Histology and histopathology Vol.33,nº10 (2018)

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Open Access
Sulfur dioxide ameliorates rat myocardial fibrosis by inhibiting endoplasmic reticulum stress
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Wang, Xin Bao; Cui, Hong; Du, Jun Bao
Myocardial remodeling occurs after myocardial infarction (MI), the leading cause of mortality worldwide. Although myocardial fibrosis plays an important role in the process of myocardial remodeling, there is not yet an effective method of reducing it. The aim of the present study was to determine the effects of sulfur dioxide (SO2) on myocardial fibrosis and the possible mechanisms of these effects. SO2 treatment reduced the extent of myocardial fibrosis and post-MI levels of collagens I and III in the left-ventricular myocardium. SO2 also improved MI-induced thinning of the left ventricular wall while enlarging the left ventricular internal diameter. SO2 was able to reduce matrix metalloproteinase (MMP)-9 activity and increase tissue matrix metalloproteinase inhibitor (TIMP)-1 content in myocardium after MI. However, the mechanism underlying these effects of SO2 on myocardial fibrosis are unknown. Western blot analysis of endoplasmic reticulum (ER) stress-related proteins showed that glucose-regulated protein 78, C/EBP homologous protein, caspase-12, and phosphorylated eukaryotic initiation factor 2α expression levels were significantly increased in MI rats and decreased by SO2 treatment. The ER stress promoter dithiothreitol reversed these effects of SO2. In conclusion, SO2 alleviated myocardial fibrosis in MI rats through a mechanism related to inhibition of excessive ER stress.
Open Access
Histomorphometric analysis with a proposed tissue lesion index in ischemia-reperfusion induced gastric mucosa damage
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Peña Mercado, Eduardo; Garcia Lorenzana, Mario; Beltran, Nohra E.
Damage to the gastrointestinal mucosa caused by ischemia - reperfusion is a significant clinical problem associated with various physiopathological conditions. Our group has conducted various studies in patients in critical conditions and in animal models to identify early damage to the gastric mucosa under ischemia using impedance spectroscopy. It is important to perform a quantitative histopathological analysis which can be linked to changes in impedance of the gastric mucosa under conditions of ischemia and I/R. Aim. To propose a tissue lesion index which considers pathological alterations inherent to the inflammatory process and cell damage which may be directly related to changes in impedance under conditions of ischemia and I/R. Methods. The animals were randomly distributed into 4 groups: control, ischemia (30 min), and I/R (30 and 60 min). Qualitative histopathological analysis was performed; the vascular area, glandular lumen area, the number of damaged cells, and the depth of the erosion were also quantified to obtain a scale to propose a tissue lesion index (TLI). Results. Under ischemic conditions, histopathological analysis showed edema and necrosis in epithelial cells, and vascular congestion. In I/R (30 and 60 min) conditions, areas of epithelial erosion were generated. Damage was classified based on the TLI. A TLI threshold of 3 showed a predictive value of tissue lesion. Conclusion. The proposed gastric lesion index allows us to objectively quantify and classify damage to the gastric mucosa produced by I/R.
Open Access
Identification of protein kinase C α- and tyrosine hydroxylase-immunoreactive cells in the microbat retina
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Park, Eun Bee; Jeon, Joo Yeong; Jeon, Chang Jin
A growing number of studies have revealed the functional neuroarchitecture of the microbat retina and suggested that microbats can see using their eyes. To better understand the organization of the microbat retina, quantitative analysis of protein kinase C alpha (PKCα)- and tyrosine hydroxylase (TH)-immunoreactive (IR) cells was conducted on the greater horseshoe bat (Rhinolophus ferrumequinum) retina. As a result, PKCα immunoreactivity was observed in rod bipolar cells, consistent with previous studies on other mammalian retinas. PKCα-IR cell distribution in the inner nuclear layer showed regional differences in density, with the highest density found in the nasal retina. The average density of PKCα-IR cells was 10,487±441 cells/mm2 (mean ± SD; n=4), with a total of 43,077±1,843 cells/retina. TH-IR cells in the Rhinolophus ferrumequinum retina could be classified into four types based on soma location and ramification in the inner plexiform layer: conventional amacrine, displaced amacrine, interplexiform, and intercalated cells. The majority of TH-IR cells were conventional amacrine cells. TH-IR cells were nonrandomly distributed at low density over the retina. The average density was 29.7±3.1 cells/mm2 (mean ± SD; n=3), with a total of 124.0±11.3 cells/retina. TH-IR processes showed varicosities and formed ring-like structures encircling AII amacrine cells. Our study provides the foundation for understanding the neurochemical architecture of the microbat retina and supports the notion that the eyes do play a role in the visual system of microbats.
Open Access
In search for a gold-standard procedure to count motor neurons in the spinal cord
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Ferrucci, Michela; Lazzeri, Gloria; Flaibani, Marina; Biagioni, Francesca; Cantini, Federica; Madonna, Michele; Bucci, Domenico; Limanaqi, Fiona; Soldani, Paola; Fornai, Francesco
Counting motor neurons within the spinal cord and brainstem represents a seminal step to comprehend the anatomy and physiology of the final common pathway sourcing from the CNS. Motor neuron loss allows to assess the severity of motor neuron disorders while providing a tool to assess disease modifying effects. Counting motor neurons at first implies gold standard identification methods. In fact, motor neurons may occur within mixed nuclei housing a considerable amount of neurons other than motor neurons. In the present review, we analyse various approaches to count motor neurons emphasizing both the benefits and bias of each protocol. A special emphasis is placed on discussing automated stereology. When automated stereology does not take into account sitespecificity and does not distinguish between heterogeneous neuronal populations, it may confound data making such a procedure a sort of “guide for the perplex”. Thus, if on the one hand automated stereology improves our ability to quantify neuronal populations, it may also hide false positives/negatives in neuronal counts. For instance, classic staining for antigens such as SMI-32, SMN and ChAT, which are routinely considered to be specific for motor neurons, may also occur in other neuronal types of the spinal cord. Even site specificity within Lamina IX may be misleading due to neuronal populations having a size and shape typical of motor neurons. This is the case of spinal border cells, which often surpass the border of Lamina VII and intermingle with motor neurons of Lamina IX. The present article discusses the need to join automated stereology with a dedicated knowledge of each specific neuroanatomical setting.
Open Access
Upregulation of autophagy and glycolysis markers in keloid hypoxic-zone fibroblasts: Morphological characteristics and implications
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Okuno, Ryoko; Ito, Yuko; Eid, Nabil; Otsuki, Yoshinori; Kondo, Yoichi; Ueda, Koichi
Keloid is a fibro-proliferative skin disorder with tumor-like behavior and dependence on anaerobic glycolysis (the Warburg effect), but its exact pathogenesis is unknown. Although autophagy is widely accepted as a lysosomal pathway for cell survival and cellular homeostasis (specifically upon exposure to stressors such as hypoxia), very few studies have investigated the involvement of autophagy and related glycolytic effectors in keloidogenesis. Here the authors examined the expression and cellular localization of autophagy proteins (LC3, pan-cathepsin), glycolytic markers (LDH, MCT1, MCT4) and the transcription factor HIF isoforms in human keloid samples using immunohistochemical analysis and double-labeling immunofluorescence methods. Based on H&E staining and expression of CD31, keloids were compartmentalized into hypoxic central and normoxic marginal zones. Vimentin-expressing fibroblasts in the central zone exhibited greater autophagy than their marginalzone counterparts, as evidenced by increased LC3 puncta formation and co-localization with lysosomal pan-cathepsin. LDH (a lactate stimulator), MCT4 (a lactate exporter) and HIF-1α expression levels were also higher in central-zone fibroblasts. Conversely, HIF-2α expression was upregulated in fibroblasts and endothelial cells of the peripheral zone, while MCT1 was expressed in both zones. Taken together, these observations suggest that upregulation of autophagy and glycolysis markers in keloid hypoxic-zone fibroblasts may indicate a prosurvival mechanism allowing the extrusion of lactate to marginal-zone fibroblasts via metabolic coupling. The authors believe this is the first report on differential expression of autophagic and glycolytic markers in keloid-zone fibroblasts. The study results indicate that autophagy inhibitors and MCT4 blockers may have therapeutic implications in keloid treatment
Open Access
The expression of CD markers in solid tumors: Significance in metastasis and prognostic value
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Rezaeeyan, Hadi; Shahrabi, Saeid; McKee, Trevor D.; Saki, Najmaldin
Objective. The clusters of differentiations (CDs) are among the surface markers expressed on different cells in the body, which are involved in the communication of cells with each other and the induction of signaling. Moreover, the evaluation of the ectopic expression of these markers in solid tumors has led to the detection of disease in early stages. In this paper, we have examined the effect of CD markers expression on the function of cancer cells, as well as their importance as the diagnostic and prognostic factors for monitoring the progression of solid tumors. Materials and methods Relevant literature was identified by a PubMed search (1988-2017) of English the language papers using the terms “CD markers”, “diagnostic”, “prognostic”, “predictive marker” and “solid tumors.” Discussion. Finally, it can be stated that the evaluation of CDs is not only of diagnostic value at disease onset, but these markers can be used as prognostic and predictive markers to contribute to the treatment of disease and predict its relapse. Conclusion. Monitoring of tumors progression through CDs expressed on circulating tumor cells could be a new diagnostic and prognostic factor in the future.
Open Access
Butylated hydroxytoluene induces type-V collagen and overexpression of remodeling genes/proteins in experimental lung fibrosis
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Martins, Vanessa; Teodoro, Walcy Rosolia; Pereira Velosa, Ana Paula; Andrade, Priscila; Farhat, Cecília; Fabro, Alexandre Todorovic; Capelozzi, Vera Luiza
Anomalous histoarchitecture with increased levels of type-V collagen (Col V) in lungs of human idiopathic pulmonary fibrosis (IPF) and bleomycin (BLM) airway-centered interstitial fibrosis suggest that this collagen can be a possible trigger involved in the pathogenesis of these diseases. Butylated hydroxytoluene (BHT) injury model revealed a distal involvement of lung parenchyma with significant endothelial injury and fibrotic response, contrasting with the BLM airway-centered insult. We undertook this study to analyze whether BHT alters distal airway/alveolar epithelial cells (AECs) and extracellular matrix (ECM) signaling involved in the initiation and progression of pulmonary fibrosis in a different pathway concerning overexpression of Col V. Female mice C57BL/6 (n=6) were instilled intraperitoneally with 400mg/kg of BHT dissolved in 1 mL of corn oil and euthanized at day 14 or 21 after BHT administration. Morphometry, immunohistochemistry and transmission electron microscopy were performed to characterize microscopic and submicroscopic changes of AECs and endothelial cells through transforming growth factor beta (TGF-β) basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) expression. Immunofluorescence and immunogold electron microscopy were performed to characterize Col V. Quantitative polymerase chain reaction (qPCR) was used to confirm differential levels of RNA messenger. BHT lungs showed marked fibrotic areas and hyperplastic AECs. The alveolar damage caused destruction of elastic fibers and a critical increase of Col V in ECM of distal lung parenchyma. Fibrogenesis-promoting markers TGF-β, bFGF and VEGF were also overexpressed in situ, coinciding with up-regulation in remodeling enzymes, growth factors, cytokines, transduction and transcription genes. BHT alters distal lung parenchyma signaling involved in pulmonary fibrosis highlighted similarities to human IPF in a pathway involving Col V arising as a promissory model to identify effective therapeutic targets.
Open Access
Claudin-1 role in colon cancer: An update and a review
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Ouban, Abderrahman
Tight junction proteins are essential for sealing the cellular sheets and controlling para-cellular ion flux. Our understanding of the role that tight junction proteins, particularly claudins, play in cellular functions and pathologic conditions is continuously expanding. Particularly, the role of claudin-1 in oncogenesis in multiple locations in the human body is coming to light. This review will shed light on the role of claudin-1 in colon cancer. It will address the mechanisms through which claudin-1 becomes dysregulated in colon cancer. This will provide a platform to address results of claudin-1 expression in the third most common malignant tumour worldwide. Furthermore, it will provide updates about possible use of this biomarker in the surveillance of difficult colon maladies, such as inflammatory bowel disease. The use of claudin-1 as a biomarker of diagnostic and prognostic values will provide Medicine with much needed ammunition in the fight against cancer and will bring about, with added refinements, a new chapter in the era of personalized medicine to tackle this disease and match its destructive course with equally powerful and specifically targeted therapies.
Open Access
Quantification of the heterogeneity of cytokeratin 18 immunoexpression in prostate adenocarcinoma and normal prostate: Global and local features
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2018) Santamaría, Luis; Ingelmo, Ildefonso; Sinues, Bryan; Martínez, Laura; Teba, Fernando
There are few studies comparing global versus local changes in spatial patterns in prostate cancer. In this study, stereological tools have been applied to find out if the cytokeratin18 (ck18) immunoexpression shows local changes in cancer compared to normal prostate. To verify if these changes are relevant to ascertain differences between normal (CTR) and cancer (Ca) cases, several parameters were estimated. Volume fraction of epithelium immunostained for ck18 (VV ck18), dispersion index of VV ck18, positional variance of VV ck18, and multiscale entropy analysis (MSE) to measure the tissue heterogeneity. The MSE values showing significant differences between CTR and Ca were employed in a discriminant analysis to determine if MSE was able to classify the cases in CTR and Ca groups. The findings obtained indicate that changes in the expression of ck18 by the cancer prostate are heterogeneous. The increase in local variability of ck18 immunoexpression can be related to the increase in heterogeneity of shape and size of the tumor acini. The asymmetry of distribution of the local values of VV ck18 along the axis of the space series may indicate the existence of anisotropy in the distribution of tumor acini. The increase in scale-dependent entropy for VV ck18 in cancer at the morphological level could be interpreted as the macroscopic expression of the same increase at the molecular level already described. The discriminant analysis shows that the dependence on the resolution for MSE values need to be taken into account to characterize the prostate cancer better

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