SC6: In vitro, in vivo, and in situ tissue engineering: pros and cons

Written by: Hossein Rhiahinezhad (Chair), Gad Sabbatier (Clerk), and Laura McKiel (editor)

Our last short science club addressed the expansive topic of tissue engineering strategies. Tissue engineering (TE) consists of multiple principles, techniques, and methods from engineering and life science to understand the relationships between structures and properties of normal and pathological tissues. This multidisciplinary approach allows us to design and develop biological substitutes to restore, maintain, or improve tissue functions. Traditional TE techniques take a processed material scaffold, seed it with mammalian cells, and add biological, chemical, or physical stimuli to support tissue growing and differentiation. Cells that are isolated for tissues and organs can be mature, embryonic, or induced pluripotent, or multipotent stem cells, or differentiated cell types.

These are some examples of the diverse scaffolding strategies that are currently available:

  • Pre-made porous scaffold can be processed and then cell-seeded (e.g. fiber scaffold, foams, films).
  • Human tissue can be decellularized using chemical agents and then cell-seeded with treated cells.
  • Cells can be cultured to produce ECM and cell sheets. Sheets are harvested and assembled layer-by-layer to produce tissues and organs.
  • Polymer solution can be directly mixed with cells and crosslink either physically or chemically and can be used as 3D culture environment.

 

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Figure 1. Scaffolding strategies. Source: Chan BP. European Spine Journal, 2008; 17(Suppl 4), 467-479.

Tissue engineering can use in vitro, in vivo, or in situ strategies to construct tissues and organs. In in vitro TE, cells are seeded in a bio-instructive scaffold and cultured in a static (incubator) or dynamic (bioreactor) environment. The culture media is generally supplemented with biomolecules such as growth factors to differentiate stem cells into the desired cell type. A typical example is illustrated in Figure 2 for using TE to create blood vessels. Autologous smooth muscle cells, fibroblasts, and endothelial cells are harvested from the patient, and the cells are seeded in a tubular scaffold such as collagen or nanofibers and cultured in a bioreactor for a certain amount of time.

 

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Figure 2. Blood vessel tissue engineering. Source: Seifu DG et al. Nat Rev Cardiol, 2013; 10(7), 410–21.

In In vivo TE, the scaffold is implanted usually with cells and an animal is used as an incubator to grow the tissue or the organ before being re-implanted in the same or another patient. Figure 3 shows the impressive example of a human ear scaffold seeded with cow cells implanted in an immune deprived mouse. (Editor’s note: There was a huge controversy when this image was released to the media. You can read more about it here.)

 

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Figure 3. The Vacanti mouse. Original article: Y. Cao et al. Plast Reconstr Surg, 1997; 100(2), 297-302.

In in situ TE, the scaffold is implanted or injected with or without cells into the patient’s (or animal’s) body. The tissue is expected to self-repair due to cell migration and cells growing directly in the body’s environment, such as in the example illustrated in Figure 4.

 

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Figure 4. Cell transplantation with a cell instructive ribbons structure. Source: Sahar Salehi, ACS Biomaterials Science & Engineering, 2017; 3(4), 579-589.

 

The science club attendees were then divided into two groups to discuss two different scenarios:

Group 1: You are asked to prepare safety data sheets for 20 different chemicals and you need to check if human skin cells survive or die in presence of chemicals.

  • Design an experiment (in vitro, in vivo, or both)
  • State your reasons why you prefer one to the other, or why you need both

 

Group 1 suggested to do a viability experiment (MTT, WST-1, Resazurin salt, etc) and put human cells in contact with chemicals at different concentration in a high throughput screening fashion experiment. The experiment could respect the ASTM standards F895 and ASTM F813.

In vitro testing for cell or tissue growth offers less animal testing, cheaper experiments than in vivo, is faster, and can be used extensively. However, in vitro testing is very simplified compared to the complexity of the body, leading to many inaccuracies.

 

Group 2: A patient is suffering from osteoarthritis (degeneration of cartilage in joint) and you are asked to use TE to regenerate his/her cartilage. You have two options:

  1. Using scaffold for in vitro TE followed by implantation (in vivo).
  2. Using in situ TE by injection of cells and polymer solution (using as scaffold).
  • Which one do you prefer and why?
  • What are the challenges of each method?

 

Group 2 preferred approach B. In vitro TE option offers a better control of cell invasion, but is made without considering the immune system. It must survive to severe mechanical stimulation of the joint, as well as be resistant to infection after an open surgery. Option B requires a highly cytocompatible hydrogel that is able to crosslink in seconds, in order to be properly injected though a minimally invasive surgery.

 

Following discussion of the advantages and disadvantages of different tissue engineering methods, here is a summary of our conclusions:

in vitro in vivo in situ
Pros • Able to perform extensive studies with many cell types and testing conditions

• Less animals sacrificed for studies

·       More reliable results compared to in vitro studies • Less invasive (injection)

• More accurate results compared to in vitro and in vivo studies

• Less possibility of infection

Cons •   Not an accurate simulation of the body’s environment

•   Usually using cell lines, which do not accurately portray primary cells

• Potential for contamination

• High costs

• Ethical issues (think about animal study)

• Potential for infection

• Animal cells (and body systems) are very different from humans

• No control of scaffold once injected

• Ethical issues related to trials in humans

• Difficult to monitor over time

 

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