Three-dimensional (3D) bioprinting combines biomaterials, cells and functional components into complex living tissues. Herein, we assembled function-control modules into cell-laden scaffolds using 3D bioprinting. A customized 3D printer was able to tune the microstructure of printed bone mesenchymal stem cell (BMSC)-laden methacrylamide gelatin scaffolds at the micrometer scale. For example, the pore size was adjusted to 282 ± 32 μm and 363 ± 60 μm. To match the requirements of the printing nozzle, collagen microfibers with a length of 22 ± 13 μm were prepared with a high-speed crusher. Collagen microfibers bound bone morphogenetic protein 2 (BMP2) with a collagen binding domain (CBD) as differentiation-control module, from which BMP2 was able to be controllably released. The differentiation behaviors of BMSCs in the printed scaffolds were compared in three microenvironments: samples without CBD-BMP2-collagen microfibers in the growth medium, samples without microfibers in the osteogenic medium and samples with microfibers in the growth medium. The results indicated that BMSCs showed high cell viability (>90%) during printing; CBD-BMP2-collagen microfibers induced BMSC differentiation into osteocytes within 14 days more efficiently than the osteogenic medium. Our studies suggest that these function-control modules are attractive biomaterials and have potential applications in 3D bioprinting.

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Amomum villosum is an important medicinal and edible plant with several pharmacologically active volatile oils. However, identifying A. villosum from A. villosum var. xanthioides and A. longiligulare which exhibit similar morphological characteristics to A. villosum, is difficult. The main goal of this study, therefore, is to mine genetic resources and improve molecular methods that could be used to distinguish these species. A total of eight complete chloroplasts (cp) genomes of these Amomum species which were collected from the main producing areas in China were determined to be 163,608–164,069 bp in size. All genomes displayed a typical quadripartite structure with a pair of inverted repeat (IR) regions (29,820–29,959 bp) that separated a large single copy (LSC) region (88,680–88,857 bp) from a small single copy (SSC) region (15,288–15,369 bp). Each genome encodes 113 different genes with 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. More than 150 SSRs were identified in the entire cp genomes of these three species. The Sanger sequencing results based on 32 Amomum samples indicated that five highly divergent regions screened from cp genomes could not be used to distinguish Amomum species. Phylogenetic analysis showed that the cp genomes could not only accurately identify Amomum species, but also provide a solid foundation for the establishment of phylogenetic relationships of Amomum species. The availability of cp genome resources and the comparative analysis is beneficial for species authentication and phylogenetic analysis in Amomum.

These authors contributed equally to this work. ‡ These authors are corresponding co-authors. SUMMARYAndrographis paniculata is a herbaceous dicot plant widely used for its anti-inflammatory and anti-viral properties across its distribution in China, India and other Southeast Asian countries. A. paniculata was used as a crucial therapeutic treatment during the influenza epidemic of 1919 in India, and is still used for the treatment of infectious disease in China. A. paniculata produces large quantities of the anti-inflammatory diterpenoid lactones andrographolide and neoandrographolide, and their analogs, which are touted to be the next generation of natural anti-inflammatory medicines for lung diseases, hepatitis, neurodegenerative disorders, autoimmune disorders and inflammatory skin diseases. Here, we report a chromosome-scale A. paniculata genome sequence of 269 Mb that was assembled by Illumina short reads, PacBio long reads and high-confidence (Hi-C) data. Gene annotation predicted 25 428 protein-coding genes. In order to decipher the genetic underpinning of diterpenoid biosynthesis, transcriptome data from seedlings elicited with methyl jasmonate were also obtained, which enabled the identification of genes encoding diterpenoid synthases, cytochrome P450 monooxygenases, 2-oxoglutarate-dependent dioxygenases and UDP-dependent glycosyltransferases potentially involved in diterpenoid lactone biosynthesis. We further carried out functional characterization of pairs of class-I and -II diterpene synthases, revealing the ability to produce diversified labdane-related diterpene scaffolds. In addition, a glycosyltransferase able to catalyze O-linked glucosylation of andrograpanin, yielding the major active product neoandrographolide, was also identified. Thus, our results demonstrate the utility of the combined genomic and transcriptomic data set generated here for the investigation of the production of the bioactive diterpenoid lactone constituents of the important medicinal herb A. paniculata.

Complicated vessels pervade almost all body tissues and influence the pathophysiology of the human body significantly. However, current fabrication strategies have limited success at multiscale vascular biofabrication. This study reports a methodology to fabricate soft vascularized tissue at centimeter scale using multimaterial bioprinting by a customized multistage-temperature-control printer. The printed constructs can be perfused via the branched endothelialized vasculatures to support the well-formed 3D capillary networks, which ensure cellular activities with sufficient nutrient supply and then mimic a mature and functional liver tissue in terms of synthesis of liver-specific proteins. Moreover, an inner and external pressure-bearing layer is printed to support the direct surgical anastomosis of the carotid artery to the jugular vein. In summary, a versatile platform to recapitulate the vasculature network is presented, in which case sustaining the optimal cellularization in engineered tissues is achievable.

Three-dimensional (3D) bioprinting technology offers the possibility to deliver, in a defined and organized manner, scaffolding materials and living cells, and therefore holds much promise for tissue engineering and regenerative medicine. In this work, we explored the possibility of gelatin/sodium alginate/carboxymethyl chitosan (Gel/SA/CMCS) hydrogel combining with bone mesenchymal stem cells (BMSCs) for 3Dbioprinting. Compared to the commonly used 3D bioprinting materials such as gelatin/sodium alginate (Gel/SA) hydrogel, the Gel/SA/CMCS hydrogel we developed shows excellent equilibrium water content, good mechanical properties, antibacterial activity, and a slow degradation rate. With this kind of material, we generated a scaffold with homogenous cell distribution. Cell viability after 3D printing showed that over 85% of the printed cells were viable at 0 and 2 days of culturing. Our work demonstrates the feasibility of mixing Gel/SA/CMCS hydrogel with stem cells for 3D bioprinting.

As the essential foundation of bioprinting technology, cell-laden bio-ink is confronted with the inevitable contradiction between printability and bioactivity. For example, type I collagen has been widely applied for its excellent biocompatibility; however, its relatively low self-assembly speed restricts the performance in high-precision bioprinting of cell-laden structures. In this study, we synthesize norbornene-functionalized neutral soluble collagen (NorCol) by the reaction of acid-soluble collagen (Col) and carbic anhydride in the aqueous phase. NorCol retains collagen triple-helical conformation and can be quickly orthogonally cross-linked to build a cell-laden hydrogel via a cell-friendly thiol-ene photoclick reaction. Moreover, the additional carboxyl groups produced in the reaction of carbic anhydride and collagen obviously improve the solubility of NorCol in neutral buffer and miscibility of NorCol with other polymers such as alginate and gelatin. It enables hybrid bio-ink to respond to multiple stimuli, resulting in continuous cross-linked NorCol networks in hybrid hydrogels. For the first time, the collagen with a triple helix structure and gelatin can be mixed and printed, keeping the integrity of the printed construct after gelatin’s dissolution. The molecular interaction among giant collagen molecules allows NorCol hydrogel formation at a low concentration, which leads to excellent cell spreading, migration, and proliferation. These properties give NorCol flexible formability and excellent biocompatibility in temperature-, ion-, and photo-based bioprinting. We speculate that NorCol is a promising bio-ink for emerging demands in tissue engineering, regenerative medicine, and personalized therapeutics.

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