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Biomedical subjects

G Imokawa

Publications and source records attributed to G Imokawa.

At least 91 records · Page 5Linked to original sources
PubMed1988

Induction of melanization within hair bulb melanocytes in chinchilla mutant by melanogenic stimulants.

In an attempt to clarify the mechanisms underlying the lack of melanin formation in hair bulb melanocytes of chinchilla mice (genotype a/a, cch/cch, strain PW), we studied the effect of exogenous melanogenic stimulants such as theophylline (Tp), dibutyryl cyclic AMP (db-cAMP), and alpha-melanocyte-stimulating hormone (alpha-MSH) on the induction of melanization. Skin explants excised from the dorsa of chinchilla or lethal yellow C57BL/6J, Ay/a) mice at 7 to 9 days of age were cultured in the presence of Tp (2 mM), db-cAMP (2 mM), or alpha-MSH (1.0 microgram/ml). After 2 to 5 days, melanin formation was induced in hair bulb melanocytes of chinchilla mutant in response to both Tp and db-cAMP, but alpha-MSH did not produce new melanin formation. In contrast, yellow mutant increased the melanin formation in response to all stimulants. Electron microscopic studies demonstrated that while non-treated hair bulb melanocytes of chinchilla mutant contain a large number of stage II-III melanosomes without melanin deposition, a hair bulb treated with Tp exhibits the new formation of melanin within melanosomes that appears both as typical eumelanosomes with striated longitudinal matrices and as pheomelanosomes with vacuolar melanization. Quantitative analysis of melanin has revealed that in chinchilla mutant, Tp and db-cAMP induce a severalfold increase in the formation of both eumelanin [pyrrole-2,3,5-tricarboxylic acid (PTCA)] and pheomelanin (aminohydroxyphenylalanine), whereas alpha-MSH does not stimulate production of either melanin. In yellow mutant, db-cAMP induced a remarkable increase in eumelanin (PTCA), in contrast to the fewfold increase induced by alpha-MSH and Tp. All stimulants induced a slight increase in pheomelanin to a similar extent. These different reactions to melanogenic stimulation suggest a possible defect in the tyrosinase activation system within hair bulb melanocytes in chinchilla mutants.

Animals↗
PubMed1987

Differential hypermelanosis induced by allergic contact dermatitis.

In moderately colored guinea-pig skin, UVB, PUVA (psoralen plus UVA), and allergic contact dermatitis were shown to induce visibly well-defined hyperpigmentation that resembled the pigmentary changes observed in Mongoloid human skin. To clarify mechanisms of allergen-induced hyperpigmentation, we compared the effects of allergic contact dermatitis on pigmentation by using 6 different allergens: dinitrochlorobenzene (DNCB), 1-phenylazo-2-naphthol (PAN), benzyl salicylate (BS), jasmine oil (JO), hydroxycitronella (HC), and ylang ylang oil (YYO). The PAN-, JO-, HC-, and YYO-induced allergic reactions caused a definite visible hyperpigmentation that began to appear within 14 days, reaching maximum intensity about 40 days after the induction of the allergic reaction. These hyperpigmentations were accompanied by a significant increase in the population of dopa-positive melanocytes on day 24 following allergic reactions. In contrast, BS- and DNCB-induced allergic reactions did not give rise to visibly distinct hyperpigmentation in spite of the intensive allergic reactions following their challenge application. In a nonsensitized group, primary irritant reactions were induced by the application of 100% JO, but no distinctive hyperpigmentation was found 40 days after the last application. Quantitative analysis of the number of melanophages in the dermis showed that there was a marked increase in the number of melanophages in PAN, YYO, and HC allergy-induced hyperpigmented areas, with PAN showing a significant increase compared with those in non-treated areas of the same animals, whereas JO was associated with no such increase in hyperpigmented area, despite the stimulated pigmentation. In the case of the lack of induced hyperpigmentation, as seen in BS and DNCB allergy and JO irritation, there was also no substantial increase in the number of melanophages. Our findings indicate that allergic contact dermatitis is a unique melanogenic stimulant different from UV irradiation.

Allergens↗
PubMed1986

Differential analysis of experimental hypermelanosis induced by UVB, PUVA, and allergic contact dermatitis using a brownish guinea pig model.

In moderately colored guinea-pig skin, UVB, PUVA, and allergic contact dermatitis were shown to induce hyperpigmentation that resembled the pigmentary changes observed in mongoloid human skin. Using this model, we examined the effects of chemical agents, including tyrosinase inhibitors and sunscreen agents, on the color changes induced by UV irradiation. The daily exposure of brownish guinea-pig skin to UVB irradiation at a variety of energies for 3 successive days induced clearly visible black pigmentation on the irradiated rectangular areas of the flank within a few days of irradiation, the maximum being reached about 1 week after irradiation, i.e., similar to the changes that occur in pigmented human skin. Split epidermal sheets prepared from untreated pigmented guinea pigs exhibited 200-400 melanocytes/mm2; 1 week after UV irradiation, the applied areas show an increased number of strongly dopa-positive melanocytes with stout dendrites (800-1,000 cells/mm2). UVA irradiation following an intraperitoneal injection of 8-methoxypsoralen (8-MOP) also produced black pigmentation 1 week after irradiation, and this was paralleled by a marked increase in the number of dopa-positive melanocytes in dopa-reacted split epidermal sheets. Allergic contact dermatitis produced by the application of 1-phenylazo-2-naphthol induced hyperpigmentation after an interval of about 14 days in 10 of the 21 allergy-acquiring animals examined. This induced pigmentation was accompanied by an increase in the number of dopa-positive melanocytes as compared to the number seen in controls. In contrast, allergic contact dermatitis produced by the application of dinitrochlorobenzene failed to induce such a high ratio of postpigmentation, with only 3 of the 21 allergy-acquiring animals showing hyperpigmentation and 5 showing depigmentation; in the latter, there was a slight decrease in the number of dopa-positive melanocytes. To study the preventive effect of tyrosine inhibitors on UVB-induced pigmentation, daily topical applications of these compounds were performed after three daily UVB irradiations. Treatment with 10% hydroquinone for 10 days interrupted UVB-induced pigmentation and resulted in a marked reduction in the number of epidermal melanocytes as compared to the number found in UVB-irradiated, untreated control skin.

Animals↗
PubMed1986

Selective recovery of deranged water-holding properties by stratum corneum lipids.

Selective removal of stratum corneum lipids following applications of acetone/ether to the human forearm for extremely prolonged periods of 5-20 min induced an enduring (more than 4 days), chapped and scaly appearance of the skin which was accompanied by a significant decrease in the water-holding properties of the stratum corneum. In order to further elucidate the significance of lipids in the water-holding properties, lipids, which were extracted as sebaceous-rich lipids (SL) for the first 10-min acetone/ether treatment and as stratum corneum lipids (SCL) for the additional 30-min treatment, were topically applied daily on lipid-depleted forearm skin which had been pretreated with acetone/ether for 40 min. Two daily applications of the SCL which were solubilized in squalane containing 1% alpha-monomethyl heptadecyl glyceryl ether (GE) caused a significant increase of conductance, accompanied by a marked improvement in the level of scaling as compared with nontreatment or GE/squalane base, whereas the SL in the GE/squalane base did not exhibit any significant recovery in either conductance value or scaling. To clarify which components of the SCL are primarily responsible for the observed recovery of the water-holding properties, chromatographically separated fractions of the SCL were also topically applied in the same manner for 2 successive days. Out of the following separated fractions: cholesterol, cholesterol ester, free fatty acid, glycolipids, and ceramide, 2 daily topical applications of ceramide fraction induced a significant and the highest increase in the conductance value as compared with GE/squalane base. Furthermore, glycolipids and cholesterol fractions also exhibited a significant recovery when compared with no application at all. In contrast, free fatty acid and cholesterol ester fractions did not indicate any significant increase in the conductance value. These findings strengthen the hypothesis that structural lipids present in the intercellular spaces of the stratum corneum, especially ceramide, play a critical role in the water-holding properties of the stratum corneum.

Acetone↗
PubMed1986

Importance of glycoproteins in the initiation of melanogenesis: an electron microscopic study of B-16 melanoma cells after release from inhibition of glycosylation.

Specific inhibition of core carbohydrate synthesis has been found to induce selective aberration and melanin loss in melanosomes, accompanied by alteration of the carbohydrate moiety in functioning glycoproteins, tyrosinases. In order to further clarify the biologic significance of glycoproteins in the initiation of melanization, initial melanogenesis which occurs during the recovery period following interrupted melanogenesis induced by glycosylation inhibitors, has been electron microscopically investigated. Changes in the tyrosinase activity of the corresponding melanogenic subcellular compartments have also been studied electron cytochemically. Removal of glycosylation inhibition was carried out after exposure of B-16 melanotic melanoma cells to the inhibitors for 10-20 culture days resulting in the loss of their melanization. Recovery of melanization begins visibly 48 h later, thereafter almost attaining the previous normal level by 72 h. At the electron microscopic level, re-formation of melanosomal matrix with periodicity is observed within premelanosomes 48 h after removal of glucosamine, soon followed by deposition of apparent melanin particles along their periodicity by 72 h. In tunicamycin experiments, melanization within premelanosomes starts after the concentration of the fine threadlike interior becomes less distinct followed by a prominence of multiple accumulation of microvesicles within the interior. Electron microscopic dopa reaction shows that the deposition of dopa melanin is prominent in Golgi-associated endoplasmic reticulum of lysosome and coated vesicles up to 24 h after the removal. Thereafter, predominant localization of dopa melanin in premelanosomes gradually increases and finally becomes almost uniform. These observations suggest 2 possible mechanisms for the involvement of glycosylation in melanogenesis: first, the translocation of tyrosinases may be regulated by the presence of specific carbohydrate moieties; second, melanosomal matrix proteins contain carbohydrates which may contribute to the tyrosinase-accepting function or in vivo melanizing function of tyrosinases after forming particle-bound T3 tyrosinase.

Animals↗
PubMed1985

A possible function of structural lipids in the water-holding properties of the stratum corneum.

In order to clarify the possible role of lipids in the water-holding property of stratum corneum, the forearm skin of 6 healthy male volunteers was treated with acetone/ether (1/1) for 1, 5, 10, and 20 min. A prolonged treatment period of 5-20 min produced a chapped and scaly appearance of the stratum corneum without any inflammatory reactions. Under these conditions, there was a marked decrease in the water-holding capacity of the stratum corneum accompanied by a considerable and selective loss of intercellular lipids such as cholesterol, cholesterol esters, and phospholipids. These impairments persisted until day 4 after treatment. Electron microscopic observation of the altered stratum corneum revealed that naturally occurring intercellular materials were absent, leaving the area with the appearance of a vacant space. These findings suggest an additional and essential role of the specific structural lipids for the water-holding properties of the stratum corneum.

Acetone↗
PubMed1985

Analysis of tyrosinases as asparagin-linked oligosaccharides by concanavalin A lectin chromatography: appearance of new segment of tyrosinases in melanoma cells following interrupted melanogenesis induced by glycosylation inhibitors.

The structural alteration of carbohydrate moieties of tyrosinases associated with the depigmentation process induced by glycosylation inhibition has been investigated by using concanavalin A (Con A) affinity chromatography. Con A affinity chromatography of deoxycholate-solubilized large and small granule fractions shows that while all tyrosinases found in control B-16 cells exhibit affinity to Con A lectin, there is an emergence on non-Con A binding tyrosinases in the unpigmented cells induced by glycosylation inhibitors, such as tunicamycin and glucosamine. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis, control tyrosinase activity forms 2 distinct bands consisting of T1 and T3. But tyrosinases from the unpigmented cells lose T3 tyrosinase and are resolved into a few different molecular weight components, one of which is Con A affinitive T1 tyrosinase and the others are non-Con A affinitive tyrosinases with smaller molecular weights than the T1 tyrosinase. These findings suggest that altered structures of carbohydrate moiety in tyrosinase molecules play a role in the induction of loss of membrane-binding capacity of tyrosinases, resulting in the loss of melanization in pigment cells.

Animals↗
PubMed1984

Functional analysis of tyrosinase isozymes of cultured malignant melanoma cells during the recovery period following interrupted melanogenesis induced by glycosylation inhibitors.

Multiple forms of tyrosinase, T1, T2, T3, have been shown to differ with respect to carbohydrate moieties of these isozymes. We demonstrated that, in cultured B-16 melanoma cells, melanization can be completely interrupted by glycosylation inhibitors, such as glucosamine and tunicamycin, and that these inhibitors cause a selective loss of membrane-bound T3. It is further found that inhibition of melanization induced by glucosamine occurs even in the presence of protease inhibitors, such as phenylmethylsulfonyl fluoride and leupeptin, and that melanization inhibition is reversible upon removal of the inhibitor. In this report we have also examined the process of development and recovery of the tyrosinase isozymes in cells in which the interruption of melanogenesis has been released by the removal of these glycosylation inhibitors. The recovery process, which occurs during the period after interrupted melanogenesis and is a process of remelanization, has been biochemically followed. Tyrosinases obtained from the deoxycholate-solubilized large-granule fraction of these melanoma cells have been analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Immediately after removal (0 h recovery) of the glycosylation inhibitor, loss of melanization and T3 is accompanied by T1 heterogeneity which is visualized as two electrophoretically distinct species, T1' and T1''. At this time, T1' and T1'' do not have a concanavalin A affinitive carbohydrate moiety but do possess in vitro dopa reactivity. When recovery of melanization begins visibly 24 h later, T3 is re-formed with disappearance of T1 heterogeneity. By 48 h, the previous normal level of melanization is almost attained. These results suggest that maturation of tyrosinase may occur via T1' and T1'' as precursors of T3, or possibly T1 through the addition of N-glycosydically linked oligosaccharide moieties which can be interrupted by glucosamine and tunicamycin.

Animals↗
PubMed1983

Selective aberration and pigment loss in melanosomes of malignant melanoma cells in vitro by glycosylation inhibitors: premelanosomes as glycoprotein.

We have found that glucosamine (1 mg/ml) or tunicamycin (0.2-0.4 micrograms/ml), specific inhibitors of lipid carrier-dependent glycosylation of protein, when added to cultured B-16 melanoma cells produce a marked loss of pigmentation, accompanied by distinctive biochemical as well as ultrastructural aberrations in their melanogenic compartments. Electron microscopic analysis shows that these newly induced unpigmented cells form uniquely altered melanosomes containing little or no melanin, although their population is not substantially reduced. Within the melanogenic compartments, selective aberration of melanosomes is seen, that is, deformity, bulging, and segregation of their interior membrane, as well as the intramelanosomal formation of irregularly concentric lamellar structure. No apparent structural deformity of Golgi apparatus, Golgi-associated endoplasmic reticulum of lysosome (GERL), and coated vesicles has been observed. Further, no substantial alteration is seen in mitochondria or other cellular structures. Quantitative analysis of altered and nonaltered melanosomes has revealed that the ratio of altered premelanosomes to the total number increases to 44% in glucosamine-treated cells and to 99.5% in tunicamycin-treated cels. Compared to 13% in the control. Electron microscopic dopa reaction has also revealed that these altered melanosomes seem to exhibit a weakly positive or a negative dopa reaction in both glucosamine- and tunicamycin-treated melanoma cells although a number of dopa-positive altered melanosomes are still seen. However, GERL and coated vesicles show no apparent decrease in dopa reaction. It may be concluded that glycoprotein synthesis is integral to the formation of normal melanosomes and to their specific melanizing function, which could be impaired by inhibition of the synthesis of asparagin-linked mannose-containing sugar chains. This results in retrogressive changes in the premelanosomal tyrosinase during its maturation process.

Animals↗
PubMed1982

The inhibitory effect of zinc pyrithione on the epidermal proliferation of animal skins.

The hypothesis that zinc pyrithione, a highly active anti-dandruff agent, exerts an anti-biosynthetic effect and reduces the epidermal turnover has been tested using guinea pig and hairless mouse skins. It has been found that enhanced mitosis caused by both stripping off the horny layers and sodium dodecyl sulphate, could be suppressed by approximately 50% by the topical application of zinc pyrithione. Furthermore, sodium dodecyl sulphate-induced epidermal hyperproliferation has been found suppressible by the simultaneous application of zinc pyrithione. Thymidine incorporation studies using hairless mice revealed that a single application of zinc pyrithione results in reduction of DNA synthesis. It is suggested that the anti-dandruff effect of zinc pyrithione results primarily from its action as an anti-biosynthetic agent rather than from its anti-yeast action.

Animals↗
PubMed1982

Loss of melanogenic properties in tyrosinases induced by glucosylation inhibitors within malignant melanoma cells.

Glycosylation inhibitors, glucosamine or tunicamycin, have been found to be specific inhibitory modulators for melanogenesis, which is accentuated generally in malignant melanoma cells. Exposure to glucosamine (1 mg/ml) or tunicamycin (0.2 to 0.4 micrograms/ml) induces a marked pigment loss within melanoma cells in vitro with a decrease in their grown curves. This melanogenic inhibition occurs without a substantial decrease in the synthesis of DNA, RNA, and protein in comparison with a specific, marked suppression of carbohydrate synthesis as revealed by suppressed mannose incorporation into these cells. Assay of tyrosinase of glucosamine- or tunicamycin-induced unpigmented melanoma cells has revealed a selective and marked decrease in the melanosome-rich large-granule fraction, but no substantial decrease in the total activity of remaining subcellular fractions. Electrophoresis of tyrosinase in the 30,000 X g supernatant fraction demonstrates an increase in the T1 form of soluble tyrosinase, while a disappearance of or marked decrease in membrane-bound tyrosinase, T3, is seen in the small- and large-granule fractions. Glycoprotein synthesis in the melanogenic subcellular compartments of pigment cells seems to play an integral role in melanogenesis which is principally enhanced in their carcinogenic status.

Animals↗