长波紫外线对大鼠皮肤角朊细胞的损伤及茶多酚的保护作用

目的 探讨长波紫外线（ＵＶＡ）对原代培养的大鼠皮肤角朊细胞脂质过氧化和生长状况等影响，同时探讨一种植物多酚－－茶多酚（ＴＰＰ）在此过程中所起的作用。方法 在原代培养大鼠皮肤角朊细胞基础上，经ＵＶＡ照射后，测定角朊细胞浆酶－－乳酸脱氢酶（ＬＤＨ）释放情况，脂质过氧化产物丙二醛（ＭＤＡ）、谷胱甘肽过氧化物酶（ＧＳＨ－Ｐｘ）水平，并测定培养细胞的存活率和细胞周期动力学。结果 ＵＶＡ可以引起体外培养的细胞膜通透性增强，胞浆酶ＬＤＨ释放增加（从８２７．５５Ｕ／Ｌ增至１３１２．４７Ｕ／Ｌ）；脂质过氧化产物ＭＤＡ水平升高，抗氧化酶ＧＳＨ－Ｐｘ水平降低；细胞存活率下降，细胞周期动力学表现为细胞增殖抑制；增殖指数（ＰＩ）从３４．２４％降至１７．９８％。天然提取物ＴＰＰ（质量浓度为０．１％）可以比较明显地抑制ＵＶＡ引起的上述损害。     

Parathyroid hormone-related protein expression and secretion in a skin organotypic culture system

Parathyroid hormone-related protein (PTHrP), an important factor in the pathogenesis of humoral hypercalcemia of malignancy, is produced by many normal tissues, including the epidermis, where it is thought to play a role in the regulation of keratinocyte growth and differentiation. Most in vitro studies of normal keratinocytes use monolayer cell cultures, which have limitations, including the inability to reproduce the stratified structure of the epidermis. The objective of this study was to investigate PTHrP production and secretion, and mRNA expression in skin organotypic cultures. The cultures consisted of an artificial dermis with differentiating keratinocytes grown at the air-liquid interface. Immunohistochemical assessment of cytokeratins 14 and 10/13, involucrin, and proliferative cell nuclear antigen (PCNA) demonstrated that keratinocytes differentiated in a manner similar to keratinocytes in normal epidermis. PTHrP expression was demonstrated in all viable layers of the epidermis, as well as in some fibroblasts of the collagen lattice by immunohistochemistry and in situ hybridization. Since most fibroblasts expressed α-smooth muscle actin, these cells were interpreted to be consistent with myofibroblasts. PTHrP expression by myofibroblasts suggests a possible role for PTHrP in the regulation of contractibility of these cells. PTHrP was also detected in conditioned media for 50 days. In conclusion, because of its superior tissue morphology and ability to induce organized keratinocyte differentiation, this culture system will be an excellent model to study the role of PTHrP in pathologic and physiologic processes involving the epidermis in vitro.     

Reversal of age-associated thymic atrophy: treatments, delivery, and side effects

Our ability to survive infectious agents depends on making adequate immune responses, but as we get older our thymus atrophies. Production and export of T cells bearing new antigen receptor specificities to the peripheral T cell pool declines and results in shrinkage of the repertoire. Other changes in the peripheral T cell pool include an increase in cells moving closer to their replicative limit. Age related immune dysfunction, evident through the increased susceptibility to infection, follows these changes. Improvement in immune function in the elderly may require us to rejuvenate the immune system starting first with reversing the atrophy seen in the thymus. This has been achieved experimentally with interleukin 7, growth hormone, growth hormone secretagogues, keratinocyte growth factor or through chemical or surgical castration. The widespread use of one or more of these treatments will depend upon their effectiveness, their ease of delivery and the extent of any side effects.     

KGF在成纤维细胞促进表皮细胞增殖中的作用

目的  研究成纤维细胞产生的角质细胞生长因子（KGF）对表皮细胞增殖的影响。方法  自包皮分离培养成纤维细胞和表皮细胞,采用逆转录-聚合酶链反应法(RT PCR)、酶联免疫吸附法(ELISA)、MTT法和免疫荧光等方法分别检测成纤维细胞KGF基因的转录水平、培养液中KGF的浓度、成纤维细胞培养液对表皮细胞增殖的促进作用。结果  RT-PCR结果显示,成纤维细胞中KGF转录水平显著,ELISA法可检测到成纤维细胞培养液中存在较高水平的KGF,细胞计数结果和MTT结果表明,含有KGF的成纤维细胞培养液能够促进表皮细胞增殖,保持表皮细胞的活性,免疫荧光观察到KGF与表皮细胞表面的特异性受体结合。结论  成纤维细胞能够合成和分泌较高水平的KGF,成为其促进表皮细胞增殖的重要因素。     

Treatment with keratinocyte growth factor does not improve lung allograft survival in the rat

Purpose  Lung allografts are threatened by primary graft dysfunction, infections, and rejection. Novel therapies protecting pulmonary allografts are badly needed. Keratinocyte growth factor (KGF) protects the lung against a variety of injurious stimuli and exerts anti-inflammatory effects. The aim of the study was to test the potential of recombinant truncated KGF (ΔN23-KGF, palifermin) to attenuate pulmonary allograft rejection. Materials and methods  Intratracheal instillation of 5 mg/kg ΔN23-KGF was performed twice in donor rats on days 3 and 2 before explantation of the lung. In control animals, an equivalent volume of vehicle was instilled. Left lungs were transplanted in the fully allogeneic Dark Agouti to Lewis rat strain combination and in the less stringent Fischer 344 to Wistar Kyoto combination. Allograft recipients were additionally treated with ΔN23-KGF post-transplantation. Graft outcome, leukocytic infiltration, and major histocompatibility complex (MHC) class II antigen expression was analyzed. Results  In both rat strain combinations, ΔN23-KGF treatment did not improve pulmonary allograft outcome. Graft infiltration by macrophages and T lymphocytes remained unchanged. In addition, we demonstrated that MHC class II antigens were more abundant in KGF-treated allografts compared to control-treated grafts, which probably results in an increased alloreactivity. Conclusion  In conclusion, intratracheal ΔN23-KGF treatment is not effective to prevent acute pulmonary allograft rejection.     

Human papillomavirus 16 E6 variants differ in their dysregulation of human keratinocyte differentiation and apoptosis

L83V-related variants of human papillomavirus (HPV) 16 E6, exemplified by the Asian-American variant Q14H/H78Y/L83V, were shown to be more prevalent than E6 prototype in progressing lesions and cervical cancer. We evaluated functions relevant to carcinogenesis for the E6 variants L83V, R10/L83V and Q14H/H78Y/L83V as well as the prototype in a model of human normal immortalized keratinocytes (NIKS). All E6 expressing NIKS equally abrogated growth arrest and DNA damage responses. Organotypic cultures derived from these keratinocytes demonstrated hyperplasia and aberrantly expressed keratin 5 in the suprabasal compartment. In contrast, differentiation and induction of apoptosis varied. The E6 variant rafts expressed keratin 10 in nearly all suprabasal cells while the prototype raft showed keratin 10 staining in a subset of suprabasal cells only. In addition, E6 variant NIKS expressing R10G/L83V and Q14H/H78Y/L83V were more prone to undergo cell-detachment-induced apoptosis (anoikis) than NIKS expressing E6 prototype. The combined differentiation and apoptosis pattern of high-risk E6 variants, especially of Q14H/H78Y/L83V, may reflect a phenotype beneficial to carcinogenesis and viral life cycle.     

组织工程皮肤细胞对朗格汉斯细胞表型的影响

目的:研究异体组织工程皮肤种子细胞(角质形成细胞、成纤维细胞)对朗格汉斯细胞的表型影响.方法:外周血单个核细胞在粒细胞巨噬细胞集落刺激因子(GM-CSF)、白介素4(IL-4)和转化生长因子β1(TGF-β1)的诱导下培养出朗格汉斯细胞.培养的朗格汉斯细胞与异体组织工程皮肤种子细胞共同培养后,检测其表型变化情况.随后将朗格汉斯细胞与淋巴细胞共培养后检测淋巴细胞的增殖程度.结果:诱导培养的朗格汉斯细胞低表达HLA-DR、CD80和CD86,不表达CD83.与异体组织工程皮肤种子细胞共培养后朗格汉斯细胞表型无明显变化,共培养后的朗格汉斯细胞无法刺激自体淋巴细胞增殖.结论:与异体组织工程皮肤种子细胞共培养后,朗格汉斯细胞仍保持不成熟的状态,这提示组织工程皮肤种子细胞免疫原性较低,移植后不易引起宿主的免疫排斥反应.     

Inhibitory effects of Cinnamomum cassia extract on atopic dermatitis-like skin lesions induced by mite antigen in NC/Nga mice

Aim of the study

: Cinnamomum cassia (C. cassia) has been traditionally used to treat allergic disease as well as dyspepsia, gastritis, and blood circulation disturbances. However, the antiallergic properties of C. cassia have not been fully verified using scientific tools. This study investigated the effectiveness of C. cassia extract (CCE) as an antiallergic agent in atopic dermatitis model and underlying mechanism.

Materials and methods

: The effect of CCE on mite antigen-treated NC/Nga mice was evaluated by examining skin symptom severity, levels of serum IgE, tumor necrosis factor-α (TNF-α), and histamine, skin histology, and mRNA expression of cytokines in the skin lesions. Moreover, the effect of CCE on TNF-α-and interferon-γ (IFN-γ)-induced chemokine production in human keratinocytes was investigated using ELISA.

Results

: CCE treatment of NC/Nga mice reduced the dermatitis score and the levels of serum IgE, histamine, and TNF-α. Histological examination showed inhibition of the thickening of the epidermis/dermis and reduced dermal infiltration of inflammatory cells. In skin lesions, mRNA expression of IL-4, TNF-α, and thymus and activation-regulated chemokine (TARC) was inhibited by CCE treatment. The production of TARC, macrophage-derived chemokine, and RANTES from IFN-γ-and TNF-α-stimulated human keratinocytes was suppressed by CCE treatment in a dose-dependent manner.

Conclusions

: CCE inhibits the development of atopic dermatitis-like skin lesions in NC/Nga mice by suppressing the T-helper 2 cell response.     

Immunomodulatory IL-23 receptor antagonist peptide nanocoatings for implant soft tissue healing

ObjectivePeri-implantitis, caused by an inflammatory response to pathogens, is the leading cause of dental implant failure. Poor soft tissue healing surrounding implants – caused by inadequate surface properties – leads to infection, inflammation, and dysregulated keratinocyte and macrophage function. One activated inflammatory response, active around peri-implantitis compared to healthy sites, is the IL-23/IL-17A cytokine axis. Implant surfaces can be synthesized with peptide nanocoatings to present immunomodulatory motifs to target peri-implant keratinocytes to control macrophage polarization and regulate inflammatory axises toward enhancing soft tissue healing.MethodsWe synthesized an IL-23 receptor (IL-23R) noncompetitive antagonist peptide nanocoating using silanization and evaluated keratinocyte secretome changes and macrophage polarization (M1-like “pro-inflammatory” vs. M2-like “pro-regenerative”).ResultsIL-23R antagonist peptide nanocoatings were successfully synthesized on titanium, to model dental implant surfaces, and compared to nonfunctional nanocoatings and non-coated titanium. IL-23R antagonist nanocoatings significantly decreased keratinocyte IL-23, and downstream IL-17A, expression compared to controls. This peptide noncompetitive antagonistic function was demonstrated under lipopolysaccharide stimulation. Large scale changes in keratinocyte secretome content, toward a pro-regenerative milieu, were observed from keratinocytes cultured on the IL-23R antagonist nanocoatings compared to controls. Conditioned medium collected from keratinocytes cultured on the IL-23R antagonist nanocoatings polarized macrophages toward a M2-like phenotype, based on increased CD163 and CD206 expression and reduced iNOS expression, compared to controls.SignificanceOur results support development of IL-23R noncompetitive antagonist nanocoatings to reduce the pro-inflammatory IL-23/17A pathway and augment macrophage polarization toward a pro-regenerative phenotype. Immunomodulatory implant surface engineering may promote soft tissue healing and thereby reduce rates of peri-implantitis.     

抗角蛋白自身抗体对培养角朊细胞产生白细胞介素8的影响

为探讨抗角蛋白自身抗体（AKantoAb）对角朊细胞免疫功能的影响，实验以IL-1β刺激无血清培养角朊细胞及AKautoAb作用后的角朊细胞，用夹心ELISA法检测培养上清中IL－8的产量。结果表明IL-1β可刺激角朊细胞产生IL－8，并对IL－1β有浓度依赖性，AKautoAb对角朊细胞产生IL－8有明显抑制作用。提示AKautoAb在炎性皮损角朊细胞中的沉着，可能存在有益的调节作用。