Ulinastatin attenuates oxidation, inflammation and neural apoptosis in the cerebral cortex of adult rats with ventricular fibrillation after cardiopulmonary resuscitation

OBJECTIVE: The role of Ulinastatin in neuronal injury after cardiopulmonary resuscitation has not been elucidated. We aim to evaluate the effects of Ulinastatin on inflammation, oxidation, and neuronal injury in the cerebral cortex after cardiopulmonary resuscitation. METHODS: Ventricular fibrillation was induced in 76 adult male Wistar rats for 6 min, after which cardiopulmonary resuscitation was initiated. After spontaneous circulation returned, the rats were split into two groups: the Ulinastatin 100,000 unit/kg group or the PBS-treated control group. Blood and cerebral cortex samples were obtained and compared at 2, 4, and 8 h after return of spontaneous circulation. The protein levels of tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6) were assayed using an enzyme-linked immunosorbent assay, and mRNA levels were quantified via real-time polymerase chain reaction. Myeloperoxidase and Malondialdehyde were measured by spectrophotometry. The translocation of nuclear factor-κB p65 was assayed by Western blot. The viable and apoptotic neurons were detected by Nissl and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL). RESULTS: Ulinastatin treatment decreased plasma levels of TNF-α and IL-6, expression of mRNA, and Myeloperoxidase and Malondialdehyde in the cerebral cortex. In addition, Ulinastatin attenuated the translocation of nuclear factor-κB p65 at 2, 4, and 8 hours after the return of spontaneous circulation. Ulinastatin increased the number of living neurons and decreased TUNEL-positive neuron numbers in the cortex at 72 h after the return of spontaneous circulation. CONCLUSIONS: Ulinastatin preserved neuronal survival and inhibited neuron apoptosis after the return of spontaneous circulation in Wistar rats via attenuation of the oxidative stress response and translocation of nuclear factor-κB p65 in the cortex. In addition, Ulinastatin decreased the production of TNF-α, ...

Effect of urinary trypsin inhibitor on DNA genotyping in urine samples / 法医学杂志

OBJECTIVE@#To study the effect of urinary trypsin inhibitor (UTI) on STR genotyping with urinary samples.@*METHODS@#Midstream urine samples of 5 male and 5 female volunteers were collected respectively, sub-packaged, added with different concentration of UTI and stored at -80 degrees C. Genomic DNA was extracted from those urinary samples, of which STR profiles were genotyped with IdentifilerTM kit at 8 different time points. Results of genotyping in urinary samples were compared with those of the homogenous blood control samples and the successful rate of genotyping in different group of urinary samples treated with UTI was determined.@*RESULTS@#Fifteen STR loci included in Identifiler system were all detected in control blood samples and urinary samples stored for 1 day. STR locus loss was observed and all 15 STR loci disappeared in female urinary samples untreated with UTI while those storage periods prolonged to 3 and 9 days, respectively. However, all 15 STR loci could be detected in female urinary samples treated with UTI and stored for as long as 9 days. No STR loci could be detected in male urinary samples preserved without UTI for 7 days while 9 STR loci detected preserved with UTI for 9 days. There was no significant difference among the average detection ratios of STR loci in female urinary samples treated with UTI at concentrations of 0.2, 0.4 or 0.6 microg/mL and stored for 30 days, mean of which was as high as 0.8400 +/- 0.0423, statistically higher than that in male urinary samples (0.1600 +/- 0.0423).@*CONCLUSION@#Detection rate of STR loci in urinary samples preserved with UTI was increased significantly, which results in prolonging the storage periods of urinary samples for personal identification.

Soybean trypsin inhibitor confers protection against rotavirus infection in infant mice.

OBJECTIVE: To study the effects of soybean trypsin inhibitor (TI) on glycine uptake, glutathione (GSH) levels and morphological changes of intestine in rotavirus (RV) infected infant mice. METHODS: A total of 144 infant mice (7/8 days old) were divided in 3 groups (i.e. control, RV and RV + inhibitor). Infant mice were orally inoculated with the EB strain of RV and Trypsin protease inhibitor (TI) and 8 animals each were sacrificed on days 0,1,3,5,7 and 10 post infection (p.i). Glycine uptake (in vitro), GSH levels and histological changes were assessed in the jejunum, ileum and colon. RESULTS: Glycine uptake and GSH levels were significantly reduced on days 3 and 5 p.i in jejunum and ileum of RV inoculated animals, compared to the controls. Glycine uptake and GSH levels were maintained as in controls in the RV + TI inoculated animals on days 3 and 5 p.i in jejunum and colon but not in ileum where lesser values were recorded. Histology showed vacuolar degeneration in ileum towards the apical portion whereas normal morphology was observed in jejunum, similar to controls. No histological changes were observed in colon in any of the groups. Electron microscopic study confirmed the viral infection. CONCLUSION: Administration of Trypsin protease inhibitor along with RV reverted the effects of RV infection on amino acid uptake and GSH levels completely in the jejunum and partially in the ileum.

Protection against rotavirus diarrhoea in mice by trypsin inhibitor.

To investigate the role of soyabean trypsin inhibitor (TI) during rotavirus (RV) diarrhoea, changes in enzyme activities of six relevant mucosal enzymes (lactase, sucrase, maltase, trehalase, glucoamylase and alkaline phosphatase) were assayed following inoculation of suckling mice with EB rotavirus (serotype 3) along with the TI and compared with the age-matched healthy control mice. The animals were divided into three groups i.e. group 1 (controls), group 2 (RV inoculated) and group 3 (RV + TI inoculated and sacrificed under light anaesthesia on 0, 1, 3, 5, 7 and 10 day post inoculation (dpi). Then intestines were excised and divided into two parts (jejunum and ileum). They were separately homogenized in 0.9% cold normal saline and activities of mucosal enzyme were measured. Alkaline phosphatase and disaccharidases were found to be decreased significantly in RV inoculated animals in both the anatomical portions of small intestine of mice. These enzyme levels were restored with the administration of TI i.e. in group 3 and became comparable to the controls in both intestinal portions. These studies suggest that activity of intestinal enzymes which are important in digestive absorptive functions of small intestine were restored with the addition of TI whengiven to infant mice showing its protective efficacy during rotavirus infection.

Linear competitive inhibition of human tissue kallikrein by 4-aminobenzamidine and benzamidine and linear mixed inhibition by 4-nitroaniline and aniline

Hydrolysis of D-valyl-L-leucyl-L-arginine p-nitroanilide (7.5-90.0 `M) by human tissue kallikrein (hK1) (4.58-5.27 nM) at pH 9.0 and 37ºC was studied in the absence and in the presence of increasing concentrations of 4-aminobenzamidine (96-576 `M), benzamidine (1.27-7.62 mM), 4-nitroaniline (16.5-66 `M) and aniline (20-50 mM). The kinetic parameters determined in the absence of inhibitors were: Km = 12.0 + or - 0.8 `M and k cat = 48.4 + or - 1.0 min-1. The data indicate that the inhibition of hK1 by 4-aminobenzamidine and benzamidine is linear competitive, while the inhibition by 4-nitroaniline and aniline is linear mixed, with the inhibitor being able to bind both to the free enzyme with a dissociation constant Ki yielding an EI complex, and to the ES complex with a dissociation constant Ki', yielding an ESI complex. The calculated Ki values for 4-aminobenzamidine, benzamidine, 4-nitroaniline and aniline were 146 + or - 10, 1,098 + or - 91, 38.6 + or - 5.2 and 37,340 + or - 5,400 `M, respectively. The calculated Ki' values for 4-nitroaniline and aniline were 289.3 + or - 92.8 and 310,500 + or - 38,600 `M, respectively. The fact that Ki'>Ki indicates that 4-nitroaniline and aniline bind to a second binding site in the enzyme with lower affinity than they bind to the active site. The data about the inhibition of hK1 by 4-aminobenzamidine and benzamidine help to explain previous observations that esters, anilides or chloromethyl ketone derivatives of Nalpha-substituted arginine are more sensitive substrates or inhibitors of hK1 than the corresponding lysine compounds

Reversal of T-cell unresponsiveness through serine-esterase inhibitors mediated enhanced lymphokine induced microbicidal activities in kala-azar.

After presenting processed glycoprotein of Leishmania donovani to T-cell, macrophage seeks the help of a panel of T-cells lymphokines to transform from a state that sustains intra cellular replication of parasite to an effector state for destructing parasites. But esterase and trypsin of macrophage membrane prevent T-cells to release MIF. Role of soya-bean trypsin inhibitor (STI) has been exposed in the present study with a view to alter esterase functional behaviour of macrophage for control of T-cell activation and also, if T-cells once made responsive to antigen by STI do alter macrophage response to T-cells or not. Results establish STI as potent effector molecule, which can serve as an adjuvant to candidate T-cell epitope and synthetic peptide for development of anti-Kala-azar vaccine protocol in future.

Purification and preliminary characterization of Torresea cearensis trypsin inhibitor

An inhibitor against serine proteinases was purified from Torresea cearensis by affinity chromatography on trypsin-Sepharose. The protein is a single polypeptide of molecular weight 13,600 after reduction and has a high content of cysteine residues. Both trypsin (Ki = 0.34 nM) and chymotrypsin (Ki = 0.15 micronM) are inhibited by Torresea cearensis inhibitor. Blood clotting factor XII is also inhibited (Ki = 24 micronM), but not plasma kallikrein, tissue kallikrein or thrombin. The stoichiometry of the inhibitorproteinase complex with trypsin is 1:1