Revealing the crosstalk between LOX+ fibroblast and M2 macrophage in gastric cancer by single-cell sequencing.

BACKGROUND/AIMS: Gastric cancer (GC) ranks among the prevalent types of cancer, and its progression is influenced by the tumor microenvironment (TME). A comprehensive comprehension of the TME associated with GC has the potential to unveil therapeutic targets of significance. METHODS: The complexity and heterogeneity of TME interactions were revealed through our investigation using an integrated analysis of single-cell and bulk-tissue sequencing data. RESULTS: We constructed a single-cell transcriptomic atlas of 150,913 cells isolated from GC patients. Our analysis revealed the intricate nature and heterogeneity of the GC TME and the metabolic properties of major cell types. Furthermore, two cell subtypes, LOX+ Fibroblasts and M2 Macrophages, were enriched in tumor tissue and related to the outcome of GC patients. In addition, LOX+ Fibroblasts were significantly associated with M2 macrophages. immunofluorescence double labeling indicated LOX+ Fibroblasts and M2 Macrophages were tightly localized in GC tissue. The two cell subpopulations strongly interacted in a hypoxic microenvironment, yielding an immunosuppressive phenotype. Our findings further suggest that LOX+ Fibroblasts may act as a trigger for inducing the differentiation of monocytes into M2 Macrophages via the IL6-IL6R signaling pathway. CONCLUSIONS: Our study revealed the intricate and interdependent communication network between the fibroblast and macrophage subpopulations, which could offer valuable insights for targeted manipulation of the tumor microenvironment.

Gene inactivation of lysyl oxidase in smooth muscle cells reduces atherosclerosis burden and plaque calcification in hyperlipidemic mice.

BACKGROUND AND AIMS: Lysyl oxidase (LOX) catalyzes the crosslinking of collagen and elastin to maintain tensile strength and structural integrity of the vasculature. Excessive LOX activity increases vascular stiffness and the severity of occlusive diseases. Herein, we investigated the mechanisms by which LOX controls atherogenesis and osteogenic differentiation of vascular smooth muscle cells (SMC) in hyperlipidemic mice. METHODS: Gene inactivation of Lox in SMC was achieved in conditional knockout mice after tamoxifen injections. Atherosclerosis burden and vascular calcification were assessed in hyperlipidemic conditional [Loxf/fMyh11-CreERT2ApoE-/-] and sibling control mice [Loxwt/wtMyh11-CreERT2ApoE-/-]. Mechanistic studies were performed with primary aortic SMC from Lox mutant and wild type mice. RESULTS: Inactivation of Lox in SMCs decreased > 70 % its RNA expression and protein level in the aortic wall and significantly reduced LOX activity without compromising vascular structure and function. Moreover, LOX deficiency protected mice against atherosclerotic burden (13 ± 2 versus 23 ± 1 %, p < 0.01) and plaque calcification (5 ± 0.4 versus 11.8 ± 3 %, p < 0.05) compared to sibling controls. Interestingly, gene inactivation of Lox in SMCs preserved the contractile phenotype of vascular SMC under hyperlipidemic conditions as demonstrated by single-cell RNA sequencing and immunofluorescence. Mechanistically, the absence of LOX in SMC prevented excessive collagen crosslinking and the subsequent activation of the pro-osteogenic FAK/ß-catenin signaling axis. CONCLUSIONS: Lox inactivation in SMC protects mice against atherosclerosis and plaque calcification by reducing SMC modulation and FAK/ß-catenin signaling.

A nomoscore of four genes for predicting the rupture risk in abdominal aortic aneurysm patients with osteoarthritis.

BACKGROUND: Abdominal aortic aneurysm (AAA) represents one of the most life-threatening cardiovascular diseases and is increasingly becoming a significant global public health concern. The aneurysms-osteoarthritis syndrome (AOS) has gained recognition, as patients with this syndrome often exhibit early-stage osteoarthritis (OA) and have a substantially increased risk of rupture, even with mild dilation of the aneurysm. The aim of this study was to discover potential biomarkers that can predict the occurrence of AAA rupture in patients with OA. METHODS: Two gene expression profile datasets (GSE98278, GSE51588) and two single-cell RNA-seq datasets (GSE164678, GSE152583) were obtained from the GEO database. Functional enrichment analysis, PPI network construction, and machine learning algorithms, including LASSO, Random Forest, and SVM-RFE, were utilized to identify hub genes. In addition, a nomogram and ROC curves were generated to predict the risk of rupture in patients with AAA. Moreover, we analyzed the immune cell infiltration in the AAA tissue microenvironment by CIBERSORT and validated key gene expression in different macrophage subtypes through single-cell analysis. RESULTS: A total of 105 intersecting DEGs that showed consistent changes between rAAA and OA dataset were identified. From these DEGs, four hub genes (PAK1, FCGR1B, LOX and PDPN) were selected by machine learning. High predictive performance was observed for the nomogram based on these hub genes, with an AUC of 0.975 (95 % CI: 0.942-1.000). Abnormal immune cell infiltration was detected in rAAA and correlated significantly with the hub genes. Ruptured AAA cases exhibited higher nomoscore values and lower M2 macrophage infiltration compared to stable AAA. Validation in animal models (PPE+BAPN-induced rAAA) confirmed the significant role of these biomarkers in AAA pathology. CONCLUSION: The present study successfully identified four potential hub genes (PAK1, FCGR1B, LOX and PDPN) and developed a robust predictive nomogram to assess the risk of AAA rupture. The findings also shed light on the connection between hub genes and immune cell components in the microenvironment of rAAA. These findings support future research on key genes in AAA patients with OA, providing insights for novel management strategies for AAA.

Lysyl oxidase-like 1 predicts the prognosis of patients with primary glioblastoma and promotes tumor invasion via EMT pathway.

Background: Lysyl oxidase enzymes (LOXs), as extracellular matrix (ECM) protein regulators, play vital roles in tumor progression by remodeling the tumor microenvironment. However, their roles in glioblastoma (GBM) have not been fully elucidated. Methods: The genetic alterations and prognostic value of LOXs were investigated via cBioPortal. The correlations between LOXs and biological functions/molecular tumor subtypes were explored in The Cancer Genome Atlas (TCGA) and the Chinese Glioma Genome Atlas (CGGA). After Kaplan‒Meier and Cox survival analyses, a Loxl1-based nomogram and prognostic risk score model (PRSM) were constructed and evaluated by time-dependent receiver operating characteristic curves, calibration curves, and decision curve analyses. Tumor enrichment pathways and immune infiltrates were explored by single-cell RNA sequencing and TIMER. Loxl1-related changes in tumor viability/proliferation and invasion were further validated by CCK-8, western blot, wound healing, and Transwell invasion assays. Results: GBM patients with altered LOXs had poor survival. Upregulated LOXs were found in IDH1-wildtype and mesenchymal (not Loxl1) GBM subtypes, promoting ECM receptor interactions in GBM. The Loxl1-based nomogram and the PRSM showed high accuracy, reliability, and net clinical benefits. Loxl1 expression was related to tumor invasion and immune infiltration (B cells, neutrophils, and dendritic cells). Loxl1 knockdown suppressed GBM cell proliferation and invasion by inhibiting the EMT pathway (through the downregulation of N-cadherin/Vimentin/Snai1 and the upregulation of E-cadherin). Conclusion: The Loxl1-based nomogram and PRSM were stable and individualized for assessing GBM patient prognosis, and the invasive role of Loxl1 could provide a promising therapeutic strategy.

IGF-II regulates lysyl oxidase propeptide and mediates its effects in part via basic helix-loop-helix E40.

Pulmonary fibrosis (PF) is a clinically severe and commonly fatal complication of Systemic Sclerosis (SSc). Our group has previously reported profibrotic roles for Insulin-like Growth Factor II (IGF-II) and Lysyl Oxidase (LOX) in SSc-PF. We sought to identify downstream regulatory mediators of IGF-II. In the present work, we show that SSc lung tissues have higher baseline levels of the total (N-glycosylated/unglycosylated) LOX-Propeptide (LOX-PP) than control lung tissues. LOX-PP-mediated changes were consistent with the extracellular matrix (ECM) deregulation implicated in SSc-PF progression. Furthermore, Tolloid-like 1 (TLL1) and Bone Morphogenetic Protein 1 (BMP1), enzymes that can cleave ProLOX to release LOX-PP, were increased in SSc lung fibrosis and the bleomycin (BLM)-induced murine lung fibrosis model, respectively. In addition, IGF-II regulated the levels of ProLOX, active LOX, LOX-PP, BMP1, and isoforms of TLL1. The Class E Basic Helix-Loop-Helix protein 40 (BHLHE40) transcription factor localized to the nucleus in response to IGF-II. BHLHE40 silencing downregulated TLL1 isoforms and LOX-PP, and restored features of ECM deregulation triggered by IGF-II. Our findings indicate that IGF-II, BHLHE40, and LOX-PP may serve as targets of therapeutic intervention to halt SSc-PF progression.

Lysyl Oxidase Production by Murine C3H10T1/2 Mesenchymal Stem Cells Is Increased by TGFßs and Differentially Modulated by Mechanical Stimuli.

Tendons are frequently injured and have limited regenerative capacity. This motivates tissue engineering efforts aimed at restoring tendon function through strategies to direct functional tendon formation. Generation of a crosslinked collagen matrix is paramount to forming mechanically functional tendon. However, it is unknown how lysyl oxidase (LOX), the primary mediator of enzymatic collagen crosslinking, is regulated by stem cells. This study investigates how multiple factors previously identified to promote tendon formation and healing (transforming growth factor [TGF]ß1 and TGFß2, mechanical stimuli, and hypoxia-inducible factor [HIF]-1α) regulate LOX production in the murine C3H10T1/2 mesenchymal stem cell (MSC) line. We hypothesized that TGFß signaling promotes LOX activity in C3H10T1/2 MSCs, which is regulated by both mechanical stimuli and HIF-1α activation. TGFß1 and TGFß2 increased LOX levels as a function of concentration and time. Inhibiting the TGFß type I receptor (TGFßRI) decreased TGFß2-induced LOX production by C3H10T1/2 MSCs. Low (5 mPa) and high (150 mPa) magnitudes of fluid shear stress were applied to test impacts of mechanical stimuli, but without TGFß2, loading alone did not alter LOX levels. Low loading (5 mPa) with TGFß2 increased LOX at 7 days greater than TGFß2 treatment alone. Neither HIF-1α knockdown (siRNA) nor activation (CoCl2) affected LOX levels. Ultimately, results suggest that TGFß2 and appropriate loading magnitudes contribute to LOX production by C3H10T1/2 MSCs. Potential application of these findings includes treatment with TGFß2 and appropriate mechanical stimuli to modulate LOX production by stem cells to ultimately control collagen matrix stiffening and support functional tendon formation.

Deletion of mouse lysyl oxidase in megakaryocytes affects bone properties in a sex-dependent manner.

ABSTRACT: Lysyl oxidase (LOX) is a facilitator of extracellular matrix cross-linking. Using newly developed megakaryocyte-specific LOX knockout mice, we show that LOX expressed in these scarce bone marrow cells affects bone volume and collagen architecture in a sex-dependent manner.

Crosstalk of lysyl oxidase-like 1 and lysyl oxidase prolongs their half-lives and regulates liver fibrosis through Notch signal.

BACKGROUND: Lysyl oxidase (LOX) family members (LOX and LOXL1 to 4) are crucial copper-dependent enzymes responsible for cross-linking collagen and elastin. Previous studies have revealed that LOX and LOXL1 are the most dramatically dysregulated LOX isoforms during liver fibrosis. However, the crosstalk between them and the underlying mechanisms involved in the profibrotic behaviors of HSCs, as well as the progression of liver fibrosis, remain unclear. METHODS: pCol9GFP-HS4,5Tg mice, Loxl1fl/flGfapCre mice, human HSC line, and primary HSCs were enrolled to study the dysregulation pattern, profibrotic roles, and the potential mechanisms of LOX and LOXL1 interaction involved in the myofibroblast-like transition of HSCs and liver fibrogenesis. RESULTS: LOX and LOXL1 were synergistically upregulated during liver fibrogenesis, irrespective of etiology, together orchestrating the profibrotic behaviors of HSCs. LOX and LOXL1 coregulated in HSCs, whereas LOXL1 dominated in the coregulation loop. Interestingly, the interaction between LOXL1 and LOX prolonged their half-lives, specifically enhancing the Notch signal-mediated myofibroblast-like transition of HSCs. Selective disruption of Loxl1 in Gfap+ HSCs deactivated the Notch signal, inhibited HSC activation, and relieved carbon tetrachloride-induced liver fibrosis. CONCLUSIONS: Our current study confirmed the synergistic roles and the underlying mechanisms of LOXL1 and LOX crosstalk in the profibrotic behaviors of HSCs and liver fibrosis progression, providing experimental evidence for further clear mechanism-based anti-LOXL1 strategy development in the therapy of liver fibrosis.

Inhibition of lysyl oxidase by pharmacological intervention and genetic manipulation alleviates epilepsy-associated cognitive disorder.

Epilepsy-associated cognitive disorder (ECD), a prevalent comorbidity in epilepsy patients, has so far uncharacterized etiological origins. Our prior work revealed that lysyl oxidase (Lox) acted as a novel contributor of ferroptosis, a recently discovered cell death mode in the regulation of brain function. However, the role of Lox-mediated ferroptosis in ECD remains unknown. ECD mouse model was established 2 months later following a single injection of kainic acid (KA) for. After chronic treatment with KA, mice were treated with different doses (30 mg/kg, 100 mg/kg and 300 mg/kg) of Lox inhibitor BAPN. Additionally, hippocampal-specific Lox knockout mice was also constructed and employed to validate the role of Lox in ECD. Cognitive functions were assessed using novel object recognition test (NOR) and Morris water maze test (MWM). Protein expression of phosphorylated cAMP-response element binding (CREB), a well-known molecular marker for evaluation of cognitive performance, was also detected by Western blot. The protein distribution of Lox was analyzed by immunofluorescence. In KA-induced ECD mouse model, ferroptosis process was activated according to upregulation of 4-HNE protein and a previously discovered ferroptosis in our group, namely, Lox was remarkably increased. Pharmacological inhibition of Lox by BAPN at the dose of 100 mg/kg significantly increased the discrimination index following NOR test and decreased escape latency as well as augmented passing times within 60 s following MWM test in ECD mouse model. Additionally, deficiency of Lox in hippocampus also led to pronounced improvement of deficits in ECD model. These findings indicate that the ferroptosis regulatory factor, Lox, is activated in ECD. Ablation of Lox by either pharmacological intervention or genetic manipulation ameliorates the impairment in ECD mouse model, which suggest that Lox serves as a promising therapeutic target for treating ECD in clinic.

Lysyl oxidase expression in smooth muscle cells determines the level of intima calcification in hypercholesterolemia-induced atherosclerosis. / La expresión de la lisil oxidasa en las células musculares lisas determina el nivel de calcificación de la íntima en la aterosclerosis inducida por hipercolesterolemia.

INTRODUCTION: Cardiovascular calcification is an important public health issue with an unmeet therapeutic need. We had previously shown that lysyl oxidase (LOX) activity critically influences vascular wall smooth muscle cells (VSMCs) and valvular interstitial cells (VICs) calcification by affecting extracellular matrix remodeling. We have delved into the participation of LOX in atherosclerosis and vascular calcification, as well as in the mineralization of the aortic valve. METHODS: Immunohistochemical and expression studies were carried out in human atherosclerotic lesions and experimental models, valves from patients with aortic stenosis, VICs, and in a genetically modified mouse model that overexpresses LOX in CMLV (TgLOXCMLV). Hyperlipemia and atherosclerosis was induced in mice through the administration of adeno-associated viruses encoding a PCSK9 mutated form (AAV-PCSK9D374Y) combined with an atherogenic diet. RESULTS: LOX expression is increased in the neointimal layer of atherosclerotic lesions from human coronary arteries and in VSMC-rich regions of atheromas developed both in the brachiocephalic artery of control (C57BL/6J) animals transduced with PCSK9D374Y and in the aortic root of ApoE-/- mice. In TgLOXCMLV mice, PCSK9D374Y transduction did not significantly alter the enhanced aortic expression of genes involved in matrix remodeling, inflammation, oxidative stress and osteoblastic differentiation. Likewise, LOX transgenesis did not alter the size or lipid content of atherosclerotic lesions in the aortic arch, brachiocephalic artery and aortic root, but exacerbated calcification. Among lysyl oxidase isoenzymes, LOX is the most expressed member of this family in highly calcified human valves, colocalizing with RUNX2 in VICs. The lower calcium deposition and decreased RUNX2 levels triggered by the overexpression of the nuclear receptor NOR-1 in VICs was associated with a reduction in LOX. CONCLUSIONS: Our results show that LOX expression is increased in atherosclerotic lesions, and that overexpression of this enzyme in VSMC does not affect the size of the atheroma or its lipid content, but it does affect its degree of calcification. Further, these data suggest that the decrease in calcification driven by NOR-1 in VICs would involve a reduction in LOX. These evidences support the interest of LOX as a therapeutic target in cardiovascular calcification.

Lysyl oxidase-mediated elastin upregulation promotes the proliferation and migration of human retinal endothelial cells.

BACKGROUND: Proliferative diabetic retinopathy (PDR) is a major cause of irreversible blindness in the working age population. The dysfunction of retinal vascular endothelial cells (RVECs) is the primary cause of PDR. Extracellular matrix (ECM) accumulation promotes intracellular signaling required for RVEC proliferation, migration, survival, and tube morphogenesis. OBJECTIVES: This study aimed to investigate the role of lysyl oxidase (LOX) in the cellular function of RVECs and PDR pathogenesis and to identify the underlying mechanisms. MATERIAL AND METHODS: Protein expression was determined with western blot. The interaction between LOX and elastin (ELN) was detected using a co-immunoprecipitation (Co-IP) assay, and the Cell Counting Kit-8 (CCK-8) assay evaluated cell viability. A colony formation assay was employed to assess the proliferation of human RVECs (hRVECs), and a transwell assay to determine their migration ability. Streptozotocin was used to establish PDR in mice in vivo. A histological analysis was conducted using hematoxylin and eosin (H&E) staining. RESULTS: The results showed that LOX was overexpressed in PDR patients. The LOX knockdown suppressed ECM formation and hRVEC proliferation and migration. Additionally, LOX upregulated ELN expression. However, overexpressed ELN promoted hRVEC proliferation and migration. In vivo experiments showed that curcumin-mediated LOX deficiency restored retinal tissue structure. CONCLUSIONS: The LOX-knockdown suppressed ECM formation and hRVEC proliferation and migration by inactivating ELN. Therefore, LOX/ELN signaling may be a potential PDR biomarker.

Lysyl Oxidase Regulates Epithelial Differentiation and Barrier Integrity in Eosinophilic Esophagitis.

BACKGROUND & AIMS: Epithelial disruption in eosinophilic esophagitis (EoE) encompasses both impaired differentiation and diminished barrier integrity. We have shown that lysyl oxidase (LOX), a collagen cross-linking enzyme, is up-regulated in the esophageal epithelium in EoE. However, the functional roles of LOX in the esophageal epithelium remains unknown. METHODS: We investigated roles for LOX in the human esophageal epithelium using 3-dimensional organoid and air-liquid interface cultures stimulated with interleukin (IL)13 to recapitulate the EoE inflammatory milieu, followed by single-cell RNA sequencing, quantitative reverse-transcription polymerase chain reaction, Western blot, histology, and functional analyses of barrier integrity. RESULTS: Single-cell RNA sequencing analysis on patient-derived organoids revealed that LOX was induced by IL13 in differentiated cells. LOX-overexpressing organoids showed suppressed basal and up-regulated differentiation markers. In addition, LOX overexpression enhanced junctional protein genes and transepithelial electrical resistance. LOX overexpression restored the impaired differentiation and barrier function, including in the setting of IL13 stimulation. Transcriptome analyses on LOX-overexpressing organoids identified an enriched bone morphogenetic protein (BMP) signaling pathway compared with wild-type organoids. In particular, LOX overexpression increased BMP2 and decreased the BMP antagonist follistatin. Finally, we found that BMP2 treatment restored the balance of basal and differentiated cells. CONCLUSIONS: Our data support a model whereby LOX exhibits noncanonical roles as a signaling molecule important for epithelial homeostasis in the setting of inflammation via activation of the BMP pathway in the esophagus. The LOX/BMP axis may be integral in esophageal epithelial differentiation and a promising target for future therapies.

Lysyl oxidase like-1 deficiency in optic nerve head astrocytes elicits reactive astrocytosis and alters functional effects of astrocyte derived exosomes.

Glaucoma is a multifactorial progressive ocular pathology that manifests clinically with damage to the optic nerve (ON) and the retina, ultimately leading to blindness. The optic nerve head (ONH) shows the earliest signs of glaucoma pathology, and therefore, is an attractive target for drug discovery. The goal of this study was to elucidate the effects of reactive astrocytosis on the elastin metabolism pathway in primary rat optic nerve head astrocytes (ONHA), the primary glial cell type in the unmyelinated ONH. Following exposure to static equibiaxial mechanical strain, we observed prototypic molecular and biochemical signatures of reactive astrocytosis that were associated with a decrease in lysyl oxidase like 1 (Loxl1) expression and a concomitant decrease in elastin (Eln) gene expression. We subsequently investigated the role of Loxl1 in reactive astrocytosis by generating primary rat ONHA cultures with ∼50% decreased Loxl1 expression. Our results suggest that reduced Loxl1 expression is sufficient to elicit molecular signatures of elastinopathy in ONHA. Astrocyte derived exosomes (ADE) significantly increased the length of primary neurites of primary neurons in vitro. In contrast, ADE from Loxl1-deficient ONHA were deficient of trophic effects on neurite outgrowth in vitro, positing that Loxl1 dysfunction and the ensuing impaired elastin synthesis during reactive astrocytosis in the ONH may contribute to impaired neuron-glia signaling in glaucoma. Our data support a role of dysregulated Loxl1 function in eliciting reactive astrocytosis in glaucoma subtypes associated with increased IOP, even in the absence of genetic polymorphisms in LOXL1 typically associated with exfoliation glaucoma. This suggests the need for a paradigm shift toward considering lysyl oxidase activity and elastin metabolism and signaling as contributors to an altered secretome of the ONH that may lead to the progression of glaucomatous changes. Future research is needed to investigate cargo of exosomes in the context of reactive astrocytosis and identify the pathways leading to the observed transcriptome changes during reactive astrocytosis.

A lysyl oxidase-responsive collagen peptide illuminates collagen remodeling in wound healing.

Tissue repair and fibrosis involve the dynamic remodeling of collagen, and accurate detection of these sites is of utmost importance. Here, we use a collagen peptide sensor (1) to visualize collagen formation and remodeling during wound healing in mice and humans. We show that the probe binds selectively to sites of collagen formation and remodeling at different stages of healing. Compared to conventional methods, the peptide sensor localizes preferentially to areas of collagen synthesis and remodeling at the wound edge and not in matured fibrillar collagen. We also demonstrate its applicability for in vivo wound imaging and for discerning differential remodeling in wounds of transgenic mice with altered collagen dynamics. Our findings show the value of 1 as a diagnostic tool to rapidly identify the sites of matrix remodeling in tissue sections, which will aid in the conception of new therapeutic strategies for fibrotic disorders and defective tissue repair.

Dysregulation of extracellular matrix and Lysyl Oxidase in Ehlers-Danlos syndrome type IV skin fibroblasts.

BACKGROUND: Ehlers-Danlos syndrome Type IV (aka Vascular Ehlers Danlos, or vEDS) is a dominantly inherited mutation in the Collagen 3A1 gene (COL3A1). The disease is characterized by tissue friability and age-related susceptibility to arterial aneurysm, dissection and rupture as well as uterine and bowl tears. These clinical manifestations result in major surgical intervention and decreased life expectancy. Understanding how mutations in COL3A1 impact the structure and function of the extracellular matrix (ECM) is important to managing the disease and finding treatments. RESULTS: Skin fibroblasts from vEDS subjects heterozygous for the p.G588S pathogenic variant in the COL3A1 gene and a normal individual were cultured and studied. Proteomics analysis identified dozens of upregulated proteins related to extracellular matrix dysregulation that is characteristic of fibrosis. Gene expression libraries from cultured primary fibroblasts were screened for messenger RNA (mRNA) markers of ECM degradation. The proteomics and targeted gene expression array results were largely consistent with dysregulation of the extracellular matrix in vEDS. The data show upregulation of multiple Collagen proteins and genes, other ECM components, and enzymes related to ECM processing and turn-over. vEDS fibroblasts expressed significantly more cross linked C-Telopeptide of Collagen III (CTXIII) than normal fibroblasts, indicative of Collagen III degradation and turn-over. Further, the expression and activity of Lysyl Oxidase (LOX), an enzyme that initiates covalent cross-linking of soluble collagen and elastin into protease resistant fibers, is elevated in vEDS fibroblasts compared to normal fibroblasts. CONCLUSION: Together, these findings suggest dysregulated ECM deposition and processing, reminiscent of a state of fibrosis. Therapeutics that target the dysregulated ECM proteins or help replace damaged tissue may improve clinical outcomes.

LOXL4 Shuttled by Tumor Cells-derived Extracellular Vesicles Promotes Immune Escape in Hepatocellular Carcinoma by Activating the STAT1/PD-L1 Axis.

Emerging evidence has validated that extracellular vesicles (EVs) regulate hepatocellular carcinoma (HCC) progression, while its role in HCC immune escape remains to be elucidated. This study investigates the role of EVs-encapsulated lysyl oxidase like-4 (LOXL4) derived from tumor cells in HCC immune escape. HCC-related microarray data sets GSE36376 and GSE87630 were obtained for differential analysis, followed by identifying the essential genes related to the prognosis of HCC patients. Bone marrow-derived macrophages were treated with EVs derived from mouse Hepa 1-6 cells and cocultured with CD8 + T cells to observe the CD8 + T-cell activity. At last, a mouse HCC orthotopic xenograft model was constructed to verify the effects of HCC cell-derived EVs on the immune escape of HCC cells and tumorigenicity in vivo by delivering LOXL4. It was found that ACAT1, C4BPA, EHHADH, and LOXL4 may be the essential genes related to the prognosis of HCC patients. On the basis of the TIMER database, there was a close correlation between LOXL4 and macrophage infiltration in HCC. Besides, STAT1 was closely related to LOXL4. In vitro experiments demonstrated that LOXL4 could induce programmed death-ligand 1 expression in macrophages and immunosuppression by activating STAT1. In vivo experiments also verified that HCC cell-derived EVs promoted the immune escape of HCC cells and tumorigenicity by delivering LOXL4. LOXL4 was delivered into macrophages via EVs to induce programmed death-ligand 1 by activating STAT1 and inhibiting the killing ability of CD8 + T cells to HCC cells, thus promoting immune escape in HCC.

Fibroblast-Derived Lysyl Oxidase Increases Oxidative Phosphorylation and Stemness in Cholangiocarcinoma.

BACKGROUND & AIMS: Metabolic and transcriptional programs respond to extracellular matrix-derived cues in complex environments, such as the tumor microenvironment. Here, we demonstrate how lysyl oxidase (LOX), a known factor in collagen crosslinking, contributes to the development and progression of cholangiocarcinoma (CCA). METHODS: Transcriptomes of 209 human CCA tumors, 143 surrounding tissues, and single-cell data from 30 patients were analyzed. The recombinant protein and a small molecule inhibitor of the LOX activity were used on primary patient-derived CCA cultures to establish the role of LOX in migration, proliferation, colony formation, metabolic fitness, and the LOX interactome. The oncogenic role of LOX was further investigated by RNAscope and in vivo using the AKT/NICD genetically engineered murine CCA model. RESULTS: We traced LOX expression to hepatic stellate cells and specifically hepatic stellate cell-derived inflammatory cancer-associated fibroblasts and found that cancer-associated fibroblast-driven LOX increases oxidative phosphorylation and metabolic fitness of CCA, and regulates mitochondrial function through transcription factor A, mitochondrial. Inhibiting LOX activity in vivo impedes CCA development and progression. Our work highlights that LOX alters tumor microenvironment-directed transcriptional reprogramming of CCA cells by facilitating the expression of the oxidative phosphorylation pathway and by increasing stemness and mobility. CONCLUSIONS: Increased LOX is driven by stromal inflammatory cancer-associated fibroblasts and correlates with diminished survival of patients with CCA. Modulating the LOX activity can serve as a novel tumor microenvironment-directed therapeutic strategy in bile duct pathologies.

Pan-Cancer Analysis of the LOX Family Reveals that LOX Affects Tumor Prognosis by Affecting Immune Infiltration.

The lysyl oxidase (LOX) gene family encodes for a group of copper-dependent enzymes that play a crucial role in the cross-linking of collagen and elastin fibers in the extracellular matrix (ECM). Dysregulation of LOX gene expression has been implicated in various pathological conditions, including cancer. Several studies have shown that the LOX gene family is involved in cancer progression and metastasis. The goal of this article is to conduct a comprehensive analysis of the LOX family's role in pan-cancer multiplexes. We utilized pan-cancer multi-omics sequencing data from TCGA to investigate the relationship between LOX family genes and tumors at four different levels: mutation, copy number variation, methylation, and gene expression. In addition, we also examined the relationship between LOX family genes and tumors at the cell line level using tumor cell line sequencing data from CCLE. Taking into account the impact of LOX family genes on lung cancer, we developed a LOX family lung cancer prognostic model to forecast the disease's prognosis. Our findings revealed that LOXL2 had the highest mutation frequency in tumors, while all four LOX family genes experienced some degree of copy number variation in diverse tumors. We observed that LOX, LOXL1 to LOXL3 were predominantly highly expressed in tumors including LUAD. The expression trends of LOX and LOXL1 to LOXL3 were consistent across tumor cell lines, but differed somewhat from LOXL4. Utilizing 25 LOX family-related genes, we constructed a LOX family prognostic model that performed well in predicting the prognosis of lung cancer. Through pan-cancer analysis, we gain further knowledge of the role of LOX family genes in different tumors, offering a novel pathway for future research into the relationship between LOX family genes and tumors.

Forkhead box F2/ Lysyl oxidase like 1 contribute to epithelial-mesenchymal transition and angiogenesis in thyroid cancer.

BACKGROUND: Bioinformatics analysis suggests an association between lysyl oxidase like 1 (LOXL1) and forkhead box F2 (FOXF2), both of which are found to be dysregulated in thyroid cancer. This study aims to elucidate their specific roles in thyroid cancer. METHODS: The correlation of LOXL1 expression with thyroid cancer staging and the overall survival was analyzed. LOXL1 levels were determined in several thyroid cancer cells, and its effects on poorly differentiated BCPAP cell proliferation, colony formation, malignant phenotypes, epithelial-mesenchymal transition (EMT) progression, and angiogenesis were evaluated. The relationship between LOXL1 and FOXF2 was confirmed using Luciferase reporter and ChIP assays. The impacts of FOXF2 on LOXL1 regulation along with the Wnt/ß-catenin signaling were assessed, followed by the verification of transplanted tumor in nude mice. RESULTS: Elevated LOXL1 expression was associated with advanced clinical staging and poorer overall survival. Reduced LOXL1 suppressed cell proliferation, colony formation, migration, invasion, EMT, and angiogenesis. FOXF2 was found to be down-regulated in thyroid cancer, acting as a transcription factor that recognizes the LOXL1 promoter and modulates its transcriptional expression. Moreover, the regulatory outcome of LOXL1 knockdown was partially reversed upon FOXF2 knockdown, including the modulation of the Wnt/ß-catenin signaling and tumor growth in vivo. CONCLUSION: Our findings indicate that LOXL1 is transcriptionally regulated by FOXF2 and activates the Wnt/ß-catenin to promote malignant phenotypes, EMT progression, and angiogenesis in BCPAP cells.

Targeting Lysyl Oxidase as a Potential Therapeutic Approach to Reducing Fibrotic Scars Post-operatively: Its Biological Role in Post-Surgical Scar Development.

Abdominal and pelvic surgery, or any surgical injury of the peritoneum, often leads to chronic abdominal adhesions that may lead to bowel obstruction, infertility, and pain. Current therapeutic strategies are usually ineffective, and the pathological mechanisms of the disease are unclear. Excess collagen cross-linking is a key mediator for extra-cellular matrix deposition and fibrogenesis. Lysyl oxidase is a key enzyme that catalyzes the formation of stabilizing cross-links in collagen. Dysregulation of Lysyl oxidase (Lox) expressing upregulates collagen cross-linking, leading ECM deposition. Tissue hypoxia during surgery induces molecular mechanisms and active transcription factors to promote the expression of several genes related to inflammation, oxidative stress, and fibrosis, such as transforming growth factor beta, and Lox. Studies have shown that targeting Lox improves clinical outcomes and fibrotic parameters in liver, lung, and myocardial fibrosis, therefore, Lox may be a potential drug target in the prevention of postsurgical adhesion.