砷剂对角质形成细胞生长和IL-8、TNF-α分泌的影响

目的研究不同浓度砷剂对角质形成细胞生长和IL-8、TNF-α分泌的影响。方法以体外培养的角质形成细胞株为对象,不同浓度砷剂处理细胞后,用MTT比色分析法反映细胞增殖的变化。ELISA方法测定细胞上清中IL-8分泌,用L929细胞检测上清TNF-α的生物活性。结果0.5～5μmol/L三氧化二砷对角质形成细胞株HaCaT细胞有明显抑制作用,并呈剂量依赖关系。在0.001μmol/L至0.015μmol/L药物浓度范围内,同对照组相比细胞增殖加快;未药物处理的对照组HaCaT细胞可分泌一定水平的IL-8和TNF-α,低浓度砷剂(0.001～0.015μmol/L)可刺激其分泌增加。结论低浓度砷剂可促进角质形成细胞株HaCaT细胞的生长,但高浓度能抑制增殖和影响细胞活性。并在一定浓度范围刺激IL-8和TNF-α的合成分泌,可能在砷相关皮肤病的发病机制中具有重要意义。     

Ultraviolet B radiation regulates cysteine‐rich protein 1 in human keratinocytes

Background: Cysteine‐rich protein 1 (CRP1) is a growth‐inhibitory cytoskeletal protein that is induced by ultraviolet (UV) C radiation radiation in fibroblasts. Our aim was to investigate the effects of UV radiation on CRP1 in keratinocytes, the main cell type subjected to UV radiation in the human body. Methods: The effects of physiologically relevant doses of UVB radiation on CRP1 protein levels were studied in cultured primary keratinocytes and transformed cell lines (HaCaT, A‐431) by immunoblotting. UVB‐induced keratinocyte apoptosis was assessed by flow cytometry and monitoring caspase activity. Expression of CRP1 in human skin in vivo was studied by immunohistochemistry in samples of normal skin, actinic keratosis (AK) representing UV‐damaged skin and squamous cell carcinoma (SCC), a UV‐induced skin cancer. Results: CRP1 expression increased by UVB radiation in primary but not in immortalized keratinocytes. Upon high, apoptosis‐inducing doses of UV radiation, CRP1 was cleaved in a caspase‐dependent manner. In normal skin, CRP1 was expressed in smooth muscle cells, vasculature, sweat glands, sebaceous glands and hair root sheath, but very little CRP1 was present in keratinocytes. CRP1 expression was elevated in basal cells in AK but not in SCC. Conclusion: CRP1 expression is regulated by UVB in human keratinocytes, suggesting a role for CRP1 in the phototoxic responses of human skin.     

Expression of midkine in normal human skin,dermatitis and neoplasms: Association with differentiation of keratinocytes

Midkine is a 13‐kDa heparin‐binding growth factor. It promotes growth, survival, migration and gene expression of various target cells and play roles in many diseases. In normal adult tissues, midkine expression is highly restricted; however, midkine expression levels are high in various malignant tumors. The major biological roles of midkine can be categorized into three areas, namely, the nervous system, cancer and inflammation. Thus far, midkine has not been studied extensively in diseased human skin. We performed immunohistochemistry tests by using anti‐midkine antibodies to study the expression of midkine in normal skin and skin samples of 26 different cutaneous diseases. In addition, we investigated the expression pattern of the midkine gene in cultured keratinocytes. In normal skin, midkine expression was observed in the secretory coils of the eccrine sweat glands, outer root sheath and inner root sheath. Among the cutaneous tumors, the majority of keratinocyte‐derived neoplasms were positive for midkine. Tumors that were not derived from keratinocytes were negative for midkine. In cultured keratinocytes, the midkine gene was expressed earlier than the genes required for keratinization, for example, cytokeratin 10 and transglutaminase 1. Because midkine is expressed in the keratinized areas of normal skin, neoplasms and inflammation, it may play a role as a modulator of keratinization in the skin.     

Leptin induces secretion of pro‐inflammatory cytokines by human keratinocytes in vitro – a possible reason for increased severity of psoriasis in patients with a high body mass index

Investigations about prevalence of obesity in psoriasis patients are increased nowadays. Higher serum levels of leptin in patients with psoriasis who are overweight or obese suggest that leptin may serve as a molecular link between psoriasis and metabolic comorbidities. However, the pathological functions of leptin in psoriasis are not clearly understood. We investigated the influence of being overweight or obese on the risk of psoriasis, and the relationship between serum leptin levels and the severity of psoriasis in Chinese Han patients. We also investigated biological effects of leptin on the proliferation and secretion of pro‐inflammatory cytokines by human keratinocytes in vitro. Obesity was a significant risk factor for psoriasis in the Chinese Han population; however, we did not observe a significant correlation between Psoriasis Area and Severity Index (PASI) and body mass index (BMI). We observed a positive correlation between the serum leptin level and PASI in overweight and obese male patients with psoriasis. Strong leptin immunoreactivity was detected in the epidermis of psoriatic lesions, particularly in keratinocytes. Leptin significantly increased the proliferation and secretion of pro‐inflammatory cytokines by keratinocytes in vitro. In conclusion, this study suggests leptin as a novel molecular link between psoriasis and obesity, which may help to explain the more server conditions of psoriasis in patients with obesity.     

Delphinidin,a dietary antioxidant,induces human epidermal keratinocyte differentiation but not apoptosis: studies in submerged and three‐dimensional epidermal equivalent models

Delphinidin (Del), [3,5,7,3'‐,4'‐,5'‐hexahydroxyflavylium], an anthocyanidin and a potent antioxidant abundantly found in pigmented fruits and vegetables exhibits proapoptotic effects in many cancer cells. Here, we determined the effect of Del on growth, apoptosis and differentiation of normal human epidermal keratinocytes (NHEKs) in vitro in submerged cultures and examined its effects in a three‐dimensional (3D) epidermal equivalent (EE) model that permits complete differentiation reminiscent of in vivo skin. Treatment of NHEKs with Del (10–40 μm ; 24–48 h) significantly enhanced keratinocyte differentiation. In Del‐treated cells, there was marked increase in human involucrin (hINV) promoter activity with simultaneous increase in the mRNA and protein expressions of involucrin and other epidermal differentiation markers including procaspase‐14 and transglutaminase‐1 (TGM1), but without any effect on TGM2. Del treatment of NHEKs was associated with minimal decrease in cell viability, which was not associated with apoptosis as evident by lack of modulation of caspases, apoptosis‐related proteins including Bcl‐2 family of proteins and poly(ADP‐ribose) polymerase cleavage. To establish the in vivo relevance of our observations in submerged cultures, we then validated these effects in a 3D EE model, where Del was found to significantly enhance cornification and increase the protein expression of cornification markers including caspase‐14 and keratin 1. For the first time, we show that Del induces epidermal differentiation using an experimental system that closely mimics in vivo human skin. These observations suggest that Del could be a useful agent for dermatoses associated with epidermal barrier defects including aberrant keratinization, hyperproliferation or inflammation observed in skin diseases like psoriasis and ichthyoses.     

Clinicopathological differentiation between Pityrosporum folliculitis and acneiform eruption

Distinguishing between Malassezia folliculitis (Pityrosporum folliculitis [P. folliculitis]) and acneiform eruption, based on clinicopathological features, is challenging for clinicians. In the literature, the histopathological differences between P. folliculitis and acneiform eruption lesions have been poorly described. We aimed to determine the clinicopathologic distinctions between P. folliculitis and acneiform eruption by retrospectively analyzing the histology of hematoxylin and eosin stained tissue sections obtained from 52 patients diagnosed with these lesions. The presence of fungal spores in the follicular lumen was most consistent with a P. folliculitis diagnosis (P < 0.001). However, intrafollicular inflammation (P = 0.009), irregular patterns of keratin plugging (P = 0.008), and nuclear dust in the follicular lumen (P < 0.001) favored an acneiform eruption diagnosis. These intrafollicular characteristics and inflammatory differences are believed to be caused by necrotic keratinocytes that lead to vacuolar changes in the follicular wall (P = 0.013). We did not observe any difference between P. folliculitis and acneiform eruption lesions in terms of perifollicular inflammatory cell infiltration. Our study demonstrated that significant differences exist between P. folliculitis and acneiform eruption lesions relative to the presence of necrotic keratinocytes in the follicular wall, intrafollicular characteristics, and inflammatory cell infiltrations. Necrotic keratinocytes are believed to have a key role in these differences. These findings may contribute to an improved understanding of the pathogenesis and differential diagnosis of P. folliculitis and acneiform eruption.