OBJECTIVETo elucidate how chronic prostatitis affects the expression and activity of the plasminogen activator (PA) system and relates to male infertility.

METHODSTwenty-three normal fertile males and 80 chronic prostatitis patients (40 fertile and 40 infertile) were included in this research. SDS polyacrylamide gel electrophoresis and fibrin overlay method were used to estimate the total PA, and tissue PA (tPA), urokinase type PA (uPA) in semen.

RESULTSTotal PA, tPA and uPA highly expressed in normal males, but decreased in the semen of the chronic prostatitis patients of both the fertile and infertile groups. However, there was no significant difference in total PA between the fertile and infertile patients.

CONCLUSIONChronic prostatitis reduces the secretory function and PA synthesis and secretion of the prostate, but the decrease of PA alone does not cause infertility. PA may be one of the tools for estimating the function of the prostate.

Adult ; Case-Control Studies ; Chronic Disease ; Humans ; Infertility, Male ; metabolism ; Male ; Prostatitis ; metabolism ; Semen ; metabolism ; Tissue Plasminogen Activator ; biosynthesis ; Urokinase-Type Plasminogen Activator ; biosynthesis

Actins ; biosynthesis ; genetics ; Adult ; Aged ; Female ; Hepatitis B, Chronic ; complications ; Humans ; Liver Cirrhosis ; blood ; enzymology ; virology ; Male ; Matrix Metalloproteinase 1 ; biosynthesis ; genetics ; Middle Aged ; Plasminogen Activator Inhibitor 1 ; blood ; Tissue Inhibitor of Metalloproteinase-1 ; biosynthesis ; genetics ; Urokinase-Type Plasminogen Activator ; blood

Taking a suspension adapted recombinant CHO cell line, 11G-S expressing human Pro-urokinase (Pro-UK) as the object of study, the impacts of different feeding nutrients, the start time of feeding and cell inoculation density on the growth and Pro-UK production of 11G-S cells in serum-free fed-batch culture were evaluated in 100 mL shacking flasks. The results indicated that amino acids, serum-free supplements and inorganic salts played important role in cell growth, cell viability and protein expression. And the effects of cells fed-batch culture was much better with the initial cell inoculation density at 3 x 10(5)-4 x 10(5) cells/mL and the start time of feeding set at 72 h, a maximum viable cells density of 7.8 x 10(6) cells/mL with a peak Pro-UK activity at 8570 IU/mL was achieved during 12 d fed-batch culture. Further, the mu of the 11G-S cells at the middle phase of the fed-batch culture, and both the q(glu) and q(gln) of the 11G-S cells at the middle and later phases of the fed-batch culture was higher than that of the 11G-S cells at the same phase of the batch culture, respectively.
Animals ; CHO Cells ; Cell Culture Techniques ; methods ; Cricetinae ; Cricetulus ; Culture Media, Serum-Free ; Humans ; Recombinant Proteins ; biosynthesis ; genetics ; metabolism ; Urokinase-Type Plasminogen Activator ; biosynthesis ; genetics ; metabolism

By using the cell density, cell viability, Pro-UK activity, specific consumption rate of glucose (q(glc)), specific production rate of lactate (q(lac)), yield of lactate to glucose (Y(lac/glc)) and as the evaluation indexes, the growth and metabolism characteristics of pro-urokinase (Pro-UK) expressing CHO cells in serum-free suspension batch culture were examined and compared to those in serum-containing suspension batch culture. We observed hardly differences in growth and metabolism characteristics between the CHO cell populations grown in serum-free suspension batch culture and serum-containing suspension batch culture. The optimal mathematical model parameters for the CHO cells grown in suspension batch culture were obtained by non-linear programming of data representing the growth, substrate consumption and product formation of the CHO cells during logarithmic growth phase using MATLAB software, and the kinetic model of the cell growth and metabolism in serum-free culture were established.
Animals ; Bioreactors ; CHO Cells ; Cricetinae ; Cricetulus ; Culture Media, Serum-Free ; Culture Techniques ; methods ; Kinetics ; Models, Theoretical ; Recombinant Proteins ; biosynthesis ; genetics ; metabolism ; Urokinase-Type Plasminogen Activator ; biosynthesis ; genetics ; metabolism

OBJECTIVETo investigate the content and expression of the urokinase plasminogen activator (uPA) in the ornidazole-induced asthenospermia animal model, and to probe the mechanism of ornidazole inducing asthenospermia and the possibility of using uPA for the prevention and treatment of asthenospermia.

METHODSForty-eight male rats were equally randomized into 5 medication groups (1 d, 5 d, 10 d, 15 d and 20 d) and a blank control group, and ornidazole (200 mg/kg) was given intragastrically every day to the former five while 0.5% carboxymethylcellulose Na (CMC-Na) to the latter for 20 successive days. Then the rats were sacrificed by intraperitoneal injection of pentobarbital at 1, 5, 10, 15 and 20 days respectively and the epididymides and testes harvested. The integrity of the sperm cell membrane was detected by hypoosmotic swelling experiments, the uPA expression in the testicular and epididymal tissues dynamically observed by immunohistochemistry and the level of uPA mRNA in the testis determined by RT-PCR.

RESULTSThe integrity of the sperm cell membrane was reduced at 10 days and remained low till the end of the medication, but with no statistic significance. Compared with the blank controls, the uPA expression and mRNA content in the testicular and epididymal tissues showed no conspicuous difference in the 1 d and 5 d groups, decreased insignificantly in the 10 d group, but significantly in the 15 d and 20 d groups (P < 0.05).

CONCLUSIONThe defect of sperm cell membrane and decrease of sperm motility go in parallel with the reduced expression and content of uPA, which may be one of the factors for the development of asthenospermia.

Animals ; Asthenozoospermia ; metabolism ; Male ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Sperm Motility ; Spermatozoa ; cytology ; Urokinase-Type Plasminogen Activator ; biosynthesis ; metabolism

OBJECTIVEThe aim of this study was to reveal the relations between expression and tumor behavior such as invasion, metastasis and prognosis in oral squamous cell carcinoma (SCC) through analyzing the expression of urokinase-type plasminogen activator (UPA).

METHODSWith Labelled streptavidin biotin method (LsAB), the expression of UPA was analyzed in 80 cases of oral squamous cell carcinoma, 10 cases of oral normal mucosa and 10 cases of oral leukoplakia.

RESULTSCompared to oral normal mucosa and oral leukoplakia, the expression of UPA was significantly higher in SCC. The expression of UPA in SCC cases with lymph node metastasis was significantly higher than in cases without metastasis. The expression in cases with good prognosis was significantly higher than in those with poor prognosis and the expression was significantly higher in cases with invasive growth than in those with ulcerative and papillary growth.

CONCLUSIONThe results obtained indicated that high expression of UPA in SCC might be closely related to lymph node metastasis, invasive growth and poor prognosis.

Carcinoma, Squamous Cell ; metabolism ; pathology ; Humans ; Leukoplakia, Oral ; metabolism ; Lymphatic Metastasis ; Mouth Mucosa ; metabolism ; Mouth Neoplasms ; metabolism ; pathology ; Neoplasm Invasiveness ; Prognosis ; Urokinase-Type Plasminogen Activator ; biosynthesis

Natural hirudin extracted from the secretion of medical leech salivary gland is a single-chain peptide containing 65 aminoacid residues with molecular weight of 7000 D, and exists in three isomers of HV1, HV2 and HV3. Hirudin possesses three disulfide bridges forming the structure of core cyclic peptides, which binds to the catalytic site of thrombin so as to inhibit the catalysis of thrombin. Its c-terminus rich in acidic aminoacid residues possesses hydrophilicity, and is free on the molecular surface, and can bind with fibrin recognition site of hirudin. The minimal segment of 12 - 16 C-terminal acidic residues keeps the minimal activity of anti-thrombosis. Thus, hirudin, as a potent and specific inhibitor of thrombin, can be used to protect from and to treat clinically thrombosis. As it has some disadvantages such as short half-life, bleeding side-effect and mono-function, and so on, hirudin has been fused with some other functional proteins in recent years. The obtained fusion proteins can prolong the half life of hirudin, or relieve it bleeding side effect, or bring new functions, such as thrombolysis, inhibiting the platelet aggregation, targeting specifically. The research progress in hirudin fusion protein was summarized in this review.
Anticoagulants ; pharmacology ; Delayed-Action Preparations ; Drug Delivery Systems ; Glucokinase ; biosynthesis ; genetics ; pharmacology ; Hirudins ; biosynthesis ; genetics ; pharmacology ; Humans ; Platelet Aggregation Inhibitors ; pharmacology ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacology ; Urokinase-Type Plasminogen Activator ; biosynthesis ; genetics ; pharmacology

It is very easy for the pro-UK to lose it's biological activity because of the digestion of pro-UK by the thrombin or the inhibition of pro-UK by the PAI-I. So three pro-UK mutant (pro-UK) genes were constructed in this experiment with the PCR point-mutant method. The thrombin cleavage site Arg156 in pro-UK was mutated into His156, and named as pro-UKM1; PAI binding sites Arg178, Arg179, Arg181 were mutated into Lys178, Lys179, His181, named as pro-UKM2; The mutant containing His156, Lys178, Lys179, His181 as pro-UKM3. Three mutants were expressed in CHO cells respectively and analyzed with SDS-PAGE fibrin plate assay and western blot. The results showed that the three mutants and the native pro-UK have the same single electrophoresis band indicating most of the pro-UK was single chain. In vitro plasma clot lysis assays indicated that the pro-UKM1 have the ability to resistant against thrombin digestion; pro-UK2 could resist against PAI inhibition; while pro-UK3 improved resistances against both thrombin and PAI. It looks very promising that the pro-UK3 can be a new medicine of dissolving thrombus.
Animals ; Base Sequence ; Blotting, Western ; CHO Cells ; Cloning, Molecular ; Cricetinae ; Cricetulus ; Electrophoresis, Polyacrylamide Gel ; Humans ; Molecular Sequence Data ; Mutant Proteins ; biosynthesis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Transfection ; Urokinase-Type Plasminogen Activator ; biosynthesis ; genetics

To evaluate the specific inhibition of antisense u-PAR on the u-PAR expressions in highly invasive cell subclones and to determine its blocking function in the invasion by those cells, a cDNA fragment of u-PAR obtained by RT-PCR was inserted into a plasmid vector named pcDNA3 in antisense orientation. Then the antisense u-PAR recombinant was transfected into highly invasive cell subclones. The u-PAR expression in neo-resistant cells was examined by RT-PCR and immunohistochemical assay. Compared to the control cells, the content of mRNA and protein of u-PAR in transfected cells decreased sharply, and the rate of inhibition was 53% and 73%, respectively, indicating that an antisense u-PAR might have played a specific inhibitory role in its expression in the cells, which may provide a good cell model for making further investigation of the inhibitory effects of the antisense u-PAR on invasion in highly invasive cell subclones of human prostate carcinoma.
Cell Line, Tumor ; Cloning, Molecular ; Humans ; Male ; Neoplasm Invasiveness ; Plasmids ; Prostatic Neoplasms ; metabolism ; pathology ; RNA, Antisense ; biosynthesis ; genetics ; Receptors, Cell Surface ; biosynthesis ; genetics ; Receptors, Urokinase Plasminogen Activator ; Reverse Transcriptase Polymerase Chain Reaction ; Transfection ; Urokinase-Type Plasminogen Activator ; antagonists & inhibitors ; genetics ; metabolism

OBJECTIVETo explore curative machanism of Shenle capsule on the 5/6 nephrectomy rats.

METHODFibrin plate method was applied to examine activity of urinary plasminogen activator(PA). Semi-quantitative analysis was used to observe stained intensity and area of tissue-type plasminogen activator, urokinas-type plasminogen activator/ plasminogen activator inhibitor(tPA, uPA/PAI-1)in remnant renal tissue. Northern blot was employed to analyze the expression of transforming growth factor (TGF-beta) mRNA.

RESULTIn model control group, the urinary PA activity and protein expression of tPA, uPA were down-regulated, but protein expression of PAI-1, TGF-beta mRNA was up-regulated in remnant renal tissue. In each treated group, the urinary PA activity and protein expression of tPA/uPA were enhanced,but the protein expression of PAI-1, TGF-beta mRNA decreased simultaneously.

CONCLUSIONShenle capsule can delay glomerulosclersis and tubulointerstitial fibrotic lesions of remnant kidney by improving the activity of urinary PA and modulating the expression of tPA, uPA/PAI-1 and TGF-beta mRNA.

Animals ; Capsules ; Codonopsis ; chemistry ; Drug Combinations ; Drugs, Chinese Herbal ; isolation & purification ; pharmacology ; Female ; Kidney ; metabolism ; Kidney Failure, Chronic ; drug therapy ; metabolism ; Leeches ; chemistry ; Male ; Materia Medica ; isolation & purification ; pharmacology ; Nephrectomy ; Plasminogen Inactivators ; metabolism ; Polyporales ; chemistry ; RNA, Messenger ; biosynthesis ; Rats ; Rats, Sprague-Dawley ; Tissue Plasminogen Activator ; metabolism ; Transforming Growth Factor beta ; biosynthesis ; genetics ; Urokinase-Type Plasminogen Activator ; metabolism