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Biomedical subjects

R Kamata

Publications and source records attributed to R Kamata.

At least 55 records · Page 3Linked to original sources
PubMed1985

The serratial 56K protease as a major pathogenic factor in serratial keratitis. Clinical and experimental study.

A possible cause and the difference in clinical severity of serratial keratitis were investigated. Two strains of Serratia marcescens were isolated: one from a patient with severe liquefactive keratitis, who had diabetes mellitus, and one from a patient with mild superficial keratitis, but who had no underlying disease. When the same numbers of bacteria were injected separately into corneas of the same rabbits or guinea pigs, the strain from the first patient elicited severe corneal destruction, remarkable intracorneal edema; and liquefactive necrosis, but the strain from the second caused mild keratitis with erosion or intracorneal abscess. The keratitis induced by the former strain required a longer time to heal, and the prognosis was poorer than that for the other keratitis. Therefore, the difference in severity between the two cases of experimentally induced keratitis paralleled that of the clinical cases. Thus, the severity of the serratial keratitis might be attributed more to the virulence of the bacteria than the condition of the host. The virulence factor seemed to be a heat-labile metabolic product (or products) of the bacteria. To clarify this virulence factor, the major secretory protease (56K protease) produced by these two strains of bacteria was compared by using in vitro and in vivo systems. The virulent strain produced about ten times more protease during culture than the less virulent strain. When injected into the corneas of experimental animals, the 56K protease from the virulent strain induced severe lesions similar to those caused by the living virulent strain of bacteria. These results indicated that one of the major factors causing the virulence was correlated with the tissue destructive 56K protease produced by S. marcescens.

Aged↗
PubMed1985

A serratial protease causes vascular permeability reaction by activation of the Hageman factor-dependent pathway in guinea pigs.

The 56-kilodalton (56K) protease isolated from a culture filtrate of Serratia marcescens caused vascular permeability enhancement followed by edema formation when injected into guinea pig peripheral corneas and subconjunctival space or skin. The character and the mechanism of permeability enhancement were analyzed in vivo. The enhancement was maximum at 5 to 10 min. The permeability reaction increased exponentially by the amount of enzyme used. The enhancement of permeability induced by the 56K protease was not affected by treatment with an antihistamine but was greatly augmented by simultaneous injection of a kinin potentiator, Glu-Trp-Pro-Arg-Pro-Gln-Ile-Pro-Pro-OH (SQ 20,881). Furthermore, the permeability activity of the protease, but not the amidolytic activity, was inhibited by soybean trypsin inhibitor, a well-known inhibitor of plasma kallikrein, as well as by corn trypsin inhibitor, the best inhibitor of activated Hageman factor. Results of these in vivo studies indicate that the permeability-enhancing reaction induced by the 56K protease is caused by activation of the Hageman factor-dependent pathway in the tissue. The permeability-increasing activity of the 56K protease was parallel with the enzyme activity. Serratial lipopolysaccharide did not produce a permeability enhancement reaction within 30 min when injected into guinea pig skin. These results are consistent with the results of recent in vitro experiments in which activation of the purified Hageman factor but not of prekallikrein by the 56K protease was elucidated (Matsumoto et al., J. Biochem. (Tokyo) 96:739-749, 1984). Thus, the molecular mechanism described above appears to be operative in the pathogenesis of corneal edema and chemosis, which is induced by S. marcescens, in addition to the direct tissue destruction by the protease.

Animals↗
PubMed1984

Pathogenesis of serratial infection: activation of the Hageman factor-prekallikrein cascade by serratial protease.

A serratial protease with an apparent molecular weight of 56,000 (56K protease), which had been purified from the culture supernatant of a strain of Serratia marcescens isolated from a corneal lesion of a human eye [Matsumoto, K. et al. (1984) J. Bacteriol. 157, 225-232], greatly enhanced vascular permeability when injected into guinea pig skin. The 56K protease, which requires zinc ion for activity, was found to possess plasma kallikrein-like properties in vitro as judged by (i) preferential amidolysis of carbobenzoxy-Phe-Arg-4-methylcoumaryl-7-amide and Pro-Phe-Arg-4-methylcoumaryl-7-amide, which are known substrates for plasma kallikrein; (ii) release of kinin from high-molecular-weight kininogen; and (iii) prompt activation of Hageman factor followed by generation of kallikrein from plasma prekallikrein. These results suggest that the 56K protease enhances vascular permeability through activation of a Hageman factor-kallikrein-kinin pathway in vivo, and this molecular process appears to be a rational mechanism of enhancement of permeability and serratial pathogenesis.

Animals↗
PubMed1984

Purification and characterization of four proteases from a clinical isolate of Serratia marcescens kums 3958.

Four distinct proteases were purified to homogeneity from culture filtrates of Serratia marcescens kums 3958, a fresh isolate from a patient with a severe corneal ulcer. Purification was achieved by ammonium sulfate precipitation, DEAE-cellulose ion-exchange chromatography, and Sephadex gel filtration chromatography. The proteases were differentiated from each other by polyacrylamide gel electrophoresis with or without sodium dodecyl sulfate and by immunodiffusion in agarose gels. The molecular weights of these purified proteases were estimated to be 56 X 10(3), 60 X 10(3), and 73 X 10(3) (hereafter designated 56K, 60K, and 73K proteases, respectively). The 73K protease was separated into 73Ka and 73Kb upon isoelectricfocusing. The isoelectric points of the 56K (major) and 60K, 73Ka, and 73Kb proteases (minors) were approximately 5.3, 4.4, 5.8, and 7.3, respectively. Both 56K and 60K enzymes were completely inactivated by EDTA at pH 5.0 and were reactivated by zinc ion; thus, they are metalloenzymes, whereas 73K (73Ka and 73Kb) enzymes appear to be thiol proteases. Carbohydrate, cysteine, and cystine were not detected in the 56K and 60K proteases. Amino acid compositions, partial amino acid sequence, and enzymological and immunological properties revealed that these four enzymes are distinct from each other.

Chromatography, DEAE-Cellulose↗
PubMed1984

Two types of lethal damage induced by pepleomycin in cultured Chinese hamster V79 cells.

The cytotoxic effect of pepleomycin (PEP), a derivative of bleomycin (BLM), was investigated and compared with that of BLM on cultured Chinese hamster V79 cells. The concentration-response curve of PEP was biphasic, like that of BLM: the D0 values of the first and second slopes for PEP were 32 micrograms/ml and 140 micrograms/ml, while those of BLM were 60 micrograms/ml and 140 micrograms/ml, respectively. The ratios of D0 for BLM to that for PEP were 1.88 for the first slope and 1.0 for the second one, suggesting that PEP is more cytotoxic than BLM on a unit concentration basis. The time-response curve for PEP showed that the surviving fraction dropped immediately, then rose gradually and finally declined slowly. Analysis of these curves together with the results of further experiments suggested the induction of two types of lethal damage by PEP; potentially lethal damage and non-reparable lethal damage. Potentially lethal damage was induced by PEP treatment, developed within a few minutes, was fixed as lethal damage by trypsinization and was repaired fully within 1 hr if trypsinization was not applied. Non-reparable lethal damage was another type of damage induced by PEP and led to cell death without repair. The two types of lethal damage can account for not only the various shapes of survival curves, but also the differences in cytotoxicity of PEP and BLM.

Animals↗