Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 10, 29

T Cells Cooperate With Palmitic Acid in Induction of Beta Cell Apoptosis

Affiliations

T Cells Cooperate With Palmitic Acid in Induction of Beta Cell Apoptosis

Tamara Cvjetićanin et al. BMC Immunol.

Abstract

Background: Diabetes is characterized by progressive failure of insulin producing beta cells. It is well known that both saturated fatty acids and various products of immune cells can contribute to the reduction of beta cell viability and functionality during diabetes pathogenesis. However, their joint action on beta cells has not been investigated, so far. Therefore, we explored the possibility that leukocytes and saturated fatty acids cooperate in beta cell destruction.

Results: Rat pancreatic islets or insulinoma cells (RIN) were co-cultivated with concanavalin A (ConA)-stimulated rat lymph node cells (LNC), or they were treated with cell-free supernatants (Sn) obtained from ConA-stimulated spleen cells or from activated CD3+ cells, in the absence or presence of palmitic acid (PA). ConA-stimulated LNC or Sn and PA cooperated in inducing caspase-3-dependent RIN cell apoptosis. The observed effect of PA and Sn on RIN cell viability was mediated by p38 mitogen-activated protein kinase (MAPK)-signaling and was achieved through auto-destructive nitric oxide (NO) production. The cooperative effect of Sn was mimicked with the combination of interleukin-1beta, interleukin-2, interleukin-6, interleukin-17, interferon-gamma and tumor necrosis factor-alpha.

Conclusion: These results imply that stimulated T cells produce cytokines that cooperate with saturated free fatty acids in beta cell destruction during diabetes pathogenesis.

Figures

Figure 1
Figure 1
PA and immune cells cooperate in pancreatic cell viability reduction. RIN cells were treated with various doses of PA (A), or they were cultivated alone (medium) or co-cultivated with LNC (B, C) in the absence (B) or presence (C) of tissue culture inserts, or cultivated in the presence of 40% Sn (D) or 10% Sn (F), in the absence or presence of 125 μM PA (B-D) or 32 μM PA (F). Primary pancreatic islets were cultivated alone (medium) or in the presence of 40% Sn, in the absence or presence of 125 μM PA (E). MTT assay was performed after 20 hours of cultivation and the results are presented as the percentage of control absorbance values obtained for cultures grown in medium alone. Mean values +/- SD of values obtained in 8 (A), 14 (B), 3 (C), 11 (D), 3 (E) and 6 (F) individual experiments are presented. *p < 0.05 represents a statistically significant difference between values obtained from RIN cell cultures treated with PA in the presence of LNC and any other culture of RIN cells or RIN cells treated with PA and Sn and any other culture of RIN cells.
Figure 2
Figure 2
Apoptotic cell death of RIN cells under the influence of PA and Sn. RIN cells were cultivated in the absence (medium) or presence of 125 μM PA and/or 40% Sn for 6 hours (A, C-F) or 4 hours (B). Subsequently, the cells were stained with AnnV/EtD-III and cytofluorimetric acquisition and analysis were performed (A, C-F) or they were analyzed for caspase-3 activity (B). Mean values +/- SD of values obtained in 3 (A) and 4 (B) individual experiments with similar results are presented. *p < 0.05 represents a statistically significant difference between values obtained from cultures of RIN cells treated with PA and Sn and any other culture of RIN cells. Plots of the cells stained with AnnV/EtD-III from a representative experiment are presented in the lower panel (C-F), where numbers in the lower right quadrant of the plots are the percentage of apoptotic cells and numbers in the upper right quadrant are the percentage of necrotic cells.
Figure 3
Figure 3
Cooperation of mouse Sn, heated Sn, cytokines, CD3+Sn with PA in reduction of RIN cell viability. RIN cells were cultivated in the absence (medium) or presence of 125 μM PA and/or 40% C57Bl/6 mouse Sn (SnM – A), and/or 40% Sn or Sn boiled for 10 minutes (SnB-B), and/or 10 ng/ml IL-1β, 10 ng/ml IL-6, 10 ng/ml IFN-γ, 10 ng/ml TNF-α, 50 ng/ml IL-17 and 100 ng/ml IL-2 (cytokines – C) and/or 40% Sn obtained from CD3+ LNC or CD3- LNC (SnCD3+, SnCD3- – D). MTT assay was performed after 20 hours of cultivation and the results are presented as the percentage of control absorbance values obtained in cultures grown in medium alone. Mean values +/- SD of values obtained in 11 (A), 7 (B), 5 (C) and 4 (D) individual experiments with similar results are presented. *p < 0.05 represents a statistically significant difference between values obtained from cultures of RIN cells treated with PA and SnM (A) or PA and Sn (B) or PA and cytokines (C) or PA and SnCD3+ (D) and any other culture of RIN cells.
Figure 4
Figure 4
The involvement of p38 MAPK signaling pathway in the induction of apoptosis of RIN cells under the influence of PA and Sn. RIN cells were cultivated in the absence (medium) or presence of 125 μM PA and/or 40% Sn. For apoptosis detection, RIN cells were treated with PA and Sn in the absence or presence of an inhibitor of p38 MAPK signaling – SB202109 (40 μM) for 6 hours, and subsequently stained with AnnV/EtD-III (A). Alternatively, RIN cells were treated with PA and/or Sn for 1 hour, then cell lysates were made before western blotting for p38 MAPK and phosphorylated p38 (p-p38) (B, C). Mean values +/- SD of values obtained in 5 (A) and 3 (B) individual experiments with similar results are presented, or a representative western blot (C). *p < 0.05 represents a statistically significant difference relative to the cultures grown in medium without additional treatment (A, B) and to the cultures treated with SB (A).
Figure 5
Figure 5
PA and Sn induce RIN cell apoptosis through induction of nitric oxide production. RIN cells were cultivated without treatment (medium) or treated with Sn and/or PA and/or hemoglobin (Hb, 20 mg/ml) and/or SB202109 (40 μM). After 2 or 6 hours RIN cells were fixed before cell-based ELISA for iNOS (A). Cells were stained with DAF-FM (B, D) and AnnV/EtD-III (C) after 6 hours of cultivation. Mean values +/- SD of values obtained in three individual experiments with similar results are presented (A, C). Alternatively, plots from a representative of at least three experiments with similar data are presented (B, D). *p < 0.05 represents a statistically significant difference relative to the cultures grown in medium without additional treatment (A, C) and to the cultures treated with Hb (C).

Similar articles

See all similar articles

Cited by 4 articles

References

    1. Roglic G, Unwin N, Bennett PH, Mathers C, Tuomilehto J, Nag S, Connolly V, King H. The burden of mortality attributable to diabetes: realistic estimates for the year. Diabetes Care. 2000;28:2130–2135. doi: 10.2337/diacare.28.9.2130. - DOI - PubMed
    1. Wild S, Roglic G, Green A, Sicree R, King H. Global prevalence of diabetes: estimates for the year 2000 and projections for 2030. Diabetes Care. 2004;27:1047–1053. doi: 10.2337/diacare.27.5.1047. - DOI - PubMed
    1. Kahn SE. The relative contributions of insulin resistance and beta-cell dysfunction to the pathophysiology of Type 2 diabetes. Diabetologia. 2003;46:3–19. doi: 10.1007/s00125-003-1190-9. - DOI - PubMed
    1. Donath MY, Schumann DM, Faulenbach M, Ellingsgaard H, Perren A, Ehses JA. Islet inflammation in type 2 diabetes: from metabolic stress to therapy. Diabetes Care. 2008;31:161–164. doi: 10.2337/dc08-s243. - DOI - PubMed
    1. Wilkin TJ. Changing perspectives in diabetes: their impact on its classification. Diabetologia. 2007;50:1587–1592. doi: 10.1007/s00125-007-0665-5. - DOI - PubMed

Publication types

MeSH terms

Feedback