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Anti-apoptotic BFL-1 is the major effector in activation-induced human mast cell survival.

Ekoff M, Lyberg K, Krajewska M, Arvidsson M, Rak S, Reed JC, Harvima I, Nilsson G - PLoS ONE (2012)

Bottom Line: FcεRI crosslinking promotes activation-induced survival of human mast cells and this is associated with an upregulation of the anti-apoptotic Bcl-2 family member Bfl-1.ABT-737 alone or in combination with roscovitine decreases viability of human mast cells although activation-induced survival is sustained, indicating a minor role for Bcl-X(L), Bcl-2, Bcl-w and Mcl-1.Taken together, our results highlight Bfl-1 as a major effector in activation-induced human mast cell survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Centre for Allergy Research, Karolinska Institutet, Stockholm, Sweden. maria.ekoff@ki.se

ABSTRACT
Mast cells are best known for their role in allergic reactions, where aggregation of FcεRI leads to the release of mast cell mediators causing allergic symptoms. The activation also induces a survival program in the cells, i.e., activation-induced mast cell survival. The aim of the present study was to investigate how the activation-induced survival is mediated. Cord blood-derived mast cells and the mast cell line LAD-2 were activated through FcεRI crosslinking, with or without addition of chemicals that inhibit the activity or expression of selected Bcl-2 family members (ABT-737; roscovitine). Cell viability was assessed using staining and flow cytometry. The expression and function of Bcl-2 family members BFL-1 and MCL-1 were investigated using real-time quantitative PCR and siRNA treatment. The mast cell expression of Bfl-1 was investigated in skin biopsies. FcεRI crosslinking promotes activation-induced survival of human mast cells and this is associated with an upregulation of the anti-apoptotic Bcl-2 family member Bfl-1. ABT-737 alone or in combination with roscovitine decreases viability of human mast cells although activation-induced survival is sustained, indicating a minor role for Bcl-X(L), Bcl-2, Bcl-w and Mcl-1. Reducing BFL-1 but not MCL-1 levels by siRNA inhibited activation-induced mast cell survival. We also demonstrate that mast cell expression of Bfl-1 is elevated in birch-pollen-provocated skin and in lesions of atopic dermatitis and psoriasis patients. Taken together, our results highlight Bfl-1 as a major effector in activation-induced human mast cell survival.

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Activation-induced mast cell survival following IgECL in presence of ABT-737 and roscovitine.(A) CBMC treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=6–3. (B) LAD-2 treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=3 Viability was assessed after 24 hrs using propidium iodide plus FITC-conjugated Annexin V. Change in viability is expressed as percentage viable cells after treatment deducted from untreated cells. ABT-737 = ABT, roscovitine = rosc.
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pone-0039117-g002: Activation-induced mast cell survival following IgECL in presence of ABT-737 and roscovitine.(A) CBMC treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=6–3. (B) LAD-2 treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=3 Viability was assessed after 24 hrs using propidium iodide plus FITC-conjugated Annexin V. Change in viability is expressed as percentage viable cells after treatment deducted from untreated cells. ABT-737 = ABT, roscovitine = rosc.

Mentions: Upon treating CBMCs with 0.5 μM ABT-737, mast cell viability decreased by approximately 10% and combination of 0.5 μM ABT-737 and 5 μM roscovitine decreased this viability even further by another 20% (Fig. 2A). Decreased viability could also be observed for LAD-2 cells where the viability was decreased by approximately 25% in response to ABT-737 or ABT-737 in combination with roscovitine (Fig. 2B). The results show that IgECL resulted in prolonged survival of both ABT-737 and ABT-737/Roscovitine-treated CBMCs and LAD-2 cells (Fig. 2A–B). These results demonstrate that the ability to induce survival is sustained in an experimental setting where Bcl-2, Bcl-XL, Bcl-W and Mcl-1 are inhibited, and suggest that one of the remaining pro-survival Bcl-2 family members (i.e. Bfl-1 and Bcl-B) was responsible for the activation-induced survival response. Since mice lack an obvious ortholog of Bcl-B, we focused on Bfl-1, the human ortholog of A1 which was shown to be essential for murine activation-induced mast cell survival [8].


Anti-apoptotic BFL-1 is the major effector in activation-induced human mast cell survival.

Ekoff M, Lyberg K, Krajewska M, Arvidsson M, Rak S, Reed JC, Harvima I, Nilsson G - PLoS ONE (2012)

Activation-induced mast cell survival following IgECL in presence of ABT-737 and roscovitine.(A) CBMC treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=6–3. (B) LAD-2 treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=3 Viability was assessed after 24 hrs using propidium iodide plus FITC-conjugated Annexin V. Change in viability is expressed as percentage viable cells after treatment deducted from untreated cells. ABT-737 = ABT, roscovitine = rosc.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3376125&req=5

pone-0039117-g002: Activation-induced mast cell survival following IgECL in presence of ABT-737 and roscovitine.(A) CBMC treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=6–3. (B) LAD-2 treated with 0.5 μM ABT-737 alone or in combination with 5 μM roscovitine following IgECL. N=3 Viability was assessed after 24 hrs using propidium iodide plus FITC-conjugated Annexin V. Change in viability is expressed as percentage viable cells after treatment deducted from untreated cells. ABT-737 = ABT, roscovitine = rosc.
Mentions: Upon treating CBMCs with 0.5 μM ABT-737, mast cell viability decreased by approximately 10% and combination of 0.5 μM ABT-737 and 5 μM roscovitine decreased this viability even further by another 20% (Fig. 2A). Decreased viability could also be observed for LAD-2 cells where the viability was decreased by approximately 25% in response to ABT-737 or ABT-737 in combination with roscovitine (Fig. 2B). The results show that IgECL resulted in prolonged survival of both ABT-737 and ABT-737/Roscovitine-treated CBMCs and LAD-2 cells (Fig. 2A–B). These results demonstrate that the ability to induce survival is sustained in an experimental setting where Bcl-2, Bcl-XL, Bcl-W and Mcl-1 are inhibited, and suggest that one of the remaining pro-survival Bcl-2 family members (i.e. Bfl-1 and Bcl-B) was responsible for the activation-induced survival response. Since mice lack an obvious ortholog of Bcl-B, we focused on Bfl-1, the human ortholog of A1 which was shown to be essential for murine activation-induced mast cell survival [8].

Bottom Line: FcεRI crosslinking promotes activation-induced survival of human mast cells and this is associated with an upregulation of the anti-apoptotic Bcl-2 family member Bfl-1.ABT-737 alone or in combination with roscovitine decreases viability of human mast cells although activation-induced survival is sustained, indicating a minor role for Bcl-X(L), Bcl-2, Bcl-w and Mcl-1.Taken together, our results highlight Bfl-1 as a major effector in activation-induced human mast cell survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Medicine, Centre for Allergy Research, Karolinska Institutet, Stockholm, Sweden. maria.ekoff@ki.se

ABSTRACT
Mast cells are best known for their role in allergic reactions, where aggregation of FcεRI leads to the release of mast cell mediators causing allergic symptoms. The activation also induces a survival program in the cells, i.e., activation-induced mast cell survival. The aim of the present study was to investigate how the activation-induced survival is mediated. Cord blood-derived mast cells and the mast cell line LAD-2 were activated through FcεRI crosslinking, with or without addition of chemicals that inhibit the activity or expression of selected Bcl-2 family members (ABT-737; roscovitine). Cell viability was assessed using staining and flow cytometry. The expression and function of Bcl-2 family members BFL-1 and MCL-1 were investigated using real-time quantitative PCR and siRNA treatment. The mast cell expression of Bfl-1 was investigated in skin biopsies. FcεRI crosslinking promotes activation-induced survival of human mast cells and this is associated with an upregulation of the anti-apoptotic Bcl-2 family member Bfl-1. ABT-737 alone or in combination with roscovitine decreases viability of human mast cells although activation-induced survival is sustained, indicating a minor role for Bcl-X(L), Bcl-2, Bcl-w and Mcl-1. Reducing BFL-1 but not MCL-1 levels by siRNA inhibited activation-induced mast cell survival. We also demonstrate that mast cell expression of Bfl-1 is elevated in birch-pollen-provocated skin and in lesions of atopic dermatitis and psoriasis patients. Taken together, our results highlight Bfl-1 as a major effector in activation-induced human mast cell survival.

Show MeSH
Related in: MedlinePlus