Fine filaments in lymphatic endothelial cells.

Several and various types of cells contain fine cytoplasmic filaments closely resembling the myofilaments of muscle cells (2, 18, 23, 24). In many of these cells and especially when cultured, it has been demonstrated that some of these filaments react with heavy meromyosin (HMM) in the same way as do the actin filaments of muscle cells (3, 6 7). This suggests that these filaments may be actinoid and form part of a contractile system. As fine intracytoplasmic filaments do occur in lymphatic endothelial cells (2, 14), we undertook an electron microscope investigation of their fine structure and their reaction on incubation with HMM and EDTA. We postulated that lymphatic endothelial cells possess a contractile filamentous system to which these filaments belong.

Pulmonary venous vasculature in neonatal hyaline membrane disease.

The pulmonary veins and venules in lungs of newborn infants with hyaline membrane disease can be completely filled with a barium sulfate solution. The same injection technique and combined roentgenological, histological, and histometrical investigations also reveal a defect in filling of many small muscular arteries and of most pulmonary arterioles. Hence, the latter pulmonary vessels are assumed to be important in pulmonary ischemia, as demonstrated in the respiratory distress syndrome of the newborn infant.

Serotonin producing neuroepithelial bodies in rabbit respiratory mucosa.

The intrapulmonary lining epithelium of rabbits contains newly identified corpuscles composed of argyrophil, argentaffin, yellow fluorescent, ultrastructurally granulated and innervated epithelial cellular organs. These are proved, by electron microscopic cytochemistry and microspectrography, to be a source for intrapulmonary production of serotonin. Probably they are intrapulmonary neuroreceptor organs modulated by the central nervous system which exhibit local secretory activities.

Effects of long-term sensory vs. sympathetic denervation on the distribution of calcitonin gene-related peptide and tyrosine hydroxylase immunoreactivities in the rat lung.

In this study the effect of long-term selective sensory vs. sympathetic denervation was examined in the rat lung. Immunoreactivities for calcitonin gene-related peptide (CGRP) on the one hand and tyrosine hydroxylase (TH) on the other, were used as indexes to assess the changes in the two nerve systems. Following long-term chemosympathectomy a marked increase of CGRP-immunoreactive nerve fibers was seen in the sections, while TH-immunoreactive nerve fibers appeared depleted. Inversely, long-term sensory denervation resulted in an increase of TH-immunoreactive nerve fibers with a sharp decrease of CGRP-immunostained fibers. These results suggest a peripheral interaction between these innervation systems in the lung, the mechanism of which has still to be elucidated.

Endothelin in normal lung tissue of newborn mammals: immunocytochemical distribution and co-localization with serotonin and calcitonin gene-related peptide.

We demonstrated immunoreactivity for endothelins (ET)-1, -2, and -3 and for the precursor, big-ET-1, in the pulmonary diffuse neuroendocrine system (PDNES) of newborn cat, rat, hamster, and mouse. ET-like positive neuroepithelial bodies (NEB) were numerous in the intrapulmonary airways and the alveolar parenchyma. Single neuroendocrine cells (NEC) were less often labeled and mainly localized in the larger bronchi. ET-3-reactive neuronal elements were rarely observed. The intensity and number of immunostained NEB were highest for ET-3, followed in declining order by big-ET-1, ET-1, and ET-2. ET-like possessing NEB displayed interspecies differences. We conclude that ET-3 represents a neuroendocrine form of the ET peptide family. NEB expressing several ET isoforms can be grouped into NEB containing either big-ET-1 and ET-1 or ET-3 only. ET-like immunoreactivity was present in a subpopulation of serotonin (5HT)- and/or calcitonin gene-related peptide (CGRP)-positive NEB. As ET, 5HT, and CGRP have potent pulmonary vaso- and/or bronchomotor effects, our observations suggest that they play a separate or synergistic role in regulatory function of the mammalian PDNES, exerting their influence by paracrine, endocrine, and neurocrine pathways or a combination of these.

Immunocytochemical localization of aromatic L-amino acid decarboxylase in human, rat, and mouse bronchopulmonary and gastrointestinal endocrine cells.

Aromatic L-amino acid decarboxylase (AADC) catalyzes the cellular decarboxylation of L-aromatic amino acids and is therefore involved in the synthesis of several biogenic amines. Application of the indirect immunoperoxidase method on human, rat, and mouse tissues using specific antibodies to AADC revealed all AADC-containing cells. Besides mast cells and adrenergic nerve fibers, the following cells were immunostained: neuroendocrine cells in the tracheobronchial epithelium; neuroepithelial bodies in the bronchopulmonary epithelium; Kultschitzky cells in the small intestine and appendix as well as adrenal chromaffin cells. All the latter cells belong to the so-called APUD system, the "D" in the acronym standing for the activity of the enzyme aromatic L-amino acid decarboxylase. Immunocytochemistry for AADC may become an additional tool not only to highlight APUD cells in tissue sections but also to differentiate the sites of cellular amine synthesis from those of amine storage.

Leu-7 immunoreactivity in human, monkey, and pig bronchopulmonary neuroepithelial bodies and neuroendocrine cells.

Anti-Leu 7 is a monoclonal antibody recognizing a surface antigen on human natural killer cells. By applying the indirect immunoperoxidase method, we demonstrated Leu-7 immunoreactivity in the cytoplasm of neuroepithelial bodies (NEB) and neuroendocrine cells (NEC) of human, monkey, and pig respiratory mucosa. In addition, the anti-Leu-7 monoclonal antibody stained the myelin sheaths of nerve fibers in all tissues investigated. Our findings support the hypothesis that shared antigens exist between the nervous, endocrine, and immune systems.

Chromogranin in bronchopulmonary neuroendocrine cells. Immunocytochemical detection in human, monkey, and pig respiratory mucosa.

Immunoreactive chromogranin A was demonstrated by immunocytochemistry in the cytoplasm of neuroendocrine cells (NEC) and neuroepithelial bodies (NEB) in human, monkey, and pig respiratory mucosa. Three different antisera (one monoclonal and one polyclonal to human chromogranin A, and one polyclonal to bovine chromogranin A) were applied in this study. Chromogranin immunopositivity varied in extent and intensity according to the antiserum applied and the tissue investigated. The monoclonal antibody revealed the strongest immunoreaction. Good correlation between chromogranin immunoreactivity and Grimelius silver staining was observed by comparing adjacent sections, although more cells seemed to reveal chromogranin immunoreactivity than argyrophylia. Chromogranin appears to be a useful histological marker for APUD cells in the respiratory mucosa of several species.

Pathogenesis of the prune-belly syndrome: a functional urethral obstruction caused by prostatic hypoplasia.

Abdominal muscle deficiency, urinary tract abnormalities, and cryptorchidism are the three major features of the prune-belly syndrome, also referred to as triad syndrome or Eagle-Barrett syndrome. The etiology is unclear and the pathogenesis a subject of continuing debate. Clinical and pathologic experience with seven cases of prune-belly syndrome is reviewed. Findings indicate that the urogenital anomalies can be attributed to a functional urethral obstruction which in turn is the result of prostatic hypoplasia. The histology of the abdominal wall is that of atrophy-ie, the degeneration of already formed muscle--and not of primitive muscle. This observation supports the theory that the abdominal muscle hypoplasia is a nonspecific lesion, resulting from fetal abdominal distension of various causes. Transient fetal ascites may be an important feature of the prune-belly syndrome.

Constrictive amniotic bands, amniotic adhesions, and limb-body wall complex: discrete disruption sequences with pathogenetic overlap.

In a fetopathologic evaluation of 18 cases with amniotic bands, we discerned 3 types of lesions: (1) constrictive tissue bands, (2) amniotic adhesions, and (3) more complex anomaly patterns, designated as limb-body wall complex (LBWC). Constrictive bands are caused by primary amnion rupture with subsequent entanglement of fetal parts (mostly limbs) by shriveled amniotic strands. Adhesive bands are the result of a broad fusion between disrupted fetal parts (mostly cephalic) and an intact amniotic membrane. Most of the craniofacial defects (encephaloceles and/or facial clefts) occurring in these fetuses are not caused by constrictive amniotic bands, but are the result of a vascular disruption sequence with or without cephalo-amniotic adhesion. Our observations confirm the fact that amnion rupture is not a conditio sine qua non for the development of LBWC. However, LBWC is often complicated by rupture of the unsupported amnion with ensuing formation of constrictive bands. We think that the concept that considers the 3 lesions in question as a single pathogenetic entity is erroneous and will inevitably lead to a never-ending debate between followers of the 2 prevailing theories. In our view, the theories of Streeter and Torpin are not mutually exclusive but rather apply to different types of lesions. The recognition of constrictive amniotic bands, amniotic adhesions, and LBWC as discrete but often combined disruption sequences with important pathogenetic overlap may resolve many dilemmas in interpretation when a fetus exhibits classical constrictive bands beside more severe defects.

The syndrome of diaphragmatic hernia, abnormal face and distal limb anomalies (Fryns syndrome): report of two sibs with further delineation of this multiple congenital anomaly (MCA) syndrome.

We describe 2 sibs with the syndrome of diaphragmatic hernia, abnormal face, and distal limb anomalies. Both infants died shortly after birth with severe respiratory distress. Postmortem examination showed gross internal anomalies: Dandy-Walker malformation, ventricular septal defect, and renal cystic dysplasia. This combination of anomalies, also termed the Fryns syndrome, appears to be a distinct MCA syndrome with variable expression and probable autosomal recessive inheritance. Prenatal ultrasonographic diagnosis was successful in both patients.

Congenital pulmonary lymphangiectasis with chylothorax: a heterogeneous lymphatic vessel abnormality.

We report on 7 perinatal autopsy cases of primary congenital pulmonary lymphangiectasis (CPL) with bilateral chylothorax. This study demonstrates that primary CPL is often complicated by chylous pleural effusions with ensuing pulmonary hypoplasia. Conversely, CPL appears to be a constant pathological finding in spontaneous congenital chylothorax. These observations indicate a common pathogenesis for both disorders. The basic defect is not an intrinsic lung abnormality, but a developmental error of the lymphatic system resulting in a pulmonary lymphatic obstruction sequence. The cause of CPL is heterogeneous. Apparently, most cases are sporadic occurrences. We report the second instance of CPL in sibs. This indicates that some cases are genetically determined with autosomal recessive inheritance. CPL may also be part of a multiple congenital anomalies (MCA) syndrome such as Noonan, Ullrich-Turner, and Down syndrome.

Pathogenesis of the lethal multiple pterygium syndrome.

We present autopsy studies in 4 unrelated fetuses with the lethal multiple pterygium syndrome (LMPS) with special emphasis on the neuromuscular system. The data suggest that LMPS combines the manifestations of a jugular lymphatic obstruction sequence with those of an early severe fetal akinesia sequence. The jugular lymphatic obstruction sequence with resultant edema and cystic hygroma colli causes fetal lethality usually in the second trimester of pregnancy. Generalized amyoplasia appears to be an important mechanism in the pathogenesis of fetal akinesia as part of LMPS and is not associated with dysgenesis or degeneration of the central nervous system (CNS) but is apparently the result of an early fetal muscular "dystrophy." We propose a genetically determined insult affecting the early embryonic development of both lymph vessels and muscles as the basic defect in LMPS. Placental structure, studied in all 4 cases, demonstrated that triploidy-like placental lesions are specific to LMPS. The present findings suggest that LMPS may be a less heterogeneous entity than previously proposed.

The senile lung. Comparison with normal and emphysematous lungs. 1. Structural aspects.

As part of a study of the structural-functional correlations of excised human lungs obtained at autopsy, the parenchyma and peripheral airways were examined by means of morphometric techniques. Among the 30 lungs characterized by the absence of fibrosis, ten differed from the normal and emphysematous lungs by a homogeneous dilatation of the airspaces, in excess of the dimensions predicted on the basis of age. Study of the standard deviations of the mean linear intercepts showed that the airspace dilatation was more regular than in emphysematous lungs; in addition, there was no clear-cut destruction, as estimated from the number of alveolar attachments. These lungs were characterized in addition by an increased thickening of alveolar septa, without inflammation or fibrosis, normal size of the diameter, and reduced density of the membranous bronchioles. Since these lungs were from people older than 60 years, it is assumed that they represent cases of exaggerated airspace enlargement of the aging lung, differing from emphysema by the absence of destruction of alveolar walls. The term "senile lung" is proposed or this condition.

The senile lung. Comparison with normal and emphysematous lungs. 2. Functional aspects.

Senile lungs are characterized by a homogeneous enlargement of the alveolar airspaces, without fibrosis or destruction of their walls. Study of the functional characteristics of excisea senile lungs showed an increase in minimal air and a shift to the left of the elastic recoil pressure-volume curves, less pronounced than in emphysematous lungs. Maximal expiratory volumes and flows were normal. Total lung capacity was not significantly increased, but this may be a consequence of preagonal edema. Comparison of normal, senile, and emphysematous lungs showed a close relationship between recoil pressures and mean linear intercept, Lm, and between forced expiratory volume in 1 s and diameter and density of the membranous bronchioles. It is concluded that airspace enlargement may precede emphysema and may be responsible for changes in lung elasticity. In this respect, senile lungs are an example of the functional changes caused by an isolated airspace enlargement.

NCAM expression in the pulmonary neural and diffuse neuroendocrine cell system.

Neural cell adhesion molecule (NCAM)--a membrane protein involved in cell-cell adhesion within the central and peripheral nervous systems--was demonstrated to be a sensitive and specific marker for neuroepithelial bodies (NEB) and neural tissue elements in the cat lung. Using the streptavidin-biotin immunoperoxidase method, NCAM reactive sites were investigated with monoclonal and polyclonal antibodies on serial section of Bouin fixed, paraffin embedded lung tissue. Moreover, NCAM expression was compared with that of neuron-specific enolase (NSE) on adjacent sections. The most obvious NCAM staining was obtained with the monoclonal antibody. From newborn to adult life, cell surface labeling was identified on NEB cells. In mature cat lung, they were no longer positive. Solitary neuroendocrine cells (NEC) were always negative. In contrast to the transient postnatal immunoreactivity of NEB cells, nerve fibers and ganglion cells were stained throughout all life stages and studied. The distribution of NCAM in NEB, nerve fibers and ganglion cells was similar to that NSE, except in the adult lung. This study reveals that during lung growth shared NCAM antigens exist between the pulmonary nervous and endocrine system, whereas in mature lung NCAM proteins are confined to neural tissue elements. The difference in NCAM expression between NEB and NEC might suggest an involvement of NCAM in the formation of contacts between NEB cells and nerves.

Stereological estimation of number and volume of pulmonary neuroepithelial bodies (NEBs) in neonatal hamster lungs.

The number and volume of pulmonary neuroepithelial bodies (NEBs) of 1- and 4-week-old hamsters were estimated using unbiased stereological principles and systematic sampling techniques. For comparative purposes, volume estimations were also made in the carotid body, the parathyroid gland, and the adrenal medulla. A significant decrease was found in the total number of NEBs, immunoreactive for CGRP, between 1 and 4 weeks. Individual as well as cumulative NEB volume also decreased significantly. The cumulative NEB volume in 1-week-old hamsters was in the same range as the volumes of the carotids and parathyroids in the same animals. The postnatal decrease of the NEB number suggests that the NEBs are of primary potential importance in the neonatal stage, when they may complement the chemoreceptor function of the carotid bodies, which are relatively inactive at birth. Since the cumulative NEB volume (at least at the age of 1 week) is equal to that of the carotid bodies and the parathyroids, their physiological function may be of similar importance.

Effects of unilateral vagal stimulation on intrapulmonary neuroepithelial bodies.

We studied the influence of unilateral vagal stimulation on intrapulmonary neuroepithelial bodies (NEB) in rabbits. The left vagus nerve was cut and electrically stimulated for 10 min. Animals were killed and the lungs studied with fluorescence and electron microscopy. Intensity of formaldehyde-induced fluorescence, which reflects the serotonin content in NEB, was higher on the stimulated side than on the nonstimulated side (118 +/- 7 vs. 100%, n = 8, P less than 0.001). The latter difference was found to correlate with the stimulus amplitude (r = 0.9, P less than 0.05). Ultrastructurally a decrease in the number of exocytotic dense-cored vesicle (DCV) profiles at the level of the NEB basal epithelial cell membrane was found on the stimulated side (0.32 +/- 0.10 vs. 0.45 +/- 0.16 DCV/micron of basal epithelial cell membrane, n = 8, P less than 0.05). Section of the left vagus nerve without electrical stimulation affected neither the fluorescence intensity nor the number of exocytotic DCV profiles. In animals with supranodosal or infranodosal chronic vagotomy the observed effects of unilateral vagal stimulation were no longer present. We conclude that 1) vagal stimulation increases the serotonin content of NEB; 2) it decreases the number of exocytotic DCV profiles; 3) this effect depends on the amplitude of the stimulus; 4) it is obtained through efferent vagal fibers; 5) these results are the opposite of the effects seen after exposing normal NEB to acute hypoxia; and 6) these physiological experiments corroborate a vagal innervation of NEB, which may play an important role in modulating the sensitivity and reaction of NEB to various stimuli.

Anatomy of membranous bronchioles in normal, senile and emphysematous human lungs.

Serial reconstructions of the membranous bronchioles (MB) were performed in randomly selected tissue blocks cut parallel to the pleural surface in fixated human lungs. Two to four normal, senile, and emphysematous lungs were examined. Three (2 in senile lungs) orders of MB were observed with a dichotomous branching pattern. Emphysematous lungs are characterized by an overall decrease in airway diameter with localized stenoses. Comparison with mean airway diameter (d) and density [n (no. of MBs per cm2 of lung tissue)] of MBs obtained using standard morphometric techniques (random sections approximately 1 cm from the pleura) showed that the values of d and n are biased because of the inclusion in the measurements of a number of respiratory bronchioles and bronchi. When these misclassifications are corrected for, it appears that d corresponds quite well to the mean diameter of the terminal bronchioles (TB) and n to approximately twice the density of TBs. After correction, n is not significantly reduced in emphysematous lungs (the grossly destroyed areas being excluded) compared with normal lungs. The estimate of the number of TBs obtained from the present data is markedly less than that calculated by Weibel (Morphometry of the Human Lung. Berlin: Springer-Verlag, 1963), which suggests that the number of bronchi was overestimated by Weibel by at least one generation. Finally, values of peripheral airway resistance computed from the present anatomic data correspond quite well to direct measurement performed on the same lungs before fixation (Verbeken et al., J. Appl. Physiol. 72:2343-2353, 1992).

Tissue and airway impedance of excised normal, senile, and emphysematous lungs.

We partitioned pulmonary resistance (RL) in excised normal, senile, and emphysematous human lungs at various distending pressures; peripheral resistance (Rp) was measured by means of retrograde catheters and lung tissue resistance (Rti) by means of pleural capsules. By subtracting Rp from RL and Rti from Rp, we obtained, respectively, central (Rcaw) and peripheral (Rpaw) airway resistance. We determined also lung volumes, the elastic recoil pressure-volume curve, and the forced expiratory volume in 1 s-to-vital capacity ratio (FEV1/VC). The functional data were related to morphometry: mean linear intercept (Lm), diameter (d), and density (n/cm2) of membranous bronchioles. In the three groups of lungs, Rti demonstrates a marked negative frequency dependence and increases with transplumonary pressure. In emphysematous lungs, the increase of RL is mainly due to an increase of Rpaw; in addition, Rcaw and Rti are higher than normal. In the group of senile lungs, airway resistances are within normal range, but Rti is slightly increased. FEV1/VC is related to Rpaw and elastic recoil pressure; Rpaw is related to d and n/cm2, and Rti is related to dynamic elastance and to Lm.