Sinomenine treats rheumatoid arthritis by inhibiting MMP9 and inflammatory cytokines expression: bioinformatics analysis and experimental validation.

Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease marked by inflammatory cell infiltration and joint damage. The Chinese government has approved the prescription medication sinomenine (SIN), an effective anti-inflammation drug, for treating RA. This study evaluated the possible anti-inflammatory actions of SIN in RA based on bioinformatics analysis and experiments. Six microarray datasets were acquired from the gene expression omnibus (GEO) database. We used R software to identify differentially expressed genes (DEGs) and perform function evaluations. The CIBERSORT was used to calculate the abundance of 22 infiltrating immune cells. The weighted gene co-expression network analysis (WGCNA) was used to discover genes associated with M1 macrophages. Four public datasets were used to predict the genes of SIN. Following that, function enrichment analysis for hub genes was performed. The cytoHubba and least absolute shrinkage and selection operator (LASSO) were employed to select hub genes, and their diagnostic effectiveness was predicted using the receiver operator characteristic (ROC) curve. Molecular docking was undertaken to confirm the affinity between the SIN and hub gene. Furthermore, the therapeutic efficacy of SIN was validated in LPS-induced RAW264.7 cells line using Western blot and Enzyme-linked immunosorbent assay (ELISA). The matrix metalloproteinase 9 (MMP9) was identified as the hub M1 macrophages-related biomarker in RA using bioinformatic analysis and molecular docking. Our study indicated that MMP9 took part in IL-17 and TNF signaling pathways. Furthermore, we found that SIN suppresses the MMP9 protein overexpression and pro-inflammatory cytokines, including tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in the LPS-induced RAW264.7 cell line. In conclusion, our work sheds new light on the pathophysiology of RA and identifies MMP9 as a possible RA key gene. In conclusion, the above findings demonstrate that SIN, from an emerging research perspective, might be a potential cost-effective anti-inflammatory medication for treating RA.

Complement regulatory protein CD46 promotes bladder cancer metastasis through activation of MMP9.

CD46, a transmembrane protein known for protecting cells from complement­mediated damage, is frequently dysregulated in various types of cancer. Its overexpression in bladder cancers safeguards the cancer cells against both complement and antibody­mediated cytotoxicity. The present study explored a new role of CD46 in facilitating cancer cell invasion and metastasis, examining its regulatory effect on matrix metalloproteases (MMPs) and their effect on the metastatic capability of bladder cancer cells. Specifically, CD46 alteration positively influenced MMP9 expression, but not MMP2, in several bladder cancer cell lines. Furthermore, CD46 overexpression triggered phosphorylation of p38 MAPK and protein kinase B (AKT), leading to enhanced activator protein 1 (AP­1) activity via c­Jun upregulation. The inhibition of p38 or AKT pathways attenuated the CD46­induced MMP9 and AP­1 upregulation, indicating that the promotion of MMP9 by CD46 involved activating both p38 MAPK and AKT. Functionally, the upregulation of MMP9 by CD46 translated to increased migratory and invasive capabilities of bladder cancer cells, as well as enhanced in vivo metastasis. Overall, the present study revealed a novel role for CD46 as a metastasis promoter through MMP9 activation in bladder cancers and highlighted the regulatory mechanism of CD46­mediated MMP9 promotion via p38 MAPK and AKT activation.

[The Miao medicine Sidaxue alleviates rheumatoid arthritis in rats possibly by downregulating matrix metalloproteinases].

OBJECTIVE: To explore the inhibitory effect of Sidaxue, a traditional Miao herbal medicine formula, on articular bone and cartilage destruction and synovial neovascularization in rats with collagen-induced arthritis (CIA). METHODS: In a SD rat model of CIA, we tested the effects of daily gavage of Sidaxue at low, moderate and high doses (10, 20, and 40 g/kg, respectively) for 21 days, with Tripterygium glycosides (GTW) as the positive control, on swelling in the hind limb plantar regions by arthritis index scoring. Pathologies in joint synovial membrane of the rats were observed with HE staining, and serum TNF-α and IL-1ß levels were detected with ELISA. The expressions of NF-κB p65, matrix metalloproteinase 1 (MMP1), MMP2 and MMP9 at the mRNA and protein levels in the synovial tissues were detected using real-time PCR and Western blotting. Network pharmacology analysis was conducted to identify the important target proteins in the pathways correlated with the therapeutic effects of topical Sidaxue treatment for RA, and the core target proteins were screened by topological analysis. RESULTS: Treatment with GTW and Sidaxue at the 3 doses all significantly alleviated plantar swelling, lowered arthritis index scores, improved cartilage and bone damage and reduced neovascularization in CIA rats (P<0.05), and the effects of Sidaxue showed a dose dependence. Both GTW and Sidaxue treatments significantly lowered TNF-α, IL-1ß, NF-κB p65, MMP1, MMP2, and MMP9 mRNA and protein expressions in the synovial tissues of CIA rats (P<0.05). Network pharmacological analysis identified MMPs as the core proteins associated with topical Sidaxue treatment of RA. CONCLUSION: Sidaxue alleviates articular bone and cartilage damages and reduces synovial neovascularization in CIA rats possibly by downregulating MMPs via the TNF-α/IL-1ß/NF-κB-MMP1, 2, 9 signaling pathway, and MMPs probably plays a key role in mediating the effect of Sidaxue though the therapeutic pathways other than oral administration.

Discovery of biomarkers in the psoriasis through machine learning and dynamic immune infiltration in three types of skin lesions.

Introduction: Psoriasis is a chronic skin disease characterized by unique scaling plaques. However, during the acute phase, psoriatic lesions exhibit eczematous changes, making them difficult to distinguish from atopic dermatitis, which poses challenges for the selection of biological agents. This study aimed to identify potential diagnostic genes in psoriatic lesions and investigate their clinical significance. Methods: GSE182740 datasets from the GEO database were analyzed for differential analysis; machine learning algorithms (SVM-RFE and LASSO regression models) are used to screen for diagnostic markers; CIBERSORTx is used to determine the dynamic changes of 22 different immune cell components in normal skin lesions, psoriatic non-lesional skin, and psoriatic lesional skin, as well as the expression of the diagnostic genes in 10 major immune cells, and real-time quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry are used to validate results. Results: We obtained 580 differentially expressed genes (DEGs) in the skin lesion and non-lesion of psoriasis patients, 813 DEGs in mixed patients between non-lesions and lesions, and 96 DEGs in the skin lesion and non-lesion of atopic dermatitis, respectively. Then 144 specific DEGs in psoriasis via a Veen diagram were identified. Ultimately, UGGT1, CCNE1, MMP9 and ARHGEF28 are identified for potential diagnostic genes from these 144 specific DEGs. The value of the selected diagnostic genes was verified by receiver operating characteristic (ROC) curves with expanded samples. The the area under the ROC curve (AUC) exceeded 0.7 for the four diagnosis genes. RT-qPCR results showed that compared to normal human epidermis, the expression of UGGT1, CCNE1, and MMP9 was significantly increased in patients with psoriasis, while ARHGEF28 expression was significantly decreased. Notably, the results of CIBERSORTx showed that CCNE1 was highly expressed in CD4+ T cells and neutrophils, ARHGEF28 was also expressed in mast cells. Additionally, CCNE1 was strongly correlated with IL-17/CXCL8/9/10 and CCL20. Immunohistochemical results showed increased nuclear expression of CCNE1 in psoriatic epidermal cells relative to normal. Conclusion: Based on the performance of the four genes in ROC curves and their expression in immune cells from patients with psoriasis, we suggest that CCNE1 possess higher diagnostic value.

Perineuronal nets are phagocytosed by MMP-9 expressing microglia and astrocytes in the SOD1G93A ALS mouse model.

AIMS: Perineuronal nets (PNNs) are an extracellular matrix structure that encases excitable neurons. PNNs play a role in neuroprotection against oxidative stress. Oxidative stress within motor neurons can trigger neuronal death, which has been implicated in amyotrophic lateral sclerosis (ALS). We investigated the spatio-temporal timeline of PNN breakdown and the contributing cellular factors in the SOD1G93A strain, a fast-onset ALS mouse model. METHODS: This was conducted at the presymptomatic (P30), onset (P70), mid-stage (P130), and end-stage disease (P150) using immunofluorescent microscopy, as this characterisation has not been conducted in the SOD1G93A strain. RESULTS: We observed a significant breakdown of PNNs around α-motor neurons in the ventral horn of onset and mid-stage disease SOD1G93A mice compared with wild-type controls. This was observed with increased numbers of microglia expressing matrix metallopeptidase-9 (MMP-9), an endopeptidase that degrades PNNs. Microglia also engulfed PNN components in the SOD1G93A mouse. Further increases in microglia and astrocyte number, MMP-9 expression, and engulfment of PNN components by glia were observed in mid-stage SOD1G93A mice. This was observed with increased expression of fractalkine, a signal for microglia engulfment, within α-motor neurons of SOD1G93A mice. Following PNN breakdown, α-motor neurons of onset and mid-stage SOD1G93A mice showed increased expression of 3-nitrotyrosine, a marker for protein oxidation, which could render them vulnerable to death. CONCLUSIONS: Our observations suggest that increased numbers of MMP-9 expressing glia and their subsequent engulfment of PNNs around α-motor neurons render these neurons sensitive to oxidative damage and eventual death in the SOD1G93A ALS model mouse.

3,3'-((3,4,5-trifluoropHenyl)methylene)bis(4-hydroxy-2H-chromen-2-one) inhibit lung cancer cell proliferation and migration.

Lung cancer is a major public health challenge and, despite therapeutic improvements, is the first leading cause of cancer worldwide. The current cure rate from advanced cancer treatment is excessively low. Therefore, it is of great importance to identify novel, potent and less toxic anticancer agents for the treatment of lung cancer. The aim of our research is to synthesize a new biscoumarin 3,3'-((3,4,5-trifluorop -phenyl)methylene)bis(4-hydroxy-2H-chromen-2-one) (C35) as an anticancer agent. C35 was simply prepared by 4-hydroxycoumarin and 3,4,5-trifluorobenzaldehyde under ethanol and its structure was analyzed by spectroscopic analyses. The anti-proliferation effect of C35 was detected using CCK-8 assay. Migration abilities were measured by Transwell assay. The expression of correlated proteins was determined by Western blot. The results showed that C35 displayed strong cytostatic effects on lung cancer cell proliferation. In addition, C35 possessed a significant inhibition of migration by reducing the expression of matrix metalloproteinases-2 (MMP-2) and MMP-9 in lung cancer cells. Furthermore, C35 treatment suppressed the phosphorylation of p38 in lung cancer cells. Moreover, in vivo experiments were carried out, in which we treated Lewis tumor-bearing C57 mice via intraperitoneal injection of C35. Results showed that C35 inhibited tumor growth in vivo. In conclusion, our study demonstrated the anticancer activity of C35 via suppression of lung cancer cell proliferation and migration, which is possibly involved with the inhibition of the p38 pathway.

[Knockdown mitochondrial transcription factor A (TFAM) inhibits autophagy, proliferation, invasion and migration in human cervical cancer and osteosarcoma cells].

Objective To investigate the effects of mitochondrial transcription factor A (TFAM) on mitochondrial function, autophagy, proliferation, invasion, and migration in cervical cancer HeLa cells and osteosarcoma U2OS cells. Methods TFAM small-interfering RNA (si-TFAM) was transfected to HeLa and U2OS cells for downregulating TFAM expression. Mito-Tracker Red CMXRos staining combined with laser confocal microscopy was used to detect mitochondrial membrane potential (MMP). MitoSOXTM Red labeling was used to test mitochondrial reactive oxygen species (mtROS) levels. The expression of mitochondrial DNA (mtDNA) was detected by real-time quantitative PCR. Changes in the number of autophagosomes were detected by immunofluorescence cytochemistry. Western blot analysis was used to detect the expressions of TFAM, autophagy microtubule associated protein 1 light chain 3A/B (LC3A/B), autophagy associated protein 2A (ATG2A), ATG2B, ATG9A, zinc finger transcription factor Snail, matrix metalloproteinase 2 (MMP2) and MMP9. CCK-8 assay and plate clony formation assay were used to detect cell proliferation, while TranswellTM assay and scratch healing assay were used to detect changes in cell invasion and migration. Results The downregulation of TFAM expression resulted in a decrease in MMP and mtDNA copy number, but an increase in mtROS production. The protein content of LC3A/B decreased significantly compared to the control group and the number of autophagosomes in the cytoplasm decreased significantly. The expressions of ATG2B and ATG9A in the early stage of autophagy were significantly reduced. The expressions of Snail, MMP2 and MMP9 proteins in HeLa and U2OS cells were also decreased. The proliferation, invasion and migration ability of HeLa and U2OS cells were inhibited after being interfered with TFAM expression. Conclusion Downregulation of TFAM expression inhibits mitochondrial function, delays autophagy process and reduces the proliferation, invasion and migration ability of cervical cancer cells and osteosarcoma cells.

Modulating synovial macrophage pyroptosis and mitophagy interactions to mitigate osteoarthritis progression using functionalized nanoparticles.

Synovial macrophages play an important role in the progression of osteoarthritis (OA). In this study, we noted that synovial macrophages can activate pyroptosis in a gasdermin d-dependent manner and produce reactive oxygen species (ROS), aberrantly activating the mammalian target of rapamycin complex 1 (mTORC1) pathway and matrix metalloproteinase-9 (MMP9) expression in synovial tissue samples collected from both patients with OA and collagen-induced osteoarthritis (CIOA) mouse model. To overcome this, we constructed rapamycin- (RAPA, a mTORC1 inhibitor) loaded mesoporous Prussian blue nanoparticles (MPB NPs, for catalyzing ROS) and modified the NPs with MMP9-targeted peptides (favor macrophage targeting) to develop RAPA@MPB-MMP9 NPs. The inherent enzyme-like activity and RAPA released from RAPA@MPB-MMP9 NPs synergistically impeded the pyroptosis of macrophages and the activation of the mTORC1 pathway. In particular, the NPs decreased pyroptosis-mediated ROS generation, thereby inhibiting cGAS-STING signaling pathway activation caused by the release of mitochondrial DNA. Moreover, the NPs promoted macrophage mitophagy to restore mitochondrial stability, alleviate pyroptosis-related inflammatory responses, and decrease senescent synoviocytes. After the as-prepared NPs were intra-articularly injected into the CIOA mouse model, they efficiently attenuated synovial macrophage pyroptosis and cartilage degradation. In conclusion, our study findings provide a novel therapeutic strategy for OA that modulates the pyroptosis and mitophagy of synovial macrophage by utilizing functionalized NPs. STATEMENT OF SIGNIFICANCE: Osteoarthritis (OA) presents a significant global challenge owing to its complex pathogenesis and finite treatment options. Synovial macrophages have emerged as key players in the progression of OA, managing inflammation and tissue destruction. In this study, we discovered a novel therapeutic strategy in which the pyroptosis and mitophagy of synovial macrophages are targeted to mitigate OA pathology. For this, we designed and prepared rapamycin-loaded mesoporous Prussian blue nanoparticles (RAPA@MPB-MMP9 NPs) to specifically target synovial macrophages and modulate their inflammatory responses. These NPs could efficiently suppress macrophage pyroptosis, diminish reactive oxygen species production, and promote mitophagy, thereby alleviating inflammation and protecting cartilage integrity. Our study findings not only clarify the intricate mechanisms underlying OA pathogenesis but also present a promising therapeutic approach for effectively managing OA by targeting dysregulation in synovial macrophages.

Deflamin Attenuated Lung Tissue Damage in an Ozone-Induced COPD Murine Model by Regulating MMP-9 Catalytic Activity.

Chronic obstructive pulmonary disease (COPD) is comprised of histopathological alterations such as pulmonary emphysema and peribronchial fibrosis. Matrix metalloproteinase 9 (MMP-9) is one of the key enzymes involved in both types of tissue remodeling during the development of lung damage. In recent studies, it was demonstrated that deflamin, a protein component extracted from Lupinus albus, markedly inhibits the catalytic activity of MMP-9 in experimental models of colon adenocarcinoma and ulcerative colitis. Therefore, in the present study, we investigated for the first time the biological effect of deflamin in a murine COPD model induced by chronic exposure to ozone. Ozone exposure was carried out in C57BL/6 mice twice a week for six weeks for 3 h each time, and the treated group was orally administered deflamin (20 mg/kg body weight) after each ozone exposure. The histological results showed that deflamin attenuated pulmonary emphysema and peribronchial fibrosis, as evidenced by H&E and Masson's trichrome staining. Furthermore, deflamin administration significantly decreased MMP-9 activity, as assessed by fluorogenic substrate assay and gelatin zymography. Interestingly, bioinformatic analysis reveals a plausible interaction between deflamin and MMP-9. Collectively, our findings demonstrate the therapeutic potential of deflamin in a COPD murine model, and suggest that the attenuation of the development of lung tissue damage occurs by deflamin-regulated MMP-9 catalytic activity.

PLD2 deficiency alleviates endothelial glycocalyx degradation in LPS-induced ARDS/ALI.

- Acute respiratory distress syndrome (ARDS)/acute lung injury (ALI) is a life-threatening condition marked by severe lung inflammation and increased lung endothelial barrier permeability. Endothelial glycocalyx deterioration is the primary factor of vascular permeability changes in ARDS/ALI. Although previous studies have shown that phospholipase D2 (PLD2) is closely related to the onset and progression of ARDS/ALI, its role and mechanism in the damage of endothelial cell glycocalyx remains unclear. We used LPS-induced ARDS/ALI mice (in vivo) and LPS-stimulated injury models of EA.hy926 endothelial cells (in vitro). We employed C57BL/6 mice, including wild-type and PLD2 knockout (PLD2-/-) mice, to establish the ARDS/ALI model. We applied immunofluorescence and ELISA to examine changes in syndecan-1 (SDC-1), matrix metalloproteinase-9 (MMP9), inflammatory cytokines (TNF-α, IL-6, and IL-1ß) levels and the effect of external factors, such as phosphatidic acid (PA), 1-butanol (a PLD inhibitor), on SDC-1 and MMP9 expression levels. We found that PLD2 deficiency inhibits SDC-1 degradation and MMP9 expression in LPS-induced ARDS/ALI. Externally added PA decreases SDC-1 levels and increases MMP9 in endothelial cells, hence underlining PA's role in SDC-1 degradation. Additionally, PLD2 deficiency decreases the production of inflammatory cytokines (TNF-α, IL-6, and IL-1ß) in LPS-induced ARDS/ALI. In summary, these findings suggest that PLD2 deficiency plays a role in inhibiting the inflammatory process and protecting against endothelial glycocalyx injury in LPS-induced ARDS/ALI.

Green tea extract suppresses airway inflammation via oxidative stress-driven MAPKs/MMP-9 signaling in asthmatic mice and human airway epithelial cells.

Introduction: The anti-inflammatory effect of green tea extract (GTE) has been confirmed in asthmatic mice, however, the pharmacological mechanism is not fully elucidated. Methods: To investigate the therapeutic efficacy of GTE in asthma and identify specific pathways, murine model of allergic asthma was established by ovalbumin (OVA) sensitization and the challenge for 4 weeks, with oral treatment using GTE and dexamethasone (DEX). Inflammatory cell counts, cytokines, OVA-specific IgE, airway hyperreactivity, and antioxidant markers in the lung were evaluated. Also, pulmonary histopathological analysis and western blotting were performed. In vitro, we established the model by stimulating the human airway epithelial cell line NCI-H292 using lipopolysaccharide, and treating with GTE and mitogen-activated protein kinases (MAPKs) inhibitors. Results: The GTE100 and GTE400 groups showed a decrease in airway hyperresponsiveness and the number of inflammatory cells in the bronchoalveolar lavage fluid (BALF) compared to the OVA group. GTE treatment also reduced interleukin (IL)-13, IL-5, and IL-4 levels in the BALF, and OVA-specific immunoglobulin E levels in the serum compared to those in the OVA group. GTE treatment decreased OVA-induced mucus secretion and airway inflammation. In addition, GTE suppressed the oxidative stress, and phosphorylation of MAPKs, which generally occurs after exposure to OVA. GTE administration also reduced matrix metalloproteinase-9 activity and protein levels. Conclusion: GTE effectively inhibited asthmatic respiratory inflammation and mucus hyperproduction induced by OVA inhalation. These results suggest that GTE has the potential to be used for the treatment of asthma.

Evaluation of oyster mushroom (Pleurotus ostreatus)-derived anthraquinone on the induction of apoptosis and suppression of MMP-2 and MMP-9 expression in breast cancer cells.

Introduction: Breast cancer results from tissue degradation caused by environmental and genetic factors that affect cells in the body. Matrix metalloproteinases, such as MMP-2 and MMP-9, are considered potential putative markers for tumor diagnosis in clinical validation due to their easy detection in body fluids. In addition, recent reports have suggested multiple roles for MMPs, rather than simply degeneration of the extracellular matrix, which comprises mobilizing growth factors and processing surface molecules. Methods: In this study, the chemotherapeutic effects of anthraquinone (AQ) extracted from edible mushrooms (Pleurotus ostreatus Jacq. ex Fr.) cells was examined in MCF-7 breast cancer cells. The cytotoxic potential and oxidative stress induced by purified anthraquinone were assessed in MCF-7 cells using MTT and ROS estimation assays. Gelatin Zymography, and DNA fragmentation assays were performed to examine MMP expression and apoptotic induction in the MCF-7 cells treated with AQ. The genes crucial for mutations were examined, and the mutated RNA knockout plausibility was analyzed using the CRISPR spcas9 genome editing software. Results: MCF-7 cells were attenuated in a concentration-dependent manner by the administration of AQ purified from P. ostreatus compared with the standard anticancer drug paclitaxel. AQ supplementation decreased oxidative stress and mitochondrial impairment in MCF-7 cells. Treatment with AQ and AQ with paclitaxel consistently decreased the expression of crucial marker genes such as MMP2 and MMP9. The mutated genes MMP2, MMP7, and MMP9 were assessed and observed to reveal four putative gene knockdown potentials for breast cancer treatment. Conclusions: The synergistic application of AQ and paclitaxel exerted a strong inhibitory effect on the MCF-7 breast cancer cells. Extensive studies are imperative to better understand the action of bioactive mixes on the edible oyster fungus P. ostreatus. The gene knockout potential detected by CRISPR SpCas9 will aid in elite research into anticancer treatments.

A novel long non-coding RNA MIR4500HG003 promotes tumor metastasis through miR-483-3p-MMP9 axis in triple-negative breast cancer.

Breast cancer (BC) is the most common cancer and the leading cause of cancer-related deaths in women worldwide. The 5-year survival rate is over 90% in BC patients, but once BC cells metastasis into distal organs, it is dramatically decreasing to less than 30%. Especially, triple-negative breast cancer (TNBC) patients usually lead to poor prognosis and survival because of metastasis. Understanding the underline mechanisms of TNBC metastasis is a critical issue. Non-coding RNAs, including of lncRNAs and microRNAs, are non-protein-coding transcripts and have been reported as important regulators in TNBC metastasis. However, the underline mechanisms for non-coding RNAs regulating TNBC metastasis remain largely unclear. Here, we found that lncRNA MIR4500HG003 was highly expressed in highly metastatic MDA-MB-231 TNBC cells and overexpression of MIR4500HG003 enhanced metastasis ability in vitro and in vivo and promoted MMP9 expression. Furthermore, we found MIR4500HG003 physically interacted with miR-483-3p and reporter assay showed miR-483-3p attenuated MMP9 expression. Importantly, endogenous high expressions of MIR4500HG003 were correlated with tumor recurrence in TNBC patients with tumor metastasis. Taken together, our findings suggested that MIR4500HG003 promotes metastasis of TNBC through miR-483-3p-MMP9 signaling axis and may be used as potential prognostic marker for TNBC patients.

Similar Distribution between EPA-containing Phosphatidylcholine and Mesenchymal Stem Marker Positive Cells in the Aortic Wall of Abdominal Aortic Aneurysm Model Rat Fed a Low-EPA Content Diet.

Abdominal aortic aneurysm (AAA) is a vascular disease characterized by progressive dilation of the abdominal aorta. Previous studies have suggested that dietary components are closely associated with AAA. Among those dietary components, eicosapentaenoic acid (EPA) is considered to have suppressive effects on AAA. In the AAA wall of AAA model animals bred under EPA-rich condition, the distribution of EPA-containing phosphatidylcholine (EPA-PC) has been reported to be similar to that of the markers of mesenchymal stem cells (MSCs) and M2 macrophages. These data suggest that the suppressive effects of EPA on AAA are related to preferential distribution of specific cells in the aortic wall. However, the distribution of EPA-PC in the AAA wall of AAA model animals fed a diet containing small amounts of EPA, which has not been reported to inhibit AAA, has not yet been explored. In the present study, we visualized the distribution of EPA-PCs in the AAA wall of AAA model animals fed a diet containing small amounts of EPA (1.5% EPA in the fatty acid composition) to elucidate the vasoprotective effects of EPA. Positive areas for markers of MSCs were significantly higher in the region where EPA-PC was abundant compared to the regions where EPA-PC was weakly detected, but not for markers of M2 macrophages, matrix metalloproteinase (MMP)-2, and MMP-9. The distribution of MSC markers was similar to that of EPA-PC but not that of M2 macrophages and MMPs. These data suggest preferential incorporation of EPA into MSCs under the conditions used in this study. The incorporation of EPA into certain cells may differ according to dietary conditions, which affect the development of AAA.

IGF2BP3 regulates the expression of RRM2 and promotes the progression of rheumatoid arthritis via RRM2/Akt/MMP-9 pathway.

BACKGROUND: Rheumatoid arthritis (RA) is a common inflammatory and autoimmune disease. Ribonucleotide Reductase Regulatory Subunit M2 (RRM2) is a crucial and a rate-limiting enzyme responsible for deoxynucleotide triphosphate(dNTP) production. We have found a high expression level of RRM2 in patients with RA, but the molecular mechanism of its action remains unclear. METHODS: We analyzed the expression of hub genes in RA using GSE77298 datasets downloaded from Gene Expression Omnibus database. RRM2 and insulin-like growth factor-2 messenger ribonucleic acid (mRNA)-binding protein 3 (IGF2BP3) gene knockdown was achieved by infection with lentiviruses. The expression of RRM2, IGF2BP3, matrix metalloproteinase (MMP)-1, and MMP-9 were detected via western blotting assay. Cell viability was detected via 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. MeRIP-qRT-PCR was performed to test the interaction of IGF2BP3 and RRM2 mRNA via m6A modification. Cell proliferation was determined by clone formation assay. Migration and invasion assays were performed using transwell Boyden chamber. RESULTS: RRM2 and IGF2BP3 were highly expressed in clinical specimens and tumor necrosis factor alpha (TNF-α) and interleukin (IL)-1ß-stimulated synovial cells. RRM2 and IGF2BP3 knockdown inhibited the proliferation, migration, and invasion of MH7A cells. The inhibitory effects of IGF2BP3 knockdown were effectively reversed by simultaneously overexpressing RRM2 in MH7A cells. By analyzing N6-methyladenosine (m6A)2Target database, five m6A regulatory target binding sites for IGF2BP3 were identified in RRM2 mRNA, suggesting a direct relationship between IGF2BP3 and RRM2 mRNA. Additionally, in RRM2 small hairpin (sh)RNA lentivirus-infected cells, the levels of phosphorylated Akt and MMP-9 were significantly decreased compared with control shRNA lentivirus-infected cells. CONCLUSION: The present study demonstrated that RRM2 promoted the Akt phosphorylation leading to high expression of MMP-9 to promote the migration and invasive capacities of MH7A cells. Overall, IGF2BP promotes the expression of RRM2, and regulates the migration and invasion of MH7A cells via Akt/MMP-9 pathway to promote RA progression.

Interleukin-1 beta signals through the ERK signalling pathway to modulate human placental trophoblast migration and invasion in the first trimester of pregnancy.

INTRODUCTION: Interleukin-1 beta (IL-1ß) can promote cell migration, invasion and metastasis in various cancer cells. The mechanism of its role in human trophoblast has not been fully investigated. Therefore, we aimed to investigate the expression level of IL-1ß in first trimester decidua and placenta and its potential role in regulation of extravillous trophoblast cell (EVT) invasion and migration. METHODS: First trimester placenta and decidua were collected to study the expression levels of IL-1ß and its receptors by immunohistochemical staining. Primary isolates of first trimester EVT or the HTR-8/SVneo trophoblast like cell line were used to assess migration and invasion. Matrix metalloproteinase levels were assessed by gelatin zymography and ELISA. The phosphorylation profile of signaling pathway proteins was detected with the Proteome Profiler Human Phospho-Kinase Array Kit. Differentially expressed proteins in cells was detected and verified by Western Blot. RESULTS: IL-1ß, its receptors and antagonist are expressed in first trimester placenta and decidua, exogenous IL-1ß stimulates trophoblast cell outgrowth, migration and invasion through the ERK signaling pathway. IL-1ß was significantly increased in the placenta at 6-7 weeks gestation compared with 8-9 weeks gestation (P < 0.0001). Transwell and RTCA assays indicated that IL-1ß stimulates the invasion and migration of EVT. In addition, IL-1ß promoted the phosphorylation of ERK 1/2. It also promoted the expression of MMP2 and MMP9 in EVT as demonstrated by gelatin zymography assay and enzyme linked immunosorbent assay. DISCUSSION: This study demonstrated IL-1ß expression in placenta and decidua, and that it regulates EVT invasion and migration.

Intracranial aneurysm circulating exosome-derived LncRNA ATP1A1-AS1 promotes smooth muscle cells phenotype switching and apoptosis.

Exosomal long non-coding RNAs (LncRNAs) play a crucial role in the pathogenesis of cerebrovascular diseases. However, the expression profiles and functional significance of exosomal LncRNAs in intracranial aneurysms (IAs) remain poorly understood. Through high-throughput sequencing, we identified 1303 differentially expressed LncRNAs in the plasma exosomes of patients with IAs and healthy controls. Quantitative real-time polymerase chain reaction (qRT-PCR) verification confirmed the differential expression of LncRNAs, the majority of which aligned with the sequencing results. ATP1A1-AS1 showed the most significant upregulation in the disease group. Importantly, subsequent in vitro experiments validated that ATP1A1-AS1 overexpression induced a phenotype switching in vascular smooth muscle cells, along with promoting apoptosis and upregulating MMP-9 expression, potentially contributing to IAs formation. Furthermore, expanded-sample validation affirmed the high diagnostic value of ATP1A1-AS1. These findings suggest that ATP1A1-AS1 is a potential therapeutic target for inhibiting IAs progression and serves as a valuable clinical diagnostic marker.

Immunohistochemical properties of embryonic telocytes in a myogenic microenvironment.

Telocytes are a unique interstitial cell type that functions in adulthood and embryogenesis. They have characteristic immunohistochemical phenotypes while acquiring different immunohistochemical properties related to the organ microenvironment. The present study aims to investigate the immunohistochemical features of embryonic telocytes during myogenesis and describe their morphology using light microscopy and TEM. Telocytes represent a major cellular constituent in the interstitial elements. They had distinguished telopodes and podoms and formed a 3D interstitial network in the developing muscles. They formed heterocellular contact with myoblasts and nascent myotubes. Telocytes also had distinctive secretory activity. Telocytes identified by CD34. They also express CD68 and MMP-9 to facilitate the development of new tissues. Expression of CD21 by telocytes may reveal their function in immune defense. They also express VEGF, which regulates angiogenesis. In conclusion, the distribution and immunological properties of telocytes in the myogenic tissue indicate that telocytes provide biological and structural support in the development of the myogenic tissue architecture and organization.

Nitric oxide releasing nanofiber stimulates revascularization in response to ischemia via cGMP-dependent protein kinase.

Nitric oxide (NO) promotes angiogenesis via various mechanisms; however, the effective transmission of NO in ischemic diseases is unclear. Herein, we tested whether NO-releasing nanofibers modulate therapeutic angiogenesis in an animal hindlimb ischemia model. Male wild-type C57BL/6 mice with surgically-induced hindlimb ischemia were treated with NO-releasing 3-methylaminopropyltrimethoxysilane (MAP3)-derived or control (i.e., non-NO-releasing) nanofibers, by applying them to the wound for 20 min, three times every two days. The amount of NO from the nanofiber into tissues was assessed by NO fluorometric assay. The activity of cGMP-dependent protein kinase (PKG) was determined by western blot analysis. Perfusion ratios were measured 2, 4, and 14 days after inducing ischemia using laser doppler imaging. On day 4, Immunohistochemistry (IHC) with F4/80 and gelatin zymography were performed. IHC with CD31 was performed on day 14. To determine the angiogenic potential of NO-releasing nanofibers, aorta-ring explants were treated with MAP3 or control fiber for 20 min, and the sprout lengths were examined after 6 days. As per either LDPI (Laser doppler perfusion image) ratio or CD31 capillary density measurement, angiogenesis in the ischemic hindlimb was improved in the MAP3 nanofiber group; further, the total nitrate/nitrite concentration in the adduct muscle increased. The number of macrophage infiltrations and matrix metalloproteinase-9 (MMP-9) activity decreased. Vasodilator-stimulated phosphoprotein (VASP), one of the major substrates for PKG, increased phosphorylation in the MAP3 group. MAP3 nanofiber or NO donor SNAP (s-nitroso-n-acetyl penicillamine)-treated aortic explants showed enhanced sprouting in an ex vivo aortic ring assay, which was partially abrogated by KT5823, a potent inhibitor of PKG. These findings suggest that the novel NO-releasing nanofiber, MAP3 activates PKG and promotes therapeutic angiogenesis in response to hindlimb ischemia.

MMPs-responsive silk spheres for controlled drug release within tumor microenvironment.

Silk is a biocompatible and biodegradable material that enables the formation of various morphological forms, including nanospheres. The functionalization of bioengineered silk makes it possible to produce particles with specific properties. In addition to tumor cells, the tumor microenvironment (TME) includes stromal, immune, endothelial cells, signaling molecules, and the extracellular matrix (ECM). Matrix metalloproteinases (MMPs) are overexpressed in TME. We investigated bioengineered spider silks functionalized with MMP-responsive peptides to obtain targeted drug release from spheres within TME. Soluble silks MS12.2MS1, MS12.9MS1, and MS22.9MS2 and the corresponding silk spheres carrying MMP-2 or MMP-2/9 responsive peptides were produced, loaded with doxorubicin (Dox), and analyzed for their susceptibility to MMP-2/9 digestion. Although all variants of functionalized silks and spheres were specifically degraded by MMP-2/9, the MS22.9MS2 nanospheres showed the highest levels of degradation and release of Dox after enzyme treatment. Moreover, functionalized spheres were degraded in the presence of cancer cells releasing MMP-2/9. In the 2D and 3D spheroid cancer models, the MMP-2/9-responsive substrate was degraded and released from spheres when loaded into MS22.9MS2 particles but not into the control MS2 spheres. The present study demonstrated that a silk-based MMP-responsive delivery system could be used for controlled drug release within the tumor microenvironment.