Effect of human epididymis protein 4 gene silencing on the malignant phenotype in ovarian cancer.

BACKGROUND: Human epididymis secretory protein 4 (HE4) has been proved to be a promising novel biomarker for the detection of epithelial ovarian carcinomas. Compared with CA125, HE4 assay demonstrated an improved ability to discriminate between pelvic mass with malignant and benign disease. Though it is well known that HE4 is overexpressed in ovarian cancer, however, the role of HE4 in the carcinogenesis and progression of ovarian cancer remains unkown. METHODS: In this study, we explored the role of HE4 in the carcinogenesis and progression of ovarian cancer. We screened nine ovarian cancer cell lines for HE4 expression, and using RNA interference (RNAi), we silenced HE4 gene expression in CaoV3 and SKOV3.ip1 ovarian cancer cell lines. We assessed the effect of HE4 gene silencing on the transformed phenotype by examining the cell cycle, apoptosis, proliferation and transwell migration/invasion in vitro. RESULTS: HE4 gene silencing induces G0/G1 arrest and blocks the progression from the G1 to S phase in CaoV3 and SKOV3.ip1 cells. HE4 knockdown also inhibited cell proliferation, migration and invasion in SKOV3.ip1 cells in vitro. CONCLUSION: HE4 may be involved in the regulation of the cell cycle and promote ovarian cancer migration and invasion.

Human epididymis protein 4 (HE4) as a serum tumor biomarker in patients with ovarian carcinoma.

BACKGROUND: Ovarian cancer remains a leading cause of death from gynecological malignancy. Early diagnosis is the most important determinant of survival. For more than 25 years, cancer antigen 125 (CA 125) has been the criterion standard biomarker for the diagnosis and management of women with epithelial ovarian cancer. This study evaluated human epididymis protein 4 (HE4), a novel ovarian cancer biomarker, both alone and in combination with CA 125 as a diagnostic marker for ovarian cancer in a Chinese population. METHODS: Sera from 491 Chinese women with ovarian cancer or nonmalignant disorders and healthy women were analyzed. Sensitivities and specificities for both biomarkers and the combination were determined using predefined cutoffs (HE4>150 pmol/L and CA 125>35 U/mL) and receiver operator characteristic curves to define cutoffs based on 95% and 98% sensitivities. RESULTS: At baseline, serum HE4 and CA 125 levels were significantly higher in the ovarian cancer group versus the 5 reference groups. Using predefined cutoffs, HE4 specificity for ovarian cancer ranged from 90% to 100%; CA 125 specificity ranged from 36% (benign gynecologic disease) to 99%. Combining both markers yielded specificity for ovarian cancer of 100%. Using receiver operator characteristic curve analysis, the cutoff for 95% and 98% specificity was 102.6 and 150.2 pmol/L for HE4, respectively, and 127.2 and 325.5 U/mL for CA 125, respectively; the sensitivity of CA 125 for distinguishing ovarian cancer from benign gynecologic disease was 54% (95% specificity) and 28% (98% specificity), improving to 78% and 68%, respectively, with the addition of HE4. CONCLUSIONS: Human epididymis protein 4 used in conjunction with CA 125 yields improved specificity for ovarian cancer compared with the use of CA 125 alone, generally similar to results seen in non-Chinese populations.

The epididymis as a target for male contraceptive development.

The epididymis is an excellent target for the development of a male contraceptive. This is because the process of sperm maturation occurs in this organ; spermatozoa become motile and are able to recognise and fertilise an egg once they have traversed the epididymal duct. However, a number of attempts to interfere in sperm maturation and epididymal function or both have not been successful. The use of transgenic animals has proved useful in identifying a few epididymal targets but has yet to open the doors for drug development. Continuous focus on identifying additional epididymal targets and sperm-specific and epididymal-specific drugs is key to bringing a male contraceptive acting on the epididymis to the public.

The roles of the epididymis and prostasomes in the attainment of fertilizing capacity by stallion sperm.

The epididymis is a long, tightly coiled tube within the lumen of which sperm matures. Sperm maturation involves morphological and biochemical changes in the sperm plasma membrane in response to epididymal secretions and their various proteins. Some of these proteins become outer membrane components while others become integral membrane proteins; transfer of some proteins to the sperm plasma membrane may be mediated by epididymosomes. Nevertheless, the molecular pathways by which spermatozoa acquire fertilizing capacity during their transit through the epididymis remain ambiguous. In a recent study of stallion epididymal sperm, we found that sperm harvested from different parts of the epididymis (caput, corpus and cauda) had a varying, but generally poor, ability to undergo the acrosome reaction in vitro. At ejaculation, however, sperm mix with seminal plasma which contains various components, including the small membranous vesicles known as prostasomes, that may enable the sperm to undergo physiological activation. Seminal plasma components may have a 'washing' effect and help to remove 'de-capacitation' factors that coat the sperm during storage in the cauda epididymis; alternatively seminal plasma and prostasomes may contain factors that more directly promote sperm activation. This article reviews current information on the roles of epididymal and accessory gland fluids on the acquisition of fertilizing capacity by stallion sperm.

WFDC2 (HE4): a potential role in the innate immunity of the oral cavity and respiratory tract and the development of adenocarcinomas of the lung.

BACKGROUND: The Whey Acidic Protein domain is an evolutionarily conserved motif found in a number of proteins, the best studied of which are antiproteinases involved in the innate immune defence of multiple epithelia. We recently characterised the WFDC2 gene which encodes a two WAP domain-containing protein, initially suggested as a marker for epididymis, and showed that it is highly expressed in the lung and salivary gland. The precise location of WFDC2 protein in these sites has not been described. METHODS: We used immunohistochemistry to localise WFDC2 in normal tissues of the respiratory tract, naso- and oropharynx, as well as in chronically inflamed lung from Cystic Fibrosis and a range of pulmonary carcinomas. We have complemented these studies with molecular analysis of WFDC2 gene expression in primary human lung cell cultures. RESULTS: WFDC2 is expressed in some epithelial cells of the upper airways as well as in mucous cells and ducts of submucosal glands. No staining was seen in peripheral lung. Intense staining is found in major salivary glands and in minor glands of the nose, sinuses, posterior tongue and tonsil. Studies with the related protein Secretory Leukocyte Protease Inhibitor (SLPI) show that although both proteins are expressed in similar tissues, the precise cellular localisation differs. Significant increases in expression and localisation of WFDC2 are seen in patients with Cystic Fibrosis. SLPI expression was greatly reduced in the same samples. In cultures of tracheobronchial epithelial cells, expression of WFDC2 and SLPI are differentially regulated during differentiation yet WFDC2 is not induced by pro-inflammatory mediators. The majority of adenocarcinomas stain with WFDC2 whilst a significant minority of squamous, small cell and large cell carcinomas exhibit focal staining. There is no clear association with tumour grade. CONCLUSION: We believe that these studies support the hypothesis that WFDC2 may be a component of the innate immune defences of the lung, nasal and oral cavities and suggest that WFDC2 functions in concert with related WAP domain containing proteins in epithelial host defence. We also suggest that WFDC2 re-expression in lung carcinomas may prove to be associated with tumour type and should be studied in further detail.

Studies on the participation of epididymal sperm protein DE/CRISP-1 in egg activation.

Protein DE (32 kDa) associates with sperm during epididymal maturation and participates in sperm-egg fusion through its binding to complementary sites on the egg surface. In the present work we investigated the participation of DE in two mechanisms probably involved in egg activation: the ability of DE to trigger activation by its interaction with the binding sites on the egg surface (receptor model) and its ability to regulate intracellular calcium channels (sperm factor model). The incubation of eggs with DE did not promote activation parameters such as calcium oscillations or meiosis resumption. Secondly, microinjection of DE into eggs was ineffective in either eliciting calcium release or modifying oscillations induced by an activating sperm extract. Together, these results argue against the participation of DE in egg activation, restricting the activity of this protein and its egg binding sites to the sperm-egg fusion process.

Analysis of epididymal proteins during sexual maturation in male albino mice.

Androgen dependent epididymal proteins act as antigen to produce autoantibodies and affect normal fertility. In the present study, epididymal proteins were analyzed during the time of sexual maturation and their androgen dependency was studied in male albino mice. Epididymis of 21 days (Pre-pubertal), 45 days (Pubertal), 60 days (Post-pubertal), orchidectomized (15 days after surgery) and orchidectomized with testosterone-treated (15 days after treatment) mice were dissected out and analyzed. Caput, corpus and cauda epididymidis were separated and the protein extract was prepared with 0.1 M PBS for 10% SDS-PAGE analysis. Testosterone assay was performed in the experimental groups except the testosterone treated group. The electrophoretic analysis of proteins in caput, corpus and cauda epididymidis of orchidectomized animals showed the disappearance of several proteins as compared to the adult. However, the disappeared proteins started to reappear in testosterone treated animals. The results suggest that removal of testis depletes the testosterone level and causes significant alteration in epididymal proteins. These proteins need further investigation for the purpose of immunocontraception by using them as antigens.