RESUMEN
Three dimensional (3D) in vitro neuronal cultures can better reproduce physiologically relevant phenotypes compared to 2D-cultures, because in vivo neurons reside in a 3D microenvironment. Interest in neuronal 3D cultures is emerging, with special attention to the mechanical forces that regulate axon elongation and sprouting in three dimensions. Type I collagen (Col-I) is a native substrate since it is present in the extracellular matrix and hence emulates an in vivo environment to study axon growth. The impact of its mechanical properties needs to be further investigated. Here, we generated Col-I 3D matrices of different mechanical stiffness and evaluated axon growth in three dimensions. Superior cervical ganglion (SCG) explants from neonatal rats were cultured in soft and stiff Col-I 3D matrices and neurite outgrowth was assessed by measuring: maximum neuritic extent; neuritic halo area and fasciculation. Axonal cytoskeletal proteins were examined. Axon elongation in stiff Col-I 3D matrices was reduced (31%) following 24 h in culture compared to soft matrices. In stiff matrices, neurites fasciculated and formed less dense halos. Consistently, almost no F-actin rich growth cones were recognized, and F-actin staining was strongly reduced in the axonal compartment. This study shows that stiffness negatively affects 3D neurite outgrowth and adds insights on the cytoskeletal responses upon mechanic interactions of axons with a 3D environment. Our data will serve to facilitate the development of model systems that are mechanically well-behaved but still mimic key physiologic properties observed in vivo.
Asunto(s)
Colágeno Tipo I , Conos de Crecimiento , Actinas , Animales , Axones/fisiología , Células Cultivadas , Matriz Extracelular , Neuritas , RatasRESUMEN
In Uruguay, a country with a small population, and hence a small scientific community, there were no classical embryologists as such in the past. However, in the decade of the 1950s, a cumulus of favorable conditions gave rise to highly active and modern research groups in the fields of cytology and physiology, which eventually contributed to developmental biology. The advent of a long dictatorship between the 1970's and 1980's caused two things: a strong lag in local research and the migration of young investigators who learned abroad new disciplines and technologies. The coming back to democracy allowed for the return of some, now as solid researchers, and together with those who stayed, built a previously inexistent postgraduate training program and a globally-integrated academy that fostered diversity of research disciplines, including developmental biology. In this paper, we highlight the key contributions of pioneer researchers and the significant role played by academic and funding national institutions in the growth and consolidation of developmental biology in our country.
Asunto(s)
Biología Evolutiva , Biología Evolutiva/tendencias , UruguayRESUMEN
Endometriosis is a benign estrogen-dependent chronic gynecological disease characterized by the presence of endometrial-like tissue outside the uterine cavity. In both women and experimental endometriotic rats, endometriosis lesions endow autonomic and sensory nerves, which are thought to contribute to the disease-associated pain. Some evidence indicates that the reinnervation of lesions is regulated by factors produced by the endometrial tissue as well as by environmental factors from the peritoneum. In this study, we examined the reinnervation of the rat endometrial tissue in an ectopic environment different from the peritoneum employing the anterior eye chamber model of experimental endometriosis. At 3 and 6weeks following transplantation, endometrial grafts retained many histological features of the eutopic tissue. Both sympathetic and sensory nerves reinnervated endometrial grafts and distributed in the stroma-like tissue, around blood vessels and in close proximity to the glands and lining epithelium. Sympathetic innervation was more robust than sensory innervation. No significant topographical relationship between sympathetic nerves and macrophages was observed. These results suggest that the rat endometrium possesses intrinsic neuritogenic capacities and can be reinnervated by sympathetic and sensory nerves in ectopic sites different from the peritoneum.
Asunto(s)
Endometriosis/fisiopatología , Endometrio/inervación , Endometrio/fisiopatología , Aloinjertos/inervación , Aloinjertos/patología , Aloinjertos/fisiopatología , Animales , Péptido Relacionado con Gen de Calcitonina/metabolismo , Modelos Animales de Enfermedad , Endometriosis/patología , Endometriosis/cirugía , Endometrio/patología , Endometrio/cirugía , Femenino , Inmunohistoquímica , Macrófagos/patología , Macrófagos/fisiología , Ratas Wistar , Sistema Nervioso Simpático/patología , Sistema Nervioso Simpático/fisiopatologíaRESUMEN
Estrogen inhibits the growth and causes the degeneration (pruning) of sympathetic nerves supplying the rat myometrium. Previous cryoculture studies evidenced that substrate-bound signals contribute to diminish the ability of the estrogenized myometrium to support sympathetic nerve growth. Using electron microscopy, here we examined neurite-substrate interactions in myometrial cryocultures, observing that neurites grew associated to collagen fibrils present in the surface of the underlying cryosection. In addition, we assessed quantitatively the effects of estrogen on myometrial collagen organization in situ, using ovariectomized rats treated with estrogen and immature females undergoing puberty. Under low estrogen levels, most collagen fibrils were oriented in parallel to the muscle long axis (83% and 85%, respectively). Following estrogen treatment, 89% of fibrils was oriented perpendicularly to the muscle main axis; while after puberty, 57% of fibrils acquired this orientation. Immunohistochemistry combined with histology revealed that the vast majority of fine sympathetic nerve fibers supplying the myometrium courses within the areas where collagen realignment was observed. Finally, to assess whether depending on their orientation collagen fibrils can promote or inhibit neurite outgrowth, we employed cryocultures, now using as substrate tissue sections of rat-tail tendon. We observed that neurites grew extensively in the direction of the parallel-aligned collagen fibrils in the tendon main axis but were inhibited to grow perpendicularly to this axis. Collectively, these findings support the hypothesis that collagen reorientation may be one of the factors contributing to diminish the neuritogenic capacity of the estrogen-primed myometrial substrate.
Asunto(s)
Colágeno/metabolismo , Estrógenos/metabolismo , Miometrio/metabolismo , Animales , Técnicas de Cultivo de Célula , Colágeno/ultraestructura , Estrógenos/administración & dosificación , Femenino , Inmunohistoquímica , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/metabolismo , Miometrio/citología , Miometrio/crecimiento & desarrollo , Miometrio/inervación , Proyección Neuronal/fisiología , Ovariectomía , Ratas Wistar , Maduración Sexual/fisiología , Simpatectomía , Sistema Nervioso Simpático/citología , Sistema Nervioso Simpático/crecimiento & desarrollo , Sistema Nervioso Simpático/metabolismo , Cola (estructura animal)/metabolismo , Tendones/metabolismoRESUMEN
Chemorepellent signals of the semaphorin family are known to play a crucial role in the development of the nervous system. Some semaphorins continue being expressed in the adult life when they regulate plasticity and regeneration. Increasing evidence indicates that semaphorins are implicated in the development of the autonomic nervous system as well as in the regulation of different forms of plasticity observed in the adulthood. Here we present selected examples illustrating the involvement of semaphorins in the regulation of autonomic plasticity in physiological and pathological conditions.
RESUMEN
Current evidence indicates that rises in systemic levels of estrogen create in the uterus an inhibitory environment for sympathetic nerves. However, molecular insights of these changes are far from complete. We evaluated if semaphorin 3F mRNA, a sympathetic nerve repellent, was produced by the rat uterus and if its expression was modulated by estrogen. We also analyzed whether uterine nerves express the semaphorin 3F binding receptor, neuropilin-2. Uterine levels of semaphorin 3F mRNA were measured using real time reverse transcriptase-polymerase chain reaction in prepubertal rat controls and following chronic estrogen treatment. Localization of semaphorin 3F transcripts was determined by in situ hybridization and the expression of neuropilin-2 was assessed by immunohistochemistry. These studies showed that: (1) chronic estrogen treatment led to a 5-fold induction of semaphorin 3F mRNA in the immature uterus; (2) estrogen provoked a tissue-specific induction of semaphorin 3F which was particularly localized in the connective tissue that borders muscle bundles and surrounds intrauterine blood vessels; (3) two major cell-types were recognized in the areas where transcripts were concentrated, fibroblast-like cells and infiltrating eosinophil leukocytes; and (4) some delicate nerve terminal profiles present in the estrogenized uterus were immunoreactive for neuropilin-2. Temporal and spatial expression patterns of semaphorin 3F/neuropilin-2 are consistent with a possible role of this guidance cue in the remodeling of uterine sympathetic innervation by estrogen. Though correlative in its nature, these data support a model whereby semaphorin 3F, in combination with other inhibitory molecules, converts the estrogenized myometrium to an inhospitable environment for sympathetic nerves.
Asunto(s)
Estrógenos/fisiología , Miometrio/inervación , Degeneración Nerviosa/metabolismo , Degeneración Nerviosa/fisiopatología , Proteínas del Tejido Nervioso/biosíntesis , Fibras Simpáticas Posganglionares/metabolismo , Regulación hacia Arriba/fisiología , Útero/inervación , Animales , Femenino , Péptidos y Proteínas de Señalización Intracelular/agonistas , Péptidos y Proteínas de Señalización Intracelular/genética , Miometrio/fisiología , Proteínas del Tejido Nervioso/agonistas , Proteínas del Tejido Nervioso/genética , Ratas , Ratas Wistar , Útero/fisiologíaRESUMEN
In order to evaluate the contribution of substrate-bound factors to the extent and patterning of the sympathetic innervation of rat uterus following estrogen treatment, superior cervical ganglion explants from neonatal and adult ovariectomized rats were cultured on tissue sections of fresh frozen uterus from adult ovariectomized rats treated with estrogen or a vehicle. The main findings were: (1) neurite growth was greatly influenced by histological features of the underlying section; (2) on myometrial sections, neurites followed the orientation of the main axis of the longitudinally sectioned muscle cells; (3) neurites showed limited growth on transversally sectioned smooth muscle; (4) neuritic patterning was unaffected by a reduction in migrating ganglionic non-neuronal cells; (5) neurite outgrowth, but not non-neural cell migration, was markedly reduced on myometrial sections from rats treated with estrogen. These results suggest that adult myometrium continues to provide signals allowing the organotypic patterning and growth of sympathetic axons, that estrogen treatment modifies myometrial substrate properties so that it is less supportive for sympathetic neurite growth, and that adult sympathetic neurons retain their ability to recognize substrate-bound cues present in the myometrium. On endometrial sections, neurites formed radially symmetric halos, which were reduced in size on estrogen-treated endometrial substrates. Thus, changes in the neuritogenic capacity of the uterus underlie plasticity in uterine sympathetic nerves, and alterations in substrate-bound factors contribute to the diminished receptivity of the estrogenized uterus to its sympathetic innervation.
Asunto(s)
Estrógenos/farmacología , Neuritas/efectos de los fármacos , Neuritas/metabolismo , Sistema Nervioso Simpático/metabolismo , Útero/efectos de los fármacos , Útero/inervación , Animales , Movimiento Celular/efectos de los fármacos , Endometrio/citología , Endometrio/efectos de los fármacos , Endometrio/metabolismo , Femenino , Miometrio/citología , Miometrio/efectos de los fármacos , Miometrio/metabolismo , Ovariectomía , Ratas , Ratas Wistar , Sistema Nervioso Simpático/citología , Sistema Nervioso Simpático/efectos de los fármacos , Útero/citologíaRESUMEN
Dynamic responses of uterine sympathetic nerves to changes in the circulating levels of sex hormones represent one of the most remarkable examples of physiological plasticity in the adult autonomic nervous system. The density of uterine sympathetic nerves is markedly and irreversibly reduced following puberty, and shows phases of degeneration and regeneration during the natural oestrous cycle. Even more remarkable, uterine sympathetic nerves degenerate during normal pregnancy and regenerate following delivery. Plasticity in uterine sympathetic nerves was initially interpreted as a selective effect of sex hormones on the system of paracervical short adrenergic neurons supplying the uterus. In the last decade, the alternative explanation that sex hormones might alter the ability of the uterine tissue to support its innervation began to be explored and current evidence indicates that oestrogen and pregnancy elicit changes in the neuritogenic properties of the target uterine tissue. In addition, there are indications that sex hormones may also affect the receptivity of uterine-related sympathetic neurons to target-derived signals. Although the nature of these signals is still fragmentary, there is evidence for the contribution of a range of molecules, including neurotrophins, pro-neurotrophins and chemorepulsive signals of the semaphorin family. This review summarizes some general features of plasticity in uterine sympathetic nerves and highlights recent investigations of the cellular and molecular mechanisms underlying this dramatic model of natural plasticity.
Asunto(s)
Ciclo Estral/fisiología , Hormonas Esteroides Gonadales/metabolismo , Plasticidad Neuronal/fisiología , Sistema Nervioso Simpático/metabolismo , Útero/inervación , Útero/fisiología , Animales , Femenino , Humanos , Inflamación/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Regeneración Nerviosa/fisiología , Embarazo , Semaforinas/metabolismo , Sistema Nervioso Simpático/citologíaRESUMEN
Estrogen and glucocorticoids are known to evoke opposing effects on the uterus. We analyzed the effects of dexamethasone (DEX) on uterine sympathetic denervation elicited by short- and long-term exposure to estrogen of intact prepubertal rats. We also studied the effects of DEX on the physiological degeneration of uterine sympathetic nerves at term pregnancy. Changes in innervation were assessed quantitatively by using computer-assisted methods on uterine cryostat tissue sections stained for tyrosine hydroxylase. At 24 h following treatment of prepubertal rats (25 days of age) with 1 microg or 2.5 microg estrogen, marked increases in uterine size and reductions in the percentage nerve area were observed. Co-administration of DEX (4 mg/kg) attenuated both these short-term estrogen-induced effects. Treatment of 19-day-old rats with a single dose of 25 mug estrogen provoked, at 26 days of age, a 54% reduction in the total nerve area. This reduction was abolished by the co-administration of nine doses of DEX (0.5 mg/kg) at 18-26 days of age. Treatment of rats with the same regime of DEX alone increased the total nerve area by 46% of the control values. Studies of control pregnant rats revealed the unexpected presence of intrauterine nerve fibers at term. Treatment of pregnant rats with six doses of DEX (4 mg/kg) at 16-21 days of age had no effects on the density of uterine sympathetic nerves. These results suggest that DEX has growth-promoting effects on immature uterine sympathetic nerves and may antagonize the degenerative effects elicited by long-term exposure to estrogen.
Asunto(s)
Dexametasona/farmacología , Estrógenos/farmacología , Sistema Nervioso Simpático/efectos de los fármacos , Útero/inervación , Glándulas Suprarrenales/efectos de los fármacos , Animales , Antiinflamatorios , Eosinófilos/metabolismo , Femenino , Embarazo , Ratas , Ratas Wistar , Sistema Nervioso Simpático/fisiología , Timo/efectos de los fármacos , Útero/efectos de los fármacos , Útero/crecimiento & desarrolloRESUMEN
Uterine sympathetic innervation undergoes profound remodelling in response to physiological and experimental changes in the circulating levels of sex hormones. It is not known, however, whether this plasticity results from changes in the innervating neurons, the neuritogenic properties of the target tissue or both. Using densitometric immunohistochemistry, we analysed the effects of prepubertal chronic oestrogen treatment (three subcutaneous injections of 20 microg of beta-oestradiol 17-cypionate on days 25, 27 and 29 after birth), natural peripubertal transition and late pregnancy (19-20 days post coitum) on the levels of TrkA and p75 nerve growth factor receptors in uterine-projecting sympathetic neurons of the thoraco-lumbar paravertebral sympathetic chain (T7-L2) identified using the retrograde tracer Fluorogold. For comparative purposes, levels of TrkA and p75 were assessed in the superior cervical ganglion (SCG) following prepubertal chronic oestrogen treatment. These studies showed that the vast majority of uterine-projecting neurons expressed both TrkA and p75. Both prepubertal chronic oestrogen treatment and the peripubertal transition increased the ratio p75 to TrkA in uterine-projecting neurons, whereas pregnancy elicited the opposite effect. Prepubertal chronic oestrogen treatment had no effects on levels of TrkA or p75 in sympathetic neurons of the SCG. Taken together, our data suggest that neurotrophin receptor-mediated events may contribute to regulate sex hormone-induced plasticity in uterine sympathetic nerves, and are in line with the idea that, in vivo, plasticity in uterine nerves involves changes in both the target and the innervating neurons.
Asunto(s)
Plasticidad Neuronal/fisiología , Neuronas/química , Receptor trkA/fisiología , Receptores de Factor de Crecimiento Nervioso/fisiología , Sistema Nervioso Simpático/fisiología , Útero/inervación , Animales , Estradiol/farmacología , Femenino , Inmunohistoquímica/métodos , Microscopía Fluorescente , Embarazo , Ratas , Ratas Wistar , Receptor de Factor de Crecimiento Nervioso , Receptor trkA/análisis , Receptores de Factor de Crecimiento Nervioso/análisis , Maduración SexualRESUMEN
El revestimiento epitelial de las vías respiratorias, conjuntamente con el transporte mucociliar, forman parte de la primera línea de defensa del aparato respiratorio. Las condiciones que alteran la integridad epitelial o afectan la eficiencia del transportemucociliar conducen o favorecen la recurrencia de la enfermedad respiratoria. En el presente trabajo reportamos los resultados obtenidos del estudio por histología de alta resolución y microscopía electrónica de barrido del epitelio nasal de 33 pacientes respiratorios crónicos. Todos las biopsias de cornete inferior analizadas presentaron algún tipo de anomalía epitelial, no detectándose en ningún caso el epitelio seudoestratificado cilíndrico ciliado que normalmente reviste las vías respiratorias. En 17 de los 33 pacientes se reconocieron epitelios ciliados con distintos grados de atipía, mientras que en los 16 restantes se observó la sustitución total de las células ciliadas por tipos celulares no ciliados, tales como células basales, células caliciformes y células escamosas. En 27 por ciento de los casos las alteraciones epiteliales del cornete inferior se presentaron en pacientes que portaban afecciones ciliares primarias, mientras que en 52 por ciento se presentaron en pacientes que mostraban alteraciones ciliares inespecíficas o ausencia de cilias. En 21 por ciento de los casos se detectaron afecciones epiteliales en pacientes que tenían una ultraestructura ciliar normal. Los datos obtenidos confirman el concepto de que las alteraciones epiteliales pueden presentarse a consecuencia de los desórdenes ciliares primarios o secundarios, y resultar también de la injuria prolongada provocada por diversas enfermedades respiratorias crónicas, tales como neumonías, bronquitis, rinitis, sinusitis y asma. Dado que la inflamación e infección respiratoria recurrente retrasa la regeneración del epitelio normal, la detección precoz de estas alteraciones histopatológicas epiteliales y sus afecciones ciliares asociadas es clave para evitar la instalación de formas epiteliales no ciliadas irreversibles. (AU)
Asunto(s)
Humanos , Mucosa Nasal/patología , Microscopía Electrónica de Rastreo , Enfermedades Respiratorias , Enfermedad CrónicaRESUMEN
El revestimiento epitelial de las vías respiratorias, conjuntamente con el transporte mucociliar, forman parte de la primera línea de defensa del aparato respiratorio. Las condiciones que alteran la integridad epitelial o afectan la eficiencia del transportemucociliar conducen o favorecen la recurrencia de la enfermedad respiratoria. En el presente trabajo reportamos los resultados obtenidos del estudio por histología de alta resolución y microscopía electrónica de barrido del epitelio nasal de 33 pacientes respiratorios crónicos. Todos las biopsias de cornete inferior analizadas presentaron algún tipo de anomalía epitelial, no detectándose en ningún caso el epitelio seudoestratificado cilíndrico ciliado que normalmente reviste las vías respiratorias. En 17 de los 33 pacientes se reconocieron epitelios ciliados con distintos grados de atipía, mientras que en los 16 restantes se observó la sustitución total de las células ciliadas por tipos celulares no ciliados, tales como células basales, células caliciformes y células escamosas. En 27 por ciento de los casos las alteraciones epiteliales del cornete inferior se presentaron en pacientes que portaban afecciones ciliares primarias, mientras que en 52 por ciento se presentaron en pacientes que mostraban alteraciones ciliares inespecíficas o ausencia de cilias. En 21 por ciento de los casos se detectaron afecciones epiteliales en pacientes que tenían una ultraestructura ciliar normal. Los datos obtenidos confirman el concepto de que las alteraciones epiteliales pueden presentarse a consecuencia de los desórdenes ciliares primarios o secundarios, y resultar también de la injuria prolongada provocada por diversas enfermedades respiratorias crónicas, tales como neumonías, bronquitis, rinitis, sinusitis y asma. Dado que la inflamación e infección respiratoria recurrente retrasa la regeneración del epitelio normal, la detección precoz de estas alteraciones histopatológicas epiteliales y sus afecciones ciliares asociadas es clave para evitar la instalación de formas epiteliales no ciliadas irreversibles.
Asunto(s)
Humanos , Enfermedades Respiratorias , Mucosa Nasal , Enfermedad Crónica , Microscopía Electrónica de RastreoRESUMEN
OBJECTIVE: To determine the association between long-term exposure to wood smoke from cooking and lung adenocarcinoma in non-smoking Mexican women. METHODS: We reviewed records of hospitalized patients at a chest referral hospital in Mexico City and identified 113 histologically proven lung adenocarcinoma cases in non-smoking women. Four control groups of non-smoking women were also selected: 99 patients with pulmonary tuberculosis (PTB), 110 with interstitial lung disease (ILD), 64 with miscellaneous pulmonary conditions (MISC), and the three control groups combined (COMB) (n = 273). RESULTS: Exposure was assessed on the basis of questionnaire responses at the time of hospital admission. Exposure to wood smoke for more than 50 years, but not for shorter periods, was associated with lung cancer after adjusting for age, education, socio-economic status and environmental tobacco smoke (ETS) exposure. Adjusted odds ratios from the multivariable logistic regression models were 1.4 (95%CI 0.6-2.0) for cases vs. TB controls, 1.9 (95%CI 0.9-4.0) for cases vs. ILD controls, 2.6 (95%CI 1.0-6.3) for cases vs. MISC controls and 1.9 (95%CI 1.1-3.5) for cases vs. COMB controls. CONCLUSION: These findings suggest that long-term exposure to wood smoke from cooking may contribute to the development of lung cancer.
Asunto(s)
Adenocarcinoma/etiología , Exposición por Inhalación , Neoplasias Pulmonares/etiología , Humo/efectos adversos , Madera , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Casos y Controles , Culinaria , Femenino , Humanos , México , Persona de Mediana Edad , Factores de TiempoRESUMEN
La disquinesia ciliar primaria es una enfermedad congénita que aúna un grupo heterogéneo de condiciones patológicas que se caracterizan por la presencia de alteraciones microanatómicas o funcionales, o ambas, en las cilias y los flagelos espermáticos. Las manifestaciones clínicas de este desorden son variadas y se caracterizan por la aparición temprana de infección recurrente de las vías auditivo-respiratorias. Además de los determinantes genéticos, la propia infección e inflamación recurrente -así como algunos agentes ambientales- pueden provocar alteraciones ciliares. Estas alteraciones son, sin embargo, de tipo inespecífico y conducen a la llamada disquinesia ciliar secundaria. En el presente trabajo se describen los resultados obtenidos del estudio de 40 biopsias de mucosa nasal y bronquial provenientes de 33 pacientes sospechosos de portar alguna de estas formas de disquinesia ciliar, las cuales fueron remitidas por los médicos tratantes al Laboratorio de Biología Celular del Instituto de Investigaciones Biológicas Clemente Estable para su estudio por microscopía electrónica de transmisión. En su conjunto estos estudios mostraron que 9 de los pacientes (27 por ciento) presentaban alteraciones ultra estructurales ciliares compatibles con el diagnóstico de disquinesia ciliar primaria. Tres de ellos tenían situs inversus totalis. El 52 por ciento de los pacientes presentó alteraciones ciliares secundarias. A pesar de sus manifestaciones clínicas, 21 por ciento de los pacientes restantes mostró una ultraestructura ciliar normal. En su conjunto, los resultados obtenidos en la población estudiada apuntan a confirmar la alta frecuencia con que se presentan las alteraciones ciliares en los pacientes respiratorios crónicos. Teniendo en cuenta que el diagnóstico precoz de esta enfermedad es esencial para prevenir el desarrollo de lesiones respiratorias irreversibles, se destaca la importancia de indicar el examen ultraestructural de las cilias en los pacientes portadores de situs inversus y sus hermanos, así como en los individuos afectados de enfermedad respiratoria crónica severa de inicio precoz y carentes de diagnóstico etiológico. (AU)
Asunto(s)
Trastornos de la Motilidad Ciliar/diagnóstico , Microscopía Electrónica , Síndrome de Kartagener/diagnósticoRESUMEN
La disquinesia ciliar primaria es una enfermedad congénita que aúna un grupo heterogéneo de condiciones patológicas que se caracterizan por la presencia de alteraciones microanatómicas o funcionales, o ambas, en las cilias y los flagelos espermáticos. Las manifestaciones clínicas de este desorden son variadas y se caracterizan por la aparición temprana de infección recurrente de las vías auditivo-respiratorias. Además de los determinantes genéticos, la propia infección e inflamación recurrente -así como algunos agentes ambientales- pueden provocar alteraciones ciliares. Estas alteraciones son, sin embargo, de tipo inespecífico y conducen a la llamada disquinesia ciliar secundaria. En el presente trabajo se describen los resultados obtenidos del estudio de 40 biopsias de mucosa nasal y bronquial provenientes de 33 pacientes sospechosos de portar alguna de estas formas de disquinesia ciliar, las cuales fueron remitidas por los médicos tratantes al Laboratorio de Biología Celular del Instituto de Investigaciones Biológicas Clemente Estable para su estudio ppor microscopía electrónica de transmisión. En su conjunto estos estudios mostraron que 9 de los pacientes (27 por ciento) presentaban alteraciones ultra estructurales ciliares compatibles con el diagnóstico de disquinesia ciliar primaria. Tres de ellos tenían situs inversus totalis. El 52 por ciento de los pacientes presentó alteraciones ciliares secundarias. A pesar de sus manifestaciones clínicas, 21 por ciento de los pacientes restantes mostró una ultraestructura ciliar normal. En su conjunto, los resultados obtenidos en la población estudiada apuntan a confirmar la alta frecuencia con que se presentan las alteraciones ciliares en los pacientes respiratorios crónicos. Teniendo en cuenta que el diagnóstico precoz de esta enfermedad es esencial para prevenir el desarrollo de lesiones respiratorias irreversibles, se destaca la importancia de indicar el examen ultraestructural de las cilias en los pacientes portadores de situs inversus y sus hermanos, así como en los individuos afectados de enfermedad respiratoria crónica severa de inicio precoz y carentes de diagnóstico etiológico.
Asunto(s)
Trastornos de la Motilidad Ciliar , Microscopía Electrónica , Síndrome de Kartagener/diagnósticoRESUMEN
In the present study we investigated the effects of infantile/prepubertal chronic oestrogen treatment, chemical sympathectomy with guanethidine and combined sympathectomy and chronic oestrogen treatment on developing sensory nerves of the rat uterus. Changes in sensory innervation were assessed quantitatively on uterine cryostat tissue sections stained for calcitonin gene-related peptide (CGRP). Uterine levels of NGF protein, using immunohistochemistry and ELISA, and mRNA, using Northern blots and in situ hybridization, were also measured. Finally, levels of TrkA NGF receptor in sensory neurons of T13 and L1 dorsal root ganglia (DRG), which supply the uterus, were assessed using densitometric immunohistochemistry. These studies showed that: (1) chronic oestrogen treatment led to an 83% reduction in the intercept density of CGRP-immunoreactive nerves; (2) sympathectomy had no effect on the density of uterine sensory nerves or on the pattern of oestrogen-induced changes; (3) NGF mRNA and protein increased following sympathectomy or chronic oestrogen treatment; and (4) oestrogen produced increased intensity of labelling (28%) for TrkA receptors in small-diameter sensory neurons, but decreased labelling (13%) in medium-sized neurons, which represent the large majority of the DRG neurons supplying the upper part of the uterine horn. Contrary to expectations, increased levels of NGF after sympathectomy and oestrogen treatment did not lead to increased sensory innervation of the uterus. The possibility that alterations in neuronal levels of TrkA contribute to the lack of response of uterine sensory nerves to the oestrogen-induced increase in NGF levels is discussed.
Asunto(s)
Estrógenos/farmacología , Factor de Crecimiento Nervioso/metabolismo , Plasticidad Neuronal , Neuronas Aferentes/fisiología , Receptor trkA/metabolismo , Útero/inervación , Adrenérgicos/farmacología , Animales , Péptido Relacionado con Gen de Calcitonina/metabolismo , Ensayo de Inmunoadsorción Enzimática , Estrógenos/metabolismo , Femenino , Ganglios Espinales/anatomía & histología , Ganglios Espinales/citología , Guanetidina/farmacología , Inmunohistoquímica , Hibridación in Situ , Neuronas Aferentes/efectos de los fármacos , Neuronas Aferentes/metabolismo , ARN Mensajero/biosíntesis , Ratas , Ratas Wistar , Simpatectomía Química , Útero/crecimiento & desarrollo , Útero/metabolismoRESUMEN
Chronic administration of oestrogen to rats during the infantile/prepubertal period provokes, at 28 days of age, complete loss of noradrenaline-labelled intrauterine sympathetic nerves. It is not known whether oestrogen inhibits the growth or causes the degeneration of developing uterine sympathetic nerves, or whether the uterus recovers its innervation following cessation of infantile/prepubertal oestrogen treatment. In the present study, we analysed the time-course of the effects of oestrogen on the development of uterine sympathetic nerves in the rat, using histochemical methods. In addition, the pattern of sympathetic reinnervation of the uterus of intact and ovariectomised females was assessed 3 and 6 months after cessation of chronic oestrogen treatment. The ability of sympathetic nerves to reinnervate the oestrogenized uterine tissue was assessed in intraocular transplants of uterine myometrium into ovariectomised host rats. Early exposure to oestrogen did not inhibit the approach of sympathetic nerves to the uterus, but prevented the normal growth and maturation of intrauterine sympathetic fibres and abolished the innervation that reached the organ before initiation of treatment. Three or six months following cessation of oestrogen treatment, most of the sympathetic nerves were restricted to the mesometrium and mesometrial entrance, whereas intrauterine innervation remained persistently depressed as a consequence of a sustained oestrous-like state provoked by ovarian dysfunction (polycystic ovary). An organotypic regrowth of uterine sympathetic nerves was observed in ovariectomised infantile/prepubertal oestrogen-treated animals. After 5 weeks in oculo, the innervation of oestrogenized myometrial transplants was reduced by 50%, and substantial changes in the pattern of reinnervation were observed. In control transplants, 86% of the nerves were terminal varicose myometrial and perivascular nerve fibres, whereas 14% were preterminal nerve bundles. In oestrogenized myometrial transplants, 83% of the noradrenaline-labelled intercepting nerves were enlarged preterminal bundles and only 17% were terminal fibres. These results indicate that the oestrogenized myometrium is unattractive for sympathetic nerves and inhibits organotypic sympathetic reinnervation.
Asunto(s)
Estrógenos/farmacología , Miometrio/efectos de los fármacos , Sistema Nervioso Simpático/fisiología , Animales , Animales Recién Nacidos , Estrógenos/sangre , Femenino , Inmunohistoquímica , Iris/inervación , Microscopía Fluorescente , Miometrio/inervación , Miometrio/trasplante , Fibras Nerviosas/fisiología , Regeneración Nerviosa , Norepinefrina/metabolismo , Ovariectomía/métodos , Progesterona/sangre , Ratas , Ratas Wistar , Estadísticas no Paramétricas , Sistema Nervioso Simpático/efectos de los fármacos , Factores de Tiempo , Útero/efectos de los fármacos , Útero/inervaciónRESUMEN
Oestrogen is a key factor in the remodelling of uterine sympathetic nerves during puberty and the oestrous cycle; these nerves are influenced by changes in their target uterine tissue. The magnitude of oestrogen-induced responses might however be influenced by the maturation stage of sympathetic nerve fibres, the age of the neurons and/or the developmental state of the uterus. We have therefore compared the sympathetic innervation of the uterus following chronic oestrogen treatment of infantile/prepubertal and young adult intact and ovariectomised rats. Treatment of infantile/prepubertal rats resulted in the complete loss of intrauterine noradrenaline (NA)-labelled sympathetic nerves and a marked reduction in the total NA content in the uterine horn. Chronic treatment of young adult rats had little effect. To examine whether the age of the neurons or the degree of development of the uterus determined responsiveness of nerves to oestrogen, we assessed the effects of oestrogen on the sympathetic reinnervation of intraocular transplants of young adult uterine myometrium into ovariectomised adult host rats. Early treatment (10 days post-transplantation) resulted in less sympathetic innervation than late treatment (30 days post-transplantation). Measurements of nerve growth factor (NGF) levels in the uterine horn of control rats before and after puberty and following infantile/prepubertal chronic oestrogen treatment and acute oestrogen treatment of young adult rats revealed a coordinated increase between the growth of the uterus and NGF protein levels. Thus, developing and recently regrown sympathetic nerves are more susceptible to oestrogen-induced changes in the uterus than mature nerves, differential susceptibility is not related to the age of the neurons or the developmental state of the uterus and changes in NGF protein do not account for the differential susceptibility of developing and mature uterine sympathetic nerve fibres to oestrogen. Growing sympathetic fibres are more vulnerable to oestrogen than mature fibres and nerve fibres that have been in contact for longer periods with their target become less susceptible to oestrogen.
Asunto(s)
Estrógenos/farmacología , Sistema Nervioso Simpático/crecimiento & desarrollo , Útero/inervación , Animales , Estrógenos/metabolismo , Femenino , Factor de Crecimiento Nervioso/análisis , Factor de Crecimiento Nervioso/biosíntesis , Norepinefrina/análisis , Norepinefrina/biosíntesis , Ratas , Ratas Wistar , Sistema Nervioso Simpático/anatomía & histología , Sistema Nervioso Simpático/efectos de los fármacos , Útero/anatomía & histología , Útero/efectos de los fármacosRESUMEN
The innervation of the uterus is remarkable in that it exhibits physiological changes in response to altered levels in the circulating levels of sex hormones. Previous studies by our group showed that chronic administration of estrogen to rats during the infantile/prepubertal period provoked, at 28 days of age, an almost complete loss of norepinephrine-labeled sympathetic nerves, similar to that observed in late pregnancy. It is not known, however, whether early exposure to estrogen affects uterine cholinergic nerves. Similarly, it is not known to what extent development and estrogen-induced responses in the uterine cholinergic innervation are affected by the absence of sympathetic nerves. To address this question, in this study we analyzed the effects of infantile/prepubertal chronic estrogen treatment, chronic chemical sympathectomy with guanethidine, and combined sympathectomy and chronic estrogen treatment on developing cholinergic nerves of the rat uterus. Cholinergic nerves were visualized using a combination of acetylcholinesterase histochemistry and the immunohistochemical demonstration of the vesicular acetylcholine transporter (VAChT). After chronic estrogen treatment, a well-developed plexus of cholinergic nerves was observed in the uterus. Quantitative studies showed that chronic exposure to estrogen induced contrasting responses in uterine cholinergic nerves, increasing the density of large and medium-sized nerve bundles and reducing the intercept density of fine fibers providing myometrial and perivascular innervation. Estrogen-induced changes in the uterine cholinergic innervation did not appear to result from the absence/impairment of sympathetic nerves, because sympathectomy did not mimic the effects produced by estrogen. Estrogen-induced responses in parasympathetic nerves are discussed, considering the direct effects of estrogen on neurons and on changes in neuron-target interactions.
Asunto(s)
Fibras Colinérgicas/metabolismo , Estradiol/farmacología , Proteínas de Transporte de Membrana , Sistema Nervioso Simpático/metabolismo , Útero/inervación , Proteínas de Transporte Vesicular , Acetilcolinesterasa/metabolismo , Animales , Animales Recién Nacidos , Proteínas Portadoras/metabolismo , Fibras Colinérgicas/enzimología , Fibras Colinérgicas/ultraestructura , Estradiol/análogos & derivados , Femenino , Guanetidina , Histocitoquímica , Ratas , Ratas Wistar , Simpatectomía Química , Sistema Nervioso Simpático/crecimiento & desarrollo , Sistema Nervioso Simpático/ultraestructura , Simpaticolíticos , Útero/crecimiento & desarrollo , Proteínas de Transporte Vesicular de AcetilcolinaRESUMEN
Previous studies have shown that chronic administration of oestrogen during postnatal rat development dramatically reduces the total content of noradrenaline in the uterine horn, abolishes myometrial noradrenergic innervation and reduces noradrenaline-fluorescence intensity of intrauterine perivascular nerve fibres. In the present study we analysed if this response is due to a direct and selective effect of oestrogen on the uterine noradrenaline-containing sympathetic nerves, using the in oculo transplantation method. Small pieces of myometrium from prepubertal rats were transplanted into the anterior eye chamber of adult ovariectomised host rats. The effect of systemic chronic oestrogen treatment on the reinnervation of the transplants by noradrenaline-containing sympathetic fibres from the superior cervical ganglion was analysed on cryostat tissue sections processed by the glyoxylic acid technique. In addition, the innervation of the host iris was assessed histochemically and biochemically. The histology of the transplants and irises was examined in toluidine blue-stained semithin sections. These studies showed that after 5 wk in oculo, the overall size of the oestrogen-treated transplants was substantially larger than controls, and histology showed that this change was related to an increase in the size and number of smooth muscle cells within the transplant. Chronic oestrogen treatment did not provoke trophic changes in the irideal muscle. Histochemistry showed that control transplants had a rich noradrenergic innervation, associated with both myometrium and blood vessels. Conversely, in oestrogen-treated transplants only occasional fibres were recognised, showing a reduced NA fluorescence intensity. No changes in the pattern and density of innervation or in the total content of noradrenaline of the host irises were detected after chronic exposure to oestrogen. We interpreted these results to indicate that the effects of oestrogen on uterine noradrenaline-containing sympathetic nerves are neither selective or direct, but result from an interaction between sympathetic nerve fibres with the oestradiol-primed uterine tissue. A potential effect of oestrogen on the neurotrophic capacity of the uterus is discussed.