Hydrophilic Patch Research Articles

Coencapsulating TMB Probes and Bimetallic MOF Nanozymes in a Hydrogel Patch for Fabricating Reusable Visual VC Sensors.

As a typical chromogenic probe, 3,3',5,5'-tetramethylbenzidine (TMB) has been widely applied in the field of visual detection due to its low toxicity and highly sensitive response. Due to the hydrophobic nature of TMB, encapsulating it into a hydrogel, which serves as an ideal matrix for wearable sensors, presents significant challenges that complicate the fabrication of visual wearable devices. Herein, the TMB probe and bimetallic MOF nanozymes are coencapsulated in a hydrogel patch for the fabrication of reusable visual sensors. Hydrophobic TMB is oxidized to hydrophilic ox-TMB by a bimetallic MOF (CuFe-MOF), allowing its diffusion into a hydrophilic agarose hydrogel patch, where it is reduced back to TMB. This process allows the coimmobilization and coencapsulation of CuFe-MOF and TMB within the hydrogel patch. Leveraging the color change between TMB and ox-TMB, as a proof-of-concept application, a reusable visual "On-Off-On" sensor is simply constructed and successfully applied to detect vitamin C in human sweat. Color changes can be quickly read by the naked eye or by smart devices without the need for external equipment. Meanwhile, based on the reversible conversion relationship between TMB and ox-TMB, a reusable sensor construction strategy is proposed. This approach not only facilitates the use of a TMB probe in hydrogel applications but also offers inspiration for the development of point-of-care testing equipment, demonstrating significant application potential.

Optimizing properties of translocation-enhancing transmembrane proteins

Cell membranes act as semi-permeable barriers, often restricting the entry of large or hydrophilic molecules. Nonetheless, certain amphiphilic molecules, such as antimicrobial and cell-penetrating peptides, can cross these barriers. In this study, we demonstrate that specific properties of transmembrane proteins/peptides can enhance membrane permeation of amphiphilic peptides. Using coarse-grained molecular dynamics with free-energy calculations, we identify key translocation-enhancing attributes of transmembrane proteins/peptides: a continuous hydrophilic patch, charged residues preferably in the membrane center, and aromatic hydrophobic residues. By employing both coarse-grained and atomistic simulations, complemented by experimental validation, we show that these properties not only enhance peptide translocation but also speed up lipid flip-flop. The enhanced flip-flop reinforces the idea that proteins such as scramblases and insertases not only share structural features but also operate through identical biophysical mechanisms enhancing the insertion and translocation of amphiphilic molecules. Our insights offer guidelines for the designing of translocation-enhancing proteins/peptides that could be used in medical and biotechnological applications.

Intratumoral delivery of a highly active form of XCL1 enhances antitumor CTL responses through recruitment of CXCL9-expressing conventional type-1 dendritic cells.

Conventional type 1 dendritic cells (cDC1s) play a crucial role in antitumor immunity through the induction and activation of tumor-specific CD8+ cytotoxic T cells (CTLs). The chemokine XCL1 is a major chemotactic factor for cDC1s and its receptor XCR1 is selectively expressed on cDC1s. Here, we investigated the effect of intratumoral delivery of a highly active form of murine XCL1 (mXCL1-V21C/A59C) on cDC1-mediated antitumor immunity using a hydrophilic gel patch. The hydrophilic gel patch containing mXCL1-V21C/A59C increased cDC1 accumulation in the tumor masses and promoted their migration to the regional lymph nodes, resulting in enhanced induction of tumor-specific CTLs. Tumor-infiltrating cDC1s not only expressed XCR1 but also produced CXCL9, a ligand for CXCR3 which is highly expressed on CTLs and NK cells. Consequently, CTLs and NK cells were increased in the tumor masses of mice treated with mXCL1-V21C/A59C, while immunosuppressive cells such as monocyte-derived suppressive cells and regulatory T cells were decreased. We also confirmed that anti-CXCL9 treatment decreased the tumor infiltration of CTLs. The intratumoral delivery of mXCL1-V21C/A59C significantly decreased tumor growth and prolonged survival in E.G7-OVA and B16-F10 tumor-bearing mice. Furthermore, the antitumor effect of mXCL1-V21CA59C was enhanced in combination with anti-programmed cell death protein 1 treatment. Finally, using The Cancer Genome Atlas database, we found that XCL1 expression was positively correlated with tumor-infiltrating cDC1s and a better prognosis in melanoma patients. Collectively, our findings provide a novel therapeutic approach to enhance tumor-specific CTL responses through the selective recruitment of CXCL9-expressing cDC1s into the tumor masses.

Molecular Dynamics of DHHC20 Acyltransferase Suggests Principles of Lipid and Protein Substrate Selectivity.

Lipid modification of viral proteins with fatty acids of different lengths (S-acylation) is crucial for virus pathogenesis. The reaction is catalyzed by members of the DHHC family and proceeds in two steps: the autoacylation is followed by the acyl chain transfer onto protein substrates. The crystal structure of human DHHC20 (hDHHC20), an enzyme involved in the acylation of S-protein of SARS-CoV-2, revealed that the acyl chain may be inserted into a hydrophobic cavity formed by four transmembrane (TM) α-helices. To test this model, we used molecular dynamics of membrane-embedded hDHHC20 and its mutants either in the absence or presence of various acyl-CoAs. We found that among a range of acyl chain lengths probed only C16 adopts a conformation suitable for hDHHC20 autoacylation. This specificity is altered if the small or bulky residues at the cavity’s ceiling are exchanged, e.g., the V185G mutant obtains strong preferences for binding C18. Surprisingly, an unusual hydrophilic ridge was found in TM helix 4 of hDHHC20, and the responsive hydrophilic patch supposedly involved in association was found in the 3D model of the S-protein TM-domain trimer. Finally, the exchange of critical Thr and Ser residues in the spike led to a significant decrease in its S-acylation. Our data allow further development of peptide/lipid-based inhibitors of hDHHC20 that might impede replication of Corona- and other enveloped viruses.

Transmembrane proteins could enhance translocation across cellular membranes

Efficient translocation of specific molecules across cell plasma membrane is crucial for cell viability and drug delivery. However, it remains elusive what molecular properties are necessary for spontaneous translocation and how to transport molecules without such properties. Here, we investigated the effect of transmembrane proteins on the translocation of amphiphilic peptides. Using two different coarse grained models, we consistently demonstrate that transmembrane proteins with a hydrophilic patch enhance the translocation of amphiphilic peptides by their stabilization in the membrane.

Adjuvant Activity of CpG-Oligonucleotide Administered Transcutaneously in Combination with Vaccination Using a Self-Dissolving Microneedle Patch in Mice.

In this study, we investigated the mechanism of transcutaneous adjuvant activity of the CpG-oligonucleotide (K3) in mice. Transcutaneous immunization (TCI) with an ovalbumin-loaded self-dissolving microneedle patch (OVA-sdMN) and K3-loaded hydrophilic gel patch (HG) increased OVA-specific Th2- and Th1-type IgG subclass antibody titers more rapidly and strongly than those after only OVA-sdMN administration. However, the antigen-specific proliferation of OVA-specific CD4+ T cells was similar between the OVA-only and the OVA+K3 groups. Population analysis of various immune cells in draining lymph nodes (dLNs) in the primary immune response revealed that the OVA+K3 combination doubled the number of dLN cells, with the most significant increase in B cells. Phenotypic analysis by flow cytometry revealed that B-cell activation and maturation were promoted in the OVA+K3 group, suggesting that direct B-cell activation by K3 largely contributed to the rapid increase in antigen-specific antibody titer in TCI. In the secondary immune response, a significant increase in effector T cells and effector memory T cells, and an increase in memory B cells were observed in the OVA+K3 group compared with that in the OVA-only group. Thus, K3, as a transcutaneous adjuvant, can promote the memory differentiation of T and B cells.

Enhanced translocation of amphiphilic peptides across membranes by transmembrane proteins

Cell membranes are phospholipid bilayers with a large number of embedded transmembrane proteins. Some of these proteins, such as scramblases, have properties that facilitate lipid flip-flop from one membrane leaflet to another. Scramblases and similar transmembrane proteins could also affect the translocation of other amphiphilic molecules, including cell-penetrating or antimicrobial peptides. We studied the effect of transmembrane proteins on the translocation of amphiphilic peptides through the membrane. Using two very different models, we consistently demonstrate that transmembrane proteins with a hydrophilic patch enhance the translocation of amphiphilic peptides by stabilizing the peptide in the membrane. Moreover, there is an optimum amphiphilicity because the peptide could become overstabilized in the transmembrane state, in which the peptide-protein dissociation is hampered, limiting the peptide translocation. The presence of scramblases and other proteins with similar properties could be exploited for more efficient transport into cells. The described principles could also be utilized in the design of a drug-delivery system by the addition of a translocation-enhancing peptide that would integrate into the membrane.

Initiated Chemical Vapor Deposition (iCVD) Functionalized Polylactic Acid-Marine Algae Composite Patch for Bone Tissue Engineering.

The current study aimed to describe the fabrication of a composite patch by incorporating marine algae powders (MAPs) into poly-lactic acid (PLA) for bone tissue engineering. The prepared composite patch was functionalized with the co-polymer, poly (2-hydroxyethyl methacrylate-co-ethylene glycol dimethacrylate) (p(HEMA-co-EGDMA)) via initiated chemical vapor deposition (iCVD) to improve its wettability and overall biocompatibility. The iCVD functionalized MAP–PLA composite patch showed superior cell interaction of human osteoblasts. Following the surface functionalization by p(HEMA-co-EGDMA) via the iCVD technique, a highly hydrophilic patch was achieved without tailoring any morphological and structural properties. Moreover, the iCVD modified composite patch exhibited ideal cell adhesion for human osteoblasts, thus making the proposed patch suitable for potential biomedical applications including bone tissue engineering, especially in the fields of dentistry and orthopedy.

Molecular Patchy Clusters with Controllable Symmetry Breaking for Structural Engineering.

Anisotropic patchy particles with molecular precision are exquisite building blocks for constructing diverse meso-structures of high complexity. In this research, a library of molecular patchy clusters consisting of a collection of functional polyhedral oligomeric silsesquioxane cages with exact regio-configuration and composition were prepared through a robust and modular approach. By meticulously tuning the composition, molecular symmetry, and other parameters, these patchy clusters could assemble into diverse nanostructures, including unconventional complex spherical phases (i.e., Frank-Kasper σ phase and dodecagonal quasicrystalline phase). As the size of the hydrophilic patch expands, a transition sequence from disorder to hexagonally packed cylinders and then to double gyroids was recorded, corresponding to a progressive decrease of interfacial curvature. On the other hand, regioisomers with the same composition but different regio-configuration adopt similar molecular packing but varied phase stability, as a result of the local self-sorting process to alleviate excess unfavorable interfacial contact. These precisely defined molecular patchy clusters provide a model system for a general understanding of the hierarchical structure formation and evolution based on anisotropic spherical building blocks at the nanoscale.

Energy Stored in Nanoscale Water Capillary Bridges between Patchy Surfaces.

We perform molecular dynamics (MD) simulations of a water capillary bridge (WCB) expanding between two identical chemically heterogeneous surfaces. The model surfaces, based on the structure of silica, are hydrophobic and are decorated by a hydrophilic (hydroxylated silica) patch that is in contact with the WCB. Our MD simulations results, including the WCB profile and forces induced on the walls, are in agreement with capillarity theory even at the smallest wall separations studied, h = 2.5-3 nm. Remarkably, the energy stored in the WCB can be relatively large, with an energy density that is comparable to that harvested by water-responsive materials used in actuators and nanogenerators.

Transcutaneous immunization with a highly active form of XCL1 as a vaccine adjuvant using a hydrophilic gel patch elicits long-term CD8+ T cell responses

Memory CD8+ cytotoxic T-lymphocytes (CTLs) play a key role in protective immunity against infection and cancer. However, the induction of memory CTLs with currently available vaccines remains difficult. The chemokine receptor XCR1 is predominantly expressed on CD103+ cross-presenting dendritic cells (DCs). Recently, we have demonstrated that a high activity form of murine lymphotactin/XCL1 (mXCL1-V21C/A59C), a ligand of XCR1, can induce antigen-specific memory CTLs by increasing the accumulation of CD103+ DCs in the vaccination site and the regional lymph nodes. Here, we combined a hydrophilic gel patch as a transcutaneous delivery device and mXCL1-V21C/A59C as an adjuvant to further enhance memory CTL responses. The transcutaneous delivery of ovalbumin (OVA) and mXCL1-V21C/A59C by the hydrophilic gel patch increased CD103+ DCs in the vaccination site and the regional lymph nodes for a prolonged period of time compared with the intradermal injection of OVA and mXCL1-V21C/A59C. Furthermore, the hydrophilic gel patch containing OVA and mXCL1-V21C/A59C strongly induced OVA-specific memory CTLs and efficiently inhibited the growth of OVA-expressing tumors more than the intradermal injection of OVA and mXCL1-V21C/A59C. Collectively, this type of hydrophilic gel patch and a high activity form of XCL1 may provide a useful tool for the induction of memory CTL responses.

Characteristic of K3 (CpG-ODN) as a Transcutaneous Vaccine Formulation Adjuvant.

Transcutaneous immunization (TCI) is easy to use, minimally invasive, and has excellent efficacy in vaccines against infections. We focused on toll-like receptor (TLR) ligands as applicable adjuvants for transcutaneous formulations and characterized immune responses. TCI was performed using poke-and-patch methods, in which puncture holes are formed with a polyglycolic acid microneedle on the back skin of mice. Various TLR ligands were applied to the puncture holes and covered with an ovalbumin-loaded hydrophilic gel patch. During the screening process, K3 (CpG-oligonucleotide) successfully produced more antigen-specific antibodies than other TLR ligands and induced T helper (Th) 1-type polarization. Transcutaneously administered K3 was detected in draining lymph nodes and was found to promote B cell activation and differentiation, suggesting a direct transcutaneous adjuvant activity on B cells. Furthermore, a human safety test of K3-loaded self-dissolving microneedles (sdMN) was performed. Although a local skin reaction was observed at the sdMN application site, there was no systemic side reaction. In summary, we report a K3-induced Th1-type immune response that is a promising adjuvant for transcutaneous vaccine formulations using MN and show that K3-loaded sdMN can be safely applied to human skin.

Immunogenicity of Milk Protein-Containing Hydrophilic Gel Patch for Epicutaneous Immunotherapy for Milk Allergy.

Epicutaneous immunotherapy (EPIT) involving the skin's immune system is easy to use, painless and has a low risk of systemic side effects; it can be applied to food allergies that have a high morbidity rate in children. In this study, we evaluated the safety and efficacy of hydrophilic gel patch (HG) for EPIT. Milk protein concentrate (MPC)-containing HG was applied to the skin that maintained a barrier function or formed puncture holes with microneedle, and MPC-specific antibodies were measured. The clinical study was conducted involving patients with severe milk allergy. No specific immune response was induced when immunizing to intact skin, and antibody production was observed by forming puncture holes. It was suggested that MPC contained in HG has immunogenicity and a very small amount of MPC was delivered to intact skin. In the clinical study, the symptom induction threshold increased in four of eight subjects, allowing them to consume milk and switch to oral immunotherapy. Although local skin reactions and temporary elevation of specific IgE antibodies were observed, no systemic side effects appeared throughout the study. EPIT using HG is a safe method to enable oral administration even in patients with severe milk allergies.

Development of an Immune Regulation Technology Targeting the Skin and Promotion of the Practical Applications of Transcutaneous Vaccination/Immunotherapy

In the premise of vaccination and allergen-specific immunotherapy, transcutaneous formulations have an advantage over conventional subcutaneous injections in terms of convenience, simplicity of delivery, and painless administration into the skin. Additionally, since transcutaneous formulations can be rendered cold-chain free, they do not require expert handling during transportation, storage, and stockpiling, which enables reductions in costs and distribution to distant areas. Furthermore, transcutaneous formulations are effective for improving adherence in children with phobias toward injection needles and may help in persuading them to perform self-vaccination and home immunotherapy against allergies in the future. We have been promoting the development of innovative "patch-type formulations for vaccination and immunotherapy" which regard skin as an immune organ and utilize our original transcutaneous administration devices (hydrophilic gel patch and microneedle patch) for their delivery. We have confirmed the safety and efficacy of transcutaneous formulations not only in demonstration experiments using animals but also in physician-initiated clinical studies. Additionally, in order to elucidate the mechanism for the induction of immune responses by transcutaneous formulations, we analyzed the immunological events occurring in the skin and regional lymph nodes which accompanied the application of transcutaneous administration devices or the delivery of antigens (vaccines and allergens) to the skin surface layer. This review presents our results from basic to clinical research on the development of transcutaneous formulations for vaccines and allergen-specific immunotherapy.

Shedding of a condensing droplet from beetle-inspired bumps

Inspired by Namib desert beetle’s bumpy structures, condensing droplet shedding from various super-hydrophobic bumps with a hydrophilic patch present on their top was numerically studied using the lattice Boltzmann method (LBM). The droplet grows on the top patch due to condensation and then sheds from the bump due to gravity. A parametric study has been conducted. It was revealed that the bump shape, diameter, height, inclination and wettability all affect the shedding volume (volume at which the droplet sheds from the bump) of the droplet. The shedding volume decreases with the bump height till a threshold height, defined as the critical bump height, beyond which the shedding volume does not change any more. With the same patch area, the shedding volume is smaller on the bump with a hemispherical top compared to the bump with a circular cylindrical top. The shedding volume increases with bump diameter, but decreases with the patch contact angle and surface inclination angle. The same trends were also found for the critical bump height. Furthermore, compared to milli-scale bumps, droplet removal followed by shedding on micro-scale bumps was found to be inefficient. Based on the simulation results, a scaling law was obtained by data fitting to estimate the critical bump height for bumps with a cylindrical top, which can be used to guide the design of desert-beetle-inspired bumps for efficient water collection from fog.

Sessile Drop on Oscillating Incline

Natural or industrial flows of a fluid often involve droplets or bubbles of another fluid, pinned by physical or chemical impurities or by the roughness of the bounding walls. Here we study numerically one drop pinned on a circular hydrophilic patch, on an oscillating incline whose angle is proportional to sin(ωt). The resulting deformation of the drop is measured by the displacement of its center of mass, which behaves similarly to a driven over-damped linear oscillator with amplitude A(ω) and phase lag φ(ω). The phase lag is O(ω) at small ω like a linear oscillator, but the amplitude is O(ω−1) in a wide range of large ω instead of O(ω−2) for a linear oscillator. A heuristic explanation is given for this behaviour. The simulations were performed with the software Comsol in mode Laminar Two-Phase Flow, Level Set, with fluid 1 as engine oil and fluid 2 as water.

The use of a novel adhesive tissue patch as an aid to anastomotic healing.

Introduction One of the most feared complications of colorectal surgery is anastomotic leak. Numerous techniques have been studied in the hope of decreasing leakage. This study was designed to assess the handling characteristics of a novel adhesive tissue patch (TissuePatch™; Tissuemed, Leeds, UK) applied to colorectal anastomoses in a pilot study. This was with a view to assessing its potential role in aiding anastomotic healing in subsequent trials. Methods A patch was applied to colorectal anastomoses after the surgeon had completed the anastomosis and prior to abdominal closure. Handling characteristics and patient outcomes were recorded prospectively. Results Nine patients were recruited before the study was prematurely terminated. In one patient, the patch fell off and in another patient, the surgeon omitted to apply it. Six patients had significant postoperative problems (1 confirmed leak necessitating return to theatre and excision anastomosis, 3 suspicious of leak on computed tomography delaying discharge, 2 perianastomotic collections). One patient had an uneventful recovery. Conclusions Although the handling characteristics of this novel tissue patch were deemed satisfactory, it appears that wrapping a colorectal anastomosis with an adhesive hydrophilic patch has significant deleterious effects on anastomotic healing. This could be a consequence of the creation of a microenvironment between the patch and the anastomosis that impairs healing. Further research is required to better understand the mechanisms involved. At present, the use of such patches on colorectal anastomoses should be discouraged outside the confines of a well monitored trial.

Electrode-assisted trapping and release of droplets on hydrophilic patches in a hydrophobic microchannel

In two-phase flow microfluidics, there is an increasing interest in technologies which enable the encapsulation of biological cells into aqueous drops and the subsequent study of their molecular (excretion or lysis) products. One not yet available but very promising analysis method is the use of biospecific surface patches embedded in the wall of microfluidic channels. In this paper, we tackle some technological challenges encountered in the development of such applications. In the detection protocol, each drop must be enabled to wet the designated patch, be held in contact long enough for biomolecular detection and subsequently be released. This is engineered via a combination of well-defined chemical sites in the walls of the flow channel and insulated microelectrodes. The tunability of the local electric field allows to modify the competition between chemical (pinning) forces which tend to immobilize the drop and hydrodynamic forces which oppose this process. We developed a prototype microfluidic device which offers this functionality. A channel structure is sandwiched between an actuation surface with electrowetting (EW) electrodes on one side and a detector surface with a hydrophilic patch amidst a hydrophobic environment on the other. Two pairs of carefully aligned EW electrodes are used: one for drop adherence and another one for the subsequent release. We demonstrate these operations and discuss the required voltage signals in terms of the forces on the drop. Finally, we discuss possible steps for further improvement in the device.

Structure of the transmembrane domain of human nicastrin-a component of γ-secretase.

Nicastrin is the largest component of γ-secretase that is an intramembrane protease important in the development of Alzheimer’s disease. Nicastrin contains a large extracellular domain, a single transmembrane (TM) domain, and a short C-terminus. Its TM domain is important for the γ-secretase complex formation. Here we report nuclear magnetic resonance (NMR) studies of the TM and C-terminal regions of human nicastrin in both sodium dodecyl sulfate (SDS) and dodecylphosphocholine (DPC) micelles. Structural study and dynamic analysis reveal that the TM domain is largely helical and stable under both SDS and DPC micelles with its N-terminal region undergoing intermediate time scale motion. The TM helix contains a hydrophilic patch that is important for TM-TM interactions. The short C-terminus is not structured in solution and a region formed by residues V697-A702 interacts with the membrane, suggesting that these residues may play a role in the γ-secretase complex formation. Our study provides structural insight into the function of the nicastrin TM domain and the C-terminus in γ-secretase complex.

Solution structure of the transmembrane domain of the mouse erythropoietin receptor in detergent micelles.

Erythropoiesis is regulated by the erythropoietin receptor (EpoR) binding to its ligand. The transmembrane domain (TMD) and the juxtamembrane (JM) regions of the EpoR are important for signal transduction across the cell membrane. We report a solution NMR study of the mouse erythropoietin receptor (mEpoR) comprising the TMD and the JM regions reconstituted in dodecylphosphocholine (DPC) micelles. The TMD and the C-terminal JM region of the mEpoR are mainly α-helical, adopting a similar structure to those of the human EpoR. Residues from S216 to T219 in mEpoR form a short helix. Relaxation study demonstrates that the TMD of the mEpoR is rigid whilst the N-terminal region preceding the TMD is flexible. Fluorescence spectroscopy and sequence analysis indicate that the C-terminal JM region is exposed to the solvent. Helix wheel result shows that there is hydrophilic patch in the TMD of the mEpoR formed by residues S231, S238 and T242, and these residues might be important for the receptor dimerization.