The clinical utility of miR-21 as a diagnostic and prognostic marker for renal cell carcinoma

Renal cell carcinoma (RCC) is the most common neoplasm of the kidney. Increasing evidence suggests that microRNAs are dysregulated in RCC and are important factors in RCC pathogenesis. miR-21 is a known oncogene with tumor-promoting effects in many types of cancer. In this study, we analyzed miR-21 in 121 cases of healthy kidney and different RCC subtypes, including clear cell (ccRCC), papillary (pRCC), chromophobe (chRCC), and oncocytoma. Total RNA was extracted, and the expression of miR-21 was measured with real-time quantitative RT-PCR using miR-21-specific probes. The expression of miR-21 was significantly up-regulated in RCC compared with healthy kidney. There was a significant difference in the expression levels between RCC subtypes, with the highest levels of expression in ccRCC and pRCC subtypes. miR-21 expression distinguished ccRCC and pRCC from chRCC and oncocytoma with 90% specificity (95% CI, 63.9% to 98.1%) and 83% sensitivity (95% CI, 53.5% to 97.6%). Significantly higher miR-21 levels were associated with higher stage and grade. Patients who were miR-21 positive had statistically significant shorter disease-free and overall survival rates. Thus, miR-21 is up-regulated in RCC, and its expression levels can be used as a diagnostic marker to distinguish ccRCC and pRCC from chRCC and oncocytoma. Moreover, it has potential as a prognostic marker in RCC, although it is not independent of tumor stage and grade.     

Nucleosomes in serum as a marker for cell death.

The concentration of nucleosomes is elevated in blood of patients with diseases which are associated with enhanced cell death. In order to detect these circulating nucleosomes, we used the Cell Death Detection-ELISAplus (CDDE) from Roche Diagnostics (Mannheim, Germany) (details at http:\\biochem.roche.com). For its application in liquid materials we performed various modifications: we introduced a standard curve with nucleosome-rich material, which enabled direct quantification and improved comparability of the values within (CVintraassay:3.0-4.11%) and between several runs (CVinterassay:8.6-13.5%), and tested the analytical specificity of the ELISA. Because of the fast elimination of nucleosomes from circulation and their limited stability, we compared plasma and serum matrix and investigated in detail the pre-analytical handling of serum samples which can considerably influence the test results. Careless venipuncture producing hemolysis, delayed centrifugation and bacterial contamination of the blood samples led to false-positive results; delayed stabilization with EDTA and insufficient storage conditions resulted in false-negative values. At temperatures of -20 degrees C, serum samples which were treated with 10 mM EDTA were stable for at least 6 months. In order to avoid possible interfering factors, we recommend a schedule for the pre-analytical handling of the samples. As the first stage, the possible clinical application was investigated in the sera of 310 persons. Patients with solid tumors (n=220; mean=361 Arbitrary Units (AU)) had considerably higher values than healthy persons (n=50; mean=30 AU; p=0.0001) and patients with inflammatory diseases (n=40; mean= 296 AU; p=0.096). Within the group of patients with tumors, those in advanced stages (UICC 4) showed significantly higher values than those in early stages (UICC 1-3) (p=0.0004).     

Identification by proteomic analysis of calreticulin as a marker for bladder cancer and evaluation of the diagnostic accuracy of its detection in urine

BACKGROUND: New methods for detection of bladder cancer are needed because cystoscopy is both invasive and expensive and urine cytology has low sensitivity. We screened proteins as tumor markers for bladder cancer by proteomic analysis of cancerous and healthy tissues and investigated the diagnostic accuracy of one such marker in urine. METHODS: Three specimens of bladder cancer and healthy urothelium, respectively, were used for proteome differential display using narrow-pH-range two-dimensional electrophoresis. To evaluate the presence of calreticulin (CRT) as detected by Western blotting, we obtained 22 cancerous and 10 noncancerous surgical specimens from transurethral resection or radical cystectomy. To evaluate urinary CRT, we collected 70 and 181 urine samples from patients with and without bladder cancer, respectively. Anti-CRT COOH-terminus antibody was used to detect CRT in tissue and urine. RESULTS: Proteomic analysis revealed increased CRT (55 kDa; pI 4.3) in cancer tissue. Quantitative Western blot analysis showed that CRT was increased in cancer tissue (P = 0.0003). Urinary CRT had a sensitivity of 73% (95% confidence interval, 62-83%) at a specificity of 86% (80-91%) for bladder cancer in the samples tested. CONCLUSIONS: Proteomic analysis is useful in searching for candidate proteins as biomarkers and led to the identification of urinary CRT. The diagnostic accuracy of urinary CRT for bladder cancer appears comparable to that of Food and Drug Administration-cleared urinary markers, but further studies are needed to determine its diagnostic role.     

Texture analysis as a radiomic marker for differentiating renal tumors

Purpose

To evaluate the utility of texture analysis for the differentiation of renal tumors, including the various renal cell carcinoma subtypes and oncocytoma.

Materials and methods

Following IRB approval, a retrospective analysis was performed, including all patients with pathology-proven renal tumors and an abdominal computed tomography (CT) examination. CT images of the tumors were manually segmented, and texture analysis of the segmented tumors was performed. A support vector machine (SVM) method was also applied to classify tumor types. Texture analysis results were compared to the various tumors and areas under the curve (AUC) were calculated. Similar calculations were performed with the SVM data.

Results

One hundred nineteen patients were included. Excellent discriminators of tumors were identified among the histogram-based features noting features skewness and kurtosis, which demonstrated AUCs of 0.91 and 0.93 (p < 0.0001), respectively, for differentiating clear cell subtype from oncocytoma. Histogram feature median demonstrated an AUC of 0.99 (p < 0.0001) for differentiating papillary subtype from oncocytoma and an AUC of 0.92 for differentiating oncocytoma from other tumors. Machine learning further improved the results achieving very good to excellent discrimination of tumor subtypes. The ability of machine learning to distinguish clear cell subtype from other tumors and papillary subtype from other tumors was excellent with AUCs of 0.91 and 0.92, respectively.

Conclusion

Texture analysis is a promising non-invasive tool for distinguishing renal tumors on CT images. These results were further improved upon application of machine learning, and support the further development of texture analysis as a quantitative biomarker for distinguishing various renal tumors.

     

Amyloid P component as a plasma marker for Parkinson's disease identified by a proteomic approach

ObjectivesParkinson's disease (PD) ranks the second among the neurodegenerative disorders. Proteins involved in Parkinson's disease (PD) have been investigated but none as the diagnostic markers in blood.Design and methodsIn this study, we applied a proteomic strategy, by utilizing two-dimensional electrophoresis and mass spectrometry, to analyze two sample pools of plasma from the healthy individuals and PD subjects.ResultsIgGκL and human serum amyloid P component (SAP) were found differentially expressed between these pools. SAP level increased by approximately 5-fold in PD samples, and the ELISA procedure revealed a significant (P < 0.001) increase in SAP concentration (65.9 ± 18.7 μg/mL) in the plasma of PD subjects (healthy individuals, 35.0 ± 12.5 μg/mL), with sensitivity of 94.1% and specificity of 87.5%.ConclusionOur results indicated a potential feasibility of plasma SAP as a marker to approach PD.     

The potential use of urine cell free DNA as a marker for cancer

Introduction: Although the role of circulating cell free DNA in cancer has been widely demonstrated, less is known about the role of urine cell free DNA (UcfDNA). UcfDNA can serve as a ‘liquid biopsy’ for urological and non-urological tumors, as it carries information on DNA from cells exfoliated in urine and from circulation.

Areas covered: We review the studies on UcfDNA as a source of biomarkers for cancer, focusing on the new techniques and the differences between urological and non-urological tumors. We searched Pubmed for articles published between 1998 and 2016 with the following key words and phrases: ‘urine’ and ‘cell free DNA’ or ‘liquid biopsy’ or ‘cancer’.

Expert commentary: Despite the few papers published on this topic, UcfDNA is an important component of ‘liquid biopsy’, a useful and non-invasive tool for cancer diagnosis, prognosis and treatment monitoring, containing a wide range of genetic information.     

7例转移性透明细胞性肾细胞癌的诊断及鉴别诊断

目的 描述肺、肾上腺、甲状腺转移性透明细胞性肾细胞癌（RCCs）的病理特点，探讨其诊断及鉴别诊断。方法 应用HE和免疫组化方法，对7例转移性透明细胞性RCCs及其中5例的肾原发性癌染色并观察。结果 转移性透明细胞性RCCs组织学具有实性巢状、有／无腔的腺泡状及腺管状结构，间质少且富于血管。免疫组化示全部转移性及原发性透明细胞性RCCs表达CD10、CK和vimentin。结论 在肿瘤发生部位特异性抗体阴性表达的情况下，结合肿瘤组织学特征，应考虑转移性透明细胞性RCCs。CD10是鉴别转移性透明细胞性RCCs的首选抗体。     

Troponin-T as a serum marker for myocardial infarction

We report here our experience with serum troponin T(TnT), measured with the sandwich immunoassay introduced by Boehringer Mannheim as a marker for myocardial infarction. We assayed TnT in serial serum samples from 30 patients with time courses of serum CK, CK-MB, AST, and LD that we consider typical of acute myocardial infarction (MI). In every patient but one, TnT rose in parallel with both CK-MB and AST, but remained elevated significantly longer. The ratios of the elevations of the different markers varied from patient to patient with marked variation in the ratio of TnT to CK-MB. There appeared to be a significant association between the magnitude of that ratio with the level of ALT. J. Clin. Lab. Anal. 11:125–128, 1997. © 1997 Wiley-Liss, Inc.     

肾细胞癌的MRI影像分析

目的分析肾细胞癌亚型MRI信号平扫特点及增强表现,提高对肾细胞癌的诊断水平。方法收集有完整临床资料及病理证实肾细胞癌48例,均做T_1WI、T_2WI、TRUFI序列平扫及增强扫描,其中15例行MRI动态增强扫描。结果本组48命名透明细胞型41例,嫌色细胞型4例,乳头状细胞型3例。T_1WI均匀等或低信号33例,混杂信号15例;T2WI均匀高信号14例,等信号6例,混杂信号28例。显示假包膜者10例。增强扫描6例病灶均匀强化,34例不均匀强化,5例内壁不规则环状强化,3例均匀环状强化(假包膜强化)。结论 MRI能够准确诊断肾细胞癌,并有助于判断细胞亚型。     

血清胱蛋白酶抑制剂C评估原发性高血压病肾功能检测的临床意义

目的:评价血清胱蛋白酶抑制剂C(CysC)在原发性高血压肾功能损害诊断中的临床价值。方法:80例不同程度肾功能患者及30例健康体检者分别测定尿微量蛋白定量、血尿酸、血清CysC、血肌酐值并进行比较。结果:血清CysC在GFR40～80mL/(min·1.73m2)开始明显升高,尿微量蛋白定量、血尿酸的测定在GFR正常时已经开始升高。结论:血清CysC对评价原发性高血压病肾功能损害中较血肌酐更敏感,但不比尿微量蛋白定量、血尿酸对早期原发性高血压肾损害灵敏。     

Metastatic renal cell carcinoma presenting as a duodenal mass

We report a case of renal cell carcinoma metastasis to the duodenum.     

Metastatic renal cell carcinoma presenting as a breast mass

In patients with breast cancer, the disease usually metastasizes from one side to the other. However, metastasis to a breast from other sites is possible. Dr Lesho describes a case of breast cancer in which the primary disease occurred in the kidney.     

多房性囊性肾细胞癌临床病理分析

目的探讨多房性囊性肾细胞癌（MCRCC）的临床病理特点及手术方式与预后的关系。方法对2例MCRCC进行病理形态及免疫组化观察,结合文献分析其临床特点及手术方式与预后的关系。结果2例MCRCC肿瘤直径均〈4cm。镜下见肿瘤全部由大小不等的囊腔构成,囊腔内衬透明细胞,间隔内透明细胞呈巢状分布,Fuhrman分级为Ⅰ～Ⅱ级,囊腔间隔内可见似平滑肌的梭形细胞。免疫组化标记：透明细胞CK（广谱）（＋）,CD68和Ki-67（-）;间隔内梭形细胞SMA（＋）。1例行根治性肾切除术,1例行肿瘤剜除术,随访2～26个月,均无复发及转移。结论MCRCC是具有低度恶性潜能的肿瘤,有其独特的病理形态特征,肿瘤预后良好,可通过手术切除而治愈。〈4cm的肿瘤,可以施行保守性手术治疗。     

多房性囊性肾细胞癌临床病理分析

目的 探讨多房性囊性肾细胞癌（MCRCC）临床病理特点及鉴别诊断要点,并对国内外相关文献进行复习,以提高对MCRCC的认识和病理诊断水平.方法 分析1例MCRCC患者的临床表现、组织形态学特征及免疫组化表型,并检索国内外相关文献报道共522例,总结MCRCC的特点.结果 患者男,50岁,因体检时发现左肾占位入院,MRI、彩超及CT检查考虑为错构瘤.病理检查：大体示肿瘤位于左肾上极,有完整包膜,大小4.5 cm×4.2 cm ×4.0 cm,剖面见肿瘤由大小不等的多个囊腔构成,囊腔间隔厚薄不均,囊壁尚光滑.镜检示囊腔内衬单层透明细胞,囊腔间隔胶原纤维增生,可见灶性钙化,间隔内可见巢片状分布的透明细胞,但未形成肉眼可见的结节.细胞异型性小,细胞核Fuhrman分级：Ⅰ级.免疫组化标记显示囊腔上皮及间隔内肿瘤细胞CK和EMA阳性,CD10、RCC、Ki67和CD68阴性.行后腹腔镜下左肾根治性切除术,随访8个月,一般情况好,未见复发和转移.结论 MCRCC作为肾细胞癌的一种少见的独立亚型,完全由囊腔构成,影像学和穿刺细胞学检查等难以与囊性相关性肾病区别,因此准确的病理诊断非常必要,免疫组化标记对其诊断及鉴别诊断具有肯定价值.该肿瘤具有低度恶性潜能,预后良好,保留肾单位手术值得考虑.     

Identification and confirmation of haptoglobin as a potential serum biomarker in hypertrophic cardiomyopathy using proteomic approaches

Abstract

Introduction. Aiming at identifying biomarkers for hypertrophic cardiomyopathy (HCM), the serum proteome was explored through a two-dimensional gel-based proteomic approach (2D-DIGE) coupled with mass spectrometry and database interrogation.

Methods. Serum samples from 20 male HCM patients and their sex- and age-matched controls were cleaned from interfering components. Patients and controls were pooled in five matched groups with the same age, and proteins extracts from each pool were labelled with cyanine dyes. Then, gel images were analysed using a fluorescence scanner and proteins were identified. Tryptic peptides were analysed by capillary reversed-phase liquid chromatography coupled online with tandem mass spectrometry (MS/MS).

Results. Four different proteins were observed to be differentially expressed between HCM patients and their matched controls. Of them, decreases in haptoglobin levels were confirmed to be associated with HCM in an independent set of 181 consecutive HCM patients from our monographic clinic and 114 controls with similar age and sex using a nephelometer-based technique. Moreover, a significant negative correlation was observed between haptoglobin and subaortic gradient, thus highlighting the role of haptoglobin in HCM.

Conclusion. All these observations point out the utility of the 2D-DIGE proteomic strategy for the identification of serum proteins indicative of the presence of cardiac injury.     

Comprehensive proteomic profiling identifies serum proteomic signatures for detection of hepatocellular carcinoma and its subtypes

BACKGROUND: Detection of hepatocellular carcinoma (HCC) in patients with chronic liver disease (CLD) is difficult. We investigated the use of comprehensive proteomic profiling of sera to differentiate HCC from CLD. METHODS: Proteomes in sera from 20 CLD patients with alpha-fetoprotein (AFP) <500 microg/L (control group) and 38 HCC patients (disease group) were profiled by anion-exchange fractionation (first dimension), two types (IMAC3 copper and WCX2) of ProteinChip Arrays (second dimension), and time-of-flight mass spectrometry (third dimension). Bioinformatic tests were used to identify tumor-specific proteomic features and to estimate the values of the tumor-specific proteomic features in the diagnosis of HCC. Cross-validation was performed, and we also validated the models with pooled sera from the control and disease groups, serum from a CLD patient with AFP >500 microg/L, and postoperative sera from two HCC patients. RESULTS: Among 2384 common serum proteomic features, 250 were significantly different between the HCC and CLD cases. Two-way hierarchical clustering differentiated HCC and CLD cases. Most HCC cases with advanced disease were clustered together and formed two subgroups that contained significantly more cases with lymph node invasion or distant metastasis. For differentiation of HCC and CLD by an artificial network (ANN), the area under the ROC curve was 0.91 (95% confidence interval, 0.82-1.01; P <0.0005) for all cases and 0.954 (95% confidence interval, 0.881-1.027; P <0.0005) for cases with nondiagnostic serum AFP (<500 microg/L). At a specificity of 90%, the sensitivity was 92%. Both cluster analysis and ANN correctly classified the pooled serum samples, the CLD serum sample with increased AFP, and the HCC patient in complete remission. CONCLUSION: Tumor-specific proteomic signatures may be useful for detection and classification of hepatocellular cancers.     

Human papillomavirus E6/E7 mRNA testing as a predictive marker for cervical carcinoma

Human papillomavirus (HPV) is necessary for the development of cervical carcinoma, and incorporation of molecular testing for HPV in screening and patient management has been proposed. Sufficient scientific evidence exists to recommend HPV DNA testing in the triage of women with equivocal cytology and in follow-up after the treatment of precursor lesions. However, due to a low clinical specificity and positive predictive value, HPV DNA testing has so far not been recommended as primary screening in Europe. In general, diagnostic HPV tests have to demonstrate accuracy, reproducibility and clinical utility before they can be used in patient management and implemented in cervical cancer screening programmes. In this article we give an overview of RNA-based HPV diagnostics and the role of E6/E7 mRNA detection as a predictive marker for the development of cervical carcinoma. HPV E6/E7 mRNA testing for high-risk types seems to correlate better with the severity of the lesion compared with HPV DNA testing, and is a potential marker for the identification of women at risk of developing cervical carcinoma. Commercial assays for simultaneous genotyping and detection of E6/E7 mRNA from the five most common high-risk HPV types are now available and require further evaluation for primary screening, triage and follow-up after treatment.     

单细胞反转录酶聚合链式反应技术对胰岛干细胞标记物胰岛素促进因子的鉴别

目的：利用干细胞分化获得胰岛细胞后目前还没有较理想的鉴别方法，实验通过单细胞反转录酶聚合链式反应技术鉴别胰岛干细胞标记物胰岛素促进因子，以建立一种检测胰岛干细胞的有效方法。方法：实验于2001-10／2005-08在加拿大多伦多大学和南京大学附属鼓楼医院完成。①将收集培养2～4周的单个胰岛干细胞用于实验。②胰岛活性测定实验中，选择1&;#215;10^6分化的胰腺干细胞，用低浓度和高浓度葡萄糖分别刺激1～4h，检测12，14，24d培养液中胰岛素的浓度。③单细胞反转录酶聚合链式反应检测胰岛素促进因子基因实验中，选择12个单个胰腺干细胞进行检测，另取12个单个未分化的干细胞作为对照。聚合链式反应体系为20μL。聚合链式反应扩增反应条件为94℃预变性4min，94℃变性30s，60℃退火30s，72℃延伸2min条件下循环35次。聚合链式反应产物以质量浓度为20g/L的琼脂糖凝胶电泳。结果：①分化的胰腺干细胞培养液中胰岛素浓度的检测：培养12，14，24d时培养液中胰岛素的浓度分别为42．6，68．2，73．5IU／L，随着时间的延长，呈逐渐升高的趋势。②单个胰岛干细胞胰岛素促进因子反转录酶聚合链式反应检测结果：12个单个胰岛干细胞中有7个胰岛素促进因子表达呈阳性，而作为对照的12个未分化干细胞均未检出，两者差异显著（P〈0．001）。结论：采用单细胞反转录酶聚合链式反应检测方法为胰腺干细胞的鉴定和纯化提供了技术保障，证明单细胞反转录酶聚合链式反应是一种检测胰腺干细胞标记物的有效方法。     

单细胞反转录酶聚合链式反应技术对胰岛干细胞标记物胰岛素促进因子的鉴别

目的:利用干细胞分化获得胰岛细胞后目前还没有较理想的鉴别方法,实验通过单细胞反转录酶聚合链式反应技术鉴别胰岛干细胞标记物胰岛素促进因子,以建立一种检测胰岛干细胞的有效方法。方法:实验于2001-10/2005-08在加拿大多伦多大学和南京大学附属鼓楼医院完成。①将收集培养2～4周的单个胰岛干细胞用于实验。②胰岛活性测定实验中,选择1×106分化的胰腺干细胞,用低浓度和高浓度葡萄糖分别刺激1～4h,检测12,14,24d培养液中胰岛素的浓度。③单细胞反转录酶聚合链式反应检测胰岛素促进因子基因实验中,选择12个单个胰腺干细胞进行检测,另取12个单个未分化的干细胞作为对照。聚合链式反应体系为20μL。聚合链式反应扩增反应条件为94℃预变性4min,94℃变性30s,60℃退火30s,72℃延伸2min条件下循环35次。聚合链式反应产物以质量浓度为20g/L的琼脂糖凝胶电泳。结果:①分化的胰腺干细胞培养液中胰岛素浓度的检测:培养12,14,24d时培养液中胰岛素的浓度分别为42.6,68.2,73.5IU/L,随着时间的延长,呈逐渐升高的趋势。②单个胰岛干细胞胰岛素促进因子反转录酶聚合链式反应检测结果:12个单个胰岛干细胞中有7个胰岛素促进因子表达呈阳性,而作为对照的12个未分化干细胞均未检出,两者差异显著(P<0.001)。结论:采用单细胞反转录酶聚合链式反应检测方法为胰腺干细胞的鉴定和纯化提供了技术保障,证明单细胞反转录酶聚合链式反应是一种检测胰腺干细胞标记物的有效方法。