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Outlines:

Introducing to organ culture

3–Dimensional culture

Bone culture

Chimera

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What is organ culture?

tomorrowsci 4

• Organ culture refers to the in vitro culture and maintenance of whole or part of an organ in a way that may allow differentiation and preservation of the architecture and/or function.

HHS.gov U.S. Department of Health & Human Services5

The necessity of organ culture

In the United States

Application

Aliya Fatehullah et al; SERIES ON STEM CELL BIOLOGY;2016 6

Features:• Promotion levels of cell differentiation

• Extensive cell proliferation

Types:• Scaffold-free platforms for spheroid growth

• Scaffolds

• Gels

• Bioreactors

• Microchips

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3–Dimensional culture

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Comparison between 3 cultivars

RCCS:

FrancescaCosmi et al; sciencedirect;2015 9

Comparison between 3 cultivars

Yilong Tang et al; nature; 2017 10

RCCS culture 2D & 3D culture

Grow: 2D 3D RCCS

Prolifration & differentiation in RCCS

Yilong Tangm et al; nature; 2017 11

Calcium deposition: 2D 3D RCCS

Bone culture

Fangzheng Li et al;Anatomical sciences education;2017 12

Why Bone culture?

Treatment of extensive bone defect

Limitations of autologous grafts:

morbidity at donor site

Preparation of suitable shape

Limitation of allogeneic grafts:

tissue matching

integrate following implantation

Ck12.org

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Select a suitable cell source

• Capable of forming bone/cartilage

• Have osteogenic potential in older individuals

• Usually BMSCs.

Marot et al; Stem Cell Research & Therapy ;2010 14

Experiment 1:

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• Gene –activated matrics methodology

• In situ tissue engineering

• Transplanting cells simultaneously gene vectors

Porus scaffold

Hang lin et al;scientific reports;2017 16

Robust osteogenesis in BMP group constructs

Hang lin et al;scientific reports;201717

In vivo:

Hang lin et al;scientific reports;201718

Robust osteogenesis in BMP group constructs

Hang lin et al;scientific reports;201719

Lv-GFP

Lv-BMP/GFP

Mineralization Ca deposition

Experiment 2:

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Providing the proper environmental with:

Desired nanoparticle polyethylene (glycol) diacrylate (PEGDA)

Arginine-Glycine-Aspartic acid-Serene (RGDS)

(nHA)

Suitable geometry

Low-intensity pulsed ultrasound (LIPUS)

Zuan zhou et al;scientific reports;2016 21

Xuan zhou et al;scientific reports;2016 22

PEGDA-RGDS

PEGDA-RGDS-nHA

Without LIPUS with LIPUS

PEGDA

Experiment 3 :

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Materials:Carrier:

MS/CPC porous scaffolds

Si for:

Reproduce aortic endothelial cells

Express BFGFR ، TGFBR

MS/CPC scaffold

Recombinant human bone morphogenic protein-2 (rhBMP-2)

For effective bone regeneration

Cuidi li et al;scientific reports;2017 24

vascularisation

Cuidi li et al;scientific reports; 2017 25

Tube formation

mineralization

CPC MS/CPC MS/CPC/rhBMP2

Vascularization

Cuidi li et al;scientific reports; 201726

After 4 weeks

Side of scaffold

longitudinal sections

Osteogenesis:

Cuidi li et al;scientific reports;2017

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After 12 weeks

Chimera

Juan Carlos et al ; Scientifi cAmerican;201628

Factors involved in interspecies chimaerism

A. Matched timing

B. Providing selective

advantage for donor cell

C. Evolutionary consideration

Jun wu et al;nature;2016 29

How to grow human organ in a pig

Juan Carlos et al; Scientifi cAmerican; 2016 30

How to grow human organ in a pig

Juan Carlos et al; Scientifi cAmerican; 201631

Ethical issues of chimeras& solutions

• Issue:

Contribution of stem cells to the development of tissues(neural and germ cells)

• Solutions:Using Mixl1-inducible stem cells

Using progenitor cells

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Using Mixl1-Inducible Mouse PSCs

Toshihiro Kobayashi et al; ORIGINAL RESEARCH REPORT; 2014 33

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Which one is better ?

chimerasOrgan culture

in vitro

Reference :

1.3D Cell Culture 101: An Introduction to 3D Cell Culture Tools and Techniques | Sigma-Aldrich [Internet]. [cited 2017 Sep 30]. Available from: http://www.sigmaaldrich.com/technical-documents/articles/biology/3d-biomatrix-white-paper-3d-cell-culture-101.html

2. Marot D, Knezevic M, Novakovic GV. Bone tissue engineering with human stem cells. Stem Cell Res Ther. 2010 May 4;1(2):10.

3. Huh D, Hamilton GA, Ingber DE. From Three-Dimensional Cell Culture to Organs-on-Chips. Trends Cell Biol. 2011 Dec;21(12):745–54.

4. Improved Human Bone Marrow Mesenchymal Stem Cell Osteogenesis in 3D Bioprinted Tissue Scaffolds with Low Intensity Pulsed Ultrasound Stimulation. - PubMed - NCBI [Internet]. [cited 2017 Oct 3]. Available from: https://www.ncbi.nlm.nih.gov/pubmed/?term=Improved+Human+Bone+Marrow+Mesenchymal+Stem+Cell+Osteogenesis+in+3D+Bioprinted+Tissue+Scaffolds+with+Low+Intensity+Pulsed+Ultrasound+Stimulation

5. Lin H, Tang Y, Lozito TP, Oyster N, Kang RB, Fritch MR, et al. Projection Stereolithographic Fabrication of BMP-2 Gene-activated Matrix for Bone Tissue Engineering. Sci Rep. 2017 Sep 12;7(1):11327.

.

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6. Li C, Jiang C, Deng Y, Li T, Li N, Peng M, et al. RhBMP-2 loaded 3D-printed mesoporoussilica/calcium phosphate cement porous scaffolds with enhanced vascularization and osteogenesisproperties. Sci Rep. 2017 Jan 27;7:41331

7. Stem cells and interspecies chimaeras : Nature : Nature Research [Internet]. [cited 2017 Oct 3]. Available from: http://www.nature.com/nature/journal/v540/n7631/full/nature20573.html

8. Wu J, Greely HT, Jaenisch R, Nakauchi H, Rossant J, Belmonte JCI. Stem cells and interspecies chimaeras. Nature. 2016 01;540(7631):51–9.

9. The combination of three-dimensional and rotary cell culture system promotes the proliferation and maintains the differentiation potential of rat BMSCs | Scientific Reports [Internet]. [cited 2017 Oct 3]. Available from: https://www.nature.com/articles/s41598-017-00087-x

10. Kobayashi T, Kato-Itoh M, Nakauchi H. Targeted organ generation using Mixl1-inducible mouse pluripotent stem cells in blastocyst complementation. Stem Cells Dev. 2015 Jan 15;24(2):182–9.

11. Levine S, Grabel L. The contribution of human/non-human animal chimeras to stem cell

research. Stem Cell Res. 2017 Sep 14;24:128–34.

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THANK YOU

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