Actually Seeing What Is Going on – Intravital Microscopy in Tissue Engineering

Abstract

Intravital microscopy (IVM) study approach offers several advantages over in vitro, ex vivo, and 3D models. IVM provides real-time imaging of cellular events, which provides us a comprehensive picture of dynamic processes. Rapid improvement in microscopy techniques has permitted deep tissue imaging at a higher resolution. Advances in fluorescence tagging methods enable tracking of specific cell types. Moreover, IVM can serve as an important tool to study different stages of tissue regeneration processes. Furthermore, the compatibility of different tissue engineered constructs can be analyzed. IVM is also a promising approach to investigate host reactions on implanted biomaterials. IVM can provide instant feedback for improvising tissue engineering strategies. In this review, we aim to provide an overview of the requirements and applications of different IVM approaches. First, we will discuss the history of IVM development, and then we will provide an overview of available optical modalities including the pros and cons. Later, we will summarize different fluorescence labeling methods. In the final section, we will discuss well-established chronic and acute IVM models for different organs.

Vessel grafts for tissue engineering revisited—Vessel segments show location‐specific vascularization patterns in ex vivo ring assay

“…Three‐dimensional (3D) ex vivo conditions closely resemble native tissue conditions 12 . Moreover, the cost, reproducibility, and simplicity of the 3D ex vivo condition provide important initial supportive information to reduce the number of experimental animals needed.…”

In Vivo Motility Patterns Displayed by Immune Cells Under Inflammatory Conditions

“…The next step to perform a MP-IVM protocol is to select the proper surgical model, to enable the exposure and immobilization of the targeted organ (Figure 1A) (8,9,55). Although this typically requires minimally invasive surgery, more advanced surgical setups can be employed for long-term imaging of internal organs, including gut (56), brain and spinal cord (57), primary tumors and metastasis (58,59) amongst others (60)(61)(62)(63).…”

Organ Scaffold Characterizations Using Intravital Microscopy

“…Static micrographs and time-series videos were obtained using the multi-photon microscope system. Imaging data was collected on day 0, day 1, day 2, and day 7 time points to examine cell nuclear patterns in native kidneys and scaffolds, changes in innate/decellularized tissue fluorescence levels, and microvascular integrity/function, using well-established techniques 13,15,[18][19][20][21][22][23][24]26,[29][30][31] . To conduct this analysis, we randomly selected 4 regions within a microscopic field and measured the blue and green pseudocolor fluorescence levels in nuclear and epithelial compartments, respectively.…”

“…The fluorescence of native/non-transplanted kidneys and decellularized scaffolds that were transplanted into living patients after introducing Hoechst 33342 exhibited significant differences in fluorescence. This membrane-permeant dye easily penetrates tissues [25][26][27][28][29][30] , where it identifies DNA in living and fixed cells by binding to adenine-thymine-rich sections of DNA in the minor groove, resulting in substantial fluorescence enhancements 31 . The clear imaging of nuclei in endothelia, epithelia, glomeruli, interstitial cells, and circulating leukocytes is known to benefit from such enhancements 28 .…”