A study of learners' perceived value and satisfaction for implied effectiveness of online learning systems

Online learning systems (OLS) have become center stage for corporations and educational institutions as a competitive tool in the knowledge economy. The satisfaction construct has received extensive coverage in information systems literature as an indicator of effectiveness but has been criticized for lack of validity; yet, the value construct has been largely ignored, although it has a long history in psychology, sociology, and behavioral science. The purpose of this dissertation is to investigate the value and satisfaction constructs in the context of OLS, and their perceived by learners relationship for implied effectiveness of OLS. ^ First, a qualitative phase is employed to gather OLS values from learners' focus groups, followed by a pilot phase to refine a proposed instrument, and a main phase to validate the survey. Responses were received from 75 students in four focus groups, 141 in the pilot, and 207 the main survey. Extensive data cleaning and exploratory factor analysis were done to identify factors of learners' perceived value and satisfaction of OLS. Then, Value-Satisfaction grids and the Learners' Value Index of Satisfaction (LeVIS) were developed as benchmarking tools of OLS. Moreover, Multicriteria Decision Analysis (MCDA) techniques were employed to impute value from satisfaction scores in order to reduce survey response time. ^ The results provided four satisfaction and four value factors with high reliability (Cronbach's α). Moreover, value and satisfaction were found to have low linear and nonlinear correlations, indicating that they are two distinct uncorrelated constructs. This is consistent with the literature. Value-Satisfaction grids and the LeVIS index indicated relatively high effectiveness for technology and support characteristics, relatively low effectiveness for professor's characteristics, while course and learner characteristics indicated average effectiveness. ^ The main contributions of this study include identifying, defining, and articulating the relationship between value and satisfaction constructs as assessment of users' implied IS effectiveness, as well as assessing the accuracy of MCDA procedures to predict value scores, thus reducing by half the survey questionnaire size. ^

Three essays on a longitudinal analysis of business start-ups using the Kauffman Firm Survey

This dissertation focused on the longitudinal analysis of business start-ups using three waves of data from the Kauffman Firm Survey. ^ The first essay used the data from years 2004-2008, and examined the simultaneous relationship between a firm's capital structure, human resource policies, and its impact on the level of innovation. The firm leverage was calculated as, debt divided by total financial resources. Index of employee well-being was determined by a set of nine dichotomous questions asked in the survey. A negative binomial fixed effects model was used to analyze the effect of employee well-being and leverage on the count data of patents and copyrights, which were used as a proxy for innovation. The paper demonstrated that employee well-being positively affects the firm's innovation, while a higher leverage ratio had a negative impact on the innovation. No significant relation was found between leverage and employee well-being.^ The second essay used the data from years 2004-2009, and inquired whether a higher entrepreneurial speed of learning is desirable, and whether there is a linkage between the speed of learning and growth rate of the firm. The change in the speed of learning was measured using a pooled OLS estimator in repeated cross-sections. There was evidence of a declining speed of learning over time, and it was concluded that a higher speed of learning is not necessarily a good thing, because speed of learning is contingent on the entrepreneur's initial knowledge, and the precision of the signals he receives from the market. Also, there was no reason to expect speed of learning to be related to the growth of the firm in one direction over another.^ The third essay used the data from years 2004-2010, and determined the timing of diversification activities by the business start-ups. It captured when a start-up diversified for the first time, and explored the association between an early diversification strategy adopted by a firm, and its survival rate. A semi-parametric Cox proportional hazard model was used to examine the survival pattern. The results demonstrated that firms diversifying at an early stage in their lives show a higher survival rate; however, this effect fades over time.^

Dietary Intake Of Flavonoids And HDL- And LDL- Cholesterol In Two Black Ethnicities With And Without Type 2 Diabetes

Flavonoids are a class of over 6,500 plant metabolites that have been associated with reduced mortality from cardiovascular disease. A cross-sectional analysis of dietary flavonoids and serum cholesterol in 507 Blacks with and without type 2 diabetes (258 Haitian-Americans and 249 African-Americans) showed differences by ethnicity and diabetes status. Haitian-Americans consumed more of most flavonoids as compared to African-Americans. Individuals with type 2 diabetes consumed less of most flavonoids as compared to those without diabetes. Flavonoids were differentially associated with low-density lipoprotein cholesterol (LDL) and high-density lipoprotein cholesterol (HDL) by diabetes status. Flavanones were associated with lower LDL for participants without diabetes and higher LDL for those with diabetes, independent of ethnicity and adjusted for age, gender, cholesterol medications, daily energy, dietary fat, body mass index (BMI), and smoking. Flavan-3-ols were positively related to LDL while polyflavonoids (theaflavin and polymers, proanthocyanidins) were inversely related to LDL for the group without diabetes only. Higher anthocyanidins and flavan-3-ols and lower polyflavonoids were associated with higher HDL (same adjustments) for those without diabetes, whereas no flavonoids were associated with HDL for individuals with type 2 diabetes.

Can minority language proficiency pay: a study on the return on English-Spanish fluent bilingualism in South Florida

This thesis studies the economic return for fluent-bilingualism in South Florida among native-born whites using IPUMS (Integrated Public Use Microdata Series) data for Miami-Dade County (1990). Previous mainstream-oriented theories focus on the benefit in English acquisition for immigrants and their descendants, either denying or ignoring the possible benefit of minority language retention in addition to English acquisition. An alternative literature, on the other hand, suggests that minority language retention can be beneficial in at least three areas: 1) enhancing cognitive ability; 2) accessing community-level social capital; and 3) serving as human capital. This study assesses economic returns in employment and earnings, using logistic and OLS (Ordinary Least Square) regression respectively. The results, countering the mainstream-oriented theories, suggest that fluent bilingualism does have economic consequences. Rather than fully supporting the positive effects thesis, the patterns shown are much more complicated, contingent on an individual's ethnic membership and educational level. Theoretical and substantive implications are discussed and suggestions for future research are made.

(Table T1) N-alkane, n-alcohol and n-fatty acid concentrations in ODP Site 184-1147 and 184-1148 sediments

Lipid compositions of sediments recovered during Ocean Drilling Program Leg 184 in the South China Sea have been identified and quantified. The identified lipids can be ascribed to terrigenous and marine sources. Terrigenous lipids are mainly C27, C29, C31 n-alkanes, C26, C28, C30 n-fatty acids, and n-alcohols, which were derived from leaf waxes of higher land plants and transported to the sea by airborne dust or fresh water. Marine lipids, mainly C37 and C38 alkenones, C30 diol, and C30 and C32 keto-ols, were from microalgae, notably haptophytes and eustigmatophytes. Elevated concentrations and accumulation rates of both terrigenous and marine lipids in the interval 202-245 meters composite depth (mcd) and 0-166 mcd were postulated to be related to the development of the East Asian monsoon at 6-8 Ma and enhanced variations of the developed East Asian monsoon after 3.2 Ma, respectively. The pronounced late Oligocene input of terrigenous lipids reflects the paleoenvironment of a newly opened, narrow basin, with restricted ocean waters and the proximity of continental runoff.

Comparação de modelos estáticos, otimização dos determinantes da estrutura de capital das PME's portuguesas

Dissertação, Mestrado, Contabilidade e Fiscalidade, Instituto Politécnico de Santarém, Escola Superior de Gestão e Tecnologia, 2016

Efeitos da internacionalização no desvio da estrutura de capital alvo e na velocidade de ajuste

This dissertation investigates the effects of internationalization in two gaps related to the capital structure that have not been discussed by the Brazilian literature yet. To this, were developed two independent sections. The first examined what the effects of internationalization on the deviation from the target capital structure. The second examined what the effects of internationalization on speed of adjustment (SOA) of the capital structure. It used data from Brazil, multinational and domestic companies, from 2006 to 2014. The results of the first analysis indicate that internationalization helps reduce the difference between the target and the current debt. That is, to the extent that the level of internationalization increases; whether only export or a combination of export, assets and employees abroad, the gap between the current structure and the target structure decreases. This reduction is given as a function of internationalization as a consequence of the upstream effect of the upstream-downstream hypothesis. Thus, as the Market Timing theory, it can be seen as an opportunity for adjustment of the capital structure, and with the reduction of deviation, there is also a reduction in the cost of capital of the firm. The result of the second analysis indicates that internationalization is able to significantly increase the speed adjustment, ensuring for the multinational a faster adjustment of its capital structure. Exports increase the SOA in 9 to 23%. And when also kept active assets and employees abroad the increase is 8 to 20%. In terms of time, while domestic company takes more than three years to reduce half of the deviation that has, while multinacional companies take on average one and a half year to reduce the same proportion of the deviation. The validity of the upstream-downstream hypothesis for the effect of internationalization in SOA was confirmed by comparing the results for US companies. Thus, the phenomenon of internationalization increases SOA when companies are from less stable markets, such as Brazil; and it has a less significcative effect when companies are derived from more stable markets, because they already have a high speed of adjustmennt. In addition, the adequacy analysis of the estimators also showed the model pooled OLS (Ordinary Least Squares) presents the highest quality in predicting the SOA than the system GMM (Generalized Method of Moments). For future studies it is suggested to analyze the effect of international event, by itself, and to validate the hypothesis using samples of different markets and the use of other estimators.

Multilevel Governance and Accountability: Does Decentralization Promote Good Governance?

Today, the trend towards decentralization is far-reaching. Proponents of decentralization have argued that decentralization promotes responsive and accountable local government by shortening the distance between local representatives and their constituency. However, in this paper, I focus on the countervailing effect of decentralization on the accountability mechanism, arguing that decentralization, which increases the number of actors eligible for policy making and implementation in governance as a whole, may blur lines of responsibility, thus weakening citizens’ ability to sanction government in election. By using the ordinary least squares (OLS) interaction model based on historical panel data for 78 countries in the 2002 – 2010 period, I test the hypothesis that as the number of government tiers increases, there will be a negative interaction between the number of government tiers and decentralization policies. The regression results show empirical evidence that decentralization policies, having a positive impact on governance under a relatively simple form of multilevel governance, have no more statistically significant effects as the complexity of government structure exceeds a certain degree. In particular, this paper found that the presence of intergovernmental meeting with legally binding authority have a negative impact on governance when the complexity of government structure reaches to the highest level.

Food Swamps, Obesity & Health Zoning Restrictions on Fast Food Restaurants

Protecting public health is the most legitimate use of zoning, and yet there is minimal progress in applying it to the obesity problem. Zoning could potentially be used to address both unhealthy and healthy food retailers, but lack of evidence regarding the impact of zoning and public opinion on zoning changes are barriers to implementing zoning restrictions on fast food on a larger scale. My dissertation addresses these gaps in our understanding of health zoning as a policy option for altering built, food environments.

Chapter 1 examines the relationship between food swamps and obesity and whether spatial mapping might be useful in identifying priority geographic areas for zoning interventions. I employ an instrumental variables (IV) strategy to correct for the endogeneity problems associated with food environments, namely that individuals may self-select into certain neighborhoods and may consider food availability in their decision process. I utilize highway exits as a source of exogenous variation .Using secondary data from the USDA Food Environment Atlas, ordinary least squares (OLS) and IV regression models were employed to analyze cross-sectional associations between local food environments and the prevalence of obesity. I find even after controlling for food desert effects, food swamps have a positive, statistically significant effect on adult obesity rates.

Chapter 2 applies theories of message framing and prospect theory to the emerging discussion around health zoning policies targeting food environments and to explore public opinion toward a list of potential zoning restrictions on fast-food restaurants (beyond moratoriums on new establishments). In order to explore causality, I employ an online survey experiment manipulating exposure to vignettes with different message frames about health zoning restrictions with two national samples of adult Americans age 18 and over (N1=2,768 and N2=3,236). The second sample oversamples Black Americans (N=1,000) and individuals with high school as their highest level of education. Respondents were randomly assigned to one of six conditions where they were primed with different message frames about the benefits of zoning restrictions on fast food retailers. Participants were then asked to indicate their support for six zoning policies on a Likert scale. Subjects also answered questions about their food store access, eating behaviors, health status and perceptions of food stores by type.

I find that a message frame about Nutrition and increasing Equity in the food system was particularly effective at increasing support for health zoning policies targeting fast food outlets across policy categories (Conditional, Youth-related, Performance and Incentive) and across racial groups. This finding is consistent with an influential environmental justice scholar’s description of “injustice frames” as effective in mobilizing supporters around environmental issues (Taylor 2000). I extend this rationale to food environment obesity prevention efforts and identify Nutrition combined with Equity frames as an arguably universal campaign strategy for bolstering public support of zoning restrictions on fast food retailers.

Bridging my findings from both Chapters 1 and 2, using food swamps as a spatial metaphor may work to identify priority areas for policy intervention, but only if there is an equitable distribution of resources and mobilization efforts to improve consumer food environments. If the structural forces which ration access to land-use planning persist (arguably including the media as gatekeepers to information and producers of message frames) disparities in obesity are likely to widen.

A recessionary debate: who suffered more? A study of sectoral well-being and economic insecurity in Ireland

The aim of this thesis is to identify the relationship between subjective well-being and economic insecurity for public and private sector workers in Ireland using the European Social Survey 2010-2012. Life satisfaction and job satisfaction are the indicators used to measure subjective well-being. Economic insecurity is approximated by regional unemployment rates and self-perceived job insecurity. Potential sample selection bias and endogeneity bias are accounted for. It is traditionally believed that public sector workers are relatively more protected against insecurity due to very institution of public sector employment. The institution of public sector employment is made up of stricter dismissal practices (Luechinger et al., 2010a) and less volatile employment (Freeman, 1987) where workers are subsequently less likely to be affected by business cycle downturns (Clark and Postal-Vinay, 2009). It is found in the literature that economic insecurity depresses the well-being of public sector workers to a lesser degree than private sector workers (Luechinger et al., 2010a; Artz and Kaya, 2014). These studies provide the rationale for this thesis in testing for similar relationships in an Irish context. Sample selection bias arises when a selection into a particular category is not random (Heckman, 1979). An example of this is non-random selection into public sector employment based on personal characteristics (Heckman, 1979; Luechinger et al., 2010b). If selection into public sector employment is not corrected for this can lead to biased and inconsistent estimators (Gujarati, 2009). Selection bias of public sector employment is corrected for by using a standard Two-Step Heckman Probit OLS estimation method. Following Luechinger et al. (2010b), the propensity for individuals to select into public sector employment is estimated by a binomial probit model with the inclusion of the additional regressor Irish citizenship. Job satisfaction is then estimated by Ordinary Least Squares (OLS) with the inclusion of a sample correction term similar as is done in Clark (1997). Endogeneity is where an independent variable included in the model is determined within in the context of the model (Chenhall and Moers, 2007). The econometric definition states that an endogenous independent variable is one that is correlated with the error term (Wooldridge, 2010). Endogeneity is expected to be present due to a simultaneous relationship between job insecurity and job satisfaction whereby both variables are jointly determined (Theodossiou and Vasileiou, 2007). Simultaneity, as an instigator of endogeneity, is corrected for using Instrumental Variables (IV) techniques. Limited Information Methods and Full Information Methods of estimation of simultaneous equations models are assed and compared. The general results show that job insecurity depresses the subjective well-being of all workers in both the public and private sectors in Ireland. The magnitude of this effect differs among sectoral workers. The subjective well-being of private sector workers is more adversely affected by job insecurity than the subjective well-being of public sector workers. This is observed in basic ordered probit estimations of both a life satisfaction equation and a job satisfaction equation. The marginal effects from the ordered probit estimation of a basic job satisfaction equation show that as job insecurity increases the probability of reporting a 9 on a 10-point job satisfaction scale significantly decreases by 3.4% for the whole sample of workers, 2.8% for public sector workers and 4.0% for private sector workers. Artz and Kaya (2014) explain that as a result of many austerity policies implemented to reduce government expenditure during the economic recession, workers in the public sector may for the first time face worsening perceptions of job security which can have significant implications for their well-being (Artz and Kaya, 2014). This can be observed in the marginal effects where job insecurity negatively impacts the well-being of public sector workers in Ireland. However, in accordance with Luechinger et al. (2010a) the results show that private sector workers are more adversely impacted by economic insecurity than public sector workers. This suggests that in a time of high economic volatility, the institution of public sector employment held and was able to protect workers against some of the well-being consequences of rising insecurity. In estimating the relationship between subjective well-being and economic insecurity advanced econometric issues arise. The results show that when selection bias is corrected for, any statistically significant relationship between job insecurity and job satisfaction disappears for public sector workers. Additionally, in order to correct for endogeneity bias the simultaneous equations model for job satisfaction and job insecurity is estimated by Limited Information and Full Information Methods. The results from two different estimators classified as Limited Information Methods support the general findings of this research. Moreover, the magnitude of the endogeneity-corrected estimates are twice as large as those not corrected for endogeneity bias which is similarly found in Geishecker (2010, 2012). As part of the analysis into the effect of economic insecurity on subjective well-being, the effects of other socioeconomic variables and work-related variables are examined for public and private sector workers in Ireland.

Drug prescribing patterns in the General Medical Services Scheme in Ireland and projected costs to 2026

The primary objective is to investigate the main factors contributing to GMS expenditure on pharmaceutical prescribing and projecting this expenditure to 2026. This study is located in the area of pharmacoeconomic cost containment and projections literature. The thesis has five main aims: 1. To determine the main factors contributing to GMS expenditure on pharmaceutical prescribing. 2. To develop a model to project GMS prescribing expenditure in five year intervals to 2026, using 2006 Central Statistics Office (CSO) Census data and 2007 Health Service Executive{Primary Care Reimbursement Service (HSE{PCRS) sample data. 3. To develop a model to project GMS prescribing expenditure in five year intervals to 2026, using 2012 HSE{PCRS population data, incorporating cost containment measures, and 2011 CSO Census data. 4. To investigate the impact of demographic factors and the pharmacology of drugs (Anatomical Therapeutic Chemical (ATC)) on GMS expenditure. 5. To explore the consequences of GMS policy changes on prescribing expenditure and behaviour between 2008 and 2014. The thesis is centered around three published articles and is located between the end of a booming Irish economy in 2007, a recession from 2008{2013, to the beginning of a recovery in 2014. The literature identified a number of factors influencing pharmaceutical expenditure, including population growth, population aging, changes in drug utilisation and drug therapies, age, gender and location. The literature identified the methods previously used in predictive modelling and consequently, the Monte Carlo Simulation (MCS) model was used to simulate projected expenditures to 2026. Also, the literature guided the use of Ordinary Least Squares (OLS) regression in determining demographic and pharmacology factors influencing prescribing expenditure. The study commences against a backdrop of growing GMS prescribing costs, which has risen from e250 million in 1998 to over e1 billion by 2007. Using a sample 2007 HSE{PCRS prescribing data (n=192,000) and CSO population data from 2008, (Conway et al., 2014) estimated GMS prescribing expenditure could rise to e2 billion by2026. The cogency of these findings was impacted by the global economic crisis of 2008, which resulted in a sharp contraction in the Irish economy, mounting fiscal deficits resulting in Ireland's entry to a bailout programme. The sustainability of funding community drug schemes, such as the GMS, came under the spotlight of the EU, IMF, ECB (Trioka), who set stringent targets for reducing drug costs, as conditions of the bailout programme. Cost containment measures included: the introduction of income eligibility limits for GP visit cards and medical cards for those aged 70 and over, introduction of co{payments for prescription items, reductions in wholesale mark{up and pharmacy dispensing fees. Projections for GMS expenditure were reevaluated using 2012 HSE{PCRS prescribing population data and CSO population data based on Census 2011. Taking into account both cost containment measures and revised population predictions, GMS expenditure is estimated to increase by 64%, from e1.1 billion in 2016 to e1.8 billion by 2026, (ConwayLenihan and Woods, 2015). In the final paper, a cross{sectional study was carried out on HSE{PCRS population prescribing database (n=1.63 million claimants) to investigate the impact of demographic factors, and the pharmacology of the drugs, on GMS prescribing expenditure. Those aged over 75 (ẞ = 1:195) and cardiovascular prescribing (ẞ = 1:193) were the greatest contributors to annual GMS prescribing costs. Respiratory drugs (Montelukast) recorded the highest proportion and expenditure for GMS claimants under the age of 15. Drugs prescribed for the nervous system (Escitalopram, Olanzapine and Pregabalin) were highest for those between 16 and 64 years with cardiovascular drugs (Statins) were highest for those aged over 65. Females are more expensive than males and are prescribed more items across the four ATC groups, except among children under 11, (ConwayLenihan et al., 2016). This research indicates that growth in the proportion of the elderly claimants and associated levels of cardiovascular prescribing, particularly for statins, will present difficulties for Ireland in terms of cost containment. Whilst policies aimed at cost containment (co{payment charges, generic substitution, reference pricing, adjustments to GMS eligibility) can be used to curtail expenditure, health promotional programs and educational interventions should be given equal emphasis. Also policies intended to affect physicians prescribing behaviour include guidelines, information (about price and less expensive alternatives) and feedback, and the use of budgetary restrictions could yield savings.

Alkenone accumulation rates, stable carbon isotope record, and sea surface temperature reconstruction for IODP Site 306-U1313

Here we present orbitally-resolved records of terrestrial higher plant leaf wax input to the North Atlantic over the last 3.5 Ma, based on the accumulation of long-chain n-alkanes and n-alkanl-1-ols at IODP Site U1313. These lipids are a major component of dust, even in remote ocean areas, and have a predominantly aeolian origin in distal marine sediments. Our results demonstrate that around 2.7 million years ago (Ma), coinciding with the intensification of the Northern Hemisphere glaciation (NHG), the aeolian input of terrestrial material to the North Atlantic increased drastically. Since then, during every glacial the aeolian input of higher plant material was up to 30 times higher than during interglacials. The close correspondence between aeolian input to the North Atlantic and other dust records indicates a globally uniform response of dust sources to Quaternary climate variability, although the amplitude of variation differs among areas. We argue that the increased aeolian input at Site U1313 during glacials is predominantly related to the episodic appearance of continental ice sheets in North America and the associated strengthening of glaciogenic dust sources. Evolutional spectral analyses of the n-alkane records were therefore used to determine the dominant astronomical forcing in North American ice sheet advances. These results demonstrate that during the early Pleistocene North American ice sheet dynamics responded predominantly to variations in obliquity (41 ka), which argues against previous suggestions of precession-related variations in Northern Hemisphere ice sheets during the early Pleistocene.

Biomarkers in turbidites at ODP Sites 157-951 and 157-952

Free and ester-bound lipid biomarkers were analysed in oxidised and unoxidised parts of four distinct turbidites from the Madeira Abyssal Plain (MAP), which contained 1 to 2% organic carbon homogeneously distributed throughout the turbidites at the time they were deposited. These turbidites are well suited to study the effects of oxic degradation on lipid biomarkers without the complicating influence of varying organic matter sources, sedimentation rates, or bioturbation. One sample from the oxidised turbidite was compared with two samples from the unoxidised part of each turbidite. Postdepositional oxic degradation decreased concentrations of biomarkers by several orders of magnitude. The ester-bound lipids were degraded to a far lesser extent than their free counterparts were. The extent of degradation of different compounds differed substantially. Within a specific class of biomarkers, degradation also took place to a different extent, altering their distributions. This study shows that oxic degradation of the organic matter may have a profound effect on the biomarker fingerprint and may result in a severe bias in, for example, the interpretation of organic matter sources and the estimation of the palaeoproductivity of specific groups of phytoplankton.

Decadal-to-centennial scale variations during the Holocene reconstructed from sediment core D13902

Sea surface temperature (SST), marine productivity, and fluvial input have been reconstructed for the last 11.5 calendar (cal) ka B.P. using a high-resolution study of C37 alkenones, coccolithophores, iron content, and higher plant n-alkanes and n-alkan-1-ols in sedimentary sequences from the inner shelf off the Tagus River Estuary in the Portuguese Margin. The SST record is marked by a continuous decrease from 19C, at 10.5 and 7 ka, to 15C at present. This trend is interrupted by a fall from 18C during the Roman and Medieval Warm Periods to 16C in the Little Ice Age. River input was very low in the early Holocene but increased in the last 3 cal ka B.P. in association with an intensification of agriculture and deforestation and possibly the onset of the North Atlantic Oscillation/Atlantic Multidecadal Oscillation modes of variability. River influence must have reinforced the marine cooling trend relative to the lower amplitude in similar latitude sites of the eastern Atlantic. The total concentration of alkenones reflects river-induced productivity, being low in the early Holocene but increasing as river input became more important. Rapid cooling, of 1-2C occurring in 250 years, is observed at 11.1, 10.6, 8.2, 6.9, and 5.4 cal ka B.P. The estimated age of these events matches the ages of equivalent episodes common in the NE Atlantic- Mediterranean region. This synchronicity reveals a common widespread climate feature, which considering the twentieth century analog between colder SSTs and negative North Atlantic Oscillation (NAO), is likely to reflect periods of strong negative NAO.

Effets préventifs et thérapeutiques des polyphénols dans un modèle in vitro et in vivo de maladie inflammatoire de l’intestin : caractérisation des polyphénols de la pelure de pomme et de la canneberge par spectrométrie de masse

La muqueuse intestinale est exposée à des agents oxydants provenant de l’ingestion d’aliments modifiés, de cellules immuno-inflammatoires et de la flore intestinale. Une diète élevée en fruits et légumes peut diminuer le stress oxydant (SOx) ainsi que l’inflammation via plusieurs mécanismes. Ces effets bénéfiques peuvent être attribuables à leur contenu élevé en polyphénols. La première étude de mon doctorat consistait à tester l’hypothèse que les polyphénols extraits de pelures de pomme (DAPP) pouvaient diminuer le stress oxydant et l'inflammation impliqués dans les maladies inflammatoires de l'intestin (MII). Nous avons caractérisé les polyphénols des DAPP par spectrométrie de masse (LC-MS) et examiné leur potentiel antioxydant et anti-inflammatoire au niveau des cellules intestinales. L’identification des structures chimiques des polyphénols a été effectuée par LC-MS. Le SOx a été induit par l’ajout du complexe fer/ascorbate (Fe/Asc, 200 µM/2 mM) et l’inflammation par la lipopolysaccharide (LPS, 200µg/mL) à des cellules intestinales Caco-2/15 pré-incubées avec les DAPP (250 µg/mL). L’effet du SOx est déterminé par le dosage du malondialdéhyde (MDA), de la composition des acides gras polyinsaturés et de l’activité des enzymes antioxydantes endogènes (SOD et GPx). L’impact des DAPP sur l’inflammation a été testé par l’analyse de l’expression des marqueurs inflammatoires: cyclooxygénase-2 (COX-2), le facteur de nécrose tumorale alpha (TNF-a et l’interleukine-6 (IL-6) et les facteurs de transcription NF-KB, Nrf-2 et PGC1α par immunobuvardage. Nos données ont montré que les flavonols et les flavan-3-ols constituent les composés polyphénoliques majoritaires des DAPP. L’ajout de Fer2+/Asc a provoqué une augmentation de la peroxidation lipidique comparativement aux cellules contrôles, un appauvrissement des acides gras polyinsaturés n-3 et n-6, et une modulation des enzymes antioxydantes, se traduisant par une augmentation de l’activité de la SOD et une diminution de la GPx. En contrepartie, les DAPP ont exhibé leur potentiel à corriger la plupart des perturbations, y compris l’expression protéique anormalement élevée du COX-2 et la production de la prostaglandine E2 (PGE2), ainsi que l’inflammation telle que réflétée par les facteurs NF-κB, TNF-α et IL-6. Par ailleurs, les mécanismes sous-jacents à ces changements bénéfiques des DAPP ont fait intervenir les facteurs de transcription antioxydants (Nrf-2, PGC1α). Vraisemblablement, cette première étude a permis de démontrer la capacité des DAPP à amoindrir le SOx et à réduire l’inflammation, deux processus étroitement impliqués dans les MII. Dans la deuxième étape de mon doctorat, nous avons voulu comparer les résultats de DAPP à ceux des polyphénols dérivant de la canneberge qui est considérée par la communauté scientifique comme le fruit ayant le plus fort potentiel antioxydant. À cette fin, nous avons caractérisé l’effet des composés polyphénoliques de la canneberge (CPC) sur le SOx, la défense antioxydante et l’inflammation au niveau intestinal tout en définissant leur métabolisme intraluminal. Les différents CPC ont été séparés selon leur poids moléculaire par chromatographie et leurs structures chimiques ont été identifiées par LC-MS. Suite à une pré-incubation des cellules Caco-2/15 avec les extraits CPC (250 µg/mL), le Fe/Asc et la LPS ont été administrés comme inducteurs du SOx et de l’inflammation, respectivement. La caractérisation globale des CPC a révélé que les acides phénoliques composaient majoritairement l’extrait de canneberge de petit poids moléculaire (LC) alors que les flavonoïdes et les procyanidines dimériques/trimériques représentaient l’extrait de poids moléculaire moyen (MC) tout en laissant les procyanidines oligo et polymériques à l’extrait de haut poids moléculaire (HC). Les CPC ont permis de restaurer la plupart des perturbations engendrées dans les Caco-2/15 par le Fe/Asc et le LPS. Les CPC exhibaient le potentiel d’abaisser les niveaux de MDA, de corriger la composition des acides gras polyinsaturés n-3 et n-6, d’augmenter l’activité des enzymes antioxydantes (SOD, GPx et CAT) et d’élever l’expression de Nrf2 et PGC1α. En outre, les CPC pouvaient aussi réduire les niveaux élevés des protéines inflammatoires COX-2, TNF-α et IL-6 ainsi que la production des PGE2 par un mécanisme impliquant le NF-κB. Au niveau mitochondrial, les procyanidines oligomériques ont réussi à corriger les dysfonctions reliées à la production d’énergie (ATP), l’apoptose (Bcl-2, Cyt C et AIF) et le statut des facteurs de transcription mitochondriaux (mtTFA, mtTFB1, mtTFB2). Dans le but de bien comprendre les mécanismes d’action des CPC, nous avons défini par LC-MS les composés polyphénoliques qui ont été transportés ou absorbés par l’entérocyte. Nos analyses soulignent le transport (i) des acides cinnamiques et benzoïques (LC); (ii) la quercétine glycosylée et conjuguée et les procyanidines dimériques de type A (MC); et (iii) l’épicatéchine et les procyanidines oligomériques (HC). Les processus de métabolisation (méthylation, glucuronidation et sulfatation) au niveau de l’entérocyte ont probablement permis le transport de ces CPC surtout sous leur forme conjuguée. Les procyanidines oligomériques ayant un degré de polymérisation supérieur à 2 (HC) ont semblé adhérer aux cellules Caco-2/15. L’épicatéchine suivi par les procyanidines dimériques de type A ont été trouvés majoritaires au niveau des mitochondries. Même si nous ignorons encore l’action biologique de chaque composé polyphénolique, nous pouvons suggérer que leurs effets combinatoires exercent des fonctions antioxydantes, anti-inflammatoires et mitochondriales dans le modèle intestinal Caco-2/15. Dans une troisième étape, nous avons procédé à l’évaluation des aspects préventifs et thérapeutique des DAPP tout en sondant les mécanismes sous-jacents dans une étude préclinique. À cette fin, nous avons exploité le modèle de souris avec colite expérimentale provoquée par le Dextran Sulfate de Sodium (DSS). L’induction de l’inflammation intestinale chez la souris C57BL6 a été effectuée par l’administration orale de DSS à 2.5% pendant 10 jours. Des doses physiologiques et supra-physiologiques de DAPP (200 et 400 mg/kg/j, respectivement) ont été administrées par gavage pendant 10 jours pré- et post-DSS. L’inflammation par le DSS a provoqué une perte de poids, un raccourcissement du côlon, le décollement dystrophique de l’épithélium, l’exulcération et les infiltrations de cellules mono et polynucléaires au niveau du côlon. De plus, le DSS a induit une augmentation de la peroxidation lipidique, une régulation à la baisse des enzymes antioxydantes, une expression protéique à la hausse de la myéloperoxidase (MPO), du COX-2 et de la production des PGE2. Par ailleurs, les DAPP ont permis de corriger ou du moins d’alléger la plupart de ces anomalies en situation préventive ou thérapeutique, en plus d’abaisser l’expression protéique de NF-κB et des cytokines inflammatoires (TNF-a et l’IL-6) tout en stimulant les facteurs de transcription antioxydants (Nrf-2, PGC1α). Conséquemment, les polyphénols des DAPP ont exhibé leur puissant pouvoir antioxydant et anti-inflammatoire au niveau intestinal dans un modèle in vivo. Leurs actions sont associées à la régulation des voies de signalisation cellulaire et des changements dans la composition du microbiote. Ces trois projets de recherche permettent d’envisager l’évaluation des effets préventifs et thérapeutiques des DAPP cliniquement chez les patients avec des désordres inflammatoires de l’intestin.