Standardized myrtol attenuates lipopolysaccharide induced acute lung injury in mice.

CONTEXT: Standardized myrtol, an essential oil containing primarily cineole, limonene and α-pinene, has been used for treating nasosinusitis, bronchitis and chronic obstructive pulmonary disease (COPD). OBJECTIVE: To investigate the effects of standardized myrtol in a model of acute lung injury (ALI) induced by lipopolysaccharides (LPS). MATERIALS AND METHODS: Male BALB/c mice were treated with standardized myrtol for 1.5 h prior to exposure of atomized LPS. Six hours after LPS challenge, lung injury was determined by the neutrophil recruitment, cytokine levels and total protein concentration in the bronchoalveolar lavage fluid (BALF) and myeloperoxidase (MPO) activity in the lung tissue. Additionally, pathological changes and NF-κB activation in the lung were examined by haematoxylin and eosin staining and western blot, respectively. RESULTS: In LPS-challenged mice, standardized myrtol at a dose of 1200 mg/kg significantly inhibited the neutrophile counts (from 820.97 ± 142.44 to 280.42 ± 65.45, 103/mL), protein concentration (from 0.331 ± 0.02 to 0.183 ± 0.01, mg/mL) and inflammatory cytokines level (TNF-α: from 6072.70 ± 748.40 to 2317.70 ± 500.14, ng/mL; IL-6: from 1184.85 ± 143.58 to 509.57 ± 133.03, ng/mL) in BALF. Standardized myrtol also attenuated LPS-induced MPO activity (from 0.82 ± 0.04 to 0.48 ± 0.06, U/g) and pathological changes (lung injury score: from 11.67 ± 0.33 to 7.83 ± 0.79) in the lung. Further study demonstrated that standardized myrtol prevented LPS-induced NF-κB activation in lung tissues. DISCUSSION AND CONCLUSION: Together, these data suggest that standardized myrtol has the potential to protect against LPS-induced airway inflammation in a model of ALI.

[Values of different biomarkers for diagnosis of Henoch-Schönlein purpura in children].

OBJECTIVE: To screen biomarkers which can be used as an auxiliary method in the diagnosis of Henoch-Schönlein purpura (HSP) and to evaluate their diagnostic values by receiver operating characteristic (ROC) curve analysis. METHODS: A total of 127 children diagnosed with HSP between April 2012 and March 2014 were included in the HSP group and an equal number of healthy children were included in the control group. Twelve parameters, i.e., serum amyloid protein A (SAA), interleukin-6 (IL-6), immunoglobulins (IgA, IgG, IgM, and IgE), C-reactive protein (CRP), white blood cell (WBC) count, complements C3 and C4, anti-streptolysin O, and ferritin, were analyzed. The values of the screened biomarkers for diagnosis of HSP were assessed by ROC curve analysis. RESULTS: The HSP group had significantly higher levels of SAA, IL-6, CRP, WBC, IgA, and IgM than the control group (P<0.05). The areas under the ROC curve of SAA, IL-6, WBC, IgA, and IgM for the diagnosis of HSP were higher than 0.7 (P<0.05). The optimal cut-off values of SAA, IgA, IgM, WBC, and IL-6 for the diagnosis of HSP were 3.035 µg/mL, 1579.5 mg/L, 922.5 mg/L, 8.850 × 109/L, and 7.035 pg/mL, respectively; the corresponding sensitivities of the optimal cut-off values for the diagnosis of HSP were 95.1%, 75.6%, 72.3%, 78.0%, and 63.4%, respectively, and the corresponding specificities were 90.2%, 85.4%, 82.4%, 70.7%, and 80.5%, respectively. CONCLUSIONS: SAA, IgA, IgM, WBC, and IL-6 are valuable biomarkers for clinical diagnosis of HSP and among them SAA seems to be the best one.

[Histopathologic features of degenerative aortic valve and its mechanisms].

OBJECTIVE: To explore the related pathogenesis of degenerative aortic valvular disease by observing the histopathological changes of aortic valves from patients with aortic degenerative stenosis and compare the results with those controls with normal aortic valves. METHODS: Between May 2009 and May 2010, 22 cases of degenerative calcified aortic valves from patients with aortic valve stenosis undergoing aortic valve replacement (14 males, 8 females, mean age: (66 ± 6) years) and 6 cases of normal aortic valves from those with dissection undergoing Bentall operation (4 males, 2 females, mean age: (43 ± 5) years) were collected. The results of hematoxylin and eosin staining and immunohistochemical examinations were used to observe the histological features of degenerative aortic valves and elucidate the related pathogenesis of degenerative aortic valvular disease. RESULTS: Degenerative aortic valve leaflets became thickened. Calcification appeared in aortic side of valve leaflets. Inflammatory infiltrate, angiogenesis, cholesterol crystals, foamy cell aggregation, diffuse and nodular calcification could be seen in subendocardial space of degenerative aortic valve leaflets. No expression of Osterix (OSX) or nuclear factor of activated T-cells 1 (NFATc1) was observed in normal valves. In contrast, the expressions of OSX and NFATc1 showed nuclear immunostaining in degenerative aortic valves. Immunohistochemical staining was graded from 0 to 3. And the expression of OSX was present in 1(4.5%), 1(4.5%), 8(36.4%) and 12 cases (54.5%) respectively in calcified areas, that of OSX in 4(18.2%), 6(27.3%), 7 (31.8%) and 5 cases (22.7%) respectively in non-calcified areas, that of NFATc1 in 1 (4.5%), 1 (4.5%), 8 (36.4%) and 12 cases (54.5%) respectively in calcified areas, that of NFATc1 in 4 (18.2%), 6 (27.3%), 8 (36.4%) and 4 cases (18.2%) respectively in non-calcified areas. The expressions of OSX and NFATc1 in calcified areas were higher than those in non-calcified areas (χ(2) = 8.320, P = 0.040 and χ(2) = 9.371, P = 0.025 respectively). CONCLUSIONS: Unlike in normal valves, inflammatory infiltrate, lipid deposition, angiogenesis and bone regulatory factors appear in degenerative aortic valves. And inflammatory infiltrate, lipid deposition, angiogenesis and ossification may be involved in the degenerative calcified aortic stenosis.

Tie2cre-induced inactivation of the miRNA-processing enzyme Dicer disrupts invariant NKT cell development.

MicroRNAs (miRNAs) are a class of evolutionarily conserved small noncoding RNAs that are increasingly being recognized as important regulators of gene expression. The ribonuclease III enzyme Dicer is essential for the processing of miRNAs. CD1d-restricted invariant natural killer T (iNKT) cells are potent regulators of diverse immune responses. The role of Dicer-generated miRNAs in the development and function of immune regulatory iNKT cells is unknown. Here, we generated a mouse strain with a tissue-specific disruption of Dicer, and showed that lack of miRNAs after the deletion of Dicer by Tie2-Cre (expressed in hematopoietic cells and endothelial cells) interrupted the development and maturation of iNKT cells in the thymus and significantly decreased the number of iNKT cells in different immune organs. Thymic and peripheral iNKT cell compartments were changed in miRNA-deficient mice, with a significantly increased frequency of CD4(+)CD8(+) iNKT cells in the thymus and a significantly decreased frequency of CD4(+) iNKT cells in the spleen. MiRNA-deficient iNKT cells display profound defects in alpha-GalCer-induced activation and cytokine production. Bone marrow (BM) from miRNA-deficient mice poorly reconstituted iNKT cells compared to BM from WT mice. Also, using a thymic iNKT cell transfer model, we found that iNKT cell homeostasis was impaired in miRNA-deficient recipient mice. Our data indicate that miRNAs expressed in hematopoietic cells and endothelial cells are potent regulators of iNKT cell development, function, and homeostasis.

[Association of ABCG2 gene C421A polymorphism and susceptibility of primary gout in Han Chinese males].

OBJECTIVE: To assess the association between a C421A single nucleotide polymorphism (SNP) in exon 5 of ATP-binding cassette, sub-family G (WHITE), member 2 (ABCG2) gene and susceptibility of primary gout in Han Chinese males. METHODS: For 200 male patients with primary gout and 235 controls, the genotype of C421A locus was analyzed by PCR and direct sequencing. Blood glucose, uric acid, total cholesterol, triglycerides, creatinine and urea nitrogen was measured by an automatic biochemical analyzer. RESULTS: Compared with the controls, there was a higher frequency for AA genotype and A allele of the rs2231142 SNP in gout patients (22.5% vs. 8.5% by genotype; 44.9% vs. 32.3% by allele). The association with gout reached significance (chi-square =15.91, P< 0.001, crude OR=3.02, 95% CI:1.36-4.90 and OR (adjusted by age)=1.80, 95% CI: 1.32-2.45 by dominant mode; chi-square=6.82, P=0.009, OR=1.67, 95% CI: 1.54-2.27 by recessive mode). Blood glucose, uric acid, triglycerides, creatinine and urea nitrogen levels in gout patients were significantly higher than those of controls (P< 0.001). CONCLUSION: The C421A SNP, in particular AA phenotype, may be associated with susceptibility of primary gout in Han Chinese males.

[Exploration on the establishment of animal models for gouty nephropathy complicated with chronic renal failure].

OBJECTIVE: To explore the method for establishing animal models of gouty nephropathy complicated with chronic renal failure. METHODS: Six-eight weeks old male Wistar rats were fed with 10% fodder yeast. The adenine at the daily dose of 100, 150, 200, 250, and 300 mg/kg was administrated to them by gastrogavage. The serum levels of blood urea nitrogen (BUN), creatinine (Cr), and uric acid (UA) were dynamically monitored. Meanwhile, the pathological changes of rat kidney were observed. RESULTS: Compared with the normal control group, serum BUN, Cr, and UA obviously increased in rats administered with 100 mg/kg for 7 days (P<0.05). Meanwhile, pathological changes as gouty nephropathy occurred. Along with the prolongation of the modeling time, the aforesaid biochemical indices and pathohistological changes of the kidney were more obvious. The blood Cr level just reached the chronic renal failure level on the 26th day of the administration (about the 4th week), and obviously exceeded the renal failure level on the 41st day (about the 6th week). The blood UA level increased to a higher level on the 7th day of modeling, and maintained at a higher level for a long time. It decreased rapidly from the 41st day to the 48th day. The renal pathological examination showed aggravated infiltration of lymphocytes and stromal fibrous proliferation. On the 48th day of modeling, the proliferation of the fibrous tissue and the interstitial fibrosis were obvious on the bases of the aforesaid changes. The serum BUN, Cr, and blood UA obviously increased in the rats administered with 150, 200, 250, and 300 mg/kg when compared with the normal control group, reaching the level of chronic renal failure (P<0.05). These levels obviously decreased 17 days after restoring to normal fodder feeding, and approached the normal levels till the 35th day. CONCLUSION: Ideal experimental animal models of gouty nephropathy complicated with chronic renal failure could be established in male Wistar rats by feeding with 10% fodder yeast and 100 mg/kg adenine by gastrogavage for 5 weeks.

[Association of the exon 8 and intron 8 polymorphisms of the human urate transporter 1 gene with primary hyperuricemia in Chinese Han population].

OBJECTIVE: To investigate the association of the exon 8 and intron 8 polymorphisms of the human urate transporter 1 gene SLC22A12 with primary hyperuricemia (HUA) in Chinese Han population. METHODS: Genomic DNA from 215 individuals with HUA and 323 controls was extracted. The exon 8 and intron 8 of the SLC22A12 gene was amplified by polymerase chain reaction (PCR). PCR product was sequenced directly. Single nucleotide polymorphisms (SNPs) were detected and the association of the SNPs with primary HUA was assessed. RESULTS: (1) Two SNPs were identified, they were T1309C located in exon 8 (rs7932775) and -103A to G located in intron 8. Pairwise linkage disequilibrium analysis displayed an absolute linkage disequilibrium between the two SNPs (D'= 1). (2) The minor allele frequencies for both SNPs were 51.9% in HUA patients, which were significantly different from that of controls (42.4%)(P< 0.01). (3) The genotype frequencies of GG+ GA and CC+ CT in HUA patients were significantly higher than that in controls (80.0% vs. 69.0%, P< 0.01). (4) Individuals of both GG+ GA and CC+ CT genotypes had 1.79 fold increase of HUA risk (OR= 1.794, 95%CI: 1.19-2.70). CONCLUSION: These findings indicated that T1309C and -103A to G polymorphisms of the SLC22A12 gene were associated with primary HUA in Chinese Han population.

Vibrio cholerae O139 multiple-drug resistance mediated by Yersinia pestis pIP1202-like conjugative plasmids.

A conjugative plasmid, pMRV150, which mediated multiple-drug resistance (MDR) to at least six antibiotics, including ampicillin, streptomycin, gentamicin, tetracycline, chloramphenicol, and trimethoprim-sulfamethoxazole, was identified in a Vibrio cholerae O139 isolate from Hangzhou, eastern China, in 2004. According to partial pMRV150 DNA sequences covering 15 backbone regions, the plasmid is most similar to pIP1202, an IncA/C plasmid in an MDR Yersinia pestis isolate from a Madagascar bubonic plague patient, at an identity of 99.99% (22,180/22,183 nucleotides). pMRV150-like plasmids were found in only 7.69% (1/13) of the O139 isolates tested during the early period of the O139 epidemic in Hangzhou (1994, 1996, and 1997); then the frequency increased gradually from 60.00% (3/5) during 1998 and 1999 to 92.16% (47/51) during 2000 to 2006. Most (42/51) of the O139 isolates bearing pMRV150-like plasmids were resistant to five to six antibiotics, whereas the plasmid-negative isolates were resistant only to one to three antibiotics. In 12 plasmid-bearing O139 isolates tested, the pMRV150-like plasmids ranged from approximately 140 kb to 170 kb and remained at approximately 1 or 2 copies per cell. High (4.50 x 10(-2) and 3.08 x 10(-2)) and low (0.88 x 10(-8) to 3.29 x 10(-5)) plasmid transfer frequencies, as well as no plasmid transfer (under the detection limit), from these O139 isolates to the Escherichia coli recipient were observed. The emergence of pMRV150-like or pIP1202-like plasmids in many bacterial pathogens and nonpathogens occupying diverse niches with global geographical distribution indicates an increasing risk to public health worldwide. Careful tracking of these plasmids in the microbial ecosystem is warranted.

[Effect of pericardial suction blood re-transfusion in off-pump coronary artery bypass grafting on inflammatory cytokines, myocardial injury and pulmonary function].

OBJECTIVE: To investigate the effect of pericardial suction blood re-transfusion in off-pump coronary artery bypass grafting (CABG) on inflammatory cytokines, myocardial injury and lung function. METHODS: 31 patients of off-pump CABG were divided into two study groups (OPCABG1 group and OPCABG2 group) according to the amount of pericardial suction blood re-transfusion beyond or less than 600 ml. 13 patients of on-pump CABG were control group. Serum samples from vein were collected for measurement of IL-6, IL-8, IL-10 and TNF-alpha pre-operation and 1, 4, 24, 48 hours post-operation respectively. The results of CK-MB, TnI, AaDO2 and PaO2/FiO2 were recorded. RESULTS: Patients of the three groups had no significant difference in terms of gender, age, bodyweight, history of hypertension and cardiac infarction and diabetes, EF and left ventricular end diastolic of pre-operation, the amount of bypass graft and shed blood. Of the three groups, IL-6, IL-8 and IL-10 reached peak level one hour after the operation, and dropped to the pre-operation level 72 hours after the operation. One hour after the operation, the level of IL-6 and IL-8 in OPCABG1 group was higher than in OPCABG2 group (P < 0.05) and about the same in CABG group (P > 0.05). Four hours after the operation, the level of CK-MB in OPCABG1 group was lower than that of CABG group (P < 0.05) and about the same in OPCABG2 group (P >0.05). 4 and 24 hours after the operation, the level of TnI in OPCABG1 group was lower than that of CABG group (P < 0.05) and about the same in OPCABG2 group (P > 0.05). Among the three groups, there was no significant difference in AaDO2 and PaO2/FiO2. CONCLUSIONS: Re-transfusion of large amount of pericardial suction blood can increase serum level of IL-6, IL-8, but it can not cause myocardial injury and affect the gas exchange function of lung significantly.

[Optimization of ligustrazine phosphate thermosensitive gel formulation by central composite design-response surface methodology].

OBJECTIVE: To optimize the formulation of ligustrazine phosphate thermosensitive gel by the central composite design-response surface methodology (RSM plus CCD). METHODS: In the formulation design using RSM plus CCD, independent variables were the amounts of poloxamer 407 and poloxamer 188, and gel temperature was dependent variable. Multilinear and quadratic models were used to estimate the relationship between the dependent and the independent variables, select the optimal formulations and validate. RESULTS: The quantitative relationships between two factors and evaluation index were characterized, quadratic model had better prediction capability than multilinear model. CONCLUSION: Quadratic model is performed in the optimization of formulation due to the statistical confidence. The optimization of ligustrazine phosphate thermosensitive gel formulation can be achieved by the central composite design and response surface methodology.

An Intron Variant of SLC2A9 Increases the Risk for Type 2 Diabetes Mellitus Complicated with Hyperuricemia in Chinese Male Population.

BACKGROUND: The aim of this study was to explore the associations of haplotypes of the glucose transporter 9 (SLC2A9) genes with type 2 diabetes mellitus (T2DM) complicated with hyperuricemia (HUA). METHODS: Overall, 608 Chinese males, enrolled from the Affiliated Hospital of Medical College of Qingdao University in 2009-2012, were genotyped. The subjects included 167 withT2DM (average age of onset (58.07±11.82 yr), 198 with HUA subjects (average age of onset (39.20±9.73) yr), 115 with T2DM complicated with HUA (average age of onset (51.24±10.09) yr), and 128 control subjects (average age (41.92±10.01) yr). Patients genotypes of the SNPs; including rs734553 was determined by PCR method. Each genotype was regressed assuming the co-dominant, dominant and the recessive models of inheritance with covariates of duration of total glucose, uric acid, urea nitrogen, triglyceride, cholesterol, and creatinine levels. RESULTS: Chi-square test revealed that rs734553polymorphism was both significantly associated with HUA as well as T2DM complicated HUA, but not with pure T2DM. After adjustment for age and gender, analysis showed that people with C allele had higher risk of HUA and T2DM complicated HUA than those without C allele. And none of the subjects had the homozygous genotype for SLC2A9 (CC). CONCLUSION: The SLC2A9 mutation increases the risk for T2DM complicated HUA in Chinese population, which suggested that intron variants between two relatively conserved exons could also be associated with diseases. In patients of T2DM complicated with HUA, the diagnosis and detection of SLC2A9 gene variants should be caused enough attention.

[PFGE of Shigella flexneri 4c isolates from food-poisoning outbreaks and sporadic diarrhea patients].

OBJECTIVE: To know the molecular characteristic of Shigella flexneri 4c isolates from patients in two food-poisoning outbreaks and one sporadic diarrhea case in Hangzhou, China. METHODS: S. flexneri isolates from patients in two food-poisoning outbreaks (outbreak 1 and outbreak 2, n = 13 and n = 12, respectively) and one sporadic diarrhea patient (n = 1) in Hangzhou during 2003 and 2005 were serotyped. Antibiotic resistances of these isolates were measured by the Kirby-Bauer method. Invasive plasmid antigen gene ipaH was examined by PCR. Pulse field gel electrophoresis (PFGE) was performed for molecular typing. RESULTS: In outbreak 1, all 13 isolates were S. flexneri 4c, of them 6 isolates tested were quite different in PFGE patterns with dice coefficient from 0.78 to 0.92. In outbreak 2, 10 isolates were S. flexneri 4c and 2 isolates were S. flexneri X, however their PFGE patterns were almost identical (dice coefficient > 0.8). Compared to the two outbreaks isolates, the sporadic isolate was demonstrated with a distinct PFGE pattern (dice coefficient < 0.8). The antibiotic resistance patterns with 14 kinds of antibiotics had a little difference among the isolates from outbreak 1, outbreak 2 and sporadic diarrhea patient, but the same pattern was found among 10 isolates of S. flexneri 4c and 2 isolates of S. flexneri X from outbreak 2. CONCLUSIONS: PFGE might distinguish the isolates from these two outbreaks and the sporadic diarrhea patient. Some differences in PFGE patterns, serotypes and antibiotic resistance patterns might occur among S. flexneri 4c isolates during an outbreak.

[Multiplex real-time PCR detecting Salmonella, Shigella and diarrheagenic Escherichia coli].

OBJECTIVE: To develop a multiplex real-time PCR for the detection of Salmonella invasion protein A gene (invA), enterotoxigenic Escherichia coli (ETEC) heat-labile I enterotoxin gene (elt), and Shigella or enteroinvasive E. coli (EIEC) invasive plasmid antigen H gene (ipaH). METHODS: Under the optimized reaction conditions of the multiplex real-time PCR, invA, elt, and ipaH were determined in 10-fold series of dilution of DNA extracted from Salmonella enterica serovar Typhimurium, ETEC 44815 strain and Shigella F301 strain. The three genes were examined in 90 fecal samples from diarrhea patients using the multiplex real-time PCR. When PCR-positive samples were found, the target strains were isolated and identified. RESULTS: The detectable concentration for this multiplex real-time PCR was 10 CFU/microl for Shigella F301 strain, 10(2) CFU/microl for S. enterica serovar Typhimurium and ETEC 44815 strain, respectively. Out of 90 fecal samples from diarrhea patients, thirteen were found positive for elt gene (14.4%), and five were found positive for ipaH gene (5.6%). Three E. coli strains positive for elt gene and four E. coli strains positive for ipaH gene were isolated successfully from the PCR-positive samples mentioned above. The detection of invA, elt and ipaH genes was completed in 10 h, which included an enrichment period of 6 h. CONCLUSION: The multiplex real-time PCR assay can detect invA, elt, ipaH simultaneously in a single reaction, moreover, it can detect for virulence genes in strains of Salmonella, ETEC, and Shigella or EIEC and screen these pathogens in fecal specimens from patients with diarrhea with a high specificity.

Study on the diversity of Bacteroides and Clostridium in patients with primary gout.

To analyze the diversity of both Bacteroides and Clostridium in patients with primary gout and the difference from that of normal individuals. And to investigate the relationship between the primary gout and the intestinal flora. Fecal samples of 90 cases with the primary gout and 94 cases normal comparison group were selected, together with the cases that match the filter criteria. The DNA is extracted from the feces. 16S rRNA specific primers of both Bacteroides and Clostridium were adopted for the PCR amplification. The molecular fingerprints of Bacteroides and Clostridium in both the primary gout group and the normal control group were obtained through DGGE and subjected for further analysis on both the diversity and the similarity. Compared with normal individuals, the number of bands and Shannon-Weaver (H') of Bacteroides in patients with primary gout was not reduced, but significantly decreased in Clostridium. Furthermore, the intra-group and inter-group similarity of both Bacteroides and Clostridium were lower. The primary gout has caused the structural change of both Bacteroides and Clostridium, inducing the low similarity, especially for Clostridium. It has statistic significance. The gut predominant flora may play an important role in the development of primary gout.

[Characteristics of virulence gene in Vibrio parahaemolyticus strains isolated from clinical patients and environment in Hangzhou, China].

OBJECTIVES: To investigate the characteristics of virulence gene in Vibrio parahaemolyticus strains isolated from clinical patients and environment in Hangzhou, China. METHODS: Thermostable direct hemolysin gene (tdh) and thermostable direct hemolysin-related hemolysin gene (trh) were determined in a total of 174 strains of V. parahaemolyticus isolated from patients and environment (seafood) in Hangzhou area by PCR. RESULTS: The tdh was found in 92 out of 94 V. parahaemolyticus strains from food poisoning patients and in 33 out of 34 strains from sporadic diarrhea patients, and trh was not detected in all above clinical strains. Meanwhile the tdh was negative in all V. parahaemolyticus strains from environment, and the trh was also negative except one strain with urease activity. All strains with trh negative had no the activity of urease. CONCLUSIONS: The V. parahaemolyticus strains from food poisoning patients and sporadic diarrhea patients are tdh positive and trh negative. The V. parahaemolyticus strains with tdh negative and almost trh positive in environment might be a potential pathogen in Hangzhou.

Preliminary study on the role of virtual touch tissue quantification combined with a urinary ß2-microglobulin test on the early diagnosis of gouty kidney damage.

The goal of the work described here was to evaluate the role of virtual touch tissue quantification (VTQ) combined with urinary ß2-microglobulin (ß2-MG) measurement in the early diagnosis of gouty kidney damage. Two hundred fifty-nine patients with gouty kidney damage and 200 healthy control subjects were tested. The shear wave velocity (SWV) of the renal parenchyma and sinus as determined with VTQ and the urinary ß2-MG level of the two groups were analyzed. Although there were no significant differences in age, body mass index, creatinine level and blood urea nitrogen between the two groups (all p's > 0.05), the aforementioned parameters were higher in the group with gouty kidney damage than in the control group. Urinary ß2-MG levels of the patients with kidney damage were significantly higher than those of the control subjects (t = 6.38, p < 0.01). The SWV of the renal parenchyma was higher than that of the sinus in both groups. Compared with controls, patients with kidney damage had significantly increased renal parenchyma and sinus SWVs (all p-values < 0.05). Urinary ß2-MG level was positively linearly correlated with the SWV of renal parenchyma in patients with kidney damage (r = 0.442, p < 0.0001). However, there was no correlation between urinary ß2-MG level and the SWV of the sinus in patients with kidney damage (r = 0). In the control group, there was no correlation between urinary ß2-MG level and the SWV of the renal parenchyma or sinus. The elasticity of the kidney as determined with VTQ, combined with the urinary ß2-MG level, may be helpful in the early diagnosis of gouty kidney damage.

Polymorphisms in the NLRP3 gene and risk of primary gouty arthritis.

The aim of the present study was to investigate the association between genetic variants in 17 tagSNPs of the NLRP3 gene and the susceptibility to primary gouty arthritis. A genotype-phenotype analysis of 480 primary gout and 480 control patients was performed. Samples from all the patients were collected from The Affiliated Hospital of Medical College (Qingdao, China). Seventeen tagSNPs of the NLRP3 gene were amplified using polymerase chain reaction (PCR) and MassARRAY technology was used for single nucleotide polymorphism (SNP) genotyping. The genetic frequency of rs7512998 was significantly different between the gout and control patients (P<0.05), whereas no significant differences were identified for the remaining SNPs. The 17 SNPs conformed to the Hardy-Weinberg equilibrium (HWE) in the control group (P>0.05). The haplotype association among the 17 SNPs of the NLRP3 gene indicated that no individual SNP was significantly associated with primary gouty arthritis. CTATCAGCGCCCAGTGC was the most common haplotype in the case and control groups, with a frequency of 0.224 and 0.243, respectively. However, the odds ratios (ORs) of the 8 haplotypes were not identified to be significantly associated with gouty arthritis (P>0.05 for all the 8 haplotypes). To the best of our knowledge, this is the first study to investigate the association between SNPs of the NLRP3 gene and the risk of primary gouty arthritis, although no significant association was identified. Further clinical studies and functional analysis are required to explore the potential associations between NLRP3 gene polymorphisms and the risk of primary gouty arthritis.

[Construction and identification of a eukaryotic expression vector containing human homeobox gene].

In order to investigate the special role of HOXB4 in expansion and self renewal of hematopoietic stem cells, the cDNA of HOXB4 was extracted and cloned from umbilical cord blood mononuclear cells by using RT-PCR. Then the eukaryotic expression bicistronic plasmid vector pIRES2-EGFP/HOXB4 was designed and constructed after cutting HOXB4 and pIRES2-EGFP respectively by restriction enzyme EcoRI and BamHI. The recombinant plasmid was delivered into competent cells of Escherichia coli. The successful construction of plasmid was confirmed by the identification of endonuclease cutting and sequencing. The results showed that the HOXB4 cDNA was cloned successfully from umbilical cord blood mononuclear cells and the recombinant eukaryotic expression bicistronic plasmid vector was constructed, and then introduced it into 293T cells successfully. It is concluded that a pIRES2-EGFP/HoxB1 eukaryotic expression bicistronic plasmid vector has been constructed successfully, which results provide a useful material basis for exploration of HoxB4 function in the proliferation and differentiation of hematopoietic cells.