Novel biological detection method for dioxins in serum, and a diagnostic use therefor in metabolic syndrome and related conditions

Abstract

The present invention relates to a novel biological detection method for dioxins in serum and a diagnostic use thereof in metabolic syndrome and related conditions. More particularly, according to the present invention, a significant correlation between dioxin content in serum and physical variables has been confirmed by measuring luciferase activity through the use of serum as a whole and cells transformed with a recombinant vector comprising a gene construct in which a dioxin responsive element, a promoter and a reporter gene are operably linked. And there is an improvement in this method of the present invention over the conventional methods which require a preprocessing step in order to purify dioxins from serum, by which plural samples can be analyzed easily and accurately even with a very small amount of each because of the use of serum as a whole. Therefore, the detection system of the present invention which involves total serum processing in a genetically transformed cell line comprising a recombinant reporter gene whose expression is modified by dioxin compounds can be effectively used for the biological detection of dioxins in serum. This method can also be used to research the correlation between specific POPs and patient's disease factors by accurate detection of POPs such as dioxins in the serum, and further this method can be effectively used to predict the occurrence of disease and to determine treatability.

What is claimed is: 
     
         1 . A detection method for a dioxin compound in serum comprising the following steps:
 i) preparing a transformed cell line by introducing a recombinant vector comprising a gene construct, in which at least one of dioxin-responsive elements (DRE) represented by SEQ. ID. NO: 1, a promoter, and a reporter gene are operably linked, into a host cell;   ii) culturing the transformed cell line prepared in step i) after treating the cell line with the serum of a test subject;   iii) detecting the expression of the protein expressed by the reporter gene in the transformed cell line cultured in step ii); and   iv) determining the serum to have a dioxin compound when the reporter gene expression is detected in step iii).   
     
     
         2 . A method of increasing the detection efficiency for a dioxin compound in serum comprising the following steps:
 i) obtaining a serum sample by heat-inactivating total serum obtained from a test subject;   ii) preparing a transformed cell line by introducing a recombinant vector comprising a gene construct, in which at least one dioxin-responsive element (DRE) represented by SEQ. ID. NO: 1 is operably linked to a promoter and a reporter gene, into a host cell;   iii) culturing the transformed cell line prepared in step ii) with the serum sample obtained in step i);   iv) detecting the activation of the reporter gene in the transformed cell line cultured in step iii); and   v) determining that the dioxin compound is comprised in the serum sample when the activation of the reporter gene is detected in step iv).   
     
     
         3 . The method according to  claim 2 , wherein the detection efficiency is determined based on a small amount of serum. 
     
     
         4 . The method of  claim 3 , wherein the small amount of serum is 1-10 μl. 
     
     
         5 . The method of  claim 2 , wherein the dioxin compound is selected from the group consisting of polychlorinated dibenzodioxins (PCDDs), polychlorinated dibenzo-furans (PCDFs), polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarcons (PAHs), flavonoids, and pesticides. 
     
     
         6 . The method of  claim 2 , wherein between about 3-4 of the dioxin-responsive elements of step i) are included in the gene construct. 
     
     
         7 . The method of  claim 2 , wherein the promoter of step ii) is selected from the group consisting of MMTV (Mouse Mammary Tumor Virus) promoter, SV40 promoter, and CMV (cytomegalovirus) promoter. 
     
     
         8 . The method of  claim 2 , wherein the reporter gene of step ii) is selected from the group consisting of luciferase, alkaline phosphatase, chloramphenicol acetyl transferase, and β-galactosidase. 
     
     
         9 . The method of  claim 2 , wherein the host cell of step ii) is a mammalian tumor cell line. 
     
     
         10 . The method of  claim 9 , wherein the host cell is a mouse hepatocarcinoma cell line. 
     
     
         11 . The method of  claim 2 , wherein diabetes or metabolic syndrome is predicted when the serum sample is the serum of a test subject suspected to have diabetes or metabolic syndrome. 
     
     
         12 . The method of  claim 2 , wherein a prognosis of diabetes or metabolic syndrome is monitored when the serum sample is the serum of a test subject suspected to have diabetes or metabolic syndrome.