# Current Biomedical Applications of 3D Printing and Additive Manufacturing
## 4 Healthcare Applications
### Tissue Engineering
#### 4.1.1 3D Models and Organoids  
In vitro applications of tissue engineering have paved the way for the development of physiological models representing tissue-like human and animal microenvironments for more clinically applicable testing. Complex three-dimensional organizations of different cell types can allow for the creation of organoids and functional units of different organ systems for studying disease processes and treatment response. It has been shown that cells behave quite differently in terms of gene expression and signaling within a 3D environment, as opposed to conventional 2D cultures [83; 84]. A study by [PERSON] et al. [83] comparing the response of 3D spheroids vs 2D cultures of human colon cancer cells lines observed contradictory levels of AKT/mTOR/S6K signaling activity upon treatment with the mTOR inhibitor rapamycin. They discovered that signaling was increased in 2D cultures, but decreased in 3D spheroids [83]. A subsequent xenografted mouse tumor model demonstrated that the 3D spheroid model was an accurate representation of the in vivo response to rapamycin treatment, showcasing the increased efficacy of 3D spheroids as accurate preclinical drug screening models compared to 2D cultures [83]. Although tumor spheroid models are a relatively inexpensive and a fast way of recreating in vivo-like environments [85], they are limited regarding their size and complexity [86]. Due to limitations concerning diffusion of nutrients, gases and wastes, spheroids can only grow to a certain size before hypoxia and lack of nutrients deteriorates the spheroid viability [86]. Additionally, fabrication methods for spheroid formation only allow for production of simple spheroids, without any design complexity, and they are also very fragile during handling [86]. As an AM technique, bioprinting allows the creation of complex 3D hydrogel models with great levels of control and reproducibility compared with conventional techniques such as molding and spheroid formation. While production of functional organs may still be decades away, bioprinting can already allow the creation of complex organ-on-a-chip models. With good anatomical, compositional and functional similarity to the host tissue, bioprinted 3D models can be used to conduct experiments in an in vivo-like physiological environment [87]. Organ development is a process that begins during the embryonic period, and so recreating the architectural environment and correct signaling patterns developed through this process is important for modeling both developmental and disease processes [88]. Current research efforts have been successful in reproducing functions of many different organs through differentiation and self-organization of stem cells into organoids [89; 90]. Reliance on the self-organizing capabilities of stem cells, however, provides limited control regarding the shape, composition and final size of the organoid [91; 92]. Through bioprinting, stem cells, cell lines and patient derived cells can be deposited with good spatial control to achieve any desired structural arrangement [82; 88; 91; 92; 93]. A study by [PERSON] et al. [94] utilized stereolithography to produce 4 mm 3D liver lobule models. They comprised a hepatocyte-seeded GelMA (methacylated gelatin) with degradable PEG (polyethylene glycol)-lined channels, mimicking the unique in vivo architecture of the liver. They demonstrated increased hepatocyte-specific gene expression within the 3D model as compared to 2D monolayer cultures [94]. Additionally, by embedding channels throughout the model, they created a vascular network that could be used to perfuse the model [94]. The high resolution that stereolithography technology enables allowed printing of microscopic details within this model [94]. Another study by [PERSON] et al. [95] used material extrusion bioprinting technology and a collagen-based bioink to create a vascularized thyroid gland model.