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-177- Keratinocyte(3) 9/5 Room-D
MSH RECEPTOR BINDING ACTIVITY AND a-MSH PRODUCTION IN NORMAL HUMAN KERATINOCYTES DURING DIFFERENTIATION.
Michiko Nagahama, Yoko Funasaka, Ashok K. Chakraborty, Akira Ito, Masamitsu Ichihashi.
Department of Dermatology, Kobe Univerisity School of Medicine Kobe, Japan.
We have reported that a-MSH was strongly stained in differentiated keratinocytes of the uppermost portion of malpighian layer in both normal human skin and squamous cell carcinoma specimens by immunohistochemistry study. To clarify whether a-MSH is highly produced in differentiated keratinocytes (d-KC) and d-KC could bind to a-MSH, production of a-MSH and MSH receptor binding ability were studied by radioimmunoassay, receptor binding assay to 125I labelled Nle4DPhe7 a-MSH, and Northern blotting using the melanocortin 1 receptor (MC1-R) cDNA as a probe in Ca++ stimulated d-KC. The production and secretion of a-MSH in d-KC was not increased compared with those in undifferentiated KC, however, the binding activity and MSH-R mRNA expression was higher in d-KC. Taken together, these results suggest that strong staining of a-MSH in d-KC was not due to the high production of a-MSH, but to the higher expression and binding ability of a-MSH receptor.
-178- Keratinocyte(3) 9/5 Room-D
EFFECTS OF INTERLEUKIN-6 ON THE EXPRESSION OF EPIDERMAL GROWTH FACTOR RECEPTOR AND CELL GROWTH IN KERATINOCYTES DERIVED FROM NORMAL AND PSORIATIC LESIONAL SKIN.
Noritaka Oyama1, Masayuki Sekimata2, Keiji Iwatsuki1, Yoshimi Homma2, Fumio Kaneko1.
1Department of Dermatology, 2Department of Biomolecular Sciences, Fukushima Medical College, Fukushima, Japan.
We investigated the effect of interleukin-6 (IL-6) on the expression of epidermal growth factor receptor (EGFR) and cell growth using 2 different sources of keratinocytes. EGF elicited the growth of both normal human keratinocytes (NHKs) and psoriatic lesional keratinocytes (PLKs). IL-6 potentiated the EGF-dependent growth of NHKs, but has no observable effect on PLKs, while IL-6 itself showed no growth-stimulating activities in both cell types. Immunodetection and in situ hybridization analyses revealed that IL-6 induces EGFR expression in NHKs in a time- and dose-dependent manner. On the other hand, PLKs expressed high levels of EGFR even when unstimulated and the expression level was not affected by IL-6 stimulation. These results suggest that the EGF/EGFR system is involved in the growth of NHKs and PLKs and that IL-6 potentiates NHK growth partly through the induction of EGFR. Furthermore, our data may also suggest a possible mechanism that the epidermal thickness is related to cytokines and EGFR expression in inflammatory skin diseases such as psoriasis.
-179- Keratinocyte(3) 9/5 Room-D
increased number of interferon-gamma producing cd4 and cd8 cells in patients with psoriasis.
Toshiaki Nakamura, Shigetoshi Sano, Satoshi Itami, Kunihiko Yoshikawa.
Department of Dermatology, Osaka University Medical School, Osaka, Japan.
-180- Keratinocyte(3) 9/5 Room-D
ELEVATED EXPRESSION OF ORNITHINE DECARBOXYLASE GENE EXPRESSION IN SCLERODERMA EPIDERMIS.
Tsutomu Ohtsua1, Noriyuki Koibuchi2, Shigeru Matsuzaki3, Akio Yamakage1, Souji Yamazaki1.
1Department of Dermatology, 21st Department of physiology, 3Department of Biochemistry, Dokkyo University, School of Medicine, Tochigi, Japan.
The role of epidermis in scleroderma remains unknown. We investigated the mRNA expression of ornithine decarboxylase (ODC), which is rate limiting enzyme for the synthesis of polyamines. Using in situ hybridization technique, ODC mRNAs were detected. Sections treated with the antisense probe showed that the grains were concentrated exclusively over the epidermis of SSc. As our subcloned antisense probe specifically hybridized with ODC mRNA, the findings indicated that the ODC mRNA level was elevated in the epidermis. Of 5 SSc, 3 cases showed highly elevated ODC mRNA, and 2 moderate. It is possible that polyamines play a role for the skin changes in SSc, because an increase in polyamine biosynthesis is closely associated with inflammation and cell proliferation.
-181- Keratinocyte(3) 9/5 Room-D
SENESCENCE OF CULTURED KERATINOCYTES AND THE EXPRESSION OF P21.
Koji Sayama, Yuji Shirakata, Kazushige Midorikawa, Yasushi Hanakawa, Koji Hashimoto.
Department of Dermatology, Ehime University School of Medicine, Ehime, Japan.
The protein p21 is an inhibitor of cyclin-dependent kinase and affects cell cycle progression and senescence. We have studied whether p21 was associated with cultured keratinocytes senescence. Keratinocytes were isolated from normal human skin obtained from individuals aged from 1 to 70 years, and cultured under serum free condition. The cultures have been continued until the cell growth arrest. The expression of p21 in the growing cells and the senescent cells was analyzed by Western blot. The keratinocytes obtained from younger individuals tend to survive longer periods than the cells from older persons. The p21 expression has not been changed compared to the early passage while the cells were growing, but increased just before the growth arrest. These results suggest that p21 might play an important role in growth arrest of keratinocyte senescence.
-182- Keratinocyte(3) 9/5 Room-D
ESTABLISHMENT OF A SPONTANEOUSLY IMMORTALIZED HUMAN KERATINOCYTE CELL LINE FROM NORMAL ADULT SKIN.
Tadashi Karashima, Osamu Mori, Takashi Hashimoto.
Department of Dermatology, Kurume University School of Medicine, Fukuoka , Japan.
Spontaneously immortalization of human keratinocyte has been reported, although such reports are still very few. ¡¡We have established the spontaneous immortalized of human keratinocytes from normal adult skin. This Kana cell line is established from a long-term primary culture of epidermal keratinocytes grown in D-MEM contained 10% FCS. These cells can be considered immortalized (>100passages), and exhibit karyotypical abnormality. In low Ca++ medium, the cells predominantly grew as monolayers. However in high Ca++ medium, the cells readily stratified and formed coherent horn sheets. This cell line may be a useful model for the study of human keratinocytes.