Phytosterols have been shown to be effective in reducing serum cholesterol levels in numerous human clinical studies and regular consumption is recommended as part of therapeutic lifestyle changes aimed at reducing low density lipoprotein (LDL-C) in the treatment of hyperlipidaemia, a risk factor for cardiovascular disease. Fat based spreads have been shown to be a very successful vehicle for delivery of plant sterols, readily accepted by consumers and efficacious in reducing cholesterol levels. Alfa One™ Rice Bran Oil (RBO) spread is a new product entering into the market place. It is derived from rice bran oil and contains high levels of unsaponifiable material rich in phytosterols, triterpene alcohols, ferulic acid esters ([gamma]-oryzanol) and vitamin E isomers. As such it may have the potential to lower serum cholesterol levels when consumed on a daily basis. In order to establish the effectiveness of Alfa One™ Rice Bran Oil (RBO) spread compared with Flora pro-activ® margarine, a well established brand of plant sterol margarine already proven to lower cholesterol, a randomised double blind cross-over human clinical trial over 12 weeks was conducted. The study was divided into two treatment arms. The first arm of the study was to determine whether Alfa One™ RBO spread (containing 1.5% plant sterols) could lower total and LDL cholesterol levels to a greater extent than standard Flora margarine (containing no plant sterols) or Flora Pro-activ® margarine (containing 8% plant sterols). The second study arm tested the proposition that daily consumption of Alfa One™ Rice Bran Oil (RBO) spread in conjunction with rice bran oil (containing 0.5% plant sterols) would lower total and LDL cholesterol to a greater extent than Alfa One™ RBO spread in isolation and more than Flora margarine in conjunction with sunflower oil. Eighty mildly hypercholesterolaemic individuals (total cholesterol [greater than or equal to] 5 mmol/L and [less than or equal to] 7.5 mmol/L) were recruited and randomised into two groups of forty. Participants were asked to continue with their normal dietary pattern but to replace any margarine/butter/fat consumption with the trial products. One group of 40 were then assigned to the first treatment arm of the study (margarine-only group) and were randomised to consume 20 g (4 teaspoons) Alfa One™ RBO spread daily for 4 weeks, or 20 g Flora margarine daily for 4 weeks, or 20 Flora pro-activ® daily for 4 weeks. Phytosterol levels delivered in these amounts were: RBO margarine: 118mg phytosterol and 14 mg [gamma]-oryzanol; Flora proactiv® 1600 mg phytosterol; Flora margarine 0mg phytosterol. The second group of 40 were allocated to the second arm of the trial (margarine and oil group) and consumed 20 g Alfa One™ RBO spread and 30 ml rice bran oil (RBO) daily for 4 weeks, or 20 g Flora margarine and 30 ml sunflower oil daily for 4 weeks, or 20 g Alfa One™ RBO spread daily for 4 weeks, changing treatment at the end of each 4-week period. Phytosterol amounts delivered in these amounts were: RBO margarine: 118 mg phytosterol and 14 mg [gamma] oryzanol; RBO 222mg mg phytosterol, 150 mg [gamma] oryzanol. Each participant consumed all three treatments in a random order over a 12 week period. At baseline and following each 4 week intervention period, measurements were made of weight and blood pressure. Venous blood samples were collected for analysis of total cholesterol, low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), total cholesterol: HDL-C, triglycerides and plasma phytosterols. Three-day diet records from each individual were also collected for analysis of normal dietary intake. Results showed that compared to a standard Flora margarine, Alfa One™ RBO spread significantly reduced total cholesterol by 2.2% (P=0.045), total cholesterol:HDL by 4.1% (P=0.005) and LDL-C by 3.5% (P=0.016), but was not as effective overall as Flora Pro-activ® which reduced total cholesterol by 4.4% (P=0.001), total cholesterol:HDL by 3.4% (P=0.014) and LDL-C by 5.6% (P=0.001). Consumption of Flora margarine alone produced no significant decrease from baseline figures in any of the cholesterol parameters measured. Surprisingly, in group two, the addition of rice bran oil to the Alfa One™ RBO spread produced no differences in cholesterol levels. The reason for this unexpected result is being explored further. These results confirm that Alfa One™ RBO spread is effective in lowering serum cholesterol levels when consumed as part of a normal diet. Studies have shown that a 1% reduction in LDL-C can equate to a 2% decrease in coronary heart disease (CHD) risk thus suggesting that the 3.5% reduction demonstrated by Alfa One™ RBO spread in this study could be effective in reducing CHD risk as much as 6% in a mildly hypercholesterolaemic population.

Phytosterols have been shown to be effective in reducing serum cholesterol levels in numerous human clinical studies and regular consumption is recommended as part of therapeutic lifestyle changes aimed at reducing low density lipoprotein (LDL-C) in the treatment of hyperlipidaemia, a risk factor for cardiovascular disease. Fat based spreads have been shown to be a very successful vehicle for delivery of plant sterols, readily accepted by consumers and efficacious in reducing cholesterol levels. Alfa One™ Rice Bran Oil (RBO) spread is a new product entering into the market place. It is derived from rice bran oil and contains high levels of unsaponifiable material rich in phytosterols, triterpene alcohols, ferulic acid esters ([gamma]-oryzanol) and vitamin E isomers. As such it may have the potential to lower serum cholesterol levels when consumed on a daily basis. In order to establish the effectiveness of Alfa One™ Rice Bran Oil (RBO) spread compared with Flora pro-activ® margarine, a well established brand of plant sterol margarine already proven to lower cholesterol, a randomised double blind cross-over human clinical trial over 12 weeks was conducted. The study was divided into two treatment arms. The first arm of the study was to determine whether Alfa One™ RBO spread (containing 1.5% plant sterols) could lower total and LDL cholesterol levels to a greater extent than standard Flora margarine (containing no plant sterols) or Flora Pro-activ® margarine (containing 8% plant sterols). The second study arm tested the proposition that daily consumption of Alfa One™ Rice Bran Oil (RBO) spread in conjunction with rice bran oil (containing 0.5% plant sterols) would lower total and LDL cholesterol to a greater extent than Alfa One™ RBO spread in isolation and more than Flora margarine in conjunction with sunflower oil. Eighty mildly hypercholesterolaemic individuals (total cholesterol [greater than or equal to] 5 mmol/L and [less than or equal to] 7.5 mmol/L) were recruited and randomised into two groups of forty. Participants were asked to continue with their normal dietary pattern but to replace any margarine/butter/fat consumption with the trial products. One group of 40 were then assigned to the first treatment arm of the study (margarine-only group) and were randomised to consume 20 g (4 teaspoons) Alfa One™ RBO spread daily for 4 weeks, or 20 g Flora margarine daily for 4 weeks, or 20 Flora pro-activ® daily for 4 weeks. Phytosterol levels delivered in these amounts were: RBO margarine: 118mg phytosterol and 14 mg [gamma]-oryzanol; Flora proactiv® 1600 mg phytosterol; Flora margarine 0mg phytosterol. The second group of 40 were allocated to the second arm of the trial (margarine and oil group) and consumed 20 g Alfa One™ RBO spread and 30 ml rice bran oil (RBO) daily for 4 weeks, or 20 g Flora margarine and 30 ml sunflower oil daily for 4 weeks, or 20 g Alfa One™ RBO spread daily for 4 weeks, changing treatment at the end of each 4-week period. Phytosterol amounts delivered in these amounts were: RBO margarine: 118 mg phytosterol and 14 mg [gamma] oryzanol; RBO 222mg mg phytosterol, 150 mg [gamma] oryzanol. Each participant consumed all three treatments in a random order over a 12 week period. At baseline and following each 4 week intervention period, measurements were made of weight and blood pressure. Venous blood samples were collected for analysis of total cholesterol, low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C), total cholesterol: HDL-C, triglycerides and plasma phytosterols. Three-day diet records from each individual were also collected for analysis of normal dietary intake. Results showed that compared to a standard Flora margarine, Alfa One™ RBO spread significantly reduced total cholesterol by 2.2% (P=0.045), total cholesterol:HDL by 4.1% (P=0.005) and LDL-C by 3.5% (P=0.016), but was not as effective overall as Flora Pro-activ® which reduced total cholesterol by 4.4% (P=0.001), total cholesterol:HDL by 3.4% (P=0.014) and LDL-C by 5.6% (P=0.001). Consumption of Flora margarine alone produced no significant decrease from baseline figures in any of the cholesterol parameters measured. Surprisingly, in group two, the addition of rice bran oil to the Alfa One™ RBO spread produced no differences in cholesterol levels. The reason for this unexpected result is being explored further. These results confirm that Alfa One™ RBO spread is effective in lowering serum cholesterol levels when consumed as part of a normal diet. Studies have shown that a 1% reduction in LDL-C can equate to a 2% decrease in coronary heart disease (CHD) risk thus suggesting that the 3.5% reduction demonstrated by Alfa One™ RBO spread in this study could be effective in reducing CHD risk as much as 6% in a mildly hypercholesterolaemic population.

<p>Inflammation and oxidative stress may be involved in atherogenesis. This thesis describes clinical studies of prostaglandin F_2α (PGF_2α), an inflammatory mediator, and the isoprostane 8-iso-PGF_2α, a reliable indicator of oxidative stress, and cytokine-related inflammatory mediators and indicators in healthy subjects and in a population-based cohort of Swedish men. </p><p>PGF_2α and 8-iso-PGF_2α formation in healthy subjects varied considerably between days with a mean intra-individual coefficient of variation of 41 % and 42 %, respectively. A morning urine sample reflected the basal level of 8-iso-PGF_2α formation as accurately as a 24-hour urine collection, and represents a more practical alternative to the 24-hour urine collection in clinical studies. PGF_2α formation (as measured by urinary 15-keto-dihydro-PGF_2α) was increased in patients with type 2 diabetes and in smokers independent of other cardiovascular risk factors. These results indicated an on-going cyclooxygenase (COX)-mediated inflammatory reaction related to these conditions. Further, an increased formation of isoprostanes (as measured by urinary 8-iso-PGF_2α) was found in patients with type 2 diabetes and in smokers, indicating a high level of oxidative stress in these men. The smokers had also increased levels of the cytokine interleukin-6, indicating an on-going cytokine-related inflammatory reaction. The inflammatory indicators C-reactive protein and serum amyloid A were related to overweight but not independently associated to type 2 diabetes. High levels of serum selenium in middle-aged men predicted reduced formation of PGF_2α and 8-iso-PGF_2α 27 years later.</p><p>In summary, low-grade, chronic COX-mediated and possibly cytokine-related inflammation, and oxidative stress, seem to be joint features of type 2 diabetes and smoking, two major risk factors of atherosclerosis, in elderly men. Inflammation and oxidative stress may represent a possible common pathogenetic link between established risk factors for atherosclerosis and atherogenesis.</p>

<p>Serum amyloid A protein (SAA) is an acute phase protein that has recently gained increasing interest as a potential marker for disease and treatment monitoring. We investigated SAA and CRP levels in (a) patients with various common infectious diseases (n=98), (b) patients with pyelonephritis (n=37) versus patients with cystitis (n=32), (c) healthy individuals of varying ages (n=231), (d) very immature newborn infants with or without nosocomial infections (NIs) (n=72) and (e) patients with bacterial infections treated with cefuroxime (n=81). </p><p>SAA significantly correlated with CRP in viral as well as in bacterial infections (for the total group: r²=0.757, p<0.0001) and showed a systemic inflammatory response in 90% of the patients with cystitis as compared with 23% for CRP. Equally high efficiencies (0.96 and 0.94 for SAA and CRP, respectively) were observed in discriminating between pyelonephritis and cystitis. SAA and high sensitive (hs) CRP were lower in umbilical cords (p<0.0001) and higher in elderly adults (p<0.0001-0.03) than in the other age groups; higher in immature newborn infants than in term infants; and higher in the NI group than in the non-NI group. Interindividual variabilities of the time course of the biomarkers SAA and CRP were considerable. Because of the smoothed distribution of SAA and CRP (i.e. elevations were both essentially unchanged during the first 3 days of cefuroxime treatment), these markers were not useful when deciding parenteral-oral switch of therapy, which occurred within this time period in most cases.</p><p>SAA is a sensitive systemic marker in cystitis. SAA and hsCRP in umbilical cord blood are close to the detection limit and increase with age. They increase in relation to NI in very immature newborn infants and might therefore be used in diagnosis and monitoring. Finally, SAA and CRP in adults with bacterial infections could not predict an early parenteral-oral switch of antimicrobial therapy.</p>

This thesis is concerned with synthesis, structure and biochemical analysis of chemically modified oligonucleotides with potential therapeutic applications. The three types of chemical modifications described here are: (a) A North-East locked 1',2'-azetidine nucleoside (b) A North locked 2',4'-cyanomethylene bridged nucleoside and (c) A 2',4'-aza-ENA-T nucleoside. The synthesis of the 1',2'-azetidine fused nucleosides was described using two different approaches. A highly strained 2',4'-cyanomethylene locked nucleoside was synthesized but could not be converted to the phosphoramidite derivative due to instability during derivatization. The key cyclization step in the aza-ENA-T nucleoside synthesis gave rise to two separable diastereomers due to chirality at the exocyclic nitrogen. Conversion of diastereomer 55 to 56 occurred with a large free energy of activation (ΔG‡ = 23.4 kcal mol-1 at 298 K in pyridine-d5). Of the two isomers the equatorial NH product was more stable than the axial one due to reduced 1,3 diaxial interactions. As a result, all NH axial product was converted to the equatorial isomer during subsequent steps in the synthesis. NMR and ab initio experiments confirmed the North-East structure of the 1',2'-azetidine locked nucleoside and North conformation of aza-ENA-T locked nucleosides with a chair conformation of the piperidine ring. The amino modified nucleosides were incorporated into different positions of a 15mer oligonucleotide. The azetidine modified AONs did not form stable duplexes with complementary RNA (ΔTm ~-1 to -4 °C), but they performed better than previously synthesized isosequential 1',2'-oxetane modified oligonucleotides. The 2',4'-aza-ENA-T modified oligonucleotide, on the other hand, showed excellent target affinity with complementary RNA (ΔTm ~+4 °C). The azetidine and aza-ENA-T modified oligonucleotides showed significant stability in the presence of human serum and snake venom phosphodiesterase (3'-exonuclease) as compared to the unmodified native sequence. The singly modified 15mer oligonucleotides were also subjected to RNase H promoted digestion in order to evaluate their potential as effective antisense agents. The effective enzyme activity (kcat/Km) was found to be lower in the modified AONs due to reduced enzyme-substrate binding. However, the catalytic activity of RNase H with these modified-AON:RNA duplexes were higher than observed with the native duplex.

Inflammation and oxidative stress may be involved in atherogenesis. This thesis describes clinical studies of prostaglandin F2α (PGF2α), an inflammatory mediator, and the isoprostane 8-iso-PGF2α, a reliable indicator of oxidative stress, and cytokine-related inflammatory mediators and indicators in healthy subjects and in a population-based cohort of Swedish men. PGF2α and 8-iso-PGF2α formation in healthy subjects varied considerably between days with a mean intra-individual coefficient of variation of 41 % and 42 %, respectively. A morning urine sample reflected the basal level of 8-iso-PGF2α formation as accurately as a 24-hour urine collection, and represents a more practical alternative to the 24-hour urine collection in clinical studies. PGF2α formation (as measured by urinary 15-keto-dihydro-PGF2α) was increased in patients with type 2 diabetes and in smokers independent of other cardiovascular risk factors. These results indicated an on-going cyclooxygenase (COX)-mediated inflammatory reaction related to these conditions. Further, an increased formation of isoprostanes (as measured by urinary 8-iso-PGF2α) was found in patients with type 2 diabetes and in smokers, indicating a high level of oxidative stress in these men. The smokers had also increased levels of the cytokine interleukin-6, indicating an on-going cytokine-related inflammatory reaction. The inflammatory indicators C-reactive protein and serum amyloid A were related to overweight but not independently associated to type 2 diabetes. High levels of serum selenium in middle-aged men predicted reduced formation of PGF2α and 8-iso-PGF2α 27 years later. In summary, low-grade, chronic COX-mediated and possibly cytokine-related inflammation, and oxidative stress, seem to be joint features of type 2 diabetes and smoking, two major risk factors of atherosclerosis, in elderly men. Inflammation and oxidative stress may represent a possible common pathogenetic link between established risk factors for atherosclerosis and atherogenesis.

Serum amyloid A protein (SAA) is an acute phase protein that has recently gained increasing interest as a potential marker for disease and treatment monitoring. We investigated SAA and CRP levels in (a) patients with various common infectious diseases (n=98), (b) patients with pyelonephritis (n=37) versus patients with cystitis (n=32), (c) healthy individuals of varying ages (n=231), (d) very immature newborn infants with or without nosocomial infections (NIs) (n=72) and (e) patients with bacterial infections treated with cefuroxime (n=81). SAA significantly correlated with CRP in viral as well as in bacterial infections (for the total group: r2=0.757, p&lt;0.0001) and showed a systemic inflammatory response in 90% of the patients with cystitis as compared with 23% for CRP. Equally high efficiencies (0.96 and 0.94 for SAA and CRP, respectively) were observed in discriminating between pyelonephritis and cystitis. SAA and high sensitive (hs) CRP were lower in umbilical cords (p&lt;0.0001) and higher in elderly adults (p&lt;0.0001-0.03) than in the other age groups; higher in immature newborn infants than in term infants; and higher in the NI group than in the non-NI group. Interindividual variabilities of the time course of the biomarkers SAA and CRP were considerable. Because of the smoothed distribution of SAA and CRP (i.e. elevations were both essentially unchanged during the first 3 days of cefuroxime treatment), these markers were not useful when deciding parenteral-oral switch of therapy, which occurred within this time period in most cases. SAA is a sensitive systemic marker in cystitis. SAA and hsCRP in umbilical cord blood are close to the detection limit and increase with age. They increase in relation to NI in very immature newborn infants and might therefore be used in diagnosis and monitoring. Finally, SAA and CRP in adults with bacterial infections could not predict an early parenteral-oral switch of antimicrobial therapy.

Background: Vitamin D deficiency is a concern in the US. Association between vitamin D status and metabolic syndrome (MetS), insulin resistance (IR), and inflammation is unclear in children. Objective: The relationship between serum vitamin D and MetS, C-reactive protein (CRP), and Homeostatic Model Assessment-IR (HOMA-IR) was investigated. Design: Data from 3 cycles of National Health and Nutrition Examination Survey, 2001-2006 for 3700 (1820, boys; 1880, girls) children and adolescents, aged 12-17 y were used to assess prevalence of vitamin D deficiency (<20 ng>/mL) and association between serum vitamin D and prevalence of MetS, various components of MetS, CRP, and HOMA-IR using multivariate regression models. Results: Overall, prevalences of MetS and vitamin D deficiency were 6.1% and 30.5%, respectively. Prevalence of vitamin D deficiency was higher in girls (52%), blacks (74%), non-supplement users (50%), persons who were examined in winter (56%), and persons in the low poverty income ratio group (57%) compared to their counterparts. Serum vitamin D was inversely associated with waist circumference (P<0.001), systolic blood pressure (P=0.009), and HOMA-IR (P=0.003) and positively associated with HDL-cholesterol (P<0.001). Children with lowest serum vitamin D are at increased risk for MetS (P=0.04; OR 2.26; 95% CI: 1.11, 4.61). Serum vitamin D was not related to CRP (P<0.10). Conclusions: Children with poor vitamin D status are at increased risk for MetS and IR. Because of negative health outcomes associated with MetS and poor vitamin D status when existed individually or in combination, early detection and intervention of these conditions are paramount, especially in children.

Thesis (Ph.D.)--Indiana University, 2009. / Title from screen (viewed on February 2, 2010). Department of Biochemistry and Molecular Biology, Indiana University-Purdue University Indianapolis (IUPUI). Advisor(s): Mervin C. Yoder, Jr., David A. Ingram, Jr., Lawrence A. Quilliam, Mark D. Pescovitz. Includes vitae. Includes bibliographical references (leaves 171-194).

Our understanding of humoral immune response to human papillomavirus (HPV) infection has been mainly derived from studies in women. Very little is known about humoral immune response to HPV in men. There is also a growing interest in understanding the burden of HPV exposure in the subgroups of the male population, including men who have sex with women (MSW), men who have sex with men (MSM) and men who have sex with both men and women (MSMW). This dissertation was undertaken to understand and characterize humoral immune response, measured by detectable serum antibody IgG, to HPV 6, 11, 16 and 18 infection, to estimates seroprevalence of HPV 6, 11, 16 and 18, to determine the associations of sociodemographic and sexual behavioral factors with seroprevalence of individual HPV types, and to evaluate the role of serum antibodies in the subsequent acquisition of infection with the same HPV type, genetically related and un-related HPV types. Three studies that compose of this dissertation were conducted within the framework of two longitudinal studies of HPV infection in men: a single-site natural history study of male residents of Tucson, Arizona (the 1st study: N=285); and a multinational natural history study of healthy men residing in São Paulo, Brazil, Cuernavaca, Mexico, and Tampa, Florida (the 2nd study: N=1477; the 3rd study: N=2187). Men were recruited using similar eligibility criteria in both natural history studies and followed every 6 months for a maximum of 18 months in the single-site study and 48 months in the multi-national study. HPV DNA status was assessed using the PGMY09/11 L1 consensus primer system and the Linear Array HPV Genotyping Protocol. Testing of serum antibodies to HPV 6, 11, 16 and 18 was performed with virus-like particle-based ELISA assays. Data from our studies indicate that exposure to HPV 6, 11, 16 and 18, the four HPV types targeted in the currently license HPV vaccines, is common. Of 285 male residents of Tucson, Arizona, 28.8% of them were seropositive to HPV 16 and/or 18 at study entry. Similarly, approximately one third of 1477 participants of the multi-national male HPV natural history study were seropositive to at least one vaccine HPV type, with the percentage of 21.8% in U.S. site, 33.4% in Mexico site, and 49.1% in Brazil site. It is also noted that seroprevalence of individual vaccine HPV types is greatly elevated among men of different sexual practices. Seroprevalence of HPV 6, 11, 16 and/or 18 was twice as high among MSM and MSMW compared to MSW. Likewise, seroprevalence of individual HPV types was two fold or higher among MSW and MSMW. Our findings suggest that the predominant predictors of seropositivity to HPV 6, 11, 16 and 18 are age and same-sex sexual behaviors. Seroprevalence increased with age among young-to-middle-aged men with significant upward age trends observed for HPV 11, 16 and 18. MSM, compared to MSW, more likely to be seropositive to HPV 16 or 18. Similarly, men who practiced same-sex anal sex, compared to those who did not, were significantly more likely to be seropositive to HPV 6, 11, 16 and 18, respectively. Among 276 men free of HPV 16 at enrollment in Tucson, We did not detect statistically significant associations between the baseline serum antibodies to HPV 16 and/or 18 and subsequent risk of infection with homogeneous HPV types or related-HPV types. Of 2187 men residing in three countries who tested HPV 16 negative at enrollment, the risk of subsequent HPV 16 infection was not associated with enrollment HPV 16 serum antibodies status. Our data provide important estimates of population exposure to vaccine HPV types for future studies modeling potential vaccine impact and vaccine cost effectiveness in men. Our findings also support strategic vaccination of males as an effective preventive measure for HPV-related diseases and cancers in men and their sex partners, men and women alike.