The Effects of Estradiol, Androgens and the Selective Estrogen Modulators: Ospemifene, Raloxifene and Tamoxifen on Explant Cultures of Human Breast Tissue

The impact of menopausal hormone therapy (MHT) on increasing the risk for breast cancer (BC) remains controversial. To understand MHT-elicited cellular breast effects and the potential risks, included with using this therapy, a further investigation into this controversy is the subject of this thesis. In this thesis, to study the effects of estrogen, progestin, androgens and selective estrogen receptor modulators (SERMs), a modified tissue explant culture system was used. The different types of human breast tissues (HBTs) used in this study were normal HBTs, obtained from reduction mammoplasties of premenopausal women (prem-HBTs) or postmenopausal (postm-HBTs) women and peritumoral HBTs (peritum-HBTs) which were obtained from surgeries on postmenopausal BC patients. The explants were cultured up to three weeks in the presence or absence of estradiol (E2), medroxyprogesterone acetate (MPA), testosterone (T), dihydrotestosterone (DHT) and SERMs - ospemifene (OSP), raloxifene (RAL) and tamoxifen (TAM). The cultured HBTs maintained morphological integrity and responded to hormonal treatment in vitro. E2, MPA or E2/MPA increased proliferative activity and was associated with increased cyclin-D1 and caused changes in the cell cycle inhibitors p21 and p27, whereas the androgens T and DHT inhibited proliferation and increased apoptosis in HBT epithelia and opposed E2-stimulated proliferation and cell survival. The postm-HBTs were more sensitive to E2 than prem-HBTs. The effects of OSP, RAL and TAM on HBT epithelium were antiproliferative. E2, androgens and SERMs were associated with marked changes in the proportions of epithelial cells expressing steroid hormone receptors: E2 increased ERα expressing cells and decreased androgen receptor (AR) positive cells, whereas T and DHT had opposite effects. The OSP, RAL and TAM, also decreased a proportion of ERα positive cells in HBT epithelium. At 100 nM, these compounds maintained the relative number of AR positive cells, present at control level, which may partly explain proliferative inhibition. In conclusion, the proliferative activity of E2, in the epithelium of postm-HBTs, is opposed by T and DHT, which suggests that the inclusion of androgens in MHT may decrease the risk for developing BC.

Knowledge, processes and relationships in organizational change: Understanding how to facilitate management of intellectual capital in diverse organization structures and cultures.

This study examined relationships of organizational dependencies, change management and developed intellectual knowledge resources, in different intellectual capital based development programs on ICT-sector. Study was carried out in a research context, where high degree of external organizational contingencies existed and lots of changes in several development programs had taken place in the last years. From a scientific perspective the main contribution was that evidence between relationships of organizational dependencies, change model portfolio and developed knowledge resources could be suggested. From managerial perspective the primary implication was that in situations where sustainable competitive advantage is pursued by means of increasing knowledge based productivity of labor, firms should seek to pursue organizational settings where external dependencies have minimal amount of effect.

Stock Price Reactions on M&A, Dividends and Game Releases. Evidence from Gaming Industry.

A rapidly growing gaming industry, which specializes on PC, console, online and other games, attracts attention of investors and analysts, who try to understand what drives changes of the gaming industry companies’ stock prices. This master thesis shows the evidence that, besides long-established types of events (M&A and dividend payments), the companies’ stock price changes depend on industry-specific events. I analyzed specific for gaming industry events - game releases with respect to its subdivisions: new games-sequels, games ratings and subdivision according to a developer of a game (self-developed by publisher or outsourced). The master thesis analyzes stock prices of 55 companies from gaming industry from all over the world. The research period covers 5 year, spreading from April 2008 to April 2013. Executed with an event study method, results of the research show that all the analyzed events types have significant influence on the stock prices of the gaming industry companies. The current master thesis suggests that acquisitions in the industry affect positively bidders’ and targets’ stock prices. Mergers events cause positive stock price reactions as well. But dividends payments and game releases events influence negatively on the stock prices. Game releases’ effect is up to -2.2% of cumulative average abnormal return (CAAR) drop during the first ten days after the game releases. Having researched different kinds of events and identified the direction of their impact, the current paper can be of high value for investors, seeking profits in the gaming industry, and other interested parties.

Viral marketing in a Finnish medium sized gaming company: discovering, analyzing and building an improved management process for viral marketing campaigns, case: Nitrogames

The goal of this thesis is to build a viral marketing management framework for a Finnish medium sized gaming company. This is achieved by first finding and building a theoretical five step management process framework based on literature, analyzing current model and giving recommendations for the case company to develop its own management process. In addition, viral marketing research is still in early stage resulting this study to propose its own take on the definition in the theory part. Empirical part is based on qualitative interviews, campaign material and secondary sources and is aimed to find out and analyze the case company’s current viral marketing state and to give recommendations to it. The final outcome of the study is a general, theoretical management framework for viral marketing campaigns and specified recommendations for the case company.

Value co-creation in gaming the industry: A qualitative research on virtual discussion forums

This study discusses the procedures of value co-creation that persist in gaming industry. The purpose of this study was to identify the procedures that persist in current video gaming industry which answers the main research problem how value is co-created in video gaming industry followed by three sub questions: (i) What is value co-creation in gaming industry? (ii) Who participates in value co-creation in gaming industry? (iii) What are the procedures that are involved in value co-creation in gaming industry? The theoretical background of the study consists of literature relating to the theory of marketing i.e., notion of value, conventional understanding of value creation, value chain, co-creation approach, co-production approach. The research adopted qualitative research approach. As a platform of relationship researcher used web 2.0 tool interface. Data were collected from the social networks and netnography method was applied for analyzing them. Findings show that customer and company both co-create optimum level of value while they interact with each other and within the customers as well. However mostly the C2C interaction, discussions and dialogues threads that emerged around the main discussion facilitated to co-create value. In this manner, companies require exploiting and further motivating, developing and supporting the interactions between customers participating in value creation. Hierarchy of value co-creation processes is the result derived from the identified challenges of value co-creation approach and discussion forums data analysis. Overall three general sets and seven topics were found that explored the phenomenon of customer to customer (C2C) and business to customer (B2C) interaction/debating for value co-creation through user generated contents. These topics describe how gamer contributes and interacts in co-creating value along with companies. A methodical quest in current research literature acknowledged numerous evolving flows of value in this study. These are general management perspective, new product development and innovation, virtual customer environment, service science and service dominant logic. Overall the topics deliver various realistic and conceptual implications for using and handling gamers in social networks for augmenting customers’ value co-creation process.

Comparative uptake and metabolism of 2-[14C]-2,4-dichlorophenoxyacetic acid in callus cultures of monocot (Dioscorea spp.) and dicot (Nicotiana tabacum L.) plants

(Comparative uptake and metabolism of 2-[14C]-2,4-dichlorophenoxyacetic acid in callus cultures of monocot (Dioscorea spp.) and dicot (Nicotiana tabacum L.) plants). The uptake and metabolism of 2-[14C]-2,4-dichlorophenoxyacetic acid (2,4-D) were investigated in leaf calluses of Nicotiana tabacum, tuber calluses of Dioscorea opposita and calluses derived from zygotic embryos, leaves and petioles of Dioscorea composita. Striking similarities were evident in the patterns of 2,4-D metabolites and their chemical characteristics in the three callus types of D. composita compared, but significant differences were detected among the patterns of rnetabolites in the three species studied. Preliminary investigations on the stability of various metabolites (separated using TLC) by hydrolysis showed that sugar esters appeared to be the major metabolites in tobacco whilst in yams (D. opposita) glycosides were shown to be the main ones, which indicated a similarity between plants of Gramineae and Dioscoreaceae in terms of 2,4-D metabolism. Release of 2,4-D from tobacco callus cells upon their transfer to 2,4-D-free medium was detected and the implications of this are discussed in relation to the cultural conditions necessary to induce morphogenesis in vitro.

Detection of anthraquinones and identification of 1,4-naphthohydroquinone in cell suspension cultures of Rudgea jasminoides (Cham.) Müll. Arg. (Rubiaceae)

In Rubiaceae, anthraquinones and naphthoquinones are secondary metabolites characteristic of the subfamily Rubioideae, in which Rudgea jasminoides is included. Thin-layer chromatography using specific solvent systems and spray reagents indicated the presence of anthraquinones constitutively produced by cell suspension cultures of R. jasminoides. GC/MS analysis detected 1,4-naphthohydroquinone as a product of biosynthesis only after elicitation of the cells with yeast extract (Saccharomyces cerevisiae). The latter compound is probably a phytoalexin produced by suspension cultures of R. jasminoides.

Effect of FSH and insulin on lipogenesis in cultures of Sertoli cells from immature rats

Follicle-stimulating hormone (FSH) and insulin regulate glycide metabolism in Sertoli cells, thus stimulating lactate production. These stimulatory effects of FSH and insulin do not require protein synthesis, suggesting a modulation of enzyme activity and/or regulation of glucose transport. The present investigation was performed to characterize the hormonal control of lipid metabolism in Sertoli cells. The data indicate that FSH and insulin have a regulatory effect on lipid metabolism in Sertoli cells. After 8 h of preincubation with insulin (5 µg/ml), the activity of the enzyme ATP-citrate lyase in cultured Sertoli cells was increased from 0.19 to 0.34 nmol NAD+ formed µg protein-1 min-1. FSH (100 ng/ml) had no effect on this enzyme. Glycerol phosphate dehydrogenase activity was not affected by any of the hormones tested. When Sertoli cells from 19-day old rats were incubated with [1,2­14C]acetate for 90 or 360 min, the [14C] label was present predominantly in triglyceride and phospholipid fractions with minor amounts in other lipids. In Sertoli cells pretreated for 16 h with insulin and FSH, an increase in acetate incorporation into lipids was observed. Most of the label was in esterified lipids and this percentage increased with the time of treatment; this increase was remarkable in triglycerides of control cells (18.8% to 30.6%). Since Sertoli cell triglycerides participate in the control of spermatogenesis, the present data suggest that the hormonal control of lipid metabolism in Sertoli cells is important not only for maintaining the energy of the cell itself, but also for the control of the spermatogenesis process.

Hydrolysis of xylans by enzyme systems from solid cultures of Trichoderma harzianum strains

Xylanase activity was isolated from crude extracts of Trichoderma harzianum strains C and 4 grown at 28oC in a solid medium containing wheat bran as the carbon source. Enzyme activity was demonstrable in the permeate after ultrafiltration of the crude extracts using an Amicon system. The hydrolysis patterns of different xylans and paper pulps by xylanase activity ranged from xylose, xylobiose and xylotriose to higher xylooligosaccharides. A purified ß-xylosidase from the Trichoderma harzianum strain released xylose, xylobiose and xylotriose from seaweed, deacetylated, oat spelt and birchwood xylans. The purified enzyme was not active against acetylated xylan and catalyzed the hydrolysis of xylooligosaccharides, including xylotriose, xylotetraose and xylopentaose. However, the enzyme was not able to degrade xylohexaose. Xylanase pretreatment was effective for hardwood kraft pulp bleaching. Hardwood kraft pulp bleached in the XEOP sequence had its kappa number reduced from 13.2 to 8.9 and a viscosity of 20.45 cp. The efficiency of delignification was 33%.

Cyclic AMP increases the survival of ganglion cells in mixed retinal cell cultures in the absence of exogenous neurotrophic molecules, an effect that involves cholinergic activity

Natural cell death is a well-known degenerative phenomenon occurring during development of the nervous system. The role of trophic molecules produced by target and afferent cells as well as by glial cells has been extensively demonstrated. Literature data demonstrate that cAMP can modulate the survival of neuronal cells. Cultures of mixed retinal cells were treated with forskolin (an activator of the enzyme adenylyl cyclase) for 48 h. The results show that 50 µM forskolin induced a two-fold increase in the survival of retinal ganglion cells (RGCs) in the absence of exogenous trophic factors. This effect was dose dependent and abolished by 1 µM H89 (an inhibitor of protein kinase A), 1.25 µM chelerythrine chloride (an inhibitor of protein kinase C), 50 µM PD 98059 (an inhibitor of MEK), 25 µM Ly 294002 (an inhibitor of phosphatidylinositol-3 kinase), 30 nM brefeldin A (an inhibitor of polypeptide release), and 10 µM genistein or 1 ng/ml herbimycin (inhibitors of tyrosine kinase enzymes). The inhibition of muscarinic receptors by 10 µM atropine or 1 µM telenzepine also blocked the effect of forskolin. When we used 25 µM BAPTA, an intracellular calcium chelator, as well as 20 µM 5-fluoro-2'-deoxyuridine, an inhibitor of cell proliferation, we also abolished the effect. Our results indicate that cAMP plays an important role controlling the survival of RGCs. This effect is directly dependent on M1 receptor activation indicating that cholinergic activity mediates the increase in RGC survival. We propose a model which involves cholinergic amacrine cells and glial cells in the increase of RGC survival elicited by forskolin treatment.

Increased production of tumor necrosis factor-alpha in whole blood cultures from children with primary malnutrition

Because low tumor necrosis factor-alpha (TNF-alpha) production has been reported in malnourished children, in contrast with high production of TNF-alpha in experimental protein-energy malnutrition, we reevaluated the production of TNF-alpha in whole blood cultures from children with primary malnutrition free from infection, and in healthy sex- and age-matched controls. Mononuclear cells in blood diluted 1:5 in endotoxin-free medium released TNF-alpha for 24 h. Spontaneously released TNF-alpha levels (mean ± SEM), as measured by enzyme immunoassay in the supernatants of unstimulated 24-h cultures, were 10,941 ± 2,591 pg/ml in children with malnutrition (N = 11) and 533 ± 267 pg/ml in controls (N = 18) (P < 0.0001). TNF-alpha production was increased by stimulation with lipopolysaccharide (LPS), with maximal production of 67,341 ± 16,580 pg/ml TNF-alpha in malnourished children and 25,198 ± 2,493 pg/ml in controls (P = 0.002). In control subjects, LPS dose-dependently induced TNF-alpha production, with maximal responses obtained at 2000 ng/ml. In contrast, malnourished patients produced significantly more TNF-alpha with 0.02-200 ng/ml LPS, responded maximally at a 10-fold lower LPS concentration (200 ng/ml), and presented high-dose inhibition at 2000 ng/ml. TNF-alpha production a) was significantly influenced by LPS concentration in control subjects, but not in malnourished children, who responded strongly to very low LPS concentrations, and b) presented a significant, negative correlation (r = -0.703, P = 0.023) between spontaneous release and the LPS concentration that elicited maximal responses in malnourished patients. These findings indicate that malnourished children are not deficient in TNF-alpha production, and suggest that their cells are primed for increased TNF-alpha production.

Detection of multiple mycoplasma infection in cell cultures by PCR

A total of 301 cell cultures from 15 laboratories were monitored for mycoplasma (Mollicutes) using PCR and culture methodology. The infection was detected in the cell culture collection of 12 laboratories. PCR for Mollicutes detected these bacteria in 93 (30.9%) samples. Although the infection was confirmed by culture for 69 (22.9%) samples, PCR with generic primers did not detect the infection in five (5.4%). Mycoplasma species were identified with specific primers in 91 (30.2%) of the 98 samples (32.6%) considered to be infected. Mycoplasma hyorhinis was detected in 63.3% of the infected samples, M. arginini in 59.2%, Acholeplasma laidlawii in 20.4%, M. fermentans in 14.3%, M. orale in 11.2%, and M. salivarium in 8.2%. Sixty (61.2%) samples were co-infected with more than one mycoplasma species. M. hyorhinis and M. arginini were the microorganisms most frequently found in combination, having been detected in 30 (30.6%) samples and other associations including up to four species were detected in 30 other samples. Failure of the treatments used to eliminate mycoplasmas from cell cultures might be explained by the occurrence of these multiple infections. The present results indicate that the sharing of non-certified cells among laboratories may disseminate mycoplasma in cell cultures.

Low-intensity red and infrared laser effects at high fluences on Escherichia coli cultures

Semiconductor laser devices are readily available and practical radiation sources providing wavelength tenability and high monochromaticity. Low-intensity red and near-infrared lasers are considered safe for use in clinical applications. However, adverse effects can occur via free radical generation, and the biological effects of these lasers from unusually high fluences or high doses have not yet been evaluated. Here, we evaluated the survival, filamentation induction and morphology of Escherichia coli cells deficient in repair of oxidative DNA lesions when exposed to low-intensity red and infrared lasers at unusually high fluences. Cultures of wild-type (AB1157), endonuclease III-deficient (JW1625-1), and endonuclease IV-deficient (JW2146-1) E. coli, in exponential and stationary growth phases, were exposed to red and infrared lasers (0, 250, 500, and 1000 J/cm2) to evaluate their survival rates, filamentation phenotype induction and cell morphologies. The results showed that low-intensity red and infrared lasers at high fluences are lethal, induce a filamentation phenotype, and alter the morphology of the E. coli cells. Low-intensity red and infrared lasers have potential to induce adverse effects on cells, whether used at unusually high fluences, or at high doses. Hence, there is a need to reinforce the importance of accurate dosimetry in therapeutic protocols.

Lactobacillus plantarum strains isolated from naturally fermented sausages and their technological properties for application as starter cultures

In the present study, technological properties of L. plantarum strains isolated from naturally fermented sausages manufactured in the South region of Brazil were investigated in order to obtain a starter culture. The technological properties evaluated were the following: ability to growth at different pH values, at different temperatures, in different salt concentrations and in the presence of commercial curing salt, fast production of acid, determination of D - and L - lactic acid; nitrate reductase activity; antagonistic activity and stability of the isolated cultures after fermentation, concentration, and freeze-drying process. The isolated strains showed effectiveness to improve technological properties as starter cultures.