Optimized Design of Hyaluronic Acid-Lipid Conjugate Biomaterial for Augmenting CD44 Recognition of Surface-Engineered NK Cells.

Triple-negative breast cancer (TNBC) presents treatment challenges due to a lack of detectable surface receptors. Natural killer (NK) cell-based adaptive immunotherapy is a promising treatment because of the characteristic anticancer effects of killing malignant cells directly by secreting cytokines and lytic granules. To maximize the cancer recognition ability of NK cells, biomaterial-mediated ex vivo cell surface engineering has been developed for sufficient cell membrane immobilization of tumor-targeting ligands via hydrophobic anchoring. In this study, we optimized amphiphilic balances of NK cell coating materials composed of CD44-targeting hyaluronic acid (HA)-poly(ethylene glycol) (PEG)-lipid to improve TNBC recognition and the anticancer effect. Changes in the modular design of our material by differentiating hydrophilic PEG length and incorporating lipid amount into HA backbones precisely regulated the amphiphilic nature of HA-PEG-lipid conjugates. The optimized biomaterial demonstrated improved anchoring into NK cell membranes and facilitating the surface presentation level of HA onto NK cell surfaces. This led to enhanced cancer targeting via increasing the formation of immune synapse, thereby augmenting the anticancer capability of NK cells specifically toward CD44-positive TNBC cells. Our approach addresses targeting ability of NK cell to solid tumors with a deficiency of surface tumor-specific antigens while offering a valuable material design strategy using amphiphilic balance in immune cell surface engineering techniques.

Effects of micro-sized biodegradable plastics on Microcystis aeruginosa.

Plethora of plastics are being used in current society, generating huge amounts of plastic waste. Non-biodegradability of conventional plastics is one of the main challenges to treat plastic waste. In an effort to increase the efficiency of plastic waste treatment, biodegradable plastics have gained attention. Although the use of biodegradable plastics has been increased, their potential effects on the environments are not fully elucidated yet. In this study, the impacts of micro-sized non-biodegradable plastic (i.e., polystyrene (PS)) and micro-sized biodegradable plastics (i.e., polycaprolactone (PCL) and polylactic acid (PLA)) on Microcystis aeruginosa were investigated. Regardless of microplastic (MP) types, MP treatments inhibited the growth of M. aeruginosa at the beginning (4 days) while significant dose-dependent effect was not observed in the range of 0.1 to 10 mg/L. However, after long-term exposure (12 days), micro-sized biodegradable plastics stimulated the growth of M. aeruginosa (up to 73 % increase compared to the control). The photosynthetic activity showed a similar trend to the cell growth. The MP treatments induced the production of extracellular polymeric substances (EPS). Indeed, micro-sized PCL and PLA stimulated the production of protein compounds in EPS. These might have affected the releases of chemicals from PCL and PLA, suggesting that the chemicals in biodegradable plastic leachates would promote the growth of M. aeruginosa in long-term exposure. The MP treatments also induced cyanotoxin (microcystin-LR) productions. Our results give a new insight into the cyanobacterial blooming and suggest a novel relationship between harmful algal blooms (HABs) and biodegradable plastics.

Micro-macro hybrid soft-lithography master (MMHSM) fabrication for lab-on-a-chip applications.

We present a novel micro-macro hybrid soft-lithography master (MMHSM) fabrication technique where microdevices having both microscale and macroscale features can be replicated with a single soft-lithography step. A poly(methyl methacrylate) (PMMA) master having macroscale structures was first created by a bench-top milling machine. An imprinting master mold having microscale structures was then imprinted on the PMMA surface through a hot-embossing process to obtain a PMMA master mold. A poly(dimethylsiloxane) (PDMS) master was then replicated from this PMMA master through a standard soft-lithography process. This process allowed both microscale (height: 3-20 microm, width: 20-500 microm) and macroscale (height: 3.5 mm, width: 1.2-7 mm) structures to co-exist on the PDMS master mold, from which final PDMS devices could be easily stamped out in large quantities. Microfluidic structures requiring macroscale dimensions in height, such as reservoirs or fluidic tubing interconnects, could be directly built into PDMS microfluidic devices without the typically used manual punching process. This significantly reduced alignment errors and time required for such manual fabrication steps. In this paper, we successfully demonstrated the utility of this novel hybrid fabrication method by fabricating a PDMS microfluidic device with 40 built-in fluidic interfaces and a PDMS multi-compartment neuron co-culture platform, where millimeter-scale compartments are connected via arrays of 20 microm wide and 200 microm long microfluidic channels. The resulting structures were characterized for the integrity of the transferred pattern sizes and the surface roughness using scanning electron microscopy and optical profilometry.

Engineering High-Resolution Micropatterns Directly onto Titanium with Optimized Contact Guidance to Promote Osteogenic Differentiation and Bone Regeneration.

Topographical cues play an important role in directing cell behavior, and thus, extensive research efforts have been devoted to fabrication of surface patterns and exploring the contact guidance effect. However, engineering high-resolution micropatterns directly onto metallic implants remains a grand challenge. Moreover, there still lacks evidence that allows translation of in vitro screening to in vivo tissue response. Herein, we demonstrate a fast, cost-effective, and feasible approach to the precise fabrication of shape- and size-controlled micropatterns on titanium substrates using a combination of photolithography and inductively coupled plasma-based dry etching. A titanium TopoChip containing 34 microgrooved patterns with varying geometry parameters and a flat surface as the control was designed for a high-throughput in vitro study of the contact guidance of osteoblasts. The correlation between the surface pattern dimensions, cell morphological characteristics, proliferation, and osteogenic marker expression was systematically investigated in vitro. Furthermore, the surface with the highest osteogenic potential in vitro along with representative controls was evaluated in rat cranial defect models. The results show that microgrooved pattern parameters have almost no effect on osteoblast proliferation but significantly regulate the cell morphology, orientation, focal adhesion (FA) formation, and osteogenic differentiation in vitro. In particular, a specific groove pattern with a ridge width of 3 µm, groove width of 7 µm, and depth of 2 µm can most effectively align the cells through regulating the distribution of FAs, resulting in an anisotropic actin cytoskeleton, and thereby promoting osteogenic differentiation. In vivo, microcomputed tomography and histological analyses show that the optimized pattern can apparently stimulate new bone formation. This study not only offers a microfabrication method that can be extended to fabricate various shape- and size-controlled micropatterns on titanium alloys but also provides insight into the surface structure design of orthopedic and dental implants for enhanced bone regeneration.

A multi-compartment CNS neuron-glia Co-culture microfluidic platform.

We present a novel multi-compartment neuron co-culture microsystem platform for in vitro CNS axon-glia interaction research, capable of conducting up to six independent experiments in parallel for higher-throughput. We developed a new fabrication method to create microfluidic devices having both micro and macro scale structures within the same device through a single soft-lithography process, enabling mass fabrication with good repeatability. The multi-compartment microfluidic co-culture platform is composed of one soma compartment for neurons and six axon/glia compartments for oligodendrocytes (OLs). The soma compartment and axon/glia compartments are connected by arrays of axon-guiding microchannels that function as physical barriers to confine neuronal soma in the soma compartment, while allowing axons to grow into axon/glia compartments. OLs loaded into axon/glia compartments can interact only with axons but not with neuronal soma or dendrites, enabling localized axon-glia interaction studies. The microchannels also enabled fluidic isolation between compartments, allowing six independent experiments to be conducted on a single device for higher throughput. Soft-lithography using poly(dimethylsiloxane) (PDMS) is a commonly used technique in biomedical microdevices. Reservoirs on these devices are commonly defined by manual punching. Although simple, poor alignment and time consuming nature of the process makes this process not suitable when large numbers of reservoirs have to be repeatedly created. The newly developed method did not require manual punching of reservoirs, overcoming such limitations. First, seven reservoirs (depth: 3.5 mm) were made on a poly(methyl methacrylate) (PMMA) block using a micro-milling machine. Then, arrays of ridge microstructures, fabricated on a glass substrate, were hot-embossed against the PMMA block to define microchannels that connect the soma and axon/glia compartments. This process resulted in macro-scale reservoirs (3.5 mm) and micro-scale channels (2.5 microm) to coincide within a single PMMA master. A PDMS replica that served as a mold master was obtained using soft-lithography and the final PDMS device was replicated from this master. Primary neurons from E16-18 rats were loaded to the soma compartment and cultured for two weeks. After one week of cell culture, axons crossed microchannels and formed axonal only network layer inside axon/glia compartments. Axons grew uniformly throughout six axon/glia compartments and OLs from P1-2 rats were added to axon/glia compartments at 14 days in vitro for co-culture.