Low-density Lipoprotein Receptor-related Protein 6 Expression Research Articles

MiR-424-5p Inhibits Proliferation, Migration, Invasion and Angiogenesis of the HTR-8/SVneo Cells Through Targeting LRP6 Mediated β-catenin.

The aim of this study was to investigate the effects of miR-424-5p on biological behaviors and angiogenesis of the HTR-8/SVneo Cells. Our study included 60 parturient women, which were divided into an PA group (placenta accreta, n = 30) and a normal group (normal placenta, n = 30). QPCR was used to measure the expression of miR-424-5p in placental tissues. The effects of the miR-424-5p mimic on proliferation, migration, and invasion of human HTR-8/SVneo cells and angiogenesis were analyzed. The potential modulated relationship between miR-424-5p and low-density lipoprotein receptor-related protein-6 (LRP6) was demonstrated by luciferase assay. The expression of LRP6, β-catenin, matrix metalloproteinase-2 (MMP-2), placental growth factor (PGF) and vascular endothelial growth factor (VEGF) were measured by qPCR and Western blot assays. The expression of miR-424-5p in the PA group was significantly decreased than that in the normal group. The expression of miR-424-5p has negative correlation with blood loss. Upregulation of miR-424-5p significantly suppressed the cell proliferation, migration, and invasion of HTR-8/SVneo cells in vitro, as well as the tube formation of human umbilical vein endothelial cells (HUVECs). The luciferase assay demonstrated that LRP6 was a target of miR-424-5p. The expression of LRP6, β-catenin, MMP-2, PGF and VEGF were also decreased with upregulation of miR-424-5p (p < 0.05). The inhibitory effects of miR-424-5p on HTR-8/SVneo cells and angiogenesis were enhanced by downregulation of LRP6, but were reversed by upregulation of LRP6. The present study suggests that downregulation of miR-424-5p is related to the occurrence of PA. Enhancing miR-424-5p inhibits proliferation, migration, invasion and angiogenesis of the HTR-8/SVneo cells through targeting LRP6 mediated β-catenin, providing more insights about PA.

Examining the expression of low-density lipoprotein receptor (LDLR) and low-density lipoprotein receptor-related protein 6 (LRP6) genes in breast cancer cell lines.

Cholesterol and the Wnt/β-catenin pathway have an effective role in the proliferation, survival, drug resistance, immune exhaustion, and metastasis of all types of cancer cells. Considering the role of LDLR and LRP6 proteins in cholesterol uptake by cells and activation of Wnt/β-catenin pathway, this study aims to examine the gene expression of LDLR and LRP6 in cell lines of breast cancer. Human breast cancer cell lines MCF7, MD468 and SKBR3 were cultured in suitable conditions and after extracting total RNA from them, real-Time PCR was used to measure the levels of gene expression for LDLR and LRP6. Our results showed that the expression of LDLR and LRP6 genes is significantly increased in MCF7 and MD468 cells compared to SKBR3 cells. These results suggest that LRP6 and LDLR can be considered as a therapeutic target in tumors that have a genetic profile similar to MCF7 and MD468 cells.

Kai-Xin-San Improves Cognitive Impairment via Wnt/β-Catenin and IRE1/XBP1s Signalings in APP/PS1 Mice.

Alzheimer's disease (AD) is the most common type of dementia with an insidious onset and slow progression. Kai-Xin-San (KXS) has been reported to be effective in improving cognitive impairment in AD. However, the mechanism is still confused. In this study, we employed APP/PS1 mice to explore the neuroprotective mechanism of KXS. Forty-eight male APP/PS1 mice were randomly divided into model group, KXS groups (0.7, 1.4, and 2.8 g/kg/d, p.o.) and the wild-type mice were assigned to the normal control group (n = 12 in each group). Y-maze and novel object recognition tests were carried out after continuous intragastric administration for 2 months. The abilities of learning, memory, and new object recognition in the APP/PS1 mice were enhanced significantly after KXS treatment. KXS can reduce the deposition of Aβ40 and Aβ42 in APP/PS1 mice brain. KXS decreased the levels of serum inflammatory cytokines, tumor necrosis factor-α, interleukin-1β, and interleukin-6. KXS increased the activities of superoxide dismutase and glutathione peroxidase significantly, whereas it inhibited the contents of reactive oxygen species and malondialdehyde significantly. In addition, we also detected Wnt/β-catenin signaling related proteins, such as Wnt7a, β-catenin, low-density lipoprotein receptor-related protein 6 (LRP6), glycogen synthase kinase-3β (GSK-3β), nuclear factor kappa-B (NF-κB), postsynaptic density 95 (PSD95), microtubule associated protein-2 (MAP-2), and endoplasmic reticulum stress (IRE1 pathway) related proteins, such as inositol-requiring enzyme 1 (IRE1), phosphorylated IRE1(p-IRE1), spliced X-box-binding protein 1 (XBP1s), immunoglobulin binding protein (BIP), and protein disulfide isomerase (PDI) in the hippocampus. Results showed that KXS decreased the expression of GSK-3β, NF-kB, p-IRE1/IRE1 ratio, XBP1s, and BIP; increased the expression of Wnt7a, β-catenin, LRP6, PSD95, MAP2, and PDI. In conclusion, KXS improved cognitive impairment by activating Wnt/β-catenin signaling, inhibiting the IRE1/XBP1s pathway in APP/PS1 mice.

Salinomycin sodium exerts anti diffuse large B-cell lymphoma activity through inhibition of LRP6-mediated Wnt/β-catenin and mTORC1 signaling

Low-density lipoprotein receptor-related protein-6 (LRP6) is overexpressed in various cancers. The small molecule salinomycin sodium inhibits LRP6. We observed a higher proportion of subjects with non-germinal center B (non-GCB) subtypes having high LRP6 expression than those with GCB subtypes by immunohistochemistry. The PCR and Western blot assays demonstrated increased LRP6 expression in non-GCB subtype cells. In addition, CCK-8 assays and transwell cell migration assays revealed that salinomycin sodium exhibited dose- and time-dependent inhibition of proliferation and migration in non-GCB subtype cells. Furthermore, Western blot assays showed that salinomycin sodium decreased the expression of Bcl2, while increasing the expression of Bax. Additionally, salinomycin sodium suppressed LRP6 expression, blocked LRP6 phosphorylation, and inhibited the Wnt/β-catenin and mTORC1 signaling pathways. Our results suggest that LRP6 is highly expressed in non-GCB subtype. Furthermore, salinomycin sodium inhibited LRP6 expression and the Wnt/β-catenin and mTORC1 signaling in non-GCB subtype cells, and displayed potent anticancer activity.

LRP6 Bidirectionally Regulates Insulin Sensitivity through Insulin Receptor and S6K Signaling in Rats with CG-IUGR

Intrauterine growth restriction followed by postnatal catch-up growth (CG-IUGR) increases the risk of insulin resistance-related diseases. Low-density lipoprotein receptor-related protein 6 (LRP6) plays a substantial role in glucose metabolism. However, whether LRP6 is involved in the insulin resistance of CG-IUGR is unclear. This study aimed to explore the role of LRP6 in insulin signaling in response to CG-IUGR. The CG-IUGR rat model was established via a maternal gestational nutritional restriction followed by postnatal litter size reduction. The mRNA and protein expression of the components in the insulin pathway, LRP6/β-catenin and mammalian target of rapamycin (mTOR)/S6 kinase (S6K) signaling, was determined. Liver tissues were immunostained for the expression of LRP6 and β-catenin. LRP6 was overexpressed or silenced in primary hepatocytes to explore its role in insulin signaling. Compared with the control rats, CG-IUGR rats showed higher homeostasis model assessment for insulin resistance (HOMA-IR) index and fasting insulin level, decreased insulin signaling, reduced mTOR/S6K/ insulin receptor substrate-1 (IRS-1) serine307 activity, and decreased LRP6/β-catenin in the liver tissue. The knockdown of LRP6 in hepatocytes from appropriate-for-gestational-age (AGA) rats led to reductions in insulin receptor (IR) signaling and mTOR/S6K/IRS-1 serine307 activity. In contrast, LRP6 overexpression in hepatocytes of CG-IUGR rats resulted in elevated IR signaling and mTOR/S6K/IRS-1 serine307 activity. LRP6 regulated the insulin signaling in the CG-IUGR rats via two distinct pathways, IR and mTOR-S6K signaling. LRP6 may be a potential therapeutic target for insulin resistance in CG-IUGR individuals.

The Relationship between LRP6 and Wnt/β-Catenin Pathway in Colorectal and Esophageal Cancer

High expression of low-density lipoprotein receptor-related protein 6 (LRP6), a key component of the Wnt/β-catenin signaling pathway, is reported to be associated with malignant potential in some solid tumors including breast cancer and hepatocellular carcinoma. Few reports, however, have examined its function and clinical significance in colorectal cancers (CRC) demonstrating constitutive activation of Wnt signaling. Here, we compared the expression level and function of LRP6 in CRC with that of esophageal squamous cell carcinoma (ESCC) bearing few Wnt/β-catenin pathway mutations. On immunohistochemical staining, high LRP6 expression was noted in three of 68 cases (4.4%), and high β-catenin in 38 of 67 cases (56.7%) of CRC. High LRP6 expression was found in 21 of 82 cases (25.6%), and high β-catenin expression in 29 of 73 cases (39.7%) of ESCC. In our in vitro studies, LRP6 knockdown hardly changed Wnt signaling activity in CRC cell lines with mutations in Wnt signaling downstream genes. In contrast, in ESCC cell lines without Wnt signaling-related mutations, LRP6 knockdown significantly decreased Wnt signaling activity. LRP6 function may depend on constitutive activation of Wnt signaling.

Chlorogenic Acid Inhibits Epithelial-Mesenchymal Transition and Invasion of Breast Cancer by Down-Regulating LRP6.

Epithelial-mesenchymal transition (EMT) is a crucial biologic process for breast cancer metastasis, and inhibition of EMT could be an effective approach to suppress metastatic potential of mammary cancer. High expression of low-density lipoprotein receptor-related protein 6 (LRP6) is usually observed in breast carcinoma and predicts poor prognosis. In the present study, we investigated whether chlorogenic acid (CA) can inhibit the EMT of breast cancer cells and underlying molecular mechanism. We found that CA treatment transformed MCF-7 cell morphology from spindle shape (mesenchymal phenotype) to spherical shape (epithelial phenotype). CA clearly increased epithelial biomarkers' expression (E-cadherin and ZO-1) but decreased mesenchymal proteins' expression (ZEB1, N-cadherin, vimentin, snail, and slug). In addition, CA attenuated MMP-2 and MMP-9 activities and inhibited cell migration and invasion. CA downregulated the expression of LRP6 in MCF-7 cells. Knockdown LRP6 with siRNA repressed cell mobility and invasion, wheras overexpression of LRP6 promoted EMT and antagonized the EMT inhibitory effect of CA on MCF-7 cells. Furthermore, CA directly interacted with Wnt/β-catenin signaling coreceptor LRP6 and reduced LRP6, p-LRP6, and β-catenin expression levels in MCF-7 cells. In vivo study revealed that CA notably reduced tumor volume and tumor weight. CA decreased the expression of LRP6, N-cadherin, ZEB1, vimentin, MMP2, MMP9, and increased the expression of E-cadherin and ZO-1. In conclusion, CA inhibited EMT and invasion of breast cancer by targeting LRP6. SIGNIFICANCE STATEMENT: CA, the familiar polyphenol compound in traditional Chinese medicine, repressed EMT and weakened cellular mobility and invasion in MCF-7 cells. The mechanism studies demonstrated that CA could inhibit EMT and invasion of MCF-7 cells via targeting LRP6. Additionally, CA restrained tumor growth and xenograft tumor EMT in vivo. The EMT inhibitory property of CA warrants further studies of CA as a drug candidate for the therapy of metastatic breast carcinoma.

The Low-density Lipoprotein Receptor-related Protein 6 Pathway in the Treatment of Intestinal Barrier Dysfunction Induced by Hypoxia and Intestinal Microbiota through the Wnt/β-catenin Pathway

Our study is to explore the key molecular of Low-density lipoprotein receptor-related protein 6 (LRP6) and the related Wnt/β-catenin pathway regulated by LRP6 during the intestinal barrier dysfunction. Colorectal protein profile analysis showed that LRP6 expression was decreased in dextran sulfate sodium (DSS)-induced colitis mice, and mice received fecal bacteria transplantation from stroke patients. Mice with intestinal hypoxia and intestinal epithelial cells cultured in hypoxia showed decreased expression of LRP6. Overexpression of LPR6 or its N-terminus rescued the Wnt/β-catenin signaling pathway which was inhibited by hypoxia and endoplasmic reticulum stress. In mice overexpressing of LRP6, the expression of β-catenin and DKK1 increased, Bcl2 decreased, and Bax increased. Mice with LRP6 knockout showed an opposite trend, and the expression of Claudin2, Occludin and ZO-1 decreased. Two drugs, curcumin and auranofin could alleviate intestinal barrier damage in DSS-induced colitis mice by targeting LRP-6. Therefore, gut microbiota dysbiosis and hypoxia can inhibit the LRP6 and Wnt/β-catenin pathway, and drugs targeting LRP6 can protect the intestinal barrier.

Changes of LRP6/β-catenin pathway in adipose tissue of rats with intrauterine growth restriction with catch-up growth.

To investigate the expression of low-density lipoprotein receptor-related protein 6 (LRP6)/β-catenin pathway related proteins in adipose tissue of rats with intrauterine growth restriction with catch-up growth SD rats were randomly divided into nutrition-restriction rats and normal feed rats during pregnancy. CG-IUGR model was established by reducing the number of offspring in the nutrition-restriction rats (CG-IUGR group); while the rats in the control group were offspring of the normal feed pregnant rats. In order to exclude the interference of gender, male offspring mice were selected in both the CG-IUGR group and the control group in the following studies. The CG-IUGR group and the control group were subjected to glucose tolerance test at 12 weeks of age, and the perirenal adipose tissue samples were taken to observe the adipose structure by HE staining. Expression of LRP6, β-catenin and insulin receptor substrate 1 (IRS-1) in adipocytes were examined by confocal microscopy. Protein expression of LRP6, β-catenin and IRS-1 were measured by Western blotting. Blood glucose level and the area under the cure of CG-IUGR group were significantly higher than that of control group (both <0.05). Adipocyte size in the CG-IUGR group was significantly larger than that of control group, and the expression of LRP6, β-catenin and IRS-1 protein in adipose tissue of the CG-IUGR group was significantly lower than that of control group (all <0.05). : The expression of LRP6/β-catenin pathway related proteins is reduced in the adipose tissue in CG-IUGR rats, probably contributing to the insulin resistance in these rats.

PUM1 modulates trophoblast cell proliferation and migration through LRP6.

Preeclampsia is a severe pregnancy complication characterized by hypertension and may cause maternal morbidity and mortality. A better understanding of the essential genes involved in preeclampsia pathophysiology is urgently needed. This study investigated the function and molecular mechanisms of pumilio RNA binding family member 1 (PUM1) in extravillous trophoblast cells (EVTs). The interaction between protein and mRNA was verified by RNA pull-down assays, RNA immunoprecipitation assays, and luciferase reporter assays. The mRNA and protein levels of the genes involved were determined by RT-qPCR and western blot assays, respectively. Our results demonstrated that PUM1 could bind to the 3'-untranslated region of low-density lipoprotein receptor-related protein 6 (LRP6) mRNA, resulting in reduced expression of LRP6 mRNA and protein. Repression of PUM1 resulted in enhanced colony formation, cell proliferation, migration, and invasion of EVTs. The PUM1-depletion-mediated promotion effects on EVTs could be abrogated by LRP6 knockdown. PUM1 regulates the growth and mobility of EVTs by modulating LRP6 expression. Developing strategies to balance PUM1 and LRP6 levels may be beneficial for the management of preeclampsia patients.

Hsa_circ_0074371 Regulates Proliferation, Apoptosis, Migration, and Invasion via the miR-582-3p/LRP6 Axis in Trophoblast Cells.

Fetal growth restriction (FGR) ranks second among causes of perinatal death. Circular RNA hsa_circ_0074371 (hsa_circ_0074371) is reported to be downregulated in FGR placentae. However, the role and regulatory mechanism of hsa_circ_0074371 in FGR pathogenesis are indistinct. Expression of hsa_circ_0074371, microRNA (miR)-582-3p, and low-density lipoprotein receptor-related protein 6 (LRP6) mRNA in FGR placentae and trophoblast cells (HTR-8/SVneo) was detected by quantitative real-time polymerase chain reaction (qRT-PCR). The relationship between hsa_circ_0074371 or LRP6 and miR-582-3p was verified by dual-luciferase reporter and/or RNA immunoprecipitation (RIP), RNA pull-down assays. The proliferation, cell cycle progression, apoptosis, migration, and invasion of HTR-8/SVneo cells were evaluated by Cell Counting Kit-8 (CCK-8), flow cytometry, wound healing, or transwell assays. Caspase3 activity was analyzed with a commercial kit. The protein levels of c-myc, cyclinD1, matrix metalloproteinase (MMP) 2, MMP9, and LRP6 were examined by western blotting. We observed that hsa_circ_0074371 and LRP6 were downregulated while miR-582-3p was upregulated in FGR placentae and HTR-8/SVneo cells. Hsa_circ_0074371 modulated LRP6 expression via sponging miR-582-3p. Hsa_circ_0074371 knockdown induced cell cycle arrest, apoptosis, and inhibited cell proliferation, migration, and invasion in HTR-8/SVneo cells. MiR-582-3p inhibitor reversed hsa_circ_0074371 silencing me on proliferation, migration, and invasion of HTR-8/SVneo cells. LRP6 overexpression overturned the inhibitory effect of miR-582-3p mimic on proliferation, migration, and invasion of HTR-8/SVneo cells. In conclusion, hsa_circ_0074371 downregulation inhibited proliferation, migration, and invasion of trophoblast cells via sponging miR-582-3p and decreasing LRP6 expression, providing a new mechanism related to FGR pathogenesis.

LDL receptor-related protein LRP6 senses nutrient levels and regulates Hippo signaling.

Controlled cell growth and proliferation are essential for tissue homeostasis and development. Wnt and Hippo signaling are well known as positive and negative regulators of cell proliferation, respectively. The regulation of Hippo signaling by the Wnt pathway has been shown, but how and which components of Wnt signaling are involved in the activation of Hippo signaling during nutrient starvation are unknown. Here, we report that a reduction in the level of low-density lipoprotein receptor-related protein 6 (LRP6) during nutrient starvation induces phosphorylation and cytoplasmic localization of YAP, inhibiting YAP-dependent transcription. Phosphorylation of YAP via loss of LRP6 is mediated by large tumor suppressor kinases 1/2 (LATS1/2) and Merlin. We found that O-GlcNAcylation of LRP6 was reduced, and the overall amount of LRP6 was decreased via endocytosis-mediated lysosomal degradation during nutrient starvation. Merlin binds to LRP6; when LRP6 is less O-GlcNAcylated, Merlin dissociates from it and becomes capable of interacting with LATS1 to induce phosphorylation of YAP. Our data suggest that LRP6 has unexpected roles as a nutrient sensor and Hippo signaling regulator.

A Role for the WNT Co-Receptor LRP6 in Pathogenesis and Therapy of Epithelial Cancers.

The WNT/β-catenin signaling pathway controls stem and progenitor cell proliferation, survival and differentiation in epithelial tissues. Aberrant stimulation of this pathway is therefore frequently observed in cancers from epithelial origin. For instance, colorectal and hepatic cancers display activating mutations in the CTNNB1 gene encoding β-catenin, or inactivating APC and AXIN gene mutations. However, these mutations are uncommon in breast and pancreatic cancers despite nuclear β-catenin localization, indicative of pathway activation. Notably, the low-density lipoprotein receptor-related protein 6 (LRP6), an indispensable co-receptor for WNT, is frequently overexpressed in colorectal, liver, breast and pancreatic adenocarcinomas in association with increased WNT/β -catenin signaling. Moreover, LRP6 is hyperphosphorylated in KRAS-mutated cells and in patient-derived colorectal tumours. Polymorphisms in the LRP6 gene are also associated with different susceptibility to developing specific types of lung, bladder and colorectal cancers. Additionally, recent observations suggest that LRP6 dysfunction may be involved in carcinogenesis. Indeed, reducing LRP6 expression and/or activity inhibits cancer cell proliferation and delays tumour growth in vivo. This review summarizes current knowledge regarding the biological function and regulation of LRP6 in the development of epithelial cancers—especially colorectal, liver, breast and pancreatic cancers.

Downregulation of microRNA‑629‑5p in colorectal cancer and prevention of the malignant phenotype by direct targeting of low‑density lipoprotein receptor‑related protein 6

Aberrant expression of numerous microRNAs (miRNAs/miRs) in colorectal cancer (CRC) significantly affects disease progression. Recently, miR‑629‑5p (miR‑629) was identified as a tumor‑promoting miRNA in the malignant processes of a number of human cancers. However, few studies have been conducted regarding expression profiles and detailed roles of miR‑629 in CRC. In the present study, reverse transcription‑quantitative polymerase chain reaction was used to assess miR‑629 expression in CRC tissues and cell lines. Cell Counting Kit‑8 assay, flow cytometry and Transwell assays were performed to determine the in vitro effects of miR‑629 on CRC cell proliferation, apoptosis, and metastasis, respectively. Xenograft models were employed to determine the in vivo effects of miR‑629 on tumor growth in nude mice. Molecular mechanisms underlying the activity of miR‑629 in CRC cells were explored. miR‑629 expression decreased in CRC tissues and cell lines. The decreased aberrant miR‑629 expression was significantly associated with tumor size, lymphatic metastasis and tumor‑node‑metastasis stage of CRC, and was a predictor of poor prognosis. Restoring miR‑629 expression attenuated CRC cell proliferation, migration and invasion; promoted cell apoptosis in vitro; and inhibited tumor growth in vivo. Low‑density lipoprotein receptor‑related protein 6 (LRP6) was a direct target gene of miR‑629 in CRC cells. Furthermore, the effect of LRP6 knockdown was similar to that of miR‑629 overexpression in CRC cells. Restoration of LRP6 expression neutralized the effects of miR‑629 in CRC cells. miR‑629 suppressed the activation of the Wnt/β‑catenin pathway through LRP6 regulation both in vitro and in vivo. In conclusion, miR‑629 suppressed the development and progression of CRC by directly targeting LRP6 and inhibiting the Wnt/β‑catenin pathway both in vitro and in vivo. Therefore, miR‑629 may be a novel prognostic biomarker and therapeutic target in CRC.

MicroRNA-590-3p inhibits trophoblast-dependent maternal spiral artery remodeling by repressing low-density lipoprotein receptor-related protein 6.

BackgroundThe remodeling of maternal spiral artery following embryo implantation, which relies on well‐regulated trophoblast functions, is a pivotal process to ensure a successful pregnancy. Low‐density lipoprotein receptor‐related protein 6 (LRP6) and microRNAs (miRNAs, miRs) are suggested to be involved in angiogenesis and several vascular diseases; however, their functions in the control of trophoblast remain elusive. We therefore aimed to examine the roles of LRP6 and miR‐590‐3p in the regulation of trophoblast during the remodeling of maternal spiral artery.MethodsHTR‐8/SVneo cell, a trophoblast cell line, was utilized to study the effects of LRP6 and miR‐590‐3p on apoptosis, cell proliferation, migration, invasion, as well as tube formation. Expression of angiogenic factors placental growth factor (PlGF), matrix metalloproteinases (MMPs), vascular endothelial growth factor (VEGF), and activities of canonical Wnt/β‐catenin signaling pathway, which were implicated in the process of artery remodeling, were also examined.ResultsMiR‐590‐3p directly targeted 3′ untranslated region (3′‐UTR) of LRP6 mRNA and repressed LRP6 expression, which in turn inhibited proliferation, migration, invasion, as well as tube formation, and resulted in apoptosis in HTR‐8/SVneo cells. Further, inhibition of LRP6 through miR‐590‐3p significantly suppressed the expression of PlGF, MMPs, and VEGF and reduced the activation of Wnt/β‐catenin signaling pathway.ConclusionMicroRNAs‐590‐3p may inhibit trophoblast‐dependent maternal spiral artery remodeling, via both trophoblast invasion and endovascular formation, by repressing LRP6.

MiR-381-3p inhibits proliferation, migration and invasion by targeting LRP6 in papillary thyroid carcinoma.

MiR-381-3p plays an essential role in the progression of a variety of cancers, but its expression and role in papillary thyroid carcinoma (PTC) progression have not been investigated. The aim of this study was to investigate the expression of miR-381-3p and its function in PTC. The expression levels of miR-381-3p and low-density lipoprotein receptor‑related protein 6 (LRP6) mRNA in PTC tissues and cell lines were measured using RT-PCR. Cell proliferation, migration and invasion were assessed by cell viability assay and transwell assay. Luciferase assays and Western blotting were performed to demonstrate miR-381-3p target gene. We found that miR-381-3p was significantly down-regulated in PTC tissues and cell lines. In vitro assay indicated that up-regulation of miR-381-3p significantly suppressed PTC cell proliferation, migration and invasion. Moreover, luciferase reporter gene assay demonstrated that miR-381-3p could target LRP6 by binding to the 3' UTR. Western blot and Reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) showed that miR-381-3p overexpression suppressed the expression of LRP6 at both mRNA and proteins levels. In addition, functional experiment confirmed that LRP6 was involved in the suppressive effect of miR-381-3p-mediated PTC on cell proliferation, migration and invasion. Our findings suggested, for the first time, that miR-381-3p was lowly expressed in PTC tissues, and its up-regulation inhibited tumorigenesis of PTC by targeting LRP6.

Pattern of LRP6 gene expression in tumoral tissues of colorectal cancer.

Colorectal cancer (CRC) is known as one of the most leading causes of death from cancer in the world. Wnt signaling pathway is the most important signaling pathway activated in this type of cancer. The low-density lipoprotein (LDL) receptor-related protein-6 (LRP6) functions as a co-receptor in order to trigger this signaling pathway. The expression of LRP6 was studied in cell lines, tumoral and normal adjacent tissues from patients affected with colorectal cancer. The expression of LRP6 was assayed by quantitative Real time PCR. LRP6 is overexpressed at the transcript and protein level in HCT116 and SW480 in comparison to HUVEC as the non-cancerous cell. Furthermore, LRP6 was significantly up-regulated in 61% of human malignant tissues (p-value = 0.0008). In inherited type of disease, this upregulation was lower than other types of colorectal cancer, while in rectal cancer the overexpression was significantly higher (p-value = 0.039). Gene expression was not correlated with the age, gender and the stage of disease. Due to the profound overexpression of LRP6 in sporadic and rectal types of cancer compared to normal colonic ones, antagonist related approaches can be promising for targeted therapies of cancer.

Role of Wnt Co-Receptor LRP6 in Triple Negative Breast Cancer Cell Migration and Invasion.

The low-density lipoprotein receptor-related protein 6 (LRP6) is an essential Wnt co-receptor of the Wnt/β-catenin signaling pathway. Although studies have shown an increased expression of LRP6 in several types of cancer, its function in tumor development and progression remains to be elucidated. We herein demonstrated that LRP6 expression is up-regulated in human triple negative breast cancer (TNBC) patients and human TNBC cell lines, and that knockdown of LRP6 expression and treatment of recombinant Mesd protein (a specific inhibitor of LRP6) significantly decreased cell migration and invasion of TNBC MDA-MB-231 and BT549 cells. Interestingly, the effects of LRP6 knockdown and Mesd treatment on TNBC cell migration and invasion were more prominent than on TNBC cell proliferation/viability. Mechanistically, LRP6 knockdown and Mesd treatment inhibited Wnt/β-catenin signaling and decreased the expression of S100A4, a mediator of cancer metastasis and a specific target of Wnt/β-catenin signaling, in TNBC cells. Together, our data suggest that LRP6 promotes TNBC cell migration and invasion by regulating the expression and function of S100A4 via the Wnt/β-catenin signaling pathway. J. Cell. Biochem. 118: 2968-2976, 2017. © 2017 Wiley Periodicals, Inc.

MiR-30e-5p and miR-15a Synergistically Regulate Fatty Acid Metabolism in Goat Mammary Epithelial Cells via LRP6 and YAP1.

MicroRNA (miRNA) regulates the expression of genes and influences a series of biological processes, including fatty acid metabolism. We screened the expression of miRNA in goat mammary glands during peak-lactation and non-lactating (“dry”) periods, and performed an in vitro study with goat mammary epithelial cells (GMEC) prior to sequencing analysis. Results illustrated that miR-30e-5p and miR-15a were highly expressed. Utilizing a luciferase reporter assay and Western blot, low-density lipoprotein receptor-related protein 6 (LRP6) and Yes associated protein 1 (YAP1) genes were demonstrated to be a target of miR-30e-5p and miR-15a in GMEC. Moreover, we demonstrated that the overexpression of miR-30e-5p and miR-15a in GMEC promoted fat metabolism while their knockdown impaired fat metabolism. These findings extend the discovery of a key role of miR-30e-5p and miR-15a in mediating adipocyte differentiation by suggesting a role in promoting milk fat synthesis. In conclusion, our findings indicate that miR-30e-5p, together with miR-15a, represses expression of LRP6 and promotes fat metabolism in GMEC. The data expanded our knowledge on the function of miRNAs in milk fat metabolism and synthesis in ruminant mammary cells.

Post-transcriptional regulation of Wnt co-receptor LRP6 and RNA-binding protein HuR by miR-29b in intestinal epithelial cells.

MicroRNAs (miRNAs) control gene expression by binding to their target mRNAs for degradation and/or translation repression and are implicated in many aspects of cellular physiology. Our previous study shows that miR-29b acts as a biological repressor of intestinal mucosal growth, but its exact downstream targets remain largely unknown. In the present study, we found that mRNAs, encoding Wnt co-receptor LRP6 (low-density lipoprotein-receptor-related protein 6) and RNA-binding protein (RBP) HuR, are novel targets of miR-29b in intestinal epithelial cells (IECs) and that expression of LRP6 and HuR is tightly regulated by miR-29b at the post-transcriptional level. miR-29b interacted with both Lrp6 and HuR mRNAs via their 3′-UTRs and inhibited LRP6 and HuR expression by destabilizing Lrp6 and HuR mRNAs and repressing their translation. Studies using heterologous reporter constructs revealed a greater repressive effect of miR-29b through a single binding site in the Lrp6 or HuR 3′-UTR, whereas deletion mutation of this site prevented miR-29b-induced repression of LRP6 and HuR expression. Repression of HuR by miR-29b in turn also contributed to miR-29b-induced LRP6 inhibition, since ectopic overexpression of HuR in cells overexpressing miR-29b restored LRP6 expression to near normal levels. Taken together, our results suggest that miR-29b inhibits expression of LRP6 and HuR post-transcriptionally, thus playing a role in the regulation of IEC proliferation and intestinal epithelial homoeostasis.