Histology and histopathology Vol.28, nº10 (2013)
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- PublicationOpen AccessDifferences in collagen distribution of healthy and regenerated periodontium. Histomorphometric study in dogs(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Souza, Sérgio Luis Scombatti de; Macedo, Guilherme O.; Silveira e Souza, Adriana M.M.; Taba Jr, Mário; Novaes Jr, Arthur B.; Oliver, Constance; Jamur, Maria C.; Correa, Vani M.A.Previous studies have shown that there is a relationship between periodontal disease and the distribution of collagen fibers. This study evaluated the distribution of collagen types I and III in regenerated bone and periodontal ligament, comparing them to the tissues near the regenerated area and to the healthy periodontium. In the third (P3) and fourth (P4) mandibular premolars of 5 healthy mongrel dogs, bilaterally, buccal class 2 furcation lesions were surgically created and chronified for 3 weeks. After that, full flaps were elevated and expanded polytetrafluoroethylene (e-PTFE) membranes were adapted, sutured and recovered by the flaps. Two weeks after surgery, two membranes on the same side were removed and the other membranes were removed four weeks after surgery. The dogs were euthanized at 12 weeks following placement of the e-PTFE membranes. P3 and P4 teeth as well as the second premolars (healthy control teeth) and their periodontal tissues were removed and histologically processed for Collagen Quantification (COLQ). The amount of type III collagen was higher in native bone compared to the regenerated area. For periodontal ligament, COLQ for type I collagen showed statistically significant differences (Tukeys’s Multiple Comparison, p<0.05) between the regenerated groups and the control group. These differences were not found for type III COLQ. There are significant differences in collagen distribution among the regenerated, native and control tissues. Membrane removal 2 or 4 weeks postoperatively did not influence the collagen composition.
- PublicationOpen AccessUmbilical cord revisited: from Wharton’s jelly myofibroblasts to mesenchymal stem cells(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Corrao, Simona; La Rocca, Giampiero; Lo Iacono, Melania; Corsello, Tiziana; Farina, Felicia; Anzalone, RitaThe umbilical cord (UC) is an essential part of the placenta, contributing to foetal development by ensuring the blood flow between mother and foetus. The UC is formed within the first weeks of gestation by the enclosure of the vessels (one vein and two arteries) into a bulk of mucous connective tissue, named Wharton’s jelly (WJ) and lined by the umbilical epithelium. Since their first identification, cells populating WJ were described as unusual fibroblasts (or myofibroblasts). Recent literature data further highlighted the functional interconnection between UC and the resident cells. The UC represents a reservoir of progenitor populations which are collectively grouped into MSCs (mesenchymal stem cells). Such cells have been sourced from each component of the cord, namely the subamnion layer, the WJ, the perivascular region, and the vessels. These cells mainly show adherence to the phenotype of adult MSCs (as bone marrow-derived ones) and can differentiate towards mature cell types belonging to all the three germ layers. In addition, cells from human UC are derived from an immunoprivileged organ, namely the placenta: in fact, its development and function depend on the elusion of the maternal immune response towards the semi-allogeneic embryo. This is reflected in the expression of immunomodulatory molecules by UC-derived MSCs. The present paper describes UC structural features and the cell types which can be derived, with a focus on their phenotype and the novel results which boosted the use of UC-derived cells for regenerative medicine applications.
- PublicationOpen AccessHistomorphometric and immunohistochemical study of the goat reticulum during prenatal development(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Garcia, A.; Masot, Javier; Franco, Antonio; Gazquez, Antonio; Redondo, E.This study sought to describe the morphological changes taking place in the goat reticulum during prenatal development, using histomorphometric and immunohistochemical techniques. A total of 140 goat embryos and foetuses were used, from the first stages of prenatal life until birth. Differentiation of the reticulum as a separate compartment of the primitive gastric tube was observed at 35 days of prenatal life (23% gestation). By 38 days (25% gestation) the reticular wall comprised three layers: an internal epithelial layer, a middle layer of pluripotential blastemic tissue and an external layer or serosa. Primary reticular crests were visible at 59 days (38% gestation) as evaginations of the epithelial stratum basale, marking the earliest histological differentiation of future reticular cells. Secondary reticular crests were observed at 87 days (61% gestation). Corneum papillae first became apparent on the lateral surface of primary reticular crests at 101 days (64% gestation). The muscularis mucosae was visible by 101 days (64% gestation) in primary reticular crests. Neuroendocrine cells were detected by synaptophysin at 64 days (43% gestation), while glial cell markers (glial fibrillary acidic protein and vimentin) were observed at 64 days (43% gestation) and 38 days (25% gestation), respectively. The peptidergic innervation markers such as neuropeptide Y and vasoactive intestinal polypeptide were detected at 75 days (50% gestation). In conclusion, prenatal development of the reticulum - like that of the rumen - appears to take place somewhat earlier in goats than in sheep or cattle, but at a similar rate to that reported in deer.
- PublicationOpen AccessA comprehensive morphometric analysis of the internal thoracic artery with emphasis on age, gender and left-to-right specific differences(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Labudović Borović, Milica; Borović, Saša; Marinković-Erić, Jelena; Todorović, Vera; Puškaš, Nela; Kočica, Mladen; Radak, Ðorde; Lačković, VesnaAims of the study. The aim of this analysis was the morphometric description of the internal thoracic artery (ITA) with an emphasis on age, gender and left-to-right specific differences, as well as on age and atherosclerosis related changes of the elastic skeleton. Methods. Forty eight arteries were obtained during forensic autopsies from 32 persons who had died of non-vascular causes. The following morphometric parameters were analyzed: thickness of the intima, the medial layer and the wall, the intima-to-media ratio and the elastic skeleton parameters. Results. The intima thickness increases significantly with aging (ANOVA F=34.061, p<0.001), as does the intima-to-media ratio (ANOVA F=10.831, p<0.001). With aging, there is a significant increase in the thickness of the media (F=56.519; p<0.001) and of the wall (F=34.094; p<0,001). There is a significant increase in the media thickness during the development of atherosclerosis in the ITA (ANOVA F=11.848, p<0.001). No significant difference was found when these data were analyzed based on the left-to-right principle or depending on gender of the patients. However, the analysis of the elastic skeleton parameters indicated that the combined effects of aging, atherosclerosis and male gender lead to the degeneration of the elastic skeleton of the ITA. Conclusion. The grade of atherosclerosis gradually increases with aging as shown by morphometric analysis. The increase in the medial layer thickness suggests the potential for positive remodeling of the ITA during aging and atherosclerosis. The left/right position has no influence on morphometric parameters of the ITA, while male gender affects parameters of the elastic skeleton.
- PublicationOpen AccessSynergism of imatinib mesylate and everolimus in attenuation of bronchiolitis obliterans after rat LTX(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Suesskind-Schwendi, M. von; Valenti, Verena; Haneya, Assad; Pühler, T.; Bewig, B.; Schmid, C.; Hirt, S.W.; Lehle, K.Bronchiolitis obliterans (BO) is a progressive and fatal disease after lung transplantation (LTX). Dysregulated growth factor-induced proliferation of myofibroblasts seems to be responsible for the development of BO. The aim was to confirm the efficacy of both inhibitors of receptor tyrosine kinases (RTKI) and of mammalian target of rapamycin (mTORI) after rat LTX. We used a rat model of left lung allotransplantation (F344-to-WKY) to evaluate the effect of imatinib (RTKI; 20 mg/kg/day; postoperative day (POD) 0-100) alone or in combination with everolimus (mTORI; 2.5 mg/kg/day; POD 14-100). Non-treated animals were the reference. In non-treated rats, acute rejection (AR) peaked between POD 20 and 30 (19/19) and ended in chronic rejection (CR) on POD 60/100 (12/12). Imatinib alone did not prevent AR (6/6), but attenuated the degree of degenerated bronchioles on POD 30 (non-treated, 57%; imatinib, 4%), and increased the allografts free of CR on POD 60/100 (3/12). A combination of imatinib and everolimus significantly reduced AR, attenuated fibrotic degenerated bronchioles (5%) and vessels (non-treated, 24%; combination therapy, 11%) on POD 30, and reduced fibrotic degenerated vessels (non-treated, 97%; combination therapy, 43%) and bronchioles (non-treated, 88%; combination therapy, 34%) on POD 60/100. Fifty percent of the animals were completely free of BO and vasculopathy. In conclusion, co-application of RTKI and mTORI attenuated the development of BO and vasculopathy. Thus, imatinib might be an interesting therapeutic approach after LTX.
- PublicationOpen AccessHistological approach to Bacillus subtilis colony-biofilm: evolving internal architecture and sporulation dynamics(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Gómez-Aguado, Fernando; Corcuera, María Teresa; Gómez-Lus, María Luisa; de la Parte, María Antonia; Ramos, Carmen; García-Rey, César; Alonso, María José; Prieto, JoséBacillus subtilis has been used as a classic model to study biofilm formation and sporulation process. Colonies of wild-type strains usually have a complex external morphology, but the details of their internal architecture are still undisclosed. Since bacterial biofilms fulfill the criteria to be considered tissues, the aim of this work was to analyse B. subtilis colony-biofilm internal architecture evolution and sporulation dynamics using histological techniques. Transversal sections of colony-biofilms incubated from 24 hours up to 20 days were stained using histochemical techniques to analyse the internal structure by light and electron microscopy. A morphometric study of the different structural biofilm components was performed by image analysis, and an application to quantify spores was developed. Internal biofilm architecture was characterised by a stratified pattern, which evolved from 3 strata at 24 hours, up to 5 strata at 20 days. At 48 hours, strata at the central area of the biofilm was folded, resulting in elevated structures (vein-like structures) that could reach up to 465 µm in height. Sporulation started at 48 hours, at the top of the vein-like structures, at the interface between the two uppermost strata. At 20 days spores formed a continuous central layer, representing 7.5% of the total biofilm. In summary, our results demonstrate that B. subtilis colony-biofilm has a complex and organized internal architecture, evolving over time, and taking place in different cell subpopulations with different functionalities. Furthermore, in situ spore quantification described in this work could be a good alternative to the classical chamber counting.
- PublicationOpen AccessRenal hybrid oncocytic/chromophobe tumors. A review(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Hes, Ondrej; Petersson, Fredrik; Kuroda, Naoto; Hora, Milan; Michal, MichalHybrid oncocytic/chromophobe tumors (HOCT) occur in three clinico-pathologic situations; (1) sporadically, (2) in association with renal oncocytomatosis and (3) in patients with Birt-HoggDubé syndrome (BHD). There are no specific clinical symptoms in patients with sporadic or HOCT associated with oncocytosis/oncocytomatosis. HOCT in patients with BHD are usually encountered on characteristic BHD clinicopathologic background. Sporadic HOCT are composed of neoplastic cells with eosinophilic oncocytic cytoplasm. Tumors are usually arranged in a solidalveolar pattern. Some neoplastic cells may have a perinuclear halo, with no raisinoid nuclei present. HOCT occurring in patients with oncocytomatosis are morphologically identical to sporadic HOCT. HOCT in BHD frequently display 3 morphologic patterns, either in isolation or in combination; (1) An admixture of areas typical of RO and CHRCC, respectively, (2) Scattered chromophobe cells in the background of a typical RO, (3) Large eosinophilic cells with intracytoplasmic vacuoles. The immunohistochemical profiles of HOCT in all clinicopathologic and morphologic groups differ slightly. The majority of tumors express parvalbumin, antimitochondrial antigen and CK 7. CD117 is invariably positive. HOCT show significant molecular genetic heterogeneity. The highest degree of variability in numerical chromosomal changes is present in sporadic HOCT. HOCT in the setting of oncocytomatosis have revealed a lesser degree of variability in the chromosomal numerical aberrations. HOCT in patients with BHD display FLCN gene mutations, which are absent in the other groups. HOCT (all three clinicopathologic groups) seem to behave indolently, as no evidence of aggressive behavior has been documented. However, no report with follow up longer than 10 years has been published.
- PublicationOpen AccessEndoplasmic reticulum resident heat shock protein-gp96 as morphogenetic and immunoregulatory factor in syngeneic pregnancy(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Jakovac, Hrvoje; Grebić, Damir; Grubić-Kezele, Tanja; Mrakovčić-Šutić, Ines; Rukavina, Daniel; Radošević-Stašić, BiserkaThe severe remodeling of endometrial stroma during blastocyst adhesion and trophoblast invasion initiates at maternal-fetal interface the reaction of evolutionary old heat shock response, in which heat shock proteins, as molecular chaperons, monitor the configurations of newly synthesized proteins and prevent the formation of functionless aggregates of misfolded proteins, targeting them to degradation by a the ubiquitin-proteasome system. In addition, the endoplasmic reticulum (ER)-resident HSPs, such as gp96/GRP94 may, after binding to CD91 and TLRs, elicit antigen-specific and antigen-unspecific immune responses, owing to its peptide-chaperoning capacity and ability to activate APCs. Considering these properties, we examined tissue expression of gp96 at the maternal-fetal interface and in the maternal liver and spleen on the 16th day of undisturbed syngeneic pregnancy and after the treatment with peptidoglycan monomer linked with zinc (PGMZn). The data showed that in undisturbed pregnancy the gp96, CD91 and TLR2 were markedly expressed on extravillous and villous trophoblast. PGM-Zn enhanced these findings, as well as the number of uterine natural killer cells and local NFκB immunoreactivity. Gp96 expression arose also in the maternal spleen and liver, where an accumulation of NKT cells or γδT lymphocytes was seen. The data point to roles of gp96 in maintenance of proteostasis and local and systemic immune balance in pregnancy complicated by infection.
- PublicationOpen AccessThe effect of low and high plasma levels of insulin-like growth factor-1 (IGF-1) on the morphology of major organs: studies of Laron dwarf and bovine growth hormone transgenic (bGHTg) mice(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Piotrowska, Katarzyna; Borkowska, Sylwia J.; Wiszniewska, Barbara; Laszczyńska, Maria; Słuczanowska-Głąbowska, Sylwia; Havens, Aaron M.; Kopchick, John J.; Bartke, Andrzej; Taichman, Russel S.; Kucia, Magda; Ratajczak, Mariusz Z.It is well known that somatotrophic/insulin signaling affects lifespan in experimental animals. To study the effects of insulin-like growth factor-1 (IGF-1) plasma level on the morphology of major organs, we analyzed lung, heart, liver, kidney, bone marrow, and spleen isolated from 2-year-old growth hormone receptor knockout (GHR-KO) Laron dwarf mice (with low circulating plasma levels of IGF-1) and 6-month-old bovine growth hormone transgenic (bGHTg) mice (with high circulating plasma levels of IGF-1). The ages of the two mutant strains employed in our studies were selected based on their overall ~50% survival (Laron dwarf mice live up to ~4 years and bGHTg mice up to ~1 year). Morphological analysis of the organs of long-living 2-year-old Laron dwarf mice revealed a lower biological age for their organs compared with normal littermates, with more brown adipose tissue (BAT) surrounding the main body organs, lower levels of steatosis in liver, and a lower incidence of leukocyte infiltration in different organs. By contrast, the organs of 6-month-old, short-living bGHTg mice displayed several abnormalities in liver and kidney and a reduced content of BAT around vital organs.
- PublicationOpen AccessPrimary tumor vascularity in esophagus cancer. CD34 and HIF1-α expression correlate with tumor progression(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Goscinski, Mariusz Adam; Nesland, Jahn M.; Giercksky, Karl- Erik; Dhaka, Hari PrasadObjective: Hypoxia inducible factor α (HIF1-α) is a key protein regulating the response of a variety of genes and pathways, including angiogenesis, to hypoxic stimuli. High vascularity in various carcinomas correlates with invasion and metastasis. Assessment of primary tumor vascularity and HIF1-α expression in esophageal carcinomas was an objective of this study. Methods: The vascularity in esophageal carcinomas (n=52) was quantified by Chalkley method on CD34 immunostained sections. HIF1-α expression was examined by immunohistochemistry. The relationships between CD34 Chalkley count, HIF1-α and various clinico-pathological characteristics with clinical outcome were evaluated. Results: High HIF1-α expression in squamous cell carcinoma (SCC) was significantly associated with the T3-4 group (p=0.02). A higher percentage of SCC with high HIF1-α expression compared to its expression in adenocarcinoma (AC) (p=0.005) was observed. In the SCC group, high CD34 Chalkley count and high HIF1-α expression implied a significantly reduced survival (p=0.003 and p=0.001). No such significant association was found in the AC group. Conclusions: HIF1-α expression is different in two separate tumor microenvironments: SCCs and ACs of the esophagus. This suggests that different mechanisms may be involved in HIF1-α expression- and activity between the two histological types of esophageal carcinoma.
- PublicationOpen AccessThe CCM3-GCKIII partnership(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Zalvide, Juan; Fidalgo, Miguel; Fraile, María; Guerrero, Ana; Iglesias, Cristina; Floridia, Ebel; Pombo, Celia M.Specific mutations in the CCM3 gene predispose to the development of cerebral cavernous malformations, a special type of vascular lesions. This calls for an elucidation of the precise nature of the CCM3 protein and a deep understanding of its molecular regulation. In this review, we outline our current knowledge of the different CCM3 protein complexes. We focus on the GCKIII family of kinases as partners of CCM3 and discuss the functional consequences of this partnership, putting forward a putative model for the activation of these kinases.
- PublicationOpen AccessThree-dimensional epithelial cultures: a tool to model cancer development and progression(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Eritja, Núria; Dolcet, Xavier; Matias-Guiu, XavierLoss of cell polarity is a hallmark of cancer, and although this feature is commonly observed in advanced tumors; growing evidence indicates that loss of cell-cell adhesion and cell polarity may also be important in early stages of cancer. Despite recent important advances, much remains unclear about the molecular and biophysical mechanisms involved in phenotypic changes observed in epithelial architecture during carcinogenesis. Over the past decade the use of three dimensional cultures (3D) has emerged as a valuable tool to study the functions of cancer genes and pathways in an adequate polarized context. 3D cultures are an outstanding tool to understand the morphologic consequences of molecular alterations. In other words, 3D cultures allow a much better understanding of the pathological features of tumours, with the microscope. In this review we will focus on how 3D models have provided unique insights into how basic cell biological processes impact in higher-order tissue architecture and how these models have enhanced our understanding of carcinoma biology.
- PublicationOpen AccessExpression of the chemokines CXCL12 and CX3CL1 and their receptors in human nerve sheath tumors(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Hattermann, Kirsten; Li, Gu; Hugo, Heinz-Hermann; Mentlein, Rolf; Mehdorn, Maximilian; Held-Feind, JankaPeripheral nerve sheath tumors are in most cases slowly growing neoplasms that can be adequately cured by surgical resection. However, facing the risk of a neurosurgical intervention and the trend of multiple relapses of nerve sheath tumors the development of additional therapy strategies seems to be favourable, and therefore substantiated knowledge of molecular and cellular mechanisms in nerve sheath tumors should be achieved. Here, we firstly describe the expression of the chemokines CXCL12 (SDF-1) and CX3CL1 (fractal-kine) and their respective receptors CXCR4, CXCR7 and CX3CR1 in different entities of human nerve sheath tumors and normal control tissues. Both ligands and their receptors are expressed in high to moderate levels on mRNA and protein level in benign and malignant nerve sheath tumors. While CXCL12 was mainly found in schwannoma cells (S100+) in situ, its receptor CXCR4 is also partly found on CD11b-positive macrophages / microglia and its alternative receptor CXCR7 is also expressed by endothelial cells and macrophages. CX3CL1 is expressed by parts of the schwannoma and endothelial cells, whereas its receptor CX3CR1 is expressed by nearly all tumor cells and macrophages, but not by endothelial cells. Taken together, we could show the presence of CXCL12 and CX3CL1 and their respective receptors in benign and malignant human nerve sheath tumors. Further investigations may show their functional role in health and disease.