Lysyl Oxidase Expression Levels Research Articles

LOX-mediated ECM mechanical stress induces Piezo1 activation in hypoxic-ischemic brain damage and identification of novel inhibitor of LOX

Hypoxic-ischemic encephalopathy (HIE) poses a significant challenge in neonatal medicine, often resulting in profound and lasting neurological deficits. Current therapeutic strategies for hypoxia-ischemia brain damage (HIBD) remain limited. Ferroptosis has been reported to play a crucial role in HIE and serves as a potential therapeutic target. However, the mechanisms underlying ferroptosis in HIBD remain largely unclear. In this study, we found that elevated lysyl oxidase (LOX) expression correlates closely with the severity of HIE, suggesting LOX as a potential biomarker for HIE. LOX expression levels and enzymatic activity were significantly increased in HI-induced neuronal models both in vitro and in vivo. Notably, we discovered that HI-induced brain tissue injury results in increased stiffness and observed a selective upregulation of the mechanosensitive ion channel Piezo1 in both brain tissue of HIBD and primary cortex neurons. Mechanistically, LOX increases its catalytic substrates, the Collagen I/III components, promoting extracellular matrix (ECM) remodeling and possibly mediating ECM cross-linking, which leads to increased stiffness at the site of injury and subsequent activation of the Piezo1 channel. Piezo1 senses these stiffness stimuli and then induces neuronal ferroptosis in a GPX4-dependent manner. Pharmacological inhibition of LOX or Piezo1 ameliorated brain neuronal ferroptosis and improved learning and memory impairments. Furthermore, we identified traumatic acid (TA) as a novel LOX inhibitor that effectively suppresses LOX enzymatic activity, mitigating neuronal ferroptosis and promoting synaptic plasticity. In conclusion, our findings elucidate a critical role for LOX-mediated ECM mechanical stress-induced Piezo1 activation in regulating ferroptotic cell death in HIBD. This mechanistic insight provides a basis for developing targeted therapies aimed at ameliorating neurological outcomes in neonates affected by HIBD.

Downregulation of LOX Overexpression Promotes Retinal Ganglion Cells Survival in an Acute Ocular Hypertension Model

Purpose To investigate the effect of reducing Lysyl oxidase (LOX) overexpression on retinal ganglion cells (RGCs) apoptosis in an acute ocular hypertension (AOH) rat model. Methods AOH rat model was performed by anterior chamber perfusion and either received an intravitreal injection with β-aminopropionitrile (BAPN) or normal saline. After 2wk, Quantification of survival RGCs in the retina was performed using Retrograde FluoroGold labeling. The mRNA expression levels of LOX, LOXL1-4, collagen 1a1 (Col1a1), collagen 3a1 (Col3a1), collagen4a1 (Col4a1), elastin (Eln), fibronectin1 (Fbn1), fibronectin4 (Fbn4) were determined by RT-qPCR. LOX expression was determined by Western blot (WB) analysis and immunohistochemistry. The RNA expression of LOX, Eln and Col1a1 in RGCs retrograde-labeled with 1,1’-dioctadecyl-3,3,3’,3’ tetra-methylindocarbocyanine perchlorate(DiI)that selected through FACS sorting were determined by RT-qPCR analysis. Changes of the retinal function were detected by Electroretinogram (ERG) analysis. Results Results showed that significant LOX overexpression and loss of RGCs related to IOP exposure in AOH retinas. PCR analysis indicated significant increased mRNA level of Col1a1, Col3al and Eln in AOH retinas. Significant increase mRNA expression of LOX, Col1a1 and Eln in the RGCs were observed in AOH group compared with CON group. AOH rats injected with BAPN showed a significant decrease in LOX expression, reduced the loss of RGCs and retinal function damage. Conclusions The results demonstrated that changes of LOX and specific ECM components in retina were correlated with AOH. Findings from this study indicated that preventing LOX over-expression may be protective against RGCs loss and retinal function damage in AOH animal model.

Hub gene target of glioblastoma: LOX, SERPINH1 and TGFBI.

Glioblastoma (GBM) is a malignant tumor. The long-term prognosis of the patients is poor. Therefore, it is of important clinical value to further explore the pathogenesis and look for molecular markers for early diagnosis and targeted treatment. Two expression profiling datasets [GSE50161 (GPL570 platform), GSE116520 (GPL10558 platform)] were respectively downloaded from the gene expression omnibus database. Volcano diagrams show the Differently expressed genes (DEGs) of GSE50161 and GSE116520. A Venn diagram revealed 467 common DEGs between the 2 datasets. Lysyl oxidase (LOX), serpin family H member 1 (SERPINH1) and transforming growth factor beta induced (TGFBI) were negatively correlated with the overall survival rate in patients with GBM. The hub genes are high in GBM tumor tissues. The relative expression levels of LOX, SERPINH1 and TGFBI were significantly higher in GBM samples, compared with the normal brain tissues groups. Bioinformatics technology could be a useful tool to predict progression of GBM and to explore the mechanism of GBM.LOX, SERPINH1 and TGFBI may be involved in the mechanism of the occurrence and development of GBM, and may be used as molecular targets for early diagnosis and specific treatment. DEGs identified using GEO2R. Functional annotation of DEGs using Kyoto Encyclopedia of Genes and Genomes and gene body pathway enrichment analysis. Construction of a protein-protein interaction network. The pathway and process enrichment analysis of the hub genes were performed by Metascape. Survival analysis was performed in gene expression profiling interactive analysis. Real-time fluorescent quantitative polymerase chain reaction assay was performed to verify. The animal model was established for western blot test analysis.

Lysyl oxidase promotes anaplastic thyroid carcinoma cell proliferation and metastasis mediated via BMP1.

Anaplastic thyroid carcinoma (ATC) is an extremely aggressive solid tumor with no effective treatment at present. Because of the rapid growth and aggressiveness, nearly all patients die within six months after developing ATC. Hence, more research regarding novel therapeutic targets for ATC is urgently needed. Single-cell RNA sequencing data and microarray data of ATC were retrieved from the Gene Expression Omnibus (GEO) database. Cell clustering was performed using the Seurat package. Then, differential expression and functional enrichment analyses were performed. Gene set enrichment analysis (GSEA) was further used to investigate the functional enrichment of lysyl oxidase (LOX) and bone morphogenetic protein-1 (BMP1). The expression levels of LOX and BMP1 were measured using quantitative real-time PCR and Western blot. LOX and BMP1 were knocked down using si-RNAs. Cell proliferation was evaluated by the CCK-8 and clone formation assays. Cell migration and invasion were assessed by the wound healing assay and Transwell assay, respectively. LOX was upregulated at the single-cell level, as well as in ATC tissues and cell lines. LOX knockdown significantly inhibited ATC cell proliferation. Furthermore, the migration and invasion of ATC cells were remarkably inhibited after LOX inhibition. In addition, BMP1 regulated LOX expression in 8505C cells, while BMP1 overexpression restored the LOX activity blocked by the LOX inhibitor BAPN. BMP1 could also induce the cell proliferation and metastasis of ATC. LOX/BMP1 mediates the malignant progression of ATC, highlighting the potential application of LOX/BMP1 in the treatment of ATC. This study provides new insights for efficient therapeutic agents based on the LOX/BMP1 axis.

Serum Lysyl Oxidase Is a Potential Diagnostic Biomarker for Kidney Fibrosis

Background: Kidney fibrosis is the ultimate consequence of advanced stages of chronic kidney disease (CKD); however, there are currently no reliable biomarkers or noninvasive diagnostic tests available for the detection of kidney fibrosis. Lysyl oxidase (LOX) promotes collagen cross-linking, and serum LOX levels have been shown to be elevated in patients with fibrosis of the heart, lungs, and liver. However, serum LOX levels have not been reported in patients with kidney fibrosis. We explored whether serum LOX levels are associated with kidney fibrosis. Method: Overall, 202 patients with kidney disease underwent renal biopsy, scoring of kidney fibrosis, and determination of the area of kidney fibrosis. LOX levels were measured in serum and in kidney tissues. We analyzed the association of circulating LOX and tissue LOX levels with the scores and areas of kidney fibrosis. LOX expression was also investigated with in vitro and in vivo kidney fibrosis models. Results: Serum LOX levels were higher in patients with kidney fibrosis than in those without kidney fibrosis (p < 0.001) and higher in patients with moderate-severe kidney fibrosis than in patients with mild kidney fibrosis (p < 0.001). Both serum LOX and renal tissue LOX levels correlated with the area of kidney fibrosis (r = 0.748, p < 0.001; r = 0.899, p < 0.001, respectively). Receiver operating characteristic curve analysis of serum LOX levels showed an area under the curve of 0.80 (95% CI: 0.74–0.86). The optimal serum LOX level cutoff point was 253.34 pg/mL for the prediction of kidney fibrosis and 306.56 pg/mL for the prediction of moderate-severe kidney fibrosis. LOX expression levels were significantly upregulated (2.3–2.6 and 6-fold, respectively) in in vitro and in vivo interstitial fibrosis models. Conclusions: Both serum LOX and tissue LOX levels correlated with the presence and degree of kidney fibrosis in patients with CKD. These results suggest that serum LOX levels could potentially serve as a noninvasive diagnostic biomarker for kidney fibrosis and may further potentially serve as a stratified biomarker for the identification of mild and moderate-severe kidney fibrosis.

Co-administration of resveratrol and beta-aminopropionitrile attenuates liver fibrosis development via targeting lysyl oxidase in CCl4-induced liver fibrosis in rats

Objectives: In the current study, we aimed to investigate the effect of administration of resveratrol (RES) and beta-aminopropionitrile (BAPN) separately and together on the liver fibrosis progression via regulation of the gene expression and protein level of lysyl oxidase (LOX).Materials and methods: The six-week old Wistar rats received carbon tetrachloride (CCl4) intraperitoneally and RES and BAPN were administrated orally for eight weeks. The hepatoprotective effects of RES, BAPN, and combination treatment were evaluated. Then the hepatic protein and gene expression levels of LOX were measured.Results: Both RES and BAPN showed the antifibrotic effect through the reduction of collagen fiber bundles, hepatic hydroxyproline content, and protein level of LOX. The antifibrotic effect increased when RES and BAPN up-taken together.Conclusion: The co-administration of RES and BAPN can be considered as a promising therapeutic approach via targeting LOX.

Analysis of lysyl oxidase as a marker for diagnosis of canine mammary tumors.

Lysyl oxidase (LOX) is an extracellular metalloenzyme which mediates crosslinking of collagen and elastin. It has been reported to play a pivotal role in cancer metastasis especially in women suffering from breast cancer. The present study is the first to evaluate the gene expression levels of LOX by Real time-polymerase chain reaction (Real time-PCR) in dogs with mammary tumor besides molecular cloning and expression of canine lysyl oxidase gene (lox). Real time-PCR studies showed a significant upregulation (threefold higher) of lox in mammary tumor cases as compared to healthy dogs indicating its possible diagnostic and prognostic role in canine mammary tumors (CMTs). Cloning and sequencing of lox gene revealed 1230bp CDS which is mostly conserved in C-terminal region. Sequence analysis of canine lox showed that it shares 99% homology with the predicted sequence available on NCBI and had greatest identity with the lox gene from cat. Protein structure predicted with homology modelling was validated by Ramachandran plot analysis which revealed most (approximately 95%) of the amino acids in favoured region. Additionally, recombinant lysyl oxidase expressed as His-tagged fusion protein in prokaryotic expression vector (pPROExHTa) was used in an ELISA for detection of circulating protein LOX in serum of CMT subjects. Receiver operating characteristics analysis of the ELISA revealed high sensitivity (90%) and specificity (85%) with histopathology as reference standard. Taken together, we propose LOX as a diagnostic biomarker and a putative prognostic candidate in CMT cases.

Lysyl oxidase and hypoxia-inducible factor 1α: biomarkers of gastric cancer.

BACKGROUNDGastric cancer (GC) is one of the main causes of cancer mortality worldwide. Recent studies on tumor microenvironments have shown that tumor metabolism exerts a vital role in cancer progression.AIMTo investigate whether lysyl oxidase (LOX) and hypoxia-inducible factor 1α (HIF1α) are prognostic and predictive biomarkers in GC.METHODSA total of 80 tissue and blood samples were collected from 140 patients admitted to our hospital between August 2008 and March 2012. Immunohistochemical staining was performed to measure the expression of LOX and HIF1α in tumor and adjacent tissues collected from patients with GC. Real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis was used to detect the mRNA expression levels of LOX and HIF1α in patients with GC. In addition, single-factor analysis was applied to analyze the relationship between LOX, HIF1α and prognosis of GC.RESULTSImmunohistochemical staining suggested that the expression levels of LOX and HIF1α increased in tumor tissues from patients with GC. QRT-PCR analysis indicated that mRNA expression of LOX and HIF1α was also upregulated in tumor tissues, which was in accordance with the above results. We also detected expression of these two genes in blood samples. The expression level of LOX and HIF1α was higher in patients with GC than in healthy controls. Additional analysis showed that the expression level of LOX and HIF1α was related to the clinicopathological characteristics of GC. Expression of LOX and HIF1α increased with the number of lymph node metastases, deeper infiltration depth and later tumor–node–metastasis stages. Single-factor analysis showed that high expression of LOX and HIF1α led to poor prognosis of patients with GC.CONCLUSIONLOX and HIF1α can be used as prognostic and predictive biomarkers for GC.

Hypercapnia downregulates hypoxia-induced lysyl oxidase expression in pulmonary artery smooth muscle cells via inhibiting transforming growth factor β1 signalling.

Hypoxic pulmonary arterial hypertension is characterized by elevated pulmonary vascular resistance and remodelling. Transforming growth factor-β1 (TGF-β1 ) is the master regulator in cellular response to hypoxia which can directly target lysyl oxidase (LOX). This study aimed to determine whether hypercapnia attenuates hypoxic pulmonary hypertension via regulating TGF-β1 and LOX signalling. We found that exposure to hypercapnia ameliorated the increase in mean pulmonary artery pressure (mPAP) and ratio of right ventricle to left ventricle plus septum (RV/(LV+S)) induced by hypoxia but had no effect on mPAP and RV/(LV+S) in normoxia-exposed control. In addition, exposure to hypoxia upregulated the mRNA and protein levels of LOX and TGF-β1 in rat PASMCs both in vivo and in vitro, but these effects were abrogated by concurrent exposure to hypercapnia. The downregulation of LOX in rat PASMCs induced by hypercapnia was reversed by the administration with TGF-β1 , while TGF-β1 knockdown repressed the upregulation of LOX in hypoxia-exposed rat PASMCs. In conclusion, hypoxia upregulates LOX and TGF-β1 expression in PASMCs and contributes to pulmonary hypertension. Hypercapnia downregulates hypoxia-induced LOX expression and alleviates hypoxia-associated pulmonary hypertension via inhibiting TGF-β1 signalling. SIGNIFICANCE OF THE STUDY: Hypoxia-induced upregulation of TGF-β1 , PDGF, and HIF-1α plays a pivotal role in PAH, but molecular mechanism of how hypoxia regulates LOX expression is not clear. In the present study, we showed that mRNA and protein expression levels of LOX were substantially increased when TGF-β1 was induced by hypoxia, and the effects were reversed by TGF-β1 knockdown. Our study indicates that TGF-β1 is implicated in the regulation of LOX.

LOX inhibition downregulates MMP-2 and MMP-9 in gastric cancer tissues and cells.

Objective: The objective of this study was to analyze the effects of lysyl oxidase (LOX) on the expression and enzyme activity of the matrix metalloproteinases 2 (MMP-2) and 9 (MMP-9) and to study its preliminary effect mechanisms.Methods: We collected fresh cancer specimens from 49 gastric cancer patients who underwent surgery. Immunohistochemistry was used to quantitate the protein expression levels of LOX and MMP-9 in gastric cancer tissues and to analyze their correlation. Also, six-week old nude mice were divided into a control group and a LOX inhibition group. SGC-7901 gastric cancer cells were inoculated subcutaneously into the backs of the two groups of these mice to construct a gastric cancer-bearing nude mouse model. In the LOX inhibition group, β-aminopropionitrile (BAPN) was used to inhibit LOX. Western blotting was used to quantitate the relative expression levels of MMP-2 and MMP-9 in mouse tumor tissues, and gelatin zymography was used to quantitate their enzyme activity levels. In addition, BGC-823 gastric cancer cells were cultured, then 0.1 mM, 0.2 mM, and 0.3 mM BAPN and 2.5 nM, 5 nM, and 10 nM LOX were added to treat BGC-823 cells. ELISA and gelatin zymography were used to quantitate the protein concentrations and changes in enzyme activity of MMP-2 and MMP-9 in the culture supernatant. Western blotting was used to quantitate the relative expression levels of platelet derived growth factor receptor (PDGFR) in the BGC-823 gastric cancer cells after LOX inhibition and exogenous LOX addition.Results: In the tissues from the gastric cancer patients, the relative expression levels of LOX and MMP-9 were positively correlated (r = 0.326, P < 0.05). Compared with the control group, the tumor tissues from mice in the LOX inhibition group had reduced relative expression levels and enzyme activities of MMP-2 and MMP-9 (P < 0.05). After LOX were inhibited with different concentrations of BAPN in BGC-823 gastric cancer cells, the protein concentrations and enzyme activity levels of MMP-2 and MMP-9 in the culture supernatants were decreased (P < 0.05). In addition, the relative expression level of PDGFR in gastric cancer was decreased when BAPN concentrations increased, showing a negative dose-dependent manner (rPDGFR-α = -0.964, rPDGFR-β = -0.988, P < 0.05). After exogenous LOX treating BGC-823 cells, the concentrations and enzyme activity levels of MMP-2 and MMP-9 in the cell supernatant were increased (P < 0.05). Further, the relative expression of PDGFR in gastric cancer cells was increased with the increase of exogenous LOX, showing a positive dose-dependent manner (rPDGFR-α=0.952, rPDGFR-β=0.953, P<0.05).Conclusions: LOX inhibition can inhibit the expression and enzyme activity of MMP-2 and MMP-9 in gastric cancer tissues and cells, and the probable mechanism is through its effects on the PDGF-PDGFR signaling pathway.

Inhibition of lysyl oxidase expression by dextran sulfate affects invasion and migration of gastric cancer cells.

In the present study, the effect of dextran sulfate (DS) on the metastasis and invasion of human gastric cancer cells and its key underlying mechanism were investigated. The levels of hypoxia-inducible factor 1α (HIF-1α), transforming growth factor β (TGF-β) and lysyl oxidase (LOX) expression were evaluated in human gastric cancer and peritumoral tissues by immunohistochemical analysis. Cell proliferation and apoptosis were also examined using the Cell Counting Kit-8 assay and flow cytometry. The effect of DS on the invasion and migration of BGC-823 cells was assessed using a Transwell assay. BGC-823 cells were divided into the control (phosphate-buffered saline-treated) and experimental (DS-treated) groups, and cultured for different times under hypoxic conditions. Subsequently, LOX and TGF-β expression levels in the cells were measured by immunocytochemistry, immunofluorescence, reverse transcription-quantitative polymerase chain reaction and western blot analysis. HIF-1α, TGF-β and LOX expression levels were significantly higher in human gastric cancer tissues as compared with that in adjacent tissues. DS influenced cell proliferation and apoptosis in a dose-dependent manner. Furthermore, DS reduced the number of invaded and migrated cells. Under hypoxic conditions, DS reduced HIF-1α, TGF-β and LOX expression levels in BGC-823 cells. After 12 h, the effect of combination of DS and β-aminopropionitrile (BAPN) on LOX and TGF-β protein levels was more significant compared with that of DS or BAPN alone. Therefore, DS may inhibit the invasion and migration of human gastric cancer cells under hypoxic conditions by influencing LOX.

Mechanical stretching promotes the differentiation of BMSC into pelvic floor ligament fibroblasts

// Bing Zhao 1, * , Junmin Wang 2, 1, * , Chenchen Ren 1 , Mengcai Hu 1 , Huiyan Wu 1 , Lulu Chen 1 , Xiaojun Liu 3 , Luwen Wang 1 , Feng Xu 1 , Xueqin Zheng 1 , Juan Chen 1 and Shihong Cui 1 1 Third Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan Province 450052, China 2 Department of Human Anatomy, Basic Medical College of Zhengzhou University, Zhengzhou, Henan Province 450052, China 3 Henan Medical Equipment Inspection Institute, Zhengzhou, Henan Province 450052, China * These authors have contributed equally to this work Correspondence to: Shihong Cui, email: shihongcui@126.com Keywords: mechanical stretch; BMSC; KDM4B; LOX; elastin Received: December 28, 2016 Accepted: July 24, 2017 Published: February 07, 2018 ABSTRACT Pelvic floor dysfunction (PFD) is a prevalent and debilitating condition in aging women that is associated with weakened pelvic connective tissue. Fibroblasts are the main component of pelvic connective tissue and play a vital role in the maintenance of the pelvic organ. Mechanical stretching is a known modulator leading to the proliferation and differentiation of bone marrow mesenchymal stem cells (BMSCs). In this paper, we measured the lysyl oxidase (LOX), elastin and KDM4B expression in cultured BMSCs under different cyclic mechanical stretch (CMS) conditions via Western blot assays. Our findings indicated that the expression of these proteins was significantly higher in the CMS group than in the normal control group, and 10% CMS at a 0.5 Hz frequency had the best stimulatory effect. We further evaluated the influence of KDM4B on LOX and elastin expression via knockdown and overexpression of KDM4B in BMSCs. Chromatin immunoprecipitation (ChIP)-PCR analysis was performed to determine how H3K9me3 and KDM4B affect the regulatory regions of LOX and elastin in BMSCs subjected to 10% CMS combined with KDM4B knockdown. Finally, we verified the expression of KDM4B under CMS in vivo by subcutaneously transplanting KDM4B- overexpressing BMSCs into mice via immunofluorescence assays. The results showed that activating KDM4B by mechanical stretching increased the protein and gene expression levels of LOX and elastin and induced more LOX-positive cells in the BMSCs. These data suggest that CMS contributes to the differentiation of mesenchymal stem cells (MSCs) into fibroblasts, which indicates its potential use as a cell-based therapy for PFD.

Lysyl oxidase is involved in synovial hyperplasia and angiogenesis in rats with collagen‑induced arthritis.

Lysyl oxidase (LOX) serves an important role in remodeling the extracellular matrix and angiogenesis in various types of cancer; however, whether LOX is involved in the pathogenesis of rheumatoid arthritis remains unknown. In order to investigate this in the present study, β-aminopropionitrile, an inhibitor of LOX, was injected intraperitoneally into rats with type II collagen-induced arthritis (CIA). Subsequently, synovial hyperplasia was examined by hematoxyl in and eosin staining, and the microvascular density (MVD) and expression levels of LOX, matrix metalloproteinase (MMP)-2 and MMP-9 in the synovial membrane and fluid were determined by immunohistochemistry and ELISA, respectively. The enzyme activity of LOX was evaluated by the Amplex Red Hydrogen Peroxide method. The results demonstrated an increased amount of rough synovial membranes, higher MVD in these membranes and more synovial cell layers in CIA rats compared with in the control rats. In addition, higher enzymatic activity of LOX and higher expression levels of MMP-2 and MMP-9 were revealed in CIA rats compared with in the control rats. Notably, β-aminopropionitrile inhibited paw swelling and the decreased the arthritis index, the MVD in the synovial membranes and the expression levels of MMP-2 and MMP-9. Furthermore, the expression level of LOX in the synovial membranes was positively associated with the MVD and the expression levels of MMP-2 and MMP-9, suggesting that LOX promotes synovial hyperplasia and angiogenesis and that LOX may be a potential therapeutic target for rheumatoid arthritis.

Low extracellular lysyl oxidase expression is associated with poor prognosis in patients with prostate cancer.

Remodeling of the extracellular matrix (ECM), which is induced by lysyl oxidase (LOX), has been demonstrated to accompany tumor progression; however, the association between LOX expression levels and the malignant behavior of prostate cancer (Pca) remains unclear. The present study aimed to analyze the tumor-associated expression profile of LOX in patients with Pca and to evaluate its potential prognostic value. In the form of a retrospective study, the expression patterns of LOX and collagen I were analyzed in patients with benign prostate hyperplasia and Pca by immunohistochemical examination. The results demonstrated that, with the initiation and progression of Pca, the expression levels of LOX and collagen I were closely associated with Gleason score and tumor stage. In addition, although LOX was expressed in cancer and non-cancer tissues, the differential expression pattern observed in the ECM of Pca cells may indicate that LOX is an important molecule that affects the progression of this disease. Therefore, LOX expression level in the ECM may function as an independent predictor of Pca.

Mechano growth factor-E regulates apoptosis and inflammatory responses in fibroblast-like synoviocytes of knee osteoarthritis.

This study investigated whether mechano growth factor-E (MGF-E) peptide can regulate apoptosis and inflammation responses in fibroblast-like synoviocytes of osteoarthritis (OA). A (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt) (MTS) assay was performed to evaluate cytotoxic effects of exogenous MGF-E peptide on OA fibroblast-like synoviocytes (OA-FLS). Quantitative real-time polymerase chain reaction (qRT-PCR) was used to check messenger RNA (mRNA) expression levels of lysyl oxidase (LOX) family members (LOX) after OA-FLS treatment using MGF-E peptide. A 5-ethynyl-2'-deoxyuridine (EdU) incorporation assay was performed to identify the influence of MGF-E peptide on proliferation OA-FLS proliferation. Western blot was used to detect biomarkers of endoplasmic reticulum (ER) stress and inflammatory cytokines. Exogenous MGF-E peptide has no obvious cytotoxic effects on OA-FLS and promotes LOX expression in OA-FLS, induce apoptosis and ER stress and down-regulate protein levels of tumour necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β). Our results suggest that MGF-E peptide possesses potential anti-inflammatory effects, induces cell apoptosis and facilitates repair of OA-FLS. Therefore, MGF-E peptide may have therapeutic potential in patients with OA.

Mechanism of serotonin-promoted synthesis of osteoblast type I collagen

To explore the mechanism of serotonin-promoted osteoblast differentiation. Expression levels of collagen I and lysyl oxidase (LOX) in osteoblast were measured by RT-PCR after treated by (50, 100, 200 and 400 ng/L) serotonin. LOX siRNA effect was measured by Western blot, and protein levels of collagen I were determined by ELISA after treated by serotonin. Expression levels of Smad2 and Smad3 in osteoblasts were also measured by RT-PCR after treated by serotonin.Moreover, expression levels of LOX were measured by RT-PCR after Smad3 was knockout. Serotonin promoted collagen I and LOX expression. The expression level of collagen I was significantly decreased by LOX siRNA. Furthermore, serotonin up-regulated the expression of Smad2 and Smad3 in osteoblasts, and the expression level of LOX was inhibited by Smad3 siRNA. Serotonin promoted collagen I expression by activating Smads signaling pathway and up-regulating the LOX expression.

LOX expression and functional analysis in astrocytomas and impact of IDH1 mutation.

Lysyl oxidase (LOX) is involved in vital biological processes such as cell motility, cell signaling and gene regulation. Deregulation of this protein can contribute to tumor formation and progression. Although it is known that LOX is involved in invasion, proliferation and tumor migration in other types of tumors, studies of LOX in astrocytomas of different grades are scarce. The purpose of our study was to characterize LOX, BMP1 and HIF1A expression by real-time PCR in astrocytomas with WHO grades I to IV compared to non-neoplastic brain tissue. IDH1 mutational status was determined by PCR and sequencing. LOX protein expression was also analyzed by immunohistochemistry. LOX functional analyses were performed using siRNA knockdown and the specific inhibitor BAPN in two glioblastoma cell lines. The expression levels of LOX, BMP1 and HIF1A were correlated and analyzed according to IDH1 mutation status and to the clinical end-point of overall survival of glioblastoma patients. The results demonstrate that increased expression and activity of LOX, BMP1 and HIF1A were positively correlated with the malignant grade of astrocytomas. LOX protein expression also increased according to the degree of malignancy, with localization in the cytoplasm and nucleus and staining observed in endothelial cells. Glioblastoma with a mutation in IDH1 expressed lower levels of LOX in the nucleus, and IDH1-mutated cases showed lower LOX expression levels when compared to wild-type IDH1 cases. LOX knockdown and inhibition by BAPN in U87MG and A172 cell lines affected migration, invasion and soft agar colony formation. Taken together, these results corroborate the role of LOX in the migration, invasion and angiogenesis of astrocytomas. Furthermore, LOX expression is influenced by IDH1 mutational status. This work provides new insights for researchers aiming to design targeted therapies to control astrocytomas.

Two functional loci in the promoter of EPAS1 gene involved in high-altitude adaptation of Tibetans.

EPAS1 involves in the hypoxic response and is suggested to be responsible for the genetic adaptation of high-altitude hypoxia in Tibetans. However, the detailed molecular mechanism remains unknown. In this study, a single nucleotide polymorphism rs56721780:G>C and an insertion/deletion (indel) polymorphism −742 indel in the promoter region showed divergence between Tibetans and non-Tibetan lowlanders. rs56721780:G>C regulated the transcription of EPAS1 by IKAROS family zinc finger 1 (IKZF1), which was identified as a new transcriptional repressor for EPAS1 gene. It demonstrated that the C allele of rs56721780:G>C decreased the binding of IKZF1, leading to the attenuated transcriptional repression of EPAS1 gene. The insertion at −742 indel provided a new binding site for Sp1 and was related to the activation of EPAS1 promoter. Further functional analysis revealed that lysyl oxidase (LOX) gene, which was reported to be responsible for extracellular matrix protein cross-linking of amnion previously, was a direct target of EPAS1. The CC genotype at rs56721780:G>C and the insertion genotype at −742 indel were found associated with higher EPAS1 and LOX expression levels in amnion, as well as higher birth weight of Tibetan newborns, suggesting that EPAS1 gene might play important roles in the development of amnion, fetus growth and high-altitude adaptation of Tibetans.

Expression of lysyl oxidase in human osteosarcoma and its clinical significance: A tumor suppressive role of LOX in human osteosarcoma cells

Lysyl oxidase (LOX) is an extracellular matrix (ECM) remodeling enzyme, which is involved in the development and progression of many types of tumors. LOX dysfunction is observed in colorectal, breast and ovarian cancer. However, the precise effects and molecular mechanisms of LOX action in osteosarcoma progression are still unknown. We evaluated the role of LOX in human osteosarcoma cell lines and clinical tumor samples in order to determine the function of this molecule. In our study, we showed that the expression level of LOX mRNA and protein were decreased in human osteosarcoma tissues as compared with normal tissue samples. In addition, we employed adenovirus-mediated overexpression of LOX in U-2OS and HOS cells to investigate the role of LOX in osteosarcoma cell lines. Adenovirus-mediated overexpression of LOX could efficiently increase the expression levels of LOX in osteosarcoma cell lines at both mRNA and protein levels. Increased expression of LOX inhibited the proliferation and migration of human osteosarcoma cells and promoted its apoptosis. Moreover, the Ki-67 and PCNA expression was decreased and MMP-2 and MMP-9 expression was inhibited. These findings also indicated that the effects of LOX may be mediated via the PI3K/AKT signaling pathway since LOX-mediated functions could be blocked by β-aminopropionitrile (β-APN), a LOX inhibitor. Taken together, our data indicated that LOX may be a tumor suppressor and could be regarded as a therapeutic target in human osteosarcoma.

Expression of lysyl oxidase and vascular endothelial growth factor in gastric cancer and their effects on tumor invasion and metastasis

Objective To study the expression of lysyl oxidase (LOX) and vascular endothelial growth factor (VEGF) in gastric cancer and paracancerous tissues,and their relationshipa with genesis of gastric cancer.Methods Sixty-five samples of freshly surgically resected gastric cancer tissues and the paracancerous tissues 5 cm away from the gastric cancer tissues each were collected in Wuhan Puren Hospital from Sep.2009 to Jan.2013.By using the method of Western blotting,the protein expression of LOX and VEGF in gastric cancer and paracancerous tissues was detected.The Spearman rank correlation analysis was done to analyze the correlation between the LOX and VEGF expression in gastric cancer.Results LOX relative expression in 65 cases of gastric cancer tissues and their paracancerous tissues was 0.4178 ± 0.1135 and 0.2174 ± 0.1524 respectively (P ＜ 0.05).VEGF in 65 cases of gastric cancer tissues and their paracancerous tissues was 0.4793 ± 0.1019 and 0.2116 ± 0.1146 respectively (P ＜ 0.05).Conclusion The expression levels of LOX and VEGF are obviously higher in the gastric cancer tissues than in the paracancerous tissues.LOX and VEGF may play an important role in the pathogenesis of gastric cancer. Key words: Gastric cancer; Lysyl oxidase; Vascular endothelial growth factor