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Articular cartilage is a tissue of fundamental importance for the mechanics of joints, since it provides a smooth and lubricated surface for the proper transfer of loads. From a mechanical point of view, this tissue is an anisotropic poroviscoelastic material: its characteristics at the macroscopic level depend on the complex microscopic architecture. With the ability to probe the local microscopic features, dynamic nanoindentation test is a powerful tool to investigate cartilage mechanics. In this work we focus on a length scale where the time dependent behaviour is regulated by poroelasticity more than viscoelasticity and we aim to understand the effect of the anisotropic permeability on the mechanics of the superficial layer of the articular cartilage. In a previous work, a finite element model for the dynamic nanoindentation test has been presented. In this work, we improve the model by considering the presence of an anisotropic permeability tensor that depends on the collagen fibers distribution. Our sensitivity analysis highlights that the permeability decreases with increasing indentation, thus making the tissue stiffer than the case of isotropic permeability, when solicited at the same frequency. With this improved model, a revised identification of the mechanical and physical parameters for articular cartilage is provided. To this purpose the model was used to simulate experimental data from tests performed on bovine tissue, giving a better estimation of the anisotropy in the elastic properties. A relation between the identified macroscopic anisotropic permeability properties and the microscopic rearrangement of the fiber/matrix structure during indentation is also provided. [ABSTRACT FROM AUTHOR]

We investigated the combined effect of the initial cell density (12 500, 35 000, 75 000, and 100 000 cells cm−2) and concentration of the anti-cancer drug doxorubicin on HeLa cells by performing time-dependent cytotoxicity assays using real-time electrochemical impedance spectroscopy. A correlation between the rate of cell death and the initial cell seeding density was found at 2.5 μM doxorubicin concentration, whereas this was not observed at 5 or 100 μM. By sensing the changes in the cell–substrate interaction using impedance spectroscopy under static conditions, the onset of cytotoxicity was observed 5 h earlier than when using a standard colorimetric end-point assay (MTS) which measures changes in the mitochondrial metabolism. Furthermore, with the MTS assay no cytotoxicity was observed after 15 h of incubation with 2.5 μM doxorubicin, whereas the impedance showed at this time point cell viability that was below 25%. These results indicate that impedance detection reveals cytotoxic events undetectable when using the MTS assay, highlighting the importance of combining impedance detection with traditional drug toxicity assays towards a more in depth understanding of the effect of anti-cancer drugs on in vitro assays. Moreover, the detection of doxorubicin induced toxicity determined with impedance under static conditions proved to be 6 times faster than in perfusion culture. [ABSTRACT FROM AUTHOR]

This study aimed to analyse the sarcolemma of Col6a1-/- fibers in comparison with wild type and mdx fibers, taken as positive control in view of the known structural and functional alterations of their membranes. Structural and mechanical properties were studied in single muscle fibers prepared from FDB muscle using atomic force microscopy (AFM) and conventional electrophysiological techniques to measure ionic conductance and capacitance. While the sarcolemma topography was preserved in both types of dystrophic fibers, membrane elasticity was significantly reduced in Col6a1-/- and increased in mdx fibers. In the membrane of Col6a1-/- fibers ionic conductance was increased likely due to an increased leakage, whereas capacitance was reduced, and the action potential (ap) depolarization rate was reduced. The picture emerging from experiments on fibers in culture was consistent with that obtained on intact freshly dissected muscle. Mdx fibers in culture showed a reduction of both membrane conductance and capacitance. In contrast, in mdx intact FDB muscle resting conductance was increased while resting potential and ap depolarization rate were reduced, likely indicating the presence of a consistent population of severely altered fibers which disappear during the culture preparation. [ABSTRACT FROM AUTHOR]

We use a multiwell micromechanical cantilever sensor (MCS) device to measure surface stress changes induced by specific adsorption of molecules. A multiplexed assay format facilitates the monitoring of the bending of 16 MCSs in parallel. The 16 MCSs are grouped within four separate wells. Each well can be addressed independently by different analyte liquids. This enables functionalization of MCS separately by flowing different solutions through each well. In addition, each well contains a fixed reference mirror which allows measuring the absolute bending of MCS. In addition, the mirror can be used to follow refractive index changes upon mixing of different solutions. The effect of the flow rate on the MCS bending change was found to be dependent on the absolute bending value of MCS. Experiments and finite element simulations of solution exchange in wells were performed. Both revealed that one solution can be exchanged by another one after 200 μl volume has flown through. Using this device, the adsorption of thiolated DNA molecules and 6-mercapto-1-hexanol on gold surfaces was performed to test the nanomechanical response of MCS. [ABSTRACT FROM AUTHOR]

A cohort of 69 children born to HIV-1 positive women was studied to evaluate types, prevalences and relationships to clinical stages of HIV-1-related oral lesions. In addition, relationships among C. albicans biotypes, clinical features of oral candidiasis and HIV-1 disease were investigated. C. albicans biotypes did not correlate with clinical features of oral lesions, disease stages and CD4+ lymphocyte count. Of 8 patients with recurrent oral candidiasis, 4 changed clinical features and 5 changed biotype. Our study pointed out the high frequency (28.9%) of oral lesions, especially caused by fungi and the importance of the examination of the oral cavity in children born to HIV-1 positive women. [ABSTRACT FROM AUTHOR]

Between February 1987 and February 1990, we studied 737 antibody anti HIV-1 positive (AbHIV+) subjects referred to the Infectious Diseases Institute of the University of Turin (Italy) in order to evaluate types, prevalences, relations with clinical stages, distributions in risk-groups and prognostic significances of HIV-1 related oral lesions. The study evidenced the high prevalence of oral lesions, especially mycoses, in the investigated population: 40.3% of the patients showed, in fact, HIV-1 related oral lesions. The 37 months follow-up of 55 AbHIV+ with oral hairy leukoplakia (HL) and 101 patients with oral candidiasis (OC), demonstrated that the probability of developing AIDS in patient with HL was 0.381 at 15, 0.635 at 25 and 0.824 at 37 months. In the patients with OC the probability was 0.294 at 15 months, 0.524 at 25 and 0.781 at 37 months. [ABSTRACT FROM AUTHOR]

Abstract: Greening the building envelope focusing on green façades with vegetation is a good example of a new construction practice. Plants and partly growing materials in case of living wall systems (LWS) have a number of functions that are beneficial, for example: increasing the biodiversity and ecological value, mitigation of urban heat island effect, outdoor and indoor comfort, insulating properties, improvement of air quality and of the social and psychological well being of city dwellers. This paper discusses a comparative life cycle analysis (LCA) situated in The Netherlands for: a conventional built up European brick façade, a façade greened directly, a façade greened indirectly (supported by a steel mesh), a façade covered with a living wall system based on planter boxes and a façade covered with a living wall system based on felt layers. Beside the environmental benefits of the above described greening systems, it is eventually not clear if these systems are sustainable, due to the materials used, maintenance, nutrients and water needed. A LCA is used to analyze the similarity and differences in the environmental impacts in relation with benefits estimated for two climate types for building energy saving (reduction of electrical energy used for building cooling and heating). [Copyright &y& Elsevier]

Thermoplastic polyurethane (TPU) is a polymer used in a variety of fields, including medical applications. Here, we aimed to verify if the brush and bar coater deposition techniques did not alter TPU properties. The topography of the TPU-modified surfaces was studied via AFM demonstrating no significant differences between brush and bar coater-modified surfaces, compared to the un-modified TPU (TPU Film). The effect of the surfaces on planktonic bacteria, evaluated by MTT assay, demonstrated their anti-adhesive effect on E. coli , while the bar coater significantly reduced staphylococcal planktonic adhesion and both bacterial biofilms compared to other samples. Interestingly, Pearson's R coefficient analysis showed that R a roughness and Haralick's correlation feature were trend predictors for planktonic bacterial cells adhesion. The surface adhesion property was evaluated against NIH-3T3 murine fibroblasts by MTT and against human fibrinogen and human platelet-rich plasma by ELISA and LDH assay, respectively. An indirect cytotoxicity experiment against NIH-3T3 confirmed the biocompatibility of the TPUs. Overall, the results indicated that the deposition techniques did not alter the antibacterial and anti-adhesive surface properties of modified TPU compared to un-modified TPU, nor its bio- and hemocompatibility, confirming the suitability of TPU brush and bar coater films in the biomedical and pharmaceutical fields.

Pre-operative delirium may cause delay in surgical intervention in older patients hospitalized for hip fracture. Also it has been associated with higher risk of post-surgical complications and worst functional outcomes. Aim of this retrospective cohort study was to evaluate whether the multidimensional prognostic index (MPI) at hospital admission was associated with pre-operative delirium in older individuals with hip fracture who are deemed to require surgical intervention. Consecutive older patients (≥ 65 years) with hip fracture underwent a comprehensive geriatric assessment to calculate the MPI at hospital admission. According to previously established cut-offs, MPI was expressed in three grades, i.e. MPI-1 (low-risk), MPI-2 (moderate-risk) and MPI-3 (high risk of mortality). Pre-operative delirium was assessed using the four 'A's Test. Out of 244 older patients who underwent surgery for hip fracture, 104 subjects (43%) received a diagnosis of delirium. Overall, the incidence of delirium before surgery was significantly higher in patients with more severe MPI score at admission. Higher MPI grade (MPI-3) was independently associated with higher risk of pre-operative delirium (OR 2.45, CI 1.21-4.96). Therefore, the MPI at hospital admission might help in early identification of older patients with hip fracture at risk for pre-operative delirium., (© 2022. The Author(s).)

The brain is the most complex organ of our body. Such a complexity spans from the single-cell morphology up to the intricate connections that hundreds of thousands of neurons establish to create dense neuronal networks. All these components are involved in the genesis of the rich patterns of electrophysiological activity that characterize the brain. Over the years, researchers coming from different disciplines developed in vitro simplified experimental models to investigate in a more controllable and observable way how neuronal ensembles generate peculiar firing rhythms, code external stimulations, or respond to chemical drugs. Nowadays, such in vitro models are named brain-on-a-chip pointing out the relevance of the technological counterpart as artificial tool to interact with the brain: multi-electrode arrays are well-used devices to record and stimulate large-scale developing neuronal networks originated from dissociated cultures, brain slices, up to brain organoids. In this review, we will discuss the state of the art of the brain-on-a-chip, highlighting which structural and biological features a realistic in vitro brain should embed (and how to achieve them). In particular, we identified two topological features, namely modular and three-dimensional connectivity, and a biological one (heterogeneity) that takes into account the huge number of neuronal types existing in the brain. At the end of this travel, we will show how 'far' we are from the goal and how interconnected-brain-regions-on-a-chip is the most appropriate wording to indicate the current state of the art., (© 2021 IOP Publishing Ltd.)

We have developed a novel experimental set-up that simultaneously, (i) applies static and dynamic deformations to adherent cells in culture, (ii) allows the visualization of cells under fluorescence microscopy, and (iii) allows atomic force microscopy nanoindentation measurements of the mechanical properties of the cells. The cell stretcher device relies on a dielectric elastomer film that can be electro-actuated and acts as the cell culture substrate. The shape and position of the electrodes actuating the film can be controlled by design in order to obtain specific deformations across the cell culture chamber. By using optical markers we characterized the strain fields under different electrode configurations and applied potentials. The combined setup, which includes the cell stretcher device, an atomic force microscope, and an inverted optical microscope, can assess in situ and with sub-micron spatial resolution single cell topography and elasticity, as well as ion fluxes, during the application of static deformations. Proof of performance on fibroblasts shows a reproducible increase in the average cell elastic modulus as a response to applied uniaxial stretch of just 4%. Additionally, high resolution topography and elasticity maps on a single fibroblast can be acquired while the cell is deformed, providing evidence of long-term instrumental stability. This study provides a proof-of-concept of a novel platform that allows in situ and real time investigation of single cell mechano-transduction phenomena with sub-cellular spatial resolution.

Objective: The goal of this work is to develop and characterize an innovative experimental framework to design interconnected (i.e. modular) heterogeneous (cortical-hippocampal) neuronal cultures with a three-dimensional (3D) connectivity and to record their electrophysiological activity using micro-electrode arrays (MEAs)., Approach: A two-compartment polymeric mask for the segregation of different neuronal populations (cortex and hippocampus) was coupled to the MEA surface. Glass microbeads were used as a scaffold to mimic the 3D brain micro-architecture., Main Results: We built a fully functional heterogeneous 3D neuronal network. From an electrophysiological point of view, we found that the heterogeneity induces a global increase of the activity rate, while the 3D connectivity modulates the duration and the organization of the bursting activity., Significance: In vivo, studies of network dynamics and interactions between neuronal populations are often time-consuming, low-throughput, complex, and suffer from reproducibility. On the other hand, most of the commonly used in vitro brain models are too simplified and thus far from the in vivo situation. The achieved results demonstrate the feasibility to build a more realistic and controllable experimental in vitro model of interconnected brain regions on-a-chip whose applications may have impacts on the study of neurological disorders that impair the connectivity between brain areas (e.g. Parkinson disease).

Evaluation and understanding the effect of drug delivery in in vitro systems is fundamental in drug discovery. We present an assay based on real-time electrical impedance spectroscopy (EIS) measurements that can be used to follow the internalisation and cytotoxic effect of a matrix metalloproteinase (MMP)-sensitive liposome formulation loaded with oxaliplatin (OxPt) on colorectal cancer cells. The EIS response identified two different cellular processes: (i) a negative peak in the cell index (CI) within the first 5 h, due to onset of liposome endocytosis, followed by (ii) a subsequent CI increase, due to the reattachment of cells until the onset of cytotoxicity with a decrease in CI. Free OxPt or OxPt-loaded Stealth liposomes did not show this two-stage EIS response; the latter can be due to the fact that Stealth cannot be cleaved by MMPs and thus is not taken up by the cells. Real-time bright-field imaging supported the EIS data, showing variations in cell adherence and cell morphology after exposure to the different liposome formulations. A drastic decrease in cell coverage as well as rounding up of cells during the first 5 h of exposure to OxPt-loaded (MMP)-sensitive liposome formulation is reflected by the first negative EIS response, which indicates the onset of liposome endocytosis. Graphical abstract.

The high sensitivity of silicon microcantilever sensors has expanded their use in areas ranging from gas sensing to bio-medical applications. Photochromic molecules also represent promising candidates for a large variety of sensing applications. In this work, the operating principles of these two sensing methods are combined in order to detect the reversible conformational change of a molecular switch, spiropyran. Thus, arrays of silicon microcantilever sensors were functionalized with spiropyran on the gold covered side and used as test microcantilevers. The microcantilever deflection response was observed, in five sequential cycles, as the transition from the spiropyran (SP) (CLOSED) to the merocyanine (MC) (OPEN) state and vice-versa when induced by UV and white light LED sources, respectively, proving the reversibility capabilities of this type of sensor. The microcantilever deflection direction was observed to be in one direction when changing to the MC state and in the opposite direction when changing back to the SP state. A tensile stress was induced in the microcantilever when the SP to MC transition took place, while a compressive stress was observed for the reverse transition. These different type of stresses are believed to be related to the spatial conformational changes induced in the photochromic molecule upon photo-isomerisation.

Cardiomyocytes (CM) placed on microelectrode array (MEA) were simultaneously probed with cantilever from atomic force microscope (AFM) system. This electric / nanomechanical combination in real time recorded beating force of the CMs cluster and the triggering electric events. Such "organ-on-a-chip" represents a tool for drug development and disease modeling. The human pluripotent stem cells included the WT embryonic line CCTL14 and the induced dystrophin deficient line reprogrammed from fibroblasts of a patient affected by Duchenne Muscular Dystrophy (DMD, complete loss of dystrophin expression). Both were differentiated to CMs and employed with the AFM/MEA platform for diseased CMs' drug response testing and DMD characterization. The dependence of cardiac parameters on extracellular Ca 2+ was studied. The differential evaluation explained the observed effects despite variability of biological samples. The β-adrenergic stimulation (isoproterenol) and antagonist trials (verapamil) addressed ionotropic and chronotropic cell line-dependent features. For the first time, a distinctive beating-force relation for DMD CMs was measured on the 3D cardiac in vitro model., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The cardiac excitation-contraction coupling is the cellular process through which the heart absolves its blood pumping function, and it is directly affected when cardiac pathologies occur. Cardiomyocytes are the functional units in which this complex biomolecular process takes place: they can be represented as a two-stage electro-chemo and chemo-mechanical transducer, along which each stage can be probed and monitored via appropriate micro/nanotechnology-based tools. Atomic force microscopy (AFM), with its unique nanoresolved force sensitivity and versatile modes of extracting sample properties, can represent a key instrument to study time-dependent heart mechanics and topography at the single cell level. In this work, we show how the integrative possibilities of AFM allowed us to implement an in vitro system which can monitor cardiac electrophysiology, intracellular calcium dynamics, and single cell mechanics. We believe this single cell-sensitive and integrated system will unlock improved, fast, and reliable cardiac in vitro tests in the future.

Graphene-derived materials, such as graphene oxide (GO), have been widely explored for biomedical and biological applications, including cancer research. Despite some recent works proving that GO inhibits the migration and invasion of different cancer cells, so far most of these in vitro studies have been conducted using GO sheets dispersed in solution or as a planar film. On the contrary, little is known about cellular activities, such as cell viability, adhesion, and spreading, when cancer cells interface with GO functionalized hydrogel-based surfaces, biomechanically and structurally more similar to the tumor environment. Here, we evaluate the interactions of human breast cancer cells (MDA-MB-231) with alginate (Alg)/GO hydrogel-based substrates, and compare them with a cancer cell line from human osteosarcoma (HOS) and healthy murine fibroblasts (3T3). We observed that GO addition selectively inhibits malignant breast cancer cell adhesion efficiency and spreading area, while promotes HOS and 3T3 adhesive processes. Furthermore, we did not observe the same results over Alg substrates with GO nanosheets dispersed in the medium, without embedment into the Alg. This suggests that cancer (MDA-MB-231 and HOS) and healthy (3T3) cell adhesion efficacy does not depend on the cellular tumoral nature and it is driven by the topographical cues provided by the GO-based substrates, whose physical-mechanical characteristics better mimic those of the cell native tissue. We envision that this study can provide a rational for future design and use of graphene-based nanomaterials for cancer research by deepening the knowledge of graphene-cancer cell specific interactions.

Objective: We aim to develop a novel non-invasive or minimally invasive method for neural stimulation to be applied in the study and treatment of brain (dys)functions and neurological disorders., Approach: We investigate the electrophysiological response of in vitro neuronal networks when subjected to low-intensity pulsed acoustic stimulation, mediated by piezoelectric nanoparticles adsorbed on the neuronal membrane., Main Results: We show that the presence of piezoelectric barium titanate nanoparticles induces, in a reproducible way, an increase in network activity when excited by stationary ultrasound waves in the MHz regime. Such a response can be fully recovered when switching the ultrasound pulse off, depending on the generated pressure field amplitude, whilst it is insensitive to the duration of the ultrasound pulse in the range 0.5 s-1.5 s. We demonstrate that the presence of piezoelectric nanoparticles is necessary, and when applying the same acoustic stimulation to neuronal cultures without nanoparticles or with non-piezoelectric nanoparticles with the same size distribution, no network response is observed., Significance: We believe that our results open up an extremely interesting approach when coupled with suitable functionalization strategies of the nanoparticles in order to address specific neurons and/or brain areas and applied in vivo, thus enabling remote, non-invasive, and highly selective modulation of the activity of neuronal subpopulations of the central nervous system of mammalians.

The availability of 3D biomimetic in vitro neuronal networks of mammalian neurons represents a pivotal step for the development of brain-on-a-chip experimental models to study neuronal (dys)functions and particularly neuronal connectivity. The use of hydrogel-based scaffolds for 3D cell cultures has been extensively studied in the last years. However, limited work on biomimetic 3D neuronal cultures has been carried out to date. In this respect, here we investigated the use of a widely popular polysaccharide, chitosan (CHI), for the fabrication of a microbead based 3D scaffold to be coupled to primary neuronal cells. CHI microbeads were characterized by optical and atomic force microscopies. The cell/scaffold interaction was deeply characterized by transmission electron microscopy and by immunocytochemistry using confocal microscopy. Finally, a preliminary electrophysiological characterization by micro-electrode arrays was carried out., (Copyright © 2017 Elsevier Ltd. All rights reserved.)