M.Sc. Hafiz Syed Usama bin Farrukh

Hydrogels mimicking the mechanical and biochemical features of the cellular microenvironment allow cell encapsulation and facilitate in vitro 3D culture. In addition to biocompatibility and reactivity in physiological conditions, a key criterion for crosslinking chemistry is appropriate gelation kinetics to allow mixing and homogeneous distribution of cells with the hydrogel precursors. We have previously presented aryl methylsulfone/thiol (MS/SH) reaction as a thiol-reactive cross-linking system for cell encapsulation in star polyethylene glycol (PEG4) hydrogels with a gelation kinetics in minutes time scale. Remaining experimental challenges for this system are a finer modulation of gelation kinetics and streamlining the synthesis of the prepolymer. Here we present the possibility to tune the gelation kinetics by introducing an electron-withdrawing substituent at p-position of the aryl MS ring. This variant also presents synthetic advantages. We study the influence of the p-substituent on the physicochemical properties of MS/SH crosslinked hydrogels, and their performance for cell encapsulation. We compare these properties with the PEG-MS variant containing an electron-donating linker. The new star poly(ethylene glycol)-4-(5-(methylsulfonyl)-1H-tetrazol-1-yl)benzamide (PEG4-CONH-TzMS) shows superior properties as cell encapsulating hydrogel in terms of ease of mixing polymer precursors, faster gelation, homogenous cell distribution and high enzymatic stability.

Living Therapeutic Materials (LTMs) are a promising alternative to polymeric drug carriers for long term release of biotherapeutics. LTMs contain living drug biofactories that produce the drug using energy sources from the body fluids. To clarify their application potential, it is fundamental to adapt biocompatibility and cytotoxicity assays applied from non-living biomaterials and therapeutics to evaluate how LTMs interact with host cells. Here, we have established a first step in this direction, by developing a practical workflow to parallelize in vitro assessment of minimal safety and cytocompatibility properties of bacterial LTMs. It allows systematic monitoring and quantification of the dynamic evolution of the bacterial population (growth, metabolic activity) in parallel to quantify the response of different mammalian cells to LTM supernatants with regards to cytotoxicity and release of pro-inflammatory cytokines over a period of 7 days using a maximum of 10 samples. The protocol was tested with a Pluronic-based thin film containing ClearColi. The results show no cytotoxic effects of ClearColi containing hydrogels in three mammalian cell lines, and no induction of pro-inflammatory cytokines under the tested conditions. This workflow represents a first step in establishing a roadmap for the safety assessment of LTMs, and investigation of biocompatibility potential of future living therapeutic devices.

Side-emitting optical fibers allow light to be deliberately outcoupled along the fiber. Introducing a customized side-emission profile requires modulation of the guiding and emitting properties along the fiber length, which is a particular challenge in continuous processing of soft waveguides. In this work, it is demonstrated that multimaterial extrusion printing can generate hydrogel optical fibers with tailored segments for light-side emission. The fibers are based on diacrylated Pluronic F-127 (PluDA). 1 mm diameter fibers are printed with segments of different optical properties by switching between a PluDA waveguiding ink and a PluDA scattering ink containing nanoparticles. The method allows the fabrication of fibers with segment lengths below 500 microns in a continuous process. The length of the segments is tailored by varying the switching time between inks during printing. Fibers with customized side-emission profiles along their length are presented. The functionality of the printed fibers is demonstrated by exciting fluorescence inside a surrounding 3D hydrogel. The presented technology and material combination allow unprecedented flexibility for designing soft optical fibers with customizable optical properties using simple processes and a medical material. This approach can be of interest to improve illumination inside tissues for photodynamic therapy (PDT).

Wound healing is a dynamic biological process that leads to the repair of damaged body tissues and restores their ability to function as protective barriers. There are several approaches to handling and treating skin wounds; however, new and efficient procedures must be developed to cope with the inadequacies of the current methods, such as longer recovery time. In our novel research, we used NiSe NPs ointment and NiSe-incorporated PANI/PEO fibers for wound healing. The materials-related characteristics were studied by UV/visible spectroscopy, FTIR, SEM, DLS, and XRD. SEM images illustrate the rod-like structure of the NiSe, while bead-free NiSe-based PANI/PEO fibers are conformed from SEM with an average diameter of 330 nm. An average crystal size of 6.2 nm is measured from the XRD pattern. The antibacterial and antioxidant properties showed that NiSe-incorporated PANI/PEO fibers exhibited better response than NiSe NPs with less toxicity. NiSe NPs and NiSe-based PANI/PEO fibers give excellent wound healing potential of 89.5% ± 1.18% and 95.6% ± 0.25%, respectively. Healing response and tissue regeneration by NiSe-incorporated PANI/PEO fibers were evaluated clinically by using histopathology and interleukin-6, which indicated efficient and effective wound recovery.

Biofunctionalized polyacrylamide (PAAm) hydrogels are important 2D substrates for studying cell physics and mechanobiology. In this work, an arylmethylsulfone (MS) comonomer is developed that can be incorporated into PAAm gels under aqueous radical polymerization conditions. The resulting hydrogels show similar properties to unmodified PAAm gels, indicating that the comonomer is incorporated without affecting PAAm physical properties. The MS-containing PAAm hydrogels allow efficient conjugation of thiol derivatized biomolecules and require very low comonomer content (2 mM, 0.18 mol% relative to AAm) and thiol incubation amounts (≥ 0.15 µg per gel) to achieve functional densities that elicit cell responses. Compared to carboxyl-functionalized PAAm hydrogels, a 10-fold lower comonomer concentration and a 10-fold lower ligand feed concentration are sufficient to achieve comparable cell adhesion responses. The new comonomer opens up possibilities for efficient and straightforward biofunctionalization of PAAm hydrogels used in cell biophysical studies.