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Biomedical subjects

M Manabe

Publications and source records attributed to M Manabe.

At least 37 records · Page 2Linked to original sources
PubMed1997

[Determination of malondialdehyde-modified LDL(MDA-LDL) and its potential usefulness].

Numerous studies have indicated that oxidative modification of low-density lipoprotein(LDL) plays a critical role in the pathogenesis of atherosclerosis. Malondialdehyde-modified LDL(MDA-LDL) is one of the candidates which is the oxidative product of LDL. However, the existence of MDA-LDL in the circulation has been in dispute. Therefore, for the assessment of oxidized-LDL in human serum, we developed a sensitive enzyme-linked-immunosorbent assay(ELISA) for the detection of MDA-LDL. In our method, monoclonal antibody against MDA-LDL, ML25 was used. ML25 recognized MDA-LDL as well as MDA-modified proteins by a solid-phase competitive enzyme immunoassay. Therefore, to establish an ELISA method which is specific for detection of MDA-LDL, ML25 was combined with apoB-specific antibody (AB16) as the second antibody. Using this method, MDA-LDL was detectable in the sera of 314 healthy individuals. The concentration of MDA-LDL preferably ranged from 20 to 80 units/l when the absorbance with artificially prepared MDA-LDL at the concentration of 1 mg/l was defined as 1 unit/l. Furthermore, assays for lipoprotein subfractions separated by density-gradient ultracentrifugation revealed that MDA-LDL was mainly distributed in the LDL fraction as expected, and MDA-LDL/apoB ratio showed a peak at small, dense LDL fractions. This finding seems to be quite interesting since an elevated level of small, dense LDL has been reported to be associated with an increased risk of atherosclerosis. We concluded that our method is useful for specific detection of MDA-LDL in human serum and might be an elevation method for atherogenicity.

Arteriosclerosis↗
PubMed1997

[Complete cholesteryl ester transfer protein deficiency increases oxidized-LDL in plasma].

Malondialdehyde-modified LDL(MDA-LDL) is one of the major oxidative LDL. MDA-LDL was determined by enzyme immunoassay in patients with complete and partial cholesteryl ester transfer protein(CETP) deficiency. Significantly increased serum MDA-LDL levels and MDA-LDL/apoB ratios in the LDL fraction were observed in patients with complete CETP deficiency but not for those with partial CETP deficiency. The present results may indicate that patients with complete CETP deficiency have a higher risk for atherosclerosis than those with partial CETP deficiency and normal.

Arteriosclerosis↗
PubMed1997

[Effect of controlled hypotension on cerebral oxygen delivery].

The margin of safety for controlled hypotension is still unclear especially in the central nervous system (CNS) which is one of the most sensitive organs to hypoxia and ischemia. Recently, cerebral optical spectroscopy in the infrared light range was developed as a useful tool which makes it possible to monitor cerebral oxygenation (rSO2) non-invasively and continuously during anesthesia. Resulting rSO2 mainly reflects oxygen extracts by cerebral tissue and then indicates cerebral oxygen delivery. We examined the limitation of controlled hypotension in the brain in 12 patients by monitoring rSO2 during anesthesia. rSO2 under room air breathing (control value as normal physiological condition) was 67 +/- 3% (mean +/- SEM). It significantly increased by 5.6 +/- 0.8% under 100% oxygen breathing, but decreased near to the control value under sevoflurane anesthesia (FIO2 1.0). During moderate controlled hypotension (70% of normal blood pressure) by prostaglandin E1 under sevoflurane anesthesia (FIO2 1.0). rSO2 remained at control value, indicating that cerebral oxygen delivery was still sufficiently maintained. However rSO2 decreased significantly by 9.0 +/- 1.1% in same controlled hypotension condition under FIO2 0.4. This decrease in rSO2 could be potentially harmful for CNS although any post-operative neurological disorder was not observed in our cases. We conclude that cerebral oxygen delivery may be insufficient even in the moderate controlled hypotension, and thus higher FIO2 is recommended in such procedures.

Alprostadil↗
PubMed1997

Distinctive expression of filaggrin and trichohyalin during various pathways of epithelial differentiation.

Filaggrin and trichohyalin are keratin intermediate filament-associated proteins, and are primarily expressed in the granular cells of the epidermis and in the inner root sheath cells of the hair follicles, respectively. These two proteins are, however, occasionally co-expressed in some tissues. To gain more insights into the mechanisms for expression and processing of (pro)filaggrin and trichohyalin during various pathways of epithelial differentiation, we compared their localization by double immunostaining techniques in normal and psoriatic epidermis, tongue filiform papillae and cultured human epidermal keratinocytes. In normal foreskin, trichohyalin immunoreactive cells were observed only occasionally and, when present, they always co-expressed filaggrin. Trichohyalin expression occurred, however, in filaggrin-negative cells as well as in filaggrin-positive cells in the psoriatic epidermis, tongue papillae and cultured keratinocytes. Filaggrin and trichohyalin were induced independently at different times following calcium shift in cultured keratinocytes. Immunoelectron microscopy demonstrated the distinct intracellular distribution of filaggrin and trichohyalin. Some filaggrin granules were observed in the cells where trichohyalin was diffusely distributed. Likewise some trichohyalin granules were found in the cells with diffuse filaggrin labelling. These results suggest the existence of distinct expression/processing mechanisms of filaggrin and trichohyalin in differentiating keratinocytes.

Adult↗
PubMed1996

Assembly of hair keratins in transfected epithelial cells.

To define the interactions required for the filament assembly of differentiation-specific keratins, active copies of mouse hair keratin mHa1 and mHb4 genes were introduced into a rat kangaroo kidney epithelial cell line (PtK2) and a rat stratified squamous epithelial cell line (rat epidermal keratinocyte). In PtK2 transient transfectants, when introduced individually or in combination, mHa1 and mHb4 formed aggregates of ring-like structures of various sizes at the perinuclear region with no evidence of organization into a keratin network. These aggregates altered the distribution of the endogenous keratins and vimentin. In most of the cells carrying the ring-like structures of mHa1 and mHb4 around the nucleus, the endogenous keratin network collapsed and localized around the nucleus. Furthermore, the densely accumulated endogenous keratin surrounded the ring-like aggregates with partial co-localization. However, when transfected into the rat epidermal keratinocytes, mHa1 and mHb4 were able to co-localize with the well-developed cytoskeleton of endogenous keratins. These results showed that, in contrast to keratin pairs K5/K14 and K8/K18, the mHa1/mHb4 pair is unable to develop an extensive keratin network on its own and that there are possible differential abilities among these hair keratins and other keratins to form well-developed cytoplasmic networks.

Animals↗
PubMed1996

Preferential deimination of keratin K1 and filaggrin during the terminal differentiation of human epidermis.

The upper layers of mammalian epidermis contain citrulline-containing proteins formed by enzymatic deimination of arginine residues. To study the role of protein deimination in epidermal differentiation, we identified deiminated proteins extracted from human epidermis. Major deiminated proteins were identified as partially degraded keratin K1, while those from keratin K10 and a highly heterogeneous mixture of deiminated filaggrin isomers were detected as minor components. Deiminated keratins were recovered in a fraction enriched with keratins from the cornified layers. The subsequent immunohistochemical study showed that deiminated proteins were localized mainly in the lowermost cornified layer, but not in the granular layer. These data suggested that partially degraded/disulfide-cross-linked keratin K1 was preferentially deiminated during the terminal stages of epidermal differentiation. We therefore speculated that the protein deimination might influence the interaction of basic K1 with its acidic partner K10, pre-existent K5/K14 networks or keratin-associated protein filaggrin.

Adult↗
PubMed1996

Molecular cloning of a cDNA encoding enolase from the filamentous fungus, Aspergillus oryzae.

A 1.6-kbp full-length cDNA for the Aspergillus oryzae enolase gene (enoA) was cloned. The sequenced insert contained a continuous open reading frame of 1314 bp encoding a protein of molecular weight 47 405. Among all enolases sequenced to-date, the deduced amino-acid sequence showed the highest homology (74.9%) with Candida albicans enolase (ENO1). Strong codon biases and multiple transcription start sites downstream from CT-blocks in the 5'-flanking region suggested strong expression. enoA mRNA was found to occupy approximately 3% (w/w) of total mRNA of A. oryzae by quantitative RT-PCR. This strong transcription was dependent on the carbon source in the medium and correlated with the growth rate of the mycelium.

Amino Acid Sequence↗
PubMed1996

Ultrastructure of size-regulated hetero-spheroids composed of human keratinocytes and fibroblasts.

In order to develop a new in vitro skin model multicellular floating hetero-spheroids were prepared by culturing keratinocytes and fibroblasts on hydrophilic culture dishes coated with type I collagen and a thermo-responsive polymer. Upon decreasing the substratum's temperature to an ambient temperature, the spheroids detached from the substratum and were thereafter maintained in either: Medium I, a medium mixture of keratinocyte growth medium (KGM) and supplemented Dulbecco's modified Eagle's medium (DMEM) at a ratio of 1 to 2; or medium II, KGM for the initial 24 h followed by supplemented DMEM for the remainder of the culture periods. The spheroids displayed a typical pattern of an external rim of keratinocytes with an internal core of fibroblasts. A minute space separated the keratinocytes and fibroblasts. The stratification of cells cultured in medium II was more prominent than that of the cells cultured in medium I. Markers of the advanced stages of keratinization such as keratohyalin granules, membrane coating granules and the cornified envelope were not observed. Interestingly, keratinocytes underwent the same differentiation pathway as non-keratinized stratified epithelia such as esophagus. With consideration to keratinocyte-fibroblast interactions, it may be of interest to incorporate the study of such morphological impairments when investigating the effects of growth factors and their ligands.

Cell Aggregation↗
PubMed1996

Immunolocalization of epimorphin in skin.

Epimorphin was originally identified as a mesenchymal cell surface-associated protein that modulates epithelial morphogenesis in embryonic skin and lung epithelia. A previous report which utilized embryonic mouse skin, showed that epimorphin was localized non-homogeneously in a region adjacent to the epidermis and in a mesenchymal cell condensation located in front of growing hair follicles. We report herein a further detailed localization of this protein in adult mouse skin using immunoelectron microscopy. Epimorphin was found to be localized on the undersurface of basal cells, in the cytoplasm of cell processes of fibroblasts, as well as on the plasma membrane of fibroblasts, endothelial cells, pericytes, perineurium and endomysium. Our present finding indicated that epimorphin is one of the factors involved in multiple biological functions in a variety of structures derived from various origins and that it is not a specific epithelial morphogenetic factor.

Animals↗
PubMed1996

Expression of keratohyalin-trichohyalin hybrid granules in molluscum contagiosum.

BACKGROUND: Recently, in the filiform papillae epithelium of mouse dorsal tongue, we showed the presence of hybrid granules in which filaggrin and trichohyalin were both present, but physically segregated. Further, trichohyalin was also detected in scattered granular cells of a number of hyperplastic skin diseases. METHODS: The epidermis infected with molluscum contagiosum virus (MCV) was studied by conventional electron microscopy in conjunction with light and electron-microscopic immunohistochemistry, using both antifilaggrin and antitrichohyalin antibodies as probes. RESULTS: We found that the granular cells of MCV-infected epidermis contained both filaggrin and trichohyalin. Subsequent electron-microscopic examination showed that the granular cells contained morphologically heterogeneous granules that appeared to be composed of discrete areas of distinct electron densities. Double-labeling, using antibodies to filaggrin and trichohyalin, clearly indicated that filaggrin and trichohyalin were both present in the hybrid granules and that the electron-dense regions contained trichohyalin while the more electron-lucent regions contained filaggrin. CONCLUSIONS: The expression of trichohyalin was a common feature observed in the epidermis from a heterogenous group of hyperplastic conditions, including MCV infection. This finding has led us to speculate that trichohyalin may be specifically or preferentially involved in interacting with the hyperproliferation-related keratin pair (K6/K16), whereas the function of filaggrin is more closely linked to the skin-type keratin pair (K1/K10) that are normal keratins found in the differentiated epidermis.

Cell Differentiation↗
PubMed1996

Trichohyalin expression in skin tumors: retrieval of trichohyalin antigenicity in tissues by microwave irradiation.

BACKGROUND: The antitrichohyalin antibody AE 15 is effective for identifying the cell lineage that undergoes the pathway of inner root sheath-type differentiation. Unfortunately, the AE 15 does not react with trichohyalin in tissue that is formalin-fixed and embedded in paraffin according to routine procedures. METHODS: We attempted to retrieve the trichohyalin antigenicity in formalin-fixed, paraffin-embedded biopsy specimens that included normal skin as well as skin tumors such as trichofolliculoma and pilotricoma. RESULTS: We found that the use of a metal solution in combination with microwave oven heating improves the trichohyalin immunoreactivity substantially. Further, trichohyalin was found to be expressed not only in the secondary hair structure in trichofolliculoma but also in a certain cell lineage that differentiates to squamoid cells in pilomatricoma. CONCLUSIONS: Our findings established that surgical specimens processed under routine procedures can be successfully investigated with AE 15 using the microwave irradiation method. Studies of epidermal diseases expressing trichohyalin should provide valuable insights into our understanding the functional significance of trichohyalin during abnormal keratinization.

Adjuvants, Immunologic↗
PubMed1996

Application of long-distance PCR to restriction site mapping of a cloned DNA fragment on the lambda EMBL3 phage vector.

Long-distance PCR was applied to rapidly map the restriction sites of long inserts cloned on lambda EMBL3 phage vector. The restriction sites of 9 of 15 enzymes were completely assigned in a model experiment within 14 h, including 8h for the PCR amplification. This method was found particularly useful for genomic DNA cloning when the partial sequence of the corresponding cDNA is known.

Bacteriophage lambda↗
PubMed1996

[Changes in bone mineral content and bone metabolism during pregnancy and puerperium].

In order to clarify the bone metabolism of women during both pregnancy and puerperium, we studied the changes in bone mineral content and bone biochemical parameters. Bone mineral content was measured by ultrasound bone densitometry, and serum calcium (Ca), ionized calcium (i-Ca), intact parathyroid hormone (i-PTH), intact osteocalcine (i-OC), urinary pyridinoline (Pyr) and deoxypyridinoline (D-Pyr) were measured concomitantly. 1. In the 3rd trimester bone stiffness was decreased (p < 0.05) compared the 1st and 2nd trimesters, but was slightly increased in the puerperal group. 2. Serum i-OC was decreased during pregnancy but significantly increased (p < 0.01) in the puerperium. The Pyr/Creatinine (Cre) and D-Pyr/Cre ratios were both increased in pregnant and puerperal women. 3. Serum Ca, i-Ca and i-PTH were within the normal range during pregnancy and the puerperium. These results suggest that in the 2nd and 3rd trimesters bone resorption would be increased, and in the puerperium bone formation would be increased. In pregnant and puerperal women serum Ca, i-Ca and i-PTH are within the normal range, but the turnover rate for Ca metabolism would be relatively increased.

Adult↗
PubMed1995

Altered distribution of keratinization markers in epidermolytic hyperkeratosis.

Epidermolytic hyperkeratosis (EH) is a genetic disorder of keratins associated with epidermal differentiation. Affected individuals carry gene mutations for conserved sequences of keratins K1 or K10. The structural alterations of tonofilaments in EH seem to be a direct consequence of the keratin gene mutations. EH epidermis, however, shows many other unexplained abnormalities including acanthosis, hypergranulosis, and hyperkeratosis. To further elucidate the pathogenetic mechanism of EH, we studied distribution patterns of other keratinization-associated molecules including involucrin, small proline-rich protein (SPRR) 1, loricrin and trichohyalin in the skin of four patients by light and electron microscopic immunohistochemistry in conjunction with conventional transmission electron microscopy. The middle to upper epidermal cells showed moderate to strong immunoreactivities to involucrin, SPRR1 and loricrin antibodies. Both intracellular staining and cell peripheral staining was seen for involucrin and SPRR1 antibodies. Loricrin labelling was prematurely associated with the plasma membrane of granular cells, possibly relating to abnormal keratin filament aggregation and cellular vacuolization. Some loricrin labelling was localized on the keratin aggregates, suggesting intermolecular associations between keratin and loricrin. Trichohyalin, hardly detectable in normal epidermis, was present in some granular and cornified cells in EH in association with keratin filaments, suggesting that it may function as an intermediate filament-associated protein. While cornified cell envelopes were intensely labelled only with loricrin antibodies in normal skin, they were immunoreactive to involucrin, SPRR1 and loricrin antibodies in EH. Sequential change in electron density of the cornified cell envelopes, a constant feature in normal skin, was often absent in EH. These results suggest an altered assembly process of cornified cell envelopes in EH.

Adolescent↗
PubMed1995

Ephedrine-induced increases in arterial blood pressure accelerate regression of epidural block.

We investigated the hypothesis that ephedrine-induced increases in blood pressure accelerate the regression of epidural block. In patients undergoing minor gynecologic surgery, we performed lumbar epidural blockade using 2% lidocaine containing 1:200,000 epinephrine to which was added 0.1 mg of fentanyl. Eighty minutes after the epidural injection, we started an ephedrine infusion to increase the systolic blood pressure by 20% in 10 min and maintained the value for the following 20 min. Then we compared the proximal extent of sensory analgesia at 80 min with that at 140 min. Ephedrine significantly (P = 0.001) hastened the regression of sensory analgesia. We conclude that an ephedrine-induced blood pressure increase accelerates regression of epidural blockade.

Adjuvants, Anesthesia↗
PubMed1995

Detection of deiminated proteins in rat skin: probing with a monospecific antibody after modification of citrulline residues.

We performed a systematic study on deiminated proteins present in rat epidermis. Proteins extracted from various epidermal samples were resolved by either one- or two-dimensional gel electrophoresis and Western blotted to nitrocellulose membranes. Deiminated proteins were detected by modification of citrulline residues followed by probing with an anti-modified citrulline monospecific antibody. The cornified layer of adult plantar skin gave multiple series of isoelectric variants, most of which were found to be differentially deiminated type II keratins (60 kDa, and 67 kDa or above). The whole epidermis of 5-day-old rat back skin showed isoelectric variants of 60-kDa keratin as major deiminated components, and deiminated 55-kDa keratin and deiminated filaggrin as minor spots. In addition, we found highly deiminated proteins (200-220 kDa) thought to be derived from trichohyalin. The immunoreactivity of deiminated proteins was mainly localized in the granular and cornified layers of epidermis. Co-localization of deiminated filaggrin and keratins in the granular layer suggests the possible role of protein deimination during the terminal stage of epidermal differentiation.

Animals↗
PubMed1995

Cutaneous ultrastructural features of the flaky skin (fsn) mouse mutation.

An autosomal recessive genetic disease with clinical and histopathological skin features resembling human psoriasis vulgaris occurs naturally in flaky skin mice (fsn/fsn). Affected mice are normal at birth, except for a hypochromic anemia. Subsequently, they develop hyperkeratotic plaques and acanthosis with elongation of rete ridges. Scanning electron microscopic examination revealed a greatly thickened epidermis, a sparsity of hairs and scale accumulations on the epidermal surface. Hair shafts had conspicuous pits, striations, and exophytic protrusions. Nails were bent at a 90 degrees angle with surface irregularities and accumulations of scale at the nail base. Transmission electron microscopic examination showed increased epidermal thickness, mitochondrial aberrations, and intraepidermal invasion by neutrophils. Keratohyalin abnormalities were detected using immunocytochemical staining for profilaggrin. At the dermal-epidermal junction, numerous macrophages and mast cells were seen in close proximity to focal dissolutions of the basement membrane. A high density of collagen fibers and cellular infiltrates were evident in the papillary dermis. This constellation of ultrastructural aberrations is typically found in psoriasis vulgaris and supports the theory that the flaky skin mouse mutation is a naturally occurring analog to one variety of human psoriasis vulgaris.

Anemia, Hypochromic↗