Blue Staining Research Articles

Abstract PO4-07-07: The construction and clinical research value of the prediction model of sentinel lymph node metastasis based on indocyanine green fluorescence quantification in breast cancer

Abstract Objective: To establish a prediction model of sentinel lymph node metastasis by quantitative indocyanine green fluorescence analysis in breast cancer. Methods: From May 2023 to June 2023, 20 patients with breast cancer underwent modified radical mastectomy in the General Surgery Department of Jiangsu Provincial People's Hospital were chosen for research and analysis. Sentinel lymph node were taken from all the patients. The sentinel lymph nodes were located by using the double staining method of subcutaneous injection of Methylene blue and intravenous injection of indocyanine green. The postoperative pathology is used as the standard for evaluating the presence or absence of metastatic cancer in lymph nodes. At the same time, the transcutaneous fluorescence of sentinel lymph node was recorded and the peak time of fluorescence intensity curve was plotted. To establish a logistic diagnostic prediction model and fluorescence threshold, the sentinel lymph node was incised during operation and the fluorescence of sentinel lymph node was recorded again. The criteria for ICG positive determination are determined by the Youden index, which was used to judge the lymph nodes above the threshold in the Logistic diagnostic prediction model. Lymph nodes below the threshold determined in the Logistic diagnostic prediction model are the criteria for ICG negative determination. Cases with less than 3 sentinel lymph node were excluded. Results: 19 cases were ultimately selected. Methylene blue staining data was taking as the judgment standard of sentinel lymph node. Fluorescence imaging was 100% in 14 patients, 66.7% (2/3) in 1 patient, and 125%-150% (5/4-6/4) in 4 patients. Fluorescence quantification values below 2.91 are negative lymph nodes, and values above 4.69 are positive lymph nodes. Moreover, caution is required between 2.91 and 4.69. Combining with the fluorescence dynamic attenuation curve, the positive group has a plateau curve, while the negative group mostly has a decline curve. Combining fluorescence quantification with fluorescence dynamic attenuation curve, the diagnostic rate of positive lymph nodes has a lower false negative rate compared to simple fluorescence quantification and fluorescence dynamic attenuation curve. Conclusion: The quantitative difference value of indocyanine green fluorescence angiography can be used to predict the positive metastasis of sentinel lymph node in breast cancer, making it an attempt to exempt sentinel lymph node biopsy. Citation Format: Yang Wu, Wei Zhang, Jinhai Tang. The construction and clinical research value of the prediction model of sentinel lymph node metastasis based on indocyanine green fluorescence quantification in breast cancer [abstract]. In: Proceedings of the 2023 San Antonio Breast Cancer Symposium; 2023 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2024;84(9 Suppl):Abstract nr PO4-07-07.

Past, Present, and Future Diagnostic Methods for the Early Noninvasive Detection of Oral Premalignant Lesions: A State of the Art and Systematic Review.

Objectives: To provide an in-depth analysis of noninvasive methods for the early diagnosis of oral premalignant lesions, focusing on novel biomarkers and optical technologies, and to discuss their potential in improving the prognosis of patients with oral oncological diseases. Methods: This state-of-the-art review examines various noninvasive diagnostic techniques, including the utilization of salivary microRNAs and optical technologies such as Raman spectroscopy, elastic scattering spectroscopy, diffuse reflectance spectroscopy, narrow-band imaging, autofluorescence imaging, toluidine blue staining, and microendoscopy. Results: Several noninvasive techniques have shown varying degrees of effectiveness in detecting oral cancer. Autofluorescence imaging exhibited sensitivities up to 100% but had variable specificity. toluidine blue staining reported sensitivity between 77% and 100% for high-risk lesions or cancer, with specificity around 45% to 67%. Spectroscopy techniques achieved 72% to 100% sensitivities and specificities of 75% to 98%. Microendoscopy presented a sensitivity of 84% to 95% and a specificity of 91% to 95%. Conclusion: The review highlights the strengths and limitations of each noninvasive diagnostic method and their recent advancements. Although promising results have been demonstrated, there is a need for further development of reliable strategies for early detection and intervention in oral oncology.

Training of a Neural Network System in the Task of Detecting Blue Stains in a Sawmill Wood Inspection System

This article presents the operation of an automatic pine sawn timber inspection system, which was developed at the Woodinspector company and is offered commercially. The vision inspection system is used to detect various wood defects, including knots, blue stain, and mechanical damage caused by worms. A blue stain is a defect that is difficult to detect based on the color of the wood, because it can be easily confused with wood defects or dirt that do not impair its strength properties. In particular, the issues of detecting blue stain in wood, the use of artificial neural networks, and improving the operation of the system in production conditions are discussed in this article. While training the network, 400 boards, 4 m long, and their cross-sections of 100 × 25 [mm] were used and photographed using special scanners with laser illuminators from four sides. The test stages were carried out during an 8-hour workday at a sawmill (8224 m of material was scanned) on material with an average of 10% blue stain (every 10th board has more than 30% of its length stained blue). The final learning error was assessed based on defective boards detected by humans after the automatic selection stage. The system error for 5387 boards, 550 m long, which had blue staining that was not detected by the scanner (clean) was 0.4% (25 pieces from 5387), and 0.1 % in the case of 3412 boards, 610 mm long, on which there were no blue stains, but were wrongly classified (blue stain). For 6491 finger-joint boards (180–400 mm), 48 pieces were classified as class 1 (clean), but had a blue stain (48/6491 = 0.7%), and 28 pieces did not have a blue stain, but were classified as class 2 (28/3561 = 0.7%).

Nasal solitary chemosensory cells govern daily rhythm in mouse model of allergic rhinitis

BackgroundWhile the daily rhythm of allergic rhinitis (AR) has long been recognized, the molecular mechanism underlying this phenomenon remains enigmatic. ObjectiveWe aim to investigate the role of circadian clock in AR development and to clarify the mechanism by which the daily rhythm of AR is generated. MethodsAR was induced in mice using the ovalbumin method. Toluidine blue staining, LC-MS/MS analysis, qPCR, and immunoblotting were performed with AR and control mice. ResultsOvalbumin-induced AR is diurnally rhythmic and associated with clock gene disruption in nasal mucosa. In particular, Rev-erbα is generally down-regulated, and its rhythm retained but with a near 12-h phase shift. Furthermore, global knockout of the core clock gene Bmal1 or Rev-erbα increases the susceptibility of mice to AR and blunts AR rhythmicity. Importantly, nasal SCCs (solitary chemosensory cells) are rhythmically activated, and inhibition of the SCC pathway leads to attenuated AR and a loss of its rhythm. Moreover, rhythmic activation of SCCs is accounted for by diurnal expression of ChAT (an enzyme responsible for the synthesis of acetylcholine) and temporal generation of the neurotransmitter acetylcholine. Mechanistically, REV-ERBα trans-represses Chat through direct binding to a specific response element, generating a diurnal oscillation in this target gene. ConclusionThese findings identify SCCs, under the control of REV-ERBα, as a driver of AR rhythmicity, and suggest targeting SCCs as a new avenue for AR management.

Annular Secondary Syphilis with Histopathologic Features of Granuloma Annulare

Syphilis is one of the most common sexually transmitted diseases in the world. It is known as “the great mimicker” due to its varied cutaneous presentations, which can make it challenging to diagnose. We report a case of a 20-year-old female presenting with a nonhealing, pruritic, annular plaque on the left malar area of 3-month duration. She was initially treated as a case of tinea faciei. The lesion was unresponsive to topical antifungals and steroids with continued increase in size and number. Skin biopsy revealed secondary syphilis with histopathologic features of granuloma annulare. Degenerated collagen, not a feature of secondary syphilis, was seen, prompting further studies to confirm the diagnosis of secondary syphilis. Alcian blue stain was negative, which helped rule out granuloma annulare. She was then given benzathine penicillin G 2.4 million U, which led to a significant decrease in the size of the lesion noted 1 week posttreatment. Only postinflammatory hyperpigmentation was noted after 1 month of treatment. Clinically, secondary syphilis may present as an annular lesion that can mimic tinea. This can also present with atypical histopathologic features and an in-depth investigation is needed to further confirm the diagnosis.

A novel preclinical model of fentanyl-induced cardiorespiratory collapse to investigate resuscitation outcomes

Introduction: Opioid-related morbidities have become a critical public health concern, with ultrapotent synthetic opioids (UPSO) such as fentanyl contributing to the rise in opioid-related fatalities. Respiratory depression is the cardinal trait of fentanyl overdose, leading to systemic hypoxia and subsequent cardiorespiratory collapse (CRC). There is growing recognition that fentanyl induced CRC (fiCRC) is unique from other forms of sudden CRC. Yet, the consequences of fiCRC and its reversal remain poorly understood. This is due in part to the limited preclinical studies investigating rescue from fiCRC. Objective: using a novel model of fiCRC and subsequent reversal, this ongoing study aims to define the impact of UPSO and naloxone rescue on in vivo survival outcomes. Methods and Results: Male and female C57BL/6J mice were used. UPSO fentanyl i.v. administration (30 to 500 mcg/kg) suppressed breathing and subsequently led to fiCRC. The return of spontaneous circulation (ROSC) was achieved by performing manual chest compressions in combination of epinephrine administration after 8 minutes of fiCRC. In subset of mice, naloxone was administered post ROSC. Outcomes after 72 hours were accessed. The latency of fiCRC (mean arterial pressure at 40 mmHg) exhibited a negative correlation with the doses of fentanyl administered. Resuscitation from fiCRC was unreliable across the dose spectrum, while naloxone consistently rescued the majority of fiCRC animals. Naloxone increased ROSC rate from 54% to 86% when the dose of fentanyl was 30 mcg/Kg (n=22). With naloxone rescue, we compared fiCRC with non-opioid-induced cardiac arrest followed by potassium chloride injection (0.8 mg/g). Both models showed severe lung edema as evidenced by Evans Blue staining. However, the lung wet/dry weight ratio was higher in fiCRC group than in the non-opioid treatment group (5.50±0.05 vs. 6.60±0.60, n=4, p<0.05). Survival in fiCRC group was significantly worse than in non-opioid treatment group (survival rate: 25% vs. 83%, n=17, p<0.05). Conclusions: Our study highlights two important findings. First, the reliable resuscitation of fiCRC necessitates the use of naloxone. Second, the likelihood of injury is significant following successful rescue from fiCRC by naloxone. Our continued work using this fiCRC model will lead to insights that can be leveraged for strategies to mitigate injury and improve survival following fentanyl overdose. R01 HL163965 & R01 DA057767. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Salidroside ameliorates acute liver transplantation rejection in rats by inhibiting neutrophil extracellular trap formation.

Acute rejection is an important factor affecting the survival of recipients after liver transplantation. Salidroside has various properties, including anti-inflammatory, antioxidant, and hepatoprotective properties. This study aims to investigate whether salidroside can prevent acute rejection after liver transplantation and to examine the underlying mechanisms involved. An in vivo acute rejection model is established in rats that are pretreated with tacrolimus (1 mg/kg/d) or salidroside (10 or 20 mg/kg/d) for seven days after liver transplantation. In addition, an in vitro experiment is performed using neutrophils incubated with salidroside (1, 10, 50 or 100 μM). Hematoxylin-eosin staining, terminal deoxynucleotidyl transferase dUTP nick-end labeling staining, immunosorbent assays, immunofluorescence analysis, Evans blue staining, and western blot analysis are performed to examine the impact of salidroside on NET formation and acute rejection in vitro and in vivo. We find that Salidroside treatment reduces pathological liver damage, serum aminotransferase level, and serum levels of IL-1β, IL-6, and TNF-α in vivo. The expressions of proteins associated with the HMGB1/TLR-4/MAPK signaling pathway (HMGB1, TLR-4, p-ERK1/2, p-JNK, p-P38, cleaved caspase-3, cleaved caspase-9, Bcl-2, Bax, IL-1β, TNF-α, and IL-6) are also decreased after salidroside treatment. In vitro experiments show that the release of HMGB1/TLR-4/MAPK signaling pathway-associated proteins from neutrophils treated with lipopolysaccharide is decreased by salidroside. Moreover, salidroside inhibits NETosis and protects against acute rejection by regulating the HMGB1/TLR-4/MAPK signaling pathway. Furthermore, salidroside combined with tacrolimus has a better effect than either of the other treatments alone. In summary, salidroside can prevent acute liver rejection after liver transplantation by reducing neutrophil extracellular trap development through the HMGB1/TLR-4/MAPK signaling pathway.

Study on the immune-enhancing and inhabiting transmissible gastroenteritis virus effects of polysaccharides from Cimicifuga rhizoma

Cimicifugae rhizoma is a traditional Chinese herbal medicine in China, and modern pharmacological research showed that it has obvious antiviral activity. Many polysaccharides have been proved to have immune enhancement and antiviral activity, but there are few studies on the biological activity of Cimicifuga rhizoma polysaccharide (CRP). The aim was to explore the character of CRP and its effects on improving immune activity and inhibiting transmissible gastroenteritis virus (TGEV). The monosaccharide composition, molecular weight, fourier transform infrared spectra and electron microscopy analysis of CRP was measured. The effect of CRP on immune activity in lymphocytes and RAW264.7 cells were studied by colorimetry, FITC-OVA fluorescent staining and ELISA. The effect of CRP on TGEV-infected PK-15 cells was determined using Real-time PCR, Hoechst fluorescence staining, trypan blue staining, acridine orange staining, Annexin V-FITC/PI fluorescent staining, DCFH-DA loading probe, and JC-1 staining. Network pharmacology was used to predict the targets of CRP in enhancing immunity and anti-TGEV, and molecular docking was used to further analyze the binding mode between CPR and core targets. The results showed that CRP was mainly composed of glucose and galactose, and its molecular weight was 64.28 kDa. The content of iNOS and NO in CRP group were significantly higher than the control group. CRP (125 and 62.5 μg/mL) could significantly enhance the phagocytic capacity of RAW264.7 cells, and imprive the content of IL-1β content compared with control group. 250 μg/mL of CRP possessed the significant inhibitory effect on TGEV, which could significantly reduce the apoptosis compared to TGVE group and inhibit the decrease in mitochondrial membrane potential compared to TGVE group. The mRNA expression of TGEV N gene in CRP groups was significantly lower than TGEV group. PPI showed that the core targets of immune-enhancing were AKT1, MMP9, HSP90AA1, etc., and the core targets of TGE were CASP3, MMP9, EGFR, etc. Molecular docking show that CRP has binding potential with target. These results indicated that CRP possessed the better immune enhancement effect and anti-TGEV activity.

Biological variability hampers the use of skeletal staining methods in zebrafish embryo developmental toxicity assays

Zebrafish embryo assays are used by pharmaceutical and chemical companies as new approach methodologies (NAMs) in developmental toxicity screening. Despite an overall high concordance of zebrafish embryo assays with in vivo mammalian studies, false negative and false positive results have been reported. False negative results in risk assessment models are of particular concern for human safety, as developmental anomalies may be missed. Interestingly, for several chemicals and drugs that were reported to be false negative in zebrafish, skeletal findings were noted in the in vivo studies. As the number of skeletal endpoints assessed in zebrafish is very limited compared to the in vivo mammalian studies, the aim of this study was to investigate whether the sensitivity could be increased by including a skeletal staining method. Three staining methods were tested on zebrafish embryos that were exposed to four teratogens that caused skeletal anomalies in rats and/or rabbits and were false negative in zebrafish embryo assays. These methods included a fixed alizarin red-alcian blue staining, a calcein staining, and a live alizarin red staining. The results showed a high variability in staining intensity of larvae exposed to mammalian skeletal teratogens, as well as variability between control larvae originating from the same clutch of zebrafish. Hence, biological variability in (onset of) bone development in zebrafish hampers the detection of (subtle) treatment-related bone effects that are not picked-up by gross morphology. In conclusion, the used skeletal staining methods did not increase the sensitivity of zebrafish embryo developmental toxicity assays.

Construction of GPC3-modified Lipopolymer SiRNA Delivery System.

Gene therapy has been widely concerned because of its unique therapeutic mechanism. However, due to the lack of safe and effective carries, it has not been widely used in clinical practice. Glypican 3 (GPC3) is a highly specific proteoglycan for hepatocellular carcinoma and is a potential diagnostic and therapeutic target for hepatocellular carcinoma. Herein, to monitor the effect of gene therapy and enhance the transfection efficiency of gene carriers, GPC3-modified lipid polyethyleneimine-modified superparamagnetic nanoparticle (GLPS), a type of visualized carrier for siRNA (small-interfering RNA) targeting the liver, was prepared. We performed in vitro gene silencing, cytotoxicity, and agarose gel electrophoresis to identify the optimal GLPS formulation. In vitro MRI and Prussian blue staining verified the liver-targeting function of GLPS. We also analyzed the biocompatibility of GLPS by co-culturing with rabbit red blood cells. Morphological changes were evaluated using HE staining. The GLPS optimal formulation consisted of LPS and siRNA at a mass ratio of 25:1 and LPS and DSPE-PEG-GPC3 at a molar ratio of 2:3. GLPS exhibited evident liver-targeting function. In vitro, we did not observe morphological changes in red blood cells or hemolysis after co-culture. In vivo, routine blood analysis revealed no abnormalities after GLPS injection. Moreover, the tissue morphology of the kidney, spleen, and liver was normal without injury or inflammation. GLPS could potentially serve as an effective carrier for liver-targeted MRI monitoring and siRNA delivery.

Dexmedetomidine alleviates blood-brain barrier disruption in rats after cerebral ischemia-reperfusion by suppressing JNK and p38 MAPK signaling.

Dexmedetomidine displays multiple mechanisms of neuroprotection in ameliorating ischemic brain injury. In this study, we explored the beneficial effects of dexmedetomidine on blood-brain barrier (BBB) integrity and neuroinflammation in cerebral ischemia/reperfusion injury. Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 1.5 h and reperfusion for 24 h to establish a rat model of cerebral ischemia/reperfusion injury. Dexmedetomidine (9 􀁐g/kg) was administered to rats 30 min after MCAO through intravenous injection, and SB203580 (a p38 MAPK inhibitor, 200 􀁐g/kg) was injected intraperitoneally 30 min before MCAO. Brain damages were evaluated by 2,3,5-triphenyltetrazolium chloride staining, hematoxylin-eosin staining, Nissl staining, and brain water content assessment. BBB permeability was examined by Evans blue staining. Expression levels of claudin-5, zonula occludens-1, occludin, and matrix metalloproteinase-9 (MMP-9) as well as M1/M2 phenotypes-associated markers were assessed using immunofluorescence, RT-qPCR, Western blotting, and gelatin zymography. Enzyme-linked immunosorbent assay was used to examine inflammatory cytokine levels. We found that dexmedetomidine or SB203580 attenuated infarct volume, brain edema, BBB permeability, and neuroinflammation, and promoted M2 microglial polarization after cerebral ischemia/reperfusion injury. Increased MMP-9 activity by ischemia/reperfusion injury was inhibited by dexmedetomidine or SB203580. Dexmedetomidine inhibited the activation of the ERK, JNK, and p38 MAPK pathways. Moreover, activation of JNK or p38 MAPK reversed the protective effects of dexmedetomidine against ischemic brain injury. Overall, dexmedetomidine ameliorated brain injury by alleviating BBB permeability and promoting M2 polarization in experimental cerebral ischemia/reperfusion injury model by inhibiting the activation of JNK and p38 MAPK pathways.

Time-restricted feeding does not improve nephropathy in humanized sickle cell disease mice

Sickle cell disease (SCD) is an autosomal recessive genetic disease caused by a point mutation in the hemoglobin gene. This substitution results in a “sickling” of red blood cells and subsequent vascular and organ damage. Current treatments, while increasing the lifespan of sickle cell patients, has resulted in a higher prevalence of chronic complications like SCD nephropathy. Our group has previously shown that time-restricted feeding (TRF) in chronic high fat fed mice reduces kidney damage. Given the increased kidney damage in SCD, we hypothesize that TRF in humanized SCD mice will attenuate sickle cell nephropathy. To test this hypothesis, we studied 18-22-week-old male SCD mice. Mice underwent 3 weeks of TRF, only having access to food during the active, dark period (Zeitgeber Time (ZT) 12 to 24). The control group had access to food ad libitum (ad-lib). At the end of the 3 weeks, at 21-25 week old ages, 12 hour urine was collected in metabolic cages as well as kidneys and plasma collection at ZT3 (inactive period). Excretion of urinary kidney damage markers (albumin and NGAL-1) showed no difference between TRF and ad-lib in both the active and inactive collection periods. Urinary excretion of endothelin-1 is known to be upregulated in SCD patients; however, TRF did not change this marker in SCD mice. TRF did not change plasma levels of cytokines or soluble adhesion molecules. Histological analysis of kidneys was conducted as an additional marker of kidney damage. We analyzed iron levels in the kidney with Prussian blue staining as well as fibrosis with Masson’s trichrome. TRF did not change kidney iron accumulation or kidney fibrosis. Immunohistochemical staining for CD3+ T cells did not show a difference in glomerular T cell infiltration in the TRF group compared to the ad-lib group. In conclusion, TRF does not appear to have an effect on renal inflammation or kidney function in SCD mice, but future studies need to be completed. R01 DK134562. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Supplementation of Salvia hispanica L. seed ameliorated liver function enzymes, hyperlipidemia, and oxidative stress in high fat diet fed Swiss albino mice

BackgroundSalvia hispanica L. (Chia) seed improves liver marker enzymes, lipid profile and antioxidant enzyme activity. PurposeThe aim of the research work was to evaluate the hepatic and antioxidant enzymes activity of chia seed supplementation in high fat diet fed mice. MethodsLiver marker enzymes, lipid profile, oxidative stress, antioxidant enzyme activity, and histology were evaluated for chia seed supplementation in high fat diet fed mice. Here, ALT, AST, and ALP were done for liver marker enzymes test. Cholesterol, triglyceride, HDL, and LDL were measured for lipid profiles. MDA, NO, AOPP, and MPO were done for oxidative stress test. Catalase and SOD were performed for measuring antioxidant enzyme activity. Hematoxylin and eosin and Prussian blue staining were done for histology purposes. ResultsIn high fat diet fed mice liver marker enzymes were significantly reduced by supplementation of chia seed. Lipid profiles like cholesterol, triglyceride, LDL concentration were declined and HDL concentration was increased significantly by chia seed supplementation in high fat diet fed mice. High fat diet increased oxidative stress markers such as MDA, NO, AOPP, and MPO which was significantly decreased by the supplementation of chia seed. Antioxidant enzyme activity and histology were also improved by the supplementation of chia seed. ConclusionAccording to this study, chia seed supplementation can reduce liver fibrosis, hyperlipidemia, oxidative stress, and improve antioxidant enzyme activity in mice fed a high-fat diet.

FOXP4-AS1 promotes CD8+ T cell exhaustion and esophageal cancer immune escape through USP10-stabilized PD-L1.

Esophageal cancer (EC) is the 9th most frequently diagnosed malignancy globally with unfavorable prognosis. Immune escape is one of the principal factors leading to poor survival, however, the mechanism underlying immune escape remains largely uninvestigated. The xenograft mouse model and EC cell-CD8+ cytotoxic T lymphocytes (CTLs) co-culture system were established. Immunohistochemistry, qRT-PCR or western blot were employed to detect the levels of long non-coding RNA (lncRNA) FOXP4-AS1, PD-L1, USP10 and other molecules. The abundance of T cells, cytokine production and cell apoptosis were monitored by flow cytometry. The viability of CTLs was assessed by Trypan blue staining. The binding between FOXP4-AS1 and USP10 was validated by RNA pull-down assay, and the interaction between USP10 and PD-L1, as well as the ubiquitination of PD-L1, were detected by co-immunoprecipitation. The elevation of FOXP4-AS1 in EC was associated with decreased CTL abundance, and upregulated PD-L1 facilitated CTL apoptosis in EC. FOXP4-AS1 accelerated EC tumor growth by decreasing the abundance of tumor infiltrating CTLs in vivo. FOXP4-AS1 inhibited the viability of CTLs and facilitated the cytotoxicity and exhaustion of CTLs. In Kyse 450 cell-CTL co-culture system, FOXP4-AS1 suppressed the viability and abundance of CTLs, and inhibited EC cell apoptosis via PD-L1. Mechanistically, FOXP4-AS1 regulated the ubiquitination of PD-L1 through deubiquitinating enzyme USP10. FOXP4-AS1 promoted CTL exhaustion and EC immune escape through USP10-stabilized PD-L1. HIGHLIGHTS: PD-L1 facilitated CD8+ T cell apoptosis in EC. Upregulated FOXP4-AS1 promoted EC tumor growth by inhibiting the viability and facilitating the cytotoxicity and exhaustion of tumor infiltrating CD8+ T cells. FOXP4-AS1 suppressed the viability and abundance of CD8+ T cells through USP10-mediated deubiquitination of PD-L1.

MiRNA-seq and mRNA-seq revealed the mechanism of fluoride-induced cauda epididymal injury

The widespread presence of fluoride in water, food, and the environment continues to exacerbate the impact of fluoride on the male reproductive health. However, as a critical component of the male reproductive system, the intrinsic mechanism of fluoride-induced cauda epididymis damage and the role of miRNAs in this process are still unclear. This study established a mouse fluorosis model and employed miRNA and mRNA sequencing; Evans blue staining, Oil Red O staining, TEM, immunofluorescence, western blotting, and other technologies to investigate the mechanism of miRNA in fluoride-induced cauda epididymal damage. The results showed that fluoride exposure increased the fluoride concentration in the hard tissue and cauda epididymis, altered the morphology and ultrastructure of the cauda epididymis, and reduced the motility rate, normal morphology rate, and hypo-osmotic swelling index of the sperm in the cauda epididymis. Furthermore, sequencing results revealed that fluoride exposure resulted in differential expression of 17 miRNAs and 4725 mRNAs, which were primarily enriched in the biological processes of tight junctions, inflammatory response, and lipid metabolism, with miR-742-3p, miR-141-5p, miR-878-3p, and miR-143-5p serving as key regulators. Further verification found that fluoride damaged tight junctions, raised oxidative stress, induced an inflammatory response, increased lipid synthesis, and reduced lipid decomposition and transport in the cauda epididymis. This study provided a theoretical basis for developing miRNA as potential diagnostic markers and therapeutic target drugs for this injury.

Coupling imaging mass cytometry with Alcian blue histochemical staining for a single-slide approach.

Imaging mass cytometry (IMC) is a metal mass spectrometry-based method allowing highly multiplex immunophenotyping of cells within tissue samples. However, some limitations of IMC are its 1-µm resolution and its time and costs of analysis limiting respectively the detailed histopathological analysis of IMC-produced images and its application to small selected tissue regions of interest (ROI) of one to few square millimeters. Coupling on a single-tissue section, IMC and histopathological analyses could permit a better selection of the ROI for IMC analysis as well as co-analysis of immunophenotyping and histopathological data until the single-cell level. The development of this method is the aim of the present study in which we point to the feasibility of applying the IMC process to tissue sections previously Alcian blue-stained and digitalized before IMC tissue destructive analyses. This method could help to improve the process of IMC in terms of ROI selection, time of analysis, and the confrontation between histopathological and immunophenotypic data of cells.

Cytokine Receptor-like Factor 1 (CRLF1) and Its Role in Osteochondral Repair.

Since cytokine receptor-like factor 1 (CRLF1) has been implicated in tissue regeneration, we hypothesized that CRLF1 released by mesenchymal stem cells can promote the repair of osteochondral defects. The degree of a femoral osteochondral defect repair in rabbits after intra-articular injections of bone marrow-derived mesenchymal stem cells (BMSCs) that were transduced with empty adeno-associated virus (AAV) or AAV containing CRLF1 was determined by morphological, histological, and micro computer tomography (CT) analyses. The effects of CRLF1 on chondrogenic differentiation of BMSCs or catabolic events of interleukin-1beta-treated chondrocyte cell line TC28a2 were determined by alcian blue staining, gene expression levels of cartilage and catabolic marker genes using real-time PCR analysis, and immunoblot analysis of Smad2/3 and STAT3 signaling. Intra-articular injections of BMSCs overexpressing CRLF1 markedly improved repair of a rabbit femoral osteochondral defect. Overexpression of CRLF1 in BMSCs resulted in the release of a homodimeric CRLF1 complex that stimulated chondrogenic differentiation of BMSCs via enhancing Smad2/3 signaling, whereas the suppression of CRLF1 expression inhibited chondrogenic differentiation. In addition, CRLF1 inhibited catabolic events in TC28a2 cells cultured in an inflammatory environment, while a heterodimeric complex of CRLF1 and cardiotrophin-like Cytokine (CLC) stimulated catabolic events via STAT3 activation. A homodimeric CRLF1 complex released by BMSCs enhanced the repair of osteochondral defects via the inhibition of catabolic events in chondrocytes and the stimulation of chondrogenic differentiation of precursor cells.

Titania nanoparticles finishing for smart leather surface

This study focuses on the synthesis and characterization of nanoparticles, specifically titanium dioxide (TiO2) with a uniform size of approximately 5–8 nm in diameter. The nanoparticles exhibit high dispersibility and stability in hydrophilic environments. Notably, nitrogen-doped nanoparticles demonstrate intriguing self-cleaning properties, as evidenced by the degradation of methylene blue stain under UV light. Rutile nanoparticles stabilize color and color fastness in the presence of anatase-covered surfaces. Anti-mold tests reveal enhanced antimicrobial properties of the surfaces containing titania NPs when functionalized with polyethylene glycol (PEG). Ascorbic acid functionalized nanoparticles show promising behavior, encouraging the exploration of nanoparticle functionalization for property improvement. Leather samples coated with nanoparticles undergo abrasion, wear, and scratch resistance tests. The nanoparticles significantly enhance abrasion resistance, doubling the resistance. This suggests the potential for nanoparticle-based finishing to improve fastness to dry and wet rubbing, reducing defects in the coating.

Electroacupuncture pretreatment alleviates rats cerebral ischemia-reperfusion injury by inhibiting ferroptosis

ObjectiveTo explore the preventive effect of electroacupuncture pretreatment on stroke in rats by inhibiting ferroptosis and oxidative stress. MethodsRats were randomly assigned to the sham, middle cerebral artery occlusion/reperfusion (MCAO/R), MCAO/R + EP, MCAO/R + EP + erastin, and MCAO/R + EP + ferrostatin 1 groups. Daily electroacupuncture was performed 2 weeks before establishing the MCAO/R model utilizing the modified Zea Longa suture method. Rats were sacrificed 1 day after reperfusion, and brain tissues were collected. They were prepared for hematoxylin and eosin staining, prussian blue staining, transmission electron microscope. Measurement of total iron levels using a commercial kit, detection of malondialdehyde (MDA) and superoxide dismutase (SOD) levels by ELISA, and examination of 15-lox2, GPX4, SLC7A11, ACSL4, and TFR1 by western blotting. ResultsCompared with sham rats, cerebral infarction size was dramatically larger in MCAO/R rats. Moreover, the MCAO/R group displayed damaged mitochondria with a disarranged structure of cristae; free iron, total iron levels, and oxidative stress were significantly higher. Cerebral pathological lesions, oxidative stress, total iron levels, and protein levels of ACSL4, TFR1, and 15-lox2 were significantly reduced in the MCAO/R + EP and MCAO/R + EP + ferrostatin 1 groups, while the protective effect of electroacupuncture pretreatment on cerebral ischemia-reperfusion injury was inhibited by treatment with the ferroptosis activator erastin. ConclusionElectroacupuncture pretreatment can protect rats from cerebral ischemia-reperfusion injury by reducing the area of cerebral infarction and inhibiting ferroptosis and oxidative stress.

Contribution of Red Blood Cells and Platelets to Blood Clot Computed Tomography Imaging and Compressive Mechanical Characteristics.

Thrombus computed tomography (CT) imaging characteristics may correspond with thrombus mechanical properties and thus predict thrombectomy success. The impact of red blood cell (RBC) content on these properties (imaging and mechanics) has been widely studied. However, the additional effect of platelets has not been considered. The objective of the current study was to examine the individual and combined effects of blood clot RBC and platelet content on resultant CT imaging and mechanical characteristics. Human blood clot analogues were prepared from a combination of preselected RBC volumes and platelet concentrations to decouple their contributions. The resulting clot RBC content (%) and platelet content (%) were determined using Martius Scarlet Blue and CD42b staining, respectively. Non-contrast and contrast-enhanced CT (NCCT and CECT) scans were performed to measure the clot densities. CECT density increase was taken as a proxy for clinical perviousness. Unconfined compressive mechanics were analysed by performing 10 cycles of 80% strain. RBC content is the major determinant of clot NCCT density. However, additional consideration of the platelet content improves the association. CECT density increase is influenced by clot platelet and not RBC content. Platelet content is the dominant component driving clot stiffness, especially at high strains. Both RBC and platelet content contribute to the clot's viscoelastic and plastic compressive properties. The current in vitro results suggest that CT density is reflective of RBC content and subsequent clot viscoelasticity and plasticity, and that perviousness reflects the clot's platelet content and subsequent stiffness. However, these indications should be confirmed in a clinical stroke cohort.