OBJECTIVETo develop an assay methodology for determination of lumbrukinase potency in Rongshuan capsules.

METHODThe agarose-fibrin plate assay methodology for determination of Lumbrukinase potency in Rongshuan capsules was studied including durability, specificity, linearity range, product's handling method, accuracy , repetitiveness, solution stability, recovery and statistical method. The method of parallel line assay based on quantitative responses in statistical methods for biological assays was used in the statistics of potency assay.

RESULTThe durability and specificity of assay accord with the requirement; The linearity range was 12.5 to approximately 400 U, the RSD of accuracy tests was 3.2%, the RSD of repetitiveness tests was 8.3%, the solution is stable under 4 degrees C for 72 hours, the recovery rate was 97.0% and the RSD of recovery assays was 16.5%.

CONCLUSIONThe agar-fibrin plate assay is rapidly, feasible, simple, convenient and accurate way for determining the Lumbrukinase potency. The method of parallel line assay based on quantitative responses in statistical methods for biological assays can control the error of determination.

Animals ; Biological Assay ; methods ; Capsules ; analysis ; Endopeptidases ; analysis ; Pharmaceutical Preparations ; analysis

From the culture filtrates of C. albicans, C. tropicalis and C. parapsilosis, proteinases were purified using a series of chromatographic steps consisting of DEAE-Sepharose, Sephacryl S-200 and size-exclusion HPLC which removed contaminating mannoproteins and extraneous proteins. Anti-Candida proteinase antibodies in sera from mice infected with various Candida species were detected using ELISA for serodiagnosis of candidiasis. Three proteinases were blotted by homologous and heterologous anti-proteinase antisera on Western blot analysis. All sera from six Candida species-infected mice were reactive with proteinases of C. albicans, C. tropicalis, and C. parapsilosis, although C. glabrata, C. guilliermondii, and C. krusei did not secrete proteinase. The seroreactivities of proteinase with sera from mice infected with homologous C. albicans and C. tropicalis were higher than those with sera from heterologous Candida species-infected mice. These results suggest that three proteinases have at least one common epitope, but its application for diagnosis of candidiasis should be considered with limits of specificity.
Animal ; Candida/genetics* ; Candida/enzymology* ; Candidiasis/enzymology* ; Endopeptidases/analysis* ; Female ; Mice ; Mice, Inbred ICR ; Species Specificity

OBJECTIVETo study the correlation of nucleotide polymorphism in the ubiquitin-specific protease 26 (Usp26) gene with idiopathic male infertility and its action mechanism in spermatogenesis.

METHODSBased on the WHO criteria (4th ed.), we selected 41 patients with idiopathic infertility from 150 infertile males, and enlisted 50 normal fertile men as controls. We examined the selected patients for mutations using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analysis, and determined how and where the mutations occurred by gene sequencing.

RESULTSLow sperm concentration and poor sperm motility were found in the 41 men with idiopathic infertility. Nine (22.0%) of them exhibited changes in the Usp26 gene (P = 0.01), including compound mutations of 364insACA and 460G > A in 8 (19.5%, P = 0.01) and 1 044T > A substitution in 1 (2.4%, P > 0.05). The above three variations led to changes in the coding amino acids. No other changes were found in the remaining patients and normal fertile controls.

CONCLUSIONThe nucleotide polymorphisms of the Usp26 gene might be closely related with idiopathic male infertility, and exert negative effect on the testis function.

Case-Control Studies ; Cysteine Endopeptidases ; genetics ; Humans ; Infertility, Male ; genetics ; pathology ; Male ; Polymorphism, Single Nucleotide ; Semen Analysis

Dendritic Cells ; immunology ; Granzymes ; Humans ; Immune Tolerance ; Killer Cells, Natural ; immunology ; Multivariate Analysis ; S100 Proteins ; analysis ; Serine Endopeptidases ; analysis ; Stomach Neoplasms ; immunology ; pathology ; T-Lymphocytes, Cytotoxic ; immunology

OBJECTIVE: Our previous study revealed many genes differentially expressed in human laryngeal squamous cell carcinoma tissues (LSCC) and related adjacent normal tissues. We verificated the differentiated expressions of target genes, possibly related to LSCC. And these results can be foundations of further study of these target genes function. METHOD: SENP1, CD109, Laminin alpha 2, Laminin alpha 3 were selected according to the cDNA microarray results. The expression of these genes mRNA and protein were detected by RT-PCR and Western blot in 12 cases of LSCC and related adjacent normal tissues. RESULT: The mRNA expression of SENP1, CD109 and Laminin alpha 3 were significantly higher while Laminin alpha 2 were significantly lower in LSCC tissues than in corresponding adjacent normal tissues by Semiquantitative RT-PCR. Western blot analysis revealed SENP1, CD109 protein expression were significantly higher in LSCC tissues than in corresponding adjacent normal tissues. CONCLUSION SENP1, CD109, Laminin alpha 2 and Laminin alpha 3 may correlated with tumorigenesis and development of LSCC and can provide beneficial clue for study pathogenesis of LSCC in molecular level.
Aged ; Aged, 80 and over ; Antigens, CD ; genetics ; Carcinoma, Squamous Cell ; genetics ; Cysteine Endopeptidases ; Endopeptidases ; genetics ; GPI-Linked Proteins ; genetics ; Gene Expression Regulation, Neoplastic ; Humans ; Laminin ; genetics ; Laryngeal Neoplasms ; genetics ; Male ; Middle Aged ; Neoplasm Proteins ; genetics ; Oligonucleotide Array Sequence Analysis

OBJECTIVETo explore the protective effects of mitochondrial ATP-sensitive potassium channel opener diazoxide on hyperoxia-induced apoptosis of type II alveolar epithelial cells (A549 cells) and possible mechanisms.

METHODSA549 cells were cultured in vitro and divided randomly into control, hyperoxia and diazoxide group. The hyperoxia group was exposed to a mixture of O2 (900 mL/L) and CO2 (50 mL/L) for 10 minutes, then cultured in a closed environment. The diazoxide group was pretreated with diazoxide of 100 μmol/L for 24 hrs before hyperxia induction. The cells were collected 12, 24 and 48 hrs after culture. The morphologic changes of A549 cells were observed under an inverted microscope. A549 cell apoptosis was detected by flow cytometry. The expression of Omi/HtrA2 in the endochylema of A549 cells was determined by immunohistochemistry.

RESULTSA549 cells were damaged and the changes in morphology of the cells were serious in the hyperoxia group. The apoptosis rate of A549 cells and the expression of Omi/HtrA2 in the endochylema increased in the hyperoxia group compared with the control group (P<0.05). The growth and the morphology of A549 cells were greatly improved and the cell injuries were obviously alleviated in the diazoxide group. The expression of Omi/HtrA2 in the endochylema and the apoptosis rate of A549 cells were significantly reduced in the diazoxide group compared with the hyperoxia group (P<0.05).

CONCLUSIONSDiazoxide as an opener of mitoKATP channel can reduce the expression of Omi/HtrA2 and the apoptosis rate of A549 cells, thus relieves the injury of A549 cells induced by hyperoxia.

Apoptosis ; Cells, Cultured ; Cytoprotection ; Diazoxide ; pharmacology ; High-Temperature Requirement A Serine Peptidase 2 ; Humans ; Hyperoxia ; complications ; Lung ; pathology ; Mitochondrial Proteins ; analysis ; Potassium Channels ; physiology ; Serine Endopeptidases ; analysis

OBJECTIVE: To observe the household environment dust mites allergens content distribution characteristics and influence factors of children with allergic rhinitis to dust mites in Wuhu. METHOD: Collect the surface dust in bedroom and living room floor, mattresses, pillows, sofa of 102 children with allergic rhinitis families. Dust mite allergen components 1 (Der f1) and house dust mites allergens 1 components (Der p1) of the dust samples were determined by enzyme-linked immunosorbent assay (ELISA). RESULT: One hundred and twenty samples were collected . In a domestic dust mites samples, with a median of M (Min and Max) said dust mite allergen levels, Der f1 and Der p1 content was 2.66 (0.03, 26.63), 3.48 (0, 03, 33.68), respectively. Der f1 was significantly less than Der p1, and the difference was statistically significant (P < 0.05). According to the classification of urban and township, there were 68 cases and 34 cases. Der f1 content in the samples was 2.91 (0.31, 26.63), 2.40 (0.08, 16.02), respectively. Der p1 content was 4.28 (0.03, 20.77), 3.88 (0.14, 33.68), respectively. The dust mites content of urban was significantly more than that of villages and towns, the difference was statistically significant (P < 0.05). Samples were also classified by gender. The dust mites allergens content in either boy's or girl's family were similar, there was no statistically significant difference (P > 0.05). CONCLUSION The household dust mites of children allergic rhinitisin Wuhu area is given priority to with Der p1, and urban dust mites are significantly more than village's and town's. Enhancing health education, controlling dust mites allergens contamination inside the bedroom, especially urban areas, are positive differences for the prevention and treatment of allergic diseases such as allergic rhinitis in children.
Allergens ; analysis ; Animals ; Antigens, Dermatophagoides ; analysis ; Arthropod Proteins ; analysis ; Bedding and Linens ; Child ; Cysteine Endopeptidases ; analysis ; Dermatophagoides farinae ; Dust ; analysis ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Male ; Rhinitis, Allergic ; epidemiology

Although active inflammation may be deleterious and indicate immunologic activation in chronically rejected grafts, the underlying mechanism of tissue destruction has been little studied. Twenty-four cases of chronic rejection (CR) with or without acute rejection (AR) were stained with antibodies against CD3, CD8, CD68, granzyme B and TIA-1, and the number of positive cells were counted. Eleven cases of AR served as controls. The number of CD3 and CD8 positive cells increased in the acute on CR group compared to the CR group. About a half of CD3 positive T cells were CD8 positive in both groups, however, the proportion of TIA-1 or granzyme B positive cells was higher in the acute on CR group. The numbers of CD3, CD68, granzyme B and TIA-1 positive cells were higher in the AR group than the acute on CR group, however, no significant difference was found between the two groups. Serum creatinine level and proteinuria at the time of biopsy and the percentages of late onset AR and graft failure rate were higher in the acute on CR group than the CR group. Summarizing, these results suggest that infiltration of activated T cells containing cytotoxic granules plays a role in graft destruction in acute on CR.
Adult ; Antigens, CD3/analysis ; Antigens, CD8/analysis ; Female ; Follow-Up Studies ; *Graft Rejection ; Human ; Immunohistochemistry ; *Kidney Transplantation ; Male ; Membrane Proteins/*analysis ; RNA-Binding Proteins/*analysis ; Serine Endopeptidases/*metabolism ; Transplantation, Homologous

BACKGROUNDHaikou locates in tropical island with unique mite propagation. The aim of this stuy is to determine mite allergens levels in Haikou, and to investigate the prevalence of mite specific IgE-sensitization and IgE cross-reactivity between house dust mites.

METHODSAllergen and antigen concentrations against six mite species were tested by enzyme-linked immunosorbent assay (ELISA). Specific IgE concentrations and cross-inhibitions were measured with ADVIA Centaur(®).

RESULTSAllergen or antigen Dermatophagoides pteronyssinus (Der p 1), Blomia tropicalis (Blo t) and Tyrophagus putrescentia (Tyr p) were detected in dust samples. Dermatophagoides farinae (Der f 1), Lepidoglyphus destructor (Lep d 2), and Acarus siro (Aca s) were found in very few samples. Specific IgE tests showed high prevalence of sensitizations against all tested mites with high IgE levels to Der p, Der f, and Blo t. Storage mites, Blo t, Tyr p, Lep d, and Aca s, could inhibit Der p from 0 to 50%. Storage mites could inhibit Der f between 30% and 100%. Der p IgE could be inhibited by Der f with up to 90%, and vice versa. Der p could inhibit Blo t from 40% to 80%. Blo t was able to fully inhibit IgE binding to Lep d, Tyr p, and Aca s compared to partial inhibition by Der p.

CONCLUSIONSDer p is the dominating mite and has the highest specific IgE prevalence among asthmatic children. Blo t represents an important source of storage mite sensitization and some patients may be independently sensitized to both Der p and Blo t. High prevalence of sensitization to Der f may be due to IgE-mediated cross-reactivity with Der p and Blo t.

Adolescent ; Air Pollution, Indoor ; Allergens ; analysis ; Animals ; Antigens, Dermatophagoides ; analysis ; Arthropod Proteins ; analysis ; Asthma ; immunology ; Child ; Child, Preschool ; China ; Cross Reactions ; Cysteine Endopeptidases ; analysis ; Dust ; Humans ; Immunoglobulin E ; blood ; immunology ; Mites ; immunology

OBJECTIVETo study the effect of imbalance between lysosomal enzymes and their inhibitors in blood on disturbance of the local and whole body after trauma.

METHODSThe dynamic changes of lysosomal enzymes and proteinase inhibitors were studied in 12 pigs with femoral comminuted fractures in both hind limbs caused by high velocity missiles. Four normal pigs served as controls.

RESULTSAfter injury, the activity of Cathepsin D in arterial plasma increased gradually and reached the highest level at 8 hours, acid phosphatase in serum began to increase at 12 hours and the value of serum elastase did not change significantly. The level of alpha1-antitrypsin, a proteinase inhibitor in plasma, decreased significantly in the early stage after injury [73.5%+/-6.4% and 81.0%+/-5.1% of the baseline value (1.67 micromol x ml(-1) x min(-1)+/- 0.29 micromol x ml(-1) x min(-1)) at l and 2 hours after injury, respectively, P<0.05], then increased gradually and was higher than the baseline value at 12 hours after injury.

CONCLUSIONSImbalance between lysosomal enzymes and proteinase inhibitors occurs soon after injury, which might result in continuous tissue damage and play an important role in the disturbance of general reaction after injury.

Acid Phosphatase ; blood ; Animals ; Cathepsin D ; blood ; Endopeptidases ; blood ; Female ; Lysosomes ; enzymology ; Male ; Pancreatic Elastase ; blood ; Swine ; Wounds, Gunshot ; blood ; alpha 1-Antitrypsin ; analysis