Diagnostic significance of serum HMGB1 in colorectal carcinomas

High mobility group box 1 protein (HMGB1), a nuclear protein, can be translocated to the cytoplasm and secreted in colon cancer cells. However, the diagnostic significance of HMGB1 has not been evaluated in colorectal carcinomas. For this purpose, we have screened the expression and secretion of HMGB1 in 10 colon cancer cell lines and 1 control cell line and found that HMGB1 was detected in the culture medium. To evaluate the diagnostic value of HMGB1, we performed an enzyme-linked immunosorbent assay to measure HMGB1 levels and compared them to carcinoembryonic antigen (CEA) levels in the serum samples of 219 colorectal carcinoma patients and 75 healthy control subjects. We found that the serum HMGB1 level was increased by 1.5-fold in patients with colorectal carcinoma compared to those in healthy controls. When HMGB1 and CEA levels were compared, HMGB1 had similar efficacy as CEA regarding cancer detection (the sensitivity was 20.1% for HMGB1 vs. 25.6% for CEA, and the specificity was 96% for HMGB1 vs. 90.7% for CEA). Moreover, the diagnostic accuracy of HMGB1 for stage I cancer was significantly higher than that of CEA (sensitivity: 41.2% vs. 5.9%; specificity: 96% vs. 90.7). When we combined HMGB1 and CEA, the overall diagnostic sensitivity was higher than that of CEA alone (42% vs. 25.6%), and the diagnostic sensitivity for stage I was also elevated (47% vs. 5.9%). However, the prognosis of patients was not related with serum HMGB1 concentrations. Our findings indicate that serum HMGB1 levels are increased in a subset of colorectal carcinomas, suggesting their potential utility as a supportive diagnostic marker for colorectal carcinomas.     

分化抑制因子-1在大肠癌中的表达及临床意义

目的:研究分化抑制因子-1(Id-1)在人大肠癌组织中的表达及其与临床病理特征的关系。方法:应用免疫组织化学方法(SP法)检测Id-1在56例大肠癌组织及56例远癌肠黏膜中的表达水平,并分析其与临床病理特征的关系。结果:在大肠癌组织中Id-1的过表达率为80.4%,在远癌肠粘膜中过表达率为12.5%,两组比较差异有统计学意义(P0.01)。Id-1过表达与大肠癌Dukes分期及淋巴结转移有关(P0.05),而与患者年龄、性别、肿瘤组织分化程度无关(P0.05)。结论:Id-1过表达可能参与大肠癌演进,Id-1可作为判断大肠癌恶性程度和预后的指标。     

结直肠癌组织RPN11与Ki67的表达及其临床病理学意义

目的观察去泛素化酶RPN11和增殖相关核标记物Ki67在结直肠癌组织中的表达,研究其与结直肠癌肿瘤细胞增殖的相关性及与结直肠癌临床病理特征的关系。方法采用免疫组织化学SABC法检测56例结直癌组织及20例癌旁正常组织中的RPN11和Ki67表达,结合临床病理学资料进行统计分析。结果免疫组织化学染色显示:RPN11及Ki67在结直肠癌组织的阳性表达率明显高于正常结直肠组织;RPN11和Ki67的表达均与肿瘤分化程度、TNM分期、转移有关,而与性别、年龄无明显相关;RPN11与Ki67的表达呈正相关。结论RPN11和Ki67可能共同参与结直肠癌肿瘤细胞的增殖调控,并促进结直肠癌的发生发展以及浸润转移。     

Flow-cytometric analysis of the DNA-content in paraffin-embedded tissue from colorectal carcinomas and its prognostic significance

The nuclear DNA content of 163 colorectal carcinomas was determined by flow-cytometry (FCM) on formalin-fixed, paraffin-embedded tissue. DNA-aneuploidy was found in 97 cases (59.5%), in which no statistically significant correlations with sex, mean age, tumour stage (Dukes and pTNM) and tumour grade were noted. The frequency of aneuploidy was significantly higher in patients less than 70 years of age (p < 0.01) and in tumours localized in the left colon and rectum (p< 0.002), irrespective of their stage. The tumours in which different areas could be analysed (n = 80) showed a heterogeneous DNA-ploidy pattern in 18%. Comparison of the DNA content in primary tumours and in lymph node metastases (n = 49) showed a difference in DNA-ploidy in 38% of the DNA-aneuploid tumours, but in only 6% of the DNA-diploid carcinomas (p<0.02). DNA-aneuploid carcinomas tended to show a higher rate of local recurrence and were associated with an unfavourable prognosis (p = 0.04) in those patients in which complete resection of their tumours was possible (n = 72). The significantly higher mortality of patients with DNA-aneuploid carcinomas of stage pT3, as well as those with Dukes stage A and B tumours indicates that DNA-aneuploidy may be a stage-independent additional risk factor in colorectal cancer.     

Tissue microarray-determined expression profiles of cyclooxygenase-2 in colorectal adenocarcinoma: association with clinicopathological parameters

Accumulated evidence reveals that increased cyclooxygenase-2 (COX-2) is involved in the development of colorectal cancer. Our purpose was to quantitate COX-2 expression in colorectal cancers using tissue microarray analysis and look for an association with clinicopathological stage. Immunohistochemical analysis of COX-2 was performed in tissue microarray slides containing 90 specimens including 32 well-differentiated, 35 moderately differentiated, and 23 poorly differentiated colorectal adenocarcinomas. All colorectal adenocarcinomas showed significant immunohistochemical expression of COX-2 when compared to normal colon epithelia. However, there was no significant difference in immunostaining scores between poorly, moderately, and well-differentiated tumors (195 +/- 28, 214 +/- 26 and 200 +/- 24, respectively). The COX-2 immunostaining score correlated significantly with T stage (P < 0.05) but not with N or M stage. The positive expression rates of CK20 were 97% for well-differentiated, 94% for moderately differentiated, and 65% for poorly differentiated colorectal adenocarcinomas, suggesting that CK20 may not be an effective discriminator between poorly differentiated colorectal adenocarcinoma and metastatic adenocarcinoma.     

Flow-cytometric analysis of the DNA-content in paraffin-embedded tissue from colorectal carcinomas and its prognostic significance.

The nuclear DNA content of 163 colorectal carcinomas was determined by flow-cytometry (FCM) on formalin-fixed, paraffin-embedded tissue. DNA-aneuploidy was found in 97 cases (59.5%), in which no statistically significant correlations with sex, mean age, tumour stage (Dukes and pTNM) and tumour grade were noted. The frequency of aneuploidy was significantly higher in patients less than 70 years of age (p less than 0.01) and in tumours localized in the left colon and rectum (p less than 0.002), irrespective of their stage. The tumours in which different areas could be analysed (n = 80) showed a heterogeneous DNA-ploidy pattern in 18%. Comparison of the DNA content in primary tumours and in lymph node metastases (n = 49) showed a difference in DNA-ploidy in 38% of the DNA-aneuploid tumours, but in only 6% of the DNA-diploid carcinomas (p less than 0.02). DNA-aneuploid carcinomas tended to show a higher rate of local recurrence and were associated with an unfavourable prognosis (p = 0.04) in those patients in which complete resection of their tumours was possible (n = 72). The significantly higher mortality of patients with DNA-aneuploid carcinomas of stage pT3, as well as those with Dukes stage A and B tumours indicates that DNA-aneuploidy may be a stage-independent additional risk factor in colorectal cancer.     

Tumour budding intensity in relation to cathepsin D expression and some clinicopathological features of colorectal cancer

Although it has been suggested that tumour budding at the invasive edge of colorectal cancer is an important prognostic factor its biological significance for tumour progression is still to be evaluated. The aim of the study was to correlate tumour budding intensity with cathepsin D expression and some other clinicopathological variables of presumed or established prognostic value. 48 patients with colorectal cancer at pT3 stage, G2 grade of histological differentiation and tumour budding at the invasive edge were evaluated. Colorectal tumours were investigated for cathepsin D expression by immunohistochemistry of formalin-fixed and paraffin embedded tissues. There was no statistically significant relationships between tumour budding intensity grade and primary tumour cathepsin D expression, stromal cell cathepsin D expression and histochemical immunostaining of cathepsin D in rumour budding at its invasive edge. The tumour budding intensity was not associated with lymph node status, tumour site, peritumoral inflammatory response as well as the patient's age and sex. The results of this study suggest that intensity of tumour budds formation at the invasive margin of colorectal cancer is not associated with presumed or established prognostic factors such as lymph node metastases, and peritumoural inflammatory reaction as well as cathepsin D expression.     

Molecular correlates and prognostic significance of SATB1 expression in colorectal cancer

ABSTRACT: BACKGROUND: Special AT-rich sequence-binding protein 1 (SATB1) is a global gene regulator that has been reported to confer malignant behavior and associate with poor prognosis in several cancer forms. SATB1 expression has been demonstrated to correlate with unfavourable tumour characteristics in rectal cancer, but its association with clinical outcome in colorectal cancer (CRC) remains unclear. In this study, we examined the prognostic impact of SATB1 expression in CRC, and its association with important molecular characteristics; i.e. beta-catenin overexpression, microsatellite instability (MSI) screening status, and SATB2 expression. METHODS: Immunohistochemical expression of SATB1 and beta-catenin was assessed in tissue microarrays with tumours from 529 incident CRC cases in the prospective population-based Malmo Diet and Cancer Study, previously analysed for SATB2 expression and MSI screening status. Spearman[ACUTE ACCENT]s Rho and Chi-Square tests were used to explore correlations between SATB1 expression, clinicopathological and investigative parameters. Kaplan Meier analysis and Cox proportional hazards modelling were used to explore the impact of SATB1 expression on cancer specific survival (CSS) and overall survival (OS). RESULTS: SATB1 was expressed in 222 (42%) CRC cases and negative, or sparsely expressed, in adjacent colorectal mucosa (n = 16). SATB1 expression was significantly associated with microsatellite stable tumours (p < 0.001), beta-catenin overexpression (p < 0.001) and SATB2 expression (p < 0.001). While not prognostic in the full cohort, SATB1 expression was significantly associated with poor prognosis in SATB2 negative tumours (HR = 2.63; 95% CI 1.46-4.71; pinteraction = 0.011 for CSS and HR = 2.31; 95% CI 1.32-4.04; pinteraction = 0.015 for OS), remaining significant in multivariable analysis. CONCLUSIONS: The results of this study demonstrate that SATB1 expression in CRC is significantly associated with beta-catenin overexpression, microsatellite stability and SATB2 expression. Furthermore, SATB1 expression is a factor of poor prognosis in SATB2 negative tumours. Altogether, these data indicate an important role for SATB1 in colorectal carcinogenesis and suggest prognostically antagonistic effects of SATB1 and SATB2. The mechanistic basis for these observations warrants further study.Virtual slidesThe virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1922643082772076.     

MAT2A与NDRG2基因在结直肠癌中表达情况及临床意义的研究

目的 研究MAT2A（甲硫氨酸腺苷转移酶2A）和NDRG2（N-Myc下游调节基因-2）基因在结直肠癌组织中的表达及两者的相关性,为结直肠癌的研究及治疗提供重要参考。方法 选取68例术中切除结直肠癌组织及相应的癌旁组织标本,应用RT-PCR（逆转录‒聚合酶链式反应）和Western blotting（蛋白质印迹法）检测结直肠癌中MAT2A和NDRG2表达水平。结果 结直肠癌中MAT2A和NDRG2 mRNA表达情况：MAT2A阳性表达率（66.2%,45/68）高于癌旁组织（7.4%,5/68）;NDRG2阳性表达率（36.8%,25/68）低于癌旁组织（91.2%,62/68）;两者的表达与患者年龄、性别、肿瘤生长位置等因素均无关,与癌肿临床分期、分化水平及淋巴结转移密切相关。癌肿组织中MAT2A蛋白表达量相较于癌旁组织明显增多（t=39.1152,P=0.0000）,而癌肿组织中NDRG2蛋白表达量较癌旁组织明显降低（t=46.5103,P=0.0000）;且在结直肠癌组织中两基因的表达可能有相关性（χ2=7.41,P=0.009）。结论 在结直肠癌组织中MAT2A表达增高,NDRG2表达低下,两者与肿瘤的发生发展具有一定相关性,推测可能与DNA甲基化异常有关,但具体机制仍待进一步研究。     

The role of EMMPRIN expression in ovarian epithelial carcinomas

Purpose: Extracellular matrix metalloproteinase inducer (EMMPRIN) was reported to involve in the invasion and metastasis of malignancies by regulating the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMPs) in stromal and cancer cells. The study aimed to clarify the role of EMMPRIN expression in tumorigenesis and progression of ovarian epithelial carcinomas.

Methods: EMMPRIN siRNA were transfected into ovarian carcinoma cells with the phenotypes and their related molecules examined. EMMPRIN expression was determined in ovarian normal tissue, benign and borderline tumors, and epithelial carcinomas by real-time PCR, western blot, and immunohistochemisty.

Results: EMMPRIN siRNA treatment resulted in a lower growth, G₁ arrest, apoptotic induction, decreased migration, and invasion. The transfectants showed reduced expression of Wnt5a, Akt, p70s6k, Bcl-xL, survivin, VEGF, and MMP-9 than mock and control cells at both mRNA and protein levels. According to real-time PCR and western blot, EMMPRIN mRNA or protein level was higher in ovarian borderline tumor and carcinoma than normal ovary and benign tumors (P < 0.05), and positively correlated with dedifferentiation and FIGO staging (P < 0.05). Immuhistochemically, EMMPRIN expression was positively correlated with FIGO staging, dedifferentiation, Ki-67 expression, the lower cumulative and relapse-free survival rate (P < 0.05).

Conclusions: Upregulated expression of EMMPRIN protein and mRNA might be involved in the pathogenesis, differentiation, and progression of ovarian carcinomas, possibly by modulating cellular events, such as proliferation, cell cycle, apoptosis, migration, and invasion.     

Prognostic significance of mucin expression profiles in breast carcinoma with signet ring cells: a clinicopathological study

Background

Signet ring cells (SRCs) often accompany gastrointestinal carcinoma, referred to as SRC carcinoma; however, breast cancers containing SRCs have not been well characterized, leaving the prognostic significance of SRCs undetermined. We have described clinicopathological characteristics of patients with breast cancer containing SRCs in relation to the expression levels of MUC1, MUC2, MUC4, MUC5AC, and MUC6.

Methods

Twenty-two breast cancer cases with variable degrees of SRC population were retrospectively studied. Each case was categorized as high (>31 %) or low (<30 %) SRC tumor. The SRCs were morphologically classified into the intra-cytoplasmic lumen (ICL) type, or the non-ICL type. The expression levels of MUC1, MUC2, MUC4, MUC5AC and MUC6 were determined immunohistochemically. Depending on its subcellular localization, MUC1 was categorized as the luminal and cytoplasmic (LC) type, or the cytoplasmic with circumferential membranous accentuation (CM) type. These histological findings were compared with other clinicopathological parameters.

Results

The series consisted of invasive ductal carcinoma (n?=?9), invasive lobular carcinoma (n?=?9), and mucinous carcinoma (n?=?4) cases. The SRC population accounted for 8–81 % of the tumor cells. Eight cases had ICL type SRCs, and the remaining 14 had non-ICL type SRCs. Neither the high (n?=?12) and low (n?=?10) percentage of SRCs, nor the SRC types affected the clinicopathological parameters. In the low MUC1 group (n?=?11), larger tumors, higher nuclear grade, lymph node metastasis, and negativity for estrogen receptor was more frequently identified compared to the high MUC1 group (n?=?11; p?=?0.01, p?=?0.002, p?=?0.008, and p?=?0.02, respectively). The CM group (n?=?7) had more patients with large-sized tumors, lymph node metastasis, lymphovascular invasion, and higher Ki67 indices than the LC group (n?=?15; p?=?0.04, p?=?0.001, p?=?0.006, and p?=?0.03, respectively). The expression levels of MUC2, MUC4, MUC5AC, and MUC6 showed no clinicopathological significance. Two patients with low MUC1 expression and CM patterns had tumor recurrence, resulting in death, while all the other patients survived without recurrence.

Conclusion

Our results demonstrate that in breast cancers containing SRCs, low MUC1 expression and/or its CM subcellular localization patterns are associated with unfavorable clinicopathological factors. The utility of MUC1 expression as a prognostic marker remains to be verified in future studies.

     

The roles of REIC gene and its encoding product in gastric carcinoma

REIC is downregulated in immortalized cell lines compared with the parental normal counterparts. It may inhibit colony formation, tumor growth and induce apoptosis. Here, gastric carcinoma or epithelial cells transfected with REIC-expressing plasmid, its siRNA or treated with recombinant REIC were subjected to the phenotypes’ measurement or related molecules’ detection. REIC expression was examined in gastric carcinomas by RT-PCR, western blot and immunohistochemistry. REIC overexpression or treatment resulted in a low karyoplasmic ratio and proliferation, G₁ arrest, high apoptosis, low migration, invasion or lamellipodia formation in AGS cells. REIC knockdown caused the opposite in GES-1 cells. Anti-REIC antibody blocked the effects of REIC overexpression on proliferation, G₁/S progression and apoptosis. Ectopic REIC expression downregulated the expression of β-catenin, phosphorylated S6K (Thr389), phosphorylated Akt1/2/3 (Ser473), cyclin D2 and E, WAVE2 and upregulated phosphorylated mTOR (Ser2448) expression and the mRNA level of Akt1, Akt2, mTOR, Raptor and Rictor in AGS cells. REIC expression was negatively associated with tumor size, lymph node metastasis, dedifferentiation or poor prognosis of carcinoma. The serum REIC level was significantly higher in healthy individuals than the carcinoma patients and inversely linked to tumor size by ELISA. The possible mechanisms underlying the forced REIC overexpression or recombinant REIC mediated the reversal of the aggressive phenotypes of gastric carcinoma cells are to downregulate β-catenin and WAVE2 expression and to alter other related target proteins. Downregulated REIC expression was closely linked to aggressive behaviors and poor prognosis of gastric carcinoma.     

Associations between the cyclooxygenase-2 expression in circulating tumor cells and the clinicopathological features of patients with colorectal cancer

While previous studies have shown that the number of circulating tumor cells (CTCs) alone is not sufficient to reflect tumor progression and that cyclooxygenase-2 (COX-2) expression is correlated with colorectal cancer (CRC) metastasis, COX-2 expression status and its potential functions in CTCs of CRC patients are unknown. Here, epithelial-mesenchymal transition (EMT) phenotype-based subsets of CTCs and the COX-2 expression status in CTCs were identified and their potential clinical values were assessed in 91 CRC patients. CTCs were enumerated in peripheral blood and subsets of CTCs (epithelial [eCTCs], mesenchymal [mCTCs], and biophenotypic [bCTCs]) and the COX-2 expression status were determined using the RNA in situ hybridization method. CTCs were detected in 80.2% (73 of 91) patients. Neither the total CTC nor eCTC numbers were found to significantly associate with any of the clinicopathological features. However, the number of mCTCs was significantly associated with distance metastasis (P = 0.035) and had a trend of being associated with lymph node metastasis ( P = 0.055). Among the 73 patients enrolled for evaluating COX-2 expression, 52.5% (38 of 73) were found to express COX-2 in CTCs, and COX-2 expression in CTCs was not found to associate with the clinicopathological factors. However, COX-2 expression in mCTCs tended to have a higher rate in patients with metastasis compared with those without metastasis (72.0% vs 42.8%; P = 0.072). Furthermore, COX-2 expression and mCTC marker expression correlated positively ( R = 0.287; P = 0.017). Further studies are required to investigate the clinical value of the expression of COX-2 in mCTCs, especially in CRC patients with the advanced tumor stage and distant metastasis.     

The clinicopathological significance of angiogenesis in hindgut neuroendocrine tumors obtained via an endoscopic procedure

Background

As the World Health Organization grading system for gastroenteropancreatic-neuroendocrine tumors (GEP-NETs) may not always correlate with tumor progression, it is imperative that other independent predictors of tumor progression be established. To identify such predictors, we conducted a retrospective histopathological study of hindgut NETs, obtained from endoscopic procedures, and used statistical analyses to evaluate predictive factors.

Methods

We first obtained clinicopathological data of cases of hindgut NETs. Tissue sections from tumor samples were prepared and subjected to pathological examination. In particular, we calculated the microvessel density (MVD) and lymphatic microvessel density (LMVD) values, and performed appropriate statistical analyses.

Results

A total of 42 cases of hindgut NETs were selected for the study, 41 from the rectum and 1 from the sigmoid colon. Based on the Ki-67 labeling index, 34 cases were classified as NET G1 tumors and 8 as NET G2 tumors. MVD values ranged from 1.4/mm² to 73.9/mm² and LMVD values from 0/mm² to 22.9/mm². MVD and LMVD were identified as risk factors for venous and lymphatic invasion of hindgut NETs. Moreover, MVD positively correlated with the maximum diameter of the tumor.

Conclusions

Tumor progression of NETs may cause angiogenesis and lymphangiogenesis, via an unknown mechanism, as well as lymphovascular invasion. Angiogenesis likely plays an important role in occurrence and progression in the initial phase of hindgut NETs.

     

Prospective study on prognostic significance of DNA ploidy and Ki-67 expression in colorectal cancer

The aim of the study was to correlate tumoral DNA ploidy and Ki-67 expression with therapy response, Overall Survival (OS), Disease Specific Survival (DSS) and Disease Free Survival (DFS). Three samples of colorectal cancer were collected from each patient. One sample of normal tissue was our internal control. DNA ploidy was evaluated by FACSCalibur cytometer and Ki-67 by immunohistochemistry. We studied 67 patients and we found aneuploidy in 65,7 percent of carcinoma with a Ki-67 median expression of 55 percent. After surgery and chemotherapy in 35 percent of the patients with aneuploid carcinoma and high proliferative activity (Ki-67 greater than 55 percent) there were no evidence of disease versus 100 percent of patients with DNA diploidy and low proliferative activity (Ki-67 less than 55 percent). Tumoral aneuploidy significantly correlated with lower OS, DSS and DFS (18 percent vs 86 percent at 30 months). Univariated analysis demonstrated a significant correlation between aneuploidy and develop disease progression (p=0,033, odd ratio=5.7), while the cut-off of 55 percent for Ki-67 expression did not correlate with OS, DSS and DFS. Preliminary results (the study is still in progress) seemed to suggest that DNA ploidy has a prognostic and predictive significance in colorectal carcinoma.