The objective of this study was to determine whether dietary fat supplementation and level of undegradable intake protein (UIP) could affect daily milk output and composition by influencing nutrient supply to the mammary gland. Three lactating Holstein cows (60, 68, and 74 d postpartum) were used in an incomplete 4 x 4 Latin square design (2 x 2 factorial) and fed diets (15.9% CP and 19.5% ADF) with 0% or 2.5% partially hydrogenated tallow and 33% or 41% UIP. A 5:2.5:1 mixture of dried brewer's grains, corn gluten meal, and blood meal was substituted for soybean meal to raise dietary UIP from 33% to 41% UIP. Despite similar DM intake across treatments, cows produced 9% more milk per day when fed 2.5% supplemental fat, 41% UIP, or the combination of 2.5% fat and 41% UIP when compared to the control diet. Fat supplementation depressed milk protein percentage but not daily milk protein output. Mammary blood flow was estimated using the Fick principle at 6-hr intervals for 24 h. Concentration of individual nutrients in arterial (carotid) and venous (abdominal vein) blood and corresponding blood flows were used to calculate nutrient uptakes by the mammary gland. Calculated carbon uptake was 95 to 101% of output when using estimated carbon content of nutrients, and 100 to 106% when using an elemental analyzer to determine actual carbon output in milk. Uptake of glucose, β-hydroxybutyrate, lactate, pyruvate, acetate, and O₂ were not affected by dietary treatment. Triacylglycerol concentration in arterial blood and uptake of long-chain fatty acids were elevated by fat supplementation, resulting in milk fat with a higher percentage of 18-carbon fatty acids and a lower ratio of saturated to unsaturated fatty acids. Arterial essential and total amino acid (AA) concentrations in plasma and whole blood were elevated when cows were fed 41% versus 33% UIP. However, mammary arteriovenous differences, extraction percentages, and uptakes of most AA were not significantly affected by dietary treatments. Across treatments, peptide AA accounted for ~10% of AA in arterial whole blood but no net uptake of peptide AA by the lactating gland was detected. Results indicated that dietary fat supplementation at two levels of UIP can increase milk production by altering mammary lipid metabolism, thereby improving the efficiency of milk synthesis. However, depression of milk protein percentage in response to dietary fat supplementation was not alleviated by elevating arterial essential and total AA through higher dietary UIP. / Ph. D.

Thesis (MPath)--Stellenbosch University, 2014. / ENGLISH ABSTRACT: Major Depressive Disorder (MDD) is a severe debilitating medical condition that may lead to suicide. Due to a poor understanding of the biological mechanisms underlying the disease process therapeutic decisions are usually taken using a ‘trial and error’ approach. This is not ideal since many treatments do not work as expected for all individuals. Studies have shown that only half of MDD patients receive the appropriate treatment, whereas many patients have adverse response to anti-depressants. These may include weight gain and raised homocysteine levels that may further compromise the health status of MDD patients and may partly explain the link with cardiovascular disease. The objective of the study was to identify genetic risk factors interacting with environmental factors implicated in MDD that may be of relevance to the South African population. Polymorphisms in the MTHFR (677 C>T, rs1801133 and 1298 A>C, rs1801131), COMT (472G>A, rs4680), CYP2D6 (6937G>A, rs3892097), ASMT (24436 G>A, rs4446909) and SLC6A4 (43 bp ins/del, rs4795541) genes were genotyped in 86 MDD patients and 97 population-matched controls. The specific aims were 1) to analytically validate high throughput real-time polymerase chain reaction (RT-PCR) genotyping assays for the selected SNPs against direct sequencing as the gold standard for 2) possible integration into a pathology-supported genetic testing strategy aimed at improved clinical management of MDD. A total of 183 unrelated Caucasians participated in the study, including 69 females and 17 males with MDD and 57 female and 40 male controls without a personal and family medical history of overlapping stress/anxiety and depressive disorders. All study participants were genotyped for the six selected SNPs considered clinically useful based on international data. The allelic distribution of the SNPs, single or combined into a genotype risk score after counting their minor alleles, did not differ between MDD patients and controls. Homocysteine levels were determined and correlated with body mass index (BMI) and other variables known to influence these phenotypes. The folate score assessed with use of the study questionnaire was significantly lower in the patient group compared with controls (p=0.003) and correlated significantly with BMI, particularly in females (p=0.009). BMI was on average 8% higher in the MDD patients compared with controls (p=0.015) after adjustment for age and sex. The MTHFR rs1801133 677 T-allele was associated with a 14% increase in BMI in MDD patients but not controls (p=0.032), which in turn was associated with significantly increased homocysteine levels (p<0.05). The aims of the study were successfully achieved. Identification of the MTHFR rs1801133 677 T-allele reinforces the importance of adequate folate intake in the diet due to increased risk of obesity and depression found to be associated with low dietary intake. Evidence of shared genetic vulnerability for many chronic diseases and drug response mediated by the MTHFR 677 T-allele support the clinical relevance of this low-penetrance mutation. / AFRIKAANSE OPSOMMING: Major depressie (MD) is ‘n aftakelende siektetoestand wat tot selfdood kan lei. Onkunde oor die siekte se onderliggende biologiese meganismes lei dikwels tot ‘n lukrake terapeutiese benadering. Dit is ‘n onbevredigende situasie aangesien indiwidue verskillend reageer op die middels wat voorgeskryf word. Navorsing toon dat slegs ongeveer die helfte van MD pasiënte toepaslike behandeling kry, terwyl anti-depressante ‘n nadelige uitwerking het op baie pasiënte. Dit sluit massatoename en verhoogde homosisteïenvlakke in wat MD pasiënte se gesondheid bykomend nadelig kan beïnvloed en die verband met kardiovaskulêre siekte gedeeltelik kan verklaar. Hierdie studie poog om MD verwante genetiese risikofaktore en omgewingsfaktore wat mekaar beïnvloed en moontlik op die Suid Afrikaanse bevolking betrekking het, te identifiseer. Polimorfismes in die MTHFR (677 C>T, rs1801133 en 1298 A>C, rs1801131), COMT (472G>A, rs4680), CYP2D6 (6937G>A, rs3892097), ASMT (24436 G>A, rs4446909) en SLC6A4 (43 bp ins/del, rs4795541) gene is geanaliseer in 86 MD pasiënte en 97 kontroles geselekteer van dieselfde populasie. Die spesifieke doelwitte was om 1) hoë deurset direkte polimerase kettingreaksie (RT-PCR) genotiperingstoetse vir die 6 gekose polimorfismes met direkte volgordebepaling as maatstaf analities te valideer vir 2) moontlike insluiting in ‘n patologie-ondersteunde genetiese toetsstrategie met die oog op beter kliniese hantering van MD. Altesaam 183 Kaukasiërs het aan die studie deelgeneem. Die MD pasiënte het uit 69 vroue en 17 mans bestaan. Die kontroles (57 vroue en 40 mans) het geen mediese geskiedenis (persoonlik of familie) van oorvleuelende stress/angstigheid of depressie gehad nie. Gebaseer op internasionale data, is al die deelnemers vir die 6 gekose, potensieel klinies-bruikbare polimorfismes getoets. Die alleliese verspreiding van die polimorfismes enkel of gekombineer (uitgedruk as ‘n genotipe-risiko-syfer nadat minor allele getel is), was dieselfde in MD-pasiënte en kontroles. Homosisteïenvlakke is bepaal en gekorreleer met die liggaamsmassa-indeks (BMI) en ander veranderlikes wat bekend is vir hulle invloed op hierdie fenotipes. In teenstelling met die kontroles, was die folaat telling, soos bepaal met die studievraelys, betekenisvol laer in die pasiënte (p=0.003). Die korrelasie met die liggaamsmassa-indeks, spesifiek by vroue, was ook betekenisvol (p=0.009). Na aanpassings vir ouderdom en geslag, is gevind dat die liggaamsmassa-indeks gemiddeld 8% hoër was in die die MD pasiënte teenoor die kontroles. By MD-pasiënte, maar nie by die kontroles nie, is die MTHFR rs1801133 677 T-alleel geassosieer met ‘n 14% toename in liggaamsmassa-indeks (p=0.032), wat ook geassosieer was met betekenisvolle verhoogde homosisteïenvlakke (p<0.05). Die doelwitte van die studie is bereik. Identifisering van die MTHFR rs1801133 677 T-alleel beklemtoon hoe belangrik dit is om voldoende folaat in te neem, veral omdat ‘n verhoogde risiko vir vetsug en depressie met ‘n lae folaatinname in die diet geassosieer word. Die kliniese belang van die MTHFR 677 T-alleel word beklemtoon deur toenemende bewyse wat daarop dui dat gedeelde genetiese vatbaarheid vir ‘n verskeidenheid van kroniese siektes asook middelrespons aan bemiddeling deur hierdie lae penetrasie mutasie toegeskryf kan word. / Winetech / Technology for Human Resources and Industry Program (THRIP).

The purpose of this study was to develop a support programme for facilitating the integration of nutrition and food security with HIV prevention, treatment and care. The study was organised in three phases. Phase one was a quantitative cross-sectional survey that employed a structured interview with people living with HIV among selected two public hospitals and three health centres. The second phase employed focus group discussion with senior health experts to explore their perspective and experience in integrating nutrition and food security with HIV prevention, treatment, and care. The findings indicated that malnutrition and food insecurity were highly prevalent and significantly affected the treatment outcome and quality of life of PLWHA in the region. Socio-economic, clinical features and structural factors, such as educational status, place of residence, household income, source of drinking water, kind of toilet facility, inadequate dietary diversity, poor asset possession, opportunistic infections, duration on ART, CD4 cell count, and health system-related factors such as lack of viral and CD4 analysis laboratories and inconsistent antiretroviral medication supply were found as predictors of malnutrition and food insecurity. To cope up with the dire impact of malnutrition and food insecurity, short term, erosive and unsustainable food consumption coping strategies were employed. Based on the findings, the researcher developed a support programme for facilitating the integration of nutrition and food security with HIV prevention, treatment and care as phase three of the study. The developed programme is holistic and focuses on multi- and intersectoral collaboration to improve the treatment outcome, quality of life and overall wellbeing people living with HIV. / Health Studies / D. Litt. et Phil. (Health Studies)

該前瞻性研究對香港中國裔孕婦的25羥基維生素D（25（OH）D）的水平及其影響因素進行調查，并對25（OH）D與甲狀旁腺激素（PTH）、孕期肌肉酸痛、不良妊娠結局、孕期及産後骨質流失，以及嬰兒的骨骼發育等關係進行探索，力求建立適用于香港的中國孕婦的25（OH）D正常值。 / 共有237名單胎妊娠婦女以及62名多胎妊娠的婦女在2010年8月至2011年11月間參加本研究中的隊列研究，分別在參加研究時（<20 孕周）、24-28孕周、31-36孕周以及産後6-11周進行抽血測量血清25（OH）D以及PTH水平，同時填寫一份包括對每月攝取含維生素D的食物以及營養補充劑頻度、接受日照情況及喜好、以及肌肉不適等情況的問卷，并在24-28孕周進行75克口服葡萄糖耐量試驗。參與隊列研究的單胎孕婦在20周前、31-36孕周以及産後隨訪時接受用定量超聲測量非優勢手的橈骨遠端以及中指近掌指骨的骨質超聲速率（SoS）。在産後複查時，對其嬰兒左側腓骨中部的骨質SoS進行測量。記錄婦女各次檢查時的體重、抽血月份紫外線輻射強度的歷史記錄、以及妊娠結局。另外募集一批孕婦參加病例對照研究，比較患早產（PTB）、子癇前期（PET）、妊娠糖尿病 （GDM）以及胎兒生長受限（FGR）併發癥的婦女與對照組 （體重指數以及抽血時紫外線強度配對）的血清25（OH）D水平。 / 孕婦在孕期的平均25（OH）D水平在44.7 ± 12.6 至48.9 ± 17.1 nmol/l範圍，25（OH）D水平與體重指數、維生素D營養補充劑、抽血時紫外線強度以及個人對陽光的喜好情況有關，而與胎兒數量、孕次、孕周以及終止妊娠無關。 / 單胎妊娠的孕婦三個孕期的血清25（OH）D與PTH水平均負相關，但在多胎妊娠中，二者無明顯相關性。PTH在孕期以及産後的變化相對不受25（OH）D影響。孕婦25（OH）D的水平與孕婦肌肉酸痛癥狀、産後恢復、孕期及產褥期骨質流失以及嬰兒骨質無關。患早期PTB（< 34孕周）、PET或FGR的孕婦的血清25（OH）D比對照組低，但GDM患者的25（OH）D水平與對照組無差別。血清25（OH）D低於34.3 nmol/l者的早期早產以及子癇前期的風險增高，低於50 nmol/l者發生胎兒生長受限的風險增高。服用維生素D補充劑情況可能影響25（OH）D與FGR的關係。 / 總而言之，血清25（OH）D水平不足以全面完全反映孕期維生素D的情況，對預測不良妊娠結局的作用有限。 / This prospective study explored the maternal serum level of 25(OH)D in Chinese pregnant women in Hong Kong and the factors affecting 25(OH)D level. It also explored the correlation between maternal 25(OH)D with PTH level, maternal musculoskeletal complaints, adverse pregnancy outcome, maternal bone turnover during pregnancy and postpartum, and the bone development of the offspring, aiming to explore and establish a normal range of 25(OH)D level in pregnancy for the Hong Kong Chinese women. / A total of 237 women with singleton pregnancy and 62 women with multiple pregnancies were recruited for the cohort study from August, 2010 to November, 2011. Maternal blood samplings for 25(OH)D and PTH measurements were performed at recruitment, 24-28 weeks, 31-36 weeks of gestation, and 6-11 weeks postpartum respectively. A questionnaire which included the monthly dietary and supplement intake of vitamin D, questions about sunlight exposure, and musculoskeletal complaints was administered on each visit. A 75g oral glucose tolerance test (OGTT) was performed on cohort cases at 24-28 weeks of gestation. Measurements of the speed of sound (SoS) at the distal one third of the maternal radius and the proximal phalanx of the third finger of the non-dominant side were performed with quantitative ultrasonography (QUS) measurement during the visits at the first and third trimesters, and postnatal period. The SoS at the left mid-shaft tibia of the offspring was determined during the postnatal visit. Maternal characteristics, ultraviolet radiation (UVR) intensity at blood sampling, and pregnancy outcome, were also recorded. Cases with pregnancy complications were recruited for case-control studies, and maternal 25(OH)D level was examined with respect to preterm birth (PTB), preeclampsia (PET), gestational diabetes (GDM), and fetal growth restriction (FGR, birthweight below the 10th percentile of the customized estimated birthweight). The controls were matched for booking body mass index (BMI) and UVR intensity at blood sampling. / The mean 25(OH)D level in ranged from 44.7 ± 12.6 to 48.9 ± 17.1 nmol/l in the three trimesters, and was related to BMI, vitamin D supplementation, UVR intensity at blood sampling, and the acceptance of sunlight exposure, but not the number of fetus, parity, gestational age, or the completion of pregnancy. / Inverse correlation between PTH and 25(OH)D were observed in singleton, but not in multiple, pregnancy. The change in maternal PTH level is found to be relatively independent from that of 25(OH)D. There was no correlation between maternal 25(OH)D level with musculoskeletal complaints, postnatal recovery, bone turnover during and after pregnancy, or the bone density of the offspring. Maternal 25(OH)D level was lower in women with early PTB ( < 34 weeks), PET, and FGR, but not for GDM. A maternal 25(OH)D level of lower than 34.3nmol/l and 50 nmol/l was associated with increased risk of early PTB, PET, and FGR respectively. But the correlation between maternal 25(OH)D level with FGR might be affected by supplementation. / In conclusion, serum level of 25(OH)D is insufficient in reflecting maternal vitamin D status and metabolism in pregnancy, and is of limited use in predicting adverse pregnancy outcome. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Hu, Zhiyang. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 201-223). / Abstracts and appendixes also in Chinese. / Thesis dedication --- p.i / Acknowledgments --- p.ii / Abstract --- p.v / Abstract (Chinese) --- p.viii / List of Abbreviation --- p.x / Table of contents --- p.xiii / List of Figures --- p.xxii / List of Tables --- p.xxiv / Chapter Chapter 1: --- Literature Review --- p.1 / Chapter 1.1 --- The synthesis and metabolism of vitamin D --- p.3 / Chapter 1.1.1 --- The synthesis of vitamin D --- p.3 / Chapter 1.1.2 --- The metabolism of vitamin D --- p.4 / Chapter 1.1.3 --- Vitamin D binding protein --- p.10 / Chapter 1.1.4 --- Factors related to 25(OH)D level --- p.11 / Chapter 1.2 --- Function of vitamin D --- p.13 / Chapter 1.2.1 --- Mechanism of vitamin D function --- p.13 / Chapter 1.2.2 --- Classic function --- p.14 / Chapter 1.2.3 --- Non-classic function --- p.16 / Chapter 1.2.3.1 --- Immune system --- p.17 / Chapter 1.2.3.2 --- Cardiovascular system --- p.18 / Chapter 1.2.3.3 --- Cell proliferation and differentiation --- p.18 / Chapter 1.2.3.4 --- Neurological system --- p.19 / Chapter 1.2.3.5 --- Reproductive system --- p.20 / Chapter 1.2.3.6 --- Fetal development --- p.21 / Chapter 1.3 --- The definition of vitamin D deficiency --- p.21 / Chapter 1.4 --- Vitamin D status and pregnancy --- p.24 / Chapter 1.4.1 --- Alteration in vitamin D metabolism during pregnancy --- p.24 / Chapter 1.4.2 --- Factors affecting maternal serum level of 25(OH)D --- p.25 / Chapter 1.4.3 --- Vitamin D and bone resorption during pregnancy and lactation --- p.27 / Chapter 1.4.3.1 --- Alteration of calcium metabolism, bone absorption and the role of vitamin D --- p.27 / Chapter 1.4.3.2 --- Measurement of bone density in pregnant women and babies --- p.33 / Chapter 1.4.4 --- Current studies on maternal vitamin D status and pregnancy outcome --- p.35 / Chapter 1.4.4.1 --- Birthweight --- p.35 / Chapter 1.4.4.2 --- Infection --- p.37 / Chapter 1.4.4.3 --- Preterm delivery --- p.39 / Chapter 1.4.4.4 --- Diabetes (DM) and gestational diabetes (GDM) --- p.39 / Chapter 1.4.4.5 --- Hypertension and preeclampsia --- p.41 / Chapter 1.4.4.6 --- Multiple pregnancy, muscular symptoms --- p.42 / Chapter 1.4.4.7 --- Vitamin D supplementation and pregnancy outcome --- p.44 / Chapter 1.5 --- Defining vitamin D deficiency in pregnancy --- p.45 / Chapter 1.6 --- Objective of the study --- p.46 / Chapter Chapter 2: --- Study design and methods --- p.48 / Chapter 2.1 --- Case recruitment and study design --- p.48 / Chapter 2.1.1 --- Longitudinal singleton study --- p.49 / Chapter 2.1.2 --- Cross-sectional study --- p.50 / Chapter 2.1.2.1 --- Preterm birth (PTB) --- p.51 / Chapter 2.1.2.2 --- Preeclampsia (PET) --- p.51 / Chapter 2.1.2.3 --- Gestational diabetes (GDM) --- p.52 / Chapter 2.1.3 --- Multiple pregnancy study --- p.52 / Chapter 2.2 --- Measurements --- p.53 / Chapter 2.2.1 --- Hormonal analysis of serum levels of 25(OH)D and PTH --- p.53 / Chapter 2.2.2 --- Calculation of monthly intake of vitamin D from diet --- p.55 / Chapter 2.2.3 --- SoS measurements --- p.56 / Chapter 2.2.4 --- Ultraviolet radiation strength assessment --- p.59 / Chapter 2.3 --- Statistical analysis --- p.60 / Chapter Chapter 3 --- Longitudinal Study on the Level of and Factors Affecting Vitamin D in Singleton Pregnancy --- p.62 / Chapter 3.1 --- Introduction --- p.62 / Chapter 3.2 --- Material and method --- p.63 / Chapter 3.3 --- Statistics --- p.64 / Chapter 3.4 --- Results --- p.65 / Chapter 3.4.1 --- Demographic data of the subjects --- p.65 / Chapter 3.4.2 --- Maternal levels of 25(OH)D and PTH, and the factors affecting their levels --- p.66 / Chapter 3.4.2.1 --- Distribution of 25(OH)D level and PTH level in the four visits --- p.66 / Chapter 3.4.2.2 --- Dietary intake of vitamin D and supplementation --- p.69 / Chapter 3.4.2.3 --- Seasonality and sunlight exposure --- p.73 / Chapter 3.4.2.4 --- Parity --- p.76 / Chapter 3.4.3 --- Changes of maternal levels of 25(OH)D and PTH in pregnancy --- p.78 / Chapter 3.4.4 --- Independent factors related to maternal 25(OH)D level in pregnancy --- p.79 / Chapter 3.4.5 --- Maternal and fetal 25(OH)D level at delivery --- p.80 / Chapter 3.4.6 --- Muscular symptoms and other complaints in pregnancy, pregnancy outcome, and their relationships with maternal 25(OH)D level --- p.81 / Chapter 3.4.7 --- Postnatal recovery and factors related to postnatal level of 25(OH)D and PTH --- p.86 / Chapter 3.4.7.1 --- Postnatal symptoms and relationship with 25(OH)D and PTH --- p.86 / Chapter 3.4.7.2 --- The postnatal level of 25(OH)D and PTH in women with different feeding mode --- p.88 / Chapter 3.4.7.3 --- Independent factors related to postnatal 25(OH)D and PTH level --- p.89 / Chapter 3.4.7.4 --- Factors related to the change of 25(OH)D and PTH after delivery --- p.90 / Chapter 3.4.8 --- Correlation between 25(OH)D with PTH in pregnancy and postnatal period --- p.91 / Chapter 3.5 --- Discussion --- p.92 / Chapter 3.5.1 --- 25(OH)D level in Chinese pregnant women --- p.92 / Chapter 3.5.2 --- Factors related to maternal 25(OH)D level --- p.93 / Chapter 3.5.2.1 --- Dietary and supplementation --- p.93 / Chapter 3.5.2.2 --- Seasonality and outdoor activity --- p.96 / Chapter 3.5.2.3 --- Gestational age --- p.98 / Chapter 3.5.2.4 --- Age and parity --- p.98 / Chapter 3.5.3 --- Relationship of 25(OH)D level in the cord blood with maternal 25(OH)D level --- p.99 / Chapter 3.5.4 --- 25(OH)D level and muscular complains in pregnancy --- p.100 / Chapter 3.5.5. --- Postnatal recovery and 25(OH)D level --- p.101 / Chapter 3.5.6 --- PTH level in pregnancy and postnatal period --- p.101 / Chapter 3.6 --- Conclusion --- p.102 / Chapter Chapter 4 --- Longitudinal Study on the Relationship between Maternal 25(OH)D level with Changes of Maternal Bone Density in Pregnancy and Lactation, and Factors Affecting Bone Density of newborn Infants --- p.105 / Chapter 4.1 --- Introduction --- p.105 / Chapter 4.2 --- Material and method --- p.106 / Chapter 4.3 --- Statistics --- p.108 / Chapter 4.4 --- Results --- p.108 / Chapter 4.4.1 --- Demographic data --- p.108 / Chapter 4.4.2 --- Maternal bone density and the changes in pregnancy and postnatal recovery --- p.109 / Chapter 4.4.2.1 --- Maternal bone density in the first trimester and related factors --- p.109 / Chapter 4.4.2.2 --- Maternal bone density in the three visits --- p.109 / Chapter 4.4.2.3 --- The change in maternal bone density in the three visits --- p.110 / Chapter 4.4.2.4 --- Diversity in the change of bone density in pregnant women --- p.112 / Chapter 4.4.3 --- Factors related to the changes in bone density --- p.114 / Chapter 4.4.3.1 --- Changes between the first and the third trimesters --- p.114 / Chapter 4.4.3.2 --- Change between the third trimester and postnatal visits --- p.116 / Chapter 4.4.4 --- The bone density in infants and related factors --- p.120 / Chapter 4.5 --- Discussion --- p.122 / Chapter 4.5.1 --- Maternal bone density changes in pregnancy and postnatal period --- p.122 / Chapter 4.5.2 --- Factors related to the maternal bone density changes in pregnancy and postnatal period --- p.124 / Chapter 4.5.2.1 --- Initial bone density, parity, and BMI --- p.125 / Chapter 4.5.2.2 --- 25(OH)D and PTH level --- p.126 / Chapter 4.5.2.3 --- Supplement --- p.127 / Chapter 4.5.2.4 --- Lactation --- p.128 / Chapter 4.5.2.5 --- Height --- p.129 / Chapter 4.5.3 --- Factors related to bone density of the infant. --- p.130 / Chapter 4.5.3.1 --- Maternal 25(OH)D level --- p.130 / Chapter 4.5.3.2 --- Gestational age and birthweight --- p.131 / Chapter 4.5.3.3 --- Maternal bone density change --- p.131 / Chapter 4.5.3.4 --- The gender of the offspring and feeding method --- p.132 / Chapter 4.6 --- Conclusion --- p.133 / Chapter Chapter 5 --- Maternal 25(OH)D Level in Multiple Pregnancy --- p.134 / Chapter 5.1 --- Introduction --- p.134 / Chapter 5.2 --- Material and method --- p.135 / Chapter 5.3 --- Statistics --- p.136 / Chapter 5.4 --- Results --- p.137 / Chapter 5.4.1 --- Demographic data of the subjects --- p.137 / Chapter 5.4.2 --- The level of 25(OH)D in multiple pregnancy and singleton pregnancy --- p.137 / Chapter 5.4.3 --- Supplementation in multiple pregnancy --- p.140 / Chapter 5.4.4 --- The change of maternal 25(OH)D and PTH levels in the three trimesters --- p.141 / Chapter 5.4.5 --- 25(OH)D level in cord blood and its correlation with 25(OH)D level of the sibling --- p.143 / Chapter 5.4.6 --- Correlation between 25(OH) with PTH in pregnancy --- p.143 / Chapter 5.5 --- Discussion --- p.144 / Chapter 5.5.1 --- 25(OH)D level in multiple pregnancy and singleton pregnancy --- p.144 / Chapter 5.5.2 --- Supplementation in multiple pregnancy --- p.146 / Chapter 5.5.3 --- Changes of maternal levels of 25(OH)D and PTH in the three trimesters in multiple pregnancy --- p.146 / Chapter 5.5.4 --- The PTH/25(OH) correlation --- p.147 / Chapter 5.6 --- Conclusion --- p.148 / Chapter Chapter 6 --- Maternal level of 25(OH)D in complicated pregnancy --- p.150 / Chapter 6.1 --- Introduction --- p.150 / Chapter 6.2 --- Method --- p.153 / Chapter 6.2.1 --- Preterm birth --- p.155 / Chapter 6.2.2 --- Preeclampsia --- p.155 / Chapter 6.2.3 --- Gestational diabetes --- p.156 / Chapter 6.2.4 --- Fetal growth restriction --- p.157 / Chapter 6.2.5 --- The association between 25(OH)D level with pregnancy complication --- p.158 / Chapter 6.3 --- Statistics --- p.159 / Chapter 6.4 --- Results --- p.160 / Chapter 6.4.1 --- Setting of the cutoff values of hypovitaminosis D --- p.160 / Chapter 6.4.2 --- Preterm birth --- p.160 / Chapter 6.4.3 --- Preeclampsia --- p.164 / Chapter 6.4.4 --- Gestational diabetes --- p.168 / Chapter 6.4.4.1 --- Case-control study --- p.168 / Chapter 6.4.4.2 --- Factors affecting OGTT results --- p.170 / Chapter 6.4.5 --- Fetal growth restriction --- p.173 / Chapter 6.5 --- Discussion --- p.179 / Chapter 6.5.1 --- Adjustment for confounders for case-control study --- p.179 / Chapter 6.5.2 --- PTB and 25(OH)D level --- p.181 / Chapter 6.5.3 --- PET and 25(OH)D level --- p.182 / Chapter 6.5.4 --- GDM and 25(OH)D level --- p.186 / Chapter 6.5.5 --- FGR and 25(OH)D level --- p.189 / Chapter 6.5.6 --- Defining vitamin D deficiency in pregnancy --- p.192 / Chapter 6.6 --- Conclusion --- p.195 / Chapter Chapter 7 --- Summary --- p.196 / References --- p.201 / Chapter Appendix 1 --- Antenatal questionnaire (English/Chinese) --- p.224 / Chapter Appendix 2 --- Postnatal questionnaire (English/Chinese) --- p.238

Thesis (MScMedSc (Biomedical Sciences))--University of Stellenbosch, 2010. / Thesis presented in partial fulfillment of the requirements for the degree Master of Science in Medical Biochemistry at the University of Stellenbosch. / Bibliography / ENGLISH ABSTRACT: Tuberculosis (TB) is an infectious disease which has plagued society for thousands of years. Despite public health programs, anti-TB drugs and a vaccine, the absolute numbers of people infected with TB each year continue to rise as populations grow. The high TB-burden areas are also plagued by other debilitating factors including HIV/AIDS infection, poverty and malnutrition. Nutrition has been implicated in TB susceptibility in a number of studies. While most are observational reports made during times of war, famine or natural disaster, multiple studies provide convincing evidence for poor nutritional status increasing the morbidity and mortality of TB. Numerous approaches are currently utilized in TB research, and there has been convincing evidence to support the role of host genetics in TB susceptibility. Based on previous linkage studies and a search of current literature, three genes were selected for this case-control study. Subsequently, variations located in cathepsin Z (CTSZ), melanocortin 3 receptor (MC3R) and melanocortin 4 receptor (MC4R) were genotyped in the South African Coloured (SAC) population to determine the existence of an association with TB disease. CTSZ is a lysosomal cysteine protease expressed in cells of the immune system. Interaction between this 303 amino acid protein and β2 integrin receptors lymphocyte function-associated antigen-1 (LFA-1) and macrophage antigen-1 (MAC-1) leads to altered lymphocyte proliferation. As a result, a single exonic variant in CTSZ, rs34069356, the same identified in a previous linkage study, showed strong evidence for association with TB susceptibility in cases (n = 410) and controls (n = 301) in the SAC population (p< 0.0001). MC3R and MC4R are two of 5 melanocortin receptors. MC3R has been found to be a key regulator in energy expenditure and host metabolism while activation of MC4R leads to a decrease in food intake. Activation of these two receptors is regulated by leptin, a hormone released by adipose tissue. A variant located upstream of the MC3R gene, rs6127698, also showed evidence of disease association with the less frequent allele, T, being under-represented in cases (n = 540) compared to controls (n = 541) (genotypic frequency, p = 0.0039), suggesting a possible resistance phenotype. Functional analysis of this variant revealed an increase in MC3R expression when stimulated with BCG, with individuals homozygous for the T allele exhibiting an even larger upregulation of MC3R expression than individuals homozygous for the G allele, though this difference was not statistically significant. A single haplotype in MC3R was found to be associated with TB susceptibility (p = 0.0008) and this association remained after permutation testing to correct for multiple testing (p = 0.0061) Three variants were selected for genotyping in MC4R and while none of these showed a statistically significant difference between cases (n = 510) and controls (n = 487), this gene should not be ruled out as both MC3R and MC4R have been found to work closely though not redundantly and double knockout experiments result in exacerbated obesity, suggesting that these proteins have a synergistic effect. The results of this study support both a role of host genetics and nutritional status in TB and strongly motivate further research in both of these fields. / AFRIKAANSE OPSOMMING: Tuberkulose (TB) is ‘n aansteeklike siekte wat reeds vir eeue die gesondheid van die publiek bedreig. Ten spyte van publieke gesondheidsprogramme en verskeie anti-TB medikasie middele, groei die aantal van mense wat hiermee ge-infekteer word steeds jaarliks. Dit is veral in areas waar TB steeds groei, waar ook ander neerdrukkende faktore soos HIV/Vigs, armoede en wanvoeding hoogty vier. Na aanleiding van verskeie verslae tydens oorloë, hongersnood en ander natuulike rampe is dit veral duidelik dat swak nutriënt inname morbiditeit en sterftes wat met TB gepaard gaan verhoog. Talle benaderings word tans gebruik in TB-navorsing, Bewyse is oortuigend om die rol van genetika van die gaheer met vatbaarheid vir TB te verbind. Op grond van vorige studies en die huidige literatuur, het ons drie gene gekies vir hierdie pasiënt-kontrole studie. Variante geleë in cathepsin Z (CTSZ), melanocortin 3 receptor (MC3R) en melanocortin 4 receptor (MC4R) is ge-genotipeer in die Suid-Afrikaanse Kleurling bevolking (SAK) (540 gevalle en 540 kontrole) om sodoende die assosiasie met TB te bepaal. CTSZ is ‘n lisosomale sisteïen protease wat uitgedruk word in immuunselle. Interaksie tussen hierdie 303 aminosuur protein en β2 integrin reseptore nl. LFA-1 en MAK-1 bring veranderde limfosiet proliferasie mee. ‘n Enkele eksoniese variant in CTSZ, rs34069356, dieselfde soos ge-identifiseer in ‘n vorige studie, verskaf sterk bewys vir assosiasie met TB vatbaarheid in gevalle (n = 410) en kontrole (n = 301) in die SAK bevolking. MC3R en MC4R is twee van 5 melanokortien reseptore. Daar is gevind dat MC3R 'n sleutelrol speel in die energie regulering van gasheer metabolisme, terwyl die aktivering van MC4R eindelik lei tot 'n afname in voedsel inname. Aktivering van hierdie twee reseptore word gereguleer deur Leptien, 'n hormoon wat vrygestel word deur adipose weefsel, ‘n Variant, stroomop geleë vanaf MC3R, rs6127698, is ook bewys om met TB ge-assosieer te wees, met die T-alleel meer seldsaam in gevalle (n = 540) as in kontroles (n = 541) wat dui op 'n moontlike weerstandsfenotipe. Funksionele analise van hierdie variant onthul 'n toename in MC3R uitdrukking wanneer gestimuleer met BCG, met individue homosigoties vir die T-alleel wat selfs groter opregulation veroorsaak wanneer vergelyk word met individue homosigoties vir die G allele. Hierdie resultaat was egter nie statisties beduidend nie. 'n Enkele haplotiepe in MC3R is ge-assosieer met TB vatbaarheid en die assosiasie is onveranderd nadat ‘n permutasie korreksie aangebring is (p = .0061). Voorts is drie variante gekies vir genotipering in MC4R en ten spyte daarvan dat nie een daarvan 'n statisties beduidende verskil getoon het tussen pasiënte (n = 510) en kontroles (n = 487) nie, behoort hierdie geen nie uitgesluit word nie, Die rede hiervoor is dat beide MC3R en MC4R verskeie kere gevind is om in samewerking ‘n rol te speel om vetsug te voorkom of te vererger. Die resultate van hierdie studie beaam beide 'n rol van gasheer genetika en voedingstatus in TB en motiveer veral verdere navorsing in beide van hierdie vakgebiede.

Thesis (MPhil (Rehabilitation))--University of Stellenbosch, 2011. / ENGLISH ABSTRACT: In South Africa, rapid urbanisation and epidemiological transition have left the black urban population vulnerable to diseases of lifestyle such as chronic heart failure. This is in part due to the fact that changes in dietary patterns during urbanization play an important role in the increase of risk factors of these diseases. However, there is a lack of information on dietary choices of black urban populations. Therefore the current study evolved to describe the food choices and macro-and micronutrient intake of black, urban Sowetans, newly diagnosed with chronic heart failure, who attended the outpatient cardiac clinic at Chris Hani Baragwanath Hospital. A descriptive study methodology that made use of quantitative methods of data collection was used. Study participants comprised Sowetans with chronic heart failure who attended the Chris Hani Baragwanath Hospital outpatient cardiac clinic for the first time. Consecutive sampling followed by stratified random sampling was used to identify study participants. Participants were stratified for gender. Hundred persons participated in the study. Data was collected through the Food Frequency Questionnaire, a demographic questionnaire and measuring of height and weight. Data from the FFQ's was analysed for macro- and micronutrient intake by using the MRC "Food Finder 3‟ programme. Data were analysed by a statistician using StatSoft, Inc. (2009) STATISTICA, version 9.0. A p value of 0.05 was seen as statistically significant. The most significant clinical finding is an inadequate intake of certain micro nutrients and excessive salt consumption. Study participants continued to eat the more traditional carbohydrate foods. These staples were supplemented by highly refined carbohydrate sources, such as added sugar, sweets and chocolates, cakes, biscuits and cold drinks. Women ate significantly more maltabella (p=0.04), sweets and chocolates (p=0.01) than men, while men consumed significantly more, meat (p=0.01), milk and milk products (p=0.04), additional salt (p=0.02) and take away foods (p=0.05). Both genders had inadequate intake of Vitamin D [men 4 mcg/day (p=0.00), and women, 4 mcg/day (p=0.01)], selenium, [46 mcg/day (p=0.03) and 32 mcg/day (p=0.00)], folate [215 mcg/day (p=0.00) and 179 mcg/day (p=0.00)] and Vitamin C [71 mg/day (p=0.05) and 66 mg/day (p=0.07)]. Women had an inadequate intake of iron of 9 mg/day (P=0.00). It is recommended that dietary health promotion packages are developed and targeted specifically at this high risk community. Key words: Chronic heart failure, black, urban, food choices, macro-and micronutrients. / AFRIKAANSE OPSOMMING: Die swart stedelike bevolking in Suid Afrika gaan gebuk onder 'n al groter wordende risiko vir leefstyl siektes soos kroniese hartversaking. Dit kan gedeeltelik toegeskryf word aan veranderinge in dieet patrone as gevolg van verstedeliking en die epidemiologiese oorgang. Daar is egter nie genoeg inligting oor die voedselkeuses van swart stedelike bevolkingsgroepe nie. Die huidige studie het dus ontwikkel uit die behoefte om die voedselkeuses en mikro- en makronutrient inname van swart, stedelike Soweto inwoners wat nuut gediagnoseer is met hartversaking en die buitepasiënt kardiologie kliniek by Chris Hani Baragwanath Hospitaal bygewoon het, te bepaal. Daar was gebruik gemaak van 'n beskrywende studie metodologie wat gebruik gemaak het van kwantitatiewe metodes van data insameling. Deelnemers aan die studie het bestaan uit swart inwoners van Soweto met kroniese hartversaking wat die buitepasiënt kardiologie kliniek by Chris Hani Baragwanath Hospitaal vir die eerste keer bygewoon het. 'n Opeenvolgende steekproef, gevolg deur gestratifiseerde steekproefneming was gebruik om die studie deelnemers te identifiseer. Deelnemers was gestratifiseer volgens geslag. Eenhonderd pesone het aan die studie deelgeneem. Data is ingesamel deur gebruik te maak van die Voedsel Frekwensie Vraelys, a demografiese vraelys en die meet van lengte en gewig. Data van die Voedsel Frekwensie Vraelyste was ge-analiseer vir mikro-en makronutrient inname met die MRC ”Food Finder 3” program. Data is ge-analiseer deur 'n statistikus met die „StatSoft, Inc. (2009) STATISTICA, version 9.0‟. 'n P waarde van 0.05 is gesien as statisties beduidend. Mees beduidendste kliniese bevinding was die ontoereikende inname van sekere mikro-nutriënte en die verhoogde inname van sout. Studie deelnemers het nog steeds die meer tradisionele koolhidraat voedsels geëet. Hierdie stapel voedsels was aangevul deur hoogs verfynde bronne van koolhidrate, soos ekstra suiker, lekkergoed en sjokolade, koek, koekies en koeldrank. Die vrouens het beduidend meer maltabella (p=0.01), lekkergoed en sjokolade (p=0.01) geëet as mans, terwyl mans beduidend meer vleis (p=0.01), melk en melkprodukte (p=0.04), bygevoegde sout (p=0.02) en wegneem kosse (p=0.05) ingeneem het. Beide geslagte het ontoereikende innames van vitamiene D [mans 4 mcg/dag (p=0.00), en vrouens, 4 mcg/dag (p=0.01)], selenium [46 mcg/dag (p=0.03) en 32 mcg/dag (p=0.00)], foliensuur [215 mcg/dag (p=0.00) en 179 mcg/dag (p=0.00)] en vitamiene C [71 mg/dag (p=0.05) en 66 mg/dag (p=0.07)]. Vrouens het 'n ontoereikende inname van yster van 9 mg/dag (p=0.00) gehad. Daar word aanbeveel dat gesonde voedingsprogramme ontwikkel word, spesifiek gemik op hierdie bevolkingsgroep. Sleutelwoorde: Kroniese hartversaking, swart, verstedeliking, voedselkeuses, makro- en mikronutriënte.

This thesis consists of a brief description on cancer, carcinogenesis, the changes in the type and level of dietary fat available in our diets over time and association with the development of certain diseases. The main focus of this research was on omega 6 and omega 3 essential fatty acids (EFA) and their interaction with regards to carcinogenesis.

The possibility that faulty expectations about success and relapse recovery contributed to poor adherence was examined in this study. Support for such an expectancy model was sought through comparing an index of relative task magnitude to adherence rates. Instructions designed to improve adherence through changing expectations about relapse and relapse recovery were also administered to 46 clients in two clinical nutritional programs. Their adherence rates <in days) were compared to the rates obtained from the records of 64 other clients who did not receive the instructions. To further understand the adherence phenomenon, several other measures were obtained from the treatment subjects. These data were compared to adherence rates in an attempt to identify potential co-variate relationships. Statistical procedures including analysis of variance to determine comparability of subject groups, Pearson Product Moment correlations, t tests of the difference between means, and the Lawshe—Baker Nomograph comparing per cent adherence rates were performed on the data. Obtained results did not support the predicted relationship between relative task magnitude and adherence. This may have been due to differences between subjective assessments of task magnitude and the objective measure used in this study. Although improvement in adherence was noted in both treatment groups, statistical significance was achieved only in the university based clinic. Differences in the settings, assisting nutritionists, and participating subjects could have produced these findings. However, because improvement did occur in both settings, and because the techniques may be easily and inexpensively utilized by clinical nutritionists, these instructions were recommended for inclusion as a routine component of nutritional clinic procedures. No strong co-variate relationships were found between adherence and the additional measures included in the study. The only variables which correlated with adherence more than trivially, emotional response to a verbal food stimulus, and imaging ability, did lend support for this cognitively active method of improving adherence.

The problem of this study is to determine the vitamin B-6 status of patients who have chronic obstructive pulmonary disease (COPD). Erythrocyte aspartate transaminase assay was the method for measuring vitamin B-6 status. The vitamin B-6 status was examined in thirty subjects (ten COPD subjects and twenty control subjects). An unpaired t-test was used to compare the vitamin B-6 status of the COPD group versus the control group. Four determinants (percentage stimulation, ratio of basal to stimulated activity, basal activity, and stimulated activity) were used to determine vitamin B-6 status in both groups of subjects. Percentage stimulation and ratio of basal to stimulated activity were not significantly different (control group versus COPD group) at the .05 level. However, two of ten COPD subjects had values for percentage stimulation that were two standard deviations above the mean, indicating a poor B-6 status. In contrast, basal activity and stimulated activity of erythrocyte aspartate transaminase were found to be significantly lower at the .05 level in the COPD group than the control group. Therefore, the COPD subjects as a group had some biochemical characteristics of a lower level of vitamin B-6 than the controls.

非傳染性疾病是目前全球最常見的疾病之一。不健康的食相信是導致非傳染性疾病增加的主要因素之一。因此，我們就食對乳腺癌的形成和膽固醇代謝調控的影響進行了研究。 / 在去除卵巢的祼鼠模型中，我們研究了長期和短期熱量限制對乳腺癌腫瘤增殖的影響。14週齡的小鼠被隨機分為5組：自由攝食組 (AL)；熱量攝入控制在AL80% 的20%CCR組；熱量攝入控制在AL的70% 的30%CCR組；熱量攝入控制在AL的65% 的35%CCR組和短期熱量限制 （SCR）組 (前3.5週熱量攝入控制在AL的65%，之後的13.5週自由攝食)。10週後，熱量限制組的腫瘤體積明顯較AL組小 (P < 0.05)。排除攝食對體重的影響，SCR組的腫瘤重量明顯較AL組小 (P < 0.05)。本實驗結果表明，在此動物模型中，短期熱量限制能有效抑制乳腺癌細胞的增殖。 / 此外，我們還研究了芹菜素在肝細胞中對膽固醇代謝的影響。芹菜素是一種常見的黃酮類化合物。研究發現，在WRL-68細胞中，芹菜素能夠劑量依賴性的抑制3 - 羥基-3 - 甲基 - 戊二酸單酰輔酶還原酶 (HMGCR)和固醇調節元件結合蛋白-2 (SREBP-2) 信使RNA和蛋白的表達及其啟動子的轉錄活性。綜上所述，在肝細胞中，芹菜素能有效抑制HMGCR和SREBP-2的表達，從而達到降低膽固醇的效果。 / 總括而言，本研究表明在去除卵巢的祼鼠模型中，短期熱量限制能有效抑制乳腺癌細胞的生長和芹菜素能有效抑制HMGCR和SREBP-2的表達。 / Non-communicable diseases (NCD) are one of the leading causes of mortality in the developed and under-developing countries. Diet is a major risk factor of NCD. In the present study, effects of diet modification on breast cancer development and cholesterol metabolism were investigated. / In the first part of this study, the effect of chronic and short-term calorie restriction (CR) on breast tumor growth in ovariectomized nude mice was investigated. The calorie-restricted dietary regimen limited the total fat intake only. 14 week-old ovariectomized female nude mice were randomly assigned to ad libitum fed (AL), 20%CCR (17-week 80% of AL), 30%CCR (17-week 70% of AL), 35%CCR (17-week 65% of AL) and short-term CR (3.5-week 65% of AL followed by 13.5-week 100% AL consumption) groups. Starting from 10 weeks after transplant of cells, the tumor volumes in all calorie-restricted groups were significantly smaller (P < 0.05) than that in ad libitum control. At sacrifice, the tumor weight in short-term CR was significantly smaller (P < 0.05) than that in ad-libitum control after normalized with body weight. This indicated that short-term CR could suppress tumor in this model. / In the second part of this study, the effect of apigenin on cholesterol metabolism was investigated. Apigenin is one of the most abundant flavonoids. In the present study, we investigated the effect of apigenin on several cholesterol-related gene expression in hepatic cells. In WRL-68 cells treated with apigenin, promoter transcription activity, mRNA and protein expression of HMGCR and SREBP-2 were significantly decreased in a dose-dependent manner. Taken together, we concluded that apigenin inhibited HMGCR and SREBP-2 gene expressions in hepatic cells, which might elicit the hypocholesterolemic effects. / In conclusion, our study has demonstrated that short-term CR could significantly block the breast tumor growth in a mice model and apigenin could inhibit the expression of HMGCR and SREBP-2 in liver cell lines. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Wong, Tsz Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 83-99). / Abstracts also in Chinese. / ACKNOWLEGEMENTS --- p.i / ABSTRACT --- p.ii / 摘要 --- p.iv / list of abbreviations --- p.v / list of figures --- p.vii / list of tables --- p.IX / TABLE of CONTENTS --- p.X / Chapter 1 --- CHAPTER 1 --- p.1 / General Introduction --- p.1 / Chapter 1.1 --- Calorie Restriction and the Prevention of Postmenopausal Breast Cancer --- p.2 / Chapter 1.1.1 --- Breast Cancer --- p.2 / Chapter 1.1.2 --- Epidemiology of Excess Body Weight and Cancer Risk --- p.3 / Chapter 1.1.3 --- Calorie Restriction and Cancer Prevention --- p.7 / Chapter 1.1.4 --- Mechanistic Targets of Calorie Restriction --- p.8 / Chapter 1.1.4.1 --- Effect of Calorie Restriction on Estrogen --- p.8 / Chapter 1.1.4.1 --- Effect of Calorie Restriction on Cell Cycle Regulation --- p.12 / Chapter 1.1.4.1 --- Effect of Calorie Restriction on Apoptosis --- p.14 / Chapter 1.2 --- Effect of Apigenin on Cholesterol Homeostasis --- p.17 / Chapter 1.2.1 --- Cardiovascular Disease and Blood Cholesterol --- p.17 / Chapter 1.2.2 --- Molecular Regulation of Cholesterol Metabolism --- p.21 / Chapter 1.2.2.1 --- HMG-CoA Reductase --- p.21 / Chapter 1.2.2.2 --- CYP7A1 --- p.24 / Chapter 1.2.2.3 --- Apolipoprotein A-1 --- p.26 / Chapter 1.2.2.4 --- Low Density Lipoprotein Receptor --- p.29 / Chapter 1.2.2.5 --- Sterol Regulatory Element Binding Proteins --- p.31 / Chapter 1.2.3 --- Flavonoid and its Association with Cholesterol Metabolism --- p.36 / Chapter 1.2.4 --- Apigenin: A Potential Alternative --- p.39 / Chapter 2 --- CHAPTER 2 --- p.41 / MATERIALS AND METHODS --- p.41 / Chapter 2.1 --- Chemicals and Materials --- p.41 / Chapter 2.1.1 --- Chemicals --- p.41 / Chapter 2.1.2 --- Plasmids --- p.41 / Chapter 2.2 --- Cell Culture --- p.41 / Chapter 2.2.1 --- Maintainance of Cells --- p.41 / Chapter 2.2.2 --- Preparation of Cell Stock --- p.42 / Chapter 2.2.3 --- Cell Recovery from Liquid Nitrogen Stock --- p.42 / Chapter 2.3 --- Measurement of Cell viability --- p.43 / Chapter 2.4 --- Semi-Quantitative and Quantitative RT-PCR Assay --- p.43 / Chapter 2.4.1 --- RNA Isolation and cDNA Synthesis --- p.43 / Chapter 2.4.2 --- Quantitative Real Time PCR Assay --- p.43 / Chapter 2.4.2.1 --- Real Time PCR Using TaqMan Probe --- p.43 / Chapter 2.4.2.2 --- Real Time PCR Using SYBR Green Dye --- p.44 / Chapter 2.4.2.3 --- Statistical Analysis of 2⁻ΔΔ{U+A7F0}{U+1D40} Comparative Gene Expression --- p.44 / Chapter 2.5 --- Western Blot Analysis --- p.46 / Chapter 2.6 --- Measurement of Promoter Activity --- p.46 / Chapter 2.6.1 --- Plasmid Preparation --- p.46 / Chapter 2.6.2 --- Transient Transfection and Dual-Luciferase Assay --- p.47 / Chapter 2.7 --- Animal Experiment Design --- p.47 / Chapter 2.7.1 --- Animal Model and Dietary Regimens --- p.47 / Chapter 2.7.2 --- Tissue Sample Collection --- p.50 / Chapter 2.7.3 --- Plasma Estradiol Determination --- p.50 / Chapter 2.7.4 --- Protein and RNA extraction --- p.50 / Chapter 2.8 --- Statistical Analysis --- p.50 / Chapter 3 --- Chapter 3 --- p.51 / EFFECT OF CHRONIC AND short-term calorie restriction on breast tumor growth in ovariectomized nude mice --- p.51 / Chapter 3.1 --- Introduction --- p.51 / Chapter 3.2 --- Objectives --- p.52 / Chapter 3.3 --- Results --- p.53 / Chapter 3.3.1 --- Food Intakes, Body, Liver and Uterus Wet Weights of the Mice --- p.53 / Chapter 3.3.2 --- Tumor Development --- p.57 / Chapter 3.3.3 --- Plasma Estradiol Level --- p.62 / Chapter 3.3.4 --- Estradiol Responsive Gene expression in Tumors --- p.63 / Chapter 3.3.5 --- Cell Apoptotic and Cell Cycle-Regulated Protein expression in Tumors --- p.65 / Chapter 3.4 --- Discussion --- p.67 / Chapter 4 --- CHAPTER 4 --- p.69 / Apigenin inhibits the expression of hmg-coa reductase and srebp-2 in hepatic cells --- p.69 / Chapter 4.1 --- Introduction --- p.69 / Chapter 4.2 --- Objectives --- p.70 / Chapter 4.3 --- Results --- p.70 / Chapter 4.3.1 --- Effect of Apigenin on Cell Viability --- p.70 / Chapter 4.3.2 --- Effect of Apigenin on HMGCR, CYP7A1, LDLR, ApoA-1, SREBP-1 and SREBP-2 mRNA expressions --- p.72 / Chapter 4.3.3 --- Effect of Apigenin on HMGCR, LDLR, ApoA-1 and SREBP-2 Promoter Transcription Activity --- p.75 / Chapter 4.3.4 --- Effect of Apigenin on HMGCR, SREBP-1 and SREBP-2 Protein Expression --- p.77 / Chapter 4.3.5 --- Role of Estrogen Receptor in Apigenin induced SREBP-2 Inhibition --- p.79 / Chapter 4.4 --- Discussion --- p.80 / Chapter 5 --- CHAPTER 5 --- p.82 / SUMMARY --- p.82 / References --- p.83