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Spatiotemporally controlled two-photon photodegradation of hydrogels has gained increasing attention for high-precision subtractive tissue engineering. However, conventional photolabile hydrogels often have poor efficiency upon two-photon excitation in the near-infrared (NIR) region and thus require high laser dosage that may compromise cell activity. As a result, high-speed two-photon hydrogel erosion in the presence of cells remains challenging. Here we introduce the design and synthesis of efficient coumarin-based photodegradable hydrogels to overcome these limitations. A set of photolabile coumarin-functionalized polyethylene glycol linkers are synthesized through a Passerini multicomponent reaction. After mixing these linkers with thiolated hyaluronic acid, semi-synthetic photodegradable hydrogels are formed in situ via Michael addition crosslinking. The efficiency of photodegradation in these hydrogels is significantly higher than that in nitrobenzyl counterparts upon two-photon irradiation at 780 nm. A complex microfluidic network mimicking the bone microarchitecture is successfully fabricated in preformed coumarin hydrogels at high speeds of up to 300 mm s -1 and low laser dosage down to 10 mW. Further, we demonstrate fast two-photon printing of hollow microchannels inside a hydrogel to spatiotemporally direct cell migration in 3D. Collectively, these hydrogels may open new avenues for fast laser-guided tissue fabrication at high spatial resolution., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

Hydrogels are polymeric materials with water contents similar to that of soft tissues. Due to their biomimetic properties, they have been extensively used in various biomedical applications including cell encapsulation for tissue engineering. The utilization of photopolymers provides a possibility for the temporal and spatial controlling of hydrogel cross-links. We produced three-dimensional (3-D) hydrogel scaffolds by means of the two-photon polymerization (2PP) technique. Using a highly efficient water-soluble initiator, photopolymers with up to 80 wt.% water were processed with high precision and reproducibility at a writing speed of 10  mm/s. The biocompatibility of the applied materials was verified using Caenorhabditis elegans as living test organisms. Furthermore, these living organisms were successfully embedded within a 200×200×35 μm³ hydrogel scaffold. As most biologic tissues exhibit a window of transparency at the wavelength of the applied femtosecond laser, it is suggested that 2PP is promising for an in situ approach. Our results demonstrate the feasibility of and potential for bio-fabricating 3-D tissue constructs in the micrometre-range via near-infrared lasers in direct contact with a living organism.

Spatiotemporally controlled two‐photon photodegradation of hydrogels has gained increasing attention for high‐precision subtractive tissue engineering. However, conventional photolabile hydrogels often have poor efficiency upon two‐photon excitation in the near‐infrared (NIR) region and thus require high laser dosage that may compromise cell activity. As a result, high‐speed two‐photon hydrogel erosion in the presence of cells remains challenging. Here we introduce the design and synthesis of efficient coumarin‐based photodegradable hydrogels to overcome these limitations. A set of photolabile coumarin‐functionalized polyethylene glycol linkers are synthesized through a Passerini multicomponent reaction. After mixing these linkers with thiolated hyaluronic acid, semi‐synthetic photodegradable hydrogels are formed in situ via Michael addition crosslinking. The efficiency of photodegradation in these hydrogels is significantly higher than that in nitrobenzyl counterparts upon two‐photon irradiation at 780 nm. A complex microfluidic network mimicking the bone microarchitecture is successfully fabricated in preformed coumarin hydrogels at high speeds of up to 300 mm s−1 and low laser dosage down to 10 mW. Further, we demonstrate fast two‐photon printing of hollow microchannels inside a hydrogel to spatiotemporally direct cell migration in 3D. Collectively, these hydrogels may open new avenues for fast laser‐guided tissue fabrication at high spatial resolution. [ABSTRACT FROM AUTHOR]

The article in Angewandte Chemie discusses the development of photoabbaubare hydrogels on Cumarin-Basis for two-photon controlled subtractive biofabrication at 300 mm/s. Researchers led by Xiao-Hua Qin have created cell-compatible, photolabile macromolecular networks that can be rapidly cleaved with laser exposure, allowing for the direct laser writing of complex 3D microfluidic networks. This technology enables proof-of-concept laser-guided tissue morphogenesis at micron-scale resolution, offering new possibilities for spatiotemporal tissue engineering and drug testing in the laboratory. [Extracted from the article]

The article in the journal "Angewandte Chemie International Edition" discusses the development of coumarin-based photodegradable hydrogels that allow for two-photon subtractive biofabrication at a speed of 300 mm/s. This technology enables the creation of complex 3D microfluidic networks and demonstrates laser-guided tissue morphogenesis at a micron-scale resolution. The research by Xiao-Hua Qin and colleagues focuses on creating cell-compatible macromolecular networks that can be rapidly cleaved upon laser exposure for applications in tissue engineering and drug testing. [Extracted from the article]

The natural extracellular matrix (ECM) represents a complex and dynamic environment. It provides numerous spatio-temporal signals mediating many cellular functions including morphogenesis, adhesion, proliferation and differentiation. The cell-ECM interaction is bidirectional. Cells dynamically receive and process information from the ECM and remodel it at the same time. Theses complex interactions are still not fully understood. For better understanding, it is indispensable to deconstruct the ECM up to the point of investigating isolated characteristics and cell responses to physical, chemical and topographical cues. Two-photon polymerization (2PP) allows the exact reconstruction of cell specific sites in 3D at micro- and nanometer precision. Processing biocompatible synthetic and naturally-derived hydrogels, the microenvironment of cells can be designed to specifically investigate their behavior in respect to key chemical, mechanical and topographical attributes. Moreover, 3D manipulation can be performed in the presence of cells, guiding biological tissue formation in all stages of its development. Here, advances in 2PP microfabrication of synthetic and naturally based hydrogels are reviewed. Key components of photopolymerizable hydrogel precursors, their structure-property relationships and their polymerization mechanisms are presented. Furthermore, it is shown how biocompatible 2PP fabricated constructs can act as biologically relevant matrices to study cell functions and tissue development. [ABSTRACT FROM AUTHOR]

Spatiotemporally controlled two‐photon photodegradation of hydrogels has gained increasing attention for high‐precision subtractive tissue engineering. However, conventional photolabile hydrogels often have poor efficiency upon two‐photon excitation in the near‐infrared (NIR) region and thus require high laser dosage that may compromise cell activity. As a result, high‐speed two‐photon hydrogel erosion in the presence of cells remains challenging. Here we introduce the design and synthesis of efficient coumarin‐based photodegradable hydrogels to overcome these limitations. A set of photolabile coumarin‐functionalized polyethylene glycol linkers are synthesized through a Passerini multicomponent reaction. After mixing these linkers with thiolated hyaluronic acid, semi‐synthetic photodegradable hydrogels are formed in situ via Michael addition crosslinking. The efficiency of photodegradation in these hydrogels is significantly higher than that in nitrobenzyl counterparts upon two‐photon irradiation at 780 nm. A complex microfluidic network mimicking the bone microarchitecture is successfully fabricated in preformed coumarin hydrogels at high speeds of up to 300 mm s−1 and low laser dosage down to 10 mW. Further, we demonstrate fast two‐photon printing of hollow microchannels inside a hydrogel to spatiotemporally direct cell migration in 3D. Collectively, these hydrogels may open new avenues for fast laser‐guided tissue fabrication at high spatial resolution. [ABSTRACT FROM AUTHOR]

[Display omitted] • The structured porous hydrogel scaffolds were fabricated by 3D printing. • The porous structure is more conducive to the activity and proliferation of cells in the hydrogel. • The porous structure promotes the expression of osteogenesis-related genes of BMSCs in hydrogel scaffolds. • Structured porous hydrogels produce more mineralization in vivo. Hydrogel is a suitable material for three-dimensional cell culture, and is rich in water content and can better simulate a natural extracellular matrix. However, the relatively compact pores in the interior often inhibit the growth and differentiation of containing cells. Additionally, the impact of the pore structure on the differentiation of cells remains to be unknown. In this work, porous hydrogel scaffolds with medium pore sizes (100–1000 μm) were prepared using 3D printing technology, and the hydrogel scaffolds without pores and non-printed hydrogel scaffolds were designed as control to investigate the effect of 3D printed porous structures on osteogenic differentiation of BMSCs within hydrogel scaffolds, both in vitro and in vivo. The pore structure of 3D printed structured porous scaffold was explicitly inspected by SEM. The water content and expansion rate of hydrogel did not change, despite its porous structure. The BMSCs were evenly distributed across the porous scaffold and maintained high activity, thus benefiting from the sufficient circulation and diffusion of oxygen and nutrients due to the regular pores. In vitro , the osteogenic differentiation of BMSCs in the structured porous hydrogels was more prominent compared to the non-porous groups. Moreover, the mineralization of the hydrogel scaffolds was also enhanced. After ectopic implantation in the posterior gluteal muscle punch, the structured porous hydrogel exhibited relatively good mineralization and ectopic osteogenesis by the three-dimensional reconstruction of micro-CT, collagen staining, and immunological analysis. The 3D printing-based approach provides a facile strategy for the preparation of hydrogels with intermediate pore structures. This can promote the viability and osteogenic differentiation of BMSCs in hydrogel scaffolds. [ABSTRACT FROM AUTHOR]

Subtractive two- photon biofabrication facilitates spatiotemporal tissue engineering and drug testingin the laboratory. However, it necessitates the rational design of cell- compatible, photolabile macromolecular networks that cleave rapidly upon laser exposure. In their Communication ( e202404599), Xiao- Hua Qin and co- workers develop coumarin- based photodegradable hydrogels that enable two- photon direct laser writing of complex 3D microfluidic networks at 300 mm s 1. Proof- of- concept laser- guided tissue morphogenesis is demonstrated at micron- scale resolution. [Extracted from the article]

The two-photon polymerization (2PP)of photosensitive gelatin inthe presence of living cells is reported. The 2PP technique is basedon the localized cross-linking of photopolymers induced by femtosecondlaser pulses. The availability of water-soluble photoinitiators (PI)suitable for 2PP is crucial for applying this method to cell-containingmaterials. Novel PIs developed by our group allow 2PP of formulationswith up to 80% cell culture medium. The cytocompatibility of thesePIs was evaluated by an MTT assay. The results of cell encapsulationby 2PP show the occurrence of cell damage within the laser-exposedregions. However, some cells located in the immediate vicinity andeven within the 2PP-produced structures remain viable and can furtherproliferate. The control experiments demonstrate that the laser radiationitself does not damage the cells at the parameters used for 2PP. Onthe basis of these findings and the reports by other groups, we concludethat such localized cell damage is of a chemical origin and can beattributed to reactive species generated during 2PP. The viable cellstrapped within the 2PP structures but not exposed to laser radiationcontinued to proliferate. The live/dead staining after 3 weeks revealedviable cells occupying most of the space available within the 3D hydrogelconstructs. While some of the questions raised by this study remainopen, the presented results indicate the general practicability of2PP for 3D processing of cell-containing materials. The potentialapplications of this highly versatile approach span from precise engineeringof 3D tissue models to the fabrication of cellular microarrays. [ABSTRACT FROM AUTHOR]