Single-cell transcriptome profiling reveals cell type-specific variation and development in HLA expression of human skin.

Skin, the largest organ of body, is a highly immunogenic tissue with a diverse collection of immune cells. Highly polymorphic human leukocyte antigen (HLA) molecules have a central role in coordinating immune responses as recognition molecules. Nevertheless, HLA gene expression patterns among diverse cell types within a specific organ, like the skin, have yet to be thoroughly investigated, with stromal cells attracting much less attention than immune cells. To illustrate HLA expression profiles across different cell types in the skin, we performed single-cell RNA sequencing (scRNA-seq) analyses on skin datasets, covering adult and fetal skin, and hair follicles as the skin appendages. We revealed the variation in HLA expression between different skin populations by examining normal adult skin datasets. Moreover, we evaluated the potential immunogenicity of multiple skin populations based on the expression of classical HLA class I genes, which were well represented in all cell types. Furthermore, we generated scRNA-seq data of developing skin from fetuses of 15 post conception weeks (PCW), 17 PCW, and 22 PCW, delineating the dynamic expression of HLA genes with cell type-dependent variation among various cell types during development. Notably, the pseudotime trajectory analysis unraveled the significant variance in HLA genes during the evolution of vascular endothelial cells. Moreover, we uncovered the immune-privileged properties of hair follicles at single-cell resolution. Our study presents a comprehensive single-cell transcriptomic landscape of HLA genes in the skin, which provides new insights into variation in HLA molecules and offers a clue for allogeneic skin transplantation.

Enhancing Mechanical Performance of High-Lignin-Filled Polypropylene via Reactive Extrusion.

Polypropylene (PP) is one of the most extensively used commodity plastics. In terms of eco-friendliness, it is worth considering preparing high-lignin-filled PP. This study explores the incorporation of high lignin content, derived from acetic acid lignin (AAL) and Kraft lignin (KL), into PP through twin-screw extrusion and injection molding. The challenge lies in maintaining mechanical performance. A compatibilizer-specifically, maleic anhydride-grafted polypropylene (MAPP)-is employed to enhance lignin-PP compatibility by chemically bonding with lignin and physically associating with the PP phase. Results indicate that KL maintains better dispersity than AAL. Compatibilizers with a high maleic anhydride (MA) level (≥0.8 wt.%) and moderate melt flow index (MFI) in the range of 60-100 g 10 min⁻¹ prove favorable in constructing a reinforced PP/KL network. Optimizing with 40 wt.% lignin content and 10 parts per hundred (pph) of compatibilizer yields blends with mechanical performance comparable to neat PP, exhibiting a notable increase in modulus and heat deflection temperature (HDT). Furthermore, utilizing PP/lignin blends can lead to a 20% reduction in expenses and approximately 40% reduction in PP-induced greenhouse gas (GHG) emissions. This approach not only reduces PP costs but also adds value to lignin utilization in a sustainable and cost-effective manner.

Machine learning method for predicting cadmium concentrations in rice near an active copper smelter based on chemical mass balance.

Identification the sources of heavy metals can effectively control and prevent agricultural soil pollution. Here we performed a three-year mass balance study along a gradient of soil pollution near a smelter to quantify the potential contribution and net cadmium (Cd) fluxes and predict Cd concentration in rice grains by multiple regression (MR) and back propagation (BP) neural network. The Cd inputs were mainly from the irrigation water (54.6-60.8%) in the moderately polluted and background sites but from atmospheric deposition (90.9%) in the highly polluted site. The Cd outputs were mainly from the surface runoff (55.8-59.5%) in the moderately polluted and background sites, but from Sedum plumbizincicola phytoextraction (83.6%) in the highly polluted site. The soil Cd concentrations, the annual fluxes of atmospheric deposition, pesticides and fertilizers, irrigation water, surface runoff, and leaching water were selected as the dependent factors to predict Cd concentrations in rice grains. The genetic algorithms (GA)-BP neural network model gives the best prediction accuracy compared to the BP neural network model and multivariate regression analysis. The major implication is that the health risks through the consumption of rice can be rapidly assessed based on the Cd concentrations in rice grains predicted by the model.

Platelet sonicates activate hair follicle stem cells and mediate enhanced hair follicle regeneration.

An increasing number of studies show that platelet-rich plasma (PRP) is effective for androgenic alopecia (AGA). However, the underlying cellular and molecular mechanisms along with its effect on hair follicle stem cells are poorly understood. In this study, we designed to induce platelets in PRP to release factors by calcium chloride (PC) or by sonication where platelet lysates (PS) or the supernatants of platelet lysate (PSS) were used to evaluate their effect on the hair follicle activation and regeneration. We found that PSS and PS exhibited a superior effect in activating telogen hair follicles than PC. In addition, PSS injection into the skin activated quiescent hair follicles and induced K15+ hair follicle stem cell proliferation in K14-H2B-GFP mice. Moreover, PSS promoted skin-derived precursor (SKP) survival in vitro and enhanced hair follicle formation in vivo. In consistence, protein array analysis of different PRP preparations revealed that PSS contained higher levels of 16 growth factors (out of 41 factors analysed) than PC, many of them have been known to promote hair follicle regeneration. Thus, our data indicate that sonicated PRP promotes hair follicle stem cell activation and de novo hair follicle regeneration.

Efficient lung cancer-targeted drug delivery via a nanoparticle/MSC system.

Low targeting efficiency limits the applications of nanoparticles in cancer therapy. The fact that mesenchymal stem cells (MSC) trapped in the lung after systemic infusion is a disadvantage for cell therapy purposes. Here, we utilized MSC as lung cancer-targeted drug delivery vehicles by loading nanoparticles (NP) with anti-cancer drug. MSC showed a higher drug intake capacity than fibroblasts. In addition, MSC showed predominant lung trapping in both rabbit and monkey. IR-780 dye, a fluorescent probe used to represent docetaxel (DTX) in NP, delivered via MSC accumulated in the lung. Both in vitro MSC/A549 cell experiments and in vivo MSC/lung cancer experiments validated the intercellular transportation of NP between MSC and cancer cells. In vivo assays showed that the MSC/NP/DTX drug delivery system exerted primary tumor inhibition efficiency similar to that of a NP/DTX drug system. Collectively, the MSC/NP drug delivery system is promising for lung-targeted drug delivery for the treatment of lung cancer and other lung-related diseases.

Immunochromatographic strip biosensor for the rapid detection of N-glycolylneuraminic acid based on aptamer-conjugated nanoparticle.

N-glycolylneuraminic acid (Neu5Gc) is a type of sialic acid that is not typically produced in healthy humans but detective in some visceral cancer cells. As a new carcinoma biomarker, the level change in the serum and urine from the patient could potentially have the relation to the disease progression. So the measurement of the Neu5Gc will help to take a better response to therapeutic schedule for the sufferers. A sensitive and rapid aptamer-nanoparticle immunochromatographic strip for the visual detection of Neu5Gc was developed. The assay is based on the competitive reaction of binding the DNA aptamer targeting the candidate molecule selected by SELEX between Neu5Gc and complementary DNA. The sensing results indicated that the aptamer-based strip was sufficiently sensitive to detect Neu5Gc. The visual limit of detection (LOD) for semi-quantitative detection was 30 ng/mL under the optimal conditions and a quantitative detection limit of 5.38 ng/mL could be obtained using a scanning strip reader. The average recovery of the spiked cancer cell samples was 88.86%, with a coefficient of variation (CV) of 5.27%. The detection could be performed in less than 15 min using a simple procedure without any complicated equipment, demonstrating that this aptamer-nanoparticle biosensor strip has great potential for use to Neu5Gc-related cancer diagnosis.

miRNA-21 enhances chemoresistance to cisplatin in epithelial ovarian cancer by negatively regulating PTEN.

MicroRNAs (miRNAs) are small non-coding RNAs, 8-23 nucleotides in length, which regulate gene expression at the post-transcriptional level. The present study was performed to analyze the association between microRNA-21 and cisplatin resistance in epithelial ovarian cancer (EOC) SKOV3 and SKOV3/DDP cells. In this experiment, the resistance of SKOV3 and SKOV3/DDP cells to cisplatin was evaluated using the MTT assay. Reverse transcription-quantitative polymerase chain reaction analysis was used to assess miRNA-21 levels and phosphatase and tensin homolog (PTEN) mRNA levels. Western blotting was used to assess PTEN protein levels. miRNA-21 mimics or inhibitors were transfected into SKOV3 and SKOV3/DDP cells. Prior to transfection, higher expression levels of miRNA-21 were observed in SKOV3/DDP cells compared with SKOV3 cells. Following transfection with miRNA-21 mimics, SKOV3 cells demonstrated increased sensitivity to cisplatin compared with negative control cells. Following transfection with the miRNA-21 inhibitor, SKOV3/DDP cells demonstrated decreased sensitivity to cisplatin compared with negative control cells. Furthermore, PTEN mRNA expression levels in SKOV3 cells transfected with miRNA-21 mimics was significantly lower compared with negative control cells. These results suggested that miRNA-21 may regulate cisplatin resistance by negatively targeting PTEN in EOC.

Macrophages induce AKT/ß-catenin-dependent Lgr5+ stem cell activation and hair follicle regeneration through TNF.

Skin stem cells can regenerate epidermal appendages; however, hair follicles (HF) lost as a result of injury are barely regenerated. Here we show that macrophages in wounds activate HF stem cells, leading to telogen-anagen transition (TAT) around the wound and de novo HF regeneration, mostly through TNF signalling. Both TNF knockout and overexpression attenuate HF neogenesis in wounds, suggesting dose-dependent induction of HF neogenesis by TNF, which is consistent with TNF-induced AKT signalling in epidermal stem cells in vitro. TNF-induced ß-catenin accumulation is dependent on AKT but not Wnt signalling. Inhibition of PI3K/AKT blocks depilation-induced HF TAT. Notably, Pten loss in Lgr5+ HF stem cells results in HF TAT independent of injury and promotes HF neogenesis after wounding. Thus, our results suggest that macrophage-TNF-induced AKT/ß-catenin signalling in Lgr5+ HF stem cells has a crucial role in promoting HF cycling and neogenesis after wounding.

Preparation of a Specific ssDNA Aptamer for Brevetoxin-2 Using SELEX.

The existing assays for detecting brevetoxin (BTX) depend on expensive equipment with a professional operator or on an antibody with limited stability, which requires complex processes, a high cost, and a considerable amount of time. The development of an alternative detection probe is another promising research direction. This paper reports the use of aptamers binding to BTX-2 in an analytical assay using the systematic evolution of ligands by exponential enrichment (SELEX). After 12 rounds of selection, the secondary structures of 25 sequences were predicted. Compared to other aptamers, Bap5 has relatively high affinity with the lowest dissociation constant of 4.83 µM, and IC50 is 73.81 ng mL-1. A good linear regression formula of y = 30.688x - 7.329 with a coefficient correlation of R2 = 0.9798 was obtained using a biotin-avidin ELISA. Moreover, there is no cross-reaction with the detected marine toxins, except for BTX-2. Thus, Bap5 has potential to detect BTX-2 in shellfish in the future as a substitute for the recognition probe.

Excess Integrins Cause Lung Entrapment of Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) are largely entrapped in the lungs after intravenous delivery. The underlying mechanisms have been poorly understood. Flow cytometry and Western blot analysis showed that the expression levels of many integrins such as ß1, α5, and αVß3 in MSCs increased markedly upon cultured expansion in 2D monolayers, whose ligands fibronectin and vitronectin were detected on the surface of vascular endothelial cells in the lungs by immunostaining and flow cytometry. Blockade of integrin ß1, integrin α5, or integrins αVß3 with functional blocking antibodies significantly decreased the amount of MSCs entrapped in the lungs following intravenous infusion as determined by real-time PCR and histological analysis; meanwhile, corresponding increases in the levels of circulating MSCs in the blood and MSCs homed to the ischemic myocardium and inflamed ear were found. Intriguingly, a short period of 3D spheroid culture of MSCs, which had been expanded for several passages in monolayers, substantially reduced the expression levels of many integrins and the number of MSCs entrapped in the lungs. Our results indicate that the excess expression and activation of integrins is a significant cause of lung entrapment of MSCs.

A novel analytical probe binding to a potential carcinogenic factor of N-glycolylneuraminic acid by SELEX.

N-glycolylneuraminic acid (Neu5Gc) is an abundant sialic acid in many mammals and is present in the glycoconjugates of most deuterostome animals. Neu5Gc also occurs in fresh samples of human tumors and fetuses. However, very little is known about the expression level and biologic functions of Neu5Gc due to the limitations of available analytical probes for detection methods. In this study, we first report the development of aptamers specific to Neu5Gc screened by Systematic Evolution of Ligands by Exponential Enrichment (SELEX). After 15 selection rounds, cloning, sequencing and enzyme-linked immuno sorbent assay (ELISA) analysis, 6 different selected aptamers showed specificity for Neu5Gc. Among these 6 aptamers, N8 showed the best half maximal inhibitory concentration (IC50) value (127 ng mL(-1)) and had a relatively high affinity constant (Ka=6.68 × 10(9)M(-1)). The aptamers selected in this study will provide a novel analytical probe for the development of a biosensor to detect Neu5Gc in tissues and sera from patients with tumors as well as to detect Neu5Gc in animal-derived foods. In addition, the successful aptamer candidate can solve the problem that antibody is difficult to prepare in immunological assays. Thus, the discovery of novel aptamers specific for Neu5Gc is important for developing new methods of detecting Neu5Gc for the diagnosis and prevention of cancer as well as food poisoning.

Selection and identification of a DNA aptamer that mimics saxitoxin in antibody binding.

In this article, high-affinity single-stranded DNA (ssDNA) aptamer-targeting F(ab')2 fragments of saxitoxin (STX) antibodies were selected from a random ssDNA library by the SELEX strategy. After 16 rounds of repeated selection, the enriched ssDNA library was sequenced, and all of the sequences were carefully identified by indirect enzyme-linked assay and indirect competitive enzyme-linked assay (icELISA). The candidate aptamers in the above identification were selected for further characterization by icELISA and the equilibrium filtration method. We successfully obtained an aptamer that mimics STX in antibody binding, and a substitute for STX in aptamer form has been developed. Further work is in progress aimed at using this aptamer substitute to replace the STX standard in an antibody-based, nontoxic detection method for field determination of STX in seafood products.