Metalloproteinase Activity Research Articles

Abstract A028: Precise monitoring of osteosarcoma patients by noninvasive assessment of matrix metalloproteinases activities in plasma extracellular vesicles

Abstract Osteosarcoma (OS) is the most prevalent bone cancer in children, characterized by aggressive primary tumors and high metastatic potential. Current treatment protocols involve a combination of neoadjuvant chemotherapy, surgery, and adjuvant chemotherapy, tailored based on histological grade and metastatic risk. Identifying metastatic lesions through routine imaging significantly impacts treatment outcomes, highlighting the need for innovative diagnostic tools for timely monitoring. Noninvasive in vitro diagnostics, analyzing circulating tumor components like tumor-derived extracellular vesicles (EVs), would offer promise for monitoring treatment responses in pediatric sarcomas. Matrix metalloproteinases (MMPs) play a crucial role in tumor progression by degradation of the extracellular matrix; thus, recent research has focused on understanding their functional roles by profiling their proteolytic activities. Tumor EVs are crucial in transporting MMPs to facilitate invasion and metastasis, making them suitable surrogate for noninvasive profiling of MMP activities. Herein, we introduce an innovative OS EV MMP Activity Assay to assess MMP activities in OS patients noninvasively. This assay evaluates six different combinations of surface markers on OS EVs and OS-associated MMPs, generating a unique profile for each patient. Using a logistic regression model, an OS EV MMP Activity Score is generated from the most effective combinations. This Score has shown excellent performance in distinguishing between metastatic and localized OS. In a longitudinal study of six OS patients undergoing treatment, the Score strongly correlated with treatment response as measured by routine imaging. The assay represents a significant advancement in harnessing tumor EVs for diagnostics, providing a noninvasive, quantitative, and reliable approach to assess MMP activities in solid tumors. Citation Format: Junseok Lee, You-Ren Ji, Hyoyong Kim, Zhao Chen, Ryan Zhang, Diego Banuet, Yusef Mathkour, Kenny Vo, Steven Jonas, Yazhen Zhu, Hsian-Rong Tseng. Precise monitoring of osteosarcoma patients by noninvasive assessment of matrix metalloproteinases activities in plasma extracellular vesicles [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A028.

Serum integrin accumulation during asthma exacerbation: The role of matrix metalloproteinases.

The inflammation caused by asthma exacerbation can lead to permanent changes in the airways and loss of lung function. Integrins are membrane receptors that interact with components of the extracellular matrix and cell adhesion molecules. It is known that these receptors can be found in soluble form in some conditions such as asthma, but it is unknown if exacerbation during asthma leads to soluble integrins. Our results indicated that asthma patients showed higher levels of soluble α1, α2, and β2 integrin subunits in their serum compared to controls, as confirmed by both ELISA and western blot. During asthma exacerbation, the levels of α2 and β2 integrin subunits increased even more compared to non-exacerbation and controls, while the α1 integrin subunit decreased. Western blot analysis identified two β2 integrin subunits, one at 75 kDa and another at 120 kDa; the 120 kDa subunit increased during asthma exacerbation. The activity of matrix metalloproteinase 9 (MMP9) increased during exacerbation, while MMP2 remained unchanged. Lower forced expiratory volume in 1 second (FEV1) values were associated with higher expression levels of α2, β1, and β2 integrin subunits. Active and latent MMP9 were correlated with the levels of the β2 integrin subunit, which means that at low levels of active and latent MMP9, there are lower levels of β2 integrin subunit. In conclusion, asthma exacerbation leads to the presence of soluble integrins, particularly the β2 subunit, most likely due to MMP9-induced proteolytic cleavage.

Endogenous/exogenous stimuli‐responsive smart hydrogels for diabetic wound healing

AbstractDiabetes significantly impairs the body's wound‐healing capabilities, leading to chronic, infection‐prone wounds. These wounds are characterized by hyperglycemia, inflammation, hypoxia, variable pH levels, increased matrix metalloproteinase activity, oxidative stress, and bacterial colonization. These complex conditions complicate effective wound management, prompting the development of advanced diabetic wound care strategies that exploit specific wound characteristics such as acidic pH, high glucose levels, and oxidative stress to trigger controlled drug release, thereby enhancing the therapeutic effects of the dressings. Among the solutions, hydrogels emerge as promising due to their stimuli‐responsive nature, making them highly effective for managing these wounds. The latest advancements in mono/multi‐stimuli‐responsive smart hydrogels showcase their superiority and potential as healthcare materials, as highlighted by relevant case studies. However, traditional wound dressings fall short of meeting the nuanced needs of these wounds, such as adjustable adhesion, easy removal, real‐time wound status monitoring, and dynamic drug release adjustment according to the wound's specific conditions. Responsive hydrogels represent a significant leap forward as advanced dressings proficient in sensing and responding to the wound environment, offering a more targeted approach to diabetic wound treatment. This review highlights recent advancements in smart hydrogels for wound dressing, monitoring, and drug delivery, emphasizing their role in improving diabetic wound healing. It addresses ongoing challenges and future directions, aiming to guide their clinical adoption.

Interpenetrating Polymer Network Hydrogels with Tunable Viscoelasticity and Proteolytic Cleavability to Direct Stem Cells In Vitro.

The dynamic nature of cellular microenvironments, regulated by the viscoelasticity and enzymatic cleavage of the extracellular matrix, remains challenging to emulate in engineered synthetic biomaterials. To address this, a novel platform of cell-instructive hydrogels is introduced, composed of two concurrently forming interpenetrating polymer networks (IPNs). These IPNs consist of the same basic building blocks - four-armed poly(ethylene glycol) and the sulfated glycosaminoglycan (sGAG) heparin - are cross-linked through either chemical or physical interactions, allowing for precise and selective tuning of the hydrogel's stiffness, viscoelasticity, and proteolytic cleavability. The studies of the individual and combined effects of these parameters on stem cell behavior revealed that human mesenchymal stem cells exhibited increased spreading and Yes-associated protein transcriptional activity in more viscoelastic and cleavable sGAG-IPN hydrogels. Furthermore, human induced pluripotent stem cell (iPSC) cysts displayed enhanced lumen formation, growth, and pluripotency maintenance when cultured in sGAG-IPN hydrogels with higher viscoelasticity. Inhibition studies emphasized the pivotal roles of actin dynamics and matrix metalloproteinase activity in iPSC cyst morphology, which varied with the viscoelastic properties of the hydrogels. Thus, the introduced sGAG-IPN hydrogel platform offers a powerful methodology for exogenous stem cell fate control.

Endogenous Tissue Inhibitor of Metalloproteinase-2 Levels Are Associated With High-Quality Neat Semen but Unrelated to Sperm Cryoresistance in Bulls.

Tissue Inhibitor of Metalloproteinase-2 (TIMP-2) is part of the tissue inhibitors of the metalloproteinases (TIMPs) family. Its primary function is to regulate the activity of matrix metalloproteinases (MMPs) across various tissues, including those of the reproductive system. This study aimed to quantify the natural levels of TIMP-2 in seminal plasma (SP) and sperm membrane (SM) of bulls, explore potential associations between TIMP-2 levels and semen quality parameters, and examine the relationship between TIMP-2 levels and sperm cryoresistance in bulls. Thirty semen samples from Frieswal breeding bulls were categorized into two groups based on their initial progressive motility (IPM): Good (IPM ≥ 70%; n = 21) and Poor (IPM ≤ 40%; n = 9). The samples were evaluated for their quality parameters at the fresh stage, and TIMP-2 levels were measured in SP and SM using a bovine-specific ELISA kit. Following cryopreservation of Good samples (n = 21), post-thaw motility (PTM) was used to further classify samples into Freezeable (PTM ≥ 50%; n = 14) and Non-Freezable (PTM < 50%; n = 7) groups. In frozen-thawed samples, sperm attributes, kinetics, and functional parameters were assessed, and the results were correlated with retrospective TIMP-2 levels of SP/SM. Our study revealed that the quantified levels of TIMP-2 ranged from 100.27 to 535.95 ng/L in SP and from 0 to 115.78 ng/10 million spermatozoa in SM. TIMP-2 levels in both SP and SM were significantly higher in Good ejaculates compared to Poor ejaculates (p < 0.01). Furthermore, total TIMP-2 levels in the SP/SM of semen samples from bulls showed a positive correlation with fresh semen attributes. However, SP/SM TIMP-2 levels in the Freezeable group did not show any significant differences compared to the Non-Freezable group in post-thaw semen quality attributes, kinetic parameters, and functional tests, except for a significant positive correlation (r = 0.530, p < 0.05) between sperm DNA integrity and SP-TIMP-2 levels. In conclusion, the findings suggested that TIMP-2 can be a positive regulator of semen quality at the neat stage. However, when it comes to the resilience of sperm to cryopreservation, the levels of TIMP-2 do not seem to exert any significant influence in breeding bulls.

Whole Blood Transcriptome Analysis in Dairy Ewes Fed a Dietary Grape Pomace Supplementation

The present study aims to evaluate the effect of a dietary supplementation with 10% grape pomace (GP) on the whole blood transcriptome of lactating ewes. By applying a log2FC higher than 0.5 or lower than −0.5 and a false discovery rate (FDR) &lt;0.05, the down-regulation of genes coding for plexin C1, ethanolamine kinase 1, tax1-binding protein 1, transmembrane 9 superfamily member 2, and Beclin-1 was observed in animals that received the dietary supplementation. This aspect was also accompanied by a reduction in the blood activity of matrix metalloproteinase 9 (MMP-9; p &lt; 0.05), a gelatinase commonly involved in both acute and chronic pathological events. The ELISA test on other factors involved in inflammatory processes, interleukin 1 (IL-1) and tumor necrosis factor α (TNF-α), as well as in the antioxidant response, glutathione peroxidase (GPx), and catalase (CAT), did not reveal any significant changes (p &gt; 0.05). Overall, the introduction of GP in the diet of ewes gave indications of greater efficacy in preserving animal welfare, with interesting cues regarding the valorization of a by-product with a high biological value.

The influence of interleukin-4 on transendothelial transport of low-density lipoproteins

BACKGROUND: Cytokines are important participants in the atherosclerotic process. The product of T-helpers and mast cells, interleukin-4, causes activation of endothelial cells and induces polarization of macrophages into anti-inflammatory M2 phenotype. AIM: The aim was to study the influence of interleukin-4 on the activation of transendothelial transport of low-density lipoproteins. MATERIALS AND METHODS: Low-density lipoproteins transendothelial transport was evaluated in a two-chamber model on a monolayer of human endothelial cell line EA.hy926. For that, 200 mkg/ml of low-density lipoproteins, and 1, or 10, or 100 ng/ml of interleukin-4, or 10, or 50 ng/ml of interleukin-6, or 50 ng/ml of tumor necrosis factor, or 10 ng/ml of interleukin-4 and 50 ng/ml of tumor necrosis factor, or a phosphate buffer saline were added to the cells on 24 h. The integrity of endothelial monolayer was controlled by the passage through 1 mkg/ml of fluorescein Na. The levels of mRNA and proteins in the cells were determined by reverse transcription polymerase chain reaction and Western blotting, the activity of matrix metalloproteinases-1, -2 and -9 in the culture medium — by zymography, interleukin-6 and -8 concentrations — by enzyme-linked immunosorbent assay. RESULTS: Interleukin-4 had no effect on the passage of low-density lipoproteins through the monolayer of endothelial cells in basal and tumor necrosis factor stimulation conditions. At the same time, interleukin-4 suppressed interleukin-8 secretion by cells and increased their production of interleukin-6. The latter also did not cause the changes in the permeability of endothelial monolayer. In addition, interleukin-4 did not affect the expression of CAV, SCARB1, ACVRL1 genes, encoding proteins involved in low-density lipoproteins transendothelial transport. An addition of interleukin-4 or -6 to the cells did not change the activity of the matrix metalloproteinases. CONCLUSIONS: Interleukin-4 does not affect the transendothelial transport of low-density lipoproteins, while modulating the production of other cytokines by endothelial cells. In the future, it is necessary to establish the effect of a wider range of cytokines produced by intimal cells on low-density lipoproteins transendothelial transport, as well as to clarify the molecular mechanisms of influence of tumor necrosis factor on this process in order to identify new targets for atherosclerosis therapy.

N6-methyladenosine-modified lncRNA in Staphylococcus aureus-injured bovine mammary epithelial cells.

Staphylococcus aureus-induced mastitis is a serious disease in dairy bovine, with no currently effective treatment. Antibiotics demonstrate certain therapeutic potency in dairy husbandry; they generate drug-resistant bacteria, thereby harming public health. LncRNAs and m6A have been verified as potential targets in infectious diseases and have powerful regulatory capabilities. However, the biological regulation of lncRNAs with m6A modification in mastitis needs further investigation. This study aims to determine the m6A-modified lncRNAs in bovine mammary epithelial cells and their diversity during S. aureus induction. Heat-inactivated S. aureus was used to develop the cell injury model, and we subsequently found low cell viability and different m6A modification levels. Our analysis of m6A-modified lncRNA profiles through MeRIP-seq revealed significant differences in 140 peaks within 130 lncRNAs when cells were injured by S. aureus. Furthermore, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses revealed that these differential m6A-modified lncRNAs were mainly enriched in the WNT pathway, and their functions were associated with amino acid metabolism, lipid translocation, and metalloproteinase activity. Here, we report for the first time lncRNAs with m6A modification in regulating S. aureus infection, revealing potential mechanisms and targets of infectious diseases, such as mastitis, from an epigenetics perspective.

OXY-SCORE and Volatile Anesthetics: A New Perspective of Oxidative Stress in EndoVascular Aneurysm Repair-A Randomized Clinical Trial.

An aortic aneurysm (AA) is a life-threatening condition. Oxidative stress may be a common pathway linking multiple mechanisms of an AA, including vascular inflammation and metalloproteinase activity. Endovascular aneurysm repair (EVAR) is the preferred surgical approach for AA treatment. During surgery, inflammation and ischemia-reperfusion injury occur, and reactive oxygen species (ROS) play a key role in their modulation. Increased perioperative oxidative stress is associated with higher postoperative complications. The use of volatile anesthetics during surgery has been shown to reduce oxidative stress. Individual biomarkers only partially reflect the oxidative status of the patients. A global indicator of oxidative stress (OXY-SCORE) has been validated in various pathologies. This study aimed to compare the effects of the main volatile anesthetics, sevoflurane and desflurane, on oxidative status during EVAR. Eighty consecutive patients undergoing EVAR were randomized into two groups: sevoflurane and desflurane. Plasma biomarkers of oxidative damage (protein carbonylation and malondialdehyde) and antioxidant defense (total thiols, glutathione, nitrates, superoxide dismutase, and catalase activity) were measured before surgery and 24 h after EVAR. The analysis of individual biomarkers showed no significant differences between the groups. However, the OXY-SCORE was positive in the desflurane group (indicating a shift towards antioxidants) and negative in the sevoflurane group (favoring oxidants) (p < 0.044). Compared to sevoflurane, desflurane had a positive effect on oxidative stress during EVAR. The OXY-SCORE could provide a more comprehensive perspective on oxidative stress in this patient population.

Features of Intracellular Matrix State during Development and Formation of COPD Clinical Phenotypes

The objective: to evaluate state of extracellular matrix markers and their tissue inhibitors during development and formation of clinical phenotypes of COPD, in patients facing the high risk of developing exacerbations.Subjects and Methods. 96 COPD patients (73% men and 27% women) were examined. Patients were randomized into the following groups: Group 1 included 40 COPD patients with bronchitis phenotype; Group 2 included 38 patients with emphysema phenotype; and Control Group consisted of 18 practically healthy subjects. All patients underwent a comprehensive clinical and laboratory examination. Levels of matrix metalloproteinases (total MMP-9 and MMP-2) and their tissue inhibitors TIMMP1 and TIMMP2 were determined by ELISA.Results. The development of COPD, regardless of the clinical phenotype of the disease, is accompanied by pronounced expression of extracellular matrix markers MMP-9 and MMP-2. Regardless of the COPD clinical phenotype, elevated activity of metalloproteinases is accompanied by inhibition of production of TIMMP2 and dysfunction of the MMP-9/TIMMP1 antiprotease system. In COPD patients with MMP-2 level of 214 ng/ml, the formation of bronchitis phenotype can be predicted, while with MMP-2 level of 214 ng/ml or more, the formation of emphysema phenotype can be predicted.

Escherichia coli induced matrix metalloproteinase-9 activity and type IV collagen degradation is regulated by progesterone in human maternal decidual

BackgroundEscherichia coli (E. coli) is one of the main bacteria associated with preterm premature rupture of membranes by increasing pro-matrix metalloproteinase 9 (proMMP-9) and degradation of type IV collagen in human feto-maternal interface (HFMi). proMMP-9 is regulated by progesterone (P4) but it is unclear whether P4 inhibits proMMP in human maternal decidual (MDec). This study aimed to determine a role of P4 on proMMP-2 and − 9 and type IV collagen induced by E. coli infection in MDec.MethodsNine HFMi were mounted in a Transwell system. MDec was stimulated with P4 or E. coli for 3-, 6-, or 24-hours. proMMP-2, -9 and type IV collagen were assessed.ResultsGelatin zymography revealed an increase in proMMP-9 after 3, 6, and 24 h of stimulating MDec with E. coli. Using immunofluorescence, it was confirmed the increase in the HFMi tissue and a reduction on the amount of type IV collagen leading to the separation of fetal amniochorion and MDEc. The degradative activity of proMMP-9 was reduced by 20% by coincubation with P4.ConclusionsP4 modulates the activity of proMMP-9 induced by E. coli stimulation but it was unable to completely reverse the degradation of type IV collagen in human MDec tissue.

Human iPSC-based disease modeling studies identify a common mechanistic defect and potential therapies for AMD and related macular dystrophies.

Age-related macular degeneration (AMD) and related macular dystrophies (MDs) primarily affect the retinal pigment epithelium (RPE) in the eye. A hallmark of AMD/MDs that drives later-stage pathologies is drusen. Drusen are sub-RPE lipid-protein-rich extracellular deposits, but how drusen forms and accumulates is not known. We utilized human induced pluripotent stem cell (iPSC)-derived RPE from patients with AMD and three distinct MDs to demonstrate that reduced activity of RPE-secreted matrix metalloproteinase 2 (MMP2) contributes to drusen in multiple maculopathies in a genotype-agnostic manner by instigating sterile inflammation and impaired lipid homeostasis via damage-associated molecular pattern molecule (DAMP)-mediated activation of receptor for advanced glycation end-products (RAGE) and increased secretory phospholipase 2-IIA (sPLA2-IIA) levels. Therapeutically, RPE-specific MMP2 supplementation, RAGE-antagonistic peptide, and a small molecule inhibitor of sPLA2-IIA ameliorated drusen accumulation in AMD/MD iPSC-RPE. Ultimately, this study defines a causal role of the MMP2-DAMP-RAGE-sPLA2-IIA axis in AMD/MDs.

A Novel Porcine Model of Thoracic Aortic Aneurysms Reveals Increased Endogenous Matrix Metalloproteinase Activity

A Novel Porcine Model of Thoracic Aortic Aneurysms Reveals Increased Endogenous Matrix Metalloproteinase Activity

Heparanase-induced endothelial glycocalyx degradation exacerbates lung ischemia/reperfusion injury in male mice.

The endothelial glycocalyx (eGC) is a carbohydrate-rich layer on the vascular endothelium, and its damage can lead to endothelial and organ dysfunction. Heparanase (HPSE) degrades the eGC in response to cellular stress, but its role in organ dysfunction remains unclear. This study investigates HPSE's role in lung ischemia-reperfusion (I/R) injury. A left lung hilar occlusion model was used in B6 wildtype (WT) and HPSE genetic knockout (-/-) mice to induce I/R injury invivo. The left lungs were ischemic for 1 h followed by reperfusion for 4 h prior to investigations of lung function and eGC status. Data were compared between uninjured lungs and I/R-injured lungs in WT and HPSE-/- mice. WT lungs showed significant functional impairment after I/R injury, whereas HPSE-/- lungs did not. Inhibition or knockout of HPSE prevented eGC damage, inflammation, and cellular migration after I/R injury by reducing matrix metalloproteinase activities. HPSE-/- mice exhibited compensatory regulation of related gene expressions. HPSE facilitates eGC degradation leading to inflammation and impaired lung function after I/R injury. HPSE may be a therapeutic target to attenuate graft damage in lung transplantation.

Impact of zirconia-based oxide on endothelial cell dynamics and extracellular matrix remodeling

IntroductionZirconia (ZrO2) is highly regarded in dental restoration due to its aesthetic compatibility and mechanical properties that align with biological tissues. This study explores the effects of stabilized ZrO2 on endothelial cell function and extracellular matrix (ECM) remodeling, processes critical to successful osseointegration in dental implants. MethodologyHuman Umbilical Vein Endothelial Cells (HUVECs) were cultured in ZrO2 -enriched medium under both static and shear stress conditions. Newly implemented techniques, including detailed zirconia surface characterization using scanning electron microscopy (SEM), energy-dispersive X-ray spectroscopy (EDX), and X-ray diffraction (XRD), were used to verify material properties. Gene and protein expression related to cell adhesion, proliferation, and ECM remodeling were assessed through RT-qPCR and Western blotting. Zymography was used to evaluate the activity of matrix metalloproteinases (MMP2 and MMP9) involved in ECM remodeling. ResultsCharacterization data confirmed the stability and structural properties of ZrO2, revealing a tetragonal crystalline structure and rough surface morphology conducive to cell adhesion. ZrO2 exposure led to the downregulation of Src, a key regulator of cell adhesion, while upregulating cell cycle regulators p15, CDK2, and CDK4, indicating enhanced cell proliferation. Under shear stress, ZrO2 modulated TGF-β and MAPK signaling, affecting cell proliferation and angiogenesis. MMP2 and MMP9 activity increased in static conditions but decreased under shear stress, suggesting ZrO2 dynamic role in ECM remodeling. ConclusionThis study shows that stabilized zirconia (ZrO2) modulates endothelial cell dynamics and ECM remodeling, key for osseointegration. ZrO2 downregulated Src expression and upregulated cell cycle regulators, enhancing endothelial proliferation. It also affected TGF-β and MAPK pathways, influencing angiogenesis, and differentially modulated MMP2 and MMP9 activity depending on mechanical conditions. These findings highlight ZrO2 has potential ability to enhance vascular and tissue integration in dental applications.

Evaluation of the Impact of Pistacia atlantica Desf. Oleo-gum Resin on Complete Freund’s Adjuvant-induced Arthritis in a Wistar Rat Model

Background: Pistacia atlantica Desf., also known as Atlantic pistachio or Mt. Atlas mastic tree, has been used in traditional medicine for its anti-inflammatory properties, helping to reduce inflammation in conditions like arthritis. Additionally, the tree's resin possesses antioxidant properties that can help protect against oxidative stress. The anti-arthritic efficacy of Pistacia atlantica oleo-gum resin extract (PAG) remains unvalidated, as its potential therapeutic benefits have not been substantiated. In the current investigation, we have substantiated the inhibitory properties of the PAG in mitigating the development of rheumatoid arthritis induced by Complete Freund's Adjuvant in rats. Methods: Male Wistar rats weighing 160±10 g were subjected to immunization through intradermal injection of 0.1 mL of Complete Freund's Adjuvant (CFA) into the left hind metatarsal footpad. Following the induction of adjuvant arthritis, oral administration of PAG was initiated at doses of 10, 400, and 800 mg/kg once a day. Prednisolone (10 mg/kg, daily) was employed as a positive control in this study. Various parameters, including paw swelling, arthritic index, body weight loss, spleen index, thymus index, serum cytokines, inflammatory mediators, and histological alterations, as well as nitrite glutathione and catalase level, were measured. Furthermore, the expression and activity of matrix metalloproteinase (MMP)-2 and MMP-9 were evaluated. Results: PAG exhibited noteworthy efficacy in attenuating paw swelling and reducing the arthritic index. Moreover, it resulted in enhanced body weight loss and a consequential reduction in the thymus index. PAG (400mg/kg) significantly decreased the level of serum nitrite in the intervention group, as well as the levels of serum glutathione. On the other hand, catalase activity increased, the activity of MMP-9 decreased, and the level of MMP-2 increased compared to the intervention group. The histopathological results revealed that PAG (at doses of 400 mg/kg) had the best therapeutic effects. Conclusion: In conclusion, the present study confirms the anti-arthritic activity of PAG in rats. The extract of PAG was found to prevent rheumatoid arthritis induced by Complete Freund's Adjuvant, as evidenced by the significant suppression of paw swelling and arthritic index.

Substance P alleviates liver fibrosis by modulating inflammation and mobilizing reparative stem cells

Repetitive hepatic damage resulting from viral hepatitis, toxins, and alcohol abuse induces chronic inflammation and excessive accumulation of the extracellular matrix, leading to the development of liver cirrhosis. Substance P (SP) promotes endogenous wound healing by mobilizing bone marrow stem cells and stimulating anti-inflammatory responses. This study aimed to investigate whether SP exerts a therapeutic effect on liver fibrosis by recruiting endogenous stem cells and modulating immune responses.A non-clinical model of liver cirrhosis was established through repeated injections of thioacetamide and recombinant leptin. After confirming liver fibrosis, SP was administered intravenously for 6 weeks.SP treatment decreased the formation of hepatic micronodules on the external surface of the liver and the infiltration of immune cells. Furthermore, SP treatment notably reduced the deposition of collagen and the activation of hepatic stellate cells, concomitant with decreased levels of transforming growth factor-β1 and matrix metalloproteinase activity. In the context of severe hepatic damage, SP increased the number of circulating stem cells, leading to the restoration of the reparative stem cell pool in the bone marrow.The findings of this study suggest that SP alleviates liver fibrosis by modulating the mobilization of functional stem cells and the immune response.

It takes two peroxisome proliferator-activated receptors (PPAR-β/δ and PPAR-γ) to tango idiopathic pulmonary fibrosis

BackgroundIdiopathic pulmonary fibrosis (IPF) is characterized by aberrant lung epithelial phenotypes, fibroblast activation, and increased extracellular matrix deposition. Transforming growth factor-beta (TGF-β)1-induced Smad signaling and downregulation of peroxisomal genes are involved in the pathogenesis and can be inhibited by peroxisome proliferator-activated receptor (PPAR)-α activation. However, the three PPARs, that is PPAR-α, PPAR-β/δ, and PPAR-γ, are known to interact in a complex crosstalk.MethodsTo mimic the pathogenesis of lung fibrosis, primary lung fibroblasts from control and IPF patients with comparable levels of all three PPARs were treated with TGF-β1 for 24 h, followed by the addition of PPAR ligands either alone or in combination for another 24 h. Fibrosis markers (intra- and extracellular collagen levels, expression and activity of matrix metalloproteinases) and peroxisomal biogenesis and metabolism (gene expression of peroxisomal biogenesis and matrix proteins, protein levels of PEX13 and catalase, targeted and untargeted lipidomic profiles) were analyzed after TGF-β1 treatment and the effects of the PPAR ligands were investigated.ResultsTGF-β1 induced the expected phenotype; e.g. it increased the intra- and extracellular collagen levels and decreased peroxisomal biogenesis and metabolism. Agonists of different PPARs reversed TGF-β1-induced fibrosis even when given 24 h after TGF-β1. The effects included the reversals of (1) the increase in collagen production by repressing COL1A2 promoter activity (through PPAR-β/δ activation); (2) the reduced activity of matrix metalloproteinases (through PPAR-β/δ activation); (3) the decrease in peroxisomal biogenesis and lipid metabolism (through PPAR-γ activation); and (4) the decrease in catalase protein levels in control (through PPAR-γ activation) and IPF (through a combined activation of PPAR-β/δ and PPAR-γ) fibroblasts. Further experiments to explore the role of catalase showed that an overexpression of catalase protein reduced collagen production. Additionally, the beneficial effect of PPAR-γ but not of PPAR-β/δ activation on collagen synthesis depended on catalase activity and was thus redox-sensitive.ConclusionOur data provide evidence that IPF patients may benefit from a combined activation of PPAR-β/δ and PPAR-γ.

Fibulin-3 Regulates Tissue Inhibitor of Metalloproteinases 3 to Inhibit Senescence in Intervertebral Disc Nucleus Pulposus Cells

To investigate the effect of fibulin-3 on the senescence of intervertebral disc nucleus pulposus cells (NPCs) through the regulation of tissue inhibitor of metalloproteinases 3 (TIMP-3) expression and to elucidate the molecular mechanisms involved. 1). The nucleus pulposus tissues and imaging data of 37 patients who had undergone intervertebral disc surgery were collected. The degree of degeneration of the intervertebral discs were classified according to the Pfirrmann grading system. The senescence degree of NPCs was determined using senescence-associated β-galactosidase (SA-β-gal) staining. Fibulin-3 expression levels were determined using Western blot and ELISA. The relationship between fibulin-3 and disc degeneration and NPCs senescence was investigated. 2). Human intervertebral disc NPCs were cultured in vitro. The proliferation and senescence of NPC across continuous passage were observed via CCK-8 assay and SA-β-gal staining, respectively. Fibulin-3 expression levels and the expression of inflammatory cytokines and matrix metalloproteinases were assessed. Exogenous fibulin-3 was added to verify its effect on the proliferation and senescence of NPCs. 3). The effect of fibulin-3 on the apoptosis and proliferation of NPCs was verified through gene overexpression, which was used in combination with an apoptosis inhibitor for bidirectional verification. 4). Bioinformatics analysis was performed to explore the relationship between fibulin-3 and the TIMP family. Experiments overexpressing fibulin-3 and silencing the TIMP-3 gene were performed to verify their role in NPCs senescence. 1). The intervertebral disc degeneration samples from 37 patients were classified according to the Pfirrmann grading system. The higher the degeneration grade, the lower fibulin-3 expression. Spearman correlation analysis showed that the disc grade was negatively correlated with the NPC senescence grade (r=－0.87, P<0.001) and fibulin-3 expression (r=－0.79, P<0.001). 2). As the passage number of NPCs increased, fibulin-3 expression gradually decreased, cell proliferation ability weakened, and the expression of inflammatory cytokines and matrix metalloproteinases increased. After exogenous fibulin-3 was added, cell morphology and growth status were maintained, cell senescence was significantly inhibited, and the expression of inflammatory cytokines and matrix metalloproteinases was markedly reduced. 3). Gene overexpression experiments showed that fibulin-3 reduced NPC apoptosis and promoted cell proliferation, thereby inhibiting NPC senescence. 4). Bioinformatics analysis revealed a significant association between fibulin-3 and TIMP-3 of the TIMP family. Further experiments confirmed that overexpressing fibulin-3 enhanced TIMP-3 expression, while silencing the TIMP-3 gene significantly weakened the inhibitory effect of fibulin-3 on NPCs senescence. This indicates that, through regulating TIMP-3, fibulin-3 inhibits the activity of matrix metalloproteinases, affects the synthesis and degradation of the extracellular matrix, and ultimately inhibits NPCs senescence. This study demonstrates that fibulin-3 plays a crucial role in inhibiting the senescence of intervertebral disc NPCs by regulating TIMP-3. The specific mechanisms involved are as follows, fibulin-3 upregulates TIMP-3 expression, inhibits matrix metalloproteinase activity, and reduces extracellular matrix degradation, thereby promoting extracellular matrix synthesis. Additionally, fibulin-3 inhibits NPCs senescence by reducing apoptosis and promoting cell proliferation. Therefore, fibulin-3 and TIMP-3 have potential therapeutic significance in maintaining intervertebral disc health and delaying degeneration.

Interferon lambda signaling in neutrophils enhances the pathogenesis of Bordetella pertussis infection

Abstract Interferon lambda plays diverse roles in bacterial infections. Previously, we showed that interferon lambda is induced in the lungs of Bordetella pertussis-infected adult mice and exacerbates inflammation. Here, we report that mice lacking the interferon lambda receptor 1 specifically on neutrophils (MRP8creIFNLR1fl/fl mice) exhibit reduced lung bacterial load and inflammation compared to wild-type mice during B. pertussis infection. In B. pertussis-infected wild-type mice, lung type I and III IFN responses were higher than in MRP8creIFNLR1fl/fl mice, correlating with increased lung inflammatory pathology. There was an increased proportion of interferon gamma-producing neutrophils in the lungs of MRP8creIFNLR1fl/fl mice compared to wild-type mice. IFNLR1−/− neutrophils incubated with B. pertussis exhibited higher killing compared to wild-type neutrophils. Treatment of wild-type neutrophils with interferon lambda further decreased their bacterial killing capacity and treatment of wild-type mice with interferon lambda increased lung bacterial loads. Contributing to the differential killing, we found that IFNLR1−/− neutrophils exhibit higher levels of reactive oxygen species, myeloperoxidase, matrix metalloproteinase-9 activity, neutrophil extracellular traps, and interferon gamma secretion than wild-type neutrophils, and inhibiting NADPH oxidase inhibited bacterial killing in IFNLR1−/− neutrophils. B. pertussis-induced interferon lambda secretion and IFNLR1 gene expression in mouse and human neutrophils and this was dependent on the bacterial virulence protein pertussis toxin. Pertussis toxin enhanced bacterial loads in wild type but not in MRP8creIFNLR1fl/fl or IFNLR1−/− mice. Thus, pertussis toxin disrupts neutrophil function by enhancing type III IFN signaling, which prevents neutrophils from effectively clearing B. pertussis during infection, leading to higher bacterial loads and exacerbation of lung inflammation.