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Background: Even today, a non-invasive biomarker to identify donors with enhanced risk for renal impairment is missing. Dickkopf 3 (DKK3) can cause tubulointerstitial fibrosis and is associated with rapid eGFR loss. The aim of our work was to analyze whether DKK3 can be used as a non-invasive alert marker for an increased risk of loss of kidney function in living kidney donors (LKDs). Methods: All donors who were examined between July 2022 and June 2023 (n = 117) were included. DKK3 was measured in the urine. The collected patient-related data were compared with parameters before donation. The study cohort was stratified by DKK3 values (

Abstract Background Urinary Dickkopf 3 (DKK3) excretion is a recently established biomarker of renal functional development. Its excretion into the peritoneal cavity has not been reported. We here studied DKK3 in peritoneal dialysis. Methods DKK3 was assessed in serum, urine and dialysate in a prevalent adult peritoneal dialysis cohort and its concentration analyzed in relation to creatinine and clinical characteristics. Results Highest DKK3 concentrations were found in serum, followed by urine. Dialysate concentrations were significantly lower. Dialysate DKK3 correlated with both other compartments. Serum, dialysate and urine values were stable during three months of follow-up. Continuous ambulatory dialysis (CAPD) but not cycler-assisted peritoneal dialysis (CCPD) volume-dependently increased peritoneal DKK3 in relation to creatinine. RAAS blockade significantly decreased urinary, but not serum or peritoneal DKK3. Conclusion Our data provide a detailed characterization of DKK3 in peritoneal dialysis. They support the notion that the RAAS system is essential for renal DKK3 handling.

Background: Urinary Dickkopf 3 (DKK3) excretion is a recently established biomarker of renal functional development. Its excretion into the peritoneal cavity has not been reported. We here studied DKK3 in peritoneal dialysis. Methods: DKK3 was assessed in serum, urine and dialysate in a prevalent adult peritoneal dialysis cohort and its concentration analyzed in relation to creatinine and clinical characteristics. Results: Highest DKK3 concentrations were found in serum, followed by urine. Dialysate concentrations were significantly lower. Dialysate DKK3 correlated with both other compartments. Serum, dialysate and urine values were stable during three months of follow-up. Continuous ambulatory dialysis (CAPD) but not cycler-assisted peritoneal dialysis (CCPD) volume-dependently increased peritoneal DKK3 in relation to creatinine. RAAS blockade significantly decreased urinary, but not serum or peritoneal DKK3. Conclusion: Our data provide a detailed characterization of DKK3 in peritoneal dialysis. They support the notion that the RAAS system is essential for renal DKK3 handling. [ABSTRACT FROM AUTHOR]

Retinoblastoma (RB) is a pediatric malignancy, typically diagnosed at birth or during early childhood. The pathogenesis of RB is marked by the amplification of the Basic Helix–Loop–Helix (BHLH) Transcription Factor MYCN, which serves as a transcriptional regulator capable of binding to Dickkopf 3 (DKK3). However, the precise role of DKK3 in the malignant progression of RB cells caused by MYCN remains elusive. In the present study, the expression of MYCN was either overexpressed or interfered in RB cells. Subsequently, the expression level of DKK3 was assessed through quantitative real‐time polymerase chain reaction and western blot analysis. Cell proliferation was evaluated using the Cell Counting Kit‐8 assay and 5‐ethynyl‐2′‐deoxyuridine staining, while cell cycle progression and apoptosis were analyzed by flow cytometry and western blot analysis, respectively. Additionally, the expression of proteins involved in the Wnt/β‐catenin/Fra‐1/p53 signaling pathway was evaluated via western blot analysis. To gain further insights, Wnt agonists and the P53 inhibitor PFT‐α were introduced into exploration. The current investigation revealed a negative correlation between the expression levels of MYCN and DKK3 in RB cells. Additionally, DKK3 overexpression inhibited cell proliferation, promoted cell apoptosis, and arrested cell cycle in RB cells with high expression of MYCN. Moreover, enhanced DKK3 expression inhibited proliferation, promoted cell cycle arrest and apoptosis of RB cells by modulating the wnt/βcatenin/Fra‐1/p53 signaling pathway. Furthermore, in vivo experiments revealed that overexpression of DKK3 inhibits the growth of RB tumors. Collectively, our findings elucidate that MYCN stimulates the Wnt/β‐catenin/Fra‐1 pathway by suppressing DKK3 expression, ultimately suppressing p53 activity and contributing to malignant progression of RB. [ABSTRACT FROM AUTHOR]

Sarcopenia, characterized by the progressive loss of muscle mass and function, significantly affects patients with chronic obstructive pulmonary disease (COPD) and worsens their morbidity and mortality. The pathogenesis of muscle atrophy in patients with COPD involves complex mechanisms, including protein imbalance and mitochondrial dysfunction, which have been identified in the muscle tissues of patients with COPD. DKK3 (Dickkopf-3) is a secreted glycoprotein involved in the process of myogenesis. However, the role of DKK3 in the regulation of muscle mass is largely unknown. This study investigated the role of DKK3 in COPD-related sarcopenia. DKK3 was found to be overexpressed in cigarette smoking-induced muscle atrophy and in patients with COPD. Importantly, plasma DKK3 levels in COPD patients with sarcopenia were significantly higher than those without sarcopenia, and plasma DKK3 levels could effectively predict sarcopenia in patients with COPD based on two independent cohorts. Mechanistically, DKK3 is secreted by skeletal muscle cells that acts in autocrine and paracrine manners and interacts with the cell surface-activated receptor cytoskeleton-associated protein 4 (CKAP4) to induce mitochondrial dysfunction and myotube atrophy. The inhibition of DKK3 by genetic ablation prevented cigarette smoking‐induced skeletal muscle dysfunction. These results suggest that DKK3 is a potential target for the diagnosis and treatment of sarcopenia in patients with COPD.

ABSTRACT: Mesenchymal stem cells can differentiate into adipocyte precursor cells, and the balance of stem cell differentiation determines the quantity of adipocytes. Early-stage adipose tissue expression profiling revealed abnormal expression of DKK3 in the high-fat group. Moreover, DKK3 is enriched in the Wnt/β-catenin signaling pathway, and studies have shown that DKK3 can serve as a gene involved in early regulation of adipogenesis. Therefore, this study focuses on exploring how DKK3 regulates the molecular mechanism of adipocyte differentiation through the Wnt/β-catenin signaling pathway. In this experiment, the role of DKK3 in the differentiation of bone marrow mesenchymal stem cells into adipocyte precursors was validated using in vitro cultured chicken bone marrow mesenchymal stem cells. The results showed that overexpression of DKK3 led to a significant downregulation of Wnt/β-catenin signaling pathway-related marker gene expression (P < 0.05), a significant upregulation of adipogenic differentiation-related genes (P < 0.05), an increase in lipid droplet content, a significant increase in OD value (P < 0.05), a significant upregulation of mitochondrial oxidative respiratory-related marker gene expression (P < 0.05), and a significant downregulation of mitochondrial autophagy-related marker genes (P < 0.05). Conversely, the results were opposite after interfering with DKK3 gene expression. In this study, 4 SNP sites, including g.8419139, g.8419556, g.8419560, and g.8419598, were detected in the 7th exon of DKK3, among which the g.8419598 (C > T) site was significantly correlated with abdominal fat weight and abdominal fat rate in 100-day-old Ma Huang chickens (P < 0.001).

Background Chronic kidney disease (CKD) lacks effective treatments and renal fibrosis (RF) is one of CKD’s outcomes. Dickkopf 3 (DKK3) has been identified as an agonist in CKD. However, the underlying mechanisms of DKK3 in CKD are not fully understood.Methods H2O2-treated HK-2 cells and ureteric obstruction (UUO) mice were used as RF models. Biomarkers, Masson staining, PAS staining, and TUNEL were used to assess kidney function and apoptosis. Oxidative stress and mitochondria function were also evaluated. CCK-8 and flow cytometry were utilized to assess cell viability and apoptosis. Western blotting, IHC, and qRT-PCR were performed to detect molecular expression levels. Immunofluorescence was applied to determine the subcellular localization. Dual luciferase assay, MeRIP, RIP, and ChIP were used to validate the m6A level and the molecule interaction.Results DKK3 was upregulated in UUO mouse kidney tissue and H2O2-treated HK-2 cells. Knockdown of DKK3 inhibited oxidative stress, maintained mitochondrial homeostasis, and alleviated kidney damage and RF in UUO mice. Furthermore, DKK3 silencing suppressed HK-2 cell apoptosis, oxidative stress, and mitochondria fission. Mechanistically, DKK3 upregulation was related to the high m6A level regulated by METTL3. DKK3 activated TCF4/β-catenin and enhanced MFF transcriptional expression by binding to its promoter. Overexpression of MFF reversed in the inhibitory effect of DKK3 knockdown on cell damage.Conclusion Upregulation of DKK3 caused by m6A modification activated the Wnt/β-catenin pathway to increase MFF transcriptional expression, leading to mitochondrial dysfunction and oxidative stress, thereby promoting RF progression.

Background: Endothelial barrier dysfunction is critical for the pathogenesis of sepsis-induced acute lung injury (ALI). Lipopolysaccharide (LPS)-stimulated human pulmonary microvascular endothelial cells (HPMECs) are widely used as the cell model of sepsis-associated ALI for exploration of endothelial barrier dysfunction. Dickkopf (DKK) family proteins were reported to mediate endothelial functions in various diseases. The present study explored the effect of Dickkopf-3 (DKK3) on endothelial barrier permeability, angiogenesis, and tight junctions in LPS-stimulated HPMECs. Methods: RT-qPCR was required for detecting DKK3 and miR-98-3p expression. The angiogenesis of HPMECs was evaluated by tube formation assays. Monolayer permeability of HPMECs was examined by Transwell rhodamine assays. The protein expression of DKK3 and tight junctions in HPMECs was measured via western blotting. Luciferase reporter assay was used to verify the interaction between miR-98-3p and DKK3. Results: LPS treatment inhibited angiogenetic ability while increasing the permeability of HPMECs. DKK3 expression was upregulated while miR-98-3p level was reduced in LPS-treated HPMECs. DKK3 knock-down alleviated HPMEC injury triggered by LPS stimulation. MiR-98-3p targeted DKK3 in HPMECs. Overexpression of miR-98-3p protects HPMECs from the LPS-induced endothelial barrier dysfunction, and the protective effect was reversed by DKK3 overexpression. Conclusions: MiR-98-3p ameliorates LPS-evoked pulmonary microvascular endothelial barrier dysfunction in sepsis-associated ALI by targeting DKK3. [ABSTRACT FROM AUTHOR]

Oxidative stress and fibrosis may accelerate the progression of chronic kidney disease (CKD). DKK3 is related to regulating renal fibrosis and CKD. However, the molecular mechanism of DKK3 in regulating oxidative stress and fibrosis during CKD development has not been clarified, which deserves to be investigated. Human proximal tubule epithelial cells (HK-2 cells) were treated with H2O2 to establish a cell model of renal fibrosis. The mRNA and protein expressions were analyzed using qRT-PCR and western blot, respectively. Cell viability and apoptosis were evaluated using MTT assay and flow cytometry, respectively. ROS production was estimated using DCFH-DA. The interactions among TCF4, β-catenin and NOX4 were validated using luciferase activity assay, ChIP and Co-IP. Herein, our results revealed that DKK3 was highly expressed in HK-2 cells treated with H2O2. DKK3 depletion increased H2O2-treated HK-2 cell viability and reduced cell apoptosis, oxidative stress, and fibrosis. Mechanically, DKK3 promoted formation of the β-catenin/TCF4 complex, and activated NOX4 transcription. Upregulation of NOX4 or TCF4 weakened the inhibitory effect of DKK3 knockdown on oxidative stress and fibrosis in H2O2-stimulated HK-2 cells. All our results suggested that DKK3 accelerated oxidative stress and fibrosis through promoting β-catenin/TCF4 complex-mediated activation of NOX4 transcription, which could lead to novel molecules and therapeutic targets for CKD. [ABSTRACT FROM AUTHOR]

Glioblastoma is the most common malignant primary tumor of the CNS. The prognosis is dismal, with a median survival of 15 months. Surgical treatment followed by adjuvant therapies such as radiotherapy and chemotherapy characterize the classical strategy. The WNT pathway plays a key role in cellular proliferation, differentiation, and invasion. The DKK3 protein, capable of acting as a tumor suppressor, also appears to be able to modulate the WNT pathway. We performed, in a series of 40 patients, immunohistochemical and Western blot evaluations of DKK3 to better understand how the expression of this protein can influence clinical behavior. We used a statistical analysis, with correlations between the expression of DKK3 and overall survival, age, sex, Ki-67, p53, and MGMT and IDH status. We also correlated our data with information included in the cBioPortal database. In our analyses, DKK3 expression, in both immunohistochemistry and Western blot analyses, was reduced or absent in many cases, showing downregulation. To date, no clinical study exists in the literature that reports a potential correlation between IDH and MGMT status and the WNT pathway through the expression of DKK3. Modulation of this pathway through the expression of DKK3 could represent a new tailored therapeutic strategy in the treatment of glioblastoma. [ABSTRACT FROM AUTHOR]

Backgound Autosomal dominant polycystic kidney disease (ADPKD) is the most common inherited kidney disease, and leads to a steady loss of kidney function in adulthood. The variable course of the disease makes it necessary to identify the patients with rapid disease progression who will benefit the most from targeted therapies and interventions. Currently, magnetic resonance imaging–based volumetry of the kidney is the most commonly used tool for this purpose. Biomarkers that can be easily and quantitatively determined, which allow a prediction of the loss of kidney function, have not yet been established in clinical practice. The glycoprotein Dickkopf 3 (DKK3) which is secreted in the renal tubular epithelium upon stress and contributes to tubulointerstitial fibrosis via the Wnt signaling pathway, was recently described as a biomarker for estimating risk of kidney function loss, but has not been investigated for ADPKD. This study aimed to obtain a first insight into whether DKK3 may indeed improve outcome prediction in ADPKD in the future. Methods In 184 ADPKD patients from the AD(H)PKD registry and 47 healthy controls, the urinary DKK3 (uDKK3) levels were determined using ELISA. Multiple linear regression was used to examine the potential of these values in outcome prediction. Results ADPKD patients showed significantly higher uDKK3 values compared with the controls (mean 1970 ± 5287 vs 112 ± 134.7 pg/mg creatinine). Furthermore, there was a steady increase in uDKK3 with an increase in the Mayo class (A/B 1262 ± 2315 vs class D/E 3104 ± 7627 pg/mg creatinine), the best-established biomarker of progression in ADPKD. uDKK3 also correlated with estimated glomerular filtration rate (eGFR). Patients with PKD1 mutations show higher uDKK3 levels compared with PKD2 patients (PKD1 : 2304 ± 5119; PKD2 : 506.6 ± 526.8 pg/mg creatinine). Univariate linear regression showed uDKK3 as a significant predictor of future eGFR slope estimation. In multiple linear regression this effect was not significant in models also containing height-adjusted total kidney volume and/or eGFR. However, adding both copeptin levels and the interaction term between copeptin and uDKK3 to the model resulted in a significant predictive value of all these three variables and the highest R2 of all models examined (∼0.5). Conclusion uDKK3 shows a clear correlation with the Mayo classification in patients with ADPKD. uDKK3 levels correlated with kidney function, which could indicate that uDKK3 also predicts a disproportionate loss of renal function in this collective. Interestingly, we found an interaction between copeptin and uDKK3 in our prediction models and the best model containing both variables and their interaction term resulted in a fairly good explanation of variance in eGFR slope compared with previous models. Considering the limited number of patients in these analyses, future studies will be required to confirm the results. Nonetheless, uDKK3 appears to be an attractive candidate to improve outcome prediction of ADPKD in the future. [ABSTRACT FROM AUTHOR]

Objective: We previously reported that dickkopf WNT signaling inhibitor 3 (DKK3) would modulate malignant potential of oral squamous cell carcinoma (OSCC) via activating Akt. Recently, cytoskeleton associated protein 4 (CKAP4) functions as receptor of DKK3, which activates Akt in esophageal squamous cell carcinoma, but its expression and function in OSCC were unclear. Methods: We studied DKK3 and CKAP4 protein expression in OSCC tissue and investigated the correlation between protein expression and clinical data. We also investigated whether antibodies (Ab) for DKK3 or CKAP4 could suppress malignant potential of the cancer cells. Results: DKK3/CKAP4 protein expression was observed in majority of OSCC cases and was associated with significantly higher T‐stage and TNM stage. Multivariate analysis revealed that DKK3 and CKAP4 were independent prognostic biomarkers for overall survival (OS) and disease‐free survival (DFS), respectively. Survival analyses revealed that DKK3‐positive cases and CKAP4‐positive cases showed significantly shorter OS and DFS, respectively, and that DKK3/CKAP4 double‐negative cases showed significantly favorable prognosis. Both anti‐DKK3Ab and anti‐CKAP4Ab could suppress cancer cell proliferation, migration, and invasion. Conclusion: DKK3/CKAP4 axis is thought to be important in OSCC, and it would be a promising therapeutic target. [ABSTRACT FROM AUTHOR]

Excessive inflammatory injury is the main cause of the incidence of severe neonatal pneumonia (NP) and associated deaths. Although dickkopf-3 (DKK3) exhibits anti-inflammatory activity in numerous pathological processes, its role in NP is still unknown. In this study, human embryonic lung WI-38 and MRC-5 cells were treated with lipopolysaccharide (LPS) to induce inflammatory injury of NP in vitro. The expression of DKK3 was downregulated in LPS-stimulated WI-38 and MRC-5 cells. DKK3 overexpression decreased LPS-induced inhibition of cell viability, and reduced LPS-induced apoptosis of WI-38 and MRC-5 cells. DKK3 overexpression also reduced LPS-induced production of pro-inflammatory factors such as ROS, IL-6, MCP-1, and TNF-α. Nuclear respiratory factors 1 (NRF1) knockdown was found to upregulate DKK3 and inactivate the GSK-3β/β-catenin pathway in LPS-injured WI-38 and MRC-5 cells. NRF1 knockdown also suppressed LPS-induced inhibition on cell viability, repressed LPS-induced apoptosis, and inhibited the accumulation of ROS, IL-6, MCP-1, and TNF-α in LPS-injured WI-38 and MRC-5 cells. DKK3 knockdown or re-activation of the GSK-3β/β-catenin pathway reversed the inhibitory effects of NRF1 knockdown on LPS-induced inflammatory injury. In conclusion, NRF1 knockdown can alleviate LPS-triggered inflammatory injury by regulating DKK3 and the GSK-3β/β-catenin pathway. DKK3 is decreased in LPS-challenged WI-38 cells. DKK3 ameliorates LPS-evoked inflammatory injury in WI-38 cells. NRF1 downregulates DKK3 to activate the GSK-3β/β-catenin pathway. NRF1 knockdown alleviates LPS-evoked inflammatory injury via DKK3. [ABSTRACT FROM AUTHOR]

Background: Dickkopf-3 (DKK-3) has garnered interest as a potential biomarker for diagnosing and monitoring kidney and cardiovascular diseases (CVDs). Herein, we aimed to evaluate the level of DKK-3 in individuals with chronic kidney disease (CKD) and detect its correlation with CVD. Patients and methods: The study involved 68 cases categorized into four groups. The study protocol involved conducting a comprehensive medical history assessment, a standard physical examination, laboratory investigations, and an enzyme-linked immunosorbent assay (ELISA) to quantify the serum DKK3 levels. Results: Our findings indicated a significant elevation in serum DKK-3 levels among CVDs patients who suffer from renal impairme nt and those with end-stage renal disease who were on dialysis. Additionally, our outcomes revealed a significant positive correlation between DKK-3 and various biomarkers levels, including SGPT (r = 0.336, p = 0.005), parathyroid hormone (r = 0.425, p < 0.001), creatinine (r = 0.62, p < 0.001), total cholesterol (r = 0.312, p = 0.01), low-density lipoprotein cholesterol (LDL-C) (r = 0.268, p = 0.027), and triglycerides (r = 0.256, p = 0.035). Our results indicate a significant negative correlation between DKK-3 levels and both eGFR (r = -0.507, p < 0.001) and hemoglobin levels (r = -0.33, p = 0.006). Additionally, an independent association was observed between the serum levels of DKK-3 and creatinine, LDL-C, and LVEDD. Conclusion: DKK-3 may serve as a novel biomarker for the progression of CKD. Moreover, DKK3 can potentially function as a biomarker for CVD among individuals with renal disease. [ABSTRACT FROM AUTHOR]

Malgré une grande hétérogénéité des situations cliniques, la définition de l'insuffisance rénale aiguë (IRA) est unique et son diagnostic repose essentiellement sur l'augmentation de la créatininémie et/ou la diminution de la diurèse. À l'inverse de l'évolution sur les marqueurs biologiques de l'infarctus du myocarde observée depuis plus de 50 ans, c'est toujours la créatinine "marqueur fonctionnel" qui sert de gold standard biologique pour le diagnostic de l'IRA. Cependant, sous le terme d'IRA, sont regroupées des situations cliniques très hétérogènes d'un point de vue étioIogique, physiopathologique, de durée, d'évolution et d'éventuelles complications associées. Depuis une quinzaine d'années des nouveaux biomarqueurs potentiels sont étudiés et semblent prometteurs. Contrairement à la troponine qui est augmentée dans tous les types d'infarctus du myocarde, ces nouveaux biomarqueurs présentent des sensibilités et spécificités différentes en fonction du type d'IRA, ce qui explique possiblement leur sous-utilisation en pratique courante. Despite a great heterogeneity of clinical situations, the definition of acute kidney injury (AKI) is unique and its diagnosis includes an increase in serum creatinine and/or a decrease in diuresis. Contrary to the evolution of the biological markers of myocardial infarction observed for more than 50 years, it is always the "functional marker" creatinine which serves as the biological gold standard for the diagnosis of AKI. However, under the term of acute renal failure, very heterogeneous clinical situations are grouped together from an etiological, pathophysiological mechanisms, duration, evolution and associated complications. For fifteen/twenty years, new potential biomarkers have been studied and seem promising. Unlike troponin, which is increased in all types of myocardial infarction, these new biomarkers have different sensitivities and specificities depending on the type of AKI, which possibly explains their under-use in current practice. [ABSTRACT FROM AUTHOR]

Multiciliated cells contain hundreds of cilia whose directional movement powers the mucociliary clearance of the airways, a vital host defense mechanism. Multiciliated cell specification requires canonical Wnt signaling, which then must be turned off. Next, ciliogenesis and polarized ciliary orientation are regulated by noncanonical Wnt/planar cell polarity (Wnt/PCP) signaling. The mechanistic relationship between the Wnt pathways is unknown. We show that DKK3, a secreted canonical Wnt regulator and WNT4, a noncanonical Wnt ligand act together to facilitate a canonical to noncanonical Wnt signaling switch during multiciliated cell formation. In primary human airway epithelial cells, DKK3 and WNT4 CRISPR knockout blocks, whereas ectopic expression promotes, multiciliated cell formation by inhibiting canonical Wnt signaling. Wnt4 and Dkk3 single-knockout mice also display defective ciliated cells. DKK3 and WNT4 are co-secreted from basal stem cells and act directly on multiciliated cells via KREMEN1 and FZD6, respectively. We provide a novel mechanism that links specification to cilium biogenesis and polarization for proper multiciliated cell formation. [ABSTRACT FROM AUTHOR]

Abstract Background Head and neck squamous cell carcinoma (HNSCC) is the most common malignant tumor of the head and neck. We identified cancer-specific genes in HNSCC and focused on DKK3 expression. DKK3 gene codes two isoforms of proteins (secreted and non-secreted) with two distinct cysteine rich domains (CRDs). It is reported that DKK3 functions as a negative regulator of oncogenic Wnt signaling and, is therefore, considered to be a tumor suppressor gene. However, our series of studies have demonstrated that DKK3 expression is specifically high in HNSCC tissues and cells, and that DKK3 might determine the malignant potentials of HNSCC cells via the activation of Akt. Further analyses strongly suggested that both secreted DKK3 and non-secreted DKK3 could activate Akt signaling in discrete ways, and consequently exert tumor promoting effects. We hypothesized that DKK3 might be a specific druggable target, and it is necessary to establish a DKK3 inhibitor that can inhibit both secreted and non-secreted isoforms of DKK3. Methods Using inverse polymerase chain reaction, we generated mutant expression plasmids that express DKK3 without CRD1, CRD2, or both CRD1 and CRD2 (DKK3ΔC1, DKK3ΔC2, and DKK3ΔC1ΔC2, respectively). These plasmids were then transfected into HNSCC-derived cells to determine the domain responsible for DKK3-mediated Akt activation. We designed antisense peptides using the MIMETEC program, targeting DKK3-specific amino acid sequences within CRD1 and CRD2. The structural models for peptides and DKK3 were generated using Raptor X, and then a docking simulation was performed using CluPro2. Afterward, the best set of the peptides was applied into HNSCC-derived cells, and the effects on Akt phosphorylation, cellular proliferation, invasion, and migration were assessed. We also investigated the therapeutic effects of the peptides in the xenograft models. Results Transfection of mutant expression plasmids and subsequent functional analyses revealed that it is necessary to delete both CRD1 and CRD2 to inhibit Akt activation and inhibition of proliferation, migration, and invasion. The inhibitory peptides for CRD1 and CRD2 of DKK3 significantly reduced the phosphorylation of Akt, and consequently suppressed cellular proliferation, migration, invasion and in vivo tumor growth at very low doses. Conclusions This inhibitory peptide represents a promising new therapeutic strategy for HNSCC treatment.

Abstract Background Extracellular vesicles (EVs) could mediate embryo-maternal communication to affect embryo implantation by delivering biology information, including microRNA (miRNA), protein, lipid. Our previous research shows that miR-92b-3p was differentially expressed in EVs of uterine flushing fluids during the embryo implantation period. However, the role of miR-92b-3p from EVs in embryo implantation remains elusive. Materials and methods EVs were isolated from porcine endometrial epithelial cells (EECs) by ultracentrifugation. MiR-92b-3p mimics and EVs were used to regulate the expression of miR-92b-3p in porcine trophoblast cells (PTr2 cells). Cell proliferation, migration and adhesion analyses were used to observe the phenotype. RT-qPCR, western blot and dual-luciferase reporter assay were used to assess the targets of miR-92b-3p. Results In this study, EVs derived from porcine EECs were identified and could be taken up by PTr2 cells. We found that the EVs derived from EECs transfected with miR-92b-3p mimic (EVs-miR-92b-3p) significantly promoted the proliferation, migration and adhesion of PTr2 cells. We verified that Tuberous sclerosis complex subunit (TSC1) and Dickkopf 3 (DKK3) were the target genes of miR-92b-3p. Moreover, our study showed that miR-92b-3p plays a vital role in PTr2 cells via targeting TSC1 and DKK3. Furthermore, the 3'UTR vectors of TSC1 and DKK3 can rescue the effect of miR-92b-3p on PTr2 cells. Conclusions Taken together, this study reveals a novel mechanism that EVs derived from porcine EECs treated with miR-92b-3p crosstalk with trophoblasts by targeting TSC1 and DKK3, leading to an enhanced ability for implantation.

Dickkopf-3 (Dkk3) is an atypical member of the Dkk family of Wnt inhibitors, which has been implicated in the pathophysiology of neurodegenerative disorders. Its role in the mechanisms of cellular degeneration and protection is still unknown. The aim of our work is to investigate the endogenous activation of the DKK3 pathway in a model of transient occlusion of the middle cerebral artery in rats. In particular, the animals were subjected to 1 h of ischemia followed by different reperfusion times (1 h, 6 h, 12 h and 24 h) to evaluate the downstream pathway and the time course of its activation. Western blot analysis showed increased Dkk3 expression in animals with the highest time of reperfusion. The increased levels of Dkk3 were accompanied by reduced Wnt3a, Frz1 and PIWI1a expression in the cytosol while FOXM1 and β-catenin decreased in the nucleus. These molecular changes led to an increase in the apoptotic pathway, as showed by the increased expression of Caspase 3 and Bax and the reduced levels of Bcl-2, and to a decrease in neurogenesis, as shown by the decreased expression of Tbr2, Ngn2 and Pax6. In the second part of the study, we decided to employ curcumin, an activator of the Wnt/β-catenin signaling, to investigate its effect on Dkk3. In particular, curcumin was administered 1 and 6 h after ischemia, and animals were sacrificed 24 h later when the expression of Dkk3 was higher. Our data displayed that curcumin administration decreased Dkk3 expression, and increased Wnt3a, Frz1 and PIWI1a levels. Well in line with these data, curcumin administration increased nuclear β-catenin and FOXM1 expression. The down-regulation of Dkk3 by curcumin led to reduced apoptosis and increased neurogenesis. Summarizing, our results showed that Dkk3 acts as an inhibitor of Wnt/β-catenin signaling during cerebral ischemia. Additionally, its inhibition and the contextual activation of the Wnt/β-catenin pathway are protective against ischemic stroke. [ABSTRACT FROM AUTHOR]

Introduction: Colorectal cancer (CRC) is the third most common cancer in the world with high mortality, hence, understanding the molecular mechanisms involved in the tumor progression are important for CRC diagnosis and treatment. MicroRNAs (miRNAs) are key gene expression regulators that can function as tumor suppressors or oncogenes in tumor cells, and modulate angiogenesis as a critical process in tumor metastasis. MiR-1290 has been demonstrated as an onco-miRNA in various types of cancer, however, the role of miR-1290 in CRC is not fully understood. This study aimed to investigate the oncogenic and angiogenic potential of miR-1290 in CRC. Methods: Lenti-miR-1290 was transduced into HCT116, SW480, and human umbilical vein endothelial cells (HUVECs). By bioinformatics analysis, we identified thrombospondin 1 (THBS1) as a novel predicted target for miR-1290. Quantitative real-time PCR, western blotting, and luciferase reporter assay were used to demonstrate suppression of miR-1290 target genes including THBS1, Dickkopf Wnt signaling pathway inhibitor 3 (DKK3), and suppressor of cancer cell invasion (SCAI) in HCT116 and HUVECs. Cell cycle analysis, proliferation, migration and, tube formation were determined by flow cytometry, MTT, wound healing, and tube formation assays, respectively. Results: MiR-1290 significantly decreased the expression of THBS1, DKK3, and SCAI. We demonstrated that miR-1290 enhanced proliferation, migration, and angiogenesis partially through suppression of THBS1, DKK3, and SCAI in CRC. Conclusion: These results suggest a novel function of miR-1290 which may contribute to tumorigenesis and angiogenesis in CRC.

Abstract Background Neuropathic pain is a common and severely disabling state that affects millions of people worldwide. Microglial activation in the spinal cord plays a critical role in the pathogenesis of neuropathic pain. However, the mechanisms underlying spinal microglial activation during neuropathic pain remain incompletely understood. Here, we investigated the role of Dickkopf (DKK) 3 and its interplay with microglial activation in the spinal cord in neuropathic pain. Methods In this study, we investigated the effects of intrathecal injection of recombinant DKK3 (rDKK3) on mechanical allodynia and microglial activation in the spinal cord after spared nerve injury (SNI) in rats by western blot (WB), immunofluorescence (IF), quantitative polymerase chain reaction (qPCR), and enzyme-linked immunosorbent assay (ELISA). Results We found that SNI induced a significant decrease in the levels of DKK3, Kremen-1 and Dishevelled-1 (DVL-1) and up-regulated the expression of phosphorylated apoptosis signal-regulating kinase 1 (p-ASK1), phosphorylated c-JUN N-terminal kinase (p-JNK), phosphorylated p38 (p-p38) in the spinal cord. Moreover, our results showed that exogenous intrathecal administration of rDKK3 inhibited expression of p-ASK1, p-JNK, p-p38, promoted the transformation of microglia from M1 type to M2 type, and decreased the production of pro-inflammatory cytokines compared to the rats of SNI + Vehicle. However, these effects were reversed by intrathecal administration of Kremen-1 siRNA or Dishevelled-1 (DVL-1) siRNA. Conclusions These results suggest that DKK3 ameliorates neuropathic pain via inhibiting ASK-1/JNK/p-38-mediated microglia polarization and neuroinflammation, at least partly, by the Kremen-1 and DVL-1 pathways.

Diabetic nephropathy (DN) is an important cause of death in diabetes patients, which is mainly due to its complex pathogenesis. Here, we explored the role of N6-methyladenosine (m6A) RNA methylation in DN development. Renal tubular epithelial cells from DN patients and experimental DN mice treated with streptozotocin (STZ) exhibited a considerable increase in METTL14 and WTAP expression as well as overall m6A methylation. Knocking down the expression of METTL14 and WTAP inhibited the migration and proliferation of tubular epithelial cells. MeRIP-seq analysis of the renal tissues of DN patients revealed that the genes with elevated m6A methylation were concentrated in the Wnt/β-Catenin signaling pathway. Dickkopf homolog 3 (DKK3) was screened out as the gene with the most significant increase in m6A methylation. In addition, the expression change pattern of DKK3 under DN circumstances is in line with those of METTL14 and WTAP. DKK3's m6A methylation sites were confirmed to be located in the 3′UTR region, which is how METTL14 and WTAP improved DKK3's mRNA stability. Finally, YTHDF1, a m6A reader, was demonstrated to recognize m6A-methylated DKK3 and promote DKK3 expression. • Total m6A level in the kidney is increased under diabetic nephropathy. • METTL14 and WTAP, act as m6A writer, are upregulated in tubular cells under DN. • m6A methylated DKK3 is upregulated under DN. • Urinary DKK3 is a biomarker of DN. • YTHDF1 is a reader protein of methylated DKK3 mRNA. [ABSTRACT FROM AUTHOR]

While it is considered one of the most common cancers and the leading cause of death in men worldwide, prognostic stratification and treatment modalities are still limited for patients with prostate cancer (PCa). Recently, the introduction of genomic profiling and the use of new techniques like next-generation sequencing (NGS) in many cancers provide novel tools for the discovery of new molecular targets that might improve our understanding of the genomic aberrations in PCa and the discovery of novel prognostic and therapeutic targets. In this study, we investigated the possible mechanisms through which Dickkopf-3 (DKK3) produces its possible protective role in PCa using NGS in both the DKK3 overexpression PCa cell line (PC3) model and our patient cohort consisting of nine PCa and five benign prostatic hyperplasia. Interestingly, our results have shown that DKK3 transfection-modulated genes are involved in the regulation of cell motility, senescence-associated secretory phenotype (SASP), and cytokine signaling in the immune system, as well as in the regulation of adaptive immune response. Further analysis of our NGS using our in vitro model revealed the presence of 36 differentially expressed genes (DEGs) between DKK3 transfected cells and PC3 empty vector. In addition, both CP and ACE2 genes were differentially expressed not only between the transfected and empty groups but also between the transfected and Mock cells. The top common DEGs between the DKK3 overexpression cell line and our patient cohort are the following: IL32, IRAK1, RIOK1, HIST1H2BB, SNORA31, AKR1B1, ACE2, and CP. The upregulated genes including IL32, HIST1H2BB, and SNORA31 showed tumor suppressor functions in various cancers including PCa. On the other hand, both IRAK1 and RIOK1 were downregulated and involved in tumor initiation, tumor progression, poor outcome, and radiotherapy resistance. Together, our results highlighted the possible role of the DKK3-related genes in protecting against PCa initiation and progression.

Background: Head and neck squamous cell carcinoma (HNSCC) is the most common malignant tumor of the head and neck. We identified cancer-specific genes in HNSCC and focused on DKK3 expression. DKK3 gene codes two isoforms of proteins (secreted and non-secreted) with two distinct cysteine rich domains (CRDs). It is reported that DKK3 functions as a negative regulator of oncogenic Wnt signaling and, is therefore, considered to be a tumor suppressor gene. However, our series of studies have demonstrated that DKK3 expression is specifically high in HNSCC tissues and cells, and that DKK3 might determine the malignant potentials of HNSCC cells via the activation of Akt. Further analyses strongly suggested that both secreted DKK3 and non-secreted DKK3 could activate Akt signaling in discrete ways, and consequently exert tumor promoting effects. We hypothesized that DKK3 might be a specific druggable target, and it is necessary to establish a DKK3 inhibitor that can inhibit both secreted and non-secreted isoforms of DKK3. Methods: Using inverse polymerase chain reaction, we generated mutant expression plasmids that express DKK3 without CRD1, CRD2, or both CRD1 and CRD2 (DKK3ΔC1, DKK3ΔC2, and DKK3ΔC1ΔC2, respectively). These plasmids were then transfected into HNSCC-derived cells to determine the domain responsible for DKK3-mediated Akt activation. We designed antisense peptides using the MIMETEC program, targeting DKK3-specific amino acid sequences within CRD1 and CRD2. The structural models for peptides and DKK3 were generated using Raptor X, and then a docking simulation was performed using CluPro2. Afterward, the best set of the peptides was applied into HNSCC-derived cells, and the effects on Akt phosphorylation, cellular proliferation, invasion, and migration were assessed. We also investigated the therapeutic effects of the peptides in the xenograft models. Results: Transfection of mutant expression plasmids and subsequent functional analyses revealed that it is necessary to delete both CRD1 and CRD2 to inhibit Akt activation and inhibition of proliferation, migration, and invasion. The inhibitory peptides for CRD1 and CRD2 of DKK3 significantly reduced the phosphorylation of Akt, and consequently suppressed cellular proliferation, migration, invasion and in vivo tumor growth at very low doses. Conclusions: This inhibitory peptide represents a promising new therapeutic strategy for HNSCC treatment. [ABSTRACT FROM AUTHOR]

Sarcopenia, characterized by the progressive loss of muscle mass and function, significantly affects patients with chronic obstructive pulmonary disease (COPD) and worsens their morbidity and mortality. The pathogenesis of muscle atrophy in patients with COPD involves complex mechanisms, including protein imbalance and mitochondrial dysfunction, which have been identified in the muscle tissues of patients with COPD. DKK3 (Dickkopf-3) is a secreted glycoprotein involved in the process of myogenesis. However, the role of DKK3 in the regulation of muscle mass is largely unknown. This study investigated the role of DKK3 in COPD-related sarcopenia. DKK3 was found to be overexpressed in cigarette smoking-induced muscle atrophy and in patients with COPD. Importantly, plasma DKK3 levels in COPD patients with sarcopenia were significantly higher than those without sarcopenia, and plasma DKK3 levels could effectively predict sarcopenia in patients with COPD based on two independent cohorts. Mechanistically, DKK3 is secreted by skeletal muscle cells that acts in autocrine and paracrine manners and interacts with the cell surface-activated receptor cytoskeleton-associated protein 4 (CKAP4) to induce mitochondrial dysfunction and myotube atrophy. The inhibition of DKK3 by genetic ablation prevented cigarette smoking-induced skeletal muscle dysfunction. These results suggest that DKK3 is a potential target for the diagnosis and treatment of sarcopenia in patients with COPD., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Chronic kidney disease (CKD) lacks effective treatments and renal fibrosis (RF) is one of CKD's outcomes. Dickkopf 3 (DKK3) has been identified as an agonist in CKD. However, the underlying mechanisms of DKK3 in CKD are not fully understood., Methods: H 2 O 2 -treated HK-2 cells and ureteric obstruction (UUO) mice were used as RF models. Biomarkers, Masson staining, PAS staining, and TUNEL were used to assess kidney function and apoptosis. Oxidative stress and mitochondria function were also evaluated. CCK-8 and flow cytometry were utilized to assess cell viability and apoptosis. Western blotting, IHC, and qRT-PCR were performed to detect molecular expression levels. Immunofluorescence was applied to determine the subcellular localization. Dual luciferase assay, MeRIP, RIP, and ChIP were used to validate the m6A level and the molecule interaction., Results: DKK3 was upregulated in UUO mouse kidney tissue and H 2 O 2 -treated HK-2 cells. Knockdown of DKK3 inhibited oxidative stress, maintained mitochondrial homeostasis, and alleviated kidney damage and RF in UUO mice. Furthermore, DKK3 silencing suppressed HK-2 cell apoptosis, oxidative stress, and mitochondria fission. Mechanistically, DKK3 upregulation was related to the high m6A level regulated by METTL3. DKK3 activated TCF4/β-catenin and enhanced MFF transcriptional expression by binding to its promoter. Overexpression of MFF reversed in the inhibitory effect of DKK3 knockdown on cell damage., Conclusion: Upregulation of DKK3 caused by m6A modification activated the Wnt/β-catenin pathway to increase MFF transcriptional expression, leading to mitochondrial dysfunction and oxidative stress, thereby promoting RF progression.

Renal ischemia-reperfusion injury (IRI) is one of the main causes of acute kidney injury (AKI). So far, there have been many studies on renal IRI, although an effective treatment method has not been developed. In recent years, growing evidence has shown that small noncoding RNAs play an important regulatory role in renal IRI. This article aims to explore whether microRNA-25-3p (miR-25-3p) plays a role in the molecular mechanism of renal IRI. The results showed that the expression level of miR-25-3p was significantly downregulated in a rat renal IRI model, and this result was confirmed with in vitro experiments. After the hypoxia-reoxygenation treatment, the apoptosis level of NRK-52E cells transfected with miR-25-3p mimics decreased significantly, and this antiapoptotic effect was antagonized by miR-25-3p inhibitors. In addition, we confirmed that DKK3 is a target of miR-25-3p. miR-25-3p exerts its protective effect against apoptosis on NRK-52E cells by inhibiting the expression of DKK3, and downregulating the expression level of miR-25-3p could disrupt this protective effect. In addition, we reconfirmed the role of miR-25-3p in rats. Therefore, we confirmed that miR-25-3p may target DKK3 to reduce renal cell damage caused by hypoxia and that miR-25-3p may be a new potential treatment for renal IRI.

Background: Extracellular vesicles (EVs) could mediate embryo-maternal communication to affect embryo implantation by delivering biology information, including microRNA (miRNA), protein, lipid. Our previous research shows that miR-92b-3p was differentially expressed in EVs of uterine flushing fluids during the embryo implantation period. However, the role of miR-92b-3p from EVs in embryo implantation remains elusive. Materials and methods: EVs were isolated from porcine endometrial epithelial cells (EECs) by ultracentrifugation. MiR-92b-3p mimics and EVs were used to regulate the expression of miR-92b-3p in porcine trophoblast cells (PTr2 cells). Cell proliferation, migration and adhesion analyses were used to observe the phenotype. RT-qPCR, western blot and dual-luciferase reporter assay were used to assess the targets of miR-92b-3p. Results: In this study, EVs derived from porcine EECs were identified and could be taken up by PTr2 cells. We found that the EVs derived from EECs transfected with miR-92b-3p mimic (EVs-miR-92b-3p) significantly promoted the proliferation, migration and adhesion of PTr2 cells. We verified that Tuberous sclerosis complex subunit (TSC1) and Dickkopf 3 (DKK3) were the target genes of miR-92b-3p. Moreover, our study showed that miR-92b-3p plays a vital role in PTr2 cells via targeting TSC1 and DKK3. Furthermore, the 3'UTR vectors of TSC1 and DKK3 can rescue the effect of miR-92b-3p on PTr2 cells. Conclusions: Taken together, this study reveals a novel mechanism that EVs derived from porcine EECs treated with miR-92b-3p crosstalk with trophoblasts by targeting TSC1 and DKK3, leading to an enhanced ability for implantation. [ABSTRACT FROM AUTHOR]

Dickkopf‐related protein (DKK) 3, a member of the DKK family, is a secreted glycoprotein that acts as a modulator of Wnt signaling in organogenesis and carcinogenesis. Recent studies have demonstrated that DKK3 has a variety of functions, suggesting that it plays roles not only in tumorigenesis, but also tumor neovascularization, prostatic acinus formation, cardiac vascular remodeling, renal and cardiac fibrosis, and immunological activity. The function of DKK3 is therefore of great interest, but details of the receptors and mechanisms involved have remained unclear. Here, we focused on the extracellular function of DKK3 as a secreted protein, and identified transforming growth factor beta induced protein ig‐h3 (TGFBI) as a secreted protein interacting with DKK3. To investigate the function of these secreted proteins, we employed an in vitro cell model involving human hepatocellular carcinoma cells, human embryonic kidney cells, or non‐neoplastic hepatocyte cells. We showed that DKK3 functions as an extracellular matrix‐like molecule supporting adhesion, motility, and invasion, and that its interaction with TGFBI inhibits the functions of secreted DKK3 in cells expressing both proteins. These results suggest that this extracellular interaction between DKK3 and TGFBI modulates cell adhesion and motility through focal adhesion kinase signaling, and that this might serve as a potential therapeutic target in the context of cancer invasion and metastasis. [ABSTRACT FROM AUTHOR]

The Wnt/β-catenin signaling pathway is frequently activated in hepatocellular carcinoma (HCC). A number of studies have focused on the aberrant hypermethylation of the DKK family proteins and its role in regulating the activation of specific signaling pathways. However, the exact way by which DKK regulates the signaling pathway caused by Core protein of HCV has not been reported. In the present study, we evaluated the expression level of DKK and its aberrant promoter methylation to investigate the involvement of epigenetic regulation in hepatoma cell lines. The transcription and protein expression of DKK1 was significantly increased, whereas the transcription and protein expression levels of DKK2, DKK3, and DKK4 were significantly decreased following overexpression of Core protein. Pyrosequencing indicated that hypermethylation of DKK3 was increased. This was associated with increased expression of Dnmt1. The investigation of the molecular mechanism indicated that HCV Core protein interacted with Dnmt1, which combined with the promoter of DKK3, leading to methylation of DKK3. Functional studies indicated that Core protein promoted the growth, migration and invasion of cancer cells. However, upregulation of the expression of DKK3 and/or the knockdown of the expression of Dnmt1 inhibited the growth, migration and invasion of cancer cells. Taken together, the data indicated that epigenetic silencing of DKK3 caused by Dnmt1 activated the Wnt/β-catenin pathway in HCV Core-mediated HCC. Therefore, DKK3 may be a potential diagnostic and therapeutic target for HCC. [ABSTRACT FROM AUTHOR]

Background and purpose: Breast cancer threatens the health of women all over the world. Although a large number of microRNAs (miRNAs) have been found to be abnormally expressed in breast cancer, a complete miRNA-messenger RNA (mRNA) network still needs to be constructed. Methods: The Cancer Genome Atlas (TCGA) database was used to download breast cancer related data sets and analyze the differentially expressed miRNAs between tumor tissues and normal tissues. The miRDB, miRTarBase and StarBase databases were used to analyze the genes targeted by different miRNAs. The ClusterProfiler package in R language was used to enrich and analyze the target genes. String database and Cytoscape 3.6.2 software were used to analyze protein- protein interaction (PPI) network and screen Hub gene. The miRNA-Hub mRNA regulatory network was constructed to determine the research signal axis, and then verified by cell experiments. Results: Two differential miRNAs were identified in TCGA data set; 278 target genes were predicted from the three databases. Ten Hub genes were identified. The constructed miRNA Hub gene network showed that hsa-mir-98-5p/DKK3 axis might play a key role in the progression of breast cancer. Cell functional experiments confirmed that hsa-miR-98-5p could inhibit apoptosis and promote cell proliferation, migration and invasion. The binding of hsa- miR-98-5p to DKK3 was further confirmed by dual luciferase activity assay. Conclusion: In this study, we analyzed a miRNA- mRNA network associated with breast cancer progression and identified an important miRNA mRNA axis in breast cancer.

Apogossypolone (ApoG2), a novel derivative of gossypol lacking of two aldehyde groups, exhibits anti-tumor effects. However, the mechanisms by which ApoG2 regulates cervical cancer (CC) cells remain unclear. In this study, we treated two CC cell lines (CaSki and HeLa) with an increasing concentration of ApoG2 for 24 h. Cell Counting Kit-8 (CCK-8) assay, colony formation assay, flow cytometry and transwell invasion assay were utilized to detect cell proliferation, apoptosis and invasion in vitro. We first observed that ApoG2 inhibited cell proliferation, invasion and epithelial-to-mesenchymal transition (EMT) process in CC cells, along with upregulation of Dickkopf Wnt signaling pathway inhibitor 3 (DKK3) in a dose-dependent manner. The immunohistochemistry confirmed the downregulation of DKK3 in tumor tissues. Moreover, DKK3 was correlated with FIGO stage and lymph node metastasis. Functionally, DKK3 overexpression significantly suppressed cell viability, colony formation and invasion, but promoted apoptosis in CaSki and HeLa cells. Overexpression of DKK3 upregulated the protein levels of cleaved caspase-3 and E-cadherin, but downregulated the protein levels of Bcl-2, N-cadherin and Vimentin. Furthermore, DKK3 knockdown reversed the suppressive effects of ApoG2 on CaSki cell proliferation, invasion and EMT markers, while DKK3 overexpression enhanced these effects. In addition, ApoG2 treatment inhibited CC xenograft tumor growth and upregulated the protein levels of DKK3, cleaved caspase-3 and E-cadherin. In conclusions, these findings suggested that ApoG2 could effectively inhibit the growth and invasion of CC cells at least partly by activating DKK3. [ABSTRACT FROM AUTHOR]

Introduction: Colorectal cancer (CRC) is the third most common cancer in the world with high mortality, hence, understanding the molecular mechanisms involved in the tumor progression are important for CRC diagnosis and treatment. MicroRNAs (miRNAs) are key gene expression regulators that can function as tumor suppressors or oncogenes in tumor cells, and modulate angiogenesis as a critical process in tumor metastasis. MiR-1290 has been demonstrated as an onco-miRNA in various types of cancer, however, the role of miR-1290 in CRC is not fully understood. This study aimed to investigate the oncogenic and angiogenic potential of miR-1290 in CRC. Methods: Lenti-miR-1290 was transduced into HCT116, SW480, and human umbilical vein endothelial cells (HUVECs). By bioinformatics analysis, we identified thrombospondin 1 (THBS1) as a novel predicted target for miR-1290. Quantitative real-time PCR, western blotting, and luciferase reporter assay were used to demonstrate suppression of miR-1290 target genes including THBS1, Dickkopf Wnt signaling pathway inhibitor 3 (DKK3), and suppressor of cancer cell invasion (SCAI) in HCT116 and HUVECs. Cell cycle analysis, proliferation, migration and, tube formation were determined by flow cytometry, MTT, wound healing, and tube formation assays, respectively. Results: MiR-1290 significantly decreased the expression of THBS1, DKK3, and SCAI. We demonstrated that miR-1290 enhanced proliferation, migration, and angiogenesis partially through suppression of THBS1, DKK3, and SCAI in CRC. Conclusion: These results suggest a novel function of miR-1290 which may contribute to tumorigenesis and angiogenesis in CRC. [ABSTRACT FROM AUTHOR]

Background: Neuropathic pain is a common and severely disabling state that affects millions of people worldwide. Microglial activation in the spinal cord plays a critical role in the pathogenesis of neuropathic pain. However, the mechanisms underlying spinal microglial activation during neuropathic pain remain incompletely understood. Here, we investigated the role of Dickkopf (DKK) 3 and its interplay with microglial activation in the spinal cord in neuropathic pain.Methods: In this study, we investigated the effects of intrathecal injection of recombinant DKK3 (rDKK3) on mechanical allodynia and microglial activation in the spinal cord after spared nerve injury (SNI) in rats by western blot (WB), immunofluorescence (IF), quantitative polymerase chain reaction (qPCR), and enzyme-linked immunosorbent assay (ELISA).Results: We found that SNI induced a significant decrease in the levels of DKK3, Kremen-1 and Dishevelled-1 (DVL-1) and up-regulated the expression of phosphorylated apoptosis signal-regulating kinase 1 (p-ASK1), phosphorylated c-JUN N-terminal kinase (p-JNK), phosphorylated p38 (p-p38) in the spinal cord. Moreover, our results showed that exogenous intrathecal administration of rDKK3 inhibited expression of p-ASK1, p-JNK, p-p38, promoted the transformation of microglia from M1 type to M2 type, and decreased the production of pro-inflammatory cytokines compared to the rats of SNI + Vehicle. However, these effects were reversed by intrathecal administration of Kremen-1 siRNA or Dishevelled-1 (DVL-1) siRNA.Conclusions: These results suggest that DKK3 ameliorates neuropathic pain via inhibiting ASK-1/JNK/p-38-mediated microglia polarization and neuroinflammation, at least partly, by the Kremen-1 and DVL-1 pathways. [ABSTRACT FROM AUTHOR]

Background: The gold standard for diagnosis of lupus nephritis (LN) is still represented by renal biopsy, and serological prognostic biomarkers are still lacking. Dickkopf homolog-3 (DKK3) has been suggested as a marker of tissue fibrosis in different conditions; however, its role in autoimmune diseases needs to be elucidated. Here, we investigated the prognostic role of DKK3 in systemic lupus erythematosus (SLE) patients with and without LN, assessing its changes in relation to kidney function, flares, and interstitial fibrosis. Methods: Overall, 132 SLE patients (57 with LN) were included and prospectively followed up for at least 36 months. DKK3 was measured in serum at baseline. Biopsies were evaluated for glomerular involvement, interstitial fibrosis, and tubular atrophy. Results: Patients with biopsy-proven LN had significantly higher levels of DKK3 than those without (median [min–max]: 215 ng/mL [81–341] vs. 21.1 ng/mL [1–69], p < 0.01). DKK3 levels were associated with prevalent chronic kidney diseases (OR: 4.31 [C.I. 2.01–6.61] per DKK3 doubling, p < 0.01), higher chronicity index at biopsy (1.75 [1.51–2.77] per DKK3 doubling, p < 0.01), and flares rate (OR: 1.45 [C.I. 1.1–5.71] per DKK3 doubling, p < 0.044). Conclusions: While kidney biopsy still represents the gold standard for diagnostic and prognostic assessment in LN, DKK3 could represent an additional prognostic tool to monitor SLE patients and guide therapeutic choices. [ABSTRACT FROM AUTHOR]

Several microRNAs (miRNAs) and circular RNAs (circRNAs) were reported to be involved in the Docetaxel (DTX) chemoresistance of cancer treatment, but the underlying mechanisms remain to be explored. In this study, we established cellular and animal models respectively to study the effect and underlying molecular mechanisms of the dysregulation of circRNA_0006404 and circRNA_0000735 in tumor response to DTX treatment. Quantitative real-time PCR was performed to measure the expression of circRNA_0006404, miR-346, circRNA_0000735, miR-526b, Dickkopf-related protein 3 (DKK3), and Dickkopf-related protein 4 (DKK4) mRNA. The expression of circRNA_0006404 and circRNA_0000735 was remarkably suppressed and activated in DTX-treated SKOV3-R cell lines, respectively. As revealed by luciferase assays, circRNA_0006404 and circRNA_0000735 was found to be respectively targeted by miR-346 and miR-526b, while DKK3 and DKK4 were respectively targeted by miR-346 and miR-526b. Moreover, the expression of DKK3 and DKK4, which were targets of miR-346 and miR-526b, respectively, was significantly altered along with the expression of p-GP. Furthermore, circ_0006404 shRNA and circRNA_0000735 shRNA showed remarkable efficiency in stimulating the expression of circRNA_0006404, miR-346, DKK3, circRNA_0000735, miR-526b, DKK4, and p-GP in cellular and animal models. Accordingly, the cell apoptosis and proliferation were apparently changed by circ_0006404 shRNA and circRNA_0000735 shRNA in both cellular and animal models. In summary, our study found the involvement of the circRNA_0006404/miR-346/DKK3/p-GP and circRNA_0000735/miR-546b/DKK4/p-GP axis in the tumor response to DTX. Both the up-regulation of circRNA_0006404 and down-regulation of circRNA_0000735 could inhibit the expression of p-GP in vivo and ex vivo, leading to the suppressed tumor response to DTX treatment. [ABSTRACT FROM AUTHOR]

Dickkopf-3 (Dkk3) is an atypical member of the Dkk family of Wnt inhibitors, which has been implicated in the pathophysiology of neurodegenerative disorders. Its role in the mechanisms of cellular degeneration and protection is still unknown. The aim of our work is to investigate the endogenous activation of the DKK3 pathway in a model of transient occlusion of the middle cerebral artery in rats. In particular, the animals were subjected to 1 h of ischemia followed by different reperfusion times (1 h, 6 h, 12 h and 24 h) to evaluate the downstream pathway and the time course of its activation. Western blot analysis showed increased Dkk3 expression in animals with the highest time of reperfusion. The increased levels of Dkk3 were accompanied by reduced Wnt3a, Frz1 and PIWI1a expression in the cytosol while FOXM1 and β-catenin decreased in the nucleus. These molecular changes led to an increase in the apoptotic pathway, as showed by the increased expression of Caspase 3 and Bax and the reduced levels of Bcl-2, and to a decrease in neurogenesis, as shown by the decreased expression of Tbr2, Ngn2 and Pax6. In the second part of the study, we decided to employ curcumin, an activator of the Wnt/β-catenin signaling, to investigate its effect on Dkk3. In particular, curcumin was administered 1 and 6 h after ischemia, and animals were sacrificed 24 h later when the expression of Dkk3 was higher. Our data displayed that curcumin administration decreased Dkk3 expression, and increased Wnt3a, Frz1 and PIWI1a levels. Well in line with these data, curcumin administration increased nuclear β-catenin and FOXM1 expression. The down-regulation of Dkk3 by curcumin led to reduced apoptosis and increased neurogenesis. Summarizing, our results showed that Dkk3 acts as an inhibitor of Wnt/β-catenin signaling during cerebral ischemia. Additionally, its inhibition and the contextual activation of the Wnt/β-catenin pathway are protective against ischemic stroke.

Predicting future kidney allograft function is challenging. Novel biomarkers, such as urinary Dickkopf-3 (uDKK3), may help guide donor selection and improve allograft outcomes. In this prospective multicenter pilot trial, we investigated whether donor uDKK3 reflects organ quality and is associated with future allograft function. We measured uDKK3/crea ratios (uDKK3/crea) from 95 deceased and 46 living kidney donors. Prenephrectomy uDKK3/crea levels were 100× higher in deceased than in living donors (9888 pg/mg vs 113 pg/mg; P < .001). Among deceased donor transplantations, recipients were stratified by their corresponding uDKK3/crea donor levels ranging below (group A, n = 68) or above (group B, n = 65) median. The primary end point of best estimated glomerular filtration rate (eGFR) within the first 3 months after kidney transplantation was superior in group A (56.3 mL/min/1.73 m 2 ) than that in group B (44.2 mL/min/1.73 m 2 ; P = .0139). Second, the composite clinical end point consisting of death, allograft failure or eGFR decline >50% occurred less frequent in group A. By mixed linear regression modeling, donor uDKK3/crea remained an independent predictor of eGFR after transplantation, with a slope of -4.282 mL/min/1.73 m 2 per logarithmic increase in donor uDKK3/crea. In summary, uDKK3 may serve as a noninvasive, donor-dependent biomarker for assessing organ quality and future allograft function., Competing Interests: Declaration of competing interest The authors of this manuscript have conflicts of interest to disclose as described by the American Journal of Transplantation. J.de Fallois is a member of the European Reference Network for Rare Kidney Diseases (ERKNet). A. Günzel received a doctoral stipend from the RES. C. Daniel receives funding from German Research Foundation (DFG; TR374 project number C2). K. Amann receives funding from German Research Foundation (DFG; TR374 project number C2). J. Halbritter receives funding from the German Research Foundation (DFG; project number HA 6908/4-1, HA 6908/7-1, and HA 6908/8-1) and is a member of the European Reference Network for Rare Kidney Diseases (ERKNet). Other authors of this manuscript have no conflicts of interest to disclose as described by the American Journal of Transplantation., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

ObjectiveThe aim of the study was to determine the differential expression patterns of the wingless-type (Wnt) pathway inhibitors Dkk3 (Dickkopf 3), SFRP1 (secreted frizzled-related protein 1), and SFRP4 in normal müllerian tissue and endometrial endometrioid adenocarcinoma specimens.MethodsMessenger RNA (mRNA) and protein levels of the Wnt pathway inhibitors Dkk3, SFRP1, and SFRP4 were evaluated by real-time reverse transcription-polymerase chain reaction and Western blot analysis. A total of 87 human tissue specimens were obtained from 60 women who participated in Gynecologic Oncology Group protocol 210. Twenty-seven normal müllerian tissues, 32 early-stage, and 28 advanced-stage endometrial endometrioid cancer specimens were analyzed.ResultsMedian age for this cohort was 60 years, with median body mass index of 32 kg/m. There was a difference in Dkk3 protein expression between normal müllerian tissues and primary endometrial endometrioid adenocarcinoma samples (P = 0.05). There was down-regulation of Dkk3, SFRP1, and SFRP4 mRNA expression in patients with high-grade disease (P = 0.08, 0.06, and 0.05, respectfully). Furthermore, a decrease in SFRP1 and SFPR4 mRNA expression was noted in patients with a diagnosis of locoregional and distant disease recurrence. Lastly, a trend toward decreased progression-free survival in patients with low Dkk3, SFRP1, and SFRP4 mRNA expression levels was noted.ConclusionsWnt pathway inhibitor (Dkk3, sFRP1, and/or sFRP4) expression was down-regulated in patients with high-grade disease and was associated with locoregional and distant disease recurrence. Despite sample size (power) limitations, these results support previous preclinical studies and may suggest a therapeutic role for Wnt signaling in endometrial cancer.

Increasing evidence has shown that microRNAs (miRNAs) participate in cardiac fibrosis. We aimed to elucidate the effect of miRNA miR-25-3p on cardiac fibrosis. MiRNA microarray was used to profile miRNAs in the myocardium of angiotensin-II (Ang-II)-infused mice. Effect of miR-25-3p on expression of fibrosis-related genes, including Col1a1, Col3a1, and Acta2, was investigated both in vitro and in vivo. MiR-25-3p was shown increased in the myocardium of Ang-II-infused mice and patients with heart failure. MiR-25-3p enhanced fibrosis-related gene expression in mouse cardiac fibroblasts (mCFs) and in the myocardium of Ang-II-infused mice. Dickkopf 3 (Dkk3) was identified as a target gene of miR-25-3p, and Dkk3 could ameliorate Smad3 activation and fibrosis-related gene expression via enhancing Smad7 expression in mCFs. Additionally, NF-κB signal was proven to mediate upregulation of miR-25-3p in cardiac fibrosis. Our findings suggest that miR-25-3p enhances cardiac fibrosis by suppressing Dkk3 to activate Smad3 and fibrosis-related gene expression. [ABSTRACT FROM AUTHOR]

Summary: Our previous studies revealed that MYCN downregulates the expression of DKK3, activates the Wnt/β-catenin signalling pathway at the transcriptional level, and thereby promotes the development of B cell acute lymphocytic leukaemia (B-ALL) but does not affect the methylation of the DKK3 promoter. Some studies have shown that MYCN is associated with histone acetylation. We speculate that histone deacetylase inhibitors (HDACis) can inhibit the Wnt/β-catenin signalling pathway by inhibiting MYCN and increasing the expression of DKK3. Based on previous experiments, we tested this hypothesis by analysing the changes in MYCN, DKK3 and the Wnt/β-catenin signalling pathways in B-ALL cells after treatment with the selective HDACi chidamide. The in vitro and in vivo experiments confirmed that chidamide inhibited the expression of MYCN and increased the expression of DKK3 by inhibiting the activity of histone deacetylase, and these effects resulted in inhibition of the Wnt/β-catenin signalling pathway and the proliferation of B-ALL cells. These findings indicate that chidamide might be used alone or in combination with other chemotherapy regimens for patients with B-ALL and thus provide a new approach to the treatment of B-ALL. [ABSTRACT FROM AUTHOR]

A host of novel renal biomarkers have been developed over the past few decades which have enhanced monitoring of renal disease and drug-induced kidney injury in both preclinical studies and in humans. Since chronic kidney disease (CKD) and acute kidney injury (AKI) share similar underlying mechanisms and the tubulointerstitial compartment has a functional role in the progression of CKD, urinary biomarkers of AKI may provide predictive information in chronic renal disease. Numerous studies have explored whether the recent AKI biomarkers could improve upon the standard clinical biomarkers, estimated glomerular filtration rate (eGFR), and urinary albumin to creatinine ratio, for predicting outcomes in CKD patients. This review is an introduction to alternative assays that can be utilized in chronic (>3 months duration) nonclinical safety studies to provide information on renal dysfunction and to demonstrate specific situations where these assays could be utilized in nonclinical drug development. Novel biomarkers such as symmetrical dimethyl arginine, dickkopf homolog 3, and cystatin C predict chronic renal injury in animals, act as surrogates for GFR, and may predict changes in GFR in patients over time, ultimately providing a bridge from preclinical to clinical renal monitoring. [ABSTRACT FROM AUTHOR]

Many long noncoding RNAs (lncRNAs) are enriched in pancreatic islets and several lncRNAs are linked to type 2 diabetes (T2D). Although they have emerged as potential players in β-cell biology and T2D, little is known about their functions and mechanisms in human β-cells. We identified an islet-enriched lncRNA, TUNAR (TCL1 upstream neural differentiation-associated RNA), which was upregulated in β-cells of patients with T2D and promoted human β-cell proliferation via fine-tuning of the Wnt pathway. TUNAR was upregulated following Wnt agonism by a glycogen synthase kinase-3 (GSK3) inhibitor in human β-cells. Reciprocally, TUNAR repressed a Wnt antagonist Dickkopf-related protein 3 (DKK3) and stimulated Wnt pathway signaling. DKK3 was aberrantly expressed in β-cells of patients with T2D and displayed a synchronized regulatory pattern with TUNAR at the single cell level. Mechanistically, DKK3 expression was suppressed by the repressive histone modifier enhancer of zeste homolog 2 (EZH2). TUNAR interacted with EZH2 in β-cells and facilitated EZH2-mediated suppression of DKK3. These findings reveal a novel cell-specific epigenetic mechanism via islet-enriched lncRNA that fine-tunes the Wnt pathway and subsequently human β-cell proliferation. NEW & NOTEWORTHY The discovery that long noncoding RNA TUNAR regulates β-cell proliferation may be important in designing new treatments for diabetes. [ABSTRACT FROM AUTHOR]

Dkk3 has been discovered during comparison of immortalized and parental cells. Its expression has been shown to reduce colony formation and induce apoptosis of cancer cells, acting as a tumor suppressor. Herein, we demonstrate that Dkk3 overexpression or protein treatment may inhibit colorectal cancer cell proliferation, migration, and invasion and that they may promote apoptosis and G2 phase arrest with hypoexpression of Bcl-2, cdc25B, cdc25c, N-cadherin, slug, and twist and hyperexpression of Bax and E-cadherin. This effect is consistent with that of recombinant Dkk3 exposure and blocked with anti-Dkk3 antibody. Dkk3 deletion in intestinal cells was not associated with the emergence of epithelial lesions; however, adenoma emerged after sodium desoxycholate treatment. At both mRNA and protein levels, Dkk3 expression was higher in normal than in cancer tissues (p

Background: MicroRNAs (miRNAs) are small noncoding RNAs (containing approximately 22 nucleotides), which modulate and control the expression of target genes by binding them. MiRNAs play a crucial role in tumorigenesis. Thus, alterations in the expression level of miRNAs play a key role in the pathobiology of numerous cancers. In this research, the expression level of MicroRNA-1290 (miR1290) and its target genes THBS1 and DKK3 were evaluated in colorectal cancer (CRC) patients. Methods: This case-control study was carried out on 144 paraffin-embedded tissue samples of CRC and adjacent tissues from patients who referred to Imam Khomeini Hospital, Tehran, Iran. Total RNA was isolated from the tissue using Trizol reagent following the manufacturer’s instructions and then reverse transcribed to cDNA. The expression of miR-1290 and its target genes was measured by quantitative Real-Time PCR (qRT-PCR). Statistical analyses were performed using SPSS V.20 statistical software. Results: We present evidence that the miR-1290 expression in CRC tissues was significantly higher than in the normal margin, and its targets were downregulated in tumor tissue compared to the adjacent tissue. Conclusion: This study supports the essential role of miR-1290 and its contribution to CRC invasion and metastasis through targeting THBS1 and DKK3, as biomarkers for CRC diagnosis.

Abstract Background miR-214 was demonstrated to be upregulated in models of renal disease and promoted fibrosis in renal injury independent of TGF-β signaling in vivo. However, the detailed role of miR-214 in acute kidney injury (AKI) and its underlying mechanism are still largely unknown. Methods In this study, an I/R-induced rat AKI model and a hypoxia-induced NRK-52E cell model were used to study AKI. The concentrations of kidney injury markers serum creatinine, blood urea nitrogen, and kidney injury molecule-1 were measured. The expressions of miR-214, tumor necrosis factor-α, interleukin (IL)-1β, IL-6, were detected by RT-qPCR. The protein levels of Bcl-2, Bax, Dickkopf-related protein 3, β-catenin, c-myc, and cyclinD1 were determined by western blot. Cell apoptosis and caspase 3 activity were evaluated by flow cytometry analysis and caspase 3 activity assay, respectively. Luciferase reporter assay was used to confirm the interaction between miR-214 and Dkk3. Results miR-214 expression was induced in ischemia–reperfusion (I/R)-induced AKI rat and hypoxic incubation of NRK-52E cells. Overexpression of miR-214 alleviated hypoxia-induced NRK-52E cell apoptosis while inhibition of miR-214 expression exerted the opposite effect. Dkk3 was identified as a target of miR-214. Anti-miR-214 abolished the inhibitory effects of DKK3 knockdown on hypoxia-induced NRK-52E cell apoptosis by inactivation of Wnt/β-catenin signaling. Moreover, miR-214 ameliorated AKI in vivo by inhibiting apoptosis and fibrosis through targeting Dkk3 and activating Wnt/β-catenin pathway. Conclusion miR-214 ameliorates AKI by inhibiting apoptosis through targeting Dkk3 and activating Wnt/β-catenin signaling pathway, offering the possibility of miR-214 in the therapy of ischemic AKI.

Introduction: Ovarian cancer (OV) can seriously endanger women's physical and mental health. Serum DKK3 has been used for the diagnosis and prognosis of patients with ovarian cancer. However, the specificity of antibodies may lead to errors in the detection of plasma protein. Methods: Circulating CD133+ cells from blood samples were separated by magnetic microbeads. Serum DKK3 levels were determined by ELISA. The roles of DKK3 in OV cells were analyzed in vitro. Results: In this study, we found that the CD133+ subpopulation in circulating tumor cells can indicate the overall survival rate of OV patients. Serum DKK3 levels were negatively correlated with the number of circulating CD133+ cells in OV patients. In addition, we confirmed the inhibitory effect of recombinant human DKK3 (rhDKK3) on OV cells via reversal of the epithelial–mesenchymal transition (EMT) process. Conclusion: Both serum DKK3 levels and circulating CD133+ tumor cells can be used as prognostic markers for patients with ovarian cancer. [ABSTRACT FROM AUTHOR]

Dkk3 has been discovered during comparison of immortalized and parental cells. Its expression has been shown to reduce colony formation and induce apoptosis of cancer cells, acting as a tumor suppressor. Herein, we demonstrate that Dkk3 overexpression or protein treatment may inhibit colorectal cancer cell proliferation, migration, and invasion and that they may promote apoptosis and G2 phase arrest with hypoexpression of Bcl-2, cdc25B, cdc25c, N-cadherin, slug, and twist and hyperexpression of Bax and E-cadherin. This effect is consistent with that of recombinant Dkk3 exposure and blocked with anti-Dkk3 antibody. Dkk3 deletion in intestinal cells was not associated with the emergence of epithelial lesions; however, adenoma emerged after sodium desoxycholate treatment. At both mRNA and protein levels, Dkk3 expression was higher in normal than in cancer tissues (p <0.05). Dkk3 mRNA expression was negatively associated with its promoter methylation, growth pattern, differentiation, and favorable prognosis in the patients with colorectal cancer (p <0.05). Dkk3 -related signal pathways in colorectal cancer included those of cellular adhesion and migration, melanogenesis, chemokine, Hedgehog, JAK-STAT, TOLL-like receptor, TGF-β, MAPK, and calcium signaling (p <0.05). These findings indicate that Dkk3 expression levels can help assess cancer aggressiveness and patient prognosis. It might also suppress aggressive phenotypes and tumorigenesis as a molecular target in gene therapy. [ABSTRACT FROM AUTHOR]