RESUMEN
The effect on murine immunoglobulin G (IgG) glycosylation of altering IgG production in vivo was assessed in interleukin (IL)-6 transgenic and CD4 knockout mice. C57BL/6 mice carrying the IL-6 transgene showed increased levels of circulating IgG. This was associated with decreased levels of galactose on the IgG oligosaccharides. No decrease in beta4-galactosyltransferase mRNA or in enzyme activity was seen in IL-6 transgenic mice. MRL-lpr/lpr mice normally have elevated levels of circulating IgG, again accompanied by decreased levels of IgG galactose. Disruption of the CD4 gene in MRL-lpr/lpr mice led to a substantial decrease in the concentration of circulating IgG, but IgG galactose levels remained low. Thus, an enforced decrease in IgG levels in the lymphoproliferative MRL-lpr/lpr mice did not alter the percentage of agalactosyl IgG in these mice, suggesting that agalactosyl IgG production is not simply caused by excessive IgG synthesis leading to an insufficient transit time in the trans-Golgi, but rather to a molecular defect in the interaction between galactosyltransferase and the immunoglobulin heavy chain.
Asunto(s)
Antígenos CD4/genética , Inmunoglobulina G/metabolismo , Interleucina-6/genética , Linfocitos/metabolismo , Animales , Galactosa/metabolismo , Galactosiltransferasas/genética , Galactosiltransferasas/metabolismo , Expresión Génica , Glicosilación , Inmunoglobulina G/sangre , Linfocitos/enzimología , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Ratones Transgénicos , Bazo/enzimología , Bazo/inmunologíaRESUMEN
The authors used murine pregnancy as a model to investigate the regulation of IgG glycosylation. Pregnancy is associated with decreased levels of circulating IgG. The oligosaccharides on this IgG from late (day 15), but not early (day 8), pregnant Balb/c mice exhibited increased levels of terminal galactose. The levels remained elevated 8 days post-partum in lactating mice. Nonetheless, splenic beta 1, 4-GalTase mRNA and enzyme activity remained relatively constant throughout pregnancy and into lactation. This was in contrast to a pregnancy-associated increase in mammary gland beta 1,4-GalTase mRNA. Thus the increased IgG galactose levels seen in pregnancy are regulated by mechanisms which are independent of transcriptional control of beta 1,4-GalTase expression.
Asunto(s)
Linfocitos B/enzimología , Regulación Enzimológica de la Expresión Génica , Lactancia/metabolismo , N-Acetil-Lactosamina Sintasa/genética , Preñez/metabolismo , Animales , Células Cultivadas , Femenino , Galactosa/metabolismo , Glicosilación , Inmunoglobulina G/análisis , Inmunoglobulina G/metabolismo , Lactancia/genética , Masculino , Ratones , Ratones Endogámicos BALB C , N-Acetil-Lactosamina Sintasa/metabolismo , Embarazo , Preñez/sangre , Preñez/genética , ARN Mensajero/análisis , ARN Mensajero/metabolismo , Ribonucleasas/farmacología , Bazo/citología , Transcripción GenéticaRESUMEN
Reduced galactosylation of immunoglobulin G (IgG) is well documented in rheumatoid arthritis (RA), but the reason for this defect is still unknown. There is some evidence supporting a defect in the biosynthetic pathway, and a reduction in the level of beta-1,4-galactosyltransferase (beta-1,4-GalTase) enzyme activity. Since glycosyltransferases are, in general, regulated at the level of transcription, we have measured the level of beta-1,4-GalTase gene expression in B cells from patients with RA and normal control individuals. We found no significant difference in mRNA levels for the transferase in these two groups (P > 0.7). MRL/Mp-lpr/lpr (MRL-lpr) mice develop a spontaneous arthritis with increased levels of agalactosyl IgG (G0). In spite of a significant reduction in the level of beta-1,4-GalTase mRNA in total spleen lymphocytes from MRL-lpr mice compared with the congenic MRL/Mp-(+/+) (MRL-(+/+) mice and with CBA/Ca mice, we found comparable levels of the beta-1,4-GalTase mRNA in purified B cells from both spleen and lymph nodes of the three strains. Amongst the lymphoid compartments examined, the spleen and peripheral blood were found to be the major contributors of G0 in MRL-lpr mice. These data indicate that in neither human RA, nor in an animal model of this disease, is reduced IgG galactosylation caused by impaired expression of the beta-1,4-GalTase gene in B lymphocytes. Furthermore, splenic B cells, which have normal levels of beta-1,4-GalTase mRNA, appear to be a major source of G0 in MRL-lpr mice.
Asunto(s)
Artritis Reumatoide/inmunología , Linfocitos B/inmunología , Inmunoglobulina G/biosíntesis , N-Acetil-Lactosamina Sintasa/biosíntesis , Animales , Técnicas de Cultivo de Célula , Modelos Animales de Enfermedad , Regulación de la Expresión Génica , Humanos , Inmunoglobulina G/genética , Ganglios Linfáticos/inmunología , Ratones , Ratones Endogámicos , N-Acetil-Lactosamina Sintasa/genética , ARN Mensajero/genética , Bazo/inmunologíaRESUMEN
MRL-lpr/lpr strain mice have defectively glycosylated IgG. This may be related to the rheumatoid arthritis (RA)-like disease that occurs in these mice, because a similar glycosylation defect is seen in human subjects with RA. Whilst it is known that this defect is associated with reduced activity of the beta-1,4-galactosyltransferase (beta-1,4-GalTase) enzyme, the cause of this reduced activity is at present unknown. We have therefore examined the molecular genetics of beta-1,4-GalTase in MRL-lpr/lpr mice. Using 10 different restriction endonucleases we found no evidence for a polymorphic variant of the gene in glycosylation-defective mice. However, the level of mRNA for beta-1,4-GalTase was lowest in the MRL-lpr/lpr mice, the strain with the most poorly galactosylated IgG of the four strains examined. Thus, the reduced level of IgG oligosaccharide galactosylation found in MRL-lpr/lpr strain mice appears to be related to either an altered transcriptional level of, or altered mRNA stability for, beta-1,4-GalTase in lymphocytes from these mice.
Asunto(s)
Artritis/inmunología , Galactosiltransferasas/genética , Inmunoglobulina G/metabolismo , Ratones Mutantes/inmunología , Animales , Southern Blotting , Técnicas Genéticas , Glicosilación , Ratones , Ratones Endogámicos NOD , Ratones Endogámicos , ARN Mensajero/análisis , Especificidad de la EspecieRESUMEN
The relationship between increased levels of IgG oligosaccharide chains lacking galactose (G0) and the development of rheumatoid arthritis is unclear. In order to further our understanding of the observed correlation between raised serum G0 and arthritis, we have studied G0 levels in arthritis-prone and non-susceptible (i.e. non-arthritis-prone) mice and the effects on G0 of mycobacterial antigens, which have been postulated to play a role in the early events leading to the development of arthritis. We have shown that different age-matched mouse strains have characteristic 'resting' levels of G0 which (in six out of seven strains of mice) increase with age. We have also shown that these increases can be enhanced by immunization of arthritis-prone strains of mice with an adjuvant containing mycobacteria (Freund's complete adjuvant (FCA)), suggesting that deflects in the ability to regulate these G0 changes may be related to susceptibility to arthritis.