Apoptosis In Ishikawa Cells Research Articles

Aqueous extract of pulp of maqui-berry (Aristotelia chilensis) induces apoptosis in human endometrial carcinoma ishikawa cells via mitochondrial and endoplasmic reticulum stress pathways

In Chilean traditional medicine, Aristotelia chilensis (Mol) Stuntz is commonly utilized for a variety of conditions (particularly digestive problems). We investigated the effects of an aqueous extract from the fruit pulp of A. chilensis (Maqui berry: M2) on endoplasmic reticulum stress and mitochondrial pathway-induced early apoptosis in Ishikawa cells. Using the MTT assay, the aqueous extract was found to have an inhibitory impact on human endometrial cancer Ishikawa cells, with an IC50 of 5.36 μg/mL. The changes in Ishikawa cells under the effect of M2 were detected by flow cytometry with cycle arrest in S phase, reduced mitochondrial membrane potential, ROS and calcium ion aggregation. Western blotting and RT-qPCR were used to identify the intracellular mRNA and protein alterations. The presence of M2 resulted in the up-regulation of mitochondrial pathway-related proteins Bad, Bax, and Cyto C, down-regulation of Bcl-2, and up-regulation of endoplasmic reticulum stress-related proteins IRE1α, p-eif2α, ATF4, and CHOP. The findings imply that A. chilensis pulp may be used as a possible raw material for the innovative functional applications.

Saikosaponin D exerts cytotoxicity on human endometrial cancer ishikawa cells by inducing apoptosis and inhibiting metastasis through MAPK pathways.

Saikosaponin D (SSD) is one of plant secondary metabolic active substance with effective anti-tumor ability; however, the toxicity of Saikosaponin D on human endometrial cancer Ishikawa cells is still unclear. Our results revealed that SSD displayed cytotoxicity on the Ishikawa cell with an IC50 = 15.69 μM, but was non-toxic to the human normal cell line HEK293. SSD could upregulate p21 and Cyclin B to keep cells in the G2/M stage. In addition, it activated the death receptor and mitochondrion routes to induce apoptosis in Ishikawa cells. The transwell chamber and wound healing results showed that SSD inhibited the cell migration and invasion. In addition, we found that it was closely related to the MAPK cascade pathway, and it could mediate the three classical MAPK pathways to block cell metastasis. In conclusion, SSD could be potentially beneficial as a natural secondary metabolite in preventing and treating endometrial carcinoma.

Isorhamnetin Induces Apoptosis and Suppresses Metastasis of Human Endometrial Carcinoma Ishikawa Cells via Endoplasmic Reticulum Stress Promotion and Matrix Metalloproteinase-2/9 Inhibition In Vitro and In Vivo.

Endometrial cancer (EC) is a very common female cancer which has attracted more and more attention. According to the individual patient's condition, the current treatment of EC patients is mainly based on surgery, which is supplemented by chemotherapy, radiotherapy, and endocrine intervention. However, these existing treatment strategies also have some inevitable limitations. Therefore, it is particularly important to find an active ingredient with low toxicity and a high safety profile against EC. Isorhamnetin is a flavonoid known to be present in a variety of plants, such as sea buckthorn, dry willow, and wolfberry. In recent years, the anti-tumor effects of isorhamnetin have been reported. In our study, isorhamnetin was shown to induce apoptosis in Ishikawa cells by inducing the endogenous mitochondrial apoptotic pathway and exogenous death receptor pathway, promoting the endoplasmic reticulum stress-related pathway, and activating the corresponding markers of UPR response. In addition, isorhamnetin affected the expression of MMP2 and MMP9-related proteins in vitro and in vivo and eventually repressed metastasis. Therefore, isorhamnetin can be used as a promising medicinal material for the treatment of EC.

Blocking PARP activity with the inhibitor veliparib enhances radiotherapy sensitivity in endometrial carcinoma.

ObjectiveOur study aimed to investigate the potential clinical utility of a poly(ADP‐ribose) polymerase (PARP) inhibitor, veliparib (ABT‐888), as a radiosensitizer in the medication of endometrial carcinoma (EC).MethodsHuman Ishikawa endometrial adenocarcinoma cells were treated with veliparib, radiotherapy (RT), or combination treatment. The viabilities, radiosensitivity enhancement ratio (sensitizer enhancement ratio (SER), and apoptosis of Ishikawa cells were, respectively, evaluated by Cell Counting Kit‐8 (CCK‐8), colony formation experiment, and flow cytometry. The tumor growth was assessed by xenograft mice models. Western blot assay investigated the expression of DNA damage and apoptosis‐related proteins in vivo and in vitro.ResultsCell Counting Kit‐8 revealed that the 10% inhibition concentration (IC10) and 50% inhibition concentration (IC50) values of veliparib‐treated Ishikawa cells were 1.7 and 133.5 µM, respectively. The SER of veliparib combined with RT was 1.229 in vitro. Flow cytometry analysis results indicated that the apoptosis rate of the veliparib + RT group was markedly higher than that of the RT group in vitro (p < 0.05). Furthermore, in vivo data revealed that veliparib + RT treatment significantly decreased tumor growth compared with single treatments of veliparib or RT and with the control group (p < 0.05). Then western blot confirmed the levels of anti‐phospho‐histone (γH2AX), caspase‐3, and B‐cell lymphoma 2 (Bcl‐2) associated protein X (Bax) were significantly higher in the veliparib + RT group, while the level of Bcl‐2 was lower compared with that of the RT group (p < 0.05), both in vivo and in vitro.ConclusionOur results indicate that veliparib in combination with RT markedly improved the therapeutic efficiency in human endometrial carcinoma.

Hypoxic induction of apoptosis occurs through HIF-1α and accompanies mammalian sterile 20-like kinase 2 cleavage in human endometrial adenocarcinoma Ishikawa cells

The incidence of endometrial cancer is increasing worldwide. One of the main causes of this cancer is a hormone imbalance; progesterone derivatives have been used for treatment. However, reports have shown that hypoxia plays important and possibly beneficial roles in endometrial function. Here, we show the effect of hypoxia on the proliferation of human endometrial adenocarcinoma Ishikawa cells. Hypoxia induced caspase-dependent apoptosis in Ishikawa cells. Overexpression and siRNA-mediated knockdown of hypoxia-inducible factor-1α (HIF-1α) confirmed that HIF-1α accelerates hypoxia-induced cell death. Treatment with dimethyloxalglycine, which stabilizes HIF-1α, suppressed cell proliferation. Kaplan–Meier analysis showed that the expression level of HIF-1α has a significant positive effect on the survival rate of endometrial cancer patients. In our search for cellular targets involved in hypoxic apoptosis, we noticed that mammalian sterile 20-like kinase 2 (MST2), a member of the Hippo pathway, was positively correlated with HIF-1α expression in 176 endometrial cancer patients extracted from the TCGA database. Hypoxia induced caspase-dependent MST2 cleavage. In addition, a MST2 inhibitor suppressed HIF-1α-mediated reporter activity. These results suggest HIF-1α and the Hippo signaling pathway are involved in endometrial cancer.

Knockdown of LMTK3 in the Endometrioid Adenocarcinoma Cell Line Ishikawa: Inhibition of Growth and Estrogen Receptor α.

ObjectiveThe curative effect of high-efficiency progesterone and other therapeutic drugs for endometrioid adenocarcinoma patients with preservation of reproductive capacity has not been satisfactory so far. Novel therapeutic drugs need to be explored.MethodsWe investigated the cytoplastic and nuclear expression levels of LMTK3 between endometrioid adenocarcinoma tissues and adjacent endometrial tissues by immunohistochemistry. We detected the effects of LMTK3 on cell viability of Ishikawa cells by CCK-8. We detected the effects of LMTK3 on cell cycle and apoptosis of Ishikawa cells by flow cytometry. We also detected the effects of LMTK3 knockdown on mRNA and protein levels of ERα by qRT-PCR and western blotting, respectively. We also used the cBioPortal online database to analyze the coexpression of LMTK3 and ESR1 in 1647 UCEC samples.ResultsWe used TMAs to identify that LMTK3 was mainly detected in the cytoplasm of endometrioid tissues, and cytoplasmic LMTK3 expression in endometrioid tissues was higher than that in adjacent endometrial tissues (P < 0.05). LMTK3 knockdown decreased the proliferation of Ishikawa cells through decreasing cell viability (P < 0.01), increasing G1 (P < 0.001) arrest, and promoting apoptosis (P < 0.01). There was a positive correlation between the mRNA expression levels of LMTK3 and ESR1 (Spearman: P=2.011e-5, R=0.13; Pearson: P=7.18e-8, R=0.17). Knockdown of LMTK3 also reduced the mRNA (P < 0.001) and protein (P < 0.001) levels of ERα.ConclusionsInhibitors of LMTK3 may be a possible future treatment for ERα and LMTK3 highly expressed endometrioid adenocarcinoma following appropriate studies.

DJ-1 activates the noncanonical NF-κB pathway via interaction with Cezanne to inhibit the apoptosis and promote the proliferation of Ishikawa cells.

Endometrial cancer is generally one of the most evident malignant tumours of the female reproductive system, and the mechanisms underlying its cell proliferation and apoptosis are key to research in gynaecological oncology. In the paper, the in-depth molecular mechanism by which DJ-1 protein regulates the proliferation and apoptosis of Ishikawa cells was investigated. DJ-1 knockdown and overexpressing Ishikawa stable cell lines were established by lentiviral transduction. The levels of DJ-1 and noncanonical NF-κB signaling key proteins were evaluated by Western blotting. Cell counting kit-8 (CCK-8) and flow cytometry were applied to analyze the cell viability and apoptosis. Co-immunoprecipitation experiment was utilized to assess the DJ-1-Cezanne interaction. The results showed that DJ-1 overexpression conferred apoptosis resistance and high proliferation on Ishikawa cells, while DJ-1 knockdown in Ishikawa cells produced the opposite results. These findings again suggested that DJ-1 inhibits the apoptosis and promotes the proliferation of Ishikawa cells. More crucially, further data showed that the noncanonical NF-κB activation was required for the regulation of Ishikawa cell proliferation and apoptosis by DJ-1. Meanwhile, it was found that noncanonical NF-κB pathway may be activated by DJ-1 interacting with and negatively regulating Cezanne in Ishikawa cells. Overall, this work revealed that DJ-1 associates with and negatively regulates Cezanne and consequently triggers the noncanonical NF-κB activation, thereby regulating Ishikawa cell proliferation and apoptosis.

Long Non-Coding RNA Small Nucleolar RNA Host Gene 1 and MicroRNA-100-3p Expression in Endometrial Carcinoma and Its Effect on the Proliferation and Apoptosis of Ishikawa Cells

The aim of this study was to explore LncRNA SNHG1 and miRNA-100-3p expression in endometrial carcinoma and its effect on the proliferation and apoptosis of Ishikawa cells. A qRT-PCR assay was conducted to determine SNHG1 and miRNA-100-3p expression in endometrial cancer tissues and paracancerous tissues. Human endometrial cancer Ishikawa cells were cultured in vitro. Si-NC, si-SNHG1, si-SNHG1, anti-miRNA-NC, and si-SNHG1 were transfected into Ishikawa cells with anti-miRNA-519b-3p. A qRT-PCR assay was performed to determine SNHG1 and miRNA-100-3p expression, and the CCK-8 method was used to determine cell proliferation. Flow cytometry was conducted to determine cell cycle and apoptosis rate and a dualluciferase reporter experiment was carried out to test the targeting association between SNHG1 and miRNA-100-3p. Cleave Caspase-3, CHOP, and ATF4 expression were determined with the Western Blot method. SNHG1 expression level and miRNA-519b-3p expression level were much higher and much lower, respectively, in endometrial cancer tissues than in paracancerous tissues (P &lt; 0.05). Transfection of si-SNHG1 can greatly attenuate cell viability and S cell ratio (P &lt; 0.05), and increase G0/G1 cell ratio, apoptosis rate, Cleaved Caspase-3, CHOP, and ATF4 protein level (P &lt; 0.05) compared to the si-NC group. Furthermore, the double luciferase reporter experiment confirmed that SNHG1 can competitively combine withmiRNA-100-3p. Also, co-transfection of si-SNHG1 and anti-miRNA-100-3p could significantly increase cell viability and S cell ratio (P &lt; 0.05), and decrease G0/G1 cell ratio and apoptosis rate, and Cleaved Caspase-3, CHOP, and ATF4 protein levels compared to si-SNHG1+anti-miRNA-NC (P &lt; 0.05). Interfering with SNHG1 could inhibit the proliferation of Ishikawa cells and promote apoptosis by upregulating miRNA-100-3p expression.

Effects of Gallic Acid on Endometrial Cancer Cells in Two and Three Dimensional Cell Culture Models.

Background and Aim:Cell culture studies are an indispensable tools used to understand basic physiological, biophysical and biomolecular mechanisms. Although traditional two-dimensional (2D) cell culture models are more preferred in experimental studies, three-dimensional (3D) cell culture models, attract more attention due to several advantages including mimicking tumor physiology, biochemistry and biomechanics. We aimed to investigate the effects of Gallic Acid, an antimutagenic, antioxidant and anticarcinogenic agent, on both 2D and 3D cultured endometrial cancer cells for the first time. Methods:IC50 values were determined in 2D and 3D cultured endometrial cancer cells exposed to different doses of GA. In the 2D culture model exposed to GA, Caspase 3 expression levels were analyzed. In addition, the effect of GA on the migration of 2D cultured endometrium cancer cells was investigated. Results:IC50 value in the 3D model was found significantly higher than the 2D model. In 2D culture model, Caspase 3 expression and apoptosis was increased significantly in cells of GA exposed group compared to the control group. GA did not have a significant effect on the migration profile of cells. Conclusion:Gallic Acid is shown to induce apoptosis in Ishikawa cells via Caspase 3 activation. We demonstrated a significantly higher IC50 level in 3D model which provide evidence to prefer 3D models in drug-test trials. The data obtained in the current study will provide a basis for further experiments to analyze the effects of GA on endometrial cancer and to develop strategies for clinical treatment.

Upregulation of miR-183-5p is responsible for the promotion of apoptosis and inhibition of the epithelial-mesenchymal transition, proliferation, invasion and migration of human endometrial cancer cells by downregulating Ezrin.

Endometrial cancer is a life-threatening malignancy that affects women all over the world, and it has an increasing incidence. MicroRNAs (miRNAs/miRs) have been reported to be involved in cellular activities in endometrial cancer. The present study aimed to examine the effects of miR-183-5p on the epithelial-mesenchymal transition (EMT), proliferation, invasion, migration and apoptosis of human endometrial cancer cells by targeting Ezrin. Primary endometrial cancer tissues and adjacent normal tissues were obtained for the investigation. The protein expression of Ezrin in tissues was detected by immunohistochemistry. The expression level of miR-183-5p and the mRNA and protein expression levels of Ezrin and EMT-associated genes were determined by reverse transcription-quantitative polymerase chain reaction and western blot analyses. Endometrial cancer cells were treated with miR-183-5p inhibitors, small interfering RNA targeting Ezrin or miR-183-5p inhibitors. Cell proliferation, cell cycle, apoptosis, migration and invasion were then evaluated using an MTT assay, flow cytometry, scratch test and Transwell assay, respectively. Compared with normal adjacent tissues, the expression of miR-183-5p was decreased in endometrial cancer tissues, and the expression of Ezrin was significantly increased in endometrial cancer tissues. The protein expression of Ezrin was correlated with the severity and poor prognosis of endometrial cancer. Notably, the target prediction program and the luciferase reporter gene assay confirmed that miR-183-5p targeted and negatively regulated the expression of Ezrin. In vivo experiments revealed that the increased expression of miR-183-5p and decreased expression of Ezrin inhibited EMT, cell proliferation, migration and invasion, but promoted cell apoptosis in Ishikawa cells. These results suggested that the upregulated expression of miR-183-5p promoted apoptosis and suppressed the EMT, proliferation, invasion and migration of human endometrial cancer cells by downregulating Ezrin.

CIP2A is overexpressed in human endometrioid adenocarcinoma and regulates cell proliferation, invasion and apoptosis

ObjectiveCancerous inhibitor of protein phosphatase 2A (CIP2A) is a recently identified oncoprotein that stabilizes c-Myc and promotes cell proliferation and transformation. Here, we investigated the clinical significance and biological function of CIP2A in endometrial cancer. MethodCIP2A expression was assessed in normal endometrium, endometrial hyperplasia, endometrial atypical hyperplasia, and endometrioid adenocarcinoma tissues using immunohistochemistry, western blot, and RT-PCR. The effect of reduced CIP2A expression was assessed by siRNA knockdown in Ishikawa and An3ca endometrial cell lines. The roles of CIP2A in proliferation, apoptosis, and the cell cycle were assessed using CCK-8 assays, colony formation assays, and flow cytometry, respectively. ResultsOur results show that CIP2A expression was higher in endometrioid adenocarcinoma tissues and cell lines. Furthermore, CIP2A siRNA significantly reduced the proliferation rate and invasion of Ishikawa and An3ca cells, and induced a significant level of apoptosis in Ishikawa cells. Moreover, CIP2A depletion resulted in reduced c-Myc and cyclin D1 protein levels, and increased caspase-3 expression. ConclusionsCIP2A is overexpressed in endometrioid adenocarcinoma and CIP2A promotes the malignant growth and invasion,decrease apoptosis in entometrioid adenocarcinoma cell lines. These results validate that CIP2A plays an important role in the carcinogenesis of endometrioid adenocarcinoma and establishes CIP2A as a clinically relevant oncoprotein and may presents a promising therapeutic target for cancer treatment.

Pseudolaric acid B induces endometrial cancer Ishikawa cell apoptosis and inhibits metastasis through AKT-GSK-3β and ERK1/2 signaling pathways.

Pseudolaric acid B (PAB) is the most active constituent extracted from the bark of Pseudolarix kaempferi, which has been used as an antifungal remedy in traditional Chinese medicine. It is reported to have cytotoxicity to many tumor cell lines. In this study, we investigated the effects of PAB against human endometrial cancer Ishikawa cells. We found that PAB inhibited Ishikawa cell proliferation, and induced cell apoptosis and G2/M phase arrest through a mechanism involving AKT-GSK-3β and ERK1/2 signaling pathways. PAB also suppressed the Ishikawa cell adhesion, invasion, migration, and colony formation ability by increasing the expression of E-cadherin, Ezrin, and Kiss-1, and decreasing the expression of matrix metalloproteinase-9 and vascular endothelial growth factor. Taken together, these data indicated that PAB can be expected to be a novel treatment agent for endometrial cancer therapy.

MiR-944 acts as a prognostic marker and promotes the tumor progression in endometrial cancer.

microRNA-944 (miR-944) has been reported to be deregulation and play either tumor suppressive or oncogenic function in human malignancies. Previous studies have reported that miR-944 is overexpressed in gynecological malignancies including cervical cancer and breast cancer. While, the clinical significance of miR-944 and its function in human endometrial carcinoma (EC) keep unknown. The levels of miR-944 were analyzed in 68 EC tissues and 20 normal endometrial tissues. Results showed that miR-944 was significantly overexpressed in EC tissues compared to normal endometrial tissues. In addition, increased levels of miR-944 also observed in EC cell lines. Clinicopathological analysis verified that miR-944 expression was associated with international federation of gynecology and obstetrics (FIGO) stages and pathology classification of EC. Moreover, Kaplan-Meier analysis found that miR-944 highly expressing EC patients showed a notable shorter survival. Further experiments revealed that miR-944 knockdown significantly inhibited growth and cell cycle progression while facilitated apoptosis in Ishikawa cells. In turn, miR-944 overexpression prominently promoted proliferation and cell cycle progression, and suppressed apoptosis in KLE cells. In vivo experiments further confirmed that miR-944 silencing notably restrained the tumor growth of EC in nude mice. Mechanically, cell adhesion molecule 2 (CADM2) was recognized as a direct downstream target of miR-944 in EC cells. An inverse correlation between miR-944 and CADM2 expression was observed in EC tissues. Interestingly, CADM2 overexpression showed similar effects to miR-944 knockdown in Ishikawa cells with decreased growth, cell cycle arrest at G1 phase and increased apoptosis. Taken together, this work support the first evidence that miR-944 can be potentially used as a promising biomarker and novel therapeutic target for human EC.

CRKL overexpression promotes cell proliferation and inhibits apoptosis in endometrial carcinoma.

The v-Crk avian sarcoma virus CT10 oncogene homolog-like (CRKL) protein is important in cancer progression. However, its expression pattern and biological roles in human endometrial carcinoma remain unexplored. The potential mechanism of CRKL-induced cancer progression is still unclear. The present study aimed to explore the expression pattern and biological roles of CRKL in human endometrial carcinoma. Using immunohistochemistry, it was observed that the CRKL protein was overexpressed in 50.5% (44/87) of endometrial carcinoma tissues. Plasmid transfection of CRKL into Ishikawa cells was performed, and CRKL overexpression promoted cell proliferation, colony formation and cell cycle transition in the transfected cells. In addition, CRKL overexpression inhibited cell apoptosis in Ishikawa cells treated with cisplatin, with decreased caspase-3 and caspase-9 cleavage. Further analysis revealed that CRKL upregulated the expression of cyclin D1, cyclin E, B cell lymphoma (Bcl)-2 and survivin, and downregulated Bcl-2 associated X protein expression. In conclusion, the present study demonstrated that CRKL overexpression in endometrial carcinoma contributes to malignant cell growth and resistance to apoptosis, possibly through Bcl-2.

Normal endometrial stromal cells regulate 17β-estradiol-induced epithelial-mesenchymal transition via slug and E-cadherin in endometrial adenocarcinoma cells in vitro

Stroma–tumor communication participates in the pathogenesis of endometrial carcinomas. In previous studies, we found that normal stromal cells inhibited the growth of endometrial carcinoma cells. Here, we investigated the role of normal stromal cells in the epithelial-mesenchymal transition (EMT) of endometrial carcinoma cells and explored the possible mechanism implied. We found that conditioned medium (CM) by normal endometrial stromal cells (NSC) reduced cell growth and induced cell apoptosis in Ishikawa cells. CM by NSC inhibited 17β-estradiol-induced cell growth and apoptosis decrease in Ishikawa cells. Moreover, CM by NSC inhibited the migration and invasion, and 17β-estradiol-induced migration and invasion in Ishikawa cells. Meanwhile, CM by NSC decreased Slug expression and 17β-estradiol-induced Slug expression, increased E-cadherin expression and abolished 17β-estradiol-induced E-cadherin reduction in Ishikawa cells. In conclusion, normal stromal factors can inhibit 17β-estradiol-induced cell proliferation and apoptosis inhibition, and abolished 17β-estradiol-induced EMT in endometrial cancer cell via regulating E-cadherin and Slug expression.

Black cohosh inhibits 17β-estradiol-induced cell proliferation of endometrial adenocarcinoma cells

This study was conducted to investigate the effect of black cohosh (BC) extract on the proliferation and apoptosis of Ishikawa cells. Ishikawa human endometrial adenocarcinoma cells were treated with or without BC (1, 5, 10 and 25 μM) and cell proliferation and cytotoxicity were measured by CCK-8 assays and flow cytometry analysis. Additionally, Ishikawa cells were treated with 17β-estradiol (E2), E2 + progesterone and E2 + BC (5 and 10 μM) and the effect of BC and progesterone on E2-induced cell proliferation was analyzed. BC decreased the proliferation of Ishikawa cells at a dose-dependent rate compared with the control group (p < 0.05). The proliferation of Ishikawa cells increased in the presence of E2, whereas the subsequent addition of progesterone or BC decreased proliferation to the level of the control group (p < 0.05). The inhibitory effect of BC on E2-induced cell proliferation was greater than the inhibitory effect of progesterone. In conclusion, BC induces apoptosis in Ishikawa cells and suppresses E2-induced cell proliferation in Ishikawa cells. BC could be considered a candidate co-treatment agent of estrogen-dependent tumors, especially those involving endometrial cells.

Fenretinide: a novel treatment for endometrial cancer.

Resistance to progestin treatment is a major hurdle in the treatment of advanced and reoccurring endometrial cancer. Fenretinide is a synthetic retinoid that has been evaluated in clinical trials as a cancer therapeutic and chemo-preventive agent. Fenretinide has been established to be cytotoxic to many kinds of cancer cells. In the present study, we demonstrate that fenretinide decreased cell viability and induced apoptosis in Ishikawa cells, which are an endometrial cancer cell line, in dose dependent manner in-vitro. This effect was found to be independent of retinoic acid nuclear receptor signaling pathway. Further, we have shown that this induction of apoptosis by fenretinide may be caused by increased retinol uptake via STRA6. Silencing of STRA6 was shown to decrease apoptosis which was inhibited by knockdown of STRA6 expression in Ishikawa cells. Results of an in-vivo study demonstrated that intraperitoneal injections of fenretinide in endometrial cancer tumors (created using Ishikawa cells) in mice inhibited tumor growth effectively. Immunohistochemistry of mice tumors showed a decrease in Ki67 expression and an increase in cleaved caspase-3 staining after fenretinide treatment when compared to vehicle treated mice. Collectively, our results are the first to establish the efficacy of fenretinide as an antitumor agent for endometrial cancer both in-vitro and in-vivo, providing a valuable rationale for initiating more preclinical studies and clinical trials using fenretinide for the treatment of endometrial cancer.

Expression of the ELAV-like protein HuR in the cytoplasm is associated with endometrial carcinoma progression

Human antigen R (HuR) is an mRNA-binding factor that belongs to the embryonic lethal abnormal vision/Hu protein family which may function as a tumor maintenance gene in a variety of carcinomas. However, there is no study to analyze HuR expression in endometrial carcinoma. Here, we investigated the expression of HuR in endometrial carcinoma carcinogenesis and subsequent progression. The expression of HuR and estrogen receptor alpha (ER-α) protein was examined by immunohistochemistry on paraffin embedding specimens containing endometrial carcinoma and adjacent non-cancerous tissues. Short hairpin RNA against HuR was transfected to investigate the role of HuR in regulating the expression of ER-α and progression in endometrial carcinoma. Cell viability and cycle were measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry, respectively. Apoptosis was examined by annexin V apoptosis assay. Our study result show that cytoplasmic HuR expression is more frequent in poorly differentiated carcinomas (p = 0.005), advanced stage (p = 0.020), and positive ER-α expression (p = 0.026). Three HuR short hairpin RNAs (shRNAs) were transfected into Ishikawa cells, and we selected the most effective shRNA for the following experiments. After the transfection, the ER-α protein level was decreased. Further, decreased expression of HuR resulted in the inhibition of proliferation and induced apoptosis in Ishikawa cells. Our results showed that HuR could be a causal factor of ER-α regulation in Ishikawa cells and thus may induce the hormone-dependent endometrial carcinoma.

Inhibitory effects of androstenedione on endometrial cells: implications for poor reproductive outcome among women with androgen excess

ObjectiveAndrostenedione (A4) is an androgen that can be metabolized by aromatase to estrone, but the effects of A4 on endometrial cell proliferation either as an androgen or via conversion to estrogens are unknown. The aim of this study was to investigate A4 effects on Ishikawa cells in culture. Study designIshikawa cells were treated with increasing concentrations of A4 (0–1000pmol) for 4 days. Cell proliferation was measured by the (4,5-dimethylthiaxol-2-yi)-2,5-diphenyltetraxolium bromide (MTT) assay. Apoptosis was analyzed through Annexin-V/propidium iodide (PI) staining and flow cytometry: 17β-hydroxy steroid dehydrogenase type 1 (17β-HSD1) and aromatase mRNA expression was measured by reverse transcription-polymerase chain reaction (RT-PCR). Western blotting was used to detect cell signaling expressions of Akt/MAPK. ResultsA4 treatment (1nM) decreased cell proliferation and increased apoptosis, as demonstrated by MTT and flow cytometry or related gene expression. The cellular responses induced by A4 treatment were mediated by activation of the Akt and MAPK signaling pathway. Treatment had no effect on 17β-HSD1 and aromatase expression. ConclusionA4 treatment induced growth inhibition and apoptosis of Ishikawa cells through activation of the Akt/MAPK pathway. Effects of A4 on Ishikawa cells occurred in the absence of increased 17β-HSD1 and aromatase expression. These results imply that women with excessive androgen, such as polycystic ovary syndrome, experience poor reproductive outcomes through androgen-regulated mechanisms.

MiR‐34c oligonucleotide enhances chemosensitivity of Ishikawa cell to cisplatin by inducing apoptosis

Expression of miR-34c is significantly reduced in human endometrial cancer cells and miR-34c regulates multiple genes associated with cancer cell proliferation, apoptosis and invasiveness. Thus, miR-34c can theoretically become a target that enhances chemotherapy. Upregulation of miR-34c to enhance the chemotherapeutic effect of cisplatin (DDP) has not been studied in human endometrial cancer cells before. In an in vitro study, the human endometrial cancer cell line, Ishikawa, was treated with DDP and miR-34c mimics, alone or in combination. IC50 values were dramatically decreased in cells treated with miR-34c mimics when combined with DDP, to a greater extent than those treated with DDP alone. Furthermore, miR-34c mimics significantly enhanced apoptosis in Ishikawa cells by inhibiting IL-6R expression. Therefore, a combination of miR-34c mimics and DDP could be an effective therapeutic strategy for controlling Ishikawa cell proliferation.