P2Y12 and P2Y13 receptors involved in ADPßs induced the release of IL-1ß, IL-6 and TNF-α from cultured dorsal horn microglia.

OBJECTIVE: P2 receptors have been implicated in the release of neurotransmitter and pro-inflammatory cytokines due to their response to neuroexcitatory substances in the microglia. Dorsal horn P2Y12 and P2Y13 receptors are involved in the development of pain behavior induced by peripheral nerve injury. However, it is not known whether P2Y12 and P2Y13 receptors activation is associated with the expression and the release of interleukin-1B (IL-1ß), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) in cultured dorsal spinal cord microglia. For this reason, we examined the effects of ADPßs (ADP analog) on the expression and the release of IL-1ß, IL-6, and TNF-α. METHODS AND RESULTS: In this study, we observed the effect of P2Y receptor agonist ADPßs on the expression and release of IL-1ß, IL-6 and TNF-α by using real-time fluorescence quantitative polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). ADPßs induced the increased expression of Iba-1, IL-1ß, IL-6 and TNF-α at the level of messenger RNA (mRNA). ADPßs-evoked increase in Iba-1, IL-1ß, IL-6 and TNF-α mRNA expression was inhibited only partially by P2Y12 receptor antagonist MRS2395 or P2Y13 receptor antagonist MRS2211, respectively. Similarly, ADPßs-evoked release of IL-1ß, IL-6 and TNF-α was inhibited only partially by MRS2395 or MRS2211. Furthermore, ADPßs-evoked increased expression of Iba-1, IL-1ß, IL-6 and TNF-α mRNA, and release of IL-1ß, IL-6 and TNF-α were nearly all blocked after co-administration of MRS2395 plus MRS2179. Further evidence indicated that P2Y12 and P2Y13 receptor-evoked increased gene expression of IL-1ß, IL-6 and TNF-α were inhibited by Y-27632 (ROCK inhibitor), SB203580 (P38MAPK inhibitor) and PDTC (NF-κb inhibitor), respectively. Subsequently, P2Y12 and P2Y13 receptor-evoked release of IL-1ß, IL-6 and TNF-α, were also inhibited by Y-27632, SB203580 and PDTC, respectively. CONCLUSION: These observations suggest that P2Y12 and P2Y13 receptor-evoked gene expression and release of IL-1ß, IL-6 and TNF-α are associated with ROCK/P38MAPK/NF-κb signaling pathway.

Study on the in vivo toxic mechanism of xixin based on trace elements determination by inductively coupled plasma-mass spectrometry.

BACKGROUND: Xixin has been widely used as a traditional Chinese medicine for headache, toothache and inflammatory diseases. Clinical investigation indicated that adverse drug reactions occurred with an overdose of xixin, but the toxic mechanism of xixin in vivo based on trace elements has not been researched yet. OBJECTIVE: To explore the in vivo toxic mechanism of xixin induced by trace elements. MATERIALS AND METHODS: The contents of trace elements in the serum and liver of mice were determined by inductively coupled plasma-mass spectrometry (ICP-MS) after obtaining xixin extracts. Principal component analysis (PCA) and cluster analysis (CA) were performed between the trace elements' content and dosage using the software GeneSpring 12.1 to analyze the main toxic elements in vivo. RESULTS: Trace elements' contents were obviously raised after xixin extracts were taken as a dosage of 150 mg/mL and 50 mg/mL, respectively. Na, Ca, Cu and Cd in serum and Ca and Zn in liver were the main trace elements inducing the toxic reaction of xixin. CONCLUSION: Xixin possesses the potential function of indirectly upregulating trace elements in vivo. This study, for the first time, elucidated the in vivo toxic mechanism of xixin based on trace elements. This method could also be utilized in the research of corresponding aspects.