Low Selenium and Low Protein Exacerbate Myocardial Damage in Keshan Disease by Affecting the PINK1/Parkin-mediated Mitochondrial Autophagy Pathway

Objective Keshan disease(KD)is a myocardial mitochondrial disease closely related to insufficient selenium(Se)and protein intake.PTEN induced putative kinase 1(PINK1)/Parkin mediated mitochondrial autophagy regulates various physiological and pathological processes in the body.This study aimed to elucidate the relationship between PINK1/Parkin-regulated mitochondrial autophagy and KD-related myocardial injury.Methods A low Se and low protein animal model was established.One hundred Wistar rats were randomly divided into 5 groups(control group,low Se group,low protein group,low Se+low protein group,and corn from KD area group).The JC-1 method was used to detect the mitochondrial membrane potential(MMP).ELISA was used to detect serum creatine kinase MB(CK-MB),cardiac troponin I(cTnI),and mitochondrial-glutamicoxalacetic transaminase(M-GOT)levels.RT-PCR and Western blot analysis were used to detect the expression of PINK1,Parkin,sequestome 1(P62),and microtubule-associated proteins1A/1B light chain 3B(MAP1LC3B).Results The MMP was significantly decreased and the activity of CK-MB,cTnI,and M-GOT significantly increased in each experimental group(low Se group,low protein group,low Se+low protein group and corn from KD area group)compared with the control group(P<0.05 for all).The mRNA and protein expression levels of PINK1,Parkin and MAP1LC3B were profoundly increased,and those of P62 markedly decreased in the experimental groups compared with the control group(P<0.05 for all).Conclusion Low Se and low protein levels exacerbate myocardial damage in KD by affecting the PINK1/Parkin-mediated mitochondrial autophagy pathway.

Relationship Between Myocardial Injury and Expression of PGC-1α and Its Coactivators in Chronic Keshan Disease

Objective:Keshan disease(KD)is a mitochondrial cardiomyopathy.The present study explored the roles of peroxisome proliferator-activated receptor(PPAR)-y coactivator-la(PGC-la),the key regulator of mitochondrial structure and function,and its coactivators in myocardial injury in chronic KD.Furthermore,the usefulness of these molecules in the diagnosis of chronic KD was assessed.

Circulating Adhesion Molecules in Patients with Keshan Disease and Their Relationship with Coxsackie B Virus Infection

This study determined the levels of serum soluble intercellular adhesion molecule-1 （sI-CAM-l） and soluble vascular cell adhesion molecular-1 （sVCAM-1） in patients with different types of Keshan disease （KD）, examined the relationship between Coxsackie B virus-specific IgM antibody （CBV-IgM） and slCAM-1 or sVCAM-1 in KD patients, and investigated the role of these adhesion molecules in the pathogenesis of KD and their clinical implications. The levels of serum slCAM-1, sVCAM-1 and CBV-IgM were measured by using enzyme-linked immunosorbent assay in 22 patients with chronic Keshan disease （CKD）, 27 with latent Keshan disease （LKD） and 28 healthy controis. The subjects in different groups were adjusted for sex and age. Echocardiography was adopted to determine left ventricular ejection fraction （LVEF） in 22 patients with CKD. The results showed that CKD patients had significantly higher levels of slCAM-1 and sVCAM-1 than LKD patients and healthy controls （P〈0.01 for all）. And there was significant difference in the levels of the 2 adhesion molecules between LKD patients and healthy controls （P〈0.05）. A negative correlation was found between LVEF and slCAM-1 or sVCAM-1 in CKD patients. The percentage of CBV-specific IgM positive individuals in KD patients was significantly higher than that of healthy controls. In CVB-specific IgM positive patients, the levels of serum slCAM-1 and sVCAM-1 were significantly greater than those in CBV-specific IgM negative counterpart. It was concluded that the increase in the levels of slCAM-1 and sVCAM-1 suggests the progression of inflammation in KD. slCAM-1 and sVCAM-1 can promote the development of myocardial pathology and lead to poor myocardial function. The increased serum slCAM-1 and sVCAM-1 in KD patients may be related to CBV infection.

THE STUDIES OF SERUM IL-6,INF-αAMONG CHRONIC KESHAN DISEASE PATIENTS

Objective To investigate the function of cytokine in the patients suffered from chronic Keshan disease. Methods Serum IL 6, INF α was tested among 55 patients of chronic Keshan disease and 30 health controls by the methods of ELISA, read in the unit of pg·mL -1 . Results The cytokine was significantly higher in Keshan disease patients than that in health control group. Conclusion Serum levels of IL 6, TNF α is higher in chronic Keshan disease patients than in health control group .It was showed that cytokine plays an important role during the etiopathology and the development in Chronic Keshan disease.

Serum level of galectin-3 and its clinical significance in patients with chronic Keshan disease

Keshan disease （KD） is a fatal endemic dilated cardiomyopathy with unclear etiology and pathogenesis, and a high mortality in China. Pathologic studies confna＇ned that different degree of myocardial fibrosis existed in various types of KD. Myocardial fibrosis is an important contributor to the pathophysiology of left ventricular remodeling. Recently, galectin-3 （Gal-3） as a marker of cardiac fibrosis and heart failure was approved by the US FDA. The aim of this study was to evaluate the changes of serum level of Gal-3 in chronic KD （CKD） mad their clinical implications. Methods： The levels of serum Gal-3 were measured by using enzyme-linked immunosorbent assay （ELISA） in 37 CKD patients and 32 healthy controls. Echocardiography was used to determine parameter of left ventricular ejection fraction （LVEF） and left ventricular end-diastolic dimension （LVEDD）. Results： The serum concentration of Gal-3 （[95.81：i：18.99] ng/ml versus [48.16-＋11.09] ng/ml, t=6,906, P〈0.001） and LVEDD （[60.46＋7.63] mm versus （42.69^-10.66） ram, t=3.61, P〈0.01） were significantly higher, while LVEF （[42.69-J：10.661% versus [62.16~6.381%, t=4.679, P〈0.01） were significantly lower in CKD patients compared with healthy controls. A negativecorrelation was found between elevated Gal-3 and lower LVEF （r=--0.882, P〈0.001） and a positive correlation was found between elevated Gal-3 levels and enlarged LVEDD （r=0.834, P〈0.001） or higher NYHA class （r=0.854, P〈0.01） in CKD patients. Conclusion： Serum concentration of Gal-3 is strongly correlated with poorer left ventricular systolic function, enlarged LVEDD and higher New York Heart Association （NYHA） class, which may provide indirectly diagnostic information on myocardial fibrosis and heart function in CKD patients

Further Investigation on the Role of Selenium Deficiency in the Aetiology and Pathogenesis of Keshan Disease

Selenium supplements were not able to restore the ultrastructural changes in the myocardiurn of latent Keshan disease patients taken by using cardiac catheter endomyocardial biopsy. Observations on the changes of seleniurn status and the incidence of Keshan disease showed that new latent and naturally-occurring chronic cases were found in the endemic area even after selenium levels had been elevated in the residents to the levels typical in the non-endemic area. These results indicate that although selenium deficiency might be a primary pathogenetic geogen in the occurrence of Keshan disease, it is rather a conditional predisposing factor than a specific or initiative aetiologic factor for the occurrence of Keshan disease. Selenium supplmentation could apparently alleviate the higher platelet responsiveness of residents in the endemic area, which might contribute to eliminating the basis for the occurrence of the multifocal perivascular necroses in myocardium of acute and subacute Keshan disease

THE STUDIES OF SERUM CVB-SPECIFIC IgM ANTIBODY AND NITRIC OXIDE AMONG THE PATIENTS WITH LATENT AND CHRONIC KESHAN DISEASE

Objective To investigate the serum coxsackie virus B(CVB) infection and nitric oxide (NO)level of the patients suffer from latent or chronic Keshan disease and their characteristics in the etiopathology of Keshan disease. Methods Sera were isolated from 30 patients with latent or chronic Keshan disease in Huangling county. Shaanxi Province, and the CVB-specific IgM antibody and NO were tested. Control groups were health subjects in Huangling county or Xi’an city, Shaanxi Province. Results The percentage of CVB-specific IgM positive in patients in Huangling county was significantly higher than that of both control groups in Huangling county and Xi’an city (P< 0.05). The serum level of NO In patients was significantly higher than that of the control group in Huangling county (P<0.05).however,compared with control group in Xi’an city. there was no difference (P>0.05). In CVB-specific IgM positive patients,the serum level of NO was significantly higher than that of CVB-specific 1gM negative group (P <0.05). Conclusion CVB infection and serum NO level might be related to the etiopathology and the development of Keshan disease.

Genome-Wide Study Identifies the Regulatory Glycosyltransferase Genes Networks and Signaling Pathways from Keshan Disease

KD （Keshan disease） is an endemic cardiomyopathy occurring only in China. Its pathogenesis is unclear till now. In the study, gene expression profiles of the PBMC （peripheral blood mononuclear cell） derived respectively from KD patients and healthy in KD areas were compared. Total RNA was isolated, amplified, labeled and hybridized to Agilent 4 ~ 44 K Whole Human Genome Oligonucleotide Microarray. Significant canonical pathways were analyzed by IPA （ingenuity pathway analysis） to identify differently expressed genes and pathways involved in the cardiovascular system development and function. Quantitative RT-PCR was applied to further validate our microarray results. Eighty-three up-regulated （ratios 〉 2.0） and nine down-regulated glycosyltransferase genes （ratios 〈 0.5） in PBMC in KD patients were detected by significance analysis of microarrays. Two significant canonical pathways from glycosyltransferase gene expression profiles were screened by IPA. The results of qRT-PCR show that four up-regulated （BMP 1/7/10 and FGF 18） and one down-regulated （BMP2） genes are consistent with those in microarray experiment, confirming the validity of the microarray data. Based on the results of the study, it is suggested that bone morphogenetic proteins and fibroblast growth factors might play an important role in the pathogenesis of KD. This further helps us to understand the pathogenesis of KD, as well as dilated cardiomyopathy.

Research on Autoantibodies Against Myocardialβ_1-adrenergic and M_2 Cholinergic Receptors in Patients With Chronic Keshan Disease

Objectives To explore the relationship between serum autoantibodies against myocardial β1-adrenergic, M2-cholinergic receptors and chronic Keshan disease （CKD）. Methods The second extracellular loops of β1 and M2 receptors on human cardiomyocytes were used as the antigens. Enzyme linked immunosorbent assay （ELISA） was applied to determine serum autoantibodies against myocardial β1 and ME receptors in 32 CKD patients. 31 healthy subjects from endemic area were selected as the control. Results Positive rate of autoantibodies against myocardial β1 adrenergic （51.3%, 17/32） and M2 cholinergic （56.3% , 18/32） receptors were significantly higher than those in the control （9.7%, 3/ 31; 12.9%, 4/31） （both P〈 0.01）. Both positive rate and titers of above autoantibodies in NYHA Ⅱ - Ⅲ CKD patients were significantly higher than those in NYHA Ⅳ, demonstrating an apparently positive correlation between serum antibodies against myocardial β1 and M2 receptors （r=0.95）. Conclusions Autoantibodies against myocardial β1 and M2 receptors were found in sera of CKD patients; distribution of positive rate and titers of the autoantibodies in CKD patients in various NYHA are significantly different. classes of cardiac function

The Level of Serum Intact Terminal Peptide of Procollagen and Its Clinical Significance in Patients with Chronic Keshan Disease

Objectives To evaluate the changes of serum intact terminal peptide of procollagen in patients with chronic Keshan disease (KD) and investigate their clinical significance. Methods The concentrations of serum intact N-terminal peptide of type procollagen (P NP) and intact N-terminal peptide of type procollagen were measured by radioimmunoassay in 35 patients with chronic KD and 31 normal control. Doppler ultrasounds was used to determine several parameters of left ventricular systole and diastole functions. Results The concentration of serum P NP (74.07±16.74)μg/L and the ratio of P NP/ P NP (18.02 ±4.60) in chronic KD were significantly increased as compared to the control (39.63±12.07 μg/L, 12.12±4.24; P< 0.001). Serum P NP (4.19±0.64)μg/L in chronic KD was higher than that in the control (3.36±0.65 μg/L,P < 0.001) too. The higher of serum concentration of P NP and the ratio of P NP/ P NP, the worse of cardiac function in patients with chronic KD. A negative correlation was found between serum P NP/ P NP, P NP and VE/VA, LVEF (γ=-0.4502, -0.4608, P< 0.01 and γ=-0.3936, -0.3904, respectively; P<0.05). Conclusions These findings suggested that tissue synthesis of collagen type and type was abnormally increased in chronic KD. On the other hand, our results indicated that P NP and P NP were related to several functional alterations of the left ventricle. Serum procollagen peptide measurements might therefore provide indirectly diagnostic information on myocardial fibrosis associated with chronic KD.

Comparing gene expression profiles of Kashin-Beck and Keshan diseases occurring within the same endemic areas of China

In this study, differentially expressed genes in peripheral blood from patients with Kashin-Beck disease and Keshan disease were compared to further investigate the etiology and pathogenesis of both diseases, which occur in a common endemic area of China. Twenty Kashin-Beck disease patients and 12 healthy controls, and 16 Keshan disease patients and 16 healthy controls, were grouped into four pairs. Patients and controls were selected from common endemic areas for the two diseases. Total RNA was isolated from peripheral blood mononuclear cells from all patients and controls, and gene expression profiles analyzed by oligonucleotide microarrays. Sixteen genes differentially expressed in both Kashin-Beck disease and Keshan disease (versus controls) were identified, and comprised nine genes showing synchronous and seven asynchronous expression. The Comparative Toxicogenomics Database shows that expression and biological function of these genes can be affected by multiple environmental factors, including mycotoxin and selenium content, potential environmental risk factors for the two diseases. Thus, these shared differentially expressed genes may contribute to the distinct organ lesions, caused by common environmental risk factors of Kashin-Beck disease and Keshan disease.

Geographical differentiation of selenium concentration in hair from children and youngsters in China

Selenium concentration in hair are sorted out into three classes, i.e. low (<110 ppb), medium (110-520 ppb) and high (>520 ppb) according to the principle of multiple normal distribution and a map of classification of selenium concentration in hair from children and youngsters in China has been drawn. Selenium insufficiency in relation to Keshan disease and Kashin-Beck's disease is expounded. Geographical differentiation of hair selenium concentrations and its causes are explained.

Detoxifying moniliformin in grains and water

The method to determine moniliformin content was established in our laboratory. The recovery rate is 97 7% while the moniliformin content in the sample is 8 8 mg/g. In comparison with different methods of detoxifying moniliformin in water, the best antidote is chlorinated lime. 1 5 mg active chlorine in chlorinated lime was required to detoxify 1 mg moniliformin. 5% H 2O 2 spray was the best way for detoxifying moniliformin in grains. These two methods are convenient, economic and with no secondary pollution. They can be used for preventing human and livestock from the toxicity of moniliformin.