344Laser bioprinting of human iPSC-derived neural stem cells and neurons: Effect on cell survival, multipotency, differentiation, and neuronal activity.

Generation of human neuronal networks by three-dimensional (3D) bioprinting is promising for drug testing and hopefully will allow for the understanding of cellular mechanisms in brain tissue. The application of neural cells derived from human induced-pluripotent stem cells (hiPSCs) is an obvious choice, since hiPSCs provide access to cells unlimited in number and cell types that could be generated by differentiation. The questions in this regard include which neuronal differentiation stage is optimal for printing of such networks, and to what extent the addition of other cell types, especially astrocytes, supports network formation. These aspects are the focus of the present study, in which we applied a laser-based bioprinting technique and compared hiPSC-derived neural stem cells (NSCs) with neuronal differentiated NSCs, with and without the inclusion of co-printed astrocytes. In this study, we investigated in detail the effects of cell types, printed droplet size, and duration of differentiation before and after printing on viability, as well as proliferation, stemness, differentiation potential, formation of dendritic extensions and synapses, and functionality of the generated neuronal networks. We found a significant dependence of cell viability after dissociation on differentiation stage, but no impact of the printing process. Moreover, we observed a dependence of the abundance of neuronal dendrites on droplet size, a marked difference between printed cells and normal cell culture in terms of further differentiation of the cells, especially differentiation into astrocytes, as well as neuronal network formation and activity. Notably, there was a clear effect of admixed astrocytes on NSCs but not on neurons.

Capillary-like Formations of Endothelial Cells in Defined Patterns Generated by Laser Bioprinting.

Bioprinting is seen as a promising technique for tissue engineering, with hopes of one day being able to produce whole organs. However, thick tissue requires a functional vascular network, which naturally contains vessels of various sizes, down to capillaries of ~10 µm in diameter, often spaced less than 200 µm apart. If such thick tissues are to be printed, the vasculature would likely need to be printed at the same time, including the capillaries. While there are many approaches in tissue engineering to produce larger vessels in a defined manner, the small capillaries usually arise only in random patterns by sprouting from the larger vessels or from randomly distributed endothelial cells. Here, we investigated whether the small capillaries could also be printed in predefined patterns. For this purpose, we used a laser-based bioprinting technique that allows for the combination of high resolution and high cell density. Our aim was to achieve the formation of closed tubular structures with lumina by laser-printed endothelial cells along the printed patterns on a surface and in bioprinted tissue. This study shows that such capillaries are directly printable; however, persistence of the printed tubular structures was achieved only in tissue with external stimulation by other cell types.

In Vitro Development of Human iPSC-Derived Functional Neuronal Networks on Laser-Fabricated 3D Scaffolds.

Neural progenitor cells generated from human induced pluripotent stem cells (hiPSCs) are the forefront of ″brain-on-chip″ investigations. Viable and functional hiPSC-derived neuronal networks are shaping powerful in vitro models for evaluating the normal and abnormal formation of cortical circuits, understanding the underlying disease mechanisms, and investigating the response to drugs. They therefore represent a desirable instrument for both the scientific community and the pharmacological industry. However, culture conditions required for the full functional maturation of individual neurons and networks are still unidentified. It has been recognized that three-dimensional (3D) culture conditions can better emulate in vivo neuronal tissue development compared to 2D cultures and thus provide a more desirable in vitro approach. In this paper, we present the design and implementation of a 3D scaffold platform that supports and promotes intricate neuronal network development. 3D scaffolds were produced through direct laser writing by two-photon polymerization (2PP), a high-resolution 3D laser microstructuring technology, using the biocompatible and nondegradable photoreactive resin Dental LT Clear (DClear). Neurons developed and interconnected on a 3D environment shaped by vertically stacked scaffold layers. The developed networks could support different cell types. Starting at the day 50 of 3D culture, neuronal progenitor cells could develop into cortical projection neurons (CNPs) of all six layers, different types of inhibitory neurons, and glia. Additionally and in contrast to 2D conditions, 3D scaffolds supported the long-term culturing of neuronal networks over the course of 120 days. Network health and functionality were probed through calcium imaging, which revealed a strong spontaneous neuronal activity that combined individual and collective events. Taken together, our results highlight advanced microstructured 3D scaffolds as a reliable platform for the 3D in vitro modeling of neuronal functions.

Cell Type-Specific Adhesion and Migration on Laser-Structured Opaque Surfaces.

Cytocompatibility is essential for implant approval. However, initial in vitro screenings mainly include the quantity of adherent immortalized cells and cytotoxicity. Other vital parameters, such as cell migration and an in-depth understanding of the interaction between native tissue cells and implant surfaces, are rarely considered. We investigated different laser-fabricated spike structures using primary and immortalized cell lines of fibroblasts and osteoblasts and included quantification of the cell area, aspect ratio, and focal adhesions. Furthermore, we examined the three-dimensional cell interactions with spike topographies and developed a tailored migration assay for long-term monitoring on opaque materials. While fibroblasts and osteoblasts on small spikes retained their normal morphology, cells on medium and large spikes sank into the structures, affecting the composition of the cytoskeleton and thereby changing cell shape. Up to 14 days, migration appeared stronger on small spikes, probably as a consequence of adequate focal adhesion formation and an intact cytoskeleton, whereas human primary cells revealed differences in comparison to immortalized cell lines. The use of primary cells, analysis of the cell-implant structure interaction as well as cell migration might strengthen the evaluation of cytocompatibility and thereby improve the validity regarding the putative in vivo performance of implant material.

Human stem cell based corneal tissue mimicking structures using laser-assisted 3D bioprinting and functional bioinks.

There is a high demand for developing methods to produce more native-like 3D corneal structures. In the present study, we produced 3D cornea-mimicking tissues using human stem cells and laser-assisted bioprinting (LaBP). Human embryonic stem cell derived limbal epithelial stem cells (hESC-LESC) were used as a cell source for printing epithelium-mimicking structures, whereas human adipose tissue derived stem cells (hASCs) were used for constructing layered stroma-mimicking structures. The development and optimization of functional bioinks was a crucial step towards successful bioprinting of 3D corneal structures. Recombinant human laminin and human sourced collagen I served as the bases for the functional bioinks. We used two previously established LaBP setups based on laser induced forward transfer, with different laser wavelengths and appropriate absorption layers. We bioprinted three types of corneal structures: stratified corneal epithelium using hESC-LESCs, lamellar corneal stroma using alternating acellular layers of bioink and layers with hASCs, and finally structures with both a stromal and epithelial part. The printed constructs were evaluated for their microstructure, cell viability and proliferation, and key protein expression (Ki67, p63α, p40, CK3, CK15, collagen type I, VWF). The 3D printed stromal constructs were also implanted into porcine corneal organ cultures. Both cell types maintained good viability after printing. Laser-printed hESC-LESCs showed epithelial cell morphology, expression of Ki67 proliferation marker and co-expression of corneal progenitor markers p63α and p40. Importantly, the printed hESC-LESCs formed a stratified epithelium with apical expression of CK3 and basal expression of the progenitor markers. The structure of the 3D bioprinted stroma demonstrated that the hASCs had organized horizontally as in the native corneal stroma and showed positive labeling for collagen I. After 7 days in porcine organ cultures, the 3D bioprinted stromal structures attached to the host tissue with signs of hASCs migration from the printed structure. This is the first study to demonstrate the feasibility of 3D LaBP for corneal applications using human stem cells and successful fabrication of layered 3D bioprinted tissues mimicking the structure of the native corneal tissue.

Laser bioprinting of human induced pluripotent stem cells-the effect of printing and biomaterials on cell survival, pluripotency, and differentiation.

Research on human induced pluripotent stem cells (hiPSCs) is one of the fastest growing fields in biomedicine. Generated from patient's own somatic cells, hiPSCs can be differentiated towards all functional cell types and returned to the patient without immunological concerns. 3D printing of hiPSCs could enable the generation of functional organs for replacement therapies or realization of organ-on-chip systems for individualized medicine. Printing of living cells was demonstrated with immortalized cell lines, primary cells, and adult stem cells with different printing technologies and biomaterials. However, hiPSCs are more sensitive to handling procedures, in particular, when dissociated into single cells. Both pluripotency and directed differentiation are influenced by numerous environmental factors including culture media, biomaterials, and cell density. Notably, existing literature on the effect of applied biomaterials on pluripotency is rather ambiguous. In this study, laser bioprinting of undifferentiated hiPSCs in combination with different biomaterials was performed and the impact on cells' behavior, pluripotency, and differentiation was investigated. Our findings suggest that hiPSCs are indeed more sensitive to the applied biomaterials, but not to laser printing itself. With appropriate biomaterials, such as the hyaluronic acid based solutions applied in this study, hiPSCs can be successfully laser printed without losing their pluripotency.

Laser-assisted bioprinting at different wavelengths and pulse durations with a metal dynamic release layer: A parametric study.

For more than a decade, living cells and biomaterials (typically hydrogels) are printed via laser-assisted bioprinting. Often, a thin metal layer is applied as laser-absorbing material called dynamic release layer (DRL). This layer is vaporized by focused laser pulses generating vapor pressure that propels forward a coated biomaterial. Different lasers with laser wavelengths from 193 to 1064 nanometer have been used. As a metal DRL gold, silver, or titanium layers have been used. The applied laser pulse durations were usually in the nanosecond range from 1 to 30 ns. In addition, some studies with femtosecond lasers have been published. However, there are no studies on the effect of all these lasers parameters on bioprinting with a metal DRL, and on comparing different wavelengths and pulse durations - except one study comparing 500 femtosecond pulses with 15 ns pulses. In this paper, the effects of laser wavelength (355, 532, and 1064 nm) and laser pulse duration (in the range of 8 to 200 ns) are investigated. Furthermore, the effects of laser pulse energy, intensity, and focal spot size are studied. The printed droplet volume, hydrogel jet velocity, and cell viability are analyzed.

Correlation between ECM guidance and actin polymerization on osteogenic differentiation of human adipose-derived stem cells.

The correlation between extracellular matrix (ECM) components, cell shape, and stem cell guidance can shed light in understanding and mimicking the functionality of stem cell niches for various applications. This interplay on osteogenic guidance of human adipose-derived stem cells (hASCs) was focus of this study. Proliferation and osteogenic markers like alkaline phosphatase activity and calcium mineralization were slightly increased by the ECM components laminin (LA), collagen I (COL), and fibronectin (FIB); with control medium no differentiation occurred. ECM guided differentiation was rather dependent on osterix than on Runx2 pathway. FIB significantly enhanced cell elongation even in presence of actin polymerization blockers cytochalasin D (CytoD) and ROCK inhibitor Y-27632, which generally caused more rounded cells. Except for the COL surface, both inhibitors increased the extent of osterix, while the Runx2 pathway was more sensitive to the culture condition. Both inhibitors did not affect hASC proliferation. CytoD enabled osteogenic differentiation independently from the ECM, while it was rather blocked via Y-27632 treatment; on FIB the general highest extent of differentiation occurred. Taken together, the ECM effect on hASCs occurs indirectly and selectively via a dominant role of FIB: it sustains osteogenic differentiation in case of a tension-dependent control of actin polymerization.

Novel biodegradable star-shaped polylactide scaffolds for bone regeneration fabricated by two-photon polymerization.

AIM: To assess the properties of 3D biodegradable scaffolds fabricated from novel star-shaped poly(D,L-lactide) (SSL) materials for bone tissue regeneration. MATERIALS & METHODS: The SSL polymer was synthesized using an optimized synthetic procedure and applied for scaffold fabrication by the two-photon polymerization technique. The osteogenic differentiation was controlled using human adipose-derived stem cells cultured for 28 days. The SSL scaffolds with or without murine MSCs were implanted into the cranial bone of C57/Bl6 mice. RESULTS: The SSL scaffolds supported differentiation of human adipose-derived stem cells toward the osteogenic lineage in vitro. The SSL scaffolds with murine MSCs enhanced the mineralized tissue formation. CONCLUSION: The SSL scaffolds provide a beneficial microenvironment for the osteogenic MSCs' differentiation in vitro and support de novo bone formation in vivo.

Osteogenic differentiation of human mesenchymal stem cells in 3-D Zr-Si organic-inorganic scaffolds produced by two-photon polymerization technique.

Two-photon polymerization (2PP) is applied for the fabrication of 3-D Zr-Si scaffolds for bone tissue engineering. Zr-Si scaffolds with 150, 200, and 250 µm pore sizes are seeded with human bone marrow stem cells (hBMSCs) and human adipose tissue derived stem cells (hASCs) and cultured in osteoinductive and control media for three weeks. Osteogenic differentiation of hASCs and hBMSCs and formation of bone matrix is comparatively analyzed via alkaline phosphatase activity (ALP), calcium quantification, osteocalcin staining and scanning electron microscopy (SEM). It is observed that the 150 µm pore size Zr-Si scaffolds support the strongest matrix mineralization, as confirmed by calcium deposition. Analysis of ALP activity, osteocalcin staining and SEM observations of matrix mineralization reveal that mesenchymal stem cells cultured on 3-D scaffolds without osteogenic stimulation spontaneously differentiate towards osteogenic lineage. Nanoindentation measurements show that aging of the 2PP-produced Zr-Si scaffolds in aqueous or alcohol media results in an increase in the scaffold Young's modulus and hardness. Moreover, accelerated formation of bone matrix by hASCs is noted, when cultured on the scaffolds with lower Young's moduli and hardness values (non aged scaffolds) compared to the cells cultured on scaffolds with higher Young's modulus and hardness values (aged scaffolds). Presented results support the potential application of Zr-Si scaffolds for autologous bone tissue engineering.

Impact of laser-structured biomaterial interfaces on guided cell responses.

To achieve a perfect integration of biomaterials into the body, tissue formation in contact with the interface has to be controlled. In this connection, a selective cell control is required: fibrotic encapsulation has to be inhibited, while tissue guidance has to be stimulated. As conventional biomaterials do not fulfil this specification, functionalization of the biointerface is under development to mimic the natural environment of the cells. One approach focuses on the fabrication of defined surface topographies. Thereby, ultrashort pulse laser ablation is very beneficial, owing to a large variety of fabricated structures, reduced heat-affected zones, high precision and reproducibility. We demonstrate that nanostructures in platinum and microstructures in silicon selectively control cell behaviour: inhibiting fibroblasts, while stimulating neuronal attachment and differentiation. However, the control of fibroblasts strongly correlates with the created size dimensions of the surface structures. These findings suggest favourable biomaterial interfaces for electronic devices. The mechanisms which are responsible for selective cell control are poorly understood. To give an insight, cell behaviour in dependence of biomaterial interfaces is discussed-including basic research on the role of the extracellular matrix. This knowledge is essential to understand such specific cell responses and to optimize biomaterial interfaces for future biomedical applications.

Tissue engineered skin substitutes created by laser-assisted bioprinting form skin-like structures in the dorsal skin fold chamber in mice.

Tissue engineering plays an important role in the production of skin equivalents for the therapy of chronic and especially burn wounds. Actually, there exists no (cellularized) skin equivalent which might be able to satisfactorily mimic native skin. Here, we utilized a laser-assisted bioprinting (LaBP) technique to create a fully cellularized skin substitute. The unique feature of LaBP is the possibility to position different cell types in an exact three-dimensional (3D) spatial pattern. For the creation of the skin substitutes, we positioned fibroblasts and keratinocytes on top of a stabilizing matrix (Matriderm®). These skin constructs were subsequently tested in vivo, employing the dorsal skin fold chamber in nude mice. The transplants were placed into full-thickness skin wounds and were fully connected to the surrounding tissue when explanted after 11 days. The printed keratinocytes formed a multi-layered epidermis with beginning differentiation and stratum corneum. Proliferation of the keratinocytes was mainly detected in the suprabasal layers. In vitro controls, which were cultivated at the air-liquid-interface, also exhibited proliferative cells, but they were rather located in the whole epidermis. E-cadherin as a hint for adherens junctions and therefore tissue formation could be found in the epidermis in vivo as well as in vitro. In both conditions, the printed fibroblasts partly stayed on top of the underlying Matriderm® where they produced collagen, while part of them migrated into the Matriderm®. In the mice, some blood vessels could be found to grow from the wound bed and the wound edges in direction of the printed cells. In conclusion, we could show the successful 3D printing of a cell construct via LaBP and the subsequent tissue formation in vivo. These findings represent the prerequisite for the creation of a complex tissue like skin, consisting of different cell types in an intricate 3D pattern.

Topography and coating of platinum improve the electrochemical properties and neuronal guidance.

To improve neuronal-electrode interfaces, we analyzed the influence of surface topographies combined with coating on the electrochemistry of platinum and neuronal differentiation of PC-12 cells. Surface structuring on nanoscale was realized by femtosecond laser ablation. Additional coating with laminin (LA), collagen type I (COL) or poly-d-lysine (PDL) did not change the produced topography. We further demonstrated that impedance could be improved in all cases. The pre-requisites of differentiation - viability and attachment - were fulfilled on the topography. Cell attachment of non-differentiated and differentiated cells and their formation of focal adhesion complexes were even enhanced compared to unstructured platinum. However, without the nerve growth factor (NGF) no cellular outgrowth and differentiation were possible. The topography enabled cell elongation and reduced the amount of rounded cells, but less effective than coating. Differentiation was either comparable or increased on the structures when compared with unstructured coatings. For instance, microtubule associated protein (MAP2) was detected most on the topography alone. But a combination of surface structuring and coating had the strongest impact on differentiation: the usage of COL provoked best cell elongation and beta III tubulin expression, PDL best synaptophysin. LA-coating had no noteworthy effect. These findings point out that innovative electronic devices like cochlear implants include two aspects: (a) nanotopography to improve the transmission of electrical signals and neuronal attachment; and (b) an additional coating to stimulate neuronal differentiation.

Skin tissue generation by laser cell printing.

For the aim of ex vivo engineering of functional tissue substitutes, Laser-assisted BioPrinting (LaBP) is under investigation for the arrangement of living cells in predefined patterns. So far three-dimensional (3D) arrangements of single or two-dimensional (2D) patterning of different cell types have been presented. It has been shown that cells are not harmed by the printing procedure. We now demonstrate for the first time the 3D arrangement of vital cells by LaBP as multicellular grafts analogous to native archetype and the formation of tissue by these cells. For this purpose, fibroblasts and keratinocytes embedded in collagen were printed in 3D as a simple example for skin tissue. To study cell functions and tissue formation process in 3D, different characteristics, such as cell localisation and proliferation were investigated. We further analysed the formation of adhering and gap junctions, which are fundamental for tissue morphogenesis and cohesion. In this study, it was demonstrated that LaBP is an outstanding tool for the generation of multicellular 3D constructs mimicking tissue functions. These findings are promising for the realisation of 3D in vitro models and tissue substitutes for many applications in tissue engineering.

Fabrication of fibrin scaffolds with controlled microscale architecture by a two-photon polymerization-micromolding technique.

Fabrication of three-dimensional (3D) fibrin scaffolds with tightly controllable pore sizes and interconnections has been investigated. The scaffolds were produced using a combination of two-photon polymerization (2PP) and micromolding techniques. Master structures were fabricated by 2PP and regenerated in fibrin by a two-step microreplication procedure. Scanning electron and optical microscopy observations showed that the fibrin scaffolds exhibited a highly porous and interconnected structure. Seeding of endothelial cells in fibrin scaffolds resulted in their directed lining and spreading within network of microreplicated pores, whereas encapsulation of endothelial cells in fibrin gel blocks led to their chaotic and irregular distribution within constructs. These results demonstrate that the 2PP-micromolding technique is suitable for fabrication of complex 3D structures from natural proteins for tissue engineering applications.

Proteomic analysis to display the effect of low doses of erythropoietin on rat liver regeneration.

AIMS: Several groups found different impact of erythropoietin (EPO) on liver regeneration. Both pro-proliferative as well as anti-proliferative and non-proliferative activities have been reported using high dosage of EPO. Systemic administration of high doses of this cytokine is a clinical concern due to risk of thrombosis. Herein, we applied EPO in low dosages and investigated whether it can stimulate liver regeneration after liver resection. MAIN METHODS: Parameters of liver regeneration were assessed 3 days after 70% hepatectomy by means of immunochemistry and proteomics. EPO was given twice in low dosages (200 and 600 IU/kg BW). KEY FINDINGS: We showed that EPO facilitated hepatic regeneration in rats. Enhanced hepatocyte proliferation (Ki67, BrdU-positive cells) was observed in all EPO-treated groups. By performing Differential Proteomic analysis, we identified two proteins which resulted sensitive to EPO treatment after hepatectomy: Peroxiredoxin-1 and glutathione S-transferase Mu 1. SIGNIFICANCE: Based on our results, low doses of rhEPO increase the hepatic regenerative capacity after partial hepatectomy in rats by enhancing hepatocyte proliferation and acting on antioxidant enzymes. Both proteins identified by proteomic analysis have not previously been associated with liver regeneration and will aid in the understanding of EPO's regenerative response having clinical implications to treat liver failure.

Laser printing of three-dimensional multicellular arrays for studies of cell-cell and cell-environment interactions.

Utilization of living cells for therapies in regenerative medicine requires a fundamental understanding of the interactions between different cells and their environment. Moreover, common models based on adherent two-dimensional cultures are not appropriate to simulate the complex interactions that occur in a three-dimensional (3D) cell-microenvironment in vivo. In this study, we present a computer-aided method for the printing of multiple cell types in a 3D array using laser-assisted bioprinting. By printing spots of human adipose-derived stem cells (ASCs) and endothelial colony-forming cells (ECFCs), we demonstrate that (i) these cell spots can be arranged layer-by-layer in a 3D array; (ii) any cell-cell ratio, cell quantity, cell-type combination, and spot spacing can be realized within this array; and (iii) the height of the 3D array is freely scalable. As a proof of concept, we printed separate spots of ASCs and ECFCs within a 3D array and observed cell-cell interactions in vascular endothelial growth factor-free medium. It has been demonstrated that direct cell-cell contacts trigger the development of stable vascular-like networks. This method can be applied to study complex and dynamic relationships between cells and their local environment.

Dispensing pico to nanolitre of a natural hydrogel by laser-assisted bioprinting.

BACKGROUND: Laser-assisted bioprinting of multi-cellular replicates in accordance with CAD blueprint may substantially improve our understandings of fundamental aspects of 3 D cell-cell and cell-matrix interactions in vitro. For predictable printing results, a profound knowledge about effects of different processing parameters is essential for realisation of 3 D cell models with well-defined cell densities. METHODS: Time-resolved imaging of the hydrogel jet dynamics and quantitative assessment of the dependence of printed droplet diameter on the process characteristics were conducted. RESULTS: The existence of a counterjet was visualised, proving the bubble collapsing theory for the jet formation. Furthermore, by adjusting the viscosity and height of the applied hydrogel layer in combination with different laser pulse energies, the printing of volumes in the range of 10 to 7000 picolitres was demonstrated. Additionally, the relationship between the viscosity and the layer thickness at different laser pulse energies on the printed droplet volume was identified. CONCLUSIONS: These findings are essential for the advancement of laser-assisted bioprinting by enabling predictable printing results and the integration of computational methods in the generation of 3 D multi-cellular constructs.

Laser fabrication of three-dimensional CAD scaffolds from photosensitive gelatin for applications in tissue engineering.

In the present work, 3D CAD scaffolds for tissue engineering applications were developed starting from methacrylamide-modified gelatin (GelMOD) using two-photon polymerization (2PP). The scaffolds were cross-linked employing the biocompatible photoinitiator Irgacure 2959. Because gelatin is derived from collagen (i.e., the main constituent of the ECM), the developed materials mimic the cellular microenvironment from a chemical point of view. In addition, by applying the 2PP technique, structural properties of the cellular microenvironment can also be mimicked. Furthermore, in vitro degradation assays indicated that the enzymatic degradation capability of gelatin is preserved for the methacrylamide-modified derivative. An in depth morphological analysis of the 2PP-fabricated scaffolds demonstrated that the parameters of the CAD model are reproduced with great precision, including the ridge-like surface topography on the order of 1.5 µm. The developed scaffolds showed an excellent stability in culture medium. In a final part of the present work, the suitability of the developed scaffolds for tissue engineering applications was verified. The results indicated that the applied materials are suitable to support porcine mesenchymal stem cell adhesion and subsequent proliferation. Upon applying osteogenic stimulation, the seeded cells differentiated into the anticipated lineage. Energy dispersive X-ray (EDX) analysis showed the induced calcification of the scaffolds. The results clearly indicate that 2PP is capable of manufacturing precisely constructed 3D tissue engineering scaffolds using photosensitive polymers as starting material.

Laser Fabrication of 3D Gelatin Scaffolds for the Generation of Bioartificial Tissues.

In the present work, the two-photon polymerization (2PP) technique was applied to develop precisely defined biodegradable 3D tissue engineering scaffolds. The scaffolds were fabricated via photopolymerization of gelatin modified with methacrylamide moieties. The results indicate that the gelatin derivative (GelMod) preserves its enzymatic degradation capability after photopolymerization. In addition, the developed scaffolds using 2PP support primary adipose-derived stem cell (ASC) adhesion, proliferation and differentiation into the anticipated lineage.