Response of rat alveolar macrophages to ozone: quantitative assessment of population size, morphology, and proliferation following acute exposure

The purpose of this study was to evaluate the in vivo effects of an acute exposure to low levels of ozone on rat pulmonary alveolar macrophages (PAM). Fisher 344 rats exposed to 0.0, 0.12, 0.8, or 1.5 ppm O3 for 6 h were killed immediately after and 3, 18, 42, or 66 h after ozone exposure and their lungs were lavaged. Compared to sham-exposed (control) rats, exposure to 0.12 ppm O3 had no measurable effect on the total number, labeling index (LI), mitotic index (MI), or morphology of rat alveolar macrophages. The number of neutrophils was significantly (p less than or equal to 0.001) greater than in controls at 3, 18, and 42 h after exposure to 1.5 ppm O3 and 42 h after exposure to 0.8 ppm O3. The number of PAM was approximately twice that of controls 42 and 66 h after exposure to 0.8 and 1.5 ppm O3. There was a significant (p less than or equal to 0.001) increase in PAM MI 42 and 66 h after exposure to 1.5 ppm O3 and 42 h after 0.8 ppm O3. The increase in the number of PAM in mitosis was preceded by an increase in PAM LI. The PAM LI was significantly (p less than or equal to 0.001) greater than controls 18 and 42 h after exposure but returned to near normal levels by 66 h after exposure. There was a transient decrease in the mean nuclear/cytoplasmic ratio of PAM from rats exposed to 1.5 ppm O3 18 and 42 h after exposure due to an increase in the mean PAM cytoplasmic area. Comparison of the PAM population doubling time (Dt) and cell cycle time (Ct) suggest that PAM proliferation played a significant role in the observed increase in PAM following exposure to 0.8 and 1.5 ppm O3. These results highlight the dynamic response of PAM to an acute exposure to ozone and suggest that the proliferative response of pulmonary alveolar macrophages may be a useful indicator of pulmonary damage following inhalation of an irritant oxidant.     

Postnatal changes in lung phospholipids and alveolar macrophages in term newborn monkeys

In order to better understand the postnatal sequence of surfactant secretion and establishment of the alveolar macrophage (AM) population in newborn primates, healthy Macaca nemestrina monkeys were sacrificed during fetal life at term gestation (n = 5), or at 2 days (n = 5) or 3-4 weeks (n = 5) after term vaginal delivery. Excised lung tissue and left lung lavage were analyzed for phospholipid (PL) content, surface active material (SAM) extract, PL components, surface activity, pressure-volume characteristics, and AM number. Compared to term fetal animals, 2 day old term newborn monkeys were found to have a several-fold increase in lavage PL and SAM, and this was associated with greater maximal lung volume and drier lungs, but not improved deflation stability. During the subsequent 3-4 weeks of life, a 42% reduction in lung tissue stores of PL and SAM, and an 87% reduction in lavage PL and SAM were noted. Despite these major changes in quantity, there were relatively minor changes in the composition of the PL synthesized and released. The reduced quantity of SAM in the 3-4 week old animals led to a small decline in deflation stability. The several-fold increase in lavage PL and SAM during the first 2 days of life was accompanied by a 33-fold increase in AM; there was an additional 4-fold increase in AM number by 3-4 weeks of age. The abundance of lavage surfactant at 2 days of age may play a role in the influx of AM.     

Epithelial-mesenchymal interactions in postnatal rat lung growth

Epithelial-mesenchymal interactions have been implicated in epithelial cell differentiation in fetal lung. The role of such intercellular communication during postnatal lung growth is now evaluated to correlate intercellular contacts with cell division and with biochemical function of epithelial and interstitial cells. Sexed newborn rats, killed at frequent intervals to 8 weeks, received 3H-thymidine 1 h before death. A period of rapid cell proliferation between 3 and 10 days involved a five-fold increase in labeling index of both interstitial and type 2 epithelial cells. Hydroxyproline levels increased rapidly as interstitial cell division slowed, and epithelial growth was associated with elevated levels of disaturated phosphatidylcholine. Compared to the time of birth when epithelial cell division was slow, continuous basement membrane (BM) was more frequently found beneath type 2 cells during the postnatal proliferative phase, and fewer foot processes (FP) penetrated the BM; type 1 cells had few FP and uninterrupted BM. The number of intercellular contacts between type 2 cells and interstitial cells, one per two epithelial cell profiles at birth, decreased rapidly during the postnatal proliferative phase. There was a subsequent increase between 2 and 4 weeks that may reflect postmitotic epithelial differentiation. These observations support the hypothesis that transfer of mesenchymal factors may be important in the control of pulmonary epithelial growth and differentiation. The low incidence of cell-cell contacts seen after 4 weeks, one per five epithelial cell profiles, may reflect base levels of communication necessary to retain type 2 cell function in a resting cell population.     

Changes in lung tissue fibronectin content and synthesis during postnatal lung growth

We studied changes in lung tissue fibronectin content and synthesis during postnatal lung growth in rats. We reasoned that fibronectin, which is important in cell differentiation, migration, and adhesion, and in the organization of the extracellular matrix, might play a role in the rapid cell proliferation and alveolar septal formation that occurs postnatally in mammalian lungs. Newborn rats were sacrificed at 4, 7, 11, 14, and 21 days after birth. The lungs were perfused and lavaged, tissue fibronectin was extracted using urea and heparin (Bray et al, Science 1981; 214:793) and the extracted fibronectin was measured by enzyme-linked immunoassay. Tissue fibronectin synthesis was measured by the in vivo incorporation of 35S-methionine into fibronectin that was extracted from lung tissue and immunoprecipitated. Lavage fibronectin and albumin content and lung tissue collagen (hydroxyproline) content were also determined. Lung tissue fibronectin content per g dry lung almost doubled between days 4 and 7 after birth, was slightly higher at day 14 than at day 7, and decreased sharply between days 14 and 21. Lung tissue fibronectin synthesis per g dry lung increased steadily between days 4 and 14 to reach a peak value of about 2.5 times the 4-day value; it then decreased sharply between days 14 and 21. The period of increased fibronectin content and synthesis (4 to 14 days) coincided with the period during which lung cell proliferation and secondary alveolar septa formation are known to be the most active, and the sharp decrease in fibronectin content and synthesis (between 14 and 21 days) coincided with the period during which lung growth and remodeling markedly decrease.(ABSTRACT TRUNCATED AT 250 WORDS)     

Hepatocyte growth factor is required for alveologenesis in the neonatal rat

RATIONALE: Our core hypothesis is that growth factors that have dysregulated expression during experimental neonatal lung injury are likely to be involved in normal postnatal lung growth and alveologenesis.OBJECTIVES: To determine if hepatocyte growth factor (HGF) is upregulated in neonatal lung injury and is essential for postnatal alveologenesis.METHODS: A neonatal lung injury, in which there were patchy areas of interstitial thickening with a relative increase in the proportion of epithelial cells, was induced in newborn rats by exposing them to 60% oxygen for 14 days. Air-exposed pups had binding of endogenous HGF to its natural receptor, c-Met, inhibited by the intraperitoneal injection of either neutralizing antibody to HGF, or a truncated soluble c-Met receptor.MEASUREMENTS AND MAIN RESULTS: The 60% oxygen-mediated lung injury was associated with increased lung mRNAs for hepatocyte growth factor and c-Met, relative to air-exposed control lungs, at Day 7 after birth. After exposure to 60% oxygen, immunoreactive HGF was increased at Days 4 and 7, and immunoreactive c-Met was increased at Day 14. In air-exposed pups, intraperitoneal injections of neutralizing antibody to HGF inhibited DNA synthesis in alveoli-forming secondary crests, and reduced the number of alveoli in 6-day-old pups. Intraperitoneal injections of a truncated soluble c-Met receptor inhibited DNA synthesis in secondary crests in 4-day-old air-exposed rat pups.CONCLUSIONS: HGF and its c-Met receptor are required for normal postnatal alveolar formation from secondary crests, and are upregulated during 60% oxygen-induced neonatal lung injury.     

Inhaled ethyl nitrite prevents hyperoxia-impaired postnatal alveolar development in newborn rats

RATIONALE: Inhaled nitric oxide (NO) has been used to prevent bronchopulmonary dysplasia, but with variable results. Ethyl nitrite (ENO) forms S-nitrosothiols more readily than does NO, and resists higher-order nitrogen oxide formation. Because S-nitrosylation is a key pathway mediating many NO biological effects, treatment with inhaled ENO may better protect postnatal lung development from oxidative stress than NO. OBJECTIVES: To compare inhaled NO and ENO on hyperoxia-impaired postnatal lung development. METHODS: We treated newborn rats beginning at birth to air or 95% O(2) +/- 0.2-20.0 ppm ENO for 8 days, or to 10 ppm NO for 8 days. Pups treated with the optimum ENO dose, 10 ppm, and pups treated with 10 ppm NO were recovered in room air for 6 more days. MEASUREMENTS AND MAIN RESULTS: ENO and NO partly prevented 95% O(2)-induced airway neutrophil influx in lavage, but ENO had a greater effect than did NO in prevention of lung myeloperoxidase accumulation, and in expression of cytokine-induced neutrophil chemoattractant-1. Treatment with 10 ppm ENO, but not NO, for 8 days followed by recovery in air for 6 days prevented 95% O(2)-induced impairments of body weight, lung compliance, and alveolar development. CONCLUSIONS: Inhaled ENO conferred protection superior to inhaled NO against hyperoxia-induced inflammation. ENO prevented hyperoxia impairments of lung compliance and postnatal alveolar development in newborn rats.     

Lung Macrophage Differentiation Antigens in Developing Fetal and Newborn Rat Lungs: A Quantitative Flow Cytometric Analysis with Immunohistochemistry

Relative deficiencies in the number and function of alveolar macrophages (AMs) are present immediately after birth and contribute to increased susceptibility to infection. We used immunohistochemical localization with a panel of monoclonal antibodies to macrophage differentiation markers to characterize lung macrophage antigens relevant to development, and we present here the first study to quantitate these markers using flow cytometric analysis. Rat lung macrophages undergo immunophenotypic maturation seen by a changing number (OX-1, ED-1, ED-2, and ED-9) and distribution (ED-2) of certain lung macrophage differentiation antigens. Quantification of these antibodies revealed that labeling for ED-1 and ED-9 was less on lavageable AMs from early postnatal days than on mature adult AMs. In vitro treatment of adult rat AMs with murine granulocyte-macrophage colony-stimulating factor produces a proliferating population of AMs with a high proliferation index and an immature phenotype, similar to that of newborn AMs. This in vitro model was useful in validating our quantitation of neonatal lung macrophage differentiation antigens. Our quantitative studies of potential markers of AM differentiation in the developing lung may help to focus the study of macrophages in the developing lung on specific cellular and molecular pathways that may control immunologic events during this critical period. Accepted for publication: 4 February 1999     

Quantitative analysis of smooth endoplasmic reticulum proliferation in hepatocytes of early postnatal and adult mice treated with phenobarbital

The smooth endoplasmic reticulum in hepatocytes of early postnatal and adult ddY male mice injected with phenobarbital was analyzed stereologically. The animals received daily intraperitoneal injections of phenobarbital and were killed about 24 h after the last injection. In early postnatal animals (injected with 35 mg/kg of the drug at days 1 and 2 after birth, or injected with 35 mg/kg at day 2 and with 50 mg/kg at days 3 and 4 after birth), the smooth endoplasmic reticulum proliferated both in perivenular and periportal hepatocytes; there was no significant difference in amount of smooth endoplasmic reticulum between the cells of the two zones. In adult animals, however, proliferation occurred only in perivenular cells after the administration of 35 mg/kg of phenobarbital for 2 days, and predominantly in perivenular cells even after the administration of 150 mg/kg of the drug for 7 days. These results suggest that the conspicuous proliferation of the smooth endoplasmic reticulum in perivenular hepatocytes in response to phenobarbital administration, which is characteristic of adult liver, does not occur in early postnatal mice but becomes manifest during postnatal development.     

Postnatal development of rat lung following retarded fetal lung growth

Postnatal lung development was examined in rats born with smaller than normal lungs after either prenatal exposure to glucocorticoid or to an inhibitor of collagen synthesis. At birth, treated animals had lower than normal lung weights, lung to body weight ratios, hydroxyproline (HYP) levels, total DNA; and rates of DNA synthesis. Rats exposed to steroid showed a rapid recovery in growth during the normal postnatal cell proliferative phase from 4 to 11 days, though collagen levels did not return to normal until 3 weeks of age. Rats exposed to a prenatal proline analog showed a much slower rise in lung weight and total DNA, and these levels were still much below normal at 2-3 weeks when the cell proliferation phase was completed. Levels of disaturated phosphatidylcholine were significantly below normal up to 11 days, whereas total HYP was significantly reduced and less fibrillar collagen was seen in the lung throughout the study. The results indicate that the smaller but mature lungs at birth after antenatal steroid show a growth rebound and quickly become structurally normal. In contrast, inhibition of fibroblast growth and collagen deposition produces small lungs at birth, which continue to show inhibited growth and development at least up to 3 weeks of age, when the cell division phase is over.     

Transforming growth factor alpha (TGF(alpha)) is increased during hyperoxia and fibrosis

Transforming growth factor alpha (TGF(alpha)) stimulates type II alveolar epithelial cell proliferation and also is associated with fibrosis. We studied the changes in bronchoalveolar lavage (BAL) TGF(alpha) protein in a neonatal rabbit hyperoxia-fibrosis model (100% O(2) for 8 to 9 days, followed by 60% O(2) to 36 days of age). Hyperoxia increased TGF(alpha) protein and delayed the appearance of mature lower molecular weight (MW) TGF(alpha) isoforms at postnatal days 6 and 8 during the acute injury period. At 3 and 5 weeks, after chronic hyperoxia exposure, there was an increase in lower MW TGF(alpha) peptides during the fibrotic period. Keratinocyte growth factor (KGF) is also a type II cell mitogen. In vitro studies of keratinocytes suggest that KGF-induced proliferation is mediated through TGF(alpha). Intratracheal KGF instillation into adult wild-type and TGF(alpha)-null mice demonstrated that the KGF induced equivalent robust levels of proliferation in both TGF(alpha) deficient and wild-genotype mice. In conclusion, there are both quantitative and qualitative changes in TGF(alpha) protein in a hyperoxia-induced fibrosis neonatal rabbit model during periods of type II cell proliferation and fibrosis.     

Early postnatal development of pituitary intermediate lobe control in the rat by dopamine neurons

In fetal and postnatal rats, serum melanophore-stimulating hormone (MSH) levels, pituitary 3H-spiperone binding, pituitary catecholamine content, and other parameters were measured to characterize the development of pars intermedia pituitary control by the tubero-hypophyseal dopamine system. Bioassay of serum for MSH revealed low but significant levels 6 days before birth (gestational day 17), rising to a peak 4 days after birth (postnatal day 5). Levels declined after PN 6, reached a low 2 weeks after birth, and then rose gradually throughout the next several weeks until adult values were attained. From excised pituitaries, significant catecholamine fluorescence in the neurointermediate lobe was first demonstrated on the 2nd postnatal day, with the highest density of fibers being attained at 2 weeks after birth. Thereafter, the density of terminals decreased in the pars intermedia until the adult pattern of innervation was observed at day 60. Total dopamine content of the neurointermediate lobe roughly paralleled the morphological observations through the first 2 weeks after birth. Later, content was seen to stabilize, while concentration decreased in line with the reduction in terminal density. Dopamine levels exceeded norepinephrine concentration by a factor of 10. 3H-spiperone binding sites were present in the pituitary before birth. Their density increased rapidly in the neurointermediate lobe during the 2 postnatal weeks, then diminished until adult densities were reached at day 30. Binding in the anterior pituitary slowly rose throughout the first 30 days after birth, although values were only about one-quarter of that found in the neurointermediate lobe. In vitro incubation of neurointermediate lobes with dopamine showed a dose-dependent inhibition of spontaneous MSH secretion as early as 2 days after birth. Blockade of dopaminergic binding sites by i.p. administration of cis-flupenthixol (1 mg/kg) to rat pups had no effect until days 8, 15, and 22 where there was a significant rise in serum MSH levels as compared with controls. These data indicate that the intermediate lobe is responsive to dopamine at around birth, while the inhibitory dopaminergic control develops only during the 1st postnatal week, in conjunction with marked changes in circulating MSH levels.     

Regulation of neonatal and adult mammalian heart regeneration by the miR-15 family

We recently identified a brief time period during postnatal development when the mammalian heart retains significant regenerative potential after amputation of the ventricular apex. However, one major unresolved question is whether the neonatal mouse heart can also regenerate in response to myocardial ischemia, the most common antecedent of heart failure in humans. Here, we induced ischemic myocardial infarction (MI) in 1-d-old mice and found that this results in extensive myocardial necrosis and systolic dysfunction. Remarkably, the neonatal heart mounted a robust regenerative response, through proliferation of preexisting cardiomyocytes, resulting in full functional recovery within 21 d. Moreover, we show that the miR-15 family of microRNAs modulates neonatal heart regeneration through inhibition of postnatal cardiomyocyte proliferation. Finally, we demonstrate that inhibition of the miR-15 family from an early postnatal age until adulthood increases myocyte proliferation in the adult heart and improves left ventricular systolic function after adult MI. We conclude that the neonatal mammalian heart can regenerate after myocardial infarction through proliferation of preexisting cardiomyocytes and that the miR-15 family contributes to postnatal loss of cardiac regenerative capacity.     

Characterization of the bronchoalveolar cellular response in experimental asbestosis. Different reactions depending on the fibrogenic potential

Analysis of bronchoalveolar cell types and structure was performed during the development of asbestos-induced lung injury in the rat. Animals received single intratracheal injections of one of the following: saline (control), UICC chrysotile B asbestos (5 mg), or very short 4T30 chrysotile fibers (5 mg). Bronchoalveolar lavage (BAL) was performed at various intervals after instillation. Analysis of BAL fluid showed significant increase in inflammatory cells in response to asbestos, which persisted longer in animals treated with chrysotile B. Presence of numerous mitotic figures in BAL fluid of treated animals suggests that macrophage replication may contribute in part to this response. Differential cellular analysis indicated that after injection of long chrysotile fibers, which causes fibrotic lesions within 7 days, polymorphonuclear leukocytes (PMN) appear as early as Day 1 in significant concentration (40%) in the bronchoalveolar compartment and persist through Day 7 after treatment. From Day 7 to Day 21, multinucleated cells (MGC) were found in lavage fluid (5 to 8%). Most of these cells were binucleated, and none had more than 3 nuclei. By contrast, exposure to very short chrysotile fibers caused only a very transient influx of PMN on Day 1. By Day 7, there was a significant increase in MGC, which persisted through Day 21, at which time no fibrosis was apparent. Although most of these cells were binucleated, many cells had 3 or more nuclei. The giant cells were predominantly of the foreign body type, with MGC of the Langhans type also present.(ABSTRACT TRUNCATED AT 250 WORDS)     

Altered breathing pattern after prenatal nicotine exposure in the young lamb

Maternal smoking during pregnancy is a risk factor for sudden fetal and infant death as well as obstructive airway disease in childhood. Fetal nicotine exposure affects organ development. The aim of the present study was to investigate effects of fetal nicotine exposure on lung function in young lambs. Nine unanesthetized, awake, prenatally nicotine-exposed lambs (N) (approximate maternal dose: 0.5 mg/kg) and 12 nonexposed control lambs (C) were studied repeatedly for 5 weeks after birth using a pneumotachograph and a computerized method for breath-by-breath determinations. N and C lambs had similar minute ventilation but a markedly different breathing pattern. At both 5 and 21 days, average age, N lambs had significantly lower tidal volumes and higher respiratory rates than C lambs. Inspiratory drive (P(0.1)) and effective impedance were significantly higher in N lambs compared with C lambs only at 5 days. Prenatal nicotine exposure appears to have long-term effects on the postnatal breathing pattern, suggesting altered lung function, e.g., increased airway resistance, decreased lung compliance, or both. The increased inspiratory drive is most likely secondary to increased impedance of the respiratory system. These changes are most marked close to birth but persist during the initial postnatal period.     

Collagen degradation during postnatal lung growth in rats.

Postnatal lung growth involves remodeling of the structure seen at birth as new alveoli are formed. To determine the role of collagen degradation in this process, in particular of the basement membrane component, we studied the synthesis of total collagen and the degradation of collagen types I or IV in a series of rats from birth to 29 days of age. During the period of rapid cell proliferation to day 11, the collagen level per mg lung did not change though the rate of synthesis increased. Up to 40% of new collagen was rapidly degraded. At the end of the growth phase, the interstitium became thinner and less cellular. Collagen synthesis slowly decreased as the total collagen content increased in the lung, and less than 20% of newly synthesized collagen was degraded. Type I collagenase activity was highest during the cell proliferation phase, though less than 20% was due to active enzyme. In contrast, type IV collagen breakdown, also maximal in the first 11 days, was almost all due to enzymes present in the active form. The results demonstrate that rapid degradation of collagen, particularly the type IV form present in basement membranes, occurs during the early phase of postnatal lung growth.     

Improvement of lung compliance during postnatal adaptation correlates with airway sodium transport

RATIONALE: Fetal lung liquid secretion is coupled with chloride transport into the lung lumen. The postnatal clearance of lung liquid is dependent on osmotic force generated by active sodium absorption. OBJECTIVE: To study the interaction between airway epithelial sodium transport and postnatal lung function. METHODS: We determined lung compliance and nasal transepithelial potential difference as a measure of airway ion transport and epithelial sodium channel gene expression in 41 healthy newborn infants during the first 50 h after birth. MEASUREMENTS AND MAIN RESULTS: Lung compliance improved significantly during the study period, whereas nasal potential difference remained constant. There was a significant decrease in the expressions of beta and gamma subunits of the epithelial sodium channel. A positive correlation existed between amiloride-sensitive nasal potential difference measured at 1-4 h of age and lung compliance at 21-27 h of age. We found no correlation between the molecular data and functional measurements. CONCLUSIONS: An important part of pulmonary adaptation takes place during the first hour after birth. The improvement of lung compliance continues over the first postnatal days and coincides with down-regulation of epithelial sodium channel beta and gamma subunit expression.     

Effects of fetal growth restriction on lung development before and after birth: a morphometric analysis

Our aim was to determine the effects of fetal growth restriction (FGR) during late gestation on the structure of the lungs in the fetus near term and at 8 weeks after birth. The studies were performed using two groups of pregnant sheep and their offspring. In both groups, FGR was induced by umbilico-placental embolisation (UPE); for fetal studies, UPE was performed from 120 days of gestation until 140 days (term, approximately 146 days), when fetuses were killed for tissue analysis. For postnatal studies, UPE continued from 120 days until delivery at term; postnatal lambs were killed at 8 weeks after birth for tissue analysis. UPE led to a thicker pulmonary blood-air barrier at 140 days of gestation and this difference, which was due to a thickened basement membrane, was still present at 8 weeks after birth. At 8 weeks, we also observed a smaller number of alveoli per respiratory unit, thicker interalveolar septa, and a greater volume density of lung tissue in FGR lambs compared to controls. These changes would be expected to impair gas exchange and alter the mechanical properties of the lungs. Our data show that structural alterations in the lungs induced by placental insufficiency were more evident at 8 weeks of postnatal age than near term, indicating that the effects of FGR on the lung may become more serious with age and may affect respiratory health later in life.     

A comparison of lymphocyte populations in lung tissue and in bronchoalveolar lavage fluid of rats at various times during the development of bleomycin-induced pulmonary fibrosis

The purpose of this study was to compare lymphocyte populations in lung lavage fluid and in lung tissue during the development of bleomycin-induced pulmonary fibrosis. At various times after bleomycin treatment, animals were killed and lavaged, and lung tissue cells were extracted. Lymphocyte populations were identified by immunofluorescent techniques, using monoclonal antibodies, and quantitated on the fluorescence-activated cell sorter. Normal control rat lung tissue was comprised of 91% T-cells (helper-to-suppressor cell ratio of approximately 1:1) and 9% B-cells. After bleomycin administration, a significant increase in the percentage of B-cells in lung tissue was observed within 3 days, with peak levels (approximately 28%) occurring at 7 days. Changes in the T-cell subset populations were also observed in lung tissue; at 14 days after the administration of bleomycin, the helper-to-suppressor T-cell ratio was 2:1, at 30 and at 120 days, the ratio was reversed to 1:2. These represent significant changes from the 1:1 ratio found in control animals. No lymphocytes were observed in the lavage fluid of control animals or in animals at 30 and 120 days after bleomycin treatment. Significant populations of lymphocytes were found in the lavage fluid at 3, 7, and 14 days after bleomycin treatment. These lymphocytes consisted of approximately 90% T-cells (helper-to-suppressor cell ratio of 1:1) and 10% B-cells. There were no significant changes in the lymphocyte populations found in the lavage fluid after the administration of bleomycin. The results demonstrate that specific lymphocyte populations are changing in lung tissue during the development of the fibrotic process, whereas this shift in populations does not occur in the lymphocyte populations found in lavage fluid.     

Comparison of the pulmonary surfactant content in alveolar macrophages of newborn, young, and adult rats.

The phospholipid composition of the 150-g pellet containing macrophages from neonatal lung lavages resembles that of surfactant. To study whether this composition reflects the surfactant content of the macrophage, we isolated the alveolar phospholipids and macrophages from the lavage fluids of fetal, newborn, young, and adult Wistar rats. The alveolar surfactant phospholipids increased from fetal levels of 2.8 nmol/mg dry lung weight (DLW) to 39 nmol/mg DLW at day 1, decreased sharply within the first week, and stabilized at a level of 2-4 nmol/mg DLW after day 15. The number of alveolar macrophages increased significantly during the first postnatal day from approximately 750 to more than 5000 (per mg DLW), decreased during the next 4 days, and varied strongly at older ages. We estimated the surfactant content in the macrophages semiquantitatively by polarization microscopy. Birefringence augmented significantly during the first 1.5 days after birth and decreased after that concurrently with the amount of alveolar surfactant. However, only cells without birefringent inclusions sedimented at 150g, whereas the phospholipid composition of the pellets falsely suggested that large amounts of intracellular surfactant were present in its cells. At least two populations of macrophages (surfactant-rich and surfactant-poor) are present in the growing animal. We suggest that differences in function of these various types of macrophages also might depend on surfactant congestion.