OBJECTIVETo study the expression level and role of apoptosis-associated speck-like protein containing a caspase recruitment domain (PYCARD) gene transcript variant mRNA in peripheral blood mononuclear cells (PBMCs) of primary gout (PG) patients with different Chinese medicine (CM) syndromes.

METHODSThe expressions of PYCARD gene transcript variant mRNA and interleukin-1β (IL-1β) mRNA in PBMCs were investigated in 96 PG patients with acute phase (APPG, 44 cases) and non-acute phase (NAPPG, 52 cases) and 30 healthy controls (HCs) by reverse transcription-polymerase chain reaction (PCR) and/or realtime quantitative PCR. PYCARD and nuclear factor-κB (p50) [NF-κB (p50)] protein was detected by Western blot in PBMCs respectively. IL-1β, IL-4 and IL-10 protein levels in plasma of HCs and PG patients were measured by enzyme-linked immuno sorbent assay.

RESULTSThe main CM syndromes in APPG patients were obstruction of dampness and heat syndrome (ODHS, 36.36%) and intermingled phlegm-blood stasis syndrome (IPBSS, 27.27%), while in NAPPG patients were Pi (Spleen)-deficiency induced dampness syndrome (PDIDS, 40.38%) and qi-blood deficiency syndrome (QBDS, 26.92%). It showed statistical significances of the expressions of PYCARD gene and its transcript variant mRNA, the protein of PYCARD and NF-κB (p50) and the plasma IL-1β, IL-4 and IL-10 in APPG, NAPPG, ODHS, IPBSS, PDIDS and QBDS groups, compared with the HC group respectively (P<0.05 or P<0.01). There were also significant differences of mRNA expressions of PYCARD-1 and PYCARD-2 as well as protein expressions of IL-1β, IL-4 and IL-10 among the 4 CM syndromes groups (P<0.05 or P<0.01). Correlation analysis showed positive correlation between the mRNA expressions of PYCARD-1 gene transcript variant and IL-1β in APPG patients (r=0.3088, P=0.0183).

CONCLUSIONPYCARD gene and its transcript variant may play a critical and regulative role in the inflflammatory response of PG patients with different phases and CM syndromes.

OBJECTIVE: To determine the effects of berberine (BBR) on the activation of toll-like receptor 4 (TLR4), nuclear factor (NF)κB (NF-κB) signaling and NLRP3 inflammasome in patients with gout. METHODS: Peripheral blood mononuclear cells (PBMCs) were obtained from 24 acute (AP) and 41 non-acute (NAP) phases of primary gout patients, respectively, as well as 30 healthy controls (HC). TLR4, NF-κB (p65), NLRP3, apoptosis-associated specklike protein containing a CARD (PYCARD), cysteinyl aspartate specific proteinase-1 (CASP1), and interleukin-1β (IL-1β) mRNA expression levels in PBMCs were measured by quantitative reverse transcriptase polymerase chain reaction. The protein levels of TLR4, myeloid differentiation factor 88 (MyD88), NF-κB (p50/65), inhibitor of kappa B kinase α/β (IKKα/β), NF-κB inhibitor α (IKBα), phospho-IKKα/β (p-IKKα/β), NLRP3, PYCARD, and CASP1 were monitored by Western blotting. Serum IL-1β protein level was measured using enzyme-linked immunosorbent assay (ELISA). In addition, PBMCs from HC and macrophages derived from a spontaneously immortalized monocyte-like cell line (THP-1) were stimulated using monosodium urate (MSU, 100 µg/mL), 0.1% dimethyl sulfoxide, 25 µmol/L BBR, and 10, 25, and 50 µmol/L BBR+100 µg/mL MSU for different time periods. The protein levels of IL-1β and IL-18 in cell culture supernatants was measured by ELISA, and the protein expressions of TLR4, MyD88, NF-κB (p50/p65), IKKα/β, I κBβ, p-IKKα/β, NLRP3, PYCARD, and CASP1 in macrophages were analyzed by Western blotting. RESULTS: (1) TLR4, NF-κB (p65), PYCARD, CASP1, and IL-1β mRNA levels in PBMCs were significantly higher in the AP group than in the HC group (P<0.05). The NLRP3 mRNA expression levels in PBMCs were found to be significantly lower in the AP and NAP groups than in the HC group (P<0.05, P<0.01). (2) The protein levels of TLR4, IKKβ, MyD88, NF-κB, p-IKKα/β, PYCARD, and CASP1 in PBMCs were significantly higher, and those of IκBα, IKKα, and NLRP3 were found to be significantly lower in the AP group than in the HC group (P<0.05 or P<0.01). (3) The serum IL-1β protein levels were significantly higher in the AP and NAP groups than in the HC group (P<0.01). (4) The IL-1β protein level was significantly lower in the culture supernatants of the PBMCs stimulated with MSU for 3 and 6 h in the 25 and 50 µmoL/L BBR groups compared with that in the MSU group (P<0.01). (5) The protein levels of IL-1β and IL-18 were also significantly lower in the culture supernatants of macrophages stimulated with MSU for 3 and 6 h in BBR groups compared with those in the MSU group (P<0.01). (6) The protein levels of TLR4, MyD88, NF-κB (p50, p65, p105), IKKα/β, p-IκBα, p-IKKα/β, PYCARD, and CASP1 were significantly differed between the macrophages stimulated with MSU for 0.5 and 6 h in BBR groups compared with those in the MSU group (P<0.05 or P<0.01). CONCLUSIONS Activation of TLR4-NFκB signaling and NLRP3 inflammasome by MSU crystals drives the progression of gout inflammation. BBR ameliorates gouty inflammation, which is mechanistically associated with its regulation of TLR4-NF-κB signaling and NLRP3 inflammasome expression.

OBJECTIVETo understand the difference in clinical indicators of gout patients of different Chinese medical syndromes and its clinical significance.

METHODSForm November 2011 to December 2012, syndrome typed were 257 male gout in-/outpatients from Affiliated Hospital of Chuanbei Medical College. Another 50 healthy male subjects were recruited as the control. Their clinical and laboratory data were collected. All were excluded from infections and other inflammatory diseases.

RESULTSFour syndrome types existed in gout patients, i.e., intermingled phlegm-stasis blood syndrome (IPSBS), obstruction of dampness and heat syndrome (ODHS), Pi-deficiency induced dampness syndrome (PDIDS), qi-blood deficiency syndrome (QBDS). Of them, 53 acute phase gout patients suffered from IPSBS, 41 from ODHS, 25 from QBDS, and 17 from PDIDS; 41 non-acute phase gout patients suffered from QBDS, 40 from PDIDS, 24 from ODHS, and 16 from IPSBS. Statistical analysis of clinical data showed that, when compared with the normal control group, there was statistical difference in blood routines (WBC, GR, LY, MO) and blood biochemical indices (UA, Ur, Cr, ALT, AST, ALB, GLOB, TG, HDL-C, VLDL-C, apoA, apoB100) of gout patients of different syndromes (P < 0.05, P < 0.01). There was also statistical difference or correlation among different syndromes (P < 0.05).

CONCLUSIONSIn the acute phase gout patients, IPSBS and ODHS were dominated, while in the non-acute phase gout patients, QBDS and PDIDS were often seen. In patients of IPSBS and ODHS, inflammation and immune response were more obvious, indicating that better efficacy might be achieved by clearing heat and removing blood stasis associated anti-inflammatory and immune regulation therapies. In patients of QBDS and PDIDS, impaired renal functions were more significant, indicating that better efficacy might be achieved by invigorating Pi and tonifying Shen dominated treatment.

Adult ; Aged ; Case-Control Studies ; Gout ; diagnosis ; Humans ; Male ; Medicine, Chinese Traditional ; methods ; Middle Aged ; Yang Deficiency ; diagnosis ; Yin Deficiency ; diagnosis ; Young Adult