RESUMEN
Podoplanin (D2-40) was shown to be expressed in cancer-associated fibroblasts (CAFs) of various malignancies. The study aimed at examining its impact on angiogenesis and lymphangiogenesis markers in invasive ductal carcinoma of the breast (IDC). The studies were performed on 104 archival cases of IDC using immunohistochemical technique. Podoplanin expression in CAFs correlated positively with cancer cell VEGF-C expression (r=0.19, p=0.0495) and intratumoral microvessel count (MVC) of CD31 positive vessels (r=0.30, p=0.0018), whereas negative correlations were observed with peritumoral MVC of D2-40 and Lyve-1 positive lymphatic vessels (r=-0.26, p=0.008 and r=-0.27, p=0.0058, respectively). Podoplanin expression in CAFs did not correlate with VEGF-A and VEGF-D expression in cancer cells, nor exerted any prognostic significance. Podoplanin expression in CAFs may have impact on angio- and lymphangiogenesis processes in IDC.
Asunto(s)
Neoplasias de la Mama/metabolismo , Carcinoma Ductal de Mama/metabolismo , Fibroblastos/metabolismo , Glicoproteínas de Membrana/biosíntesis , Factor C de Crecimiento Endotelial Vascular/biosíntesis , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/análisis , Neoplasias de la Mama/patología , Carcinoma Ductal de Mama/patología , Femenino , Humanos , Inmunohistoquímica , Linfangiogénesis/fisiología , Persona de Mediana Edad , Neovascularización Patológica/metabolismo , Neovascularización Patológica/patologíaRESUMEN
BACKGROUND: It was shown recently on the level of gene expression that UGT8, coding UDP-galactose:ceramide galactosyltransferase, is one of six genes whose elevated expression correlated with a significantly increased the risk of lung metastases in breast cancer patients. In this study primary tumours and their lung metastases as well as breast cancer cell lines were analysed for UGT8 expression at the protein level. METHODS: Expression of UGT8 in breast cancer tissue specimens and breast cancer cell lines was analysed using IHC, real-time PCR and Western blotting. RESULTS: Comparison of the average values of the reaction intensities (IRS scale) showed a significant difference in UGT8 expression between (1) primary and metastatic tumours (Mann-Whitney U, P<0.05), (2) tumours of malignancy grades G3 and G2 (Mann-Whitney U, P<0.01) as well as G3 and G1 (Mann-Whitney U, P<0.001) and (3) node-positive and node-negative tumours (Mann-Whitney U, P<0.001). The predictive ability of increased expression of UGT8 was validated at the mRNA level in three independent cohorts of breast cancer patients (721). Similarly, breast cancer cell lines with the 'luminal epithelial-like' phenotype did not express or weakly expressed UGT8, in contrast to malignant, 'mesenchymal-like,' cells forming metastases in nude mice. CONCLUSION: Our data suggest that UGT8 is a significant index of tumour aggressiveness and a potential marker for the prognostic evaluation of lung metastases in breast cancer.
Asunto(s)
Biomarcadores de Tumor/genética , Neoplasias de la Mama/genética , Carcinoma Ductal de Mama/genética , Balactosiltransferasa de Gangliósidos/genética , Neoplasias Pulmonares/genética , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias de la Mama/patología , Carcinoma Ductal de Mama/patología , Línea Celular Tumoral , Femenino , Expresión Génica , Humanos , Neoplasias Pulmonares/secundario , Persona de Mediana Edad , PronósticoRESUMEN
This study aims at presenting histology of growing and mature antlers in red deer stag (Cervus elaphus). Growing antlers constitute a model organ for examining regeneration processes of tissues because they are the only mammalian appendages capable of regeneration. Histological study revealed that the tip of a growing antler consists of hairy skin, perichondrium, mesenchyme and chondroprogenitors area. By performing immunochistochemistry, we found that cell expressing Ki-67 and PCNA antigens were localized in basal layer of epidermis, skin glands and beneath their secretory sections, mesenchyme as well as within and in the vicinity of central blood vessels. Ultrastructurally, cells from chondroprogenitors zone have chondroblast-like morphology and take part in producing of collagen fibres followed by the process of cartilage mineralization. However, mature antlers also consist of lamellar osseous tissue.
Asunto(s)
Cuernos de Venado/fisiología , Ciervos , Regeneración/fisiología , Animales , Cuernos de Venado/anatomía & histología , Cuernos de Venado/citología , Cuernos de Venado/ultraestructura , Inmunohistoquímica/veterinaria , Antígeno Ki-67/análisis , Antígeno Nuclear de Célula en Proliferación/análisisRESUMEN
AIM: Inhaled nitric oxide (iNO) is a selective pulmonary vasodilator. We hypothesized that those piglets exposed to prolonged iNO react with a modified renal function. METHODS: Randomized, placebo-controlled exposure to 40 p.p.m. iNO (30 h) in piglets (n = 20). Plasma and urine were sampled during three periods (first and second 12 h periods, and finally a 6 h period). We measured urine volumes, plasma and urine electrolytes (UNa, UK, UCl), plasma creatinine and urea. We calculated creatinine clearance (Ccr), and fractional excretions of sodium and potassium (FENa, FEK) and urinary excretions of electrolytes (UENa, UEK, UECl). Haemodynamic data were recorded and renal tubular apoptosis detected. RESULTS: For the first 12 h, certain parameters significantly increased in the iNO group (mean +/- SD): UNa (mmol L(-1)), 87.7 (+/-35.0) vs. 39.3 (+/-22.9), UCl (mmol L(-1)) 80.4 (+/-32.8) vs. 48.0 (+/-26.7), FENa (%) 2.1 (+/-0.8) vs. 0.7 (+/-0.5), FEK (%) 31.7 (+/-7.0) vs. 20.7 (+/-12.3), as well as UENa (mmol) 61.0 (+/-21.1) vs. 27.6 (+/-17.9) and UECl (mmol) 57.3 (24.5) vs. 37.6 (29.0). These changes were absent in the second and third periods of the study. Significant differences in percentage of apoptotic cell nuclei in the renal cortex and medulla were found after iNO exposure: 39% vs. 15%. CONCLUSION: Exposure to 40 p.p.m. iNO in healthy anaesthetized piglets has a transient natriuretic effect that disappears after 12 h. We also found evidence of renal tubular apoptosis promotion after 30 h of iNO.
Asunto(s)
Apoptosis/efectos de los fármacos , Túbulos Renales/efectos de los fármacos , Túbulos Renales/fisiología , Óxido Nítrico/administración & dosificación , Administración por Inhalación , Animales , Núcleo Celular/fisiología , Cloruros/orina , Esquema de Medicación , Corteza Renal/fisiología , Médula Renal/fisiología , Natriuresis/efectos de los fármacos , Óxido Nítrico/efectos adversos , Potasio/orina , Porcinos , Factores de TiempoRESUMEN
Excessive physical exercise may lead to disturbance of the entire homeostasis in the body, including damage not only in skeletal muscles but also in many distant organs. The mechanisms responsible for the exercise-induced changes could include oxidative stress or angiotensin II. We previously showed that acute exercise led to apoptosis in kidney but not as a result of oxidative stress. In this study, we examined the role of angiotensin II and its AT1 and AT2 receptors in mediation of exercise-induced apoptosis in kidney. We clearly demonstrated that acute physical exercise induced apoptosis in renal cells of distal convoluted tubuli and cortical and medullary collecting ducts. Moreover, the cells displayed an increased expression of both AT1 and AT2 angiotensin II receptors and of p53 protein. The results suggest that angiotensin II could upregulate p53 expression in renal distal convoluted tubular cells and in the cells collecting ducts via both AT1 and AT2 receptors, which might be the crucial apoptosis-mediating mechanism in kidneys after excessive exercise.
Asunto(s)
Apoptosis/fisiología , Riñón/citología , Condicionamiento Físico Animal/fisiología , Receptor de Angiotensina Tipo 1/fisiología , Receptor de Angiotensina Tipo 2/fisiología , Angiotensina II/fisiología , Animales , Regulación de la Expresión Génica/fisiología , Inmunohistoquímica , Riñón/química , Riñón/fisiología , Túbulos Renales Colectores/química , Túbulos Renales Colectores/citología , Túbulos Renales Colectores/fisiología , Túbulos Renales Distales/química , Túbulos Renales Distales/citología , Túbulos Renales Distales/fisiología , Masculino , Estrés Oxidativo/fisiología , Ratas , Ratas Wistar , Receptor de Angiotensina Tipo 1/análisis , Receptor de Angiotensina Tipo 1/genética , Receptor de Angiotensina Tipo 2/análisis , Receptor de Angiotensina Tipo 2/genética , Proteína p53 Supresora de Tumor/análisis , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/fisiologíaRESUMEN
PURPOSE: To evaluate the effect of endurance exercise on the activity changes of selected lysosomal enzymes in particular types of rat muscle fibers, occurring by 0-4 days following the trial. MATERIAL AND METHODS: The experiment was performed on 3 month old male Wistar rats with body mass 250 +/- 25 g, exposed to single physical exercise on moving track (speed 17 m x min(-1), decline 0 degree, duration 87.5 +/- 27.5 min). Biochemical analyses were performed on homogenized fast-twitch FTa and FTb (m. gastrocnemius) and slow-twitch ST (m. soleus) muscle fibers of animals sacrificed 2 h (group II), 6 h (III) or 96 h (IV) after exercise and control group. The measurements considered protein concentration and the activities of beta-glucuronidase (beta-GRS), N-acetyl-beta-D-glucosaminidase (NAG), and arylsulphatase A (ASA). RESULTS: In FTa fibers, ASA and beta-GRS activities were elevated in all the exercised groups, with the most evident changes in animals tested 96 h post trial (group IV), while the peak of NAG activity was demonstrated 2 h after exercise (group II). In contrast, in FTb and ST fibers the levels of all the enzymes studied peaked 96 h after exercise, following the transient decrease in activity. CONCLUSIONS: The present study demonstrated that maximal running exercise, without the eccentric components, affects the activities of lysosomal enzymes in all types of rat muscular fibers. The lack of uniform activity profile for the lysosomal enzymes studied probably reflects the variety of their cellular functions.
Asunto(s)
Enzimas/metabolismo , Músculo Esquelético/química , Resistencia Física/fisiología , Acetilglucosaminidasa/análisis , Animales , Arilsulfatasas/análisis , Enzimas/análisis , Prueba de Esfuerzo , Glucuronidasa/análisis , Lisosomas/metabolismo , Masculino , Músculo Esquelético/metabolismo , Ratas , Ratas WistarRESUMEN
Intensive physical exercise disturbs the entire homeostasis in the body and leads to changes in haemodynamic and metabolic alterations not only in skeletal muscles but also in many distant organs. In response to acute physical exercise, a decrease of the glomerular filtration may occur, followed by stimulation of the renin-angiotensin system (RAS). Recent studies have shown that both AT1 and AT2 angiotensin receptors may play a role in mediating the apoptotic process in the kidney. Our previous studies have demonstrated an occurrence of apoptosis in rat renal tubular cells after an excessive exercise. The aim of the present study was to determine the possible mechanism of exercise-induced apoptosis in rat kidney. The analysis was performed on kidneys of rats, subjected to treadmill running until exhaustion. Apoptosis was detected in paraffin sections by the TUNEL technique. The expression of AT1 and AT2 receptors in renal tubular cells was examined by immunohistochemistry and Western blot. Our results confirmed that apoptosis after physical exercise is present in renal distal tubular cells. Moreover, there was an increased expression of AT1 and AT2 receptors in distal tubular cells. These studies suggest that physical exercise may induce apoptosis by a mechanism, involving the activation of angiotensin AT1 and AT2 receptors.
Asunto(s)
Apoptosis/fisiología , Corteza Renal/fisiología , Túbulos Renales/fisiología , Condicionamiento Físico Animal , Receptor de Angiotensina Tipo 1/metabolismo , Receptor de Angiotensina Tipo 2/metabolismo , Animales , Corteza Renal/citología , Túbulos Renales/citología , Masculino , Modelos Animales , Ratas , Ratas WistarRESUMEN
Metallothionein (MT) is a low molecular weight protein, which participates in differentiation and proliferation of normal and tumour cells. In some malignant tumours (mammary, renal, ovarian cancers), its increased expression is thought to represent an unfavourable prognostic factor. Non-small-cellular lung cancers (mainly squamocellular cancer and adenocarcinoma) are characterised by ill-defined prognosis, which poses problems in the selection of effective post-surgical therapy. The present study aimed at demonstration of the prognostic significance of MT expression in cells of non-small cell lung cancers, attempting to correlate the intensity of MT expression with G grade and with the intensity of proliferation-associated antigen, Ki-67 expression. The studies were performed on archival paraffin blocks with samples of 25 cases of non-small cell lung cancers (5 squamous cell cancers, 20 adenocarcinomas). In paraffin sections of the studied tumours, immunocytochemical reactions were performed, using mouse monoclonal anti-MT and anti-Ki-67 antibodies. The expressions of MT and Ki-67 were demonstrated in all the studied tumours. An analysis of correlation between the expression of MT, Ki-67 antigen and G grade demonstrated a strong positive relation between the latter two parameters (r=0.70; p<0.05). Less pronounced positive correlations were disclosed between MT expression and G grade (r=0.44; p<0.05) and between MT expression and the expression of Ki-67 antigen (r=0.41; p<0.05). In addition, in 15 cases of examined tumours, survival analysis was performed, which disclosed a shorter survival in patients with high MT expression. The obtained results confirmed the relationship between MT expression and Ki-67 antigen expression, indicating an involvement of the proteins in processes of tumour cell proliferation. In turn, the shorter survival of patients with high expression of MT pointed to prognostic significance of the protein in non-small cell lung cancers.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/patología , Neoplasias Pulmonares/patología , Metalotioneína/metabolismo , Adenocarcinoma/patología , Carcinoma de Pulmón de Células no Pequeñas/mortalidad , Carcinoma de Células Escamosas/mortalidad , Carcinoma de Células Escamosas/patología , Humanos , Antígeno Ki-67/metabolismo , Neoplasias Pulmonares/mortalidad , Análisis de SupervivenciaAsunto(s)
Apoptosis/fisiología , Fibras Musculares Esqueléticas/citología , Músculo Esquelético/fisiología , Esfuerzo Físico/fisiología , Proteínas Proto-Oncogénicas c-bcl-2 , Animales , Fragmentación del ADN , Etiquetado Corte-Fin in Situ , Ratones , Fibras Musculares Esqueléticas/fisiología , Músculo Esquelético/química , Músculo Esquelético/citología , Proteínas Proto-Oncogénicas/análisis , Proteína X Asociada a bcl-2 , Receptor fas/análisisRESUMEN
Apoptosis is well accepted as a type of cell death occurring in the development of mammalian muscles, but the death of adult myofibres in neuromuscular disorders and exercise-induced muscle damage is usually explained in terms of muscle necrosis. The current view that apoptosis precedes necrosis in death of dystrophin-deficient muscle fibres of mdx mouse has been well substantiated. Moreover, apoptotic myonuclei have been reported to increase in mdx mice 2 days after spontaneous exercise. To investigate the contribution of apoptosis to exercise-induced damage of normal muscle fibre a time-course analysis has been performed in adult C57BL/6 mice. Groups of five mice were sacrificed immediately after the end of the exercise, and after a rest period of 6 or 96 h. The amount of apoptosis in leg muscles was assessed by electron microscopy, by the terminal deoxynucleotidyl transferase assay and by electrophoretic detection of fragmented DNA; the expression of Bcl-2, Bax, Fas, ICE, p53 and ubiquitin was examined by immunohistochemistry and Western blot. Absent in muscles of normal 'sedentary' mice, apoptotic myonuclei peak in muscles of normal mice after a night of spontaneous wheel-running (4% +/- 3.5, immediately and 2.5% +/- 1.8 after 6 h rest, P < 0.05 vs non-runner mice); they then decrease but are present 4 days later (0.8% +/- 1.5). Satellite cells are also involved in the apoptotic process. Myofibre content of Bcl-2 decreases whereas Bax, Fas, ICE and ubiquitin modify their pattern of expression in correlation with the changes in apoptotic myonuclei. Apoptosis of endothelial cells is present after the night of wheel-running and with a twofold increase 4 days later (1.5 +/- 2.3 and 4.8 +/- 4.4 P < 0.05, respectively). Satellite cells are also involved in the apoptotic process. Thus, spontaneous running in unaccustomed mice increases the number of apoptotic nuclei in adult muscle fibres and in endothelial cells. It remains to be established whether muscle apoptosis is restricted to the repair mechanisms, as often suggested in many pathologic processes, or it is also part of pathogenesis of muscle damage. Regardless of whether these results are extended to human dystrophies, the clinical implications in terms of secondary pathogenetic mechanisms and muscle training are obvious.
Asunto(s)
Apoptosis/fisiología , Fibras Musculares Esqueléticas/patología , Músculo Esquelético/patología , Carrera/fisiología , Animales , Western Blotting , Núcleo Celular/patología , Modelos Animales de Enfermedad , Inmunohistoquímica , Etiquetado Corte-Fin in Situ , Ratones , Ratones Endogámicos C57BL , Microscopía Electrónica , Fibras Musculares Esqueléticas/química , Proteínas Musculares/análisis , Músculo Esquelético/fisiología , Regeneración/fisiología , Ubiquitinas/análisisRESUMEN
By using a polyclonal antibody raised against smooth muscle Myosin Light Chain Kinase of adult chicken we show that the 135 kDa smooth muscle Myosin Light Chain Kinase isoform is present in neonatal and regenerating rat skeletal muscle, as well as in adult atrial myocardium. No reaction was evident in adult skeletal muscle fibres. In neonatal and in early regenerating muscle smooth muscle Myosin Light Chain Kinase is associated with embryonic myosin as revealed by their co-presence in muscle fibres. Experiments in vitro show the same results in myotubes. In atrial myocardium there is a patchy positivity in certain group of myocytes. Immunoblotting experiments show in muscle cell cultures, in neonatal and in regenerating skeletal muscle a protein band with electrophoretic mobility corresponding to that of smooth muscle Myosin Light Chain Kinase. These results suggest that the expression of smooth muscle Myosin Light Chain Kinase is not fully tissue-specific and that regulation of the contractile machinery could be different during myogenesis and in adulthood, in relation to the peculiar dynamic characteristics of developing muscles.
Asunto(s)
Músculo Esquelético/fisiología , Músculo Liso/química , Quinasa de Cadena Ligera de Miosina/metabolismo , Animales , Animales Recién Nacidos , Células Cultivadas , Pollos , Técnica del Anticuerpo Fluorescente Indirecta , Immunoblotting , Músculo Esquelético/embriología , Músculo Esquelético/metabolismo , Miocardio/metabolismo , Ratas , Ratas Wistar , RegeneraciónRESUMEN
Apoptosis plays a major role in several diseases, including viral infections, autoimmune diseases, cancer, cardiac infarct, and neurological disorders. To investigate the role of apoptosis in muscular dystrophy, dystrophin-deficient (mdx) mice were subjected to spontaneous exercise and skeletal muscles were analyzed for apoptosis and ubiquitin. The increase of apoptotic myonuclei after exercise was detected by TUNEL, by electron microscopy, and by DNA analyses for high molecular weight and for ladder fragments. Expression of ubiquitin correlated with exercise and with positive myonuclei for apoptosis. Biochemical analysis confirmed a high level of ubiquitination both in sarcoplasmic and myofibrillar proteins. Muscles from sedentary congenit control mice (C57B ) were negative for apoptosis, while after exercise some nuclei were positive. We also revealed that normal myoblasts committed to apoptosis in vitro showed an increased expression of ubiquitin. Western blot for bcl-2, FasL, and BAG1 showed a significant decrease of bcl-2 product only in mdx mice after exercise. Thus, spontaneous exercise results in the increase of ubiquitin expression and in the reduction of bcl-2 tightly related to programmed cell death in mdx mice. These findings confirm that DNA fragmentation, absent in muscles of sedentary normal mice but present in mdx mice at rest, dramatically increases after exercise, shedding new light on exercise-induced muscle damage and on its progression in dystrophinopathies.