Histology and histopathology Vol.25, nº6 (2010)

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Open Access
Investigation of AGE, their receptor and NF-KB activation and apoptosis in patients with ATTR and Gelsolin amyloidosis
(Murcia : F. Hernández, 2010) Anan, Intissar; Kiuru-Enari, Sari; Obayashi, Konen; Ranløv, Poul Jørgen; Ando, Yukio
Background: Transthyretin (TTR) and gelsolin amyloidoses represent two types of hereditary amyloidosis in which point mutations in the respective protein lead to conformational changes of the protein with subsequent amyloid fibril formation. Material and methods: Tissues from Finnish gelsolin amyloid patients, Danish, Japanese and Swedish ATTR patients were immunostained for AGE, RAGE, NF-κB, PARP, and caspases 3 and 8. Results: Amyloid was heavily deposited in myocard, kidney and gastrointestinal tract of all patients. Immunoreactive areas to AGE and RAGE were detected in the heart, kidney, rectum, gut and appendix. AGE and RAGE were well co-localised with amyloid deposits. In five out of 14 patients neither NF-κB activation nor induction of apoptosis marked by positive immunostaining for NF-κB, PARP, or caspases 3 and 8 was found, and markers of apoptosis were detected in some samples without accompanying NF-κB activation. Conclusion: Our results suggest that both AGE and RAGE may have a common role in evolution of TTR and gelsolin-related amyloidoses. Apart from AGE-RAGE interactions both amyloid proteins may directly bind to RAGE and result in cellular perturbations; but in view of this study cytotoxic effects other than those triggered by activation of NF-κB or apoptosis should be considered
Open Access
Defining adipose tissue-derived stem cells in tissue and in culture
(Murcia: F. Hernández, 2010) Lin, Ching-Shwun; Xin, Zhong-Cheng; Deng, Chun-Hua; Ning, Hongxiu; Lin, Guiting; Lue, Tom F.
Adipose tissue-derived stem cells (ADSC) are routinely isolated from the stromal vascular fraction (SVF) of homogenized adipose tissue. Similar to other types of mesenchymal stem cells (MSC), ADSC remain difficult to define due to the lack of definitive cellular markers. Still, many types of MSC, including ADSC, have been shown to reside in a perivascular location, and increasing evidence shows that both MSC and ADSC may in fact be vascular stem cells (VSC). Locally, these cells differentiate into smooth muscle and endothelial cells that are assembled into newly formed blood vessels during angiogenesis and neovasculogenesis. Additionally, MSC or ADSC can also differentiate into tissue cells such as adipocytes in the adipose tissue. Systematically, MSC or ADSC are recruited to injury sites where they participate in the repair/regeneration of the injured tissue. Due to the vasculature’s dynamic capacity for growth and multipotential nature for diversification, VSC in tissue are individually at various stages and on different paths of differentiation. Therefore, when isolated and put in culture, these cells are expected to be heterogeneous in marker expression, renewal capacity, and differentiation potential. Although this heterogeneity of VSC does impose difficulties and cause confusions in basic science studies, its impact on the development of VSC as a therapeutic cell source has not been as apparent, as many preclinical and clinical trials have reported favorable outcomes. With this understanding, ADSC are generally defined as CD34+CD31- although loss of CD34 expression in culture is well documented. In adipose tissue, CD34 is localized to the intima and adventitia of blood vessels but not the media where cells expressing alpha-smooth muscle actin (SMA) exist. By excluding the intima, which contains the CD34+CD31+ endothelial cells, and the media, which contains the CD34-CD31- smooth muscle cells, it leaves the adventitia as the only possible location for the CD34+ ADSC. In the capillary, CD34 and CD140b (a pericyte marker) are mutually exclusively expressed, thus suggesting that pericytes are not the CD34+ ADSC. Many other cellular markers for vascular cells, stem cells, and stem cell niche have also been investigated as possible ADSC markers. Particularly the best-known MSC marker STRO-1 has been found either expressed or not expressed in cultured ADSC. In the adipose tissue, STRO-1 appears to be expressed exclusively in the endothelium of certain but not all blood vessels, and thus not associated with the CD34+ ADSC. In conclusion, we believe that ADSC exist as CD34+CD31-CD104b-SMA- cells in the capillary and in the adventitia of larger vessels. In the capillary these cells coexist with pericytes and endothelial cells, both of which are possibly progenies of ADSC (or more precisely VSC). In the larger vessels, these ADSC or VSC exist as specialized fibroblasts (having stem cell properties) in the adventitia.
Open Access
Microanatomy of lymphocyte-endothelial interactions at the high endothelial venules of lymph nodes
(Murcia: F. Hernández, 2010) Tohya, Kazuo; Umemoto, Eiji; Miyasaka, Masayuki
Lymphocyte trafficking into lymph nodes and Peyer’s patches is mediated primarily by specifically differentiated venules, called high endothelial venules (HEVs), located in the tissue parenchyma. HEVs have a unique morphology and phenotype, which enables them to interact with circulating lymphocytes efficiently. That is, the HEV endothelial cells have a tall and plump appearance, and constitutively express multiple adhesion molecules and chemokines on their surface. These molecules can interact with cognate receptors on circulating lymphocytes, thereby mediating the stepwise and sequential lymphocyte adhesion and transendothelial migration (TEM) at the HEV endothelial luminal surface. This review summarizes the fine morphological aspects of the unique HEV endothelial cells, with special reference to the spatial distribution of the adhesion molecules and chemokines that regulate lymphocyte migration.
Open Access
Oxidized regenerated cellulose does not prevent the formation of experimental postoperative perineural fibrosis assessed by digital analysis
(Murcia: F. Hernández, 2010) Hernández Cortés, Pedro; Peregrina, Magdalena; Aneiros Fernández, José; Tassi, Mohamed; Pajares López, Miguel; Toledo, Miguel; O’Valle, Francisco
Introduction: It is difficult to prevent and treat intra- and peri-neural fibrosis after peripheral nerve surgery. Many authors have attempted to develop and verify the effectiveness of substances to decrease the formation of adherences in different tissues. Material and Methods: this study aimed to assess the effectiveness of a barrier of oxidized regenerated cellulose (ORC) to reduce adherence and perineural fibrosis in a model of surgical perineural induced fibrosis in rat sciatic nerve in 40 rats. After tissue aggression, the nerve of the right rear limb was wrapped in ORC and the left limb served as control. Animals were killed at 3 and 6 weeks, and nerves and muscle mass were extracted en bloc. Connective tissue was quantified by conventional histopathological techniques and Fibrosis HR® automatic image analysis. Results: No significant differences were found in intra- or peri-neural induced fibrosis between control nerves (6.88% and 8.90%, respectively) and treated nerves (6.57% and 9.90%) at 3 or 6 weeks (10.41% and 12.51% in controls; 11.85% and 15.72% in treated nerves). Inflammatory phenomena and granulomatous reactions were more frequent in treated animals. Conclusions: ORC conferred no advantage in prevention of nerve fibrosis and might have interfered with healing.
Open Access
Dynorphin expression, processing and receptors in the alveolar macrophages, cancer cells and bronchial epithelium of lung cancer patients
(Murcia: F. Hernández, 2010) Mousa, Shaaban A.; Krajnik, Malgorzata; Sobanski, Piotr; Kowalewski, Janusz; Bloch-Boguslawska, Elzbieta; Zylicz, Zbigniew; Schafer, Michael
Functional evidence suggests that opioid peptides such as dynorphin are involved in the regulation of airway macrophage functions and of human cancer growth. However, anatomical evidence for components of a putative dynorphin network within lung cancer patients is scarce. Tissue from lung cancer patients was examined immunohistochemically for all components of a local dynorphin (DYN) network. Double immunofluorescence microscopy analysis revealed colocalization of the opioid precursor PDYN with its end-product DYN, and key processing enzymes prohormone convertases 1 and 2 and carboxypeptidase E, as well as the kappa-opioid receptor (KOR) within alveolar macrophages and cancerous cells in varying degrees among patients. Moreover, chromograninAimmunoreactive pulmonary neuroendocrine cells expressing DYN were close to substance P- and KORimmunoreactive sensory nerves. Our findings give a first hint of a neuroanatomical basis for a peripheral DYN network, conceivably regulating pulmonary, immune and cell-proliferative functions within the human lung, most likely in a paracrine/autocrine fashion.
Open Access
Podoplanin expression in tumor-free resection margins of oral squamous cell carcinomas: an immunohistochemical and fractal analysis study
(Murcia: F. Hernández, 2010) Margaritescu, C.; Raica, M.; Pirici, D.; Simionescu, C.; Mogoanta, L.; Stinga, A.C.; Stinga, A.S.; Ribatti, Doménico
Podoplanin is involved in tumorigenesis and cancer progression in head and neck malignancies and its expression is not restricted to lymphatic vessel endothelium. The aim of this study was to establish podoplanin expression in the tumor-free resection margins of oral squamous cell carcinomas (OSCCs) and to evaluate the geometric complexity of the lymphatic vessels in oral mucosa by utilizing fractal analysis. As concerns the podoplanin expression in noncancerous tissue, forty tumor-free resection margins from OSCCs were investigated utilizing immunohistochemistry for D2-40 antibody and image densitometry analysis. Podoplanin expression was extremely low in basal cells, especially in resection margins of OSCCs developed in the lower lip regions. However, a highly variable D2-40 expression in tumor-free resection margins associated with hyperplastic or dysplastic lesions was identified. Moreover, podoplanin expression also extended to the basal layer of the lower lip skin appendages, the myoepithelial cells of acini and ducts of minor salivary glands, and other structures from the oral cavity. As concerns the study of the density and complexity of oral lymphatic vessels architecture by means of immunohistochemistry (D2-40, CD31 and Ki-67 antibodies) and fractal analysis, we demonstrated that in normal oral mucosa the geometry of the lymphatic vessels was less complex at the level of the lower lip compared to the anterior part of the oral floor mucosa or the tongue. A comparative analysis between the normal and pathological aspects revealed statistically significant differences between the fractal dimension (FD) of the vessels’ outline, especially in the tongue. Fractal analysis proved an increasing lymphatic network complexity from normal to premalignant oral mucosal lesions, providing additional prognostic information in oral malignant tumors
Open Access
Diagnostic and prognostic value of T-cell receptor gamma alternative reading frame protein (TARP) expression in prostate cancer
(Murcia: F. Hernández, 2010) Fritzsche, Florian R.; Stephan, Carsten; Gerhardt, Josefine; Lein, Michael; Hofmann, Irina; Jung, Klaus; Dietel, Manfred; Kristiansen, Glen
T-cell receptor gamma chain alternative reading frame protein (TARP) has recently been proposed as being up-regulated in prostate cancer (PCA). Additionally, TARP has been proposed as a potential therapeutic target for cancer therapy. We analysed the protein expression of TARP in a large well characterised prostate cancer cohort to assess its diagnostic and prognostic value. Methodologically, we constructed a tissue microarray comprising more than 600 PCA cases including matching benign prostate tissue. TARP protein expression was carefully analysed and associated with clinico-pathological parameters, PSA-relapse free survival and expression data of established and proposed diagnostic markers (AMACR, p63, GOLPH2). Our results show that TARP is significantly over-expressed in the vast majority (~85%) of PCA in comparison to non neoplastic prostate tissue. Its expression was associated with conventional markers of unfavourable and more aggressive tumour behaviour. However, a prognostic value of TARP could not be found. The diagnostic value of TARP is limited in comparison to AMACR, p63 or GOLPH2. Since TARP specific immunologic therapy regimen are currently being tested, the high frequency of TARP overexpression in PCA conveys a high potential for a predictive and potentially therapeutic use of this biomarker.
Open Access
Treadmill training increases the size of A cells from the L5 dorsal root ganglia in diabetic rats
(Murcia : F. Hernández, 2010) do Nascimento, Patricia S.; Malysz, Tais; Ilha, Jocemar; Araujo, Rafaela T.; Hermel, Erica E.S.; Kalil-Gaspar, Pedro I.; Faccioni-Heuser, María C.; Schaan, Beatriz D.; Achaval, Matilde
The aim of this study was to evaluate the effects of physical training on the L5 dorsal root ganglion (DRG) cells in streptozotocin diabetic rats. Male adult rats were divided into 3 groups: (control, diabetic and trained diabetic). Treadmill training was performed for 10 weeks (5 days/week, twice a day). Blood glucose concentrations and body weight were evaluated 48h after diabetes induction and every 30 days thereafter. Then, animals were killed and the right L5 DRG removed. Histological and morphometric analysis consisted of evaluating nuclear and cellular volumes and areas in A and B cells at light and ultrastructural levels. Blood glucose concentrations were higher in both diabetic groups vs controls at all periods. Body weights were lower in all diabetics vs controls at all periods after diabetes induction, with a significant time vs group interaction. In A cells, the cellular and nuclear volumes were lower than control animals only in the diabetic group; control and trained diabetic animals did not differ; in B cells the cellular and nuclear volumes were lower in diabetic and trained diabetic rats. The cellular areas of A cells were smaller in diabetic rats than in control and trained diabetic rats, while the cellular areas of B cells were smaller in the diabetic and trained groups. In conclusion, treadmill training was able to increase the size of A cells from the DRG in diabetic rats and improved the morphological features of these cells
Open Access
Melanocytic matricoma. Report of a further case with clinicopathological and immunohistochemical findings, differential diagnosis and review of the literature
(Murcia: F. Hernández, 2010) Cartaginese, F.; Sidoni, Angelo
. Melanocytic matricoma is a rare recently described lesion. It usually presents as a pigmented, dark-papular, crusted lesion on sun-damaged skin of adult patients. Histopathologically, these lesions are characterized by well-circumscribed nodules composed of matrical and supramatrical cells with clustered shadow cells, and admixed pigmented dendritic melanocytes. It differs from matricomas and pilomatricomas by its lack of calcification, cyst formation, granulomas, and connections to the epidermis and other adnexal structures. The clinical differential diagnosis includes hemangioma, pigmented basal cell carcinoma, and melanoma. Melanocytic matricoma presumably is the representation of an epithelialmelanocytic interaction in the anagen phase of the hair cycle. An extensive search of the medical literature revealed 11 reports of benign melanocytic matricomas and 5 malignant counterparts. We report an additional case of melanocytic matricoma with discussion of clinicopathologic features and differential diagnosis.
Open Access
Role of neutrophil-derived matrix metalloproteinase-9 in tissue regeneration
(Murcia: F. Hernández, 2010) Heissig, Beate; Nishida, Chiemi; Tashiro, Yoshihiko; Sato, Yayoi; Ishihara, Makoto; Ohki, Makiko; Gritli, Ismael; Rosenkvist, Jeanette; Hattori, Koichi
Ischemic tissue regeneration depends on neovascularization, the growth of new blood vessels. Bone marrow (BM)-derived cells, including neutrophils, have been shown to contribute to neovascularization during hind limb ischemia and inflammation. Neutrophils produce a broad array of angiogenic growth factors and proteases, which promote remodeling of arterioles into arteries through proteolytic mechanisms. Matrix metalloproteinases (MMPs) have been shown to play a role in the recruitment of neutrophils to sites of inflammation, which requires the extravascular migration of neutrophils through the extracellular matrix. Neutrophils control critical steps during angiogenesis and neutrophil-derived MMPs can promote neoangiogenesis, and collateral growth and perfusion recovery, in part by liberating vital angiogenic growth factors, including vascular endothelial growth factor-A (VEGF-A). This review focuses on the role of neutrophils as key players in the control of the angiogenic process during ischemic tissue regeneration. Aspects of neutrophil regulation, in particular regulation by its major growth factor granulocyte colonystimulating factor (G-CSF), the role of the unique, readily available, neutrophil-derived MMP-9, and the functional consequences of this MMP-9 activation for angiogenesis, such as MMP-mediated release of biologically relevant cytokines from the matrix and cell surfaces, will be discussed.
Open Access
Differential tissue expression of enhanced green fluorescent protein in ‘Green mice’
(Murcia: F. Hernández, 2010) Ma, De-Fu; Tezuka, Hideo; Kondo, Tetsuo; Sudo, Katsuko; Niu, Dong-Feng; Nakazawa, Tadao; Kawasaki, Tomonori; Yamane, Tetsu; Nakamura, Nobuki; Katoh, Ryohei
In order to clarify tissue expression of enhanced green fluorescent protein (EGFP) in ‘green mice’ from a transgenic line having an EGFP cDNA under the control of a chicken beta-actin promoter and cytomegalovirus enhancer, we studied the expression of EGFP in various organs and tissues from these ‘green mice’ by immunohistochemistry with anti- EGFP antibody in conjunction with direct observation for EGFP fluorescence using confocal laser scanning microscopy. On immunohistochemical examination and on direct observation by confocal laser scanning microscopy, the level of EGFP expression varied among organs and tissues. EGFP expression was diffusely and strongly observed in the skin, pituitary, thyroid gland, parathyroid gland, heart, gall bladder, pancreas, adrenals and urinary bladder. There was only sporadic and weak expression of EGFP in the epithelium of the trachea, bronchus of the lung, stratified squamous epithelium and gastric glands of the stomach, hepatic bile ducts of the liver, glomeruli and renal tubules of the kidney and endometrial glands of the uterus. Furthermore, EGFP was only demonstrated within the goblet and paneth cells in the colon and small intestine, the tall columnar cells in the ductus epididymis, and the leydig cells in the testis. In conclusion, our results show that EGFP is differentially expressed in organs and tissues of ‘green mice’, which indicates that ‘green mice’ may prove useful for research involving transplantation and tissue clonality.
Open Access
Neurotrophic keratopathy; its pathophysiology and treatment
(Murcia: F. Hernández, 2010) Okada, Yuka; Reinach, Peter S.; Kitano, Ai; Shirai, Kumi; Kao, Winston W.-Y.; Saika, Shizuya
Neurotropic keratopathy is one of the major refractory corneal disorders, which was first recognized in 1824. This category of diseases is caused by damage to the trigeminal nerve and the consequent loss of corneal sensation. It leads to various types of corneal disorder, including superficial keratopathy, persistent epithelial defects and corneal ulcers. In the present article, we review the pathobiology and prevention/ treatment of neurotrophic corneal diseases. The goals of treatment of neurotrophic keratopathy are to prevent the progression of corneal damage to promote epithelial healing in case that the original damage in the trigeminal nerve or its braches is difficult. The therapy should be prompt and based on the clinical stage of the disease. Although at present, there are no accurate treatment methods for neurotrophic keratopathy, several effective treatments have been reported. Signals derived from TRP channels are involved in homeostasis of tissues by controlling cell behaviors, i. e., gene expression for inflammation, cell migration, or cell survival/cell death. Targeting TRP channel in the cornea is expected to contribute to the development of a new treatment method for neurotrophic keratopathy. Further study and clinical trial are need to establish this new treatment strategy
Open Access
The CCN proteins: important signaling mediators in stem cell differentiation and tumorigenesis
(Murcia: F. Hernández, 2010) Zuo, Guo-Wei; Kohls, Christopher D.; He, Bai-Cheng; Chen, Liang; Zhang, Wenli; Shi, Qiong; Zhang, Bing-Qiang; Kang, Quan; Luo, Jinyong; Luo, Xiaoji; Wagner, Eric R.; Kim, Stephanie H.; Restegar, Farbod; Haydon, Rex C.; Deng, Zhong-Liang; Luu, Hue H.; He, Tong-Chuan; Luo, Qing
The CCN proteins contain six members, namely CCN1 to CCN6, which are small secreted cysteine-rich proteins. The CCN proteins are modular proteins, containing up to four functional domains. Many of the CCN members are induced by growth factors, cytokines, or cellular stress. The CCNs show a wide and highly variable expression pattern in adult and in embryonic tissues. The CCN proteins can integrate and modulate the signals of integrins, BMPs, VEGF, Wnts, and Notch. The involvement of integrins in mediating CCN signaling may provide diverse contextdependent responses in distinct cell types. CCN1 and CCN2 play an important role in development, angiogenesis and cell adhesion, whereas CCN3 is critical to skeletal and cardiac development. CCN4, CCN5 and CCN6 usually inhibit cell growth. Mutations of Ccn6 are associated with the progressive pseudorheumatoid dysplasia and spondyloepiphyseal dysplasia tarda. In stem cell differentiation, CCN1, CCN2, and CCN3 play a principal role in osteogenesis, chondrogenesis, and angiogenesis. Elevated expression of CCN1 is associated with more aggressive phenotypes of human cancer, while the roles of CCN2 and CCN3 in tumorigenesis are tumor type-dependent. CCN4, CCN5 and CCN6 function as tumor suppressors. Although CCN proteins may play important roles in fine-tuning other major signaling pathways, the precise function and mechanism of action of these proteins remain undefined. Understanding of the biological functions of the CCN proteins would not only provide insight into their roles in numerous cellular processes but also offer opportunities for developing therapeutics by targeting CCN functions.

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