The electron microscopic characteristics of stress lung

The influence of long-term (6 hours) immobilization stress on morphofunctional state of lung air-blood barrier was studied in experiments of the rats. It was shown that stress provoked the marked ultrastructural changes in the lungs, which were as follows: lung tissue oedema, pronounced thickening of lung air-blood barrier and its separate layers, edema-hemorrhagic syndrome, alveolar epithelial injury, disturbance of lung surfactant systems. Such a pathological complex may be designated as a "stress lung".     

Neuromediator action on surfactant system activity and the functioning area of the lung

Experiments on rats have demonstrated the effect of neurotransmitter function on the activity of the surfactant system and functioning area of the lung (FAL). Excess transmitters gave rise to activation of the lung surfactant system and increase in the FAL, whereas the deficiency of transmitters lowered surfactant activity and the area of the air-blood barrier.     

State of the surface activity of pulmonary surfactant in rats at different periods after left-sided pneumonectomy

The surface activity of the seven-fold washing of the right lung was measured on the modified Wilhelmy's balance after the leftside pneumonectomy in rats. It appeared to be normal (gamma min-23--24 dynes/cm) up to the 5th day, and at the remote postoperative periods. The intracellular edema of the air-blood barrier components and the release of the edema fluid into the alveolar lumen in the "vesicle" composition failed to influence the surface properties of the lung surfactant. A sharp increase of the alveolar dimensions on the 5th--7th postoperative day was followed by an increase of the surface-active properties of the lung washings (gamma-min-11--15 dynes/cm) and by the intensified secretion of the material of the osmiophilic lamellar bodies from the alveolar cells of the 2nd type into the alveolar lumen. The cytological mechanisms providing the intensified production of the surfactant in the hypertrophic alveoli are activation of the lipid synthesis in the alveolar cells of the 2nd type, their hypertrophy, and also the appearance of binuclear cells.     

Pulmonary surfactant system of newborn rats in alcoholic intoxication of pregnant rats

Data, received in investigation of the lungs of 45 newborn rats, show, that there is the suppression of the surface active properties of surfactant in animals, born from female rats with simulated alcoholic intoxication in pregnancy period. The decrease of the surface activity of surfactant may be connected with direct injury influence of alcohol on surfactant as well as with inactivation of surfactant with serum proteins, which appear in the alveolar space because of the increase of the permeability of components of air-haematic barrier. The suppression of the surface active properties of surfactant is accompanied by reinforcement of the functional activity of the 2nd type pneumocytes and appearance of the hypertrophic forms of these cells.     

Ultrastructural and morphometric study of the lung of the European salamander,Salamandra salamandra L.

Ultrastructural and morphometric investigations were performed on the lung of the European salamander, Salamandra salamandra L. Folds of first and second order are covered with a ciliated epithelium containing goblet cells. The respiratory surface of the lung is lined by a single type of cell which, in amphibians, combines features of type I and type II alveolar cells of the mammalian lung. In the salamander the respiratory and ciliated epithelial cells as well as goblet cells possess electron dense and lucent vesicles in their cytoplasm as well as lamellar bodies. A small amount of surfactant, composed most probably of phospholipids and mucopolysaccharides, was observed covering the entire inner surface of the lung. Morphometric methods were used to determine the dimensions of the perinuclear region of pneumocytes, the thickness of the air-blood barrier and lung wall, and also the diameter of capillaries. The thickness of the respiratory air-blood barrier was found to be considerably higher than that of the corresponding barrier in mammals.     

Changes in surface activity of surfactant and ultrastructure of the air-blood barrier in alcoholic intoxication in experimental animals

In the results of complex investigation of the lungs of 26 white rats, it was established, that there is the suppression of surface active properties of surfactant under influence of ethanol. In acute poisoning this suppression is associated with direct injury of surfactant with ethanol and inactivation of surfactant with serum proteins, which appear in the alveolar space because of the edema of air-haematic barrier. In prolonged influence the suppression of the surface activity of surfactant is due to the increase of its catabolism with alveolar macrophages.     

Effect of exogenic hypercapnia on external respiration and oxygen transport function of lungs

The functioning of the respiratory system and oxygen-transport function of the lungs were estimated in experiments on healthy people under the effect of exogenic hypercapnia. The external respiration is activated under conditions of hypercapnia of the given degree, an increase in the total and alveolar ventilation of the lungs testifies to this fact. Age differences are found in diffusive and specific diffusive capacity of the lungs for O2, which indicates to changes in the oxygen-transport function of the lungs. In teenagers the diffusive capacity of the lungs changes due to the gas-exchange surface area and in people of middle age - due to changes in the diffusive properties of the lungs air-blood barrier.     

Effect of the antihypoxic agent ionol on the morphofunctional state of the air-blood barrier of the lungs in hypoxic anoxia

It is shown that antihypoxic ionol has promoted normalization of the air-blood lung barrier ultrastructure, activation of the surfactant system under acute hypoxic hypoxia effect as well as compensatory redistribution of the thickness of separate barrier layers due to intensified synthesis of phospholipids which are the components of cytoplasmic membranes and pulmonary surfactant.     

A morphometric examination of type II alveolar epithelial cells in normal and isolated-perfused dog lungs

The volume densities of type II alveolar cell cytoplasmic organelles and alveolar surface densities were estimated by established stereologic procedures. The morphometric measurements were obtained from normal dog lungs (in situ) and isolated dog lungs perfused for 30-minute, 1-hour, and 2-hour periods. The type II cell lamellar body volume densities and the alveolar surface densities progressively decreased as the times of perfusion were increased. The volume densities of the granular and agranular endoplasmic reticulum progressively increased during the periods of perfusion. These morphometric parameters from lungs in situ and isolated lungs suggest that changes occur in pulmonary surfactant synthesis and activity during perfusion. It is further postulated that progressive increases in the rates of surfactant removal and/or inactivation during perfusion may contribute to spontaneous edema in lungs isolated for periods exceeding two hours. The morphologic and physiologic integrity of isolated perfused lung preparations, widely used as models of lungs in vivo, in situ requires further evaluation.     

Ultrastructure of the respiratory epithelium in the lungs of the tortoise,Testudo graeca

The respiratory epithelium in the lungs of the tortoise (Testudo graeca) has been studied by electron microscopy. The epithelium consists of a mosaic of two different cell types (here called "pneumonocytes"). Type I pneumonocytes are roughly squamous and possess attenuated flanges of cytoplasm which extend over the septal capillaries. Localized cytoplasmic expansions are often present near the periphery of these flanges. Most of the organelles are concentrated in the perinuclear region; the most prominent of these are the mitochondria and osmiophilic inclusions. In contrast, type II pneumonocytes are cuboidal and are richly endowed with organelles including large Golgi complexes, extensive endoplasmic reticulum and numerous inclusion bodies. The morphological evidence suggests that type I pneumonocytes are involved in the secretion of osmiophilic material (presumed to be pulmonary surfactant) and in maintaining the integrity of the air-blood barrier. Type II pneumonocytes appear to be concerned solely with the production of surfactant.     

Air-blood barrier of the lungs in acute hypoxic hypoxia as affected by liposomes

Phospholipids injected into the organism under an acute hypoxic hypoxia prevent development of ultrastructural disorders in the air-blood barrier of the lungs, thus averting development of the pulmonary acute hypoxic hypoxia.     

The state of the "glycocalyx" of rat lung cells following left-sided pneumonectomy]

Edema in the tissue of the right lung developed 24 hours after the leftsided pneumonectomy. It was found that: 1) the layer of acid mucopolysaccharide on the surface of the alveolar and endothelial cells became thicker; 2) accumulation of the product of reaction with the RR on the surface of the alveolar cells of type 1 and of the endothelial cells took place; 3) "blisters" coated with the product of reaction with the RR (Red Ruthenium) appeared. These "blisters" were connected with the plasmolemma of the alveolar and endothelial cells. All these findings suggest that acid mucopolysaccharides of the lung surfactant system participated in the accumulation and elimination of the water from the tissues of the air-blood barrier.     

Exposure to Polymers Reverses Inhibition of Pulmonary Surfactant by Serum,Meconium, or Cholesterol in the Captive Bubble Surfactometer

Dysfunction of pulmonary surfactant in the lungs is associated with respiratory pathologies such as acute respiratory distress syndrome or meconium aspiration syndrome. Serum, cholesterol, and meconium have been described as inhibitory agents of surfactant’s interfacial activity once these substances appear in alveolar spaces during lung injury and inflammation. The deleterious action of these agents has been only partly evaluated under physiologically relevant conditions. We have optimized a protocol to assess surfactant inhibition by serum, cholesterol, or meconium in the captive bubble surfactometer. Specific measures of surface activity before and after native surfactant was exposed to inhibitors included i), film formation, ii), readsorption of material from surface-associated reservoirs, and iii), interfacial film dynamics during compression-expansion cycling. Results show that serum creates a steric barrier that impedes surfactant reaching the interface. A mechanical perturbation of this barrier allows native surfactant to compete efficiently with serum to form a highly surface-active film. Exposure of native surfactant to cholesterol or meconium, on the other hand, modifies the compressibility of surfactant films though optimal compressibility properties recover on repetitive compression-expansion cycling. Addition of polymers like dextran or hyaluronic acid to surfactant fully reverses inhibition by serum. These polymers also prevent surfactant inhibition by cholesterol or meconium, suggesting that the protective action of polymers goes beyond the mere enhancement of interfacial adsorption as described by depletion force theories.     

Effect of exogenous pulmonary surfactant preparations on the structure of pulmonary alveoli in newborn rats

Treatment of pre-term newborns with exogenous surfactant preparation is a well established part of the therapy for respiratory distress syndrome of the newborns (RDS). Since the introduction of surfactant into clinical practice in 1980, hundreds of studies have been published describing beneficial effects of such treatment. There is only limited number of morphological publications reporting adverse effects of surfactant administration. The aim of the present study is to describe morphological changes in the lung after surfactant administration to healthy newborn rats. Two types of surfactant were used: Exosurf (Glaxo Wellcome, England) and Survanta (Abbott Laboratories, USA). Surfactant preparation were given intratracheally in single dose (bolus) (100 mg of lipids per kg b.w.). Animals from control group received 0.9% saline in equivalent volume. Lung specimens were taken 15, 20, 25 and 30 minutes after drug administration and evaluated by light and electron microscopy. There was no damage in lungs from the control group. Tissue specimens from the Exosurf group revealed severe pathological changes: foci of atelectasis, frank edema in the parenchyma, focal disruption of air-blood barrier, hemorrhages in many alveoli, surfactant particles in many alveolar capillaries, and strongly activated alveolar macrophages. In this group changes appeared as early as 15 min after surfactant administration and intensity of lung injury increased with time. Also, Survanta administration caused damage to the lung tissue. However, the changes were less intense and appeared later (20-25 minutes after Survanta treatment). In conclusion, the presented morphological findings proved that exogenous surfactant administration to healthy rat newborns caused lung damage. Comparing two different surfactant preparation, Exosurf and Survanta, it was shown that the former one produced stronger and faster damage to lung alveoli than the latter one.     

Scanning and transmission electron microscopic study of the lung of the newt,Triturus alpestris Laur.

Summary The lungs of Triturus alpestris Laur. were investigated with the scanning and transmission electron microscopes. Dimensions of the cell bodies of pneumocytes and ciliated cells, as well as the thickness of the air-blood barrier, were determined. The lungs of the newt form two simple sacs without septa. A ciliated epithelium containing goblet cells lines the pulmonary vein and partially the pulmonary artery. The remainder of the lung surface is covered internally by respiratory epithelium consisting of one type of cell and only occasionally showing the presence of single ciliated cells. All cells, ciliated, goblet and pneumocytes, contain in their cytoplasm lamellar bodies. Multivesicular bodies and numerous vesicles of variable electron density also occur in the cytoplasm of pneumocytes. Atypical mitochondria can be found in all cell types of the lung. Fixation with addition of tannic acid reveals the surface lining film. Tubular myelin figures were not observed.     

Electron microscopic demonstration of pulmonary surfactant proteins using procion brilliant blue H5GS

A new electron microscopy technique is described for detection of lung surfactant proteins with the copper-containing phthalocyanine dye, procion brilliant blue H5GS. The protein structures were stained concurrently with the fixation during perfusion through the pulmonary artery of a fixative-staining mixture containing glutaric aldehyde and a dye in the kakodilate buffer, pH 5.6-6, and in the course of a subsequent immersion of lung tissue pieces into the same mixture. Then the material was treated with thiosemicarbazide and post-fixed with OsO4. The dye did not penetrate intact cells. The electron-dense products of the histochemical reaction were located inside and on the surface of the surfactant membrane, in the hypophase of the surfactant complex, on the plasmalemma of air-blood barrier cells and in its micropinocytosis vesicles, as well as on the membranes of osmophilic plate-like bodies as their contents egressed into the alveolar lumen.     

Morpho-functional analysis of lung tissue in mild interstitial edema

Mild pulmonary interstitial edema was shown to cause fragmentation of interstitial matrix proteoglycans. We therefore studied compartmental fluid accumulation by light and electron microscopy on lungs of anesthetized rabbits fixed in situ by vascular perfusion after 0.5 ml.kg(-1).min(-1) iv saline infusion for 180 min causing approximately 6% increase in lung weight. Morphometry showed that a relevant portion (44%) of extravascular fluid is detected early in the alveolar septa, 85% of this fluid accumulating in the thick portion of the air-blood barrier. The arithmetic mean thickness of the barrier increased in interstitial edema from 1.06 +/- 0.05 (SE) to 1.33 +/- 0.06 microm. The harmonic mean thickness increased from 0.6 +/- 0.03 to 0.86 +/- 0.07 microm, mostly due to thickening of the thin portion causing an increase in gas diffusion resistance. Despite some structural damage, the air-blood barrier displays a relatively high structural resistance providing a safety factor against the development of severe edema. It is suggested that the increase in extra-alveolar perivascular space occurs as a consequence of fluid accumulation in the air-blood barrier.     

Interplay between the morphometry of the lungs and the mode of locomotion in birds and mammals

We studied the lung diffusion parameters of two species of birds and two species of mammals to explore how structural and functional features may be paralleled by differences in life style or phylogenetic origin. We used two fast-flying species (one mammal and one bird), one running mammal and one bird species that flies only occasionally as models. The harmonic mean thickness of the air-blood barrier was very thin in the species we studied. An exception was the Chilean tinamou Notoprocta perdicaria, which only flies occasionally. It showed an air-blood barrier as thick as that of flightless Galliformes. We found that the respiratory surface density was significantly greater in flying species compared to running species. The estimated values for the oxygen diffusion capacity, DtO2 follow the same pattern: the highest values were obtained in the flying species, the bat and the eared dove. The lowest value was in N. perdicaria. Our findings suggest that the studied species show refinements in their morphometric lung parameters commensurate to their energetic requirements as dictated by their mode of locomotion, rather than their phylogenetic origin. The air-blood barrier appears to be thin in most birds and small mammals, except those with low energetic requirements such as the Chilean tinamou. In the species we studied, the respiratory surface density appears to be the factor most responsive to the energetic requirements of flight.     

Pulmonary and gastric surfactants. A comparison of the effect of surface requirements on function and phospholipid composition

Surfactant is present in the alveoli and conductive airways of mammalian lungs. The presence of surface active agents was, moreover, demonstrated for avian tubular lungs and for the stomach and intestine. As the surface characteristics of these organs differ from each other, their surfactants possess distinct biochemical and functional characteristics. In the stomach so-called 'gastric surfactant' forms a hydrophobic barrier to protect the mucosa against acid back-diffusion. For this purpose gastric mucosal cells secrete unsaturated phosphatidylcholines (PC), but no dipalmitoyl-PC (PC16:0/16:0). By contrast, surfactant from conductive airways, lung alveoli and tubular avian lungs contain PC16:0/16:0 as their main component in similar concentrations. Hence, there is no biochemical relation between gastric and pulmonary surfactant. Alveolar surfactant, being designed for preventing alveolar collapse under the highly dynamic conditions of an oscillating alveolus, easily reaches values of <5 mN/m upon cyclic compression. Surfactants from tubular air-exposed structures, however, like the conductive airways of mammalian lungs and the exclusively tubular avian lung, display inferior compressibility as they only reach minimal surface tension values of approximately 20 mN/m. Hence, the highly dynamic properties of alveolar surfactant do not apply for surfactants designed for air-liquid interfaces of tubular lung structures.     

Structure-function relationships in pulmonary surfactant membranes: From biophysics to therapy

Pulmonary surfactant is an essential lipid–protein complex to maintain an operative respiratory surface at the mammalian lungs. It reduces surface tension at the alveolar air–liquid interface to stabilise the lungs against physical forces operating along the compression–expansion breathing cycles. At the same time, surfactant integrates elements establishing a primary barrier against the entry of pathogens. Lack or deficiencies of the surfactant system are associated with respiratory pathologies, which treatment often includes supplementation with exogenous materials. The present review summarises current models on the molecular mechanisms of surfactant function, with particular emphasis in its biophysical properties to stabilise the lungs and the molecular alterations connecting impaired surfactant with diseased organs. It also provides a perspective on the current surfactant-based strategies to treat respiratory pathologies. This article is part of a Special Issue entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy.