Tissue Surface Research Articles

Highly Sensitive Spatial Glycomics at Near-Cellular Resolution by On-Slide Derivatization and Mass Spectrometry Imaging.

Glycans on proteins and lipids play important roles in maturation and cellular interactions, contributing to a variety of biological processes. Aberrant glycosylation has been associated with various human diseases including cancer; however, elucidating the distribution and heterogeneity of glycans in complex tissue samples remains a major challenge. Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) is routinely used to analyze the spatial distribution of a variety of molecules including N-glycans directly from tissue surfaces. Sialic acids are nine carbon acidic sugars that often exist as the terminal sugars of glycans and are inherently difficult to analyze using MALDI-MSI due to their instability prone to in- and postsource decay. Here, we report on a rapid and robust method for stabilizing sialic acid on N-glycans in FFPE tissue sections. The established method derivatizes and identifies the spatial distribution of α2,3- and α2,6-linked sialic acids through complete methylamidation using methylamine and PyAOP ((7-azabenzotriazol-1-yloxy)tripyrrolidinophosphonium hexafluorophosphate). Our in situ approach increases the glycans detected and enhances the coverage of sialylated species. Using this streamlined, sensitive, and robust workflow, we rapidly characterize and spatially localize N-glycans in human tumor tissue sections. Additionally, we demonstrate this method's applicability in imaging mammalian cell suspensions directly on slides, achieving cellular resolution with minimal sample processing and cell numbers. This workflow reveals the cellular locations of distinct N-glycan species, shedding light on the biological and clinical significance of these biomolecules in human diseases.

Three-dimensional changes in the soft tissue chin after mandibular setback surgery in adult Japanese patients with skeletal class III malocclusion

ABSTRACT Purpose The purpose of the present study is to improve the quality of prediction of soft tissue morphology by clarifying the relationship between the amount of three-dimensional (3-D) changes of hard tissue and the morphological differences of the soft tissue chin before and after orthognathic surgery in skeletal class III malocclusion. Materials and methods Preoperative and six-month postoperative CT was used to analyze the changes in 26 adult Japanese skeletal class III patients. The mean age was 25.5 years. Lateral cephalograms were analyzed to evaluate the initial antero-posterior jaw relationship and the changes before and after treatment. Three-D masks and bone structures were reconstructed using 3-D modelling software, and the final chin models were defined for further analysis following voxel-based registration. Correlation and linear regression analyses were performed to clarify the relationship between the movement of the soft tissue chin and the underlying bone. Results The soft tissue chin volume and surface area significantly increased after surgery. The thickness of the chin significantly increased only at pogonion and menton after surgery. Furthermore, a statistically significant correlation between the movement of soft tissue chin and underlying bone was found. Consequently, the regression model predicted the changes in soft tissue chin from the movement of the underlying bone. Conclusion Three-D analysis of the soft tissue chin revealed a significant change in response to surgical orthodontic treatment in skeletal class III adult patients, and soft tissue chin movement could be predicted by using 3-D CT data when planning orthognathic treatment.

Unusual Extension of Superficial Spreading Squamous Cell Carcinoma of Cervix to Uterus, Paraovarian, Paratubal Tissue, and Vaginal Surface: A Unique Case Report

ABSTRACT Superficial spreading cervical squamous cell carcinoma (SCC) is a rare phenomenon with few cases reported in the literature. The present case report briefs the findings of superficial spreading cervical SCC in postmenopausal women. A 65-year-old postmenopausal woman presented with bleeding per-vaginum for 1–2 months. On examination, the cervix was healthy with no visible growth. She was diagnosed with an endometrial malignancy on magnetic resonance imaging. On endometrial and cervical biopsy, histopathology revealed cervical intraepithelial neoplasia III in ectocervix with SCC in endocervical and endometrial curetting. The patient underwent modified radical hysterectomy with bilateral pelvic lymphadenectomy and histopathology revealed superficially spreading cervical SCC with tumor cells involving endometrium, myometrium, paratubal, paraovarian, vaginal margins without extending to pelvic, and para-aortic lymph nodes. The patient was further managed successfully with adjuvant chemoradiotherapy. Superficial spreading cervical SCC is very rare phenomenon. Its spread is not only limited to endometrium but can extend to involve myometrium, paratubal, paraovarian, and vaginal tissues.

Chondroprotection of articular cartilage integrity: Utilizing ultrasonic scalpel and hyperosmolar irrigation solution during cutting

ObjectivesUltrasonic (US) cutting of cartilage in orthopaedic surgery has received little attention despite its potential to reduce chondrocyte death which could enhance cartilage repair. We aimed to investigate whether an ultrasonically-vibrating scalpel to cut human articular cartilage could reduce chondrocyte death, and to determine if hyper-osmolarity could provide chondroprotection during the procedure. MethodsA scalpel (no. 15) was mounted on an ultrasonic transducer to resonate at 35 ​kHz with 30 ​μm vibrational displacement. Thirty-six fresh human femoral cartilage samples were divided into four groups based on ultrasonic activation (US or non-US) and saline osmolarity (300 or 600 mOsm/L). Cell viability was assessed using a live/dead cell assay and analysed quantitatively by confocal microscopy. Histology illustrated tissue surface changes at the cut site. ResultsThe overall chondrocyte death percentage at both the US and non-US cut sites showed comparable results (p ​> ​0.05) in both osmolarities. However, the zone of chondrocyte death was reduced by 31 ​± ​5% and 36 ​± ​6%, respectively, when comparing US cutting at 300 mOsm/L and 600 mOsm/L to the control group (non-US cutting; 300 mOsm/L) (p ​< ​0.05). The width of the cut was consistent at both sites, regardless of the method of cutting. ConclusionCutting human cartilage with US in the presence of 300 or 600 mOsm/L media was chondroprotective compared to normal (non-US) scalpel cutting in 300 mOsm/L medium. These results suggest chondroprotection can be achieved while cutting using a US scalpel and raised osmolarity, potentially improving cartilage regeneration and repair following injury.

Fresh Meat Classification Using Laser-Induced Breakdown Spectroscopy Assisted by LightGBM and Optuna.

To enhance the accuracy of identifying fresh meat varieties using laser-induced breakdown spectroscopy (LIBS), we utilized the LightGBM model in combination with the Optuna algorithm. The procedure involved flattening fresh meat slices with glass slides and collecting spectral data of the plasma from the surfaces of the fresh meat tissues (pork, beef, and chicken) using LIBS technology. A total of 900 spectra were collected. Initially, we established LightGBM and SVM (support vector machine) models for the collected spectra. Subsequently, we applied information gain and peak extraction algorithms to select the features for each model. We then employed Optuna to optimize the hyperparameters of the LightGBM model, while a 10-fold cross-validation was conducted to determine the optimal parameters for SVM. Ultimately, the LightGBM model achieved higher accuracy, macro-F1, and Cohen's kappa coefficient (kappa coefficient) values of 0.9370, 0.9364, and 0.9244, respectively, compared to the SVM model's values of 0.8888, 0.8881, and 0.8666. This study provides a novel method for the rapid classification of fresh meat varieties using LIBS.

Metabolomics research on treatment of primary liver cancer with Cortex Juglandis Mandshuricae on LC-MS/MS technology

Diethylnitrosamine (DEN) was applied to create the primary liver cancer (PLC) animal model. In the study, the normal group, model group, cyclophosphamide (CTX) group, Cortex Juglandis Mandshuricae (CJM) extract group, myricetin group and myricitrin group were divided. LC-MS/MS technology was applied to determine the metabolites of liver tissue samples from different locations (nodular and non-nodular parts of liver tissue) in each group of rats. Through metabolomics research, the connection and difference of anti-PLC induced by the CJM extract, myricetin and myricitrin was analyzed. The surface of the liver tissues of rats in the model group was rough, dimly colored, inelastic, on which there were scattered gray white cancer nodules and blood stasis points. The number of cancer nodules was significantly reduced, and the degree of cell malignancy was low, but there were some inflammatory cell infiltrations, necrosis area and karyokinesis in the CJM extract group, myricetin group, myricitrin group and CTX group. The result of metabolic research indicated that 45 potential biomarkers of the PLC were found, as gamma-aminoisobutyrate, taurochenodeoxycholate, xanthurenic acid, etc. There were 22 differential metabolites in the CTX group, 16 differential metabolites in the CJM extract group, 14 differential metabolites in the myricetin group, 14 differential metabolites in the myricitrin group.

Application of lactobacillus casei and lactobacillus plantarum to develop dried functional apple and banana.

The goal of this research was to create dried fruits loaded with probiotic microorganisms (Lactobacillus casei and Lactobacillus plantarum). In separate bottles for each probiotic microbe, apple and banana pieces have been submerged into the impermeability solution with gentle shaking. The vacuum pressure was applied. By the conclusion of the incubation time, L. casei and L. plantarum colonies were enumerated (CFU/g). The scanning electron microscope method was applied to confirm the penetration of impregnation solutions into the intercellular spaces of fruit tissue. On day 28, the population of L. plantarum was 5 log CFU/g for apples and 5.5 log CFU/g for bananas. After storage, the number of L. casei in apples was 5 log CFU/g and 5.5 log CFU/g, respectively. L. casei was found on the surface of apple and banana tissue. After one-week, whole phenolic content of probiotic-enriched bananas and apples augmented. After storage, the antioxidant activity of all samples decreased greatly. The sensory qualities of the samples were excellent throughout storage in terms of color, quality, scent, sensitivity, chewiness, and general adequacy. As a result, dried apples and bananas infused with L. plantarum and L. casei might be a novel probiotic meal. RESEARCH HIGHLIGHTS: Dried apples and bananas infused with L. plantarum and L. casei are novel probiotic meal. After one-week, whole phenolic content of probiotic-enriched bananas and apples augmented. The sensory qualities of the samples were excellent throughout storage in terms of color, quality, scent, sensitivity, chewiness, and general adequacy.

Formulation and dermal delivery of a new active pharmaceutical ingredient in an in vitro wound model for the treatment of chronic ulcers

The aim of this study was to investigate dermal delivery of the new active pharmaceutical ingredient (API) TOP-N53 into diabetic foot ulcer using an in vitro wound model consisting of pig ear dermis and elucidate the impact of drug formulation and wound dressing taking into consideration clinical relevance in the home care setting and possible bacterial infection. Different formulation approaches for the poorly water-soluble API including colloidal solubilization, drug micro-suspension and cosolvent addition were investigated; moreover, the effect of (micro-)viscosity of hydrogels used as primary wound dressing on delivery was assessed. Addition of Transcutol® P as cosolvent to water improved solubility and was significantly superior to all other approaches providing a sustained three-day delivery that reached therapeutic drug levels in the tissue. Solubilization in micelles or liposomes, on the contrary, did not boost delivery while micro-suspensions exhibited sedimentation on the tissue surface. Microbial contamination was responsible for considerable metabolism of the drug leading to tissue penetration of metabolites which may be relevant for therapeutic effect. Use of hydrogels under semi-occlusive conditions significantly reduced drug delivery in a viscosity-dependent fashion. Micro-rheologic analysis of the gels using diffusive wave spectroscopy confirmed the restricted diffusion of drug particles in the gel lattice which correlated with the obtained tissue delivery results. Hence, the advantages of hydrogel dressings from the applicatory characteristic point of view must be weighed against their adverse effect on drug delivery. The employed in vitro wound model was useful for the assessment of drug delivery and the development of a drug therapy concept for chronic diabetic foot ulcer. Mechanistic insights about formulation and dressing performance may be applied to drug delivery in other skin conditions such as digital ulcer.

Self-assembled multicolor fluorescent nanoparticles derived from dopamine analogues: A versatile platform for biomedical applications

Fluorescent polydopamine (PDA) nanoparticles (FPNPs) have gained recognition as invaluable resources for a wide range of biomedical applications. Typically, these FPNPs are synthesized solely from dopamine (DA). In this study, our focus lies in the design of PDA-based multicolor fluorescent nanoparticles (m-FNPs) utilizing DA and its analogues, including levodopa (LVD), norepinephrine (NPP), 6-hydroxydopamine (HDA), and epinephrine (EPP) monomers, thereby paving the way for the next generation of FPNPs. The m-FNPs, respectively labeled as L-FNPs, N-FNPs, H-FNPs, E-FNPs, and D-FNPs, exhibit distinct band gaps influenced by the monomer’s structure, allowing versatile multicolor fluorescence bioimaging across a broad fluorescence emission spectrum ranging from 410 to 680 nm. Significantly, the catechol groups present on the surfaces of m-FNPs enable chelation with various theranostic metal ions and subsequent release under endo/lysosome pH conditions in cancer cells, thereby eliciting a cancer-specific “OFF-ON” fluorescence signal in vivo. Furthermore, integrating m-FNPs into poly(dimethylsiloxane) (PDMS) produces a tissue-adhesive fluorescence sheet that permits fluorescence monitoring of internal tissue surfaces within living organisms for 15 days. These m-FNPs-incorporated PDMS (m-FNPs-PDMS) sheets, enriched with catechol, carboxyl, and amine groups on their surfaces, exhibit significantly enhanced adhesion to biological tissues compared to conventional PDMS sheets containing PDA. Our findings suggest that m-FNPs pave the way for advancing PDA-based fluorescent nanoparticles with diverse applications in the biomedical field, supported by comprehensive in vitro and in vivo evaluations.

Clonal dynamics of surface-driven growing tissues.

The self-organization of cells into complex tissues relies on a tight coordination of cell behavior. Identifying the cellular processes driving tissue growth is key to understanding the emergence of tissue forms and devising targeted therapies for aberrant growth, such as in cancer. Inferring the mode of tissue growth, whether it is driven by cells on the surface or by cells in the bulk, is possible in cell culture experiments but difficult in most tissues in living organisms (in vivo). Genetic tracing experiments, where a subset of cells is labeled with inheritable markers, have become important experimental tools to study cell fate in vivo. Here we show that the mode of tissue growth is reflected in the size distribution of the progeny of marked cells. To this end, we derive the clone size distributions using analytical calculations in the limit of negligible cell migration and cell death, and we test our predictions with an agent-based stochastic sampling technique. We show that for surface-driven growth the clone size distribution takes a characteristic power-law form with an exponent determined by fluctuations of the tissue surface. Our results propose a possible way of determining the mode of tissue growth from genetic tracing experiments.

Harnessing bi-layered supramolecular Janus tissue-adhesive/anti-adhesive fibrous hydrogel for efficient hemostasis, wound healing, and suppressing postoperative tissue adhesion

Inadequate progression of primary (bleeding/infection) and secondary (rebleeding/postoperative tissue adhesion) complications at the wound site pose a substantial burden of morbidity and mortality worldwide. Therefore, developing a multifunctional biomaterial dressing for abrupt tissue adhesion and suppressed postoperative adhesion simultaneously is deemed essential. Herein, we designed a bi-layered Janus fibrous hydrogel (J-FH). The bottom layer comprises zinc-incorporated mesoporous silica nanogranules (ZnMSG) reinforced catechol-modified gelatin methacrylate (GC) hydrogel. GC@ZnMSG supramolecular hydrogel easily adheres to wet wounded tissue and possesses hemostatic, mechanical, and antibacterial properties. On the other hand, the top layer comprises dopamine-modified cellulose acetate nanofibrous membrane (DC) sequentially deposited with polydopamine-conjugated zwitterionic sulfobetaine methacrylate (PDA-SBMA). DC@PDA-SBMA fibrous membrane performs the anti-adhesive function by suppressing fouling and preventing tissue adhesion. The findings of animal experiments show that the bottom layer of the J-FH adheres firmly to the tissue surface, preventing bleeding and supporting tissue closure/regeneration. In contrast, the top layer provides mechanical stability, preventing secondary bleeding and suppressing postoperative tissue adhesion. Overall, J-FH demonstrated excellent biocompatibility, antibacterial properties, superior mechanical strength, asymmetric adhesion, robust hemostasis, and accelerated wound healing. It can replace traditional sutures, non-degradable dressings, and double-sided adhesive scaffolds for hemostasis, external, and internal postoperative wound healing.

Hydroxypropyl methylcellulose reinforced bilayer hydrogel dressings containing L-arginine-modified polyoxometalate nanoclusters to promote healing of chronic diabetic wounds

Diabetes-related slow healing of wounds is primarily driven by bacterial infections and angiogenesis disorder and presents a substantial hurdle in clinical treatment. To solve the above problems, an advanced multifunctional hydrogel system based on natural polymer was created here to facilitate wound healing in patients with chronic diabetes. The prepared dressing was composed of an outer hydrogel containing polyvinyl alcohol and hydroxypropyl methyl cellulose in dimethyl sulfoxide and water as binary solvents, and an inner hydrogel containing chitosan quaternary ammonium salt, flaxseed gum, and polyvinyl alcohol. Thus, a polysaccharide based bilayer hydrogel (BH) with superior mechanical strength and biocompatibility was created. This bilayer hydrogel could easily bind to dynamic tissue surfaces, thereby generating a protective barrier. Meanwhile, L-arginine-modified polyoxometalate (POM@L-Arg) nanoclusters were loaded in the inner hydrogel. They released NO when stimulated by the peroxide microenvironment of diabetic wounds. NO as a signal molecule regulated vascular tension and promoted cell proliferation and migration. Additionally, because of the synergistic effect of NO and the chitosan quaternary ammonium salt, the hydrogel system exhibited excellent antibacterial performance. The NO released reduced the levels of proinflammatory factors IL-6 and TNF-α in the diabetic wounds, which thus accelerated wound healing. In short, BH + POM@L-Arg is expected to serve as an ideal wound dressing as it exerts a good promotion effect on diabetes-related wound healing.

Line-field dynamic optical coherence tomography platform for volumetric assessment of biological tissues.

Dynamic optical coherence tomography (dOCT) utilizes time-dependent signal intensity fluctuations to enhance contrast in OCT images and indirectly probe physiological processes in cells. Majority of the dOCT studies published so far are based on acquisition of 2D images (B-scans or C-scans) by utilizing point-scanning Fourier domain (spectral or swept-source) OCT or full-field OCT respectively, primarily due to limitations in the image acquisition rate. Here we introduce a novel, high-speed spectral domain line-field dOCT (SD-LF-dOCT) system and image acquisition protocols designed for fast, volumetric dOCT imaging of biological tissues. The imaging probe is based on an exchangeable afocal lens pair that enables selection of combinations of transverse resolution (from 1.1 µm to 6.4 µm) and FOV (from 250 × 250 µm2 to 1.4 × 1.4 mm2), suitable for different biomedical applications. The system offers axial resolution of ∼ 1.9 µm in biological tissue, assuming an average refractive index of 1.38. Maximum sensitivity of 90.5 dB is achieved for 3.5 mW optical imaging power at the tissue surface and maximum camera acquisition rate of 2,000 fps. Volumetric dOCT images acquired with the SD-LF-dOCT system from plant tissue (cucumber), animal tissue (mouse liver) and human prostate carcinoma spheroids allow for volumetric visualization of the tissues' cellular and sub-cellular structures and assessment of cellular motility.

Molecular Basis for Surface-Initiated Non-Thrombin-Generated Clot Formation Following Viral Infection.

In this paper, we propose a model that connects two standard inflammatory responses to viral infection, namely, elevation of fibrinogen and the lipid drop shower, to the initiation of non-thrombin-generated clot formation. In order to understand the molecular basis for the formation of non-thrombin-generated clots following viral infection, human epithelial and Madin-Darby Canine Kidney (MDCK, epithelial) cells were infected with H1N1, OC43, and adenovirus, and conditioned media was collected, which was later used to treat human umbilical vein endothelial cells and human lung microvascular endothelial cells. After direct infection or after exposure to conditioned media from infected cells, tissue surfaces of both epithelial and endothelial cells, exposed to 8 mg/mL fibrinogen, were observed to initiate fibrillogenesis in the absence of thrombin. No fibers were observed after direct viral exposure of the endothelium or when the epithelium cells were exposed to SARS-CoV-2 isolated spike proteins. Heating the conditioned media to 60 °C had no effect on fibrillogenesis, indicating that the effect was not enzymatic but rather associated with relatively thermally stable inflammatory factors released soon after viral infection. Spontaneous fibrillogenesis had previously been reported and interpreted as being due to the release of the alpha C domains due to strong interactions of the interior of the fibrinogen molecule in contact with hydrophobic material surfaces rather than cleavage of the fibrinopeptides. Contact angle goniometry and immunohistochemistry were used to demonstrate that the lipids produced within the epithelium and released in the conditioned media, probably after the death of infected epithelial cells, formed a hydrophobic residue responsible for fibrillogenesis. Hence, the standard inflammatory response constitutes the ideal conditions for surface-initiated clot formation.

Nuclear position and local acetyl-CoA production regulate chromatin state

Histone acetylation regulates gene expression, cell function and cell fate1. Here we study the pattern of histone acetylation in the epithelial tissue of the Drosophila wing disc. H3K18ac, H4K8ac and total lysine acetylation are increased in the outer rim of the disc. This acetylation pattern is controlled by nuclear position, whereby nuclei continuously move from apical to basal locations within the epithelium and exhibit high levels of H3K18ac when they are in proximity to the tissue surface. These surface nuclei have increased levels of acetyl-CoA synthase, which generates the acetyl-CoA for histone acetylation. The carbon source for histone acetylation in the rim is fatty acid β-oxidation, which is also increased in the rim. Inhibition of fatty acid β-oxidation causes H3K18ac levels to decrease in the genomic proximity of genes involved in disc development. In summary, there is a physical mark of the outer rim of the wing and other imaginal epithelia in Drosophila that affects gene expression.

Three-Dimension Epithelial Segmentation in Optical Coherence Tomography of the Oral Cavity Using Deep Learning.

This paper aims to simplify the application of optical coherence tomography (OCT) for the examination of subsurface morphology in the oral cavity and reduce barriers towards the adoption of OCT as a biopsy guidance device. The aim of this work was to develop automated software tools for the simplified analysis of the large volume of data collected during OCT. Imaging and corresponding histopathology were acquired in-clinic using a wide-field endoscopic OCT system. An annotated dataset (n = 294 images) from 60 patients (34 male and 26 female) was assembled to train four unique neural networks. A deep learning pipeline was built using convolutional and modified u-net models to detect the imaging field of view (network 1), detect artifacts (network 2), identify the tissue surface (network 3), and identify the presence and location of the epithelial-stromal boundary (network 4). The area under the curve of the image and artifact detection networks was 1.00 and 0.94, respectively. The Dice similarity score for the surface and epithelial-stromal boundary segmentation networks was 0.98 and 0.83, respectively. Deep learning (DL) techniques can identify the location and variations in the epithelial surface and epithelial-stromal boundary in OCT images of the oral mucosa. Segmentation results can be synthesized into accessible en face maps to allow easier visualization of changes.

The Development of Robust Antibodies to Sarcospan, a Dystrophin- and Integrin-Associated Protein, for Basic and Translational Research.

Sarcospan (SSPN) is a 25-kDa transmembrane protein that is broadly expressed at the cell surface of many tissues, including, but not limited to, the myofibers from skeletal and smooth muscles, cardiomyocytes, adipocytes, kidney epithelial cells, and neurons. SSPN is a core component of the dystrophin-glycoprotein complex (DGC) that links the intracellular actin cytoskeleton with the extracellular matrix. It is also associated with integrin α7β1, the predominant integrin expressed in skeletal muscle. As a tetraspanin-like protein with four transmembrane spanning domains, SSPN functions as a scaffold to facilitate protein-protein interactions at the cell membrane. Duchenne muscular dystrophy, Becker muscular dystrophy, and X-linked dilated cardiomyopathy are caused by the loss of dystrophin at the muscle cell surface and a concomitant loss of the entire DGC, including SSPN. SSPN overexpression ameliorates Duchenne muscular dystrophy in the mdx murine model, which supports SSPN being a viable therapeutic target. Other rescue studies support SSPN as a biomarker for the proper assembly and membrane expression of the DGC. Highly specific and robust antibodies to SSPN are needed for basic research on the molecular mechanisms of SSPN rescue, pre-clinical studies, and biomarker evaluations in human samples. The development of SSPN antibodies is challenged by the presence of its four transmembrane domains and limited antigenic epitopes. To address the significant barrier presented by limited commercially available antibodies, we aimed to generate a panel of robust SSPN-specific antibodies that can serve as a resource for the research community. We created antibodies to three SSPN protein epitopes, including the intracellular N- and C-termini as well as the large extracellular loop (LEL) between transmembrane domains 3 and 4. We developed a panel of rabbit antibodies (poly- and monoclonal) against an N-terminal peptide fragment of SSPN. We used several assays to show that the rabbit antibodies recognize mouse SSPN with a high functional affinity and specificity. We developed mouse monoclonal antibodies against the C-terminal peptide and the large extracellular loop of human SSPN. These antibodies are superior to commercially available antibodies and outperform them in various applications, including immunoblotting, indirect immunofluorescence analysis, immunoprecipitation, and an ELISA. These newly developed antibodies will significantly improve the quality and ease of SSPN detection for basic and translational research.

Optimalisation of in vitro sterilisation methods for North Sumatra local garlic (Allium Sativum L.)

Abstract The establishment stage is critical to the success of plant tissue culture. Each plant tissue has unique tissue surface characteristics that interact with various natural microbes. The goal of this study was to find a sterilisation method that is effective at removing contamination while not damaging or inhibiting explant regeneration. In this study, Completely Randomized Design (CRD) factorial with 2 factors was used. The first factor was the disinfectant, namely T1 (Benzalkonium chloride 0.5%), T2 (Nordox 6 g/L 0.6%), and T3 (Clorox 10%). The second factor is the type of media, consisting of M1 (BDS supplemented with 1.8 μM/L 2,4-D and 9.2 μM/L Kinetin), M2 (BDS supplemented with 9 μM/L 2,4-D and 9.2 μM/L Kinetin), M3 (MS supplemented with 9 μM/L 2,4-D and 4.6 μM/L Kinetin), and M4 (MS media supplemented with 1.3 μM/L 2,4-D and 9.2 μM/L Kinetin. The observed parameters were the percentage of contamination, the survival rate, the type of contamination, the percentage of callus growing, weight and diameter. The T2 treatment was the most effective in reducing contamination and had the highest survival rate. T3 treatment resulted in the highest callus regeneration percentage, callus weight, and callus diameter, but also the highest level of contamination.

Protective effect of Albizia chinensis saponin on ethanol-induced gastric mucosal injury in rats focus on mucus and mucosal barriers

This study aims to explore the protective effect of Albizia chinensis saponin on ethanol-induced acute gastric ulcer in rats and elucidate its mechanisms. SD rats were deprived of water for 24 hours before the experiment. The control group and model group were administered water by gavage, and the positive drug group received rabeprazole sodium solution(40 mg·kg~(-1)) by gavage. The experimental groups were given different doses of Albizia chinensis saponin solution(3, 10, and 30 mg·kg~(-1)). After 30 minutes, the control group received 1.5 mL of water by gavage, while the other groups were administered an equal volume of 95% ethanol for modeling. After six hours, the rats were killed by cervical dislocation, and the stomachs were collected. The ulcer area was measured, and the ulcer index was calculated. Hematoxylin-eosin(HE) staining was performed to assess histopathological changes in gastric tissue. Periodic acid-Schiff(PAS) staining was used to evaluate the distribution of gastric mucosal surface mucus. Enzyme-linked immunosorbent assay(ELISA) was employed to measure the levels of phospholipids and aminohexose in the gastric mucosa. Western blot was performed to determine the expression levels of the bicarbonate transporter, matrix metalloproteinase, and tight junction-associated proteins in gastric tissue. Immunohistochemistry(IHC) staining was conducted to quantify the number of positive cells for secreted mucin and tight junction-associated proteins. The results showed that the gastric tissue surface of rats in the control group was smooth without ulceration, and the gastric ulcer index of rats in the model group was 35±11. Albizia chinensis saponin at doses of 3, 10, and 30 mg·kg~(-1) resulted in inhibition rates of gastric ulcer of 46%(P&lt;0.01), 85%(P&lt;0.001), and 100%(P&lt;0.001), respectively. Severe disruption of gastric mucosal structure and absence of the mucus layer were observed in the model group. Compared with the model group, the Albizia chinensis saponin group showed intact gastric mucosal surface mucus layer, significantly increased levels of phospholipids and aminohexose in the mucus, increased number of MUC5AC positive cells, and upregulated expression levels of the bicarbonate transporter SLC26A3 and CFTR. It also showed decreased phosphorylation of JNK and c-Jun, reduced expression levels of MMP-8, elevated expression of TIMP-1, and increased expression levels of Occludin and ZO-1. In conclusion, Albizia chinensis saponin enhances the function of the mucus-bicarbonate barrier by upregulating the content of MUC5AC, phospholipids, and aminohexose and increasing the expression levels of the bicarbonate transporter SLC26A3 and CFTR. Moreover, Albizia chinensis saponin exerts its protective effects on gastric ulcers by inhibiting the JNK signaling pathway to prevent excessive activation of MMP-8, thereby reducing the degradation of Occludin and ZO-1 and enhancing the mucosal barrier function. In summary, Albizia chinensis saponin exerts its anti-gastric ulcer effects by simultaneously enhancing the mucus barrier and the mucosal barrier.

The MOCART (magnetic resonance observation of cartilage repair tissue) 2.0 Ankle Score

ObjectivesThe aim of this study was to introduce the MOCART 2.0 ankle score and evaluate its utility and reproducibility for the radiological assessment of cartilage repair tissue in the ankle joint.MethodsThe MOCART 2.0 ankle score evaluates seven individual variables, including “volume fill of (osteo)chondral defect,” “Integration into adjacent cartilage and bone,” “surface of the repair tissue,” “signal intensity of the repair tissue,” “bony defect and bony overgrowth,” “presence of edema-like-marrow signal,” and “presence of subchondral cysts.” Overall, a MOCART 2.0 ankle score between 0 and 100 points may be reached. Two independent readers assessed the 3-T MRI examinations of 48 ankles, who had undergone cartilage repair of a talar cartilage defect using the new MOCART 2.0 ankle score. One of the readers performed two readings. Intra- and interrater reliability were assessed using intraclass correlation coefficients (ICCs) for the overall MOCART 2.0 ankle score.ResultsForty-eight ankles (mean age at surgery 30.2 ± 11.2 years) were evaluated. The overall interrater (ICC = 0.75; 95%CI 0.60–0.85), as well as the intrarater (ICC = 0.83; 95%CI 0.72–0.90) reliability of the MOCART 2.0 ankle score was good. For individual variables the interrater reliability ranged from a kappa value of 0.29 (95%CI 0.01–0.57) for “surface of the repair tissue” to 0.83 (95%CI 0.71–0.95) for “presence of subchondral cysts”.ConclusionsThe newly introduced MOCART 2.0 ankle score, which encompasses the distinct anatomy of the ankle joint, demonstrates good intra- and interrater reliability.Critical relevance statementThe newly introduced MOCART 2.0 ankle score may facilitate the standardized assessment of cartilage repair in the ankle joint and allow an objective comparison of the morphological outcome between alternative treatment options and between different studies.Key PointsThis study introduces the MOCART 2.0 ankle score.The MOCART 2.0 ankle score demonstrated good intra- and interrater reliability.Standardized reporting may improve communication between radiologists and other physicians.Graphical