Expression Of LRIG1 Research Articles

IPSC-NSCs-derived exosomal let-7b-5p improves motor function after spinal cord Injury by modulating microglial/macrophage pyroptosis

BackgroundFollowing spinal cord injury (SCI), the inflammatory storm initiated by microglia/macrophages poses a significant impediment to the recovery process. Exosomes play a crucial role in the transport of miRNAs, facilitating essential cellular communication through the transfer of genetic material. However, the miRNAs from iPSC-NSCs-Exos and their potential mechanisms leading to repair after SCI remain unclear. This study aims to explore the role of iPSC-NSCs-Exos in microglia/macrophage pyroptosis and reveal their potential mechanisms.MethodsiPSC-NSCs-Exos were characterized and identified using transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and Western blot. A mouse SCI model and a series of in vivo and in vitro experiments were conducted to investigate the therapeutic effects of iPSC-NSCs-Exos. Subsequently, miRNA microarray analysis and rescue experiments were performed to confirm the role of miRNAs in iPSC-NSCs-Exos in SCI. Mechanistic studies were carried out using Western blot, luciferase activity assays, and RNA-ChIP.ResultsOur findings revealed that iPSC-NSCs-derived exosomes inhibited microglia/macrophage pyroptosis at 7 days post-SCI, maintaining myelin integrity and promoting axonal growth, ultimately improving mice motor function. The miRNA microarray showed let-7b-5p to be highly enriched in iPSC-NSCs-Exos, and LRIG3 was identified as the target gene of let-7b-5p. Through a series of rescue experiments, we uncovered the connection between iPSC-NSCs and microglia/macrophages, revealing a novel target for treating SCI.ConclusionIn conclusion, we discovered that iPSC-NSCs-derived exosomes can package and deliver let-7b-5p, regulating the expression of LRIG3 to ameliorate microglia/macrophage pyroptosis and enhance motor function in mice after SCI. This highlights the potential of combined therapy with iPSC-NSCs-Exos and let-7b-5p in promoting functional recovery and limiting inflammation following SCI.

Immunoreactivity of LMO7 and other molecular markers as potential prognostic factors in oropharyngeal squamous cell carcinoma

BackgroundDespite the better prognosis associated with human papillomavirus (HPV)-positive oropharyngeal squamous cell carcinoma (OPSCC), some patients experience relapse and succumb to the disease; thus, there is a need for biomarkers identifying these patients for intensified treatment. Leucine-rich repeats and immunoglobulin-like domain (LRIG) protein 1 is a negative regulator of receptor tyrosine kinase signaling and a positive prognostic factor in OPSCC. Studies indicate that LRIG1 interacts with the LIM domain 7 protein (LMO7), a stabilizer of adherence junctions. Its role in OPSCC has not been studied before.MethodsA total of 145 patients diagnosed with OPSCC were enrolled. Immunohistochemical LMO7 expression and staining intensity were evaluated in the tumors and correlated with known clinical and pathological prognostic factors, such as HPV status and LRIG1, CD44, Ki67, and p53 expression.ResultsOur results show that high LMO7 expression is associated with significantly longer overall survival (OS) (p = 0.044). LMO7 was a positive prognostic factor for OS in univariate analysis (HR 0.515, 95% CI: 0.267–0.994, p = 0.048) but not in multivariate analysis. The LMO7 expression correlated with LRIG1 expression (p = 0.048), consistent with previous findings. Interestingly, strong LRIG1 staining intensity was an independent negative prognostic factor in the HPV-driven group of tumors (HR 2.847, 95% Cl: 1.036–7.825, p = 0.043).ConclusionsWe show for the first time that high LMO7 expression is a positive prognostic factor in OPSCC, and we propose that LMO7 should be further explored as a biomarker. In contrast to previous reports, LRIG1 expression was shown to be an independent negative prognostic factor in HPV-driven OPSCC.

Dysregulation and antimetastatic function of circLRIG1 modulated by miR-214-3p/LRIG1 axis in bladder carcinoma

CircLRIG1, a newly discovered circRNA, has yet to have its potential function and biological processes reported. This study explored the role of circLRIG1 in the development and progression of bladder carcinoma and its potential molecular mechanisms. Techniques such as qRT-PCR, Western blot, various cellular assays, and in vivo models were used to investigate mRNA and protein levels, cell behavior, molecular interactions, and tumor growth. The results showed that both circLRIG1 and LRIG1 were significantly reduced in bladder carcinoma tissues and cell lines. Low circLRIG1 expression was associated with poor patient prognosis. Overexpressing circLRIG1 inhibited bladder carcinoma cell growth, migration, and invasion, promoted apoptosis, and decreased tumor growth and metastasis in vivo. Importantly, circLRIG1 was found to sponge miR-214-3p, enhancing LRIG1 expression, and its overexpression also modulated protein levels of E-cadherin, N-cadherin, Vimentin, and LRIG1. Similar effects were observed with LRIG1 overexpression. Notably, a positive correlation was found between circLRIG1 and LRIG1 expression in bladder carcinoma tissues. Additionally, the tumor-suppressing effect of circLRIG1 was reversed by overexpressing miR-214-3p or silencing LRIG1. The study concludes that circLRIG1 suppresses bladder carcinoma progression by enhancing LRIG1 expression via sponging miR-214-3p, providing a potential strategy for early diagnosis and treatment of bladder carcinoma.

FoxC1 activates limbal epithelial stem cells following corneal epithelial debridement

Limbal epithelial stem cells are not only critical for corneal epithelial homeostasis but also have the capacity to change from a relatively quiescent mitotic phenotype to a rapidly proliferating cell in response to population depletion following corneal epithelial wounding. Pax6+/− mice display many abnormalities including corneal vascularization and these aberrations are consistent with a limbal stem cell deficiency (LSCD) phenotype. FoxC1 has an inhibitory effect on corneal avascularity and a positive role in stem cell maintenance in many tissues. However, the role of FoxC1 in limbal epithelial stem cells remains unknown. To unravel FoxC1's role(s) in limbal epithelial stem cell homeostasis, we utilized an adeno-associated virus (AAV) vector to topically deliver human FOXC1 proteins into Pax6+/− mouse limbal epithelium. Under unperturbed conditions, overexpression of FOXC1 in the limbal epithelium had little significant change in differentiation (PAI-2, Krt12) and proliferation (BrdU, Ki67). Conversely, such overexpression resulted in a marked increase in the expression of putative limbal epithelial stem cell markers, N-cadherin and Lrig1. After corneal injuries in Pax6+/− mice, FOXC1 overexpression enhanced the behavior of limbal epithelial stem cells from quiescence to a highly proliferative status. Overall, the treatment of AAV8-FOXC1 may be beneficial to the function of limbal epithelial stem cells in the context of a deficiency of Pax6 function.

Transitory alkali exposure on meibomian gland orifices induces meibomian gland dysfunction

PurposeTo determine pathological changes of meibomian glands (MGs) after transient exposure of the rat eyelid margin to alkali solution. MethodsFilter paper infiltrated with 1 N sodium hydroxide solution was applied to the eyelid margin of Sprague-Dawley rats for 30 s under general anesthesia, without touching the conjunctiva, after which the ocular surface and eyelid margin were examined by slit-lamp microscopy. In vivo confocal microscopy and stereomicroscopy were subsequently applied to observe MG morphology on day 5, day 10 and day 30 post alkali injury. Eyelid cross-sections were processed for H&E staining, Oil red O staining and immunofluorescent staining. ResultsAfter alkali injury, there was marked plugging of MG orifices, telangiectasia and hypertrophy of the eyelid margin, while corneal epithelium was intact at post-injury days 5 and 10. However, 30 days after alkali injury, mild corneal epithelial damage was observed. Degeneration of MG acini was observed at days 5 and became aggravated at days 10 and 30, along with MG duct dilation and acini loss. Oil red O staining showed lipid accumulation in the dilated duct. Inflammatory cell infiltration and the presence of apoptotic cells was seen in the MG loci 5 days post injury, but diminished at days 10 and 30. Cytokeratin 10 expression was increased in dilated duct, while cytokeratin 14, PPAR-γ, Ki67 and LRIG1 expression were decreased in the acini of injured loci. ConclusionsTransitory alkali exposure of the rat eyelid margin obstructs the MG orifice and induces pathological changes of MG dysfunction.

Abstract 3748: Epstein-Barr virus BART lncRNAs induce IKZF3/Aiolos to maintain EBV latency and promote tumorigenicity in nasopharyngeal carcinoma

Abstract Epstein-Barr virus (EBV) maintains latency in its associated tumors, including nasopharyngeal carcinoma (NPC), expressing very few viral proteins but abundant levels of noncoding RNAs (mainly EBERs and BARTs) in NPC cells. We found that IKZF3/Aiolos is a downstream target of BART lncRNAs in NPC cells. The functions of BART lncRNAs and IKZF3 in EBV latency and pathogenesis of NPC remain elusive. In this study, meta-analysis of eight EBV-associated studies showed that IKZF3/Aiolos was consistently upregulated in EBV-infected NPC cells. Our study further showed that transcription of IKZF3/Aiolos is highly upregulated in EBV-infected NPC cells and clinical tissue samples due to the action of EBV encoded BART lncRNAs, with IKZF3/Aiolos was expressed at higher levels in late stage (stage III & IV) NPC patients than in early stage (stage I & II) disease. Secondly, we found that IKZF3/Aiolos was upregulated in BART-activated NPC cells but downregulated in BART-knockdown NPC cells. In addition, this study further demonstrated that IKZF3/Aiolos modulates transcription of EBV BZLF1 and the cellular gene, LRIG1, to maintain EBV latency and promote NPC tumorigenesis in vitro and in vivo. Our results showed that the EBV lytic reactivator BZLF1 was upregulated in IKZF3/Aiolos knockout or Aiolos inhibitor-treated NPC cells. ChIP-qPCR, immunoprecipitation and immunofluorescence analyses further revealed that IKZF3/Aiolos induces H3K27 deacetylation to silence expression of BZLF1 and maintain EBV latency in NPC cells. Moreover, functional analyses and western blotting showed that IKZF3/Aiolos inhibited the tumor suppressor, LRIG1, to upregulate expression and phosphorylation of the cellular proto-oncogene Erb-B2 for NPC pathogenesis. Sphere- and colony-formation assays demonstrated that IKZF3/Aiolos enhances growth of NPC cells in vitro, and in vivo experiments further showed that IKZF3/Aiolos promotes tumorigenicity of NPC cells in NOD mice. Mechanically, we found that the expression of EBV lytic reactivator BZLF1 and cellular LRIG1 were negatively regulated by BART lncRNA/IKZF3/Aiolos regulatory machinery in NPC cells, while Erb-B2 was positively regulated, which indicated that IKZF3/Aiolos is a new biomarker for NPC and may lead to the development of novel diagnostic tests and treatments for NPC. Citation Format: Songtao He, Jiayan Liu, Bobo Wing-Yee Mok, Sai Wah Tsao, Honglin Chen. Epstein-Barr virus BART lncRNAs induce IKZF3/Aiolos to maintain EBV latency and promote tumorigenicity in nasopharyngeal carcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3748.

Glioma-derived LRIG3 interacts with NETO2 in tumor-associated macrophages to modulate microenvironment and suppress tumor growth

Tumor-associated macrophages (TAMs) account for 30–50% of glioma microenvironment. The interaction between glioma tumor cells and TAMs can promote tumor progression, but the intrinsic mechanisms remain unclear. Herein, we reported that soluble LRIG3 (sLRIG3) derived from glioma tumor cells can block the M2 polarization of TAMs via interacting with NETO2, thus suppressing GBM malignant progression. The expression or activity of ADAM17 in glioma cells was positively correlated with the expression of sLRIG3 in cell supernatant. Soluble LRIG3 can suppress the M2-like polarity transformation of TAMs and inhibit the growth of tumor. High expression of LRIG3 predicts a good prognosis in patients with glioma. Mass spectrometry and Co-immunoprecipitation showed that sLRIG3 interacts with the CUB1 domain of NETO2 in TAMs. Silencing or knockout of NETO2 could block the effect of sLRIG3, which inhibited the M2-like polarity transformation of TAMs and promoted GBM tumor growth. However, overexpressing His-target NETO2 with CUB1 deletion mutation does not fully recover the suppressive effects of sLRIG3 on the TAM M2-polarization in NETO2-Knockout TAMs. Our study revealed vital molecular crosstalk between GBM tumor cells and TAMs. Glioma cells mediated the M2 polarization of TAM through the sLRIG3-NETO2 pathway and inhibited the progression of GBM, suggesting that sLRIG3-NETO2 may be a potential target for GBM treatment.

Lrig1 expression identifies quiescent stem cells in the ventricular-subventricular zone from postnatal development to adulthood and limits their persistent hyperproliferation

BackgroundWe previously identified Leucine-rich repeats and immunoglobulin-like domains 1 (Lrig1) as a marker of long-term neurogenic stem cells in the lateral wall of the adult mouse brain. The morphology of the stem cells thus identified differed from the canonical B1 type stem cells, raising a question about their cellular origin. Thus, we investigated the development of these stem cells in the postnatal and juvenile brain. Furthermore, because Lrig1 is a known regulator of quiescence, we also investigated the effect(s) of its deletion on the cellular proliferation in the lateral wall.MethodsTo observe the development of the Lrig1-lineage stem cells, genetic inducible fate mapping studies in combination with thymidine analog administration were conducted using a previously published Lrig1T2A-iCreERT2 mouse line. To identify the long-term consequence(s) of Lrig1 germline deletion, old Lrig1 knock-out mice were generated using two different Lrig1 null alleles in the C57BL/6J background. The lateral walls from these mice were analyzed using an optimized whole mount immunofluorescence protocol and confocal microscopy.ResultsWe observed the Lrig1-lineage labeled cells with morphologies consistent with neurogenic stem cell identity in postnatal, juvenile, and adult mouse brains. Interestingly, when induced at postnatal or juvenile ages, morphologically distinct cells were revealed, including cells with the canonical B1 type stem cell morphology. Almost all of the presumptive stem cells labeled were non-proliferative at these ages. In the old Lrig1 germline knock-out mice, increased proliferation was observed compared to wildtype littermates without concomitant increase in apoptosis.ConclusionsOnce set aside during embryogenesis, the Lrig1-lineage stem cells remain largely quiescent during postnatal and juvenile development until activation in adult age. The absence of premature proliferative exhaustion in the Lrig1 knock-out stem cell niche during aging is likely due to a complex cascade of effects on the adult stem cell pool. Thus, we suggest that the adult stem cell pool size may be genetically constrained via Lrig1.

MicroRNA-218-5p accelerates malignant behaviors of breast cancer through LRIG1

ObjectiveMicroRNAs play crucial roles in the pathogenesis of cancers. MiRNA-218-5p may act as either an oncogene or a tumor suppressor, but its role in the pathogenesis of Breast Cancer (BC) remains unclear. MethodsInfiltrative breast ductal carcinoma as well as corresponding adjacent normal samples were collected from 30 patients. Mimics and inhibitors of miRNA-218-5p or corresponding negative controls were transfected into BC cells. miRNA-218-5p expression was detected by quantitative PCR. The effects of miRNA-218-5p on the malignant behaviors of BC were assessed. Dual-luciferase reporter assay was employed to evaluate the binding of miRNA-218-5p to LRIG1. ResultsBC tissues showed higher miRNA-218-5p expression as compared to the adjacent normal tissues. Ectopic miRNA-218-5p expression accelerated the cell cycle, cell growth and migration of BC, while repressed cell apoptosis. Interestingly, ectopic miRNA-218-5p expression down-regulated LRIG1 expression, and miRNA-218-5p could bind to LRIG1. Also, our study indicated that miRNA-218-5p up-regulated ErbB2 and EGFR expression by targeting LRIG1, suggesting that the LRIG1-mediated signaling pathway contributed to the pro-tumor effects of miRNA-218-5p on BC. ConclusionMiRNA-218-5p up-regulates ErbB2 and EGFR expression by suppressing LRIG1 expression, thus promoting the malignant behaviors of BC. miRNA-218-5p may exert a pro-tumor effect on BC and serve as a therapeutic target for BC treatment.

LRIG2 promotes glioblastoma progression by modulating innate antitumor immunity through macrophage infiltration and polarization

BackgroundGlioblastoma (GBM) is the most common malignant brain tumor with poor clinical outcomes. Immunotherapy has recently been an attractive and promising treatment of extracranial malignancies, however, most of clinical trials...

Evaluation of LRIG1 Expression in Larynx Pathologies

Objective: Studies have been performed on many biomolecules to determine the prognosis of LSCC and predict the course of the disease. However, a molecular marker that can be used clinically has not yet been found. Therefore, in this study, we aimed to investigate the expression levels of LRIG 1 in laryngeal cancer. Materials and Methods: In our study, 219 cases who underwent surgery due to LSCC and 88 randomly selected patients whose pathologic result were benign and premalignant lesions in Marmara University Pendik Education and Research Hospital between 2003 and 2018 were analyzed. Patients’ data were obtained from the medical records. The tissue microarray method was used to evaluate specimens. Results: There was a statistically significant difference between the tumor differentiation, diagnosis, and the expression of LRIG1 (respectively p=0.045, p<0.001). Also, an increase in the degree of dysplasia in premalignant lesions correlates with a decrease in LRIG1 expression (p=0.015). Conclusion: Our findings suggest that LRIG1 plays a role in the early tumorigenesis of LSCC. Therefore, LRIG1 can be a target molecule for treatment approaches. However, LRIG1 was not correlated with overall survival of the LSCC.

PO-1143 High LRIG1 expression is associated with better prognosis for patients with Glioblastoma Multiforme

PO-1143 High LRIG1 expression is associated with better prognosis for patients with Glioblastoma Multiforme

Aberrant promoter methylation contributes to LRIG1 silencing in basal/triple-negative breast cancer

BackgroundLRIG1, the founding member of the LRIG (leucine-rich repeat and immunoglobulin-like domain) family of transmembrane proteins, is a negative regulator of receptor tyrosine kinases and a tumour suppressor. Decreased LRIG1 expression is consistently observed in cancer, across diverse tumour types, and is linked to poor patient prognosis. However, mechanisms by which LRIG1 is repressed are not fully understood. Silencing of LRIG1 through promoter CpG island methylation has been reported in colorectal and cervical cancer but studies in breast cancer remain limited.MethodsIn silico analysis of human breast cancer patient data were used to demonstrate a correlation between DNA methylation and LRIG1 silencing in basal/triple-negative breast cancer, and its impact on patient survival. LRIG1 gene expression, protein abundance, and methylation enrichment were examined by quantitative reverse-transcription PCR, immunoblotting, and methylation immunoprecipitation, respectively, in breast cancer cell lines in vitro. We examined the impact of global demethylation on LRIG1 expression and methylation enrichment using 5-aza-2’-deoxycytidine. We also examined the effects of targeted demethylation of the LRIG1 CpG island, and transcriptional activation of LRIG1 expression, using the RNA guided deadCas9 transactivation system.ResultsAcross breast cancer subtypes, LRIG1 expression is lowest in the basal/triple-negative subtype so we investigated whether differential methylation may contribute to this. Indeed, we find that LRIG1 CpG island methylation is most prominent in basal/triple-negative cell lines and patient samples. Use of the global demethylating agent 5-aza-2’-deoxycytidine decreases methylation leading to increased LRIG1 transcript expression in basal/triple-negative cell lines, while having no effect on LRIG1 expression in luminal/ER-positive cell lines. Using a CRISPR/deadCas9 (dCas9)-based targeting approach, we demonstrate that TET1-mediated demethylation (Tet1-dCas9) along with VP64-mediated transcriptional activation (VP64-dCas9) at the CpG island, increased endogenous LRIG1 expression in basal/triple-negative breast cancer cells, without transcriptional upregulation at predicted off-target sites. Activation of LRIG1 by the dCas9 transactivation system significantly increased LRIG1 protein abundance, reduced site-specific methylation, and reduced cancer cell viability. Our findings suggest that CRISPR-mediated targeted activation may be a feasible way to restore LRIG1 expression in cancer.ConclusionsOur study contributes novel insight into mechanisms which repress LRIG1 in triple-negative breast cancer and demonstrates for the first time that targeted de-repression of LRIG1 in cancer cells is possible. Understanding the epigenetic mechanisms associated with repression of tumour suppressor genes holds potential for the advancement of therapeutic approaches.

Prognostic significance of LRIG2 and LRIG3 proteins in urothelial bladder carcinoma

ABSTRACT Bladder carcinoma is the second most frequent cancer in Egyptian males. Leucine-rich and immunoglobulin-like domains (LRIGs) are usually dysregulated in various human tumors. The aim of this study is to explore the immunohistochemical expression of LRIG2 and LRIG3 in urothelial bladder carcinoma (UBC) and their relationship to patients clinicopathological data including survival. The study cohort included 79 UBC cases (14 non muscle invasive (NMI) and 65 muscle invasive (MI)). We assessed the associations of LRIG2 and LRIG3 expression with clinicopathological data, as well as progression-free and overall survival. Most of studied cases (>50%) express LRIG2 and LRIG3. Statistically significant association was observed between positivity for LRIG3 and muscle invasion (P = 0.001), high grade (P = 0.03), and female gender (P = 0.02). Moreover, positive LRIG2 staining was associated with early stage (T2) (P = 0.03), lymphovascular invasion (P = 0.004), and tendency to non-muscle invasive stage (P = 0.07). Grouping of cases according to positivity/negativity of both markers showed that cases with dual positivity for both proteins are associated with muscle invasion (P = 0.001) and paradoxically with prolonged overall survival (P = 0.037). We conclude that although the association of LRIG3 with MI and high-grade tumors, its expression is related to better survival. LRIG3 has the dominant role even if it coexists with LRIG2. The role of LRIG2 remains to be further investigated.

Lrig1 and Lrig3 cooperate to control Ret receptor signaling, sensory axonal growth and epidermal innervation.

Negative feedback loops represent a regulatory mechanism that guarantees that signaling thresholds are compatible with a physiological response. Previously, we established that Lrig1 acts through this mechanism to inhibit Ret activity. However, it is unclear whether other Lrig family members play similar roles. Here, we show that Lrig1 and Lrig3 are co-expressed in Ret-positive mouse dorsal root ganglion (DRG) neurons. Lrig3, like Lrig1, interacts with Ret and inhibits GDNF/Ret signaling. Treatment of DRG neurons with GDNF ligands induces a significant increase in the expression of Lrig1 and Lrig3. Our findings show that, whereas a single deletion of either Lrig1 or Lrig3 fails to promote Ret-mediated axonal growth, haploinsufficiency of Lrig1 in Lrig3 mutants significantly potentiates Ret signaling and axonal growth of DRG neurons in response to GDNF ligands. We observe that Lrig1 and Lrig3 act redundantly to ensure proper cutaneous innervation of nonpeptidergic axons and behavioral sensitivity to cold, which correlates with a significant increase in the expression of the cold-responsive channel TrpA1. Together, our findings provide insights into the in vivo functions through which Lrig genes control morphology, connectivity and function in sensory neurons.

Hsa_circ_0000006 induces tumorigenesis through miR-361-3p targeting immunoglobulin-like domains protein 1 (LRIG1) in osteosarcoma.

BackgroundOsteosarcoma (OS) is considered to be the most highly prevalent bone tumor. In the progression of different human cancers, the role of circular RNAs (circRNAs) has been extensively studied. Microarray analysis has indicated that hsa_circ_0000006 expression was lower in OS, but the mechanism of hsa_circ_0000006 in regulating the progression of OS remains elusive.MethodsThe expression of cancer-related genes at the transcriptional and translational levels was assessed by RT-qPCR and western blotting (WB). Colony formation and Cell Counting Kit-8 (CCK-8) assays were used to evaluate the proliferative potential of cells. The transwell assay was used to examine the invasive and migratory potential of cells. Furthermore, dual-luciferase reporter (DLR) and RNA pull-down assays were performed for the validation of the targeting sites of hsa_circ_0000006, miR-361-3p, and the 3'-untranslated region (3'-UTR) of immunoglobulin-like domains protein 1 (LRIG1) mRNA. Moreover, the protein levels of epithelial-to-mesenchymal transition (EMT) markers were analyzed by WB.ResultsThe expression of hsa_circ_0000006 and LRIG1 were found to be down-regulated in OS tissues and cells, while miR-361-3p was up-regulated. Knockdown of hsa_circ_0000006 promoted the progression and development of OS, as well as EMT. Furthermore, hsa_circ_0000006 was revealed as a sponge of miR-361-3p, which negatively regulates miR-361-3p expression. LRIG1 was found to be an miR-361-3p target. In OS cells, the LRIG1 expression level was decreased, with elevated expression of miR-361-3p. Advanced studies demonstrated that hsa_circ_0000006 regulates LRIG1 expression through sponging miR-361-3p, then promotes the tumorigenesis of OS.Conclusionshsa_circ_0000006 is associated with the progression and development of OS through miR-361-3p by target LRIG1, which is a significant biomarker and effective therapeutic target for patients with OS.

LRIG1 is a positive prognostic marker in Merkel cell carcinoma and Merkel cell carcinoma expresses epithelial stem cell markers

Merkel cell carcinoma (MCC) is a rare and aggressive neuroendocrine malignancy of the skin. The cell of origin of MCC is thus far unknown and proposed cells of origin include Merkel cells, pro-/pre- or pre-B cells, epithelial stem cells, and dermal stem cells. In this study, we aimed to shed further light on the possibility that a subset of MCC tumors arise from epithelial stem cells of the skin by examining the expression of hair follicle and epidermal stem cell markers in MCC and normal human skin. We also aimed to elucidate any correlation between the expression of these markers and tumor Merkel cell polyomavirus (MCPyV) status or other clinicopathological characteristics or patient survival. Expression of CK19, SOX9, LGR5, and LRIG1 in MCC and normal human skin was studied by immunohistochemistry, and the staining patterns or intensities were statistically correlated with patient, tumor, MCPyV, and survival parameters. In a cohort of 137 cases of MCC, we observed dot-like immunoexpression of CK19 in 30 cases (22.1%) and homogeneous expression in 103 cases (75.7%). We also observed positive immunoexpression of SOX9 in 21 cases (15.3%), LGR5 in 118 cases (86.1%), and LRIG1 in 117 cases (86.0%). Immunoexpression of LRIG1 was found to correlate with better overall and MCC-specific survival. We observed frequent immunoexpression of several hair follicle and epidermal stem cell markers in MCC and found LRIG1 to be a positive prognostic marker in MCC.

Abstract 2360: Identifying differentially expressed miRNAs in CRC stem cell subpopulations

Abstract Our research goal is to discover the molecular mechanisms behind tumor heterogeneity in CRC. We previously showed that CRCs contain multiple subpopulations of cancer stem cells (CSCs) which may explain the occurrence tumor heterogeneity and resistance to treatment. To determine how CSC sub-populations might arise, we are studying miRNA expression in CRC SCs. MicroRNAs are known to regulate SC phenotype and are found to be dysregulated in many cancers. Hypothesis: Tumor heterogeneity results from existence of multiple CSC subpopulations that are regulated by distinct miRNAs. Accordingly, we are using bioinformatics and miRNA profiling to identify miRNAs that target SC genes in CRCs. Indeed, our miRNA expression profiling of normal and malignant ALDH+ human colonic SCs showed that miRNA92a targets the SC gene LRIG1 and upregulation of miRNA92a leads to decreased LRIG1 expression. We also discovered that miRNA23b targets the SC gene LGR5 and miRNA23b is upregulated in ALDH+ CSCs. We have identified several other candidate miRNAs that are predicted to target CD166, ALDH1A1, BMI1, LRG5, and LRIG1 SC genes. We are currently in the process of validating whether these miRNAs contribute to emergence of specific CSC sub-populations in CRCs. Thus, identifying miRNAs that regulate CSC subpopulations should provide new strategies to modulate CSC composition in order to sensitize tumors to treatments. Citation Format: Victoria A. Stark, Vignesh Viswanathan, Caroline O. Facey, Lynn M. Opdenaker, Bruce M. Boman. Identifying differentially expressed miRNAs in CRC stem cell subpopulations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2360.

Member Domain 3 (LRIG3) Activates Hypoxia-Inducible Factor-1 α/Vascular Endothelial Growth Factor (HIF-1α/VEGF) Pathway to Inhibit the Growth of Bone Marrow Mesenchymal Stem Cells in Glioma

Member domain 3 (LRIG3) of the LRIG gene family is down-regulated in several cancers. However, its role in bone marrow mesenchymal stem cells (BMSCs) in gliomas and the related mechanisms is unknown. The qRT-PCR assessed LRIG3 mRNA level. Rat BMSCs were randomly assigned into glioma group (BMSCs cultured in glioma microenvironment); LRIG3 overexpression group; and si-LRIG3 inhibitor group followed by analysis of LRIG3 expression, cell proliferation, PCNA and Ki-67 apoptosis, TNF-α; and HIF-1α/VEGF mRNA level. LRIG3 mRNA expression was decreased in gliomas patients (P < 0.05). BMSCs cultured in glioma microenvironment showed decreased LRIG3, increased cell proliferation, decreased PCNA, Ki-67 and TNF-α secretion as well as elevated HIF-1α and VEGF level (P < 0.05). Transfection of LRIG3 siRNA further promoted the above changes. Conversely, LRIG3 plasmid transfection significantly promoted its expression in glioma BMSCs (P < 0.05), inhibited cell proliferation, promoted PCNA, Ki-67, and TNF-α secretion, and increased HIF-1α and VEGF level (P < 0.05). LRIG3 in rat BMSCs cultured in the glioma microenvironment is decreased. Down-regulation of LRIG3 inhibits TNF-α secretion by activating HIF-1α/VEGF pathway regulating BMSCs proliferation and apoptosis.

CircRNA LRIG3 knockdown inhibits hepatocellular carcinoma progression by regulating miR-223-3p and MAPK/ERK pathway

IntroductionEmerging evidence suggests that circular RNAs (circRNAs) play critical roles in tumorigenesis. However, the roles and molecular mechanisms of circRNA leucine-rich repeat immunoglobulin domain-containing protein 3 (circ_LRIG3) in hepatocellular carcinoma (HCC) has not been investigated.Material and methodsThe expression levels of circ_LRIG3, miR-223-3p, and mitogen-activated protein kinase kinase 6 (MAP2K6) were determined by qRT-PCR. Flow cytometry was applied to determine the cell cycle distribution and apoptosis. Cell proliferation, migration and invasion were assessed by MTT, colony formation, and transwell assays. Western blot assay was employed to measure the protein levels of the snail, E-cadherin, MAP2K6, mitogen-activated protein kinase (MAPK), phospho-MAPK (p-MAPK), extracellular signal-regulated kinases (ERKs), and phospho-ERKs (p- ERKs). The relationship between miR-223-3p and circ_LRIG3 or MAP2K6 was predicted by bioinformatics tools and verified by dual-luciferase reporter assay. A xenograft tumor model was established to confirm the functions of circ_LRIG3 in vivo.ResultsCirc_LRIG3 and MAP2K6 expression were enhanced while miR-223-3p abundance was reduced in HCC tissues and cells. Knockdown of circ_LRIG3 inhibited cell proliferation, metastasis, and increasing apoptosis. MiR-223-3p was a target of circ_LRIG3, and its downregulation reversed the inhibitory effect of circ_LRIG3 knockdown on the progression of HCC cells. Moreover, MAP2K6 could bind to miR-223-3p, and MAP2K6 upregulation also abolished the suppressive impact of circ_LRIG3 interference on progression of HCC cells. Additionally, the silence of circ_LRIG3 suppressed the activation of the MAPK/ERK pathway and tumor growth by upregulating miR-223-3p and downregulating MAP2K6.ConclusionsCirc_LRIG3 knockdown inhibited HCC progression through regulating miR-223-3p/MAP2K6 axis and inactivating MAPK/ERK pathway.