ABSTRACT
This study examined the effect of insulin on the expression of very low density lipoprotein receptor (VLDLR) subtypes of SGC7901 cells and discussed its biological implication. In vitro,moderately or poorly-differentiated human gastric adenocarcinoma cell line SGC7901 was incubated with insulin for different lengths of time, and then the expression of protein and RNA level in VLDLR subtypes were detected by Western blotting and real-time PCR, respectively. The results showed that, at certain time interval, insulin could down-regulate expression of type Ⅰ VLDLR and up-regulate the expression of type Ⅱ VLDLR in SGC7901 cells, at both protein and RNA level.We are led to conclude that insulin serves as a regulator in maintaining the balance between glucose and lipid metabolism in vivo, possibly through its effect on the differential expression of VLDLR subtypes.
ABSTRACT
Se estudió la regulación del receptor de lipoproteína de muy baja densidad (VLDLR) en dos tipos de células que intervienen en la respuesta inflamatoria: hepatocitos (HepG2) y células linfocitarias de bazo de ratón (CMB) por agentes implicados en la inflamación: VLDL o Gemfibrozil, lipopolisacárido bacteriano (LPS) para HepG2 o Concanavalina A (ConA) para CMB. Se determinaron por citometría de flujo las subpoblaciones de CMB y de HepG2 en distintas fases del ciclo celular (G1 y G2/M) con ioduro de propidio y las VLDLR+ con VLDL fluorescente. Entre 10 a 60 por ciento de células expresaron VLDLR dependiendo de las condiciones experimentales. El enfrentamiento con LPS o ConA produjo un aumento de células VLDLR+. El tratamiento con Gemfibrozil disminuyó el número de hepatocitos en reposo VLDLR+ pero incrementó significativamente (más de dos veces) los hepatocitos VLDLR+ en fase G2/M. En los cultivos de CMB Gemfibrozil aumentó por igual el porcentaje de células VLDLR+ quiescentes y en G2/M. El comportamiento de estos tipos celulares con VLDL fue distinto: CMB no mostró cambios en las subpoblaciones VLDLR+, los hepatocitos mostraron disminución de VLDLR+ tanto en fase G1 como en fase G2/M. Se concluyó que el estudio de la regulación de VLDLR en distintas células facilitará el diseño de fármacos específicos para el tratamiento de enfermedades de etiología inflamatoria.
The regulation of the VLDL receptor (VLDLR) by molecules involved in the inflammation process (lipopolysaccharide, LPS; Concanavalin A, ConA, VLDL or Gemfibrozil) in two cellular types implied in the inflammatory response, hepatocytes (HepG2) and lymphocitary cells from mice spleen (CMB), was studied. Different subpopulations of CMB and HepG2 in different cell cycle phases (G1 and G2/M) were analyzed using flow citometry techniques with propidium iodide, and the VLDLR+ with fluorescent VLDL. In cultures of both cell types, it was observed that 10 to 60% of the cells expressed the VLDLR (VLDLR+ cells) indistinctly of the culture conditions. VLDLR+ cells belonged equally to cells in the quiescent and in the synthesis or mitosis phase of the cell cycle. Challenging them with LPS or ConA an increase in the percentage of VLDLR+ cells was produced. Gemfibrozil treatment decreased the number of resting hepatocytes VLDLR+ but increased significantly (more than twice) the number of hepatocytes VLDLR+ in phase G2/M. In hepatocytes there was almost the same proportion of VLDLR+ cells that were in the G1 or in the G2/M phase of the cell cycle. It is concluded that the study of VLDLR regulation will facilitate the design of new drugs to treat inflammatory diseases.
Subject(s)
Gemfibrozil , Inflammation , LipoproteinsABSTRACT
Summary: To explore the functions of very low density lipoprotein receptor (VLDL-R) subtype II in lipoprotein metabolism and foam cells formation, the recombinant plasmid with the two subtypes cDNA was constructed respectively, the ldl-A7 cell lines were transfected and two cell lines expressing VLDL-R were obtained: one stably expressing the VLDLR with the O-linked sugar region (type I VLDLR) and the other without the O-linked sugar region (type II VLDLR). In the study on binding of VLDLR to their nuclein labeled natural ligands (VLDL and β-VLDL), it was found that surface binding of 125I-VLDL or 125I-β-VLDL of ldl-A7 cells transfected with type I VLDLR recombinant (ldl-A7-VRI) was more higher than that of ldl-A7 cells transfected with type II VLDLR recombinant (ldl-A7-VRII). After being incubated with VLDL for different time, the contents of triglyceride and total cholesterol in cells were mensurated, and the formation of foam cells and accumulation of lipid in cells was observed by oil-red O staining. The results showed that the contents of triglyceride and total cholesterol in ldl-A7-VR I were much higher than those in ldl-A7-VR II, and ldl-A7-VR I could transform into foam cells notably. It was suggested that type I VLDLR binds with relative higher affinity to VLDL and β-VLDL, and internalizes much more lipoprotein into cells. As a result, we can conclude that type I VLDLR plays a more important role in lipoprotein metabolism and foam cells formation than type II VLDLR.