A Hybrid Imaging Platform(CT/PET/FMI) for Evaluating Tumor Necrosis and Apoptosis in Real-Time.

Multimodality imaging is an advanced imaging tool for monitoring tumor behavior and therapy in vivo. In this study, we have developed a novel hybrid tri-modality system that includes two molecular imaging methods: positron emission computed tomography (PET) and fluorescence molecular imaging (FMI) and the anatomic imaging modality X-ray computed tomography (CT). The following paper describes the system development. Also, its imaging performance was tested in vitro (phantom) and in vivo, in Balb/c nude mice bearing a head and neck tumor xenograft treated with novel gene therapy [a new approach to the delivery of recombinant bacterial gene (IL-24-expressing strain)]. Using the tri-modality imaging system, we simultaneously monitored the therapeutic effect, including the apoptotic and necrotic induction within the tumor in vivo. The apoptotic induction was examined in real-time using an 18F-ML-10 tracer; the cell death was detected using ICG. A CT was used to evaluate the anatomical situation. An increased tumor inhibition (including tumor growth and tumor cell apoptosis) was observed in the treatment group compared to the control groups, which further confirmed the therapeutic effect of a new IL-24-expressing strain gene therapy on the tumor in vivo. By being able to offer concurrent morphological and functional information, our system is able to characterize malignant tissues more accurately. Therefore, this new tri-modality system (PET/CT/FMI) is an effective imaging tool for simultaneously investigating and monitoring tumor progression and therapy outcomes in vivo.

In vivo long-term investigation of tumor bearing mKate2 by an in-house fluorescence molecular imaging system.

BACKGROUND: Optical imaging is one of the most common, low-cost imaging tools used for investigating the tumor biological behavior in vivo. This study explores the feasibility and sensitivity of a near infrared fluorescent protein mKate2 for a long-term non-invasive tumor imaging in BALB/c nude mice, by using a low-power optical imaging system. METHODS: In this study, breast cancer cell line MDA-MB-435s expressing mKate2 and MDA-MB-231 expressing a dual reporter gene firefly luciferase (fLuc)-GFP were used as cell models. Tumor cells were implanted in different animal body compartments including subcutaneous, abdominal and deep tissue area and closely monitored in real-time. A simple and low-power optical imaging system was set up to image both fluorescence and bioluminescence in live animals. RESULTS: The presence of malignant tissue was further confirmed by histopathological assay. Considering its lower exposure time and no need of substrate injection, mKate2 is considered a superior choice for subcutaneous imaging compared with fLuc. On the contrary, fLuc has shown to be a better option when monitoring the tumor in a diffusive area such as abdominal cavity. Furthermore, both reporter genes have shown good stability and sensitivity for deep tissue imaging, i.e. tumor within the liver. In addition, fLuc has shown to be an excellent method for detecting tumor cells in the lung. CONCLUSIONS: The combination of mKate2 and fLuc offers a superior choice for long-term non-invasive real-time investigation of tumor biological behavior in vivo.

Validation of Bevacizumab Therapy Effect on Colon Cancer Subtypes by Using Whole Body Imaging in Mice.

PURPOSE: Preclinical imaging offers a useful tool for monitoring cancer biological behavior and therapy in vivo without the necessity of animal surgery. The following paper describes our examination of tumor progress and anti-angiogenic therapy with Bevacizumab on colon cancer subtypes (SW480 and SW620) by using different non-invasive real-time in vivo imaging techniques. PROCEDURES: Color Doppler ultrasound imaging (CDUI) was used to observe the formation of new blood vessels; a homemade fluorescence reflectance imaging (FRI) apparatus was mainly used to test the difference in VEGFR2 expression between the tumor subtypes. Briefly, 15 Balb/c nude mice bearing subcutaneous SW480 and SW620 xenografts were randomly divided into Control and Drug groups. Bevacizumab treatment lasted for 3 weeks. All images were captured pre- and post-treatment. At the end of experiment, all mice were euthanized, and tumor tissue was collected and analyzed by immunohistochemical staining. RESULTS: Expression of VEGFR2 was found to be slightly (10 %) but significantly higher for the SW620 cells than for SW480 cells. In addition, SW620 has shown to be more vascularized than SW480 subtype. After 3-week Bevacizumab therapy, no blood vessels were found within 83 % of SW620, while it was 67 % in SW480; the increase of SW620 tumor volume post-treatment was only 3.17-fold compared with the tumor volume pre-treatment, and 4.51-fold higher in SW480. CONCLUSION: Our data suggest that SW480 and SW620 cell lines respond differently to Bevacizumab therapy in vivo. Because of higher vascularization, and subsequently higher reduction by drug of new blood vessels and tumor growth rate, xenografts derived from the metastatic SW620 cell line have a better chance of being successfully treated with Bevacizumab compared with those derived from the primary tumor SW480 cell line.

Overcoming foreign-body reaction through nanotopography: Biocompatibility and immunoisolation properties of a nanofibrous membrane.

Implantable immunoisolation membranes need to possess superior biocompatibility to prohibit the fibrotic deposition that would reduce the nutrient supply and impair the viability/function of the encapsulated cells. Here, electrospun membranes based on thermoplastic polyurethane (TPU) were fabricated to contain microfibers (PU-micro) or nanofibers (PU-nano). The two types of membranes were compared in terms of their interaction with macrophage cells and the host tissues. It was found that the fibrous membranes of different topographies possess distinct material properties: PU-nano caused minimal macrophage responses in vitro and in vivo and induced only mild foreign body reactions compared to PU-micro membranes. A flat macroencapsulation device was fabricated using PU-nano membranes and its immunoisolation function investigated in subcutaneous transplantation models. The nanofibrous device demonstrated the capability to effectively shield the allografts from the immune attack of the host. Nanotopography may confer biocompatibility to materials and nanofibrous materials warrant further study for development of "invisible" immunoisolation devices for cell transplantation.

Establishment of an mKate2-Expressing Cell Line for Non-Invasive Real-Time Breast Cancer In Vivo Imaging.

PURPOSE: Non-invasive real-time in vivo imaging experiments using mice as animal models have become crucial for understanding cancer development and treatment. In this study, we have developed and validated a new breast cancer cell line MDA-MB-435s that stably express a far-red fluorescence protein (mKate2) and that could serve as a highly valuable cell model for studying breast cancer detection and therapy using in vivo fluorescence imaging in nude mice. PROCEDURES: The new cell line (MDA-MB-435s-mKate2) was constructed by plasmid transfection. The stability and sensitivity of mKate2, and the cell biological activities, were tested in vitro using different experimental approaches. For its potential use in tumor growth research and drug therapy in vivo, MDA-MB-435s-mKate2 was validated using the immunocompromised Balb/c nude mice tumor model. In addition, the new cell line has been characterized as a luteinizing hormone-releasing hormone receptor (LHRHR) positive cell line. RESULTS: Firstly, MDA-MB-435s-mKate2 has shown a stable chromosomal integration of the amplified mKate2 gene and good fluorescence sensitivity for detection using a fluorescence reflectance imaging (FRI) device. Compared to its parental cell line, no significant difference in cell migration, proliferation, and clone formation was observed in vitro. Secondly, using the quantification of tumor-fluorescence surface area in live animals, we were able to monitor and detect the tumor progress or tumor inhibition rate (by Paclitaxel treatment) non-invasively and in real-time. Furthermore, MDA-MB-435s-mKate2 has been positively tested for LHRHR; these findings open the possibility to use this cell line for future studies of breast cancer therapy based on LHRH analogs in vivo. CONCLUSION: In the present research, we have successfully built the MDA-MB-435s-mKate2 cell line that can be used as a suitable cell model for breast cancer therapy and anti-cancer drug evaluation by non-invasive fluorescence imaging in mice.

Prospective assessment of the gastroesophageal microbiome in VLBW neonates.

BACKGROUND: The distal GI microbiota of hospitalized preterm neonates has been established to be unique from that of healthy full-term infants; the proximal GI, more specifically gastroesophageal colonization has not been systematically addressed. We prospectively evaluated early colonization of gastroesophageal portion of the GI tract of VLBW infants. METHODS: This study involved 12 infants admitted to a level III NICU with gestational age (GA) 27 +/- 0.5 weeks and birth weight 1105 +/- 77 grams. The gastroesophageal microbial flora was evaluated using 16S rDNA analysis of aspirates collected in a sterile manner during the first 28 days of life. RESULTS: Bacteria were detected in 9 of the 12 neonates. Ureaplasma was the dominant species in the first week of life, however, staphylococci were the predominant bacteria overall. By the fourth week, Gram (-) bacteria increased in abundance to account for 50% of the total organisms. Firmicutes were present in the majority of the neonates and persisted throughout the 4 weeks comprising nearly half of the sequenced clones. Noticeably, only two distinct species of Staphylococcus epidermidis were found, suggesting acquisition from the environment. CONCLUSIONS: In our neonates, the esophagus and stomach environment changed from being relatively sterile at birth to becoming colonized by a phylogenetically diverse microbiota of low individual complexity. By the fourth week, we found predominance of Firmicutes and Proteobacteria. Bacteria from both phyla (CONS and Gram (-) organisms) are strongly implicated as causes of hospital-acquired infections (HAI). Evaluation of the measures preventing colonization with potentially pathogenic and pathogenic microorganisms from the hospital environment may be warranted and may suggest novel approaches to improving quality in neonatal care.