METHODS: BV2 microglial cells c for 24 h, pre-treated with EPA for 24 h prior to LPS induction for another 24 h. Surface expression of CD11b and CD40 on BV2 cells was analyzed by flow cytometry. ELISA was employed to measure the production of pro-inflammatory mediators i.e. nitric oxide (NO) and tumor necrosis factor (TNF)-α. Western blotting technique was used to determine the expression of inducible nitric oxide synthase (iNOS), myeloid differentiation protein 88 (MYD88), nuclear factor kappa B (NF-κB), caspase-1, and mitogen activated protein kinase (MAPK).
RESULTS: Qualitative and quantitative analyses of the EPA using a validated ultra-high pressure liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method indicated the presence of phyllanthin, hypophyllanthin, niranthin, ellagic acid, corilagin, gallic acid, phyltetralin, isolintetralin and geraniin. EPA suppressed the production of NO and TNFα in LPS-activated BV2 microglial cells. Moreover, EPA attenuated the expression of MyD88, NF-κB and MAPK (p-P38, p-JNK and p-ERK1/2). It also inhibited the expression of CD11b and CD40. EPA protected against LPS-induced microglial activation via MyD88 and NF-κB signaling in BV2 microglial cells.
CONCLUSIONS: EPA demonstrated neuroprotective effects against LPS-induced microglial cells activation through the inhibition of TNFα secretion, iNOS protein expression and subsequent NO production, inhibition of NF-κB and MAPKs mediated by adapter protein MyD88 and inhibition of microglial activation markers CD11b and CD40.
METHODS: Haemoglobin variants were identified by HbA1c analysis in 93 of 3522 samples sent to our laboratory in a period of 1 month. Haemoglobin analysis identified HbE trait in 81 of 93 samples. To determine the influence of HbE trait on HbA1c analysis by Variant II Tubo 2.0, boronate affinity high performance liquid chromatography (HPLC) method (Primus PDQ) was used as the comparison method. Two stage linear regression analysis, Bland Altman plot and Deming regression analysis were performed to analyse whether the presence of HbE trait produced a statistically significant difference in the results. The total allowable error for HbA1c by the Royal Australasian College of Pathologists (RCPA) external quality assurance is 5%. Hence clinically significant difference is more than 5% at the medical decision level of 6% and 9%.
RESULTS: Statistically and clinically significant higher results were observed in Variant II Turbo 2.0 due to the presence of HbE trait. A positive bias of ∼10% was observed at the medical decision levels.
CONCLUSION: Laboratories should be cautious when evaluating HbA1c results in the presence of haemoglobin variants.