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101.
Koistinen H Soini T Leinonen J Hyden-Granskog C Salo J Halttunen M Stenman UH Seppälä M Koistinen R 《Biology of reproduction》2002,66(3):624-628
Semenogelin plays an important role in sperm clotting and is degraded into smaller fragments by prostate-specific antigen (PSA) during clot liquefaction. Semenogelin and its fragments inhibit sperm motility in vitro. We studied the expression of semenogelin I mRNA and its localization in various tissues of the male genital tract. We also studied semenogelin concentrations with respect to sperm parameters and the outcome of in vitro fertilization. Semenogelin protein was detected by immunohistochemical staining and semenogelin I mRNA was detected by Northern blot analysis in the seminal vesicles and ampullary part of the vas deferens, whereas specimens from the prostate, epididymis, testis, and the female genital tract were negative. Using monoclonal antibodies against semenogelin, an immunofluorometric assay was developed to measure semenogelin levels in seminal plasma and to evaluate possible correlations with sperm parameters and fertilization in vitro. No correlation was found between the semenogelin concentration and the volume of the ejaculate, sperm concentration, sperm motility, or in vitro fertilization rate. Semenogelin levels were positively correlated with the total protein concentration in seminal plasma, and there was an inverse correlation between the concentration of semenogelin and that of PSA. The levels of semenogelin appear to bear no relationship to the in vitro fertilization capacity of the spermatozoa. 相似文献
102.
103.
Simultaneous detection of several oligonucleotides by time-resolved fluorometry: the use of a mixture of categorized microparticles in a sandwich type mixed-phase hybridization assay. 总被引:1,自引:0,他引:1
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Porous, uniformly sized (50 micrometer) glycidyl methacrylate/ethylene dimethacrylate particles (SINTEF) were used as a solid phase to construct a sandwich type hybridization assay that allowed simultaneous detection of up to six oligonucleotides from a single sample. The assay was based on categorization of the particles by two organic prompt fluorophores, viz. fluorescein and dansyl, and quantification of the oligonucleotide hybridization by time-resolved fluorometry. Accordingly, allele-specific oligodeoxyribonucleotide probes were assembled on the particles by conventional phosphoramidite strategy using a non-cleavable linker, and the category defining fluorescein and/or dansyl tagged building blocks were inserted in the 3'-terminal sequence. An oligonucleotide bearing a photoluminescent europium(III) chelate was hybridized to the complementary 3'-terminal sequence of the target oligonucleotide, and the resulting duplex was further hybridized to the particle-bound allele-specific probes via the 5'-terminal sequence of the target. After hybridization each individual particle was subjected to three different fluorescence intensity measurements. The intensity of the prompt fluorescence signals of fluorescein and dansyl defined the particle category, while the europium(III) chelate emission quantified the hybridization. The length of the complementary region between the target oligonucleotide and the particle-bound probe was optimized to achieve maximal selectivity. Furthermore, the kinetics of hybridization and the effect of the concentration of the target oligomer on the efficiency of hybridization were evaluated. By this approach the possible presence of a three base deletion (DeltaF508), point mutation (G542X) and point deletion (1078delT) related to cystic fibrosis could unequivocally be detected from a single sample. 相似文献
104.
105.
Poliovirus type 1, coxsackievirus type A9, and echovirus type 7 were inactivated by sodium bisulfite and ascorbic acid. Inactivation rates depended upon concentration, temperature, and pH. RNA infectivity was lost during inactivation; the capsid was also altered by these inactivating agents, as determined by enzyme sensitivity assays and by tests of adsorption to cells. Structural modifications of the virus particles were not identical, suggesting that the mechanism of inactivation by ascorbic acid differs from that of sodium bisulfite. 相似文献
106.
107.
We describe a generalized PCR method that will amplify fragments of DNA without any knowledge of sequence using a single primer. Although we are presently using this method to amplify DNA fragments isolated from ancient preserved tissues, in effect, producing PCR libraries, it may prove to have other applications. 相似文献
108.
M M?kel? H Larjava E Piril? P Maisi T Salo T Sorsa V J Uitto 《Experimental cell research》1999,251(1):67-78
The role of matrix metalloproteinases (MMPs) in cell migration was studied by measuring cell growth, migration, and production of MMP-2 and -9 in oral mucosal and skin keratinocytes cultured in the presence of synthetic MMP inhibitors. MMP-2 was the major gelatinolytic MMP produced by these cells while MMP-9 was produced at a low basal level. Inhibitor effects on MMP-9 production were therefore studied in keratinocytes stimulated by tumor necrosis factor alpha (TNFalpha). Tetracycline analogues at concentrations that inhibited the production of MMP-2 but not MMP-9 were able to drastically inhibit migration of both mucosal and skin keratinocytes. Tetracycline analogues also inhibited keratinocyte growth, an effect not found for the other inhibitors tested. Heterocyclic carbonate-derived compounds (LWs) that inhibited MMP-9 but not MMP-2 production had no effect on cell migration. Batimastat, a potent MMP inhibitor, did not have any effect on MMP production or cell growth but did inhibit keratinocyte migration. Tumor growth factor beta (TGFbeta) increased keratinocyte migration as well as both cell-associated and secreted MMP-2 production in wounded cell cultures. The secreted enzyme was partially converted into an active form. In this model batimastat totally blocked TGFbeta-promoted keratinocyte migration. Immunostaining of keratinocytes advancing into the wound revealed that MMP-2 was localized in extracellular matrix contactlike structures against the endogenously produced laminin-5-rich matrix. MMP-9 was localized diffusely along the cell membranes. Using in situ hybridization we observed that in chronically inflamed human gingiva MMP-2 is expressed in epithelium extending into subepithelial connective tissue. These results suggest that MMP-2 plays a specific role in epithelial migration, possibly by detaching the advancing cells from the pericellular matrix or by activating other MMPs. 相似文献
109.
Overexpression of TIMP-1 under the MMP-9 promoter interferes with wound healing in transgenic mice 总被引:2,自引:0,他引:2
Salonurmi T Parikka M Kontusaari S Pirilä E Munaut C Salo T Tryggvason K 《Cell and tissue research》2004,315(1):27-37
We have generated transgenic mice harboring the murine matrix metalloproteinase 9 (MMP-9) promoter cloned in front of human TIMP-1 cDNA. The transgenic mice were viable and fertile and exhibited normal growth and general development. During wound healing the mice were shown to express human TIMP-1 in keratinocytes that normally express MMP-9. However, the healing of skin wounds was significantly retarded with slow migration of keratinocytes over the wound in transgenic mice. In situ zymography carried out on wound tissues revealed total blockage of gelatinolytic activity (i.e., MMP-9 and MMP-2). The results confirm studies with MMP-9 knockout mice showing that MMP-9 is not essential for general development, but they also demonstrate an important role of keratinocyte MMP-9, as well that of other keratinocyte MMPs that are inhibited by TIMP-1, in wound healing. The transgenic mice generated in this study provide a model for the role of MMPs in MMP-9-producing cells in other challenging situations such as bone fracture recovery and cancer invasion.The expert technical assistance of M. Jarva, L. Ollitervo, S. Kangas, and R. Jokisalo is gratefully acknowledged. This work was supported in part by grants from the Finnish Academy of Science, the Swedish Cancer Foundation, the Novo Nordisk Foundation and EC contract QLG1-CT-2000-01131 (K.T.), the Finnish Dental Society Apollonia and the Northern Finland Cancer Foundation (M.P.), as well as the K. Albin Johansson Foundation and the Einar and Karin Stroems Foundation (E.P.) 相似文献
110.
Cloning of the Cdc42 gene from Schizophyllum commune enabled investigation of the role of ScCdc42 in the regulation of vegetative growth and sexual reproduction in this fungus, which has a well-characterized hyphal cell structure, cytoskeleton, and mating system. Ectopic expression of the constitutively active Sccdc42(G12V) or Sccdc42(Q61L) alleles from native or inducible ScCel1 promoters in haploid hyphae had dramatic effects on hyphal morphology, cytoskeletal structure, and Cdc42 localization. For transformants with constitutively active Sccdc42, polar tip growth of apical cells in the leading hyphae was normal but polar tip growth in side branches was altered, implying different regulation of polarity establishment in the two groups of apical cells. Branch emergence at exceptional sites and isotropic growth of cells near the septum indicated that ScCdc42 regulates branch site selection and subsequent hyphal development. Poor dikaryotization along with irregular clamp connections in mates expressing Sccdc42(G12V) or Sccdc42(Q61L) suggested that Cdc42 also contributes to efficient mating in S. commune. 相似文献