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Critical role of gap junction communication, calcium and nitric oxide signaling in bystander responses to focal photodynamic injury.

Calì B, Ceolin S, Ceriani F, Bortolozzi M, Agnellini AH, Zorzi V, Predonzani A, Bronte V, Molon B, Mammano F - Oncotarget (2015)

Bottom Line: Here we show that photosentizer activation in a single cell triggers apoptosis in bystander cancer cells, which are electrically coupled by gap junction channels and support the propagation of a Ca2+ wave initiated in the irradiated cell.The latter also acts as source of nitric oxide (NO) that diffuses to bystander cells, in which NO levels are further increased by a mechanism compatible with Ca(2+)-dependent enzymatic production.Pharmacological blockade of connexin channels significantly reduced the extent of apoptosis in bystander cells, consistent with a critical role played by intercellular communication, Ca2+ and NO in the bystander effects triggered by photodynamic therapy.

View Article: PubMed Central - PubMed

Affiliation: Foundation for Advanced Biomedical Research, Venetian Institute of Molecular Medicine, Padua, Italy.

ABSTRACT
Ionizing and nonionizing radiation affect not only directly targeted cells but also surrounding "bystander" cells. The underlying mechanisms and therapeutic role of bystander responses remain incompletely defined. Here we show that photosentizer activation in a single cell triggers apoptosis in bystander cancer cells, which are electrically coupled by gap junction channels and support the propagation of a Ca2+ wave initiated in the irradiated cell. The latter also acts as source of nitric oxide (NO) that diffuses to bystander cells, in which NO levels are further increased by a mechanism compatible with Ca(2+)-dependent enzymatic production. We detected similar signals in tumors grown in dorsal skinfold chambers applied to live mice. Pharmacological blockade of connexin channels significantly reduced the extent of apoptosis in bystander cells, consistent with a critical role played by intercellular communication, Ca2+ and NO in the bystander effects triggered by photodynamic therapy.

No MeSH data available.


Related in: MedlinePlus

Percentage of apoptotic cells at three time points following focal photodynamic injury in control conditions and in the presence of FFAData are mean ± s.e.m. from n = 3 cultures in each condition.
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Figure 7: Percentage of apoptotic cells at three time points following focal photodynamic injury in control conditions and in the presence of FFAData are mean ± s.e.m. from n = 3 cultures in each condition.

Mentions: The results presented so far support the notion that coupling through gap junctions significantly contributes to the ΔCa2+ and ΔNO signals evoked by focal photodynamic injury, by permitting cell-to-cell propagation of ER− and IP3R−related Ca2+ signals through the network of bystander cells. To evaluate the relevance of gap junction communication also for the apoptotic effects triggered by focal photodynamic injury, we performed additional experiments in C26GM cultures co-loaded with AlClPc and fura-2. At the end of laser irradiation, we switched from Ca2+ imaging to a time-lapse protocol based on staining with propidium iodide and pSIVA-IANBD (Figure 6), an annexin-based polarity sensitive probe for the spatiotemporal or kinetic analysis of apoptosis [57]. Under control conditions, the irradiated cell exhibited detectable pSIVA-IANBD signals as soon as 30 min after focal photodynamic injury. Both pSIVA-IANBD and propidium iodide signals became detectable in the irradiated cells and the nearest neighbours within 1 h, and reached the limits of the field of view within 3 h (Figure 6a). These processes were greatly attenuated and slowed down by the gap junction blocker FFA (Figure 6b). No toxicity was observed in C26GM cultures loaded with AlClPc but not exposed to laser irradiation at 671 nm (Figure 6c). The fraction of C26GM cells exhibiting apoptotic signals as a consequence of focal photodynamic injury increased almost linearly with time, exceeding 15% of the population within 3h; this apoptotic rate was significantly reduced, by a factor >2.5, in the presence of FFA (Figure 7).


Critical role of gap junction communication, calcium and nitric oxide signaling in bystander responses to focal photodynamic injury.

Calì B, Ceolin S, Ceriani F, Bortolozzi M, Agnellini AH, Zorzi V, Predonzani A, Bronte V, Molon B, Mammano F - Oncotarget (2015)

Percentage of apoptotic cells at three time points following focal photodynamic injury in control conditions and in the presence of FFAData are mean ± s.e.m. from n = 3 cultures in each condition.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4496347&req=5

Figure 7: Percentage of apoptotic cells at three time points following focal photodynamic injury in control conditions and in the presence of FFAData are mean ± s.e.m. from n = 3 cultures in each condition.
Mentions: The results presented so far support the notion that coupling through gap junctions significantly contributes to the ΔCa2+ and ΔNO signals evoked by focal photodynamic injury, by permitting cell-to-cell propagation of ER− and IP3R−related Ca2+ signals through the network of bystander cells. To evaluate the relevance of gap junction communication also for the apoptotic effects triggered by focal photodynamic injury, we performed additional experiments in C26GM cultures co-loaded with AlClPc and fura-2. At the end of laser irradiation, we switched from Ca2+ imaging to a time-lapse protocol based on staining with propidium iodide and pSIVA-IANBD (Figure 6), an annexin-based polarity sensitive probe for the spatiotemporal or kinetic analysis of apoptosis [57]. Under control conditions, the irradiated cell exhibited detectable pSIVA-IANBD signals as soon as 30 min after focal photodynamic injury. Both pSIVA-IANBD and propidium iodide signals became detectable in the irradiated cells and the nearest neighbours within 1 h, and reached the limits of the field of view within 3 h (Figure 6a). These processes were greatly attenuated and slowed down by the gap junction blocker FFA (Figure 6b). No toxicity was observed in C26GM cultures loaded with AlClPc but not exposed to laser irradiation at 671 nm (Figure 6c). The fraction of C26GM cells exhibiting apoptotic signals as a consequence of focal photodynamic injury increased almost linearly with time, exceeding 15% of the population within 3h; this apoptotic rate was significantly reduced, by a factor >2.5, in the presence of FFA (Figure 7).

Bottom Line: Here we show that photosentizer activation in a single cell triggers apoptosis in bystander cancer cells, which are electrically coupled by gap junction channels and support the propagation of a Ca2+ wave initiated in the irradiated cell.The latter also acts as source of nitric oxide (NO) that diffuses to bystander cells, in which NO levels are further increased by a mechanism compatible with Ca(2+)-dependent enzymatic production.Pharmacological blockade of connexin channels significantly reduced the extent of apoptosis in bystander cells, consistent with a critical role played by intercellular communication, Ca2+ and NO in the bystander effects triggered by photodynamic therapy.

View Article: PubMed Central - PubMed

Affiliation: Foundation for Advanced Biomedical Research, Venetian Institute of Molecular Medicine, Padua, Italy.

ABSTRACT
Ionizing and nonionizing radiation affect not only directly targeted cells but also surrounding "bystander" cells. The underlying mechanisms and therapeutic role of bystander responses remain incompletely defined. Here we show that photosentizer activation in a single cell triggers apoptosis in bystander cancer cells, which are electrically coupled by gap junction channels and support the propagation of a Ca2+ wave initiated in the irradiated cell. The latter also acts as source of nitric oxide (NO) that diffuses to bystander cells, in which NO levels are further increased by a mechanism compatible with Ca(2+)-dependent enzymatic production. We detected similar signals in tumors grown in dorsal skinfold chambers applied to live mice. Pharmacological blockade of connexin channels significantly reduced the extent of apoptosis in bystander cells, consistent with a critical role played by intercellular communication, Ca2+ and NO in the bystander effects triggered by photodynamic therapy.

No MeSH data available.


Related in: MedlinePlus