Levensvatbaarheid van Bioprinted Cellular constructen met behulp van een drie Dispenser cartesiaanse Printer

Summary

A Cartesian bioprinter was designed and fabricated to allow multi-material deposition in precise, reproducible geometries, while also allowing control of environmental factors. Utilizing the three-dimensional bioprinter, complex and viable constructs may be printed and easily reproduced.

Abstract

Tissue engineering has centralized its focus on the construction of replacements for non-functional or damaged tissue. The utilization of three-dimensional bioprinting in tissue engineering has generated new methods for the printing of cells and matrix to fabricate biomimetic tissue constructs. The solid freeform fabrication (SFF) method developed for three-dimensional bioprinting uses an additive manufacturing approach by depositing droplets of cells and hydrogels in a layer-by-layer fashion. Bioprinting fabrication is dependent on the specific placement of biological materials into three-dimensional architectures, and the printed constructs should closely mimic the complex organization of cells and extracellular matrices in native tissue. This paper highlights the use of the Palmetto Printer, a Cartesian bioprinter, as well as the process of producing spatially organized, viable constructs while simultaneously allowing control of environmental factors. This methodology utilizes computer-aided design and computer-aided manufacturing to produce these specific and complex geometries. Finally, this approach allows for the reproducible production of fabricated constructs optimized by controllable printing parameters.

Introduction

Tissue engineering gebruikt de principes van de biologie en techniek in de ontwikkeling van functionele substituten te behouden, herstellen of versterken natief weefsel en. Het vermogen van het opwekken van drie-dimensionale biomimetische constructen on demand zou wetenschappelijke en technologische vooruitgang in tissue engineering en in cel-gebaseerde sensoren, geneesmiddel / toxiciteitsscreening, weefsel of tumormodellen en andere vergemakkelijken. De driedimensionale organisatie van weefselengineering constructs is een fundamentele component van de fabricagemethode omdat het nauw de zeer georganiseerde interactie van cellen en extracellulaire matrix in natief wee....

Protocol

1. Voorbereiding van de Gelatine bevattend substraat voor driedimensionale Bioprinting alginaat Hydrogels

1. Bereid de calcium / gelatine substraat volgens de calcium / gelatine substraat beschreven door Pataky et al ¹¹ tot verminderde levensvatbaarheid geassocieerd met een hoog gehalte voorkomen. De calcium / gelatine substraat methode hieronder.
   1. Combineer calciumchloride dehydraat (1,5 gew%), natriumchloride (0,9 gew%) en varkensgelatine (2 gew%) in gedestilleerd water en kook 2 min een 100 mM gelatine-oplossing te creëren.
2. Giet 5 ml gelatine / calciumoplossing in 100 mm standaard petrischalen, zwenk de oplos....

Results

De resultaten tonen de bioprinter kan afzetten cellen beladen hydrogels in bepaalde driedimensionale locaties nauwkeurig en consistent gebruik van computerondersteunde software. Deze software bepaalt de plaatsing van elke druppel en controleren vele van de parameters voor het afgeven (figuur 3,4). De herhaalbaarheid van de bioprinter om adequaat biomaterialen storten is van fundamenteel belang voor het succes ervan in tissue engineering-toepassingen.

Levensvatbaarheid van de.......

Discussion

De primaire focus van tissue engineering is om de kloof tussen de tekorten orgel en transplantatie behoeften te overbruggen door het ontwikkelen van biologische substituten te kunnen herstellen, handhaven of verbeteren van inheemse weefsel functio. Dit heeft geleid tot de directe vervaardiging van steigers met een complex anatomisch juiste externe geometrie en nauwkeurige controle over de interne geometr. Driedimensionale bioprinting is een methode voor het genereren van driedimensionale constructies van verschillende m.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Positioning Robot (JR2000 XYZ)	Janome
Dispensers: SDAV Linear Drive SmartDispensers	Fishman Corporation
Optical Light Sensors:	Keyensce
Displacement Laser: OD Mini	SICK
Recirculating Water Bath: Polystat	Cole-Parmer	EW-12122-02
USB Cameras: Dino-Lite Premier 5MP	AnMo Electrionics/YSC Technologies	AD7013MT
Printer-Compatible Computer Design Software: JR-C Points	Janome		Comes with purchase of Janome Robot
Computer-Aided Design Drawing Software: Visual PathBuilder	RatioServ		Can be downloaded at: www.ratioserv.com/index.php/downloads
Printer 3 cc Syringes:	Fishman Corporation	122051
22 G Dispenser Tips	Fishman Corporation	Z520122
Calcium Chloride Dihydrate	Sigma-Aldrich	10035-04-8
Sodium Chloride	Sigma-Aldrich	7647-14-5
Porcine Gelatin	Sigma-Aldrich	9000-70-8
Titanium Dioxide	Sigma-Aldrich	13462-67-7
Protanal LF 20/40 Alginate (Sodium Alginate)	FMC BioPolymer	9005-38-3
Hydrochloric Acid	Sigma-Aldrich	7647-01-0
Ethylene Glycol	Mallinckrodt Baker, Inc	9300-01
Sodium Periodate	Sigma-Aldrich	7790-28-5
hADSC	Lonza	PT-5006	Store in vials in liquid nitrogen until use.
Dulbecco's Modified Eagle's Medium	Gibco Life Technologies	11965-092	Warm in 37 °C water before use.
Trypsin/EDTA	Lonza	CC-5012	Warm in 37 °C water before use.
Calcein AM	Gibco Life Technologies	C3100MP	Store in the dark at -80 °C until use.
Live/Dead Mammalian Viability Assay Kit	Invitrogen Life Technologies	L-3224	Store in the dark at -80 °C until use.
MES Hydrate	Sigma-Aldrich	M2933
N-Hydroxysuccinimide	Sigma-Aldrich	130672
1-ethyl-(dimethylaminopropyl) carbodiimide (EDC)	Sigma-Aldrich	E1769	10 G
Dulbecco's Phosphate-Buffered Saline, +Calcium, +Magnesium	Life Technologies	14040133	Warm in 37 °C water before use.
Dulbecco's Phosphate-Buffered Saline, -Calcium, -Magnesium	Life Technologies	14190144	Warm in 37 °C water before use.
RGD Peptides	International Peptides
Alexa Fluor 546 Phalloidin Stain	Invitrogen Life Technologies	A22283	Store at -20 °C until use
(4’, 6-Diamidino-2-Phenylindole, Dihydrochloride) (DAPI) Stain	Life Technologies	R37606	Store at -20 °C until use