Combination of Irreversible Electroporation and STING Agonist for Effective Cancer Immunotherapy.

Recently, cancer immunotherapy has received attention as a viable solution for the treatment of refractory tumors. However, it still has clinical limitations in its treatment efficacy due to inter-patient tumor heterogeneity and immunosuppressive tumor microenvironment (TME). In this study, we demonstrated the triggering of anti-cancer immune responses by a combination of irreversible electroporation (IRE) and a stimulator of interferon genes (STING) agonist. Optimal electrical conditions inducing damage-associated molecular patterns (DAMPs) by immunogenic cell death (ICD) were determined through in vitro 2D and 3D cell experiments. In the in vivo syngeneic lung cancer model, the combination of IRE and STING agonists demonstrated significant tumor growth inhibition. We believe that the combination strategy of IRE and STING agonists has potential for effective cancer immunotherapy.

Antimony(III/V) removal from industrial wastewaters: treatment of spent catalysts formally used in the SOHIO acrylonitrile process.

A treatment and volume reduction process for a spent uranium-antimony catalyst has been developed. Targeted removal, immobilization and disposal of the uranium component has been confirmed, thus eliminating the radiological hazard. However, significant concentrations of antimony ([Sb] ≥ 25-50 mg L-1) remain in effluent from the process, which require removal in compliance with Korean wastewater regulations. Antimony(III/V) removal via co-precipitation with iron has been considered with optimal pH, dose and kinetics being determined. The effect of selected anions - Cl-, SO4 2- and PO4 3- - have also been considered, the latter present due to a prior uranium removal step. Removal of Sb(III) from both Cl- and SO4 2- media and Sb(V) removal from Cl- media to below release limits were found to be effective within 5 minutes at an iron dose of 8 mM (molar ratio, [FeIII]/[Sb] = 20) and a target pH of 5.0. However, Sb(V) removal from SO4 2- was significantly hampered requiring significantly higher iron dosages for the same removal performance. Phosphate poses significant challenges for the removal of Sb(V) due to competition between PO4 3- and Sb(OH)6 - species for surface binding sites, attributed to similarities in chemistries and a shared preference for an inner vs outer binding mechanism.

Linear Micro-patterned Drug Eluting Balloon (LMDEB) for Enhanced Endovascular Drug Delivery.

In-stent restenosis (ISR) often occurs after applying drug eluting stents to the blood vessels suffering from atherosclerosis or thrombosis. For treatment of ISR, drug eluting balloons (DEB) have been developed to deliver anti-proliferative drugs to the lesions with ISR. However, there are still limitations of DEB such as low drug delivery efficiency and drug loss to blood flow. Although most researches have focused on alteration of drug formulation for more efficient drug delivery, there are few studies that have attempted to understand and utilize the contact modality of DEB drug delivery. Here, we developed a linear micro-patterned DEB (LMDEB) that applied higher contact pressure to enhance drug stamping to vascular tissue. Ex vivo and in vivo studies confirmed that higher contact pressure from micro-patterns increased the amount of drug delivered to the deeper regions of vessel. Finite element method simulation also showed significant increase of contact pressure between endothelium and micro-patterns. Quantitative analysis by high performance liquid chromatography indicated that LMDEBs delivered 2.3 times higher amount of drug to vascular tissue in vivo than conventional DEBs. Finally, efficacy studies using both atherosclerotic and ISR models demonstrated superior patency of diseased vessels treated with LMDEB compared to those treated with DEB.

Electrospun Silk Fibroin Nanofibrous Scaffolds with Two-Stage Hydroxyapatite Functionalization for Enhancing the Osteogenic Differentiation of Human Adipose-Derived Mesenchymal Stem Cells.

The development of functional scaffolds with improved osteogenic potential is important for successful bone formation and mineralization in bone tissue engineering. In this study, we developed a functional electrospun silk fibroin (SF) nanofibrous scaffold functionalized with two-stage hydroxyapatite (HAp) particles, using mussel adhesive-inspired polydopamine (PDA) chemistry. HAp particles were first incorporated into SF scaffolds during the electrospinning process, and then immobilized onto the electrospun SF nanofibrous scaffolds containing HAp via PDA-mediated adhesive chemistry. We obtained two-stage HAp-functionalized SF nanofibrous scaffolds with improved mechanical properties and capable of providing a bone-specific physiological microenvironment. The developed scaffolds were tested for their ability to enhance the osteogenic differentiation of human adipose-derived mesenchymal stem cells (hADMSCs) in vitro and repair bone defect in vivo. To boost their ability for bone repair, we genetically modified hADMSCs with the transcriptional coactivator with PDZ-binding motif (TAZ) via polymer nanoparticle-mediated gene delivery. TAZ is a well-known transcriptional modulator that activates the osteogenic differentiation of mesenchymal stem cells (MSCs). Two-stage HAp-functionalized SF scaffolds significantly promoted the osteogenic differentiation of TAZ-transfected hADMSCs in vitro and enhanced mineralized bone formation in a critical-sized calvarial bone defect model. Our study shows the potential utility of SF scaffolds with nanofibrous structures and enriched inorganic components in bone tissue engineering.

Microchannel system for rate-controlled, sequential, and pH-responsive drug delivery.

Controlled delivery of drug at a constant rate, in a sequential order, or responsive to environment conditions has been pursued for a long time to enhance the efficacy of therapeutic molecules and to minimize side effects of highly potent drugs. However, achieving such delicately-controlled delivery of a drug molecule is non-trivial and still remains a challenge. We propose the use of microchannels to control the rate, sequence, and pH-responsiveness of drug delivery for high precision and predictability. In this study, we introduce elementary drug delivery units consisting of micro-reservoirs and microchannels that have variations in their lengths, widths, numbers, and straightness. The release study demonstrates that the release rates of model drugs can be modulated by the design of microchannels. Finite element modeling of drug release predicts the performance of the drug delivery units with high accuracy. The possibility of sequential drug delivery is also demonstrated using biodegradable polymer plug in microchannels. Finally, pH-responsive delivery of drugs in microfluidic units is also discussed and demonstrated via cell viability tests. STATEMENT OF SIGNIFICANCE: In this work, we developed microchannel-based drug delivery devices whose release rate could be accurately calculated and controlled by design of microchannel geometry. Although there have been many advances in microfabricated drug delivery systems, in particular, reservoir-based systems, no systematic investigation has been made to utilize the release channels. In our work, an equivalent electrical circuit concept was applied to the microfluidic systems for more detailed design and analysis. A microfluidic channel was regarded as an electrical resistor; their diffusion/electrical flux could be tuned with geometric factors such as length, width, a number of channel/resistor and their connections. Furthermore, from delivery rate control using channel geometry, multifunctional channel-based release systems for sequential and pH-responsive were demonstrated.

Varicose and cheerio collaborate with pebble to mediate semaphorin-1a reverse signaling in Drosophila.

The transmembrane semaphorin Sema-1a acts as both a ligand and a receptor to regulate axon-axon repulsion during neural development. Pebble (Pbl), a Rho guanine nucleotide exchange factor, mediates Sema-1a reverse signaling through association with the N-terminal region of the Sema-1a intracellular domain (ICD), resulting in cytoskeletal reorganization. Here, we uncover two additional Sema-1a interacting proteins, varicose (Vari) and cheerio (Cher), each with neuronal functions required for motor axon pathfinding. Vari is a member of the membrane-associated guanylate kinase (MAGUK) family of proteins, members of which can serve as scaffolds to organize signaling complexes. Cher is related to actin filament cross-linking proteins that regulate actin cytoskeleton dynamics. The PDZ domain binding motif found in the most C-terminal region of the Sema-1a ICD is necessary for interaction with Vari, but not Cher, indicative of distinct binding modalities. Pbl/Sema-1a-mediated repulsive guidance is potentiated by both vari and cher Genetic analyses further suggest that scaffolding functions of Vari and Cher play an important role in Pbl-mediated Sema-1a reverse signaling. These results define intracellular components critical for signal transduction from the Sema-1a receptor to the cytoskeleton and provide insight into mechanisms underlying semaphorin-induced localized changes in cytoskeletal organization.

Differential ligand regulation of PlexB signaling in motor neuron axon guidance in Drosophila.

Plexins (Plexs) are a large family of phylogenetically conserved guidance receptors that bind specifically to semaphorins (Semas), another large family of guidance molecules. In the Drosophila embryonic central nervous system (CNS), the secreted semaphorins Sema-2a and Sema-2b both act as ligands for PlexB, but mediate mutually independent and opposite functions (repulsive and attractive guidance, respectively). PlexB is also known to regulate motor axon guidance in the embryonic peripheral nervous system (PNS). However, it is unclear whether the mechanisms of ligand regulation of PlexB seen in the CNS are similar or the same as those that exist in PNS motor axon guidance. Here, we find that two distinct modes of ligand regulation underlie differential roles of PlexB in PNS motor axon pathfinding during embryonic development. Epistasis analyses in the intersegmental nerve b (ISNb) pathway suggest that PlexB serves as a receptor for both Sema-2a and Sema-2b and integrates their mutually dependent but opposite guidance functions. Furthermore, we present evidence that PlexB mediates not only Sema-2a/2b-dependent guidance functions, but also Sema-2a/2b-independent target recognition in establishing the segmental nerve a (SNa) motor axon pathway. These results demonstrate that a single guidance receptor can elicit diverse effects on the establishment of neuronal connectivity via regulation of its ligands themselves.

The axon guidance function of Rap1 small GTPase is independent of PlexA RasGAP activity in Drosophila.

Plexins (Plexs) comprise a large family of cell surface receptors for semaphorins (Semas) that function as evolutionarily conserved guidance molecules. GTPase activating protein (GAP) activity for Ras family small GTPases has been implicated in plexin signaling cascades through its RasGAP domain. However, little is known about how Ras family GTPases are controlled in vivo by plexin signaling. Here, we found that Drosophila Rap1, a member of the Ras family of GTPases, plays an important role controlling intersegmental nerve b motor axon guidance during neural development. Gain-of-function studies using dominant-negative and constitutively active forms of Rap1 indicate that Rap1 contributes to axonal growth and guidance. Genetic interaction analyses demonstrate that the Sema-1a/PlexA-mediated repulsive guidance function is regulated positively by Rap1. Furthermore, neuronal expression of mutant PlexA robustly restored defasciculation defects in PlexA null mutants when the catalytic arginine fingers of the PlexA RasGAP domain critical for GAP activity were disrupted. However, deleting the RasGAP domain abolished the ability of PlexA to rescue the PlexA guidance phenotypes. These findings suggest that PlexA-mediated motor axon guidance is dependent on the presence of the PlexA RasGAP domain, but not on its GAP activity toward Ras family small GTPases.

Three-Dimensional Rapid Prototyping of Multidirectional Polymer Nanoprobes for Single Cell Insertion.

Three-dimensional (3D) thermal drawing at nanoscale as a novel rapid prototyping method was demonstrated to create multidirectional polymer nanoprobes for single cell analysis. This 3D drawing enables simple and rapid fabrication of polymeric nanostructures with high aspect ratio. The effect of thermal drawing parameters, such as drawing speeds, dipping depths, and contact duration on the final geometry of polymer nanostructures was investigated. Vertically aligned and L-shaped nanoprobes were fabricated and their insertion into living single cells such as algal cells and human neural stem cells was demonstrated. This technique can be extended to create more complex 3D structures by controlling drawing steps and directions on any surface.