Bioprinting, defined as depositing cells, extracellular matrices and other biologically relevant materials in user-defined patterns to build tissue constructs de novo or to build upon pre-fabricated scaffolds, is among one of the most promising techniques in tissue engineering. Among the various technologies used for Bioprinting, pressure driven systems are most conducive to preserving cell viability. Herein, we explore the abilities of a novel bioprinter - Digilab, Inc.'s prototype cell printer. The prototype cell printer (Digilab Inc., Holliston, MA) is an automated liquid handling device capable of delivering cell suspension in user-defined patterns onto standard cell culture substrates or custom-designed scaffolds. In this work, the feasibility of using the cell printer to deliver cell suspensions to biological sutures was explored. Cell therapy using stem cells of various types shows promise to aid healing and regeneration in various ailments, including heart failure. Recent evidence suggests that delivering bone-marrow derived mesenchymal stem cells to the infarcted heart reduces infarct size and improves ventricular performance. Current cell delivery systems, however, have critical limitations such as inefficient cell retention, poor survival, and lack of targeted localization. Our laboratories have developed a method to produce discrete fibrin microthreads that can be bundled to form a suture and attached to a needle. These sutures can then be seeded with bone-marrow derived mesenchymal stem cells to deliver these cells to a precise location within the heart wall, both in terms of depth and surface localization. The efficiency of the process of seeding cells onto fibrin thread bundles (sutures) has previously been shown to be 11.8 ± 3.9 %, suggesting that 88% of the cells in suspension are not used. Considering that the proposed cell-therapy model for treatment of myocardial infarction contemplates use of autologous bone-marrow derived stem cells, an improvement in the efficiency of seeding cells onto the fibrin sutures is highly desirable. The feasibility of using Digilab's prototype cell printer to deliver concentrated cell suspension containing human mesenchymal stem cells (hMSCs) directly onto a fibrin thread bundle was explored in this work, in order to determine if this technology could be adapted to seed cells onto such biological sutures. First the effect of the printing process on the viability of hMSCs was assessed by comparing to cells dispensed manually using a hand-held pipette. The viability of hMSCs 24 hours post-dispensing using the cell printer was found to be 90.9 ± 4.0 % and by manual pipetting was 90.6 ± 8.2 % (p = ns). Thereafter a special bioreactor assembly composed of sterilizable Delrin plastic and stainless steel pins was designed to mount fibrin thread bundles onto the deck of the cell printer, to deliver a suspension containing hMSCs on the bundles. Highly targeted delivery of cell suspension directly onto fibrin thread bundles (average diameter 310 µm) was achieved with the bundle suspended in mid-air horizontally parallel to the printer's deck mounted on the bioreactor assembly. To compare seeding efficiency, fibrin thread bundles were simultaneously seeded with hMSCs using either the cell printer or the current method (tube-rotator method) and incubated for 24 hours. Seeded thread bundles were visualized using confocal microscopy and the number of cells per unit length of the bundle was determined for each group. The average seeding efficiency with the tube rotator method was 7.0 ± 0.03 % while the cell printer was 3.46 ± 2.24% (p = ns). In conclusion, the cell printer was found to handle cells as gently as manual pipetting, preserve their viability, with the added abilities to dispense cells in user-defined patterns in an automated manner. With further development, such as localized temperature, gas and humidity control on the cell printer's deck to aid cell survival, the seeding efficiency is likely to improve. The feasibility of using this automated liquid handling technology to deliver cells to biological scaffolds in specified patterns to develop vehicles for cell therapy was shown in this study. Seeding other cell types on other scaffolds along with selectively loading them with growth factors or multiple cell types can also be considered. In sum, the cell printer shows considerable potential to develop novel vehicles for cell therapy. It empowers researchers with a supervision-free, gentle, patterned cell dispensing technique while preserving cell viability and a sterile environment. Looking forward, de novo biofabrication of tissue replicates on a small scale using the cell printer to dispense cells, extracellular matrices, and growth factors in different combinations is a very realistic possibility.
Worcester Polytechnic Institute
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Kanani, Chirantan, "Cell Printing: A novel method to seed cells onto biological scaffolds" (2012). Masters Theses (All Theses, All Years). 332.
fibrin, cell therapy, stem cell therapy, Bioprinting, Cell printing