ETS transcription factors regulate precise matrix metalloproteinase expression and follicle rupture in Drosophila.

Drosophila matrix metalloproteinase 2 (MMP2) is specifically expressed in posterior follicle cells of stage-14 egg chambers (mature follicles) and is crucial for the breakdown of the follicular wall during ovulation, a process that is highly conserved from flies to mammals. The factors that regulate spatiotemporal expression of MMP2 in follicle cells remain unknown. Here, we demonstrate crucial roles for the ETS-family transcriptional activator Pointed (Pnt) and its endogenous repressor Yan in the regulation of MMP2 expression. We found that Pnt is expressed in posterior follicle cells and overlaps with MMP2 expression in mature follicles. Genetic analysis demonstrated that pnt is both required and sufficient for MMP2 expression in follicle cells. In addition, Yan was temporally upregulated in stage-13 follicle cells to fine-tune Pnt activity and MMP2 expression. Furthermore, we identified a 1.1 kb core enhancer that is responsible for the spatiotemporal expression of MMP2 and contains multiple pnt/yan binding motifs. Mutation of pnt/yan binding sites significantly impaired the Mmp2 enhancer activity. Our data reveal a mechanism of transcriptional regulation of Mmp2 expression in Drosophila ovulation, which could be conserved in other biological systems.

The bHLH-PAS transcriptional complex Sim:Tgo plays active roles in late oogenesis to promote follicle maturation and ovulation.

Across species, ovulation is a process induced by a myriad of signaling cascades that ultimately leads to the release of encapsulated oocytes from follicles. Follicles first need to mature and gain ovulatory competency before ovulation; however, the signaling pathways regulating follicle maturation are incompletely understood in Drosophila and other species. Our previous work has shown that the bHLH-PAS transcription factor Single-minded (Sim) plays important roles in follicle maturation downstream of the nuclear receptor Ftz-f1 in Drosophila. Here, we demonstrate that Tango (Tgo), another bHLH-PAS protein, acts as a co-factor of Sim to promote follicle cell differentiation from stages 10 to 12. In addition, we discover that re-upregulation of Sim in stage-14 follicle cells is also essential to promote ovulatory competency by upregulating octopamine receptor in mushroom body (OAMB), matrix metalloproteinase 2 (Mmp2) and NADPH oxidase (NOX), either independently of or in conjunction with the zinc-finger protein Hindsight (Hnt). All these factors are crucial for successful ovulation. Together, our work indicates that the transcriptional complex Sim:Tgo plays multiple roles in late-stage follicle cells to promote follicle maturation and ovulation.

Dendrite regeneration of adult Drosophila sensory neurons diminishes with aging and is inhibited by epidermal-derived matrix metalloproteinase 2.

Dendrites possess distinct structural and functional properties that enable neurons to receive information from the environment as well as other neurons. Despite their key role in neuronal function, current understanding of the ability of neurons to regenerate dendrites is lacking. This study characterizes the structural and functional capacity for dendrite regeneration in vivo in adult animals and examines the effect of neuronal maturation on dendrite regeneration. We focused on the class IV dendritic arborization (c4da) neuron of the Drosophila sensory system, which has a dendritic arbor that undergoes dramatic remodeling during the first 3 d of adult life and then maintains a relatively stable morphology thereafter. Using a laser severing paradigm, we monitored regeneration after acute and spatially restricted injury. We found that the capacity for regeneration was present in adult neurons but diminished as the animal aged. Regenerated dendrites recovered receptive function. Furthermore, we found that the regenerated dendrites show preferential alignment with the extracellular matrix (ECM). Finally, inhibition of ECM degradation by inhibition of matrix metalloproteinase 2 (Mmp2) to preserve the extracellular environment characteristics of young adults led to increased dendrite regeneration. These results demonstrate that dendrites retain regenerative potential throughout adulthood and that regenerative capacity decreases with aging.

Flavonifractor plautii Protects Against Elevated Arterial Stiffness.

BACKGROUND: Dysbiosis of gut microbiota plays a pivotal role in vascular dysfunction and microbial diversity was reported to be inversely correlated with arterial stiffness. However, the causal role of gut microbiota in the progression of arterial stiffness and the specific species along with the molecular mechanisms underlying this change remain largely unknown. METHODS: Participants with elevated arterial stiffness and normal controls free of medication were matched for age and sex. The microbial composition and metabolic capacities between the 2 groups were compared with the integration of metagenomics and metabolomics. Subsequently, Ang II (angiotensin II)-induced and humanized mouse model were employed to evaluate the protective effect of Flavonifractor plautii (F plautii) and its main effector cis-aconitic acid. RESULTS: Human fecal metagenomic sequencing revealed a significantly high abundance and centrality of F plautii in normal controls, which was absent in the microbial community of subjects with elevated arterial stiffness. Moreover, blood pressure only mediated part of the effect of F plautii on lower arterial stiffness. The microbiome of normal controls exhibited an enhanced capacity for glycolysis and polysaccharide degradation, whereas, those of subjects with increased arterial stiffness were characterized by increased biosynthesis of fatty acids and aromatic amino acids. Integrative analysis with metabolomics profiling further suggested that increased cis-aconitic acid served as the main effector for the protective effect of F plautii against arterial stiffness. Replenishment with F plautii and cis-aconitic acid improved elastic fiber network and reversed increased pulse wave velocity through the suppression of MMP-2 (matrix metalloproteinase-2) and inhibition of MCP-1 (monocyte chemoattractant protein-1) and NF-κB (nuclear factor kappa-B) activation in both Ang II-induced and humanized model of arterial stiffness. CONCLUSIONS: Our translational study identifies a novel link between F plautii and arterial function and raises the possibility of sustaining vascular health by targeting gut microbiota.

Mechanism of pro-MMP9 activation in co-culture of pro-inflammatory macrophages and cardiomyocytes.

OBJECTIVE: A wide range of cardiac diseases is associated with inflammation. "Inflamed" heart tissue is infiltrated with pro-inflammatory macrophages which extensively secrete matrix metalloproteinase 9 (MMP9), a regulator of extracellular matrix turnover. As MMP9 is released from macrophages in a latent form, it requires activation. The present study addresses the role of cardiomyocytes in the course of this activation process. METHODS AND RESULTS: In mono- and co-cultures of pro-inflammatory rat macrophages (bone marrow-derived and peritoneal) and cardiomyocytes (H9C2 cell line) gelatin zymography demonstrated that activated macrophages robustly secreted latent pro-MMP9, whereas cardiomyocytes could not produce the enzyme. Co-culturing of the two cell species was critical for pro-MMP9 activation and was also accompanied by processing of cardiomyocyte-secreted pro-MMP2. A cascade of pro-MMP9 activation was initiated on macrophage membrane with pro-MMP2 cleavage. Namely, pro-inflammatory macrophages expressed an active membrane type 1 MMP (MT1MMP), which activated pro-MMP2, which in turn converted pro-MMP9. Downregulation of MT1MMP in macrophages by siRNA abolished activation of both pro-MMP2 and pro-MMP9 in co-culture. In addition, both cell species secreted MMP13 as a further pro-MMP9 activator. In co-culture, activation of pro-MMP13 occurred on membranes of macrophages and was enhanced in presence of active MMP2. Using incubations with recombinant MMPs and isolated macrophage membranes, we demonstrated that while both MMP2 and MMP13 individually had the ability to activate pro-MMP9, their combined action provided a synergistic effect. CONCLUSION: Activation of pro-MMP9 in a co-culture of pro-inflammatory macrophages and cardiomyocytes was the result of a complex interaction of several MMPs on the cell membrane and in the extracellular space. Both cell types contributed critically to pro-MMP9 processing.

Epithelial-mesenchymal plasticity in kidney fibrosis.

Epithelial-mesenchymal transition (EMT) is an important biological process contributing to kidney fibrosis and chronic kidney disease. This process is characterized by decreased epithelial phenotypes/markers and increased mesenchymal phenotypes/markers. Tubular epithelial cells (TECs) are commonly susceptible to EMT by various stimuli, for example, transforming growth factor-ß (TGF-ß), cellular communication network factor 2, angiotensin-II, fibroblast growth factor-2, oncostatin M, matrix metalloproteinase-2, tissue plasminogen activator (t-PA), plasmin, interleukin-1ß, and reactive oxygen species. Similarly, glomerular podocytes can undergo EMT via these stimuli and by high glucose condition in diabetic kidney disease. EMT of TECs and podocytes leads to tubulointerstitial fibrosis and glomerulosclerosis, respectively. Signaling pathways involved in EMT-mediated kidney fibrosis are diverse and complex. TGF-ß1/Smad and Wnt/ß-catenin pathways are the major venues triggering EMT in TECs and podocytes. These two pathways thus serve as the major therapeutic targets against EMT-mediated kidney fibrosis. To date, a number of EMT inhibitors have been identified and characterized. As expected, the majority of these EMT inhibitors affect TGF-ß1/Smad and Wnt/ß-catenin pathways. In addition to kidney fibrosis, these EMT-targeted antifibrotic inhibitors are expected to be effective for treatment against fibrosis in other organs/tissues.

WISP1 induces the expression of macrophage migration inhibitory factor in human lung fibroblasts through Src kinases and EGFR-activated signaling pathways.

Wnt1-inducible signaling protein 1 (WISP1/CCN4) is a secreted matricellular protein that is implicated in lung and airway remodeling. The macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine that has been associated with chronic lung diseases. In this study, we aimed to investigate the WISP1 signaling pathway and its ability to induce the expression of MIF in primary cultures of fibroblasts from normal human lungs (HLFs). Our results showed that WISP1 significantly stimulated the expression of MIF in a concentration- and time-dependent fashion. In WISP1-induced expression of MIF, αvß5-integrin and chondroitin sulfate proteoglycans as well as Src tyrosine kinases, MAP kinases, phosphatidylinositol 3-kinase/Akt, PKC, and NF-κB were involved. WISP1-induced expression of MIF was attenuated in the presence of the Src kinase inhibitor PP2 or the MIF tautomerase activity inhibitor ISO-1. Moreover, WISP1 significantly increased the phosphorylation and activation of EGF receptor (EGFR) through transactivation by Src kinases. WISP1 also induced the expression of MIF receptor CD74 and coreceptor CD44, through which MIF exerts its effects on HLFs. In addition, it was found that MIF induced its own expression, as well as its receptors CD74/CD44, acting in an autocrine manner. Finally, WISP1-induced MIF promoted the expression of cyclooxygenase 2, prostaglandin E2, IL-6, and matrix metalloproteinase-2 demonstrating the regulatory role of WISP1-MIF axis in lung inflammation and remodeling involving mainly integrin αvß5, Src kinases, PKC, NF-κB, and EGFR. The specific signaling pathways involved in WISP1-induced expression of MIF may prove to be excellent candidates for novel targets to control inflammation in chronic lung diseases.NEW & NOTEWORTHY The present study demonstrates for the first time that Wnt1-inducible signaling protein 1 (WISP1) regulates migration inhibitory factor (MIF) expression and activity and identifies the main signaling pathways involved. The newly discovered WISP1-MIF axis may drive lung inflammation and could result in the design of novel targeted therapies in inflammatory lung diseases.

Activation of the alternative complement pathway modulates inflammation in thoracic aortic aneurysm/dissection.

Thoracic aortic aneurysm/dissection (TAAD) is a lethal vascular disease, and several pathological factors participate in aortic medial degeneration. We previously discovered that the complement C3a-C3aR axis in smooth muscle cells promotes the development of thoracic aortic dissection (TAD) through regulation of matrix metalloproteinase 2. However, discerning the specific complement pathway that is activated and elucidating how inflammation of the aortic wall is initiated remain unknown. We ascertained that the plasma levels of C3a and C5a were significantly elevated in patients with TAD and that the levels of C3a, C4a, and C5a were higher in acute TAD than in chronic TAD. We also confirmed the activation of the complement in a TAD mouse model. Subsequently, knocking out Cfb (Cfb) or C4 in mice with TAD revealed that the alternative pathway and Cfb played a significant role in the TAD process. Activation of the alternative pathway led to generation of the anaphylatoxins C3a and C5a, and knocking out their receptors reduced the recruitment of inflammatory cells to the aortic wall. Moreover, we used serum from wild-type mice or recombinant mice Cfb as an exogenous source of Cfb to treat Cfb KO mice and observed that it exacerbated the onset and rupture of TAD. Finally, we knocked out Cfb in the FBN1C1041G/+ Marfan-syndrome mice and showed that the occurrence of TAA was reduced. In summary, the alternative complement pathway promoted the development of TAAD by recruiting infiltrating inflammatory cells. Targeting the alternative pathway may thus constitute a strategy for preventing the development of TAAD.NEW & NOTEWORTHY The alternative complement pathway promoted the development of TAAD by recruiting infiltrating inflammatory cells. Targeting the alternative pathway may thus constitute a strategy for preventing the development of TAAD.

Data-Independent Acquisition Proteomics and N-Terminomics Methods Reveal Alterations in Mitochondrial Function and Metabolism in Ischemic-Reperfused Hearts.

Myocardial ischemia-reperfusion (IR) (stunning) injury triggers changes in the proteome and degradome of the heart. Here, we utilize quantitative proteomics and comprehensive degradomics to investigate the molecular mechanisms of IR injury in isolated rat hearts. The control group underwent aerobic perfusion, while the IR injury group underwent 20 min of ischemia and 30 min of reperfusion to induce a stunning injury. As MMP-2 activation has been shown to contribute to myocardial injury, hearts also underwent IR injury with ARP-100, an MMP-2-preferring inhibitor, to dissect the contribution of MMP-2 to IR injury. Using data-independent acquisition (DIA) and mass spectroscopy, we quantified 4468 proteins in ventricular extracts, whereby 447 proteins showed significant alterations among the three groups. We then used subtiligase-mediated N-terminomic labeling to identify more than a hundred specific cleavage sites. Among these protease substrates, 15 were identified following IR injury. We identified alterations in numerous proteins involved in mitochondrial function and metabolism following IR injury. Our findings provide valuable insights into the biochemical mechanisms of myocardial IR injury, suggesting alterations in reactive oxygen/nitrogen species handling and generation, fatty acid metabolism, mitochondrial function and metabolism, and cardiomyocyte contraction.

Exosomes isolated from TNF-α-treated bone marrow mesenchymal stem cells ameliorate pelvic floor dysfunction in rats.

Exosomes derived from bone marrow-derived mesenchymal stem cells (BMSCs) can alleviate the symptoms of pelvic floor dysfunction (PFD) in rats. However, the potential therapeutical effects of exosomes derived from BMSCs treated with tumour necrosis factor (TNF)-α on the symptoms of PFD in rats are unknown. Exosomes extracted from BMSCs treated with or without TNF-α were applied to treat PFD rats. Our findings revealed a significant elevation in interleukin (IL)-6 and TNF-α, and matrix metalloproteinase-2 (MMP2) levels in the vaginal wall tissues of patients with pelvic organ prolapse (POP) compared with the control group. Daily administration of exosomes derived from BMSCs, treated either with or without TNF-α (referred to as Exo and TNF-Exo), resulted in increased void volume and bladder void pressure, along with reduced peak bladder pressure and leak point pressure in PFD rats. Notably, TNF-Exo treatment demonstrated superior efficacy in restoring void volume, bladder void pressure and the mentioned parameters compared with Exo treatment. Importantly, TNF-Exo exhibited greater potency than Exo in restoring the levels of multiple proteins (Elastin, Collagen I, Collagen III, IL-6, TNF-α and MMP2) in the anterior vaginal walls of PFD rats. The application of exosomes derived from TNF-α-treated BMSCs holds promise as a novel therapeutic approach for treating PFD.

Irradiated riboflavin over nonradiated one: Potent antimigratory, antiproliferative and cytotoxic effects on glioblastoma cells.

Riboflavin is a water-soluble yellowish vitamin and is controversial regarding its effect on tumour cells. Riboflavin is a powerful photosensitizer that upon exposure to radiation, undergoes an intersystem conversion with molecular oxygen, leading to the production of ROS. In the current study, we sought to ascertain the impact of irradiated riboflavin on C6 glioblastoma cells regarding proliferation, cell death, oxidative stress and migration. First, we compared the proliferative behaviour of cells following nonradiated and radiated riboflavin. Next, we performed apoptotic assays including Annexin V and caspase 3, 7 and 9 assays. Then we checked on oxidative stress and status by flow cytometry and ELISA kits. Finally, we examined inflammatory change and levels of MMP2 and SIRT1 proteins. We caught a clear antiproliferative and cytotoxic effect of irradiated riboflavin compared to nonradiated one. Therefore, we proceeded with our experiments using radiated riboflavin. In all apoptotic assays, we observed a dose-dependent increase. Additionally, the levels of oxidants were found to increase, while antioxidant levels decreased following riboflavin treatment. In the inflammation analysis, we observed elevated levels of both pro-inflammatory and anti-inflammatory cytokines. Additionally, after treatment, we observed reduced levels of MMP2 and SIRT. In conclusion, radiated riboflavin clearly demonstrates superior antiproliferative and apoptotic effects on C6 cells at lower doses compared to nonradiated riboflavin.

Chlorotoxin binds to both matrix metalloproteinase 2 and neuropilin 1.

Chlorotoxin (CTX), a scorpion venom-derived 36-residue miniprotein, binds to and is taken up selectively by glioblastoma cells. Previous studies provided controversial results concerning target protein(s) of CTX. These included CLC3 chloride channel, matrix metalloproteinase 2 (MMP-2), regulators of MMP-2, annexin A2, and neuropilin 1 (NRP1). The present study aimed at clarifying which of the proposed binding partners can really interact with CTX using biochemical methods and recombinant proteins. For this purpose, we established two new binding assays based on anchoring the tested proteins to microbeads and quantifying the binding of CTX by flow cytometry. Screening of His-tagged proteins anchored to cobalt-coated beads indicated strong interaction of CTX with MMP-2 and NRP1, whereas binding to annexin A2 was not confirmed. Similar results were obtained with fluorophore-labeled CTX and CTX-displaying phages. Affinity of CTX to MMP-2 and NRP1 was assessed by the "immunoglobulin-coated bead" test, in which the proteins were anchored to beads by specific antibodies. This assay yielded highly reproducible data using both direct titration and displacement approach. The affinities of labeled and unlabeled CTX appeared to be similar for both MMP-2 and NRP1 with estimated KD values of 0.5 to 0.7 µM. Contrary to previous reports, we found that CTX does not inhibit the activity of MMP-2 and that CTX not only with free carboxyl end but also with carboxamide terminal end binds to NRP1. We conclude that the presented robust assays could also be applied for affinity-improving studies of CTX to its genuine targets using phage display libraries.

miR-146a regulates emphysema formation and abnormal inflammation in the lungs of two mouse models.

miR-146a, a microRNA (miRNA) that regulates inflammatory responses, plays an important role in many inflammatory diseases. Although an in vitro study had suggested that miR-146a is involved in abnormal inflammatory response, being a critical factor in the pathogenesis of chronic obstructive pulmonary disease (COPD), in vivo evidence of its pathogenic role in COPD remains limited. Eight-week-old male B6(FVB)-Mir146tm1.1Bal/J [miR-146a knockout (KO)] and C57BL/6J mice were intratracheally administered elastase and evaluated after 28 days or exposed to cigarette smoke (CS) and evaluated after 5 mo. miR-146a expression was significantly increased in C57BL/6J mouse lungs due to elastase administration (P = 0.027) or CS exposure (P = 0.019) compared with that in the control group. Compared with C57BL/6J mice, elastase-administered miR-146a-KO mice had lower average computed tomography (CT) values (P = 0.017) and increased lung volume-to-weight ratio (P = 0.016), mean linear intercept (P < 0.001), and destructive index (P < 0.001). Moreover, total cell (P = 0.006), macrophage (P = 0.001), neutrophil (P = 0.026), chemokine (C-X-C motif) ligand 2/macrophage inflammatory protein-2 [P = 0.045; in bronchoalveolar lavage fluid (BALF)], cyclooxygenase-2, and matrix metalloproteinase-2 levels were all increased (in the lungs). Following long-term CS exposure, miR-146a-KO mice showed a greater degree of emphysema formation in their lungs and inflammatory response in the BALF and lungs than C57BL/6J mice. Collectively, miR-146a protected against emphysema formation and the associated abnormal inflammatory response in two murine models.NEW & NOTEWORTHY This study demonstrates that miR-146a expression is upregulated in mouse lungs because of elastase- and CS-induced emphysema and that the inflammatory response by elastase or CS is enhanced in the lungs of miR-146a-KO mice than in those of control mice, resulting in the promotion of emphysema. This is the first study to evaluate the protective role of miR-146a in emphysema formation and the associated abnormal inflammatory response in different in vivo models.

Inhibition of heme oxygenase 1 alleviates thoracic aortic aneurysm via restoration of extracellular matrix.

BACKGROUND: Thoracic aortic aneurysm (TAA) is a silent but life-threatening cardiovascular disease. Heme oxygenase 1 (HO-1) plays an important role in the cardiovascular diseases but is poorly understood in TAA. This study aims at investigating the role of HO-1 in TAA. METHODS: Single-cell RNA sequencing, Western blot and histological assay were performed to identify specific cellular expression of HO-1 in both human and ß-aminopropionitrile (BAPN)-induced mice TAA. Zinc protoporphyrin (ZnPP), a pharmacological inhibitor of HO-1, was used to investigate whether inhibition of HO-1 could attenuate BAPN-induced TAA in rodent model. Histological assay, Western blot assay, and mRNA sequencing were further performed to explore the underlying mechanisms. RESULTS: Single-cell transcriptomic analyses of 113,800 thoracic aortic cells identified an increase of HO-1(+) macrophage in aneurysmal thoracic aorta from BAPN-induced TAA mice and TAA patients. Histological assay verified HO-1 overexpression in clinical TAA specimens, which was co-localized with CD68(+) macrophage. HO-1(+) macrophage was closely associated with pro-inflammatory response and immune activation. Inhibition of HO-1 through ZnPP significantly alleviated BAPN-induced TAA in mice and restored extracellular matrix (ECM) in vivo. Further experiments showed that ZnPP treatment suppressed the expression of matrix metalloproteinases (MMPs) in aneurysmal thoracic aortic tissues from BAPN-induced TAA mice, including MMP2 and MMP9. Macrophages from myeloid specific HO-1 knockout mice displayed weakened pro-inflammatory activity and ECM degradation capability. CONCLUSION: HO-1(+) macrophage subgroup is a typical hallmark of TAA. Inhibition of HO-1 through ZnPP alleviates BAPN-induced TAA in mice, which might work through restoration of ECM via suppressing MMP2 and MMP9 expression.

Metabolically healthy/unhealthy obesity and breast cancer: A possible role of plasma-derived extracellular vesicles on the cancerous behavior of triple-negative breast cancer.

PURPOSE: Obesity has known detrimental effects on breast cancer (BC) development and progression. However, it's essential to consider the obesity phenotype based on metabolic health. This study aims to evaluate the impact of circulating extracellular vesicles (EVs) from women with metabolically healthy or unhealthy normal weight, overweight, and obesity on MDA-MB-231 cell migration, invasion, and apoptosis. METHODS: Plasma EVs were isolated from different obesity phenotypes in women. EVs were characterized and EVs uptake by MDA-MB-231 cells was assessed. MDA-MB-231 cell lines were treated with EVs obtained from various studied groups, and migration, invasion, MMP-2 and MMP-9 activity, Bax and Bcl-2 mRNA expression, p-53 and Thr55 p-p53 protein expression, and apoptosis were assessed. RESULTS: EVs from obese individuals, regardless of phenotype, increased invasion and MMP-2 activity compared to healthy normal-weight EVs. Normal-weight EVs led to higher invasion under unhealthy conditions. BC cell migration was enhanced by EVs from healthy obese individuals compared to healthy normal-weight EVs. EVs from unhealthy obese women exhibited significantly lower p53/p-p53 levels and reduced apoptosis compared to healthy obese groups. CONCLUSION: It appears that EVs from both normal-weight women with unhealthy conditions and those with obesity or overweight, irrespective of metabolic status, worsened the cancerous behavior of TNBC cells. Therefore, considering metabolic health, in addition to BMI, is crucial for understanding obesity-related disorders.

SPOCK2 modulates neuropathic pain by interacting with MT1-MMP to regulate astrocytic MMP-2 activation in rats with chronic constriction injury.

BACKGROUND: Neuropathic pain (NP) is a kind of intractable pain. The pathogenesis of NP remains a complicated issue for pain management practitioners. SPARC/osteonectin, CWCV, and Kazal-like domains proteoglycan 2 (SPOCK2) are members of the SPOCK family that play a significant role in the development of the central nervous system. In this study, we investigated the role of SPOCK2 in the development of NP in a rat model of chronic constriction injury (CCI). METHODS: Sprague-Dawley rats were randomly grouped to establish CCI models. We examined the effects of SPOCK2 on pain hpersensitivity and spinal astrocyte activation after CCI-induced NP. Paw withdrawal threshold (PWT) and paw withdrawal latency (PWL) were used to reflects the pain behavioral degree. Molecular mechanisms involved in SPOCK2-mediated NP in vivo were examined by western blot analysis, immunofluorescence, immunohistochemistry, and co-immunoprecipitation. In addition, we examined the SPOCK2-mediated potential protein-protein interaction (PPI) in vitro coimmunoprecipitation (Co-IP) experiments. RESULTS: We founded the expression level of SPOCK2 in rat spinal cord was markedly increased after CCI-induced NP, while SPOCK2 downregulation could partially relieve pain caused by CCI. Our research showed that SPOCK2 expressed significantly increase in spinal astrocytes when CCI-induced NP. In addition, SPOCK2 could act as an upstream signaling molecule to regulate the activation of matrix metalloproteinase-2 (MMP-2), thus affecting astrocytic ERK1/2 activation and interleukin (IL)-1ß production in the development of NP. Moreover, in vitro coimmunoprecipitation (Co-IP) experiments showed that SPOCK2 could interact with membrane-type 1 matrix metalloproteinase (MT1-MMP/MMP14) to regulate MMP-2 activation by the SPARC extracellular (SPARC_EC) domain. CONCLUSIONS: Research shows that SPOCK2 can interact with MT1-MMP to regulate MMP-2 activation, thus affecting astrocytic ERK1/2 activation and IL-1ß production to achieve positive promotion of NP.

BATF promotes extramedullary infiltration through TGF-ß1/Smad/MMPs axis in acute myeloid leukemia.

Acute myeloid leukemia (AML) is one of the most prevalent types of leukemia and is challenging to cure for most patients. Basic Leucine Zipper ATF-Like Transcription Factor (BATF) has been reported to participate in the development and progression of numerous tumors. However, its role in AML is largely unknown. In this study, the expression and prognostic value of BATF were examined in AML. Our results demonstrated that BATF expression was upregulated in AML patients, which was significantly correlated with poor clinical characteristics and survival. Afterward, functional experiments were performed after knocking down or overexpressing BATF by transfecting small interfering RNAs and overexpression plasmids into AML cells. Our findings revealed that BATF promoted the migratory and invasive abilities of AML cells in vitro and in vivo. Moreover, the target genes of BATF were searched from databases to explore the binding of BATF to the target gene using ChIP and luciferase assays. Notably, our observations validated that BATF is bound to the promoter region of TGF-ß1, which could transcriptionally enhance the expression of TGF-ß1 and activate the TGF-ß1/Smad/MMPs signaling pathway. In summary, our study established the aberrantly high expression of BATF and its pro-migratory function via the TGF-ß1-Smad2/3-MMP2/9 axis in AML, which provides novel insights into extramedullary infiltration of AML.

Resistance training modifies of serum levels of matrix metalloproteinase 2 and tissue inhibitor of matrix metalloproteinases in multiple sclerosis women - a randomized controlled trail.

The objectives of the present study was to investigate the effects of resistance training (RT) on serum levels of controlling blood-brain barrier (BBB) permeability indices and cognitive performance in MS women (MS-W). In this randomized control trail study (IRCT registration code: IRCT20120912010824N3, 07.09.2023), twenty-five MS-W were randomly divided into sedentary (MS) and resistance exercise (12 weeks/3 times per week/ 60-80% of 1RM) (MS + RT) groups. Fifteen healthy aged-matched women participated as a control group (HCON). The serum level of matrix metalloproteinase-2 (MMP-2), matrix metallopeptidase-9 (MMP-9), tissue metalloproteinase inhibitors-1 (TIMP-1), tissue metalloproteinase inhibitors-2 (TIMP-2), and S100 calcium-binding protein B (S100B) were assessed. In addition, cognitive performance was assessed pre- and post- intervention with the Brief International Cognitive Assessment for MS (BICAMS). A significant reduction in MMP-2, TIMP-2 serum levels, and MMP-2/TIMP-2 ratio were observed in post-test for MS + RT group (p < 0.01) in comparison to the HCON and MS groups; however, no changes were observed in MMP-9, TIMP-1, S100B and MMP-9/TIMP-1 ratio after RT (p > 0.05). The verbal learning was improved in post-test for MS + RT group (p < 0.01), although no change were observed for visuospatial memory and information processing speed (p > 0.05). These findings suggest that resistance training can modify some indices of BBB permeability and improve verbal learning in MS-W. The findings may also be beneficial as a non-pharmacological intervention to reduce inflammation.

Autophagy Induced by Low Shear Stress Leads to Endothelial Glycocalyx Disruption.

INTRODUCTION: Previous studies have confirmed that low shear stress (LSS) induces glycocalyx disruption, leading to endothelial dysfunction. However, the role of autophagy in LSS-induced glycocalyx disruption and relevant mechanism are not clear. In this study, we hypothesized that LSS may promote autophagy, disrupting the endothelium glycocalyx. METHODS: Human umbilical vein endothelial cells were subjected to physiological shear stress and LSS treatments, followed by the application of autophagy inducers and inhibitors. Additionally, cells were treated with specific matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) inhibitor. The expression of autophagic markers, glycocalyx, MMP-2, and MMP-9 was measured. RESULTS: LSS impacted the expression of endothelium autophagy markers, increasing the expression of LC3II.LC3I-1 and Beclin-1, and decreasing the levels of p62, accompanied by glycocalyx disturbance. Moreover, LSS upregulated the expression of MMP-2 and MMP-9 and downregulated the levels of syndecan-1 and heparan sulfate (HS). Additionally, expression of MMP-2 and MMP-9 was increased by an autophagy promoter but was decreased by autophagy inhibitor treatment under LSS. Autophagy and MMP-2 and MMP-9 further caused glycocalyx disruption. CONCLUSION: LSS promotes autophagy, leading to glycocalyx disruption. Autophagy increases the expression of MMP-2 and MMP-9, which are correlated with the glycocalyx destruction induced by LSS.

Protein S-palmitoylation enhances profibrotic signaling in response to cadmium.

Cadmium (Cd) is a naturally occurring, toxic environmental metal found in foods. Humans do not have an efficient mechanism for Cd elimination; thus, Cd burden in humans increases with age. Cell and mouse studies show that Cd burden from low environmental levels of exposure impacts lung cell metabolism, proliferation signaling and cell growth as part of disease-promoting profibrotic responses in the lungs. Prior integrative analysis of metabolomics and transcriptomics identified the zDHHC11 transcript as a central functional hub in response to Cd dose. zDHHC11 encodes a protein S-palmitoyltransferase, but no evidence is available for effects of Cd on protein S-palmitoylation. In the present research, we studied palmitoylation changes in response to Cd and found increased protein S-palmitoylation in human lung fibroblasts that was inhibited by 2-bromopalmitate (2-BP), an irreversible palmitoyltransferase inhibitor. Mass spectrometry-based proteomics showed palmitoylation of proteins involved in divalent metal transport and in fibrotic signaling. Mechanistic studies showed that 2-BP inhibited palmitoylation of divalent metal ion transporter ZIP14 and also inhibited cellular Cd uptake. Transcription analyses showed that Cd stimulated transforming growth factor (TGF)-ß1 and ß3 expression within 8 h and lung fibrotic markers α-smooth muscle actin, matrix metalloproteinase-2, and collagen 1α1 gene expression and that these effects were blocked by 2-BP. Because 2-BP also blocked palmitoylation of proteins controlled by TGFß1, these results show that palmitoylation impacts Cd-dependent fibrotic signaling both by enhancing cellular Cd accumulation and by supporting post-translational processing of TGFß1-dependent proteins.