DigitalUM :: Browsing by Subject "Fibronectin"

Browsing by Subject "Fibronectin"

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Altered distribution of extracellular matrix proteins in the periodontal ligament of periostin-deficient mice
(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2014) Tabata, Chihiro; Hongo, Hiromi; Sasaki, Muneteru; Hasegawa, Tomoka; Luiz de Freitas, Paulo Henrique; Yamada, Tamaki; Yamamoto, Tomomaya; Suzuki, Reiko; Yamamoto, Tsuneyuki; Oda, Kimimitsu; Li, Minqi; Kudo, Akira; Iida, Junichiro; Amizuka, Norio
Verifying whether periostin affects the distribution of type I collagen, fibronectin and tenascin C in the periodontal ligament (PDL) is important to contribute to a more thorough understanding of that protein’s functions. In this study, we have histologically examined incisor PDL of mandibles in 20 week-old male wild-type and periostin-deficient (periostin-/-) mice, by means of type I collagen, fibronectin, tenascin C, proliferating cell nuclear antigen, matrix metalloproteinase (MMP)-1 and F4/80-positive monocyte/ macrophage immunostaining, transmission electron microscopy and quantitative analysis of cell proliferation. Wild-type PDL featured well-arranged layers of collagen bundles intertwined with PDL cells, whose longitudinal axis ran parallel to the collagen fibers. However, cells in the periostin-/- PDL were irregularly distributed among collagen fibrils, which were also haphazardly arranged. Type I collagen and fibronectin reactivity was seen throughout the wild-type PDL, while in the periostin-/- PDL, only focal, uneven staining for these proteins could be seen. Similarly, tenascin C staining was evenly distributed in the wildtype PDL, but hardly seen in the periostin-/- PDL. MMP1 immunoreactivity was uniformly distributed in the wild-type PDL, but only dotted staining could be discerned in the periostin-/- PDL. F4/80-positive monocyte/macrophages were found midway between tooth- and bone-related regions in the wild-type PDL, a pattern that could not be observed in the periostin-/- PDL. In summary, periostin deficiency may not only cause PDL collagen fibril disorganization, but could also affect the distribution of other major extracellular matrix proteins such as fibronectin and tenascin C.
Open Access
Effect of rehabilitation protocols on muscle function and morphology following hindlimb disuse in weanling rats
(Universidad de Murcia. Departamento de Biología Celular e Histología, 2016) Leite Nogut, Keite i; Bianchi, Eduardo; Chesca Simões, Deise Lúcia; Mattiello-Sverzut, Ana Claudia; de Moura-Jucá, Renata Viana Brígido
Background: Primary or secondary disorders in developing skeletal muscles are prevalent in physical therapy practice. Assessment of gait functional changes and morphological aspects of hindlimb muscles of weanling rats have not been reported simultaneously in the literature. Rehabilitation by active (eccentric training) and passive (stretching) exercises after hypomobility needs to be investigated. Methods: After ten days of immobilisation in a plantar flexion-shortened position, animals underwent eccentric training on treadmills, intermittent (a single series of ten exercises of 30 seconds each, with a 30-s interval) or continuous stretching protocols for 40 minutes, or had free cage activity for three days. Analysis of gait variables and muscle morphology (immunohistochemical staining of soleus and plantar muscles for fibronectin and types I and III collagen and immunofluorescence staining for dystrophin, laminin, Pax-7, and CD68) were performed. Results: On the third day, the rehabilitated animals touched the ground surface with their toes, except for the group undergoing continuous stretching. The total amount of extracellular macrophages was higher in the rehabilitated animals. The number of satellite cells was not significantly different between groups. Conclusion: Three days of active training (eccentric exercise) showed greater effectiveness compared to the other rehabilitation programs. Weanling rats seem to respond differently to external stimuli such as disuse and remobilisation.
Open Access
Electron and immunoelectron microscopy on healing process of the rat anterior cruciate ligament after partial transection: the roles of multipotent fibroblasts in the synovial tissue
(Murcia : F. Hernández, 1996) Maekawa, K.; Furukawa, H.; Kanazawa, Y.; Hijioka, A.; Suzuki, K.; Fujimoto, Sunao
The healing process of the rat anterior cruciate ligament (ACL) after partial transection was examined. In sham operated samples, the synovial tissue near the infrapatellar fat pad includes an abundance of young fibroblasts which can be classified into two types: A type cells often exist along the surface of the synovial tissue and contain numerous lysosomes, while B type cells are often associated with small vessels and are actively involved in the production of fibronectin and laminin. At 1 week after transection of the ACL, B type cells frequently undergo mitotic proliferations and are eventually incorporated into the endothelium of the growing capillaries extending from the proximal remnants of the synovial tissue to the transected lesion. The transformation of B to A type cells is indicated by our electron micrographs. After 2 weeks, the replacement of the transected lesion by regenerated soft tissue becomes pronounced. After 4 weeks, B type cells in the deeper layer of the regenerated tissue are first involved in the production of type 111 and then in that of type 1 collagens as revealed by the immunocytochemistry. The present study indicates that B type cells are a kind of stem cell: they possess the ability to transform to vasoformative cells involved in a manner suggestive of vasculogenesis, to phagocytic A type cells and to the synthetic fibroblasts in the regeneration of the ACL.
Open Access
Fibronectin expression in cancer tissues from patients undergoing radiation therapy
(Murcia : F. Hernández, 1993) Nishioka, A.; Ogawal, Y.; Inomata, T.; Maeda, T.; Seguchi, H.
Fibronectin expression and distribution were examined in cancer tissues from 19 patients with cancer of the head and neck regions. Samples taken before and after irradiation of approximately l0 Gy, 20 Gy or 30 Gy were analyzed by the avidin-biotin-horseradish peroxidase method using mouse monoclonal antibodies against human fibronectin. The results were correlated with the patient's prognosis after radiation therapy. No remarkable changes in the fibronectin expression or distribution were found between tissue specimens taken before and after each dose of irradiation. The prognosis, however, varied according to the degree of expression and the distribution pattern of fibronectin. Seven patients in which the cancer tissue was encircled by a thick fibronectin network are still alive without recurrence 4.5-6 years after treatment, whereas 6 patients in which fibronectin was only faintly expressed or focally distributed died or developed recurrence soon after treatment. The present findings den~onstrate that fibronectin expression and distribution in cancer tissue are intimately related to the patient's prognosis, and that the analysis of these two parameters is applicable as a predictive assay in radiotherapy of cancer of the head and neck regions.
Open Access
Heterogeneity of mesenchymal cells in human amniotic membrane at term
(Universidad de Murcia, Departamento de Biologia Celular e Histiologia, 2024) Cortés Sandoval, Salvador; Serrano Sánchez, Mª Isabel; Ferrer, Concepción; Delgado, Juan L.; Insausti, Carmen L.; Blanquer, Miguel; Beltrán Frutos, Ester; Martínez Hernández, Jesús; Pastor García, Luis Miguel; Seco Rovira, Vicente
There is increasing interest in understanding the tissue biology of human amniotic membrane (hAM) given its applications in medicine. One cellular component is mesenchymal cells, which can be extracted, cultured and differentiated "in vitro" into various cell types. These studies show that there is heterogeneity among mesenchymal cells. The aim of this work is to study the membrane "in situ" to determine whether this cellular heterogeneity exists. The hAMs were obtained from caesarean deliveries at term and analyzed by histological techniques. Types I-III mesenchymal cells and Hofbauer were distinguished by light microscopy. Histochemically, mesenchymal cell types showed successively increasing positivity to: PAS, vimentin, fibronectin, and Concanavalin-A; VGEF, TGF-β2, PDGF-C, FGF-2. By the semiquantitative point of view, the percentage of Type II cells was 60%, significantly higher than the other types. With transmission electron microscopy, an intermediate cell type between II-III was observed. Strong vesiculation of the rough endoplasmic reticulum (RER) with exocytosis was observed. In addition, an accumulation of a similar material to the extracellular matrix in the RER caused its dilation especially in type IIITEM cells. Some of this material acquired a globular structure. These structures were also found free in the extracellular matrix. In conclusion, the mesenchymal cells of the fibroblastic layer of the hAMs studied are heterogeneous, with some undifferentiated and others with a probably senescent fibroblastic phenotype with accumulation in their RER of fibronectin. These results may be of interest to extract mesenchymal cells from hAMs for use in regenerative medicine and to better understand the mechanisms of fetal membrane rupture
Open Access
Immunohistochemical study of the upper surface layer in rat mandibular condylar cartilage
(Murcia : F. Hernández, 2004) Zea-Aragón, Z.; Ohtsuki, K.; Ohnishi, M.; Ohno, S.
Both hyaluronic acid and fibronectin localizations were examined in the upper surface layer of rat mandibular condylar cartilages by immunohistochemical techniques. Their delicate structure was successfully preserved by preparation procedures of joint condyles with disks. Paraformaldehyde-fixed cartilaginous tissues were cut in a cryostat, and cryosections were analyzed using streptavidinperoxidase and indirect immunofluorescence methods. Another immunogold method with conventional preparation procedures and a quick-freezing method was performed for their ultrastructural analyses. Both hyaluronic acid-binding protein and anti-fibronectin antibody were used to localize hyaluronic acid and fibronectin in the mandibular condylar cartilage, respectively. Some cryosections were pre-treated with hyaluronidase and chondroitinase before such labeling. The upper surface layer was composed of double laminar structures. One bordered with the cartilage matriceal surface, which was positive for fibronectin. The hyaluronic acid was localized over the fibronectin layer. Therefore, the hyaluronic acid in vivo was bound with fibronectin in the cartilaginous matrix, performing lubrication for the mandibular joint movement.
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Immunohistochemistry as a tool to characterize human skin wounds of hanging marks
(Romanian Society of Legal Medicine, 2019-03) Pérez Cárceles, María Dolores; Giménez, M.; Martínez Díaz, F.; Osuna, Eduardo; Luna, Aurelio; Legaz Pérez, Isabel; Ciencias Sociosanitarias
Estimation of age and vitality of human skin wounds both in the living and dead is essential in forensic practice. The use of immunohistochemical parameters for the age estimation and vitality of human skin wounds remains difficult. Forensic literature describes different biomarkers and methods for the differential diagnosis of vital and post-mortem wounds, but to this day it is unclear its utility. The aim of this study was analysed wounds vital origin in a series of suicidal hangings were vitality had been demonstrated using fibronectin, cathepsin D and P-selectin, in order to discover the morphological changes that occur in a vital wound, and consequently, find useful vital injury diagnosis markers. A total of 15 human vital skin wounds from ligature marks from deaths by suicidal hanging and skin controls from same cadaver were analyzed in a postmortem interval between 19-36 hours. Fibronectine, cathepsin D and P-selectin were detected by immunohistochemistry. Our result shows a strongly fibronectin-positive reaction in basement membranes and interstitial connective tissue in all specimen of wounds of ligature mark. Granular staining pattern characteristic of cathepsin D was observed mainly in the basal layer of the epidermis in normal and wound skin. Cathepsin D analysis in ligature mark showed moderate positive and strong positive cells. A weak positive immunoreactivity P-selectin were found in vital wound compared with undamaged skin. In conclusion, our data show an increase of fibronectin and cathepsin D immunoreactivity expression and a decrease of P-selectin immunoreactivity in skin wounds from ligature marks from deaths by suicidal hanging of a postmortem interval between 19-36 hours.
Open Access
lmmunocytochemistry of perinatal rat livers with a special reference to the roles of mesenchymal cells in hepatic differentiation
(Murcia : F. Hernández, 1996) Hayashida, T.; Nagata, Kengo; Doi, Y.; Ozaka, T.; Itoh, Hiroyuki
To investigate the roles of extracellular matrix produced by hepatic mesenchymal cells in the organization of hepatic cell cords, perinatal rat livers were examined with immunocytochemistry of fibronectin (FN) and larninin (LM). Some hepatocytes in a free state at prenatal day 15 actively produced FN and LM in the rough endoplasmic reticulum but lost this synthetic activity when such cells were incorporated into hepatic cell cords. On the other hand, hepatic mesenchymal cells, especially those associated with the perisinusoidal space, retained this synthetic activity throughout the stages examined. In the differentiating hepatic cell cords, positive imrnunoreactions for FN and LM were preferentially seen on the cell surface facing both sinusoidal space and differentiating bile canaiiculus concomitant with the expression of the tight junction protein, ZO-1, from prenatal day 17. Since such hepatocytes have lost or reduced their synthetic activities of both glycoproteins in the rER, the immunoreactions appear to be mainly due to hepatic mesenchymal cells which seem to play a role in the formation of the hepatic cell cords and the bile canaliculi.
Open Access
Molecular Segmentation of the Spinal Trigeminal Nucleus in the Adult Mouse Brain
(Frontiers Media, 2021-12-10) García-Guillén, Isabel M.; Martínez-de-la-Torre, Margaret; Puelles, Luis; Aroca Tejedor, Pilar; Marín San Leandro, Faustino; Anatomía Humana y Psicobiología
The trigeminal column is a hindbrain structure formed by second order sensory neurons that receive afferences from trigeminal primary (ganglionic) nerve fibers. Classical studies subdivide it into the principal sensory trigeminal nucleus located next to the pontine nerve root, and the spinal trigeminal nucleus which in turn consists of oral, interpolar and caudal subnuclei. On the other hand, according to the prosomeric model, this column would be subdivided into segmental units derived from respective rhombomeres. Experimental studies have mapped the principal sensory trigeminal nucleus to pontine rhombomeres (r) r2-r3 in the mouse. The spinal trigeminal nucleus emerges as a plurisegmental formation covering several rhombomeres (r4 to r11 in mice) across pontine, retropontine and medullary hindbrain regions. In the present work we reexamined the issue of rhombomeric vs. classical subdivisions of this column. To this end, we analyzed its subdivisions in an AZIN2-lacZ transgenic mouse, known as a reference model for hindbrain topography, together with transgenic reporter lines for trigeminal fibers. We screened as well for genes differentially expressed along the axial dimension of this structure in the adult and juvenile mouse brain. This analysis yielded genes from multiple functional families that display transverse domains fitting the mentioned rhombomeric map. The spinal trigeminal nucleus thus represents a plurisegmental structure with a series of distinct neuromeric units having unique combinatorial molecular profiles.
Open Access
Morphological and molecular characterization of healthy human ascending aorta
(F. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histología, 2012) Forte, A.; Della Corte, A.; Grossi, M.; Finicelli, M.; Bancone, C.; Provenzano, R.; Pepino, P.; Nappi, G.A.; De Feo, M.; Galderisi, U.; Cotrufo, M.; Cipollaro, M.
Knowledge of the characteristics of the normal human aorta has been constrained by lack of data on fresh aortic tissue, especially from healthy individuals. In this study, the gene expression and morphological characteristics of the thoracic ascending aorta (AA) of healthy organ donors have been evaluated, with the aim of providing reference data for the analysis of pathological AAs. We analysed by RT-PCR the differential expression of mRNAs coding for myocardin, smoothelin, alpha-smooth muscle actin (alpha-SMA) and the ED-A isoform of fibronectin (ED-A FN) in AA specimens from donors, integrating the results with immunohistochemical analysis of the same targets. Morphological and morphometric characteristics of the AAs were also evaluated. In order to account for possible regional variations in wall structure, the convexity of the aortic profile was compared to the concavity. No differences in gene expression occurred for any of the target genes between the concavity and the convexity of AAs. Immunohistochemistry revealed a different distribution of total FN and of its ED-A isoform in the media and in the intima. Smoothelin is expressed by the majority of cells in the media, with some positive cells also in the intima. Alpha-SMA is expressed in all the tunicae. Immunohistochemistry also revealed in the convexity of 50% of AAs the presence of discrete areas in the subadventital media with altered structure and cell morphology and with altered gene expression, resulting positive for ED-A FN and alpha-SMA, but not for smoothelin, indicating the occurrence of early lesions also in macroscopically healthy AAs
Open Access
Paracrine role for TGF-ß-induced CTGF and VEGF in mesangial matrix expansion in progressive glomerular disease
(F. Hernández y Juan F. Madrid. Universidad de Murcia. Departamento de Biología Celular e Histología, 2012) Soon Lee, Hyun
Transforming growth factor-ß (TGF-ß) is a key regulator of extracellular matrix (ECM), and may mediate the development of glomerulosclerosis with accumulation of mesangial matrix. Mesangial cells secrete TGF-ß in response to common in vitro fibrogenic stimuli. Yet mesangial immunostaining for active TGF-ß1 is frequently negative in chronic glomerular disease. TGF-ß is rather expressed and/or activated by podocytes in both mesangial and podocyte diseases. Activated TGF-ß/Smad signaling by podocytes may induce connective tissue growth factor (CTGF or CCN2) and vascular endothelial growth factor (VEGF) expression. Podocyte CTGF seems to have paracrine effects on mesangial cells to stimulate CTGF expression. CTGF appears to stimulate the fibronectin-matrix assembly via enhanced cell-surface expression of α5ß1 integrin in the mesangium of diseased glomeruli. Podocyte VEGF-A overexpression also seems to play a paracrine role on mesangial cells to upregulate VEGF/VEGF receptor systems and to overproduce matrix proteins. Thus, paracrine CTGF and VEGF may contribute to mesangial matrix accumulation in chronic glomerular disease, culminating in the development of glomerulosclerosis. Together, these data bring new mechanistic insights into our understanding of the pathogenic role of TGF-ß-induced CTGF and VEGF in mesangial matrix expansion in chronic progressive glomerular disease
Open Access
Phenotypic modulation of smooth muscle cells during formation of neointimal thickenings following vascular injury
(Murcia : F. Hernández, 1998) Thyberg, J.
Smooth muscle cells build up the media of mammalian arteries and constitute one of the principal cell types in atherosclerotic and restenotic lesions. Accordingly, they show a high degree of plasticity and are able to shift from a differentiated, contractile phenotype to a less differentiated, synthetic phenotype, and then back again. This modulation occurs as a response to vascular injury and includes a prominent structural reorganization with loss of myofilaments and formation of an extensive endoplasmic reticulum and a large Golgi complex. At the same time, the expression of cytoskeletal proteins and other gene products is altered. As a result, the cells lose their contractility and become able to migrate from the media to the intima, proliferate, and secrete extracellular matrix components, thereby contributing to the formation of intimal thickenings. The mechanisms behind this change in morphology and function of the smooth muscle cells are still incompletely understood. A crucial role has been ascribed to basement membrane proteins such as laminin and collagen type IV and adhesive proteins such as fibronectin. A significant role is also played by mitogenic proteins such as platelet-derived growth factor (PDGF) and basic fibroblast growth factor (bFGF). An improved knowledge of the regulation of smooth muscle differentiated properties represents an important part in the search for new methods of prevention and treatment of vascular disease.
Open Access
Quantitative in situ hybridization for the evaluation of gene expression in asynchronous and synchronized cell cultures and in tissue sections
(Murcia : F. Hernández, 1999) Barlati, S.; Zoppi, N.; Copeta, A.; Tavian, D.; De Petro, G.; Colombi, M.
We describe an image analysis (IA) system that has been applied for the quantitative evaluation of mRNAs evidenced by in situ hybridization (ISH) with radiolabelled probes in cultured cells and in tissue sections. The ISH-IA method was used for the evaluation of cultured cell morphological parameters such as cell and nucleous area (CA and NA, respectively) in parallel with the levels of mRNAs detected as hybridization grains areas (GA). The evaluation of these parameters, together with the analysis of the levels of mRNAs (c-jun, cyclin A) specific for given cell cycle phases (i.e. G1 and S/G2), allowed the identification, in asynchronous cultures of human skin fibroblasts, of cells in G1 and SlG2 phases. The mRNA levels measured by ISH-AI were comparable with those detected by RT-PCR. This method was also applied for the analysis of fibronectin (FN) gene expression in control skin fibroblasts in relationship with the different phases of the cell cycle and in comparison with a tumor cell line (Sk-Hepl), heterogeneous either for morphometric parameters or for the levels of this transcript. Finally, the ISH-AI was applied for the semiquantitative evaluation of the expression, localization and alternative splicing pattern of FN mRNA in normal liver and in hepatocellular carcinoma (HCC) tissue sections.
Open Access
Role of multipotent fibroblasts in the healing colonic mucosa of rabbits. Ultrastructural and immunocytochemical study
(Murcia : F. Hernández, 1992) Mori, Naoki; Doi, Y.; Hara, K.; Yoshizuka, M.; Ohsato, K.; Fujimoto, Sunao
Light- and electron microscopy and immunocytochemistry were used to study the healing colonic mucosa of rabbits after experimental excision. Between 3 and 5 days, abundant young fibroblasts which retained many features of mesenchymal cells invaded the growing capillaries into the loose connective tissue of the healing colonic mucosa. Our electron microscopy revealed the transformation of these young fibroblasts into smooth muscle cells, into histiocyte-like cells involved in phagocytotic activity, and into vasoformative cells incorporated into the growing capillaries. The mitotic proliferation of pre-existing smooth muscle cells at the ulcer margin did not seem to be the major reason for re-establishment of the muscular tissue. The present immunocytochemistry revealed an active production of fibronectin in rough endoplasmic reticulum in the young fibroblasts. This may mean that this glycoprotein is involved in the re-establishment of both connective and muscular tissues by enhancement of adhesion and chemoattractant activities of such cells. In addition, the immunoreaction of endothelial celis of the growing capillaries suggests a role of this glycoprotein in the acceleration of the neocapillarization.
Open Access
The glomerular distribution of laminin and fibronectin in glomerulonephritis
(Murcia : F. Hernández, 1993) Nakopoulou, Lydia; Stefanak, K.; Zeis, P.M.; Papadakis, J.; Boletis, J.; Vosnidis, G.; Davaris, P.
Laminin (LAM) and fibronectin (FI) are regarded as major components of the glomerular extracellular matrix The aim of this study was to define the distribution of LAM and F1 in primary glomerulonephritis (GN) and GN of systemic lupus erythematosus (SLE) and to correlate the type of glomerular disorders with possible changes in the expression of these components. Normal portions of kidney tissue from 10 patients with renal tumors and sixty-six renal biopsies obtained from patients with GN were studied by the immunoperoxidase- antiperoxidase (PAP) method for the detection of LAM and FI. Twelve patients had membranous GN (MGN), 8 mesangiocapillary GN (MCGN), 2 1 mesangioproliferative GN (MPGN), including I l cases of IgA nephropathy, 11 focal segmental glomerulosclerosis (FSGS) and 14 had SLE. In MGN, LAM was detected more intensely than F1 along the glomerular basement membranes (GBM), in subepithelial GBM protrusions and in the newly-formed GBM. On the contrary, F1 was intensely expressed in the mesangium. LAM and FI expression was pronounced in stages I1 and 111 of MGN. In MCGN, LAM and F1 were diffusely expressed along the GBM and in the mesangium. The distribution of the two antigens in MPGN and FSGS was similar to that seen in normal glomeruli. However, the F1 staining reaction was more intense in severe mesangioproliferative lesions, mainly observed in the cases of IgA-nephropathy, There were no differences in the distribution of LAM and F1 between primary and SLE GN. The antigen staining pattern was pronounced in the membranous and mesangiocapillary lesions of SLE GN. The crescents observed in 7 cases contained increased amounts of both LAM and FI, while the adhesions with Bowman's capsule seen in 9 cases demonstrated increased amounts of LAM. In contrast, the large adhesions observed in 2 cases and the sclerotic lesions in 4 cases contained only small amounts of LAM. Offprint requests to: Dr. Lydia Nakopoulou. Department of Pathology. Medical School, University of Athens, 138-140 Grigoriou Afxediou, Alhens 15772, Greece In conclusion, the increased LAM and F1 glomerular expression mainly in MGN and MCGN, and the F1 overexpression in severe mesangioproliferative lesions of IgA nephropathy suggest that the disturbance of extracellular glomerular matrix is probably due to the damage of glomerular cells or the involvement of the above components in immune-complex formation.

Browsing by Subject "Fibronectin"

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