Exploring the Interplay between Polyphenols and Lysyl Oxidase Enzymes for Maintaining Extracellular Matrix Homeostasis.

Collagen, the most abundant structural protein found in mammals, plays a vital role as a constituent of the extracellular matrix (ECM) that surrounds cells. Collagen fibrils are strengthened through the formation of covalent cross-links, which involve complex enzymatic and non-enzymatic reactions. Lysyl oxidase (LOX) is responsible for catalyzing the oxidative deamination of lysine and hydroxylysine residues, resulting in the production of aldehydes, allysine, and hydroxyallysine. These intermediates undergo spontaneous condensation reactions, leading to the formation of immature cross-links, which are the initial step in the development of mature covalent cross-links. Additionally, non-enzymatic glycation contributes to the formation of abnormal cross-linking in collagen fibrils. During glycation, specific lysine and arginine residues in collagen are modified by reducing sugars, leading to the creation of Advanced Glycation End-products (AGEs). These AGEs have been associated with changes in the mechanical properties of collagen fibers. Interestingly, various studies have reported that plant polyphenols possess amine oxidase-like activity and can act as potent inhibitors of protein glycation. This review article focuses on compiling the literature describing polyphenols with amine oxidase-like activity and antiglycation properties. Specifically, we explore the molecular mechanisms by which specific flavonoids impact or protect the normal collagen cross-linking process. Furthermore, we discuss how these dual activities can be harnessed to generate properly cross-linked collagen molecules, thereby promoting the stabilization of highly organized collagen fibrils.

Phosphorylation of AKT by lysyl oxidase-like 2 activates the PI3K/AKT signaling pathway to promote proliferation, invasion and metastasis in esophageal squamous carcinoma.

OBJECTIVE: Esophageal squamous cell carcinoma (ESCC) is a common and aggressive malignancy of the gastrointestinal tract for which therapeutic options are scarce. This study screens for LOXL2, a key gene in ESCC, and explains the molecular mechanism by which it promotes the progression of ESCC. METHODS: Immunohistochemical staining was performed to detect the expression level of LOXL2 in ESCC tissues and paraneoplastic tissues. CCK-8 and Transwell assays were performed to assess the effects of LOXL2 knockdown and overexpression on the proliferation, apoptosis, migration and invasion ability of ESCC cells. High-throughput sequencing analysis screens for molecular mechanisms of action by which LOXL2 promotes ESCC progression. Western blotting and qRT-PCR were used to determine the expression levels of relevant markers. RESULTS: LOXL2 is positively expressed in ESCC and highly correlated with poor prognosis. Silencing LOXL2 significantly inhibited the proliferation, migration and invasive ability of ESCC cells, whereas overexpression showed the opposite phenotype. High-throughput sequencing suggested that LOXL2-associated differentially expressed genes were highly enriched in the PI3K/AKT signaling pathway. In vitro cellular assays confirmed that silencing LOXL2 significantly reduced PI3K, p-AKTThr308 and p-AKTSer473 gene and protein expression levels, while overexpression increased all three gene and protein levels, while AKT gene and protein expression levels were not significantly different. CONCLUSION: This study found that LOXL2 may regulate the PI3K/AKT signaling pathway and exert protumor effects on ESCC cells through phosphorylation of AKT. LOXL2 may be a key clinical warning biomarker or therapeutic target for ESCC.

Implications of lysyl oxidase-like protein 3 expression in the periodontium of diabetic rats.

OBJECTIVES: Diabetes has been strongly associated with periodontal diseases. The periodontal ligament (PDL) has an abundant extracellular matrix (ECM). Lysyl oxidases (LOXs) are closely associated with various diseases caused by abnormal ECM functions, however, the role of LOXs in periodontal diseases induced by diabetes remains unclear. METHODOLOGY: In this study, 8-week-old Zucker diabetic fatty rats were used to establish a type 2 diabetes mellitus (T2DM) model. After 9 and 16 weeks, hematoxylin and eosin (H&E), Masson's trichrome, and immunohistochemical staining were performed. RESULTS: After 9 weeks, loose collagen fibers were found in the interradicular area of the diabetic group, in opposition to the control group. There were no significant differences in LOX expression between the diabetic and control groups (p>0.05). However, after 16 weeks, the diabetic group presented a disordered arrangement of the PDL, showing decreased collagen content and significantly increased lysyl oxidase-like protein 3 (LOXL3) expression when compared with the control group (p<0.05). This suggests that LOXL3 plays a significant role in periodontal histopathological changes in diabetic rats. CONCLUSION: Our study showed elevated LOXL3 expression in the PDL of diabetic rats after 16 weeks, suggesting that LOXL3 may be involved in the occurrence and development of periodontal histopathological changes in diabetic rats. LOXL3 could be further used as an indicator for the early diagnosis of diabetic periodontitis in T2DM patients in clinical settings.

Lysyl Oxidase Like-4 (LOXL4) as a tumor marker and prognosticator in advanced stage laryngeal cancer.

INTRODUCTION: Lysyl oxidase-like 4 is an amine oxidase from the lysyl oxidase family that was previously shown to be overexpressed in head and neck cancer and upregulated in response to hypoxia. The possible role of lysyl oxidase-like 4 as a tumor marker in advanced stage larynx cancer was investigated. OBJECTIVE: To investigate the expression of lysyl Oxidase-Like 4 protein in advanced stage laryngeal cancer and elucidate its possible role as a tumor marker, predictor of treatment response and prognosticator. METHODS: Diagnostic specimens of 72 patients treated for stage III-IV laryngeal squamous cell carcinoma were evaluated for lysyl oxidase-like 4 expression by immunohistochemistry. RESULTS: Lysyl oxidase-like 4 expression was correlated with advanced tumor stage (p = 0.041) and better differentiation (p = 0.025) but was independent of tumor diameter (p = 0.456). Response to induction chemotherapy or the need for salvage laryngectomy were not affected by lysyl oxidase-like 4 expression (p = 0.999, p = 0.070 respectively). Increased lysyl oxidase-like 4 expression was associated with better 2 year overall survival in both univariate (p = 0.036) and multivariate analyses (p = 0.014). CONCLUSION: Lysyl oxidase-like 4 expression emerges with advancing stages, is lost with worsening differentiation, and may have tumor suppressive properties in larynx cancer.

Dysregulation of Lysyl Oxidases Expression in Diabetic Nephropathy and Renal Cell Carcinoma.

Lysyl oxidases (LOXs) are amino oxidase enzymes that catalyze the oxidative deamination of lysine and hydroxylysine residues to form allysine, the first step towards the development of the final cross-linking reaction in collagens, a crucial macromolecule that reinforces extracellular matrices. Basement membranes are specialized extracellular matrices that are essential components of the glomerular filtration barrier, which also support tubular epithelial cells. Lysyl oxidases are post-translational enzymes indispensable for tissue architecture, participating actively in the development and function of kidneys. The differential expression and dysregulation of these enzymes promote diabetic nephropathy, one of the major complications observed in end-stage renal diseases. In addition, these enzymes act as transcription factors that trigger the epithelial-mesenchymal transition responsible for the generation of different cancers. In the kidney, the expression studies in physiological conditions identified LOXL1 and LOXL2 as constituent proteins of glomerular basement membranes. Besides, LOX and LOXL2 are upregulated in fibrosis and renal cell carcinoma. The current review summarizes the physiological expression of LOXs enzymes in the nephrons, including glomerulus and tubules. Their roles in renal diseases are particularly highlighted in diabetic nephropathy and renal cell carcinoma, two pathophysiological conditions where these enzymes have been demonstrated to participate. The focus of the present study is to describe and discuss the current understanding in this field. The current potential of LOXs enzymes as a biomarker and pharmacological target to kidney diseases that involves extracellular matrix cross-linking enzymes is also discussed. LOXs isoforms and their capacity as therapeutic targets could be used for diagnostic and prognostic purposes and in treating these renal complications.

The Dynamic Interaction between Extracellular Matrix Remodeling and Breast Tumor Progression.

Desmoplastic tumors correspond to a unique tissue structure characterized by the abnormal deposition of extracellular matrix. Breast tumors are a typical example of this type of lesion, a property that allows its palpation and early detection. Fibrillar type I collagen is a major component of tumor desmoplasia and its accumulation is causally linked to tumor cell survival and metastasis. For many years, the desmoplastic phenomenon was considered to be a reaction and response of the host tissue against tumor cells and, accordingly, designated as "desmoplastic reaction". This notion has been challenged in the last decades when desmoplastic tissue was detected in breast tissue in the absence of tumor. This finding suggests that desmoplasia is a preexisting condition that stimulates the development of a malignant phenotype. With this perspective, in the present review, we analyze the role of extracellular matrix remodeling in the development of the desmoplastic response. Importantly, during the discussion, we also analyze the impact of obesity and cell metabolism as critical drivers of tissue remodeling during the development of desmoplasia. New knowledge derived from the dynamic remodeling of the extracellular matrix may lead to novel targets of interest for early diagnosis or therapy in the context of breast tumors.

Fibrostenotic eosinophilic esophagitis might reflect epithelial lysyl oxidase induction by fibroblast-derived TNF-α.

BACKGROUND: Fibrosis and stricture are major comorbidities in patients with eosinophilic esophagitis (EoE). Lysyl oxidase (LOX), a collagen cross-linking enzyme, has not been investigated in the context of EoE. OBJECTIVE: We investigated regulation of epithelial LOX expression as a novel biomarker and functional effector of fibrostenotic disease conditions associated with EoE. METHODS: LOX expression was analyzed by using RNA-sequencing, PCR assays, and immunostaining in patients with EoE; cytokine-stimulated esophageal 3-dimensional organoids; and fibroblast-epithelial cell coculture, the latter coupled with fluorescence-activated cell sorting. RESULTS: Gene ontology and pathway analyses linked TNF-α and LOX expression in patients with EoE, which was validated in independent sets of patients with fibrostenotic conditions. TNF-α-mediated epithelial LOX upregulation was recapitulated in 3-dimensional organoids and coculture experiments. We find that fibroblast-derived TNF-α stimulates epithelial LOX expression through activation of nuclear factor κB and TGF-ß-mediated signaling. In patients receiver operating characteristic analyses suggested that LOX upregulation indicates disease complications and fibrostenotic conditions in patients with EoE. CONCLUSIONS: There is a novel positive feedback mechanism in epithelial LOX induction through fibroblast-derived TNF-α secretion. Esophageal epithelial LOX might have a role in the development of fibrosis with substantial translational implications.

Regulation of extracellular matrix elements and sarcomerogenesis in response to different periods of passive stretching in the soleus muscle of rats.

Stretching is a common method used to prevent muscle shortening and improve limited mobility. However, the effect of different time periods on stretching-induced adaptation of the extracellular matrix and its regulatory elements have yet to be investigated. We aimed to evaluate the expression of fibrillar collagens, sarcomerogenesis, metalloproteinase (MMP) activity and gene expression of the extracellular matrix (ECM) regulators in the soleus (SOL) muscle of rats submitted to different stretching periods. The soleus muscles were submitted to 10 sets of passive stretching over 10 (St 10d) or 15 days (St 15d) (1 min per set, with 30 seconds' rest between sets). Sarcomerogenesis, muscle cross-sectional area (CSA), and MMP activity and mRNA levels in collagen (type I, III and IV), connective tissue growth factor (CTGF), growth factor-beta (TGF-ß), and lysyl oxidase (LOX) were analyzed. Passive stretching over both time periods mitigated COL-I deposition in the SOL muscle of rats. Paradoxically, 10 days of passive stretching induced COL-I and COL-III synthesis, with concomitant upregulation of TGF-ß1 and CTGF at a transcriptional level. These responses may be associated with lower LOX mRNA levels in SOL muscles submitted to 10 passive stretching sessions. Moreover, sarcomerogenesis was observed after 15 days of stretching, suggesting that stretching-induced muscle adaptations are time-dependent responses.

Lysyl oxidase isoforms in gastric cancer.

Gastric cancer (GC) is the fifth most frequent cancer in the world and shows the highest incidence in Latin America and Asia. An increasing amount of evidence demonstrates that lysyl oxidase isoforms, a group of extracellular matrix crosslinking enzymes, should be considered as potential biomarkers and therapeutic targets in GC. In this review, we focus on the expression levels of lysyl oxidase isoforms, its functions and the clinical implications in GC. Finding novel proteins related to the processing of these extracellular matrix enzymes might be helpful in the design of new therapies, which, in combination with classic pharmacology, could be used to delay the progress of this aggressive cancer and offer a wider temporal window for clinical intervention.

Genes responsible for vaginal extracellular matrix metabolism are modulated by women's reproductive cycle and menopause.

OBJECTIVES: To analyze the expression of genes involved in extracellular matrix (ECM) biogenesis and remodeling in vaginal tissue of women with clinically normal pelvic floor support (defined as controls) according to the phase of menstrual cycle and postmenopausal women with and without pelvic organ prolapse (POP). MATERIALS AND METHODS: This study examined the expression of matrix metalloproteinases (MMPs), their tissue inhibitors (TIMPs), and the Lysyl oxidase (LOX) family genes in the anterior vaginal wall of Caucasian women by real-time RT-PCR. Initially, mRNA expression was assessed in premenopausal controls in the secretory (group 1, n = 10) vs. proliferative (group 2, n = 8) phase of menstrual cycle. In addition, we compared premenopausal controls in the proliferative phase (group 2) vs. postmenopausal controls (group 3, n = 5). Finally, we analyzed postmenopausal controls (group 3) vs. postmenopausal women with advanced POP (group 4, n = 13). RESULTS: According to the phase of menstrual cycle, MMP1 was significantly reduced (p = 0.003), whereas the expression of TIMP1 and LOXL4 was significantly up-regulated during proliferative phase (both p < 0.01) when compared to the secretory phase in premenopausal control women. Regarding menopausal status/ageing, all MMPs were down-regulated, while TIMP3, TIMP4 and LOXL2 were significantly up-regulated in postmenopausal control women when compared to premenopausal controls (p = 0.005, p = 0.01 and p < 0.001, correspondingly). TIMP4 and LOXL2 mRNA levels were significantly decreased in postmenopausal POP patients compared to asymptomatic postmenopausal controls (p < 0.01 for both). CONCLUSIONS: Our results indicate that ovarian cycle and age-related changes influence the expression of genes encoding proteins responsible for ECM metabolism in human vagina. Moreover, POP is associated with alteration in vaginal ECM components after menopause.

Chromatin supraorganization, mitotic abnormalities and proliferation in cells with increased or down-regulated lox expression: Indirect evidence of a LOX-histone H1 interaction in vivo.

Lysyl oxidases (LOXs) are enzymes that permit the covalent crosslinking of the component chains of collagen and elastin. These enzymes are present inside the nuclei of certain mammalian cells. Previous studies have proposed LOX binding to histone H1 in vitro, and histone H1 is known to control global chromatin compaction and mitotic chromosome architecture. Therefore, in the present study, we analyzed chromatin supraorganizational changes, mitotic abnormalities, mitotic indices and cell death ratios in COS-7 and NRK-49F cells with high and low lox expression levels, respectively. The objective was to support biochemical data of LOX-H1 interaction, by providing evidence of chromatin remodeling in vivo, under different lox expressions. Chromatin decondensation assessed by image analysis was observed in COS-7 cells with increased lox expression. This decondensation is suggested to be promoted by LOX actions on histone H1, which loosens the DNA-H1 complex. In NRK-49F cells transfected with antisense lox or subjected to treatment with beta-aminopropionitrile (BAPN), chromatin condensation and nuclear phenotypic variability were found, which may be due to reduced LOX-H1 interaction. When lox expression was increased in COS-7 cells, the frequency of irregular chromosome plates was not affected, but cell proliferation decreased and "cell death preceded by multinucleation" increased. In NRK-49F cells there was accelerated proliferation induced by transfection with the antisense lox, and confirmed when cells were treated with BAPN. Apoptosis increased in NRK-49F cells only with BAPN treatment whereas cell death preceded by multinucleation increased only after antisense lox transfection. The data presented herein regarding chromatin remodeling indirectly support the hypothesis that LOX binds to histone H1 in vivo. Cell proliferation in COS-7 and NRK-49F cells and cell death at least in COS-7 cells agree with predicted effects of LOX interference in these processes.