Relationships Between Immunological Characteristics,Influence On FFA Storage Rate And Insulin Resistance In Obese Patients

Objective:Obesity is associated with the developing and progressive process of insulin resistance.However,the mechanism of insulin resistance in obesity is not fully understood and far more complicated and complex than we suspected in human.This study hopes to study the immunological characteristics,including human adipose tissue macrophage（ATM）burden in different depots,the characteristics and their changes of peripheral blood mononuclear cells（PBMCs）subtypes in bariatric surgery patients as well as healthy controls,and mechanism of free fatty acid（FFA）storage of obese patients,and to explore the relationships between these factors and insulin resistance in obese patients.Materials and Methods:（1）97 obese or non-obese volunteers,and 27 obese patients undergoing bariatric surgery were included,respectively.Abdominal subcutaneous adipose tissue and blood samples were obtained from all subjects.The omental adipose tissue samples were also obtained from the 27 bariatric surgery patients.Adipose macrophage burden was measured by immunohistochemistry,using CD68 as the marker for total macrophage burden,CD14 as the marker for the M1 macrophage burden,and CD206as the marker for the M2 macrophage burden.Fat cell size was measured by methylene staining and Ad Count program.Plasma palmitate concentrations were measured by LC/MS.Homeostasis model assessment of insulin resistance（HOMA-IR）and adipose tissue insulin resistance measured by palmitate(ADIPO_palmitate-IR)were used to assess system insulin resistance and adipose tissue insulin resistance,respectively.（2）16 obese patients undergoing bariatric surgery and 15 healthy controls were enrolled.PBMCs were obtained,stained,and detected by flow cytometry（T lymphocytes,regulatory T cells,B cells,and NK cells）.HOMA-IR was used to assess system insulin resistance.（3）16 obese patients were enrolled and underwent diet/lifestyle interventions for weight loss.Abdominal and femoral subcutaneous adipose tissue and blood samples were obtained before and after weight loss.FFA storage rates in adipose tissue were measured by administering radioactive isotope tracers into participants and detecting their portion in the adipose tissue by liquid scintillation counter.Adipose tissue CD36 levels were measured by western-blot.Adipose tissue ACS and DGAT activity were measured by liquid scintillation counter.Fat cell size was measured by methylene staining and Ad Count program.Plasma palmitate concentrations were measured by LC/MS.HOMA-IR and ADIPO_palmitate-IR were used to assess system insulin resistance and adipose tissue insulin resistance,respectively.Results:（1）We found significant univariate correlations between plasma insulin,triglycerides,fasting blood glucose,plasma IL-6 and ATM per 100 adipocytes,but not ATM per gram tissue.Adipocyte size was a confounding variable.For the groups concordant for adipocyte size but discordant for CD206 ATM,we found higher fasting insulin and IL-6 concentrations in the high CD206 ATM group.However,for groups concordant for ATM burden but discordant for adipocyte size,increased visceral fat,plasma triglyceride level,fasting blood glucose and TNF-αconcentrations were found in larger adipocyte size group.Indexes of insulin resistance and TG level were predicted by body composition.Only CD206 ATM per gram tissue was significantly higher in omental adipose tissue than in subcutaneous fat tissue.Plasma insulin,fasting blood glucose levels,and insulin resistance indexes were correlated with omental ATM per 100 adipocytes,while insulin resistance index was can only be predicted by omental adipocyte size and CD206 per gram tissue.（2）HOMA-IR was significantly decreased after bariatric surgery-induced weight loss in obese patients.Activated CD8⁺T cells were significantly higher in obese patients than in healthy controls,and decreased after surgery-induced weight loss.Although Treg cells were significantly lower in obese patients than in healthy controls,they increased after weight loss,yet still lower than normal controls.Obese patients have a higher frequency of mature NK cells,lower frequency of inhibitory NK cells,and lower frequency of activatory NK cells than healthy controls,without any changes after weight loss.Na?ve B cells were significantly higher in obese patients than in healthy controls,and the frequency dropped after weight loss.T cells,Treg cells,B cells,NK cells,and their subgroups were correlated with BMI,WHR,LDL-C,or cholesterol levels.But only B cells and the subgroups were correlated with insulin resistance index,fasting blood glucose,and insulin.（3）Plasma FFA concentrations were closely correlated with direct FFA storage rates in abdominal and femoral subcutaneous adipose tissue.Under high FFA conditions,FFA storage rates in femoral adipose tissue were associated with plasma FFA concentrations,not adipocyte fatty acid storage factors.Under low FFA conditions,direct FFA storage rates in femoral adipose tissue were related to adipocyte CD36 content,not tissue level content of fatty acid storage factors.Weight loss did not change these relationships.Insulin resistance index was predicted by cellular FFA storage rates before weight loss,and cellular CD36 content or DGAT activities after weight loss.Conclusion:（1）The relationship between ATM burden and metabolic/inflammatory variables is confounded by adipocyte size/body composition.ATM does not predict insulin resistance,systemic inflammation,or dyslipidemia.But ATM may primarily play a role in tissue remodeling rather than in metabolic pathology.（2）The change of circulating lymphocyte subgroups frequency in PBMCs indicated impaired immune status in obese patients,and weight loss can alleviate their dysfunction status.Systematic insulin resistance was more correlated with B lymphocytes,while other lymphocyte subgroups were correlated with BMI,waist-hip-ratio,and plasma lipid panel.（3）The regulation of direct FFA storage under low FFA concentration conditions appeared to be at the level of the cell and the cellular content of CD36whereas under high FFA concentration conditions direct FFA storage at the tissue level was predicted by plasma FFA concentrations,independent of adipocyte size or fatty acid storage factors.Insulin resistance in obese patients was predicted by FFA storage rates at the cellular level before weight loss,and FFA storage factors（CD36content and DGAT activity）after weight loss.