Objective:To investigate the effect and mechanism of 6-formylindolo[3,2-b]carbazole (FICZ) on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice.Methods:Male C57BL/6J mice aged 8-12 weeks were divided into 4 groups with 8 mice in each group, according to the method of simple random sampling. Sepsis-induced ALI mice model was established by intraperitoneal injection of LPS 5 mg/kg (LPS group), and phosphate buffer saline (PBS) control group (PBS group) was injected with equal volume of PBS. The LPS+FICZ group was intervened by intraperitoneal injection of 1 μg FICZ 1 hour after LPS stimuli, while the FICZ control group (FICZ group) was given the same amount of FICZ 1 hour after intraperitoneal injection of PBS. Serum and lung tissue were collected 24 hours after LPS stimuli, and the pathological changes of lung tissue were analyzed by hematoxylin-eosin (HE) staining and wet/dry weight (W/D) ratio of lung tissue. The concentrations of inflammatory factors in serum and lung tissue were detected by enzyme linked immunosorbent assay (ELISA). The expression levels of endoplasmic reticulum stress signaling pathway related molecules were detected by real-time fluorescent quantitative polymerase chain reaction (RT-qPCR) and Western blotting.Results:Compared with PBS group, inflammatory cell infiltration, alveolar collapse and obvious alveolar exudative lesions had increased, lung tissue W/D ratio was significantly increased, serum interleukin-6 (IL-6) level, lung tissue IL-6 mRNA expression, and the mRNA expressions of glucose-regulated protein 78 (GRP78), protein kinase R-like endoplasmic reticulum kinase (PERK), CCAAT/EBP homologous protein (CHOP), and the protein expressions of GRP78, PERK, activating transcription factor 6 (ATF6), CHOP in lung tissue were significantly increased in LPS group. However, the indexes of FICZ group were not affected. Compared with LPS group, LPS+FICZ group had less inflammatory cell infiltration, relatively intact alveolar structure. Lung W/D weight ratio in LPS+FICZ group was significantly decreased (5.38±0.10 vs. 6.60±0.30, P < 0.01), so as serum IL-6 (ng/L: 15.55±3.77 vs. 32.22±3.84) and lung IL-6 mRNA expression (2 -ΔΔCt: 0.79±0.21 vs. 6.89±0.92, both P < 0.01). The mRNA expressions of GRP78, PERK and CHOP were also significantly decreased [GRP78 mRNA (2 -ΔΔCt): 1.90±0.16 vs. 7.55±1.29, PERK mRNA (2 -ΔΔCt): 1.68±0.20 vs. 4.54±0.89, CHOP mRNA (2 -ΔΔCt): 1.13±0.24 vs. 4.44±1.13, all P < 0.05], and the protein expressions of GRP78, PERK, ATF6 and CHOP were significantly decreased (GRP78/GAPDH: 0.59±0.02 vs. 0.77±0.01, PERK/GAPDH: 0.48±0.03 vs. 1.04±0.05, ATF6/GAPDH: 0.51±0.03 vs. 0.65±0.01, CHOP/GAPDH: 0.91±0.05 vs. 1.11±0.07, all P < 0.05). Conclusion:FICZ protects LPS-induced ALI possibly via suppressing endoplasmic reticulum stress and reducing IL-6 expression in blood and lung tissue.

OBJECTIVETo construct a subtracted cDNA library of differentially expressed genes in eosinophils from asthma patients.

METHODSSuppression subtractive hybridization (SSH) was used to isolate the cDNA fragments of differentially expressed genes in the eosinophils of asthma patients before and after treatment. The cDNA fragments were directly inserted into T/A cloning vector to establish the subtractive library, followed by amplification of the library through E. coli transformation with calcium chloride and screening of blue and white clones of the transformants. One hundred positive bacterial clones were randomly picked and identified by colony PCR.

RESULTSThe amplified library contained more than 3,000 positive bacterial clones. Analysis of the randomly selected 100 white clones by PCR showed that 90% of the clones contained 100-500 bp inserts, which might be the cDNA fragments of differentially expressed genes in eosinophils of asthma patients before treatment.

CONCLUSIONA subtracted cDNA library of differentially expressed genes in the eosinophils of asthma patients before and after treatment is constructed successfully by SSH and T/A cloning techniques, which lays a solid foundation for screening and cloning new specific differentially.expressed genes in the eosinophils of asthma patients.

Asthma ; blood ; genetics ; DNA, Complementary ; genetics ; Eosinophils ; metabolism ; Gene Expression Profiling ; Gene Expression Regulation ; genetics ; Gene Library ; Humans ; Male ; Middle Aged ; Nucleic Acid Hybridization

OBJECTIVETo investigate the feasibility of promoting nerve regeneration by using microcapsulated NGF-expressing cells transplantation.

METHODSThe plasmid pcDNA3. 1 + /NGF, containing rat NGF gene, was transfected into the NIH 3T3 cell by using FuGENE6 transfection reagent. The genetically modified cells expressing NGF gene were enclosed within the alginate-polylysine-alginate (APA) microcapsules and then cultured in vitro. The growth and NGF secretion of the cells were measured periodically. At the same time, these microcapsulated NGF-expressing cells were transplanted into the injured sciatic nerve. The regeneration and functional recovery of the nerve were evaluated in 4 weeks, 8 weeks and 12 weeks after the operation.

RESULTSThe microcapsulated cells had survived and secreted the NGF in three months in vitro. In the group with microcapsulated NGF-expressing cells, the number of the regenerated axons was in large and the nerve fibers were arranged regularly. Compared to other groups, there was less scar , edema and monocytes found at the stoma in the goup. The moter nerve conductive velocity, nerve muscle-action potential and SFI were improved.

CONCLUSIONSThe microcapsulated NGF-expressing cells could significantly enhance the nerve regeneration and reduce inflammatory response of xenograft.

Animals ; Cell Transplantation ; Female ; Male ; Mice ; NIH 3T3 Cells ; Nerve Growth Factor ; biosynthesis ; genetics ; Nerve Regeneration ; Rats ; Rats, Sprague-Dawley ; Transfection

OBJECTIVETo explore the relationship between nitrite levels in the exhaled breath condensates (EBC) and the severity of asthma.

METHODSSixty asthmatic patients with exacerbation (including 23 with mild, 21 with moderate, and 16 with severe asthma) and 23 healthy nonsmokers were enrolled in this study. The lung function tests were performed and nitrite levels measured in the EBC by the spectrophotometry and nitric oxide assay kit in these subjects. The percentage of eosinophils was also measured in induced sputum by Wright staining.

RESULTSThe concentrations of nitrites in the EBC and the percentage of eosinophils in induced sputum in the asthmatic patients were significantly higher than those of the healthy subjects (P<0.01), and showed positive correlations to the disease severity. A significant positive correlation was found between nitrites in the EBC and percentage of eosinophils in induced sputum (r=0.706, P<0.01). The concentration of exhaled nitrites was inversely correlated to MEF50% (r=-0.806, P<0.01) and FEV1% (r=-0.724, P<0.01).

CONCLUSIONNitrite level in the EBC may serve as useful indicators for estimating the severity of asthma.

Adult ; Asthma ; metabolism ; Breath Tests ; methods ; Case-Control Studies ; Exhalation ; Female ; Forced Expiratory Volume ; Humans ; Male ; Middle Aged ; Nitrites ; analysis ; Respiratory Function Tests ; Severity of Illness Index ; Young Adult

Objective:To observe the impact of diabetes mellitus (DM)on pacing parameters and postoperative com-plications in patients With implantation of permanent artificial cardiac pacemaker.Methods:A total of 80 patients With sick sinus syndrome,Who received implantation of permanent artificial cardiac pacemaker from Jun 2008 to Jun 2011,Were enrolled.According to complicated With DM or not,they Were divided into DM group (n=40)and non-DM control group (n=40).Pacing parameters and postoperative complications Were compared betWeen tWo groups.Results:There Were no significant difference in atrial and ventricular pacing threshold,sensing and of pace-maker impedance in baseline betWeen tWo groups (P>0.05).All parameters of pacemaker increased in tWo groups after implantation 12 months;compared With non-DM control group,there Were significant increase in pacing threshold [atrial:(0.59±0.23)V vs.(0.67±0.25)V,ventricular:(0.47±0.28)V vs.(0.54±0.35)V],sens-ing [atrial:(2.33±1.16)mV vs.(2.92±1.36)mV,ventricular:(12.21±4.82)mV vs.(12.77±5.36)mV], impedance [atrial:(537.12±115.32)Ωvs.(662.48±235.26)Ω,ventricular:(602.48±222.46)Ωvs.(762.41± 235.38)Ω]of pacemaker in DM group,P<0.05 or <0.01;and incidence rate of postoperative complications (12.5%)in DM group Was significantly higher than that of non-DM control group (5%),P<0.05.Conclusion:Electrocardiographic reconstruction is more severe in SSS patients complicated DM,in these patients postoperative complication incidence significantly elevates.

OBJECTIVETo investigate UbcH10 expression in hepatocellular carcinoma and explore its clinicopathological implications.

METHODSWe detected UbcH10 mRNA expression using RT-PCR in normal liver cell line, cancer cell lines, surgically removed hepatocellular carcinoma tissue and corresponding adjacent non-tumor tissue and evaluated the clinicopathological significance of UbcH10. Immunohistochemistry was performed to investigate UbcH10 protein expression in hepatocellular carcinoma tissue, the adjacent tissue, and normal liver tissue specimens.

RESULTSNormal liver cell line L02 showed significantly lower UbcH10 mRNA expression levels than the cancer cell lines BEL-7402, Hep3B, HepG2 and SMMC-7721 (P<0.05). UbcH10 mRNA expression was also was significantly higher in hepatocellular carcinoma tissues than in the corresponding non-tumor tissues (P<0.05). Clinicopathological evaluation suggested that UbcH10 expression was associated with tumor invasion of the portal vein, tumor size, TNM staging, and tumor differentiation (P<0.05). Immunohistochemistry identified stronger UbcH10 expression in hepatocellular carcinoma tissues than in the adjacent tissues and normal liver tissues (68.6%, 28.6%, and 26.7%, respectively).

CONCLUSIONUbcH10 is over-expressed in hepatocellular carcinoma and may serve as a novel biomarker as well as a therapeutic target of hepatocellular carcinoma.

Adult ; Aged ; Biomarkers, Tumor ; Carcinoma, Hepatocellular ; metabolism ; pathology ; Female ; Hep G2 Cells ; Humans ; Liver Neoplasms ; metabolism ; pathology ; Male ; Middle Aged ; RNA, Messenger ; genetics ; metabolism ; Ubiquitin-Conjugating Enzymes ; genetics ; metabolism

Objective To study the clinical effect of heterogeneity (cattle) acellulur deunal matrix in repairing mucosa defect in laryngeal surgery. Methods Eighteen cancer patients with mucosa defect in central vocal area accepted treatment with heterogeneity acellular deunal matrix after surgery. There were two methods to repair mucosa defect. One was simple use of acellular deunal matrix, the second was combined use of acellular deunal matrix and muscle lamella or muscle and tendon film lameUa. 18 cases had cancer in central vocal area: T2N0M0(8), T3N1M0(5), T3N2M0(4), T4N2M0(1). All were squamous cell carcinoma. Ten cancer patients accepted radiation after surgery. The radiotherapy volume was 60 -80 Cry. After the operation, the patients were checked by fibrolaryngoscope four or five times after half a year, observing the dynamic development. Results All 18 patients were healed, rechecked by endoscope after 0. 5 -6 months, heterogeneity acellular deunal matrix mingled with mucosa within 30 -60 d, no allergy and irritation were found. The laryngeal function, including breathing, pronouncing and swallowing, was recovered. The survival rate (1 year) was 100%, and 10 patients survived after 2 years. After radiotherapy, the process of recovery was not affected. Conclusions Heterogeneity acellular deual matrix can be easily obtained and it is a new method to repair mucosa defect. The operative procedure is easy to perform and worthwhile to use clinically.

The study was aimed to investigate the biological behavior of stromal cell-derived factor-1 (SDF-1) in migration, adhesion and apoptosis as well as the related signaling transduction pathways in different kinds of acute myeloid leukemia (AML) cell lines. The expression of surface molecules on AML (KG1a, ML1 and U937) cells were analyzed by flow cytometry. The cell adhesion was detected by MTT assay. The cell migration was checked by transwell assay. Bcl-xl was checked by immunoblotting after activation of phosphionositide-3 kinase (PI3K) in AML cells treated with SDF-1. The results indicated that the expressions of the surface molecules on AML (KG1a, ML1 and U937) cells were different. The list of the expression showed CD34 (KG1a = 95.6%, ML1 = 4.6%, U937 = 4.8%), CD45 (KG1a = 98.3%, U937 = 97.5%, ML1 = 17.8%), CXCR4 (ML1 = 85.4%, U937 = 43.6%, KG1a = 3.8%), ICAM (KG1a = 75.8%, U937 = 41.8% and ML1 = 46.3%). SDF-1 could not upregulate their expression, but could trigger the establishment of polarized morphology of the cells which expressed CXCR4 high. SDF-1 promoted ML1 and U937 cell adhesion to the stroma cells (HS5, HS27), stimulated PI3K in the cells. It was also confirmed that SDF-1 could increase the leukemic cell survival by stimulate this pathway. After addition of wortmaninn or PTX, the cell death increased. It is concluded that the SDF-1 increases the leukemic cell adhesion, migration and survival by stimulating the PI3K pathway. These functions can be depressed by the PI3K inhibitor and also the inhibitor of G protein as well.
Apoptosis ; drug effects ; Cell Adhesion ; drug effects ; Cell Movement ; drug effects ; Chemokine CXCL12 ; physiology ; Humans ; Leukemia, Myeloid, Acute ; pathology ; Phosphatidylinositol 3-Kinases ; metabolism ; Signal Transduction ; Tumor Cells, Cultured ; U937 Cells

OBJECTIVETo detect the disease-causing point mutations in the dystrophin gene of Duchenne muscular dystrophy (DMD) patients.

METHODSThe approach of denaturing high performance liquid chromatography (DHPLC) coupling with sequencing was used to screen the point mutations of 79 exons and the untranslated regions of dystrophin gene without large deletions/duplications, which was in 6 unrelated DMD probands from 6 DMD families.

RESULTSFive disease-causing mutations, 697-698insGT, C616T, G1255T, C4279T, and C2302T, were ides created the new stop codons in downstream sites of mutations, respectively. In addition to the disease-causing point mutations, a point mutation T5586+61A in intron 39 was also found at patient 3, and a missense mutation A694T in exon 8 was detected at patient 5. Four point mutations, C2168+13T, 5740-13dupG, G5234A and C5280T, were also detected at patient 6 whose causative point mutation was unavailable. Seven point mutations have not been reported previously. Bi-directional PCR amplification of specific alleles (Bi-PASA) method was established to distinguish the haplotypes of heterozygote or homozygote in a single PCR reaction.

CONCLUSIONVia automated DHPLC screening or detecting the subexonic mutations in dystrophin gene is feasible to clinical laboratories, and also is a superior method in terms of sensitivity and efficiency.

Base Sequence ; Chromatography, High Pressure Liquid ; DNA Mutational Analysis ; Dystrophin ; genetics ; Gene Duplication ; Humans ; Male ; Muscular Dystrophy, Duchenne ; genetics ; Point Mutation ; Polymerase Chain Reaction ; Sequence Deletion

BACKGROUNDTalaromyces (Penicillium) marneffei (TM) is an emerging dimorphic human pathogenic fungus that is endemic to Southeast Asia. TM mostly occurs as an opportunistic infection in patients with human immunodeficiency virus (HIV). The objective of this study was to compare the clinical and laboratory parameters of patients with TM infections who were HIV-positive and HIV-negative and to assess therapies and outcomes.

METHODSThis was a retrospective analysis of 26 patients diagnosed with disseminated TM infection from September 2005 to April 2014 at Fujian Provincial Hospital, China.

RESULTSPatients with TM infection tend to present with fever, weight loss, and anemia. The time from symptom onset to confirmed diagnosis was greater for HIV-negative patients (n = 7; median: 60 days, range: 14-365 days) than for HIV-positive patients (n = 19; median: 30 days, range: 3-90 days, Mann-Whitney U = 31.50, P= 0.041). HIV-negative patients were more likely to have dyspnea (57.1% vs. 5.3%, χ2 = 8.86, P= 0.010), low neutrophil count (Mann-Whitney U = 27.00, P= 0.029), high CD4 count (Mann-Whitney U = 0.00, P= 0.009), and high lymphocyte count (Mann-Whitney U = 21.00, P= 0.009). There were no significant differences in other demographic, clinical, or biochemical characteristics. Among all the patients, 12 HIV-positive patient and 1 HIV-negative patient received amphotericin and fluconazole treatment, 9 of whom improved, 1 died, 2 had kidney damage, 1 had hypokalemia due to exceeded doses.

CONCLUSIONSHIV-negative patients with TM infections tend to have a longer diagnostic interval, a higher percentage of dyspnea, higher levels of CD4 and lymphocytes, and lower neutrophil counts than TM infection in HIV-positive patients. Treatment programs with amphotericin and fluconazole are mostly effective.

Adult ; Aged ; CD4 Lymphocyte Count ; Female ; HIV Infections ; complications ; Humans ; Male ; Middle Aged ; Mycoses ; diagnosis ; drug therapy ; immunology ; Retrospective Studies ; Talaromyces ; drug effects