Identification of squamous cell carcinoma associated proteins by proteomics and loss of beta tropomyosin expression in esophageal cancer

AIM： To assess the proteome of normal versus tumor tissue in squamous cell carcinoma of the esophagus （SCCE） in Iranian patients and compare our results with former reports by using proteomics. METHODS： Protein was extracted from normal and tumor tissues. Two dimensional electrophoresis was carried out and spots with differential expression were identified with mass spectrometry. RNA extraction and RT-PCR along with immunodetection were performed. RESULTS： Fourteen proteins were found whose expression levels differed in tumor compared to normal tissues. Mass spectrometric analysis resulted in the identification of β-tropomyosin （TMβ）, myosin light chain 2 （and its isoform）, myosin regulatory light chain 2, peroxyredoxin 2, annexin I and an unknown polypeptide as the down regulated polypeptides in tumor tissue. Heat shock protein 70 （HSP70）, TPM4-ALK fusion oncoprotein 2, myosin light polypeptide 6, keratin I, GH16431p and calreticulin were the up-regulated polypeptides found in tumor tissue. Several of these proteins, such as TMβ, HSP70, annexin Ⅰ, calreticulin, TPM4-ALK and isoforms of myosins, have been well recognized in tumorigenesis of esophageal or other types of cancers. CONCLUSION： Our study not only supports the involvement of some of the formerly reported proteins in SCCE but also introduces additional proteins found to be lost in SCCE, including TMβ.

Secondary Structure and Neurotrophic Effect of a 33.1 kDa Specific Protein (SSP-33.1) in Spinal Sensory Ganglia

Aim To analyze the secondary structure and neurotrophic effect of a specific protein in sensory neurons. Methods Comparison of the proteins expressed in the rat spinal sensory neurons and motor neurons was made by two dimensional electrophoresis. One specific protein in sensory neurons was isolated and purified by DEAE Sephacel ion exchange chromatography and high performance liquid chromatography. A primary analysis of its secondary structure by circular dichroism, and its neurotrophic effects were investigated using the model of dorsal root ganglia(DRG) cultured in vitro . Results The molecular weight and isoelectric point of the protein were 33 1 kDa and 5 52, respectively. Its circular dichroism showed that there were 20 8% α helix, 54 8% β sheet, 7 3% turn, and 17 1% random coil in its secondary structure. Biological experiments showed that the protein could promote the neurite outgrowth of DRG. Conclusion A specific protein in spinal sensory tissue with molecular weight of 33 1 kDa has been purified. There is mainly β sheet in the secondary structure of the protein. And the protein has neurotrophic effects in the model of DRG.

Comparative Study on the Infectivity and Spore Surface Protein of Nosema bombycis and Its Morphological Variant Strain

A new morphological variant strain of microsporidium was produced by infecting the mulberry looper, Hemerophilaatrilineata [Phthonandria atrilineata], with Nosema bombycis successively for 24 times, and named 24Nbh. Comparativestudies on morphology, infectivity and spore surface protein were conducted. 24Nbh was short and wide, and had asignificant difference (P<0.01) over the Nb spores. The infectivity tests conducted on second instar silkworm larvaeshowed that IC50 of 24Nbh was 1.98104 spores mL-1 and of Nb was 1.72103 spores mL-1, thus indicating that the infectivityof Nb decreased 11.5 times after multiplying in mulberry looper for 24 times. The IC50 of spores from silkworm infected with24 Nbh was 6.9 times less than Nb, showing that the infectivity of 24Nbh spores rejuvenated very fast when reinfected tosilkworms, further more, the length and width of such spore was larger than 24Nbh (P<0.01) and smaller than Nb (P<0.05).The SDS-PAGE profiles of Nb and 24Nbh were generally the same, 4 distinct proteins of 12, 17, 30, 33 kDa were obtainedwith difference in quantity. When 120 g of protein was applied for 2D-PAGE, five suspected different proteins withdifference in quantity were observed. These results demonstrate that these differential proteins maybe associated withvariation in infectivity of the spores.

胰腺癌血浆蛋白质组学研究

目的利用双向电泳(2-DE)的方法进行胰腺癌的比较蛋白质组学研究,寻找和鉴定与胰腺癌相关或特异的生物标志分子。方法选择12例胰腺癌患者以及与之性别年龄匹配的12例正常人和10例慢性胰腺炎患者的血浆标本进行研究。对去除ALB和IgG等预分离方法进行探索,确定最佳的血浆2-DE方案。通过比较胰腺癌(术前、术后)、正常人和慢性胰腺炎患者的血浆2-DE图谱,寻找差异蛋白并进行质谱鉴定。结果去除ALB和IgG以及超滤的预处理仅使血浆2-DE图谱的局部分辨率提高。共发现并鉴定了5种与胰腺癌或慢性胰腺炎相关的差异蛋白:Haptoglobin(Hp),Hemoglobin(Hb),α1-antitrypsin(α1AT),Immunoglobin J chain(Ig J) and Leucine-rich α2-glycoprotein(LRG)。Hp β链在胰腺癌患者中轻度升高,胰腺癌患者的Hp2-2表型的比例高于正常人,而缺少Hp1-1表型。胰腺癌和慢性胰腺炎患者的血浆Hb高于正常人。α1AT的变化趋势为慢性胰腺炎>胰腺癌>正常人。慢性胰腺炎患者的IgJ链高于胰腺癌患者和正常人,可能来源于血分泌型IgA(sIgA)。胰腺癌患者的LRG水平较正常人和慢性胰腺炎轻度升高。术后Hp、α1AT和LRG均明显升高,未发现术后明显下降的蛋白。结论去除ALB和IgG以及超滤去除大分子的预处理方法对于血浆蛋白的差异比较并无太大优势,但在质谱鉴定方面可以与未处理的方法互相补充。在发现的5种差异蛋白中,Hp的表型、Hb、IgJ链和LRG与胰腺癌或慢性胰腺炎的关系在以往的文献中未见报道,这些差异蛋白对于胰腺癌或慢性胰腺炎的意义及诊断价值还需要进一步的验证。

它莫西芬诱导乳腺癌MCF-7细胞自噬的比较蛋白质组学研究

The differentially expressed proteins between normal and tamoxifen-induced autophagic MCF-7 cells were studied by 2-DE combining with mass spectrometric analysis.Totally,there were 668±99 and 550±97 protein spots detected in the 2-DE maps from normal and autophagic MCF-7 cells respectively,among which 5 changed protein spots were identified by the Q-TOF MS/MS and database search.

Effect of arsenite on the proteome of earthworms Eisenia fetida

Arsenic(As)is broadly distributed due to natural and anthropogenic sources,and it is toxic to organisms.This study aimed to investigate the proteomic response in earthworm exposed to As^(3+).Earthworms were exposed to As^(3+)in soil at 5-80 mg kg1,and samples were collected after 60 days exposure.Two-dimensional electrophoresis(2-DE)was used to separate the proteins in earthworm homogenate,then differentially expressed proteins(DEPs)were identified using MALDI-TOF/TOF-MS analysis.After 2-DE,36 DEPs were found and 24 of them were successfully identified.79.2%of DEPs were upregulated compared to the control group.The maximum fold change reached 53.8 in spot 3108 in the 80 mg kg^(-1)As group.Two proteins were not found in the control group but found in the As treated groups.Proteins were grouped into metabolism,signal transduction,stress-related,transport,regulation,and predicted/hypothetical protein categories based on their function.The protein-protein interaction between the DEPs indicated that serum albumin(ALB)is very important,related to 6 other proteins.Proteins were then verified by western blot,the results were in agreement with the proteomic analyses.The identification of induced or repressed proteins because of As^(3+)in earthworms is helpful to explore the underlying mechanisms of soil arsenic ecotoxicity.

Difference of gene expression between the central and the peripheral epithelia of the bovine lens

Background Equatorial lens epithelial cells proliferate and differentiate into fiber cells throughout life, while central lens epithelial cells proliferate little and do not form fiber ceils. This study aimed to investigate the differences in gene expression between the central and the peripheral epithelial cells of the bovine lens. Methods Lens epithelia were dissected into central （≤11.5 mm diameter, cLEC） and peripheral regions （pLEC）. The differences in gene expression and protein accumulation between these two regions were assayed by microarray analysis and two-dimensional electrophoresis （2-DE） combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS）. Differently expressed proteins were validated by immunoanalyses. Results By microarray analysis, 67 transcripts were at least two-fold lower and 269 at least two-fold higher in pLEC compared with that in cLEC. Thirty-four protein spots, including 20 in cLEC and 14 in pLEC, were identified by two dimensional electrophoresis and mass spectrometry. Of these 34 protein products, 28 were represented by probe sets on the microarray. Nine transcripts changed in the same direction and four transcripts in the opposite direction to their protein products. Immunoanalyses revealed that three （mitogen-activated protein kinase 1 （MAPK1）, nidogen （NID）, small nuclear ribonucleoprotein N （SNRPN）） out of four transcripts with opposite change between 2-DE and microarray assay showed the same changes as the results of 2-DE gel analyses. The genes differently expressed between cLEC and pLEC mainly include those related to the MAPK, transforming growth factor β （TGFβ） signaling and glycolysis pathways. Conclusion The results suggested that there were distinctly different genome activities, including a specific group of pathways, between central and peripheral lens epithelial cells.

Application of proteomics in environmental science

Proteomics involves the separation of proteins,identification of the amino acid sequence of the interested or target proteins,study of the function of the proteins,modification,structure and ultimate assignments to functional pathways in the cell.The proteomic investigations have contributed greatly to human diseases studies,new drugs discovery researches,and environmental science in recent years.This article provides a review on the development of the main proteomic technologies,including both the gel based and non-gel based technologies,and their applications in environmental science.Proteomic technologies have been utilized in the environmental stresses studies to analyze the induction or reduction of proteins at expression level and identify the target proteins to investigate their function in response to environmental stresses,such as high or low pH,oxidation stress,and toxic chemicals.Such protein responses are also helpful to understand the mechanisms of some cellular activities and the functions of some proteins.