Antibiofilm Activity of Eugenol-Loaded Chitosan Coatings against Common Medical-Device-Contaminating Bacteria.

The formation of pathogenic biofilms on medical devices is a major public health concern accounting for over 65% of healthcare-associated infections and causing high infection morbidity, mortality, and a great burden to patients and the healthcare system due to its resistance to treatment. In this study, we developed a chitosan-based antimicrobial coating with embedded mesoporous silica nanoparticles (MSNs) to load and deliver eugenol, an essential oil component, to inhibit the biofilm formation of common bacteria in medical-device-related infections. The eugenol-loaded MSNs were dispersed in a chitosan solution, which was then cross-linked with glutaraldehyde and drop-casted to obtain coatings. The MSNs and coatings were characterized by dynamic light scattering, Brunauer-Emmett-Teller analysis, attenuated-total-reflectance Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, 3D optical profilometry, and scanning electron microscopy. The release behavior of eugenol-loaded MSNs and coatings and the antibiofilm and antimicrobial activity of the coatings against adherent Staphylococcus aureus, methicillin-resistant S. aureus, and Pseudomonas aeruginosa were investigated. Eugenol was released from the MSNs and coatings in aqueous conditions in a controlled manner with an initial low release, followed by a peak release, a decrease, and a plateau. While the chitosan coatings alone or with unloaded MSNs demonstrated limited antimicrobial effects and still supported biofilm formation after 24 h, the coating containing eugenol not only reduced biofilm formation but also killed the majority of the attached bacteria. It also showed biocompatibility in indirect contact with NIH/3T3 fibroblasts and a high percentage of live cells in direct contact. However, further investigations into cell proliferation in direct contact are recommended. The findings indicated that the chitosan-based coating with eugenol-loaded MSNs could be developed into an effective strategy to inhibit biofilm formation on medical devices.

Strategic Replication of the Hepatic Zonation In Vitro Employing a Biomimetic Approach.

The varied functions of the liver are dependent on the metabolic heterogeneity exhibited by the hepatocytes within the liver lobule spanning the porto-central axis. This complex phenomenon plays an important role in maintaining the physiological homeostasis of the liver. Standard in vitro culture models fail to mimic this spatial heterogeneity of hepatocytes, assuming a homogeneous population of cells, which leads to inaccurate translation of results. Here, we demonstrate the development of an in vitro model of hepatic zonation by mimicking the microarchitecture of the liver using a 3D printed mini bioreactor and decellularized liver matrix to provide the native microenvironmental cues. There was a differential expression of hypoxic and metabolic markers across the developed mini bioreactor, showing the establishment of gradients of oxygen, Wnt/ß-catenin pathway, and other metabolic pathways. The model also showed the establishment of zone-dependent toxicity on treatment with acetaminophen. The developed model would thus be a promising avenue in the field of tissue engineering for understanding the liver physiology and pathophysiology and for drug screening to evaluate the potential of new pharmaceutical interventions.

Highly active nisin coated polycaprolactone electrospun fibers against both Staphylococcus aureus and Pseudomonas aeruginosa.

In this study, a wound dressing of electrospun polycaprolactone (PCL) fibers incorporating the antimicrobial peptide (AMP) nisin was fabricated. Nisin was physically adsorbed to the PCL fibers and tested for antibacterial activity against both Staphylococcus aureus (S. aureus) and Pseudomonas aeruginosa (P. aeruginosa). The PCL fibers had an average diameter of 1.16 µm ± 0.42 µm and no significant change in diameter occurred after nisin adsorption. X-ray photoelectron spectroscopy (XPS) analysis of the fibers detected nitrogen indicative of adsorbed nisin and the signal was used to quantify the levels of coverage on the fiber surfaces. In vitro nisin release studies showed a burst release profile with 80 % of the nisin being released from the fibers within 30 min. Air plasma pre-treatment of the PCL fibers to render them hydrophilic improved nisin loading and release. Antibacterial testing was performed using minimum inhibitory concentration (MIC) and surface attachment assays. The released nisin remained active against both Gram positive S. aureus and Gram negative P. aeruginosa, which has previously been difficult to achieve with single polymer fiber systems. Mammalian cell culture of the nisin coated fibers with L-929 mouse fibroblasts and human epidermal keratinocytes (HEKa) showed that the nisin did not have a significant effect on the biocompatibility of the PCL fibers. The results presented here demonstrate that the physical adsorption, which is a post-treatment, overcomes the potential limitations of harsh chemicals and fabrication conditions of electrospinning from organic solvents and provides a drug loading system having effective antibacterial properties in wound dressings.

Enhancing the Activity of Surface Immobilized Antimicrobial Peptides Using Thiol-Mediated Tethering to Poly(ethylene glycol).

Considering the need for versatile surface coatings that can display multiple bioactive signals and chemistries, the use of more novel surface modification methods is starting to emerge. Thiol-mediated conjugation of biomolecules is shown to be quite advantageous for such purposes due to the reactivity and chemoselectivity of thiol functional groups. Herein, the immobilization of poly(ethylene glycol) (PEG) and antimicrobial peptides (AMPs) to silica colloidal particles based on thiol-mediated conjugation techniques, along with an assessment of the antimicrobial potential of the functionalized particles against Pseudomonas aeruginosa and Staphylococcus aureus is investigated. Immobilization of PEG to thiolated Si particles is performed by either a two-step thiol-ene "photo-click" reaction or a "one-pot" thiol-maleimide type conjugation using terminal acrylate or maleimide functional groups, respectively. It is demonstrated that both immobilization methods result in a significant reduction in the number of viable bacterial cells compared to unmodified samples after the designated incubation periods with the PEG-AMP-modified colloidal suspensions. These findings provide a promising outlook for the fabrication of multifunctional surfaces based upon the tethering of PEG and AMPs to colloidal particles through thiol-mediated biocompatible chemistry, which has potential for use as implant coatings or as antibacterial formulations that can be incorporated into wound dressings to prevent or control bacterial infections.

Magnetically Cured Macroradical Epoxy as Antimicrobial Coating.

The radical-bearing epoxy monomer could be the ideal embodiment of multifunctionality in epoxy-based materials. This study demonstrates the potential of macroradical epoxies as surface coating materials. A diepoxide monomer derivatized with a stable nitroxide radical is polymerized with a diamine hardener under the influence of a magnetic field. The magnetically oriented and stable radicals in the polymer backbone render the coatings antimicrobial. The unconventional use of magnets during polymerization proved crucial in correlating the structure-property relationships with antimicrobial performance inferred from oscillatory rheological technique, polarized macro-attenuated total reflectance - infrared (macro-ATR-IR) spectroscopy and X-ray photoelectron spectroscopy (XPS). The magnetic thermal curing influenced the surface morphology, resulting in a synergy of the coating's radical nature with microbiostatic performance assessed using the Kirby-Bauer test and liquid chromatography - mass spectroscopy (LC-MS). Further, the magnetic curing of blends with a traditional epoxy monomer demonstrates that radical alignment is more critical than radical density in imparting biocidal behavior. This study shows how the systematic use of magnets during polymerization could pave for probing more significant insights into the mechanism of antimicrobial action in radical-bearing polymers.

Recent Advances in Using Natural Antibacterial Additives in Bioactive Wound Dressings.

Wound care is a global health issue with a financial burden of up to US $96.8 billion annually in the USA alone. Chronic non-healing wounds which show delayed and incomplete healing are especially problematic. Although there are more than 3000 dressing types in the wound management market, new developments in more efficient wound dressings will require innovative approaches such as embedding antibacterial additives into wound-dressing materials. The lack of novel antibacterial agents and the misuse of current antibiotics have caused an increase in antimicrobial resistance (AMR) which is estimated to cause 10 million deaths by 2050 worldwide. These ongoing challenges clearly indicate an urgent need for developing new antibacterial additives in wound dressings targeting microbial pathogens. Natural products and their derivatives have long been a significant source of pharmaceuticals against AMR. Scrutinising the data of newly approved drugs has identified plants as one of the biggest and most important sources in the development of novel antibacterial drugs. Some of the plant-based antibacterial additives, such as essential oils and plant extracts, have been previously used in wound dressings; however, there is another source of plant-derived antibacterial additives, i.e., those produced by symbiotic endophytic fungi, that show great potential in wound dressing applications. Endophytes represent a novel, natural, and sustainable source of bioactive compounds for therapeutic applications, including as efficient antibacterial additives for chronic wound dressings. This review examines and appraises recent developments in bioactive wound dressings that incorporate natural products as antibacterial agents as well as advances in endophyte research that show great potential in treating chronic wounds.

Surface Characteristics and Bone Biocompatibility of Cold-Sprayed Porous Titanium on Polydimethylsiloxane Substrates.

A variant of the cold spray (CS) technique was applied for the functionalization of polymer-based materials such as polydimethylsiloxane (PDMS) to improve the extent of mammalian cell interactions with these substrates. This was demonstrated by the embedment of porous titanium (pTi) into PDMS substrates using a single-step CS technique. CS processing parameters such as gas pressure and temperature were optimized to achieve the mechanical interlocking of pTi in the compressed PDMS to fabricate a unique hierarchical morphology possessing micro-roughness. As evidenced by the preserved porous structure, the pTi particles did not undergo any significant plastic deformation upon impact with the polymer substrate. The thickness of the particle embedment layer was determined, by cross-sectional analysis, ranging from 120 µm to over 200 µm. The behavior of osteoblast-like cells MG63 coming into contact with the pTi-embedded PDMS was examined. The results showed that the pTi-embedded PDMS samples promoted 80-96% of cell adhesion and proliferation during the early stages of incubation. The low cytotoxicity of the pTi-embedded PDMS was confirmed, with cell viability of the MG63 cells being above 90%. Furthermore, the pTi-embedded PDMS facilitated the production of alkaline phosphatase and calcium deposition in the MG63 cells, as demonstrated by the higher amount of alkaline phosphatase (2.6 times) and calcium (10.6 times) on the pTi-embedded PDMS sample fabricated at 250 °C, 3 MPa. Overall, the work demonstrated that the CS process provided flexibility in the parameters used for the production of the modified PDMS substrates and is highly efficient for the fabrication of coated polymer products. The results obtained in this study suggest that a tailorable porous and rough architecture could be achieved that promoted osteoblast function, indicating that the method has promise in the design of titanium-polymer composite materials applied to biomaterials used in musculoskeletal applications.

Biomimicking nanofibrous gelatin microspheres recreating the stem cell niche for their ex-vivo expansion and in-vivo like differentiation for injectable stem cell transplantation.

Stem cells based novel treatment modality for degenerative and immune dysfunction diseases created a huge demand of suitable carriers to support ex-vivo production of quality stem cells, and effective in-vivo transplantation of stem cells and their fate. In spite of promising candidature of nanofibrous microspheres (NFM) to recreate native stem cell niches to be used for possible scaling-up for ex-vivo stem cells expansion, it remains fairly unexplored. A systematic study on the stem cell-NFM interaction comparative with commercial microspheres (CM) has been performed for the first time. Gelatin NFM with variable physicochemical properties such as size, surface properties, surface chemistry, and variable degradability were prepared using microemulsion coupled with thermally induced phase separation (TIPS) method. Effect of physicochemical properties of NFM and their cellular interaction such as binding, morphology, metabolic activity and proliferation studies were performed using human bone marrow-derived mesenchymal stem cells (hBMSCs), human dental follicle stem cells (hDFSCs) and human gingival fibroblast (HGF) cells and compared with the commercial and solid microspheres. Gelatin NFM supports excellent cell binding, proliferation, metabolic activities and chemical cues specific differentiation. All out-turns indicate that NFM stand to be an outstanding candidate for ex-vivo cells' expansion and injectable carriers for stem cell transplantation.

Galactose Tethered Decellularized Liver Matrix: Toward a Biomimetic and Biofunctional Matrix for Liver Tissue Engineering.

The major challenge in liver tissue engineering is the replication of the microenvironment and microarchitecture of the liver tissue at the nanoscale. Decellularized liver matrix (DLM) provides an ideal material for scaffold preparation, as it retains the relevant structural and biochemical composition. However, the loss of bioactive factors during decellularization needs to be taken into account when using DLM and should be supplemented accordingly for an expected outcome. This study reports on the modification of DLM by the addition of galactose residues using a two-step thiol-ene-mediated photoclick chemistry for the coupling of galactose moieties to the DLM. Modification with galactose enhanced the function of hepatocytes and provides many advantages over currently used DLM and DLM-based materials. The galactose modified DLM enhanced the initial HepG2 cell adhesion to the substrate with changes in dynamics over time such as spheroid formation and further migration on the matrix. Our observation is that the galactose ligand decoration can also enhance the liver-specific metabolism of HepG2 compared to unmodified DLM. Galactosylated DLM also showed a better establishment of cellular polarity which also contributes to the function of HepG2 cells. Together our results demonstrate the advantages of adding galactose residues to currently available biomaterials, which makes this approach an attractive method for ECM-based liver tissue engineering.

The Mechanisms of Restenosis and Relevance to Next Generation Stent Design.

Stents are lifesaving mechanical devices that re-establish essential blood flow to the coronary circulation after significant vessel occlusion due to coronary vessel disease or thrombolytic blockade. Improvements in stent surface engineering over the last 20 years have seen significant reductions in complications arising due to restenosis and thrombosis. However, under certain conditions such as diabetes mellitus (DM), the incidence of stent-mediated complications remains 2-4-fold higher than seen in non-diabetic patients. The stents with the largest market share are designed to target the mechanisms behind neointimal hyperplasia (NIH) through anti-proliferative drugs that prevent the formation of a neointima by halting the cell cycle of vascular smooth muscle cells (VSMCs). Thrombosis is treated through dual anti-platelet therapy (DAPT), which is the continual use of aspirin and a P2Y12 inhibitor for 6-12 months. While the most common stents currently in use are reasonably effective at treating these complications, there is still significant room for improvement. Recently, inflammation and redox stress have been identified as major contributing factors that increase the risk of stent-related complications following percutaneous coronary intervention (PCI). The aim of this review is to examine the mechanisms behind inflammation and redox stress through the lens of PCI and its complications and to establish whether tailored targeting of these key mechanistic pathways offers improved outcomes for patients, particularly those where stent placement remains vulnerable to complications. In summary, our review highlights the most recent and promising research being undertaken in understanding the mechanisms of redox biology and inflammation in the context of stent design. We emphasize the benefits of a targeted mechanistic approach to decrease all-cause mortality, even in patients with diabetes.

A nanofiber based antiviral (TAF) prodrug delivery system.

HIV and hepatitis B are two of the most prevalent viruses globally, and despite readily available preventive treatments unforgiving treatment regimens still exist, such as daily doses of medicine that are challenging to maintain especially in poorer countries. More advanced and longer-lasting delivery vehicles can potentially overcome this problem by reducing maintenance requirements and significantly increase access to medicine. Here, we designed a technology to control the delivery of an antiviral drug over a long timeframe via a nanofiber based delivery scaffold that is both easy to produce and use. An antiviral prodrug containing tenofovir alafenamide (TAF) was synthesized by initial conjugation to glycerol monomethacrylate followed by polymerization to form a diblock copolymer (pTAF) using reversible addition-fragmentation chain transfer (RAFT). In order to generate an efficient drug delivery system this copolymer was fabricated into an electrospun nanofiber (ESF) scaffold using blend electrospinning with poly(caprolactone) (PCL) as the carrier polymer. SEM images revealed that the pTAF-PCL ESFs were uniform with an average diameter of (787 ± 0.212 nm), while XPS analysis demonstrated that the pTAF was overrepresented at the surface of the ESFs. Additionally, the pTAF exhibited a sustained release profile over a 2 month period in human serum (HS), suggesting that these types of copolymer-based drugamers can be used in conjunction with electrospinning to produce long-lasting drug delivery systems.

Influence of Liver Extracellular Matrix in Predicting Drug-Induced Liver Injury: An Alternate Paradigm.

In vitro drug-induced liver injury (DILI) models are promising tools for drug development to predict adverse events during clinical usage. However, the currently available DILI models are not specific or not able to predict the injury accurately. This is believed to be mainly because of failure to conserve the hepatocyte phenotype, lack of longevity, and difficulty in maintaining the tissue-specific microenvironment. In this study, we have assessed the potential of decellularized liver extracellular matrix (DLM) in retaining the hepatic cellular phenotype and functionality in the presence of a tissue-specific microenvironment along with its role in influencing the effect of the drug on hepatic cells. We show that DLM helps maintain the phenotype of the hepatic cell line HepG2, a well-known cell line for secretion of human proteins that is easily available. Also, the DLM enhanced the expression of a metabolic marker carbamoyl phosphate synthetase I (CPS1), a regulator of urea cycle, and bile salt export pump (BSEP), a marker of hepatocyte polarity. We further validated the DLM for its influence on the sensitivity of cells toward different classes of drugs. Interestingly, the coculture model, in the presence of endothelial cells and stellate cells, exhibited a higher sensitivity for both acetaminophen and trovafloxacin, a toxic compound that does not show any toxicity on preclinical screening. Thus, our results demonstrate for the first time that a multicellular combination along with DLM can be a potential and reliable DILI model to screen multiple drugs.

Advancing of 3D-Printed Titanium Implants with Combined Antibacterial Protection Using Ultrasharp Nanostructured Surface and Gallium-Releasing Agents.

This paper presents the development of advanced Ti implants with enhanced antibacterial activity. The implants were engineered using additive manufacturing three-dimensional (3D) printing technology followed by surface modification with electrochemical anodization and hydrothermal etching, to create unique hierarchical micro/nanosurface topographies of microspheres covered with sharp nanopillars that can mechanically kill bacteria in contact with the surface. To achieve enhanced antibacterial performance, fabricated Ti implant models were loaded with gallium nitrate as an antibacterial agent. The antibacterial efficacy of the fabricated substrates with the combined action of sharp nanopillars and locally releasing gallium ions (Ga3+) was evaluated toward Staphylococcus aureus and Pseudomonas aeruginosa. Results confirm the significant antibacterial performance of Ga3+-loaded substrates with a 100% eradication of bacteria. The nanopillars significantly reduced bacterial attachment and prevented biofilm formation while also killing any bacteria remaining on the surface. Furthermore, 3D-printed surfaces with microspheres of diameter 5-30 µm and interspaces of 12-35 µm favored the attachment of osteoblast-like MG-63 cells, as confirmed via the assessment of their attachment, proliferation, and viability. This study provides important progress toward engineering of next-generation 3D-printed implants, that combine surface chemistry and structure to achieve a highly efficacious antibacterial surface with dual cytocompatibility to overcome the limitations of conventional Ti implants.

Decontamination-Induced Modification of Bioactivity in Essential Oil-Based Plasma Polymer Coatings.

Plasma polymer coatings fabricated from Melaleuca alternifolia essential oil and its derivatives have been previously shown to reduce the extent of microbial adhesion on titanium, polymers, and other implantable materials used in dentistry. Previous studies have shown these coatings to maintain their performance under standard operating conditions; however, when used in e.g., a dental implant, these coatings may inadvertently become subject to in situ cleaning treatments, such as those using an atmospheric pressure plasma jet, a promising tool for the effective in situ removal of biofilms from tissues and implant surfaces. Here, we investigated the effect of such an exposure on the antimicrobial performance of the Melaleuca alternifolia polymer coating. It was found that direct exposure of the polymer coating surface to the jet for periods less than 60 s was sufficient to induce changes in its surface chemistry and topography, affecting its ability to retard subsequent microbial attachment. The exact effect of the jet exposure depended on the chemistry of the polymer coating, the length of plasma treatment, cell type, and incubation conditions. The change in the antimicrobial activity for polymer coatings fabricated at powers of 20-30 W was not statistically significant due to their limited baseline bioactivity. Interestingly, the bioactivity of polymer coatings fabricated at 10 and 15 W against Staphylococcus aureus cells was temporarily improved after the treatment, which could be attributed to the generation of loosely attached bioactive fragments on the treated surface, resulting in an increase in the dose of the bioactive agents being eluted by the surface. Attachment and proliferation of Pseudomonas aeruginosa cells and mixed cultures were less affected by changes in the bioactivity profile of the surface. The sensitivity of the cells to the change imparted by the jet treatment was also found to be dependent on their origin culture, with mature biofilm-derived P. aeruginosa bacterial cells showing a greater ability to colonize the surface when compared to its planktonic broth-grown counterpart. The presence of plasma-generated reactive oxygen and nitrogen species in the culture media was also found to enhance the bioactivity of polymer coatings fabricated at power levels of 10 and 15 W, due to a synergistic effect arising from simultaneous exposure of cells to reactive oxygen and nitrogen species (RONS) and eluted bioactive fragments. These results suggest that it is important to consider the possible implications of inadvertent changes in the properties and performance of plasma polymer coatings as a result of exposure to in situ decontamination, to both prevent suboptimal performance and to exploit possible synergies that may arise for some polymer coating-surface treatment combinations.

Colloidal Self-Assembled Patterns Maintain the Pluripotency and Promote the Hemopoietic Potential of Human Embryonic Stem Cells.

The generation of blood cells in a significant amount for clinical uses is still challenging. Human pluripotent stem cells-derived hemopoietic cells (hPSC-HCs) are a promising cell source to generate blood cells. Previously, it has been shown that the attached substrates are crucial in the maintenance or differentiation of hPSCs. In this study, a new family of artificial extracellular matrix (ECM) called colloidal self-assembled patterns (cSAPs: #1-#5) was used for the expansion of mouse and human PSCs. The optimized cSAP (i.e., #4 and #5) was selected for subsequent hemopoietic differentiation of human embryonic stem cells (hESCs). Results showed that the hematopoietic potential of hESCs was enhanced approx 3-4 folds on cSAP #5 compared to the flat control. The cell population of hematopoietic progenitors (i.e., CD34+CD43+ cells) and erythroid progenitors (i.e., CD71+GPA+ cells) were enhanced 4 folds at day 8 and 3 folds at day 14. RNA sequencing analysis of cSAP-derived hESCs showed that there were 300 genes up-regulated and 627 genes down-regulated compared to the flat control. The enriched signaling pathways, including up-regulation (i.e., Toll-like receptor, HIF-1a, and Notch) or down-regulation (i.e., FAs, MAPK, JAK/STAT, and TGF-ß) were classic in the maintenance of hESC phenotype Real time PCR confirmed that the expression of focal adhesion (PTK2, VCL, and CXCL14) and MAPK signaling (CAV1) related genes was down-regulated 2-3 folds compared to the flat control. Altogether, cSAP enhances the pluripotency and the hematopoietic potential of hESCs that subsequently generates more blood-like cells. This study reveals the potential of cSAPs on the expansion and early-stage blood cell lineage differentiation of hPSCs.

Conductive, Acid-Doped Polyaniline Electrospun Nanofiber Gas Sensing Substrates Made Using a Facile Dissolution Method.

A novel dissolution method that allows for the total solvation of high-concentration, high-molecular-weight polyaniline (PANi) doped with (+)-camphor-10-sulfonic acid (CSA) is reported. Preparation of 12-16 wt % 65,000 Da PANi solutions in N,N-dimethylformamide is achievable using a simple one-pot method. Doped polyaniline solutions in common organic solvents were processed into nanofibers using a convenient single-nozzle electrospinning technique. The electrospinning of PANi-CSA into nanofibrous membranes generated substrates that were subsequently employed in colorimetric gas sensing. These substrates demonstrated linearity of response upon exposure to 50-5500 ppm ammonia at ambient (50 ± 10% RH) and high (80% RH) humidity.

Harnessing Colloidal Self-Assembled Patterns (cSAPs) to Regulate Bacterial and Human Stem Cell Response at Biointerfaces In Vitro and In Vivo.

The generation of complex physicochemical signals on the surface of biomedical materials is still challenging despite the fact that a broad range of surface modification methods have been developed over the last few decades. Colloidal self-assembled patterns (cSAPs) are combinations of unique colloids differing in size and surface chemistry acting as building blocks that can be programmed to generate surface patterns with exquisite control of complexity. This study reports on producing a variety of pre-modified colloids for the fabrication of cSAPs as well as post-assembly modifications to yield complex surfaces. The surface of cSAPs presents hierarchical micro- and nanostructures, localized hydrophilic/hydrophobic characteristics, and tunable surface functionality imparted by the individual colloids. The selected cSAPs can control bacterial adhesion (S. aureus, P. aeruginosa, and E. coli) and affect the cell cycle of human bone marrow stem cells (hBMSCs). Moreover, in a mouse subcutaneous model, cSAPs with selective [2-(methacryloyloxy)ethyl]dimethyl-(3-sulfopropyl)ammonium (SBMA) modification can reduce the inflammatory response after being challenged with bacteria. This study reveals that functionalized cSAPs are versatile tools for controlling cellular responses at biointerfaces, which is instructive for biomaterials or biodevices.

Indirect co-culture of lung carcinoma cells with hyperthermia-treated mesenchymal stem cells influences tumor spheroid growth in a collagen-based 3-dimensional microfluidic model.

BACKGROUND: Mesenchymal stem cells (MSCs) have paradoxically been reported to exert either pro- or anti-tumor effects in vitro. Hyperthermia, in combination with chemotherapy, has tumor-inhibiting effects; however, its role, together with MSCs, so far is not well understood. Furthermore, a lot of research is conducted using conventional 2-dimensional in vitro models that do not mimic the actual tumor microenvironment. AIM: In light of this fact, an indirect method of co-culturing human amniotic membrane-derived MSCs (AMMSCs) with collagen-encapsulated human lung carcinoma cells (A549) was performed using a 3-dimensional (3D) tumor-on-chip device. METHODS: The conditioned medium of AMMSCs (AMMSC-CM) or heat-treated AMMSCs (heat-AMMSC-CM) was utilized to create indirect co-culture conditions. Tumor spheroid growth characterization, immunocytochemistry and cytotoxicity assays, and anti-cancer peptide (P1) screening were performed to determine the effects of the conditioned medium. RESULTS: The A549 cells cultured inside the 3D microfluidic chip developed into multicellular tumor spheroids over five days of culture. The AMMSC-CM, contrary to previous reports claiming its tumor-inhibiting potential, led to significant proliferation of tumor spheroids. Heat-AMMSC-CM led to reductions in both spheroid diameter and cell proliferation. The medium containing the P1 peptide was found to be the least cytotoxic to tumor spheroids in co-culture compared with the monoculture and heat-co-culture groups. CONCLUSIONS: Hyperthermia, in combination with the anticancer peptide, exhibited highest cytotoxic effects. This study highlights the growing importance of 3D microfluidic tumor models for testing stem-cell-based and other anti-cancer therapies.

Laminin coated diamond electrodes for neural stimulation.

The performance of many implantable neural stimulation devices is degraded due to the loss of neurons around the electrodes by the body's natural biological responses to a foreign material. Coating of electrodes with biomolecules such as extracellular matrix proteins is one potential route to suppress the adverse responses that lead to loss of implant functionality. Concurrently, however, the electrochemical performance of the stimulating electrode must remain optimal to continue to safely provide sufficient charge for neural stimulation. We have previously found that oxygen plasma treated nitrogen included ultrananocrystalline diamond coated platinum electrodes exhibit superior charge injection capacity and electrochemical stability for neural stimulation (Sikder et al., 2019). To fabricate bioactive diamond electrodes, in this work, laminin, an extracellular matrix protein known to be involved in inter-neuron adhesion and recognition, was used as an example biomolecule. Here, laminin was covalently coupled to diamond electrodes. Electrochemical analysis found that the covalently coupled films were robust and resulted in minimal change to the charge injection capacity of diamond electrodes. The successful binding of laminin and its biological activity was further confirmed using primary rat cortical neuron cultures, and the coated electrodes showed enhanced cell attachment densities and neurite outgrowth. The method proposed in this work is versatile and adaptable to many other biomolecules for producing bioactive diamond electrodes, which are expected to show reduced the inflammatory responses in vivo.

Binary Colloidal Crystal (BCC) Substrates for Controlling the Fate of Mouse Embryonic Stem Cells.

Understanding the interactions of stem cells with surface topography can give us an invaluable tool in controlling stemness and fate of stem cells for further use in biomedical applications. In this study, we have fabricated topographical features using a class of cell culture substrates called binary colloidal crystals (BCCs), that are made by self-assembly of mixtures of spherical micron sized silica (Si) and nanometer sized polystyrene (PS) or poly (methyl methacrylate) (PMMA) particles. The substrates formed are arrays of ordered, hexagonally packed large Si particles inter-dispersed with the PS particles that are stabilized by gentle heating, which melts the PS or PMMA forming substrates suitable for cell culture. BCC substrates were used for culture of mouse embryonic stem cells (mESCs). Compared to tissue culture plates, COM1 (Si5-PMMA0.4), COM2 (Si5-PS0.4) and COM4 (Si2-PSC0.22) have shown to provide a better support for mESC proliferation in the presence of the cytokine leukemia inhibitory factor (LIF). The behavior of mESCs with the BCCs in presence and absence of LIF, was further explored and it was found that interaction of mESCs with the culture substrate can be controlled by tuning surface topography and roughness, which is determined by the size and type of particles used in making BCCs. Furthermore, it was shown that limiting cell-surface interactions and controlling colony shape can promote stemness maintenance on COM1 and COM2 substrates as indicated by better proliferation and higher expression of pluripotency genes including Nanog both in presence and in absence of LIF. Together with higher expression of GATA6 gene, it can be stated that these surfaces can be used for endodermic priming of mESCs. Therefore, we believe that these surfaces, especially COM1 and COM2 surfaces can be beneficial as stem cell culture systems for further use in biomedical research.