Engineering vascularized skin-mimetic phantom for non-invasive Raman spectroscopy.

Recent advances in Raman spectroscopy have shown great potential for non-invasive analyte sensing, but the lack of a standardized optical phantom for these measurements has hindered further progress. While many research groups have developed optical phantoms that mimic bulk optical absorption and scattering, these materials typically have strong Raman scattering, making it difficult to distinguish metabolite signals. As a result, solid tissue phantoms for spectroscopy have been limited to highly scattering tissues such as bones and calcifications, and metabolite sensing has been primarily performed using liquid tissue phantoms. To address this issue, we have developed a layered skin-mimetic phantom that can support metabolite sensing through Raman spectroscopy. Our approach incorporates millifluidic vasculature that mimics blood vessels to allow for diffusion akin to metabolite diffusion in the skin. Furthermore, our skin phantoms are mechanically mimetic, providing an ideal model for development of minimally invasive optical techniques. By providing a standardized platform for measuring metabolites, our approach has the potential to facilitate critical developments in spectroscopic techniques and improve our understanding of metabolite dynamics in vivo.

Functional heterogeneity of IFN-γ-licensed mesenchymal stromal cell immunosuppressive capacity on biomaterials.

Mesenchymal stromal cells (MSCs) are increasingly combined with biomaterials to enhance their therapeutic properties, including their immunosuppressive function. However, clinical trials utilizing MSCs with or without biomaterials have shown limited success, potentially due to their functional heterogeneity across different donors and among different subpopulations of cells. Here, we evaluated the immunosuppressive capacity, as measured by the ability to reduce T-cell proliferation and activation, of interferon-gamma (IFN-γ)-licensed MSCs from multiple donors on fibrin and collagen hydrogels, the two most commonly utilized biomaterials in combination with MSCs in clinical trials worldwide according to ClinicalTrials.gov Variations in the immunosuppressive capacity between IFN-γ-licensed MSC donors on the biomaterials correlated with the magnitude of indoleamine-2,3-dioxygenase activity. Immunosuppressive capacity of the IFN-γ-licensed MSCs depended on the αV/α5 integrins when cultured on fibrin and on the α2/ß1 integrins when cultured on collagen. While all tested MSCs were nearly 100% positive for these integrins, sorted MSCs that expressed higher levels of αV/α5 integrins demonstrated greater immunosuppressive capacity with IFN-γ licensing than MSCs that expressed lower levels of these integrins on fibrin. These findings were equivalent for MSCs sorted based on the α2/ß1 integrins on collagen. These results demonstrate the importance of integrin engagement to IFN-γ licensed MSC immunosuppressive capacity and that IFN-γ-licensed MSC subpopulations of varying immunosuppressive capacity can be identified by the magnitude of integrin expression specific to each biomaterial.

SETD2-mediated H3K14 trimethylation promotes ATR activation and stalled replication fork restart in response to DNA replication stress.

Ataxia telangiectasia and Rad3 related (ATR) activation after replication stress involves a cascade of reactions, including replication protein A (RPA) complex loading onto single-stranded DNA and ATR activator loading onto chromatin. The contribution of histone modifications to ATR activation, however, is unclear. Here, we report that H3K14 trimethylation responds to replication stress by enhancing ATR activation. First, we confirmed that H3K14 monomethylation, dimethylation, and trimethylation all exist in mammalian cells, and that both SUV39H1 and SETD2 methyltransferases can catalyze H3K14 trimethylation in vivo and in vitro. Interestingly, SETD2-mediated H3K14 trimethylation markedly increases in response to replication stress induced with hydroxyurea, a replication stress inducer. Under these conditions, SETD2-mediated H3K14me3 recruited the RPA complex to chromatin via a direct interaction with RPA70. The increase in H3K14me3 levels was abolished, and RPA loading was attenuated when SETD2 was depleted or H3K14 was mutated. Rather, the cells were sensitive to replication stress such that the replication forks failed to restart, and cell-cycle progression was delayed. These findings help us understand how H3K14 trimethylation links replication stress with ATR activation.

Toward Single Cell Tattoos: Biotransfer Printing of Lithographic Gold Nanopatterns on Live Cells.

Lithographic nanopatterning techniques such as photolithography, electron-beam lithography, and nanoimprint lithography (NIL) have revolutionized modern-day electronics and optics. Yet, their application for creating nanobio interfaces is limited by the cytotoxic and two-dimensional nature of conventional fabrication methods. Here, we present a biocompatible and cost-effective transfer process that leverages (a) NIL to define sub-300 nm gold (Au) nanopattern arrays, (b) amine functionalization of Au to transfer the NIL-arrays from a rigid substrate to a soft transfer layer, (c) alginate hydrogel as a flexible, degradable transfer layer, and (d) gelatin conjugation of the Au NIL-arrays to achieve conformal contact with live cells. We demonstrate biotransfer printing of the Au NIL-arrays on rat brains and live cells with high pattern fidelity and cell viability and observed differences in cell migration on the Au NIL-dot and NIL-wire printed hydrogels. We anticipate that this nanolithography-compatible biotransfer printing method could advance bionics, biosensing, and biohybrid tissue interfaces.

The living interface between synthetic biology and biomaterial design.

Recent far-reaching advances in synthetic biology have yielded exciting tools for the creation of new materials. Conversely, advances in the fundamental understanding of soft-condensed matter, polymers and biomaterials offer new avenues to extend the reach of synthetic biology. The broad and exciting range of possible applications have substantial implications to address grand challenges in health, biotechnology and sustainability. Despite the potentially transformative impact that lies at the interface of synthetic biology and biomaterials, the two fields have, so far, progressed mostly separately. This Perspective provides a review of recent key advances in these two fields, and a roadmap for collaboration at the interface between the two communities. We highlight the near-term applications of this interface to the development of hierarchically structured biomaterials, from bioinspired building blocks to 'living' materials that sense and respond based on the reciprocal interactions between materials and embedded cells.

Structurally Dynamic Hydrogels for Biomedical Applications: Pursuing a Fine Balance between Macroscopic Stability and Microscopic Dynamics.

Owing to their unique chemical and physical properties, hydrogels are attracting increasing attention in both basic and translational biomedical studies. Although the classical hydrogels with static networks have been widely reported for decades, a growing number of recent studies have shown that structurally dynamic hydrogels can better mimic the dynamics and functions of natural extracellular matrix (ECM) in soft tissues. These synthetic materials with defined compositions can recapitulate key chemical and biophysical properties of living tissues, providing an important means to understanding the mechanisms by which cells sense and remodel their surrounding microenvironments. This review begins with the overall expectation and design principles of dynamic hydrogels. We then highlight recent progress in the fabrication strategies of dynamic hydrogels including both degradation-dependent and degradation-independent approaches, followed by their unique properties and use in biomedical applications such as regenerative medicine, drug delivery, and 3D culture. Finally, challenges and emerging trends in the development and application of dynamic hydrogels are discussed.

Different Regulatory Effects of Heated Products and Maillard Reaction Products of Half-Fin Anchovy Hydrolysates on Intestinal Antioxidant Defense in Healthy Animals.

The oxidative state of intestinal tracts of healthy animals were investigated after short-term intake of half-fin anchovy hydrolysates (HAHp) and their thermal or Maillard reaction products (MRPs). After one month of continuous oral gavage of HAHp, HAHp-heated products (HAHp-H), the MRPs of HAHp with 3% of glucose (HAHp-3%G MRPs), and the MRPs of HAHp with 3% of fructose (HAHp-3%F MRPs) at a dose of 1.0 g/kg of body weight per day into healthy ICR male mice, the concentrations of serum low-density and high-density lipoprotein cholesterol did not significantly change compared to the control group (CK, gavage with saline). Similar results were found for the interleukin-6 concentrations of all groups. By comparison, HAHp-H, HAHp-3%G MRPs, and HAHp-3%F MRPs administration decreased serum tumor necrosis factor-α concentration as compared to the CK group (p < 0.05). No histological damage was observed in the jejunum, ileum, and colonic tissues of all groups. However, HAHp-H treatment induced higher upregulation of Kelch-like ECH-associated protein 1, transcription factors Nrf-2, associated protective phase-II enzymes of NAD(P)H: quinine oxidoreductase-1, and hemoxygenase-1 in colon tissue, as well as higher upregulation of endogenous antioxidant enzymes, including copper/zinc superoxide dismutase, manganese superoxide dismutase, catalase, and glutathione peroxidase 2 than other groups (p < 0.05). Additionally, increases in Nε-carboxymethyllysine expression in the colonic tissues of all groups were consistent with their increased oligopeptide transporter 1 expressions. Our results suggest that the thermal products of HAHp might have a broad application prospect in improving antioxidant defense in vivo in healthy animals.

Antibacterial Effect of Shrimp By-Products Hydrolysate on Specific Spoilage Organisms of Squid.

In order to further develop and utilize shrimp processing by-products, in this study, a novel antibacterial hydrolysate of shrimp by-products by pepsin hydrolysis (SPH) was prepared. The antibacterial effect of SPH on specific spoilage organisms of squid after end storage at room temperature (SE-SSOs) was investigated. SPH showed an antibacterial effect on the growth of SE-SSOs, with (23.4 ± 0.2) mm of inhibition zone diameter. The cell permeability of SE-SSOs was enhanced after SPH treatment for 12 h. Some bacteria were twisted and shrunk, while pits and pores formed and intracellular contents leaked under scanning electron microscopy observation. The flora diversity of SE-SSOs treated with SPH was determined by a 16S rDNA sequencing technique. Results showed that SE-SSOs were mainly composed of the phyla of Firmicutes and Proteobacteria, among which Paraclostridium (47.29%) and Enterobacter (38.35%) were dominant genera. SPH treatment resulted in a significant reduction in the relative abundance of the genus Paraclostridium and increased the abundance of Enterococcus. Linear discriminant analysis (LDA) of LEfSe conveyed that SPH treatment had a significant impact on altering the bacterial structure of SE-SSOs. The 16S PICRUSt of Cluster of Orthologous Group (COG) annotation revealed that SPH treatment for 12 h could significantly increase the function of transcription level [K], while SPH treatment for 24 h could downregulate post-translational modifications, protein turnover, and chaperone metabolism functions [O]. In conclusion, SPH has a proper antibacterial effect on SE-SSOs and can change the flora structure of SE-SSOs. These findings will provide a technical basis for the development of inhibitors of squid SSOs.

Ultra-thin multifocal integral LED-projector based on aspherical microlens arrays.

Multifocal imaging has been a challenging and rewarding research focus in the field of imaging optics. In this paper, an ultra-thin multifocal integral LED-projector based on aspherical microlens array (MLA) is presented. A two-layer aspherical sub-lens with NA = 0.3 is proposed as a sub-channel projector and the optimization design ensures high optical integration precision and improves optical efficiency. To avoid the tailoring loss of the projected images between multi-plane projections, the central-projection constraints between size and projection distance for the multifocal projection are defined. The depth of focus (DOF) analysis for MLA and sub-lens is also introduced to proof the sufficiency of realizing multifocal projection. Combined with the radial basis function image warping method, multifocal sub-image arrays were acquired, and three types of multifocal integral projection were realized, breaking through the traditional limitations of the single-focal DOF. A prototype with thickness of less than 4 mm is developed. Substantial simulations and experiments are conducted to verify the effectiveness of the method and the design.

RNF8-ubiquitinated KMT5A is required for RNF168-induced H2A ubiquitination in response to DNA damage.

Histone modifications play critical roles in DNA damage repair to safeguard genome integrity. However, how different histone modifiers coordinate to build appropriate chromatin context for DNA damage repair is largely unknown. Here, we report a novel interplay between the histone methyltransferase KMT5A and two E3 ligases RNF8 and RNF168 in establishing the histone modification status for DNA damage repair. KMT5A is a newly identified substrate of RNF8 in vitro and in vivo. In response to DNA double-strand breaks (DSBs), RNF8 promotes KMT5A recruitment onto damaged chromatin in a ubiquitination-dependent manner. RNF8-induced KMT5A ubiquitination increases the binding capacity of KMT5A to RNF168. Interestingly, KMT5A not only drives a local increase in H4K20 monomethylation at DSBs, but also promotes RNF168's activity in catalyzing H2A ubiquitination. We proved that the interaction between the H2A acidic patch and KMT5A R188/R189 residues is critical for KMT5A-mediated regulation of H2A ubiquitination. Taken together, our results highlight a new role for KMT5A in linking H4K20 methylation and H2A ubiquitination and provide insight into the histone modification network during DNA damage repair.

Digestive properties of half-fin anchovy hydrolysates/glucose Maillard reaction products and modulation effects on intestinal microbiota.

BACKGROUND: The consumption of dietary Maillard reaction products (MRPs) might lead to positive or negative effects on health. The digestibility of half-fin anchovy hydrolysates/glucose MRPs (HAHp(9.0)-G MRPs) was therefore determined. The intestinal microbiota modulation of HAHp(9.0)-G MRPs in mice was also evaluated after administration for 14 days (1 g kg-1 •bodyweight). RESULTS: Different levels of digestibility of MRPs of fructosamine and advanced glycation products of Nε -carboxymethyllysine were detected in HAHp(9.0)-G MRPs during simulated gastrointestinal digestion. An increased relative proportion of soluble fluorescent melanoidins (SFMs) was observed during gastric digestion as compared to that in the original HAHp(9.0)-G MRPs, followed by decreases in SFMs in intestinal digestion. After feeding with HAHp(9.0)-G MRPs for 14 days, increased goblet cells were observed in the ileum regions of female and male mice. High-throughput 16S ribosomal RNA gene sequencing of fecal samples revealed that HAHp(9.0)-G MRPs administration increased the density of the phylum Bacteriodetes and reduced the density of the phylum Firmicutes in male mice. By comparison, a relatively higher density of members of the phylum Saccharibacteria was observed in female mice. A consistent increase in the abundance of Bacteroidales_S24-7_group_norank was found in female and male groups fed with HAHp(9.0)-G MRPs. Female and male mice treated with HAHp(9.0)-G MRPs also showed higher levels of propionic and butyric acids in feces than their corresponding controls. CONCLUSION: Half-fin anchovy hydrolysates/glucose MRPs can be partly hydrolyzed in the simulated gastrointestinal digestion system. Treatment with HAHp(9.0)-G MRPs induced sex-related differences in bacterial abundance and diversity in mice; however, the up-regulation of anti-inflammatory activity was predicted in both female and male mice. © 2021 Society of Chemical Industry.

GLP-catalyzed H4K16me1 promotes 53BP1 recruitment to permit DNA damage repair and cell survival.

The binding of p53-binding protein 1 (53BP1) to damaged chromatin is a critical event in non-homologous DNA end joining (NHEJ)-mediated DNA damage repair. Although several molecular pathways explaining how 53BP1 binds damaged chromatin have been described, the precise underlying mechanisms are still unclear. Here we report that a newly identified H4K16 monomethylation (H4K16me1) mark is involved in 53BP1 binding activity in the DNA damage response (DDR). During the DDR, H4K16me1 rapidly increases as a result of catalyzation by the histone methyltransferase G9a-like protein (GLP). H4K16me1 shows an increased interaction level with 53BP1, which is important for the timely recruitment of 53BP1 to DNA double-strand breaks. Differing from H4K16 acetylation, H4K16me1 enhances the 53BP1-H4K20me2 interaction at damaged chromatin. Consistently, GLP knockdown markedly attenuates 53BP1 foci formation, leading to impaired NHEJ-mediated repair and decreased cell survival. Together, these data support a novel axis of the DNA damage repair pathway based on H4K16me1 catalysis by GLP, which promotes 53BP1 recruitment to permit NHEJ-mediated DNA damage repair.

Material microenvironmental properties couple to induce distinct transcriptional programs in mammalian stem cells.

Variations in a multitude of material microenvironmental properties have been observed across tissues in vivo, and these have profound effects on cell phenotype. Phenomenological experiments have suggested that certain of these features of the physical microenvironment, such as stiffness, could sensitize cells to other features; meanwhile, mechanistic studies have detailed a number of biophysical mechanisms for this sensing. However, the broad molecular consequences of these potentially complex and nonlinear interactions bridging from biophysical sensing to phenotype have not been systematically characterized, limiting the overall understanding and rational deployment of these biophysical cues. Here, we explore these interactions by employing a 3D cell culture system that allows for the independent control of culture substrate stiffness, stress relaxation, and adhesion ligand density to systematically explore the transcriptional programs affected by distinct combinations of biophysical parameters using RNA-seq. In mouse mesenchymal stem cells and human cortical neuron progenitors, we find dramatic coupling among these substrate properties, and that the relative contribution of each property to changes in gene expression varies with cell type. Motivated by the bioinformatic analysis, the stiffness of hydrogels encapsulating mouse mesenchymal stem cells was found to regulate the secretion of a wide range of cytokines, and to accordingly influence hematopoietic stem cell differentiation in a Transwell coculture model. These results give insights into how biophysical features are integrated by cells across distinct tissues and offer strategies to synthetic biologists and bioengineers for designing responses to a cell's biophysical environment.

Probing Membrane Protein Association Using Concentration-Dependent Number and Brightness.

We introduce concentration-dependent number and brightness (cdN&B), a fluorescence fluctuation technique that can be implemented on a standard confocal microscope and can report on the thermodynamics of membrane protein association in the native plasma membrane. It uses transient transfection to enable measurements of oligomer size as a function of receptor concentration over a broad range, yielding the association constant. We discuss artifacts in cdN&B that are concentration-dependent and can distort the oligomerization curves, and we outline procedures that can correct for them. Using cdN&B, we characterize the association of neuropilin 1 (NRP1), a protein that plays a critical role in the development of the embryonic cardiovascular and nervous systems. We show that NRP1 associates into a tetramer in a concentration-dependent manner, and we quantify the strength of the association. This work demonstrates the utility of cdN&B as a powerful tool in biophysical chemistry.

Design of an ultra-thin, wide-angle, stray-light-free near-eye display with a dual-layer geometrical waveguide.

The field of view (FOV) of a geometrical waveguide display is limited by the total internal reflection (TIR) condition (related with the index of glass) and the stray light generated inside the waveguide. A novel concept of an ultra-thin, wide-angle, stray-light-free, optical see-through near-eye display (NED) with a dual-layer geometrical waveguide is proposed in this paper. In the dual-layer waveguide, the two waveguides have different structures and are responsible for two different FOVs which are spliced together to form the entire FOV. The stray light of the dual-layer waveguide is analyzed and an optimized structure to suppress the stray light is designed. An optimized coupling-in structure is designed and a progressive optimization method is proposed for optimizing the illuminance uniformity of the entire FOV across the exit pupil. A dual-layer waveguide with a total thickness of 3.0 mm and stray light of less than 1% is designed. The FOV is 62° in the pupil-expanding direction, and the diameter of the exit pupil (EPD) is 10 mm at an eye relief (ER) of 18 mm. A compact projection optic is designed and finally is integrated with the dual-layer waveguide.

Design and fabrication of an off-axis four-mirror system for head-up displays.

Head-up displays (HUDs), as one kind of the augmented reality (AR) applications, enable users to enhance the situational awareness. In this paper, we propose a design of a HUD employing the freeform off-axis four-mirror construction, in which the eyebox and eye relief as large as 120mm×80mm and 400 mm, respectively, are derived. In the part of system design, the biocular parallax is evaluated and corrected. An optimization strategy in forward ray-tracing mode is presented to directly govern the biocular parallax, and the convergence, divergence, and dipvergence are finally constrained within 3.5 mrad, 1.5 mrad, and 2.0 mrad, respectively. An overall design procedure of a HUD, including initial configuration, structure constraints, optimization method, and parallax correction, is illustrated in detail. After analyzing the tolerance, a proof-of-concept prototype with the field of 24∘×15∘ is developed to demonstrate the image quality.

MICAL-L2 potentiates Cdc42-dependent EGFR stability and promotes gastric cancer cell migration.

Enhanced migration potential is a common characteristic of cancer cells induced by mechanisms that are incompletely defined. The present study was designed to investigate relationship of a new discovered cytoskeleton regulator MICAL-L2 and the endogenous epidermal growth factor receptor (EGFR) signalling pathways in gastric cancer cell migration. Increased expression of MICAL-L2 in gastric cancer cells up-regulated EGFR protein level, accompanied by the increase of cell migration, whereas silencing MICAL-L2 down-regulated EGFR and inhibited cell migration. Expression of MICAL-L2 was also shown positively correlated with the activation of HSP27/cytoskeleton and HSP27/ß-catenin signalling pathways that provide key mechanisms controlling cell migration. The up-regulating effect of MICAL-L2 on EGFR is mediated through a transcription-independent mechanism that involves inhibiting EGFR protein degradation in lysosome. Further analysis indicated that Cdc42 activation contributed in maintaining the effect of MICAL-L2 on EGFR stability. Furthermore analysis of clinic specimens revealed increased expression of MICAL-L2 in carcinoma tissues and a positive correlation between MICAL-L2 and EGFR expression levels. The above results indicate that MICAL-L2 potentiates gastric cell migration via inhibiting EGFR degradation in lysosome via a Cdc42-dependent manner that leads to the activation of EGFR/HSP27 signalling pathways.

Effect of melatonin on EGF- and VEGF-induced monolayer permeability of HUVECs.

Melatonin is a natural hormone involved in the regulation of circadian rhythm, immunity, and cardiovascular function. In the present study, we focused on the mechanism of melatonin in the regulation of vascular permeability. We found that melatonin could inhibit both VEGF- and EGF-induced monolayer permeability of human umbilical vein endothelial cells (HUVECs) and change the tyrosine phosphorylation of vascular-endothelial (VE-)cadherin, which was related to endothelial barrier function. In addition, phospho-AKT (Ser473) and phospho-ERK(1/2) played significant roles in the regulation of VE-cadherin phosphorylation. Both the phosphatidylinositol 3-kinase/AKT inhibitor LY49002 and MEK/ERK inhibitor U0126 could inhibit the permeability of HUVECs, but with different effects on tyrosine phosphorylation of VE-cadherin. Melatonin can influence the two growth factor-induced phosphorylation of AKT (Ser473) but not ERK(1/2). Our results show that melatonin can inhibit growth factor-induced monolayer permeability of HUVECs by influencing the phosphorylation of AKT and VE-cadherin. Melatonin can be a potential treatment for diseases associated with abnormal vascular permeability. NEW & NOTEWORTHY We found that melatonin could inhibit both EGF- and VEGF-induced monolayer permeability of human umbilical vein endothelial cells, which is related to phosphorylation of vascular-endothelial cadherin. Blockade of phosphatidylinositol 3-kinase/AKT and MEK/ERK pathways could inhibit the permeability of human umbilical vein endothelial cells, and phosphorylation of AKT (Ser473) might be a critical event in the changing of monolayer permeability and likely has cross-talk with the MEK/ERK pathway.

Design of a uniform-illumination two-dimensional waveguide head-up display with thin plate compensator.

Nonuniform illumination is one of the factors in degrading image performance of geometrical waveguide-based head-up display. The two-dimensional (2-D) waveguide expanding the exit pupil along two orthogonal directions makes illumination uniformization more intractable. To solve this problem, the reasons for nonuniform illumination in 2-D geometrical waveguide were probed and a novel waveguide structure incorporating illumination compensator was proposed. A bilayer illumination compensator was first presented to change the period of rays propagating in waveguide to improve the inter-pupil illumination uniformity and inter-field illumination uniformity. Then optimized coating design for different partially reflective mirrors in horizontal waveguide and the vertical one allowed further improvement of these two kinds of illumination uniformity, raising both up to 70%. A matched catadioptric projection optics using freeform surface was designed and integrated with the resultant 2-D geometrical waveguide, achieving a head-up display with an eye relief of 400 mm, an eyebox of 80 mm × 80 mm and a field of 24° × 15°. A prototype of the proposed 2-D geometrical waveguide display was developed and demonstrated.

Design and evaluation of a biocular system.

In this study, a biocular system is established and a biocular analysis method is proposed by analyzing the biocular parallax principle to determine the relationship between convergence, dipvergence, azimuth, and elevation. Based on the biocular analysis method, a looped optimization method for reducing biocular parallax is presented. After optimizing the biocular system, convergence of the system is between -6 mrad-6 mrad, and dipvergence is between -1.5 mrad-1.5 mrad. This satisfies the requirements of the human eye for observation with a biocular system. Experimental results are approximately consistent with theoretical analysis. This proves the accuracy of the proposed biocular analysis method.