J Korean Neurosurg Soc.  2014 Mar;55(3):131-135. 10.3340/jkns.2014.55.3.131.

Targeting Orthotopic Glioma in Mice with Genetically Engineered Salmonella typhimurium

• Wen M
• Jung S
• Moon KS
• Jiang SN
• Li SY
• Min JJ

• KMID: 2191077
• DOI: http://doi.org/10.3340/jkns.2014.55.3.131

Abstract

OBJECTIVE
With the growing interests of bacteria as a targeting vector for cancer treatment, diverse genetically engineered Salmonella has been reported to be capable of targeting primary or metastatic tumor regions after intravenous injection into mouse tumor models. The purpose of this study was to investigate the capability of the genetically engineered Salmonella typhimurium (S. typhimurium) to access the glioma xenograft, which was monitored in mouse brain tumor models using optical bioluminescence imaging technique.
METHODS
U87 malignant glioma cells (U87-MG) stably transfected with firefly luciferase (Fluc) were implanted into BALB/cAnN nude mice by stereotactic injection into the striatum. After tumor formation, attenuated S. typhimurium expressing bacterial luciferase (Lux) was injected into the tail vein. Bioluminescence signals from transfected cells or bacteria were monitored using a cooled charge-coupled device camera to identify the tumor location or to trace the bacterial migration. Immunofluorescence staining was also performed in frozen sections of mouse glioma xenograft.
RESULTS
The injected S. typhimurium exclusively localized in the glioma xenograft region of U87-MG-bearing mouse. Immunofluorescence staining also demonstrated the accumulation of S. typhimurium in the brain tumors.
CONCLUSION
The present study demonstrated that S. typhimurium can target glioma xenograft, and may provide a potentially therapeutic probe for glioma.

Keyword

Cancer targeting; Mouse glioma model; Optical bioluminescence imaging; Salmonella typhimurium; U87-MG

MeSH Terms

Animals
Bacteria
Brain Neoplasms
Fireflies
Fluorescent Antibody Technique
Frozen Sections
Glioma*
Heterografts
Injections, Intravenous
Luciferases
Mice*
Mice, Nude
Salmonella
Salmonella typhimurium*
Veins
Luciferases

Figure

• 

  Fig. 1 Visualization of tumor development on optical bioluminescence imaging. To evaluate tumor growth in U87-Fluc-bearing nude mouse, a cooled CCD camera was used to analyze the mouse brain tumor model after intraperitoneal injection of 3 mg of D-luciferin per animal. A : The signal gradually increased in brain up to a peak at approximately 30 min, after which time, the signal slowly decayed. B : Bioluminescence images of U87-Fluc tumor developed in mouse over POD 10 and gradually increased over the time. POD : post-operative day, CCD : charge-coupled device.

• 

  Fig. 2 Bioluminescence imaging of S. typhimurium-ΔppGpp-Lux targeting in the U87-Fluc mouse glioma model. A : In vivo bioluminescence imaging was performed on a nude mouse with U87-Fluc stably expressing firefly luciferase in the left stiatum. (a) After 15 days of celluluar inoculation, tumor growth was demonstrated by intraperitonealy injection of D-lucferin. (b, c, and d) And then the S. typhimurium-ΔppGpp-Lux (3×107 CFU) was administered intravenously via the tail vein in POD 15, and exclusively proliferated in the tumor region at 5 dpi (POD 20). Animals were killed 9 days post-beacteral injection and brain tissue was examined histologically using Hematoxylin & Eosin (B) and immunofluorescence (C) staining. Hematoxylin & Eosin staining shows brain tumor and opposite normal brain. Immunofluorescence staining demonstated S. typhimurium in the corresponding area (small black box in B) of tumor growth, not in normal brain (small white box in B) (a and e : H&E, b and f : DAPI, c and g : anti-salmonella, d and h : merged ×200). POD : post-operative day, dpi : days after post-inoculation.

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