Anti-Histone H2A.X (Ab-139) Antibody (A34576)

Many pro-apoptotic factors, such as nuclear factor-kappa B (NF-κB) and Fas, play crucial roles in the process of Leydig cell apoptosis, ultimately leading to male sterility, such as in Sertoli cell only syndrome (SCO) and hypospermatogenesis. However, the molecular mechanism of such apoptosis is unclear. Recent reports on N-myc downstream-regulated gene 2 (ndrg2) have suggested that it is involved in cellular differentiation, development, and apoptosis. The unique expression of NDRG2 in SCO and hypospermatogenic testis suggests its pivotal role in those diseases. In this study, we analyzed NDRG2 expression profiles in the testes of normal spermatogenesis patients, hypospermatogenesis patients, and SCO patients, as well as in vivo and in vitro models, which were Sprague-Dawley rats and the Leydig cell line TM3 treated with the Leydig cell-specific toxicant ethane-dimethanesulfonate (EDS). Our data confirm that NDRG2 is normally exclusively located in the cytoplasm of Leydig cells and is up-regulated and translocates into the nucleus under apoptotic stimulations in human and murine testis. Meanwhile, transcription factor NF-κB was activated by EDS administration, bound to the ndrg2 promoter, and further increased in expression, effects that were abolished by NF-κB inhibitor Pyrrolidine dithiocarbamate (PDTC). Furthermore, siRNA knock-down of ndrg2 led to increased proliferative or decreased apoptotic TM3 cells, while over-expression of ndrg2 had the reverse effect. This study reveals that ndrg2 is a novel gene that participates in Leydig cell apoptosis, with essential functions in testicular cells, and suggests its possible role in apoptotic Leydig cells and male fertility.

ING4 (inhibitor of growth gene 4) is a new member of the ING gene family and is implicated in chromatin remodeling and repression of cell growth. In order to explore the roles of ING4 in head and neck squamous cell carcinoma (HNSCC), ING4 expression was assessed in 214 cases of HNSCC by immunohistochemistry using tissue microarray, and in three oral SCC cell lines by immunohistochemistry and Western blotting. Expression of ING4 was also compared to clinicopathological variables, TUNEL assay staining, and the expression of several tumorigenic markers. We found nuclear expression of ING4 was gradually decreased from non-cancerous epithelium and dysplasia to HNSCC and was negatively correlated with a poorly-differentiated status, T staging, and TNM staging in HNSCC. In contrast, cytoplasmic expression of ING4 was significantly increased in HNSCC and was significantly associated with lymph node metastasis and 14-3-3η expression. In addition, nuclear expression of ING4 was positively correlated with p21 and p300 expression and with the apoptotic index. These results suggest that the decreases in nuclear ING4 may play important roles in tumorigenesis, progression and tumor differentiation in HNSCC. Increases in cytoplasmic ING4 may be due to 14-3-3η binding and may also be involved in malignant progression. Nuclear ING4 may modulate the transactivation of target genes, promoting apoptosis and cell cycle arrest through interactions with p300 and p21.

PURPOSE:

This study aimed to assess the protein level of inhibitor of growth gene 5 (ING5) in head and neck squamous cell carcinoma (HNSCC) and to explore its roles in tumorigenesis and cancer progression.

METHODS:

ING5 expression was assessed in 172 cases of HNSCC by immunohistochemistry using tissue microarray, and in 3 oral SCC cell lines by immunohistochemistry and Western blot. Expression of ING5 was compared with clinicopathological variables, TUNEL assay staining, and the expression of several tumorigenic markers. In addition, double immunofluorescence labeling was performed in order to analyze the colocalization of ING5 with p300 and p21.

RESULTS:

ING5 expression was primarily observed in the nuclei, but was also occasionally found in the cytoplasm of both SCC cell lines and tissue samples of HNSCC. Nuclear expression of ING5 in HNSCC was significantly lower than that of non-cancerous epithelium, and was positively correlated with a well-differentiated status. In contrast, cytoplasmic expression of ING5 was significantly increased in HNSCC, and was inversely correlated with a well-differentiated status and nuclear ING5 expression. In addition, nuclear expression of ING5 was positively correlated with p21 and p300 expression, and with the apoptotic index. In contrast, cytoplasmic expression of ING5 was negatively correlated with the expression of p300, p21, and PCNA. Although no statistical association was found between the expression of nuclear ING5 and mutant p53 in HNSCC, patients with high expression of nuclear ING5 tended to have converse prognoses when grouped according to mutant p53 expression.

CONCLUSIONS:

Our results suggest that a decrease in nuclear ING5 localization and cytoplasmic translocation are involved in tumorigenesis and tumor differentiation in HNSCC. Nuclear ING5 may modulate the transactivation of target genes, and may promote apoptosis and cell cycle arrest by interacting with the p300 and p21 proteins. ING5 may function as a tumor suppressor gene or oncogene tightly linked with p53 status, and may play an important role in the prognosis of HNSCC patients. Therefore, we propose that ING5 represents a novel potential molecular therapeutic target for HNSCC.