Honokiol induces apoptosis and autophagy in dexamethasone-resistant T-acute lymphoblastic leukemia CEM-C1 cells.

Objective: To study whether and, if so, how honokiol overcome dexamethasone resistance in DEX-resistant CEM-C1 cells. Methods: We investigated the effect of honokiol (0-20 µM) on cell proliferation, cell cycle, cell apoptosis and autophagy in DEX-resistant CEM-C1 cells and DEX-sensitive CEM-C7 cells. We also determined the role of c-Myc protein and mRNA in the occurrence of T-ALL associated dexamethasone resistance western blot and reverse transcription-qPCR (RT-qPCR) analysis. Results: Cell Counting Kit (CCK)-8 assay shows that DEX-resistant CEM-C1 cell lines were highly resistant to dexamethasone with IC50 of 364.1 ± 29.5 µM for 48 h treatment. However, upon treatment with dexamethasone in combination with 1.5 µM of honokiol for 48 h, the IC50 of CEM-C1 cells significantly decreased to 126.2 ± 12.3 µM, and the reversal fold was 2.88. Conversely, the IC50 of CEM-C7 cells was not changed combination of dexamethasone and honokiol as compared to that of CEM-C7 cells treated with dexamethasone alone. It has been shown that honokiol induced T-ALL cell growth inhibition by apoptosis and autophagy via downregulating cell cycle-regulated proteins (Cyclin E, CDK4, and Cyclin D1) and anti-apoptotic proteins BCL-2 and upregulating pro-apoptotic proteins Bax and led to PARP cleavage. Honokiol may overcome dexamethasone resistance in DEX-resistant CEM-C1 cell lines via the suppression of c-Myc mRNA expression. Conclusion: The combination of honokiol and DEX were better than DEX alone in DEX-resistant CEM-C1 cell lines. Honokiol may regulate T-ALL-related dexamethasone resistance by affecting c-Myc.

[A sericin hydrogel scaffold for sustained dexamethasone release modulates macrophage polarization to promote mandibular bone defect repair in rats].

OBJECTIVE: To evaluate the efficacy of a modified sericin hydrogel scaffold loaded with dexamethasone (SMH-CD/DEX) scaffold for promoting bone defect healing by stimulating anti-inflammatory macrophage polarization. METHODS: The light-curable SMH-CD/DEX scaffold was prepared using dexamethasone-loaded NH2-ß-cyclodextrin (NH2-ß-CD) and sericin hydrogel and characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), biocompatibility assessment and drug release test. THP-1 macrophages incubated with the scaffold were examined for protein expressions of iNOS and Arg-1, mRNA expressions of IL-6, Il-10, Arg-1 and iNOS, and surface markers CD86 and CD206 using Western blotting, RT-qPCR, and flow cytometry. In a co-culture system of human periodontal ligament stem cells (HPDLSCs) and THP-1 macrophages, the osteogenic ability of the stem cells incubated with the scaffold was evaluated by detecting protein expressions of COL1A1 and Runx2 and expressions of ALP, Runx2, OCN and BMP2 mRNA, ALP staining, and alizarin red staining. In a rat model of mandibular bone defect, the osteogenic effect of the scaffold was assessed by observing bone regeneration using micro-CT and histopathological staining. RESULTS: In THP-1 macrophages, incubation with SMH-CD/DEX scaffold significantly enhanced protein expressions of Arg-1 and mRNA expressions of IL-10 and Arg-1 and lowered iNOS protein expression and IL-6 and iNOS mRNA expressions. In the co-culture system, SMH-CD/DEX effectively increased the protein expressions of COL1A1 and Runx2 and mRNA expressions of ALP and BMP2 in HPDLSCs and promoted their osteogenic differentiation. In the rat models, implantation of SMH-CD/DEX scaffold significantly promoted bone repair and bone regeneration in the bone defect. CONCLUSION: The SMH-CD/DEX scaffold capable of sustained dexamethasone release promotes osteogenic differentiation of stem cells and bone defect repair in rats by regulating M2 polarization.

Liraglutide and Naringenin relieve depressive symptoms in mice by enhancing Neurogenesis and reducing inflammation.

Depression is a debilitating mental disease that negatively impacts individuals' lives and society. Novel hypotheses have been recently proposed to improve our understanding of depression pathogenesis. Impaired neuroplasticity and upregulated neuro-inflammation add-on to the disturbance in monoamine neurotransmitters and therefore require novel anti-depressants to target them simultaneously. Recent reports demonstrate the antidepressant effect of the anti-diabetic drug liraglutide. Similarly, the natural flavonoid naringenin has shown both anti-diabetic and anti-depressant effects. However, the neuro-pharmacological mechanisms underlying their actions remain understudied. The study aims to evaluate the antidepressant effects and neuroprotective mechanisms of liraglutide, naringenin or a combination of both. Depression was induced in mice by administering dexamethasone (32 mcg/kg) for seven consecutive days. Liraglutide (200 mcg/kg), naringenin (50 mg/kg) and a combination of both were administered either simultaneously or after induction of depression for twenty-eight days. Behavioral and molecular assays were used to assess the progression of depressive symptoms and biomarkers. Liraglutide and naringenin alone or in combination alleviated the depressive behavior in mice, manifested by decrease in anxiety, anhedonia, and despair. Mechanistically, liraglutide and naringenin improved neurogenesis, decreased neuroinflammation and comparably restored the monoamines levels to that of the reference drug escitalopram. The drugs protected mice from developing depression when given simultaneously with dexamethasone. Collectively, the results highlight the usability of liraglutide and naringenin in the treatment of depression in mice and emphasize the different pathways that contribute to the pathogenesis of depression.

Channel Expansion in the Ligand-Binding Domain of the Glucocorticoid Receptor Contributes to the Activity of Highly Potent Glucocorticoid Analogues.

Glucocorticoids (GCs) act through the glucocorticoid receptor (GR) and are commonly used as anti-inflammatory and immunosuppressant medications. Chronic GC use has been linked with unwanted complications such as steroid-induced diabetes mellitus (SIDM), although the mechanisms for these effects are not completely understood. Modification of six GC parent molecules with 2-mercaptobenzothiazole resulted in consistently less promoter activity in transcriptional activation assays using a 3xGRE reporter construct while constantly reducing inflammatory pathway activity. The most selective candidate, DX1, demonstrated a significant reduction (87%) in transactivation compared to commercially available dexamethasone. DX1 also maintained 90% of the anti-inflammatory potential of dexamethasone while simultaneously displaying a reduced toxicity profile. Additionally, two novel and highly potent compounds, DX4 and PN4, were developed and shown to elicit similar mRNA expression at attomolar concentrations that dexamethasone exhibits at nanomolar dosages. To further explain these results, Molecular Dynamic (MD) simulations were performed to examine structural changes in the ligand-binding domain of the glucocorticoid receptor in response to docking with the top ligands. Differing interactions with the transcriptional activation function 2 (AF-2) region of the GR may be responsible for lower transactivation capacity in DX1. DX4 and PN4 lose contact with Arg611 due to a key interaction changing from a stronger hydrophilic to a weaker hydrophobic one, which leads to the formation of an unoccupied channel at the location of the deacylcortivazol (DAC)-expanded binding pocket. These findings provide insights into the structure-function relationships important for regulating anti-inflammatory activity, which has implications for clinical utility.

The glucocorticoid dexamethasone alleviates allergic inflammation through a mitogen-activated protein kinase phosphatase-1-dependent mechanism in mice.

Glucocorticoids are widely used in the treatment of allergic and inflammatory diseases. Glucocorticoids have a widespread action on gene expression resulting in their pharmacological actions and also an array of adverse effects which limit their clinical use. It remains, however, to be studied which target gene effects are essential for the anti-allergic activity of glucocorticoids. Mitogen-activated protein kinase phosphatase-1 (MKP-1) inhibits proinflammatory signalling by suppressing the activity of mitogen activated protein kinase (MAP kinase) pathways. MKP-1 is one of the anti-inflammatory genes whose expression is enhanced by glucocorticoids. In the present study, we aimed to investigate the role of MKP-1 in the therapeutic effects of the glucocorticoid dexamethasone in acute allergic reaction. The effects of dexamethasone were studied in wild-type and MKP-1 deficient mice. The mice were first sensitized to ovalbumin, and the allergic reaction was then induced by a subcutaneous ovalbumin injection in the hind paw. Inflammatory edema was quantified with plethysmometer and expression of inflammatory factors was measured by quantitative reverse transcription polymerase chain reaction (RT-PCR). Dexamethasone reduced the ovalbumin-induced paw edema at 1.5, 3 and 6 h time points in wild-type mice by 70%, 95% and 89%, respectively. The effect was largely abolished in MKP-1 deficient mice. Furthermore, dexamethasone significantly attenuated the expression of ovalbumin-induced inflammatory factors cyclooxygenase-2 (COX-2); inducible nitric oxide synthase (iNOS); interleukins (IL) 1ß, 6 and 13; C-C motif chemokine 11 (CCL-11); tumour necrosis factor (TNF) and thymic stromal lymphopoietin (TSLP) in wild-type mice by more than 40%. In contrast, in MKP-1 deficient mice dexamethasone had no effect or even enhanced the expression of these inflammatory factors. The results suggest that dexamethasone alleviates allergic inflammation through an MKP-1-dependent mechanism. The results also demonstrate MKP-1 as an important conveyor of the favourable glucocorticoid effects in ovalbumin-induced type I allergic reaction. Together with previous findings, the present study supports the concept of MKP-1 enhancing compounds as potential novel anti-inflammatory and anti-allergic drugs.

Differential Effects of Benzalkonium Chloride on Human Trabecular Meshwork Cells Not Treated or Treated with Transforming Growth Factor-ß2 or Dexamethasone.

Purpose: The objective of the present study was to evaluate the effects of low concentrations of benzalkonium chloride (BAC) (10-7%, 10-6%, or 10-5%) on healthy and glaucomatous human trabecular meshwork (HTM) cells. For this purpose, we used in vitro models replicating a healthy HTM and HTM with primary open-angle glaucoma (POAG) or steroid-induced glaucoma (SG) using two-dimensional (2D) cultures of HTM cells not treated or treated with a 5 ng/mL solution of transforming growth factor-ß2 or 250 nM dexamethasone (DEX). Methods: Analyses were carried out for (1) the intercellular affinity function of 2D HTM monolayers, as determined by transepithelial electrical resistance (TEER) measurements; (2) cell viability; (3) cellular metabolism by using a Seahorse bioanalyzer; and (4) expression of extracellular matrix (ECM) molecules, an ECM modulator, and cell junction-related molecules. Results: In the absence and presence of BAC (10-7% or 10-5%), intercellular affinity function determined by TEER and cellular metabolic activities were significantly and dose dependently affected in both healthy and glaucomatous HTM cells despite the fact that there was no significant decrease in cell viabilities. However, the effects based on TEER values were significantly greater in the healthy HTM. The mRNA expression of several molecules that were tested was not substantially modulated by these concentrations of BAC. Conclusions: The findings reported herein suggest that low concentrations of BAC may have unfavorable adverse effects on cellular metabolic capacity by inducing increases in the intercellular affinity properties of the HTM, but those effects of BAC were different in healthy and glaucomatous HTM cells.

Synergistic effects of mesenchymal stem cell-derived extracellular vesicles and dexamethasone on macrophage polarization under inflammatory conditions.

The undesirable inflammation and the excessive M1 macrophage activity may lead to inflammatory diseases. Corticosteroids and stem cell therapy are used in clinical practice to promote anti-inflammatory responses. However, this protocol has limitations and is associated with numerous side effects. In this study, the synergistic anti-inflammatory effects of dexamethasone (Dex) and mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) were evaluated to enhance the polarization of M1 inflammatory macrophages into the anti-inflammatory (M2) phenotype. Hence, we designed different combinations of Dex and EVs using three methods, including EVs isolated from Dex-preconditioned MSCs (Pre-Dex-EVs), EVs loaded with Dex (L-Dex-EVs), and EVs and Dex co-administration (Dex + EVs). All designed EVs had a significant effect on reducing the expression of M1-related genes (iNOS, Stat1, and IRF5), cytokines (IL6 and TNF-a), and CD markers (CD86) in lipopolysaccharide-stimulated macrophages. On the other hand, these combinations promoted the expression of alternative-activated M2-related genes (Arg-1, Stat6, and IRF4), cytokine (IL10), and CD markers (CD206).The combination of Dex and MSC-EVs enhances the effectiveness of both and synergistically promotes the conversion of inflammatory macrophages into an anti-inflammatory state.

Ameliorated Skin Inflammation through the Synergistic Effect of Gold Nanorod-Dexamethasone and Photothermal Therapy.

Psoriasis, a prevalent chronic inflammatory skin ailment affecting approximately 2-3% of the global population, is characterized by persistent symptoms. Dexamethasone, a primary corticosteroid for treating psoriasis, demonstrates notable efficacy; however, its limited skin permeation results in documented adverse effects. To address this, the presented study employed a novel strategy to conjugate gold nanorod and dexamethasone and evaluate their potential for mitigating psoriatic inflammation using an imiquimod-induced mouse model and human skin cells. Our findings revealed enhanced cutaneous penetration of gold nanorod and dexamethasone conjugates compared with that of dexamethasone, owing to superior skin penetration. Gold nanorod and dexamethasone conjugates demonstrated an optimal pharmacological impact at minimal dosages without toxicity during extended use. To further enhance the effectiveness of gold nanorod and dexamethasone conjugates, 808 nm near-infrared laser irradiation, which reacts to gold, was additionally applied to achieve thermal elevation to expedite drug skin penetration. Supplementary laser irradiation at 808 nm significantly ameliorated psoriatic symptoms following deep gold nanorod and dexamethasone conjugates penetration. This corresponded with restored peroxisome proliferator-activated receptor-γ levels and accelerated dexamethasone release from the gold nanorod and dexamethasone conjugates complex. These findings highlight the potential of gold nanorod and dexamethasone conjugates to enhance drug penetration through dermal layers, thereby aiding psoriasis treatment. Moreover, its compatibility with photothermal therapy offers prospects for novel therapeutic interventions across various inflammatory skin disorders.

Dexamethasone Inhibits White Adipose Tissue Browning.

White adipose tissue (WAT) regulates energy balance through energy storage, adipokines secretion and the thermogenesis process. Beige adipocytes are responsible for WAT thermogenesis. They are generated by adipogenesis or transdifferentiation during cold or ß3-adrenergic agonist stimulus through a process called browning. Browning has gained significant interest for to its preventive effect on obesity. Glucocorticoids (GCs) have several functions in WAT biology; however, their role in beige adipocyte generation and WAT browning is not fully understood. The aim of our study was to determine the effect of dexamethasone (DXM) on WAT thermogenesis. For this purpose, rats were treated with DXM at room temperature (RT) or cold conditions to determine different thermogenic markers. Furthermore, the effects of DXM on the adipogenic potential of beige precursors and on mature beige adipocytes were evaluated in vitro. Our results showed that DXM decreased UCP-1 mRNA and protein levels, mainly after cold exposure. In vitro studies showed that DXM decreased the expression of a beige precursor marker (Ebf2), affecting their ability to differentiate into beige adipocytes, and inhibited the thermogenic response of mature beige adipocytes (Ucp-1, Dio2 and Pgc1α gene expressions and mitochondrial respiration). Overall, our data strongly suggest that DXM can inhibit the thermogenic program of both retroperitoneal and inguinal WAT depots, an effect that could be exerted, at least partially, by inhibiting de novo cell generation and the thermogenic response in beige adipocytes.

Aerosol inhalation of inflammatory cells-targeted dendrimer-dexamethasone conjugate for efficient allergic asthma therapy.

Allergic asthma (AA) is a common breathing disorder clinically characterized by the high occurrence of acute and continuous inflammation. However, the current treatment options for AA are lacking in effectiveness and diversity. In this study, we determined that the cell membrane receptor of gamma-glutamyl transferase (GGT) was highly overexpressed on the inflammatory cells that infiltrate the pulmonary tissues in AA cases. Therefore, we developed a GGT-specific dendrimer-dexamethasone conjugate (GSHDDC) that could be administered via aerosol inhalation to treat AA in a rapid and sustained manner. The GSHDDC was fabricated by the covalent attachment of 6-hydroxyhexyl acrylate-modified dexamethasone to polyamidoamine dendrimers via a carbonic ester linkage and the amino Michael addition, followed by the surface modification of the dendrimers with the GGT substrate of glutathione. After aerosol inhalation by the AA mice, the small particle-sized GSHDDC could easily diffuse into pulmonary alveoli and touch with the inflammatory cells via the glutathione ligand/GGT receptor-mediated recognition. The overexpressed GGT on the surface of inflammatory cells then triggers the gamma-glutamyl transfer reactions of glutathione to generate positively charged primary amines, thereby inducing rapid cationization-mediated cellular endocytosis into the inflammatory cells. The dexamethasone was gradually released by the intracellular enzyme hydrolysis, enabling sustained anti-inflammatory effects (e.g., reducing eosinophil infiltration, decreasing the levels of inflammatory factors) in the ovalbumin-induced AA mice. This study demonstrates the effectiveness of an inhalational and active inflammatory cells-targeted dendrimer-dexamethasone conjugate for efficient AA therapy.

Pinoresinol diglucoside mitigates dexamethasone-induced osteoporosis and chondrodysplasia in zebrafish.

BACKGROUND: The global increase in the aging population has led to a higher incidence of osteoporosis among the elderly. OBJECTIVE: This study aimed to evaluate the protective properties of pinoresinol diglucoside (PDG), an active constituent of Eucommia ulmoides, against dexamethasone-induced osteoporosis and chondrodysplasia. METHODS: A zebrafish model of osteoporosis was established by exposing larval zebrafish to dexamethasone. The impact of PDG on bone mineralization was assessed through alizarin red and calcein staining. Alkaline phosphatase activity was quantified to evaluate osteoblast function. The influence of PDG on chondrogenesis was estimated using alcian blue staining. Fluorescence imaging and motor behavior analysis were employed to assess the protective effect of PDG on the structure and function of dexamethasone-induced skeletal teratogenesis. qPCR determined the expression of osteogenesis and Wnt signaling-related genes. Molecular docking was used to assess the potential interactions between PDG and Wnt receptors. RESULTS: PDG significantly increased bone mineralization and corrected spinal curvature and cartilage malformations in the zebrafish model. Furthermore, PDG enhanced swimming abilities compared to the model group. PDG mitigated dexamethasone-induced skeletal abnormalities in zebrafish by upregulating Wnt signaling, showing potential interaction with Wnt receptors FZD2 and FZD5. CONCLUSION: PDG mitigates dexamethasone-induced osteoporosis and chondrodysplasia by promoting bone formation and activating Wnt signaling.

Intracellular Zn2+ promotes extracellular matrix remodeling in dexamethasone-treated trabecular meshwork.

Given the widespread application of glucocorticoids in ophthalmology, the associated elevation of intraocular pressure (IOP) has long been a vexing concern for clinicians, yet the underlying mechanisms remain inconclusive. Much of the discussion focuses on the extracellular matrix (ECM) of trabecular meshwork (TM). It is widely agreed that glucocorticoids impact the expression of matrix metalloproteinases (MMPs), leading to ECM deposition. Since Zn2+ is vital for MMPs, we explored its role in ECM alterations induced by dexamethasone (DEX). Our study revealed that in human TM cells treated with DEX, the level of intracellular Zn2+ significantly decreased, accompanied by impaired extracellular Zn2+ uptake. This correlated with changes in several Zrt-, Irt-related proteins (ZIPs) and metallothionein. ZIP8 knockdown impaired extracellular Zn2+ uptake, but Zn2+ chelation did not affect ZIP8 expression. Resembling DEX's effects, chelation of Zn2+ decreased MMP2 expression, increased the deposition of ECM proteins, and induced structural disarray of ECM. Conversely, supplementation of exogenous Zn2+ in DEX-treated cells ameliorated these outcomes. Notably, dietary zinc supplementation in mice significantly reduced DEX-induced IOP elevation and collagen content in TM, thereby rescuing the visual function of the mice. These findings underscore zinc's pivotal role in ECM regulation, providing a novel perspective on the pathogenesis of glaucoma.NEW & NOTEWORTHY Our study explores zinc's pivotal role in mitigating extracellular matrix dysregulation in the trabecular meshwork and glucocorticoid-induced ocular hypertension. We found that in human trabecular meshwork cells treated with dexamethasone, intracellular Zn2+ significantly decreased, accompanied by impaired extracellular Zn2+ uptake. Zinc supplementation rescues visual function by modulating extracellular matrix proteins and lowering intraocular pressure, offering a direction for further exploration in glaucoma management.

Improvement of inhaled paraquat induced lung and systemic inflammation, oxidative stress and memory changes by safranal.

The effects of safranal and pioglitazone alone and their combination on inhaled paraquat (PQ)-induced systemic oxidative stress and inflammation as well as behavioral changes were examined in rats. In this study, animals were exposed to saline (Ctrl) or PQ (PQ groups) aerosols. PQ exposed animals were treated with dexamethasone, 0.8 and 3.2 mg/kg/day safranal (Saf-L and Saf-H), 5 mg/kg/day pioglitazone (Pio), and Saf-L + Pio for 16 days during PQ exposure period. PQ group showed increased numbers of total and differential WBCs in blood and bronchoalveolar lavage fluid (BALF), increased malondialdehyde (MDA), in the serum BALF and brain reduced thiol, catalase (CAT), and superoxide dismutase (SOD) levels compared to the control group (for all, p < 0.001). The escape latency and traveled distance were enhanced, but the time spent in the target quadrant in the probe day and the latency to enter the dark room 3, 24, 48, and 72 h after receiving an electrical shock, (in the shuttle box test) were decreased in the PQ group (p < 0.05 to P < 0.001). In all treated groups, all measure values were improved compared to PQ group (p < 0.05 to p < 0.001). In combination treated group of Saf-L + Pio, most measured values were more improved than the Saf-L and Pio groups (p < 0.05 to p < 0.001). Saf and Pio improved PQ-induced changes similar to dexamethasone but the effects produced by combination treatments of Saf-L + Pio were more prominent than Pio and Saf-L alone, suggesting a potentiating effect for the combination of the two agents.

Anti-fibrotic and anti-stricture effects of biodegradable biliary stents braided with dexamethasone-impregnated sheath/core structured monofilaments.

Endoscopic biliary stent insertion has been widely used for the treatment of benign biliary stricture (BBS). Thus, the development of stent materials in the perspectives of structure, mechanical properties, and biocompatibility has been also studied. However, conventional metal and plastic stents have several disadvantages, such as repeated procedures to remove or exchange them, dislodgment, restenosis, biocompatibility, and poor mechanical properties. Sustainable effectiveness, attenuation and prevention of fibrosis, and biocompatibility are key factors for the clinical application of stents to BBS treatment. In addition, loading drugs could show synergistic effects with stents' own performance. We developed a dexamethasone-eluting biodegradable stent (DBS) consisting of a sheath/core structure with outstanding mechanical properties and sustained release of dexamethasone, which maintained its functions in a BBS duct over 12 weeks in a swine model. The insertion of our DBS not only expanded BBS areas but also healed secondary ulcers as a result of the attenuation of fibrosis. After 16 weeks from the insertion, BBS areas were totally improved, and the DBS was degraded and thoroughly disappeared without re-intervention for stent removal. Our DBS would be an effective clinical tool for non-vascular diseases. STATEMENT OF SIGNIFICANCE: This study describes the insertion of a drug-eluting biodegradable stent (DBS) into the bile duct. The sheath/core structure of DBS confers substantial durability and a sustained drug release profile. Drug released from the DBS exhibited anti-fibrotic effects without inflammatory responses in both in vitro and in vivo experiments. The DBS maintained its function over 12 weeks after insertion into the common bile duct, expanding benign biliary stricture (BBS) and reducing inflammation to heal secondary ulcers in a swine BBS model. After 16 weeks from the DBS insertion, the DBS thoroughly disappeared without re-intervention for stent removal, resulting in totally improved BBS areas. Our findings not only spotlight the understanding of the sheath/core structure of the biodegradable stent, but also pave the way for the further application for non-vascular diseases.

The effect of lentinan on dexamethasone-induced immunosuppression in mice.

The immune system acts as a vital defense barrier against pathogenic invasions, and its stable operation is crucial for maintaining body health. Nevertheless, various natural or artificial factors can compromise the body's immune function, leading to immunosuppression, which may interfere with the efficacy of vaccination and increase the susceptibility of the body to disease-causing pathogens. In an effort to ensure successful vaccinations and improve overall physical well-being, the search for appropriate immune regulators to enhance immunity is of paramount importance. Lentinan (LNT) has a significant role in immune regulation and vaccine adjuvants. In the present study, we constructed an immunosuppressive model using dexamethasone (DEX) and demonstrated that LNT could significantly improved antibody levels in immunosuppressive mice and stimulated T-lymphocyte proliferation and differentiation in intestinal Peyer's patches. LNT also increased the production of secretory immunoglobulin A (sIgA) in the duodenal fluid, the number of goblet cells, and the proportion of mucin area. Moreover, LNT modulated the intestinal microbiota and increased the production of short-chain fatty acids. Additionally, LNT promoted the proliferation, differentiation, and pro-inflammatory cytokines production of DEX-treated splenic T lymphocytes in vitro. Thus, the present study highlights the potential of LNT in reversing immunosuppression and avoiding the failure of vaccination.

Dexamethasone protects against asthma via regulating Hif-1α-glycolysis-lactate axis and protein lactylation.

PURPOSE: Asthma can not be eradicated till now and its control primarily relies on the application of corticosteroids. Recently, glycolytic reprogramming has been reportedly contributed to asthma, this study aimed to reveal whether the effect of corticosteroids on asthma control is related to their regulation of glycolysis and glycolysis-dependent protein lactylation. METHODS: Ovalbumin (OVA) aeroallergen was used to challenge mice and stimulate human macrophage cell line THP-1 following dexamethasone (DEX) treatment. Airway hyperresponsiveness, airway inflammation, the expressions of key glycolytic enzymes and pyroptosis markers, the level of lactic acid, real-time glycolysis and oxidative phosphorylation (OXPHOS), and protein lactylation were analyzed. RESULTS: DEX significantly attenuated OVA-induced eosinophilic airway inflammation, including airway hyperresponsiveness, leukocyte infiltration, goblet cell hyperplasia, Th2 cytokines production and pyroptosis markers expression. Meanwhile, OVA-induced Hif-1α-glycolysis axis was substantially downregulated by DEX, which resulted in low level of lactic acid. Besides, key glycolytic enzymes in the lungs of asthmatic mice were notably co-localized with F4/80-positive macrophages, indicating metabolic shift to glycolysis in lung macrophages during asthma. This was confirmed in OVA-stimulated THP-1 cells that DEX treatment resulted in reductions in pyroptosis, glycolysis and lactic acid level. Finally, protein lactylation was found significantly increased in the lungs of asthmatic mice and OVA-stimulated THP-1 cells, which were both inhibited by DEX. CONCLUSION: Our present study revealed that the effect of DEX on asthma control was associated with its suppressing of Hif-1α-glycolysis-lactateaxis and subsequent protein lactylation, which may open new avenues for the therapy of eosinophilic asthma.

DT-13 attenuates inflammation by inhibiting NLRP3-inflammasome related genes in RAW264.7 macrophages.

Plant derived saponins or other glycosides are widely used for their anti-inflammatory, antioxidant, and anti-viral properties in therapeutic medicine. In this study, we focus on understanding the function of the less known steroidal saponin from the roots of Liriope muscari L. H. Bailey - saponin C (also known as DT-13) in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages in comparison to the well-known saponin ginsenoside Rk1 and anti-inflammatory drug dexamethasone. We proved that DT-13 reduces LPS-induced inflammation by inhibiting nitric oxide (NO) production, interleukin-6 (IL-6) release, cycloxygenase-2 (COX-2), tumour necrosis factor-alpha (TNF-α) gene expression, and nuclear factor kappa-B (NFκB) translocation into the nucleus. It also inhibits the inflammasome component NOD-like receptor family pyrin domain containing protein 3 (NLRP3) regulating the inflammasome activation. This was supported by the significant inhibition of caspase-1 and interleukin-1 beta (IL-1ß) expression and release. This study demonstrates the anti-inflammatory effect of saponins on LPS-stimulated macrophages. For the first time, an in vitro study shows the attenuating effect of DT-13 on NLRP3-inflammasome activation. In comparison to the existing anti-inflammatory drug, dexamethasone, and triterpenoid saponin Rk1, DT-13 more efficiently inhibits inflammation in the applied cell culture model. Therefore, DT-13 may serve as a lead compound for the development of new more effective anti-inflammatory drugs with minimised side effects.

An injectable thermosensitive hyaluronic acid/pluronic F-127 hydrogel for deep penetration and combination therapy of frozen shoulder.

Frozen shoulder (FS) is a common and progressive shoulder disorder that causes glenohumeral joint stiffness, characterized by inflammation and fibrosis. The treatment options are quite limited, and the therapeutic response is hindered by the fibrous membrane formed by excessive collagen and the rapid removal by synovial fluid. To address these challenges, we designed a hyaluronic acid/Pluronic F-127 (HP)-based injectable thermosensitive hydrogel as a drug carrier loaded with dexamethasone and collagenase (HPDC). We screened for an optimal HP hydrogel that can sustain drug release for approximately 10 days both in vitro and in vivo. In the meanwhile, we found that HP hydrogel could inhibit the proliferation and diminish the adhesion capacity of rat synovial cells induced by transforming growth factor-ß1. Furthermore, using an established immobilization rat model of FS, intra-articular injection of HPDC significantly improved joint range of motion compared to medication alone. Relying on sustained drug release, the accumulated collagen fibers were degraded by collagenase to promote the deep delivery of dexamethasone. These findings showed a positive combined treatment effect of HPDC, providing a novel idea for the comprehensive treatment of FS.

Echinacoside alleviates glucocorticoid induce osteonecrosis of femoral head in rats through PI3K/AKT/FOXO1 pathway.

Steroid-induced osteonecrosis of the femoral head (SONFH), caused by glucocorticoid (GC) administration, is known to exhibit a high incidence worldwide. Although osteoblast apoptosis has been reported as an important cytological basis of SONFH, the precise mechanism remains elusive. Echinacoside (Ech), a natural phenylethanoid glycoside, exerts multiple beneficial effects, such as facilitation of cell proliferation and anti-inflammatory and anticancer activities. Herein, we aimed to explore the regulatory mechanism underlying glucocorticoid-induced osteoblast apoptosis and determine the protective efficacy of Ech against SONFH. We comprehensively surveyed multiple public databases to identify SONFH-related genes. Using bioinformatics analysis, we identified that the PI3K/AKT/FOXO1 signaling pathway was most strongly associated with SONFH. We examined the protective effect of Ech against SONFH using in vivo and in vitro experiments. Specifically, dexamethasone (Dex) decreased p-PI3K and p-AKT levels, which were reversed following Ech addition. Validation of the PI3K inhibitor (LY294002) and molecular docking of Ech and PI3K/AKT further indicated that Ech could directly enhance PI3K/AKT activity to alleviate Dex-induced inhibition. Interestingly, Dex upregulated the expression of FOXO1, Bax, cleaved-caspase-9, and cleaved-caspase-3 and enhanced MC3T3-E1 apoptosis; application of Ech and siRNA-FOXO1 reversed these effects. In vitro, Ech decreased the number of empty osteocytic lacunae, reduced TUNEL and FOXO1 positive cells, and improved bone microarchitecture. Our results provide robust evidence that PI3K/AKT/FOXO1 plays a crucial role in the development of SONFH. Moreover, Ech may be a promising candidate drug for the treatment of SONFH.

Dexamethasone upregulates macrophage PIEZO1 via SGK1, suppressing inflammation and increasing ROS and apoptosis.

The side effects of high-dose dexamethasone in anti-infection include increased ROS production and immune cell apoptosis. Dexamethasone effectively activates serum/glucocorticoid-regulated kinase 1 (SGK1), which upregulates various ion channels by activating store-operated calcium entry (SOCE), leading to Ca2+ oscillations. PIEZO1 plays a crucial role in macrophages' immune activity and function, but whether dexamethasone can regulate PIEZO1 by enhancing SOCE via SGK1 activation remains unclear. The effects of dexamethasone were assessed in a mouse model of sepsis, and primary BMDMs and the RAW264.7 were treated with overexpression plasmids, siRNAs, or specific activators or inhibitors to examine the relationships between SGK1, SOCE, and PIEZO1. The functional and phenotypic changes of mouse and macrophage models were detected. The results indicate that high-dose dexamethasone upregulated SGK1 by activating the macrophage glucocorticoid receptor, which enhanced SOCE and subsequently activated PIEZO1. Activation of PIEZO1 resulted in Ca2+ influx and cytoskeletal remodelling. The increase in intracellular Ca2+ mediated by PIEZO1 further increased the activation of SGK1 and ORAI1/STIM1, leading to intracellular Ca2+ peaks. In the context of inflammation, activation of PIEZO1 suppressed the activation of TLR4/NFκB p65 in macrophages. In RAW264.7 cells, PIEZO1 continuous activation inhibited the change in mitochondrial membrane potential, accelerated ROS accumulation, and induced autophagic damage and cell apoptosis in the late stage. CaMK2α was identified as a downstream mediator of TLR4 and PIEZO1, facilitating high-dose dexamethasone-induced macrophage immunosuppression and apoptosis. PIEZO1 is a new glucocorticoid target to regulate macrophage function and activity. This study provides a theoretical basis for the rational use of dexamethasone.