Expression Of Markers Research Articles

Enhancing osteoblast differentiation and bone repair: The priming effect of photobiomodulation on adipose stromal cells

Cellular therapy using adipose tissue–derived mesenchymal stromal cells (at-MSCs) has garnered attention for the treatment of bone defects. Therefore, preconditioning strategies to enhance the osteogenic potential of at-MSCs could optimize cell therapy outcomes, and photobiomodulation (PBM) therapy has emerged as an effective, noninvasive, and low-cost alternative. This study explored the impacts of PBM on at-MSCs differentiation and the subsequent repair of bone defects treated with cell injection. Rat at-MSCs were cultured and irradiated (at-MSCsPBM) following the PBM protocol (660 nm; 20 mW; 0.714 W/cm2; 0.14 J; 5 J/cm2). Cellular differentiation was assessed based on the expression of gene and protein markers. Reactive oxygen species (ROS) were detected using fluorescence. At-MSCsPBM were injected into 5-mm calvarial lesions, and bone formation was analyzed using micro-CT and histological evaluations. At-MSCs were used as control. Data were analyzed using the ANOVA or t-test. At-MSCsPBM exhibited high levels of gene and protein runt-related transcription factor-2 (Runx2) and alkaline phosphatase (Alp) expression. PBM increased ALP activity and significantly reduced ROS levels. In addition, PBM increased the expression of Wnt pathway–associated genes. In vivo, there was an increase in the morphometric parameters, including bone volume, percentage of bone volume, bone surface area, and trabecular number, in at-MSCsPBM-treated defects compared with those in the control. These findings suggest that PBM enhances the osteogenic potential of at-MSCs, thereby supporting the advancement of improved cellular therapies for bone regeneration.

Macrophage erythropoietin signaling promotes macrophage-myofibroblast transformation and fibroblast-myofibroblast differentiation

While myofibroblasts are the key cause of abnormal extracellular matrix accumulation, the origin of which has not yet been fully elucidated. Recently, it has been found that macrophage-myofibroblast transformation (MMT) defined by the expression of both macrophage markers (F4/80 or CD68) and myofibroblast markers (α-SMA) is one of its important sources. In the process of MMT, it is unclear whether epor is involved. In this study, when BMDM was induced by tgf-β1, the number of F4/80+α-SMA+ cells increased, the cells polarized toward M2, and the expression of tgf-β1 increased. After the activation of epor, the number of F4/80 +α-SMA + cells and the polarization level of M2 were further increased. At the same time, we found that the conditioned medium from MMT cells could induce the activation of 3T3 cells with increased the expression of α-SMA and col-1. In contrast, the number of F4/80+α-SMA + cells, the polarization of M2, and the expression of Tgf-β1 decreased after epor was inhibited by siRNA. Our results demonstrate that the activation of epor in BMDMs could promote the transformation of macrophage-myofibroblast induced by TGF-β1.

Characterising the expression of the organic cation transporter OCT3 in cutaneous papillomas of dogs.

The identification of the activation of the mammalian target of rapamycin (mTOR) signalling pathway as a frequent molecular event in canine cutaneous papillomas (CPs) has provided the rational foundation to explore novel molecular-targeted therapies. Recent evidence indicates that metformin reduces the size of CPs in mice by inhibiting the mTOR signalling pathway. These effects require the expression of the organic cation transporter 3 (OCT3/SLC22A3), a well-known metformin uptake transporter. The aim of the present study was to characterise the expression pattern of the metformin uptake transporter OCT3 in canine samples of CP that have shown activation of the mTOR signalling pathway in order to predict if this hyperplastic epidermal lesion is potentially sensitive to metformin. The expression of OCT3 was evaluated by immunohistochemical investigation in sections of a previously constructed tissue microarray containing 28 samples of canine CP and compared with that previously evaluated for the mTOR activation marker pS6. OCT3 was highly expressed in the membrane and cytoplasm of the basal and suprabasal epidermal cells in all samples of canine CP. This OCT3 expression was localised at similar epidermal compartments to those observed for pS6. These results show that canine CPs exhibit the expression of surrogate markers that suggest sensitivity to metformin, such as upregulated OCT3 and pS6 expression. Taken together, these findings provide the rationale for the early assessment of the use of metformin as a mechanism-based therapeutic approach for treating canine patients with persistent or multiple CPs.

Pressure loading regulates the stemness of liver cancer stem cells via YAP/BMF signaling axis.

Cancer stem cells (CSCs) are considered the major cause of the occurrence, progression, chemoresistance/radioresistance, recurrence, and metastasis of cancer. Increased interstitial fluid pressure (IFP) is a key feature of solid tumors. Our previous study showed that the distribution of liver cancer stem cells (LCSCs) correlated with the mechanical heterogeneity within liver cancer tissues. However, the regulation of liver cancer's mechanical microenvironment on the LCSC stemness is not fully understood. Here, we employed a cellular pressure-loading device to investigate the effects of normal IFP (5 mmHg), as well as increased IFP (40 and 200 mmHg) on the stemness of LCSCs. Compared to the control LCSCs (exposure to 5 mmHg pressure loading), the LCSCs exposed to 40 mmHg pressure loading exhibited significantly upregulated expression of CSC markers (CD44, EpCAM, Nanog), enhanced sphere and colony formation capacities, and tumorigenic potential, whereas continuously increased pressure to 200 mmHg suppressed the LCSC characteristics. Mechanistically, pressure loading regulated Yes-associated protein (YAP) activity and Bcl-2 modifying factor (BMF) expression. YAP transcriptionally regulated BMF expression to affect the stemness of LCSCs. Knockdown of YAP and overexpression of BMF attenuated pressure-mediated stemness and tumorgenicity, while YAP-deficient and BMF-deletion recused pressure-dependent stemness on LCSCs, suggesting the involvement of YAP/BMF signaling axis in this process. Together, our findings provide a potential target for overcoming the stemness of CSCs and elucidate the significance of increased IFP in cancer progression.

Targeting CREB-binding protein (CBP) abrogates colorectal cancer stemness through epigenetic regulation of C-MYC.

Colorectal cancer (CRC) is a common cancer worldwide with an increasing annual incidence. Cancer stem cells (CSCs) play important roles in the occurrence, development, recurrence, and metastasis of CRC. The molecular mechanism regulating the development of colorectal CSCs remains unclear. The discovery of human induced pluripotent stem cells (hiPSCs) through somatic cell reprogramming has revolutionized the fields of stem cell biology and translational medicine. In the present study, we converted hiPSCs into cancer stem-like cells by culture with conditioned medium (CM) from CRC cells. These transformed cells, termed hiPSC-CSCs, displayed cancer stem-like properties, including a spheroid morphology and the expression of both pluripotency and CSC markers. HiPSC-CSCs showed tumorigenic and metastatic abilities in mouse models. The epithelial-mesenchymal transition phenotype was observed in hiPSC-CSCs, which promoted their migration and angiogenesis. Interestingly, upregulation of C-MYC was observed during the differentiation of hiPSC-CSCs. Mechanistically, CREB binding protein (CBP) bound to the C-MYC promoter, while histone deacetylase 1 and 3 (HDAC1/3) dissociated from the promoter, ultimately leading to an increase in histone acetylation and C-MYC transcriptional activation during the differentiation of hiPSC-CSCs. Pharmacological treatment with a CBP inhibitor or abrogation of CBP expression with a CRISPR/Cas9-based strategy reduced the stemness of hiPSC-CSCs. This study demonstrates for the first time that colorectal CSCs can be generated from hiPSCs. The upregulation of C-MYC via histone acetylation plays a crucial role during the conversion process. Inhibition of CBP is a potential strategy for attenuating the stemness of colorectal CSCs.

The Optejet Technology Minimizes Preservative-Mediated Cytotoxicity of Conjunctival Epithelial Cells Treated with Latanoprost In Vitro.

Purpose: Benzalkonium chloride (BAK) is a commonly used preservative to maintain sterility for multiuse eye drops such as latanoprost. One option to minimize the deleterious effects of BAK in eye drops may be to reduce the volume administered. The aim of this study was to assess the response of cells from the ocular surface to latanoprost+BAK administered by the Optejet technology, which dispenses a microdose (∼8 µL) ophthalmical spray. Methods: Cultured human conjunctival epithelial cells were exposed to the following treatments: (1) no treatment, (2) drop form of latanoprost without BAK (∼35 µL), (3) drop form of latanoprost with 0.01% BAK (∼35 µL), (4) ophthalmical spray form of latanoprost with 0.01% BAK delivered by the Optejet technology (∼8 µL). After 5 h, cells were assessed for changes in cytotoxicity, morphology, and inflammatory marker expression. Results: Latanoprost+BAK delivered by a drop induced cytotoxicity, cytoplasmic shrinkage, and loss of cell-cell contact, and expression of chemokine (C-C motif) ligand 2 and interleukin-6. In contrast, latanoprost+BAK delivered by the Optejet technology was both well tolerated and similar to no treatment controls and BAK-free latanoprost treatment. Conclusions: A microdose of latanoprost+BAK ophthalmical spray administered with the Optejet technology prevented the cytotoxicity associated with larger volumes found in eye drops. Precision dosing by the Optejet technology has the potential to decrease ocular surface disorder typically associated with eye drops containing preservatives.

Lens capsule advanced glycation end products induce senescence in epithelial cells: Implications for secondary cataracts.

Posterior capsule opacification (PCO) is a common complication after cataract surgery. Residual lens epithelial cells (LECs) on the anterior lens capsule, after cataract surgery, migrate to the posterior lens capsule and undergo transdifferentiation into myofibroblast-like cells. Those cells synthesize excessive amounts of extracellular matrix and contribute to fibrosis during PCO. Cellular senescence, a phenomenon that increases with aging, has been implicated in several fibrotic diseases. Here, we have investigated the prevalence of senescent LECs within the lens posterior capsule and the ability of advanced glycation end products (AGEs) in lens capsules to induce senescence, contributing to PCO. Aged lens capsules from pseudophakic human cadaver eyes showed the presence of senescent LECs. In human capsular bags, LECs showed an age-dependent increase in senescence after 28 days of culture. Human LECs cultured on aged lens capsules for 3 days underwent senescence; this effect was not seen in LECs cultured on young lens capsules. Human LECs cultured on an AGE-modified extracellular matrix (ECM-AGEs) showed an AGE-concentration-dependent increase in the expression of senescence markers and reactive oxygen species (ROS) levels. Treatment with a RAGE antagonist and ROS inhibitor reduced the expression of senescence and fibrotic markers. Additionally, conditioned media from ECM-AGEs-treated cells induced the expression of fibrotic markers in naïve LECs. Together, these suggest that AGEs in the capsule induce senescence of LECs, which triggers the mesenchymal transition of neighboring non-senescent LECs and contributes to PCO.

PPARγ/ETV2 axis regulates endothelial-to-mesenchymal transition in pulmonary hypertension.

Endothelial-to-mesenchymal transition (EndoMT) plays an important role in pulmonary hypertension (PH) but the molecular mechanisms regulating EndoMT remain to be defined. We demonstrate that the axis of the transcription factors PPARγ (Peroxisome Proliferator-Activated Receptor gamma) and ETV2 (ETS variant 2) play important roles in the pathogenesis of PH. Decreased levels of the expression of PPARγ and ETV2 along with reduced endothelial and increased EndoMT markers are consistently observed in lungs and pulmonary artery endothelial cells (PAECs) of idiopathic pulmonary arterial hypertension patients, in hypoxia-exposed mouse lungs, human PAECs, and in induced-EndoMT cells. Etv2 +/- mice spontaneously developed PH and right ventricular hypertrophy (RVH), associated with increased EndoMT markers and decreased EC markers. Interestingly, chronic hypoxia exacerbated right ventricular systolic pressure and RVH in Etv2 +/- mice. PPARγ transcriptionally activates the ETV2 promoter. Consistently, while mice overexpressing endothelial PPARγ increases the expression of ETV2 and endothelial markers with reduced EndoMT markers, endothelial PPARγ KO mice show decreased ETV2 expression and enhanced EndoMT markers. Inducible overexpression of ETV2 under induced-EndoMT cell model reduces number of cells with mesenchymal morphology and decreases expression of mesenchymal markers with increased EC makers, compared to control. Therefore, our study suggests that PPARγ-ETV2 signaling regulates PH pathogenesis through EndoMT.

Prenatal exposure to di-n-butyl phthalate promotes RIPK1-regulated necroptosis of uroepithelial cells and induces hypospadias through the epithelial-mesenchymal transition process in newborn male rats

Maternal exposure to di-n-butyl phthalate (DBP) has been linked to the induction of hypospadias; however, the underlying mechanism remains unclear. Necroptosis is reported to be implicated in developmental malformations. This study aimed to investigate the underlying mechanism of necroptosis in the development of hypospadias. DBP was dissolved in corn oil, and pregnant rats were administered a precisely measured dose of DBP (750mg/kg/day) via gastric intubation from gestation day 14 to 18. Control rats received only corn oil. The day of birth was considered postnatal day (PND) 1. Male hypospadias rats were identified on PND 7. Genital tubercle tissues were collected and stored at −80°C for subsequent PCR analysis, cryopreserved in liquid nitrogen for western blot, or fixed in formalin for immunohistochemistry (IHC) staining. IHC staining and western blot analysis revealed increased expression of RIPK1 and necroptosis markers in genital tubercle (GT) tissue compared to the control group. Additionally, higher levels of EMT and impaired androgen receptor expression were observed in GT tissue. Exposure to increased DBP concentrations in rat primary uroepithelial cells (PUCs) led to elevated ROS production. Necroptosis markers and EMT expression levels were upregulated in PUCs following DBP incubation. Notably, treatment with DBP combined with necrostatin-1, a necroptosis inhibitor, reduced the expression of EMT markers and ROS production compared to DBP treatment alone. In vitro studies further revealed that DBP-induced necroptosis promoted the degradation of E-cadherin through the ubiquitin-proteasome pathway in PUCs. Our findings suggest that maternal exposure to DBP promotes necroptosis in uroepithelial cells by elevating ROS level and EMT status. Thus, necroptosis may play an essential role in the development of hypospadias.

Didang decoction attenuates cancer-associated thrombosis by inhibiting PAD4-dependent NET formation in lung cancer.

This research aims to investigate the impact of Didang decoction (DD) on the formation of neutrophil extracellular traps (NETs) and cancer-associated thrombosis in lung cancer. BALB/c nude mice were used to establish xenograft models for inducing deep vein thrombosis. Tumor growth and thrombus length were assessed. The impact of DD on NET generation was analyzed using enzyme-linked immunosorbent assay, immunofluorescence staining, quantitative real-time PCR, and western blot analysis, both in vivo and in vitro. CI-amidine, a PAD4 inhibitor, was employed to evaluate the role of PAD4 in the generation of NETs. In vivo studies demonstrated that treatment with DD reduced tumor growth, inhibited thrombus formation, and decreased the levels of NET markers in the serum, tumor tissues, neutrophils, and thrombus tissues of mice. Additional data indicated that DD could suppress neutrophil counts, the release of tissue factor (TF), and the activation of thrombin-activated platelets, all of which contributed to increased formation of NETs in mouse models. In vitro, following incubation with conditioned medium (CM) derived from Lewis lung carcinoma cells, the expression of NET markers in neutrophils was significantly elevated, and an extracellular fibrous network structure was observed. Nevertheless, these NET-associated changes were partially counteracted by DD. Additionally, CI-amidine reduced the expression of NET markers in CM-treated neutrophils, consistent with the effects of DD. Collectively, DD inhibits cancer-associated thrombosis in lung cancer by decreasing PAD4-dependent NET formation through the regulation of TF-mediated thrombin-platelet activation. This presents a promising therapeutic strategy for preventing and treating venous thromboembolism in lung cancer.

TGF-β2, EGF and FGF21 influence the suckling rat intestinal maturation

Some of the growth factors present in breast milk, such as transforming growth factor-β (TGF-β), epidermal growth factor (EGF) and fibroblast growth factor 21 (FGF21), play important roles in the development of the intestinal tract. The aim of this study was to determine the effect of a supplementation with TGF-β2, EGF and FGF21 on suckling rats intestinal maturation. For this purpose, Wistar rats were supplemented daily with TGF-β2, EGF or FGF21 throughout the suckling period. We evaluated the functionality of the intestinal epithelial barrier through an in vivo dextran permeability assay, and by a histomorphometric and immunohistochemical study. In addition, the intestinal gene expression of tight junction-associated proteins, mucins, toll-like receptors, and maturation markers was analyzed. Moreover, the intraepithelial lymphocyte (IEL) phenotypical composition was established.. During the suckling period, the supplementation with TGF-β2, EGF and FGF21 showed important signs of intestinal maturation. These results suggest that these molecules, present in breast milk, play a modulatory role in the maturation of the intestinal barrier function and the IEL composition during the suckling period.

Establishment of an induced pluripotent cell line (ABCRIi001-A) from an elderly female for ageing research

Human induced pluripotent stem cells (hiPSCs) hold promises to model and understand human diseases, including those associated with ageing. Here, we describe ABCRIi001-A, a hiPSC line generated from peripheral blood mononuclear cells (PBMCs) of a 79-year-old female enrolled in a study for development of an ageing score (ALFA Score). PBMCs were reprogrammed using three Sendai virus-based reprogramming vectors (hKOS, hc-Myc, and hKlf4). ABCRIi001-A showed normal morphology and karyotype, viral clearance, absence of genomic aberrations, and their pluripotency was confirmed by expression of pluripotency-related markers and their ability to differentiate into the three germ layers. ABCRIi001-A is valuable for ageing-related studies.

A phase 2a trial of brepocitinib for cicatricial alopecia

BackgroundCicatricial alopecias (CA) are chronic, progressive scarring hair-loss conditions. Molecular dysregulation is not fully understood, hindering treatment development. Th1/IFNγ signaling and JAK dysregulation has shown involvement, providing rationale for this phase 2a trial with TYK2/JAK1 inhibitor brepocitinib. MethodsRandomized, placebo-controlled phase 2a trial spanning 52 weeks. Adults (18≥years of age) with lichen planopilaris, frontal fibrosing alopecia, or central centrifugal cicatricial alopecia diagnosis were randomized 3:1 to brepocitinib 45mg daily or placebo for 24 weeks, after which all patients received brepocitinib for another 24 weeks, with a safety follow up 4 weeks later. Lesional scalp biopsies were collected at baseline, week 24, and week 48. Co-primary endpoints were changes in lesional expression of CCL5, changes in lesional expression of fibrosis-related markers, and safety at week 24. ResultsPatients receiving brepocitinib showed significant downregulation in CCL5 expression at week 24 (p=0.004). Enrichment analysis of a subset of fibrosis markers showed trending upregulation in placebo patients (p<0.1). Brepocitinib was well tolerated and improved clinical severity scores. LimitationsSingle-dose regimen, small placebo group. ConclusionBrepocitinib significantly reduces CCL5 expression and was well tolerated at week 24, meeting co-primary endpoints. Brepocitinib reduces inflammatory biomarker expression and improves clinical severity, while maintaining favorable safety profile.

Enzymatic activity of fibroblast activation protein-α is essential for TGF-β1-induced fibroblastic differentiation of human periodontal ligament cells

Human periodontal ligament cells (hPDLCs) contain multipotent postnatal stem cells that can differentiate into PDL fibroblasts, osteoblasts, and cementoblasts. Interaction between the extracellular environment and stem cells is an important factor for differentiation into other progenitor cells. To identify cell surface molecules that induce PDL fibroblastic differentiation, we developed a series of monoclonal antibodies against membrane/ECM molecules. One of these antibodies, an anti-PDL25 antibody, recognizes approximately a 100 kDa protein, and this antigenic molecule accumulates in the periodontal ligament region of tooth roots. By mass spectrometric analysis, we found that the antigenic molecule recognized by the anti-PDL25 antibody is fibroblast activation protein α (FAPα). The expression level of FAPα/PDL25 increased in TGF-β1-induced PDL fibroblasts, and this protein was localized in the cell boundaries and elongated processes of the fibroblastic cells. Ectopic expression of FAPα induced fibroblastic differentiation. In contrast, expression of representative markers for PDL differentiation was decreased by knock down and antibody blocking of FAPα/PDL25. Inhibition of dipeptidyl peptidase activity by a potent FAPα inhibitor dramatically inhibited PDL fibroblastic marker expression but did not affect in cell proliferation and migration.

Experimental and proteomics evidence revealed the protective mechanisms of ShemaZhichuan Liquid in attenuating neutrophilic asthma

BackgroundNeutrophilic asthma (NA) is one of the most important phenotypes of non-Th2 asthma and is often insensitive to glucocorticoid therapy, making current treatment difficult. Shemazhichuan Liquid (SMZCL), a Chinese medicine compound preparation, has unique advantages in the treatment of asthma. However, the underlying mechanisms of SMZCL in treating NA are not fully understood. PurposeThe efficacy and underlying mechanisms of SMZCL on NA were investigated by TMT-labeled quantitative proteomics analysis and in vivo and in vitro experiments. MethodsNA mouse model was constructed by OVA/CFA sensitization followed by a 10-day challenge with 5% OVA. Lung histopathology, leukocyte counts and cell sorting counts, inflammatory cytokines levels, as well as expression of autophagy markers were then assessed. The specific pathways and proteins of SMZCL for treating NA were further illustrated through TMT-based quantitative proteomics. In addition, RAW264.7 cells were induced by LPS to further explore the mechanism of the main active ingredient of SMZCL on autophagy pathway. ResultsIn vivo, SMZCL contributed to attenuating airway inflammation and collagen disposition, markedly reduced the number of leukocytes, especially neutrophils in bronchoalveolar lavage fluid (BALF), as well as decreased IgE and inflammatory cytokine levels (TNF-α, IL-1β, IL-6 and IL-8) in BALF and serum. Besides, SMZCL elevated the levels of LC3 and ATG5 while inhibiting the expression of p62 and mTOR. Mzb1 and Rab3ip were identified as the critical overlapping DEPs whose expression was inhibited by SMZCL and rapamycin. KEGG enrichment analysis showed that necroptosis process was a key pathway for SMZCL to treat NA airway inflammation. IHC and WB results confirmed that SMZCL and rapamycin inhibited the phosphorylation of RIPK1, PIPK3 and MLKL. In vitro, ATG5 and LC3 proteins were obviously increased while p-mTOR expression was inhibited after amygdalin treatment. ConclusionSMZCL attenuated airway inflammation in NA mainly through inhibition of the mTOR pathway, along with inhibition of the necroptosis pathway regulated by the RIPK1/RIPK3/MLKL axis and inhibition of Mzb1 and Rab3ip expression.

Niraparib Enhances and Synergizes with Ganglioside GD2 to Potentiate its Inhibitory Effect on Bladder Cancer Cells.

This study aimed to study the inhibitory effect of niraparib alone or in combination with GD2 specific antibody on Bladder Cancer (BCa). The migration ability of BCa cells was assessed through a scratch assay. CCK-8 assay was performed to evaluate the viability of BCa cells, and Transwell invasion assays were utilized to examine invasive capacity. The expression levels of E-cadherin and vimentin in BCa cells were measured using QRT-PCR. Western blot showed the EMT level to be the lowest in the niraparib+GD2 group. The transwell invasion assay suggested that the invasion ability of BCa cells was weakened in the niraparib+ GD2 group. CCK8 assay indicated that the proliferation ability of BCa cells was decreased. Scratch test suggested that the migration ability of BCa cells was weakened. PCR result showed that the niraparib + GD2 group had the most significant inhibitory effect on mRNA expression of EMT markers. Niraparib combined with a GD2-specific antibody exerted a more prominent inhibitory effect on BCa.

Synergy Between NK Cells and Monocytes in Potentiating Cardiovascular Disease Risk in Severe COVID-19.

Evidence suggests that COVID-19 predisposes to cardiovascular diseases (CVDs). While monocytes/macrophages play a central role in the immunopathogenesis of atherosclerosis, less is known about their immunopathogenic mechanisms that lead to CVDs during COVID-19. Natural killer (NK) cells, which play an intermediary role during pathologies like atherosclerosis, are dysregulated during COVID-19. Here, we sought to investigate altered immune cells and their associations with CVD risk during severe COVID-19. We measured plasma biomarkers of CVDs and determined phenotypes of circulating immune subsets using spectral flow cytometry. We compared these between patients with severe COVID-19 (severe, n=31), those who recovered from severe COVID-19 (recovered, n=29), and SARS-CoV-2-uninfected controls (controls, n=17). In vivo observations were supported using in vitro assays to highlight possible mechanistic links between dysregulated immune subsets and biomarkers during and after COVID-19. We performed multidimensional analyses of published single-cell transcriptome data of monocytes and NK cells during severe COVID-19 to substantiate in vivo findings. During severe COVID-19, we observed alterations in cardiometabolic biomarkers including oxidized-low-density lipoprotein, which showed decreased levels in severe and recovered groups. Severe patients exhibited dysregulated monocyte subsets, including increased frequencies of proinflammatory intermediate monocytes (also observed in the recovered) and decreased nonclassical monocytes. All identified NK-cell subsets in the severe COVID-19 group displayed increased expression of activation and tissue-resident markers, such as CD69 (cluster of differentiation 69). We observed significant correlations between altered immune subsets and plasma oxidized-low-density lipoprotein levels. In vitro assays revealed increased uptake of oxidized-low-density lipoprotein into monocyte-derived macrophages in the presence of NK cells activated by plasma of patients with severe COVID-19. Transcriptome analyses confirmed enriched proinflammatory responses and lipid dysregulation associated with epigenetic modifications in monocytes and NK cells during severe COVID-19. Our study provides new insights into the involvement of monocytes and NK cells in the increased CVD risk observed during and after COVID-19.

Go with the flow: An in vitro model of a mature endothelium for the study of the bioresponse of IV injectable nanomedicines

The first exposure of intravenously (IV) administered nanomedicines in vivo is to endothelial cells (ECs) lining blood vessels. While it is known that in vitro endothelium models to assess responses to circulating nanoparticles require shear stress, there is no consensus on when and how to include it in the experimental design. Our experimental workflow integrates shear stress by featuring a flow-induced mature endothelium (14 days) and a flow-mediated nanoparticle treatment. The mature endothelium model exhibited distinct features that indicated a structurally stable and quiescent monolayer. Upon treatment with iron sucrose under dynamic conditions, there was a lower nanoparticle uptake, lower cytotoxicity, and decreased expression of activation markers compared to the static control. This response was attributed to glycocalyx expression, predominantly observed on the mature endothelium. In conclusion, our proposed in vitro endothelium model can be leveraged to understand the dynamics of IV injectable nanomedicines at the initial nano-bio interface in veins immediately post-injection.

6:2 chlorinated polyfluoroalkyl ether sulfonate (F–53B) induced nephrotoxicity associated with oxidative stress, inflammation and fibrosis in mice

6:2 Chlorinated polyfluoroalkyl ether sulfonate (trade name F–53B) is a substitute for perfluorooctane sulfonate (PFOS) used in the plating industry, and has been found in a range of environmental matrices and livings. There are numerous ways by which it is biotoxic to mammals. The kidneys are critical for maintaining homeostasis. However, little research has been conducted on how F–53B affects the kidneys. In this work, we investigated the renal toxicity of long-term oral F–53B treatment in C57BL/6J mice. Mice were allowed to drink F–53B freely at concentrations of 0, 0.057, 0.57, and 5.7 mg/L for 8 weeks. Renal oxidative stress, inflammation, and fibrosis were detected in mice exposed to F–53B, and the expression of related biochemical markers was significantly altered. Further investigations revealed that the TGF-β1/Smad3 and NF-κB signaling pathways may be associated with F–53B-induced renal fibrotic damage and inflammation. Overall, this study suggested that F–53B causes renal injury possibly via oxidative stress, activating the TGF-β1/Smad3 and NF-κB signaling pathways. This provides a foundation for further research into the harmful mechanism of F–53B in mammals.

Phillyrin prevents sepsis-induced acute lung injury through inhibiting the NLRP3/caspase-1/GSDMD-dependent pyroptosis signaling pathway.

Acute lung injury (ALI) is a severe pulmonary disorder of sepsis with high clinical incidence and mortality. Nucleotide-binding oligomerization domain-like receptor family pyrin domain containing 3 (NLRP3)-cysteinyl aspartate specific proteinase 1-gasdermin D (GSDMD)-dependent pyroptosis of alveolar epithelial cells (AECs) has emerged as a crucial contributor to ALI during sepsis. Phillyrin (PHI), a natural lignan isolated from the traditional Chinese herbal medicine Forsythia suspensa, has been shown to have anti-inflammatory, antioxidant and antiviral properties. However, little is known about the protective role and potential mechanism of PHI in sepsis-induced ALI, and it is uncertain whether the protective effect of PHI in sepsis-induced ALI is connected to pyroptosis. This study aims to examine the preventive effects of PHI on sepsis-induced ALI via the inhibition of NLRP3/caspase-1/GSDMD-mediated pyroptosis in AECs. Our findings demonstrate that preadministration of PHI successfully reduces sepsis-induced pulmonary edema, systemic/pulmonary inflammation, and pulmonary histological damage in lung tissues, bronchoalveolar lavage fluid, and the serum of septic mice. Intriguingly, PHI preadministration suppresses sepsis-induced protein expressions of pyroptosis-specific markers, especially their active forms. In vitro assays show that PHI pretreatment also protects type II AECs (MLE-12) from lipopolysaccharide-induced pyroptosis by preventing the activation of the pyroptosis signaling pathway. The results from molecular docking and surface plasmon resonance reveal that PHI has a significant affinity for direct binding to the GSDMD protein, suggesting that GSDMD is a potential pharmacological target for PHI. In conclusion, PHI can prevent sepsis-triggered ALI by effectively suppressing the activation of the canonical pyroptosis signaling pathway and pyroptosis of AECs.