HIST1H2BK蛋白与慢性鼻窦炎发病机制的相关性研究

目的　探索HISH2BK基因及其编码的蛋白在慢性鼻窦炎发病机制中的作用。方法　选取慢性鼻窦炎伴鼻息肉（chronic rhinosinusitis with nasal polyps,CRSwNP）患者6例,鼻中隔偏曲、脑脊液鼻漏修补等非鼻腔炎症行鼻内镜手术患者7例作为对照,获取鼻息肉及黏膜组织进行二代基因测序,应用Reactome通路数据库进行通路分析后发现目标基因HIST1H2BK,应用PCR技术验证HIST1H2BK基因的表达水平,应用免疫组化染色定位HIST1H2BK蛋白在鼻息肉及正常黏膜组织中的表达位置。结果　HIST1H2BK蛋白主要表达于鼻腔黏膜上皮细胞层,HIST1H2BK基因在慢性鼻窦炎鼻息肉患者中低表达。结论　HIST1H2BK基因及其编码的蛋白可能是参与慢性鼻窦炎发病机制的重要因子。     

鼻息肉组织免疫相关基因的表达谱

目的　运用基因芯片技术研究鼻息肉组织免疫相关基因的表达谱 ,并探讨相关基因在鼻息肉免疫发病机制中的作用。方法　将按微距阵排列的 4 91条免疫相关全长基因制成表达谱芯片。分别抽提 4例鼻息肉组织和下鼻甲组织的总RNA ,逆转录成互补DNA(complementaryDNA ,cDNA)一链 ,并分别用cy5和cy3标记制备成杂交探针 ,混合后在 2张免疫相关基因芯片上进行杂交 ,用扫描仪扫描芯片荧光信号图像 ,并用软件对扫描图像进行数字化处理和分析。结果　鼻息肉组织免疫基因表达谱中差异表达的基因共 87条 ,其中上调基因 4 5条 ,下调基因 4 2条 ;2张芯片共存的差异基因 15条 ,其中上调基因 5条 ,下调基因 10条。在差异表达的基因中 ,主要包括细胞因子及其受体、趋化因子及其受体、黏附分子、白细胞分化抗原、免疫信号传导分子 ,还包括一些补体分子及其受体、免疫转录调节分子、天然免疫分子和神经免疫相关基因。结论　鼻息肉免疫相关基因表达谱中差异表达基因为研究鼻息肉免疫学发病机理提供了线索和理论依据 ,其中IL 17可能在鼻息肉发病中发挥一定的作用 ,天然免疫和免疫信号传导分子在鼻息肉发病中的作用还需要进一步研究。     

鼻息肉组织免疫相关基因的表达谱

目的运用基因芯片技术研究鼻息肉组织免疫相关基因的表达谱，并探讨相关基因在鼻息肉免疫发病机制中的作用。方法将按微距阵排列的491条免疫相关全长基因制成表达谱芯片。分别抽提4例鼻息肉组织和下鼻甲组织的总RNA，逆转录成互补DNA(complementary DNA，cDNA)一链，并分别用cy5和cy3标记制备成杂交探针，混合后在2张免疫相关基因芯片上进行杂交，用扫描仪扫描芯片荧光信号图像，并用软件对扫描图像进行数字化处理和分析。结果鼻息肉组织免疫基因表达谱中差异表达的基因共87条，其中上调基因45条，下调基因42条；2张芯片共存的差异基因15条，其中上调基因5条，下调基因10条。在差异表达的基因中，主要包括细胞因子及其受体、趋化因子及其受体、黏附分子、白细胞分化抗原、免疫信号传导分子，还包括一些补体分子及其受体、免疫转录调节分子、天然免疫分子和神经免疫相关基因。结论鼻息肉免疫相关基因表达谱中差异表达基因为研究鼻息肉免疫学发病机理提供了线索和理论依据，其中IL-17可能在鼻息肉发病中发挥一定的作用，天然免疫和免疫信号传导分子在鼻息肉发病中的作用还需要进一步研究。     

转化生长因子β受体Ⅰ型及Ⅱ型在慢性鼻-鼻窦炎和鼻息肉组织中的表达

目的:探讨转化生长因子β受体(TGFβR)Ⅰ型及Ⅱ型在慢性鼻-鼻窦炎、鼻息肉及正常下鼻甲组织中表达的差异性,以及Ⅰ、Ⅱ型受体在慢性鼻窦炎、鼻息肉发病机制中可能的作用。方法:采用免疫组织化学方法检测TGFβRⅠ、TGFβRⅡ在25例慢性鼻-鼻窦炎、21例鼻息肉、17例下鼻甲组织中的表达,并比较TGFβRⅠ、TGFβRⅡ在慢性鼻-鼻窦炎、鼻息肉、正常下鼻甲组织中表达的差异。结果:TGFβRⅠ、TGFβRⅡ表达的平均灰度值在正常下鼻甲黏膜分别为175.78±7.06、165.00±1.79;在慢性鼻-鼻窦炎组织中分别为147.33±8.15、147.77±4.62;而在鼻息肉组织中分别为125.91±11.26、129.82±1.46。慢性鼻-鼻窦炎及鼻息肉组织中TGFβRⅠ、TGFβRⅡ的表达均比正常下鼻甲黏膜中的表达高,均差异有统计学意义(均P<0.01);且TGFβRⅠ、TGFβRⅡ在鼻息肉组织中的表达比在慢性鼻-鼻窦炎病变黏膜中的表达高,均差异有统计学意义(均P<0.01)。结论:TGFβRⅠ、TGFβRⅡ在慢性鼻-鼻窦炎、鼻息肉组织中具有不同的表达水平及分布特点,提示其可能在慢性鼻-鼻窦炎、鼻息肉的发生发展过程中发挥不同的作用。     

慢性鼻窦炎鼻息肉基底干细胞转录组生物信息学分析

目的 通过生物信息学技术对慢性鼻窦炎鼻息肉基底干细胞增殖分化过程中的差异表达基因分析,为慢性鼻窦炎鼻息肉上皮屏障损伤机制及治疗提供新的思路和方向。方法 从基因表达数据库(GEO数据库)下载慢性鼻窦炎鼻息肉基底干细胞增殖分化转录组芯片数据集,使用R软件构建加权基因共表达网络分析(WGCNA)网络,筛选与正常对照组间与疾病相关的差异表达基因,通过网络在线工具DAVID进行差异基因的基因本体论(GO)分析和京都基因与基因组百科全书(KEGG)通路富集分析,通过STRING数据库构建差异表达基因的蛋白相互作用网络,并应用Cytoscape中的MCODE插件对蛋白相互作用网络进行分析。最后,使用NetworkAnalys整合转录因子数据库构建核心基因的转录因子网络。结果 研究发现基底干细胞在慢性鼻窦炎伴鼻息肉组与正常对照组间共有175个与疾病相关的差异表达基因(P<0.05,∣logFC∣>1)。GO分析和KEGG通路分析显示,这些差异基因主要富集在内肽酶活性的负调控、丝氨酸型内肽酶抑制剂活性和Wnt信号通路等。通过蛋白相互作用网络的构建及分析,筛选出IVL和SPRR2A等核心基因,...     

TLR2和TLR4在慢性鼻窦炎鼻息肉中的表达及临床意义

目的检测Toll样受体2(toll-like receptor 2,TLR2)和Toll样受体4(toll-like receptor 4,TLR4)蛋白在慢性鼻窦炎鼻息肉黏膜组织中的表达,并探讨其在鼻窦炎鼻息肉发病机制中的作用。方法采用免疫组织化学技术检测62例鼻窦炎鼻息肉黏膜组织中TLR2和TLR4蛋白的表达和分布,同期选取5例正常筛窦黏膜组织进行对照。按照1997年海口制定的"慢性鼻窦炎鼻息肉临床分型分期及鼻内镜下鼻窦手术疗效评定标准"将其分组,比较各组间TLR2和TLR4表达程度的差异。结果①TLR2、TLR4主要表达在鼻腔黏膜上皮细胞、固有层炎性细胞及黏膜下层腺体的胞浆和胞膜上。②慢性鼻窦炎鼻息肉各型各期均较对照组高;并且随着分型的升高,TLR2、TLR4的表达量也随之增加,差异具有统计学意义(P<0.05)。结论 TLR2和TLR4在鼻窦炎鼻息肉中的表达可能与鼻窦炎鼻息肉的发病机制存在着一定的相互关联,TLR在该机制中可能扮演重要角色。     

鼻息肉组织及上皮细胞的形态学特征与嗜酸粒细胞阳离子蛋白韵表达

目的：研究鼻息肉组织及其上皮细胞的形态学特征，检测嗜酸粒细胞阳离子蛋白（ECP）在其中的表达，探讨其发病机制及病理特点、ECP的作用。方法：慢性鼻-鼻窦炎不伴鼻息肉（鼻窦炎组）5例，慢性鼻-鼻窦炎伴鼻息肉（鼻息肉组）5例，正常下鼻甲黏膜（对照组）5例，标本均分成2份，一份放入4％的戊二醛液中固定，常规电镜制片；另一份放入10％甲醛中固定，石蜡包埋行ECP原位杂交检测。结果：电镜下见鼻息肉组黏膜上皮排列紊乱，基底膜增厚，纤毛排列不整齐，倒伏，其中基底层有脱颗粒现象，鼻窦炎组上皮层破坏、基底膜水肿；对照组上皮层连续，基底层排列整齐。鼻窦炎组、鼻息肉组ECP阳性表达于上皮层、黏膜下层及血管周，呈棕褐色，但鼻窦炎组表达较鼻息肉组低。结论：鼻息肉组织及上皮细胞内有活化的嗜酸粒细胞，脱颗粒现象，鼻息肉组织中ECP高表达，说明鼻窦炎和（或）鼻息肉组织中有活跃的炎症反应，引起组织上皮损伤和脱落，最终形成鼻息肉。     

鼻息肉基因芯片检测及基因表达谱的研究

目的:应用寡聚核苷酸基因表达谱芯片研究鼻息肉(NP)基因表达谱的变化.方法:应用包含14500条人类全长基因的寡聚核苷酸芯片HG-U133A2.0检测6例NP组织,6例正常鼻黏膜组织的基因表达谱,分析差异表达基因,并选择其中有差异表达的部分基因进行realtime RT-PCR验证.结果:基因芯片筛选出大量差异表达的基因,其中包括炎性反应、免疫应答、免疫调控及信号传导的相关基因及其受体,如IL-8、RGS1、GRK4、CCL20、子宫球蛋白(uteroglobin)等.实时荧光定量PCR测定差异表达基因IL-8、RGS1与基因芯片结果一致.结论;NP基因表达谱存在差异,RGS1可能通过影响细胞信号传导通路发挥作用,IL-8可能通过促使炎性细胞释放多种炎性递质参与鼻息肉发病.     

白介素-2信号通路在鼻息肉发病中的作用

目的：研究白介素2（IL-2）信号通路相关因子及调节性T细胞在鼻息肉患者组织中的表达情况,探讨二者之间的关联及其在息肉发病中的可能作用机制。方法：集取鼻息肉及正常对照组共30例患者的息肉组织及鼻甲黏膜,采用ELISA法检测局部组织内IL-2及IL-2R,免疫印迹检测组织中的磷酸化STAT5,实时荧光定量PCR法检测Foxp3mRNA在局部组织的表达情况;采用流式细胞术检测局部组织内Treg细胞的比率;同时分析IL-2通路相关因子及Treg细胞的相关性。结果：鼻息肉患者局部组织中IL-2、IL-2R及pSTAT5的含量较正常对照组显著降低,差异有统计学意义（均P〈O．05）;同时,与正常对照组相比,鼻息肉患者的Foxp3mRNA及Treg细胞的比率显著下调,差异有统计学意义（均P〈O．05）;且鼻息肉局部组织内IL-2信号通路相关因子的表达与Treg细胞比率及Foxp3mRNA水平呈正相关（均P〈O．01）。结论：IL-2信号通路的活化状态在鼻息肉患者局部组织中发生了变化,且与Treg细胞的表达呈正相关,提示IL-2信号通路在鼻息肉的发病中起重要作用,鼻息肉中Treg细胞的下调可能是由IL-2信号通路的活化下调所引起的。     

Altered expression of genes associated with innate immunity and inflammation in recalcitrant rhinosinusitis with polyps

BACKGROUND: The role of the innate immune system in the pathophysiology of chronic rhinosinusitis (CRS) is poorly understood. In this study, we compared sinonasal expression of toll-like receptors (TLRs), complement components, serum amyloid A, and inflammatory genes (chemokines and cytokines) in control subjects and patients undergoing sinus surgery for CRS. METHODS: Eleven control subjects and 30 subjects with CRS unresponsive to medical management were enrolled prospectively before undergoing endoscopic sinus surgery. Ethmoid mucosal specimens were obtained surgically and processed for RNA extraction. Real-time polymerase chain reaction was used to quantitate the level of expression of messenger RNA (mRNA) for TLR, acute phase proteins, and cytokine genes. Subjects were followed for a minimum of 6 months postoperatively with nasal endoscopy to assess for recurrence of polyps. RESULTS: mRNA for all target genes was detected in the ethmoid mucosa of both control and CRS subjects. The level of gene expression was normalized to the housekeeping genes 18s RNA and glyceraldehyde-3-phosphate dehydrogenase. As compared with controls, CRS was associated with significantly higher expression of TLR2 and the inflammatory genes macrophage-inflammatory protein alpha, RANTES, and granulocyte-macrophage colony-stimulating factor. Patients with early recurrence of polyps after surgery had significantly decreased expression of TLR2, 9, and serum amyloid A and increased expression of macrophage-inflammatory protein alpha compared with surgery-responsive patients. CONCLUSION: This study shows the increased levels of expression of TLR2 and a variety of inflammatory genes in sinonasal mucosa of CRS patients compared with controls. Whether these differences play a role in pathogenesis or are merely manifestations of disease activity is worthy of investigation.     

Expression profile of immune-associated genes in nasal polyps

OBJECTIVES: We performed this study to investigate the expression profile of immune-associated genes and to probe the role of related genes in the immune pathogenesis of nasal polyps. METHODS: Microarray analysis was used to find the expression profile of 491 immune-associated genes in nasal polyps. In validation studies, immunohistochemical staining and Western blot analysis were used to detect interleukin (IL)-17 and IL-17 receptor (IL-17R) in nasal polyps and controls. RESULTS: Eighty-seven genes were differentially expressed in the immune-associated gene profile of nasal polyps, and 15 genes showed differential expression in both chips. In nasal polyp tissues, IL-17 was expressed mainly in the cytoplasm of plasma cells and to a lesser degree in the prickle cell layer of the epithelium and the acinus of the serous gland. In turbinates, IL-17 was also expressed in the same location, but the expression of IL-17 in nasal polyps and that in turbinates differed significantly (p < .05). Both IL-17 and IL-17R displayed specific bands in nasal polyps and turbinates, but the bands of IL-17 and IL-17R in nasal polyps were stronger than those in turbinates. CONCLUSIONS: The differentially expressed genes in immune-associated gene chips will provide clues about, and a theoretical foundation for, the pathogenesis of nasal polyps. Furthermore, IL-17 may play an important role in the occurrence of nasal polyps by overexpression.     

Epithelial genes in chronic rhinosinusitis with and without nasal polyps

BACKGROUND: Genetic studies on chronic inflammatory diseases have resulted in an emphasis on the epithelial interface with the environment and the genes that influence this interaction. This study examines the expression of key epithelial genes implicated in the pathogenesis of other inflammatory disorders for their role in chronic rhinosinusitis (CRS). METHODS: Epithelial cells were collected from the inferior turbinate, middle turbinate, and/or uncinate from 62 subjects undergoing sinonasal surgery. Patient groups included 21 CRS patients with nasal polyposis, 23 CRS patients without nasal polyposis, and 18 controls. Samples were analyzed for S100A7, S100A8, S100A9, SLC9A3R1, G-protein-coupled receptor for asthma, and serine protease inhibitor kazal type 5 (SPINK5) by quantitative real-time polymerase chain reaction. Immunohistochemistry (IHC) was performed to analyze expression of SPINK5 lympho epithelial kazal-type inhibitor (LEKTI) in sinonasal samples. RESULTS: Expression of S100A7 and S100A8 was significantly decreased in CRS with and without nasal polyps when compared with controls. S100A9 expression was significantly decreased in CRS without nasal polyps, and SPINK5 expression was significantly decreased in CRS with nasal polyps. SPINK5 (LEKTI) protein was detected in sinonasal tissue and was significantly decreased in polyp samples using IHC. CONCLUSION: This study shows marked reductions in the level of expression of several genes involved in epithelial barrier maintenance and repair in the inflammatory state of CRS.     

Inflammatory pathway gene expression in chronic rhinosinusitis

BACKGROUND: The main objective in this preliminary experiment was to compare gene expression in the sinus mucosa of patients with chronic hyperplastic rhinosinusitis (CRS) against normal subjects, using gene microarray technology. The specific aim was to examine alterations in inflammatory mediator expression in patients with CRS. We performed a prospective experimental study. METHODS: Total RNA samples were obtained from the sinus mucosa biopsies of 14 patients with chronic hyperplastic sinusitis and 4 normal controls, using the Affymetrix recommended protocol. The data for 22,000 genes on the GeneChip U133A were generated from 18 hybridizations. Affymetrix GeneChip 5.0 was used as the image acquisition software for the U133A chips. Data normalization, log transformation, statistical analysis, and pattern study were performed with GeneSpring software. Comparison between patients with CRS and normal controls were performed using the Welch t-test, with log transformed data. RESULTS: There were a total of 1283 genes scored as differentially expressed between groups. The value of p, the probability of a false positive, was set to <0.05. Hierarchical clustering was applied to study coexpression patterns of the 1283 significant genes. The inflammatory pathway was overlaid with the differential expressed gene list. Four genes involved in the inflammatory pathway, interleukin (IL)-6, IL-12A, IL-13, and tumor necrosis factor (TNF) alpha (2) were consistently overexpressed in patients with hyperplastic CRS. CONCLUSION: There is overexpression of four major genes of the inflammatory pathway (IL-6, IL-12A, IL-13, and TNF-alpha (2)) in patients with CRS compared with the normal population. Defining gene expression profiles may help elucidate new key factors in the pathogenesis of CRS and perhaps aid in the development of new therapeutic modalities.     

Mucin gene expression in nasal polyps

CONCLUSIONS: A large set of mucin genes is expressed in nasal polyps. The expression pattern is complex and may reflect the wide spectrum of variables involved in polyp formation and progression. Prospective studies including subgroups of nasal polyps and involving substantial numbers of cases in each subgroup will be required to elucidate these variables and to understand how they affect mucus secretion. OBJECTIVE: At present, 15 of the 19 known mucin genes are expressed in the human airways. Nasal polyps might be expected to have a mucin expression pattern comparable to that of the airways. The aim of this study was to investigate mucin expression in nasal polyps. MATERIAL AND METHODS: Nasal polyp samples were obtained from 20 patients during functional endoscopic sinus surgery. Normal (control) sphenoid sinus mucosa was obtained from patients undergoing trans-sphenoid hypophysectomy. The expression of eight mucin genes (MUC1-4, -5AC, -5B, -6 and -7) was studied by in situ hybridization utilizing digoxigenin-labelled oligonucleotide probes. RESULTS: MUC6 and -7 were not expressed in sphenoid sinus mucosa, while all the studied mucin genes were expressed in nasal polyps. Expression patterns varied widely between individual polyps. The predominant epithelial mucin genes were MUC4, -5AC and -3, while MUC5B and -7 were mainly of glandular origin. MUC1, -2 and -6 were weakly expressed. The major alteration in gene expression in nasal polyps was found in the submucosal glands. MUC4 and -5AC represent a major component of both submucosal glands and epithelial cells in nasal polyps.     

Serum amyloid A, properdin, complement 3, and toll-like receptors are expressed locally in human sinonasal tissue

BACKGROUND: There is a growing appreciation of the role that nasal mucosa plays in innate immunity. In this study, the expression of pattern recognition receptors known as toll-like receptors (TLRs) and the effector molecules complement factor 3 (C3), properdin, and serum amyloid A (SAA) were examined in human sinonasal mucosa obtained from control subjects and patients with chronic rhinosinusitis (CRS). METHODS: Sinonasal mucosal specimens were obtained from 20 patients with CRS and 5 control subjects. Messenger RNA (mRNA) was isolated and tested using Taqman real-time polymerase chain reaction with primer and probe sets for C3, complement factor P, and SAA. Standard polymerase chain reaction was performed for the 10 known TLRs. Immunohistochemistry was performed on the microscopic sections using antibodies against C3. RESULTS: Analysis of the sinonasal sample mRNA revealed expression of all 10 TLRs in both CRS samples and in control specimens. Expression of the three effector proteins was detected also, with the levels of mRNA for C3 generally greater than SAA and properdin in CRS patients. No significant differences were found in TLR or innate immune protein expression in normal controls. Immunohistochemical analysis of sinonasal mucosal specimens established C3 staining ranging from 20 to 85% of the epithelium present. CONCLUSION: These studies indicate that sinonasal mucosa expresses genes involved in innate immunity including the TLRs and proteins involved in complement activation. We hypothesize that local production of complement and acute phase proteins by airway epithelium on stimulation of innate immune receptors may play an important role in host defense in the airway and, potentially, in the pathogenesis of CRS.     

克拉霉素对离体鼻息肉组织中环氧合酶-2及核因子κB表达的影响

目的：检测用不同浓度梯度的克拉霉素干预后,离体鼻息肉组织中环氧合酶-2（COX-2）及核因子κB（NF-κB）的表达状况,探讨其在鼻黏膜炎症机制中的作用。方法：将鼻息肉组织块分别与克拉霉素（0、10^-6、10^-5及10^-4mol／L）于体外共同培养1d,应用Western blot和荧光实时定量PCR技术检测COX-2和NF-κB亚基的表达水平,观察克拉霉素干预后COX-2和NF-κB表达的变化规律。结果：对照组（0mol／L克拉霉素组）鼻息肉组织中COX-2、NF-κBp50和NF-κcBp65的蛋白质和核酸表达水平最高;随着克拉霉素干预剂量的增加,COX-2、NF-κBp50和NF—κBp65的表达水平呈剂量依赖性下降。相关分析表明,同一组鼻息肉组织中,COX-2核酸表达量分别与NF-κBp50、NF-κBp65呈直线正相关。结论：COX-2异常表达参与了鼻-鼻窦黏膜慢性炎症反应,克拉霉素可能通过阻断NF-κB通路来下调COX-2的合成,发挥其抗炎作用。