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Vascular disrupting action of electroporation and electrochemotherapy with bleomycin in murine sarcoma.

Sersa G, Jarm T, Kotnik T, Coer A, Podkrajsek M, Sentjurc M, Miklavcic D, Kadivec M, Kranjc S, Secerov A, Cemazar M - Br. J. Cancer (2008)

Bottom Line: The vascular effect was determined by laser Doppler flowmetry, power Doppler ultrasonographic imaging and Patent blue staining.Consequently, electrochemotherapy has, besides the immediate vascular disrupting action, also a delayed one (after 24 h), as a consequence of endothelial cell swelling and apoptosis demonstrated by extensive tumour necrosis, tumour hypoxia, prolonged reduction of tumour blood flow and significant tumour growth delay, and tumour cures.Our results demonstrate that in addition to the well-established direct cytotoxic effect on tumour cells, electrochemotherapy also has an indirect vascular disrupting action resulting altogether in extensive tumour cell necrosis leading to complete regression of tumours.

View Article: PubMed Central - PubMed

Affiliation: Institute of Oncology Ljubljana, Zaloska 2, SI-1000 Ljubljana, Slovenia. gsersa@onko-i.si

ABSTRACT
Electrochemotherapy has a direct cytotoxic effect on tumour cells, and presumably, a vascular disrupting effect. In this study, on the basis of the prediction of the mathematical model, histological evaluation and physiological measurements of the tumours were carried out to confirm that electroporation and electrochemotherapy of tumours have a vascular disrupting action. In the study, SA-1 solid subcutaneous sarcoma tumours in A/J mice were treated by bleomycin (BLM) given intravenously (1 mg kg(-1)), application of electric pulses (8 pulses, 1040 V, 100 micros, 1 Hz) or a combination of both - electrochemotherapy. The vascular effect was determined by laser Doppler flowmetry, power Doppler ultrasonographic imaging and Patent blue staining. The extent of tumour hypoxia was determined immunohistochemically by hypoxia marker pimonidazole and partial pressure of oxygen (pO(2)) in tumours by electron paramagnetic resonance oximetry. Electrochemotherapy with BLM induced good antitumour effect with 22 days, tumour growth delay and 38% tumour cures. The application of electric pulses to the tumours induced instant but transient tumour blood flow reduction (for 70%) that was recovered in 24 h. During this tumour blood flow reduction, we determined an increase in hypoxic tumour area for up to 30%, which was also reflected in reduced tumour oxygenation (for 70%). According to the described mathematical model, endothelial cells lining in tumour blood vessels are exposed to a approximately 40% higher electric field than the surrounding tumour cells, and therefore easily electroporated, allowing access of high BLM concentration to the cytosol. Consequently, electrochemotherapy has, besides the immediate vascular disrupting action, also a delayed one (after 24 h), as a consequence of endothelial cell swelling and apoptosis demonstrated by extensive tumour necrosis, tumour hypoxia, prolonged reduction of tumour blood flow and significant tumour growth delay, and tumour cures. Our results demonstrate that in addition to the well-established direct cytotoxic effect on tumour cells, electrochemotherapy also has an indirect vascular disrupting action resulting altogether in extensive tumour cell necrosis leading to complete regression of tumours.

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Representative tumour sections 90 min after treatment with bleomycin, application of electric pulses or electrochemotherapy. Brown regions are the cells stained with pimonidazole, a marker of tumour hypoxia. Note marked differences in the extent of hypoxic regions between groups without application of electric pulses (control and bleomycin) and those with the application of electric pulses (electric pulses and electrochemotherapy).
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fig7: Representative tumour sections 90 min after treatment with bleomycin, application of electric pulses or electrochemotherapy. Brown regions are the cells stained with pimonidazole, a marker of tumour hypoxia. Note marked differences in the extent of hypoxic regions between groups without application of electric pulses (control and bleomycin) and those with the application of electric pulses (electric pulses and electrochemotherapy).

Mentions: Tumour sections were immunohistochemically stained for protein adduct of reductively activated pimonidazole. Positive staining was found in the cytoplasm of tumour cells and some positive nuclei of tumour cells were also determined. As pimonidazole is known to be preferentially bound by hypoxic tumour cells, the detection of pimonidazole adducts using monoclonal antibodies can serve as a method for measuring tumour hypoxia. In most control tumours, discrete foci of pimonidazole positivity were found. In BLM-treated tumours, a rim of positive cells was found mainly around the necrotic area. In electric pulses and in the electrochemotherapy-treated group of tumours, reticular staining patterns were observed at some distance from blood vessels, with some diffuse pimonidazole-positive area. In some slides from the electrochemotherapy-treated group of tumours, the endothelial cells of blood vessels were also positive. The tumour cells around these pimonidazole-labelled vessels did not show pimonidazole immunoreactivity (Figure 7).


Vascular disrupting action of electroporation and electrochemotherapy with bleomycin in murine sarcoma.

Sersa G, Jarm T, Kotnik T, Coer A, Podkrajsek M, Sentjurc M, Miklavcic D, Kadivec M, Kranjc S, Secerov A, Cemazar M - Br. J. Cancer (2008)

Representative tumour sections 90 min after treatment with bleomycin, application of electric pulses or electrochemotherapy. Brown regions are the cells stained with pimonidazole, a marker of tumour hypoxia. Note marked differences in the extent of hypoxic regions between groups without application of electric pulses (control and bleomycin) and those with the application of electric pulses (electric pulses and electrochemotherapy).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2361464&req=5

fig7: Representative tumour sections 90 min after treatment with bleomycin, application of electric pulses or electrochemotherapy. Brown regions are the cells stained with pimonidazole, a marker of tumour hypoxia. Note marked differences in the extent of hypoxic regions between groups without application of electric pulses (control and bleomycin) and those with the application of electric pulses (electric pulses and electrochemotherapy).
Mentions: Tumour sections were immunohistochemically stained for protein adduct of reductively activated pimonidazole. Positive staining was found in the cytoplasm of tumour cells and some positive nuclei of tumour cells were also determined. As pimonidazole is known to be preferentially bound by hypoxic tumour cells, the detection of pimonidazole adducts using monoclonal antibodies can serve as a method for measuring tumour hypoxia. In most control tumours, discrete foci of pimonidazole positivity were found. In BLM-treated tumours, a rim of positive cells was found mainly around the necrotic area. In electric pulses and in the electrochemotherapy-treated group of tumours, reticular staining patterns were observed at some distance from blood vessels, with some diffuse pimonidazole-positive area. In some slides from the electrochemotherapy-treated group of tumours, the endothelial cells of blood vessels were also positive. The tumour cells around these pimonidazole-labelled vessels did not show pimonidazole immunoreactivity (Figure 7).

Bottom Line: The vascular effect was determined by laser Doppler flowmetry, power Doppler ultrasonographic imaging and Patent blue staining.Consequently, electrochemotherapy has, besides the immediate vascular disrupting action, also a delayed one (after 24 h), as a consequence of endothelial cell swelling and apoptosis demonstrated by extensive tumour necrosis, tumour hypoxia, prolonged reduction of tumour blood flow and significant tumour growth delay, and tumour cures.Our results demonstrate that in addition to the well-established direct cytotoxic effect on tumour cells, electrochemotherapy also has an indirect vascular disrupting action resulting altogether in extensive tumour cell necrosis leading to complete regression of tumours.

View Article: PubMed Central - PubMed

Affiliation: Institute of Oncology Ljubljana, Zaloska 2, SI-1000 Ljubljana, Slovenia. gsersa@onko-i.si

ABSTRACT
Electrochemotherapy has a direct cytotoxic effect on tumour cells, and presumably, a vascular disrupting effect. In this study, on the basis of the prediction of the mathematical model, histological evaluation and physiological measurements of the tumours were carried out to confirm that electroporation and electrochemotherapy of tumours have a vascular disrupting action. In the study, SA-1 solid subcutaneous sarcoma tumours in A/J mice were treated by bleomycin (BLM) given intravenously (1 mg kg(-1)), application of electric pulses (8 pulses, 1040 V, 100 micros, 1 Hz) or a combination of both - electrochemotherapy. The vascular effect was determined by laser Doppler flowmetry, power Doppler ultrasonographic imaging and Patent blue staining. The extent of tumour hypoxia was determined immunohistochemically by hypoxia marker pimonidazole and partial pressure of oxygen (pO(2)) in tumours by electron paramagnetic resonance oximetry. Electrochemotherapy with BLM induced good antitumour effect with 22 days, tumour growth delay and 38% tumour cures. The application of electric pulses to the tumours induced instant but transient tumour blood flow reduction (for 70%) that was recovered in 24 h. During this tumour blood flow reduction, we determined an increase in hypoxic tumour area for up to 30%, which was also reflected in reduced tumour oxygenation (for 70%). According to the described mathematical model, endothelial cells lining in tumour blood vessels are exposed to a approximately 40% higher electric field than the surrounding tumour cells, and therefore easily electroporated, allowing access of high BLM concentration to the cytosol. Consequently, electrochemotherapy has, besides the immediate vascular disrupting action, also a delayed one (after 24 h), as a consequence of endothelial cell swelling and apoptosis demonstrated by extensive tumour necrosis, tumour hypoxia, prolonged reduction of tumour blood flow and significant tumour growth delay, and tumour cures. Our results demonstrate that in addition to the well-established direct cytotoxic effect on tumour cells, electrochemotherapy also has an indirect vascular disrupting action resulting altogether in extensive tumour cell necrosis leading to complete regression of tumours.

Show MeSH
Related in: MedlinePlus