Selection and Validation of Reference Genes for Normalization of RT-qPCR Analysis in Developing or Abiotic-Stressed Tissues of Loquat (Eriobotrya japonica)

Loquat(Eriobotrya japonica Lindl.)is a subtropical evergreen fruit tree that produces fruits with abundant nutrients and medicinal components.Confirming suitable reference genes for a set of loquat samples before qRT-PCR experiments is essential for the accurate quantification of gene expression.In this study,eight candidate reference genes were selected from our previously published RNA-seq data,and primers for each candidate reference gene were designed and evaluated.The Cq values of the candidate reference genes were calculated by RT-qPCR in 31 different loquat samples,including 12 subgroups of developing or abiotic-stressed tissues.Different combinations of stable reference genes were screened according to a comprehensive rank,which was synthesized from the results of four algorithms,including the geNorm,NormFinder,BestKeeper andΔCt methods.The screened reference genes were verified by normalizing EjLGA1 in each subgroup.The obtained suitable combinations of reference genes for accurate normalization were GAPDH,EF1αand ACT for floral development;GAPDH,UBCE and ACT for fruit setting;EF1α,GAPDH and eIF2B for fruit ripening;ACT,EF1αand UBCE for leaves under heat stress;eIF2B,UBCE and EF1αfor leaves under freezing stress;EF1α,TUA and UBCE for leaves under salt stress;ACT,EF1αand eIF2B for immature pulp under freezing stress;ACT,UBCE and eIF2B for immature seeds under freezing stress;EF1α,eIF2B and UBCE for both immature pulp and seeds under freezing stress;UBCE,TUB and TUA for red-fleshed fruits under cold-storage stress;eIF2B,RPS3 and TUB for white-fleshed fruits under coldstorage stress;and eIF2B,UBCE and RPS3 for both red-and white-fleshed fruits under cold-storage stress.This study obtained different combinations of stable reference genes for accurate normalization in twelve subgroups of developing or abiotic-stressed tissues in loquat.To our knowledge,this is the first report to obtain stable reference genes for normalizing gene expression of abiotic-stressed tissues in E.japonica.The use of the three most stable reference genes could increase the reliability of future quantification experiments.

Novel methylation gene panel in adjacent normal tissues predicts poor prognosis of colorectal cancer in Taiwan

BACKGROUND It is evident that current clinical criteria are suboptimal to accurately estimate patient prognosis.Studies have identified epigenetic aberrant changes as novel prognostic factors for colorectal cancer(CRC).AIM To estimate whether a methylation gene panel in different clinical stages can reflect a different prognosis.METHODS We enrolled 120 CRC patients from Tri-Service General Hospital in Taiwan and used the candidate gene approach to select six genes involved in carcinogenesis pathways.Patients were divided into two groups based on the methylation status of the six evaluated genes,namely,the<3 aberrancy group and≥3 aberrancy group.Various tumor stages were divided into two subgroups(local and advanced stages)on the basis of the pathological type of the following tissues:Tumor and adjacent normal tissues(matched normal).We assessed DNA methylation in tumors and adjacent normal tissues from CRC patients and analyzed the association between DNA methylation with different cancer stages and the prognostic outcome including time to progression(TTP)and overall survival.RESULTS We observed a significantly increasing trend of hazard ratio as the number of hypermethylated genes increased both in normal tissue and tumor tissue.The 5-year TTP survival curves showed a significant difference between the≥3 aberrancy group and the<3 aberrancy group.Compared with the<3 aberrancy group,a significantly shorter TTP was observed in the≥3 aberrancy group.We further analyzed the interaction between CRC prognosis and different cancer stages(local and advanced)according to the methylation status of the selected genes in both types of tissues.There was a significantly shorter 5-year TTP for tumors at advanced stages with the promoter methylation status of selected genes than for those with local stages.We found an interaction between cancer stages and the promoter methylation status of selected genes in both types of tissues.CONCLUSION Our data provide a significant association between the methylation markers in normal tissues with advanced stage and prognosis of CRC.We recommend using these novel markers to assist in clinical decision-making.

Paraben Content in Adjacent Normal-malignant Breast Tissues from Women with Breast Cancer

Objective Accumulation of estrogenic compounds and other carcinogens in normal breast tissues contributes to unpredictable breast cancer incidence during adolescence and throughout life.To assess the role of parabens in this phenomenon,the paraben content of adjacent normal-malignant breast tissues is measured in women with breast cancer living in Isfahan Province,Iran.Methods Adjacent normal-malignant breast tissue samples were obtained from 53 subjects.The parabens including methyl-paraben(Me PB),ethyl-paraben(Et PB),propyl-paraben(Pr PB),and butylparaben(Bu PB)were extracted from the sample supernatant and then subjected to gas chromatography analysis.Results Some risk factors for breast cancer were stimulated by parabens in adjacent malignant-normal breast tissues among young and middle-aged women with breast cancer.We observed a significant association for dose-response pattern of Me PB[OR=98.34(11.43–185.2),P=0.027]for both ER+and PR+women and Me PB[OR=164.3(CI:112.3–216.3),P<0.001]for HER2+women than women with negative receptors.The risk of 95-fold increase in Me PB dose and 164-fold increase in∑PBs dose were significant for women with hereditary breast cancer in first-degree relatives.Conclusions These results may promote future epidemiology studies and strategies to improve women's lifestyle and consume paraben-free products.

Determination of optical properties of normal and adenomatous human colon tissues in vitro using integrating sphere techniques

AIM: The purpose of the present study is to compare the optical properties of normal human colon mucosa/submucosa and muscle layer/chorion, and adenomatous human colon mucosa/submucosa and muscle layer/chorion in vitro at 476.5, 488, 496.5, 514.5 and 532 nm. We believe these differences in optical properties should help differential diagnosis of human colon tissues by using optical methods.METHODS: In vitro optical properties were investigated for four kinds of tissues: normal human colon mucosa/submucosa and muscle layer/chorion, and adenomatous human colon mucosa/submucosa and muscle layer/chorion. Tissue samples were taken from 13 human colons (13 adenomatous, 13 normal). From the normal human colons a total of 26 tissue samples, with a mean thickness of 0.40 mm, were used (13 from mucosa/submucosa and 13 from muscle layer/chorion), and from the adenomatous human bladders a total of 26 tissue samples, with a mean thickness of 0.40 mm, were used (13 from mucosa/submucosa and 13 from muscle layer/chorion). The measurements were performed using a double-integratingsphere setup and the optical properties were assessed from these measurements using the adding-doubling method that was considered reliable.RESULTS: The results of measurement showed that there were significant differences in the absorption coefficients and scattering coefficients between normal and adenomatous human colon mucosa/submucosa at the same wavelength,and there were also significant differences in the two optical parameters between both colon muscle layer/chorion at the same wavelength. And there were large differences in the anisotropy factors between both colon mucosa/submucosa at the same wavelength, there were also large differences in the anisotropy factors between both colon muscle layer/chorion at the same wavelength.There were large differences in the value ranges of the absorption coefficients, scattering coefficients and anisotropy factors between both colon mucosa/submucosa,and there were also large differences in these value ranges between both colon muscle layer/chorion. There are the same orders of magnitude in the absorption coefficients for four kinds of colon tissues. The scattering coefficients of these tissues exceed the absorption coefficients by at least two orders of magnitude.CONCLUSION: There were large differences in the three optical parameters between normal and adenomatous human colon mucosa/submucosa at the same laser wavelength, and there were also large differences in these parameters between both colon muscle layer/chorion at the same laser wavelength. Large differences in optical parameters indicate that there were large differences in compositions and structures between both colon mucosa/submucosa, and between both colon muscle layer/chorion.Optical parameters for four kinds of colon tissues are wavelength dependent, and these differences would be useful and helpful in clinical applications of laser and tumors photodynamic therapy (PDT).

Study on normal ophthalmic tissues among ocular volume and extraocular muscles structure measurement by HR-MRI

Objective: To measure the ocular volume and extraocular muscles of healthy people using the high-resolution magnetic reso-nance imaging (HR-MRI) technology, to provide parameters support for eye research, teaching and diagnosis. Methods: The study collected 100 cases of normal subjects (200 eyes) from November 2012 to December 2013, including 50 males and 50 females, accepted line HR-orbital MRI, and image processing software was applied to morphology and statistical analysis. The extraocular muscles, muscle belly level horizontal diameter and maximum diameter vertical diameter were mea-sured. Results: The volume of the vitreous cavity (4.51 ± 0.26) cm3 had no significant difference with the anatomic normal value (p= .148);The volumes of the ocular globe, the anterior chamber and the lens cavity were (5.05 ± 0.21) cm3, (0.24 ± 0.26) cm3, (0.23 ± 0.14) cm3, with no statistic differences compared with those of the anatomic normal values. The extraocular muscles of normal Chinese were symmetrical on the two sides. The relationships of the short diameters of extraocular muscles in the maximum planes were: inferior rectus muscle (IR) > medial rectus muscle (MR) > superior rectus (SR) > lateral rectus muscle (LR). Those of the long diameters of extraocular muscles were: LR > MR > SR > IR. Conclusions: MRI can be used for measuring ocular volume and sizes of extraocular muscles. Compared with the anatomic normal value, the sizes of extraocular muscles can also be assessed by the symmetry on the two sides and by observing the usual rule of extraocular muscles.

Gene expression arrays as a tool to unravel mechanisms of normal tissue radiation injury and prediction of response

Over the past 5 years there has been a rapid increase in the use of microarray technology in the field of cancer research, The majority of studies use microarray analysis of tumor biopsies for profiling of molecular characteristics in an attempt to produce robust classifiers for prognosis. There are now several published gene sets that have been shown to predict for aggressive forms of breast cancer, where patients are most likely to benefit from adjuvant chemotherapy and tumors most likely to develop distant metastases, or be resistant to treatment. The number of publications relating to the use of microarrays for analysis of normal tissue damage, after cancer treatment or genotoxic exposure, is much more limited. A PublVled literature search was conducted using the following keywords and combination of terms： radiation, normal tissue, microarray, gene expression profiling, prediction. With respect to normal tissue radiation injury, microarrays have been used in three ways： （1） to generate gene signatures to identify sensitive and resistant populations （prognosis）; （2） to identify sets of biomarker genes for estimating radiation exposure, either accidental or as a result of terrorist attack （diagnosis）; （3） to identify genes and pathways involved in tissue response to injury （mechanistic）. In this article we will review all （relevant） papers that covered our literature search criteria on microarray technology as it has been applied to normal tissue radiation biology and discuss how successful this has been in defining predisposition markers for radiation sensitivity or how it has helped us to unravel molecular mechanisms leading to acute and late tissue toxicity. We also discuss some of the problems and limitations in application and interpretation of such data.

A NEW METHOD TO CONSTRUCT A FULL-LENGTH cDNA LIBRARY OF HUMAN NORMAL BLADDER TISSUE

Objective Using template switch mechanism at the 5’ end of mRNA technique (SMART) to construct a full length cDNA library of human normal bladder tissue. Methods The novel procedures used the template switching activity of powerscript reverse transcriptase to synthesize and anchor first strand cDNA in one step. Following reverse transcription, 5 cycles of PCR were performed using a modified oligo(dT) primer and an anchor primer to enrich the full length cDNA population with 1.0 g human normal bladder poly(A) + RNA, then double strand cDNA was synthesized. After digestion with sfiI and size fractionation by CHROMA SPIN 400 columns, double strand cDNA was ligated into λ TripIEx 2 vector and was packaged. We determined the titer of the primary library and the percentage of recombinant clones and finally amplified the library. Results The titer of the cDNA library constructed was 2.1×10 6 pfu·mL -1 , and the amplified cDNA library was 6×10 11 pfu·mL -1 , the percentage of recombination clones was 99%. Conclusion Using SMART technique helps us to construct full length cDNA library with high efficiency and high capacity which lays solid foundation for screening target genes of bladder diseases with probes and antibodies.

Differential Expression of Genes Involved in Cell Polarity, EMT and Cell-Fate in Breast Cancer and Corresponding Normal Tissue

Objectives: Cell polarity and epithelial morphology are peculiar features of cells forming the terminal ductal lobular unit, and they are early lost during neoplastic transformation because of an epithelial-mesenchymal transition (EMT). To understand these early events we analyzed a set of 125 genes related to cell polarity, EMT and cell-fate decision in 26 breast cancer specimens and corresponding patient-matched normal tissue. Methods: The difference of gene expression was explored by t-paired test. In addition, to evidence latent variables accounting for genes correlations, a Factor Analysis was applied as exploratory technique. Results: Among the 90 differentially expressed genes, those coding for cell polarity complexes, apical-junctional components and luminal cytokeratins were overexpressed in tumor samples (suggesting a terminally differentiated phenotype) whereas those coding for stemness-associated features or related with EMT were expressed in normal tissues but not in tumor samples, suggesting the persistence of stem/progenitor cells. Factor analysis confirmed these findings and indicated that the difference between tumors and normal tissues can be synthesized in three main features representative of specific molecular/morphological alterations. Conclusions: The a priori definition of a selected panel of genes and the application of an exploratory statistical approach, greatly contribute to reduce the intrinsic biological complexity of tumor specimens and to describe the difference between tumor specimens and corresponding histologically normal tissues.

Doppler Tissue Imaging Assessment of Left Ventricular Systolic Dyssynchrony in Severe Heart Failure Patients With a Normal QRS Duration

Objectives To assess the prevalence of systolic dyssynchrony of the left ventricular （LV） walls in patients of heart failure（HF） with a normal QRS duration by Doppler tissue imaging （DTI）. Methods 20 patients of HF with a normal QRS duration and 20 healthy individuals were investigated with DTI to quantitatively analyze their pulsed-wave Doppler spectrum of basal and middle segments in six walls of left ventricle. The time between the onset of the QRS complex of the surface ECG and the onset of the systolic wave of pulsed-wave Doppler spectrum was measured （TS）. LV systolic synchronization was assessed by the maximal difference （MD） in time of TS, the standard deviation （SD） and the coefficient of variation （CV） of TS in the all 12 LV segments. Results When a TS-MD of TS〉 53.08 ms, a TS-SD of TS 〉18.08 ms and a TS-CV of TS 〉 0.91 （＋1.65 SD of normal controls） was used to define significant systolic dyssynchrony, the prevalence of systolic dyssynchrony was 55.0 %, 55.0 % and 55.0 %, respectively, in the HF patients group, significantly higher than those in the normal control and the locations of delayed contraction of these patients were different. Conclusions LV systolic dyssynchrony could be commonly demonstrated by DTI in HF patients with a normal QRS duration. This finding will support the view about the possibility that more HF patients could benefit from cardiac resynchronization therapy.

胃癌患者组织中XPNPEP2、EZRIN的表达及意义

目的通过对胃癌及癌旁正常组织氨肽酶P2(XPNPEP2)、埃兹蛋白(EZRIN)的表达情况,探讨二者在胃癌发生、发展时的表达情况及临床意义。方法选取2020年2月至2022年12月于枣庄市立医院胃肠外科就诊的患者,就诊前无任何治疗措施,经组织病理诊断胃癌组96例及正常对照组64例。应用免疫组织化学方法检测胃癌及癌旁正常组织中的XPNPEP2和EZRIN的表达水平。结合临床资料分析其表达情况与患者年龄、性别、肿瘤大小、TNM分期、浸润转移情况的关系。结果病例组XPNPEP2、EZRIN阳性表达率分别为67.71%、63.54%,均高于对照组的26.56%、29.69%,差异有统计学意义(P<0.05);在不同TNM分期、是否发生浸润转移的患者中XPNPEP2阳性表达率组间比较,差异有统计学意义(P<0.05);XPNPEP2、EZRIN阳性表达率与患者年龄、性别、肿瘤大小无关,差异无统计学意义(P>0.05)。结论胃癌组织中XPNPEP2、EZRIN蛋表达水平升高,二者高表达与胃癌TNM分期、肿瘤浸润转移显著相关,在胃癌发生、侵袭、转移过程中发挥了重要作用,可能提示预后不良。

Close relationship between mediators of inflammation and pancreatic cancer:Our experience

In this editorial,we focus specifically on the mechanisms by which pancreatic inflammation affects pancreatic cancer.Cancer of the pancreas remains one of the deadliest cancer types.The highest incidence and mortality rates of pancreatic cancer are found in developed countries.Trends of pancreatic cancer incidence and mortality vary considerably worldwide.A better understanding of the etiology and identification of the risk factors is essential for the primary prevention of this disease.Pancreatic tumors are characterized by a complex microenvironment that orchestrates metabolic alterations and supports a milieu of interactions among various cell types within this niche.In this editorial,we highlight the foundational studies that have driven our understanding of these processes.In our experimental center,we have carefully studied the mechanisms of that link pancreatic inflammation and pancreatic cancer.We focused on the role of mast cells(MCs).MCs contain pro-angiogenic factors,including tryptase,that are associated with increased angiogenesis in various tumors.In this editorial,we address the role of MCs in angiogenesis in both pancreatic ductal adenocarcinoma tissue and adjacent normal tissue.The assessment includes the density of c-Kit receptor-positive MCs,the density of tryptase-positive MCs,the area of tryptasepositive MCs,and angiogenesis in terms of microvascularization density.

蝉花优良菌株的选育

蝉花(Cordyceps chanhua)是一种药食两用虫草科真菌,是食品和保健品领域极具开发潜力的重要新原料。为探究野生蝉花不同分离方式对生物学特性和活性成分的影响,以大别山地区采集的5株野生蝉花为研究对象,采用分生孢子分离和组织分离(菌核分离)两种方法对野生蝉花进行分离,获取蝉花菌株10株,比较分析不同蝉花菌株的菌落形态、生长速度、子实体产量和活性物质含量等,使用归一化法进行品质综合评价和聚类热图分析。结果表明:分生孢子分离菌株与组织分离的菌株相比,除菌落生长速度外,子实体鲜重、子实体长度、子实体直径、生物学效率和活性成分等各项指标普遍占优,蝉花的菌落生长速度并不能作为菌株优劣的筛选依据,子实体产孢量与生物学效率和活性成分含量呈负相关;经比较分析发现4株综合品质较好的菌株,分别为IC1-1、IC3-1、IC1-2和IC2-1,子实体湿重在12.17~15.31 g之间,子实体长度在6.20~7.00 cm之间,子实体直径在2.15~2.75 mm之间,子实生物学效率在40.57%~51.03%之间,腺苷含量在1.76~1.89 mg·g^(-1)之间,多糖含量在46.1~50.1 mg·g^(-1)之间,N~6-(2-羟乙基)腺苷含量在1.21~1.33 mg·g^(-1)之间。该研究为蝉花资源保护和品种创制提供了理论依据。

自动uARC计划在左侧乳腺癌同步加量调强放疗中的可行性研究

目的 比较联影计划系统中左侧乳腺癌同步加量调强放疗自动优弧(uARC)计划和人工计划的剂量学和正常组织并发症概率(normal tissue complication probability,NTCP)差异,为临床应用提供依据。方法 回顾性收集2021年5月至2022年5月在河南省人民医院已完成同步加量调强放疗的左侧保乳术后女性乳腺癌患者21例。应用联影计划系统分别设计人工计划和自动uARC计划。评估两组计划靶区的平均剂量(mean dose,D_(mean))、D_(2%)、D_(98%)、均匀性指数(homogeneity index,HI)、适形性指数(conformity index,CI)、危及器官剂量、计划时间和机器跳数的差异,计算并比较患侧肺、健侧肺及心脏的NTCP差异。结果 自动计划的靶区HI、CI、D_(mean)及V_(50)均优于人工计划(均P<0.05),而人工计划的处方剂量为60 Gy的计划靶区(planning target volume,PTV)(PTV_(60))的D_(98%)优于自动计划(P<0.05)。自动计划患侧肺和心脏D_(mean)均优于人工计划[(9.35±0.47)Gy vs (9.61±0.37)Gy,(7.66±1.18)Gy vs (8.50±1.10)Gy,均P<0.01]。两组计划心脏V_(30)和患侧肺V_(30)比较,差异均无统计学意义(均P>0.05)。自动计划和人工计划的计划时间分别为(25.00±2.30) min和(64.57±7.17) min(P<0.01)。两组计划机器跳数比较,差异无统计学意义(P=0.207)。自动计划肺和心脏的NTCP均低于人工计划(均P<0.01)。结论 使用联影智能计划模块实现乳腺癌自动uARC计划具有一定可行性,并且能够提高靶区剂量覆盖,减少计划设计时间。

鼻咽癌调强放疗期间重度口腔黏膜炎相关参数的预测因素分析

目的探讨接受调强放疗(IMRT)的鼻咽癌患者发生严重口腔黏膜炎(SOM)的影响因素,建立预测SOM的正常组织并发症概率(NTCP)模型并进行验证。方法对2021年6月至2022年3月接受评估和治疗的95例鼻咽癌患者的临床变量和辐射剂量学参数进行回顾性分析。使用基于最小绝对值收缩和选择算子(LASSO)回归模型建立NTCP模型,确定SOM的最佳预测因素。建立单变量截止剂量NTCP模型,以确定剂量-体积限制,并对NTCP模型的辨别能力进行评价。结果95例患者中有43例(45.3%)出现SOM。利用LASSO法筛选变量,IMRT后鼻咽癌患者V 35、T分期、年龄和诱导化疗周期数是影响SOM的独立预测因素(P<0.05)。NTCP模型为S=-16.220+(V_(35)×0.059)+(T分期×0.655)+(年龄×0.083)+(诱导化疗周期数×0.537)。基于V 35拟合体积-反应曲线,拟合参数包括T分期、年龄、诱导化疗周期数,当鼻咽癌患者口腔黏膜V 35约为44.5%时,SOM风险为50.0%。NTCP的受试者工作特征曲线下面积为0.89(95%CI:0.83~0.96)。结论本研究建立并验证了预测SOM的NTCP模型,尤其是V 35,可用于预测鼻咽癌患者放疗后SOM的发病风险。

通量平滑度对食管癌放疗计划的影响

目的 比较Monaco计划系统4种通量平滑度对食管癌放疗计划的影响。方法 选取我院23例食管癌患者为研究对象,按照计划靶区50 Gy/25次的处方剂量,分别使用4种通量平滑模式(Off、Low、Medium和High)设计4组放疗计划,将处方剂量归一至95%的靶区体积后,统计各评价指标,并使用基于等效均匀剂量的参数化正常组织并发症概率(Normal Tissue Complication Probability,NTCP)模型进行评估。使用SPSS 27.0软件进行单因素方差分析,比较各组之间的差异,并使用优劣解距离法(Technique for Order Preference by Similarity to Ideal Solution,TOPSIS)进行综合评估。结果 4组计划在靶区剂量(D_(98)、D_(2)、D_(mean)、适形度指数、均匀性指数)、危及器官剂量(双肺V_(30)、V_(20)、V_(10)、V_(5)、D_(mean)、NTCP;心脏V30、Dmean、NTCP;脊髓Dmax)以及计划执行时间和计划总控制点数目方面差异均无统计学意义(P>0.05),在治疗机器跳数(Monitor Units,MU)方面差异显著(P<0.05),且随着通量平滑程度增加,MU呈降低趋势。TOPSIS综合分析结果显示LOW组综合评分最高。结论 在食管癌放疗计划设计中,4种通量平滑模式均能满足临床治疗要求,通量平滑度的提高可降低治疗MU;考虑计划整体指标的情况下,可选择Low模式达到最优的剂量结果。

Comparative Proteome Analysis of Human Lung Squamous Carcinoma Tissue

Objective： To establish the two-dimensional electrophoresis profiles with high resolution and reproducibility from human lung squamous carcinoma tissue and paired normal tumor-adjacent bronchial epithelial tissue, and to identify differential expression tumor-associated proteins by using proteome analysis. Methods： Comparative proteome analysis with 20 human lung squamous carcinoma tissues and the paired normal bronchial epithelial tissues adjacent to tumors was carried out. The total proteins of human lung squamous carcinoma tissue and paired normal tumor-adjacent bronchial epithelial tissue were separated by means of immobilized pH gradient-based two-dimensional gel electrophoresis （2-DE） and silver staining. The differential expression proteins were analyzed and then identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry （MALDI-TOF-MS）. Results： （1） Well-resolved, reproducible 2-DE patterns of human lung squamous carcinoma and adjacent normal bronchial epithelial tissues were obtained. For tumor tissue, average spots of 3 gels were 1567±46, and 1436±54 spots were matched with an average matching rate of 91.6%. For control, average spots of 3 gels were 1349±58, and 1228±35 spots were matched with an average matching rate of 91.03%. The average position deviation of matched spots was 0.924±0.128 mm in IEF direction, and 1.022±0.205 mm in SDS-PAGE direction; （2） A total of 1178±56 spots were matched between the eleetrophoretie maps of 20 human lung squamous carcinoma tissues and paired normal tumor-adjacent bronchial epithelial tissues. Seventy-six differentially expressed proteins were screened; （3） Sixty-eight differential proteins were identified by PMF, some proteins were the products of oneogenes, and others involved in the regulation of cell cycle and signal transduetion; （4） In order to validate the reliability of the identified results, the expression of 3 proteins mdm2, c-jun and EGFR, which was correlated with lung squamous carcinoma, was detected by immunohistoehemieal staining and Western blot analysis. The results revealed that mdm2, c-jun and EGFR were up-regulated in lung squamous carcinomas, whereas they were down-regulated in adjacent normal bronchial epithelial tissues, normal lung tissues and inflammatory pseudotumor, which was consistent with our proteome analysis results. Conclusion： The well-resolved, reproducible 2-DE patterns of human lung squamous carcinoma and adjacent normal bronchial epithelial tissues were established and 68 differential proteins were characterized by applying comparative proteome analysis successfully. These results will provide scientific foundation for screening the molecular biomarker used to diagnose and treat lung squamous carcinoma, as well as to improve the patient＇s prognosis and provide new clue for the research of lung squamous carcinogenic mechanism.

Sparing lung tissue with virtual block method in VMAT planning for locally advanced non-small cell lung cancer

This study aimed to exploit a new virtual block method to spare normal lung tissue in VMAT planning for patients with locally advanced non-small cell lung cancer(LA-NSCLC).The previous method was used to manually restrict the angle of the beam passing through,which ignored the location and shape of large targets that varied between different slices and did not block the beamlets precisely.Unlike the previous method,this new virtual block method was used to block the beamlets when necessary by closing the multi-leaf collimator(MLC)at prerequisite angles.The algorithm for closing the MLC depended on the thickness of the beamlets passing through the lungs and avoided only the entrance radiation beamlet.Moreover,this block can be automatically contoured.A retrospective study was performed to compare the VMAT plans with and without the virtual block method for 17 LANSCLC patients,named the block plan(B-plan)/non-block plan(N-plan).All cases were selected in this study because of the large tumor size and unmet dose constraints of the lungs.In addition to the maximum dose constraint for the virtual block,B-plans adopted identical optimization parameters to N-plans for each patient.These two types of plans were compared in terms of dosimetric indices and plan scores.The results were statistically analyzed using the Wilcoxon nonparametric signed-rank test.B-plans have advantages in the following dosimetric metrics that have statistical significance(p<0.05):(1)lower V_(5)/V_(10)/D_(mean)/normal tissue complication probability(NTCP)of total lungs;(2)reductions in V_(5)/V_(10)for the contralateral lung;(3)decrease in Dmean/V_(40)of the heart;(4)decrease in esophagus V_(40);(5)reductions in Dmean,V_(5)/V_(10) of normal tissue.B-plans(82.51±7.07)achieved higher-quality scores than N-plans(80.74±7.22).The new virtual block spared the lungs as well as other normal structures in VMAT planning for LA-NSCLC.Thus,the block method may decrease the risk of radiation-related toxicity in patients.

Glycosylated and non-glycosylated quantum dot-displayed peptides trafficked indiscriminately inside lung cancer cells but discriminately sorted in normal lung cells: An indispensable part in nanoparticle-based intracellular drug delivery

Difference in sub-cellular trafficking of glycosylated and naked peptides, between normal and lung cancer cells, was established. Normal lung tissue discriminately sorted glycosylated from non-glycosylated peptides by allowing golgi localization of the glycosylated peptides while restricting golgi entry of the naked peptides. This mechanism was surprisingly not observed in its cancer cell counterpart. Lung cancer cells tend to allow unrestricted localization of both glycosylated and naked peptides in the golgi apparatus. This newly discovered difference in sub-cellular trafficking between normal and lung cancer cells could potentially be used as an effective strategy in targeted intracellular delivery, especially targeting golgi-resident enzymes for possible treatment of diseases associated with glycans and glycoproteins, such as, congenital disease of glycosylation(CDG). This very important detail in intracellular trafficking inside normal and cancer cells is an indispensable part in nanoparticle-based intracellular drug delivery.

不同来源条件培养基对人牙髓干细胞增殖的影响

背景:利用条件培养基与人牙髓干细胞共培养,初步探索可以快速促进人牙髓干细胞增殖的方法,为今后细胞治疗、扩增高质量种子细胞提供研究基础。目的:初步探究不同来源条件培养基对人牙髓干细胞增殖的影响。方法:体外分离培养人牙髓干细胞,并用流式细胞术鉴定。然后将胎牛血清超高速低温离心产物、骨折患者尿液超高速低温离心产物、骨髓来源树突状细胞体外培养上清液、人皮肤成纤维细胞培养上清液与低糖DMEM培养液配置成条件培养基后,再分别与人牙髓干细胞共培养,使用细胞无标记培养观察装置(BioStation-T)连续拍摄各组细胞生长72 h的照片、使用实时无标记细胞分析仪(RTCA)动态监测150 h,比较各组细胞的标准化细胞指数值的差异。结果与结论:①人牙髓干细胞为典型的间充质干细胞形态,表达间充质干细胞表面标志物CD90和CD105,不表达CD34和CD45;②细胞无标记培养观察装置动态监测时,发现胎牛血清超高速低温离心组的颗粒物质被吸收时间要显著早于其他实验组和空白对照组;③在实时无标记细胞分析仪检测时,与空白对照组相比,胎牛血清超高速低温离心组、骨髓来源树突状细胞体外培养上清液组、人皮肤成纤维细胞培养上清液组的标准化细胞指数值均显著升高(P<0.001),骨折患者尿液超高速低温离心组的标准化细胞指数值显著降低(P<0.001);④结果表明,胎牛血清超高速低温离心产物、骨髓来源树突状细胞体外培养上清液、人皮肤成纤维细胞培养上清液所配置的条件培养基能促进人牙髓干细胞增殖,其中胎牛血清超高速低温离心产物所配置的条件培养基对细胞还有一定保护作用。

人表皮生长因子受体2在结直肠癌中的表达及其临床意义

目的探讨人表皮生长因子受体2(HER-2)在结直肠癌中的表达及其与临床病理特征的关系。方法选取2019年4月至2022年1月徐州市肿瘤医院病理科100例结直肠癌标本设为结直肠癌组,将其中距肿瘤5 cm以上的60例正常肠黏膜设为正常肠黏膜组,另选取同期的60例结直肠腺瘤标本设为结直肠腺瘤组。三组均采用免疫组织化学(IHC)方法和荧光原位杂交(FISH)技术检测。分析HER-2在三组中的表达情况,比较三组HER-2(2+)基因扩增结果、三组HER-2的表达、结直肠癌不同临床病理特征的HER-2表达。结果结直肠癌组中,HER-2(3+)15例,HER-2(2+)45例,HER-2(0)或(1+)40例。结直肠腺瘤组中,HER-2(3+)7例,HER-2(2+)22例,HER-2(0)或(1+)31例。正常肠黏膜组中HER-2(2+)7例,HER-2(0)或(1+)53例。三组的HER-2(2+)基因扩增结果比较,差异无统计学意义(P>0.05)。结直肠癌组、结直肠腺瘤组的HER-2阳性表达均显著高于正常黏膜组,差异有统计学意义(P<0.05);结直肠癌组与结直肠腺瘤组的HER-2阳性表达比较,差异无统计学意义(P>0.05)。不同性别、年龄、肿瘤直径、TNM分期的HER-2阳性表达比较,差异无统计学意义(P>0.05);“高+中分化”的HER-2阳性表达低于“低分化”,“Tis+T1+T2”浸润深度的HER-2阳性表达低于“T3+T4”浸润深度,有淋巴结转移的HER-2阳性表达高于无淋巴结转移,差异有统计学意义(P<0.05)。结论HER-2在结直肠癌和腺瘤中均存在高表达,与癌组织的分化程度、浸润深度及淋巴结转移有密切关系。