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Biomedical subjects

E Kominami

Publications and source records attributed to E Kominami.

At least 109 records · Page 6Linked to original sources
PubMed1995

A family with adult type ceroid lipofuscinosis (Kufs' disease) and heart muscle disease: report of two autopsy cases.

Two cases in a family with Kufs' disease had lethal arrhythmias and heart muscle disease. Autopsy findings showed an abundant accumulation of lipofuscin-like lipopigments in most neurons in the central nervous system (CNS). The heart showed a slight increase in the accumulation of the lipofuscin-like lipopigments in the myocardial fibers, slight to severe fibrosis and infiltration of fat cells in the myocardium. The lipopigments both in the heart and in neurons of the CNS had curvilinear profiles on electron microscope and reacted immunohistochemically to polyclonal antibodies against subunit c of mitochondrial adenosine triphosphate (ATP) synthase. The degenerative process in this heart muscle disease might be attributable to the same metabolic abnormality as seen in the neuronal degeneration associated with Kufs' disease.

Arrhythmias, Cardiac↗
PubMed1995

Delayed neuronal death in the CA1 pyramidal cell layer of the gerbil hippocampus following transient ischemia is apoptosis.

The CA1 pyramidal neurons in the hippocampus are selectively vulnerable to transient ischemic damage. In experimental animals, the CA1 pyramidal neurons undergo cell death several days after brief forebrain ischemia. It remains, however, unknown whether this delayed neuronal death is necrosis or apoptosis. To investigate the degenerating processes of the CA1 pyramidal neurons in gerbil hippocampus after brief ischemia, lysosomal and nuclear alterations in the cells were examined using immunocytochemistry, in situ nick-end labeling, and Southern blotting. By light and electron microscopy, immunoreactivity for cathepsins B, H, and L, representative lysosomal cysteine proteinases, increased in the CA1 pyramidal neurons 3 d after ischemic insult, which showed cell shrinkage. By morphometric analysis, the volume density of cathepsin B-positive lysosomes markedly increased 3 d after ischemic insult, while that of autophagic vacuole-like structures also increased at this stage, suggesting that cathepsin B-immunopositive lysosomes increasing in the neurons after ischemic insult are mostly autolysosomes. Nuclei of the CA1 neurons were nick-end labeled by biotinylated dUTP mediated by terminal deoxytransferase 3 and 4 d after ischemic insult, but not in the prior stages. Simultaneously, dense chromatin masses appeared in nuclei of the neurons. By Southern blotting, laddering of DNA occurred only in CA1 hippocampal tissues obtained 4 d after ischemic insult. Confocal laser scanning microscopy demonstrated that the fragmented DNA in the CA1 pyramidal layer was phagocytosed by microglial cells. The results suggest that delayed death of the CA1 pyramidal neurons after brief ischemia is not necrotic but apoptotic.

Animals↗
PubMed1995

Cysteine proteinases in GH4C1 cells, a rat pituitary tumor cell line, are secreted by the constitutive and regulated secretory pathways.

Secretory granules of GH4C1 cells, a rat pituitary tumor cell line, are known to be induced by the treatment of estradiol (E2), insulin, and epidermal growth factor (EGF). We examined changes in the localization of cathepsins B, H, and L, lysosomal cysteine proteinases, in GH4C1 cells before and after hormonal treatment. Northern blotting and immunofluorescence microscopy showed that both mRNAs and intracellular protein concentrations of these enzymes were increased in the hormone-induced cells. By immunoelectron microscopy, immunogold particles indicating cathepsins B, H, and L were localized not only in lysosomes but also in some secretory granules. To further examine the molecular forms of these proteinases in secretory granules, radiolabeling and immunoprecipitation methods were applied to the media of the cells incubated with or without secretagogues (100 nM 12-O-tetradecanoylphorbol-13-acetate and 50 microM forskolin); the proforms of cathepsins B, H, and L were secreted from the cells by the constitutive pathway, whereas the mature forms of cathepsins B and H, and the proform and mature form of cathepsin L were secreted by the regulated pathway. These results suggest that in hormone-induced GH4C1 cells, cathepsins B, H, and L are sorted from the Golgi complex not only into lysosomes but also into secretory granules, in which proforms of cathepsins B and H, and a part of procathepsin L are processed into mature forms.

Animals↗
PubMed1995

[Batten disease (Neuronal ceroid lipofuscinoses)--accumulation of ATP synthase subunit c caused by the delay of lysosomal degradation].

The neuronal ceroid lipofuscinoses (NCLs) represent a group of recessively inherited neurogenerative diseases of infants, children, and young adults that leads to blindness, seizures, dementia, and premature death. These diseases are pathologically characterized by a massive lysosomal storage of autofluorescent lipopigments in neurons and a wide variety of extraneuronal cells. Linkage studies have shown localization of the infantile disease to chromosome region 1p32 the juvenile onset disease to chromosome 16p12.1-p11.2 and a variant form of late infantile form to chromosome 13q21.1-q32. Recently, protein sequencing and immunochemical studies have identified subunit c of the mitochondrial ATP synthase as a major component of the storage material in the late infantile and juvenile types of NCL, and SAPs in infantile type of NCL. Immunolocalization studies demonstrated a dot-like staining of subunit c in the cells with NCL and the staining pattern of subunit c was similar to that of a lysosomal membrane marker, 1gp120. Pulse-chase experiments revealed that a specific failure occurs in the degradation of subunit c in lysosomes whereas its transport into mitochondria and subsequent sequestration into lysosomes are apparently normal.

Adult↗
PubMed1994

Multi-step processing of procathepsin L in vitro.

The proteolytic processes involved in the conversion of procathepsin L to cathepsin L on a negatively charged surface, dextran sulfate, were studied. Upon incubation for 30 min at 37 degrees C, pH 5.5 with dextran-sulfate and dithiothreitol, purified procathepsin L showed maximal activation and, correspondingly, the complete conversion to the 30 kDa, single chain mature form of enzyme was observed. In contrast, incubation under the same conditions on ice rather than at 37 degrees C for 30 or 60 min resulted in partial proteolysis to produce a 31 kDa form without a significant increase in activity. Amino terminal amino acid sequence analyses showed that the 30 kDa form obtained by incubation at 37 degrees C corresponds to the purified form of mature cathepsin L with a 2 amino acid extension at the amino terminal, and that the 31 kDa form generated by incubation on ice possesses a 6 amino acid amino terminal extension, suggesting that the activation and processing of procathepsin L are different processes, and that 4 amino acid residues (Glu-Pro-Leu-Met) at the carboxyterminal in the propeptide function to prevent the activation of processed cathepsin L.

3T3 Cells↗
PubMed1994

Muscle fiber degradation in distal myopathy with rimmed vacuoles.

Late-onset distal myopathy showed numerous rimmed vacuoles with the same properties as autophagic vacuoles. Electron microscopy showed numerous degenerated mitochondria, glycogen, or cell membranes in rimmed vacuoles, but no evidence that these vacuoles engulfed and contained intact or partially disrupted myofibrils. Immunostaining for myosin, alpha-actinin, and actin, however, was sometimes positive within the vacuoles. Compared to the control muscle, there was increased staining activity by calpain around the rimmed vacuoles or in the cytoplasm of mainly atrophic fibers. The result seems to indicate an increase of calpain activity in these muscle fibers. We hypothesize that the myofibrils as well as mitochondria, glycogen, or cell membranes in this myopathy are degraded finally through a lysosomal autophagic process. However, the breakdown of the myofibrils may be not initiated by lysosomal activation; rather it may be the result of extralysosomal processes such as the calpain system.

Actinin↗
PubMed1994

Both cathepsin B and cathepsin D are necessary for processing of ovalbumin as well as for degradation of class II MHC invariant chain.

The effect of highly selective inhibitors of cathepsins on the processing of ovalbumin (OVA) and the presentation of an OVA-derived antigenic peptide (OVA323-339) by antigen presenting cells (APC) was investigated. Both CA-074 (a specific inhibitor of cathepsin B) and pepstatin A (a specific inhibitor of cathepsin D) showed an inhibitory effect on the IL-2 production from an OVA-specific, I-Ad-restricted helper T (Th) cell clone upon stimulation with OVA presented by the I-Ad-positive APC. In contrast, the presentation of the antigenic epitope, OVA323-339, to the same Th clone was not inhibited by either CA-074 or pepstatin A alone, nor even by the mixture of both inhibitors. When APC were treated with cathepsin inhibitor for 24 h, and then antigen and Th were added to the culture, the presentation of not only OVA but also an OVA-derived antigenic peptide was inhibited by either cathepsin inhibitor alone. In addition, the expression of invariant chain on APC was significantly augmented by the pretreatment of APC with either cathepsin inhibitor. Two main conclusions are drawn from these results. First, not only aspartyl protease, such as cathepsin D, but also thiol protease, such as cathepsin B, is involved in antigen processing by APC. Second, both cathepsin B and cathepsin D are necessary for degradation of the invariant chain (Ii) from the MHC class II alpha beta heterodimer in endosomes in order to express functional MHC class II molecules for binding antigenic peptides.

Animals↗
PubMed1994

Lysosomal cysteine proteinases in rat epididymis.

To examine the precise localization of lysosomal cysteine proteinases, cathepsins B, H, and L in rat epididymal epithelial cells, immunohistochemistry and enzyme assay were applied to the epididymal tissue. Granular immunodeposits for cathepsins B and H were detected in epididymal epithelial cells, whereas faint or no immunoreactivity for cathepsin L was found. Moreover, immunoreactivity for cathepsin B appeared mainly in principal cells and was more intense in the head of the epididymis than in the tail, whereas that for cathepsin H appeared in both principal and clear cells and was more intense in the tail than the head. By enzyme assay, activities of cathepsins B and H showed a similar distribution to that of the immunoreactivity. The cathepsin L-specific activity was distributed evenly in each part of the epididymis and was also detected in epididymal fluids obtained from the body and tail parts. By immunoblotting, proforms of cathepsins B, H, and L were present in the seminal fluid. The results suggest that cathepsins B and H are involved in the intracellular degradation system of epididymal epithelial cells, and proforms of cathepsins B, H, and L may be secreted into the epididymal duct lumen.

Animals↗
PubMed1993

Procathepsin L-specific antibodies that recognize procathepsin L but not cathepsin L.

Procathepsin L was purified to apparent homogeneity from the culture medium of v-Ha-ras transformed NIH3T3 (Ras-NIH) cells in three steps; anion-exchange chromatography, gel filtration, and re-gel filtration. SDS-PAGE analyses revealed that the purified samples contained only the precursor form, procathepsin L, but not the mature enzyme, cathepsin L. Antibodies against purified procathepsin L were raised. These recognized both rat cathepsin L and the purified procathepsin L. To isolate procathepsin L-specific antibodies that did not recognize cathepsin L, sequential affinity chromatography procedures were carried out. Immunoblot analyses showed that the procathepsin L-specific antibodies recognized only procathepsin L, but not cathepsin L.

3T3 Cells↗
PubMed1993

Cysteine proteinases in rat dorsal root ganglion and spinal cord, with special reference to the co-localization of these enzymes with calcitonin gene-related peptide in lysosomes.

To examine the localization of lysosomal cysteine proteinases, cathepsins B, H, and L, and their correlation with a neuropeptide, calcitonin gene-related peptide (CGRP) in nerve cells, immunocytochemistry was applied to rat dorsal root ganglion (DRG) and spinal cord (SC) of lumbar segments. By light microscopy, granular immunodeposits for cathepsins B and L were localized in small and large nerve cells in DRG and SC. The cathepsin B- and L-positive smaller nerve cells in DRG and large nerve cells in the ventral horn of SC were also immunostained by anti-CGRP. By electron microscopy, nerve cells in DRG and SC contained lysosomes labeled by immunogold particles indicating cathepsins B and L, while immunogold particles for cathepsin L labeled lysosomes of glial cells. Double immunostaining co-localized immunogold particles for cathepsin B or L and CGRP in lysosomes of small nerve cells in DRG and motoneurons in SC. Moreover, cathepsin H was immunocytochemically demonstrated in lysosomes of satellite cells in DRG, while lysosomes of perivascular microglia-like cells were labeled by immunogold particles for cathepsins B, H, and L. The results suggest the possibility that lysosomes in nerve cells participate in regulation of intraneuronal CGRP level by degrading CGRP.

Animals↗
PubMed1993

Processing and transport of the precursor of cathepsin C during its transfer into lysosomes.

The biosynthesis and processing of a lysosomal cysteine proteinase, cathepsin C (dipeptidylaminopeptidase I), was investigated by pulse-chase experiments in cultured rat macrophages. Cathepsin C is first synthesized as procathepsin C with a molecular mass of 55 kDa. Procathepsin C is then cleaved and modified within 1 h into mature cathepsin C with two chains of 25 and 7.8 kDa. A combination of pulse-chase experiments and the subcellular fractionation analysis showed that procathepsin C and cathepsin C are located in low-buoyant-density organelles and lysosomes, respectively. The reactivity of endoglycosidase H and N-glycanase and analysis of phosphorylation indicated that both precursor and mature cathepsin C are phosphorylated and N-glycosylated to give a high-mannose-type. The addition of 300-kDa mannose 6-phosphate receptor antiserum to the chase medium caused extensive release of procathepsin C into the medium, whereas the addition of control serum did not. The membrane association of procathepsin C was tested by successive extraction of cells pulse labeled for 75 min with hypotonic buffer, alkaline solution, and Triton X-100. Procathepsin C was totally extracted by hypotonic solution, whereas procathepsin D was a membrane-associated form requiring Triton X-100 for its extraction. Gel-filtration chromatography analysis of the pulse-labeled products revealed that the precursor product exists as an oligomeric form. It is suggested that the oligomerization of cathepsin C occurs before its entry into lysosomes.

Amino Acid Sequence↗
PubMed1993

Proenzyme from of cathepsin L produced by thymic epithelial cells promotes proliferation of immature thymocytes in the presence of IL-1, IL-7, and anti-CD3 antibody.

A thymic epithelial cell line (TEC1-4) established from newborn mouse thymus produced a cofactor to promote proliferation of double-negative (CD4-8-) thymocytes in the presence of IL-1, IL-7, and anti-CD3 antibody. Based upon the sequence of 8 amino acid residues near the N terminal portion, the factor was identified as the mouse proenzyme form of cathepsin L which is almost identical to the rat proenzyme form of cathepsin L (95% in homology). Production of the proenzyme form of cathepsin L was confirmed in the TEC1-4 and mouse embryonic thymus by Western blotting analysis and Northern blotting analysis. Furthermore, immunohistologically, the presence of cathepsin L was observed in the epithelial cells of the subcapsular layer and around the small vessels in the thymic cortex. It has been concluded that a subpopulation of thymic cortical epithelial cells produces the proenzyme form of cathepsin L which can act as a cofactor to promote proliferation of DN immature thymocytes.

Amino Acid Sequence↗
PubMed1993

Abnormal distribution of cathepsin proteinases and endogenous inhibitors (cystatins) in the hippocampus of patients with Alzheimer's disease, parkinsonism-dementia complex on Guam, and senile dementia and in the aged.

The immunolocalization of cathepsins B(CB), H and L and cystatins alpha(C alpha) and beta(C beta) were examined in the hippocampus of cases of sporadic Alzheimer's disease (12 cases), parkinsonism-dementia complex on Guam (eight cases), senile dementia of Alzheimer type (two cases), aged subjects with marked senile change (one case) and controls (12 cases, including six normal subjects). CB was lower in most nerve cells in patients than in controls, but markedly increased at the sites of intracellular neurofibrillary tangles (NFTs) and degenerative neurites and/or dendrites in and outside senile plaques (SPs), indicating its close involvement in the metabolisms of various proteins in NFT and SPs. Abundant C alpha and C beta were demonstrated in SP amyloid, suggesting that they are amyloid constituents or co-exist with amyloid. The present study indicated that CB, C alpha and C beta are closely involved in abnormal protein metabolism in NFTs and SP amyloid and suggested that degeneration or denaturation of intracellular proteins, including substrates for proteases and lysosomes, from some acquired cause, results in absolute and/or relative overload for these proteolytic systems, including their inhibitors. This results in incomplete and/or abnormal proteolysis related to NFT and/or amyloid formation.

Adult↗
PubMed1993

Cathepsin L activity is increased in alveolar macrophages and bronchoalveolar lavage fluid of smokers.

Elastinolytic enzymes derived from alveolar macrophages (AM) are considered to play an important role in the development of emphysema associated with cigarette smoking. In this study, the enzyme activity and mRNA expression of cathepsin L were quantitated in AM and bronchoalveolar lavage (BAL) fluid obtained from current smokers and compared with those from nonsmokers. Activity was measured with the synthetic substrate Z-Phe-Arg-MCA combined with a novel cathepsin B inhibitor, CA-074. We found that the specific activity of cathepsin L was significantly elevated in BAL cells from smokers (7.1 +/- 0.7 mumol/mg protein/h, mean +/- SEM) compared with cells from nonsmokers (2.9 +/- 0.3) (p < 0.01). The expression of cathepsin L mRNA in BAL cells as determined by dot-blot analysis was also higher in BAL cells from smokers, which was comparable to the increase in the enzyme activity. About 5 to 6% of the specific activity of cathepsin L in BAL cell lysates was detected in unconcentrated BAL fluid; specific activity was also significantly higher in samples from smokers (0.38 +/- 0.04 mumol/mg protein/h) than from nonsmokers (0.14 +/- 0.02). In addition, procathepsin L (42 kD) and the mature form of cathepsin L (33 kD) were demonstrated in BAL fluid by immunoblot analyses. These data suggest that cigarette smoking induces mRNA expression and the synthesis of cathepsin L in AM and the release of procathepsin from AM into extracellular milieu. Furthermore, increased activity levels of cathepsin L in extracellular compartments may contribute to the proteolysis of elastin in the process of lung destruction associated with cigarette smoking.

Adult↗
PubMed1993

Cysteine proteinases in rat parathyroid cells with special reference to their correlation with parathyroid hormone (PTH) in storage granules.

To further understand the roles of storage granules in parathyroid cells, we examined by immunocytochemistry the localization of cathepsins B and H and of PTH in rat parathyroid gland. In semi-thin sections, small and large granular immunodeposits for cathepsins B and H appeared in the cells, whereas those for PTH were detected throughout the cells, especially in perinuclear regions. By electron microscopy, immunogold particles indicating cathepsins B and H labeled lysosomes and storage granules, whereas those showing PTH were localized in storage granules, small secretory granules, and the trans-Golgi network. Small vesicles labeled by immunogold particles showing these proteinases often appeared close to the storage granules. By double immunostaining, immunogold particles indicating these proteinases were co-localized with those for PTH in storage granules. By EDTA treatment, immunoreactivity for cathepsins B and H and for PTH was notably reduced in the cells, but immunoreactivity for the proteinases was still seen in lysosomes. These results suggest that storage granules in the rat parathyroid cells fuse with small vesicles containing cathepsins B and H, which may participate in regulating the intracellular PTH levels by degrading PTH in the granules.

Amino Acid Sequence↗
PubMed1993

Coexistence of renin and cathepsin B in secretory granules of granular duct cells in male mouse submandibular gland.

Cathepsin B, a representative lysosomal cysteine proteinase, has been demonstrated to coexist with renin in secretary granules of rat pituitary LH/FSH cells and renal juxtaglomerular cells. We investigated immunocytochemically the localization of cathepsins B, H, and L in the submandibular gland of male mice, in which active renin is also produced. By light microscopy, granular immunodeposits for cathepsin B were detected in epithelial cells of the gland, particularly in granular duct cells and interstitial cells. Immunoreactivity for cathepsins H and L was mainly found in interstitial cells, although that for cathepsin H was weakly seen in acinar cells. By electron microscopy, immunogold particles indicating cathepsin B intensely labeled small granules near the Golgi complex of granular duct cells and weakly labeled large secretory granules, whereas those showing renin labeled both granules. Double immunostaining co-localized immunogold particles showing renin and cathepsin B in small perinuclear granules near the Golgi complex. Some immunopositive granules seemed to be closely associated with the Golgi elements. These results indicate that the co-localization of renin and cathepsin B is also seen in secretory granules of granular duct cells in the mouse submandibular gland, as seen in rat juxtaglomerular and LH/FSH cells. This suggests that cathepsin B is one of the possible candidates for the renin-processing enzyme.

Animals↗