And Metalloproteinase With Thrombospondin Motifs Research Articles

A Disintegrin and Metalloproteinase with Thrombospondin Motifs 4 Regulates Pulmonary Vascular Hyperpermeability through Destruction of Glycocalyx in Acute Respiratory Distress Syndrome.

Acute respiratory distress syndrome (ARDS) has no specific and effective treatment, and there is an urgent need to understand its pathogenesis. Therefore, based on the hypothesis that molecules whose expression is upregulated in injured pulmonary vascular endothelial cells (VECs) are involved in the pathogenesis of ARDS, we conducted a study to elucidate the molecular mechanisms and identify target factors for treatment. Primary human lung microvascular endothelial cells (HMVEC-Ls) were stimulated with lipopolysaccharide (LPS) or poly (I:C) and analyzed via a microarray to identify target genes for ARDS. We found that a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) was induced in murine lung VECs in an LPS-mediated ARDS model. Elevated ADAMTS4 was also observed by the immunostaining of lung samples from ARDS patients. The suppression of ADAMTS4 by siRNA in VECs ameliorated LPS-stimulated vascular permeability. The impairment of the cell surface expression of syndecan-1, a marker of the glycocalyx that is an extracellular matrix involved in vascular permeability, was dramatically inhibited by ADAMTS4 suppression. In addition, the suppression of ADAMTS4 protected against LPS-induced reductions in syndecan-1 and the adherens junction protein vascular endothelial cadherin. These results suggest that ADAMTS4 regulates VEC permeability in ARDS and may be a predictive marker and therapeutic target for ARDS.

ADAMTS-1 has nuclear localization in cells with epithelial origin and leads to decreased cell migration

In the study of tumorigenesis, the involvement of molecules within the extracellular matrix (ECM) is crucial. ADAMTSs (A Disintegrin and Metalloproteinase with Thrombospondin motifs), a group of secreted proteases known for their role in ECM remodeling, were primarily considered to be extracellular proteases. However, our research specifically detected ADAMTS-1, a member of this family, predominantly within the nucleus of mammary cells. Our main objective was to understand the mechanism of ADAMTS-1 translocation to the nucleus and its functional significance in this cellular compartment. Our investigation uncovered that nuclear ADAMTS-1 was present in cells exhibiting an epithelial phenotype, while cells of mesenchymal origin contained the protease in the cytoplasm. Moreover, disruption of ADAMTS-1 secretion, induced by Monensin treatment, resulted in its accumulation in the cytoplasm. Notably, our research indicated that alterations in the secretory pathways could influence the protease's compartmentalization. Additionally, experiments with conditioned medium from cells containing nuclear ADAMTS-1 demonstrated its internalization into the nucleus by HT-1080 cells and fibroblasts. Furthermore, heightened levels of ADAMTS-1 within the ECM reduced the migratory potential of mesenchymal cells. This highlights the potential significance of nuclear ADAMTS-1 as a critical component within the tumor microenvironment due to its functional activity in this specific cellular compartment.

Molecular analyses of ADAMTS-1, -4, -5, and IL-17 a cytokine relationship in patients with ulcerative colitis

BackgroundUlcerative colitis (UC) is a chronic inflammatory bowel disease that develops due to the impaired immune response in genetically susceptible individuals, and its etiopathogenesis is not fully elucidated. IL-17 A is a cytokine that is produced by a type of immune cell called Th17 cells and is involved in the immune response and inflammation. On the other hand, ADAMTS-1, -4, and − 5 are enzymes that are involved in the breakdown of extracellular matrix proteins, including proteoglycans, which are important components of the intestinal wall. This study aimed to evaluate the relationship between interleukin 17 (IL-17 A) cytokine, which plays a role in the pathogenesis of ulcerative colitis, and the inflammation-controlled a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-1, -4, and − 5 protein members.MethodsBowel tissue samples and blood serum from 51 patients with UC and 51 healthy controls were included in this study. mRNA expression levels of the ADAMTS-1, -4, -5, and IL-17 A were analyzed by RT-qPCR, and immunohistochemical analyses were performed to evaluate ADAMTS-1, -4, -5, and IL-17 A proteins in tissue samples. In addition, ELISA analysis determined serum levels of the ADAMTS-1, -4, -5, and IL-17 A.ResultsRT-qPCR results reveal that the expression of ADAMTS-1, -4, -5, and IL-17 A genes in the UC tissue samples were significantly high according to the control tissue samples. Also, ADAMTS-1, -4, -5, and IL-17 A proteins revealed enhanced expression pattern UC groups according to the control. Also, ADAMTS-1, -4, -5, and IL-17 A protein showed cytoplasmic localization patterns in both control and UC groups. The serum levels of ADAMTS-1,-5, and IL-17 A were significantly higher in UC samples than in the control group.ConclusionsWe observed a positive correlation between the ADAMTS-1, -5 and IL17A cytokine expression in UC samples. These results provide a new understanding of controlling crucial ADAMTS family protein members by IL-17 A cytokines with UC.

Cangxi Tongbi Capsules promote chondrocyte autophagy by regulating circRNA＿0008365/miR-1271/p38 MAPK pathway to inhibit development of knee osteoarthritis

To investigate the mechanism by which Cangxi Tongbi Capsules promote chondrocyte autophagy to inhibit knee osteoarthritis(KOA) progression by regulating the circRNA＿0008365/miR-1271/p38 mitogen-activated protein kinase(MAPK) pathway. The cell and animal models of KOA were established and intervened with Cangxi Tongbi Capsules, si-circRNA＿0008365, si-NC, and Cangxi Tongbi Capsules combined with si-circRNA＿0008365. Flow cytometry and transmission electron microscopy were employed to determine the level of apoptosis and observe autophagosomes, respectively. Western blot was employed to reveal the changes in the protein levels of microtubule-associated protein light chain 3(LC3)Ⅱ/Ⅰ, Beclin-1, selective autophagy junction protein p62/sequestosome 1, collagen Ⅱ, a disintegrin and metalloproteinase with thrombospondin motifs 5(ADAMTS-5), and p38 MAPK. The mRNA levels of circRNA＿0008365, miR-1271, collagen Ⅱ, and ADAMTS-5 were determined by qRT-PCR. Hematoxylin-eosin staining was employed to reveal the pathological changes of the cartilage tissue of the knee, and enzyme-linked immunosorbent assay to measure the levels of interleukin-1β(IL-1β) and tumor necrosis factor-alpha(TNF-α). The chondrocytes treated with IL-1β showed down-regulated expression of circRNA＿0008365, up-regulated expression of miR-1271 and p38 MAPK, lowered autophagy level, increased apoptosis rate, and accelerated catabolism of extracellular matrix. The intervention with Cangxi Tongbi Capsules up-regulated the expression of circRNA＿0008365, down-regulated the expression of miR-1271 and p38 MAPK, increased the autophagy level, decreased the apoptosis rate, and weakened the catabolism of extracellular matrix. However, the effect of Cangxi Tongbi Capsules was suppressed after interfering with circRNA＿0008365. The in vivo experiments showed that Cangxi Tongbi Capsules dose-dependently inhibited the p38 MAPK pathway, enhanced chondrocyte autophagy, and mitigated articular cartilage damage and inflammatory response, thereby inhibiting the progression of KOA in rats. This study indicated that Cangxi Tongbi Capsules promoted chondrocyte autophagy by regulating the circRNA＿0008365/miR-1271/p38 MAPK pathway to inhibit the development of KOA.

Krüppel-like factor 15 deficiency exacerbates osteoarthritis through reduced expression of peroxisome proliferator-activated receptor gamma signaling in mice

Krüppel-like zinc finger transcription factors (KLFs) play diverse roles in mammalian cell differentiation and development. In this study, we investigated the function of KLF15 in the progression of osteoarthritis (OA). 0Destabilization of the medial meniscus (DMM) surgery was performed in 10-week-old male wild-type control (WT) mice and cartilage-specific KLF15 knockout (KO) mice. Histological analysis, immunohistochemistry, and terminal deoxynucleotidyl transferase deoxyuridine triphosphate nick-end labeling staining were performed. Morphological changes were measured using microcomputed tomography. Six mice from each group were analyzed (total number of mice analyzed: 60). In vitro, immunofluorescence, quantitative reverse transcription-polymerase chain reaction, and western blot analyses were performed. KLF15 KO DMM mice exhibited significant cartilage degradation compared to WT mice. According to the Osteoarthritis Research Society International cartilage OA-histopathology scoring system, the mean sum score in KLF15 KO mice was significantly higher than that in WT mice at 8 weeks after surgery. Immunohistochemistry results revealed KLF15 KO mice exhibited reducedperoxisome proliferator-activated receptor gamma (PPARγ) expression, increased pIKKα/β, a disintegrin-like and metalloproteinase with thrombospondin motifs (ADAMTS) 5, and Matrix metalloproteinases (MMP13) expression, and reduced Forkhead box O (FOXO1) and Light chain 3B (LC3B) expression. Inhibition of PPARγ phosphorylation accelerated the effects of interleukin (IL) 1β-treatment in both KLF15 KO and WT chondrocytes, and activation of PPARγ expression canceled the IL1β-induced catabolic effects. Our results indicated that the OA phenotype of KLF15 KO DMM mice was influenced by reduced PPARγ expression, including enhanced pIKKα/β, ADAMTS5, and MMP13 expression, reduced autophagy, and increased apoptosis. KLF15 regulation may constitute a possible therapeutic strategy for the treating OA.

Role of Syndecan-4 in the Inhibition of Articular Cartilage Degeneration in Osteoarthritis.

Despite its widespread existence, there are relatively few drugs that can inhibit the progression of osteoarthritis (OA). Syndecan-4 (SDC4) is a transmembrane heparan sulfate proteoglycan that modulates cellular interactions with the extracellular matrix. Upregulated SDC4 expression in articular cartilage chondrocytes correlates with OA progression. In the present study, we treated osteoarthritic cartilage with SDC4 to elucidate its role in the disease's pathology. In this in vitro study, we used real-time polymerase chain reaction (PCR) to investigate the effects of SDC4 on anabolic and catabolic factors in cultured chondrocytes. In the in vivo study, we investigated the effect of intra-articular injection of SDC4 into the knee joints of an OA mouse model. In vitro, SDC4 upregulated the expression of tissue inhibitor of metalloproteinase (TIMP)-3 and downregulated the expression of matrix metalloproteinase (MMP)-13 and disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-5 in chondrocytes. Injection of SDC4 into the knee joints of OA model mice prevented articular cartilage degeneration 6 and 8 weeks postoperatively. Immunohistochemical analysis 8 weeks after SDC4 injection into the knee joint revealed decreased ADAMTS-5 expression and increased TIMP-3 expression. The results of this study suggest that the treatment of osteoarthritic articular cartilage with SDC4 inhibits cartilage degeneration.

Characteristic Inflammatory Biomarkers in an Equine Model of Persistent Synovitis Induced By the Intra-Articular Administration of Monoiodoacetic Acid.

Persistent synovitis damages the articular cartilage in horses. To evaluate the effectiveness of treatment for synovitis using a model induced by intra-articular administration of monoiodoacetic acid (MIA), it is necessary to identify inflammatory biomarkers characteristic of the MIA model. Synovitis was induced by administering MIA into the unilateral antebrachiocarpal joints of five horses, and saline was injected into the contralateral joints as a control on day 0. Clinical and ultrasonographic examinations and synovial fluid collection were performed on days 0, 1, 2, 7, 14, 21, 28, and 35. Leukocyte, lactate dehydrogenase (LDH), tumor necrosis factor-α (TNF-α), interleukin-1 receptor antagonist (IL-1Ra), interleukin-6 (IL-6), and transforming growth factor-β1 (TGF-β1) concentrations in the synovial fluid were measured. Synovium was obtained after euthanasia on day 42 and histologically examined before quantification of the gene expression of inflammatory biomarkers by real-time PCR. Acute inflammatory symptoms persisted for approximately 2 weeks before returning to control levels. However, some indicators of chronic inflammation remained elevated until day 35. On day 42, synovitis continued histologically, with osteoclasts. The expressions of matrix metalloproteinase 13 (MMP13), a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4), receptor activator of nuclear factor kappa-Β ligand (RANKL), and collagen type I α2 chain (Col1a2) were significantly higher in the MIA model than in the control. In the MIA model, representative inflammatory biomarkers in the chronic inflammatory stage were persistently expressed in both synovial fluid and tissue, suggesting that they may be useful for the assessment of the anti-inflammatory effect of drugs.

The Role of ADAMTS Proteoglycanases in Thoracic Aortic Disease.

Thoracic aortic aneurysm and dissection (TAAD) are complex disease states with high morbidity and mortality that pose significant challenges to early diagnosis. Patients with an aneurysm are asymptomatic and typically present to the emergency department only after the development of a dissection. The extracellular matrix (ECM) plays a crucial role in regulating the aortic structure and function. The histopathologic hallmark termed medial degeneration is characterised by smooth muscle cell (SMC) loss, the degradation of elastic and collagen fibres and proteoglycan (PG) accumulation. Covalently attached to the protein core of PGs are a number of glycosaminoglycan chains, negatively charged molecules that provide flexibility, compressibility, and viscoelasticity to the aorta. PG pooling in the media can produce discontinuities in the aortic wall leading to increased local stress. The accumulation of PGs is likely due to an imbalance between their synthesis by SMCs and decreased proteolysis by A Disintegrin-like and Metalloproteinase with Thrombospondin motifs (ADAMTS) proteoglycanases in the ECM. Mouse models of TAAD indicated that these proteases exert a crucial, albeit complex and not fully elucidated, role in this disease. This has led to a mounting interest in utilising ADAMTS proteoglycanases as biomarkers of TAAD. In this review, we discuss the role of ADAMTSs in thoracic aortic disease and their potential use in facilitating the clinical diagnosis of TAAD and disease progression.

ADAMTS5 promotes neovascularization via autophagic degradation of PEDF in proliferative diabetic retinopathy

Proliferative diabetic retinopathy (PDR) adversely affects visual function. Extracellular matrix proteins (ECM) contribute significantly to the development of PDR. A Disintegrin and Metalloproteinase with Thrombospondin motifs 5 (ADAMTS5) is a member of ECM proteins. ADAMTS5 participates in angiogenesis and inflammation in diverse diseases. However, the role of ADAMTS5 in PDR remains elusive. Multiplex beam array technology was used to analyze vitreous humor of PDR patients and normal people. ELISA and Western blot were used to detect the expression of ADAMTS5, PEDF and autophagy related factors. Immunofluorescence assay was used to mark the expression and localization of ADAMTS5 and PEDF. The neovascularization was detected by tube formation test. Our results revealed that ADAMTS5 expression was increased in the vitreous humor of PDR patients and oxygen-induced retinopathy (OIR) mice retinas. Inhibiting ADAMTS5 alleviated pathological angiogenesis and upregulated PEDF expression in the OIR mice. In addition, ADAMTS5 inhibited PEDF secretion in ARPE-19 cells in vitro studies, thereby inhibiting the migration of HMEC-1. Mechanically, ADAMTS5 promoted the autophagic degradation of PEDF. Collectively, inhibition of ADAMTS5 during OIR suppresses pathological angiogenesis. Our study provides a new approach for resolving pathological angiogenesis in PDR.

Neonatal Zika virus infection causes transient perineuronal net degradation.

Perineuronal nets (PNNs) form a specialized extracellular matrix that predominantly surrounds parvalbumin (PV)-expressing GABAergic inhibitory interneurons and help regulate neuronal activity. Their formation early in the postnatal period is regulated by neuronal signaling and glial activation raising concerns that part of the long-term effects ascribed to perinatal viral infections could be mediated by altered PNN formation. Previously, we developed a model of neonatal Zika virus (ZIKV) infection where mice have lifelong neurological sequelae that includes motor disfunction and reduced anxiety coupled with a persistent low-grade expression in proinflammatory markers despite resolving the acute infection. Here, we demonstrate that ZIKV infection to P1 neonatal mice results in a reduction of PNN formation during the acute disease with significant reduction in Wisteria floribunda agglutinin (WFA) staining at the peak of infection [15 days post infection (dpi)] that persisted after the symptoms resolved (30 dpi). At 60 dpi, when there is residual inflammation in the CNS, the number of WFA+ cells and the level of WFA staining as well as levels of aggrecan and brevican in the brains of convalescent mice were not different from those in uninfected controls, however, there was increased frequency of PNNs with an immature phenotype. Over time the impact of the perinatal infection became less evident and there were no clear differences in PNN morphology between the groups at 1 year post infection. Of note, the reduction in PNNs during acute ZIKV infection was not associated with decreased mRNA levels of aggrecan or brevican, but increased levels of degraded aggrecan and brevican indicating increased PNN degradation. These changes were associated with increased expression of matrix metalloproteinase 12 (MMP12) and MMP19, but not MMP9, a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) or ADAMTS5. Together our findings indicate that infection at the time of PNN development interferes with PNN formation, but the nets can reform once the infection and inflammation subside.

Synergetic enrichment of aggrecan in nucleus pulposus cells by scAAV6-shRNA-mediated knockdown of aggrecanase-1 and aggrecanase-2.

Degenerative disk disease (DDD) that aggravates structural deterioration of intervertebral disks (IVDs) can be accompanied by painful inflammation and immunopathological progressions. Current surgical or pharmacological therapies cannot repair the structure and function of IVDs. Nucleus pulposus (NP) cells are crucial for the preservation or restoration of IVDs by balancing the anabolic and catabolic factors affecting the extracellular matrix. Imbalanced anabolic and catabolic factors cause increased degradation of aggrecan. Aggrecanases A Disintegrin And Metalloproteinase with ThromboSpondin motifs (ADAMTS)4 and ADAMTS5 are the main degrading enzymes of aggrecan. Previously, we characterized adeno-associated virus (AAV6) as the most suitable serotype with marked NP cellular tropism and demonstrated that ADAMTS4 could be silenced by self-complementary adeno-associated virus grade 6 small helix ribonucleic acid (scAAV6-shRNA) in NP cells of degeneration grade III, which resulted in enrichment of aggrecan. Nonetheless, neither scAAV6-shRNA-mediated inhibition of ADAMTS5 nor joint inhibitions of ADAMTS4 and ADAMTS5 have been investigated, although both enzymes are regulated by analogous proinflammatory cytokines and have the same cleavage sites in aggrecan. Therefore, we attempted scAAV6-shRNA-mediated inhibitions of both enzymes in NP cells of degeneration grade IV to increase efficacies in treatments of DDD. The degeneration grade of IVDs in patients was determined by magnetic resonance imaging (MRI) before surgical operations. After isolation and culturing of NP cells, cells were transduced with scAAV6-shRNAs targeting ADAMTS4 or ADAMTS5. Transduced cells were analyzed by reverse transcription quantitative polymerase chain reaction (RT-qPCR), fluorescence microscopy, flow cytometry-assisted cell sorting (FACS), MTT assay (3-(4, 5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide assay), immunoblotting, and enzyme-linked immunosorbent assay (ELISA). Joint transduction of NP cells exhibited high transduction efficacies (98.1%), high transduction units (TU) (1381 TU/Cell), and no effect on cell viability or proliferation. Above all joint treatments resulted in effective knockdown of ADAMTS4 (92.8%) and ADAMTS5 (93.4%) along with additive enrichment of aggrecan (113.9%). Treatment effects were significant for more than 56 days after transduction (P < 0.001). In conclusion, scAAV6-shRNA-mediated combined molecular therapy could be very valuable for more effective, durable, and less immunogenic treatment approaches in DDD.

Identification of Andrographolide as a novel FABP4 inhibitor for osteoarthritis treatment

Background and PurposeFatty acid binding protein 4 (FABP4) has been identified as a contributor to cartilage degradation in osteoarthritis (OA) patients, and inhibiting FABP4 using small molecules has emerged as a promising approach for developing OA drugs. Our previous research showed that Andrographis paniculata, a medicinal plant, strongly inhibits FABP4 activity. This led us to hypothesize that Andrographis paniculata ingredients might have protective effects on OA cartilage through FABP4 inhibition. MethodsWe analyzed scRNA-seq data from joint tissue of OA patients (GSE152805; GSE145286) using Scanpy 1.9.1 and Single Cell Portal. We conducted docking analysis of FABP4 inhibitors using Autodock Vina v.1.0.2. We evaluated the anti-FABP4 activity using a fluorescence displacement assay and measured the fatty acid oxidation (FAO) activity using the FAOBlue assay. We used H2DCF-DA to measure reactive oxygen species (ROS) levels. We studied signaling pathways using bulk RNA sequencing and western blot analysis in human C28/I2 chondrocytes. We evaluated anti-OA activity in monosodium iodoacetate (MIA)-induced rats. ResultsWe identified Andrographolide (AP) as a novel FABP4 inhibitor. Bulk RNA-sequencing analysis revealed that FABP4 upregulated FAO and ROS in chondrocytes, which was inhibited by AP. ROS generation activated the NF-κB pathway, leading to overexpression of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4), which is a responsible factor for cartilage degradation in OA patients. AP inhibited FABP4, thereby reducing the overexpression of ADAMTS4 by inhibiting the NF-κB pathway. In MIA rats, AP treatment reduced the overexpression of ADAMTS4, repaired cartilage and subchondral bone, and promoted cartilage regeneration. ConclusionOur results indicate that the inhibition of FABP4 activity by AP explains the anti-OA properties of Andrographis paniculata by protecting against cartilage degradation in OA patients. Additionally, our findings suggest that AP may be a promising therapeutic agent for OA treatment due to its ability to alleviate cartilage damage and bone erosion.

Secoisolariciresinol diglucoside Ameliorates Osteoarthritis via Nuclear factor-erythroid 2-related factor-2/ nuclear factor kappa B Pathway: In vitro and in vivo experiments

Osteoarthritis (OA) is an age-related joint disease in which inflammation and extracellular matrix (ECM) degradation play a crucial role in the destruction of articular cartilage. Secoisolariciresinol diglucoside (SDG), the main lignan in wholegrain flaxseed, which has been reported to remarkably suppress inflammation and oxidative stress, may have potential therapeutic value in OA. In this study, the effect and mechanism of SDG against cartilage degeneration were verified in the destabilization of the medial meniscus (DMM) and collagen-induced (CIA) arthritis models and interleukin-1β (IL-1β)-stimulated osteoarthritis chondrocyte models. From our experiments, SDG treatment downregulated the expression of pro-inflammatory factors induced by IL-1β in vitro, including inducible nitric oxide synthase (INOS), cyclooxygenase-2 (COX2), tumor necrosis factor (TNF-α), and interleukin 6 (IL-6). Additionally, SDG promoted the expression of collagen II (COL2A1) and SRY-related high-mobility-group-box gene 9(SOX9), while suppressing the expression of a disintegrin and metalloproteinase with thrombospondin motifs 5(ADAMTS5) and matrix metalloproteinases 13(MMP13), which leads to catabolism. Consistently, in vivo, SDG has been identified to have chondroprotective effects in DMM-induced and collagen-induced arthritis models. Mechanistically, SDG exerted its anti-inflammation and anti-ECM degradation effects by activating the Nrf2/HO-1 pathway and inhibiting the nuclear factor kappa B (NF-κB) pathway. In conclusion, SDG ameliorates the progression of OA via the Nrf2/NF-κB pathway, which indicates that SDG may have therapeutic potential for OA.

HTRA1 from OVX rat osteoclasts causes detrimental effects on endplate chondrocytes through NF-κB

Endplate osteochondritis is considered one of the major causes of intervertebral disc degeneration (IVDD) and low back pain. Menopausal women have a higher rate of endplate cartilage degeneration than similarly aged men, but the related mechanisms are still unclear. Subchondral bone changes, mainly mediated by osteoblasts and osteoclasts, are considered an important reason for the degeneration of cartilage. This work explored the role of osteoclasts in endplate cartilage degeneration, as well as its underlying mechanisms. A rat ovariectomy (OVX) model was used to induce estrogen deficiency. Our experiments indicated that OVX significantly promoted osteoclastogenesis and anabolism and catabolism changes in endplate chondrocytes. OVX osteoclasts cause an imbalance between anabolism and catabolism in endplate chondrocytes, as shown by a decrease in anabolic markers such as Aggrecan and Collagen II, and an increase in catabolic markers such as a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS5) and matrix metalloproteinases (MMP13). Osteoclasts were also confirmed in this study to be able to secrete HtrA serine peptidase 1 (HTRA1), which resulted in increased catabolism in endplate chondrocytes through the NF-κB pathway under estrogen deficiency. This study demonstrated the involvement and mechanism of osteoclasts in the anabolism and catabolism changes of endplate cartilage under estrogen deficiency, and proposed a new strategy for the treatment of endplate osteochondritis and IVDD by targeting HTRA1.

ADAMTS Proteases: Importance in Animal Reproduction.

Many reproductive physiological processes, such as folliculogenesis, ovulation, implantation, and fertilization, require the synthesis, remodeling, and degradation of the extracellular matrix (ECM). The ADAMTS (A Disintegrin and Metalloproteinase with Thrombospondin Motifs) family genes code for key metalloproteinases in the remodeling process of different ECM. Several genes of this family encode for proteins with important functions in reproductive processes; in particular, ADAMTS1, 4, 5 and 9 are genes that are differentially expressed in cell types and the physiological stages of reproductive tissues. ADAMTS enzymes degrade proteoglycans in the ECM of the follicles so that the oocytes can be released and regulate follicle development during folliculogenesis, favoring the action of essential growth factors, such as FGF-2, FGF-7 and GDF-9. The transcriptional regulation of ADAMTS1 and 9 in preovulatory follicles occurs because of the gonadotropin surge in preovulatory follicles, via the progesterone/progesterone receptor complex. In addition, in the case of ADAMTS1, pathways involving protein kinase A (PKA), extracellular signal regulated protein kinase (ERK1/2) and the epidermal growth factor receptor (EGFR) might contribute to ECM regulation. Different Omic studies indicate the importance of genes of the ADAMTS family from a reproductive aspect. ADAMTS genes could serve as biomarkers for genetic improvement and contribute to enhance fertility and animal reproduction; however, more research related to these genes, the synthesis of proteins encoded by these genes, and regulation in farm animals is needed.

The Comprehensive Analysis of Specific Proteins as Novel Biomarkers Involved in the Diagnosis and Progression of Gastric Cancer.

Gastric cancer (GC) cases are predicted to rise by 2040 to approximately 1.8 million cases, while GC-caused deaths to 1.3 million yearly worldwide. To change this prognosis, there is a need to improve the diagnosis of GC patients because this deadly malignancy is usually detected at an advanced stage. Therefore, new biomarkers of early GC are sorely needed. In the present paper, we summarized and referred to a number of original pieces of research concerning the clinical significance of specific proteins as potential biomarkers for GC in comparison to well-established tumor markers for this malignancy. It has been proved that selected chemokines and their specific receptors, vascular endothelial growth factor (VEGF) and epidermal growth factor receptor (EGFR), specific proteins such as interleukin 6 (IL-6) and C-reactive protein (CRP), matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs), a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS), as well as DNA- and RNA-based biomarkers, and c-MET (tyrosine-protein kinase Met) play a role in the pathogenesis of GC. Based on the recent scientific literature, our review indicates that presented specific proteins are potential biomarkers in the diagnosis and progression of GC as well as might be used as prognostic factors of GC patients' survival.

CHANGES TO LEVELS OF CHONDROCYTE-ASSOCIATED ADAMTS-4 IN EARLY HUMAN OSTEOARTHRITIS

Early changes within articular cartilage during human idiopathic osteoarthritis are poorly understood. However alterations to chondrocyte morphology occur with the development of fine cytoplasmic processes and cell clusters, potentially playing a role in cartilage degeneration. The aggrecanase ADAMTS-4 (A disintegrin and metalloproteinase with thrombospondin motifs-4) has been implicated as an important factor in cartilage degradation, so we investigated the relationship between chondrocyte morphology and levels of ADAMTS-4 in both non-degenerate and mildly osteoarthritic human cartilage.Human femoral heads were obtained following consent from patients undergoing hip arthroplasty following femoral neck fracture. Cartilage explants of normal (grade 0; G0) and mildly osteoarthritic (grade 1; G1) cartilage were labelled with the cytoplasmic dye CMFDA (5-chloromethylfluorescein-diacetate). Explants were cryosectioned (30μm sections), and labelled for ADAMTS-4 by fluorescence immunohistochemistry. Sections were imaged with confocal microscopy, allowing the semi-quantitative analysis of ADAMTS-4 and 3D visualisation of in situ cell morphology.With cartilage degeneration from G0 to G1, there was a decrease in the proportion of chondrocytes with normal rounded morphology (P&lt;0.001) but an increase in the proportion of cells with processes (P&lt;0.01) and those in clusters (P&lt;0.001;[4(1653)]; femoral heads:cells). Although average levels of ADAMTS-4 for all cells was the same between G0 and G1 (P&gt;0.05), a change was evident in the distribution curves for cell-specific ADAMTS-4 labelling. Cell-by-cell analysis showed that ADAMTS-4 levels were higher in chondrocytes with cytoplasmic processes compared to normal cells (P=0.044) however cells in clusters had lower levels than normal cells (P=0.003;[3(436)]). Preliminary data suggested that ADAMTS-4 levels increased with larger chondrocyte clusters.These results suggest complex heterogeneous changes to levels of cell-associated ADAMTS-4 with early cartilage degeneration – increasing in cells with processes and initially decreasing in clusters. Increased levels of ADAMTS-4 are likely to produce focal areas of matrix weakness potentially leading to early cartilage degeneration.

IRISIN STIMULATES ANABOLISM AND MATRIX SYNTHESIS IN HUMAN NUCLEUS PULPOSUS CELLS IN VITRO: NEW INSIGHTS INTO A CROSS-TALK BETWEEN THE MUSCLE AND THE INTERVERTEBRAL DISC

This study aimed to investigate the effect of irisin on human nucleus pulposus cells (hNPCs) in vitro. Our hypothesis was that irisin would improve hNPC metabolism and proliferation.hNPCs were isolated from intervertebral discs and cultured in alginate beads. hNPCs were exposed to phosphate-buffered saline (PBS) or recombinant irisin (r-irisin) at 5, 10 and 25 ng/mL (n=4). Each experiment was performed in triplicate. Cell proliferation was assessed with trypan blue staining-automated cell counting and PicoGreen assay. Glycosaminoglycan (GAG) content was measured using the DMMB assay. Metabolic activity was assessed with the MTT assay and the Griess Reagent System. Gene expression of collagen type II (COL2), matrix metalloproteinase (MMP)-13, tissue inhibitor of matrix metalloproteinase (TIMP)-1 and −3, aggrecan, interleukin (IL)-1β, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-5 was measured by RT-PCR. MTT assay and ADAMTS-5, COL2, TIMP-1 and IL-1β gene expression were evaluated following incubation with 5, 10 and 25 ng/mL r-irisin for 24 hours and subsequent culture with 10 ng/ml IL-1β and vice versa (incubation for 24 hours with IL-1β and subsequent culture with r-irisin).Irisin increased hNPC proliferation (p&lt;0.001), metabolic activity (p&lt;0.05), GAG content (p&lt;0.01), as well as COL2 (p&lt;0.01), aggrecan (p&lt;0.05), TIMP-1 and −3 (p&lt;0.01) gene expression, while decreasing MMP-13 (p&lt;0.05) and IL-1β (p&lt;0.001) mRNA levels. r-irisin pretreatment of hNPCs cultured in pro-inflammatory conditions resulted in a rescue of metabolic activity (p&lt;0.001) and a decrease of IL-1β (p&lt;0.05) levels. Similarly, incubation of hNPCs with IL-1β and subsequent exposure to r-irisin increased hNPC metabolic activity (p&lt;0.001), COL2 gene expression (p&lt;0.05) and decreased IL-1β (p&lt;0.05) and ADAMTS-5 levels (p&lt;0.01).Irisin stimulates hNPC proliferation, metabolic activity, and anabolism by reducing IL-1β and catabolic enzyme expression while promoting matrix synthesis.

Load-induced blood marker kinetics in patients with medial knee compartment osteoarthritis are associated with accumulated load and patient reported outcome measures

Background: This study aimed to quantify the mechanoresponse of 10 blood marker candidates for joint metabolism to a walking stress test in patients with knee osteoarthritis and to determine the association among marker kinetics and with accumulated load and patient reported outcomes. Methods: 24 patients with knee osteoarthritis completed questionnaires, and a walking stress test with six blood serum samples and gait analysis. Concentrations of cartilage oligomeric matrix protein (COMP), matrix metalloproteinases (MMP)-1, -3, and -9, epitope resulting from cleavage of type II collagen by collagenases (C2C), type II procollagen (CPII), interleukin (IL)-6, proteoglycan (PRG)-4, A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4, and resistin were determined by enzyme-linked immunosorbent assays, Joint load (moments and compartmental forces) was estimated using musculoskeletal modeling using gait analysis data. Results: COMP and MMP-3 showed an immediate increase after the walking stress followed by a decrease. MMP-9 and resistin showed a delayed decrease below pre-stress levels. ∆COMP correlated with ∆MMP-3 for most time points. ∆MMP-9 correlated with ∆resistin for most time points. The load-induced increase in blood marker levels correlated among blood markers and time points. C2C and resistin correlated positively and C2C/CPII and MMP2 correlated negatively with load during gait. Immediate relative ∆CPII and ∆MMP1 and delayed relative ∆COMP, ∆IL6, ∆C2C, ∆CPII, ∆MMP1 and ∆MMP3 correlated with the load accumulated during the walking stress. Baseline C2C levels correlated with Knee Osteoarthritis Outcome Score (KOOS) subscales and load-induced changes in MMP-3 with KOOS and Short Form 36 quality of life subscores (P&lt;0.05). Conclusions: The distinct and differentiated physiological response to the walking stress depends on accumulated load and appears relevant for patient reported osteoarthritis outcome and quality of life and warrants further investigation in the context of disease progression. ClinicalTrials.gov registration: NCT02622204

Changes in the Extracellular Matrix in Endometrial and Cervical Cancer: A Systematic Review

Endometrial and cervical cancers are the two most common gynaecological malignancies and among the leading causes of death worldwide. The extracellular matrix (ECM) is an important component of the cellular microenvironment and plays an important role in developing and regulating normal tissues and homeostasis. The pathological dynamics of the ECM contribute to several different processes such as endometriosis, infertility, cancer, and metastasis. Identifying changes in components of ECM is crucial for understanding the mechanisms of cancer development and its progression. We performed a systematic analysis of publications on the topic of changes in the extracellular matrix in cervical and endometrial cancer. The findings of this systematic review show that matrix metalloproteinases (MMP) play an important role impacting tumour growth in both types of cancer. MMPs degrade various specific substrates (collagen, elastin, fibronectin, aggrecan, fibulin, laminin, tenascin, vitronectin, versican, nidogen) and play a crucial role in the basal membrane degradation and ECM components. Similar types of MMPs were found to be increased in both cancers, namely, MMP-1, MMP-2, MMP-9, and MMP-11. Elevated concentrations of MMP-2 and MMP-9 were correlated with the FIGO stage and are associated with poor prognosis in endometrial cancer, whereas in cervical cancer, elevated concentrations of MMP-9 have been associated with a better outcome. Elevated ADAMTS levels were found in cervical cancer tissues. Elevated disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) levels were also found in endometrial cancer, but their role is still unclear. Following these findings, this review reports on tissue inhibitors of ECM enzymes, MMPs, and ADAMTS. The present review demonstrates changes in the extracellular matrix in cervical and endometrial cancers and compared their effect on cancer development, progression, and patient prognosis.