The Correlation Between CD14 Gene Polymorphism And Cell-mediated Immunity In Extensively Burned Patients

Objective: In recent years, genetic polymorphism has became a promising tool to explain interindividual variations in gene products and susceptibility to sepsis. The present study was performed to investigate the correlation between CD14 gene polymorphism and the cell-mediated immunity in extensively burned patients. In this study, the polymorphisms of the CD14 promoter region -159 were determined in the severely burned patients, and their relations to the development of sepsis or multiple organ dysfunction syndrome (MODS) were analyzed, thereby providing new methods to predict and prevent postburn sepsis and MODS. Meanwhile, we studied the potential mechanism underlying the effect of CD14-159C/T polymorphism on cell-mediated immunity as well as late cytokine (high mobility group box-1 protein, HMGB1) expression following major burns.Methods: (1) Blood samples were obtained from 77 patients with burns covering more than 30% of the total body surface area on postburn days (PBD) 1, 3, 5, 7, 14, 21, and 28, respectively. the CD14 polymorphism was determined by polymerase chain reaction (PCR) and subsequent restriction fragment length polymorphism (RFLP) analysis. The proliferation of T lymphocytes in periphery blood was detected by MTT, and sCD14 levels as well as IL-2 production were measured by ELISA. The percentage of CD4+/CD8+ lymphocytes, apoptosis of CD4+ lymphocytes, and the amounts of binding HLA-DR on mononuclear cell surface were determined by flow cytometry (using Monoclonal antibody, QuantiBRITETM Anti-HLA-DR PE*/Anti-Monotype PerCP-Cy5.5). (2) Blood samples were collected from 35 patients with burns covering more than 30% of the total body surface area, and plasma HMGB1 levels and HMGB1 mRNA expression were measured by ELISA and RT-PCR, respectively. The correlation between HMGB1 levels and parameters of cellular immunity was analyzed in burned patients.Results: (1) The genetype of CD14 was observed in 77 severely burned patients, and the frequence of sepsis occurred in homozygous for the C allele (CC) (42.9%) was much lower than in heterozygote for the TC allele (TC) (77.6%) and homozygous for the T allele (TT) (71.4%). In three septic patients for CC alleles, only one developed MODS, while in 38 patients of TC alleles, there were 19 patients appeared MODS (50%), and in 15 patients of TT alleles, there were 10 patients developed MODS (66.7%). (2) In patients of three genetypes, the expression of CD14 mRNA was markedly higher in TC heterozygote and TT homozygote than in CC homozygote on PBD 7 to 21, and expression of CD14 mRNA among three genetypes was obviously different (P<0.05 or P<0.01), but the difference of CD14 mRNA between TT and TC was not significantly (P>0.05). As same as CD14 mRNA expression, high sCD14 levels were observed in TC, TT patients, and sCD14 levels in CC patients was lower than TC homozygote on PBD 5 (P<0.01)(.3)The ability of T lymphocyte proliferation and the production of IL-2 were in low level in TC, TT patients, but they increased in CC patients on PBD 14. Compared with CC homozygote patients, ratios of T lymphocytes in TT and TC patients was decreased, extremely on PBD 1, 3, 14, 21, and 28 (P<0.01). The apoptosis of CD3+/CD4+ T lymphocyte in TT, TC patients was in a high value, and it was much higher in TT patients than in CC patients on PBD 5, 7, and 21 (P<0.05), and this ratio in TC patients was also higher than CC patients on PBD 7 and 14 (P<0.05). Furthermore, the amounts of binding HLA-DR on mononuclear cell surface were obviously higher in CC homozygote patients than in TT patients on PBD 3, 14, and 21 (P<0.05), without statistical significance compared to TC heterozygote (P>0.05). (4) The plasma HMGB1 concentrations was markedly higher in TT and TC patients than in CC homozygote on PBD 14, 21, and 28 (P<0.05). Also, mRNA expression of HMGB1 was obviously difference in TT, TC and CC groups on PBD 14 (P<0.05), while no significant differences were found at other time points. (5) Plasma HMGB1 concentrations were negatively correlated with T lymphocyte proliferation activity, IL-2 release, the ratio of CD4+/CD8+ lymphocyte, and the monocyte HLA-DR expression, respectively (P<0.05 or 0.01), while they were positively correlated with apotosis of CD4+ T lymphocyte as well as plasma sCD14 levels, respectively (P<0.05 or 0.01).Conclusion: (1) CD14 gene promoter -159 position polymorphism TT homozygote might be a high risk marker for the development of MODS in septic burned patients. (2) The polymorphism of the CD14 gene promoter -159 position was closely related to changes in cellular immune function after burns, suggesting it could markedly influence the host immunological respose following acute insult. (3) It is likely that CD14 gene polymorphism might influence the expression as well as release of HMGB1, subsequently contributing to the dysfunction of cell-mediated immunity associated with major burns.