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Tumor invasion of Salmonella enterica serovar Typhimurium is accompanied by strong hemorrhage promoted by TNF-alpha.

Leschner S, Westphal K, Dietrich N, Viegas N, Jablonska J, Lyszkiewicz M, Lienenklaus S, Falk W, Gekara N, Loessner H, Weiss S - PLoS ONE (2009)

Bottom Line: A rapid increase of TNF-alpha in blood was observed at that time, in addition to other pro-inflammatory cytokines.In addition, blood influx was followed by necrosis formation, bacterial growth, and infiltration of neutrophilic granulocytes.Depletion of TNF-alpha retarded blood influx and delayed bacterial tumor-colonization.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology, HZI-Helmholtz Centre for Infection Research, Braunschweig, Germany. sara.leschner@helmholtz-hzi.de

ABSTRACT

Background: Several facultative anaerobic bacteria with potential therapeutic abilities are known to preferentially colonize solid tumors after systemic administration. How they efficiently find and invade the tumors is still unclear. However, this is an important issue to be clarified when bacteria should be tailored for application in cancer therapy.

Methodology/principal findings: We describe the initial events of colonization of an ectopic transplantable tumor by Salmonella enterica serovar Typhimurium. Initially, after intravenous administration, bacteria were found in blood, spleen, and liver. Low numbers were also detected in tumors associated with blood vessels as could be observed by immunohistochemistry. A rapid increase of TNF-alpha in blood was observed at that time, in addition to other pro-inflammatory cytokines. This induced a tremendous influx of blood into the tumors by vascular disruption that could be visualized in H&E stainings and quantified by hemoglobin measurements of tumor homogenate. Most likely, together with the blood, bacteria were flushed into the tumor. In addition, blood influx was followed by necrosis formation, bacterial growth, and infiltration of neutrophilic granulocytes. Depletion of TNF-alpha retarded blood influx and delayed bacterial tumor-colonization.

Conclusion: Our findings emphasize similarities between Gram-negative tumor-colonizing bacteria and tumor vascular disrupting agents and show the involvement of TNF-alpha in the initial phase of tumor-colonization by bacteria.

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Related in: MedlinePlus

Time course of bacterial accumulation in different organs and bacterial colonization of solid CT26 tumors.Tumor bearing mice were infected i.v. with S. Typhimurium SL7207. (a) 2 h, 6 h, 12 h, 24 h and 48 h p.i. tumor and spleen were homogenized. Tumor, spleen and blood were plated and the CFUs per total organ were determined. (b) Non-invasive in vivo imaging of bacterial bioluminescence. Tumor-bearing mice were infected with S. Typhimurium expressing the luxCDABE operon of Photorhabdus luminescens under the control of the β-Lactamase promoter. Arrows show sites of high bacterial accumulation. Images were taken at the indicated time points. (c) High magnification images of tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. In all pictures, bacteria are stained in green, blood vessel endothelial cells are stained in blue and neutrophilic granulocytes are stained in red. White arrows point at individual Salmonella. White bars correspond to 10 µm in all pictures. (d) Overviews of CT26 tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. Stainings are described in (c). White bars correspond to 100 µm in all pictures. Experiments were repeated at least three times with identical results.
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pone-0006692-g001: Time course of bacterial accumulation in different organs and bacterial colonization of solid CT26 tumors.Tumor bearing mice were infected i.v. with S. Typhimurium SL7207. (a) 2 h, 6 h, 12 h, 24 h and 48 h p.i. tumor and spleen were homogenized. Tumor, spleen and blood were plated and the CFUs per total organ were determined. (b) Non-invasive in vivo imaging of bacterial bioluminescence. Tumor-bearing mice were infected with S. Typhimurium expressing the luxCDABE operon of Photorhabdus luminescens under the control of the β-Lactamase promoter. Arrows show sites of high bacterial accumulation. Images were taken at the indicated time points. (c) High magnification images of tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. In all pictures, bacteria are stained in green, blood vessel endothelial cells are stained in blue and neutrophilic granulocytes are stained in red. White arrows point at individual Salmonella. White bars correspond to 10 µm in all pictures. (d) Overviews of CT26 tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. Stainings are described in (c). White bars correspond to 100 µm in all pictures. Experiments were repeated at least three times with identical results.

Mentions: Although it is well known that several bacterial strains are able to colonize solid tumors and exert potential anti-tumoricidal effects, the mechanism by which bacteria enter tumors is not clear so far. As this knowledge is essential to render bacteria safe and efficient for anti-cancer therapy we investigated the early time points after systemic Salmonella administration in a murine tumor model. To this end, BALB/c mice bearing the commonly used murine colon carcinoma CT26 were infected i.v. (intravenously) with 5×106 S. Typhimurium SL7207 a strain that is widely used as carrier for vaccine experiments. At different time points p.i. (post infection), colony forming units (CFU) in blood, tumor, and spleen were determined (Fig. 1a). In blood, bacterial numbers quickly decreased within the first 6 h p.i. but then remained constant until the end of the observation period. In spleen, the number of Salmonellae reached a plateau already at 2 h p.i. and showed only minor variations over the observation period. In tumors, bacterial numbers increased constantly from 2 h to 48 h p.i. most likely due to proliferation.


Tumor invasion of Salmonella enterica serovar Typhimurium is accompanied by strong hemorrhage promoted by TNF-alpha.

Leschner S, Westphal K, Dietrich N, Viegas N, Jablonska J, Lyszkiewicz M, Lienenklaus S, Falk W, Gekara N, Loessner H, Weiss S - PLoS ONE (2009)

Time course of bacterial accumulation in different organs and bacterial colonization of solid CT26 tumors.Tumor bearing mice were infected i.v. with S. Typhimurium SL7207. (a) 2 h, 6 h, 12 h, 24 h and 48 h p.i. tumor and spleen were homogenized. Tumor, spleen and blood were plated and the CFUs per total organ were determined. (b) Non-invasive in vivo imaging of bacterial bioluminescence. Tumor-bearing mice were infected with S. Typhimurium expressing the luxCDABE operon of Photorhabdus luminescens under the control of the β-Lactamase promoter. Arrows show sites of high bacterial accumulation. Images were taken at the indicated time points. (c) High magnification images of tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. In all pictures, bacteria are stained in green, blood vessel endothelial cells are stained in blue and neutrophilic granulocytes are stained in red. White arrows point at individual Salmonella. White bars correspond to 10 µm in all pictures. (d) Overviews of CT26 tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. Stainings are described in (c). White bars correspond to 100 µm in all pictures. Experiments were repeated at least three times with identical results.
© Copyright Policy
Related In: Results  -  Collection

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pone-0006692-g001: Time course of bacterial accumulation in different organs and bacterial colonization of solid CT26 tumors.Tumor bearing mice were infected i.v. with S. Typhimurium SL7207. (a) 2 h, 6 h, 12 h, 24 h and 48 h p.i. tumor and spleen were homogenized. Tumor, spleen and blood were plated and the CFUs per total organ were determined. (b) Non-invasive in vivo imaging of bacterial bioluminescence. Tumor-bearing mice were infected with S. Typhimurium expressing the luxCDABE operon of Photorhabdus luminescens under the control of the β-Lactamase promoter. Arrows show sites of high bacterial accumulation. Images were taken at the indicated time points. (c) High magnification images of tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. In all pictures, bacteria are stained in green, blood vessel endothelial cells are stained in blue and neutrophilic granulocytes are stained in red. White arrows point at individual Salmonella. White bars correspond to 10 µm in all pictures. (d) Overviews of CT26 tumor cryosections at the indicated time points p.i. with S. Typhimurium SL7207. Stainings are described in (c). White bars correspond to 100 µm in all pictures. Experiments were repeated at least three times with identical results.
Mentions: Although it is well known that several bacterial strains are able to colonize solid tumors and exert potential anti-tumoricidal effects, the mechanism by which bacteria enter tumors is not clear so far. As this knowledge is essential to render bacteria safe and efficient for anti-cancer therapy we investigated the early time points after systemic Salmonella administration in a murine tumor model. To this end, BALB/c mice bearing the commonly used murine colon carcinoma CT26 were infected i.v. (intravenously) with 5×106 S. Typhimurium SL7207 a strain that is widely used as carrier for vaccine experiments. At different time points p.i. (post infection), colony forming units (CFU) in blood, tumor, and spleen were determined (Fig. 1a). In blood, bacterial numbers quickly decreased within the first 6 h p.i. but then remained constant until the end of the observation period. In spleen, the number of Salmonellae reached a plateau already at 2 h p.i. and showed only minor variations over the observation period. In tumors, bacterial numbers increased constantly from 2 h to 48 h p.i. most likely due to proliferation.

Bottom Line: A rapid increase of TNF-alpha in blood was observed at that time, in addition to other pro-inflammatory cytokines.In addition, blood influx was followed by necrosis formation, bacterial growth, and infiltration of neutrophilic granulocytes.Depletion of TNF-alpha retarded blood influx and delayed bacterial tumor-colonization.

View Article: PubMed Central - PubMed

Affiliation: Molecular Immunology, HZI-Helmholtz Centre for Infection Research, Braunschweig, Germany. sara.leschner@helmholtz-hzi.de

ABSTRACT

Background: Several facultative anaerobic bacteria with potential therapeutic abilities are known to preferentially colonize solid tumors after systemic administration. How they efficiently find and invade the tumors is still unclear. However, this is an important issue to be clarified when bacteria should be tailored for application in cancer therapy.

Methodology/principal findings: We describe the initial events of colonization of an ectopic transplantable tumor by Salmonella enterica serovar Typhimurium. Initially, after intravenous administration, bacteria were found in blood, spleen, and liver. Low numbers were also detected in tumors associated with blood vessels as could be observed by immunohistochemistry. A rapid increase of TNF-alpha in blood was observed at that time, in addition to other pro-inflammatory cytokines. This induced a tremendous influx of blood into the tumors by vascular disruption that could be visualized in H&E stainings and quantified by hemoglobin measurements of tumor homogenate. Most likely, together with the blood, bacteria were flushed into the tumor. In addition, blood influx was followed by necrosis formation, bacterial growth, and infiltration of neutrophilic granulocytes. Depletion of TNF-alpha retarded blood influx and delayed bacterial tumor-colonization.

Conclusion: Our findings emphasize similarities between Gram-negative tumor-colonizing bacteria and tumor vascular disrupting agents and show the involvement of TNF-alpha in the initial phase of tumor-colonization by bacteria.

Show MeSH
Related in: MedlinePlus