Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells

Oscar K. Nihei, Paula C. Fonseca, Nara M. Rubim, Andre G. Bonavita, Jurandy S P O Lyra, Sandra Neves-dos-Santos, Antonio C C de Carvalho, David C. Spray, Wilson Savino, Luiz A. Alves

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Abstract

Background: We investigated the effects of the signaling molecules, cyclic AMP (cAMP) and protein-kinase C (PKC), on gap junctional intercellular communication (GJIC) between thymic epithelial cells (TEC).Results: Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC.Conclusions: Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.

Original languageEnglish (US)
Article number3
JournalBMC Cell Biology
Volume11
DOIs
StatePublished - Jan 15 2010

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Adenylate Kinase
Cyclic AMP
Protein Kinase C
Epithelial Cells
8-Bromo Cyclic Adenosine Monophosphate
Connexin 43
Thymocytes
Epinephrine
Gap Junctions
Cyclic Nucleotides
Tetradecanoylphorbol Acetate
Colforsin
Giant Cells
Coculture Techniques
Adenosine
Thymus Gland
Flow Cytometry
Coloring Agents
Epithelium
Fluorescence

ASJC Scopus subject areas

  • Cell Biology

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Nihei, O. K., Fonseca, P. C., Rubim, N. M., Bonavita, A. G., Lyra, J. S. P. O., Neves-dos-Santos, S., ... Alves, L. A. (2010). Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells. BMC Cell Biology, 11, [3]. https://doi.org/10.1186/1471-2121-11-3

Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells. / Nihei, Oscar K.; Fonseca, Paula C.; Rubim, Nara M.; Bonavita, Andre G.; Lyra, Jurandy S P O; Neves-dos-Santos, Sandra; de Carvalho, Antonio C C; Spray, David C.; Savino, Wilson; Alves, Luiz A.

In: BMC Cell Biology, Vol. 11, 3, 15.01.2010.

Research output: Contribution to journalArticle

Nihei, OK, Fonseca, PC, Rubim, NM, Bonavita, AG, Lyra, JSPO, Neves-dos-Santos, S, de Carvalho, ACC, Spray, DC, Savino, W & Alves, LA 2010, 'Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells', BMC Cell Biology, vol. 11, 3. https://doi.org/10.1186/1471-2121-11-3
Nihei, Oscar K. ; Fonseca, Paula C. ; Rubim, Nara M. ; Bonavita, Andre G. ; Lyra, Jurandy S P O ; Neves-dos-Santos, Sandra ; de Carvalho, Antonio C C ; Spray, David C. ; Savino, Wilson ; Alves, Luiz A. / Modulatory effects of cAMP and PKC activation on gap junctional intercellular communication among thymic epithelial cells. In: BMC Cell Biology. 2010 ; Vol. 11.
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abstract = "Background: We investigated the effects of the signaling molecules, cyclic AMP (cAMP) and protein-kinase C (PKC), on gap junctional intercellular communication (GJIC) between thymic epithelial cells (TEC).Results: Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC.Conclusions: Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.",
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AU - Fonseca, Paula C.

AU - Rubim, Nara M.

AU - Bonavita, Andre G.

AU - Lyra, Jurandy S P O

AU - Neves-dos-Santos, Sandra

AU - de Carvalho, Antonio C C

AU - Spray, David C.

AU - Savino, Wilson

AU - Alves, Luiz A.

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N2 - Background: We investigated the effects of the signaling molecules, cyclic AMP (cAMP) and protein-kinase C (PKC), on gap junctional intercellular communication (GJIC) between thymic epithelial cells (TEC).Results: Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC.Conclusions: Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.

AB - Background: We investigated the effects of the signaling molecules, cyclic AMP (cAMP) and protein-kinase C (PKC), on gap junctional intercellular communication (GJIC) between thymic epithelial cells (TEC).Results: Treatment with 8-Br-cAMP, a cAMP analog; or forskolin, which stimulates cAMP production, resulted in an increase in dye transfer between adjacent TEC, inducing a three-fold enhancement in the mean fluorescence of coupled cells, ascertained by flow cytometry after calcein transfer. These treatments also increased Cx43 mRNA expression, and stimulated Cx43 protein accumulation in regions of intercellular contacts. VIP, adenosine, and epinephrine which may also signal through cyclic nucleotides were tested. The first two molecules did not mimic the effects of 8-Br-cAMP, however epinephrine was able to increase GJIC suggesting that this molecule functions as an endogenous inter-TEC GJIC modulators. Stimulation of PKC by phorbol-myristate-acetate inhibited inter-TEC GJIC. Importantly, both the enhancing and the decreasing effects, respectively induced by cAMP and PKC, were observed in both mouse and human TEC preparations. Lastly, experiments using mouse thymocyte/TEC heterocellular co-cultures suggested that the presence of thymocytes does not affect the degree of inter-TEC GJIC.Conclusions: Overall, our data indicate that cAMP and PKC intracellular pathways are involved in the homeostatic control of the gap junction-mediated communication in the thymic epithelium, exerting respectively a positive and negative role upon cell coupling. This control is phylogenetically conserved in the thymus, since it was seen in both mouse and human TEC preparations. Lastly, our work provides new clues for a better understanding of how the thymic epithelial network can work as a physiological syncytium.

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