Objective:A multi-center and large sample volume study was conducted on the verification and improvement of the early established criteria for intelligent routine urinalysis validation (including the microscopic review rules and manual validation rules, referred to as intelligent criteria for short), in order to improve the clinical application of this intelligent criteria.Methods:A total of 31 456 urine specimens were collected from the inpatients and outpatients in six hospitals in China, from March to September 2019. Firstly, 3105 specimens were analyzed for preliminary verification and improvement of the intelligent criteria based on the results of the microscopic examination and manual validation. Secondly, 28 351 specimens were used to verify the clinical application of the improved intelligent criteria. All samples were manually validated as reference.Results:The approval inconsistency rate of the manual validation rules in the original intelligent criteria was 8.59% (202/2 352), and the interception inconsistency rate was 8.84% (208/2 352). The false negative rate and the microscopic review rate of the microscopic review rules were similar to the previous results. Based on an in-depth analysis of big data and the discussions by senior technicians from eight hospitals, one microscopic review rules and four manual validation rules were added, meanwhile two manual validation rule was deleted. The manual validation standards were unified. Finally, the intelligent criteria was improved. Based on the improved intelligent criteria, for microscopic review rules, the false positive rate, false negative rate (misdiagnosis rate), and microscopic review rate did not change significantly, which were 14.72% (457/3 105), 4.06% (126/3 105), and 24.73% (768/3 105), respectively. The approval inconsistency rate and the interception inconsistency rate of manual validation rules were both reduced to 0; the total manual validation rate of the intelligent criteria was 50.89% (1 580/3 105), and the auto-validation rate was 49.11% (1 525/3 105). The large sample volume verification results were consistent with the preliminary verification results of the improved intelligent criteria.Conclusion:This multi-center and large sample volume study had shown that the improved intelligent criteria had better clinical performance.

Objective To investigate the protective effects of N-acetyl-L-tryptophan (L-NAT) on intestinal damage after rat hepatic ischemia reperfusion.Methods Twenty-four healthy adult rats were divided into the sham operation group (Sham),ischemia reperfusion group (IR),ischemia reperfusion and N-acetyl-L-tryptophan group(IR+L-NAT).The hepatic ischemia reperfusion model was established by occluding the afferent vessels of the left and middle lobes.The morphological structures of the small intestine were observed by hematoxylin-eosin (HE) staining.The expressions of active caspase-3,Bax and Bcl-2 were detected by immunohistochemistry staining.Results (1) In the IR group,the structure of intestinal villis was destroyed,the intestinal mucosa showed congestion and exfoliation,the epithelial cells had degeneration and necrosis,and infiltration of inflammatory cells appeared;which could be alleviated by L-NAT.(2)The immunohistochemistry showed that compared with the Sham group,the expression of active caspase-3,Bcl-2 and Bax in the IR group was increased,after L-NAT intervention,the Bax and caspase-3 expression was decreased,while the Bcl-2 expression was further increased.Conclusion L-NAT could inhibit the apoptosis of small intestinal epithelial cells caused by liver ischemic reperfusion and attenuates intestinal epithelial damage.

Objective: Acoustic pharyngealmetry technology is utilized to evaluate the change and clinical significance of obstructive sleep apnea-hypopnea syndrome (OSAHS) patients caused by non-upper airway structural factor and normal individuals′ PWF(pharyngeal wall floppiness). Methods: Acoustic pharyngealmetry instrument of Ecconvision was utilized to examine 102 OSAHS patients and 50 normal individuals, separately recorded their volume of pharyngeal cavity in sit or supine position, calculated PWF in sit or supine position, and SPSS 12.0 of tware was used to analyze data. Results: PWF was 0.14±0.09 in sit position and PWF was 0.21±0.10, (t=5.96, t=9.63, P<0.001)in supine position of OSAHS group, which were all significantly higher than those of control group. PWFs in supine position of OSAHS group and control group were evidently higher than PWF(t=-11.91, P<0.001; t=-2.32, P=0.025) in sit position. ΔPWF(PWF_supine-PWF_sit)was 0.063±0.054 in OSAHS group which was significantly greater than in control(F=41.173, P<0.01). PWF in sit position and supine position were all positively related with age(r=0.714, r=0.735, P<0.001)while irrelevant with BMI(P>0.05). Conclusions PWF can be utilized to be an index to reflect the physiological feature of upper airway. PWF can more precisely reflect upper airway collapsibility of OSAHS patients on the condition of PWF in supine position. Pharyngeal wall floppiness quantified as a high PWF index is a non-structure vital factor of OSAHS patients and plays a role of guiding us to make personal treatment plans for OSAHS patients.

Objective To study the putative mechanisms underlying fetal alcohol syndrome by comparative protein-profile analysis between normal and ethanol-treated zebrafish embryo with twodimensional electrophoresis (2-DE).Methods Zebrafish embryos were exposed in 400 mmol/L ethanol at dome stage for 3 hours,and then ethanol-induced abnormalities were observed.Proteomes of zebrafish embryos at early stages including zygote stage,dome stage,shield stage and 5-somite stage,were separated by 2-DE.The subtraction analysis method was applied to eliminate the interference from maternal derived proteins.The ethanol-treated embryos at 5-somite stage was analyzed by 2-DE,and the protein profile was compared with that generated from control embryos at the same stage.The data obtained from 2-DE analysis were verified by in-situ hybridization.Results 400 mmol/L ethanol treatment caused axial malformation (62%) and cyclopia (60%) in zebrafish embryos.The 2-DE analysis showed that the expression of Collagen2al (Col2a1) and TAR DNA binding protein (TDP) was decreased in 12 hours post fertilization (12 hpf) ethanol-treated embryos by 81% and 73%,respectively.The in-situ hybridization also demonstrated that the expression of Col2al in axial mesoderm was reduced by ethanol treatment at the same stage.But for 24 hpf ethanoltreated embryos,the expression of Col2al in axis recovered to a comparable level to that in control embryos,while the structure of neural tube was disrupted severely by ethanol exposure.Conclusions It is suggested that the expressions of Col2al and TDP were disrupted by ethanol during early stage,which might induce the zebrafish developmental abnormalities.The ethanol interference on early expression of Col2al is supposed to be one of the major reasons leading to later abnormalities of axis and neutral tube.

Objective: To investigate the expression of ATP-Binding Cassette Proteins including P-gp (P-glycoprotein), MRP1 (multidrug resistance associated protein 1) and BCRP (breast cancer resistance protein) in hepatocellular carcinoma and its relationship with pathological features. Methods: The expression of P-gp/MDR1 (multidrug resistance gene 1), MRP1 and BCRP in hepatocellular carcinoma was examined by RT-PCR and immunohistochemistry in 34 cases of hepatocellular carcinoma and 19 cases of paraneoplastic hepatic tissues. Results: The expression of MDR1, MRP1 and BCRP mRNA (messenger ribonucleic acid) was 1.15±0.24, 0.64±0.33, and 1.07±0.32 in hepatocellular carcinoma and 0.36±0.14, 0.19±0.06, and 0.31±0.09 in paraneoplastic hepatic tissues. The expression of MDR1, MRP1 and BCRP mRNA was 1.38±0.26, 0.73±0.35, and 1.34±0.21 in poorly differentiated hepatocellular carcinoma and 0.74±0.32, 0.30±0.11, and 0.45±0.13 in well differentiated hepatic tissues. The immunohistochemical positive substance was detected in the plasma membrane and cytoplasm. The positive rates of P-gp, MRP1 and BCRP were 82.35%, 58.82%, and 79.41% in hepatocellular carcinoma and 42.11%, 26.32%, and 36.84% in paraneoplastic hepatic tissues, respectively. The positive rates of P-gp, MRP1 and BCRP were 100.00%, 81.25%, and 100.00% in poorly differentiated hepatocellular carcinoma and 66.67%, 38.89%, and 61.11% in well differentiated hepatic tissues. The expression of three indicies in hepatocellular carcinoma was higher than that in paraneoplastic hepatic tissues (P<0.05). The expression of P-gp/MDR1, MRP1 and BCRP in poorly differentiated hepatocellular carcinoma was higher than that in well differentiated hepatic tissues (P<0.05). No correlation was found among the three indices. Conclusion: Intrinsic multidrug resistance exsists in hepatocellular carcinoma, with various mechanisms. The multidrug resistance of HCC (hepatic cell carcinoma) is related to P-gp/MDR1, MRP1 and BCRP. MRP1 and BCRP may be targets for reversing multidrug resistance.

Objective To select the tag SNPs of TLR2 gene in the Chinese population with bioinformatics techniques.Methods We ascertained the assayed scope of the TLR2 gene with the aid of NCBI database and downloaded SNP genotype data of TLR2 gene in the Chinese population from Hapmap database.Then Haploview (version 4.0) was used to calculate linkage disequilibrium (LD) statistics.Haplotype blocks were constructed throughout the TLR2 gene according to the upper and lower 95% confidence bound of the D'value.Meanwhile,we selected the tag SNPs based on r~2 values and the LOD value between SNPs and picked up the representative haplotypes in accordance with the proportion of each haplotype in the haplotype blocks,respectively.Results We constructed 2 haplotype blocks within the TLR2 gene and selected 3 tag SNPs containing 3013 A/G,19216 T/C and 22215 G/T in the Chinese population.Meanwhile,we identified the representative haplotypes of which the tag SNP would be on behalf of every haplotype block.Conclusion The SNPs of 3013 A/G,19216 T/C and 22215 G/T,the most representative SNPs in the whole TLR2 gene in the Chinese population,could be selected as tag SNPs to guide their association studies between the TLR2 gene and sepsis.

BACKGROUND: There are few studies conceming morphological charactedstics, space-time distribution and differentiation of hepatic stem cells during embryo liver development.OBJECTIVE: To understand the action of alpha fetoprotein, cytokeratin-19 (CK19) and c-Met in the liver through observing the expression of them.DESIGN, TIME AND SETTING: The in vitro cytological observational study was performed at the Central Laboratory of Weifang Medical College from June 2005 to December 2006.MATERIALS: A total of 40 embryo samples were obtained from 3-rnonth aborted fetus, which were supplied by Hospital Affiliated to Weifang Medical College.METHODS: Aborted embryo was collected and made into sections within 30 minutes. Fetal age was defined according to embryonic layer formation, somite number and organ development under a microscope. Sample sections with fetal age of 3-12weeks were selected. One was collected from every eleven sections and underwent immunohistochemical staining.MAIN OUTCOME MEASURES: Expression of alpha fetoprotein, CK19 and c-Met was measured in embryo hepatic stem cells aged 3-12 weeks.RESULTS: At 3-5 weeks, samples were positive for alpha fetoprotein and c-Met, which were indicated as hepatic stem ceils. At 10-12 weeks, alpha fetoprotein- and c-Met-positive cells were mainly distributed surrounding the header, which suggested that hepatic stem cells were mainly located at hepatic cord of the header. This had similar distribution as adult hepatic oval cells (adult hepatic stem cells). CK19-positive reaction was found at week 7, and mainly at hepatic cord cells, bile duct sheet cells and bile duct epithelial cells at 10-11 weeks. CKlg-positive reaction was only seen at the bile duct sheet and bile duct epithelial cells at week 12. At this time, all bile duct sheet cells and bile duct epithelial cells were positive for alpha fetoprotein, c-Met and CK19.CONCLUSION: CK19-positive reaction was not found in hepatic stem cells, but only detected in bile duct epithelial cells and progenitor calls. CK19 may be not fit for a marker of hepatic stem cells. All bile duct sheet and bile duct epithelial cells are positive for alpha fetoprotein, c-Met and CK19. It is assumed that alpha fetoprotein+/c-Met+/CK19+ may be bile duct progenitor calls.

Objective To explore the association of CD22 gene T＞A locus (SNPrs2267574) with the development of systemic lupus erythematosus (SLE) and SLE phenotypes in Southern Chinese Han people.Methods Two hundreds fifteen cases and 216 normal controls were enrolled with the aim of case-control design,and the genotype was determined by PCR-RFLP.We calculated X2 and ORs for association study.Results In CD22 gene T＞A locus,there was significant difference of genotypes distribution between cases and controls (X2=6.086,P＜0.05).The frequency of AT genotype was higher in cases than in controls (OR=1.68,95%CI:1.08～2.60,P=0.021 ),and A allele had a higher proportion in cases (OR=1.58,95%CI:1.09～2.29,P=0.015).Meanwhile,the frequency of A allele in patients with positive anti-SSA was higher than in patients with negative anti-SSA (OR=3.69,95%CI:2.08～6.52,P＜0.01 ).Conclusion In Southern Chinese Han population,CD22 gene T＞A locus is associated with the development of SLE and the A allele has positive association with anti-SSA.

BACKGROUND: Studies of biological characteristics of mesenchymal stem cells (MSCs) and regulatory factors that influenced the differentiation of MSCs have shown that the proportion of the natural differentiation from in vitro primarily cultured MSCs into hepatocytes was low, and to select a suitable inductor is important to enhance the differentiation of MSCs into hepatocytes.OBJECTIVE: To verify the feasibility of induced differentiation of rat bone marrow MSCs (BMSCs) into hepatocytes using the combination of hepatocyte growth factor (HGF), epidermal growth factor (EGF) and fibroblast growth factor (FGF-4).DESIGN, TIME AND SETTING: The cytological in vitro study was performed at the Experimental Center, Weifang Medical College in August 2007.MATERIALS: Totally 40 Sprague-Dawley rats were supplied by the Experimental Animal Center, Weifang Medical College.METHODS: Rat BMSCs were incubated by adherent method. BMSCs at passage 3 were assigned to 2 groups. BMSCs in the blank control group were treated with L-DMEM containing 10% fetal bovine serum. BMSCs in the combination group were treated with 10 μg/L FGF, 8 μg/L HGF and 8 μg/L EGF following above-mentioned procedures.MAIN OUTCOME MEASURES: Inverted microscope was used to observe the morphological changes in cells.Immunofluorescence method was used to observe the expression of alpha-fetoprotein (AFP) and albumin (ALB). PAS was employed to detect the expression of glycogen. Fox green intake experiment was conducted. Enzymology was utilized to test the contents of alanine aminotransferase (ALT), aspartate aminotransferase (AST), and alkaline phosphatase (ALP).RESULTS: BMSCs in the combination group presented polygonal, orbicular or round shape. BMSCs in the blank control group remained spindle. BMSCs in the combination group were positive for AFP and ALB at day 14 following culture, and a few PAS-positive and fox green-positive cells were found at day 7. Positive cells became more over time. Synthesis of ALT, AST and ALP was detected at day 14, reached a peak at 21 days, and then decreased. Above-described indexes were negative in the blank control group.CONCLUSION: After induced by the FGF, HGF and EGF, BMSCs have the ability to differentiate into hepatocytes in vitro.

Objective To explore the possibility that rat bone mesenchymal cells (BMSCs) can differentiate into hepatocytes under the affection of fetal liver filtrate. Methods PAS and green indigo dye were used to detect glycogen and differential level of hepatocytes, respectively. The concentration of ALT, AST, ALP in the culture supernatant were served as markers of hepaocyte function. Results Fourteen days after induced by the fetal liver filtrate, BMSCs changed their shapes into polygon, oval or round. Some of BMSCs were positive for AFP and ALB at 7 days after induction, then the number of positive cells increased, and most of BMSCs expressed AFP and ALB till 21days. The PAS reaction and indocyanine green(ICG) intaking also appeared at 7days. Enzyme in supernatant such as ALT, AST, ALP were fristly detected at 7days and peaked at 14days,then the level declined. Conclusion The fetal rat liver filtrate was able to induce BMSCs into cells with function and characteristics of hepatocytes.