Claudin蛋白家族在泌尿系肿瘤中的研究进展北大核心

Claudin蛋白家族是构成细胞紧密连接(tight junctions,TJs)的重要骨架蛋白,在细胞极性、上皮屏障特性、细胞运动性和细胞间的稳定性等方面发挥着关键作用。最近研究发现,Claudin蛋白家族在人类多种肿瘤中都有表达失调,起着癌基因或者抑癌基因的作用。本文综述了近年来Claudin蛋白家族在泌尿系统肿瘤(膀胱癌、前列腺癌、肾癌)中的研究进展。     

Epithelial physical barrier defects in chronic rhinosinusitis

Introduction: Chronic rhinosinusitis (CRS) is a common upper airway disease with a prevalence of greater than 10% of the general population. Although the pathogenesis of CRS remains poorly understood, there is growing evidence indicating that epithelial physical barrier defects play an important role in CRS pathogenesis.

Areas covered: Epithelial physical barriers are maintained by various intercellular junctions, especially tight junctions (TJs). Recent studies suggest that the expression of TJ molecules and epithelial barrier function in human nasal epithelium are modulated by various internal and external factors. This review summarizes recent advances regarding the structure, function, and regulating mechanisms of the epithelial physical barrier in the context of CRS.

Expert opinion: Available data indicate that epithelial physical barrier defects in CRS can result from inhaled allergens, microbial or virus infections, cytokines, hypoxia, or zinc deficiency, among other causes. Several genes/molecules, such as SPINK5, S100A7, S100A8/9, PCDH1, NDRG1, SPRR, and p63 are involved in modulating the physical barrier function in the context of CRS. The exact mechanisms and molecular pathways that lead to these barrier defects, however, require additional study. Additional work is necessary to further explore the epithelial physical barrier function in normal and pathologic sinonasal mucosa.     

Bioinformatic identification of key candidate genes and pathways in axon regeneration after spinal cord injury in zebrafish

Zebrafish and human genomes are highly homologous;however,despite this genomic similarity,adult zebrafish can achieve neuronal proliferation,regeneration and functional restoration within 6–8 weeks after spinal cord injury,whereas humans cannot.To analyze differentially expressed zebrafish genes between axon-regenerated neurons and axon-non-regenerated neurons after spinal cord injury,and to explore the key genes and pathways of axonal regeneration after spinal cord injury,microarray GSE56842 was analyzed using the online tool,GEO2R,in the Gene Expression Omnibus database.Gene ontology and protein-protein interaction networks were used to analyze the identified differentially expressed genes.Finally,we screened for genes and pathways that may play a role in spinal cord injury repair in zebrafish and mammals.A total of 636 differentially expressed genes were obtained,including 255 up-regulated and 381 down-regulated differentially expressed genes in axon-regenerated neurons.Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment results were also obtained.A protein-protein interaction network contained 480 node genes and 1976 node connections.We also obtained the 10 hub genes with the highest correlation and the two modules with the highest score.The results showed that spectrin may promote axonal regeneration after spinal cord injury in zebrafish.Transforming growth factor beta signaling may inhibit repair after spinal cord injury in zebrafish.Focal adhesion or tight junctions may play an important role in the migration and proliferation of some cells,such as Schwann cells or neural progenitor cells,after spinal cord injury in zebrafish.Bioinformatic analysis identified key candidate genes and pathways in axonal regeneration after spinal cord injury in zebrafish,providing targets for treatment of spinal cord injury in mammals.     

养血解毒方对银屑病角质形成细胞紧密连接的调节作用

目的:观察银屑病样小鼠皮损紧密连接蛋白中水闸蛋白(claduin-1,claudin-7),闭锁蛋白(occludin)的表达,明确养血解毒方对银屑病表皮通透屏障的修复作用,为养血解毒方治疗银屑病提供科学依据。方法:将C57BL/6J小鼠随机分为空白组、模型组、甲氨喋呤组、养血解毒方组,制备甲氨喋呤溶液、养血解毒方水煎剂对应灌胃干预,同时小鼠背部剃毛后给予咪喹莫特涂抹诱导银屑病样皮损模型。每日拍照记录皮损形态并对严重程度指数(PASI)评分;水油测试笔检测皮损表皮含水量;苏木素-伊红(HE)染色观察其病理改变、测量表皮厚度;免疫荧光法检测增殖相关的核抗原(Ki67);免疫组化法检测表皮兜甲蛋白(loricrin),真皮中CD3^+T淋巴细胞浸润和紧密连接蛋白claduin-1,claudin-7,occludin的表达;蛋白免疫印迹法(Western blot)检测皮损中claudin-7,occludin的表达。模拟银屑病皮损微环境,建立白细胞介素-17(IL-17,1 mg·L^-1)刺激的角质形成细胞(Hacat)模型,制作养血组分、解毒组分、养血解毒方喷干粉进行干预。采用细胞增殖毒性检测试剂盒-8(CCK-8)法检测药物对Hacat细胞的毒性;细胞免疫荧光法检测药物对角质形成细胞claudin-1,claudin-7,occludin表达的干预作用。结果:与模型组比较,养血解毒方可显著减轻小鼠银屑病样皮损表现,降低PASI评分及皮损表皮厚度(P<0.01),增加皮损区表皮水分含量(P<0.01),减少表皮ki67,loricrin的异常表达和真皮CD3+T细胞浸润(P<0.01),并增加紧密连接蛋白claudin-1,claudin-7,occludin的表达(P<0.05),增加紧密连接结构的完整性;体外研究发现,与模型组比较,养血解毒方组和养血组分组明显升高紧密连接蛋白claudin-1,claudin-7,occludin的表达(P<0.05);与模型组比较,解毒组分组蛋白表达水平无统计学差异。结论:养血解毒方通过调节角质形成细胞间紧密连接的表达抑制其异常的增殖分化过程,进一步恢复破坏的表皮通透屏障,可能是其治疗银屑病的作用机制之一。其中养血解毒方的养血组分对调节紧密连接的修复起主要作用。     

紧密连接蛋白Claudin-3与三阴性乳腺癌患者临床特征的相关性

目的:探讨紧密连接蛋白Claudin-3与三阴性乳腺癌患者临床特征的相关性。方法:自2012年3月至2017年3月，收集我院收治的三阴性乳腺癌患者108例，行乳腺癌根治术，术中取乳腺癌标本和癌旁标本，检测组织中紧密连接蛋白Claudin-3表达情况，分析紧密连接蛋白Claudin-3与患者临床病理参数的相关性。对患者随访1年，分析术后1年复发患者与不复发患者组织中紧密连接蛋白Claudin-3的差异。结果:乳腺癌组织和癌旁组织中紧密连接蛋白Claudin-3表达无统计学差异(P=0.331)。与紧密连接蛋白Claudin-3阴性的患者相比，紧密连接蛋白Claudin-3阳性的患者TNM分期为Ⅲ期的比例显著增高(52.46% vs 29.79%,P=0.018),淋巴结转移比例增高(55.74% vs 29.79%,P=0.007),术后复发率显著增高(26.23% vs 10.64%,P=0.042)。结论:紧密连接蛋白Claudin-3与淋巴结转移和TNM分期有关，对预测患者术后复发具有一定价值。     

GDF15 promotes simultaneous astrocyte remodeling and tight junction strengthening at the blood–brain barrier

Perivascular astrocyte processes (PAP) surround cerebral endothelial cells (ECs) and modulate the strengthening of tight junctions to influence blood–brain barrier (BBB) permeability. Morphologically altered astrocytes may affect barrier properties and trigger the onset of brain pathologies. However, astrocyte-dependent mediators of these events remain poorly studied. Here, we show a pharmacologically driven elevated expression and release of growth/differentiation factor 15 (GDF15) in rat primary astrocytes and cerebral PAP. GDF15 has been shown to possess trophic properties for motor neurons, prompting us to hypothesize similar effects on astrocytes. Indeed, its increased expression and release occurred simultaneously to morphological changes of astrocytes in vitro and PAP, suggesting modulatory effects of GDF15 on these cells, but also neighboring EC. Administration of recombinant GDF15 was sufficient to promote astrocyte remodeling and enhance barrier properties between ECs in vitro, whereas its pharmacogenetic abrogation prevented these effects. We validated our findings in male high anxiety-related behavior rats, an animal model of depressive-like behavior, with shrunk PAP associated with reduced expression of the junctional protein claudin-5, which were both restored by a pharmacologically induced increase in GDF15 expression. Thus, we identified GDF15 as an astrocyte-derived trigger of astrocyte process remodeling linked to enhanced tight junction strengthening at the BBB.     

Increased activity of Rho kinase contributes to hemoglobin-induced early disruption of the blood-brain barrier in vivo after the occurrence of intracerebral hemorrhage

This study is to examine whether the activation of Rho kinase (ROCK) accounts for hemoglobin (Hb)-induced disruption of blood-brain barrier (BBB) after the occurrence of intracerebral hemorrhage. A model of intracerebral injection of Hb was established in rats. Changes in the levels of mRNA of RhoA, ROCK2 and matrix metalloproteinase-9 (MMP-9) were measured using quantitative real-time polymerase chain reaction. Protein expression of RhoA, ROCK2, claudin-5 and MMP-9, as well as ROCK activity, were determined using Western blotting. Immunohistochemical assay was performed to visualize the expression of RhoA, ROCK2, claudin-5 and MMP-9 in endothelial cells. Hb injection produced a significant increase in BBB permeability and water content in the brain. Significant reduction of claudin-5 expression was detected by Western blotting and immunofluorescence in Hb group. The levels of RhoA and ROCK2 were significantly up-regulated from 6 h to 12 h after Hb injection and were concomitant with the increase in ROCK activity. Immunofluorescence double staining showed enhanced p-myosin light chain immunoreactivity but diminished claudin-5 staining in endothelial cells. Significant up-regulation of MMP-9 expression was detected after Hb injection, and statistical analyses further confirmed a positive correlation of MMP-9 expression with ROCK activity. The results showed that ROCK was activated in endothelial cells by Hb. This may account for the early disruption of the BBB via up-regulation of p-myosin light chain expression and aggravation of injuries to TJ proteins. The activation of ROCK may also increase MMP-9 expression, thereby leading to further BBB disruption.     

The effect of retinal pigment epithelial cell patch size on growth factor expression

The spatial organization of retinal pigment epithelial (RPE) cells grown in culture was controlled using micropatterning techniques in order to examine the effect of patch size on cell health and differentiation. Understanding this effect is a critical step in the development of multiplexed high throughput fluidic assays and provides a model for replicating disease states associated with the deterioration of retinal tissue during age-related macular degeneration (AMD). Microcontact printing of fibronectin on polystyrene and glass substrates was used to promote cell attachment, forming RPE patches of controlled size and shape. These colonies mimic the effect of atrophy and loss-of-function that occurs in the retina during degenerative diseases such as AMD. After 72 h of cell growth, levels of vascular endothelial growth factor (VEGF), an important biomarker of AMD, were measured. Cells were counted and morphological indicators of cell viability and tight junction formation were assessed via fluorescence microscopy. Up to a twofold increase of VEGF expression per cell was measured as colony size decreased, suggesting that the local microenvironment of, and connections between, RPE cells influences growth factor expression leading to the initiation and progression of diseases such as AMD.     

Requirement of the F‐actin‐binding activity of l‐afadin for enhancing the formation of adherens and tight junctions

The apical junctional complex consists of adherens junctions (AJs) and tight junctions (TJs) in polarized epithelial cells, which are attached to each other to form a sheet. Actin filaments (F‐actin) are associated with AJs and TJs and required for the formation and maintenance of this complex. l‐Afadin is an F‐actin‐binding protein, which is localized at AJs through binding to the cell adhesion molecule nectin, and regulates the formation of AJs and TJs. However, the role of the F‐actin‐binding activity of l‐afadin for the formation of the apical junctional complex remains unknown. We generated here the cultured EpH4 mouse mammary epithelial cells in which afadin was genetically ablated. In the Ca²⁺ switch assay, the formation of both AJs and TJs was markedly impaired in the afadin‐deficient cells. Re‐expression of l‐afadin in the afadin‐deficient cells fully restored the formation of both AJs and TJs, but the re‐expression of the l‐afadin mutant lacking the FAB domain did not completely restore the formation of AJs or TJs. These results indicate that the F‐actin‐binding activity of l‐afadin is required for enhancing the formation of both AJs and TJs.