Histology and histopathology Vol.28, nº 8 (2013)
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- PublicationOpen AccessThe peculiar apoptotic behavior of skeletal muscle cells(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Salucci, Sara; Burattini, Sabrina; Baldassarri, ValentinaApoptosis plays an active role in maintaining skeletal muscle homeostasis. Its deregulation is involved in several skeletal muscle disorders such as dystrophies, myopathies, disuse and sarcopenia. The aim of this work was to study in vitro the apoptotic behavior induced by etoposide, staurosporine and hydrogen peroxide in the C2C12 skeletal muscle cell line, comparing myoblast vs myotube sensitivity, investigated by means of morphological and cytofluorimetric analyses. Myotubes appeared more resistant than myoblasts to apoptotic induction. In myoblasts treated with etoposide, nuclei with chromatin condensation were observed, in the presence of a diffuse DNA fragmentation, as shown by confocal microscopy. The latter also appeared in myotubes, where apoptotic and normal nuclei coexisted inside the same syncytium. After staurosporine treatment, myobalsts evidenced late apoptotic features and a high number of TUNEL-positive nuclei. Secondary necrosis appeared in myotubes, where myonuclei with cleaved DNA again coexisted with normal myonuclei. After H2O2 exposure, myotubes, differently from myoblasts, showed a poor sensitivity to cell death. Intriguingly, autophagic granules appeared abundantly in myotubes after each treatment. In myotubes, mitochondria were better preserved than in myoblasts since those which were damaged were probably degraded through autophagic processes. These findings demonstrate a scarce sensitivity of myotubes to apoptotic stimuli due to acquisition of an apoptosis-resistant phenotype during differentiation. The presence of nuclear-dependent “territorial” death domains in the syncytium could explain a slower death of myotubes compared to mononucleated cells. In addition, autophagy could preserve and protect muscle cell integrity against chemical stimuli, making C2C12 cells, in particular myotubes, more resistant to apoptosis induction.
- PublicationOpen AccessEffect of regenerative factor rich plasma, P substance and fetal calf serum on the growth of epithelial cells in the cornea. Comparative experimental study(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Márquez de Aracena del Cid, R.; Pérez Ordoñez, A.Purpose. The goal of this study was to evaluate the experimental effectiveness of Regenerative Factor Rich Plasma (RFRP) of human blood versus Fetal Bovim Serum (FBS) and neuropeptide Substance P (SP) on corneal epithelium cell proliferation. Method. Rabbit corneal epithelium cell (CCL-60) growth was compared between different RFRP fractions, FBS and with the neuropeptide Substance P. The ability of the RFRP fractions and SP to revert the inhibitory effect of the CsA was also evaluated. Results. All groups showed an increase (p<0.001) in corneal epithelial cell growth compared with the control group. The maximum capacity of cell growth was obtained with dilutions of 50% in the FBS, RFRP-I, RFRP -II, RFRP-III groups and with 100nM of SP. The highest growth was observed with 50% FBS, RFRP-I and RFRP-II. The group with SP and RFRP-III had significantly lower growth (p<0.001). When the NK1 receptor antagonist CsA was added at a dose of IC50, we found a significant decrease in cell growth (p<0.001) in all culture conditions, including the control group. The decrease was similar in all groups, but was especially pronounced in RFRP-II. RFRP I, II and III promoted growth more than SFB 10%. Conclusion. The RFRP of human blood promotes the growth of corneal epithelial cells in a significantly more efficient manner than FSB and SP. RFRP can be effective both in cell cultures and stem cell cultures.
- PublicationOpen AccessSpatial-temporal protein expression of inhibitor of differentiation-1 (Id1) during fetal embryogenesis and in different mouse and human cancer types(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Redrado, Miriam; Bodegas, Elena; Villaro, Ana Cristina; Nguewa, Paul A.; Lopez, Ines; Gil-Bazo, Ignacio; Calvo, AlfonsoInhibitor of differentiation-1 (Id1) plays a role in cell proliferation, acquisition of epithelial to mesenchymal transition (EMT) features and angiogenesis. Id1 was shown to be expressed in some tumor types, mainly in advanced dedifferentiated stages. However, recent studies using a validated and highly specific monoclonal antibody against Id1 have challenged many of the results obtained by immunohistochemistry. The goal of our work was to perform a thorough analysis of Id1 expression in mouse embryos and adult tissues, as well as healthy and malignant mouse and human samples using this validated antibody (Perk et al., 2006). Our results show that Id1 was highly expressed in the oropharyngeal cavity, lung, cartilage and skin of E14 and E15 mouse embryos, but expression was progressively reduced in more developed embryos. Immunostaining only remained in epithelial cells of the gut and uterus of adult mice. Mammary MMTV-Myc and MMTV-Myc/VEGF transgenic mouse tumors, and squamous cell carcinomas of the lung induced by N-nitroso-tris-chloroethylurea (NTCU) were highly positive for Id1, unlike their respective healthy counterparts. Id1 immunostaining in a human tissue microarray (TMA) revealed strong expression in cancers of the oral cavity, bladder and cervix. Some tumor specimens of esophagus, thyroid and breast were also strongly positive. Our results suggest that Id1 is an oncofetal protein highly expressed in particular tumor types that should be reanalyzed in future studies using large cohorts of patients to reassess its diagnostic/prognostic value. Moreover, MMTV-Mycand NTCU-induced tumors could serve as appropriate mouse models to study Id1 functions in breast and lung cancer, respectively.
- PublicationOpen AccessMorphological spectrum and clinical features of myopathies with tubular aggregates(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Funk, Fabian; Ceuterick-de Groote, Chantal; Martin, Jean-Jacques; Meinhardt, Axel; Taratuto, Ana L.; De Bleecker, Jan; Coster, Rudy Van; De Paepe, Boel; Schara, Ulrike; Vorgerd, Matthias; Häusler, Martin; Koppi, Stefan; Maschke, Matthias; De Jonghe, Peter; Maldergem, Lionel Van; Noel, Stéphane; Zimmermann, Christoph W.; Wirth, Stefan; Isenmann, Stefan; Stadler, Rudolf; Schröder, J. Michael; Schulz, Jörg B.; Weis, Joachim; Claeys, Kristl G.Tubular aggregates (TAs) are aggregates of densely packed tubules in human skeletal muscle fibers with particular histochemical and ultrastructural features that most probably arise from the sarcoplasmic reticulum. Some studies have shown an additional mitochondrial origin of TAs. We studied the histopathological spectrum and clinical features in a large cohort of patients with TAs in their muscle biopsy (106 biopsies), derived from our muscle biopsy archive (15,412 biopsies in total). In particular, we examined light microscopic, enzyme histochemical, immunohistochemical and ultrastructural features in the muscle biopsies, as well as the patients’ clinical data. We found TAs in 0.5% of all muscle biopsies. Based on the size of TAs, we identified two sub-groups: (1) myopathies with large TAs (29 biopsies) in type 2 fibers and sometimes also in type 1 fibers, absence of any other associated disorder, and a familial history in half of the cases, and (2) myopathies with small TAs (77 biopsies), exclusively in type 2 fibers, presence of another associated disease in the majority of patients and mostly no familial history. In the sub-group with large TAs, we observed a high variability of ultrastructural changes. The most frequent clinical symptom in both groups was limb muscle weakness. No significant differences in clinical presentation, age at onset or disease duration at the time of biopsy were found between the two groups. In conclusion, myopathies with TAs can be sub-divided into a group with large TAs, probably corresponding to the so-called primary TA myopathies, and into a group with small TAs as a feature of another underlying condition.
- PublicationOpen AccessEffects of the relationship between 65Zn and blood cells. A dynamic and morphological study(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2013) Vera-Gil, A.; Pérez Castejón, M.J.; Whyte, J.; Cisneros, A.; Recreo, P.; Gascón, M.A.; Whyte, A.; Lahoz, M.; Pérez Castejón, M.C.We have studied the dynamic pathway of 65Zn and its autoradiographic location in blood cells, even at the ultra-structural level. We have found evidence that tends to confirm the old biochemical postulates about the capacity of this isotope to displace iron in the haemoglobin molecule. Recently, the bibliography has demonstrated that 57Co is also able to perform this displacement, but unlike 65Zn it does not invalidate the Redox function of the molecule. In the case of 65Zn, the mentioned displacement invalidates this function because the radionuclide can only use valence 2. We have also contributed evidence of erythrocytes destruction by the spleen after the incorporation of 65Zn, as well as the clearly marked degradation of haematic pigments inside the spleen.