Gelatin methacryloyl (GelMA) is a versatile material for a wide range of bioapplications. There is an intense interest in developing effective chemical strategies to prepare GelMA with a high degree of batch-to-batch consistency and controllability in terms of methacryloyl functionalization and physiochemical properties. Herein, we systematically investigated the batch-to-batch reproducibility and controllability of producing GelMA (target highly and lowly substituted versions) via a one-pot strategy. To assess the GelMA product, several parameters were evaluated, including the degree of methacryloylation, secondary structure, and enzymatic degradation, along with the mechanical properties and cell viability of GelMA hydrogels. The results showed that two types of target GelMA with five batches exhibited a high degree of controllability and reproducibility in compositional, structural, and functional properties owing to the highly controllable one-pot strategy.

Gelatin methacryloyl (GelMA) has been increasingly considered as an important bioink material due to its tailorable mechanical properties, good biocompatibility, and ability to be photopolymerized in situ as well as printability. GelMA can be classified into two types: type A GelMA (a product from acid treatment) and type B GelMA (a product from alkali treatment). In current literature, there is little research on the comparison of type A GelMA and type B GelMA in terms of synthesis, rheological properties, and printability for bioink applications. Here, we report the synthesis, rheological properties, and printability of types A and B GelMA. Types A and B GelMA samples with different degrees of substitution (DS) were prepared in a controllable manner by a time-lapse loading method of methacrylic anhydride (MAA) and different feed ratios of MAA to gelatin. Type B GelMA tended to have a slightly higher DS compared to type A GelMA, especially in a lower feed ratio of MAA to gelatin. All the type A and type B GelMA solutions with different DS exhibited shear thinning behaviours at 37 °C. However, only GelMA with a high DS had an easy-to-extrude feature at room temperature. The cell-laden printed constructs of types A and B GelMA at 20% w/v showed around 75% cell viability.

5-Hydroxymethylfurfural (5-HMF) is found in many food products including honey, dried fruits, coffee and black garlic extracts. Here, we investigated the anti-inflammatory activity of 5-HMF and its underlying mechanisms in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. 5-HMF pretreatment ranging from 31.5 to 126.0 μg/mL reduced the production of nitric oxide (NO), prostaglandin E2 (PGE2) and pro-inflammatory cytokines (TNF-α, IL-1β and IL-6) in a concentration-dependent manner in LPS-stimulated cells. Moreover, 5-HMF-pretreated cells significantly down-regulated the mRNA expression of two major inflammatory mediators, nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and suppressed the production of pro-inflammatory cytokines, as compared with the only LPS-stimulated cells. 5-HMF suppressed the phosphorylation of extracellular regulated protein kinases (ERK1/2), c-Jun N-terminal kinase (JNK), IκBα, NF-κB p65, the mammalian target of rapamycin (mTOR) and protein kinase B (Akt). Besides, 5-HMF was proved to inhibit NF-κB p65 translocation into nucleus to activate inflammatory gene transcription. These results suggest that 5-HMF could exert the anti-inflammatory activity in the LPS-induced inflammatory response by inhibiting the MAPK, NF-κB and Akt/mTOR pathways. Thus, 5-HMF could be considered as a therapeutic ingredient in functional foods.

Gelatin methacryloyl (GelMA) is a versatile biomaterial that has been shown to possess many advantages such as good biocompatibility, support for cell growth, tunable mechanical properties, photocurable capability, and low material cost. Due to these superior properties, much research has been carried out to develop GelMA as a bioink for bioprinting. However, there are still many challenges, and one major challenge is the control of its rheological properties to yield good printability. Herein, this study presents a strategy to control the rheology of GelMA through partial enzymatic crosslinking. Unlike other enzymatic crosslinking strategies where the rheological properties could not be controlled once reaction takes place, we could, to a large extent, keep the rheological properties stable by introducing a deactivation step after obtaining the optimized rheological properties. Ca2+-independent microbial transglutaminase (MTGase) was introduced to partially catalyze covalent bond formation between chains of GelMA. The enzyme was then deactivated to prevent further uncontrolled crosslinking that would render the hydrogel not printable. After printing, a secondary post-printing crosslinking step (photo crosslinking) was then introduced to ensure long-term stability of the printed structure for subsequent cell studies. Biocompatibility studies carried out using cells encapsulated in the printed structure showed excellent cell viability for at least 7 d. This strategy for better control of rheological properties of GelMA could more significantly enhance the usability of this material as bioink for bioprinting of cell-laden structures for soft tissue engineering.

Triple-negative breast cancer (TNBC) accounts for 15% of overall breast cancer. A lack of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2 receptor) makes TNBC more aggressive and metastatic. Wnt signaling is one of the important pathways in the cellular process; in TNBC it is aberrantly regulated, which leads to the progression and metastasis. In this study, we designed a therapeutic strategy using a combination of a low dose of paclitaxel and a Wnt signaling inhibitor (XAV939), and examined the effect of the paclitaxel-combined XAV939 treatment on diverse breast cancer lines including TNBC cell lines (MDA-MB-231, MDA-MB-468, and BT549) and ER+ve cell lines (MCF-7 and T-47D). The combination treatment of paclitaxel (20 nM) and XAV939 (10 µM) exerted a comparable therapeutic effect on MDA-MB-231, MDA-MB-468, BT549, MCF-7, and T-47D cell lines, relative to paclitaxel with a high dose (200 nM). The paclitaxel-combined XAV939 treatment induced apoptosis by suppressing Bcl-2 and by increasing the cleavage of caspases-3 and PARP. In addition, the in vivo results of the paclitaxel-combined XAV939 treatment in a mice model with the MDA-MB-231 xenograft further confirmed its therapeutic effect. Furthermore, the paclitaxel-combined XAV939 treatment reduced the expression of β-catenin, a key molecule in the Wnt pathway, which led to suppression of the expression of epithelial-mesenchymal transition (EMT) markers and angiogenic proteins both at mRNA and protein levels. The expression level of E-cadherin was raised, which potentially indicates the inhibition of EMT. Importantly, the breast tumor induced by pristane was significantly reduced by the paclitaxel-combined XAV939 treatment. Overall, the paclitaxel-combined XAV939 regimen was found to induce apoptosis and to inhibit Wnt signaling, resulting in the suppression of EMT and angiogenesis. For the first time, we report that our combination approach using a low dose of paclitaxel and XAV939 could be conducive to treating TNBC and an external carcinogen-induced breast cancer.