Ultra high frequency ultrasound enables real-time visualization of blood supply from chorioallantoic membrane to human autosomal dominant polycystic kidney tissue.

Ultra high frequency (UHF) ultrasound enables the visualization of very small structures that cannot be detected by conventional ultrasound. The utilization of UHF imaging as a new imaging technique for the 3D-in-vivo chorioallantoic membrane (CAM) model can facilitate new insights into tissue perfusion and survival. Therefore, human renal cystic tissue was grafted onto the CAM and examined using UHF ultrasound imaging. Due to the unprecedented resolution of UHF ultrasound, it was possible to visualize microvessels, their development, and the formation of anastomoses. This enabled the observation of anastomoses between human and chicken vessels only 12 h after transplantation. These observations were validated by 3D reconstructions from a light sheet microscopy image stack, indocyanine green angiography, and histological analysis. Contrary to the assumption that the nutrient supply of the human cystic tissue and the gas exchange happens through diffusion from CAM vessels, this study shows that the vasculature of the human cystic tissue is directly connected to the blood vessels of the CAM and perfusion is established within a short period. Therefore, this in-vivo model combined with UHF imaging appears to be the ideal platform for studying the effects of intravenously applied therapeutics to inhibit renal cyst growth.

Utilisation of Chick Embryo Chorioallantoic Membrane as a Model Platform for Imaging-Navigated Biomedical Research.

The fertilised chick egg and particularly its chorioallantoic membrane (CAM) have drawn continuing interest in biomedicine and bioengineering fields, especially for research on vascular study, cancer, drug screening and development, cell factors, stem cells, etc. This literature review systemically introduces the CAM's structural evolution, functions, vascular features and the circulation system, and cell regulatory factors. It also presents the major and updated applications of the CAM in assays for pharmacokinetics and biodistribution, drug efficacy and toxicology testing/screening in preclinical pharmacological research. The time course of CAM applications for different assays and their advantages and limitations are summarised. Among these applications, two aspects are emphasised: (1) potential utility of the CAM for preclinical studies on vascular-disrupting agents (VDAs), promising for anti-cancer vascular-targeted therapy, and (2) modern imaging technologies, including modalities and their applications for real-time visualisation, monitoring and evaluation of the changes in CAM vasculature as well as the interactions occurring after introducing the tested medical, pharmaceutical and biological agents into the system. The aim of this article is to help those working in the biomedical field to familiarise themselves with the chick embryo CAM as an alternative platform and to utilise it to design and optimise experimental settings for their specific research topics.

Effects of topical prostaglandin drops on angiogenesis in an in ovo chick chorioallantoic membrane model.

BACKGROUND: To investigate the effects of bimatoprost, latanoprost and travoprost on angiogenesis in the chick chorioallantoic membrane (CAM) model in ovo. MATERIALS AND METHODS: Fifty fertilized specific-pathogen-free chick eggs were used in this preclinical, prospective, experimental embryo study. Eggs were randomly distributed into 5 groups of ten eggs. Eggs were placed in the incubator after disinfection of their shells with alcohol and monitored appropriate temperature and humidity. On the 3rd day of incubation, a small window was opened on the eggshell. Bimatoprost in group 1, latanoprost in group 2, travoprost in group 3, bevacizumab in group 4, phosphate-buffered-saline (PBS) used in group 5 was applied by injection to CAM. The sterile film was glued onto the broken part of the shell and the eggs were placed in the incubator again. On the 8th day of incubation, eggs were opened and vascular structures on CAMs were examined. Digital photographs were taken, analysed in the ImageJ open source image processing software and differences between groups were evaluated. Thereafter, VEGF (Vascular endothelial growth factor) levels were measured appropriately in the embryo samples. RESULTS: All embryos in the prostaglandin groups and the PBS control group were observed to have life signs confirmed by heart rate. In 8 embryos in the bevacizumab group, no life signs were confirmed, while 2 embryos with life signs showed severe hypoplasia. Vascular density, number of vessels and VEGF levels in the bimatoprost, latanoprost and travoprost groups, there were statistically significantly higher than the PBS control group. CONCLUSION: This study demonstrates that topical prostaglandin drops increase angiogenesis in the chick CAM model in ovo.

Tumor grafts grown on the chicken chorioallantoic membrane are distinctively characterized by MRI under functional gas challenge.

Recently, a tumor model based on the chorioallantoic membrane (CAM) was characterized structurally with Magnetic Resonance Imaging (MRI). Yet, capability of MRI to assess vascular functional reserve and potential of oxygenation-sensitive MRI remain largely unexplored in this model. For this purpose, we compared MC-38 colon and A549 lung adenocarcinoma cell grafts grown on the CAM, using quantitative T1 and T2* MRI readouts as imaging markers. These are associated with vascular functionality and oxygenation status when compared between periods of air and carbogen exposure. Our data show that in A549 lung adenocarcinoma cell grafts T2* values increased significantly upon carbogen exposure (p < 0.004, Wilcoxon test; no change in T1), while MC-38 grafts displayed no changes in T1 and T2*), indicating that the grafts differ in their vascular response. Heterogeneity with regard to T1 and T2* distribution within the grafts was noted. MC-38 grafts displayed larger T1 and T2* in the graft centre, while in A549 they were distributed more towards the graft surface. Finally, qualitative assessment of gadolinium-enhancement suggests that A549 grafts display more prominent enhancement compared to MC-38 grafts. Furthermore, MC-38 grafts had 65% larger volumes than A549 grafts. Histology revealed distinct underlying phenotypes of the two tumor grafts, pertaining to the proliferative status (Ki-67) and cellularity (H&E). In sum, a functional gas challenge with carbogen is feasible through gas exchange on the CAM, and it affects MRI signals associated with vascular reactivity and oxygenation status of the tumor graft planted on the CAM. Different grafts based on A549 lung adenocarcinoma and MC-38 colon carcinoma cell lines, respectively, display distinct phenotypes that can be distinguished and characterized non-invasively in ovo using MRI in the living chicken embryo.

The CAM cancer xenograft as a model for initial evaluation of MR labelled compounds.

Non-invasive assessment of the biodistribution is of great importance during the development of new pharmaceutical compounds. In this contribution, the applicability of in ovo MRI for monitoring the biodistribution of MR contrast agent-labelled compounds was investigated in mamaria carcinomas xentotransplanted on the chorioallantoic membrane (CAM) exemplarily for Gd-DOTA and cHSA-PEO (2000)16-Gd after systemic injection of the compounds into a chorioallantoic capillary vein. MRI was performed directly prior and 30 min, 3 h, 5 h, 20 h, and 40 h after injection of the compound. The biodistribution of injected compounds could be assessed by MRI in different organs of the chicken embryo as well as in xenotransplanted tumors at all time points. A clearly prolonged enhancement of the tumor substrate could be shown for cHSA-PEO (2000)16-Gd. In conclusion, high-resolution in ovo MR imaging can be used for assessment of the in vivo biodistribution of labelled compounds, thus enabling efficient non-invasive initial testing.

Methods for Analyzing Tumor Angiogenesis in the Chick Chorioallantoic Membrane Model.

Models of tumor angiogenesis have played a critical role in understanding the mechanisms involved in the recruitment of vasculature to the tumor mass, and have also provided a platform for testing antiangiogenic potential of new therapeutics that combat the development of malignant growth. In this regard, the chorioallantoic membrane (CAM) of the developing chick embryo has proven to be an elegant model for investigation of angiogenic processes. Here, we describe methods for effectively utilizing the preestablished vascular network of the chick CAM to investigate and quantify tumor-associated angiogenesis in a breast tumor model.

In vivo PET/CT in a human glioblastoma chicken chorioallantoic membrane model: a new tool for oncology and radiotracer development.

UNLABELLED: For many years the laboratory mouse has been used as the standard model for in vivo oncology research, particularly in the development of novel PET tracers, but the growth of tumors on chicken chorioallantoic membrane (CAM) provides a more rapid, low cost, and ethically sustainable alternative. For the first time, to our knowledge, we demonstrate the feasibility of in vivo PET and CT imaging in a U87 glioblastoma tumor model on chicken CAM, with the aim of applying this model for screening of novel PET tracers. METHODS: U87 glioblastoma cells were implanted on the CAM at day 11 after fertilization and imaged at day 18. A small-animal imaging cell was used to maintain incubation and allow anesthesia using isoflurane. Radiotracers were injected directly into the exposed CAM vasculature. Sodium (18)F-fluoride was used to validate the imaging protocol, demonstrating that image-degrading motion can be removed with anesthesia. Tumor glucose metabolism was imaged using (18)F-FDG, and tumor protein synthesis was imaged using 2-(18)F-fluoro-l-tyrosine. Anatomic images were obtained by contrast-enhanced CT, facilitating clear delineation of the tumor, delineation of tracer uptake in tumor versus embryo, and accurate volume measurements. RESULTS: PET imaging of tumor glucose metabolism and protein synthesis was successfully demonstrated in the CAM U87 glioblastoma model. Catheterization of CAM blood vessels facilitated dynamic imaging of glucose metabolism with (18)F-FDG and demonstrated the ability to study PET tracer uptake over time in individual tumors, and CT imaging improved the accuracy of tumor volume measurements. CONCLUSION: We describe the novel application of PET/CT in the CAM tumor model, with optimization of typical imaging protocols. PET imaging in this valuable tumor model could prove particularly useful for rapid, high-throughput screening of novel radiotracers.