Kinetin Improves Barrier Function of the Skin by Modulating Keratinocyte Differentiation Markers

BACKGROUND: Kinetin is a plant hormone that regulates growth and differentiation. Keratinocytes, the basic building blocks of the epidermis, function in maintaining the skin barrier. OBJECTIVE: We examined whether kinetin induces skin barrier functions in vitro and in vivo. METHODS: To evaluate the efficacy of kinetin at the cellular level, expression of keratinocyte differentiation markers was assessed. Moreover, we examined the clinical efficacy of kinetin by evaluating skin moisture, transepidermal water loss (TEWL), and skin surface roughness in patients who used kinetin-containing cream. We performed quantitative real-time polymerase chain reaction to measure the expression of keratinocyte differentiation markers in HaCaT cells following treatment. A clinical trial was performed to assess skin moisture, TEWL, and evenness of skin texture in subjects who used kinetin-containing cream for 4 weeks. RESULTS: Kinetin increased involucrin, and keratin 1 mRNA in HaCaT cells. Moreover, use of a kinetin-containing cream improved skin moisture and TEWL while decreasing roughness of skin texture. CONCLUSION: Kinetin induced the expression of keratinocyte differentiation markers, suggesting that it may affect differentiation to improve skin moisture content, TEWL, and other signs of skin aging. Therefore, kinetin is a potential new component for use in cosmetics as an anti-aging agent that improves the barrier function of skin.

Bullous ichthyosiform erythroderma: case presentation

Bullous Ichthyosiform Erythroderma [BIE] is a rare disorder of keratinization [mutation in either keratin 1 or 10]. It typically presents with fragile skin, which gives way to gradual evolution of hyperkeratosis. Flaccid blisters, peeling and superficial erosions at sites of minor trauma or friction are apparent within the first few hours of life. Yellow-brown, waxy, ridged or corrugated scale builds up in skin creases, sometimes forming spiny [Hystrix] outgrowths. Cobblestone-like keratoses occur at other sites such as the dorsal hands and feet and over the trunk. We report an 11-year-old boy with a generalized hyperkeratosis on the neck, trunk, extremities and scalp

Screening of differential proteins binding to Nox1 promoter in A549 cell model of inflammation and oxidative stress / 南方医科大学学报

<p><b>OBJECTIVE</b>To screen the regulatory proteins involved in Nox1 promoter activation in a cell model of inflammation and oxidative stress.</p><p><b>METHODS</b>A cell model of inflammation and oxidative stress was established by stimulating A549 cells with tumor necrosis factor-α (TNF-α). The differential proteins binding to Nox1 promoter were screened by DNA pull-down and the binding proteins were separated by 2D electrophoresis and selected according to the their differential expression levels (with over 1.5-fold changes relative to the control level). The screened proteins were finally identified by MALDI-TOF/TOF-MS.</p><p><b>RESULTS</b>Seven differentially expressed protein spots (all upregulated in the cell model) were obtained, among which GLE1, DDX19A, KRT1 and KRT10 were identified by mass spectrometry.</p><p><b>CONCLUSION</b>GLE1, DDX19A, KRT1 and KRT10 participate in the activation of Nox1 promoter in TNF-α-induced A549 cells, and this result provides new insights into the biological roles of the regulatory proteins of Nox1 promoter in inflammation and oxidative stress.</p>

A case of isolated metastatic hepatocellular carcinoma arising from the pelvic bone / 대한간학회지

Reports of metastatic hepatocellular carcinoma (HCC) without a primary liver tumor are rare. Here we present a case of isolated HCC that had metastasized to the pelvic bone without a primary focus. A 73-year-old man presented with severe back and right-leg pain. Radiological examinations, including computed tomography (CT) and magnetic resonance imaging (MRI), revealed a huge mass on the pelvic bone (13x10 cm). He underwent an incisional biopsy, and the results of the subsequent histological examination were consistent with metastatic hepatocellular carcinoma. The tumor cells were positive for cytokeratin (AE1/AE3), hepatocyte paraffin 1, and glypican-3, and negative for CD56, chromogranin A, and synaptophysin on immunohistochemical staining. Examination of the liver by CT, MRI, positron-emission tomography scan, and angiography produced no evidence of a primary tumor. Radiotherapy and transarterial chemoembolization were performed on the pelvic bone, followed by systemic chemotherapy. These combination treatments resulted in tumor regression with necrotic changes. However, multiple lung metastases developed 1 year after the treatment, and the patient was treated with additional systemic chemotherapy.

Mutation analysis of a Uighur family with epidermolytic palmoplantar keratoderma / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To map and identify the disease gene for the epidermolytic palmoplantar keratoderma (EPPK) in a Uighur family of China.</p><p><b>METHODS</b>Blood samples were collected and genomic DNA was extracted from 48 members of the Xinjiang Uighur family. Six microsatellite repeat sequences on chromosome region 17q12-q21 and 12q13 were selected based on the two known candidate genes KRT9 and KRT1. Two-point linkage analysis and haplotype analysis were performed. Exons and their flanking intronic sequence of the KRT9 gene were amplified by polymerase chain reaction (PCR) and sequenced.</p><p><b>RESULTS</b>Data from the marker D17S1787 suggested linkage and yielded a Lod score of 8.65 at theta=0 by using MLINK software. Genotypes and haplotypes were acquired. The disease gene of the EPPK family is located between markers 17/TG/36620115 and D17S846. Chromosome 12q13 region was excluded with the negative Lod score obtained in marker D12S96 (Lod=-infinity at theta=0). No pathogenic mutation was detected in the KRT9 gene.</p><p><b>CONCLUSION</b>The disease gene of the EPPK family is located on chromosome region 17q21.2. The keratin 9 gene might not be the disease gene.</p>

The Effects of Adipose Tissue-Derived Mesenchymal Stem Cells on the Formation ofEpidermis and Basement Membrane in Artificial Skin Models / 대한피부과학회지

BACKGROUND: There is an increasing need for making a more ideal artificial skin model. OBJECTIVE: To evaluate the effects of adipose tissue-derived mesenchymal stem cells (ATMSC) on the formation of epidermis and basement membrane in artificial skin models. METHODS: ATMSC were isolated from lipo-aspirated fat tissues, and their phenotypes were confirmed by cell surface markers. Three kinds of artificial skin models were made using three different dermal substitutes. The dermal substitutes in the three models contained fibroblasts only, fibroblasts together with ATMSC or ATMSC only. The formation of epidermis and basement membrane was evaluated by immunohistochemical stains and transmission electron microscopy. RESULTS: Among the three models, the model with both fibroblasts and ATMSC in the dermal substitute showed the most excellent formation of epidermis and, especially, basement membrane. In this model, the basement membrane proteins, laminin and type IV collagen, were expressed most apparently at the dermo-epidermal junction and, lamina lucida, lamina densa and anchoring fibrils were most evidently observed under transmission electron microscopy. Whereas, the model with only ATMSC did not show keratin 1 expression, suggesting that the 'skin-type' stratified squamous epithelium was not formed well. CONCLUSION: ATMSC together with fibroblasts can be used effectively in constructing artificial skin models.

Organotypic Culture of HaCaT cells: Use of Dermal Substrate that Combines de-epidermized Dermis with Fibroblast-populated Collagen Matrix

BACKGROUND: The immortalized human keratinocyte line, HaCaT cells have been widely used as substitutes for normal epidermal keratinocytes. Recently, reconstruction of a skin equivalent using HaCaT cells showed a multilayered epithelium,but somewhat different tissue architecture as compared with normal epidermis. OBJECTIVE: In this study, using HaCaT cells we tried to reconstruct an epidermis resembling more closely to normal epidermis than the previous results. MATERIALS AND METHODS: HaCaT cells were cultured in air-liquid interface on a recently developed dermal substrated in our laboratory, de-epidermized dermis (DED) raised on fibroblast-populated collagen matrix and the result was compared with those on DED or fibroblast-populated collagen matrix alone. RESULTS: HaCaT cells on the new dermal substrate formed a multilayered epithelium with rete ridges, showing rather orderly cellular organization compared with those on fibroblast-populated collagen matrix. However, horny and granular layers were not observed contrary to normal epidermis. Immunohistochemical studies revealed that differentiation markers such as keratin 1, keratin 6 and involucrin showed the similar pattern to those in HaCaT cells cultured on fibroblast-populated collagen matrix. Markers of terminal differentiation, loricrin and filaggrin were not expressed contrary to normal epidermis. CONCLUSION: These results suggest that organotypic culture HaCaT cells on the dermal substrate combines DED with fivroblast-populated collagen matrix results in incomplete differentiation of HaCaT cells contrary to normal keratinocytes.

Is epithelial membrane antigen a better prognostic factor than keratin proteins in colorectal carcinoma?: a comparative immunohistochemical study

The heterogeneous nature of tumour antigen expression may require the selection of monoclonal antibodies on an individual tumour to allow its adequate localization. Epithelial Membrane Antigen [EMA] and Cytokeratins [CK] expressions are not previously been compared in colorectal cancer patients. Sections of colorectal cancer [n= 32] were examined by monoclonal antibodies to EMA; CK. In the normal mucosa adjacent to the tumours, EMA was weakly expressed in 8 cases [25%], and negative in 24 cases [75%] recording mean weighted score of 0.43[ +/- 0.84]. CK expression was positive in the normal mucosa in all cases [100%] recording mean weighted score of 10.4 [ +/- 2.01]. This indicates that EMA is more specific marker than CK in colorectal carcinomas [80% and 50% respectively]. All primary colorectal cancers expressed EMA [100%] while 30 of 32 expressed CK [93.7%]. These results suggest that EMA is more sensitive than CK expression for colorectal cancer. The mean weighted score of EMA staining density was 6.4 [ +/- 4.2] in all grades while it recorded 6.8 [ +/- 4.5] for CK staining density. These results showed no significant difference between EMA and CK expression in all grades of differentiation of the tumours, and highly significant differences in their expression in the normal mucosa when compared with that in the malignant tissue [P<0.005]. Regarding tumour staging; the mean weighted score of EMA staining density was 8.57 [ +/- 4.07] in Dukes stage A; it recorded 4.75[ +/- 4.55] in stage B7 and 6.29[ +/- 4.07] in stage B2. The mean weighted score of CK staining density was 4.14 [ +/- 4.33], 5.87 [ +/- 4.35] and 8.35[ +/- 4.22] respectively. There were no significant differences in EMA expression in the different stages of colorectal carcinoma while CK expression in stages A and B1 and stage B2 was significantly different. EMA expression is negatively correlated with the tumour grade of differentiation and stage, while CK expression was negatively correlated with the tumour grade of differentiation and positively correlated with the stage of colorectal cancer. The combinations of monoctonal antibodies directed against distinct tumour - associated antigens such as EMA and CK may overcome the problem of heterogeneity of antigen expression and improve both the immunolocalization and potential for targeted therapy of monoclonal antibodies to patients with colorectal cancer. These findings lead us to recommend the selection of both Cytokeratins and Epithelial Membrane Antigen as prognostic factors in colorectal carcinoma. Thus selecting monoclonal antibodies markers based on tumour biopsies allow improved tumour localization for imaging or therapy in patients with colorectal cancer

Effects of Calcipotriol ( MC 903 ) on Proliferation and Differentiation of Cultured Human Epidermal Keratinocytes / 대한피부과학회지

BACKGROUND: Calcipotriol(MC903), a new vitamin D(3) analogue, has been reported to be effective in the treatment of patients with psoriasis. OBJECTIVE: The purpose of this study is to examine the effects of calcipotriol on proliferation and differentiation of the keratinocytes in monolayer cultures and three-dimensional cultures. METHODS: Using moriolayer cultures, we examined morphological changes of keratinocytes and performed [(3)H]thymidine incorporation after calcipotriol was added into the medium. Using three dimensional cultures, we performed two experiments: one with cultures treated with calcipotriol immediately after the keratinocytes had been exposed to the air and another set of cultures treated with calcipotriol after three dimensional morphogenesis of the keratinocytes. We examined morphological changes of keraitinocytes and performed a immunohistochemical study for proliferation differentiation markers RESULTS: In monolayer cultures, at calcipotriol concentrations of 10(-9)M-10(-6)M, keratinocytes became larger, more irregular, and flattened in a dose-dependent manner. At 10(-9)M-10(-6)M, [3Hl thymidine incorporatiorn was decreased dose-dependently as compared to the control culture. In the first experiment using three-dimensional cultures, at 10(-9)M-10(-6)M, total epidermal layers were thinned. This was associated with thinnings of nucleated and horny layers in a dose dependent manner. In the seconcd experiment using three-dimensional cultures, at 10(-8)M-10(-6)M, nucleated layers were thinned in a dose dependent manner, but the horny layer was slightly thickened, as compared to the control culture. Immunohistochemical studies showed a reduction of differentiation markers such as keratin 1, involucrin, filaggrin, loricrin consistent with a thinning of nucleated layers in the epidermal architecture in both experiments. In the basal layer, at 10(-9)M-10(-6), PCNA-positive cells were and BrdU-positive cells were decreased dose-dependently as compared to the control culture. CONCLUSIONS: In this study, we demonstrated that at 10(-9)M-10(-6) calcipotriol inhibited keratinocytes proliferation and stimulated keratinocytes differentiation in a dose-dependent manner.

Expression of Epidermal Protein Antigens and Basement Membrane Components in Human Epidermis Reconstructed by Culture / 대한피부과학회지

BACKGROUND: A number of in vitro skin models have been developed for the purpose of the screening of cosmetics, pharmaceuticals, and environmental chemicals. To mimic the skin in vivo, a model should resemble morphologically and biochemically the parent, tissue. OBJECTIVE: The purpos of this study is to study the differentiation and organization of the artificial epidermis in comparsion with epidermis in vivo based on the expression of epidermal protein antigens and basement membrane components. METHODS: Human keratinocytes were cultured on deepidermidized dermis (RE-DED) or on fibroblast-populated collag-,n matrix (LSE). After 10 days culture, the sections of RE-DED and LSE were stained with hematoxylin and eosin. An immunohistochemical study was also performed with the sections of RE-DED and LSE using antibodies recognizing proliferating cell nuclear antigens (PCNA), epidermal growth factor receptor (EGFR), keratin 1, involucrin, filaggrin, loricrin, keratin 13, type IV collagen, and laminin. RESULTS: In both culture systems(RE-DED and LSE) a multilayered epidermis with a horny layer was observed. In the human epidermis reconstructed by both culture systems, differentiation markers appeared but with a topography slightly different from that of epidermis in vivo, and components of the basement membrane was also expressed. CONCLUSION: Our findings suggest the epidermis obtained in both culture systems(RE-DED and LSE) resembled in vivo epidermis morphologically and biochemically, although it was not the same.

Expression of Epidermal Protein Antigens and Basement Membrane Components in Human Epidermis Reconstructed by Culture / 대한피부과학회지

BACKGROUND: A number of in vitro skin models have been developed for the purpose of the screening of cosmetics, pharmaceuticals, and environmental chemicals. To mimic the skin in vivo, a model should resemble morphologically and biochemically the parent, tissue. OBJECTIVE: The purpos of this study is to study the differentiation and organization of the artificial epidermis in comparsion with epidermis in vivo based on the expression of epidermal protein antigens and basement membrane components. METHODS: Human keratinocytes were cultured on deepidermidized dermis (RE-DED) or on fibroblast-populated collag-,n matrix (LSE). After 10 days culture, the sections of RE-DED and LSE were stained with hematoxylin and eosin. An immunohistochemical study was also performed with the sections of RE-DED and LSE using antibodies recognizing proliferating cell nuclear antigens (PCNA), epidermal growth factor receptor (EGFR), keratin 1, involucrin, filaggrin, loricrin, keratin 13, type IV collagen, and laminin. RESULTS: In both culture systems(RE-DED and LSE) a multilayered epidermis with a horny layer was observed. In the human epidermis reconstructed by both culture systems, differentiation markers appeared but with a topography slightly different from that of epidermis in vivo, and components of the basement membrane was also expressed. CONCLUSION: Our findings suggest the epidermis obtained in both culture systems(RE-DED and LSE) resembled in vivo epidermis morphologically and biochemically, although it was not the same.