Related in: MedlinePlus

CX3CL1 treatment prevents the impact of TNFα on glucose-stimulated insulin secretion and on the insulin signaling pathway in rat primary β-cells. Rat primary β-cells were cultured in the presence of CX3CL1 (48 h, 25 ng/ml) and/or TNFα (20 ng/ml for the last 24 h) and then incubated for 60 min at 2.8 mM glucose (open bars) followed by 60 min at 16.7 mM glucose (closed bars) (n = 5). A: Insulin secretion (expressed as percentage of cell content/h). B and C: Representative western blots and quantification showing Akt Ser-473 (B) or AS160 (C) phosphorylation. D: Representative western blots and quantification showing IRS-2 protein expression. *p < 0.05 vs. 2.8 mM glucose; #p < 0.05 for indicated comparison as tested by ANOVA followed by Bonferroni post hoc test.
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fig4: CX3CL1 treatment prevents the impact of TNFα on glucose-stimulated insulin secretion and on the insulin signaling pathway in rat primary β-cells. Rat primary β-cells were cultured in the presence of CX3CL1 (48 h, 25 ng/ml) and/or TNFα (20 ng/ml for the last 24 h) and then incubated for 60 min at 2.8 mM glucose (open bars) followed by 60 min at 16.7 mM glucose (closed bars) (n = 5). A: Insulin secretion (expressed as percentage of cell content/h). B and C: Representative western blots and quantification showing Akt Ser-473 (B) or AS160 (C) phosphorylation. D: Representative western blots and quantification showing IRS-2 protein expression. *p < 0.05 vs. 2.8 mM glucose; #p < 0.05 for indicated comparison as tested by ANOVA followed by Bonferroni post hoc test.

Mentions: The impact of CX3CL1 on insulin secretion was further investigated in primary rat sorted β-cells following 24 h of treatment with TNFα to induce insulin resistance and decrease GSIS [8]. Neither CX3CL1 (48 h pre-treatment) nor TNFα (24 h treatment) had an impact on subsequent short-term (1 h) basal (2.8 mM glucose) insulin secretion, whereas TNFα treatment dramatically reduced GSIS (1 h, 16.7 mM glucose) and this was prevented by CX3CL1 (Figure 4A). As observed previously for TNFα [10], the total insulin content was not influenced by the different conditions (data not shown). We have previously shown that several proteins from the canonical insulin signaling pathway are involved in GSIS in primary β-cells [30]. Therefore, we next explored the impact of CX3CL1 and/or TNFα on Akt, AS160, and IRS-2 protein expression in the basal condition (2.8 mM glucose) or after glucose stimulation (16.7 mM glucose). In rat primary sorted β-cells treated with TNFα for 24 h, glucose action on Akt Ser473 and AS160 phosphorylation was completely prevented (Figure 4B and C). By contrast, in cells pretreated with CX3CL1, glucose-induced Akt and AS160 phosphorylation was unaltered by TNFα exposure (Figure 4B and C). IRS-2 protein expression was increased by 48 h CX3CL1 treatment. In that condition IRS-2 protein degradation induced by TNFα was prevented (Figure 4D).


Fractalkine (CX3CL1), a new factor protecting β-cells against TNFα.

Rutti S, Arous C, Schvartz D, Timper K, Sanchez JC, Dermitzakis E, Donath MY, Halban PA, Bouzakri K - Mol Metab (2014)

CX3CL1 treatment prevents the impact of TNFα on glucose-stimulated insulin secretion and on the insulin signaling pathway in rat primary β-cells. Rat primary β-cells were cultured in the presence of CX3CL1 (48 h, 25 ng/ml) and/or TNFα (20 ng/ml for the last 24 h) and then incubated for 60 min at 2.8 mM glucose (open bars) followed by 60 min at 16.7 mM glucose (closed bars) (n = 5). A: Insulin secretion (expressed as percentage of cell content/h). B and C: Representative western blots and quantification showing Akt Ser-473 (B) or AS160 (C) phosphorylation. D: Representative western blots and quantification showing IRS-2 protein expression. *p < 0.05 vs. 2.8 mM glucose; #p < 0.05 for indicated comparison as tested by ANOVA followed by Bonferroni post hoc test.
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fig4: CX3CL1 treatment prevents the impact of TNFα on glucose-stimulated insulin secretion and on the insulin signaling pathway in rat primary β-cells. Rat primary β-cells were cultured in the presence of CX3CL1 (48 h, 25 ng/ml) and/or TNFα (20 ng/ml for the last 24 h) and then incubated for 60 min at 2.8 mM glucose (open bars) followed by 60 min at 16.7 mM glucose (closed bars) (n = 5). A: Insulin secretion (expressed as percentage of cell content/h). B and C: Representative western blots and quantification showing Akt Ser-473 (B) or AS160 (C) phosphorylation. D: Representative western blots and quantification showing IRS-2 protein expression. *p < 0.05 vs. 2.8 mM glucose; #p < 0.05 for indicated comparison as tested by ANOVA followed by Bonferroni post hoc test.
Mentions: The impact of CX3CL1 on insulin secretion was further investigated in primary rat sorted β-cells following 24 h of treatment with TNFα to induce insulin resistance and decrease GSIS [8]. Neither CX3CL1 (48 h pre-treatment) nor TNFα (24 h treatment) had an impact on subsequent short-term (1 h) basal (2.8 mM glucose) insulin secretion, whereas TNFα treatment dramatically reduced GSIS (1 h, 16.7 mM glucose) and this was prevented by CX3CL1 (Figure 4A). As observed previously for TNFα [10], the total insulin content was not influenced by the different conditions (data not shown). We have previously shown that several proteins from the canonical insulin signaling pathway are involved in GSIS in primary β-cells [30]. Therefore, we next explored the impact of CX3CL1 and/or TNFα on Akt, AS160, and IRS-2 protein expression in the basal condition (2.8 mM glucose) or after glucose stimulation (16.7 mM glucose). In rat primary sorted β-cells treated with TNFα for 24 h, glucose action on Akt Ser473 and AS160 phosphorylation was completely prevented (Figure 4B and C). By contrast, in cells pretreated with CX3CL1, glucose-induced Akt and AS160 phosphorylation was unaltered by TNFα exposure (Figure 4B and C). IRS-2 protein expression was increased by 48 h CX3CL1 treatment. In that condition IRS-2 protein degradation induced by TNFα was prevented (Figure 4D).

Bottom Line: CX3CL1 decreased human (but not rat) β-cell apoptosis.Moreover, CX3CL1 decreases basal apoptosis of human β-cells.We further demonstrate that CX3CL1 protects β-cells from the adverse effects of TNFα on their function by restoring the expression and phosphorylation of key proteins of the insulin secretion pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Genetic Medicine and Development, Geneva University, Geneva, Switzerland.

ABSTRACT

Objective: We have previously shown the existence of a muscle-pancreas intercommunication axis in which CX3CL1 (fractalkine), a CX3C chemokine produced by skeletal muscle cells, could be implicated. It has recently been shown that the fractalkine system modulates murine β-cell function. However, the impact of CX3CL1 on human islet cells especially regarding a protective role against cytokine-induced apoptosis remains to be investigated.

Methods: Gene expression was determined using RNA sequencing in human islets, sorted β- and non-β-cells. Glucose-stimulated insulin secretion (GSIS) and glucagon secretion from human islets was measured following 24 h exposure to 1-50 ng/ml CX3CL1. GSIS and specific protein phosphorylation were measured in rat sorted β-cells exposed to CX3CL1 for 48 h alone or in the presence of TNFα (20 ng/ml). Rat and human β-cell apoptosis (TUNEL) and rat β-cell proliferation (BrdU incorporation) were assessed after 24 h treatment with increasing concentrations of CX3CL1.

Results: Both CX3CL1 and its receptor CX3CR1 are expressed in human islets. However, CX3CL1 is more expressed in non-β-cells than in β-cells while its receptor is more expressed in β-cells. CX3CL1 decreased human (but not rat) β-cell apoptosis. CX3CL1 inhibited human islet glucagon secretion stimulated by low glucose but did not impact human islet and rat sorted β-cell GSIS. However, CX3CL1 completely prevented the adverse effect of TNFα on GSIS and on molecular mechanisms involved in insulin granule trafficking by restoring the phosphorylation (Akt, AS160, paxillin) and expression (IRS2, ICAM-1, Sorcin, PCSK1) of key proteins involved in these processes.

Conclusions: We demonstrate for the first time that human islets express and secrete CX3CL1 and CX3CL1 impacts them by decreasing glucagon secretion without affecting insulin secretion. Moreover, CX3CL1 decreases basal apoptosis of human β-cells. We further demonstrate that CX3CL1 protects β-cells from the adverse effects of TNFα on their function by restoring the expression and phosphorylation of key proteins of the insulin secretion pathway.

No MeSH data available.


Related in: MedlinePlus