LASU: An efficient and stable phthalocyanine dye with tolerable safety profile for self-disinfecting anti-COVID textiles activated by ambient light.

BACKGROUND: Recent COVID crisis has demonstrated that modern society urgently needs an accessible protection against mass infections, especially viruses, as the new strains are appearing at an ever-increasing pace and cause severe harm to the population and the world economy. METHODS: We have developed an efficient phthalocyanine photosensitizer LASU, that is suitable for dyeing textiles and allows to prepare reusable self-disinfecting fabrics with strong antiviral properties. The safety profile of LASU was evaluated in accredited laboratories by several in vitro assays according to the OECD-guidelines. RESULTS: The textiles impregnated with LASU phthalocyanine showed a significant antiviral photodynamic effect even under moderate indoor and outdoor light. The dye did not show any genotoxic potential in human lymphocyte micronucleus assay. It showed a possible indication for eye irritation in human EpiOcular™ model and was phototoxic when tested in mouse BALB/c 3T3 cell test in the presence and absence of UVA-irradiation. CONCLUSION: Novel phthalocyanine-dyed textiles are suitable for general use as self-disinfecting antiviral barriers and materials in hospitals, households, and public places. The safety profile of LASU is the phototoxic effect which is related to LASU´s mode of action.

A Human Cell-based Assay to Assess the Induction of Vasculature Formation for Non-genotoxic Carcinogenicity Testing Purposes: A Pilot Study.

The induction of vasculature formation is proposed to be a significant mechanism behind the non-genotoxic carcinogenicity of a chemical. The vasculature formation model used in this study is based on the coculture of human primary HUVECs and hASCs. This model was used to develop an assay to assess the induction of vasculature formation. Three assay protocols, based on different conditions, were developed and compared in order to identify the optimal conditions required. Some serum supplements and growth factors were observed to be essential for initiating vasculature formation. Of the studied putative positive reference chemicals, aspartame, sodium nitrite, bisphenol A and nicotine treatment led to a clear induction of vasculature formation, but arsenic and cadmium treatment only led to a slight increase. This human cell-based assay has the potential to be used as one test within a next generation testing battery, to assess the non-genotoxic carcinogenicity of a chemical through the mechanism of vasculature formation induction.

In Search of Clinical Markers: Indicators of Exposure in Dampness and Mold Hypersensitivity Syndrome (DMHS).

Potential markers were sought to diagnose mold hypersensitivity. Indoor air condensed water and human macrophage THP-1 test were applied to evaluate the buildings. Basophil activation tests (BAT) were conducted and mold-specific immunoglobulins (IgE, IgG, IgA, and IgD) were measured in study subjects' serum and feces. Exposed subjects reported markedly more symptoms from occupational air than controls. Basophils from exposed subjects died/lost activity at 225 times lower concentrations of toxic extracts from the target building than recommended in the common BAT protocol. Fecal IgG and IgD levels against Acrostalagmus luteoalbus and Aspergillus versicolor produced receiver operating curves (ROC) of 0.928 and 0.916, respectively, when plotted against the inflammation marker MRP8/14. Assaying serum immunoglobulin concentrations against the toxic Chaetomium globosum (MTAV35) from another building, a test control, did not differentiate study individuals. However, if liver metabolism produced the same core molecule from other Chaetomium globosum strains, this would explain the increased response in fecal immunoglobulins in the exposed. The altered immunoglobulin values in the samples of exposed when compared to controls revealed the route of mold exposure. The toxicity of indoor air condensed water samples, BAT and serology confirmed the severity of symptoms in the target building's employees, supporting earlier findings of toxicity in this building.

New approach methods for assessing indoor air toxicity.

Indoor air is typically a mixture of many chemicals at low concentrations without any adverse health effects alone, but in mixtures they may cause toxicity and risks to human health. The aim of this study was by using new approach methods to assess the potential toxicity of indoor air condensates. In specific, different in vitro test methods including cyto-and immunotoxicity, skin sensitization and endocrine disruption were applied. In addition to biological effects, the indoor air samples were subjected to targeted analysis of 25 volatile organic compounds (VOCs) and Genapol X-80 (a nonionic emulsifier) suspected to be present in the samples, and to a non-targeted "total chemical scan" to find out whether the chemical composition of the samples is associated with the biological effects. The results confirm that assessing health risks of indoor air by analysing individual chemicals is not an adequate approach: We were not able to detect the VOCs and Genapol X-80 in the indoor air samples, yet, several types of toxicity, namely, cytotoxicity, immunotoxicity, skin sensitization and endocrine disruption were detected. In the non-targeted total chemical scan of the indoor air samples, a larger number of compounds were found in the cytotoxic samples than in the non-cytotoxic samples supporting the biological findings. If only one biological method would be selected for the screening of indoor air quality, THP-1 macrophage/WST-1 assay would best fit for the purpose as it is sensitive and serves as a good representative for different sub-toxic end points, including immunotoxicity, (skin) sensitization and endocrine disruption.

Functional human cell-based vascularised cardiac tissue model for biomedical research and testing.

Cardiomyocytes derived from human induced pluripotent stem cells (hiPSC) are widely used in in vitro biomedical research and testing. However, fully matured, adult cardiomyocyte characteristics have not been achieved. To improve the maturity and physiological relevance of hiPSC-derived cardiomyocytes, we co-cultured them with preconstructed vascular-like networks to form a functional, human cell-based cardiac tissue model. The morphology and gene expression profiles indicated advanced maturation in the cardiac tissue model compared to those of a cardiomyocyte monoculture. The cardiac tissue model's functionality was confirmed by measuring the effects of 32 compounds with multielectrode array and comparing results to human data. Our model predicted the cardiac effects with a predictive accuracy of 91%, sensitivity of 90% and specificity of 100%. The correlation between the effective concentration (EC50) and the reported clinical plasma concentrations was 0.952 (R2 = 0.905). The developed advanced human cell-based cardiac tissue model showed characteristics and functionality of human cardiac tissue enabling accurate transferability of gained in vitro data to human settings. The model is standardized and thus, it would be highly useful in biomedical research and cardiotoxicity testing.

Direct Contraction Force Measurements of Engineered Cardiac Tissue Constructs With Inotropic Drug Exposure.

Contractility is one of the most crucial functions of the heart because it is directly related to the maintenance of blood perfusion throughout the body. Both increase and decrease in contractility may cause fatal consequences. Therefore, drug discovery would benefit greatly from reliable testing of candidate molecule effects on contractility capacity. In this study, we further developed a dual-axis piezoelectric force sensor together with our human cell-based vascularized cardiac tissue constructs for cardiac contraction force measurements. The capability to detect drug-induced inotropic effects was tested with a set of known positive and negative inotropic compounds of isoprenaline, milrinone, omecamtiv mecarbil, propranolol, or verapamil in different concentrations. Both positive and negative inotropic effects were measurable, showing that our cardiac contraction force measurement system including a piezoelectric cantilever sensor and a human cell-based cardiac tissue constructs has the potential to be used for testing of inotropic drug effects.

Toxic Indoor Air Is a Potential Risk of Causing Immuno Suppression and Morbidity-A Pilot Study.

We aimed to establish an etiology-based connection between the symptoms experienced by the occupants of a workplace and the presence in the building of toxic dampness microbiota. The occupants (5/6) underwent a medical examination and urine samples (2/6) were analyzed by LC-MS/MS for mycotoxins at two time-points. The magnitude of inhaled water was estimated. Building-derived bacteria and fungi were identified and assessed for toxicity. Separate cytotoxicity tests using human THP-1 macrophages were performed from the office's indoor air water condensates. Office-derived indoor water samples (n = 4/4) were toxic to human THP-1 macrophages. Penicillium, Acremonium sensu lato, Aspergillus ochraceus group and Aspergillus section Aspergillus grew from the building material samples. These colonies were toxic in boar sperm tests (n = 11/32); four were toxic to BHK-21 cells. Mycophenolic acid, which is a potential immunosuppressant, was detected in the initial and follow-up urine samples of (2/2) office workers who did not take immunosuppressive drugs. Their urinary mycotoxin profiles differed from household and unrelated controls. Our study suggests that the presence of mycotoxins in indoor air is linked to the morbidity of the occupants. The cytotoxicity test of the indoor air condensate is a promising tool for risk assessment in moisture-damaged buildings.

Association between [68Ga]NODAGA-RGDyK uptake and dynamics of angiogenesis in a human cell-based 3D model.

Radiolabeled RGD peptides targeting expression of αvß3 integrin have been applied to in vivo imaging of angiogenesis. However, there is a need for more information on the quantitative relationships between RGD peptide uptake and the dynamics of angiogenesis. In this study, we sought to measure the binding of [68Ga]NODAGA-RGDyK to αvß3 integrin in a human cell-based three-dimensional (3D) in vitro model of angiogenesis, and to compare the level of binding with the amount of angiogenesis. Experiments were conducted using a human cell-based 3D model of angiogenesis consisting of co-culture of human adipose stem cells (hASCs) and of human umbilical vein endothelial cells (HUVECs). Angiogenesis was induced with four concentrations (25%, 50%, 75%, and 100%) of growth factor cocktail resulting in a gradual increase in the density of the tubule network. Cultures were incubated with [68Ga]NODAGA-RGDyK for 90 min at 37 °C, and binding of radioactivity was measured by gamma counting and digital autoradiography. The results revealed that tracer binding increased gradually with neovasculature density. In comparison with vessels induced with a growth factor concentration of 25%, the uptake of [68Ga]NODAGA-RGDyK was higher at concentrations of 75% and 100%, and correlated with the amount of neovasculature, as determined by visual evaluation of histological staining. Uptake of [68Ga]NODAGA-RGDyK closely reflected the amount of angiogenesis in an in vitro 3D model of angiogenesis. These results support further evaluation of RGD-based approaches for targeted imaging of angiogenesis.

Qualitative and Quantitative Analysis of Certain Aspects of the Cytotoxic and Genotoxic Hazard of Hospital Wastewaters by Using a Range of In Vitro Assays.

Health care facilities and hospitals generate significant amounts of wastewater which are released into the sewage system, either after a preliminary treatment or without any further treatment. Hospital wastewater may contain large amounts of hazardous chemicals and pharmaceuticals, some of which cannot be eliminated entirely by wastewater treatment plants. Moreover, hospital effluents may be loaded with a plethora of pathogenic microorganisms or other microbiota and microbiome residues. The need to monitor hospital effluents for their genotoxic hazard is of high importance, as detailed information is scarce. DNA-based information can be acquired directly from samples through the application of various molecular methods, while cell-based biomonitoring assays can provide important information about impaired cellular pathways or mechanisms of toxicity without prior knowledge of the identity of each toxicant. In our study, we evaluated samples of chlorinated hospital wastewater discharged into the sewage system after this disinfection process. The assessment of cytotoxicity, genotoxicity and mutagenicity of the hospital effluents was performed in vitro by using a broad battery of biomonitoring assays that are relevant for human health effects. All the tested hospital wastewater samples could be classified as potentially genotoxic, and it is concluded that the microbiota present in hospital wastewater might contribute to this genotoxic potential.

Angiogenic Effect of Pravastatin Alone and with Sera from Healthy and Complicated Pregnancies Studied by in vitro Vasculogenesis/Angiogenesis Assay.

OBJECTIVE: To determine the direct effect of pravastatin on angiogenesis and to study the interaction between pravastatin and maternal sera from women with early- or late-onset pre-eclampsia (PE), intrauterine growth restriction, or healthy pregnancy. METHODS: We collected 5 maternal serum samples from each group. The effect of pravastatin on angiogenesis was assessed with and without maternal sera by quantifying tubule formation in a human-based in vitro assay. Pravastatin was added at 20, 1,000, and 8,000 ng/mL concentrations. Concentrations of angiogenic and inflammatory biomarkers in serum and in test medium after supplementation of serum alone and with pravastatin (1,000 ng/mL) were measured. RESULTS: Therapeutic concentration of pravastatin (20 ng/mL) did not have significant direct effect on angiogenesis, but the highest concentrations inhibited angiogenesis. Pravastatin did not change the levels of biomarkers in the test media. There were no changes in angiogenesis when therapeutic dose of pravastatin was added with maternal sera, but there was a trend to wide individual variation towards enhanced angiogenesis, particularly in the early-onset PE group. CONCLUSIONS: At therapeutic concentration, pravastatin alone or with maternal sera has no significant effect on angiogenesis, but at high concentrations the effect seems to be anti-angiogenic estimated by in vitro assay.

Sensitization potential of medical devices detected by in vitro and in vivo methods.

Medical devices must be tested before marketing in accordance with ISO EN 10993-10 in order to avoid skin sensi­tization. This standard predominantly refers to the in vivo test but does not exclude the use of in vitro methods that have been sufficiently technically and scientifically validated for medical device testing. It is foreseen that, due to the complexity of the sensitization endpoint, a combination of several methods will be needed to address all key events occurring in the sensitization process. The objective of this pilot study was to evaluate the sensitization potential of selected medical devices using a combination of in chemico (DPRA, OECD TG 442C) and in vitro (LuSens, OECD TG 442D) methods in comparison with the in vivo (LLNA DA, OECD TG 442A) method and to suggest a possible testing strategy for the safety assessment of medical device extracts. Overall, one of the 42 tested samples exhibited positive results in all employed test methods, while 33 samples were predicted as non-sensitizing in all three performed methods. This study demonstrated good agreement between in vitro and in vivo results regarding non-sensitizing samples; however, some discrepancies in positive classification were recorded. A testing strategy is suggested in which negative results are accepted and any positive results in the in chemico or in vitro tests are followed up with a third in vitro test and evaluated in accordance with the "2 out of 3 approach". This strategy may reduce and replace animal use for testing the sensitization potential of medical devices.

Families' Experiences Living with Acquired Brain Injury: "Thinking Family"-A Nursing Pathway for Family-Centered Care.

The objective of this study was to examine families' experiences living with acquired brain injury (ABI) using a research approach that included both the affected individual family member and the family together as a family group. A narrative inquiry study, informed by the life-stage approach of Lieblich, Tuval-Mashiach, and Zilber, was used to obtain family stories. Families experiencing an ABI event were purposefully selected from different regions in a western Canadian province. Centered on the life stages of before the ABI event, now living with the ABI, and the future, thematic findings included: Families, a grounding force; Losses, individual and family; Family adaptive capacities; Experiences with the healthcare system-hospital to home; and A patchwork future-entering the unknown. Themes affirmed the significant impacts of ABI on individual and family members and acknowledged ABI as an ambiguous loss event. The findings also illuminated families' strengths and resiliencies in coping with living with ABI. The study results suggest by "thinking family" nurses can contribute towards a healthcare model that focuses on "family" as the central unit of care.

Therapeutic doses of metformin do not have impact on angiogenesis in presence of sera from pre-eclamptic, IUGR and healthy pregnancies.

Recent evidence suggests that metformin may prevent pre-eclampsia by reverting the angiogenic imbalance in maternal sera. In this study, we investigated effect of metformin on angiogenesis by quantifying tubule formation in a human-based in vitro test with co-culture of human adipose stromal cell (hASC) and human umbilical vein endothelial cell (HUVEC). A total of 20 pregnant women were recruited in the study. Serum samples were obtained from women with early- and late-onset pre-eclampsia and from women with pregnancies complicated by intrauterine growth restriction (IUGR) without pre-eclampsia (N = 5 in each of the three groups). Serum samples from women with healthy pregnancies served as controls (N = 5). The direct effect of metformin on angiogenesis was first assessed without maternal sera. Secondly, we investigated the impact of metformin on angiogenesis in the present of maternal sera. Metformin was used at 5, 50 and 600 µg/ml concentrations. Angiogenic and inflammatory biomarkers in maternal sera were analyzed by immunoassays. When the direct effect of metformin was studied, the two lowest concentrations of metformin did not affect tubule formation (angiogenesis), but the highest concentration inhibited angiogenesis. When metformin was supplemented at therapeutic concentrations of 5 and 50 µg/ml along with serum samples, there was no change in tubule formation in comparison to maternal sera alone. However, strong inhibitory effect on tubule formation was observed in all groups with the highest, non-therapeutic (600 µg/ml), concentration of metformin.

Development of novel human in vitro vascularized adipose tissue model with functional macrophages.

Inflammation has been proven significant factor in development of type 2 diabetes. So far, most of the adipose tissue related research has been performed in animals, mainly rodent models. The relevance of translation of animal results to humans is questionable. However, in vitro model with relevant human cell source, such as human adipose tissue stromal cells (hASC), can be developed and should be utilized for human adipose tissue research. We developed in vitro models of human adipose tissue utilizing hASC, endothelial cells and monocytes/macrophages. By isolating endothelial cells and macrophages from same adipose tissue as hASC, we were able to provide method for constructing personalized models of adipose tissue. With these models, we studied the effect of macrophages on adipogenesis and protein secretion, with and without vasculature. The models were analyzed for immunocytochemical markers, cell number, triglyceride accumulation and protein secretion. We found that lipid accumulation was greater in adipocytes in the presence of macrophages. Interferon gamma increased this difference between adipocyte culture and Adipocyte-Macrophage co-culture. Protein secretion was affected more by macrophages when vasculature was not present compared to the mild effect when vasculature was present. The vascularized adipose model with macrophages is valuable tool for human adipose tissue research, especially for the personalized medicine approaches; for choosing the right treatments and for studying rare medical conditions.

A cross-platform approach to characterize and screen potential neurovascular unit toxicants.

Development of the neurovascular unit (NVU) is a complex, multistage process that requires orchestrated cell signaling mechanisms across several cell types and ultimately results in formation of the blood-brain barrier. Typical high-throughput screening (HTS) assays investigate single biochemical or single cell responses following chemical insult. As the NVU comprises multiple cell types interacting at various stages of development, a methodology combining high-throughput results across pertinent cell-based assays is needed to investigate potential chemical-induced disruption to the development of this complex cell system. To this end, we implemented a novel method for screening putative NVU disruptors across diverse assay platforms to predict chemical perturbation of the developing NVU. HTS assay results measuring chemical-induced perturbations to cellular key events across angiogenic and neurogenic outcomes in vitro were combined to create a cell-based prioritization of NVU hazard. Chemicals were grouped according to similar modes of action to train a logistic regression literature model on a training set of 38 chemicals. This model utilizes the chemical-specific pairwise mutual information score for PubMed MeSH annotations to represent a quantitative measure of previously published results. Taken together, this study presents a methodology to investigate NVU developmental hazard using cell-based HTS assays and literature evidence to prioritize screening of putative NVU disruptors towards a knowledge-driven characterization of neurovascular developmental toxicity. The results from these screening efforts demonstrate that chemicals representing a range of putative vascular disrupting compound (pVDC) scores can also produce effects on neurogenic outcomes and characterizes possible modes of action for disrupting the developing NVU.

Erratum: Multilabel segmentation of cancer cell culture on vascular structures with deep neural networks (Erratum).

[This corrects the article DOI: 10.1117/1.JMI.7.2.024001.].

Toxicological testing of a photoactive phthalocyanine-based antimicrobial substance.

The aim of the study was toxicological testing of an innovative and efficient antimicrobial agent based on photoactive phthalocyanine (Pc) derivative. A promising Aluminium phthalocyanine (AlPc) with efficient and stable antimicrobial effects was subjected to a battery of toxicological tests to avoid local and systemic toxicity hazard. In compliance with the current European legislation restricting the use of experimental animals, the methods comprised exclusively in vitro procedures based on cellular and tissue models of human origin or mimicking human tissues. The battery of toxicological tests to identify local toxicity included skin corrosion/irritation, eye irritation, and phototoxicity. The basic systemic toxicity tests included acute toxicity, skin sensitization, genotoxicity, and endocrine disruption. The results showed that AlPc induced skin and eye irritation, exhibited borderline sensitization potential and mutagenic potential in one test strain of the Ames test, which was not confirmed in the chromosome aberration test. The AlPc was found to be phototoxic. The results from the cytotoxicity test designed for acute oral toxicity estimation were not conclusive, the acute toxicity potential has to be determined by conventional tests in vivo. Regarding endocrine disruption, no agonistic activity of the AlPc on human estrogen receptor α, nor human androgen receptor was observed. The skin penetration/absorption test revealed that the AlPc has not penetrated into the dermis and receptor fluid, confirming no risk of systemic exposure via the bloodstream.

Multilabel segmentation of cancer cell culture on vascular structures with deep neural networks.

New increasingly complex in vitro cancer cell models are being developed. These new models seem to represent the cell behavior in vivo more accurately and have better physiological relevance than prior models. An efficient testing method for selecting the most optimal drug treatment does not exist to date. One proposed solution to the problem involves isolation of cancer cells from the patients' cancer tissue, after which they are exposed to potential drugs alone or in combinations to find the most optimal medication. To achieve this goal, methods that can efficiently quantify and analyze changes in tested cell are needed. Our study aimed to detect and segment cells and structures from cancer cell cultures grown on vascular structures in phase-contrast microscope images using U-Net neural networks to enable future drug efficacy assessments. We cultivated prostate carcinoma cell lines PC3 and LNCaP on the top of a matrix containing vascular structures. The cells were imaged with a Cell-IQ phase-contrast microscope. Automatic analysis of microscope images could assess the efficacy of tested drugs. The dataset included 36 RGB images and ground-truth segmentations with mutually not exclusive classes. The used method could distinguish vascular structures, cells, spheroids, and cell matter around spheroids in the test images. Some invasive spikes were also detected, but the method could not distinguish the invasive cells in the test images.