Jae Eun Lee

Claudin-4 overexpression is associated with epigenetic derepression in gastric carcinoma

The tight junction (TJ) protein claudin-4 is aberrantly upregulated in gastric cancer, but its clinical significance and the molecular mechanisms underlying claudin-4 overexpression in gastric cancer remain unclear. Here, we investigated its roles and epigenetic mechanisms regulating CLDN4 expression in gastric cancer. We show that increased membranous expression of claudin-4 in gastric carcinoma is associated with better patient prognosis, whereas cytoplasmic claudin-4 expression did not show a significant association with prognosis. Consistent with the correlation of increased membranous claudin-4 with favorable clinicopathological factors, claudin-4 overexpression inhibited the migration and invasion of gastric cancer cells; in contrast, it did not affect cell growth. Claudin-4 expression also increased the barrier function of TJs. Claudin-4 upregulation was strongly correlated with DNA hypomethylation in both gastric tissues and gastric cancer cells. Moreover, CLDN4 expression was repressed in normal gastric tissues in association with bivalent histone modifications, and loss of repressive histone methylations and gain of active histone modifications were associated with CLDN4 overexpression in gastric cancer cells. Interestingly, CLDN4 repression could be markedly derepressed by combined treatments that simultaneously target both histone modifications and DNA demethylation in CLDN4-hypermethylated cells, whereas concomitant changes in histone methylations and acetylations are required for CLDN4 induction in CLDN4-repressed cells with low DNA methylation. Taken together, this study reveals that membranous claudin-4 expression is associated with gastric cancer progression and that it is an independent positive prognosis marker in gastric carcinoma. Furthermore, our findings suggest that epigenetic derepression may be a possible mechanism underlying CLDN4 overexpression in gastric cancer and that claudin-4 may have potential as a promising target for the treatment of gastric cancer.

Claudin-11 expression increased in spermatogenic defect in human testes

Expression of claudin-11, which builds up blood-testis barrier (BTB), was increased in decreased spermatogenesis, including in hypospermatogenesis, spermatocytic and maturation arrest, and Sertoli cell only (SCO) testes. Increased claudin-11 immunoreactivity was observed at the inter-Sertoli tight junctions in maturation arrest and in the cytoplasm of Sertoli cells in SCO. The late spermatogenic wave may negatively regulate claudin-11 gene activation and the subcellular localization of claudin-11 in Sertoli cells, thus altering the BTB in the human testis.

Hematopoetic Prostaglandin D Synthase: An ESR1-Dependent Oviductal Epithelial Cell Synthase

Oviductal disease is a primary cause of infertility, a problem that largely stems from excessive inflammation of this key reproductive organ. Our poor understanding of the mechanisms regulating oviductal inflammation restricts our ability to diagnose, treat, and/or prevent oviductal disease. Using mice, our objective was to determine the spatial localization, regulatory mechanism, and functional attributes of a hypothesized regulator of oviductal inflammation, the hematopoietic form of prostaglandin D synthase (HPGDS). Immunohistochemistry revealed specific localization of HPGDS to the oviducts epithelium. In the isthmus, expression of HPGDS was consistent. In the ampulla, expression of HPGDS appeared dependent upon stage of the estrous cycle. HPGDS was expressed in the epithelium of immature and cycling mice but not in the oviducts of estrogen receptor α knockouts. Two receptor subtypes bind PGD₂: PGD₂ receptor and G protein-coupled receptor 44. Expression of mRNA for Ptgdr was higher in the epithelial cells (EPI) than in the stroma (P < 0.05), whereas mRNA for Gpr44 was higher in the stroma than epithelium (P < 0.05). Treatment of human oviductal EPI with HQL-79, an inhibitor of HPGDS, decreased cell viability (P < 0.05). Treatment of mice with HQL-79 increased mRNA for chemokine (C-C motif) ligands 3, 4, and 19; chemokine (C-X-C motif) ligands 11 and 12; IL-13 and IL-17B; and TNF receptor superfamily, member 1b (P < 0.02 for each mRNA). Overall, these results suggest that HPGDS may play a role in the regulation of inflammation and EPI health within the oviduct.

Expression of coxsackievirus and adenovirus receptor isoforms in developing mouse bladder uroepithelium.

OBJECTIVESnTight junctions are important for uroepithelial paracellular permeability barriers. In the present study, we examined the developmental changes in the expression of coxsackievirus and adenovirus receptor (CAR) isoforms in mouse bladder uroepithelium.nnnMETHODSnMultiplex reverse transcriptase polymerase chain reaction using CAR isoform-specific primer sets and Western blotting were conducted on gestational day 19 and postnatal days 1, 7, and 55. Subcellular localization of CAR was examined, together with occludin and zonula occludens-1, in neonatal and adult bladder using light microscopy and immunofluorescence microscopy.nnnRESULTSnThe total CAR and short CAR isoform mRNA were significantly increased from gestational day 19 to birth. Long CAR isoform mRNA was transiently decreased on postnatal day 7 and had recovered during adulthood. On Western blotting, molecular weight 46-kDa CAR was abundant in the mucosa and increased postnatally. In the neonatal, as well as the adult, bladder uroepithelium, CAR immunoreactivity was observed, together with occludin and zonula occludens-1 at the apical tight junctions and basolateral contacts between the adjacent uroepithelial cells. In adult bladder uroepithelium, CAR was increased at the interface between the basal cells and basal lamina.nnnCONCLUSIONSnThe expression patterns of CAR isoforms changed during the late fetal to adult development of the mouse bladder. CAR at the apical tight junctions and cellular adhesions between the uroepithelial cells and the interfaces between the basal cells and basal lamina might support the paracellular permeability barrier and structural integrity of the uroepithelium in the mouse bladder. The expression of CAR in the uroepithelial cells can be integrated as a part of the strategy for virus-mediated gene therapy in the bladder uroepithelium.

Expression of claudin-1 and -11 in immature and mature pheasant (Phasianus colchicus) testes

The expression of claudin-1 and -11, tight junctions (TJs) proteins was examined in immature and adult pheasant (Phasianus colchicus) testes. Claudin-1 and -11 cDNA were highly similar to those of human, mice, and chicken. Claudin-1 mRNA and protein (21 kDa) levels in immature testes were higher than those of adult testis. In immature testes until 6 weeks of age, Claudin-1 was found at contacts between adjacent Sertoli cells and between Sertoli cells and germ cells. In adult testis, Claudin-1 was found in early spermatocytes migrating the blood testis barrier (BTB). Blood vessels were positive for claudin-1. Claudin-11 mRNA and protein (21 kDa) increased during adulthood development of testis. In immature testis, Claudin-11 was found in apicolateral contacts between adjacent Sertoli cells, indicating its involvement in cell adhesion in immature testis. In adult testis, strong wavy Claudin-11 immunoreactivity was parallel to basal lamina at the basal part of seminiferous epithelium, indicating that Claudin-11 at the inter-Sertoli TJs may act as a structural element of the BTB. Weak Claudin-1 and -11 immunoreactivity at contacts between Sertoli cells to elongating/elongated spermatids, meiotic germ cells, and basal lamina suggests that they also participate in the cell-cell and cell-extracellular matrix adhesion in pheasant testis. Testosterone increased claudin-11 mRNA in testis organ culture and Sertoli cell primary culture, suggesting positive regulation of claudin-11 gene by androgen in Sertoli cells of pheasant testis. This is the first report on the claudins expression at BTB in avian testis.

Postnatal changes in the expression of claudin-11 in the testes and excurrent ducts of the domestic rabbit (Oryctolagus cuniculus domesticus).

We examined the expression of claudin-11 (CLDN11) in the testes and male reproductive tracts of rabbits. The rabbit CLDN11 cDNA sequences were nearly identical with human, mouse, and bovine CLDN11. The levels of CLDN11 mRNA and protein (22 kDa) were markedly increased in the testis during adult development. On postnatal day (PND) 10, CLDN11 was colocalized with ZO-1 at the lateral contacts between adjacent Sertoli cells and was perpendicular to the basal lamina. In adult testis on PND 180, CLDN11 was codistributed with ZO1, and the pattern of immunoreactivity consisted of wavy linear tracts parallel to the basal lamina, which was different according to the spermatogenic stage. These results suggest that CLDN11 participates in inter-Sertoli cell tight junctions (TJs) at the blood-testis barrier in adult rabbits. CLDN11 was also found in the basal regions of Sertoli cells adjacent to the basal lamina in adult testis, suggesting that CLDN11 also participates in the adhesion between Sertoli cells and the basal lamina. CLDN11 mRNA and protein expressions were decreased in the adult epididymis compared with those in immature animals. In adults, CLDN11 mRNA levels were relatively high in the efferent duct, followed by those in the vas deferens, proximal corpus, and distal cauda, although low levels were observed in the initial segment and caput. On PND 10, CLDN11 immunoreactivity was identified at the apicolateral contacts between adjacent epithelial cells in the epididymis and vas deferens. In adults, CLDN11 was found in the nonciliated cells in the efferent duct and at the lateral contacts in the epithelial cells in the epididymal segments. In the caput, CLDN11 was found at the apicolateral contacts between adjacent epithelial cells, but expression was weak to negligible in the corpus of the vas deferens. CLDN11 may play an important role in TJs and cell adhesion in immature rabbit excurrent duct epithelia. In adult rabbits, CLDN11 in efferent duct epithelium and epididymal epithelium may contribute to the specific environment for sperm maturation.

Claudin 11 Inter-Sertoli Tight Junctions in the Testis of the Korean Soft-Shelled Turtle (Pelodiscus maackii)

ABSTRACT Expression of claudin 11 (CLDN11), a tight junction (TJ) protein, was examined in the Korean soft-shelled turtle (Pelodiscus maackii) testis. Spermatogenesis began during the breeding season and peaked at the end of the breeding season. Spermiation started in summer and peaked in autumn. The deduced amino acid sequence of P. maackii CLDN11 was similar to those of avian and mammalian species. During the nonbreeding season when spermatogenesis and testosterone production were active, testicular Cldn11 levels were high. In the seminiferous epithelium, strong, wavy CLDN11 strands parallel to the basement membrane delaminate the spermatogonia, and early spermatocytes are in the open compartment. Otherwise, CLDN11 was found beneath the early spermatocytes and in the Sertoli cell cytoplasm. Punctate zonula occludens 1 (ZO-1) immunoreactivity was found within the CLDN11 strands parallel to the basement membrane or at the outermost periphery of the seminiferous epithelium close to the basal lamina. During the breeding season, when circulating testosterone levels and spermatogenic activity was low, testicular CLDN11 level was lower than those during the nonbreeding season. CLDN11 was found at apicolateral contact sites between adjacent Sertoli cells devoid of the postmeiotic germ cells. At this time, lanthanum tracer diffused to the adluminal compartment of seminiferous epithelium. In cultured testis tissues, testosterone propionate significantly increased the level of Cldn11 mRNA. In P. maackii testis, CLDN11 participates in the development of the blood-testis barrier (BTB), where the CLDN11 expression was coupled with spermatogenic activity and circulating androgen levels, indicating the conserved nature of TJs expressing CLDN11 at the BTB in amniotes.

Abstract P2-10-39: Does E-cadherin or N-cadherin or epithelial-mesenchymal transition have a probability of clinical implication of the prognostic marker in invasive ductal carcinoma?

Purpose: Epithelial to mesenchymal transition (EMT) is widely accepted hypothesis of progression and metastasis in carcinoma. EMT can be explained as a loss of epithelial features and gain of mesenchymal properties, and in the same manner, it can be characterized by down-regulation of epithelial cellular markers, such as E-cadherin (EC) and up-regulation of mesenchymal cellular markers, such as N-cadherin (NC). Breast cancer has a diverse clinical spectrum, treatments and prognosis according to clinicopathologic characteristics and immunohistochemical features. Identifying the presence of EMT also would be able to predict the prognosis and design better and customized therapy. Our objective was to evaluate expression of the EC/NC and EMT and correlation between EMT and clinicopathologic, immunohistochemical features and to investigate clinical implication of the EC/NC and EMT as prognostic marker in breast cancer. Methods: Using the clinicopathologic data of 480 patients who underwent operation for invasive ductal carcinoma (IDC) during February 2001 to December 2008. 282 patients with good quality of paraffin embedded tissue block were selected. The ER and PR immunohistochemistry was scored using the Allred Score. EC and NC expression were classified on the basis of intensity of immunohistochemistry staining. The gain of a mesenchymal marker or loss of an epithelial marker was interpreted as EMT-positive. In this study, breast cancer was classified into four types according to the immunohistochemical expression of ER, PR and HER2 expression on immunohistochemical analysis and/or with HER2 gene by FISH analysis: luminal A, luminal B, HER2-enriched, and triple negative type. Results: Median follow-up period was 52.6 months (range 1–113) and median age was 49 years (range 27–79). Of total 282 cases, 152 (53.9%) cases were classified as luminal A, 32 (11.3%) classified as luminal B, 43 (15.2%) cases were HER2-enriched and 55 (19.5%) cases were triple negative type. Expression of a mesenchymal marker, NC was observed in 22 (7.8%) patients and loss of an epithelial marker, EC was observed in 57 (20.2%) cases. A total of 75 (26.6%) were interpreted as EMT-positive. NC score was associated with HER2-enriched, and triple negative molecular subtype (P = 0.0234). NC status was associated with HER2 positive status (P = 0.0456) and HER2-enriched, triple negative molecular subtype (P = 0.0029). EMT correlated with ER negative status (P = 0.0199). T stage (P Conclusion: NC status was associated with HER2 positive status and HER2-enriched, triple negative molecular subtype. And EMT correlated with ER negative status. EC/NC and EMT status have relevance to a specific molecular subtype or a probability of clinical implication of the poor prognostic marker in breast cancer. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P2-10-39.

Abstract 101: Claudin-4 overexpression is associated with epigenetic derepression in gastric carcinoma

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FLnnThe tight junction protein claudin-4 is aberrantly up-regulated in gastric cancer, but its clinical significance and the molecular mechanism underlying claudin-4 overexpression in gastric cancer remain unclear. Here, we investigated its roles and epigenetic mechanism regulating CLDN4 expression in gastric cancer. We show that increased membranous expression of claudin-4 in gastric carcinoma is associated with better patient prognosis, whereas cytoplasmic claudin-4 expression didnt show a significant association with prognosis. Consistent with the correlation of increased membranous claudin-4 with favorable clinicopathological factors, claudin-4 overexpression inhibited the migration and invasion of gastric cancer cells; in contrast, it didnt affect cell growth. Claudin-4 expression also increased the barrier function of tight junctions. Claudin-4 up-regulation was strongly correlated with DNA hypomethylation in both gastric tissues and gastric cancer cells. Moreover, loss of repressive histone methylations and gain of active histone modifications were associated with CLDN4 overexpression in gastric cancer cells. Interestingly, CLDN4 repression could be markedly derepressed by combined treatments that simultaneously target both histone modifications and DNA demethylation in CLDN4-hypermethylated cells, whereas concomitant changes in histone methylations and acetylations are required for CLDN4 induction in CLDN4-repressed cells with low DNA methylation. Taken together, this study reveals that membranous claudin-4 expression is associated with gastric cancer progression as well as it is an independent positive prognosis marker in gastric carcinoma. Furthermore, our findings suggest that epigenetic derepression may be a possible mechanism underlying CLDN4 overexpression in gastric cancer and that claudin-4 may have potential as a promising target for the treatment of gastric cancer.nnCitation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 101. doi:10.1158/1538-7445.AM2011-101