Histology and histopathology Vol.28, nº 3 (2013)

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Open Access
Immunohistochemical study of enteric nervous system in hirschsprung’s disease and intestinal neuronal dysplasia
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Geramizadeh, Bita; Akbarzadeh, Elahe; Izadi, Babak; Foroutan, Hamid-Reza; Heidari, Tahere
Background. Hirschsprung’s disease (HD) is one of the most common motility disorders in pediatric age groups and it is very important that it be differentiated from other types of motility disorders, especially intestinal neuronal dysplasia B (IND B). Although many studies regarding the differences between the two disorders by immunohistochemical studies exist, there is as yet no consistent result. The purpose of this research was to study the immunohistochemical findings of enteric nervous system in these two motility disorders in comparison with colectomies without motility disorder. Methods. Full wall thickness specimens of three groups of patients (HD, IND B and non motility disorders) were included in the study to be evaluated by immunohistochemistry (IHC). Markers were specific for neuronal cells and pace maker cells composed of PGP 9.5, c-kit, synaptophysin, S100 and CD56. The number of cells was evaluated in the muscularis properia, and myenteric plexus. Results. The number of all the IHC markers i.e. PGP9.5, c-kit, synaptophysin, S100 and CD56 was completely different in HD from the two other groups, while IND B was quite similar to control group. Conclusion. Our finding suggests that there is a marked and significant difference between HD and IND B by IHC markers, which can be used as an additional test for the diagnosis of HD with more accuracy. Further multicenter studies with a greater number of cases would be necessary to find a cut-off point for every IHC marker to differentiate HD and IND B.
Open Access
Levels of acyl-Coenzyme A synthetase 5 in urothelial cells and corresponding neoplasias reflect cellular differentiation
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Gaisa, Nadine T.; Reinartz, Andrea; Schneider, Ursula; Klaus, Christina; Heidenreich, Axel; Jakse, Gerhard; Kaemmerer, Elke; Klinkhammer, Barbara Mara; Knuechel, Ruth; Gassler, Nikolaus
Metabolic components like fatty acids and acyl-Coenzyme A (acyl-CoA) thioesters have been implicated in the pathogenesis of various tumours. The activation of fatty acids to acyl-CoAs is catalysed by long chain acyl-CoA synthetases (ACSLs), and impairment of ACSL expression levels has been associated with tumourigenesis and progression. Since ACSLs have never been investigated in bladder tissues, the study aims to characterize ACSL expression and acyl-CoA synthesis in normal and neoplastic bladder tissues, as well as cell lines. ACSL isoforms 1, 3, 4 and 5 and synthesis of acyl-CoAs were analysed using qRTPCR, western blot analysis, immunohistochemistry and lipid mass spectrometry. In normal urothelium, expression of ACSL1, 3, 4 and 5, with highest levels of ACSL isoform 5 was found. However, ACSL5 expression was reduced in corresponding neoplastic tissues and urothelial cell lines depending on the grade of cellular differentiation. AntiACSL5 immunostainings showed expression in normal urothelium and a gradual loss of ACSL5 protein via preinvasive lesions to invasive carcinomas. High expression of ACSL5 correlated with increased α-galactosidase activity and positive Uroplakin III staining in tumours. In contrast, synthesis of acyl-CoAs was enhanced in neoplastic bladder tissues compared to normal urothelium, and reflected an increase with respect to cellular differentiation. These results confirm an expression of ACSLs, especially isoform 5, in human urothelium, prove enzymatic/lipidomic changes in bladder cancer tissues, and suggest an involvement of ACSL5 in cellular maturation and/or senescence with possible effects onto induction of tumour formation or progression. Further work may identify responsible pathway alterations, and attempting to re-balance the metabolic equilibrium of the urothelium may offer a further opportunity for tumour treatment and prevention.
Open Access
DKC1 gene mutations in human sporadic cancer
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Penzo, Marianna; Casoli, Lucia; Ceccarelli, Claudio; Treré, Davide; Ludovini, Vienna; Crinò, Lucio; Montanaro, Lorenzo
Introduction: Germline mutations in the tumour suppressor gene dyskeratosis congenit 1 (DKC1) cause the cancer prone syndrome called X-linked dyskeratosis congenita. The present study aims to determine whether mutations of the DKC1 gene may also be present in frequent human sporadic cancers (breast, colon and lung cancers), thus potentially contributing to the neoplastic phenotype. Materials and methods: mutation analysis of the DKC1 gene was performed on DNA from 110 primary human lung, 54 breast, and 35 colon cancers, focusing on gene regions where pathogenic germline mutations have been described previously (promoter and exons 1, 3, 9, 10, 11, and 14). Results: Out of a total of 199 primary tumours of different origins, only 5 turned out to have sequence variations in the DKC1 gene. These variations were of two kinds, C8120T and C13554T, which are both classified as synonymous mutations and do not affect DKC1 mRNA splicing. Conclusion: direct DKC1 gene mutations are not a frequent event in tumourigenesis, at least in the tumour types investigated and for the DKC1 gene portions considered in this study.
Open Access
Clear cell papillary cystadenocarcinoma of the epididymis: a case report and immunohistochemistry of markers for renal cell carcinoma
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Nozawa, Tatsuru; Konda, Ryuichiro; Ohsawa, Taisuke; Yoshida, Makoto; Komatsu, Masayo; Iwama, Takahide; Fujioka, Tomoaki
Neoplasms of the epididymis are uncommon, and malignant tumors are extremely rare. We report a case of clear cell papillary cystadenocarcinoma of the epididymis presenting with a long history of painless scrotal mass on the left side. Immunohistochemical markers for clear cell renal cell carcinoma (RCC) were examined to distinguish between clear cell papillary cystadenocarcinoma of the epididymis and metastatic clear cell renal cell carcinoma. The present case was positive for cytokeratin-7, PAX2, vinculin, vimentin and carbonic anhydrase IX. Expression of CD10 was focally observed. In contrast, no immunoreactivities for α- methylacyl-CoA racemase, RCC marker, glutathione Stransferase α or C-KIT were detected. The immunophenotypic profile of clear cell papillary cystadenocarcinoma of the epididymis closely resembles that of clear cell papillary RCC, although the immunohistochemical markers tested in this study are useful to make a differential diagnosis between clear cell papillary cystadenocarcinoma of the epididymis and metastatic clear cell RCC.
Open Access
Chromosome 16q loss– a genetic key to the understanding of breast carcinogenesis
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Bürger, Horst; Boer, Mirthe de; Van Diest, Paul J.; Korsching, Eberhard
In the last decade the concepts of breast cancer dedifferentiation and progression have undergone a significant and substantial change. In the past it was widely believed that the detailed associations between genetic and morphological changes defined in the Vogelstein model of colorectal cancer pathogenesis could be transferred to breast carcinogenesis. A multitude of studies seemed to verify this a priori hypothesis. However, with the introduction of global screening techniques, predominantly at the DNA level, it became obvious that this linear model might be oversimplified for breast cancer. It is now widely accepted that losses of chromosomal 16q characterize in-situ and invasive breast cancer tumours with predominantly low tumour grade and estrogen receptor (ER) positivity (luminal breast cancers). In contrast, high grade breast cancers of the HER2, the basal or the non expressor phenotype with 16q-losses are rarely seen and in consequence a concept of multiple, parallel pathways with defined precursor lesions emerged. As a consequence, it became obvious that the hunt for oncogenes/tumour suppressor genes in invasive breast cancer is pathway specific. Whereas high grade breast cancers have been relatively well characterized by several recurrent changes in oncogenes/tumour suppressor genes located on various chromosomal regions (e.g. egfr, p53, HER2), the characterization of a 16q-specific tumour suppressor gene in ER-positive breast cancer is still a tremendous challenge. This review will focus on the role of 16q in breast cancer and aims to give insights into actual research efforts, e.g. alternative explanations in order to unravel the central role of 16q in breast cancer.
Open Access
Role of podoplanin expression in squamous cell carcinoma of upper aerodigestive tract
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Chuang, Wen-Yu; Chang, Yu-Sun; Yeh, Chi-Ju; Wu, Yi-Chin; Hsueh, Chuen
Podoplanin, a type-1 transmembrane glycoprotein, was originally named due to its expression in renal podocytes of rats. It was subsequently detected in a variety of normal human tissues, including lymphatic endothelium. Although podoplanin has been identified as the endogenous ligand of C-type lectin-like receptor 2 (CLEC-2) on platelets, its physiological functions and pathways remain largely unknown. A role in lymphangiogenesis has been suggested, since podoplanin-deficient mice were found to die at birth with a phenotype of dilated, malfunctioning lymphatic vessels and lymphedema. Podoplanin is invariably expressed in some tumors, such as lymphangioma, seminoma and follicular dendritic cell tumor, but tumor cell expression of podoplanin is highly variable in squamous cell carcinoma (SCC). It has been found that high podoplanin expression is associated with lymph node metastasis and poor prognosis in SCC of the upper aerodigestive tract. Now there is growing evidence that podoplanin is also involved in carcinogenesis, cell motility, tumor invasiveness, platelet aggregation and hematogenous metastasis. Additionally, animal studies confirmed some in vivo effects of podoplaninoverexpressing tumors, including formation of more tumor lymphatic vessels, larger lymph node metastases, more platelet aggregation, and more pulmonary metastases. Several recently developed anti-podoplanin antibodies, such as NZ-1, P2-0 and hP2-0, have been shown to attenuate podoplanin-induced platelet aggregation and prevent experimental hematogenous metastasis in nude mice. These antibodies may be applied in preclinical and clinical studies to evaluate the possibility of podoplanin-targeted therapy.
Open Access
Histological examination on osteoblastic activities in the alveolar bone of transgenic mice with induced ablation of osteocytes
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Li, Minqi; Hasegawa, Tomoka; Hogo, Hiromi; Tatsumi, Sawako; Liu, Zhusheng; Guo, Ying; Sasaki, Muneteru; Tabata, Chihiro; Yamamoto, Tsuneyuki; Ikeda, Kyoji; Amizuka, Norio
The purpose of this study was to examine histological alterations on osteoblasts from the alveolar bone of transgenic mice with targeted ablation of osteoctyes. Eighteen weeks-old transgenic mice based on the diphtheria toxin (DT) receptor-mediated cell knockout (TRECK) system were used in these experiments. Mice were injected intraperitoneally with 50 µg/kg of DT in PBS, or only PBS as control. Two weeks after injections, mice were subjected to transcardiac perfusion with 4% paraformaldehyde in 0.1M phosphate buffer (pH 7.4), and the available alveolar bone was removed for histochemical analyses. Approximately 75% of osteocytes from alveolar bones became apoptotic after DT administration, and most osteocytic lacunae became empty. Osteoblastic numbers and alkaline phosphatase (ALP) activity were markedly reduced at the endosteum of alveolar bone after DT administration compared with the control. Osteoblastic ALP activity in the periodontal ligament region, on the other hand, hardly showed any differences between the two groups even though numbers were reduced in the experiment group. Silver impregnation showed a difference in the distribution of bone canaliculi between the portions near the endosteum and the periodontal ligament: the former appeared regularly arranged in contrast to the latter’s irregular distribution. Under transmission electron microscopy (TEM), the osteoblasts in the periodontal ligament showed direct contact with the Sharpey’s fibers. Thus, osteoblastic activity was affected by osteocyte ablation in general, but osteoblasts in contact with the periodontal ligament were less affected than endosteal osteoblasts.
Open Access
Different influence of ovine estrus synchronization treatments on caruncular early angiogenesis
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Ruiz-González, Irene; Sánchez, M.A.; García-Fernández, R.A.; García-Palencia, P.; Sánchez, B.; González-Bulnes, A.; Flores, J.M.
The present study compares two protocols for ovine estrus synchronization by assessing the caruncular angiogenic response to the establishment of pregnancy. The analysis consisted of the immunohistochemical evaluation of Vascular Endothelial Growth Factor (VEGF), Platelet Endothelial Cell Adhesion Molecule-1 (PECAM-1, CD31) and Von Willebrand Factor (vWF) in ovine caruncular stroma. A flock of thirty-eight adult ewes was divided in two groups and synchronized with either progestagens (Group P) or prostaglandin analogues (Group PG). Immunohistochemistry was performed in uterine samples obtained from pregnant ewes (P, n=15; PG, n=13) on days 15 post coitus (pc), 17pc and 21pc (day 0 =day of estrus). Each factor was assessed by total vascular density (TVD, total positive blood vessels/mm2), capillary vascular density (CVD, positive blood capillaries/mm2) and arteriolar vascular density (AVD, positive arterioles/mm2). Group P demonstrated higher VEGF-CVD (P=0.045) when compared to prostaglandin treated animals. Vascular CD31- expression decreased on days 15pc and 21pc (TVD, P=0.007 and CVD, P=0.014) in both groups. vWF analysis did not show significant differences between groups or days of study. These results demonstrate a different influence of progestagen-based and prostaglandin analogues-based synchronization treatments over VEGF vascular expression during caruncular development taking place in response to pregnancy establishment. In addition, observations pointed out in this study support the involvement of CD31 in the angiogenic stimulus that occurs during early maternal placentation in the ewe.
Open Access
Malignant transformation of nasal chondromesenchymal hamartoma in adult: a case report and review of the literature
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Li, Yang; Yang, Qing-xu; Tian, Xiao-ying; Li, Bin; Li, Zhi
Nasal chondromesenchymal hamartoma (NCMH) is an extremely rare benign tumor arising in the sinonasal tract, predominantly involving infants and children. To date, only 27 cases are reported in the international literature and there have been no reported cases of malignant transformation. We present a 40-yearold female patient with nasal obstruction and bloody rhinorrhea. Computed tomography (CT) of the nose and paranasal sinuses confirmed a heterogeneous polypoid soft-tissue mass filling the nasal cavity and extending into the maxillary and ethmoid sinus. The patient underwent a complete radical resection. Histological and immunohistochemical analyses showed a portion of the mass was consistent with typical NCMH. However, some areas of mass exhibited cytological atypia, marked mitotic activity and foci of necrosis. The atypical mesenchymal spindle cells were immunoreactive for vimentin, CD99 and smooth muscle actin (SMA) diffusely. The cartilaginous cells were immuno-positive for S-100 protein. Ki-67 index was high in atypical areas, accounting for 50%. A rapid mass recurrence was observed at the original site only 3 months after surgery. The final diagnosis of NCMH with malignant transformation was made. To our knowledge, this is the first report of malignant transformation occurring in an adult with NCMH. Although NCMH commonly develops in the neonate or young infants and exhibits benign histological appearance and favorable prognosis, there is a possibility of malignant transformation in adult patients. Thoroughly histological inspections are suggested to be necessary to accurately diagnose this tumor when it is encountered in adults
Open Access
Metastasis suppressor genes
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Yan, Jinchun; Yang, Qin; Huang, Qihong
Metastasis is a major cause of cancer mortality. Metastasis is a complex process that requires the regulation of both metastasis-promoting and metastasis suppressor genes. The discovery of metastasis suppressor genes contributes significantly to our understanding of metastasis mechanisms and provides prognostic markers and therapeutic targets in clinical cancer management. In this review, we summarize the methods that have been used to identify metastasis suppressors and the potential clinical impact of these genes.
Open Access
Postnatal neurogenesis in the cow pineal gland: an immunohistochemical study
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Gómez Esteban, M.B.; Muñoz Mosqueira, Mª.I.; Arroyo, A.A.; Muñoz Barragán, L.
In the pineal gland of cows and rats structures designated rosettes have been described both during embryonic development and in adult animals. In order to investigate the possible nature of the cells comprising such structures, in the present work we studied the pineal glands from 10 cows of one- or fouryears-old using conventional immunocytochemical and confocal microscopy techniques. As markers of glial cells, we used anti-vimentin (Vim) and glial fibrillary acidic protein (GFAP) and anti-S-100 sera, and the pinealocytes were labelled with ß-III tubulin. As a marker of stem cells, we used an antinestin serum, while an anti-PCNA serum was employed to label proliferating cells. To explore the neuronal nature of some cells of the rosettes, we used an anti-SRIF serum. The rosettes were seen to be present throughout the glandular parenchyma and displayed a central cavity surrounded by cells, most of which expressed all or just some of the above glial labels and nestin, although there were also some rosettes with cells that expressed ß-III tubulin and other cells that expressed SRIF. Likewise, in the cells of the rosettes the cell nucleus showed strong expression of PCNA. Confocal microscopy revealed that the walls of the rosettes contained cells that coexpressed Vim/S-100, Vim/GFAP and Vim/nestin. The number of rosettes was significantly greater in the animals of one year of age with respect to the four-year-old cows. The present findings allow us to suggest that rosettes are evolving structures and that most of the cells present in their walls should be considered stem cells, and hence responsible for the postnatal neurogenesis occurring in the pineal gland of cows.
Open Access
Neurobiological toxicity of radiation in hippocampal cells
(2013) Kim, Joong-Sun; Yang, Miyoung; Kim, Sung-Ho; Shin, Taekyun; Moon, Changjong
Worldwide radiation exposure is increasing due to recent nuclear accidents, space travel, atomic weapons testing and use, and medical treatments. In adult animals, ionizing radiation can significantly impact hippocampal neurogenesis and negatively affect hippocampal functions such as cognition. However, there is considerable uncertainty regarding the mechanisms underlying these effects. This article reviews in vivo and in vitro studies on the effects of irradiation on hippocampal neurogenesis and function in order to gain new mechanistic insights. This information will provide complementary views of our understanding of the normal brain’s tolerance to radiation exposure, the potentially serious implications of radiation exposure to cognition, and lead to a discussion of potential strategies for pharmacotherapy and behavioral intervention.

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