Four dimensional (4D) printing is an emerging technology with great capacity for fabricating complex, stimuli-responsive 3D structures, providing great potential for tissue and organ engineering applications. Although the 4D concept was first highlighted in 2013, extensive research has rapidly developed, along with more-in-depth understanding and assertions regarding the definition of 4D. In this review, we begin by establishing the criteria of 4D printing, followed by an extensive summary of state-of-the-art technological advances in the field. Both transformation-preprogrammed 4D printing and 4D printing of shape memory polymers are intensively surveyed. Afterwards we will explore and discuss the applications of 4D printing in tissue and organ regeneration, such as developing synthetic tissues and implantable scaffolds, as well as future perspectives and conclusions.

4D printing represents one of the most advanced fabrication techniques for prospective applications in tissue engineering, biomedical devices, and soft robotics, among others. In this study, a novel multiresponsive architecture is developed through stereolithography-based 4D printing, where a universal concept of stress-induced shape transformation is applied to achieve the 4D reprogramming. The light-induced graded internal stress followed by a subsequent solvent-induced relaxation, driving an autonomous and reversible change of the programmed configuration after printing, is employed and investigated in depth and details. Moreover, the fabricated construct possesses shape memory property, offering a characteristic of multiple shape change. Using this novel multiple responsive 4D technique, a proof-of-concept smart nerve guidance conduit is demonstrated on a graphene hybrid 4D construct providing outstanding multifunctional characteristics for nerve regeneration including physical guidance, chemical cues, dynamic self-entubulation, and seamless integration. By employing this fabrication technique, creating multiresponsive smart architectures, as well as demonstrating application potential, this work paves the way for truly initiation of 4D printing in various high-value research fields.

Our previous study has demonstrated that tissue factor-factor VIIa (TF/FVIIa) complex promotes the proliferation and migration of colon cancer cell line SW620 through the activation of protease-activated receptor 2 (PAR2). In the current study, the underlying molecular mechanisms of TF/FVIIa/PAR2 signaling in SW620 cells were further explored, with the focus on the role of activator protein-1 (AP-1) subunit c-Jun. The results revealed that PAR2-AP and FVIIa could upregulate c-Jun expression and c-Jun phosphorylation in SW620 cells in a time-dependent manner. The effect of FVIIa was significantly blocked by anti-TF and anti-PAR2 antibodies. Protein kinase Cα (PKCα) inhibitor safingol and extracellular signal-regulated kinase 1 and 2 (ERK1/2) inhibitor U0126 abrogated the activation of c-Jun. In contrast, Ca(2+) chelators EGTA and thapsigargin, and p38MAPK inhibitor SB203580 had no effect. Suppression of c-Jun/AP-1 activation using a natural inhibitor curcumin decreased the expression of caspase-3, MMP-9, and TF, as well as the proliferation and migration of SW620 cells induced by PAR2-AP or FVIIa. Collectively, our findings suggest that c-Jun/AP-1 activation is required for TF/FVIIa/PAR2-induced SW620 cell proliferation and migration. PKCα and ERK1/2 are located upstream of c-Jun/AP-1 in this signaling pathway. Pharmacological inhibition of this pathway might be a novel strategy for colon cancer therapy.

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Chemotherapeutic agents for treating cancers show considerable side effects, toxicity, and drug resistance. To mitigate the problems, we designed nucleus-targeted, echogenic, stimuli-responsive polymeric vesicles (polymersomes) to transport and subsequently release the encapsulated anticancer drugs within the nuclei of pancreatic cancer cells. We synthesized an alkyne-dexamethasone derivative and conjugated it to N-polyethylene glycol (PEG)-polylactic acid (PLA) copolymer employing the Cu catalyzed "Click" reaction. We prepared polymersomes from the dexamethasone-PEG-PLA conjugate along with a synthesized stimuli-responsive polymer PEG-S-S-PLA. The dexamethasone group dilates the nuclear pore complexes and transports the vesicles to the nuclei. We designed the polymersomes to release the encapsulated drugs in the presence of a high concentration of reducing agents in the nuclei of pancreatic cancer cells. We observed that the nucleus-targeted, stimuli-responsive polymersomes released 70% of encapsulated contents in the nucleus-mimicking environment in 80 min. We encapsulated the cancer stemness inhibitor BBI608 in the vesicles and observed that the BBI608 encapsulated polymersomes reduced the viability of the BxPC3 cells to 43% in three-dimensional spheroid cultures. The polymersomes were prepared following a special protocol so that they scatter ultrasound, allowing imaging by a medical ultrasound scanner. Therefore, these echogenic, targeted, stimuli-responsive, and drug-encapsulated polymersomes have the potential for trackable, targeted carrier of chemotherapeutic drugs to cancer cell nuclei.