A new instrument for hematocrit measurement is introduced in the paper, which can get the same accurate Hct values while its centrifuging speed is reduced. It integrates high speed centrifuge, automatic data processing and automatic analysis of hematocrit into a system, having more advantages than the traditional measuring devices, such as lower noise, less test errors and better accuracy. Therefore, it can satisfy well clinical applications.
Automatic Data Processing ; Centrifugation ; Hematocrit ; instrumentation

AIM: The purpose of the present study was to detect intracellular Ca 2+ changes in living brain slices during focal cerebral ischemia/reperfusion (I/R) and reveal the role of intracellular Ca 2+ in the cerebral I/R injury. METHODS: The model of focal cerebral I/R was established in rats by reversible inserting a nylon thread, and dynamic change of intracellular Ca 2+ in brain slices was determined using laser confocal imaging system. RESULTS: ① Ca 2+ gradually enhanced with increase in ischemic time in cortex and striatum. ②At 1 h ischemia/ 10 min reperfusion, Ca 2+ increased significantly in striatum, but Ca 2+ decreased at 3 h reperfusion compared with 10 min reperfusion. ③ Ca 2+ markedly enhanced at 6 h ischemia compared with 1 h ischemia, and after 3 h reperfusion Ca 2+ decreased, but was still higher than that in sham-operation group. ④The striatum is more sensitive than cortex to ischemia/reperfusion. CONCLUSION: Ca 2+ overload in the area of cortex and striatum may play an important role in cerebral ischemia/reperfusion injury in rats.

Abstract: Objective　To investigate the diagnostic efficacy of rifampin-resistant real-time fluorescent quantitative nucleic acid amplification detection technology (GeneXpert MTB/RIF) in bronchoalveolar lavage fluid (BALF) combined with peripheral blood tuberculosis infection T cell spot test (T-SPOT) and tuberculosis antibody (TB-Ab) in smear-negative pulmonary tuberculosis. Methods　The clinical data of 114 cases of clinically diagnosed smear-negative pulmonary tuberculosis, 80 cases of non-tuberculous pulmonary diseases and 22 cases of smear-positive pulmonary tuberculosis in our hospital from January 2019 to January 2021 were retrospectively analyzed. The detection results of peripheral blood T-SPOT, TB-Ab and BALF GeneXpert in the three groups were analyzed. The sensitivity, specificity, negative predictive value, positive predictive value, false negative rate, false positive rate and Youden index of the three detection methods were compared. The differences in the positive detection rate of smear-negative pulmonary tuberculosis between the separate detection and the combined detection of the three methods were compared. The receiver operating characteristic curve (ROC) was performed to calculate the area under the curve (AUC). Results　The sensitivity of BALF GeneXpert and peripheral blood T-SPOT and TB-Ab was 66.91%, 80.88% and 90.44%, respectively. The specificity was 98.75%, 73.75% and 41.25%, respectively; the diagnostic coincidence rates were 78.70%, 78.24% and 72.22%, respectively, which were higher than 70.00%. In the smear-negative pulmonary tuberculosis group, the positive detection rates of these three methods in the smear-negative pulmonary tuberculosis group were 63.15%, 79.82% and 90.35%, respectively, and the differences were statistically significant compared with those in the non-tuberculosis pulmonary disease group (all P<0.01). The positive detection rate of the three combined methods in the smear-negative pulmonary tuberculosis group was 96.49 %, which was significantly higher than that of TB-GeneXpert method and T-SPOT, and the differences were statistically significant (χ2=37.283, P<0.01; χ2=13.612, P<0.01); the Youden index of combined detection was significantly higher than that of single detection, and the AUC of combined detection was 0.977, which was significantly higher than that of single detection. Conclusion　BALF GeneXpert combined with peripheral blood T-SPOT and TB-Ab can significantly improve the diagnostic rate of bacterial-negative pulmonary tuberculosis, providing a strong basis for guiding clinical treatment.

Objective This article is aim to test the safety and the in vitro antibacterial activity of the Lentiviral-nedicated humanβ-defensin 3 (hβD-3) transfecion synovium-derived mesenchymal stem cells (SMSCs).Methods Tissues with SMSCs were obtained by surgical operations.Cells were explant cultured and purified by magnetic cell separation system.Cells were identified by microscopic observation, immunofluorescence and cell surface markers.Through inducing the cells into fat, osteoblasts and chondroblasts to respectively determine the multi-directional differentiation potential.Construct a hβD-3 contained lentivirus and transfected into SMSCs.Got the cell growth curve and determine the DNA content by using flow cytometry.The NOD/SCID mice were applied to verify the tumorigenicity of SMSCs.Agar diffusion test was applied to doing antibacterial activity test of posttransfecton SMSCs.The rabbit model of knee joint in staphylococcus aureus infections verify its bacteriostatic action in vivo.Results: Purified SMSCs had the structure and surface signatures of MSCs.It had the potential of multi-differentiation.Immunofluorescence had verified that SMSCs transfected by lentivirus could stably express hβD-3.Cell proliferation kinetics, karyotype analysis and Tumorigenic type analysis verified the safety of SMSCs after transfection.The in vivo and vitro antibacterial activity test of the recombinant SMSCs showed that after cell passages, the subcultured SMSCs (P5 cells were used in this article) could express hβD-3 stably and had antibacterial activity.The result of the antibacterial circle showed that low concentration group inhibit bacterial activity while the medium and high group could enlarge compared with the negative and positive control group.Conclusion Lentiviral-medicated hβD-3 gene express SMSCs could successfully subcultured and stably express hβD-3, meanwhile it had apparent in vitro antibacterial activity.

Objective To examine expression of PTEN gene in ovarian cancer cisplatin-sensitive cell line OV2008 cells and cisplatin-resistant cell line C13K cells,and evaluate the effect of wild-type PTEN gene on reversing cisplatin-resistance of C13K cells and underlying mechanisms.Methods The expression of PTEN mRNA and protein in OV2008 and C13K cells were detected by semi-quantitative RT-PCR and western blot.Recombinant eukaryotic expression plasmid containing human wild-type PTEN gene was transfected into C13K cells by lipofectamine 2000.The expression of PTEN mRNA was monitored by RT- PCR and the expression of PTEN,protein kinase B(AKT),phospho-AKT(p-AKT)protein were analyzed by western blot in PTEN transfected and untransfected C13K cells.Proliferation and chemosensitivity of cells to cisplatin were measured by methyl thiazolyl tetrazolium(MTT),and cell apoptosis was detected by flow cytometry after treatment with cisplatin.Results(1)The expression of PTEN mRNA and protein(1.02 ?0.05,1.02?0.07)in OV2008 cells were significantly higher than those in C13K cells,which were 0.45 ?0.03 and 0.55?0.03 respectively(P

Objective To establish a radioresistant cell subline from a human A549 lung cancer cell line and investigate the mechanism of radioresistance. Methods Two proposals were applied for the non-small cell lung cancer A549 cells irradiated with X-rays:A group of A549 cell line was irradiated five times, the fractionated dose was 600 cGy, and the other group was exposed 15 times, the fractionated dose was 200 cGy. After the completion of irradiation, two monoclones were obtained from the survival of cells and named the subline A549-S1 and A549-S2. The radiosensitivity and cell cycle distribution of these two clones,together with its parental A549 cells were measured by clone formation assay and flow cytometry.The mRNA and protein levels of Notch1 in A549 cell line and the sublines were determined by RT-PCR and Western-blots. Results Compared with the parental A549 cells, A549-S1 cells showed significant resistance to radiation with D0, Dq and N values increased, and a broader initial shoulder as well as 1.38-fold increased value of SF2. The A549-S1 subline also showed higher percentage of cells in S phase and G2/M phase, but lower percentages in G0/G1 phase (P<0.05). The expression of Notch1 in A549-S1 was enhanced obviously than in A549 cells. But for A549-S2 the radioseasitivity was slightly increased compared with the parental cells with D0, Dq and N values decreased and a curve initial shoulder. The ratio of cells in S and G0/G1 phase ratio was lower than that in parental A549 cells, but that in G2/M phase ratio was higher significantly (P<0.05) .The expression of Notch1 had no marked change compared to A549 cell. Conclusions The radioresistance of the A549 cell subline is correlated with the irradiation program. The cell subline shows a different cell cycle distribution from their parental line. The cell cycle distribution has a close correlation with the expression of Notch1.

Objective To evaluate changes of regional cerebral metabolism by proton MR spectroscopy (~1H-MRS) in patients with minimal hepatic encephalopathy (MHE) and to correlate these changes with the neuropsychological test. Methods Fifty-four patients with cirrhosis including nine patients with hepatic encephalopathy (HE),23 patients with MHE,22 patients without HE and 13 controls underwent neuropsychological tests and ~1H-MRS scanning. The volumes of interest included occipital gray matter and left parietal white matter regions. Ratio of spectral peak areas of N-acetylaspartate (NAA),choline (Cho),myo-inositol (mI),and glutamine/glutamate (Glx) relative to creatine (Cr) were acquired. Statistical analysis was conducted using independent t test and one-way analysis of variance. The results of different groups were compared by using the nonparametric Mann-Whitney U test with Bonferroni correction. Correlations among the ~1H-MRS ratios, the grade of HE, neuropsychological test and ammonia data were calculated with Spearman correlation test. Results The ratios of NAA/Cr,Cho/Cr,mI/Cr,Glx/Cr of the occipital gray matter and left parietal white matter regions in patients with cirrhosis are 1.55±0.12,0.48±0.10,0.42±0.14,2.52±0.48 and 1.73±0.17,0.75±0.16,0.42±0.16,2.75±0.59respectively,and they are 1.53±0.10,0.48±0.09,0.51±0.11,2.20±0.39 and 1.69±0.15,0.82±0.14,0.53±0.12,2.40±0.40 in patients without HE,1.58±0.13,0.48±0.08,0.38±0.13,2.62±0.39 and 1.78±0.18,0.74±0.14,0.38±0.15,2.84±0.58 in patients with MHE,1.54±0.12,0.50±0.13,0.29±0.07,3.04±0.31 and 1.70±0.19,0.62±0.16,0.29±0.07,3.37±0.38 inpatients with HE.Compared with controls, decreased mI/Cr and Cho/Cr ratios and elevated Glx/Cr ratios were found in patients with cirrhosis (t=3.196,9.394,-6.527,P＜0.01,occipital gray matter. t=5.592,9.717,-6.681,P＜0.01,left parietal white matter＝ and in subgroup of patients without HE, with MHE and HE (F=5.097,25.896,20.204,P＜0.01,occipital gray matter.F=16.435,28.660,21.283,P＜0.01,left parietal white matter).Significant difference in these metabolic alterations was also found among the different groups of cirrhosis especially the ratios of Glx/Cr in occipital gray matter and left parietal white matter (P＜0.0084).The ratios of mI/Cr also significantly altered between patients without HE and with MHE (P＜0.0084).There was a significant negative correlation between the ratios of Cho/Cr,mI/Cr and the grade of HE (P＜0.01＝ and a significant positive correlation between the ratios of Glx/Cr and the grade of HE (r=0.709,P＜0.01,occipital gray matter; r=0.720,P＜0.01,left parietal white matter＝.NCT-A and DST of controls is (49±8) s and 39±6.They are (134±37),(83±26),(64±22) second and 15±2,25±9,35±8 in patients with HE,MHE and without HE respectively.The metabolic alterations of Cho/Cr,mI/Cr,Glx/Cr correlated significantly with neurepsychological tests in all subjects (P＜0.01＝.There was a significant positive and a negative correlation between the ratios of Glx/Cr and the data of NCT-A and DST respectively (r=0.570,-0.642,occipital gray matter; r=0.541,-0.632,left parietal white matter).The metabolic alterations of Glx/Cr had no correlation with ammonia data as well as other metabolic alterations.Conclusions ~1H-MRS study shows cerebral metabolic alterations of gray and white matter in patients with cirrhosis,especially the reduction in mI/Cr ratio and increase in Glx/Cr ratio. These changes correlate well with the neuropsychological tests and may be useful in predicting the presence of MHE.

Objective To investigate the dynamic expressions of CXCL11 and its role in the pathogenesis of acute necrotizing pancreatitis (ANP).Methods Forty-eight SD rats were randomly divided into control group and ANP group,with 24 rats in each group.ANP model was induced by retrograde injection of 4％ sodium taurocholate (1 ml/kg body weight) into the biliary and pancreatic duct.The rats were sacrificed at 1,3,6,12 hours.Serum level of amylase was determined,pathological changes in pancreatic tissue were routinely observed and scored.The expression of CXCL11 mRNA and proteon in pancreas was measured by fluorescence quantitative polymerase chain reaction and immunohistochemical method.The serum levels of CXCL11 were measured by enzyme-linked immunoadsorbent assay.Results The serum levels of amylase in ANP rats were significantly higher than those in control group [(6153 ± 355)U/L vs (185 ± 32)U/L at 6 h,P ＜0.05],pathological changes in pancreatre tisues were more significant in ANP rats,and the pathological score was significantly higher than that in control group [(9.00 ± 0.63) vs (0.33 ± 0.12) points at 6 h,P ＜ 0.05] ; the expressions of CXCL11 mRNA and protein in pancreatic tissue were significantly increased than those in control group (3.13 ± 0.43 vs 0.99 ± 0.24,2.76 ± 0.27 vs 0.33 ± 0.12 at 6 h,P ＜ 0.05).The serum level of CXCL11 was significantly higher than that in control group [(112.1 ± 14.2)ng/L vs (56.8 ±4.3) ng/L at 6 h,P ＜0.05)].Conclusions CXCL11 is an early inflammatory mediator in acute pancreatitis,and involved in the pathogenesis of ANP in rats.

AIM: To study apoptosis and bcl-2 mRNA gene expression of cardiomyocytes in donor hearts of immature rabbits underwent prolonged protection by 11, 12-epoxyeicosatrienoic acid (11, 12-EET), and further probe into the possible mechanisms. METHODS: 24 isolated immature rabbit hearts were performed to the model in a Langendorff perfusion apparatus and randomly assigned to normal control group,ST control group and EET group. The isolated rabbit hearts in ST control group and EET group were stored for 24 hours with 4 ℃ hypothermia, and underwent 30 minutes of reperfusion (37 ℃). TUNEL and in situ hybridization (ISH) methods were applied in the present study and apoptotic cells and bcl-2 mRNA gene expression were observed. RESULTS: The numbers of apoptotic cardiomyocytes in ST group and EET group were higher than that in normal control group, and the numbers of apoptotic cardiomyocytes were significantly decreased in EET group and bcl-2 mRNA positive expression were higher than that in ST control group, respectively. CONCLUSIONS: There were apoptosis during the prolonged protection of donor heart in our study, and we proved that: ①11,12-EET could decrease cardiomyocyte apoptosis significantly. ②Up-regulation of the bcl-2 mRNA expression in cardiomyocytes may be one of the mechanism responsible for inhibition of cardiomyocyte apoptosis by 11, 12-EET.

Objective To investigate the diagnostic technique of Alport syndrome(AS)by immunohistochemical staining of type Ⅳ collagen ? chains on paraffin-embedded renal sections.Methods Renal biopsies were obtained from 2 patients with autosomal recessive form of AS,2 female patients and 2 male patients with X-linked dominant form of AS and 2 patients with hematuria(1male and 1 female).AS was diagnosed according to symptoms,family history,pathology,immunofluorescence staining of type Ⅳ collagen ? chains on renal and skin biopsies and gene analysis.Normal portions of nephrectomized kidneys from 2 patients with renal tumor were used as controls.Type Ⅳ collagen ? chains were stained by two-step immunohistochemistry staining method on paraffin-embedded renal sections.Three antigen retrieval methods including autoclave heating,pepsin digestion and proteinase were investigated to find the best antigen retrieval method for type Ⅳ collagen ? chains.The findings were compared with those examined by immunofluorescence staining on fresh frozen sections.Results By immunohistochemistry staining,type Ⅳ collagen ?3 and ?5 chains showed continuous linear pattern along glomerular basement membrane on sections from the controls and the hematuria patients,intermittent linear pattern for X-linked dominant female AS patients,negative for X-linked dominant male AS patient.For patients with autosomal recessive AS,the staining of type Ⅳ collagen ?3 and ?5 chains were negative on glomerular basement membrane,but ?5 chain was positive on glomerular capsules and partial tubular basement membrane.The results were the same as those examined by immunofluorescence staining.Conclusion AS can be diagnosed by immunohistochemistry staining of type Ⅳ collagen on paraffin-embedded renal sections,which is a new technique for diagnosis of AS in China.