In Vitro Assessment of Cardiac Fibroblast Activation at Physiologic Stiffness.

Cardiac fibroblasts (CF) are an essential cell type in cardiac physiology, playing diverse roles in maintaining structural integrity, extracellular matrix (ECM) synthesis, and tissue repair. Under normal conditions, these cells reside in the interstitium in a quiescent state poised to sense and respond to injury by synthesizing and secreting collagen, vimentin, hyaluronan, and other ECM components. In response to mechanical and chemical stimuli, these "resident" fibroblasts can undergo a transformation through a continuum of activation states into what is commonly known as a "myofibroblast," in a process critical for injury response. Despite progress in understanding the contribution of fibroblasts to cardiac health and disease, much remains unknown about the signaling mediating this activation, in part owing to technical challenges in evaluating CF function and activation status in vitro. Given their role in monitoring the ECM, CFs are acutely sensitive to stiffness and pressure. High basal activation of isolated CFs is common due to the super-physiologic stiffness of traditional cell culture substrates, making assays dependent on quiescent cells challenging. To overcome this problem, cell culture parameters must be tightly controlled, and the use of dishes coated with biocompatible reduced-stiffness substrates, such as 8-kPa polydimethylsiloxane (PDMS), has shown promise in reducing basal activation of fibroblasts. Here, we describe cell culture protocol for maintaining CF quiescence in vitro to enable a dynamic range for the assessment of activation status in response to fibrogenic stimuli using PDMS-coated coverslips. Our protocol provides a cost-effective tool to study fibroblast signaling and activity, allowing researchers to better understand the underlying mechanisms involved in cardiac fibrosis. © 2024 The Authors. Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: Generation of 8-kPa polydimethylsiloxane (PDMS)/gelatin-coated coverslips for cardiac fibroblast cell culture Basic Protocol 2: Isolation of adult cardiac fibroblasts and plating onto PDMS coverslips Basic Protocol 3: Assessment of cardiac fibroblast activation by α smooth muscle actin (αSMA) immunocytochemistry.

The effect of educational intervention on efficacy of 1% permethrin shampoo and 4% dimeticone lotion to treat head lice infestation using propensity score matching (PSM).

BACKGROUND: Head lice are a main public health problem and the most important human ectoparasites and the use of pediculicides is the most common way to control it. One of the possible causes of treatment failure is the lack of improper application of pediculicide. The aim of this study was to assess the effect of education on efficacy of 1% permethrin or 4% dimeticone lotion to treat head lice infestation. METHODS: This quasi-experimental study included 100 individuals with head lice infestation from comprehensive urban health centers in Ardabil as the intervention group, and 400 individuals from East Azerbaijan and West Azerbaijan provinces as the control group, from April to March 2019. The data collection tools included a demographic questionnaire and an examination recording sheet, which documented the presence of adult lice or nits. Due to the inability to perform random assignment and control for numerous observed covariates, propensity score matching (PSM) was used. RESULTS: The outcome of treatment included elimination of head lice infestation on is 7, and in the case of recurrence, it was considered on days 14 and 30 after treatment. The results showed that the educational intervention program had a significant positive effect on the efficacy of both treatments. The likelihood of improvement was approximately three times greater in the intervention group compared to the control group. CONCLUSION: Participants who received the training intervention (OR = 3.29; CI 95%: 2.21-4.88) were more likely to have a successful treatment than control group. In the case of providing proper training on the use of pediculicides and observing hygiene tips to patients with pediculosis, could help to successful treatment of pediculosis.

Guttapercha Improves In Vitro Bioactivity and Dentin Remineralization Ability of a Bioglass Containing Polydimethylsiloxane-Based Root Canal Sealer.

Guttapercha (GP, trans-1,4-polyisoprene) is the most used tooth root filling material, and it must be used with an appropriate cement (typically a polydimethylsiloxane (PDMS)-based sealer) to ensure an adequate canal obturation. This study aimed to assess the bioactivity and dentin remineralization ability of a bioglass containing PDMS commercial endodontic sealer, BG-PDMS (GuttaFlow Bioseal), and to evaluate the possible influence of a GP cone (Roeko GP point) on the mineralization process. To this end, BG-PDMS disks were aged alone or in the presence of a GP cone in Hank's Balanced Salt Solution (28 d, 37 °C). Dentin remineralization experiments were carried out under the same conditions. Micro-Raman and IR analyses demonstrated that BG-PDMS is bioactive, thanks to the formation of a silica-rich layer with nucleation sites for B-type carbonated apatite deposition. This phase was thicker when BG-PDMS was aged in the presence of GP. The two materials influenced each other because GP, which alone did not show any bioactivity, nucleated a calcium phosphate phase under these conditions. Analogously, dentin remineralization experiments showed that BG-PDMS is able to remineralize dentin, especially in the presence of GP. Under the experimental conditions, GP acted as a templating agent for calcium phosphate deposition.

Stable hydrogel adhesion to polydimethylsiloxane enables cyclic mechanical stimulation of 3D-bioprinted smooth muscle constructs.

During normal urination, smooth muscle cells (SMCs) in the lower urinary tract (LUT) are exposed to mechanical signals that have a critical impact on tissue structure and function. Nevertheless, the mechanisms underlying the maintenance of the contractile phenotype of SMCs remain poorly understood. This is due, in part, to a lack of studies that have examined the effects of mechanical loading using three-dimensional (3D) models. In this study, surface modifications of polydimethylsiloxane (PDMS) membrane were evaluated to investigate the effects of cyclic mechanical stimulation on SMC maturation in 3D constructs. Commercially available cell stretching plates were modified with amino or methacrylate groups to promote adhesion of 3D constructs fabricated by bioprinting. After 6 days of stimulation, the effects of mechanical stimulation on the expression of contractile markers at the mRNA and protein levels were analyzed. Methacrylate-modified surfaces supported stable adhesion of the 3D constructs to the membrane and facilitated cyclic mechanical stimulation, which significantly increased the expression of contractile markers at the mRNA and protein levels. These effects were found to be mediated by activation of the p38 MAPK pathway, as inhibition of this pathway abolished the effects of stimulation in a dose-dependent manner. These results provide valuable insights into the role of mechanical signaling in maintaining the contractile phenotype of bladder SMCs, which has important implications for the development of future treatments for LUT diseases.

Multifunctional Surface Modification of PDMS for Antibacterial Contact Killing and Drug-Delivery of Polar, Nonpolar, and Amphiphilic Drugs.

Medical device-associated infections pose major clinical challenges that emphasize the need for improved anti-infective biomaterials. Polydimethylsiloxane (PDMS), a frequently used elastomeric biomaterial in medical devices, is inherently prone to bacterial attachment and associated infection formation. Here, PDMS surface modification strategy is presented consisting of a cross-linked lyotropic liquid crystal hydrogel microparticle coating with antibacterial functionality. The microparticle coating composed of cross-linked triblock copolymers (diacrylated Pluronic F127) was deposited on PDMS by physical immobilization via interpenetrating polymer network formation. The formed coating served as a substrate for covalent immobilization of a potent antimicrobial peptide (AMP), RRPRPRPRPWWWW-NH2, yielding high contact-killing antibacterial effect against Staphylococcus epidermidis and Staphylococcus aureus. Additionally, the coating was assessed for its ability to selectively host polar, amphiphilic, and nonpolar drugs, resulting in sustained release profiles. The results of this study put forward a versatile PDMS modification strategy for both contact-killing antibacterial surface properties and drug-delivery capabilities, offering a solution for medical device-associated infection prevention.

Bone surface mimicked PDMS membranes stimulate osteoblasts and calcification of bone matrix.

Cellular microenvironments play a crucial role in cell behavior. In addition to the biochemical cues present in the microenvironments, biophysical and biomechanical properties on surfaces have an impact on cellular functionality and eventually cellular fate. Effects of surface topography on cell behavior are being studied extensively in the literature. However, these studies often try to replicate topographical features of tissue surfaces by using techniques such as chemical etching, photolithography, and electrospinning, which may result in the loss of crucial micro- and nano- features on the tissue surfaces such as bone. This study investigates the topographical effects of bone surface by transferring its surface features onto polydimethylsiloxane (PDMS) membranes using soft lithography from a bovine femur. Our results have shown that major features on bone surfaces were successfully transferred onto PDMS using soft lithography. Osteoblast proliferation and calcification of bone matrix have significantly increased along with osteoblast-specific differentiation and maturation markers such as osteocalcin (OSC), osterix (OSX), collagen type I alpha 1 chain (COL1A1), and alkaline phosphatase (ALP) on bone surface mimicked (BSM) PDMS membranes in addition to a unidirectional alignment of osteoblast cells compared to plain PDMS surfaces. This presented bone surface mimicking method can provide a versatile native-like platform for further investigation of intracellular pathways regarding osteoblast growth and differentiation.

Contact-Active Layer-by-Layer Grafted TPU/PDMS Blends as an Antiencrustation and Antibacterial Platform for Next-Generation Urological Biomaterials: Validation in Artificial and Human Urine.

Urinary tract infections and urinary encrustation impede the long-term clinical performance of urological implants and medical devices. Together, biofilm formation and encrustation constitute serious complications, driving the development of next-generation urological biomaterials. The currently available bioengineered solutions have limited success during long-term usage in the urinary environment. In addressing this unmet clinical challenge, contact-active, antiencrustation surface grafting were conceived onto a dynamically cross-linked polydimethylsiloxane (PDMS) modified thermoplastic polyurethane (TPU) blend using the layer-by-layer (LbL) assembly route. To the best of the authors' knowledge, the present study is the first to investigate the LbL grafting in developing an antiencrustation platform. These multilayered assemblies strategically employed covalent cross-linking and electrostatic interaction-assisted progressive depositions of branched polyethyleneimine and poly(2-ethyl-2-oxazoline). While polyethyleneimine conferred the contact-killing bactericidal activity, the much-coveted antiencrustation properties were rendered by incorporating a partially hydrolyzed derivative of poly(2-ethyl-2-oxazoline). The performance of the resultant surface-modified TPU/PDMS blends was benchmarked against the conventional urological alloplasts, in a customized lab-scale bioreactor-based dynamic encrustation study and in human urine. After 6 weeks of exposure to an artificial urine medium, simulating urease-positive bacterial infection, the surface-modified blends exhibited a remarkable ability to suppress Ca and Mg encrustation. In addition, these blends also displayed superior grafting stability and antibacterial efficacy against common uropathogens. As high as 4-fold log reduction in the planktonic growth of Gram-negative P. mirabilis and Gram-positive MRSA was recorded with the LbL platform vis-à-vis medical-grade TPU. In conjunction, the in vitro cellular assessment with human keratinocytes (HaCaT) and human embryonic kidney cells (HEK) established the uncompromised cytocompatibility of the multilayered grafted blends. Finally, the physiologically relevant functionality of the LbL grafting has been validated using clinical samples of human urine collected from 129 patients with a broad spectrum of disease conditions. The phase-I pre-clinical study, entailing 6 week-long incubation in human urine, demonstrated significantly improved encrustation resistance of the blends. The collective findings of the present work clearly establish the success of LbL strategies in the development of stable, multifunctional new-generation urological biomaterials.

The effect of AKT in extracellular matrix stiffness induced osteogenic differentiation of hBMSCs.

Extracellular matrix (ECM) stiffness is an important biophysical factor in human bone marrow mesenchymal stem cells (hBMSCs) differentiation. Although there is evidence that Yes-associated protein (YAP) plays an important role in ECM elasticity induced osteogenesis, but the regulatory mechanism and signaling pathways have not been distinctly uncovered. In this study, hBMSCs were cultured on collagen-coated polydimethylsiloxane hydrogels with stiffness corresponding to Young's moduli of 0.5 kPa and 32 kPa, and gene chip analyses revealed the phosphoinositide 3-kinase (PI3K)-AKT pathway was highly correlated with ECM stiffness. Following western blots indicated that AKT phosphorylation was evidently affected in 5th-7th days after ECM stiffness stimulation, while PI3K showed little difference. The AKT activator SC79 and inhibitor MK2206 were utilized to modulate AKT phosphorylation. SC79 and MK2206 caused alteration in the mRNA expression and protein level of alkaline phosphatase (ALP), collagen type I alpha 1 (COL1A1) and runt related transcription factor 2 (RUNX2). On 32 kPa substrates, YAP enrichment in nucleus were significantly promoted by SC79 and remarkably decreased by MK2206. Besides, the ratio of YAP/p-YAP is upregulated by SC79 on both 32 kPa and 0.5 kPa substrates. In conclusion, these findings suggest that AKT is involved in the modulation of ECM stiffness induced osteogenesis, and AKT phosphorylation also influences the subcellular localization and activation of YAP.

Bonding antimicrobial rhamnolipids onto medical grade PDMS: A strategy to overcome multispecies vascular catheter-related infections.

In clinic there is a demand to solve the drawback of medical devices multispecies related infections. Consequently, different biomaterial surfaces, such as vascular catheters, urgently need improvement regarding their antifouling/antimicrobial properties. In this work, we covalently functionalized medical grade polydimethylsiloxane (PDMS) with antimicrobial rhamnolipids to investigate the biomaterial surface activity towards mono and dual species biofilms. Preparation of surfaces with "piranha" oxidation, followed by APTES bonding and carbodiimide reaction with rhamnolipids effectively bonded these compounds to PDMS surface as confirmed by FTIR-ATR and XPS analysis. Generated surfaces were active towards S. aureus biofilm formation showing a 4.2 log reduction while with S. epidermidis and C. albicans biofilms a reduction of 1.2 and 1.0 log reduction, respectively, was observed. Regarding dual-species testing the higher biofilm log reduction observed was 1.9. Additionally, biocompatibility was assessed by cytocompatibility towards human fibroblastic cells, low platelet activation and absence of vascular irritation. Our work not only sheds light on using covalently bonded rhamnolipids towards dual species biofilms but also highlights the biocompatibility of the obtained PDMS surfaces.

Biomimetic approach to articular cartilage tissue engineering using carbon nanotube-coated and textured polydimethylsiloxane scaffolds.

There is a significant need to understand the complexity and heterogeneity of articular cartilage to develop more effective therapeutic strategies for diseases such as osteoarthritis. Here, we show that carbon nanotubes (CNTs) are excellent candidates as a material for synthetic scaffolds to support the growth of chondrocytes-the cells that produce and maintain cartilage. Chondrocyte morphology, proliferation, and alignment were investigated as nanoscale CNT networks were applied to macroscopically textured polydimethylsiloxane (PDMS) scaffolds. The application of CNTs to the surface of PDMS-based scaffolds resulted in an up to 10-fold increase in cell adherence and 240% increase in proliferation, which is attributable to increased nanoscale roughness and hydrophilicity. The introduction of macroscale features to PDMS induced alignment of chondrocytes, successfully mimicking the cell behavior observed in the superficial layer of cartilage. Raman spectroscopy was used as a noninvasive, label-free method to monitor extracellular matrix production and chondrocyte phenotype. Chondrocytes on these scaffolds successfully produced collagen, glycosaminoglycan, and aggrecan. This study demonstrates that introducing physical features at different length scales allows for a high level of control over tissue scaffold design and, thus, cell behavior. Ultimately, these textured scaffolds can serve as platforms to improve the understanding of osteoarthritis and for early-stage therapeutic testing.

New Carbon-Mineral Sorbent Based on Aluminum Oxide, Polydimethylsiloxane, and Single-Wall Carbon Nanotubes: Assessment of the Effect on Erythrocytes In Vitro.

A comparative study of the effect of a sorbent with nanotubes (Al2O3@ WCNT-PDMS) and a carbon-mineral sorbent (Al2O3@C) on the parameters of human erythrocytes was carried out. Using scanning flow cytometry, the morphological and functional parameters of venous blood erythrocytes as well as drainage blood after its perfusion through columns with sorbents were determined. The compared samples Al2O3@SWCNT-PDMS and Al2O3@C are similar by their effect on the morphological and functional parameters of erythrocytes. The maximum membrane extensibility increased to a greatest extent after contact with Al2O3@C, the amount of hemoglobin in erythrocytes decreased to the greatest extent after perfusion through a column with Al2O3@SWCNT-PDMS sorbent. The scanning flow cytometry is promising for assessing the effect on erythrocytes of new sorption materials intended for blood detoxification. Changes in the parameters of erythrocytes of blood collected in a sterile drainage system for subsequent reinfusion were revealed.

Slippery Liquid-Like Solid Surfaces with Promising Antibiofilm Performance under Both Static and Flow Conditions.

Biofilms are central to some of the most urgent global challenges across diverse fields of application, from medicine to industries to the environment, and exert considerable economic and social impact. A fundamental assumption in anti-biofilms has been that the coating on a substrate surface is solid. The invention of slippery liquid-infused porous surfaces─a continuously wet lubricating coating retained on a solid surface by capillary forces─has led to this being challenged. However, in situations where flow occurs, shear stress may deplete the lubricant and affect the anti-biofilm performance. Here, we report on the use of slippery omniphobic covalently attached liquid (SOCAL) surfaces, which provide a surface coating with short (ca. 4 nm) non-cross-linked polydimethylsiloxane (PDMS) chains retaining liquid-surface properties, as an antibiofilm strategy stable under shear stress from flow. This surface reduced biofilm formation of the key biofilm-forming pathogens Staphylococcus epidermidis and Pseudomonas aeruginosa by three-four orders of magnitude compared to the widely used medical implant material PDMS after 7 days under static and dynamic culture conditions. Throughout the entire dynamic culture period of P. aeruginosa, SOCAL significantly outperformed a typical antibiofilm slippery surface [i.e., swollen PDMS in silicone oil (S-PDMS)]. We have revealed that significant oil loss occurred after 2-7 day flow for S-PDMS, which correlated to increased contact angle hysteresis (CAH), indicating a degradation of the slippery surface properties, and biofilm formation, while SOCAL has stable CAH and sustainable antibiofilm performance after 7 day flow. The significance of this correlation is to provide a useful easy-to-measure physical parameter as an indicator for long-term antibiofilm performance. This biofilm-resistant liquid-like solid surface offers a new antibiofilm strategy for applications in medical devices and other areas where biofilm development is problematic.

Substrate stiffness regulates the differentiation profile and functions of osteoclasts via cytoskeletal arrangement.

OBJECTIVES: Aging and common diseases alter the stiffness of bone tissue, causing changes to the microenvironment of the mechanosensitive bone cells. Osteoclasts, the sole bone-resorbing cells, play a vital role in bone remodeling. This study was performed to elucidate the mechanism through which osteoclasts sense and react to substrate stiffness signals. MATERIALS AND METHODS: We fabricated polydimethylsiloxane (PDMS) substrates of different stiffness degrees for osteoclast formation progressed from osteoclast precursors including bone marrow-derived macrophages (BMMs) and RAW264.7 monocytes. Osteoclast differentiation in response to the stiffness signals was determined by examining the cell morphology, fusion/fission activities, transcriptional profile, and resorption function. Cytoskeletal changes and mechanosensitive adhesion molecules were also assessed. RESULTS: Stiffer PDMS substrates accelerated osteoclast differentiation, firstly observed by variations in their morphology and fusion/fission activities. Upregulation of canonical osteoclast markers (Nfatc1, Acp5, Ctsk, Camk2a, Mmp9, Rela, and Traf6) and the fusion master regulator DC-stamp were detected on stiffer substrates, with similar increases in their bone resorption functions. Additionally, the activation of cytoskeleton-associated adhesion molecules, including fibronectin and integrin αvß3, followed by biochemical signaling cascades of paxillin, FAK, PKC, and RhoA, was detected on the stiffer substrates. CONCLUSIONS: This is the first study to provide evidence proving that extracellular substrate stiffness is a strong determinant of osteoclast differentiation and functions. Higher stiffness upregulated the differentiation profile and activity of osteoclasts, revealing the mechanical regulation of osteoclast activity in bone homeostasis and diseases.

On/off switchable physical stimuli regulate the future direction of adherent cellular fate.

The utilization of cell-manipulating techniques reveals information about biological behaviors suited to address a wide range of questions in the field of life sciences. Here, we introduced an on/off switchable physical stimuli technique that offers precise stimuli for reversible cell patterning to allow regulation of the future direction of adherent cellular behavior by leveraging enzymatically degradable alginate hydrogels with defined chemistry and topography. As a proof of concept, targeted muscle cells adherent to TCP exhibited a reshaped structure when the hydrogel-based physical stimuli were applied. This simple tool offers easy manipulation of adherent cells to reshape their morphology and to influence future direction depending on the characteristics of the hydrogel without limitations of time and space. The findings from this study are broadly applicable to investigations into the relationships between cells and physiological extracellular matrix environments as well as has potential to open new horizons for regenerative medicine with manipulated cells.

Influence of Culture Substrates on Morphology and Function of Pulmonary Alveolar Cells In Vitro.

Cell's microenvironment has been shown to exert influence on cell behavior. In particular, matrix-cell interactions strongly impact cell morphology and function. The purpose of this study was to analyze the influence of different culture substrate materials on phenotype and functional properties of lung epithelial adenocarcinoma (A549) cells. A549 cells were seeded onto two different biocompatible, commercially available substrates: a polyester coverslip (Thermanox™ Coverslips), that was used as cell culture plate control, and a polydimethylsiloxane membrane (PDMS, Elastosil® Film) investigated in this study as alternative material for A549 cells culture. The two substrates influenced cell morphology and the actin cytoskeleton organization. Further, the Yes-associated protein (YAP) and its transcriptional coactivator PDZ-binding motif (TAZ) were translocated to the nucleus in A549 cells cultured on polyester substrate, yet it remained mostly cytosolic in cells on PDMS substrate. By SEM analysis, we observed that cells grown on Elastosil® Film maintained an alveolar Type II cell morphology. Immunofluorescence staining for surfactant-C revealing a high expression of surfactant-C in cells cultured on Elastosil® Film, but not in cells cultured on Thermanox™ Coverslips. A549 cells grown onto Elastosil® Film exhibited morphology and functionality that suggest retainment of alveolar epithelial Type II phenotype, while A549 cells grown onto conventional plastic substrates acquired an alveolar Type I phenotype.

Dynamically crosslinked polydimethylsiloxane-based polyurethanes with contact-killing antimicrobial properties as implantable alloplasts for urological reconstruction.

A large population of patients is reported to suffer from urinary bladder-associated irreversible physiological disorders, rationalizing a continuous surge for structural and functional substitutes of urinary tissues, including ureters, bladder-wall, and urethra. The current gold standard for bladder reconstruction, an autologous gastrointestinal graft, is proven not to be an ideal substitute in the clinic. While addressing this unmet clinical need, a unique platform of antimicrobial polydimethyl siloxane-modified polyurethanes (TPU/PDMS) is designed and developed for its potential application as a urological implant. To the best of our knowledge, this study reports for the first time the successful integration of varying contents of PDMS within the molten polyurethane matrix using in situ crosslinking methodology. Thus, compatibilized binary blends possess clinically relevant viscoelastic properties to sustain high pressure, large distensions, and surgical manipulation. Furthermore, different chemical strategies are explored to covalently incorporate quaternized moieties, including 4-vinyl pyridine (4-VP), branched-polyethyleneimine (bPEI) as well as bPEI-grafted-(acrylic acid-co-vinylbenzyltriphenyl phosphonium chloride) (PAP), and counter urinary tract infections. The modified compositions, endowed with contact killing surfaces, reveal nearly three log reduction in bacterial growth in pathogenically infected artificial urine. Importantly, the antimicrobial TPU/PDMS blends support the uninhibited growth of mitochondrially viable murine fibroblasts, in a manner comparable to the medical-grade polyurethane. Collectively, the obtained results affirmed the newly developed polymers as promising biomaterials in reconstructive urology. STATEMENT OF SIGNIFICANCE: The clinical procedure for end-stage bladder disease remains replacement or augmentation of the bladder wall with a section of the patient's gastrointestinal tract. However, the absorptive and mucus-producing epithelium of intestinal segment is liable to short- and long-term complications. The dynamically crosslinked polydimethyl siloxane-based polyurethanes proposed herein, and the associated synthesis strategies to induce polycation grafted non-exhaustive contact-killing surfaces against uropathogents, have a significant clinical prospect in reconstructive urology. As an 'off-the-shelf' available alloplastic substitute, these blends offer the potential to circumvent the challenges associated with non-urinary autografts or scaffold based regenerative engineering and, thereby, shorten as well as simplify the surgical treatment. The targeted application has been conceived for a bladder patch to assist in various urinary diseases including, bladder carcinoma, refractory overactive bladder, interstitial cystitis, etc. However, given the ease of fabrication, moldability and the wide spectrum of mechanical properties that could be encompassed, these blends also present the possibility to be manifested into artificial ureteral or urethral conduits.

Straightforward neuron micropatterning and neuronal network construction on cell-repellent polydimethylsiloxane using microfluidics-guided functionalized Pluronic modification.

Neuronal cell microengineering involving micropatterning and polydimethylsiloxane (PDMS) microfluidics enables promising advances in microscale neuron control. However, a facile methodology for the precise and effective manipulation of neurons on a cell-repellent PDMS substrate remains challenging. Herein, a simple and straightforward strategy for neuronal cell patterning and neuronal network construction on PDMS based on microfluidics-assisted modification of functionalized Pluronic is described. The cell patterning process simply involves a one-step microfluidic modification and routine in vitro culture. It is demonstrated that multiple types of neuronal cell arrangements with various spatial profiles can be conveniently produced using this patterning tool. The precise control of neuronal cells with high patterning fidelity up to single cell resolution, as well as high adhesion and differentiation, is achieved too. Furthermore, neuronal network construction using the respective cell population and single cell patterning prove to be applicable. This achievement provides a convenient and feasible methodology for engineering neuronal cells on PDMS substrates, which will be useful for applications in many neuron-related microscale analytical research fields, including cell engineering, neurobiology, neuropharmacology, and neuronal sensing.

High density culture of pancreatic islet-like 3D tissue organized in oxygen-permeable porous scaffolds with external oxygen supply.

Transplantation of macroencapsulated pancreatic islets within semipermeable membranes is a promising approach for the treatment of type 1 diabetes. Encapsulation beneficially isolates the implants from the host immune system. Deleteriously however, it also limits oxygen supply to the cells. This creates challenges in loading islets at the amount and density required to meet the practical demands of clinical usage. To overcome this challenge, we investigated the feasibility of using macroporous scaffolds made of an oxygen-permeable polymer, poly(dimethylsiloxane) (PDMS) by culturing pancreatic islet-like three-dimensional tissue made of a rat pancreatic beta cell line on the scaffolds. With external oxygenation, the density and function of cells on the PDMS scaffold were more than three times and almost two times higher than those without oxygenation, respectively. This suggests that the oxygenation afforded by the PDMS scaffolds allows for high-density loading of islet tissue into the devices.

GuttaFlow® Bioseal Cytotoxicity Assessment: In Vitro Study.

The sealers used for root canal treatment should be biocompatible for the peri-radicular tissues, to evaluate the cytotoxic effects of GuttaFlow® bioseal sealer and to compare them with AH26® epoxy resin. Culture media were conditioned with the GuttaFlow® bioseal and AH26® pellets. MDPC-23 odontoblast cell cultures were treated with conditioned medium and serial dilutions. To evaluate the metabolic activity and cellular viability, the MTT and SRB assays were performed. To determine the production of reactive oxygen species, the DHE and DCF-DA probes were used. Cell cycle and cell-death types were assessed by cytometry, and to evaluate the mineralization capacity, the Alizarin Red S coloration was used. Statistical analysis was performed using analysis of variance (ANOVA) when normality was found and Kruskal-Wallis on the opposite case. For the comparison with normality values, the Student t-test was used. Cells exposed to the GuttaFlow® bioseal conditioned medium maintained high metabolic activities, except at higher concentrations. Likewise, viability was maintained, but a significant decrease was observed after exposure to the highest concentration (p < 0.001), associated with cell death by late apoptosis and necrosis. When cell cultures were exposed to AH26®, metabolic activity was highly compromised, resulting in cell death. An imbalance in the production of peroxides and superoxide anion was observed. GuttaFlow® bioseal showed higher biocompatibility than AH26®.

Enhanced Biocompatibility and Differentiation Capacity of Mesenchymal Stem Cells on Poly(dimethylsiloxane) by Topographically Patterned Dopamine.

Controlling the behavior of mesenchymal stem cells (MSCs) through topographic patterns is an effective approach for stem cell studies. We, herein, reported a facile method to create a dopamine (DA) pattern on poly(dimethylsiloxane) (PDMS). The topography of micropatterned DA was produced on PDMS after plasma treatment. The grid-topographic-patterned surface of PDMS-DA (PDMS-DA-P) was measured for adhesion force and Young's modulus by atomic force microscopy. The surface of PDMS-DA-P demonstrated less stiff and more elastic characteristics compared to either nonpatterned PDMS-DA or PDMS. The PDMS-DA-P evidently enhanced the differentiation of MSCs into various tissue cells, including nerve, vessel, bone, and fat. We further designed comprehensive experiments to investigate adhesion, proliferation, and differentiation of MSCs in response to PDMS-DA-P and showed that the DA-patterned surface had good biocompatibility and did not activate macrophages or platelets in vitro and had low foreign body reaction in vivo. Besides, it protected MSCs from apoptosis as well as excessive reactive oxygen species (ROS) generation. Particularly, the patterned surface enhanced the differentiation capacity of MSCs toward neural and endothelial cells. The stromal cell-derived factor-1α/CXantiCR4 pathway may be involved in mediating the self-recruitment and promoting the differentiation of MSCs. These findings support the potential application of PDMS-DA-P in either cell treatment or tissue repair.