Targeting KDM4A epigenetically activates tumor-cell-intrinsic immunity by inducing DNA replication stress.

Developing strategies to activate tumor-cell-intrinsic immune response is critical for improving tumor immunotherapy by exploiting tumor vulnerability. KDM4A, as a histone H3 lysine 9 trimethylation (H3K9me3) demethylase, has been found to play a critical role in squamous cell carcinoma (SCC) growth and metastasis. Here we report that KDM4A inhibition promoted heterochromatin compaction and induced DNA replication stress, which elicited antitumor immunity in SCC. Mechanistically, KDM4A inhibition promoted the formation of liquid-like HP1γ puncta on heterochromatin and stall DNA replication, which activated tumor-cell-intrinsic cGAS-STING signaling through replication-stress-induced cytosolic DNA accumulation. Moreover, KDM4A inhibition collaborated with PD1 blockade to inhibit SCC growth and metastasis by recruiting and activating CD8+ T cells. In vivo lineage tracing demonstrated that KDM4A inhibition plus PD1 blockade efficiently eliminated cancer stem cells. Altogether, our results demonstrate that targeting KDM4A can activate anti-tumor immunity and enable PD1 blockade immunotherapy by aggravating replication stress in SCC cells.

Compressive force-induced LincRNA-p21 inhibits mineralization of cementoblasts by impeding autophagy.

The mineralization capability of cementoblasts is the foundation for repairing orthodontic treatment-induced root resorption. It is essential to investigate the regulatory mechanism of mineralization in cementoblasts under mechanical compression to improve orthodontic therapy. Autophagy has a protective role in maintaining cell homeostasis under environmental stress and was reported to be involved in the mineralization process. Long noncoding RNAs are important regulators of biological processes, but their functions in compressed cementoblasts during orthodontic tooth movement remain unclear. In this study, we showed that compressive force downregulated the expression of mineralization-related markers. LincRNA-p21 was strongly enhanced by compressive force. Overexpression of lincRNA-p21 downregulated the expression of mineralization-related markers, while knockdown of lincRNA-p21 reversed the compressive force-induced decrease in mineralization. Furthermore, we found that autophagy was impeded in compressed cementoblasts. Then, overexpression of lincRNA-p21 decreased autophagic activity, while knockdown of lincRNA-p21 reversed the autophagic process decreased by mechanical compression. However, the autophagy inhibitor 3-methyladenine abolished the lincRNA-p21 knockdown-promoted mineralization, and the autophagy activator rapamycin rescued the mineralization inhibited by lincRNA-p21 overexpression. Mechanistically, the direct binding between lincRNA-p21 and FoxO3 blocked the expression of autophagy-related genes. In a mouse orthodontic tooth movement model, knockdown of lincRNA-p21 rescued the impeded autophagic process in cementoblasts, enhanced cementogenesis, and alleviated orthodontic force-induced root resorption. Overall, compressive force-induced lincRNA-p21 inhibits the mineralization capability of cementoblasts by impeding the autophagic process.

Compressive force regulates orthodontic tooth movement via activating the NLRP3 inflammasome.

Mechanical stress regulates various cellular functions like cell inflammation, immune responses, proliferation, and differentiation to maintain tissue homeostasis. However, the impact of mechanical signals on macrophages and the underlying mechanisms by which mechanical force regulates bone remodeling during orthodontic tooth movement remain unclear. NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome has been reported to promote osteoclastic differentiation to regulate alveolar bone resorption. But the relationship between the compressive force and NLRP3 inflammasome in macrophages remains unknown. In this study, immunohistochemical staining results showed elevated expression of NLRP3 and interleukin-1ß, as well as an increased number of macrophages expressing NLRP3, on the compression side of the periodontal tissues, after force application for 7 days. Furthermore, the number of tartrate-resistant acid phosphatase-positive osteoclasts, and the mRNA and protein expression levels of osteoclast-related genes in the periodontal tissue decreased in the Nlrp3-/- mice compared to the WT mice group after orthodontic movement. In vitro mechanical force activates the NLRP3 inflammasome and inhibits autophagy. Intraperitoneal injection of the autophagy inhibitor 3-methyladenine in Nlrp3-/- mice promoted orthodontic tooth movement. This result indicates that the absence of NLRP3 inflammasome activation can be partially compensated for by autophagy inhibitors. Mechanistically, force-induced activation of the NLRP3 inflammasome in macrophages via the cGAS/P2X7R axis. In conclusion, compressive force regulates orthodontic tooth movement via activating the NLRP3 inflammasome.

Long non-coding RNA SNHG5 mediates periodontal inflammation through the NF-κB signalling pathway.

AIM: We investigated the role of long non-coding RNAs and small nucleolar RNA host gene 5 (SNHG5) in the pathogenesis of periodontitis. MATERIALS AND METHODS: A ligature-induced periodontitis mouse model was established, and gingival tissues were collected from patients with periodontitis and healthy controls. Inflammatory cytokines were detected using quantitative reverse transcription-polymerase chain reaction and western blotting analyses. Direct interactions between SNHG5 and p65 were detected by RNA pull-down and RNA immunoprecipitation assays. Micro-computed tomography, haematoxylin and eosin staining, and immunohistochemical staining were used to measure periodontal bone loss. RESULTS: SNHG5 expression was down-regulated in human and mouse periodontal tissues compared to that in the healthy controls. In vitro experiments demonstrated that SNHG5 significantly ameliorated tumour necrosis factor α-induced inflammation. Mechanistically, SNHG5 directly binds to the nuclear factor-kappa B (NF-κB) p65 subunit and inhibits its translocation, thereby suppressing the NF-κB signalling pathway activation and reducing the nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing three inflammasome expression. Locally injecting si-SNHG5 aggravated the periodontal destruction. CONCLUSION: This study revealed that SNHG5 mediates periodontal inflammation through the NF-κB signalling pathway, providing a potential therapeutic target for periodontitis treatment.

The emerging roles of circular RNAs in regulating the fate of stem cells.

Circular RNAs(circRNAs) are a large family of RNAs shaping covalently closed ring-like molecules and have become a hotspot with thousands of newly published studies. Stem cells are undifferentiated cells and have great potential in medical treatment due to their self-renewal ability and differentiation capacity. Abundant researches have unveiled that circRNAs have unique expression profile during the differentiation of stem cells and could serve as promising biomarkers of these cells. There are key circRNAs relevant to the differentiation, proliferation, and apoptosis of stem cells with certain mechanisms such as sponging miRNAs, interacting with proteins, and interfering mRNA translation. Moreover, several circRNAs have joined in the interplay between stem cells and lymphocytes. Our review will shed lights on the emerging roles of circRNAs in regulating the fate of diverse stem cells.

Mechanical force inhibited hPDLSCs proliferation with the downregulation of MIR31HG via DNA methylation.

OBJECTIVE: This study aimed to investigate how mechanical force affects the proliferation of human periodontal ligament stem cells (hPDLSCs). METHODS: CCK-8 assays and staining of ki67 were performed to evaluate hPDLSCs proliferation. qRT-PCR, ELISA, or Western blot analysis were used to measure the expression levels of interleukin (IL)-6, miR-31 host gene (MIR31HG), DNA methyltransferase 1 (DNMT1), and DNA methyltransferase 3B (DNMT3B). Dual-luciferase reporter assays and chromatin immunoprecipitation (ChIP) assays were conducted to determine whether MIR31HG was targeted by DNMT1 and DNMT3B. MassARRAY mass spectrometry was used to quantify DNA methylation levels of the MIR31HG promoter. RESULTS: Mechanical force inhibited hPDLSCs proliferation with the downregulation of MIR31HG and upregulation of IL-6, DNMT1 and DNMT3B. Knockdown of MIR31HG suppressed hPDLSCs proliferation, and knockdown of DNMT1 or DNMT3B reversed mechanical force-induced downregulation of MIR31HG. Dual-luciferase and ChIP assays revealed DNMT1 and DNMT3B bound MIR31HG promoter in the region 1,015 bp upstream of the transcriptional start site. Treatment with 5'-aca-2'-deoxycytidine downregulated DNA methylation level in MIR31HG gene promoter, while mechanical force promoted the methylation of MIR31HG gene promoter. CONCLUSIONS: These findings elucidated how mechanical force affects proliferation via MIR31HG in hPDLSCs, providing clues for possible MIR31HG-based orthodontic therapeutic approaches.

Exosomes derived from maxillary BMSCs enhanced the osteogenesis in iliac BMSCs.

OBJECTIVE: Secondary alveolar bone grafting is an essential part in the treatment of alveolar cleft deformity. Autologous iliac bone is the most favorable grafting source. However, the factors regulating postoperative bone formation are unclear. Investigations are needed to found whether the alveolar bone niche and bone marrow mesenchymal stem cells (BMSCs) derived from the jaw bone (BMSCs-J) affected the osteogenesis of BMSCs from the ilium (BMSCs-I). MATERIALS AND METHODS: The effect of BMSCs-J on BMSCs-I was investigated using a co-culture model. The exosomes were purified by sequential centrifugation. The osteoblastic differentiation of BMSCs was analyzed in vitro and in vivo. RESULTS: Co-culture with BMSCs-J increased the alkaline phosphatase (ALP) activity, Alizarin Red S (ARS) staining, and osteogenic gene expression in BMSCs-I. Transmission electron microscopy and nanoparticle tracking analysis verified the presence of exosomes in the culture supernatants of BMSCs. Exosomes secreted by BMSCs-J enhanced the ALP activity, ARS staining, osteogenic gene expression of BMSCs-I in vitro, and new bone formation in vivo. Blocking the secretion of exosomes using siRNA for Rab27a inhibited the effect of BMSCs-J. CONCLUSION: Exosomes played a role in the interaction between BMSCs-J and BMSCs-I, thereby leading to the enhanced osteogenic capacity of BMSCs-I and bone formation.

Genome-wide DNA-methylation profiles in human bone marrow mesenchymal stem cells on titanium surfaces.

The characteristics of titanium (Ti) have been shown to influence dental implant fixation. Treatment of surfaces using the sandblasted, large-grit, acid-etched (SLA) method is widely used to provide effective osseointegration. However, the DNA methylation-associated mechanism by which SLA surface treatment affects osseointegration of human bone marrow mesenchymal stem cells (hBMSCs) remains elusive. Genome-wide methylation profiling of hBMSCs on SLA-treated and machined smooth Ti was performed using Illumina Infinium Methylation EPIC BeadChip at day 7 of osteogenic induction. In total, 2,846 CpG sites were differentially methylated in the SLA group compared with the machined group. Of these sites, 1,651 (covering 1,066 genes) were significantly hypermethylated and 1,195 (covering 775 genes) were significantly hypomethylated. Thirty significant enrichment pathways were observed, with Wnt signaling being the most significant. mRNA expression was identified by microarray and combined with DNA-methylation profiles. Thirty-seven genes displayed negative association between mRNA expression and DNA-methylation level, with the osteogenesis-related genes insulin-like growth factor 2 (IGF2) and carboxypeptidase X, M14 Family Member 2 (CPXM2) showing significant up-regulation and down-regulation, respectively. In summary, our results demonstrate differences between SLA-treated and machined surfaces in their effects on genome-wide DNA methylation and enrichment of osteogenic pathways in hBMSCs. We provide novel insights into genes and pathways affected by SLA treatment in hBMSCs at the molecular level.

Subgingival Microbial Changes During the First 3 Months of Fixed Appliance Treatment in Female Adult Patients.

Although periodontal diseases during fixed appliance treatment are a common issue, few studies have focused on the clinical and microbial factors associated with orthodontic appliances. Hence, we investigated changes in the subgingival microbial community and their association with periodontal changes at the early stage of fixed appliance treatment. Subgingival plaques from ten female patients with fixed appliances were obtained at three time points: before, 1 month and 3 months after the placement of the brackets (T0, T1 and T2). The 16S rRNA gene sequencing was used to analyze the microbial community of the subgingival plaque. The Plaque Index (PI) and Gingival Bleeding Index (GBI) were also recorded. The GBI significantly increased at T2, and the PI showed a temporary increase without a significant difference. The alpha diversity indices were stable. However, the beta diversity was significantly higher at T2 compared to T0 and T1. The relative abundance of core microbiomes at the genus level was relatively stable. Four periodontal pathogens at the species level, including Prevotella intermedia (Pi), Campylobacer rectus (Cr), Fusobacterium nucleatum (Fn), and Treponema denticola (Td), increased without significant differences. The subgingival microbial community affected by fixed appliance treatment might cause transient mild gingival inflammation.

The long non-coding RNA landscape of periodontal ligament stem cells subjected to compressive force.

OBJECTIVE: The role of long non-coding ribonucleic acids (lncRNAs) during orthodontic tooth movement remains unclear. We explored the lncRNA landscape of periodontal ligament stem cells (PDLSCs) subjected to compressive force. MATERIALS AND METHODS: PDLSCs were subjected to static compressive stress (2 g/cm2) for 12 hours. Total RNA was then extracted and sequenced to measure changes in lncRNA and messenger RNA (mRNA) expression levels. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to validate the expression levels of certain lncRNAs. Differential expression analysis as well as Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were also performed. RESULTS: In total, 90 lncRNAs and 519 mRNAs were differentially expressed in PDLSCs under compressive stress. Of the lncRNAs, 72 were upregulated and 18 downregulated. The levels of eight lncRNAs of interest (FER1L4, HIF1A-AS2, MIAT, NEAT1, ADAMTS9-AS2, LUCAT1, MIR31HG, and DHFRP1) were measured via qRT-PCR, and the results were found to be consistent with those of RNA sequencing. GO and KEGG pathway analyses showed that a wide range of biological functions were expressed during compressive loading; most differentially expressed genes were involved in extracellular matrix organization, collagen fibril organization, and the cellular response to hypoxia. CONCLUSIONS: The lncRNA expression profile was significantly altered in PDLSCs subjected to compressive stress. These findings expand our understanding of molecular regulation in the mechanoresponse of PDLSCs.

Inhibition of lncRNA MIR31HG Promotes Osteogenic Differentiation of Human Adipose-Derived Stem Cells.

Osteogenic differentiation and bone formation is suppressed under condition of inflammation induced by proinflammation cytokines. A number of studies indicate miRNAs play a significant role in tumor necrosis factor-α-induced inhibition of bone formation, but whether long non-coding RNAs are also involved in this process remains unknown. In this study, we evaluated the role of MIR31HG in osteogenesis of human adipose-derived stem cells (hASCs) in vitro and in vivo. The results suggested that knockdown of MIR31HG not only significantly promoted osteogenic differentiation, but also dramatically overcame the inflammation-induced inhibition of osteogenesis in hASCs. Mechanistically, we found MIR31HG regulated bone formation and inflammation via interacting with NF-κB. The p65 subunit bound to the MIR31HG promoter and promoted MIR31HG expression. In turn, MIR31HG directly interacted with IκBα and participated in NF-κB activation, which builds a regulatory circuitry with NF-κB. Targeting this MIR31HG-NF-κB regulatory loop may be helpful to improve the osteogenic capacity of hASCs under inflammatory microenvironment in bone tissue engineering. Stem Cells 2016;34:2707-2720.

Long non-coding RNA MEG3 inhibits adipogenesis and promotes osteogenesis of human adipose-derived mesenchymal stem cells via miR-140-5p.

lncRNAs are an emerging class of regulators involved in multiple biological processes. MEG3, an lncRNA, acts as a tumor suppressor, has been reported to be linked with osteogenic differentiation of MSCs. However, limited knowledge is available concerning the roles of MEG3 in the multilineage differentiation of hASCs. The current study demonstrated that MEG3 was downregulated during adipogenesis and upregulated during osteogenesis of hASCs. Further functional analysis showed that knockdown of MEG3 promoted adipogenic differentiation, whereas inhibited osteogenic differentiation of hASCs. Mechanically, MEG3 may execute its role via regulating miR-140-5p. Moreover, miR-140-5p was upregulated during adipogenesis and downregulated during osteogenesis in hASCs, which was negatively correlated with MEG3. In conclusion, MEG3 participated in the balance of adipogenic and osteogenic differentiation of hASCs, and the mechanism may be through regulating miR-140-5p.

Long non-coding RNA MIAT knockdown promotes osteogenic differentiation of human adipose-derived stem cells.

Recently, long non-coding RNAs (lncRNAs) have emerged as critical players in gene regulation for multiple biological processes. However, their roles and functions in human adipose-derived stem cells (hASCs) differentiation remain unclear. In the present study, we investigated the role of lncRNA myocardial infarction-associated transcript (MIAT) in the osteogenic differentiation of hASCs. We found that the expression of MIAT was downregulated in a time-dependent manner during hASCs osteoinduction. MIAT knockdown promoted osteogenic differentiation of hASCs both in vitro and in vivo. Moreover, MIAT expression was increased upon tumor necrosis factor-α treatment and MIAT knockdown reversed the negative effects of inflammation on osteoblastic differentiation. This study improves our knowledge of lncRNAs in governing the osteogenic differentiation of hASCs and may provide novel therapeutic strategies for treating bone diseases.

Long Noncoding RNA H19 Promotes Osteoblast Differentiation Via TGF-ß1/Smad3/HDAC Signaling Pathway by Deriving miR-675.

Long noncoding RNAs (lncRNAs) are emerging as important regulatory molecules at the transcriptional and post-transcriptional levels and may play essential roles in the differentiation of human bone marrow mesenchymal stem cell (hMSC). However, their roles and functions remain unclear. Here, we showed that lncRNA H19 was significantly upregulated after the induction of osteoblast differentiation. Overexpression of H19 promoted osteogenic differentiation of hMSCs in vitro and enhanced heterotopic bone formation in vivo, whereas knockdown of H19 inhibited these effects. Subsequently, we found that miR-675, encoded by exon1 of H19, promoted osteoblast differentiation of hMSCs and was partially responsible for the pro-osteogenic effect of H19. Investigating the underlying mechanism, we demonstrated that H19/miR-675 inhibited mRNA and protein expression of transforming growth factor-ß1 (TGF-ß1). The downregulation of TGF-ß1 subsequently inhibited phosphorylation of Smad3. Meanwhile, H19/miR-675 downregulated the mRNA and protein levels of histone deacetylase (HDAC) 4/5, and thus increased osteoblast marker gene expression. Taken together, our results demonstrated that the novel pathway H19/miR-675/TGF-ß1/Smad3/HDAC regulates osteogenic differentiation of hMSCs and may serve as a potential target for enhancing bone formation in vivo.

[Relationships between microRNA expressions and prognosis in patients with tongue squamous cell carcinoma and the mechanisms microRNA regulating tongue squamous cell carcinoma biological behavior].

Tongue squamous cell carcinoma (TSCC) is the most common type of oral cancer and is well known for its high rate of proliferation and lymph nodal metastasis. Exploring the underlying pathways regulating TSCC could provide novel ideas for diagnosis and prognosis of TSCC patients, as well as molecular targets for treatment of TSCC. MicroRNAs (miRNAs) are small noncoding RNAs that inhibit gene expression through the 3' untranslated regions (3'UTRs) of their target messenger RNAs. They play crucial roles in numerous biological processes, including cancer progression. Although great efforts have been made, what role miRNAs may play in the early detection and diagnosis of TSCC is not fully understood. Recently, our team has performed a series of basic and clinical researches in an attempt to investigate the relationships between miRNA expressions and prognosis of patients with TSCC and the mechanisms under regulation of TSCC. The results showed that miR-195, miR-34a, miR-29b, miR-375 and miR-26a could inhibit TSCC cells progression and development via a sophisticated network of genes. Specifically, the anti-tumor effects of miR-195 in TSCC may be partially mediated by its inhibition of CyclinD1 and Bcl-2 expression. The expression of miR-34a could inhibit migration and invasion of TSCC cell lines via targeting MMP9 and MMP14. The function of miR-29b may be through the miR-29b/Sp1/PTEN/AKT axis. Overexpression of miR-375 inhibited Sp1 expression by targeting the 3' untranslated region of the Sp1 transcript. MEG3 and miR-26a inhibited TSCC cell proliferation, cycle progression and promoted cell apoptosis and miR-26a could increase the MEG3 expression through reduction of the expression of DNMT3B in TSCC. In light of the role of those miRNAs in diagnosis and prognosis of TSCC, we reported that decreased miR-195 and miR-375 expression was associated with poor overall survival rate of the TSCC patients, while miR-34a expression was negatively correlated with cervical lymph node metastases. Furthermore, combined low expression levels of miR-26a and MEG3 emerged as an independent prognostic factor for poor clinical outcomes in TSCC patients, suggesting that combined miR-26a and MEG3 expression might prove useful as an independent biomarker of clinical prognosis among TSCC patients.

Expression, regulation and roles of miR-26a and MEG3 in tongue squamous cell carcinoma.

MicroRNA miR-26a and long noncoding RNA (lncRNA) MEG3 gene have been independently reported to be tumor suppressor genes in various cancers, but neither has been previously associated with tongue squamous cell carcinoma (TSCC). We report here that miR-26a and lncRNA MEG3 gene expression were both strongly reduced in TSCC compared with levels in matched nonmalignant tissues, and combined low expression levels of both miR-26a and MEG3 emerged as an independent prognostic factor for poor clinical outcome in TSCC patients. Assays in the human TSCC cell lines SCC-15 and CAL27 showed that miR-26a targets the DNA methyltransferase 3B transcript and that its inhibition may result in the upregulation of MEG3, providing a plausible link between the observed reduction of miR-26a and MEG3 in TSCC tissue. Furthermore, the overexpression of miR-26a or MEG3 in SCC-15 and CAL27 cells inhibited cell proliferation and cell cycle progression, and promoted cell apoptosis. Considering the poor prognostic outcomes associated with reduced miR-26a and MEG3, our findings imply that these factors likely play important antitumor effects in TSCC pathogenesis. Furthermore, they represent potential prognostic biomarkers for stratification of TSCC patients.

Pharmacological inhibition of MYC to mitigate chemoresistance in preclinical models of squamous cell carcinoma.

Rationale: Cisplatin-based chemotherapy is the first-line treatment for late-stage head and neck squamous cell carcinoma (HNSCC). However, resistance to cisplatin has become a major obstacle for effective therapy. Cancer stem cells (CSCs) are critical for tumor initiation, growth, metastasis, and chemoresistance. How to effectively eliminate CSCs and overcome chemoresistance remains a key challenge. Herein, we confirmed that MYC plays critical roles in chemoresistance, and explored targeting MYC to overcome cisplatin resistance in preclinical models. Methods: The roles of MYC in HNSCC cisplatin resistance and cancer stemness were tested in vitro and in vivo. The combined therapeutic efficiency of MYC targeting using the small molecule MYC inhibitor MYCi975 and cisplatin was assessed in a 4­nitroquinoline 1-oxide-induced model and in a patient-derived xenograft model. Results: MYC was highly-expressed in cisplatin-resistant HNSCC. Targeting MYC using MYCi975 eliminated CSCs, prevented metastasis, and overcame cisplatin resistance. MYCi975 also induced tumor cell-intrinsic immune responses, and promoted CD8+ T cell infiltration. Mechanistically, MYCi975 induced the DNA damage response and activated the cGAS-STING-IRF3 signaling pathway to increase CD8+ T cell-recruiting chemokines. Conclusions: Our findings suggested that targeting MYC might eliminate CSCs, prevent metastasis, and activate antitumor immunity to overcome cisplatin resistance in HNSCC.

PVT1 inhibition stimulates anti-tumor immunity, prevents metastasis, and depletes cancer stem cells in squamous cell carcinoma.

Cancer stem cells (CSCs) cause tumor metastasis and immune evasion by as-yet-unknown molecular mechanisms. In the present study, we identify a long noncoding RNA (lncRNA), termed PVT1, which is highly expressed in CSCs and correlated closely with lymph node metastasis of head and neck squamous cell carcinoma (HNSCC). PVT1 inhibition eliminates CSCs, prevents metastasis, and stimulates anti-tumor immunity, while inhibiting HNSCC growth. Moreover, PVT1 inhibition promotes the infiltration of CD8+ T cells into the tumor microenvironment, thereby enhancing immunotherapy by PD1 blockade. Mechanistically, PVT1 inhibition stimulates the DNA damage response, which induces CD8+ T cell-recruiting chemokines, while preventing CSCs and metastasis via regulating the miR-375/YAP1 axis. In conclusion, targeting PVT1 might potentiate the elimination of CSCs via immune checkpoint blockade, prevent metastasis, and inhibit HNSCC growth.

A Novel LncRNA MASCC1 Regulates the Progression and Metastasis of Head and Neck Squamous Cell Carcinoma by Sponging miR-195.

The altered expression of long noncoding RNAs (lncRNAs) is associated with human carcinogenesis. We performed a high-throughput analysis of lncRNA expression in strictly selected pairs of metastatic head and neck squamous cell carcinoma (HNSCC) and non-metastatic HNSCC samples. We identified a novel lncRNA, which was highly expressed in metastatic HNSCC, named Metastasis Associated Squamous Cell Carcinoma 1 (MASCC1), for further study. Using qRT-PCR, we further compared MASCC1 expression in 60 HNSCC samples. The results show that high expression of MASCC1 in patients with HNSCC was related to poor prognosis. In vitro, MASCC1 knockdown (KD) inhibited HNSCC proliferation, migration, invasion, and tumor sphere formation, while promoting apoptosis. In vivo, MASCC1 KD inhibited HNSCC growth and lymph node metastasis. Mechanistically, MASCC1 acted as a competing endogenous RNA (ceRNA) by binding to miR-195, subsequently regulating the expression of Cyclin D1, BCL-2, and YAP1. Moreover, miR-195 overexpression rescued the effects of MASCC1 on the biological behaviors of HNSCC. Taken together, our results suggest that MASCC1 is a novel oncogene that can predict the prognosis of patients with HNSCC and is a potential therapeutic target for HNSCC intervention.

Long non-coding RNA SNHG5 promotes the osteogenic differentiation of bone marrow mesenchymal stem cells via the miR-212-3p/GDF5/SMAD pathway.

BACKGROUND: The treatment of bone loss has posed a challenge to clinicians for decades. Thus, it is of great significance to identify more effective methods for bone regeneration. However, the role and mechanisms of long non-coding RNA small nucleolar RNA host gene 5 (SNHG5) during osteogenic differentiation remain unclear. METHODS: We investigated the function of SNHG5, Yin Yang 1 (YY1), miR-212-3p and growth differentiation factor 5 (GDF5) in osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) in vitro and in vivo. Molecular mechanisms were clarified by chromatin immunoprecipitation assay and dual luciferase reporter assay. RESULTS: We found SNHG5 expression was upregulated during osteogenesis of hBMSCs. Knockdown of SNHG5 in hBMSCs inhibited osteogenic differentiation while overexpression of SNHG5 promoted osteogenesis. Moreover, YY1 transcription factor directly bound to the promoter region of SNHG5 and regulated SNHG5 expression to promote osteogenesis. Dual luciferase reporter assay confirmed that SNHG5 acted as a miR-212-3p sponge and miR-212-3p directly targeted GDF5 and further activated Smad1/5/8 phosphorylation. miR-212-3p inhibited osteogenic differentiation, while GDF5 promoted osteogenic differentiation of hBMSCs. In addition, calvarial defect experiments showed knockdown of SNHG5 and GDF5 inhibited new bone formation in vivo. CONCLUSION: Our results demonstrated that the novel pathway YY1/SNHG5/miR-212-3p/GDF5/Smad regulates osteogenic differentiation of hBMSCs and may serve as a potential target for the treatment of bone loss.