[Effect of subglottic air insufflation on subglottic pressure during swallowing]. / Efecto de la insuflación de aire subglótico sobre la presión subglótica durante la deglución.

OBJECTIVE: To determine whether there are differences between subglottic pressure during swallowing with and without air insufflation via a subglottic catheter in tracheostomized patients. DESIGN: A prospective, randomized cross-over study was made. SETTING: Adult Intensive Care Units. PARTICIPANTS: Patients requiring mechanical ventilation and tracheostomy with a subglottic catheter, and with tolerance to deflation of the balloon and a speaking valve placed over the opening of the tracheostomy tube. INTERVENTIONS: Subglottic pressure was measured during swallowing of a thickened solution with and without the delivery of airflow through the subglottic catheter. MAIN VARIABLE: Subglottic pressure during swallowing. RESULTS: Twelve out of 14 patients showed higher subglottic pressure values during swallowing with air insufflation. Two patients showed no differences between both conditions. Median (Med) values of subglottic pressure for the first, second and third swallow were 5, 4 and 4.5 cmH2O (Med 4.5 cmH2O) without air insufflation, and 8, 5.5 and 7.5 cmH2O (Med 5.5 cmH2O) with air insufflation, respectively (Wilcoxon, Z=-3.078; p=.002). CONCLUSION: In a group of tracheostomized patients, air insufflation via a subglottic catheter increased subglottic pressure levels measured during swallowing.

How to measure the increase in elastic system fibres in the lamina propria of the uterine cervix of pregnant rats.

As the uterus enlarges to accommodate the growing fetus during pregnancy, the cervix behaves essentially as a barrier. During ripening and delivery, it needs to become soft and distensible to allow dilation and the passage of the conceptus. As the transformations of the collagen-containing fibres are known to be essential for ripening and delivery, it has been hypothesized that the elastic system fibres, owing to their intrinsic mechanical properties (reversible extensibility), could be involved in the shape-recovering process immediately after delivery. In sections stained by Weigert's resorcin-fuchsin (with previous oxidation), we describe the elastic system fibres in the lamina propria of the rat uterine cervix. They are distributed following different patterns when in the endocervix or in the ectocervical-vaginal region. A third distinctive pattern (named the 'elastic tendon') is described here for the first time in the uterine-cervical transition. A special morphometrical protocol has been designed in order to overcome problems during the quantification process. Using the so-called intercept counting method, it was possible to demonstrate that the elastic system fibres are increased in the cervix at the end of pregnancy. They may be involved in the immediate shape-recovering of the cervix after delivery as well as in helping to strengthen the anchoring of the epithelium to the lamina propria, thus minimizing birth trauma.

Phenotypic modulation of fibroblastic cells in the mucous layer of the human uterine cervix at term.

The uterine cervix is a dynamic structure with a high capacity to adapt to different, even opposing, roles during the sequence of physiological events of gestation (for example, acting as a barrier to retain the fetus during pregnancy and dilating to allow delivery at term). Histoarchitectural changes of the uterine cervix allow its successful adaptation. The aim of this study was to investigate whether fibroblastic cell plasticity, described in the lamina propria of the rat uterine cervix at term, could be observed in women too. Biopsy specimens of non-pregnant and intrapartum human cervices were studied under the transmission electron microscope, and cytoskeletal differentiation markers were identified by immunohistochemistry under the light microscope. Desmin-positive cells were present in the mucous layer of the cervix during labour. These cells displayed cytoplasmic processes (typical of myofibroblasts) that also stained positively for vimentin. The main ultrastructural features for defining the myofibroblast under the electron microscope were also observed in these cells. However, cervices of non-pregnant women contained resident fibroblasts at the same location. Examination of the differentiation repertoire of fibroblastic cells in the mucous layer of the uterine cervix resulted in the characterization of myofibroblasts at term. The implications of the plasticity of fibroblastic-myofibroblastic cells in the physiological changes displayed in the uterine cervix during pregnancy, labour and postpartum involution require further investigation.

Characterization of fibroblastic cell plasticity in the lamina propria of the rat uterine cervix at term.

Different organs contain fibroblasts with specific features and functions, indicating the complexity of fibroblast biology. In the rat cervical stroma, fibroblasts are preferentially located in the fibrous ring that surrounds the mucous layer. The purpose of this study was to investigate the morphological features and immunophenotype of fibroblastic cells of the uterine cervix in cycling, pregnant, and postpartum rats. Expression of the cytoskeletal proteins desmin, vimentin, and alpha-smooth muscle actin (alpha-SMA) were studied by immunohistochemistry. The optical density of immunohistochemical staining was quantified by image analysis. The ultrastructural features of fibroblastic cells were observed under transmission electron microscopy. Cervical fibroblastic cells always expressed vimentin and desmin but never alpha-SMA. During the first half of pregnancy (Day 5 [D5] to D14), desmin intensity values were similar to those of cycling and postpartum fibroblasts. In contrast, a strong expression of desmin was found from D15 to D22, with maximal expression at term (D23). Immunohistochemical expression for vimentin was constant throughout pregnancy and showed no differences with cycling and postpartum uterine cervices. Stromal cells from cycling and early pregnant rats displayed ultrastructural features characteristic of typical fibroblasts. In contrast, at the end of pregnancy, fibroblasts differentiated and showed increased secretory characteristics, reaching the ultrastructural features of a myofibroblast. Based on the differential expression of desmin and the electron microscopic observations, the foregoing results showed a modulation of the fibroblastic phenotype in the uterine cervix during pregnancy. To our knowledge, this is the first report that addresses the presence of myofibroblasts derived from resident fibroblasts in the fibrous ring of the rat uterine cervix. Fibroblastic-myofibroblastic cell plasticity may have implications in the physiological changes displayed in the uterine cervix during pregnancy, parturition, and postpartum involution.

Differential distribution of the fibres of the collagenous and elastic systems and of glycosaminoglycans in the rat pubic joint.

The pubic joint of male and female rats was studied at the light- and electron microscopical levels using methods that selectively disclose the extracellular matrix fibres and glycosaminoglycans. The interpubic tissue showed no difference between sexes (including pregnant and intrapartum females). The medial ends of the pubic bones were covered by articular caps of hyaline cartilage that blended in the midline. The whole articular cartilage was covered dorsally and ventrally (as well as craneally and caudally) by a typical perichondrium. The differential distribution of the fibres of the collagenous and elastic systems in the pubic joint agreed with the results reported in the literature for other rat cartilages. Collagen fibres, composed mainly of type-I collagen, were localised to the fibrous perichondrium and bone. Type-II collagen was localised to the central nucleus of hyaline cartilage, whereas reticulin fibres (rich in type-III collagen) were found in the adventitial loose connective tissue adherent to the most superficial layer of the perichondrium. The central nucleus of hyaline cartilage possessed the two types of elastic-related fibres: elaunin fibres were localised mainly to the chondrogenic layer of the perichondrium, whereas oxytalan fibres were found in the matrix that surrounded the chondrocytes. The bulk of the glycosaminoglycans present in the pubic joint cartilage corresponded to hyaluronic acid and chondroitin sulphate. The propriety of classification of the rat pubic joint as a true synchondrosis (instead of symphysis), and the fact that the unaltered pelvis of the rat seems to be adequate for normal parturition, are discussed.

Relaxin has a minor role in rat mammary gland growth and differentiation during pregnancy.

Growth and differentiation of mammary gland is associated with numerous hormones and a variety of cell-cell, cell-matrix interactions. This study addressed the role of relaxin (Rlx) on these processes. Morphologic and biochemical changes that occur throughout the second half of pregnancy are reported. Temporal patterns and spatial distributions of markers useful to evaluate proliferation, secretion, and collagen remodeling were established. To evaluate the role of Rlx, an ablation/replacement animal model was used. Considering Rlx secretion pattern, two periods were selected: d 11 through d 13, and d 20 through d 23. In the stroma, the extracellular compartment showed changes associated with the lack of Rlx. Collagen remodeling within the lobuloalveolar structure, measured by a significant increase in collagen birefringence, decreased at d 12, d 21, and d 22. Parenchymal structures were less sensitive to the absence of Rlx than stroma. Epithelial cell proliferation was lower in Rlx-deficient rats only at d 12, and alpha-lactalbumin expression decreased at d 21 and d 22. Both lobuloalveolar diameter and percentage of area occupied by these structures showed no changes. In the absence of Rlx, some of the studied markers showed statistically significant differences in scattered days; these do not make clear trends. No differences were found on d 23 on any of the studied parameters suggesting that compensatory mechanisms might be activated to overcome the effects of the absence of Rlx. Unlike the critical role of Rlx either in uterine cervix dilation or in nipple development during rat pregnancy, Rlx had a minor role in growth and differentiation of rat mammary gland.

Architectural remodelling in acute and chronic interstitial lung disease: fibrosis or fibroelastosis?

AIMS: Pulmonary fibrosis in acute and chronic lung disease has been much investigated, but little attention has been directed at the elastic tissue in these situations. Our aim was to verify whether elastic deposition accompanies collagen deposition in the repairing process of acute and chronic lung injury. METHODS AND RESULTS: We measured, by image analysis, the content of fibres of the collagenous and elastic systems of the alveolar septum in histological slides sampled from autopsied lungs, using the picrosirius-polarization method and Weigert's resorcin-fuchsin stain, respectively. Five groups were studied: I, 10 normal patients; II, 10 patients with cardiogenic pulmonary oedema; III, 23 adult respiratory distress syndrome (ARDS) patients in the early phase; IV, 14 ARDS patients in the late fibroproliferative phase; and V, 10 idiopathic pulmonary fibrosis patients. The first two groups were used as controls. The content of fibres of the collagenous and elastic systems was significantly increased in groups IV and V as compared to the other groups. CONCLUSIONS: Our results indicate that deposition of elastic system fibres is present in the fibroproliferative phase of ARDS and in usual interstitial pneumonia and suggest that this event may contribute to the alveolar mechanical dysfunction and remodelling that occur in acute and chronic interstitial lung disease.

Collagen-affecting drugs impair regeneration of teleost tail fins.

Regenerating tail fins were studied in two species of teleosts, Tilapia rendalli and Cyprinus carpio, treated with indomethacin, aspirin, dexamethasone, penicillamine, and beta-aminoproprionitrile, drugs known to disrupt collagen metabolism in mammals. Collagen was studied under the light microscope by the Picrosirius-polarization method and also under the electron microscope. In general, these drugs disturbed the deposition and organization of collagen fibrils leading to abnormally thin or practically absent lepidotrichia and actinotrichia, and also to disorganized fibrous connective tissue. The resulting disorganization of the collagenous scaffolding of the regenerating dermoskeleton was probably responsible for a secondary effect on blastema distalization and on the general fin ray patterning that were also observed. The foregoing observations suggest that the stromal histoarchitecture of the regenerate plays a vital role in fin regeneration and indicate that these drugs may be useful in studying the extracellular matrix-cell interactions at the cellular and molecular level. In addition, the present findings provide a basis for developing different biological models by using teleost fin regeneration.

[Distribution of financial resources according to the productivity (determined by bibliometry) in the Medical Research Laboratories of the Hospital das Clínicas da Faculdade de Medicina da Universidad de São Paulo (Brazil)]. / Distribución de los recursos financieros de acuerdo con la productividad (determinada por bibliometría) en los Laboratorios de Investigación Médica del Hospital de Clínicas, de la Facultad de Medicina de la Universidad de San Pablo (Brasil).

A challenge that research managers have to face is how to "reallocate" agency budgets in order to bring them in line with the results of performance reviews. Research policies must develop a strategic plan describing their goals, devise yardsticks to measure their progress, and tie that performance to allocate research funds with some degree of priority. Though Brazil already has a substantial presence in world's science, scientific enterprise must be used to strengthen it. The first step should be to raise standards in Brazilian science by concentrating the resources on supporting excellence. A strategy to judge biomedical research productivity should include tactics to disclose whether the resulting publications have appeared in the field's most respected, peer-reviewed journals. A pilot project to road-test the above-discussed ideas on performance measurements was conducted at the Laboratories of Medical Research (Clinical Hospital, University of São Paulo School of Medicine). These Laboratories perform a vast proportion of biomedical research at the country's largest University. This article illustrates that confidence in fairness and consistency with which funds are now being allocated has helped to improve productivity, thus showing that this strategy is fruitful.

Estrogen and progesterone modulation of eosinophilic infiltration of the rat uterine cervix.

Ripening of the rat cervix involves widespread collagenolysis that follows an eosinophilic leukocyte infiltration. The hormonal control of these events is not well understood. The aims of this study were to investigate the mechanism through which progesterone (P) and 17beta-estradiol (E(2)) modulate eosinophilic invasion and to determine if this event is protein synthesis mediated. Cervical eosinophilic invasion was measured in intact rats during the second half of pregnancy and compared with values from ovariectomized (O) pseudopregnant (PSP) rats treated with P and E(2) in doses that mimicked the levels of pregnancy. Other O-PSP rats were treated with an E(2) antagonist (tamoxifen) and the antiprogestin RU-486. To study the role of protein synthesis in eosinophilic invasion of the cervix, rats were treated with actinomycin-D (an inhibitor of mRNA synthesis), and animals were sacrificed on D21 or D22 to evaluate eosinophilic invasion. Rats treated with E(2) showed high levels of infiltration and tamoxifen blocked this E(2) effect. On the other hand, P antagonized the stimulatory effects of E(2) on eosinophilic invasion, however when the P and E(2) treated rats were injected with RU-486 the inhibitory effect of P was reversed. In intact pregnant rats a sharp rise in eosinophilic infiltration was detected on D23, 20 h after the fall of serum P. Finally, E(2) treated rats injected with actinomycin-D had no invasion of eosinophils. In conclusion, the estrogen-triggered eosinophil invasion is affected by the classic estrogen receptor antagonist tamoxifen and by the mRNA synthesis blocker actinomycin-D suggesting a genomic action of E(2). Furthermore, the estrogen effect is blocked by P and this inhibition is reversed by RU-486.

Morphometric study of the regeneration of individual rays in teleost tail fins.

The results obtained using morphometric variables which describe fin ray regeneration patterns are reported for individual fin ray amputations in the goldfish (Carassius auratus) and zebrafish (Brachydanio rerio). Classical and updated experiments are compared to verify previous morphogenetic models of cell tractions (Oster et al. 1983) or epidermis-mesenchyme induction (Saunders et al. 1959) applied to the limb of other vertebrates. Position-dependent patterns within the fin of Carassius auratus are analysed under a comparative protocol using morphometric methods. Conditions in which the apical epidermis is separated from blastema may differentiate small fin rays, thus suggesting this epidermis is involved in blastemal formation. Blastemal cells differentiating as lepidotrichia forming cells (LFCs) may also be related to morphological changes in covering epidermis. Long-range interactions from neighbouring fin ray blastemas or short-range interactions within the blastema, may be postulated through the analysis of segmentation.

Use of the Picrosirius-polarization method to age fibrotic lesions in the hepatic granulomas produced in experimental murine schistosomiasis.

The state of collagen aggregation in hepatic granulomas produced in experimental murine schistosomiasis and harvested at different phases of the disease was estimated by staining histological sections in Picrosirius and studying them under polarized light. Two different types of collagenous fibres appeared successively during granuloma evolution, the first population, of thin, weakly birefringent, greenish fibres being replaced by thicker collagen fibres which display a stronger yellow-red birefringence. A simple morphometric technique (a point-counting procedure) was used to quantify the two distinct populations of collagenous fibres. Curve-fitting procedures were then employed, using the state of collagen aggregation as the dependent variable and time as the independent variable. The results showed that there is a progressive organization of the collagenous scaffolding of hepatic granulomas, such that quantitative study of the degree of collagen organization allows an adequate determination of the age of each granuloma. The combination of the Picrosirius-polarization method with simple and efficient morphometric approaches will clearly provide useful information on the natural history of schistosomal granuloma scarring.

Relation between interstitial myocardial collagen and the degree of clinical impairment in Chagas' disease.

Cardiac performance is negatively associated with interstitial collagen in Chagas' cardiomyopathy. The magnitude of dysfunction is related to the degree of collagen, and this association seems to exhibit a threshold above it if definite cardiac deterioration occurs.

Differential distribution of elastic system fibers in control and bronchoconstricted intraparenchymatous airways in the guinea-pig lung.

The elastic system fibers were studied at the light microscopic level by using Weigert's resorcin-fuchsin method after oxidation. This study was designed to describe the distribution of these fibers in intrapulmonary guinea-pig airways and to characterize their conformational changes during bronchoconstriction induced by methacholine aerosol. Airways present a palisade of elastic system fibers just beneath the epithelial basement membrane; these fibers are also present in the adventitial connective tissue. Thin fibers link the fibers located in the palisade among themselves and also connect them to those fibers located in the bronchial adventitial tissue, by traversing the airway smooth muscle. During bronchoconstriction, the fibers located beneath the epithelial basement membrane are divided into two components: one follows the epithelial invaginations towards airway lumen, while the other population remains attached through airway smooth muscle to the fibers located in the adventitial connective tissue. At the ultrastructural level, the findings corroborated those of the light microscopy and in addition, disclosed that typical mature elastic fibers and also elaunin fibers attach directly to the basal lamina, a feature that has not been reported previously in other tissues studied. This configuration is compatible with the idea that fibers of the elastic system restrict the mucosal folding during bronchoconstriction, and may also provide energy to restore airway configuration to its normal status after contraction.

Organization of collagen and elastic fibers studied in stretch preparations of whole mounts of human visceral pleura.

Fibers of the collagenous and elastic systems are most relevant in the double mechanical action of visceral pleura (VP), i.e. volume limitation and the generation of elastic recoil pressure. In this work we studied the organization of these fibrous components of VP in two situations: normal lungs and bullous disease. We employed histochemical methods on conventional histological slides and on thin spreads of whole mounts of visceral pleura. In addition, the scanning electron microscope was also used. According to our results, pleural function is made possible by the combination of both the elastic and collagenous fiber systems, each one having as intrinsic organizational pattern. Marked alterations of pleural bullous structure are observed with changes in lung volume. Fibers of the elastic and collagenous systems are clearly interdependent elements. Collagenous fibers are interwoven in a plaited structure that closely resembles the osiers of a wicker basket, indicating that collagen fibers allow for lung volume increase up to a point of maximal stretching of the system. The pleural contribution to lung elastic recoil pressure originates from the elastic network which turns back to its resting position when inspiratory pressures are negligible. The pleural immobility in bullous disease is associated with an almost complete absence of elastic fibers and the presence of very thick collagen fibers, suggestive of a cicatricial process, devoid of any characteristic pattern of distribution.

Urban air pollution enhances the formation of urethane-induced lung tumors in mice.

This paper investigates the association between air pollution and lung neoplasia in an animal model. The experimental exposures were done in two locations with different air pollution profiles: a polluted area (downtown São Paulo) and a "clean" environment (Atibaia). Swiss mice were employed and urethane (3 g/kg) was used as carcinogenic substance. Two experiments were performed: Experiment I was designed to verify whether air pollution acts as initiator and/or promoter of lung cancer, using 300 mice; Experiment II employed 250 animals and aimed to verify if the effects of air pollution on the development of lung tumors was dose dependent. A significant effect of air pollution in augmenting lung carcinogenecity induced by urethane was observed. This effect was shown to be dose-dependent and reproducible on two different occasions. In addition, morphometric studies revealed that pollution may influence tumor phenotype. These results support the hypothesis that air pollution plays a significant role in the development of lung tumors.

Dating of fibrotic lesions by the Picrosirius-polarization method. An application using the lesions of Lechiguana (bovine focal proliferative fibrogranulomatous panniculitis).

This work was designed to verify if a simple quantitative procedure to estimate the state of collagen aggregation is useful in describing the natural history of a fibrous process. For this purpose sixteen cases of Lechiguana lesion were used. Histochemical evaluation of the collagen content and its state of aggregation was done by the Picrosirius-polarization method. Morphometric studies were done by means of a point-counting procedure, which allowed the determination of the areal fraction of thin and thick collagenous fibers within Lechiguana lesions collected at different times of clinical evolution (14 days through 8 months). Early lesions are characterized by thin collagenous fibers. This population of slender fibers decreases later on, when thick fibers become more prevalent. Curve fitting procedures were employed using the state of collagen aggregation as the dependent variable and time as the independent variable. The best fitting was obtained by linear and exponential functions. Statistical analysis indicates that the quantitative study of the degree of collagen organization allows an adequate determination of the time course of Lechiguana lesions. We concluded that simple determinations of collagen aggregation provide numerical data that may be useful to build mathematical models relating time of evolution of the disease to fibrosis.

Leukocyte infiltration and collagenolysis in cervical tissue from intrapartum sheep.

Sheep uterine cervices and cervical mucus were heavily infiltrated by neutrophils during labour, whereas samples of cervices obtained from non-pregnant controls had no infiltrate. The neutrophilic infiltrate of the sheep uterine cervix at term was not homogeneously distributed throughout the organ: luminal mucus contained more neutrophils than tissues which, in turn, displayed a differential distribution, the superficial subepithelial layer being more heavily infiltrated than the deeper submucous layers. A widespread collagenolysis was observed in the sheep uterine cervix at term. The homogeneous morphological aspect of degradation of collagen fibres throughout the whole cervical stroma contrasted with the above-mentioned differential distribution of neutrophils. On the basis of previous reports showing that collagenolysis follows the leukocytic invasion of human and rat cervices at term, a possible role for the neutrophilic infiltrate of the sheep uterine cervix is discussed.

The lysosomal-associated membrane protein LAMP-1 is a novel differentiation marker for HC11 mouse mammary epithelial cells.

HC11 cells are a model for mammary epithelial cell differentiation. Following treatment with the lactogenic hormones glucocorticoids, insulin and prolactin the HC11 cells synthesize milk proteins. Stereological analysis at the ultrastructural level suggested that lysosomal biogenesis was activated following lactogenic hormone treatment of HC11 cells. Differentiation was also accompanied by an increase in the cellular content of tri- and tetra-antennary oligosaccharides, which were reactive with isolectin L4 from Phaseolus vulgaris (L-PHA). The lysosomal-associated membrane glycoproteins LAMP-1 and LAMP-2 are the major carriers of this glycosylation pattern. An analysis of LAMP-1 and LAMP-2 expression levels showed that there was a dramatic increase in LAMP-1 following lactogenic hormone treatment of HC11 cells. The control of LAMP-1 expression is mainly post-transcriptional since the level of LAMP-1 RNA is not affected by lactogenic hormones. Stereological analysis also showed an increase in intermediate filament control of differentiated cells. Analysis of the cytokeratins expressed in differentiated cells suggests that HC11 cells have characteristics of a mammary-specific stem cell. Increase in lysosomal vesicles and their contents might play a role in intra- and extra-cellular remodeling, which is characteristic of cell differentiation.

The role of the collagenous and elastic system fibers in modulating bronchoconstriction.

The distribution and conformational changes of the fibers of the collagenous and elastic systems in guinea pig airways after a contractile agonist challenge are described. We observed a distinct pattern of behavior within the mucosal fibers during bronchoconstriction. Part of the fibers of the two systems tend to follow the epithelial invaginations towards the airway lumen, while the remaining ones seem to be attached to the internal smooth muscle. These layers of fibers in the mucosa are interconnected to one another and to the adventitial network by slender fibers. We suggest that the configuration and behavior of these fibers during bronchoconstriction may contribute to airway reopening after the contractile stimulus has ceased. The possible role of this mechanism in the pathophysiology of human asthma is discussed.