Association of colorectal cancer with pathogenic Escherichia coli: Focus on mechanisms using optical imaging

AIM: To investigate the molecular or cellular mechanisms related to the infection of epithelial colonic mucosa by pks-positive Escherichia coli(E. coli) using optical imaging.METHODS: We choose to evaluate the tumor metabolic activity using a fluorodeoxyglucose analogue as 2-deoxyglucosone fluorescent probes and to correlate it with tumoral volume(mm^3). Inflammation measuring myeloperoxidase(MPO) activity and reactive oxygen species production was monitored by a bioluminescent(BLI) inflammation probe and related to histological examination and MPO levels by enzyme-linked immunosorbent assay(ELISA) on tumor specimens. The detection and quantitation of these two signals were validated on a xenograft model of human colon adenocarcinoma epithelial cells(HCT116) in nude mice infected with a pks-positive E. coli. The inflammatory BLI signal was validated intra-digestively in the colitisCEABAC10 DSS models, which mimicked Crohn's disease. RESULTS: Using a 2-deoxyglucosone fluorescent probe, we observed a high and specific HCT116 tumor uptake in correlation with tumoral volume(P = 0.0036). Using the inflammation probe targeting MPO, we detected a rapid systemic elimination and a significant increase of the BLI signal in the pks-positive E. coli-infected HCT116 xenograft group(P < 0.005). ELISA confirmed that MPO levels were significantly higher(1556 ± 313.6 vs 234.6 ± 121.6 ng/m L P = 0.001) in xenografts infected with the pathogenic E. coli strain. Moreover, histological examination of tumor samples confirmed massive infiltration of pks-positive E. coli-infected HCT116 tumors by inflammatory cells compared to the uninfected group. These data showed that infection with the pathogenic E. coli strain enhanced inflammation and ROS production in tumors before tumor growth. Moreover, we demonstrated that the intra-digestive monitoring of inflammation is feasible in a reference colitis murine model(CEABAC10/DSS).CONCLUSION: Using BLI and fluorescence optical imaging, we provided tools to better understand hostpathogen interactions at the early stage of disease, such as inflammatory bowel disease and colorectal cancer.

In vivo bioluminescence imaging of hyperglycemia exacerbating stem cells on choroidal neovascularization in mice

AIM： To investigate the influence of hyperglycemia on the severity of choroidal neovascularization（CNV）,especially the involvement of bone marrow-derived cells（BMCs） and underlying mechanisms.·METHODS： BMCs from firefly luciferase（Fluc）/green fluorescent protein（GFP） double transgenic mice were transplanted into C57BL/6J wide-type mice. The recipient mice were injected intraperitoneally with streptozotocin（STZ） daily for 5 consecutive days to induce diabetes mellitus（DM）, followed by CNV laser photocoagulation.The BMCs recruitment in CNV exposed to hyperglycemia was firstly examined in Fluc/GFP chimeric mice by in vivo optical bioluminescence imaging（BLI） and in vitro Fluc assays. The CNV severity was evaluated by H＆E staining and choroidal flatmount. The expression of vascular endothelial growth factor（VEGF） and stromal cell derived factor-1（SDF-1） was detected by Western blot.·RESULTS： BLI showed that the BMCs exerted dynamic effects in CNV model in Fluc/GFP chimeric mice exposed to hyperglycemia. The signal intensity of transplanted Fluc＋GFP＋BMCs in the DM chimeric mice was significantly higher than that in the control chimeric mice with CNV induction at days 5, 7, 14 and 21（121861.67 ±9948.81 vs 144998.33 ±13787.13 photons/second/cm2/sr for control and DM mice, P5d〈0.05; 178791.67±30350.8 vs240166.67 ±22605.3, P7d〈0.05; 124176.67 ±16253.52 vs196376.67 ±18556.79, P14d〈0.05; 97951.60 ±10343.09 vs119510.00 ±14383.76, P21d〈0.05）, which was consistent with in vitro Fluc assay at day 7 [relative light units of Fluc（RLU1）], 215.00±52.05 vs 707.33±88.65, P 〈0.05; RLU1/relative light units of renilla luciferase（RLU2）, 0.90 ±0.17 vs 1.83 ±0.17, P 〈0.05]. The CNVs in the DM mice were wider than those in the control group at days 5, 7, 14 and21（147.83±17.36 vs 220.33±20.17 μm, P5d〈0.05; 212.17 ±24.63 vs 326.83 ±19.49, P7d〈0.05; 163.17 ±18.24 vs265.17 ±20.55, P14d〈0.05; 132.00 ±10.88 vs 205.33 ±12.98,P21d〈0.05）. The average area of CNV in the DM group was larger at 7d（20688.67±3644.96 vs 32218.00±4132.69 μm2,P 〈0.05）. The expression of VEGF and SDF-1 was enhanced in the DM mice.·CONCLUSION： Hyperglycemia promots the vasculo-genesis of CNV, especially the contribution of BMCs,which might be triggered by VEGF and SDF-1 production.

In vivo biodistribution of topical low molecular weight heparin-taurocholate in a neovascularized mouse cornea

AIM： To investigate the ocular biodistribution and clearance of topically administered 7-taurocholic acid conjugated low-molecular weight heparin（LHT7） in a neovascularized mouse cornea using an in vivo optical imaging system. METHODS： A total of 10 eyes of 6 to 8-week-old BALB/c mice were analyzed. Corneal neovascularization（CoNV） was induced in the inferior cornea（IC） of each animal by penetrating the stroma with two interrupted sutures. The development of CoNV was verified after one week and the area of each neovascularized region was measured. A near-infrared fluorescent probe of 20 μmol/L Cy5.5 labeled LHT7（LHT7-Cy5.5） in 0.02 mL solution was topically instilled onto the cornea in the experimental group（n=5）. Free-Cy5.5 of 20 μmol/L in 0.02 mL was instilled in the control group（n=5）. In vivo optical images were obtained before instillation and 5 min, 2, 4, and 6 h after instillation. The intensities were separately measured at the superior cornea（SC） and the IC. RESULTS： The mean CoNV areas were 1.97±0.17 mm^2 and 1.92±0.96 mm^2 in the experimental and control groups, respectively（P=0.832）. The SC remained normal in all 10 subject animals. The IC intensity of the LHT7-Cy5.5 was greater than the SC intensity at 5 min（P=0.038）, 2 h（P=0.041）, and 4 h（P=0.041） after application. The IC intensity fell to less than half of its initial value（42.9%±8.6%） at 6 h in the experimental group. In the control mice, here were no significant differences in the free-Cy5.5 intensity between the IC and SC. CONCLUSION： Topically administered LHT7 shows a high biodistribution in CoNV areas for 4 h and should be reapplied accordingly to maintain its effects. In vivo optical imaging can be a useful tool for evaluating the ocular biodistribution of a drug in an animal model.

Optical coherence tomography imaging of cranial meninges post brain injury in vivo

We report a new application of optical coherence tomography（OCT） to investigate the cranial meninges in an animal model of brain injury in vivo. The injury is induced in a mouse due to skull thinning, in which the repeated and excessive drilling exerts mechanical stress on the mouse brain through the skull, resulting in acute and mild brain injury. Transcranial OCT imaging reveals an interesting virtual space between the cranial meningeal layers post skull thinning, which is gradually closed within hours. The finding suggests a promise of OCT as an effective tool to monitor the mechanical trauma in the small animal model of brain injury.

Bimorph deformable mirror based adaptive optics scanning laser ophthalmoscope for retina imaging in vivo

A bimorph deformable mirror （DM） with a large stroke of more than 30 μm using 35 actuators is presented and characterized for an adaptive optics （AO） confocal scanning laser ophthalmoscope application. Facilitated with a Shack-Hartmann wavefront sensor, the bimorph DM-based AO operates closed-loop AO corrections for hu- man eyes and reduces wavefront aberrations in most eyes to below 0.1 μm rms. Results from living eyes, including one exhibiting ~5D of myopia and ~2D of astigmatism along with notable high-order aberrations, reveal a prac- tical efficient aberration correction and demonstrate a great benefit for retina imaging, including improving resolution, increasing brightness, and enhancing the contrast of images.