## 3 Stimuli-Responsive Mechanisms: Avenues for 4D (Bio-)printing
### Temperature-Responsive Materials  
The medical interest in thermoresponsive materials relies on the fact that physiological temperature is constantly set around 37 \({}^{\circ}\)C in humans, besides small variations due to pathologic conditions. As thermoresponsive materials change their properties at a predefined temperature, they can be handled and delivered to the human body in a different state (e.g., liquid), to finally change after implantation (e.g., solid). In addition to this, thermoresponsive materials can also provide a useful means of monitoring storage temperature of drugs and bioactive devices, as a prolonged time of exposure to high temperatures can affect their outcomes. In this view, new SU-8 nanofiber-based intelligent indicators for perishable supply chain aim to monitor both temperature and time of temperature change duration in order to ensure reliability of bioactive properties [64]. Furthermore, some diseases induce changes in body temperature that can be identified as a planned stimulus for temperature-sensitive polymers [65]. Temperature responsiveness is one of the most frequently studied mechanisms to obtain user-defined functionality [16], and the most used stimulus to deform bioprinted constructs [24; 66; 67; 68]. Thermoresponsive materials may employ exogenous temperature changes as stimuli to achieve shape transformation. These materials can bend, shrink, or swell depending on temperature changes, as some polymers have the phase transition temperature similar to the physiological temperature [24]. The sensitivity to temperature is caused by the equilibrium between hydrophobic and hydrophilic segments, such as monomeric units or polymer blocks [69]; an upper critical solution temperature (UCST) or a lower critical solution temperature (LCST) can promote interactions between the polymer and the aqueous medium, providing the interruption of the electrostatic (intra- and intermolecular) and hydrophobic actions, which result in collapse or expansion of the polymeric chains [65]. By copolymerizing hydrophobic monomers or adapting the molecular weight of the polymer, LCST can be adjusted to the desired temperature. For instance, increasing the monomers along with a high molecular weight will promote a reduction of LSCT, whereas adding hydrophilic monomers (forminghydrogen bonds) with thermosensitive monomers will cause an increase of the LCST [69]. A number of temperature-sensitive polymers are reported in Table 2. A candidate material adequate for drug delivery or tissue regeneration is poly(\(N\)-isopropylacrylamide) (PNIPAAm), which can be composed with PCL and applied in cell encapsulation and/or release [24; 67; 70]. For example, star-like bilayer poly-(NIPAM-ABP) films remained undeformed at T \(>\) 28 \({}^{\circ}\)C, but by decreasing the temperature below the cloud point of poly-(NIPAM-ABP), the arms of the bilayer started to bend (Figure 4a) [70]. Figure 4b shows the shrinking process of hydrogel after being incubated at 40 \({}^{\circ}\)C water for 35 s, which led to the formation of a tubelike gel reaching \(\approx\)80% of the shape change [71]. Among all thermoresponse materials, PNIPAAm, which is composed of both hydrophilic amide (-CONH-) groups and hydrophobic isopropyl [-CH(CH\({}_{3}\))\({}_{2}\)] side chains, is the best known, as it exhibits a LCST around 32 \({}^{\circ}\)C, has a sol state that can switch to gel state by approaching the body temperature (37 \({}^{\circ}\)C). This property makes PNIPAM a very good candidate for biomedical applications such as scaffolds for tissue engineering, drug carriers, and skin dressings for wound treatment [72]. Cytotoxicity tests have already been performed on PNIPAAm, and results showed low levels of toxicity for the cells [73; 74]. A temperature-responsive hydrogel based on PNIPAAm-_co_-gelatin copolymer was used to produce hepatoma cell line (Hepa/8F5) spheroids, showing that, by virtue of the 3D cellular aggregation enabled by this copolymer, they possessed higher resistance to different drugs [75].