Search

INTRODUCTION: Mast cells present in the connective tissue stroma release pro inflammatory and mitogenic cytokines. The mast cell in tumor act as a host immunologic anti tumor response. More than 90% of oral cancers are squamous cell carcinomas with oral epithelial dysplasia being the most common potentially malignant disorder. Mast cells play an important role in tumourigenesis. In some studies, with different granules of mast cell is closely related with angiogenesis and tumor invasion. In this study we used toluidine blue for the staining because it reveals mast cells as large, purple, oval and granulated cells. MATERIAL AND METHODS: 30 cases of oral epithelial dysplasia, 30 cases of Oral squamous cell carcinoma (OSCC) and 10 cases of Normal oral mucosa (NOM) were studied for mast cell number using toluidine blue. Mast cells were counted using an Olympus CX41 microscope fitted with an Olympus oculometer grid. Counting was carried out at ×40 by two independent observers and was done in six non overlapping fields in each slide. Mast cells were identified on the basis of the purple color attained by the granules after toluidine blue staining, but the nucleus of these cells appeared blue. RESULTS: Highly significant increase of mast cells in oral epithelial dysplasia on comparison with OSCC whereas there was only a significant increase in mast cells in OSCC on comparison with NOM. CONCLUSION: Mast cells can be used an indicator of increased angiogenesis and can help in the prediction of carcinogenesis, its progression, and also in the prognosis of the malignant lesions. [ABSTRACT FROM AUTHOR]

Phrenic nerves (PNs) play an important role in respiration; however, very few morphological studies have assessed them. This study aimed to provide control reference values, including the density of large and small myelinated PN fibers, for future pathological studies. We assessed a total of nine nerves from eight cases among consecutive autopsy cases registered to the Brain Bank for Aging Research between 2018 and 2019 (five men and three women, mean age 77.0 ± 7.0 years). The nerves were sampled distally, and their structures were analyzed using semi‐thin sections stained with toluidine blue. The mean and standard deviation of the density of each myelinated fiber of the PN was 6908 ± 1132 fibers/mm2 (total myelinated fiber), 4095 ± 586 fibers/mm2 (large diameter myelinated fiber; diameter ≥7 μm), and 2813 ± 629 fibers/mm2 (small diameter myelinated fiber; diameter <7 μm). There was no correlation between myelinated fiber density and age. This study provides the density measurement of the human PN myelinated fiber, and these findings can be used as reference values for the PN in elderly individuals. [ABSTRACT FROM AUTHOR]

The main target of bone tissue engineering is to design biomaterials that support bone regeneration and vascularization. Nanostructured membranes of (MMA)1-co-(HEMA)1/(MA)3-co-(HEA)2 loaded with 5% wt of SiO2-nanoparticles (Si-M) were doped with zinc (Zn-Si-M) or doxycycline (Dox-Si-M). Critical bone defects were effectuated on six New Zealand-bred rabbit skulls and then they were covered with the membranes. After six weeks, a histological analysis (toluidine blue technique) was employed to determine bone cell population as osteoblasts, osteoclasts, osteocytes, M1 and M2 macrophages and vasculature. Membranes covering the bone defect determined a higher count of bone cells and blood vessels than in the sham group at the top regions of the defect. Pro-inflammatory M1 appeared in a higher number in the top regions than in the bottom regions, when Si-M and Dox-Si-M were used. Samples treated with Dox-Si-M showed a higher amount of anti-inflammatory and pro-regenerative M2 macrophages. The M1/M2 ratio obtained its lowest value in the absence of membranes. On the top regions, osteoblasts were more abundant when using Si-M and Zn-Si-M. Osteoclasts were equally distributed at the central and lateral regions. The sham group and samples treated with Zn-Si-M attained a higher number of osteocytes at the top regions. A preferential osteoconductive, osteoinductive and angiogenic clinical environment was created in the vicinity of the membrane placed on critical bone defects. [ABSTRACT FROM AUTHOR]

Copyright of Wool Textile Journal is the property of National Wool Textile Science & Technology Information Center and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Objective: In order to establish a non-decalcified pathological section preparation method for bone tissue specimens of lumbar vertebral body of miniature pigs implanted with bone repair materials. Methods: In this study, lumbar vertebral bone tissue specimens containing bone repair materials were segmented to expose tissue sections. Dehydration with graded ethanol followed by infiltration with Technovit 7200 VLC photopolymer resin. Photopolymerization embedding of lumbar vertebral body tissue specimens containing bone repair material by co-irradiation with yellow and blue light, Preparation of non-decalcified pathological tissue sections with bone-containing repair materials by means of hard tissue pathological cutting and grinding system. Results: The results show that the pathological tissue sections prepared by the above method, After HE staining and toluidine blue staining, observation under a light microscope can better show various tissue and cell structures of bone, The direction and connection of trabecular bone can be clearly observed. Conclusions: A method for preparing pathological tissue sections of bone tissue specimens containing bone repair materials was established, Realized the preparation of histopathological sections of non-decalcified bone with bone repair material, Histological observation with implants achieved after pathological staining, It provides a new pathological detection method and histological evaluation method for the animal test research of biological materials and medical devices. [ABSTRACT FROM AUTHOR]

A semi-interpenetrating network consisting of karaya gum-graft-polyacrylamide and poly(vinyl alcohol) has been developed as an adsorbent material for dye removal applications. The system was made by microwave irradiation of the aqueous mixture of karaya gum, acrylamide, N,N′-methylenebisacrylamide, and poly (vinyl alcohol); and was characterized by FTIR, TGA, SEM, and XRD techniques. It was evaluated as an adsorbent material for removal of ionic dyes from aqueous solutions. Appreciably high adsorption capacity is exhibited by the material towards cationic dyes as indicated by values of maximum adsorption capacity of 82.28, 72.94, 48.75 and 34.67 mg/g towards adsorption of cationic dyes, namely methylene blue (MB), crystal violet (CV), rhodamine B (Rh B), and toluidine blue (TB). Relatively low value of 21.66 mg/g is obtained for anionic dye indigo carmine (IC). The kinetic data of dye adsorption best fitted with the pseudo-first-order kinetic model with R2 values of 0.979, 0.989, 0.946, 0.991, and 0.986 for TB, MB, CV, Rh B and IC, respectively. The isotherm data fit well with the Freundlich model with R2 values of 0.986, 0.955, 0.990, 0.980 and 0.993 for TB, MB, CV, Rh B, and IC, respectively, indicating the heterogeneous and multilayer adsorption. The thermodynamic studies showed dye adsorption to be an exothermic and spontaneous process. [ABSTRACT FROM AUTHOR]

Background/Aims: Alzheimer's disease is a progressive neurological disorder characterized by the intracellular accumulation of Tau protein aggregates. In the present work, we studied the effect of Toluidine Blue and photo-excited Toluidine Blue on the aggregation of repeat Tau using in vitro assays. Methods: The in vitro experiments were carried out on recombinant repeat Tau which was purified by cation exchange chromatography. The ThS fluorescence analysis was used to study the aggregation kinetics of Tau. CD spectroscopy and electron microscopy were used to study the secondary structure and morphology of Tau respectively. The actin cytoskeleton modulation was studied in Neuro2a cells with help of immunofluorescent microscopy. Results: Results showed that Toluidine Blue efficiently inhibited the formation of higher-order aggregates, which was evidenced by Thioflavin S fluorescence assay, SDSPAGE, and TEM. Immunofluorescence studies on the cytoskeleton of Neuro2a cells showed that Toluidine Blue and photo-excited Toluidine Blue treatment at a non-toxic concentration of 0.5 µM stimulated the formation of actin-rich lamellipodia and filopodia structures. Tubulin networks were also differentially modulated after the treatment of Toluidine Blue and photoexcited Toluidine Blue. End-binding protein 1 (EB1) levels were observed to increase after Toluidine Blue and photo-excited Toluidine Blue treatment indicating accelerated microtubule polymerization. Conclusion: The overall study suggested that Toluidine Blue inhibited the aggregation of soluble Tau and photo-excited Toluidine Blue disaggregated the pre-formed Tau filaments. In our study, TB and PE-TB were observed to be potent against Tau aggregation. We observed a distinctive modulation of actin, tubulin networks, and EB1 levels after TB and PE-TB treatment, which suggested that TB and PE-TB have potency against cytoskeleton deformities. [ABSTRACT FROM AUTHOR]

Introduction: Chronic diseases have become major health issues encountering people globally and on top of the list comes "Diabetes Mellitus (DM)". One of its serious complications is "Diabetic Nephropathy (DN)". Aim of the Work: Demonstration of the possible effect of pentoxifylline (PTX) on induced diabetic nephropathy in rats as a cure or prophylactic therapy. Materials and Methods: Seventy adult male albino rats were randomly distributed into five groups; group I (control group), group II (PTX group), group III (diabetic nephropathy group), group IV (diabetic rats received pentoxifylline as a prophylactic treatment one week after induction of diabetes) and group V (diabetic rats received pentoxifylline as a curative treatment starting after sixth week of induction of diabetes). Induction of diabetes was done by intraperitoneal injection of a single dose of streptozotocin (50 mg/kg). A single dose of PTX (200 mg/kg) was given daily by gastric intubation. At the end of experiment (after 8 weeks), renal specimens were collected and processed for paraffin blocks and stained with H&E and immunohistochemical stains and examined by light microscope. Other samples were prepared to obtain semithin sections stained with toluidine blue to be examined by light microscope and ultrathin sections to be examined by transmission electron microscope (TEM). Results: In diabetic nephropathy, there were shrunken glomeruli. The epithelial cells of renal tubules showed vacuolations, degenerative changes and complete shedding. There was anti-NF-κb strong positive reaction of renal tubules. In TEM examination, there were thickening of the glomerular filtration barrier, fusion of foot processes of podocytes, damage of renal tubules. Histopathological examination of the kidney showed improvement in the prophylactic and the curative groups. There was a significant decrease in anti-NF-κb positive reaction in groups II, IV and V. Conclusion: Pentoxifylline had both prophylactic and curative effects against diabetic nephropathy. [ABSTRACT FROM AUTHOR]

Due to the similar pattern of ghrelin localisation in human and porcine intestine, swine are described as a suitable animal model for experiments related to ghrelin and its receptors. In a previous study of ours, the presence of ghrelin immunoreactive endocrine cells in common bile duct (Ductus choledochus) (CBD) of domestic swine was established. However, data regarding the distribution of mast cell types in the CBD in this species are missing. The current study aimed to determine the distribution of tryptase-, ghrelin- and toluidine blue positive mast cells in the wall of extra- and intramural parts of porcine CBD. For this purpose, immunohistochemical staining for detection of tryptase was performed in order to identify ghrelin reactivity in mast cells. Additionally, toluidine blue staining was carried out to compare the number of metachromatic mast cells with that of tryptase positive cells. The comparative quantitative analysis showed that the number of tryptase immunorective cells was higher than that of metachromatic ones in the CBD wall. Ghrelin immunoreactive cells were the most numerous, therefore it can be assumed that not only mast cells but other cells also contain ghrelin. The three mast cells types were more abundant in the propria and subserosa of the intramural part than in the same layers of extramural CBD part. In the muscle layer, mast cell number was higher in the extramural than in the intramural part of the duct. In conclusion, taking into account the results obtained, an important role of mast cells play in controlling CBD motility and biliary gland function was suggested. [ABSTRACT FROM AUTHOR]

• Synovitis was more severe in knees subjected to synovium harvesting than in knees not subjected to harvesting. • Healing was better in the menisci that underwent mesenchymal stem cell (MSC) transplantation than in the control menisci that did not undergo MSC transplantation. • Menisci treated with autologous MSCs showed no red granulation at the tear of the meniscus, but menisci not treated with MSCS showed red granulation. • Macroscopic scores, inflammatory cell infiltration scores, and matrix scores assessed by toluidine blue staining were all significantly better in the autologous MSC group than in the control group without MSCs. • Autologous synovial MSC transplantation suppressed the inflammation caused by synovial harvesting and promoted healing of the repaired menisci in micro minipigs. Allogeneic synovial mesenchymal stem cells (MSCs) effectively promote meniscus healing in micro minipigs. We investigated the effect of autologous synovial MSC transplantation on meniscus healing in a micro minipig model of meniscus repair showing synovitis after synovial harvesting. Synovium was harvested from the left knee of the micro minipigs after arthrotomy and used to prepare synovial MSCs. The left medial meniscus in the avascular region was injured, repaired, and transplanted with synovial MSCs. First, synovitis was compared after 6 weeks in knees with and without synovial harvesting. Second, the repaired meniscus was compared for the autologous MSC group and the control group (in which synovium was harvested but MSCs were not transplanted) 4 weeks after transplantation. Synovitis was more severe in knees subjected to synovium harvesting than in knees not subjected to harvesting. Menisci treated with autologous MSCs showed no red granulation at the tear of the meniscus, but menisci not treated with MSCS showed red granulation. Macroscopic scores, inflammatory cell infiltration scores, and matrix scores assessed by toluidine blue staining were all significantly better in the autologous MSC group than in the control group without MSCs (n = 6). Autologous synovial MSC transplantation suppressed the inflammation caused by synovial harvesting in micro minipigs and promoted healing of the repaired meniscus. [Display omitted] [ABSTRACT FROM AUTHOR]

Copyright of Journal of Guangxi Normal University - Natural Science Edition is the property of Gai Kan Bian Wei Hui and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Photodynamic therapy (PDT) provides an effective therapeutic option for different types of cancer in addition to surgery, radiation, and chemotherapy. The treatment outcome of PDT is largely determined by both the light and dark toxicity of photosensitizers (PSs), which can be technically improved with the assistance of a drug delivery system, especially the nanocarriers. Toluidine blue (TB) is a representative PS that demonstrates high PDT efficacy; however, its application is largely limited by the associated dark toxicity. Inspired by TB’s noncovalent binding with nucleic acids, in this study, we demonstrated that DNA nanogel (NG) could serve as an effective TB delivery vehicle to facilitate anticancer PDT. The DNA/TB NG was constructed by the simple self-assembly between TB and short DNA segments using cisplatin as a crosslinker. Compared with TB alone, DNA/TB NG displayed a controlled TB-releasing behavior, effective cellular uptake, and phototoxicity while reducing the dark toxicity in breast cancer cells MCF-7. This DNA/TB NG represented a promising strategy to improve TB-mediated PDT for cancer treatments.

Collagenous tissues exhibit anisotropic optical properties such as birefringence and linear dichroism (LD) as a result of their structurally oriented supraorganization from the nanometer level to the collagen bundle scale. Changes in macromolecular order and in aggregational states can be evaluated in tendon collagen bundles using polarization microscopy. Because there are no reports on the status of the macromolecular organization in tendon explants, the objective of this work was to evaluate the birefringence and LD characteristics of collagen bundles in rat calcaneal tendons cultivated in vitro on substrates that differ in their mechanical stiffness (plastic vs. glass) while accompanying the expected occurrence of cell migration from these structures. Tendon explants from adult male Wistar rats were cultivated for 8 and 12 days on borosilicate glass coverslips (n = 3) and on nonpyrogenic polystyrene plastic dishes (n = 4) and were compared with tendons not cultivated in vitro (n = 3). Birefringence was investigated in unstained tendon sections using high-performance polarization microscopy and image analysis. LD was studied under polarized light in tendon sections stained with the dichroic dyes Ponceau SS and toluidine blue at pH 4.0 to evaluate the orientation of proteins and acid glycosaminoglycans (GAG) macromolecules, respectively. Structural remodeling characterized by the reduction in the macromolecular orientation, aggregation and alignment of collagen bundles, based on decreased average gray values concerned with birefringence intensity, LD and morphological changes, was detected especially in the tendon explants cultivated on the plastic substrate. These changes may have facilitated cell migration from the lateral regions of the explants to the substrates, an event that was observed earlier and more intensely upon tissue cultivation on the plastic substrate. The axial alignment of the migrating cells relative to the explant, which occurred with increased cultivation times, may be due to the mechanosensitive nature of the tenocytes. Collagen fibers possibly played a role as a signal source to cells, a hypothesis that requires further investigation, including studies on the dynamics of cell membrane receptors and cytoskeletal organization, and collagen shearing electrical properties. [ABSTRACT FROM AUTHOR]

Objectives: The transtibial pull‐out repair (TP) is a relatively new method for treating meniscal root tear; however, the clinical evaluation of its healing effect remains controversial. Due to ethical constraints and limitations of imaging techniques in humans, here we dynamically observe the healing effects of TP and TP with platelet‐rich plasma gel (PRG) at the histological level using an animal model. Methods: Platelet‐rich plasma (PRP) and PRG of rabbits were prepared. Platelet‐derived growth factor (PDGF) and transforming growth factor‐β1 (TGF‐β1) levels in PRP and PRG were determined using an enzyme‐linked immunosorbent assay. A rabbit model of anterior horn tear of the medial meniscus and TP surgery were created. PRG was injected between the anterior horn of the medial meniscus and the tibial tunnel. Rabbits were divided into three groups: the anterior horn tear group (Tear group), the anterior horn tear + TP group (TP group), and the anterior horn tear + TP + PRG group (TP + PRG group). The healing effect was observed dynamically using histopathological studies and biomechanical experiments. Results: The platelet content in PRP significantly increased to approximately 4.57 times that of whole blood. PDGF and TGF‐β1 concentrations in PRG increased to 2.46 and 4.15 times those in PRP, respectively. Hematoxylin and eosin (H&E) and Masson staining showed that the number of inflammatory cells in healing tissue decreased and the collagen fibers significantly increased in TP and TP + PRG groups at 4, 8, and 12 weeks postoperatively compared to those in Tear group. Neatly arranged, interlaced, and dense collagen fibers were found between the anterior horn and bone at 12 weeks. H&E and toluidine blue staining showed that the injury to the femoral condyle cartilage was alleviated. The healing performance in TP + PRG group was better and faster than that in TP group. The maximum tensile fracture strength of the meniscus progressively increased at 8 and 12 weeks postoperatively. Conclusions: Anterior horn injury of the medial meniscus in rabbits can be repaired using the TP technique, and the addition of autologous PRG to the bone tunnel promotes early healing of the meniscus and bone postoperatively. Meanwhile, both treatments can reduce the secondary damage to the cartilage due to osteoarthritis. [ABSTRACT FROM AUTHOR]

Toluidine blue O (TBO) is a phenothiazine dye that, due to its photochemical characteristics and high affinity for biomembranes, has been revealed as a new photosensitizer (PS) option for antimicrobial photodynamic therapy (PDT). This points to a possible association with membranous organelles like mitochondrion. Therefore, here we investigated its effects on mitochondrial bioenergetic functions both in the dark and under photostimulation. Two experimental systems were utilized: (a) isolated rat liver mitochondria and (b) isolated perfused rat liver. Our data revealed that, independently of photostimulation, TBO presented affinity for mitochondria. Under photostimulation, TBO increased the protein carbonylation and lipid peroxidation levels (up to 109.40 and 119.87%, respectively) and decreased the reduced glutathione levels (59.72%) in mitochondria. TBO also uncoupled oxidative phosphorylation and photoinactivated the respiratory chain complexes I, II, and IV, as well as the FoF1-ATP synthase complex. Without photostimulation, TBO caused uncoupling of oxidative phosphorylation and loss of inner mitochondrial membrane integrity and inhibited very strongly succinate oxidase activity. TBO's uncoupling effect was clearly seen in intact livers where it stimulated oxygen consumption at concentrations of 20 and 40 μM. Additionally, TBO (40 μM) reduced cellular ATP levels (52.46%) and ATP/ADP (45.98%) and ATP/AMP (74.17%) ratios. Consequently, TBO inhibited gluconeogenesis and ureagenesis whereas it stimulated glycogenolysis and glycolysis. In conclusion, we have revealed for the first time that the efficiency of TBO as a PS may be linked to its ability to photodynamically inhibit oxidative phosphorylation. In contrast, TBO is harmful to mitochondrial energy metabolism even without photostimulation, which may lead to adverse effects when used in PDT. [ABSTRACT FROM AUTHOR]

Introduction: Sepsis-associated encephalopathy (SAE) is a diffuse cerebral dysfunction resulting from a systemic inflammatory response to infection; however, its pathophysiology remains unclear. Sepsis-induced neuroinflammation and blood-brain barrier (BBB) disruption are crucial factors in brain function disturbance in SAE. Mast cells (MCs) activation plays an important role in several neuroinflammation models; however, its role in SAE has not been comprehensively investigated. Methods: We first established a SAE model by cecal ligation puncture (CLP) surgery and checked the activation of MCs. MCs activation was checked using immumohistochemical staining and Toluidine Blue staining. We administrated cromolyn (10mg/ml), a MC stabilizer, to rescue the septic mice. Brain cytokines levels were measured using biochemical assays. BBB disruption was assessed by measuring levels of key tight-junction (TJ) proteins. Cognitive function of mice was analyzed by Y maze and open field test. Transwell cultures of brain microvascular endothelial cells (BMVECs) co-cultured with MCs were used to assess the interaction of BMVECs and MCs. Results: Results showed that MCs were overactivated in the hippocampus of CLPinduced SAE mice. Cromolyn intracerebroventricular (i.c.v) injection substantially inhibited the MCs activation and neuroinflammation responses, ameliorated BBB impairment, improved the survival rate and alleviated cognitive dysfunction in septic mice. In vitro experiments, we revealed that MCs activation increased the sensitivity of BMVECs against to lipopolysaccharide (LPS) challenge. Furthermore, we found that the histamine/histamine 1 receptor (H1R) mediated the interaction between MCs and BMVECs, and amplifies the LPS-induced inflammatory responses in BMVECs by modulating the TLR2/4-MAPK signaling pathway. Conclusions: MCs activation could mediate BBB impairment and cognitive dysfunction in septic mice in a histamine-dependent pathway. [ABSTRACT FROM AUTHOR]

Background. Atopic dermatitis is a common dermatological disease, and mast cell degranulation is believed to be related with the progression of atopic dermatitis. Mas-related G protein-coupled receptor-X2 (MRGPRX2), and calcium release-activated calcium channel protein 1-2 (ORAI-1, ORAI-2) are involved in mast cell degranulation. Celastrol is an active monomer of Tripterygium wilfordii, and it presents an antiatopic role. Methods. 2,4-Dinitrofluorobenzene (DNFB) and compound 48/80 (C 48/80) were used to establish a slow and acute scratching animal model, respectively. Hematoxylin-eosin and toluidine blue staining was used to investigate tissue injury. Inflammatory factor concentration was measured with ELISA. The expression of MRGPRX2, ORAI-1, and ORAI-2 was detected with immunohistochemistry (IHC) staining. Gene expression profiling and microRNA array were performed to investigate gene differential expression. Results. Celastrol greatly inhibited atopic dermatitis-related tissues injury, mast cell production, histamine release, scratching level, inflammatory factor expression, and activation of MRGPRX2/ORAI axis in the DNFB-induced atopic dermatitis model. The influence of Celastrol on atopic dermatitis was remarkably reversed by overexpression of MRGPRX2. Conclusion. We found that the improvements of atopic dermatitis caused by Celastrol were reversed by treatment with MRGPRX2OE, indicating that Celastrol might affect atopic dermatitis through MRGPRX2. This study might provide a novel thought for the prevention and treatment of atopic dermatitis by regulating MRGPRX2. [ABSTRACT FROM AUTHOR]

Calcium phosphate forms particles under excessive urinary excretion of phosphate in the kidney. While the formation of calcium phosphate particles (CaPs) has been implicated in the damage to renal tubular cells and renal dysfunction, clarifying the ultrastructural information and the elemental composition of the small CaPs in the wide areas of kidney tissue has been technically difficult. This study introduces correlative and sequential light as well as electron microscopic CaP observation in the kidney tissue by combining fluorescent staining for CaPs and energy-dispersive X-ray spectroscopy (EDS) in scanning electron microscopy (SEM) on resin sections prepared using high-pressure freezing and freeze substitution. CaPs formed in mouse kidneys under long-term feeding of a high-phosphate diet were clearly visualized on resin sections by fluorescence-conjugated alendronate derivatives and toluidine blue metachromasia. These CaPs were verified by correlative observation with EDS. Furthermore, small CaPs formed in the kidney under short-term feeding were detected using fluorescent probes. The elemental composition of the particles, including calcium and magnesium, was identified following EDS analyses. These results suggest that the correlative microscopy approach is helpful for observing in situ distribution and elemental composition of CaPs in the kidney and contributing to studies regarding CaP formation-associated pathophysiology. [ABSTRACT FROM AUTHOR]

The surface sulfate groups of cotton-derived cellulose nanowhiskers (CNWs) were quantified using three different methods: conductometric titration, adsorption of toluidine blue O (TBO), and sulfur elemental analysis. The former two methods indicated comparable values, whereas the last method indicated a considerable overestimation of up to 1.4 times the sulfate content compared to the values obtained by titration. This overestimation may be due to the inaccessibility of the sulfates inside the CNWs to the titrant and TBO, and/or the presence of non-sulfate sulfur elements. In addition to the quantitativeness of the TBO adsorption method, the present results also demonstrate the desulfation efficiency under several different desulfation conditions, where the sulfate content decreased by only half after alkali-mediated desulfation even with 2 M NaOH, whereas quite effective desulfation (ca. 90% decrease) was achieved with dilute HCl (0.25 M). [ABSTRACT FROM AUTHOR]

Cellulose nanofiber (CNF) derived from woody bioresources is a fascinating natural nanomaterial. In this work, spherical microparticles were fabricated by using TEMPO-oxidized CNF (TOCNF) and cellulose diacetate (CDA) via Pickering emulsion templating. The CDA-dissolved organic solvents were emulsified stably with TOCNF, followed by removing the solvents to form microspheres with core-shell structures, where the CDA cores were covered with the TOCNF shells. The prepared spherical microparticles possessed an average diameter and sphericity index of 6.4 µm and 0.96, respectively. The zeta-potential value was approximately-48 mV, indicating the stable colloidal system in water. The CDA/TOCNF microparticles were stained with toluidine blue dye for negatively-charged TOCNF. Besides, furry nanofiber-like morphology was observed on the particle surface by scanning electron microscopy. Woodderived CDA/TOCNF microspheres are a promising alternative to fossil resource-derived, non-biodegradable microbeads in cosmetic applications. [ABSTRACT FROM AUTHOR]

Numerous tools and techniques such as vital stain, autofluorescence, exfoliative cytology, histomorphometry, etc. were explored in the detection of dysplasia, but have variable sensitivity and specificity. The present study evaluated the effect of toluidine blue in increasing the accuracy of histomorphometry analysis in detection of the dysplasia in OPMDs on exfoliative cytology. An experimental, observational, cross-sectional research was performed on patients attending dental hospitals and in oral health check-up camps suspected of having OPMDs. Cytology smear was collected using the standard protocol before and after the application of a toluidine blue stain over the lesion. Two pathologists performed the dysplasia grading and histomorphometry analysis. Chi-square and t-tests were carried out (p<0.05). The study observed a positive correlation between toluidine blue positivity and dysplasia in OPMDs. The toluidine blue stain sensitivity, specificity, Positive Predictive Value, and Negative predictive value were 39.53%, 97.56, 97.14, and 43.58% respectively. Toluidine blue before the cytology smear does not improve the diagnostic accuracy of cytomorphometry. Further, there was no significant variation in cellular and nuclear morphometry parameters as compared to normal mucosa. Toluidine blue stain does not improve the efficacy of cytomorphometry in discriminating dysplasia on cytosmears. However, toluidine blue stain can assist clinicians in identifying potential areas of having a suspicion of dysplasia for scalpel biopsy. [ABSTRACT FROM AUTHOR]

In tissue engineering bioresorbable polymers are commonly used to mimetize a damaged tissue characteristics. In spite of that, we did not find a comparative study of the response of the same cell to dense, porous, and fibrous biomaterials. The present work aimed to study the in vitro of pure polymers [poly (ε-caprolactone) (PCL) and poly (hydroxybutyrate-co-hydroxyvalerate) (PHBV)] and their 75:25 blend in dense, porous, and fibrous forms in Vero cells, a fibroblast cell line. The biomaterials were characterized morphologically by scanning electron microscopy (SEM). The MTT assay was used for analysis of possible cytotoxicity. Vero cells were cultured on biomaterials for 120 h and submitted to SEM, morphological and cytochemical analyses. The dyes used were crystal violet (morphology), toluidine blue at pH 4.0 (for nucleic acids and glycosaminoglycans) and 2.5 (for glycosaminoglycans only), periodic acid-Schiff reactive (for neutral sugars), xylidine ponceau at pH 2.5 (for total proteins), and picrosirius/hematoxylin (for selective collagen analysis). The biomaterials studied were not considered citocytotoxic. The cells were able to adhere to and proliferate on the substrates. The cytochemical data indicate that the cells have high functional activity, with high protein synthesis on the scaffolds but do not accumulate acid or neutral sugars. The cells on the fibrous polymers appear to be more elongated, following the shape of the fibers. Regarding the scaffold form (porous or fibrous), the in vitro data do not show great superiority of one type to the other. Then, all the polymers tested are suitable for tissue engineering. [ABSTRACT FROM AUTHOR]

Research background. Despite clearly recognized links between increased body mass and increased risk for various pathological conditions, therapeutic options to treat obesity are still very limited. The aim of the present study is to explore the effect of low-molecular-mass collagen fragments obtained from the scales of Antarctic wild marine fish on rats' visceral and subcutaneous white adipose tissue in a high-calorie diet-induced obesity model. Experimental approach. The study was conducted on outbred rats, which were divided into 3 experimental groups: (i) control, consuming standard food (3.81 kcal/g), (ii) obese group, consuming a high-calorie diet (5.35 kcal/g), and (iii) obese group, consuming a high-calorie diet (5.35 kcal/g) with intragastric administration of low-molecular-mass collagen fragments (at a dose 1 g/kg of body mass during 6 weeks). Low-molecular-mass collagen fragments were obtained by a procedure that included collagen extraction from fish scales and enzymatic hydrolysis with pepsin. Apart from hematoxylin and eosin staining, fibrosis level was assessed by histochemical Van Gieson's trichrome picrofuchsin staining, and mast cells were analysed by toluidine blue O staining. Results and conclusions. Group treated with low-molecular-mass fragments of collagen exhibited decreased rate of mass gain, relative mass, area occupied by collagen fibre of both visceral and subcutaneous adipose tissue, and cross-sectional area of both visceral and subcutaneous adipocytes. Treatment with low-molecular-mass fragments of collagen reduced the infiltration of immune cells, number of mast cells and their redistribution back to the septa. This was also accompanied by a decreased number of the crown-like structures formed by the immune cells, which are markers of chronic inflammation that accompanies obesity. Novelty and scientific contribution. This is the first study that reports the anti-obesity effect of low-molecular-mass fragments produced as a result of controlled hydrolysis of collagen from the scales of Antarctic wild marine fish in the in vivo model. Another novelty of this work is the observation that the tested collagen fragments not only reduce the body mass, but also improve the morphological and inflammatory parameters (decrease in the number of crown-like structures, immune cell infiltration, fibrosis and mast cells). Altogether, our work suggests that low-molecular-mass collagen fragments are a promising candidate for amelioration of some comorbidities linked to obesity. [ABSTRACT FROM AUTHOR]

Carbon nanotubes (CNTs) are known to be a suitable adsorbent of organic contaminants due to their large surface area and hollow structures; but at the nanoscale, CNTs have poor solubility and can be difficult to remove from solution. Studies have shown that the use of peptides can control the size, shape, and structure of materials during synthesis. In this research, magnetic CNTs were synthesized in a co‐precipitation reaction in the presence of the M6A−B1 peptide to correct these deficiencies in CNTs. Characterization of nanoparticles was facilitated by analytical methods including scanning electron microscopy, infrared spectroscopy, x‐ray diffraction, elemental analysis, and more. Water remediation studies were conducted to study the effect of dye, pH, and adsorbent on the removal of Toluidine Blue O from an aqueous solution. Pseudo‐second order kinetics were used in describing the rate of adsorption while a Langmuir isotherm model was used to study the interaction between the peptide‐functionalized material and the adsorbate. The separation factor, RL, calculated for the studied dye concentrations showed that adsorption with the peptide‐functionalized material was favorable. [ABSTRACT FROM AUTHOR]

The relevance of the work is related to the need to develop methods for determining individual opportunities to minimize the harmful effects of ionizing radiation. Experiments were conducted to search for informative indicators of the structure of mast cells to predict the individual effectiveness of the radioprotective effect of hypoxia. A study was performed on white male rats. In 81 animals, before the main experiment, a hypoxic sample was carried out once, by placing the rats in a gas mixture containing 8% oxygen and 92% nitrogen (GGS-8). In each of these rats, a biopsy of the skin of the auricle was performed immediately before the test and at the time of its completion. In the biopsies, the content of various forms of mast cells detected by the method of metachromasia with toluidine blue was determined. After a week, all animals of the main group (81 rats) were exposed to total γ-irradiation at a dose of 11 Gy under the protection of GGS-8. Rats in the control group (63 rats) were irradiated in the same dose in the air. The life expectancy after the exposure was assessed. The radioprotective effect of the gas hypoxic mixture was confirmed, which is expressed in an increase in the average life expectancy of irradiated rats by 2.2 times. A weak relationship between individual quantitative characteristics of mast cells and the life expectancy of irradiated animals is shown. A formal expression of the equation of regression of the number of mast cells on the life expectancy of animals irradiated under the protection of hypoxia has been created. On the basis of regression analysis, groups of animals in which the highest radioprotective effect of hypoxia was manifested were revealed. The minimum life expectancy after irradiation is characteristic of rats with extreme values of the mast cell reactivity index, and all surviving cases are noted in rats with average values of the indicator. Among them there was not a single animal with an intestinal form of death. This indicates the possibility of predicting the radioprotective effect of hypoxia based on an assessment of the functional activity of mast cells. [ABSTRACT FROM AUTHOR]

Background: Gastrobronchial fistulas mostly occur as a result of postoperative complications, including those of bariatric, esophageal, and spleno-pancreatic surgery. Other causes are pneumonia, neoplasm, gastric ulcer, and subphrenic abscess. Traumatic fistulous communications between the stomach and the lung tissue are rare, with only 8 cases reported in the English-language literature (PubMed search) until now. Case Report: We report a 49-year-old female patient with a gastrobronchial fistula secondary to diaphragm rupture 7 years prior, with intrathoracic herniation of the gastric fundus. She underwent thoracotomy for surgical repair. She presented in our Emergency Department with recurrent hemoptysis and painful cough. The diagnosis of the gastrobronchial fistula was confirmed by computed tomography and simultaneous bronchoscopy and esophagogastroscopy, with injection of toluidine blue. As a multidisciplinary team, we opted for surgical repair owing to the fistula extent and severity and the need of repair of the diaphragm hernia. The patient underwent leftsided thoracoscopy. However, owing to dense adhesions and chronic inflammation, we converted to an open procedure. The herniated gastric fundus was repaired by wedge resection. The affected lung tissue was debrided and reconstructed by suture repair. The diaphragmatic defect was closed by sutures with mesh augmentation. The patient's postoperative course was uncomplicated, and she was discharged in good clinical condition on postoperative day 7. Conclusions: Owing to the scarcity of the disease, the management of a gastrobronchial fistula is not standardized. The establishment of the diagnosis of the disease is often challenging. Therapeutic options include conservative measures, endoscopic options, and surgical repair. Our case showed that a multidisciplinary workup is essential for successful treatment. [ABSTRACT FROM AUTHOR]

MicroRNAs are associated with pivotal post-transcriptional gene regulation in bone formation. Human differentiated embryonic chondrocyte expressed gene 1 (Dec1) is also involved in regulating osteoblastogenesis. In the present study, we aimed to investigate the distinctive role of miR-21–5p and Dec1 in osteoblast function and to determine their biological functions. MC3T3-E1 pre-osteoblastic cells were used for in vitro analyses. miR-21–5p knockout (KO) mice, Dec1 KO mice and age-matched wild-type (WT) mice were used to characterize the influence of miR-21–5p and Dec1 deficiencies on bone formation. Morphological analyses [micro-computed tomography (micro-CT)] were performed, and measurements were collected to validate miR-21-5pKO mice. Histopathological changes in mouse femur tissues were assessed by H-E staining, Azan staining, Masson's Trichrome staining, and Toluidine Blue staining. Quantitative real-time RT-PCR, western blotting and immunohistochemical staining were used to characterize the expression levels of Alkaline Phosphatase, Runx2, Osterix, Osteopontin, Dec1 and miR-21–5p. Bioinformatics analyses and dual-luciferase reporter assays were performed to confirm Dec1 as a target of miR-21–5p. Dec1 expression was gradually increased from day 7 of osteoblast induction, while miR-21–5p showed a peak at day 21. In non-induced osteoblasts, a mechanistically gain-of-function transfection study with a miR-21–5p mimic enhanced Runx2 and Osterix expression but suppressed Dec1. miR-21-5pKO mice had reduced bone growth. Dec1-deficient mice showed advanced bone formation at the age of 12 weeks compared to WT mice. The Dec1 deficiency upregulated Runx2 and Osterix expression in Dec1 KO mouse femurs. Those changes, however, were reversed in miR-21-5pKO mouse femurs compared to WT mouse femurs. Dual-luciferase reporter assays showed that Dec1 is a possible downstream target of miR-21–5p. These findings showed that the reduced osteogenic potential due to a miR-21–5p deficiency is achieved by enhanced Dec1 expression and that the miR-21–5p/Dec1 axis is involved in regulating osteoblast function. [Display omitted] • miR-21–5p induced osteoblast differentiation. • Dec1 inhibited osteoblast differentiation via downregulating Runx2 and Osx. • miR-21–5p antagonized Dec1 expression by binding to the 3′UTR region. [ABSTRACT FROM AUTHOR]

The use of doxorubicin (DOX) in clinical practice continues to be challenged by its severe toxicity. DOX cytotoxic activity is not only directed against malignant tumours, given that the treatment will damage healthy tissues as well leading to irreversible injuries. This study aimed to address the in vivo effects of DOX and its co‐administration with a new analog of thioamide; thiocyanoacetamide (TA) on the germinal epithelium. Thus, male rats received either intravenous injection (iv) of 0.03 mg/kg of body weight/week, 0.9% NaCl and were regarded as the control group (CTR), treated with DOX (3.7 mg/kg/week iv), TA [10 mg/kg/day intragastrically (ig)] or a co‐supplementation of DOX and TA. After 50 days, the left testes were dissected and used for toluidine blue, periodic acid‐Schiff (PAS) staining (to evaluate the change in polysaccharides/glycoproteins content), and transmission electron microscopy (TEM) (to assess the morphological damages). To estimate the impact of the test compounds on mitochondrial biogenesis, the expression of NAD‐dependent deacetylase sirtuin‐3 (SIRT‐3) and proliferator‐activated receptor gamma coactivator 1‐alpha (PGC‐1α) were evaluated by immunofluorescence. Apoptotic cells were observed using Hoechst 33324 fluorescent staining. Data showed testicular injuries in the DOX‐treated group, manifested by a significant decrease in total germ cell (GC) number, alteration of Sertoli cell (SC) nucleolus, anchoring junction, along with modifications of the basement membrane (BM) regularity and increase in apoptotic cell count. Mitochondrial aspect and SIRT‐3 and PGC‐1α expression in the testis were unaffected by the DOX. Co‐therapy increased GC number, decreased apoptotic cell count, and restored the BM and anchoring junction regular aspects. This study provides novel insights into understanding DOX‐mediated impairment in rats' testis and might offer some basis for the emerging new alternative therapeutic schemes in male patients undergoing chemotherapy. [ABSTRACT FROM AUTHOR]

Th e purpose of this study was to observe the eff ect of diff erent thawing methods on semen parameters such as motility and morphology and sperm chromatin integrity as assessed by toluidine blue (TB) staining. A total of 20 frozen sperm straws from the same Holstein bull were used. While the 30 sec thawing protocol at 37°C, which is used for thawing frozen sperm straws, constitutes our slow thawing group (n=10), the 6 sec thawing protocol at 70°C constitutes our fast-thawing group (n=10). Th e motility, viability, morphology, plasma membrane integrity, and sperm chromatin condensation parameters of all thawed sperm were investigated. Th ere was a significant diff erence (P

Imaging, silencing cancer-related microRNA, and chemotherapy-phototherapy (CTPT) combination therapy are crucial for cancer diagnosis and drug resistance overcoming. In this study, we designed a multifunctional DNA tetrahedron (MB-MUC1-TD) for the targeted delivery of combined daunorubicin (DAU) + toluidine blue O (TBO). The detection limit of miRNA-21 was determined to be 0.91 nM. The intercalation of DAU and TBO into MB-MUC1-TD was proved by spectroscopic and calorimetric methods. The thermodynamic parameters for the interactions of DAU and/or TBO with MB-MUC1-TD confirmed high drug loading. The first addition of TBO in the ternary system achieved a higher loading of both drugs and a more stable complex structure. Deoxyribonuclease I (DNase I) accelerated the release of DAU and/or TBO loaded in MB-MUC1-TD. Confocal laser scanning microscope demonstrated that MB-MUC1-TD exhibited good imaging ability for miRNA-21 to accurately identify cancer cells, and DAU/TBO was predominantly distributed within the nucleus of cancer cells. In vitro cytotoxicity showed better gene therapy efficacy of MB on MCF-7 cells, better biocompatibility of loaded DAU and TBO on LO2 cells, and stronger synergistic cytotoxicity of DAU + TBO on MCF-7/ADR cells. This study may establish a theoretical foundation for co-loading CTPT combination drugs based on multifunctional DNA nanostructures. [Display omitted] • A multifunctional DNA tetrahedron (MB-MUC1-TD) for combination therapy was designed. • MB-MUC1-TD had good imaging and silencing ability for miRNA-21. • The first addition of TBO facilitated the co-loading of DAU and TBO. • The first-order release mechanism of DAU and TBO was confirmed. • The combination of DAU and TBO loaded by MB-MUC1-TD inhibited drug resistance. [ABSTRACT FROM AUTHOR]

Sonodynamic antimicrobial chemotherapy (SACT) is an effective antimicrobial treatment that can avoid the production of drug-resistant bacteria. Design and development of new high-efficiency sonosensitizers play a key role in the practical application of SACT. The bacteriostatic effects of two phenothiazine compounds, toluidine blue (TB) and azure A (AA) combined with ultrasonic (US) on Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were studied, and the sonodynamic antibacterial activities of TB and AA were compared. The reactive oxygen species (ROS) and the types of ROS produced in the sonodynamic system were detected and the sonodynamic mechanisms of TB and AA were proposed. The sonodynamic bacteriostasis mediated by TB and AA increased with the increasing concentration of sonosensitizer, the extension of sonication time and the increase of reaction temperature. The production of ROS was the main reason that TB and AA had excellent sonodynamic antibacterial performance. Singlet oxygen (1O 2) and hydroxyl radical (•OH) were the main ROS types in the sonodynamic antibacterial system. The ROS produced by the combined action of AA and US was higher than that of TB. Both TB and AA displayed excellent sonodynamic antibacterial activities. Moreover, AA had a higher sonodynamic activity than TB. The electron donation effect and steric hindrance effect of the methyl group of phenothiazine parent nucleus of TB might be the cause of the decrease of its sonodynamic activity. These results would provide a valuable reference for the further study of phenothiazines sonosensitizers and their clinical application in SACT. • TB and AA had excellent sonodynamic antibacterial performance. • 1O 2 and •OH were the main ROS in the sonodynamic antibacterial process. • The sonodynamic activity of AA was higher than that of TB. [ABSTRACT FROM AUTHOR]

The robustness and sensitivity of the surface-enhanced Raman spectroscopy (SERS) technique heavily relies on the development of SERS active materials. A hybrid of semiconductor and plasmonic metals is highly effective as a SERS substrate, which enables the trace level detection of various organic pollutants. This approach demonstrates the photodeposition of plasmonic gold nanoparticles (Au-NPs) on the surface of semiconductor-zinc sulfide nanoflowers (ZnS NFs), grown via the hydrothermal route. The synergistic contribution of the charge-transfer phenomenon and localized surface plasmon resonance of the Au-NPs/ZnS NFs makes it an ideal SERS substrate for the detection of organic pollutants, toluidine blue (TB). The proposed material has a high SERS enhancement factor (109), low limit of detection (10−11 M), good reproducibility, selectivity and strong anti-interference ability. Furthermore, the practicability of the Au-NPs/ZnS NFs is explored in real-time water samples, which are obtained with the satisfactory recovery rates. Additionally, the UVC light illumination on the Au-NPs/ZnS NFs has efficiently degraded TB within a time period of 150 min. These finding demonstrate the significance of the proposed Au-NPs/ZnS NFs for SERS based detection and degradation of organic pollutants in real-time samples, highlighting their potential in monitoring and treating water pollutants in wastewater. [Display omitted] • The Au-NPs/ZnS NFs is prepared by a facile hydrothermal and photoreduction route. • Efficient SERS enhancement is achieved by the synergistic effect of Au-NPs and ZnS. • Excellent performance includes low LOD, high EF, reproducibility and sensitivity. • Real-time analysis of TB in water samples are obtained with good recovery rates. • The degradation ability of Au-NPs/ZnS NFs are demonstrated under UVC irradiation. [ABSTRACT FROM AUTHOR]

The presence of nitrite in food products has generated significant public concern. A simple and rapid dual-mode surface-enhanced Raman spectroscopy (SERS)/colorimetric detection of nitrite is proposed based on a diazo reaction and multifunctional gold nanoparticle-doped covalent organic framework (Au@COF) composite. Under acidic conditions, the reaction between toluidine blue and nitrite yielded a colorless diazo salt, simultaneously attenuating its characteristic absorption peak and Raman signal. The multifunctional Au@COF materials enhanced the Raman signal and ensured good reproducibility. Additionally, the reaction rates improved, and the sensitivity was enhanced due to the excellent adsorption capacity of the COF. The proposed method demonstrated high sensitivity and excellent recovery rates for nitrite detection in food samples. This approach shows potential for precisely detecting nitrite content in real-world food samples by integrating the simplicity of colorimetric analysis with the enhanced sensitivity of SERS. We have developed a novel dual-mode SERS/colorimetric detection of nitrite in foods utilizing multifunctional Au NPs@COF composite. This innovative fabrication of Au NPs@COF composite showed exceptional stability, remarkable adsorption capacity, as well as outstanding SERS performance in terms of both enhancement and reproducibility. Thus, this dual-mode biosensor exhibited an ultra-low limit of detection, excellent reproducibility, high specificity, and satisfactory recoveries in food samples. [Display omitted] • A novel dual-mode SERS/colorimetric method was developed for highly sensitive, specific, and efficient nitrite detection. • The Au@COF composite showed excellent stability and adsorption capacity, facilitating colorimetric applications. • The multifunctional Au@COF demonstrated exceptional SERS performance in terms of both enhancement and reproducibility. [ABSTRACT FROM AUTHOR]

Unusual findings during an autopsy may come from peculiarities in the position, shape, size, weight, consistency, smell or color of organs. The following study was triggered by an autopsy case in which an unusual blue-green discoloration of organs, which changed during the autopsy, was noticed. A review of the local autopsy database, selected cases including the antemortem clinical documentation and the literature has been performed to clarify the etiology of these conspicuous discolorations and to evaluate their diagnostic value. The study showed that certain xenobiotics may lead to such discoloration. After systemic administration of methylene blue, darkening blue-green discoloration of organs, especially the brain and heart, can be observed. In addition, the systemic administration of toluidine blue also appears to be capable of causing such discoloration. Beyond that, drugs (like Rohypnol®) or other foreign substances (like detergents) containing warning colors, i.e. indigocarmin (E132) or Brilliant Blue FCF (E133) may cause discolorations of the upper gastrointestinal tract or the urinary bladder respectively. A blue-green, possibly darkening discoloration of organs during autopsy may point towards an antemortem administration of certain xenobiotics. The affected organs give an indication of the possible route of application and the type of substance. A differentiated interpretation of the etiology of such conspicuous discolorations at autopsy should only be made considering the (medical) history and, if necessary, complementary (toxicological) examinations. [ABSTRACT FROM AUTHOR]

In this study, adsorption of two cationic dyes, Toluidine Blue (TB) and Methyl violet 2B (MV 2B) from an aqueous solution was achieved by using multifunctional composite material. The formulation of the composite (MO@HA) was obtained by using Moringa oleifera seed hull powder, calcium chloride (CaCl2), and ammonium hydrogenophosphate (NH4)2HPO4 salts. Surface morphology, functional groups, specific surface area, and surface charge of the composite were explored using Scanning electron microscopy (SEM), Fourier Transform infrared spectroscopy (FTIR), BET analysis, and point of zero charge (PZC), respectively. The composite material resulted in a structural change in the surface of the adsorbents, increased oxygen vacancies, enhancement of active sites, and a specific surface area of 735.55 m2 g−1. Different adsorption parameters such as pH, contact time, adsorbent dosage, and initial concentration were evaluated. The adsorption study showed that equilibrium was reached after 60 min, and the optimum adsorption pH for both dyes (TB and MV 2B) was 6. Langmuir, Freundlich, Liu, and Temkin were fitted to describe the adsorption isotherm, both TB and MV 2B had best correlation with Liu isotherm. The maximum adsorption capacity of TB and MV 2B were 341.488 and 182.453 mg g−1, respectively. Adsorption-desorption cycling studies on the adsorbent confirmed its regeneration and reusability after 5 cycles. A possible adsorption mechanism involving electrostatic interactions, n-π bonding, and hydrogen bonding was suggested. These findings highlight a new direction in the development of efficient and sustainable adsorbent in environmental remediation, specifically in the removal of dyes from aqueous solution. [ABSTRACT FROM AUTHOR]

Long-chain acyl-CoA synthetases (LACSs), belonging to the acyl-activating enzyme superfamily, play crucial roles in lipid biosynthesis and fatty acid catabolism. Here, we identified 11 LACS genes in the tomato reference genome, and these genes were clustered into six subfamilies. Gene structure and conserved motif analyses indicated that LACS s from the same subfamily shared conserved gene and protein structures. Expression analysis revealed that SlLACS1 was highly expressed in the outer epidermis of tomato fruits and leaves. Subcellular localization assay results showed that SlLACS1 was located in the endoplasmic reticulum. Compared with wild-type plants, the wax content on leaves and fruits decreased by 22.5–34.2 % in SlLACS1 knockout lines, confirming that SlLACS1 was involved in wax biosynthesis in both leaves and fruits. Water loss, chlorophyll extraction, water-deficit, and toluidine blue assays suggested that cuticle permeability was elevated in SlLACS1 knockout lines, resulting in reduction in both drought stress resistance and fruit shelf-life. Overall, our analysis of the LACSs in tomato, coupled with investigations of SlLACS1 function, yielded a deeper understanding of the evolutionary patterns of LACS members and revealed the involvement of SlLACS1 in wax accumulation contribute to drought resistance and extended fruit shelf-life in tomato. [ABSTRACT FROM AUTHOR]

Environmental pollution poses a major problem now a day. Several dyes, in the form of industrial waste, pollute water body and may cause adverse effects to human health. In this paper ADME and toxicity of fives Phenothiazinium group of dyes Methylene blue (MB), Azure A (AA), Azure B (AB), Azure C (AC) and Toluidine Blue O (TBO) were predicted using Swiss ADME and Protox II tools. Results showed these dyes may herm for living organism due to their carcinogenic, mutagenic and hepatotoxic properties. Removal efficiency of these dyes using okra plant product were determined using spectroscopic, thermodynamic and molecular modeling study. It was revealed that these dyes adsorb on the surface of okra leaf mostly at pH 7.0 and the adsorption isotherms were found to fit in Langmuir and Freundlich isotherm model, while Temkin model fails to do this. Mucilage present in different parts of okra plant plays a significant role on removal of these dyes and is able to remove near about 71–92 % of dyes from water body by itself. As this process did not fit in any of above said adsorption isotherm model, it may be suggested that some other mechanism may happen. Further studies explore that these dyes bound to the hydrophobic pocket of mucilage with binding affinity in the order of 105 M−1 and the bindings were exothermic in nature with enthalpy change in the range of − 2.94 to − 4.28 kcal/mole. Molecular docking study validate all the experimental results obtained from spectroscopic and thermodynamic study and enlighten the role of structure of dyes on their binding affinity to mucilage. This paper will help to systematically understand the role of okra plant products on removal efficiency of Phenothiazinium group of dyes with their structural variations. [Display omitted] • Swiss ADME and Protox II showed phenothiazinium group of dyes may be hazardous to human health. • These dyes can be removed from water body using okra plant product and the optimum pH of removal is 7.0. • Dyes adsorb on okra leaf and the adsorption fit to Langmuir, and Freundlich model. • Removal of dyes using okra plant product follow pseudo 1st order kinetics. • The dyes can bound to mucilage through exothermic reaction process, and the binding controlled by overall structure of dyes. [ABSTRACT FROM AUTHOR]

• Electrochemical treatment effectively addresses complex dye mixtures in water. • Photoelectro-Fenton meets effluent color disposal standards. • Significant Chemical Oxygen Demand reduction achieved with photoelectron-Fenton. • Enhanced performance observed in industrial effluent, aided by active chlorine for additional removal. Many industries use complex synthetic organic dye mixtures in their dyeing processes, producing wastewater effluents with complex dye mixtures. In this study, electrochemical advanced oxidation processes like electrochemical oxidation (ECO), ECO with electrogenerated H 2 O 2 (ECO-H 2 O 2), electro-Fenton (EF), and photo electro-Fenton (PEF) with UVC irradiation were compared to discolor and mineralize a mixture of Methyl Orange, Methyl Red, and Toluidine Blue O with a boron-doped diamond anode and a gas-diffusion cathode. While ECO and ECO-H 2 O 2 required long times to achieve a complete discoloration, the dyes were quickly degraded by EF and PEF due to the oxidation with ∙OH formed from Fenton's reaction. Almost complete mineralization was rapidly achieved by PEF for a solution with 424 mg O 2 L−1 of chemical oxygen demand (COD) at a current density of 66.6 mA cm−2 due to the synergistic action of the UVC irradiation. Additionally, the treated effluent was evaluated using the DFZ (Deutsche Farb Zah) parameter for water condition, offering a new perspective and distinguishing this work from previous studies focused on electrified technologies for dye removal. The same behavior has been found for the mixture of such dyes in an industrial effluent upon PEF treatment, even showing faster discoloration and COD reduction as result of the additional removal with active chlorine formed from Cl− oxidation. Ion-exclusion HPLC analysis of the treated synthetic effluent revealed the formation of final carboxylic acids like malic, acetic, oxalic, and formic. Released nitrate, ammonia, and nitrite were detected as nitrogenated by-products from the initial N of the dyes. [ABSTRACT FROM AUTHOR]

[Display omitted] • In ASD, the total volume of brain increases, while the volume of thalamus decreases. • The number of neurons and non-neuron cells in the thalamus is reduced in ASD. • Low numbers of neuron and non-neuron in thalamus are related to ASD-like behavior. The contribution of the thalamus to the development and behavioural changes in autism spectrum disorders (ASD), a neurodevelopmental syndrome, remains unclear. The aim of this study was to determine the changes in thalamic volume and cell number in the valproic acid (VPA)-induced ASD model using stereological methods and to clarify the relationship between thalamus and ASD-like behaviour. Ten pregnant rats were administered a single dose (600 mg/kg) of VPA intraperitoneally on G12.5 (VPA group), while five pregnant rats were injected with 5 ml saline (control group). Behavioural tests were performed to determine appropriate subjects and ASD-like behaviours. At P55, the brains of the subjects were removed. The sagittal sections were stained with cresyl violet and toluidine blue. The thalamic and hemispheric volumes with their ratios, the total number of thalamic cells, neurons and non-neuronal cells were calculated using stereological methods. Data were compared using a t -test and a Pearson correlation analysis was performed to examine the relationship between behaviour and stereological outcomes. VPA-treated rats had lower sociability and sociability indexes. There was no difference in social novelty preference and anxiety. The VPA group had larger hemispheric volume, lower thalamic volume, and fewer neurons. The highest percentage decrease was in non-neuronal cells. There was a moderate positive correlation between the number of non-neuronal cells and sociability, thalamic volume and the number of neurons as well as the time spent in the light box. The correlation between behaviour and stereological data suggests that the thalamus is associated with ASD-like behaviour. [ABSTRACT FROM AUTHOR]

The propagation of oil palm through somatic embryogenesis is the most effective method of cloning this palm tree; however, in vitro cultivation can lead to abnormalities in plant tissue, such as hyperhydricity. The present study aimed to evaluate the difference in anatomical, morphological, and histochemical characteristics, and gene expression in normal (Nm) and hyperhydric (Hh) somatic embryos of oil palm. For this purpose, Nm and Hh somatic embryos were collected from the differentiation medium and were submitted to anatomical and histochemical analyses to assess the nucleus/cytoplasm ratio (toluidine blue), starch (Lugol), and proteins (XP), as well as ultrastructural analyses via transmission electron microscopy. Additionally, gene expression analyses were performed to gain a better understanding on the molecular aspect of hyperhydric abnormality. A higher quantity of differentiated Nm somatic embryos per explant was observed, with a germination rate close to zero in Hh somatic embryos. Additionally, a higher accumulation of proteins and starch was found in Nm somatic embryos when compared to Hh embryos. It was also noted that in Nm somatic embryos, protein reserves were primarily located in the proximal region (embryonic axis), whereas starch reserves were mainly accumulated in the distal region of the somatic embryos. Hh somatic embryos exhibit insignificant starch reserves, and a greater number of intercellular spaces were observed compared to Nm somatic embryos. However, some Hh somatic embryos displayed histochemical characteristics similar to Nm, which could explain the occurrence of reversions from the Hh state to the Nm state observed in this study. Regarding molecular analyses, the gene expression results obtained showed that out of the 19 genes analyzed, 17 were upregulated in hyperhydric embryos when compared to the control condition (normal somatic embryos). Genes involved in stress response, energy metabolism, defense, membrane transport, hormonal regulation, and development were positively regulated, especially those involved in ethylene synthesis and energetic metabolism. To the best of our knowledge, this is the first in-depth study addressing hyperhydricity in oil palm during somatic embryogenesis. [ABSTRACT FROM AUTHOR]

Oral squamous cell carcinoma (OSCC) is considered the most common type of head and neck squamous cell carcinoma (HNSCC) as it holds 90 % of HNSCC cases that arise from multiple locations in the oral cavity. The last three decades witnessed little progress in the diagnosis and treatment of OSCC the aggressive tumor. However, in-depth knowledge about OSCC's pathogenesis, staging & grading, hallmarks, and causative factors is a prime requirement in advanced diagnosis and treatment for OSCC patients. Therefore present review was intended to comprehend the OSCCs' prevalence, staging & grading, molecular pathogenesis including premalignant stages, various hallmarks, etiology, diagnostic methods, treatment (including FDA-approved drugs with the mechanism of action and side effects), and theranostic agents. The current review updates that for a better understanding of OSCC progress tumor-promoting inflammation, sustained proliferative signaling, and growth-suppressive signals/apoptosis capacity evasion are the three most important hallmarks to be considered. This review suggests that among all the etiology factors the consumption of tobacco is the major contributor to the high incidence rate of OSCC. In OSCC diagnosis biopsy is considered the gold standard, however, toluidine blue staining is the easiest and non-invasive method with high accuracy. Although there are various therapeutic agents available for cancer treatment, however, a few only are approved by the FDA specifically for OSCC treatment. The present review recommends that among all available OSCC treatments, the antibody-based CAR-NK is a promising therapeutic approach for future cancer treatment. Presently review also suggests that theranostics have boosted the advancement of cancer diagnosis and treatment, however, additional work is required to refine the role of theranostics in combination with different modalities in cancer treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

This article discusses the advances in non-invasive techniques for the early detection of oral potentially malignant disorders (OPMDs) and oral squamous cell carcinoma (OSCC). The study reviews various tools and methods, such as autofluorescence, chemiluminescence, optical spectroscopy, narrow band imaging, and vital stain colourants, and evaluates their sensitivity and specificity in diagnosing OPMDs. While these non-invasive techniques show promise in early detection, the article concludes that biopsy remains the gold standard for diagnosis. Further research is needed to explore the potential of artificial intelligence and imaging analysis in developing non-invasive screening methods. [Extracted from the article]

Background: To date, no simple and reliable method can replace oral biopsy site detection using Toluidine blue for early identification and treatment of potentially malignant oral disorders (OPMDs). Aims and Objectives: The study was conducted to assess the feasibility of using the colposcope and compare the findings with the toluidine blue staining method for selecting the biopsy site. Materials and Methodology: Study group included 30 patients; five patients dropped out as regression was seen in the lesion. Twenty patients with homogenous leucoplakia and five patients with nonhomogenous Leukoplakia formed a study group. An area with the most extensive cell change in the colposcopic examination was selected by the first version, Image J 1. x Software. After toluidine, blue staining was done. Dark blue-stained sites were considered positive, and site selection for the biopsy was made. Results: Normal vascular pattern in six (24%) cases, 10 (40%) cases showing punctuate vascular pattern, four (16%) of cases showed the mosaic vascular pattern, five (20%) of cases were with the atypical vascular pattern were assessed in colposcopic examination with statistically significant P value (0.009). Fourteen cases out of 20 cases of homogenous and all five cases of nonhomogenous leukoplakia were correctly diagnosed for the biopsy site with both toluidine blue and colposcopic techniques. Conclusion: In this study, Toluidine blue and colposcopic technique showed 76% accuracy for detecting the biopsy site. So colposcope can be an alternative diagnostic aid to toluidine blue in detecting biopsy sites for PMD.

Introduction: In this paper, MgO@AgO/Ag2O nanoparticles were greenly synthesized, the current idea is to replace the harmful chemical technique with an ecofriendly synthesis of metal oxide nanoparticles (NPs) utilizing biogenic sources.Methods: The current investigation was conducted to create silver oxide NPs decorated by MgO NPs (namely, MgO@AgO/Ag2O nanocom-posite) using the leaves extract of Purslane (Portulaca Oleracea) as the reducing and capping agent. The nanopowder was investigated by means of X-ray diffraction, scanning electron mi-croscope, BET surface area, Fourier transform infrared, and UV-vis spectrophotom-eter studies. XRD studies reveal the monophasic nature of these highly crystalline silver nano-particles. SEM studies the shape and morphology of the synthesis AgO/Ag2O and MgO@AgO/Ag2O NPs. The presence of magnesium and oxygen was further confirmed by EDS profile.Results and discussion: The surface area was found to be 9.1787 m2/g and 7.7166 m2/g, respectively. FTIR analysis showed the presence of specific functional groups. UV-vis spectrophotometer studies show the absorption band at 450 nm due to surface plasmon resonance. The results have also indicated the high performance of the greenly synthesized AgO/Ag2O NPs and MgO@AgO/Ag2O NPs for photocatalytic activity dye degradation (methylene blue and toluidine blue).

BACKGROUND: Subarachnoid injection is a commonly used intervention method in animal experiments of spinal cord injury. However, at present, there is a lack of methods for one-time large dose of subarachnoid injection that can cooperate with sustained-release/controlled-release dosage forms to reduce the frequency of administration and avoid subarachnoid catheterization. OBJECTIVE: To investigate a method of subarachnoid injection for one-time large dose administration in a rat spinal cord injury model. METHODS: Sprague-Dawley rats were randomly divided into six groups: a spinal cord injury group, a subarachnoid injection group, a spinal cord injury+subarachnoid injection group, a subarachnoid injection sham operation group, a spinal cord injury+subarachnoid injection sham operation group, and a control group. In the spinal cord injury group, New York University spinal cord impactor was used to establish the rat spinal cord injury model. In the subarachnoid injection group, 200 μL of simulated liquid (stroke-physiological saline solution) was injected into the subarachnoid space of rats via the posterior atlanto-occipital interspace. Rats in the spinal cord injury+subarachnoid injection group received subarachnoid injection after modeling. Rats in the control group were given no treatment. In the subarachnoid injection sham operation group, the posterior atlanto-occipital membrane was exposed and the incision was then sutured. In the spinal cord injury+subarachnoid injection sham operation group, the posterior atlanto-occipital membrane was exposed and the incision was then sutured after modeling. Toluidine blue staining was used to detect drug diffusion and distribution in the subarachnoid space at 1.5 hours after injection. Open field test, inclined plane test, hot-plate test, and body mass measurement were used to verify the occurrence of subarachnoid injection-related complications. RESULTS AND CONCLUSION: (1) Subarachnoid injection via the posterior atlanto-occipital interspace did not cause abnormal results of Basso-Beattie- Bresnahan score, hot-plate test, and inclined plane test in each group. (2) Rats in the subarachnoid injection sham operation and the subarachnoid injection groups regained their preoperative body mass at 1 week after operation (P > 0.05). Rats in the spinal cord injury group, the spinal cord injury+subarachnoid injection sham operation group, and the spinal cord injury+subarachnoid injection group regained their preoperative body mass at 2 weeks after operation (P > 0.05). (3) No special signs of central nervous system injury caused by subarachnoid injection were observed intraoperatively and postoperatively. No leakage of cerebrospinal fluid/simulated solution was found. After the operation, all rats could raise their head normally, and no listlessness or self-mutilating behavior occurred. Rats in the subarachnoid injection sham operation and the subarachnoid injection groups did not have dysuria and dysporia. (4) At 1.5 hours after subarachnoid injection, toluidine blue solution was widely diffused in the subarachnoid space and distributed throughout the whole spinal cord. (5) The above data indicate that subarachnoid injection via the posterior atlanto-occipital interspace is safe and reliable. A single dose of 200 μL administrated can reach the injured segment of the spinal cord quickly and has a less risk of serious complications in the rat model of spinal cord injury. Therefore, it is an alternative method of subarachnoid administration in the rat model of spinal cord injury. BACKGROUND: Subarachnoid injection is a commonly used intervention method in animal experiments of spinal cord injury. However, at present, there is a lack of methods for one-time large dose of subarachnoid injection that can cooperate with sustained-release/controlled-release dosage forms to reduce the frequency of administration and avoid subarachnoid catheterization. OBJECTIVE: To investigate a method of subarachnoid injection for one-time large dose administration in a rat spinal cord injury model. METHODS: Sprague-Dawley rats were randomly divided into six groups: a spinal cord injury group, a subarachnoid injection group, a spinal cord injury+subarachnoid injection group, a subarachnoid injection sham operation group, a spinal cord injury+subarachnoid injection sham operation group, and a control group. In the spinal cord injury group, New York University spinal cord impactor was used to establish the rat spinal cord injury model. In the subarachnoid injection group, 200 μL of simulated liquid (stroke-physiological saline solution) was injected into the subarachnoid space of rats via the posterior atlanto-occipital interspace. Rats in the spinal cord injury+subarachnoid injection group received subarachnoid injection after modeling. Rats in the control group were given no treatment. In the subarachnoid injection sham operation group, the posterior atlanto-occipital membrane was exposed and the incision was then sutured. In the spinal cord injury+subarachnoid injection sham operation group, the posterior atlanto-occipital membrane was exposed and the incision was then sutured after modeling. Toluidine blue staining was used to detect drug diffusion and distribution in the subarachnoid space at 1.5 hours after injection. Open field test, inclined plane test, hot-plate test, and body mass measurement were used to verify the occurrence of subarachnoid injection-related complications. RESULTS AND CONCLUSION: (1) Subarachnoid injection via the posterior atlanto-occipital interspace did not cause abnormal results of Basso-Beattie- Bresnahan score, hot-plate test, and inclined plane test in each group. (2) Rats in the subarachnoid injection sham operation and the subarachnoid injection groups regained their preoperative body mass at 1 week after operation (P > 0.05). Rats in the spinal cord injury group, the spinal cord injury+subarachnoid injection sham operation group, and the spinal cord injury+subarachnoid injection group regained their preoperative body mass at 2 weeks after operation (P > 0.05). (3) No special signs of central nervous system injury caused by subarachnoid injection were observed intraoperatively and postoperatively. No leakage of cerebrospinal fluid/simulated solution was found. After the operation, all rats could raise their head normally, and no listlessness or self-mutilating behavior occurred. Rats in the subarachnoid injection sham operation and the subarachnoid injection groups did not have dysuria and dysporia. (4) At 1.5 hours after subarachnoid injection, toluidine blue solution was widely diffused in the subarachnoid space and distributed throughout the whole spinal cord. (5) The above data indicate that subarachnoid injection via the posterior atlanto-occipital interspace is safe and reliable. A single dose of 200 μL administrated can reach the injured segment of the spinal cord quickly and has a less risk of serious complications in the rat model of spinal cord injury. Therefore, it is an alternative method of subarachnoid administration in the rat model of spinal cord injury. [ABSTRACT FROM AUTHOR]

BACKGROUND: Knee cartilage loss certainly occurs in aged mice, so it is difficult to extract the articular cartilage from the mice. In particular, it is difficult and rarely reported to extract chondrocytes from the small joint on the articular cartilage surface of the ankle joint. OBJECTIVE: To explore the effect of an improved two-step enzyme digestion method to extract chondrocytes from the ankle joint of aged mice, and to observe and evaluate their biological characteristics in vitro. METHODS: The ankle joints of 46-week-old mice were selected, separated and extracted sequentially by the improved two-step enzymatic digestion method in the laboratory with 0.25% pancreatin EDTA and 2g/L type II collagenase. Extracted cells were cultured in vitro and cell growth morphology was observed with an inverted microscope. After trypan blue staining, the number of primary adherent cells was calculated. Toluidine blue and type II collagen immunofluorescence staining was conducted. Chromogenic method using dimethyl methylene blue was used to determine the content of glycosaminoglycan sulfate in the primary adherent cell culture and digestive fluid. Proliferation of chondrocytes was detected using cell counting kit-8 to identify and evaluate the biological characteristics of chondrocytes. RESULTS AND CONCLUSION: After the two-step enzymatic digestion of the articular cartilage, all cells were dissociated and released. An average of 9×105 primary adherent cells were harvested from each ankle joint of aged mice. Primary cells and the first- and second-generation cells displayed spindle and polygonal morphology. Toluidine blue staining showed blue-purple particles that were easily dyed. Type II collagen immunofluorescence staining revealed the positive expression of red fluorescence. The contents of glycosaminoglycan sulfate in the primary adherent cell culture fluid and digestive fluid were (42.41±15.00) and (65.63±10.00) mg/L, which were (0.35±3.78) and (0.21±2.56) mg/L in the negative control culture fluid and digestive fluid, respectively. The number of primary cells reached the maximum on the 6th day, and the number of the first- and second-generation cells reached the maximum on the 5th day. Meanwhile, the cell viability of the first-generation cells was greater than that of the second-generation cells and the primary cells. To conclude, the improved two-step enzymatic digestion method can successfully isolate chondrocytes from the ankle joint of aged mice. Large amounts of adherent cells with high purity and good biological characteristics can successfully synthesize and secrete type II collagen and proteoglycan. The first-generation chondrocytes have the highest viability, followed by the second-generation cells, which can be used for experimental research. [ABSTRACT FROM AUTHOR]

To investigate photoactivated disinfection (PAD) to treat acute pseudomembranous stomatitis, an animal model was established. Six‐week‐old male ICR mice were inoculated with Candida albicans under immunosuppression then divided into three groups (15 mice per group). Pseudomembranous area was measured, then mice had 1 mg mL−1 toluidine blue solution spread on the area, left for 1 min (PAD‐1 group) or 2 min (PAD‐2 group), then irradiated with a 750 mW LED red light for 1 min, a control group received no treatment. Fungal load was measured immediately; after 48 h of observation pseudomembranous area and fungal load were measured. The mice were sacrificed and histopathological examination was performed. Before treatment, pseudomembranous area scores were similar (3 to 4 points) in all groups; 48 h after treatment, the treatment groups' scores were lower (1 to 2 points) than the control group (3 to 4 points, P < 0.05). Immediately after treatment and 48 h later, the fungal loads of the treatment groups were lower than the control group (both P < 0.05). Histopathology of the treatment groups improved more than controls. The treatment groups' results were similar. Therefore, this method of PAD, with short treatment time, reduced the fungal load and pseudomembranous area scores in a mouse model of acute pseudomembranous stomatitis and may have clinical potential. [ABSTRACT FROM AUTHOR]

Four genera of the Boidae family are found in Brazil: Eunectes, Corallus, Epicrates, and Boa. Male copulatory organs in snakes are located inside the tail and are called hemipenes. They are double structures in an inverted position that are exposed during copulation. This study describes the macroscopic and histological aspects of hemipenes in Boidae snakes. One Boa constrictor, three Epicrates cenchria, one Corallus hortulanus hemipenes, and one fragment of the muscular retractor penis magnus (m. retractor penis magnus) from Eunectes murinus were included in this study. The structures were evaluated macroscopically and photo‐documented using a stereo microscope. Tissue fragments were included in glycol methacrylate plastic resin, cut into 3‐μm‐thick slices, and stained with 1% toluidine blue for microscopic evaluation. The macroscopic analysis showed that the arrangement and diameter occupied by the m. retractor penis magnus in the hemipenis and the internal and external cavernous bodies differ between species. The histological analysis showed that the hemipenis of B. constrictor, E. cenchria, and C. hortulanus has stratified epithelium in the external portion supported by dense connective tissue with blood vessels and muscle tissue. Although the literature reports the presence of skeletal musculature in the hemipenis, striated musculature with a centrally located nucleus in the fibre bundle, typical of cardiac musculature in mammals, was observed in the studied species. [ABSTRACT FROM AUTHOR]

Cuticular waxes comprise the hydrophobic layer that protects crops against nonstomatal water loss and biotic and abiotic stresses. Expanding on our current knowledge of the genes that are involved in cuticular wax biosynthesis and regulation plays an important role in dissecting the processes of cuticular wax metabolism. In this study, we identified the Cer-GN1 barley (Hordeum vulgare L.) mutant that is generated by ethyl methanesulfonate mutagenesis with a glossy spike phenotype that is controlled by a single recessive nuclear gene. A physiological analysis showed that the total cuticular wax loads of Cer-GN1 were one-third that of the progenitor wild-type (WT), and its water loss rate was significantly accelerated (p < 0.05). In addition, Cer-GN1 was defective in the glume's cuticle according to the toluidine blue dye test, and it was deficient in the tubule-shaped crystals which were observed on the glume surfaces by scanning electron microscopy. Using metabolomics and transcriptomics, we investigated the impacts of cuticular wax composition and waxy regulatory genes on the loss of the glaucous wax in the spikes of Cer-GN1. Among the differential metabolites, we found that 16-hydroxyhexadecanoic acid, which is one of the predominant C16 and C18 fatty acid-derived cutin monomers, was significantly downregulated in Cer-GN1 when it was compared to that of WT. We identified two novel genes that are located on chromosome 4H and are downregulated in Cer-GN1 (HvMSTRG.29184 and HvMSTRG.29185) that encode long-chain fatty acid omega-monooxygenase CYP704B1, which regulates the conversion of C16 palmitic acid to 16-hydroxyhexadecanoic acid. A quantitative real-time PCR revealed that the expression levels of HvMSTRG.29184 and HvMSTRG.29185 were downregulated at 1, 4, 8, 12, and 16 days after the heading stage in Cer-GN1 when it was compared to those of WT. These results suggested that HvMSTRG.29184 and HvMSTRG.29185 have CYP704B1 activity, which could regulate the conversion of C16 palmitic acid to 16-hydroxyhexadecanoic acid in barley. Their downregulation in Cer-GN1 reduced the synthesis of the cuticular wax components and ultimately caused the loss of the glaucous wax in the spikes. It is necessary to verify whether HvMSTRG.29184 and HvMSTRG.29185 truly encode a CYP704B1 that regulates the conversion of C16 palmitic acid to 16-hydroxyhexadecanoic acid in barley. [ABSTRACT FROM AUTHOR]

Background: Macrophages are the only inflammatory cells that can penetrate the closed nucleus pulposus and their polarization plays an important role in intervertebral disc degeneration (IVDD). This paper attempted to investigate the pathogenesis of IVDD by altering the polarization state of macrophages. Methods: Macrophage RAW264.7 cells were induced by interferonγ (IFN‐γ) and lipopolysaccharide (LPS). The polarization of RAW264.7 cells was estimated by western blot and immunofluorescence. The expressions of inflammatory factors were detected by ELISA. Subsequently, RAW264.7 cells were treated with different concentrations of minocycline (Mino) and sinomenine (Sino), followed by the assessment of cell viability with cell counting kit‐8 kit. Then, RAW264.7 cell culture medium was collected for the culture of human nucleus pulposus cells (NPCs). Toluidine blue staining and type II collagen staining were applied to assay the level of type II collagen. The cell apoptosis, oxidative stress, and nitric oxide (NO) level were appraised by TUNEL, oxidative stress kits and NO kit, respectively. Western blot was employed to test the levels of apoptosis‐ and oxidative stress‐related proteins. Results: IFN‐γ and LPS could induce M1 polarization of RAW264.7 cells. Mino and Sino could reduce the polarization of RAW264.7 cells toward M1. M1‐polarized medium inhibited LPS‐induced activity, inflammation, and damage of NPCs, which were enhanced by Mino and Sino in medium. Conclusion: M1 polarization of macrophages promoted LPS‐induced inflammation and damage of NPCs. [ABSTRACT FROM AUTHOR]