Metabonomics Characteristics Of Hyperuricemia And Common TCM Constitution Type In Middle-aged And Old People

BackgroundHyperuricemia is a disease characterized by significantly higher uric acid (UA) level in the blood of patients. Recent epidemiological studies have suggested that Hyperuricemia may be an independent risk factor for kidney disease. Beyond inducing kidney disease, Hyperuricemia is closely related with the occurrence and growth of gout, coronary heart disease, myocardial infarction and cerebral infarction, and has become a significant factor of metabolic syndrome. With the development of economy and the change of living style, the prevalence of senile Hyperuricemia shows an increasing tendency. A2011study showed that the prevalence rate of Hyperuricemia was9.8%in Beijing community elderly, and the prevalence rate in females (9.9%) is significantly higher than that in males (9.6%). However, Health-Examination results of middle-aged and elderly people in2012showed that prevalence rate of Hyperuricemia was23.45%, and the prevalence rate in males is higher than that in females by nearly2-to-1. Recent foreign studies reported that prevalence rate of Hyperuricemia in the elderly has reached50%. Above all, Hyperuricemia in the elderly population has become a major public health problem.The phenomenon of constitution is an important manifestation of life activity in human being and it is closely related to disease and health. Traditional Chinese Medical (TCM) Constitution guided by traditional Chinese medicine theory is used to study various physical characteristics, physiological and pathological features, as well as analyze disease state, the nature of the lesion and development trends. Many scholars have studied the TCM constitution of patients with Hyperuricemia. Results of one study on123cases of patients with Hyperuricemia constitutional analysis showed that 104cases were qi deficiency and phlegm constitution,10cases were damp-heat constitution, and9cases were blood stasis constitution. Accounting for TCM constitution type,26patients with gout were classified. Results showed17cases with physical phlegm constitution,4cases with mildly constitution,3cases with mildly and dampness constitution,1case with yang deficiency,1case with Qi deficiency, revealing that the incidence of gout has the certain relations with TCM constitution. Studies on the TCM constitution of patients with Hyperuricemia in Xinjiang have found that the common types are phlegm-dampness constitution, damp-heat constitution and blood stasis. Studies have reported that patients with gout are mainly middle-aged men and post-menopausal women, and most patients are characterized by Phlegm-dampness constitution manifesting as body fat and food addiction, therefore the conclusion can be made that constitution is associated with gout.Metabolomics is a rapidly developing, aiming at small molecule metabolites science. Being an essential component of systematic biology, metabolomics is a post genomic technology aimed at seeking to provide a comprehensive profile of all the metabolites present in a biological sample in a particular physiological period, such as organism, cells, blood or body fluids. Results of some studies on combining Hyperuricemia and metabolomics showed that abnormal metabolism of glucose and lipoid were found in patients with Hyperuricemia (HUA), these metabolites change may be associated with the pathogenesis, progress, and prognosis of HUA. Uric acid is closely related to the metabolic levels of blood glucose. Although the material metabolism paths of triglyceride (TG) are not the same to those of Uric acid (UA), the degradation process of triglyceride generated free fatty acid and esterification into other organizations will increase the utilization rate of adenosine triphosphate (ATP), accelerate ATP decomposition and increased production of uric acid.Therefore, combining metabolomics and TCM constitution of middle-aged and aged patients with Hyperuricemia is an important method to study prevalence and pathogenesis of Hyperuricemia in middle-aged and aged population, and it will provide theoretical basis for clinical treatment. Research on this subject has broad prospects and practical significance.ObjectiveThe objectives of this study:①To explore the incidence of Hyperuricemia in middle-aged and aged population and the impact of gender, age;②To explore TCM constitution of middle-aged and aged patients with Hyperuricemia and the impact of gender, age;③To analyze the metabolite composition of samples and find out metabolic markers of common TCM constitution from middle-aged and aged patients with Hyperuricemia using liquid chromatography-mass spectrometry technology, it will provide theoretical basis for the study of pathogenesis.MethodsIn the clinical studies, a cross-section epidemiological investigation was carried out on community residents in Guangzhou. Correct differentiation of constitutional pattern referring to the constitution of TCM criterion of2009, and drew blood for analysis. All the subjects were interviewed with a questionnaire by specially trained professional person in a standardized way. Calculate sample size, and the guideline of choosing and excluding case was established. The mildly constitution is considered normal, while other8kinds of constitution were unhealthy constitution. All original data were examined by professionals, and unqualified questionnaire should be eliminated. Epidata3.1software was used to build data record system; and the raw data was placed into the system by two operators. Statistical analysis was accomplished with SPSS17.0soft package. According to the research needs, we carried out descriptive analysis of the distribution feature, and then analyzed the characteristic variables such as age, gender and so on; Chi-square test were applied when more than one objective indicators discussed in various differences between groups were significant; We intend to adopt unconditioned Logistic regression analysis to explore the internal distribution and characteristics, while we didn’t do it as the incomplete data collection. In experimental aspect, patients were grouped according to the results of investigation:①Hyperuricemia group;②Control group;③Hyperuricemia with mildly constitution group;④Hyperuricemia with qi-insufficiency constitution group. Hyperuricemia group and Hyperuricemia with qi-insufficiency constitution group have no significant differences compared with control group and Hyperuricemia with mildly constitution group, respectively. Collect, classify and preserve the serum at-80degrees and make a clear sign of everyone’s information. Freeze thawing at4degrees overnight.400μL methanol is added into the serum (per100μL), then the solutions should be vortexes for4minutes and centrifuge at4degrees (15min,13000rpm/min), thus supernatant is obtained and put into in96-well plates for chemical analysis. HPLC/MS were used to do the metabolomics experiment, compounds with significant difference between groups(P<0.05) were screened out according to VIP (Variable Importance in the Projection) value(≧1) got from PLS-DA analysis combined with t test. Then identify potential biomarkers according to formula searchable database,(http://www.genome.jp/kegg/ligand.html and http://redpoll. pharmacy.ualberta. ca/～aguo/www_hmdb_ca/HMDB/)ResultsIn the clinical research, the study involved463participants, including235patients with Hyperuricemia(172females and63males) and228controls; The prevalence of Hyperuricemia is50.76%and significantly higher in women(53.58%) than in men (44.37%); The prevalence of Hyperuricemia in the age groups of55-64years,65-74years and75-84years were53.40%,48.51%and52.80%, respectively; And in the age groups of55-69years,70-84years were50%and51.54%, respectively. People with mildly constitution account for30.67%among elderly people; subjects with normouricemia (32.46%) were significantly higher than those with Hyperuricemia (28.81%). Common TCM Constitutions among elderly people with Hyperuricemia were mildly constitution, qi-insufficiency constitution and yang-insufficiency constitution, accounting for28.81%,9.7%and7.63%, respectively. In experimental aspect, the metabolic markers of Hyperuricemia included: acylglycerol phospholipid; hemolytic lecithin; lecithin; triglycerides; diglyceride; acyldehydrogenase; Uridine-5’-diphosphoglucose; acetyl CoA; pentosan; stearoyl-CoA; dehydrocholesterol; homocysteine; phosphatidylserine; dihydroxyacetone phosphate and acyl monoethanolamide. The metabolic markers of Hyperuricemia with qi-insufficiency constitution included:acyl glycine, cardiolipin, glycerine phosphatide, glycosphingolipids, phospholipids, inositol phospholipids, ceramide, lysophosphatide acid, sulfate, DHT dihydroxy acetone phosphate, lysolecithin, pure ammonia acyl alanine, leukotriene, aspartic acid, etc. Common metabolic markers of Hyperuricemia and Hyperuricemia with qi-insufficiency constitution included:Glycerine phosphatide, hemolytic lecithin and dihydroxyacetone phosphate.Conclusion In the clinical research, the prevalence of Hyperuricemia is significantly higher in women than in men among middle aged and elderly people; when stratified by age, different stratified sampling method obtained different results. If we classified subject participants into three categories (55-64years,65-74years and75-84years), prevalence of Hyperuricemia were descend by the increase of age; however, when we classified subject participants into two categories, prevalence of Hyperuricemia increased with age. The proportion of participants with Hyperuricemia is higher than those with normouricemia in the middle-aged and old women, while the proportion of participants with Hyperuricemia is lower than those with normouricemia in the middle-aged and old men. For the participants with mildly constitution, the proportion of participants with Hyperuricemia is higher than those with normouricemia. Sorting by TCM Constitution:mildly constitution>qi-insufficiency constitution>yang-insufficiency.In the experimental aspect, Middle aged and elderly people with Hyperuricemia differ in several aspects, such as lipid metabolism, glucose metabolism and energy metabolism. And there were difference in lipid, protein, sugar, energy and steroid metabolism for patients with qi-insufficiency constitution. Lipid and glucose metabolism were common metabolic markers for Hyperuricemia and Hyperuricemia with qi-insufficiency constitution.There are some shortcuts in this thesis. In the epidemiological cross-sectional investigation, limitations of this study include:small sample size, although it is greater than the minimum requirement; Low repossessions rate of constitution questionnaire, although it is greater than the minimum requirement (71%); inadequate data of middl—aged and elderly men. Therefore, further research on epidemiological cross-sectional investigation is necessary. In the experimental aspect, although we has carried on the statistical analysis of the metabonomics markers content, the accurate molecular structure of these metabolic marker also need to be further studied and confirmed. All in all, the setting up of correlative experiment analysis made good foundation for deeper research work.