Search

Binesis is a process whereby the membrane of the insuline secretory vesicle in the beta cell forms a lingula that indents the vesicular membrane of an adjoining secretory vesicle of the plasma membrane. Vesicular binesis in beta cells increases when islets of Langerhans are incubated at a stimulatory glucose concentration (300 miligrams per 100 milliliters). These vesicular membrane alterations may be the morphological concomitants of activation of the insulin secretory vesicle, and indicate an active role for the vesicle and its membrane in the release mechanism.

Radiolabeled phosphate, acetate, and glycerol are incorporated into strain L-fibroblast phospholipids. The acetate and glycerol specifically label the fatty acid and glycerol moieties, respectively, of the phospholipids. To study the metabolic fate of the various moieties of phospholipids, cells incubated with the above radiolabeled compounds were transferred to unlabeled medium, and the rate at which phospholipid radioactivity per 10(6) cells decreased was determined. The rate of decrease expected on the basis of cell division alone was estimated either by monitoring increases in cell number, or by measuring the rate at which radiolabeled DNA per 10(6) cells decreased. Both phospholipid phosphorus and glycerol are lost at a rate greater than can be accounted for by cell division alone. By contrast, nearly all phospholipid acyl chains were retained by the cell to the same extent as radiolabeled DNA. While presence of nonradioactive glycerol in the medium increased the rate at which glycerol was lost from phospholipid, the addition of exogenous fatty acid was without effect on the retention of phospholipid acyl groups. The acyl-glycerol bond of phosphatidylcholine is metabolically more labile than that of phosphatidylethanolamine. Together the data suggest that although L-fibroblast phospholipids undergo deacylation-reacylation reactions, the acyl chains do not equilibrate with either extracellular or intracellular pools of unesterified fatty acid.

Two patients with a persistent mesangial proliferative glomerulonephritis and reduced serum C3 levels have been followed for six and 10 years. Both have had a mild course with normal renal function and spontaneous morphologic improvement. Levels of the fourth component (C4) and second component (C2) of complement, and properdin factor B have been normal; the third component (C3) of complement nephritic factor has not been detected. Ultrastructurally, irregular intramembranous electron-dense deposits are present, primarily within the lamina densa. These deposits are separated by varying lengths of normal-appearing glomerular basement membrane. Intramembranous and subendothelial electron-lucent areas, containing vesicles, are also seen. Subendothelial deposits, mesangial interposition, splitting of the glomerular basement membrane and ribbon-like intramembranous deposits are not found. The disease in these patients was clinically indistinguishable at onset from mrmbranoproliferative glomerulonephritis (MPGN). The possibility of a variant with mild clinical course, no progression and spontaneous morphologic improvement is important in planning and assessing treatment programs in this disease group.

The integrity of epithelial tight junctions in foetal mammalian lungs is essential to maintain the unique ionic composition of lung liquid, and to prevent leakage of serum proteins into peripheral air spaces. In the present study the development of intercellular junctions of the lining epithelium of foetal lamb lungs during gestation was examined by light and electron microscopy. Both thin sections and freeze-fracture replicas were examined by electron microscopy. By 39 days of gestation, epithelial tight junctions consist of a minimum of 3.1 +/- 1.6 (s.D.) and a maximum of 5.8 +/- 2.0 discontinuous rows of particles and short segments of strands on P face ridges and in complementary E face grooves, while from 58 to 76 days they are composed of a network of 4.3 +/- 1.6 to 7.7 +/- 1.9 focally interrupted P face strands. Complementary replicas show that many of the discontinuities on the P face are due to separation of junctional particles on to the E face during fracturing, and not to an absence of junctional particles. From 76 days to term, epithelial tight junctions (exclusive of upper airway epithelium which was not examined) resemble those of adult lungs, and consist of a continuous network of 4.5 +/- 2.0 to 7.5 +/- 2.5 P face strands and complementary particle-free grooves. Permeability measurements, published elsewhere, indicate that the epithelium is functionally 'tight' from 69 days onwards. Tight junctions in peripheral air-space epithelium, therefore, are structurally continuous and functionally 'tight' early in foetal lung development, and form seals at one end of long, narrow intercellular spaces; these features may be important for coupled ion and water transport. When the bounding epithelial cells become flattened, these narrow intercellular spaces remain intact as a result of complex interdigitations of adjacent cell membranes. Desmosomes were present throughout gestation near the abluminal side of the tight junctions and occasionally near the base of the intercellular space. These junctions may serve to connect cells to each other at a time when tight junctions may be mechanically weak. In addition, gap junctions are associated with tight junctions from the glandular through the canalicular stages of lung development. They disappear by 120 days when the epithelial cells are differentiated.

In the present study morphology of tight junctions was related to the various cell types lining extrapulmonary and intrapulmonary airways of the rat. Freeze fracture replicas were prepared from extrapulmonary airway epithelium derived from the cartilagenous and membranous sides of upper, middle, and lower thirds of the trachea. Intrapulmonary airway epithelium was obtained from airways less than 1 mm in diameter. Tight junction fibrils on the P fracture face were organized into three types of patterns. Type 1: parallel sparsely interconnected lumenal fibrils with large ablumenal fibril loops. Type 2: richly interconnected lumenal fibrils with large ablumenal fibril loops. Type 3: narrow network of interconnected fibrils. On the E fracture face complementary grooves were organized in a similar pattern. Ciliated cells on both sides and all levels of the trachea were associated with type 1 junctions. In intrapulmonary airways, however, the junctional pattern of ciliated cells changed to type 2. Brush cells at all levels of the airways were bounded by type 2 and occasionally by type 1 junctions. Secretory cell junctions displayed the following patterns: Mucous cells were bounded solely by type 3, serous cells by either types 2 or 3, and Clara cells predominantly by type 2. Cells tentatively identified as intermediate cells displayed all three junctional patterns. The number of parallel fibrils comprising tight junctions was higher in extrapulmonary as compared to intrapulmonary airways. No difference was seen in the various locations sampled in the trachea. Gap junctions were observed between secretory cells of extrapulmonary but not intrapulmonary airways. These observations are discussed in relation to current physiologic data.

Experiments are described, using isolated perfused capillaries, fluorocarbon exchanged rats, and endothelial cells in culture, that examine the interaction of circulating proteins with the endothelial surface. A consequence of the interaction of circulating proteins with the endothelial surface is to render the endothelium less permeable to water, small water-soluble molecules, as well as macromolecules. Indirect evidence suggests that this is the result of adsorption of proteins to the glycoprotein coat of endothelial cells. An understanding of the physiochemical basis for this interaction awaits further studies of the chemical composition and properties of molecules constituting the endothelial cell surface.

Neonates have an increased susceptibility to infections. Because optimal phagocytosis of offending organisms by alveolar macrophages (AM) requires recognition and attachment of opsonized organisms to the AM cell membrane, the expression of opsonic receptors on the surface of newborn and adult rat AM was investigated using immunologic techniques, cell culture, and flow cytometry. We investigated the expression of Fc, C3b, fibronectin, and lectin receptors on newborn (1 to 5 wk) AM and compared them with those of adult AM. The expression of Fc receptors (FcR) was significantly less on the surface of newborn AM, particularly during the first week of life, as determined by their binding of aggregated IgG, E(IgG), and opsonized 125I-Listeria monocytogenes. A similar depressed receptor function was observed for C3b, fibronectin, and some lectins. The possible effect of cell size on receptor expression was examined by morphometry and flow cytometry. The results indicated that, while mean AM size was approximately 12% smaller during the first week of life, it attained adult levels by the second week. Thus, a difference in size did not account for either the magnitude of decreased receptor expression or the diminished intensity of staining for surface-aggregated IgG that was detectable for up to 5 wk. Binding of a variety of lectins to the AM surface was decreased during the first week after birth, but approximated adult levels by 4 wk. By contrast, soybean and peanut agglutinin, lectins that bind to D-GaINAC moieties, showed a transient increase in binding to AM during the second and third weeks of life.(ABSTRACT TRUNCATED AT 250 WORDS)

Morphologic alterations of the slit diaphram and glomercular epithelium were examined in rats with experimental autologous immune complex nephritis following perfusion fixation with tannic acid-glutaraldehyde. The changes observed included a displacemnt by immune complex deposits of foot processes and associated slit diaphragms towards the urinary space. With the onset of proteinuria, foot processes spreas, resulting in a decrease in spithelial cell surface area and a diminution in the length of interpithelial slits. Concomiantly, the redundant slit diaphragm became folded and pleated in the residual interpithelial spaces. No primary defect in the slit diaphragm was observed to account fo the severe proteinuria seen in this disease. Instead areas of detached epithelial cells and an occasional slit pore lacking a slit diaphragm may consitute sites for substantial protein loss in the urinary space in autologous immune complex nephritis.

Cellular expression and extracellular deposition of fibronectin and fibrin/fibrinogen in experimental allergic encephalomyelitis (EAE) in guinea pigs (GP) were studied by light and electron microscopic immunohistochemistry of the central nervous system (CNS). Normal GPs had few fibronectin+ and fibrin/fibrinogen+ CNS vessels. Positively stained vessels were more numberous in sensitized GPs prior to and after onset of EAE, and there were extracellular deposits of these molecules in inflammatory foci. Staining was greater in EAE-susceptible strain 13 GPs than in sensitized EAE-resistant strain 2 GP. On ultrastructural analysis both molecules were localized on endothelial cell luminal surfaces and on intravascular mononuclear cells. Fibronectin and the macrophage fibronectin receptor (A6F10) were found on mononuclear cells in parenchyma. Endothelial cell luminal surface fibronectin and fibrin/fibrinogen may mark and/or contribute to early immune events in EAE, including mononuclear cell margination and emigration in inflammatory sites. Genetic factors, cytokines, and components of coagulation may regulate these interactions in vivo.

The intravenous injection into mice of small volumes (less than 0.1 ml) of peroxidatic enzymes of molecular weight of 40 000 daltons or greater results in little if any penetration of these probe molecules into endothelial junctions. The injection of cytochrome c (12 000 daltons), on the other hand, results in the localization of this tracer in some but not all endothelial junctions. When horseradish peroxidase (EC 1.11.1.7) is injected in a large volume of saline (0.5 ml), reaction product is present in endothelial junctions and basement membrane, but is prevented from entering the alveolar space by zonulae occludentes between epithelial cells. These experiments indicate that although endothelial junctions, under physiological conditions, are largely impermeable to molecules the size of horseradish peroxidase, and presumably most serum proteins, they are labile and susceptible to stretching if intravascular pressure is increased. Freeze-fracture studies show that pulmonary capillary endothelial junctions are composed of one or at the most two strands which show areas of discontinuity. Epithelial junctions, by contrast, are composed of a continuous, complex network of anastomosing fibres. These observations confirm physiological experiments which indicate that it is the pulmonary epithelium rather than the endothelium which determines the permeability properties of the alveolar-capillary membrane to lipid-insoluble molecules. Bidirectional pinocytic transport is an additional mechanism whereby lipid-insoluble molecules are transported across both endothelial and epithelial layers. The relative contribution of this transport mechanism to the total amount transported remains to be established.

A review of the histologic findings in 27 patients originally classified as having some form of membranoproliferative glomerulonephritis (MPGN) revealed 13 with MPGN Type I and 14 with dense deposit disease (DDD). In all 14 cases where electron microscopy was performed, the histologic diagnosis was confirmed. In nine cases the diagnosis of DDD was easily made in histologic sections on the basis of ribbon-like, brightly PAS positive thickening of the GBM, without "splitting" and with relatively slight mesangial proliferation. However, in five cases the picture closely resembled MPGN Type I, with hypercellularity, "splitting" and only focal ribbon-like thickening of the GBM, which required oil immersion for recognition. There was no correlation between the serum C3 levels and the morphologic diagnosis: nine (4 MPGN Type I, 5DDD) had persistently low C3 levels, two (1 MPGN Tye I, 1DDD) were normocomplementemic, and in 16, the C3 levels varied. C3 levels increased with time in nearly all patients. The clinical course was similar in patients with MPGN Type I and DDD. Significant correlations between the rate of development of renal failure and sex, age of onset, nephrotic syndrome or therapy could not be made. The five year survival rate was 87%; 12 developed renal insufficiency by five years. Although morphologically distinct, these findings suggest that DDD is clinically indistinguishable from MPGN Type I.

Alveolar macrophages (AM) from adult and newborn rats were studied by flow cytometry and ultrastructural morphometry. We observed that the laser scatter and autofluorescent properties of newborn macrophages were different from those of adult cells. Relative to the adult AM, the forward-angle laser scatter obtained with the newborn AM was reduced; this optical measurement appeared to correlate with the smaller mean size, as determined by ultrastructural and electronic volume measurements. The diminished right-angle laser scatter (90 degrees angle) correlated with the presence of fewer small, irregularly shaped lysosomal structures in the newborn AM, compared with AM from adult animals. AM from 1-2-day-old rats displayed large vacuoles containing multilamellar structures, which proved to be less effective at scattering light. Cells from newborn rats were less autofluorescent, a finding that appeared to correlate best with the numbers of secondary lysosomes. Flow cytometry may be used to discern structural alterations that occur during the maturation of AM. These changes correlate well with quantitative ultrastructural analyses of these cells.

A variety of benzamidine and pyridinium compounds were examined for their ability to inhibit irreversibly C1s-induced vascular leakage in guinea pig skin. Vascular leakage was compared with esterolysis of N-Z-L-Tyr-Np and catalysis of EAC42 formation by C1s. Vascular leakage correlated significantly better with esterolytic activity than with EAC42 formation. The presence of a sulfonyl fluoride moiety in the compounds is important in the inhibition of C1s-induced vascular leakage.

Freeze-fracture studies have shown that there is a segmental differentiation of the structure of intercellular junctions of intraacinar pulmonary vessels. Tight junctions vary in complexity, being composed in the arterial segment of the most highly interconnected and most numerous (two to seven) rows of tight junction particles, which are present in E face grooves. In capillaries, they consist of one to three rows of particles, which, in the venular segment, show some discontinuities. Tight junctions in intraacinar veins consist of one to five rows of particles in the E face. Large gap junctions are numerous and are intimately associated with arterial tight junctions. They are absent from capillaries, and are fewer in number and smaller in size in veins than in arteries. The above observations suggest that, as in the systemic circulation, the venular segment is likely to be the most permeable to small, water-soluble solutes. The presence of numerous gap junctions, particularly in the arteries, suggests that in this segment there is a high degree of electrotonic and/or metabolic coupling between endothelial cells.

Thymic epithelial cells have a critical influence on T-cell differentiation. In order to characterize these cells in humans, a serum-free growth medium was developed for their long-term culture. Important components of this medium included transferrin, epidermal growth factor, prostaglandin E1, and selenious acid. The presence of a keratin cytoskeleton, tonofilaments, and desmosomes confirmed the epithelial nature of these cells. Indirect immunofluorescence study of these epithelial cells demonstrated the presence of Ia and B-2 microglobulin antigens. The availability of highly enriched thymic epithelial cultures should simplify the functional characterization of this cell.

The structural basis for the permeability of the alveolar-capillary membrane to water-soluble solutes rests in part on the structure and function of its intercellular junctions and the pinocytotic vesicles within its cells. Intercellular junctions between endothelial cells of the pulmonary capillary bed differ both in permeability to enzyme tracers and in their structure. As determined by freeze fracture, the junctions in the arteriolar, capillary, and venular portion of the capillary network vary in complexity, and in the number of rows of particles constituting the junction. Because there are few particles associated with the junctions in the venular end of the capillary bed, these are considered to be the most permeable of the three types of vascular junctions. Epithelial junctions, in contrast, are impermeable to all enzyme tracers studied, and they are composed of a continuous, complex network of junctional fibrils. While intercellular junctions form seals of varying 'tightness,' pinocytotic vesicles provide a means for the transport of water-soluble macromolecules across the alveolar-capillary membrane.

The substructure of the human glomerular slit diaphragm is examined after perfusion fixation of a human kidney with tannic acid-glutaraldehyde. The present study shows that the substructure of the slit diaphragm in man is very similar to that described in rats and mice, and that the dimensions of the contained pores (50 times 120 A) in the human differ only slightly from those reported for the murine kidney. It is unlikely that rectangular pores of the dimensions of those measured in the human kidney would permit, under physiological conditions, the penetration of significant quantities of serum protein molecules. It is possible, therefore, that the slit diaphragm in the human glomerulus could function as a fine filter for serum proteins, although this remains to be definitely proved.

The peripheral blood lymphocytes of nine patients with hyper immunoglobulin (Ig)M immunodeficiency were studied in an attempt to define the cellular basis of this disorder. B cells were normal in number but qualitatively abnormal in all patients. Approximately one-half of the B cell consisted of small lymphocytes (7-9 mum in diameter) bearing surface IgM and IgD, as well as C3 receptors. These cells were driven to secrete IgM but not IgG after in vitro stimulation by pokeweed mitogen. In the blood there were also large lymphocytes (10-14 mum in diameter) that possessed surface as well as intracytoplasmic IgM but lacked C3 receptors. These cells spontaneously secreted large amounts of IgM in vitro and on electron microscopy were found to be rich in rough endoplasmic reticulum. Such a subpopulation of lymphoid cells was not detected in normal peripheral blood and was unique for all patients with hyper IgM immunodeficiency studied.T cells from all patients were normal in number and in function both in vivo and in vitro and were able to generate adequate T-cell help to support IgG synthesis by normal B cells. No evidence was obtained for T cells capable of suppressing normal IgG synthesis in any of the patients after coculture with normal peripheral blood lymphocytes. The defect in hyper IgM immunodeficiency is intrinsic to B cells, which fail to switch from IgM to IgG synthesis.

The fluorocarbon exchange transfused rat was used to examine the effect of varying concentrations of circulating proteins on the permeability of pulmonary endothelium to inject ferritin. There is a significant inverse relationship between protein concentration and the relative number of vesicles containing ferritin as well as the amount of ferritin in the basement membrane. A hysteresis effect of the amount of circulating protein on the permeability of endothelium to ferritin was noted. In repletion experiments almost 2.5 times more protein was required to maintain capillary permeability at normal levels than during depletion experiments.

The accessory cell functions of Ia+ alveolar and peritoneal macrophages were compared to those of splenic cells in the rat. Whereas splenic mononuclear cells and dendritic cells were excellent supporters of both MHC-restricted and nonrestricted T-cell mitogenic responses, Ia+ macrophages were inefficient antigen-presenting cells and poor supporters of lectin mitogenic responses. Binding of antigen-primed T-cell blasts by splenic cells in the presence of Con A or antigen occurred within 30 min and subsequently led to the formation of nonadherent clusters of "dendritic-like cells" and proliferating T-cell blasts. Unstimulated Ia- macrophages failed to bind T cells during 30 min of coculture but formed conjugates with T-cell blasts within 24 hr. Delayed binding did not require the presence of antigen or lectin, or the expression of Ia antigens by the macrophage, and did not lead to T-cell proliferation. Antigen-specific binding and antigen presentation, but not lectin mitogenesis, were enhanced by treating antigen-pulsed Ia+ macrophages with neuraminidase for 30 min at 37 degrees C. Neuraminidase did not augment splenic accessory cell function. Antigen-specific binding of T cells to Ia+ macrophages and accessory cell function may be enhanced by desialation of glycoproteins on the cell surface membrane.

The extent to which exogenous 18:3(n-3) and 18:3(n-6) were desaturated and elongated and the degree to which they and their derivatives altered the unsaturation index of cell glycerolipids were compared using clone 4 MDCK cells grown in lipid- and serum-free medium. Despite differences in the degree of unsaturation of the individual polyunsaturated fatty acids produced from 18:3(n-3) or 18:3(n-6), the unsaturation index of phospholipids increased similarly from 0.7 in control cells grown in serum- and lipid-free medium to ca. 1.6 in those supplemented with fatty acid. The added fatty acids had little effect on cell growth. The conversion of 18:3(n-6) to 20:3(n-6) and 20:4(n-6) was more rapid than that of 18:3(n-3) to 20:4(n-3) and 20:5(n-3). No significant quantities of 20:3(n-3) or 18:4(n-3) were noted. When both 18:3 isomers were supplied simultaneously, marked differences in the amounts of some species of n-3 and n-6 polyunsaturated fatty acids were observed. The presence of 18:3(n-6) and/or its derivatives suppressed levels of 20:4(n-3) and 20:5(n-3), perhaps through inhibition of the delta 6 and delta 5 desaturases responsible for their synthesis from 18:3(n-3). Similarly 18:3(n-3), and/or its longer more unsaturated derivatives, diminished the formation of 20:4(n-6) from 18:3(n-6). No marked effect on the products derived from elongation alone were observed.

The availability of techniques for establishment of primary, long term monolayers of breast milk macrophages and blood monocytes permitted a direct comparison of biosynthetic functions of human tissue macrophage and its progenitor. In addition to previously described morphological differences, four specific characteristics of the breast milk macrophage were identified: (i) a reduced rate of total protein secretion relative to the monocyte; (ii) abrogation of the 3 day lag in complement secretion regularly observed in blood monocyte cultures; (iii) an increase in the secretion of complement proteins C2 and factor B; and (iv) an increase in the ratio of C2 : factor B secretion. These differences did not result from cell-cell interactions between monocytes and macrophages, stable factors elucidated by monocyte or macrophage monolayers, or heat-stable factors in breast milk. In addition, both monocytes and macrophages synthesized and secreted C3 in an apparently native but haemolytically inactive form. The differences observed suggest that macrophages may modulate local availability of complement proteins in tissues or in the early phase of an inflammatory response.

Plasma proteins interact with vascular endothelium in such a way as to render it less permeable to other macromolecules. Evidence from a variety of sources indicates that this may result from interaction of the circulating macromolecules with the negatively charged glycoprotein layer on the surface of endothelial cells, and that this layer may be responsible for some of the known molecular sieving properties attributed to the endothelium. Experiments with the fluorocarbon exchange-transfused rat are described, which suggest that there may be mechanisms other than vesicular translocation that facilitate the passage of macromolecules across endothelium. Such mechanisms include, among others, the formation of transient transendothelial channels that appear to be less sensitive than pinocytotic vesicles to the concentration of ambient protein. Recent evidence suggests that, in addition to molecular size and charge, glycosylation of protein molecules and cell membranes themselves may facilitate vesicular uptake.

Histological studies have been performed on experimental acute renal failure induced by intravenous injection of hemoglobin in rats. These have been correlated with alterations in renal excretory function, assessed by the measurement of inulin clearance, at various stages of the lesion. The most prominent morphological changes during the first 24 hr after hemoglobin injection, when inulin clearance is most markedly suppressed, are: the presence of hemoglobin within the lumen of small intrarenal vessels, particularly the vasa recta; hemoglobin cast formation involving predominantly the thick ascending limbs of the loops of Henle; and evidence of injury of the epithelium of the proximal tubules and thick ascending limbs. Notably absent during this stage of the lesion are marked tubular dilatation, interstitial edema, and cast formation in the distal collecting ducts. The considerable recovery of function which occurs at 72 hr is accompanied by a marked reduction in involvement of the vasa recta. Standard sections and microdissection reveal many markedly dilated proximal tubules at this stage of the lesion, suggesting obstruction of filtering nephrons. These data have led to a tentative hypothesis regarding the pathogenesis of renal failure in this experimental lesion. It is suggested that renal ischemia and failure of glomerular filtration are the primary factors responsible for the early and severe impairment of renal function, and that these are related to intravascular aggregation of hemoglobin pigment. As this defect recedes, tubular obstruction by hemoglobin casts prevents restitution of excretory function in a variable fraction of the nephrons. The latter accounts for the relatively prolonged, moderate reduction in inulin clearance associated with the late stages of this lesion. These hypotheses form the basis for a continuing study of this renal lesion.

The permeability of the alveolar-capillary membrane of newborn and adult mice to horseradish peroxidase (HRP) and catalase was studied by means of ultrastructural cytochemistry, and the permeability to ferritin was studied by electron microscopy. The influence of varying volumes of intravenously injected fluid on the rate of leakage of the tracers from pulmonary capillaries was examined. The tracers were injected intravenously and the mice were sacrificed at timed intervals. Experiments on newborn mice with intranasally instilled HRP were also done. The tissues were fixed in formaldehyde-glutaraldehyde fixative. Chopped sections were incubated in Graham and Karnovsky's medium for peroxidase and in a modification of this medium for catalase. Tissues were postfixed in OsO(4) and processed for electron microscopy. In both newborn and adult mice, the ready passage of peroxidase through endothelial clefts was dependent on the injection of the tracer in large volumes of saline. When the tracer was injected in small volumes of saline, its passage through endothelial clefts was greatly reduced. Endothelial junctions of newborn mice were somewhat more permeable to HRP than those of adult mice. In all animals, alveolar epithelial junctions were impermeable to HRP. Catalase and ferritin did not pass through endothelial junctions. Intranasally instilled HRP in newborn mice was taken up by pinocytotic vesicles and tubules of flat alveolar cells.