ILC1 drive intestinal epithelial and matrix remodelling.

Organoids can shed light on the dynamic interplay between complex tissues and rare cell types within a controlled microenvironment. Here, we develop gut organoid cocultures with type-1 innate lymphoid cells (ILC1) to dissect the impact of their accumulation in inflamed intestines. We demonstrate that murine and human ILC1 secrete transforming growth factor ß1, driving expansion of CD44v6+ epithelial crypts. ILC1 additionally express MMP9 and drive gene signatures indicative of extracellular matrix remodelling. We therefore encapsulated human epithelial-mesenchymal intestinal organoids in MMP-sensitive, synthetic hydrogels designed to form efficient networks at low polymer concentrations. Harnessing this defined system, we demonstrate that ILC1 drive matrix softening and stiffening, which we suggest occurs through balanced matrix degradation and deposition. Our platform enabled us to elucidate previously undescribed interactions between ILC1 and their microenvironment, which suggest that they may exacerbate fibrosis and tumour growth when enriched in inflamed patient tissues.

Mental transportation mediates nostalgia's psychological benefits.

Nostalgizing confers social, existential, and self-oriented psychological benefits or functions. But how does the experience of nostalgia conduce to these functions? We propose that it does so, in part, through mental transportation, which involves mentally leaving one's current space and transporting oneself into a past event. We addressed the role of mental transportation in one daily diary study and two experiments (N = 514). By assessing daily experiences of nostalgia in Study 1, we found that, on days in which participants felt more nostalgic, they were more likely to experience mental transportation. Following a narrative induction of nostalgia, we assessed mental transportation (Studies 2 and 3) and the three putative nostalgia functions: social, existential, self-oriented (Study 3). Nostalgic (vs. control) participants reported greater mental transportation (Studies 2 and 3), which in turn was associated with stronger functions (Study 3). The findings portray mental transportation as a key mechanism underlying the psychological benefits of nostalgia.

Collective Cell Behavior in Mechanosensing of Substrate Thickness.

Extracellular matrix stiffness has a profound effect on the behavior of many cell types. Adherent cells apply contractile forces to the material on which they adhere and sense the resistance of the material to deformation-its stiffness. This is dependent on both the elastic modulus and the thickness of the material, with the corollary that single cells are able to sense underlying stiff materials through soft hydrogel materials at low (<10 µm) thicknesses. Here, we hypothesized that cohesive colonies of cells exert more force and create more hydrogel deformation than single cells, therefore enabling them to mechanosense more deeply into underlying materials than single cells. To test this, we modulated the thickness of soft (1 kPa) elastic extracellular-matrix-functionalized polyacrylamide hydrogels adhered to glass substrates and allowed colonies of MG63 cells to form on their surfaces. Cell morphology and deformations of fluorescent fiducial-marker-labeled hydrogels were quantified by time-lapse fluorescence microscopy imaging. Single-cell spreading increased with respect to decreasing hydrogel thickness, with data fitting to an exponential model with half-maximal response at a thickness of 3.2 µm. By quantifying cell area within colonies of defined area, we similarly found that colony-cell spreading increased with decreasing hydrogel thickness but with a greater half-maximal response at 54 µm. Depth-sensing was dependent on Rho-associated protein kinase-mediated cellular contractility. Surface hydrogel deformations were significantly greater on thick hydrogels compared to thin hydrogels. In addition, deformations extended greater distances from the periphery of colonies on thick hydrogels compared to thin hydrogels. Our data suggest that by acting collectively, cells mechanosense rigid materials beneath elastic hydrogels at greater depths than individual cells. This raises the possibility that the collective action of cells in colonies or sheets may allow cells to sense structures of differing material properties at comparatively large distances.

Polymersome nanoparticles for delivery of Wnt-activating small molecules.

Spatiotemporal control of drug delivery is important for a number of medical applications and may be achieved using polymersome nanoparticles (PMs). Wnt signalling is a molecular pathway activated in various physiological processes, including bone repair, that requires precise control of activation. Here, we hypothesise that PMs can be stably loaded with a small molecule Wnt agonist, 6-bromoindirubin-3'-oxime (BIO), and activate Wnt signalling promoting the osteogenic differentiation in human primary bone marrow stromal cells (BMSCs). We showed that BIO-PMs induced a 40% increase in Wnt signaling activation in reporter cell lines without cytotoxicity induced by free BIO. BMSCs incubated with BIO-PMs showed a significant up-regulation of the Wnt target gene AXIN2 (14 ±â€¯4 fold increase, P < 0.001) and a prolonged activation of the osteogenic gene RUNX2. We conclude that BIO-PMs could represent an innovative approach for the controlled activation of Wnt signaling for promoting bone regeneration after fracture.

Single-Molecule Imaging of Wnt3A Protein Diffusion on Living Cell Membranes.

Wnt proteins are secreted, hydrophobic, lipidated proteins found in all animals that play essential roles in development and disease. Lipid modification is thought to facilitate the interaction of the protein with its receptor, Frizzled, but may also regulate the transport of Wnt protein and its localization at the cell membrane. Here, by employing single-molecule fluorescence techniques, we show that Wnt proteins associate with and diffuse on the plasma membranes of living cells in the absence of any receptor binding. We find that labeled Wnt3A transiently and dynamically associates with the membranes of Drosophila Schneider 2 cells, diffuses with Brownian kinetics on flattened membranes and on cellular protrusions, and does not transfer between cells in close contact. In S2 receptor-plus (S2R+) cells, which express Frizzled receptors, membrane diffusion rate is reduced and membrane residency time is increased. These results provide direct evidence of Wnt3A interaction with living cell membranes, and represent, to our knowledge, a new system for investigating the dynamics of Wnt transport.

Transient Canonical Wnt Stimulation Enriches Human Bone Marrow Mononuclear Cell Isolates for Osteoprogenitors.

Activation of the canonical Wnt signaling pathway is an attractive anabolic therapeutic strategy for bone. Emerging data suggest that activation of the Wnt signaling pathway promotes bone mineral accrual in osteoporotic patients. The effect of Wnt stimulation in fracture healing is less clear as Wnt signaling has both stimulatory and inhibitory effects on osteogenesis. Here, we tested the hypothesis that transient Wnt stimulation promotes the expansion and osteogenesis of a Wnt-responsive stem cell population present in human bone marrow. Bone marrow mononuclear cells (BMMNCs) were isolated from patients undergoing hip arthroplasty and exposed to Wnt3A protein. The effect of Wnt pathway stimulation was determined by measuring the frequency of stem cells within the BMMNC populations by fluorescence-activated cell sorting and colony forming unit fibroblast (CFU-F) assays, before determining their osteogenic capacity in in vitro differentiation experiments. We found that putative skeletal stem cells in BMMNC isolates exhibited elevated Wnt pathway activity compared with the population as whole. Wnt stimulation resulted in an increase in the frequency of skeletal stem cells marked by the STRO-1(bright) /Glycophorin A(-) phenotype. Osteogenesis was elevated in stromal cell populations arising from BMMNCs transiently stimulated by Wnt3A protein, but sustained stimulation inhibited osteogenesis in a concentration-dependent manner. These results demonstrate that Wnt stimulation could be used as a therapeutic approach by transient targeting of stem cell populations during early fracture healing, but that inappropriate stimulation may prevent osteogenesis.

The role of intracellular calcium phosphate in osteoblast-mediated bone apatite formation.

Mineralization is a ubiquitous process in the animal kingdom and is fundamental to human development and health. Dysfunctional or aberrant mineralization leads to a variety of medical problems, and so an understanding of these processes is essential to their mitigation. Osteoblasts create the nano-composite structure of bone by secreting a collagenous extracellular matrix (ECM) on which apatite crystals subsequently form. However, despite their requisite function in building bone and decades of observations describing intracellular calcium phosphate, the precise role osteoblasts play in mediating bone apatite formation remains largely unknown. To better understand the relationship between intracellular and extracellular mineralization, we combined a sample-preparation method that simultaneously preserved mineral, ions, and ECM with nano-analytical electron microscopy techniques to examine osteoblasts in an in vitro model of bone formation. We identified calcium phosphate both within osteoblast mitochondrial granules and intracellular vesicles that transported material to the ECM. Moreover, we observed calcium-containing vesicles conjoining mitochondria, which also contained calcium, suggesting a storage and transport mechanism. Our observations further highlight the important relationship between intracellular calcium phosphate in osteoblasts and their role in mineralizing the ECM. These observations may have important implications in deciphering both how normal bone forms and in understanding pathological mineralization.

Use of oxygen-loaded nanobubbles to improve tissue oxygenation: Bone-relevant mechanisms of action and effects on osteoclast differentiation.

Gas-loaded nanobubbles have potential as a method of oxygen delivery to increase tumour oxygenation and therapeutically alleviate tumour hypoxia. However, the mechanism(s) whereby oxygen-loaded nanobubbles increase tumour oxygenation are unknown; with their calculated oxygen-carrying capacity being insufficient to explain this effect. Intra-tumoural hypoxia is a prime therapeutic target, at least partly due to hypoxia-dependent stimulation of the formation and function of bone-resorbing osteoclasts which establish metastatic cells in bone. This study aims to investigate potential mechanism(s) of oxygen delivery and in particular the possible use of oxygen-loaded nanobubbles in preventing bone metastasis via effects on osteoclasts. Lecithin-based nanobubbles preferentially interacted with phagocytic cells (monocytes, osteoclasts) via a combination of lipid transfer, clathrin-dependent endocytosis and phagocytosis. This interaction caused general suppression of osteoclast differentiation via inhibition of cell fusion. Additionally, repeat exposure to oxygen-loaded nanobubbles inhibited osteoclast formation to a greater extent than nitrogen-loaded nanobubbles. This gas-dependent effect was driven by differential effects on the fusion of mononuclear precursor cells to form pre-osteoclasts, partly due to elevated potentiation of RANKL-induced ROS by nitrogen-loaded nanobubbles. Our findings suggest that oxygen-loaded nanobubbles could represent a promising therapeutic strategy for cancer therapy; reducing osteoclast formation and therefore bone metastasis via preferential interaction with monocytes/macrophages within the tumour and bone microenvironment, in addition to known effects of directly improving tumour oxygenation.

Remembering our first date brings back those fuzzy feelings: The role of romantic nostalgia in relationship functioning.

People often reflect nostalgicallyon the momentous occasions of their relationships. Experiencing romantic nostalgia-nostalgia for shared experiences with one's current romantic partner-likely confers relationship-specific benefits. Indeed, prior research indirectly hints at this assumed outcome by looking at relationship narratives. More recent work provides direct evidence that romantic nostalgia is both positively correlated with and causally enhances relationship-specific benefits. We describe such research and discuss avenues for future investigations that address mechanisms and potential downsides of romantic nostalgia while exploring translational possibilities.

An Acoustic Device for Ultra High-Speed Quantification of Cell Strain During Cell-Microbubble Interaction.

Microbubbles utilize high-frequency oscillations under ultrasound stimulation to induce a range of therapeutic effects in cells, often through mechanical stimulation and permeabilization of cells. One of the largest challenges remaining in the field is the characterization of interactions between cells and microbubbles at therapeutically relevant frequencies. Technical limitations, such as employing sufficient frame rates and obtaining sufficient image resolution, restrict the quantification of the cell's mechanical response to oscillating microbubbles. Here, a novel methodology was developed to address many of these limitations and improve the image resolution of cell-microbubble interactions at high frame rates. A compact acoustic device was designed to house cells and microbubbles as well as a therapeutically relevant acoustic field while being compatible with a Shimadzu HPV-X camera. Cell viability tests confirmed the successful culture and proliferation of cells, and the attachment of DSPC- and cationic DSEPC-microbubbles to osteosarcoma cells was quantified. Microbubble oscillation was observed within the device at a frame rate of 5 million FPS, confirming suitable acoustic field generation and ultra high-speed image capture. High spatial resolution in these images revealed observable deformation in cells following microbubble oscillation and supported the first use of digital image correlation for strain quantification in a single cell. The novel acoustic device provided a simple, effective method for improving the spatial resolution of cell-microbubble interaction images, presenting the opportunity to develop an understanding of the mechanisms driving the therapeutic effects of oscillating microbubbles upon ultrasound exposure.

Tracking cellular uptake, intracellular trafficking and fate of nanoclay particles in human bone marrow stromal cells.

Clay nanoparticles, in particular synthetic smectites, have generated interest in the field of tissue engineering and regenerative medicine due to their utility as cross-linkers for polymers in biomaterial design and as protein release modifiers for growth factor delivery. In addition, recent studies have suggested a direct influence on the osteogenic differentiation of responsive stem and progenitor cell populations. Relatively little is known however about the mechanisms underlying nanoclay bioactivity and in particular the cellular processes involved in nanoclay-stem cell interactions. In this study we employed confocal microscopy, inductively coupled plasma mass spectrometry and transmission electron microscopy to track the interactions between clay nanoparticles and human bone marrow stromal cells (hBMSCs). In particular we studied nanoparticle cellular uptake mechanisms and uptake kinetics, intracellular trafficking pathways and the fate of endocytosed nanoclay. We found that nanoclay particles present on the cell surface as µm-sized aggregates, enter hBMSCs through clathrin-mediated endocytosis, and their uptake kinetics follow a linear increase with time during the first week of nanoclay addition. The endocytosed particles were observed within the endosomal/lysosomal compartments and we found evidence for both intracellular degradation of nanoclay and exocytosis as well as an increase in autophagosomal activity. Inhibitor studies indicated that endocytosis was required for nanoclay upregulation of alkaline phosphatase activity but a similar dependency was not observed for autophagy. This study into the nature of nanoclay-stem cell interactions, in particular the intracellular processing of nanosilicate, may provide insights into the mechanisms underlying nanoclay bioactivity and inform the successful utilisation of clay nanoparticles in biomaterial design.

Depth profiling via nanoindentation for characterisation of the elastic modulus and hydraulic properties of thin hydrogel layers.

The accurate determination of the mechanical properties of hydrogels is of fundamental importance for a range of applications, including in assessing the effect of stiffness on cell behaviour. This is a particular issue when using thin hydrogel layers adherent to stiff substrate supports, as the apparent stiffness can be significantly influenced by the constraint of the underlying impermeable substrate, leading to inaccurate measurements of the elastic modulus and permeability of thin hydrogel layers. This study used depth profiling nanoindentation and a poroelastic model for spherical indentation to identify the elastic moduli and hydraulic conductivity of thin polyacrylamide (PAAm) hydrogel layers (∼27 µm-782 µm thick) on impermeable substrates. The apparent stiffness of thin PAAm layers increased with indentation depth and was significantly greater than those of thicker hydrogels, which showed no influence of indentation depth. The hydraulic conductivity decreased as the geometrical confinement of hydrogels increased, indicating that the fluid became more constrained within the confinement areas. The impact of geometrical confinement on the apparent modulus and hydraulic conductivity of thin PAAm hydrogel layers was then established, and their elastic moduli and intrinsic permeability were determined in relation to this effect. This study offers valuable insights into the mechanical characterisation of thin PAAm hydrogel layers used for the fundamental study of cell mechanobiology.

Self-Assembly of Structured Colloidal Gels for High-Resolution 3D Micropatterning of Proteins at Scale.

Self-assembly, the spontaneous ordering of components into patterns, is widespread in nature and fundamental to generating function across length scales. Morphogen gradients in biological development are paradigmatic as both products and effectors of self-assembly and various attempts have been made to reproduce such gradients in biomaterial design. To date, approaches have typically utilized top-down fabrication techniques that, while allowing high-resolution control, are limited by scale and require chemical cross-linking steps to stabilize morphogen patterns in time. Here, a bottom-up approach to protein patterning is developed based on a novel binary reaction-diffusion process where proteins function as diffusive reactants to assemble a nanoclay-protein composite hydrogel. Using this approach, it is possible to generate scalable and highly stable 3D patterns of target proteins down to sub-cellular resolution through only physical interactions between clay nanoparticles and the proteins and ions present in blood. Patterned nanoclay gels are able to guide cell behavior to precisely template bone tissue formation in vivo. These results demonstrate the feasibility of stabilizing 3D gradients of biological signals through self-assembly processes and open up new possibilities for morphogen-based therapeutic strategies and models of biological development and repair.

Changes in embryonic stem cell colony morphology and early differentiation markers driven by colloidal crystal topographical cues.

The use of materials properties to guide cell behaviour is an attractive option for regenerative medicine, where controlling stem cell behaviour is important for the establishment of a functioning cell population. A wide range of materials properties have been shown to influence many types of cells but little is known about the effects of topography on embryonic stem cells (ESCs). In order to advance this knowledge, we synthesised and characterised substrates formed of silica colloidal crystal (SCC) microspheres to present highly ordered and reproducible topographical features from 120-600 nm in diameter. We found that, compared to cells cultured on flat glass, cells cultured on the SCC substrates retained transcription of stem cell (Dppa5a, Nanog, and Pou5f1) and endoderm (Afp, Gata4, Sox17, and Foxa2) markers more similar to undifferentiated ESCs, suggesting the substrates are restricting differentiation, particularly towards the endoderm lineage. Additionally, five days after seeding, we observed strikingly different colony morphology, with cells on the SCC substrates growing in spherical colonies approximately ten cells thick, while cells on glass were growing in flat monolayers. Colonies on the SCC substrates developed a central pit, which was never observed in cells cultured on glass, and expressed proteins related to epithelialisation. Together, these data demonstrate the potential of using topographical cues to control stem cell behaviour in vitro.

Gene-expression analysis reveals that embryonic stem cells cultured under osteogenic conditions produce mineral non-specifically compared to marrow stromal cells or osteoblasts.

Pluripotent cells, such as embryonic stem cells (ESCs), divide indefinitely and can differentiate to form mineralised nodules in response to osteogenic supplements. This suggests that they may be used as a cell source for bone replacement strategies. Here, we related the expression of osteogenic and chondrogenic genes in cultures of murine ESCs, marrow stromal cells (MSCs) and calvarial osteoblasts (OBs) cultured under osteogenic conditions to the biochemical composition and quantity of mineral formed. Mineralisation, measured by calcium sequestration, was >2-fold greater in ESC cultures than in either MSCs or OBs. Micro-Raman spectroscopy and spectral mapping revealed a lower mineral-to-matrix ratio and confirmed a more diffuse pattern of mineralisation in ESCs compared to MSCs and OBs. Baseline expression of chondrogenic and osteogenic genes was between 1 and 4 orders of magnitude greater in MSCs and OBs than in ESCs. Osteogenic culture of MSCs and OBs was accompanied by increases in osteogenic gene expression by factors of ~100 compared to only ~10 in ESCs. Consequentially, peak expression of osteogenic and chondrogenic genes was greater in MSCs and OBs than ESCs by factors of 100-1000, despite the fact that mineralisation was more extensive in ESCs than either MSCs or OBs. We also observed significant cell death in ESC nodules. We conclude that the mineralised material observed in cultures of murine ESCs during osteogenic differentiation may accumulate non-specifically, perhaps in necrotic cell layers, and that thorough characterisation of the tissue formed by ESCs must be achieved before these cells can be considered as a cell source for clinical applications.

What shall we call God? An exploration of metaphors coded from descriptions of God from a large U.S. undergraduate sample.

People use numerous metaphors to describe God. God is seen as a bearded man, light, and love. Based on metaphor theories, the metaphors people use to refer to God reflect how people think about God and could, in turn, reflect their worldview. However, little work has explored the common metaphors for God. This was the purpose of the current investigation. Four trained raters coded open-ended responses from predominantly Christian U.S. undergraduates (N = 2,923) describing God for the presence or absence of numerous metaphoric categories. We then assessed the frequency of each of the metaphor categories. We identified 16 metaphor categories that were present in more than 1% of the responses. The top categories were "GOD IS POWER," "GOD IS HUMAN," and "GOD IS MALE." These findings were similar across religious affiliations. We attempted to support our coding analysis using top-down and bottom-up automated language analysis. Results from these analyses provided added confidence to our conclusions. We discuss the implications of our findings and the potential for future studies investigating important psychological and behavioral outcomes of using different metaphors for God.

The role of lithium in the osteogenic bioactivity of clay nanoparticles.

LAPONITE® clay nanoparticles are known to exert osteogenic effects on human bone marrow stromal cells (HBMSCs), most characteristically, an upregulation in alkaline phosphatase activity and increased calcium deposition. The specific properties of LAPONITE® that impart its bioactivity are not known. In this study the role of lithium, a LAPONITE® degradation product, was investigated through the use of lithium salts and lithium modified LAPONITE® formulations. In contrast to intact particles, lithium ions applied at concentrations equivalent to that present in LAPONITE®, failed to induce any significant increase in alkaline phosphatase (ALP) activity. Furthermore, no significant differences were observed in ALP activity with modified clay structures and the positive effect on osteogenic gene expression did not correlate with the lithium content of modified clays. These results suggest that other properties of LAPONITE® nanoparticles, and not their lithium content, are responsible for their bioactivity.

Tailoring the size of ultrasound responsive lipid-shelled nanodroplets by varying production parameters and environmental conditions.

Liquid perfluorocarbon nanodroplets (NDs) are an attractive alternative to microbubbles (MBs) for ultrasound-mediated therapeutic and diagnostic applications. ND size and size distribution have a strong influence on their behaviour in vivo, including extravasation efficiency, circulation time, and response to ultrasound stimulation. Thus, it is desirable to identify ways to tailor the ND size and size distribution during manufacturing. In this study phospholipid-coated NDs, comprising a perfluoro-n-pentane (PFP) core stabilised by a DSPC/PEG40s (1,2-distearoyl-sn-glycero-3-phosphocholine and polyoxyethylene(40)stearate, 9:1 molar ratio) shell, were produced in phosphate-buffered saline (PBS) by sonication. The effect of the following production-related parameters on ND size was investigated: PFP concentration, power and duration of sonication, and incorporation of a lipophilic fluorescent dye. ND stability was also assessed at both 4 °C and 37 °C. When a sonication pulse of 6 s and 15% duty cycle was employed, increasing the volumetric concentration of PFP from 5% to 15% v/v in PBS resulted in an increase in ND diameter from 215.8 ± 16.8 nm to 408.9 ± 171.2 nm. An increase in the intensity of sonication from 48 to 72 W (with 10% PFP v/v in PBS) led to a decrease in ND size from 354.6 ± 127.2 nm to 315.0 ± 100.5 nm. Increasing the sonication time from 20 s to 40 s (using a pulsed sonication with 30% duty cycle) did not result in a significant change in ND size (in the range 278-314 nm); however, when it was increased to 60 s, the average ND diameter reduced to 249.7 ± 9.7 nm, which also presented a significantly lower standard deviation compared to the other experimental conditions investigated (i.e., 9.7 nm vs. > 49.4 nm). The addition of the fluorescent dye DiI at different molar ratios did not affect the ND size distribution. NDs were stable at 4 °C for up to 6 days and at 37 °C for up to 110 min; however, some evidence of ND-to-MB phase transition was observed after 40 min at 37 °C. Finally, phase transition of NDs into MBs was demonstrated using a tissue-mimicking flow phantom under therapeutic ultrasound exposure conditions (ultrasound frequency: 0.5 MHz, acoustic pressure: 2-4 MPa, and pulse repetition frequency: 100 Hz).

Investigation of the Acoustic Vaporization Threshold of Lipid-Coated Perfluorobutane Nanodroplets Using Both High-Speed Optical Imaging and Acoustic Methods.

A combination of ultrahigh-speed optical imaging (5 × 106 frames/s), B-mode ultrasound and passive cavitation detection was used to study the vaporization process and determine both the acoustic droplet vaporization (ADV) and inertial cavitation (IC) thresholds of phospholipid-coated perfluorobutane nanodroplets (PFB NDs, diameter = 237 ± 16 nm). PFB NDs have not previously been studied with ultrahigh-speed imaging and were observed to form individual microbubbles (1-10 µm) within two to three cycles and subsequently larger bubble clusters (10-50 µm). The ADV and IC thresholds did not statistically significantly differ and decreased with increasing pulse length (20-20,000 cycles), pulse repetition frequency (1-100 Hz), concentration (108-1010 NDs/mL), temperature (20°C-45°C) and decreasing frequency (1.5-0.5 MHz). Overall, the results indicate that at frequencies of 0.5, 1.0 and 1.5 MHz, PFB NDs can be vaporized at moderate peak negative pressures (<2.0 MPa), pulse lengths and pulse repetition frequencies. This finding is encouraging for the use of PFB NDs as cavitation agents, as these conditions are comparable to those required to achieve therapeutic effects with microbubbles, unlike those reported for higher-boiling-point NDs. The differences between the optically and acoustically determined ADV thresholds, however, suggest that application-specific thresholds should be defined according to the biological/therapeutic effect of interest.

Synthetic Nanoclay Gels Do Not Cause Skin Irritation in Healthy Human Volunteers.

Synthetic clays are promising biomaterials for delivery of therapeutic molecules in regenerative medicine. However, before their use can be translated into clinical applications, their safety must be assessed in human volunteers. The aim of this study was to test the hypothesis that a synthetic nanoclay (LAPONITE) does not cause irritation to the human skin. To achieve this, a nanoclay gel at two different concentrations (1.5 and 3% w/v) was applied on the forearm of healthy volunteers for 24 h. 1% sodium lauryl sulfate (SLS) and 3% (w/v) polyacrylic acid were used as the positive and negative controls, respectively. The compromise in the skin barrier function was measured by trans-epidermal water loss (TEWL), erythema by spectroscopic measurements, and skin inflammatory biomarkers (IL-1α and IL-1RA) by the enzyme-linked immunosorbent assay. We found that the nanoclay caused no prolonged increase in TEWL, erythema, or induction of inflammatory cytokines. This was in contrast to 1% SLS, a known irritant, which induced significant increases in both skin erythema and TEWL. We conclude that the nanoclay is not an irritant and is thus suitable for therapeutic interventions at the skin surface.