Purpose: There is growing evidence that stress and other behavioral factors may affect cancer progression and patient survival.The underlying mechanisms for this association are poorly understood.The purpose of this study is to determine the effects of stress-associated hormones norepinephrine, epinephrine, and cortisol on the invasive potential of ovarian cancer cells. Experimental Design:The ovarian cancer cells EG, SKOV3, and 222 were exposed to increasing levels of either norepinephrine, epinephrine, or cortisol, and the in vitro invasive potential was determined using the membrane invasion culture system. Additionally, the effects of these stress hormones on matrix metalloproteinase-2 (MMP-2) and MMP-9 were determined by ELISA. The effects of the h-adrenergic agonist isoproterenol on in vivo tumor growth were determined using nude mice. Results: Stress levels of norepinephrine increased the in vitro invasiveness of ovarian cancer cells by 89% to 198%. Epinephrine also induced significant increases in invasion in all three cell lines ranging from 64% to 76%. Cortisol did not significantly affect invasiveness of the EG and 222 cell lines but increased invasion in the SKOV3 cell line (P = 0.01). We have previously shown that ovarian cancer cells express h-adrenergic receptors. The h-adrenergic antagonist propanolol (1 Amol/L) completely blocked the norepinephrine-induced increase in invasiveness. Norepinephrine also increased tumor cell expression of MMP-2 (P = 0.02 for both SKOV3 and EG cells) and MMP-9 (P = 0.01and 0.04, respectively), and pharmacologic blockade of MMPs abrogated the effects of norepinephrine on tumor cell invasive potential. Isoproterenol treatment resulted in a significant increase in tumor volume and infiltration in the SKOV3ip1 in vivo model, which was blocked by propranolol. Conclusions: These findings provide direct experimental evidence that stress hormones can enhance the invasive potential of ovarian cancer cells. These effects are most likely mediated by stimulation of MMPs.There is extensive evidence supporting stress-immune relationships in healthy adults (1) and a growing body of literature demonstrating these relationships in cancer patients (2 -4). Meta-analyses and reviews have reported alterations in cellular immunity (decreased T-cell response to mitogen stimulation, decreased natural killer cell cytotoxicity, and altered production of cytokines) in association with chronic stress and/or depressed affect (5, 6). Among cancer patients, behavioral factors may serve as predictors of clinical outcome, such as response to therapy and overall survival (7 -11). These findings suggest that psychosocial stress factors not only affect the immune system adversely but also contribute to poor outcome in cancer patients. However, no study has shown that stressinduced changes in cancer outcomes are mediated by changes in immune system function. Here, we consider the alternative hypothesis that stress hormones directly affect tumor cells to alter their malignant potential.Immune s...

At the midblastula transition, the Xenopus laevis embryonic cell cycle is remodeled from rapid alternations between S and M phases to become the complex adult cell cycle. Cell cycle remodeling occurs after zygotic transcription initiates and is accompanied by terminal downregulation of maternal cyclins A1 and B2. We report here that the disappearance of both cyclin A1 and B2 proteins is preceded by the rapid deadenylation of their mRNAs. A specific mechanism triggers this deadenylation. This mechanism depends upon discrete regions of the 3 untranslated regions and requires zygotic transcription. Together, these results strongly suggest that zygote-dependent deadenylation of cyclin A1 and cyclin B2 mRNAs is responsible for the downregulation of these proteins. These studies also raise the possibility that zygotic control of maternal cyclins plays a role in establishing the adult cell cycle.In Xenopus laevis, the first 12 cell cycles following fertilization occur in the absence of transcription. Maternal mRNAs and proteins that are synthesized and stored in the growing oocyte control these early embryonic cell divisions. Zygotic transcription begins upon completion of the 12th cell cycle and, in Xenopus, is referred to as the midblastula transition (MBT) (25). Prior to the MBT, gene expression is controlled by posttranscriptional mechanisms including regulation of the adenylation state of maternal mRNAs. Polyadenylated mRNAs are recruited into polysomes and translated, while deadenylated mRNAs are released from polysomes and translationally silenced (27).In Xenopus, adenylation control regulates both progression through meiosis (oocyte maturation) and the transition from the meiotic to the mitotic cell cycle by regulating levels of cell cycle proteins, including cyclins. The mRNAs encoding cyclins A1, B1, B2, and E1 are synthesized during oogenesis and stored untranslated as poly(A) Ϫ mRNAs until oocyte maturation (5, 35). In response to progesterone, cytoplasmic polyadenylation elements (CPEs) in the 3Ј untranslated region (UTR) trigger polyadenylation and translational activation of these mRNAs, allowing accumulation of cyclin protein and progression through meiosis. After fertilization, cyclin mRNAs are further adenylated and continuously translated (35). However, periodic degradation of cyclin A1, B1, and B2 proteins during each cell cycle allows progression through the first 12 cell divisions (18).The 12th cell division is completed approximately 6 h postfertilization (p.f.) and is followed by remodeling of the cell cycle between cell cycles 13 and 15 (24). The cell cycle is remodeled from a rapid alternation between DNA synthesis and mitosis to become a cell cycle containing gap phases and checkpoint controls. The mechanism of cell cycle remodeling is not well understood, but it is accompanied by temporally specific degradation of maternal cyclin A1, B2, and E1 proteins (14, 33). We define this event as the terminal disappearance of the maternal cyclin protein.Because cyclins A1 and B2 are normally degraded ...

Metformin, an antidiabetic agent, potentiates insulin action and reduces insulin resistance. We examined the antihypertensive effects and vascular effects of metformin in spontaneously hypertensive rats (SHR). Wistar-Kyoto normotensive (WKY) and SHR were injected with metformin (100 mg/kg) or saline subcutaneously twice daily for 4 weeks. Blood pressure was recorded by a tail-cuff plethesmographic method. Metformin treatment significantly attenuated (P < .05) the increase in blood pressure in metformin treated SHR versus untreated control SHR. At the end of the experimental period of 4 weeks, metformin-treated SHR had a mean blood pressure that was 34 mm lower than that of untreated SHR. Metformin treatment had no significant effect on blood pressure in WKY rats. Treatment of SHR aortic smooth muscle (SM) cells with metformin (2 micrograms/mL) for 24 h significantly decreased (P < .05) arginine vasopressin- and thrombin- stimulated increase in [Ca2+]i. However, metformin treatment did not have a significant effect on the basal [Ca+]i. Incubation of SHR aortic SM cells with OH-L-arginine (25 to 100 mumol/L) for 24 h increased nitrite production in a dose dependent manner. Metformin (5 micrograms/mL) treatment of SM cells increased nitrite production at all concentrations of OH-L-arginine; however, differences were significant (P < .05) only at 25 and 50 mumol/L OH-L-arginine. These results suggest that metformin may be decreasing arterial pressure in the SHR, at least in part, by attenuating the agonist-stimulated [Ca2+]i response in SHR vascular smooth muscle cells.

Epidemiological evidence and estrogen replacement studies suggest that estrogen has a protective effect on the cardiovascular system against coronary artery disease. Vascular smooth muscle (VSM) cell replication has been shown to play a causative role in the pathogenesis of atherosclerosis. Therefore, in this study, we investigated the effect of chronic treatment of cultured guinea pig coronary artery VSM cells with physiological concentrations of 17beta-estradiol (E2) on thymidine incorporation, cell proliferation, and bradykinin-stimulated cytosolic calcium concentration ([Ca2+]i). Bradykinin at physiological concentrations causes contraction of endothelium-denuded guinea pig coronary artery rings in a concentration-dependent manner. VSM cells were first treated with low doses of E2 (10 pg/ml) for 1-2 days followed by treatment for 4-6 days with 50 pg/ml of E2, a concentration similar to that found in pregnancy. Using these protocols, we consistently observed the presence of E2-receptor mRNA in VSM cells by a ribonuclease protection assay. Fetal calf serum-stimulated [3H]thymidine incorporation was significantly reduced (P < 0.05) in E2-treated cells compared with untreated control cells. Similarly, E2 treatment significantly inhibited fetal calf serum-stimulated VSM cell proliferation compared with untreated control cells (P < 0.05). We also tested the hypothesis that E2 treatment attenuates agonist-stimulated [Ca2+]i in VSM cells because acute E2 treatment has been shown to produce relaxation of precontracted isolated coronary artery preparations. E2 treatment of VSM cells resulted in a significant decrease in bradykinin-stimulated [Ca2+]i compared with untreated cells (P < 0.05). In conclusion, our data demonstrate that estrogen at physiological concentrations directly regulates coronary VSM cell function.

The subfornical organ (SFO) is sensitive to both ANG II and ACh, and local application of these agents produces dipsogenic responses and vasopressin release. The present study examined the effects of cholinergic drugs, ANG II, and increased extracellular osmolarity on dissociated, cultured cells of the SFO that were retrogradely labeled from the supraoptic nucleus. The effects were measured as changes in cytosolic calcium in fura 2-loaded cells by using a calcium imaging system. Both ACh and carbachol increased intracellular ionic calcium concentration ([Ca2+]i). However, in contrast to the effects of muscarinic receptor agonists on SFO neurons, manipulation of the extracellular osmolality produced no effects, and application of ANG II produced only moderate effects on [Ca2+]i in a few retrogradely labeled cells. The cholinergic effects on [Ca2+]i could be blocked with the muscarinic receptor antagonist atropine and with the more selective muscarinic receptor antagonists pirenzepine and 4-diphenylacetoxy-N-methylpiperdine methiodide (4-DAMP). In addition, the calcium in the extracellular fluid was required for the cholinergic-induced increase in [Ca2+]i. These findings indicate that ACh acts to induce a functional cellular response in SFO neurons through action on a muscarinic receptor, probably of the M1 subtype and that the increase of [Ca2+]i, at least initially, requires the entry of extracellular Ca2+. Also, consistent with a functional role of M1 receptors in the SFO are the results of immunohistochemical preparations demonstrating M1 muscarinic receptor-like protein present within this forebrain circumventricular organ.