Alexandra Krettek

Hypoxia Increases LDL Oxidation and Expression of 15-Lipoxygenase-2 in Human Macrophages

Objective—Macrophage-mediated oxidation of low-density lipoprotein (LDL) by enzymes, such as the lipoxygenases, is considered of major importance for the formation of oxidized LDL during atherogenesis. Macrophages have been identified in hypoxic areas in atherosclerotic plaques. Methods and Results—To investigate the role of hypoxia in macrophage-mediated LDL oxidation, we incubated human monocyte-derived macrophages with LDL under normoxic (21% O2) or hypoxic (0% O2) conditions. The results showed that hypoxic macrophages oxidized LDL to a significantly higher extent than normoxic cells. Interestingly, the mRNA and protein expression of 15-lipoxygenase-2 (15-LOX-2) as well as the activity of this enzyme are elevated in macrophages incubated at hypoxia. Both the unspliced 15-LOX-2 and the spliced variant 15-LOX-2sv-a are found in macrophages. In addition, 15-LOX-2 was identified in carotid plaques in some macrophage-rich areas but was only expressed at low levels in nondiseased arteries. Conclusions—In summary, these observations show for the first time that 15-LOX-2 is expressed in hypoxic macrophages and in atherosclerotic plaques and suggest that 15-LOX-2 may be one of the factors involved in macrophage-mediated LDL oxidation at hypoxia.

Androgen Receptor-Dependent and Independent Atheroprotection by Testosterone in Male Mice

The atheroprotective effect of testosterone is thought to require aromatization of testosterone to estradiol, but no study has adequately addressed the role of the androgen receptor (AR), the major pathway for the physiological effects of testosterone. We used AR knockout (ARKO) mice on apolipoprotein E-deficient background to study the role of the AR in testosterone atheroprotection in male mice. Because ARKO mice are testosterone deficient, we sham operated or orchiectomized (Orx) the mice before puberty, and Orx mice were supplemented with placebo or a physiological testosterone dose. From 8 to 16 wk of age, the mice consumed a high-fat diet. In the aortic root, ARKO mice showed increased atherosclerotic lesion area (+80%, P < 0.05). Compared with placebo, testosterone reduced lesion area both in Orx wild-type (WT) mice (by 50%, P < 0.001) and ARKO mice (by 24%, P < 0.05). However, lesion area was larger in testosterone-supplemented ARKO compared with testosterone-supplemented WT mice (+57%, P < 0.05). In WT mice, testosterone reduced the presence of a necrotic core in the plaque (80% among placebo-treated vs. 12% among testosterone-treated mice; P < 0.05), whereas there was no significant effect in ARKO mice (P = 0.20). In conclusion, ARKO mice on apolipoprotein E-deficient background display accelerated atherosclerosis. Testosterone treatment reduced atherosclerosis in both WT and ARKO mice. However, the effect on lesion area and complexity was more pronounced in WT than in ARKO mice, and lesion area was larger in ARKO mice even after testosterone supplementation. These results are consistent with an AR-dependent as well as an AR-independent component of testosterone atheroprotection in male mice.

Obesity Prevalence in Nepal: Public Health Challenges in a Low-Income Nation during an Alarming Worldwide Trend

The future toll of the obesity epidemic will likely hit hardest in low- and middle-income countries. Ongoing urbanization promotes risk factors including sedentary lifestyle and fat- and sugar-laden diets. Low-income countries like Nepal experience a double disease burden: infectious diseases as well as rising incidence of noncommunicable diseases (e.g., cardiovascular disease and diabetes mellitus) frequently characterized by obesity. Nepal currently directs efforts towards curing disease but pays little attention to preventive actions. This article highlights obesity prevalence in Nepal, delineates the challenges identified by our pilot study (including low health literacy rates), and suggests strategies to overcome this trend.

Cardiovascular health knowledge, attitude and practice/behaviour in an urbanising community of Nepal: a population-based cross-sectional study from Jhaukhel-Duwakot Health Demographic Surveillance Site

Objectives This study determined the knowledge, attitude and practice/behaviour of cardiovascular health in residents of a semiurban community of Nepal. Design To increase the understanding of knowledge, attitude and practice/behaviour towards cardiovascular health, we conducted in-home interviews using a questionnaire based on the WHO STEPwise approach to surveillance and other resources, scoring all responses. We also recorded blood pressure and took anthropometric measurements. Setting Our study was conducted as part of the Heart-Health-Associated Research and Dissemination in the Community project in the Jhaukhel-Duwakot Health Demographic Surveillance Site in two urbanising villages near Kathmandu. Participants The study population included 777 respondents from six randomly selected clusters in both villages. Results Seventy per cent of all participants were women and 26.9% lacked formal education. The burden of cardiovascular risk factors was high; 20.1% were current smokers, 43.3% exhibited low physical activity and 21.6% were hypertensive. Participants showed only poor knowledge of heart disease causes; 29.7% identified hypertension and 11% identified overweight and physical activity as causes, whereas only 2.2% identified high blood sugar as causative. Around 60% of respondents did not know any heart attack symptoms compared with 20% who knew 2–4 symptoms. Median percentage scores for knowledge, attitude and practice/behaviour were 79.3, 74.3 and 48, respectively. Nearly 44% of respondents had insufficient knowledge and less than 20% had highly satisfactory knowledge. Among those with highly satisfactory knowledge, only 14.7% had a highly satisfactory attitude and 19.5% and 13.9% had satisfactory and highly satisfactory practices, respectively. Conclusions Our study demonstrates a gap between cardiovascular health knowledge, attitude and practice/behaviour in a semiurban community in a low-income nation, even among those already affected by cardiovascular disease.

Bacterial cellulose modified with xyloglucan bearing the adhesion peptide RGD promotes endothelial cell adhesion and metabolism—a promising modification for vascular grafts

Today, biomaterials such as polytetrafluorethylene (ePTFE) are used clinically as prosthetic grafts for vascular surgery of large vessels (>5 mm). In small diameter vessels, however, their performance is poor due to early thrombosis. Bacterial‐derived cellulose (BC) is a new promising material as a replacement for blood vessels. This material is highly biocompatible in vivo but shows poor cell adhesion. In the native blood vessel, the endothelium creates a smooth non‐thrombogenic surface. In order to sustain cell adhesion, BC has to be modified. With a novel xyloglucan (XG) glycoconjugate method, it is possible to introduce the cell adhesion peptide RGD (Arg‐Gly‐Asp) onto bacterial cellulose. The advantage of the XG‐technique is that it is an easy one‐step procedure carried out in water and it does not weaken or alter the fiber structure of the hydrogel. In this study, BC was modified with XG and XGRGD to asses primary human vascular endothelial cell adhesion, proliferation, and metabolism as compared with unmodified BC. This XG‐RGD‐modification significantly increased cell adhesion and the metabolism of seeded primary endothelial cells as compared with unmodified BC whereas the proliferation rate was affected only to some extent. The introduction of an RGD‐peptide to the BC surface further resulted in enhanced cell spreading with more pronounced stress fiber formation and mature phenotype. This makes BC together with the XG‐method a promising material for synthetic grafts in vascular surgery and cardiovascular research. Copyright

Physical activity level and its sociodemographic correlates in a peri-urban Nepalese population: a cross-sectional study from the Jhaukhel-Duwakot health demographic surveillance site.

BackgroundPhysical inactivity is a leading risk factor for cardiovascular and other noncommunicable diseases in high-, low- and middle-income countries. Nepal, a low-income country in South Asia, is undergoing an epidemiological transition. Although the reported national prevalence of physical inactivity is relatively low, studies in urban and peri-urban localities have always shown higher prevalence. Therefore, this study aimed to measure physical activity in three domains—work, travel and leisure—in a peri-urban community and assess its variations across different sociodemographic correlates.MethodsAdult participants (n = 640) from six randomly selected wards of the Jhaukhel-Duwakot Health Demographic Surveillance Site (JD-HDSS) near Kathmandu responded to the Global Physical Activity Questionnaire. To determine total physical activity, we calculated the metabolic equivalent of task in minutes/week for each domain and combined the results. Respondents were categorized into high, moderate or low physical activity. We also calculated the odds ratio for low physical activity in various sociodemographic variables and self-reported cardiometabolic states.ResultsThe urbanizing JD-HDSS community showed a high prevalence of low physical activity (43.3%; 95% CI 39.4–47.1). Work-related activity contributed most to total physical activity. Furthermore, women and housewives and older, more educated and self-or government-employed respondents showed a greater prevalence of physical inactivity. Respondents with hypertension, diabetes or overweight/obesity reported less physical activity than individuals without those conditions. Only 5% of respondents identified physical inactivity as a cardiovascular risk factor.ConclusionsOur findings reveal a high burden of physical inactivity in a peri-urban community of Nepal. Improving the level of physical activity involves sensitizing people to its importance through appropriate multi-sector strategies that provide encouragement across all sociodemographic groups.

Establishing a health demographic surveillance site in Bhaktapur district, Nepal: initial experiences and findings.

BackgroundA health demographic surveillance system (HDSS) provides longitudinal data regarding health and demography in countries with coverage error and poor quality data on vital registration systems due to lack of public awareness, inadequate legal basis and limited use of data in health planning. The health system in Nepal, a low-income country, does not focus primarily on health registration, and does not conduct regular health data collection. This study aimed to initiate and establish the first HDSS in Nepal.ResultsWe conducted a baseline survey in Jhaukhel and Duwakot, two villages in Bhaktapur district. The study surveyed 2,712 households comprising a total population of 13,669. The sex ratio in the study area was 101 males per 100 females and the average household size was 5. The crude birth and death rates were 9.7 and 3.9/1,000 population/year, respectively. About 11% of births occurred at home, and we found no mortality in infants and children less than 5 years of age. Various health problems were found commonly and some of them include respiratory problems (41.9%); headache, vertigo and dizziness (16.7%); bone and joint pain (14.4%); gastrointestinal problems (13.9%); heart disease, including hypertension (8.8%); accidents and injuries (2.9%); and diabetes mellitus (2.6%). The prevalence of non-communicable disease (NCD) was 4.3% (95% CI: 3.83; 4.86) among individuals older than 30 years. Age-adjusted odds ratios showed that risk factors, such as sex, ethnic group, occupation and education, associated with NCD.ConclusionOur baseline survey demonstrated that it is possible to collect accurate and reliable data in a village setting in Nepal, and this study successfully established an HDSS site. We determined that both maternal and child health are better in the surveillance site compared to the entire country. Risk factors associated with NCDs dominated morbidity and mortality patterns.

The androgen receptor confers protection against diet-induced atherosclerosis, obesity, and dyslipidemia in female mice

Androgens have important cardiometabolic actions in males, but their metabolic role in females is unclear. To determine the physiologic androgen receptor (AR)‐dependent actions of androgens on atherogenesis in female mice, we generated female AR‐knockout (ARKO) mice on an atherosclerosis‐prone apolipoprotein E (apoE)‐deficient background. After 8 weeks on a high‐fat diet, but not on a normal chow diet, atherosclerosis in aorta was increased in ARKO females (+59% vs. control apoE‐deficient mice with intact AR gene). They also displayed increased body weight (+18%), body fat percentage (+62%), and hepatic triglyceride levels, reduced insulin sensitivity, and a marked atherogenic dyslipidemia (serum cholesterol, +52%). Differences in atherosclerosis, body weight, and lipid levels between ARKO and control mice were abolished in mice that were ovariectomized before puberty, consistent with a protective action of ovarian androgens mediated via the AR. Furthermore, the AR agonist dihydrotestosterone reduced atherosclerosis (‐41%; thoracic aorta), subcutaneous fat mass (‐44%), and cholesterol levels (‐35%) in ovariectomized mice, reduced hepatocyte lipid accumulation in hepatoma cells in vitro, and regulated mRNA expression of hepatic genes pivotal for lipid homeostasis. In conclusion, we demonstrate that the AR protects against diet‐induced atherosclerosis in female mice and propose that this is mediated by modulation of body composition and lipid metabolism.—Fagman, J. B., Wilhelmson, A. S., Motta, B. M., Pirazzi, C., Alexanderson, C., De Gendt, K., Verhoeven, G., Holmäng, A., Anesten, F., Jansson, J.‐O., Levin, M., Borén, J., Ohlsson, C., Krettek, A., Romeo, S., Tivesten, A. The androgen receptor confers protection against diet‐induced atherosclerosis, obesity, and dyslipidemia in female mice. FASEB J. 29, 1540‐1550 (2015). www.fasebj.org

Perceived risks and benefits of cigarette smoking among Nepalese adolescents: a population-based cross-sectional study

BackgroundThe perceived risks and benefits of smoking may play an important role in determining adolescents’ susceptibility to initiating smoking. Our study examined the perceived risks and benefits of smoking among adolescents who demonstrated susceptibility or non susceptibility to smoking initiation.MethodsIn October–November 2011, we conducted a population-based cross-sectional study in Jhaukhel and Duwakot Villages in Nepal. Located in the mid-hills of Bhaktapur District, 13 kilometers east of Kathmandu, Jhaukhel and Duwakot represent the prototypical urbanizing villages that surround Nepal’s major urban centers, where young people have easy access to tobacco products and are influenced by advertising. Jhaukhel and Duwakot had a total population of 13,669, of which 15% were smokers. Trained enumerators used a semi-structured questionnaire to interview 352 randomly selected 14- to 16-year-old adolescents. The enumerators asked the adolescents to estimate their likelihood (0%–100%) of experiencing various smoking-related risks and benefits in a hypothetical scenario.ResultsPrincipal component analysis extracted four perceived risk and benefit components, excluding addiction risk: (i) physical risk I (lung cancer, heart disease, wrinkles, bad colds); (ii) physical risk II (bad cough, bad breath, trouble breathing); (iii) social risk (getting into trouble, smelling like an ashtray); and (iv) social benefit (looking cool, feeling relaxed, becoming popular, and feeling grown-up). The adjusted odds ratio of susceptibility increased 1.20-fold with each increased quartile in perception of physical Risk I. Susceptibility to smoking was 0.27- and 0.90-fold less among adolescents who provided the highest estimates of physical Risk II and social risk, respectively. Similarly, susceptibility was 2.16-fold greater among adolescents who provided the highest estimates of addiction risk. Physical risk I, addiction risk, and social benefits of cigarette smoking related positively, and physical risk II and social risk related negatively, with susceptibility to smoking.ConclusionTo discourage or prevent adolescents from initiating smoking, future intervention programs should focus on communicating not only the health risks but also the social and addiction risks as well as counteract the social benefits of smoking.

Quantitation of Platelet-Derived Growth Factor Receptors in Human Arterial Smooth Muscle Cells In Vitro

Platelet-derived growth factor (PDGF) is suggested to play an important role in the development of atherosclerosis as a migratory and mitogenic stimulus to arterial smooth muscle cells (ASMCs). Stimulated and unstimulated ASMCs were studied with respect to PDGF receptor (PDGF-R) mRNA and protein expression. Quantitative RT-PCR was developed for simultaneous evaluation of both PDGF-R alpha and -R beta mRNA expression and a quantitative ELISA for estimation of corresponding PDGF-R subunits. On the mRNA level, the overall PDGF-R beta expression was approximately 100 times lower than that of PDGF-R alpha. Furthermore, although PDGF-R alpha mRNA levels were high irrespective of hASMC phenotype, PDGF-R beta mRNA was influenced by serum stimulation with lower copy numbers in proliferating and confluent cells compared with quiescent cells. On the protein level, quiescent hASMCs expressed 10 times more PDGF-R beta than PDGF-R alpha. Serum stimulation decreased cell surface PDGF-Rs, with most prominent loss of PDGF-R alpha (ELISA and immunohistochemistry). Our results suggest a differential regulatory pattern for PDGF-R alpha and -R beta and are compatible with the usage of alternative promoters for regulation of -R alpha expression. Further, it seems that the number of available receptor subunits is not the only determinant of variations in cell stimulation with different PDGF isoforms.