本文選題：銀屑病 + 鈣離子��； 參考：《河北大學(xué)》2017年碩士論文

【摘要】：銀屑病為皮膚科常見病,其發(fā)病機(jī)制復(fù)雜,多認(rèn)為與遺傳、免疫、感染、外界環(huán)境等因素有關(guān)。臨床上銀屑病的治療方法雖多,但也只能達(dá)到暫時(shí)緩解的目的,且銀屑病病程較長(zhǎng),反復(fù)發(fā)作不易根治,嚴(yán)重危害人們的身心健康。白細(xì)胞介素(interleukin,IL)6是一種重要的細(xì)胞因子,在銀屑病患者皮損處表達(dá)增高并可被角質(zhì)形成細(xì)胞合成分泌。鈣離子作為細(xì)胞內(nèi)重要的第二信使參與機(jī)體凝血、肌肉收縮、心臟節(jié)律正常等多種生理功能外,對(duì)角質(zhì)形成細(xì)胞的增殖、分化也有一定影響。為了探討鈣離子與IL-6之間是否存在相關(guān)性,本研究通過觀察不同濃度鈣離子對(duì)體外培養(yǎng)人角質(zhì)形成細(xì)胞株HaCaT增殖的影響及對(duì)IL-6 mRNA表達(dá)、蛋白分泌的影響初步分析兩者間的相關(guān)性。目的:探究鈣離子對(duì)人角質(zhì)形成細(xì)胞株HaCaT增殖的影響及對(duì)IL-6mRNA表達(dá)、蛋白分泌的影響。方法:用加入不同濃度鈣離子的洛斯維RPMI1640培養(yǎng)液[鈣離子終濃度分別為0.3毫摩爾/升(mmol/L)、0.5mmol/L、1mmol/L]與RPMI1640培養(yǎng)液(對(duì)照組)分別培養(yǎng)HaCaT細(xì)胞。培養(yǎng)HaCaT細(xì)胞24h、48h時(shí),二甲基噻唑二苯基四鹽(MTT)法檢測(cè)細(xì)胞增殖情況;熒光定量聚合酶鏈反應(yīng)(qPCR)法檢測(cè)鈣離子對(duì)HaCaT細(xì)胞IL-6信使核糖核酸(messenger ribonucleic acid,mRNA)表達(dá)的影響;酶聯(lián)免疫吸附法(ELISA)檢測(cè)不同鈣離子濃度培養(yǎng)條件下HaCaT細(xì)胞上清液中IL-6的分泌量;免疫細(xì)胞化學(xué)法觀察細(xì)胞中IL-6的表達(dá),其結(jié)果用專業(yè)圖像分析軟件(Image Pro Plus)6.0進(jìn)行分析。采用獨(dú)立樣本t檢驗(yàn)、單因素方差分析和皮爾遜(Pearson)相關(guān)分析進(jìn)行統(tǒng)計(jì)學(xué)處理。結(jié)果:MTT結(jié)果顯示,在設(shè)定濃度下HaCaT細(xì)胞的增殖率隨著鈣離子濃度的增加而增高;與對(duì)照組相比,各濃度組培養(yǎng)24h時(shí)HaCaT細(xì)胞增殖率分別為9.65%?0.57%、29.47%?0.73%、52.18%?1.13%(t=29.08,69.12,79.52,P均0.01);培養(yǎng)48h時(shí)各濃度組HaCaT細(xì)胞增殖率分別為10.23%?0.51%、34.17%?1.14%、57.96%?0.21%(t=35.36,51.67,87.37,P均0.01)。各濃度組培養(yǎng)HaCaT細(xì)胞24h時(shí),IL-6 mRNA表達(dá)量及其蛋白分泌量隨鈣離子濃度增加而升高(F=133.07,121.68,P0.01);培養(yǎng)HaCaT細(xì)胞48h時(shí),IL-6 mRNA表達(dá)量及其蛋白分泌量隨鈣離子濃度增加而升高(F=151.17,78.64,P0.001)。免疫細(xì)胞化學(xué)結(jié)果顯示,培養(yǎng)HaCaT細(xì)胞24h、48h時(shí),IL-6平均光密度隨鈣離子濃度增加而升高(F=113.22,202.05,P0.01)。相關(guān)性分析結(jié)果顯示,培養(yǎng)HaCaT細(xì)胞24h時(shí),鈣離子與IL-6 mRNA表達(dá)和蛋白分泌均呈正性相關(guān)(r=0.97、r=0.83,P0.01),培養(yǎng)HaCaT細(xì)胞,48h時(shí),鈣離子與IL-6 mRNA表達(dá)和蛋白分泌均呈正性相關(guān)(r=0.98、0.91,P0.01)。結(jié)論:在設(shè)定濃度下,鈣離子可促進(jìn)HaCaT細(xì)胞的體外增殖,并可在mRNA和蛋白水平促進(jìn)IL-6的表達(dá)和分泌。這可能是銀屑病皮損處鈣濃度增高導(dǎo)致角質(zhì)形成細(xì)胞增殖和炎癥反應(yīng)加重的原因。
[Abstract]:Psoriasis is a common disease in the Department of dermatology. Its pathogenesis is complicated, and most of them are related to the factors of heredity, immunity, infection and external environment. Although the treatment of psoriasis is more and more, it can only achieve the purpose of temporary relief, and the course of psoriasis is long, repeated attacks are not easy to cure, and it seriously endangers people's physical and mental health. Interleukin (inter) Leukin, IL) 6 is an important cytokine, which is expressed in the skin lesions of patients with psoriasis and can be synthesized and secreted by keratinocytes. Calcium ions, as an important second messenger in the cells, participate in the body's coagulation, muscle contraction, and normal cardiac rhythm, and have a certain effect on the proliferation and differentiation of keratinocytes. To investigate whether there is a correlation between calcium ion and IL-6, the effect of calcium ions on the proliferation of human keratinocyte cell line HaCaT in vitro and the effect of IL-6 mRNA expression and protein secretion on the proliferation of human keratinocyte strain were investigated. Influence and effect on the expression of IL-6mRNA and protein secretion. Methods: the culture solution of RPMI1640 (0.3 milmole / liter (mmol/L), 0.5mmol/L, 1mmol/L] and RPMI1640 culture solution) was cultured with different concentration of calcium ions, and HaCaT fine cells were cultured respectively. HaCaT cells 24h, 48h, two methyl thiazole two phenyl four salt (MTT) assay of cell proliferation; fluorescence quantitative polymerase chain reaction (qPCR) method to detect the effect of calcium ion on the expression of IL-6 messenger RNA (messenger ribonucleic acid, mRNA) in HaCaT cells; enzyme linked immunosorbent assay (ELISA) detection of IL-6 secretion in the supernatant of HaCaT cells under different calcium concentration cultures; immunization The expression of IL-6 in the cells was observed by the method of study. The results were analyzed with professional image analysis software (Image Pro Plus) 6. Independent sample t test, single factor ANOVA analysis and Pearson (Pearson) correlation analysis were used for statistical processing. Results: MTT results showed that the proliferation rate of HaCaT cells increased with the increase of calcium ion concentration under the set concentration. The proliferation rate of HaCaT cells in each concentration group was 9.65%? 0.57%, 29.47%? 0.73%, 52.18%? 1.13% (t=29.08,69.12,79.52, P 0.01), compared with the control group, and the proliferation rates of HaCaT cells in each concentration group were 10.23%? 0.51%, 34.17%? 1.14%, 57.96%? 0.21% (t=35.36,51.67,87.37, P are all 0.01) at the time of 48h culture. Each concentration group was cultured for HaCaT fine. When the cell 24h, the expression of IL-6 mRNA and its protein secretion increased with the increase of calcium ion concentration (F=133.07121.68, P0.01). When the HaCaT cell 48h was cultured, the mRNA expression of IL-6 and its protein secretion increased with the increase of calcium ion concentration (F=151.17,78.64, P0.001). The density increased with the increase of calcium ion concentration (F=113.22202.05, P0.01). The results of correlation analysis showed that the calcium ions were positively correlated with the expression of IL-6 mRNA and protein secretion (r=0.97, r=0.83, P0.01) when cultured HaCaT cells 24h (r=0.97, r=0.83, P0.01), and 48h, calcium ions were positively correlated with the expression of IL-6 and protein secretion. 1). Conclusion: at the set concentration, calcium ions can promote the proliferation of HaCaT cells in vitro, and promote the expression and secretion of IL-6 at the level of mRNA and protein. This may be the cause of the increase of calcium concentration in the skin lesions of psoriasis and the aggravation of the proliferation of keratinocytes and the aggravation of the inflammatory reaction.
【學(xué)位授予單位】：河北大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R758.63

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