Bioprinting for osteochondral tissue regeneration · •A model of an organ is not a functional...

Bioprinting for

skeletal tissue regeneration

Current strategies and future perspectives

Veerle Bloemen

Biofabrication Lab

Faculty of Engineering Technology

Prometheus - Division of Skeletal TissueEngineering

Faculty of Engineering Technology2

Will we soon be printing organs?

• A model of an organ is not a functional organ

• Bioprinting involves biological material such as living cells or

proteins

• An organ has a complex architecture related to its function

A few important remarks

YES? -> NO

There is a need

Data from optn.transplant.hrsa.gov and OPTN/SRTR Annual Report.

Critical skeletal defects

Critical bone defects

Osteochondral defects

Ho-Shui-Ling et al., 2018

Current treatments

• Autografts

• Allografts

• Xenografts

Tissue transplants

Material-based implants

• Metal implants

• Ceramic implants

Several limitations:

- Minimal tissue availability

- Donor side morbidity

- Wear -> 2nd surgery

- Limited implant integration

(replacement strategies)

From replacement towards regeneration

The traditional concept of Tissue Engineering

The development of

cell-based implants for

tissue regeneration

Skeletal tissue engineering

Dvir et.al., 2011

Increase robustness

• Implanted cells significantly contribute to tissue regeneration

• A biomimetic approach improves tissue formation: using engineering

strategies based on principles in developmental biology

• Large variability in the in vivo outcome

• One size does not fit all

Insights from traditional TE-results

Advanced Therapy Medicinal Products

Tissue Engineering approaches are challenged in terms of reproducibility

and clinical relevance of the cell-based product

Journal of market access & health policy,

sCTMP: somatic cell therapy medicinal product

GTMP: gene therapy medicinal product

TEP: tissue engineered product

Combined products: cellular/tissue part + medical device

Top-down versus bottom-up Tissue Engineering

Bioprinting as a tool for the precise fabrication of

complex architectures

The process of bioprinting

Bioprinting for

osteochondral regeneration

Osteochondral tissue

Nukavarapu et al. 2013 Biotechnol Adv

Subchondral

Articular

cartilage

! Articular cartilage is avascular and aneural !

• Articular cartilage

• Superficial zone (A)

• Middle zone (B)

• Deep zone (C)

Calcified cartilage

• Subchondral bone (D)

Osteochondral tissue

Matrix stiffness

2000 kPa

100 kPa

Cell density

24 ∙ 106 cells/mL

7 ∙ 106 cells/mL

Schinagl et al. 1997 , Hunziker et al. 2002

Superficial

zone (A)

Middle (B)

Deep zone (C)

Calcified

cartilage

Subchondral (D)

Compressive modulus 380 kPa

10 million cells/mL

Bioprinting for osteochondral regeneration

Inkjet-based bioprinting of a cartilage construct

Modified HP Deskjet 500 thermal inkjet printer

10 % PEGDMA in PBS

0,05 % I-2959

5 x 106 cells/mL human chondrocytes

ᴓ 4 mm x 2 mm

Cui et al , 2012

Lower cytotoxicity: cell viability 89% vs 63%

Maintained position and phenotype

Distribution of fluorescently labeled chondrocytes in PEG

Scalebar = 100µm

Safranin-O staining after 6 weeks in chondrogenic medium.

Scale bar = 200µm

Construct printed in mold

Scalebar = 2 mm

• Simultaneous photopolymerisation

• Printed in bovine osteochondral plugs more GAG/DNA than without plug

Cui et al , 2012

Multilayered constructs containing human MSCs for osteochondral tissue regeneration in rabbits

In-house built extrusion bioprinter

4 % alginate OR 5% CB[6]DAH-HA in DMEM

3% atelocollagen in DMEM

2 or 1 x106 cells/mL human MSCs

100 ng/mL TGF-β or 5 µg/mL rhBMP-2

ᴓ 5 x 5 mm

4 experimental groups, cultured in DMEM for 24h, then 8 weeks in vivo

• Cell viability after printing atelocollagen

CB[6]/DAH-HA

• Mechanically stabilized atelocollagen gel and new crosslinked HA hydrogel with MSCs and

growth factors promote heterogeneous neotissue formation in vivo

Commercial systems

Organovo – Novogen MMX EnvisionTec – 3D-Bioplotter RegenHu - Biofactory

Cellink Bio-X Allevi (former biobots)

Biofabrication of spatially organised tissues

Melt electrowriting (MEW) for the reinforcement

of hydrogels with 3D printed microfibers

Bio-ink development for osteochondral

regeneration: preliminary in-house data3 mm/s 5 mm/s 7 mm/s

145kPa

160kPa

175kPa

chondrocytes

Bioprinting for long bone healing

Developmentally engineered callus organoid

bioassemblies for long bone healing

3D printing as a route for upscaling?

McMaster et al , 2019

Arai et al , 2018

The “Kenzan” method

Sheet-like tissue constructs

from spheroids and MEW-scaffolds

From bioprinting to biofabrication

‘the automated generation of biologically functional products with structural organization

from living cells, bioactive molecules, biomaterials, cell aggregates such as micro-tissues,

or hybrid cell-material constructs, through bioprinting or bioassembly and subsequent

tissue maturation processes’

Moroni et al , 2018

BIOFABRICATION

BIOASSEMBLY BIOPRINTING

https://doi.org/10.1016/j.tibtech.2017.10.015

aggregates

micro-tissues

Hybrid cell-material

constructs

Single cells

(living materials)(cell-driven

self-organization)

In summary

Traditional TE

needed…

- Scaling up

- Anatomically accurate

and mechanically

functional implants upon

implantation

- A robust and controllable

process that diminishes

variability in the biological

outcome

- Personalised approach

- The gradient deposition

of cells, proteins,

materials…

- An automated,

controlled process from

design to fabrication

allowing scale up

- Combined technologies

that have shown

potential to develop

improved constructs

- Vascularisation

- Material optimisation to

increase shape fidelity

- Complex architectures

- Post-fabrication

processing

- Regulatory and ethical

challenges

Biofabrication

technologies offer…

Challenges still

remain…

State-of-the-art and future perspectives

Possible applications

THANK YOU

Veerle.Bloemen@kuleuven.be

Bioprinting for osteochondral tissue regeneration · •A model of an organ is not a functional...

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