A genome-wide analysis of the molecular alterations occurring in the adenomatous and carcinomatous components of the same tumor based on the adenoma–carcinoma sequence

Identification of molecular alterations occurring in the adenomatous and carcinomatous components within the same tumor would greatly enhance understanding of the neoplastic progression of colorectal cancer. We examined somatic copy number alterations (SCNAs) and mRNA expression at the corresponding loci involved in the adenoma–carcinoma sequence in the isolated adenomatous and cancer glands of the same tumor in 15 cases of microsatellite-stable “carcinoma in adenoma,” using genome-wide SNP and global gene expression arrays. Multiple copy-neutral loss of heterozygosity events were detected at 4q13.2, 15q15.1, and 14q24.3 in the adenomatous component and at 4q13.2, 15q15.1, and 14q24.3 in the carcinomatous component. There were significant differences in the copy number (CN) gain frequencies at 20q11.21–q13.33, 8q13.3, 8p23.1, and 8q21.2–q22.2 between the adenomatous and carcinomatous components. Finally, we found a high frequency of five genotypes involving CN gain with upregulated expression of the corresponding gene (RPS21, MIR3654, RSP20, SNORD54, or ASPH) in the carcinomatous component, whereas none of these genotypes were detected in the adenomatous component. This finding is interesting in that CN gain with upregulated gene expression may enhance gene function and play a crucial role in the progression of an adenoma into a carcinomatous lesion.     

Changes in the expression of syndecan-1 in the colorectal adenoma–carcinoma sequence

 Syndecan-1, a transmembrane heparan sulphate proteoglycan (HSPG), functions as a matrix receptor on the basal surface of epithelial cells. It also co-localizes with E-cadherin at the lateral cell surface where its function is uncertain. Tumour development in the large bowel is associated with loss of normal epithelial adhesion and altered patterns of expression of cell adhesion molecules, possibly including syndecan-1. To evaluate changes in syndecan-1 expression during the development of colorectal neoplasia, 59 adenomas and 20 carcinomas arising from adenomas were investigated by immunohistochemistry. The staining intensity and distribution of syndecan-1 and E-cadherin in sequential sections was examined, semi-quantified and compared. Staining of syndecan-1 and E-cadherin was uniform in normal colorectal epithelial cells, and located at the basolateral surface. No significant change was seen in either molecule in mildly or moderately dysplastic adenomas. A significant reduction in expression of both syndecan-1 and E-cadherin was seen in severely dysplastic epithelium as compared to moderate dysplasia (P=0.001 and P=0.004 respectively). Similarly, there was a significant reduction of both molecules in carcinomas compared with associated adenomas (syndecan-1 P=0.00003; E-cadherin P=0.002). In both cases the loss of syndecan-1 expression was more striking than that of E-cadherin. Previous in vitro studies have shown that epithelial cells made deficient in syndecan-1 cease to express E-cadherin, suggesting a causal association. Our results support these findings and indicate that disruption of cell-matrix adhesion is critical in colorectal carcinogenesis, probably preceding changes in the purely homotypic cell-cell adhesion mediated by E-cadherin. Received: 11 May 1998 / Accepted: 14 September 1998     

Numerical and structural centrosome aberrations are an early and stable event in the adenoma–carcinoma sequence of colorectal carcinomas

Aims Numerical and structural centrosome changes have been described for and linked with genetic instability in solid tumors. Here, we specifically address centrosome aberrations in the adenoma–carcinoma sequence of colorectal cancer by detailed evaluation of -tubulin staining patterns.Methods Formalin-fixed and paraffin-embedded specimens (normal colonic epithelium n=21; low-grade intraepithelial neoplasia n=27, high-grade intraepithelial neoplasia n=16 and invasive adenocarcinomas n=33) were stained by an anti--tubulin antibody using standard immunofluorescence. Three-dimensional image stacks of the stainings were recorded (Zeiss LSM510 confocal microscope), followed by numerical and structural data analysis (DIAS software package) and statistical evaluation (NCSS-software).Results The mean centrosome signal per cell differed significantly (P<0.0001) between normal colonic epithelium (0.8775) and each low-grade intraepithelial neoplasia (1.787), high-grade intraepithelial neoplasia (2.259) and invasive carcinomas (2.267). Similarly, both the centrosomes structural entropy (SE) and minimal spanning tree (MST) differed significantly (P<0.001) between normal (SE=3.956, MST=38.78) and each low- (SE=6.39, MST=26) and high-grade intraepithelial neoplasia (SE=5.75, MST=26.97) and invasive carcinoma (SE=6.86, MST=28.08).Conclusion Numerical and structural centrosome dysregulation is seen as early as in low-grade dysplastic lesions of the adenoma–carcinoma sequence of colorectal carcinomas and may, as such, play an initial role in the carcinogenic process.     

Pitfalls in diagnostic molecular pathology – significance of sampling error

Today, molecular diagnostic tests are widely used in clinical medicine with polymerase chain reaction (PCR)-based techniques being of particular interest. In tissue specimens, however, false-positive and false-negative results can be obtained if pathomorphological and processing aspects are not considered. We therefore studied the impact of tissue sampling in three widely used diagnostic tests: (1) assessment of clonality in B-cell non-Hodgkin's lymphoma, (2) analysis of microsatellite instability (MSI) in colorectal neoplasia, and (3) demonstration of mycobacterium tuberculosis. Tissue sections of routinely formalin-fixed and paraffin-embedded diagnostic specimens were taken, and the significance of sampling was systematically investigated using laser microdissection or processing of the entire section. PCR analyses were done according to standard protocols. False-positive pseudo-monoclonality was obtained in small gastrointestinal biopsies and in laser microdissected lymph follicles of non-neoplastic tonsils. False negativity (pseudo-stability) could be demonstrated in a case with hereditary rectal adenoma if whole tissue sections were taken without microdissection of the most dysplastic subareas. Demonstration of mycobacteria failed in tissue sections of a formalin-fixed lymph node that was positive after complete digestion or in fresh frozen material of the same patient. In diagnostic molecular pathology, sampling error is an important source of false-positive and false-negative results, particularly if disease- and tissue-specific morphological features, such as sample size, type of fixation, and intralesional heterogeneity, are ignored.     

Activity of the leukotriene pathway in Barrett’s metaplasia and oesophageal adenocarcinoma

Objective

Leukotriene (LT) B₄ is a lipid inflammatory mediator implicated in tumorigenesis in animal models of Barrett’s oesophagitis, but little is known about the cysteinyl-leukotrienes (LTC₄, LTD₄, LTE₄), which have distinct inflammatory and tumorigenic actions in other tissues. We recently showed that the terminal enzymes for the synthesis of both LT families are highly expressed in human oesophageal adenocarcinoma (OA) tissues. This study therefore examined the capacity of Barrett’s metaplasia (BM) and OA tissues to synthesise LTs in vitro.

Subjects and methods

Oesophageal biopsies from patients with BM (n?=?14), high-grade dysplasia (n?=?2), OA (n?=?11), and squamous control tissues (n?=?11) were cultured with calcium ionophore A32187 (2?μM) for 60?min. LTB₄ and cysteinyl-leukotrienes were extracted and measured by specific enzyme immunoassays.

Results

Levels of LTB₄ and cysteinyl-leukotrienes were 8.6-fold (P?<?0.01) and 2.4-fold (P?<?0.02) higher, respectively, in OA tissues than in squamous control tissues, but levels in BM tissues (n?=?14) were not altered. Production of the two LT families correlated across all tissue types (r?=?0.62, p?<?0.00005).

Conclusions

Increased synthesis of LTB₄ and cysteinyl-leukotrienes has not previously been shown in human OA tissue and our results may indicate a role of these lipids in Barrett’s disease progression.     

Clinical implications of hepatitis B viral infection in Epstein–Barr virus-associated nasopharyngeal carcinoma

BackgroundLittle is known about the clinical implication of hepatitis B virus (HBV) infection in Epstein–Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC).ObjectiveThis study aimed to investigate the clinical characteristics and prognostic factors in patients with newly-diagnosed NPC with HBV infection.Study designA total of 722 patients with pathologically-diagnosed NPC who received comprehensive treatment at First People's Hospital of Foshan between June 2006 and December 2011 were enrolled in this retrospective study; 79 and 643 patients were HBsAg(+) and HBsAg(−), respectively. The correlations between HBV (HBsAg status and HBV DNA load) and EBV DNA were analyzed, further long-term survival and prognostic factors also were explored.ResultsWe reported NPC patients with HBsAg(+) represented worse outcome, and distant-failure especially liver metastasis was more common in these patients. HBV infection was more frequent in younger patients and male patients. No correlation was observed between the pre-treatment plasma EBV DNA load (cutoff, 1500 copies/ml) and HBsAg status (positive or negative; r = −0.036, P = 0.392), or the pre-treatment plasma EBV DNA load and HBV DNA load (r = 0.042, P = 0823).ConclusionsBoth HBV and EBV infection is an independent negative prognostic factor for long-term survival, distant metastasis, especially liver metastasis, was more common in NPC patients with HBsAg(+), and it seemed no link between EBV DNA load and HBsAg status in NPC.     

Genome–environment interactions in the molecular pathogenesis of dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a heart muscle disease characterized by impaired contractility and dilation of the ventricles. In a subset of DCM patients, classical inheritance patterns occur (familial DCM), which have led to the identification of specific genomic loci and gene defects causing monogenic DCM subtypes. In the majority of DCM patients, however, there is no evidence for a monogenic etiology of the disorder (sporadic DCM), and in the absence of other recognizable etiological factors, these cases were classified as idiopathic. Recent research suggests that cardiotropic viruses are important environmental factors in the pathogenesis of idiopathic cases and that DCM commonly results from interactions between genetic and environmental factors, whereas pure genetic forms are rather rare. Regarding genetics, the clinical cardiomyopathic phenotype associated with single gene defects may be highly variable for unknown reasons. Furthermore, a novel class of genetic defects was identified recently which provide a molecular basis for abnormal reactions of cardiomyocytes to environmental stress. These defects are paradigms of specific molecular links between genome and environment during the pathogenesis of DCM. Regarding environmental factors, a recent molecular virological study based on myocardial biopsies in a large series of sporadic DCM patients has detected cardiac viral infections in the majority of patients, with a broad spectrum of virus species being involved. Apparently, DCM does not only occur as a late sequela of acute viral myocarditis, but also in patients without clinical history of cardiac viral disease. Cardiotropic viruses thus emerge as prevalent environmental factors which may cause or influence the course of DCM in a large fraction of cases. Synopsis of current data suggests that a comprehensive picture of DCM pathogenesis can only be drawn if both genetic and environmental pathogenetic factors are considered. The course of cardiac viral infections depends strongly on genetic host factors and may range from rapid and complete virus elimination or silencing without clinical symptoms, to rapidly progressive or fatal disease. Viruses interact not only with genetically heterogenous host systems of virus uptake, migration, and antiviral immunity, but, due to their prevalence in DCM hearts, are also likely to encounter multiple structural proteins of cardiac cells known to be defective in familial DCM. The combined knowledge on DCM-associated gene defects and viruses therefore suggests in-depth studies on genome–environment interactions in DCM pathogenesis which may underlie the high clinical variability observed both in monogenic and virus-associated DCM and have implications for the clinical management of DCM patients.     

The genetic and molecular basis of muscular dystrophy: roles of cell–matrix linkage in the pathogenesis

Muscular dystrophies are a heterogeneous group of genetic disorders. In addition to genetic information, a combination of various approaches such as the use of genetic animal models, muscle cell biology, and biochemistry has contributed to improving the understanding of the molecular basis of muscular dystrophy's etiology. Several lines of evidence confirm that the structural linkage between the muscle extracellular matrix and the cytoskeleton is crucial to prevent the progression of muscular dystrophy. The dystrophin–glycoprotein complex links the extracellular matrix to the cytoskeleton, and mutations in the component of this complex cause Duchenne-type or limb-girdle-type muscular dystrophy. Mutations in laminin or collagen VI, muscle matrix proteins, are known to cause a congenital type of muscular dystrophy. Moreover, it is not only the primary genetic defects in the structural or matrix proteins, but also the primary mutations of enzymes involved in the protein glycosylation pathway that are now recognized to disrupt the matrix–cell interaction in a certain group of muscular dystrophies. This group of diseases is caused by the secondary functional defects of dystroglycan, a transmembrane matrix receptor. This review considers recent advances in understanding the molecular pathogenesis of muscular dystrophies that can be caused by the disruption of the cell–matrix linkage.     

Roles of estrogen receptor α and β in the regulation of proliferation in endometrial carcinoma

Endometrial carcinoma (EC), an estrogen-dependent gynecological malignancy, is prevalent worldwide. Estrogen receptor α (ERα) and estrogen receptor β (ERβ) are two main estrogen receptor isoforms, which mediate estrogen-induced proliferation in EC. However, the dynamic changes of ERα and ERβ subtype expression and their functions on proliferation in EC remain elusive. In this study, we aimed to investigate the expression of ERα and ERβ in para-tumor eutopic endometrium, endometrial atypical hyperplasia and EC by immunohistochemistry and then analyse their clinical significance. Subsequently, Ishikawa cells with ERα or ERβ knockdown by lentivirus transfection were used to explore the relationship between ERα/ERβ and cell proliferation, and preliminarily evaluate whether metformin’s inhibitory effect on estrogen-induced cell proliferation was mediated by ERα and ERβ. We found that the expression of ERα and ERβ were markedly changed in endometrial hyperplasia and EC compared with that in para-tumor eutopic endometrium and exhibited different expression trends. Through further analysis, we discovered that ERα presented higher expression in endometrial atypical hyperplasia and early stage of EC than that in para-tumor eutopic endometrium, while the expression of ERβ gradually decreased from para-tumor eutopic endometrium to EC. Additionally, the cell cycle-related protein, CyclinD1 was gradually increased but p21 decreased. Furthermore, knockdown of ERα and ERβ severally in Ishikawa cells either inhibited or promoted estrogen-induced cell proliferation through regulating CyclinD1 and p21 expression. Meanwhile, the inhibitory effect of metformin on estrogen-induced cell proliferation was respectively blunted or partly reversed by knockdown of ERα or ERβ. Altogether, ERα and ERβ have different expression patterns in the progression of EC either facilitating or suppressing cell proliferation through regulating the expression of CyclinD1 and p21 in EC cells, and may also mediate the inhibitory effect of metformin on estrogen-induced EC cells proliferation.     

Expression and clinical significance of PI3Kp110β in human colorectal carcinoma and colorectal adenoma with different grades of dysplasia

目的 探讨PI3Kp110β表达与结直肠癌发生、发展的相关性及其临床意义.方法 应用免疫组化EnVision法检测PI3Kp110β在138例结直肠腺癌、29例低级别上皮内瘤变(low-grade intraepithelial neoplasia,LGIN)腺瘤、35例高级别上皮内瘤变(high-grade intraepithelial neoplasia,HGIN)腺瘤、10例结肠正常黏膜组织中的表达情况.结果 (1)PI3Kp110β在结直肠癌组织中的表达明显高于结直肠LGIN腺瘤、HGIN腺瘤及正常黏膜组织,其差异有统计学意义(P<0.05),PI3Kp110β在结直肠HGIN腺瘤组织中的表达与结直肠LGIN腺瘤、正常黏膜组织中的表达相比,差异无显著性(P>0.05);(2)结直肠癌组织中PI3Kp110β的表达与肿瘤的发生部位、分化程度、临床分期及淋巴结转移等临床因素间具有相关性(P<0.05),与患者的性别、年龄及肿瘤大小等因素无关(P>0.05).结论 PI3Kp110β异常表达可能在结直肠癌的发生、发展中发挥着重要作用,检测PI3Kp110β对评判结直肠癌的生物学行为及预后有一定临床意义.     

Evolution of the term and definition of dysplasia of the hip – a review of the literature

There is no consensus on the definition of dysplasia of the hip (DH). Past and present concepts used to describe DH do not form a complete view of the pathology. Moreover, some authors still present the disease as congenital, not developmental. This prompted authors to analyze the evolution of the definition of DH. Based on the biomedical databases 500 articles and books in the field of hip dysplasia were found and analyzed. Fifteen definitions of hip dysplasia met inclusion criteria, subsequently were analyzed and presented in chronological order. The analysis revealed that currently there is no single, universal definition of hip dysplasia in the aspect of morphological, clinical, and radiological studies. Despite the widely-used term of DH, it is described imprecisely and in different ways. Therefore, it is necessary to develop a multidisciplinary definition of this pathology covering all aspects of hip disorders considered valid in modern orthopaedics.     

Enterovirus infections in England and Wales, 2000–2011: the impact of increased molecular diagnostics

There have recently been significant changes in diagnostic practices for detecting enterovirus (EV) infections across England and Wales. Reports of laboratory-confirmed EV infections submitted by National Health Service (NHS) hospital laboratories to Public Health England (PHE) over a 12-year period (2000–2011) were analysed. Additionally, the PHE Virus Reference Department (VRD) electronic database containing molecular typing data from 2004 onwards was interrogated. Of the 13 901 reports, there was a decline from a peak of 2254 in 2001 to 589 in 2006, and then an increase year-on-year to 1634 in 2011. This increase coincided with increasing PCR-based laboratory diagnosis, which accounted for 36% of reported cases in 2000 and 92% in 2011. The estimated annual incidence in 2011 was 3.9/100 000 overall and 238/100 000 in those aged <3 months, who accounted for almost one-quarter of reported cases (n = 2993, 23%). During 2004–2011, 2770 strains were submitted for molecular typing to the VRD, who found no evidence for a predominance of any particular strain. Thus, the recent increase in reported cases closely reflects the increase in PCR testing by NHS hospitals, but is associated with a lower proportion of samples being submitted for molecular typing. The high EV rate in young infants merits further investigation to inform evidence-based management guidance.     

Nuclear factor κB and cyclooxygenase-2 immunoexpression in oral dysplasia and oral squamous cell carcinoma

Oral leukoplakia is the main potentially malignant oral lesion, and oral squamous cell carcinoma accounts for more than 95% of all malignant neoplasms in the oral cavity. Therefore, the aim of this study was to verify the immunoexpression of nuclear factor κB (NF-κB) and cyclooxygenase-2 (COX-2) proteins in dysplastic oral lesions and oral squamous cell carcinoma. Immunohistochemical reactions were performed on 6 inflammatory fibrous hyperplasia, 28 oral leukoplakia, and 15 oral squamous cell carcinoma paraffin-embedded samples. Immunoperoxidase reaction for NF-κB and COX-2 was applied on the specimens, and the positivity of the reactions was calculated for 1000 epithelial cells. Using the analysis of variance and the Tukey post hoc statistical analyses, a significantly increased immunoexpression for NF-κB was observed when oral squamous cell carcinoma samples were compared with the other groups studied. However, using the Kruskal-Wallis and the Dunn post hoc tests, a statistically significant result for COX-2 expression was obtained only when the moderate dysplasia group was compared with the inflammatory fibrous hyperplasia group. Nuclear factor κB may participate in the malignant phenotype acquisition process of the oral squamous cell carcinoma in its late stages, whereas COX-2 may be involved in the early stages of oral carcinogenesis process.     

Intrauterine diagnosis and pathology of fetal choroid plexus carcinoma – A case study

This is a case report of a 39-year-old pregnant woman whose fetus was found to have a large hydrocephalus on routine prenatal ultrasound at the 29th gestational week. A 56 mm × 73 mm mass was detected in the fetal brain arising from the brainstem and invading the third cerebral ventricle. On the subsequent fetal cranial MRI, T2-weighted image the tumor measured 55 mm × 50 mm × 48 mm and had a non-homogeneous consistency and irregular contours. Elective cesarean section was performed during gestational week 32, delivering a male fetus with a cranial circumference of 46 cm (normal circumference, 30 cm) and a birth weight of 2920 g. The infant expired 4 h following delivery. Autopsy revealed a carcinoma of the choroid plexus. Our case, like others, suggests that MRI is more accurate than prenatal ultrasound in prenatal brain tumor diagnosis. More precise morphological detail is provided by MRI, which improves surgical planning and survival.