Transformation of different human breast epithelial cell types leads to distinct tumor phenotypes.

We investigated the influence of normal cell phenotype on the neoplastic phenotype by comparing tumors derived from two different normal human mammary epithelial cell populations, one of which was isolated using a new culture medium. Transformation of these two cell populations with the same set of genetic elements yielded cells that formed tumor xenografts exhibiting major differences in histopathology, tumorigenicity, and metastatic behavior. While one cell type (HMECs) yielded squamous cell carcinomas, the other cell type (BPECs) yielded tumors closely resembling human breast adenocarcinomas. Transformed BPECs gave rise to lung metastases and were up to 10(4)-fold more tumorigenic than transformed HMECs, which are nonmetastatic. Hence, the pre-existing differences between BPECs and HMECs strongly influence the phenotypes of their transformed derivatives.

[Reasons for older adults independently continuing exercise after a supervised Square-Stepping Exercise intervention].

OBJECTIVE: Square-Stepping Exercise (SSE), composed of movements similar to walking, involves varied movements in multiple directions and is performed on a thin mat (100 x 250 cm) that is partitioned into 40 squares (25 cm each). We introduced SSE to a group of older adults for three months as a supervised intervention. After this intervention period, the participants continued SSE without supervision for four years. The current study was conducted to determine why the participants independently continued SSE. METHODS: Among 52 older adults who attended the SSE intervention, 40 continued SSE (continued group) and 12 discontinued (discontinued group). Seven in the continued group were excluded from analyses because of low attendance rates. Each of the remainder (n = 33) was independently interviewed and asked why he/she had continued SSE. The average interview time for the continued group was 12 minutes. Twelve in the discontinued group were investigated for exercise habits by postal questionnaire. RESULTS: The participants in the continued group noted two to six reasons for continuation of SSE. After analyzing data inductively, the answers were categorized as follows: (1) friends and social communication; (2) equitable management of group activity; (3) expectation of health from exercise; (4) simple-easy exercise; and (5) family support for exercise. The participants in the discontinued group reported that 89% of them continually did walking, muscular strength exercise, and calisthenics. CONCLUSION: We found that reasons why adoption of SSE as an intervention program for older adults enhance their exercise adherence in the long term.

Inhibition of phosphatidylinositol 3-kinase increases efficacy of cisplatin in in vivo ovarian cancer models.

The phosphatidylinositol 3-kinase (PI3K)/Akt cascade has an important role in the resistance of ovarian cancer cells to cisplatin in vitro; however, there have been no reports about whether blocking the PI3K/Akt cascade enhances the sensitivity to cisplatin in vivo. We investigated whether inhibition of PI3K increased the efficacy of cisplatin in an in vivo ovarian cancer model. Blocking the PI3K/Akt cascade with a PI3K inhibitor (wortmannin) increased the efficacy of cisplatin-induced inhibition of intraabdominal dissemination and production of ascites in athymic nude mice inoculated ip with the Caov-3 human ovarian cancer cell line. In addition, wortmannin increased the efficacy of cisplatin-induced apoptosis in tumors cells. There were no detectable side effects in mice treated with wortmannin. Moreover, the antitumor effect of cisplatin detected in mice inoculated with Caov-3 cells stably transfected with empty vector was significantly attenuated, compared with mice inoculated with Caov-3 cells stably transfected with a dominant-negative Akt, K179M-Akt. We confirmed that wortmannin blocked Akt phosphorylation and the downstream targets of the PI3K/Akt cascade, such as BAD (Bcl-2-associated death protein) and nuclear factor-kappaB in vivo by immunohistochemical staining and Western blotting. In accordance with the previously reported in vitro results, these in vivo results support the idea that combination therapy with cisplatin and a PI3K inhibitor would increase the therapeutic efficacy of cisplatin.

Medroxyprogesterone acetate induces cell proliferation through up-regulation of cyclin D1 expression via phosphatidylinositol 3-kinase/Akt/nuclear factor-kappaB cascade in human breast cancer cells.

The mechanism of medroxyprogesterone acetate (MPA)-induced cell proliferation in human breast cancer cells remains elusive. We examined the mechanism by which MPA affects the cyclin D1 expression in progesterone receptor (PR)-positive T47D human breast cancer cells. MPA (10 nM) treatment for 48 h induced proliferation of the cells (1.6-fold induction). MPA induced cyclin D1 expression (3.3-fold induction), and RU486, a selective PR antagonist, blocked the MPA-induced cell proliferation and cyclin D1 expression (23% inhibition). MPA increased both the protein level (2.2-fold induction) and promoter activity (2.7-fold induction) of cyclin D1 in MCF-7 cells transfected with PRB but not with PRA. Although MPA transcriptionally activated cyclin D1 expression, cyclin D1 promoter does not have progesterone-responsive element-related sequence. We further examined the mechanism for the regulation of the cyclin D1 expression. Because the cyclin D1 promoter contains three putative nuclear factor-kappaB (NFkappaB)-binding motifs and NFkappaB is a substrate of Akt, we investigated the effect of the phosphatidylinositol 3-kinase (PI3K)/Akt/NFkappaB cascade on the responses of cyclin D1 to MPA. MPA induced the transient phosphorylation of Akt (2.7-fold induction at 5 min), and treatment with PI3K inhibitor (wortmannin) attenuated the MPA-induced up-regulation of cyclin D1 expression (40% inhibition) and cell proliferation (40% inhibition). MPA also induced phosphorylation of inhibitor of NFkappaBalpha (IkappaBalpha) (2.3-fold induction), and treatment with wortmannin attenuated the MPA-induced IkappaBalpha phosphorylation (60% inhibition). Treatment with an IkappaBalpha phosphorylation inhibitor (BAY 11-7085) or a specific NFkappaB nuclear translocation inhibitor (SN-50) attenuated the MPA-induced up-regulation of both cyclin D1 expression (80 and 50% inhibition, respectively) and cell proliferation (55 and 34% inhibition, respectively). Because MPA induced a transient phosphorylation of Akt and the cyclin D1 promoter contains no progesterone-responsive element-related sequence, the MPA-induced cell proliferation through PRB by up-regulation of cyclin D1 expression via the PI3K/Akt/NFkappaB cascade may be a nongenomic mechanism.

Growth factors change nuclear distribution of estrogen receptor-alpha via mitogen-activated protein kinase or phosphatidylinositol 3-kinase cascade in a human breast cancer cell line.

In the present study, to examine the dynamic changes in the localization of nuclear estrogen receptor (ER)alpha induced by growth factors, we used time-lapse confocal microscopy to directly visualized ERalpha fused with green fluorescent protein (GFP-ERalpha) in single living cells treated with epidermal growth factor (EGF) or IGF-I. We observed that 17beta-estradiol (E2) changed the normally diffuse distribution of GFP-ERalpha throughout the nucleoplasm to a hyperspeckled distribution within 10 min. Both EGF and IGF-I also changed the nuclear distribution of GFP-ERalpha, similarly to E2 treatment. However, the time courses of the nuclear redistribution of GFP-ERalpha induced by EGF or IGF-I were different from that induced by E2 treatment. In the EGF-treated cells, the GFP-ERalpha nuclear redistribution was observed at 30 min and reached a maximum at 60 min, whereas in the IGF-I-treated cells, the GFP-ERalpha nuclear redistribution was observed at 60 min and reached a maximum at 90 min. The EGF-induced redistribution of GFP-ERalpha was blocked by pretreatment with a MAPK cascade inhibitor, PD98059, whereas the IGF-I-induced redistribution of GFP-ERalpha was blocked by pretreatment with a phosphatidylinositol 3-kinase inhibitor, LY294002. Analysis using an activation function-2 domain deletion mutant of GFP-ERalpha showed that the change in the distribution of GFP-ERalpha was not induced by E2, EGF, or IGF-I treatment. These data suggest that MAPK and phosphatidylinositol 3-kinase cascades are involved in the nuclear redistribution of ERalpha by EGF and IGF-I, respectively, and that the activation function-2 domain of ERalpha may be needed for the nuclear redistribution of ERalpha.

Both estrogen and raloxifene protect against beta-amyloid-induced neurotoxicity in estrogen receptor alpha-transfected PC12 cells by activation of telomerase activity via Akt cascade.

Although estrogen is known to protect against beta-amyloid (Abeta)-induced neurotoxicity, the mechanisms responsible for this effect are only beginning to be elucidated. In addition, the effect of raloxifene on Abeta-induced neuro-toxicity remains unknown. Here we investigated whether raloxifene exhibits similar neuro-protective effects to estrogen against Abeta-induced neurotoxicity and the mechanism of the effects of these agents in PC12 cells transfected with the full-length human estrogen receptor (ER) alpha gene (PCER). Raloxifene, like 17beta-estradiol (E2), significantly inhibited Abeta-induced apoptosis in PCER cells, but not in a control line of cells transfected with vector DNA alone (PCCON). Since telomerase activity, the level of which is modulated by regulation of telomerase catalytic subunit (TERT) at both the transcriptional and post-transcriptional levels, is known to be involved in suppressing apoptosis in neurons, we examined the effect of E2 and raloxifene on telomerase activity. Although both E2 and raloxifene induced telomerase activity in PCER cells, but not in PCCON cells, treated with Abeta, they had no effect on the level of TERT expression. These results suggest that neither E2 nor raloxifene affects the telomerase activity at the transcriptional level. We therefore studied the mechanism by which E2 and raloxifene induce the telomerase activity at the post-transcriptional level. Both E2 and raloxifene induced the phosphorylation of Akt, and pre-treatment with a phosphatidylinositol 3-kinase inhibitor, LY294002, attenuated both E2- and raloxifene-induced activation of the telomerase activity. Moreover, both E2 and raloxifene induced both the phosphorylation of TERT at a putative Akt phosphorylation site and the association of nuclear factor kappaB with TERT. Our findings suggest that and raloxifene exert neuroprotective effects by E2 telomerase activation via a post-transcriptional cascade in an experimental model relevant to Alzheimer's disease.

Rapamycin-induced insulin resistance is mediated by mTORC2 loss and uncoupled from longevity.

Rapamycin, an inhibitor of mechanistic target of rapamycin complex 1 (mTORC1), extends the life spans of yeast, flies, and mice. Calorie restriction, which increases life span and insulin sensitivity, is proposed to function by inhibition of mTORC1, yet paradoxically, chronic administration of rapamycin substantially impairs glucose tolerance and insulin action. We demonstrate that rapamycin disrupted a second mTOR complex, mTORC2, in vivo and that mTORC2 was required for the insulin-mediated suppression of hepatic gluconeogenesis. Further, decreased mTORC1 signaling was sufficient to extend life span independently from changes in glucose homeostasis, as female mice heterozygous for both mTOR and mLST8 exhibited decreased mTORC1 activity and extended life span but had normal glucose tolerance and insulin sensitivity. Thus, mTORC2 disruption is an important mediator of the effects of rapamycin in vivo.

mTOR complex 2 is required for the development of prostate cancer induced by Pten loss in mice.

mTOR complex 2 (mTORC2) contains the mammalian target of rapamycin (mTOR) kinase and the Rictor regulatory protein and phosphorylates Akt. Whether this function of mTORC2 is critical for cancer progression is unknown. Here, we show that transformed human prostate epithelial cells lacking PTEN require mTORC2 to form tumors when injected into nude mice. Furthermore, we find that Rictor is a haploinsufficient gene and that deleting one copy protects Pten heterozygous mice from prostate cancer. Finally, we show that the development of prostate cancer caused by Pten deletion specifically in prostate epithelium requires mTORC2, but that for normal prostate epithelial cells, mTORC2 activity is nonessential. The selective requirement for mTORC2 in tumor development suggests that mTORC2 inhibitors may be of substantial clinical utility.

Case of a giant mucinous ovarian carcinoma with bone metastasis.

We report the case of a 51-year-old Japanese woman with a giant (50.75-kg) ovarian tumor. The histopathologic diagnosis was mucinous cystadenocarcinoma. After surgery, the patient was intubated and connected to a respirator for 8 days. Thereafter, she was diagnosed with bone metastasis to the hip bone and the femur.

Primary mucinous adenocarcinoma of the vagina: possibility of differentiating from metastatic adenocarcinomas.

Primary vaginal adenocarcinomas are rare neoplasms. Herein is reported a case of primary vaginal mucinous adenocarcinoma with an interesting mucin profile, presumably arising from a lesion of adenosis in a patient without in utero exposure to diethylstilbesterol (DES). The patient, a 44-year-old woman, had undergone vaginal total hysterectomy 10 years previously for myoma uteri corporis. The histological features of the vaginal intramural tumor found in this patient resembled those of mucinous adenocarcinoma of the endocervical type. Therefore, it was necessary to determine whether or not the tumor was metastatic from an occult cervical adenocarcinoma. However, the adenocarcinoma cells of the present case did not contain sulfomucin at all, being different from most mucinous adenocarcinoma cells of the endocervical type. Moreover, there were foci of adenosis adjacent to the adenocarcinoma foci, which also did not contain sulfomucin. These findings indicate that the mucinous adenocarcinoma arose from vaginal adenosis. Further studies are necessary to investigate whether lack of sulfomucin expression is a characteristic feature of vaginal adenosis.

Characteristic features of ovarian borderline tumors with invasive implant.

OBJECTIVE: Since ovarian borderline tumor with invasive implant behaves as carcinoma, it should be managed like carcinoma. Since its characteristic features have not been reported, its preoperative diagnosis was thought to be impossible. We evaluated the features of MRI and macroscopic appearance in two cases of ovarian borderline tumor with invasive implant. METHODS: Borderline tumor with invasive implant was evaluated in two patients by MRI and macroscopic examination. RESULTS: In these patients, MRI revealed profuse papillary projections. Although the lesion showed high signal intensity on contrast-enhanced T1-weighted images compared with that on T1-weighted ones, most of the signal intensity on T2-weighted images was high, suggesting that the lesion is an assembly of vesicles and an obvious solid part is absent. The macroscopic appearance of the tumor showed profuse papillary projections consisting of many vesicles perforating and extending far beyond the ovarian capsule, without a solid part. The histological findings indicated serous borderline tumors with invasive implant. CONCLUSION: In these two cases, we found the characteristic features of serous borderline tumor with invasive implant by MRI and macroscopic examination. Our findings may be of clinical value since the preoperative information about the possibility of invasive implant may be quite important for the management of borderline tumor with invasive implant, especially for young patients wishing to bear children.

Intravenous leiomyomatosis with cardiac extension.

Intravenous leiomyomatosis with cardiac extension is an extremely rare uterine tumor. We report here a case of a patient with a uterine leiomyoma which extended into the right atrium through the left ovarian vein, progressing into the left renal vein along the inferior vena cava. Complete one-stage removal of the tumor was performed using cardiopulmonary bypass, and the patient has shown a favorable outcome. Successful therapy for intravenous leiomyomatosis is dependent on total surgical excision of the tumor, cessation of ovarian function and avoidance of postoperative estrogen replacement therapy.

Local and systemic wound-induction of RNase and nuclease activities in Arabidopsis: RNS1 as a marker for a JA-independent systemic signaling pathway.

Induction of defense-related genes is one way in which plants respond to mechanical injury. We investigated whether RNases are involved in the wound response in Arabidopsis thaliana. As in other plant systems, several activities are induced with various timings in damaged leaves, stems and seedlings in Arabidopsis, including at least three bifunctional nucleases, capable of degrading both RNA and DNA, as well as RNS1, a member of the ubiquitous RNase T(2) family of RNases. The strong induction of RNS1 is particularly interesting because it occurs both locally and systemically following wounding. The systemic induction of this RNase indicates that members of this family may be involved in defense mechanisms in addition to their previously hypothesized functions in nutrient recycling and remobilization. Additionally, the systemic induction appears to be controlled independently of jasmonic acid, and the local induction of RNS1 and the nuclease activities are independent of both JA and oligosaccharide elicitors. Consequently, a novel systemic pathway, likely involving a third signal, appears to exist in Arabidopsis.

Raloxifene inhibits estrogen-induced up-regulation of telomerase activity in a human breast cancer cell line.

The mechanism by which raloxifene acts in the chemoprevention of breast cancer remains unclear. Because telomerase activity is involved in estrogen-induced carcinogenesis, we examined the effect of raloxifene on estrogen-induced up-regulation of telomerase activity in MCF-7 human breast cancer cell line. Raloxifene inhibited the induction of cell growth and telomerase activity by 17beta-estradiol (E2). Raloxifene inhibited the E2-induced expression of the human telomerase catalytic subunit (hTERT), and transient expression assays using luciferase reporter plasmids containing various fragments of the hTERT promoter showed that the estrogen-responsive element appeared to be partially responsible for the action of raloxifene. E2 induced the phosphorylation of Akt, and pretreatment with a phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, attenuated the E2-induced increases of the telomerase activity and hTERT promoter activity. Raloxifene inhibited the E2-induced Akt phosphorylation. In addition, raloxifene also inhibited the E2-induced hTERT expression via the PI3K/Akt/NFkappaB cascade. Moreover, raloxifene also inhibited the E2-induced phosphorylation of hTERT, association of NFkappaB with hTERT, and nuclear accumulation of hTERT. These results show that raloxifene inhibited the E2-induced up-regulation of telomerase activity not only by transcriptional regulation of hTERT via an estrogen-responsive element-dependent mechanism and the PI3K/Akt/NFkappaB cascade but also by post-translational regulation via phosphorylation of hTERT and association with NFkappaB.