MYCT1 inhibits hematopoiesis in diffuse large B-cell lymphoma by suppressing RUNX1 transcription.

BACKGROUND: The abnormality of chromosomal karyotype is one factor causing poor prognosis of lymphoma. In the analysis of abnormal karyotype of lymphoma patients, three smallest overlap regions were found, in which MYCT1 was located. MYCT1 is the first tumor suppressor gene cloned by our research team, but its studies relating to the occurrence and development of lymphoma have not been reported. METHODS: R banding analyses were employed to screen the abnormality of chromosomal karyotype in clinical specimen and MYCT1 over-expression cell lines. FISH was to monitor MYCT1 copy number aberration. RT-PCR and Western blot were to detect the mRNA and protein levels of the MYCT1 and RUNX1 genes, respectively. The MYCT1 and RUNX1 protein levels in clinical specimen were evaluated by immunohistochemical DAB staining. The interaction between MYCT1 and MAX proteins was identified via Co-IP and IF. The binding of MAX on the promoter of the RUNX1 gene was detected by ChIP and Dual-luciferase reporter assay, respectively. Flow cytometry and CCK-8 assay were to explore the effects of MYCT1 and RUNX1 on the cell cycle and proliferation, respectively. RESULTS: MYCT1 was located in one of three smallest overlap regions of diffuse large B-cell lymphoma, it altered chromosomal instability of diffuse large B-cell lymphoma cells. MYCT1 negatively correlated with RUNX1 in lymphoma tissues of the patients. MAX directly promoted the RUNX1 gene transcription by binding to its promoter region. MYCT1 may represses RUNX1 transcription by binding MAX in diffuse large B-cell lymphoma cells. MYCT1 binding to MAX probably suppressed RUNX1 transcription, leading to the inhibition of proliferation and cell cycle of the diffuse large B-cell lymphoma cells. CONCLUSION: This study finds that there is a MYCT1-MAX-RUNX1 signaling pathway in diffuse large B-cell lymphoma. And the study provides clues and basis for the in-depth studies of MYCT1 in the diagnosis, treatment and prognosis of lymphoma.

MicroRNA-144 Regulates Cardiomyocyte Proliferation and Apoptosis by Targeting TBX1 through the JAK2/STAT1 Pathway.

It is currently believed that the TBX1 gene is one of the core genes of congenital heart disease (CHD). However, there are few studies on the abnormal regulation of TBX1 gene expression. The purpose of this work was to investigate the role of miR-144 and TBX1 in cardiac development by studying the regulatory relationship and mechanism of miR-144 on TBX1/JAK2/STAT1 in cardiomyocytes. Cell proliferation was detected by MTT and clone formation assay and cell cycle and apoptosis by flow cytometry. The levels of miR-144 and TBX1 in H9c2 cells were assessed by qRT-PCR. Dual luciferase reporter assay was used to validate the direct targeting of TBX1 with miR-144. The protein expression levels of TBX1 and its downstream proteins were measured by Western blot analysis. miR-144 inhibited H9c2 cell proliferation by arresting cells in G1 phase. Furthermore, miR-144 induced H9c2 cell apoptosis and activated the JAK2/STAT1 signaling pathway. Bioinformatic predictions and luciferase reporter assay showed that miR-144 directly targets TBX1. Co-overexpression of miR-144 and TBX1 upregulated cell proliferation by accelerating G1 to S phase transition and downregulated cell apoptosis through inhibiting the JAK2/STAT1 signaling pathway. miR-144 acts as a proliferation inhibitor in cardiomyocytes via the TBX1/JAK2/STAT1 axis and is therefore a potential novel therapeutic target for CHD treatment.

Oncogenic miR-23a in Pancreatic Ductal Adenocarcinogenesis Via Inhibiting APAF1.

BACKGROUND: miR-23a, which participates in invasion of pancreatic ductal adenocarcinoma cells into the mesothelial barrier, is a critical regulator in many cancers. It, however, is still unknown whether miR-23a regulates pancreatic cell proliferation and apoptosis or not. AIMS: We sought to investigate the role of miR-23a in regulation of pancreatic cell proliferation and apoptosis. METHODS: miRNA, mRNA, and protein expressions were determined by qRT-PCR and Western blot, respectively. Dual-luciferase reporter assay was used in detection for binding ability of miR-23a to APAF1. Ectopic miR-23a and APAF 1 were introduced to pancreatic cells, and their roles in proliferation and apoptosis were detected by MTT, colony formation, and apoptosis assays, respectively. RESULTS: Up-regulation of miR-23a and down-regulation of APAF 1 were found in pancreatic ductal cancer, respectively. miR-23a significantly inhibited the luciferase activity by targeting APAF 1 3'UTR. Ectopic miR-23a significantly suppressed the APAF 1 gene expression in pancreatic cancer cells. Similar to siAPAF1, miR-23a significantly promoted pancreatic cancer cell proliferation and repressed apoptosis. Furthermore, miR-23a inhibitor and exogenous APAF 1 could recover the effects. CONCLUSIONS: It is suggested that miR-23a, acting as an oncogenic regulator by directly targeting APAF 1 in pancreatic cancer, is a useful potential biomarker in diagnosis and treatment of pancreatic cancer.

Genetic Effects on Sensorineural Hearing Loss and Evidence-based Treatment for Sensorineural Hearing Loss.

In this article, the mechanism of inheritance behind inherited hearing loss and genetic susceptibility in noise-induced hearing loss are reviewed. Conventional treatments for sensorineural hearing loss (SNHL), i.e. hearing aid and cochlear implant, are effective for some cases, but not without limitations. For example, they provide little benefit for patients of profound SNHL or neural hearing loss, especially when the hearing loss is in poor dynamic range and with low frequency resolution. We emphasize the most recent evidence-based treatment in this field, which includes gene therapy and allotransplantation of stem cells. Their promising results have shown that they might be options of treatment for profound SNHL and neural hearing loss. Although some treatments are still at the experimental stage, it is helpful to be aware of the novel therapies and endeavour to explore the feasibility of their clinical application.

MicroRNA-27a promotes proliferation and suppresses apoptosis by targeting PLK2 in laryngeal carcinoma.

BACKGROUND: miRNA-27a has been confirmed as an important regulator in carcinogenesis and other pathological processes. Whether and how it plays a role in the laryngeal carcinoma is unknown. METHODS: Mature miRNA-27a expression in laryngeal cancer was detected by qRT-PCR. Gain-of-function studies using mature miR-27a were performed to investigate cell proliferation and apoptosis in the Hep2 cells. In silico database analysis and luciferase reporter assay were applied to predict and validate the direct target, respectively. Loss-of-function assays were performed to investigate the functional significance of the miR-27a target gene. qRT-PCR and Western blot were used to evaluate mRNA and protein levels of the target, respectively. RESULTS: miR-27a was significantly up-regulated in the laryngeal tumor tissues compared to the adjacent non-tumor tissues. In silico database analysis result revealed that PLK2 is a potential target of miR-27a. luciferase reporter assay result showed the direct inhibition of miR-27a on PLK2-3'UTR. In the cases with miR-27a up-regulation, PLK2 protein expression level was significantly lower in cancer tissues than that in the adjacent non-tumor tissues, which showed a negative correlation with miR-27a expression level. Both miR-27a and knockdown of PLK2 caused the increase of the cell viability and colony formation and inhibition of the late apoptosis in the Hep2 cell lines. Moreover, miR-27a but not PLK2 also repressed the early apoptosis in the Hep2 cells. Additionally, no alteration of the Hep2 cell cycle induced by miR-27a was detected. CONCLUSIONS: miR-27a acts as an oncogene in laryngeal squamous cell carcinoma through down-regulation of PLK2 and may provide a novel clue into the potential mechanism of LSCC oncogenesis or serve as a useful biomarker in diagnosis and therapy in laryngeal cancer.

MYCT1 attenuates renal fibrosis and tubular injury in diabetic kidney disease.

Tubulointerstitial abnormalities contribute to the progression of diabetic kidney disease (DKD). However, the underlying mechanism of the pathobiology of tubulointerstitial disease is largely unknown. Here, we showed that MYCT1 expression was downregulated in in vitro and in vivo DKD models. Adeno-associated virus (AAV)-Myct1 significantly attenuated renal dysfunction and tubulointerstitial fibrosis in diabetic db/db mice and downregulated Sp1 transcription and TGF-ß1/SMAD3 pathway activation. In human proximal tubular epithelial cells, high glucose-induced high expression of SP1 and TGF-ß1/SMAD3 pathway activation as well as overaccumulation of extracellular matrix (ECM) were abrogated by MYCT1 overexpression. Mechanistically, the binding of VDR to the MYCT1 promoter was predicted and confirmed using dual-luciferase reporter and ChIP analysis. VDR transcriptionally upregulates MYCT1. Our data reveal MYCT1 as a new and potential therapeutic target in treating DKD.

Promoter hypermethylation-induced transcriptional down-regulation of the gene MYCT1 in laryngeal squamous cell carcinoma.

BACKGROUND: MYCT1, previously named MTLC, is a novel candidate tumor suppressor gene. MYCT1 was cloned from laryngeal squamous cell cancer (LSCC) and has been found to be down-regulated in LSCC; however, the regulatory details have not been fully elucidated. METHODS: Here, we sought to investigate the methylation status of the CpG islands of MYCT1 and mRNA levels by bisulfite-specific PCR (BSP) based on sequencing restriction enzyme digestion, reverse transcription and real-time quantitative polymerase chain reaction (RQ-PCR). The function of specific sites in the proximal promoter of MYCT1 in LSCC was measured by transient transfection, luciferase assays, electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation assay (ChIP). RESULTS: The results suggested hypermethylation of 12 CpG sites of the promoter in both laryngeal cancer tissues and the laryngeal cancer line Hep-2 cell. The hypermethylation of the site CGCG (-695 to -692), which has been identified as the c-Myc binding site, was identified in laryngeal cancer tissues (59/73) compared to paired mucosa (13/73); in addition, statistical analysis revealed that the methylation status of this site significantly correlated with cancer cell differentiation(p < 0.01). The mRNA level of MYCT1 increased in Hep-2 cells treated with 5-aza-C (p < 0.01). The luciferase activity from mutant transfectants pGL3-MYCT1m (-852/+12, mut-695-C > A, mut-693-C > G) was significantly reduced compared with the wild type pGL3-MYCT1 (-852/+12), while the luciferase activity from wild transfectants pGL3-MYCT1 (-852/+12) rose after 5-aza treatment in Hep-2 cells. Finally, EMSA and ChIP confirmed that the methylation of the CGCG (-695 to -692) site prevented c-Myc from binding of the site and demethylation treatment of the 5' flanking region of MYCT1 by 5-aza induced the increased occupation of the core promoter by c-Myc (p < 0.01). CONCLUSION: In summary, this study concluded that hypermethylation contributed to the transcriptional down-regulation of MYCT1 and could inhibit cancer cell differentiation in LSCC. DNA methylation of the CGCG site (-695 to -692) of MYCT1 altered the promoter activity by interfering with its binding to c-Myc in LSCC. Epigenetic therapy of reactivating MYCT1 by 5-aza should be further evaluated in clinical trails of LSCC.

MYCT1 alters the glycogen shunt by regulating selective translation of RACK1-mediated enzymes.

MYCT1 has been shown to function as a tumor suppressor in various tumors, but its role in metabolism has never been reported. Here, we showed that global inactivation of Myct1 in mice led to progressive accumulation of glycogen in the liver, which was accompanied by aberrant changes in intermediates of the glycogen metabolic pathway. Mechanistically, MYCT1 appeared to promote translation efficiency of PGM1, UGP2 and GSK3A in hepatic cells in a RACK1-dependent manner. Consequently, upregulation of the three enzymes enhanced the glycogen shunt. Our data reveal a critical role of MYCT1 as a switch for the glycogen shunt in tumor cells.

Overexpression of DOCK6 in oral squamous cell cancer promotes cellular migration and invasion and is associated with poor prognosis.

OBJECTIVE: We aimed to identify the role of DOCK6 in oral squamous cell cancer (OSCC) in this study. DESIGN: DOCK6 expression in OSCC was analyzed using TCGA and GEO datasets and was verified by quantitative real-time PCR, Western blotting, and immunohistochemistry. Statistical analyses were performed to evaluate the relationships between DOCK6 expression and the clinicopathological characteristics of OSCC patients. Wound healing and Transwell assays were performed to assess OSCC cell migration and invasion, respectively. STRING and GO analyses and gene set enrichment analysis were used to identify DOCK6-interacting proteins, their functions and their potential pathways. RESULTS: DOCK6 was significantly upregulated at both the mRNA and protein levels in OSCC tissues (all P < 0.05). DOCK6 levels were positively correlated with age (P < 0.05), lymph node metastasis status (P < 0.001), clinical stage (P < 0.001), differentiation (P < 0.05), and poor clinical outcome (P < 0.05) in OSCC patients. Furthermore, univariate and multivariate analyses revealed that high DOCK6 expression (P < 0.01) and clinical stage III-IV (P < 0.05) might serve as independent prognostic factors for OSCC patients. Functionally, DOCK6 silencing significantly suppressed OSCC cell migration and invasion (all P < 0.05). Ten proteins that interact with DOCK6, more than ten functions related to cancer, and more than six pathways related to DOCK6 in OSCC were identified via bioinformatic methods. CONCLUSION: DOCK6 is upregulated in OSCC, is associated with a poor prognosis in OSCC patients and increases OSCC cells migration and invasion. These findings suggest that DOCK6 may be a potential therapeutic target with prognostic implication in patients with OSCC.

14-3-3epsilon contributes to tumour suppression in laryngeal carcinoma by affecting apoptosis and invasion.

BACKGROUND: 14-3-3epsilon regulates a wide range of biological processes, including cell cycle control, proliferation, and apoptosis, and plays a significant role in neurogenesis and the formation of malignant tumours. However, the exact function and regulatory mechanism of 14-3-3epsilon in carcinogenesis have not been elucidated. METHODS: The expression of 14-3-3epsilon was assessed by RT-PCR and western blotting. The invasiveness and viability of Hep-2 cells were determined by the transwell migration assay and MTT assay, respectively. Cell cycle and apoptosis of Hep-2 cells were detected by flow cytometry. RESULTS: The mRNA and protein expression of 14-3-3epsilon in larynx squamous cell carcinoma (LSCC) tissues were significantly lower than those in clear surgical margin tissues. Statistical analysis showed that the 14-3-3epsilon protein level in metastatic lymph nodes was lower than that in paired tumour tissues. In addition, the protein level of 14-3-3epsilon in stage III or IV tumours was significantly lower than that in stage I or II tumours. Compared with control Hep-2 cells, the percentages of viable cells in the 14-3-3epsilon-GFP and negative control GFP groups were 36.68 +/- 14.09% and 71.68 +/- 12.10%, respectively. The proportions of S phase were 22.47 +/- 3.36%, 28.17 +/- 3.97% and 46.15 +/- 6.82%, and the apoptotic sub-G1 populations were 1.23 +/- 1.02%, 2.92 +/- 1.59% and 13.72 +/- 3.89% in the control, negative control GFP and 14-3-3epsilon-GFP groups, respectively. The percentages of the apoptotic cells were 0.84 +/- 0.25%, 1.08 +/- 0.24% and 2.93 +/- 0.13% in the control, negative control GFP and 14-3-3epsilon-GFP groups, respectively. The numbers of cells that penetrated the filter membrane in the control, negative control GFP and 14-3-3epsilon-GFP groups were 20.65 +/- 1.94, 17.63 +/- 1.04 and 9.1 +/- 0.24, respectively, indicating significant differences among the different groups. CONCLUSIONS: Decreased expression of 14-3-3epsilon in LSCC tissues contributes to the initiation and progression of LSCC. 14-3-3epsilon can promote apoptosis and inhibit the invasiveness of LSCC.

MYCT1 Inhibits the Adhesion and Migration of Laryngeal Cancer Cells Potentially Through Repressing Collagen VI.

MYCT1, a target of c-Myc, inhibits laryngeal cancer cell migration, but the underlying mechanism remains unclear. In the study, we detected differentially expressed genes (DEGs) from laryngeal cancer cells transfected by MYCT1 using RNA-seq (GSE123275). DEGs from head and neck squamous cell carcinoma (HNSCC) were first screened by comparison of transcription data from the Gene Expression Omnibus (GSE6631) and the Cancer Genome Atlas (TCGA) datasets using weighted gene co-expression network analysis (WGCNA). GO and KEGG pathway analysis explained the functions of the DEGs. The DEGs overlapped between GSE6631and TCGA datasets were then compared with ours to find the key DEGs downstream of MYCT1 related to the adhesion and migration of laryngeal cancer cells. qRT-PCR and Western blot were applied to validate gene expression at mRNA and protein levels, respectively. Finally, the cell adhesion, migration, and wound healing assays were to check cell adhesion and migration abilities, respectively. As results, 39 overlapping genes were enriched in the GSE6631 and TCGA datasets, and most of them revealed adhesion function. Thirteen of 39 genes including COL6 members COL6A1, COL6A2, and COL6A3 were overlapped in GSE6631, TCGA, and GSE123275 datasets. Similar to our RNA-seq results, we confirmed that COL6 is a target of MYCT1 in laryngeal cancer cells. We also found that MYCT1 inhibited the adhesion and migration of laryngeal cancer cells via COL6. These indicate that COL6 is a potential target of MYCT1 and participates the adhesion and migration of laryngeal cancer cells, which provides an important clue for further study on how MYCT1 regulating COL6 in laryngeal cancer progression.

MYCT1 inhibits the EMT and migration of laryngeal cancer cells via the SP1/miR-629-3p/ESRP2 pathway.

MYCT1 has an inhibitory effect on the migration of laryngeal cancer cells, although the underlying molecular mechanism remains unknown. In this study, we aimed to explore the mechanism of MYCT1 in the epithelial-mesenchymal transition (EMT) and migration of laryngeal cancer cells. We found that MYCT1 significantly decreased the expression of miR-629-3p but increased the expression of ESRP2 in laryngeal cancer cells. The expression of miR-629-3p and ESRP2 in laryngeal cancer tissues showed significantly positive and negative correlations with patient metastasis, respectively. miR-629-3p was confirmed to repress the expression of ESRP2 by targeting its 3'UTR. SP1 was verified to be a direct transcription factor for miR-629-3p and a downstream target of MYCT1. Moreover, MYCT1 inhibited the EMT and migration of laryngeal cancer cells through the SP1/miR-629-3p/ESRP2 pathway. Taken together, our results establish a novel MYCT1 signaling pathway in the EMT and migration of laryngeal cancer cells, thus providing important insights for further studying the pathway in the diagnosis and treatment of laryngeal cancer.

MYCT1 represses apoptosis of laryngeal cancerous cells through the MAX/miR-181a/NPM1 pathway.

MYCT1 is an important gene known to regulate cell viability and apoptosis of laryngeal cancer cells. However, the underlying molecular mechanism remains unclear. Here, we show that MAX enhances the expression of miR-181a by directly binding to its promoter, whereas miR-181a targets NPM1 and suppresses its expression in laryngeal cancer cells. MYCT1 and miR-181a decrease cell viability and colony formation through enhanced apoptosis, whereas NPM1 displays opposite effects in laryngeal cancer cells. Their opposing functions are further supported by the findings (a) that miR-181a is down-regulated, while NPM1 is up-regulated in laryngeal cancer, and (b) that either inhibition of miR-181a or overexpression of NPM1 can revert the pro-apoptotic effects of MYCT1 on laryngeal cancer cells through extracellular and intracellular apoptotic pathways. Our data suggest that MYCT1 may synergistically interact with MAX as a co-transcription factor or a component of MAX transcriptional complex, to transcriptionally regulate the expression of miR-181a, which, in turn, decreases NPM1 expression at post-transcriptional levels, leading to enhanced apoptosis in laryngeal cancer cells. These factors may serve as potential targets for early diagnosis and treatment of laryngeal cancer.

Genetic Alleles Associated with SLE Susceptibility and Clinical Manifestations in Hispanic Patients from the Dominican Republic.

PURPOSE: Systemic lupus erythematosus (SLE) is a complex autoimmune disease with marked disparities in prevalence and disease severity among different ethnic groups. The purpose of this study is to characterize a Latin American cohort and identify genetic risk factors for developing SLE and its end-organ manifestations in this Latin Hispanic cohort. METHODS: A total of 201 SLE cases and 205 non-diseased controls were recruited in the Dominican Republic (DR). Cases were defined according to the 1997 revised American College of Rheumatology criteria for the classification of SLE. Genomic DNA was prepared from whole blood and applied to genotyping analyses for 42 single nucleotide polymorphisms (SNPs) that have been implicated in autoimmune diseases, including SLE, in other ethnic populations. Data were analyzed by Fisher's Exact Probability Test. RESULTS: In this cohort, SNP rs9271366 (tag SNP for HLA-DRB1*15:01) confers the highest risk for SLE among the 13 MHC gene alleles that display association with SLE (p = 8.748E-10; OR = 3.5). Among the 26 non-MHC gene alleles analyzed, SNP rs2476601 in PTPN22 gene confers the highest risk for SLE (p = 0.0001; OR = 5.6). ITGAM, TNFSF4, TNIP1, STAT4, CARD11, BLK, and TNXB gene alleles were confirmed as SLE-susceptible alleles in the DR cohort. However, IRF5 and TNFAIP3 gene alleles, established risk factors for SLE in populations of European and Asian ancestry, are not significantly associated with SLE in this cohort. We also defined a novel HLA-DRA haplotype that confers an increased risk for lupus nephritis (LN) and alleles in HLA-DRA2 and TNFSF4 genes as genetic risk factors for developing neuropsychiatric (NP) SLE. CONCLUSION: Our data suggest that the Latin American population shares some common genetic risk factors for SLE as other populations, but also has distinct risk gene alleles that contribute to SLE susceptibility and development of LN and NPSLE. This is the first study focusing on genetic risk factors for SLE in the DR, a Latin American population that has never been characterized before.

[Research of HOXD13 and FHL1 in idiopathic congenital talipes equinovarus].

To investigate the relationship of HOXD13 and FHL1 in idiopathic congenital talipes equinovarus(ICTEV), 84 samples from patients with ICTEV were used in the study. Mutation in the coding region of HOXD13 was detected by denaturing gradinent electrophoresis. The mRNA and protein levels of HOXD13 and FHL1 were evaluated by RT-PCR and immunohistochemistry, respectively. The binding site of FHL1 to HOXD13 predicted by PMATCH software was validated by EMSA( Electrophoretic mobility shift assay,EMSA).No mutation was found in the coding region of HOXD13 in 84 samples from patients with ICTEV. Both HOXD13(33.3%) and FHL1(46.6%) were down-regulated in ICTEV muscle tissue. The result of EMSA showed that the special binding band appeared when HOXD13 existed. The results shows that HOXD13 gene mutation was not involved in outbreak in idiopathic congenital talipes equinovarus, but changes of HOXD13 and FHL1 gene expression related to the development of talipes equinovarus malformation. HOXD13 might play an role in ICTEV through regulating FHL1 expression.

CREB promotes laryngeal cancer cell migration via MYCT1/NAT10 axis.

PURPOSE: CREB, MYCY1 and NAT10 are involved in cancer cell migration. However, the relationship between these three proteins and their role in laryngeal cancer cell migration remains unknown. METHODS: Transient gene transfection was performed in laryngeal cancer cells. Bioinformatics analysis was used to predict the binding of CREB to MYCT1 promoter. Binding of CREB to the promoter of MYCT1 was monitored by luciferase reporter assay and chromatin immuno-precipitation method in vitro and in vivo, respectively. Real-time RT-PCR and Western bolt were applied to detect gene transcription and translation levels, respectively. Laryngeal cancer cell migration was assayed by transwell chamber experiment. RESULTS: CREB protein expression was significantly up-regulated in laryngeal cancer tissues and associated with cancer differentiation, tumor stage, and lymphatic metastasis. CREB inhibits MYCT1 expression by direct binding to its promoter. Meanwhile, MYCT1 has a negative impact on the NAT10 gene expression. Furthermore, CREB promotes NAT10 expression via down-regulating the MYCT1 gene expression. In addition, contrary to MYCT1, CREB and NAT10 enhanced laryngeal cancer cell migration. MYCT1 and NAT10 significantly rescued the effects of CREB and MYCT1 on Hep2 cell migration, respectively. CONCLUSION: CREB promotes laryngeal cancer cell migration via MYCT1/NAT10 axis, suggesting that CREB might be a potential prognostic marker in laryngeal cancer.

Study of the SH3-domain GRB2-like 2 gene expression in laryngeal carcinoma.

BACKGROUND: Laryngeal carcinoma is a common malignant tumor of the upper respiratory tract, and in 95% of cases the tumor is laryngeal squamous cell carcinoma (LSCC). The abnormity of SH3-domain GRB2-like 2 (SH3GL2) gene was found in LSCC. In order to clarify the relationship between SH3GL2 gene and LSCC, we evaluated the expression of the SH3GL2 gene in LSCC. METHOD: Real-time PCR, immunohistochemistry and Western blotting were used to detect the mRNA and protein expression and find the various rules of SH3GL2 gene in LSCC. RESULTS: The result of real-time PCR showed that the expression level of SH3GL2 mRNA in LSCC tissue was apparently down-regulated; immunohistochemical analysis showed that SH3GL2 protein was mainly located in cytoplasm, the rate of positive cells and SH3GL2 protein expression level were fluctuated with the pathological classification of LSCC; the result of Western blotting showed that SH3GL2 protein was down-regulated significantly in LSCC samples, especially in metastatic lymph nodes. CONCLUSIONS: These results suggest that SH3GL2 is a LSCC related gene and its expression level is fluctuated with the pathological classification which indicate that SH3GL2 participates in the development and progression of LSCC. And it may be considered as a novel tumor marker to find both a new anti-oncogene and relative factors of invasion and metastasis of laryngeal carcinoma.

Decreased expression of DICER1 in gastric cancer.

BACKGROUND: The role of epigenetics in gene expression regulation and development significantly enhances our understanding of carcinogenesis. All the tumor related genes may be the target of epigenetical or genetic regulation. We selected some epigenetically regulated genes for cDNA array analysis and observed variability in the expression of the DICER1 gene in distinct stages of gastric cancer. The aim of this study was to assess the correlation between the expression of DICER1, an epigenetically regulated gene, and gastric cancer. METHODS: To detect the expression of 506 tumor-associated genes, including DICER1, in the matched cancerous mucosa, pre-malignant lesion (adjacent mucosa), non-cancerous gastric mucosa and distant lymphocyte metastatic lesion in 3 cases of gastric cancers using cDNA array. DICER1 mRNA expression and DICER1 protein expression were further analyzed by Real-time PCR and Western blot in 32 cases of progressive gastric cancer. DICER1 protein expression was also detected in 33 early and 30 progressive gastric cancers by the immunohistochemistry (IHC) method. RESULTS: In 3 cases of gastric cancer cDNA array showed dramatically decreased expression of DICER1 in pre-malignant lesion, cancerous mucosa and distant lymphocyte metastatic lesions compared with matched noncancerous gastric mucosa, pre-malignant lesion and cancerous mucosa. Real-time PCR results showed that the expression level of DICER1 mRNA in gastric cancer was significantly down-regulated compared to normal gastric tissue (P < 0.05). The IHC assay also showed that the expression of DICER1 was significantly decreased in progressive gastric cancer. Among the 63 cases of gastric cancers, 13/33 early (39.4%) and 19/30 (63.3%) progressive cancers showed negative expression of DICER1 (50.8%). The difference in expression of DICER1 between early and progressive gastric cancers was significant (P < 0.01). The result of Western blotting showed that DICER1 protein was down-regulated significantly in advanced gastric cancer (P < 0.05). CONCLUSIONS: DICER1 expression is decreased during the progression of gastric cancer, especially in progressive gastric cancers, which indicating DICER1 may play an important role in the development of cancer and the epigenetical regulation involved.

[Analysis of chromosome aberrations in the cell derived from primary cell culture of laryngeal carcinoma and the Hep-2 cell line].

OBJECTIVE: To search for characteristic chromosome changes in primary laryngeal squamous cell carcinoma (LSCC) and Hep-2 cell line and to realize the relationship between the cytogenetic abnormality and the pathogenetic mechanism in LSCC. METHODS: The fresh resulted samples of LSCC were analyzed with an improved primary cell culture for chromosome preparation and G-banding technique. Hep-2 cell line was analyzed by high resolution banding technique. Molecular cytogenetics analysis was made by chromosome 6 painting probe. RESULTS: Four primary LSCC succeeded in primary cell culture and obtained metaphases, one was tetraploid, the other three were triploid. The chromosome mode of Hep-2 cell line was from 68 to 75 and fifteen marker chromosomes were found. The most structural abnormalities of chromosome in primary LSCC and HEP-2 cell line were unbalance translocation, terminal deletion and isochromosome. The complicate aberration in chromosome 6 was common in LSCC and Hep-2. CONCLUSION: 6q-, I(5p), 17p-, 5q- are considered as characteristic chomosome changs in LSCC. Fluorescence in situ hybridization (FISH) may enhance the ability of detecting complicated chromosome rearrangements and marker chromosomes, which could provide more value data to verify the chromosome characteristic aberration in LSCC.

Novel partners of S100A8 identified in laryngeal cancer cell lines.

OBJECTIVE: To explore mechanism of S100A8 in the oncogenesis and development of laryngeal cancer. METHODS: Proteins interacting with S100A8 were isolated from laryngeal cancer cell lines Hep-2 by immunoprecipitation assay with anti-S100A8 antibody. The target bands were cut out and identified by maxtrix assisted laser desorption/ionization time of flight (MALDI-TOF). The peptide mass fingerprinting data of the proteins identified were analyzed based on the Mascot database. The NF-kappa B binding sites of the proteins were predicted by P-Match software. The binding ability of one of the proteins to S100A8 was confirmed by co-immunoprecipitation and immunocytochemistry methods. RESULTS: Four proteins interacting with S100A8 were obtained, which were hypothetical protein LOC80154, MHC class I HLA-B, similar to T-box 1 isoform C and sarcolemmal associated protein 1. The four genes were predicted to have NF-kappa B binding sites. MHC class I HLA-B, which is one of targets in NF-kappa B pathway, was first confirmed to have the binding ability to S100A8. CONCLUSION: The novel partners of S100A8 identified in the study might be involved in NF-kappa B pathway. The binding ability of MHC class I HLA-B to S100A8 implies that S100A8 might function as a new member with other proteins including HLA-B in NF-kappa B pathway. These findings provide a new clue to further study on the molecular mechanism of S100A8 in the genesis of laryngeal carcinomas.