Expression of the insulin-like growth factor system in postpneumonectomy lung growth.

The insulin-like growth factors (IGF-I and IGF-II) may play an important role in postpneumonectomy compensatory lung growth by translating hormonal inputs and mechanical forces into cellular proliferation signals. We examined the mRNA abundance of IGF-I, IGF-II, and IGF binding proteins (IGFBPs) in lungs of rats on postoperative days 1, 2, 3, 5, and 7 following left pneumonectomy (PNX) or shamoperation (SC) and in normal animals (CON). There was no difference in the abundance of lung IGF-I mRNA (measured by Northern analysis) or serum IGF-I (measured by radioimmunoassay (RIA)) between SC and PNX animals. IGF-II mRNA abundance was initially decreased following PNX (73% decrease compared to SC animals on day 1, p < .05) and then rose to approach SC group values on subsequent days. Transcripts for IGFBP-2, -3, -4, -5, and -6 were decreased in both the SC and PNX groups compared to CON animals on the day following pneumonectomy, then rose back to baseline by postoperative day 2-3. Tissue IGFBPs, measured by ligand blot analyses, were not different in either the SC or PNX groups. In contrast, all serum IGFBP bands were increased on postoperative day 1 following either sham or PNX surgery. In addition, serum IGFBP-4 was increased in PNX animals compared to the SC group on days 1 and 2 (increase of 38% and 78%, respectively, p < .05). We conclude that the changes observed in lung IGF and IGFBP expression following pneumonectomy do not represent major.

Comparative biochemistry of gestational and postnatal lung growth and development in the rat and human.

We compared the ontogeny of collagen (hydroxyproline), elastin (desmosine), soluble protein, and DNA in the lungs of rate and humans during gestation and postnatal life. In humans, lung weight/body weight ratios declined faster during gestation than postnatally, whereas in rats lung weight/body weight ratio declined little during gestation and then suddenly on the first day of life. Lung weight/body weight ratios may be lower than expected around term in humans, and prediction data are given to assess human pulmonary hypoplasia. Rats and humans differed in water content of their lungs, with rats showing a sharper decline during gestation. In the human lung, collagen and elastin made their appearance at an early stage of gestation; elastin. In particular, increased rapidly during gestation, suggesting a role in intrauterine alveolar formation. In the rat, elastin accumulation is primarily a postnatal event, as is alveolar formation. Hydroxyproline concentrations increased with conceptual age and continued to increase rapidly postnatally between 4 and 7 weeks in the rat, but slowed in the human after 60 weeks of conceptual age. Desmosine concentrations level off at the end of the study period in rats, while these are still increasing, although slowly, in humans. Overall lung growth, as assessed by weight, was linear in humans, but phases of lung growth were apparent in the rat, including one of minimal growth in the immediate postnatal period.

Effect of triamcinolone acetonide on the development of the pulmonary airways in the fetal rat.

Triamcinolone acetonide (TAC) has a potent teratogenic effect on various mammalian fetal tissues as well as a steroid effect on the lung. Less well documented is the fact that it produces profound oligohydramnios. We wished to determine what effect TAC would have on branching morphogenesis and other aspects of lung development, using an in vivo model described previously. Thirty rats were randomized to receive 0.6 mg/kg of TAC or saline on days 12, 13, and 14 of gestation. At gestational days 15, 17, 18, and 21, the left lungs of 365 fetuses were studied by dissecting microscopy, histology, and morphometry. TAC produced profound pulmonary hypoplasia (dry Jung weight/body weight 0.025, compared with 0.06 in controls) on day 21. TAC decreased maternal weight gain, fetal weight, placental weight, aminiotic fluid, and pole to pole length (PTP), while it increased the peripheral airway count (PAC). The number of central and intermediate airway branches was reduced, and they were dilated. Growth of peripheral airways was enhanced. In treated fetuses epithelial cells lining these airspaces were histologically more mature and the mesenchyme thinner than in controls. These findings were confirmed by the morphometric measurements. We conclude that when TAC is administered in the early phase of fetal rat lung development, the lungs become hypoplastic, with hypoplasia of the intermediate airways, an increase in the number of peripheral airways, and increased differentiation. We speculate that these effects are primarily due to the steroid action of TAC and that the mechanisms of monopodial branching are different from those of dichotomous branching.

Two different patterns of airway branching regulated by different components of the extracellular matrix in vitro.

This study examined the effects of beta-D-xyloside (an inhibitor of proteoglycan synthesis) and cis-4-hydroxyl-L-proline (an inhibitor of collagen synthesis) on branching morphogenesis in cultures of fetal rat lung. Lungs from day 15 gestation were incubated for 4 days in (1) the control medium (Dulbecco's Modified Eagle Medium + 10% Fetal Bovine Serum) alone (control), (2) control medium plus 2 mM beta-D-xyloside (beta-XYL), (3) control medium plus 2 mM alpha-D-xyloside (alpha-XYL), (4) control medium plus 50 micrograms/mL cis-4-hydroxy-L-proline (cis-HYP). The number of peripheral buds of left lungs was counted daily. Histological examination was performed on lungs on days 2 and 4. beta-XYL inhibited proximal monopodial branching on day 2 without affecting lung size, but produced numerous peripheral buds on day 4 which were of abnormal appearance, suggesting that lung airway branching and growth may be regulated by different mechanisms. Histology and morphometry showed significantly enlarged airspaces and diminished mesenchyme. cis-HYP did not affect proximal branching on 2 days in culture, but inhibited further dichotomous branching and lung growth after 2 days. On day 4, diminished branching and lung growth was accompanied by a proportional decrease in mesenchyme. The difference between effects of beta-XYL and cis-HYP on the branching pattern suggests that proteoglycans and collagen are involved in different patterns of branching morphogenesis.

Lung morphometric changes after exposure to hypobaria and/or hypoxia and undernutrition.

Lung morphometry was studied in rats between 4 and 7 weeks of age. The animals were divided into 5 groups: general controls (fed ad libitum), hypobaric normoxic, normobaric hypoxic, hypobaric hypoxic, and weight-matched controls (weight matched to the hypobaric hypoxic group). In both hypobaric and normobaric hypoxia, lung volume, alveolar surface area and total alveolar number increased compared to weight-matched controls. In normobaric hypoxia, mean linear intercept, mean chord length of alveoli increased and number of alveoli/unit volume decreased compared to weight-matched animals. In hypobaric hypoxia, only mean chord length increased. Dysanaptic index decreased in both. In hypobaric normoxia, alveolar size and lung volume diminished compared to general controls. Lung growth was impaired in weight-matched controls without affecting airspace dimensions. Hypobaric and normobaric hypoxia increase lung growth overcoming nutritional effects but is dysanaptic. Lung growth in hypobaric hypoxia is mainly determined by low oxygen but low pressure may also produce subtle structural alterations.

Biochemical, clinical, and morphologic studies on lungs of infants with bronchopulmonary dysplasia.

We correlated clinical, biochemical, and morphologic findings in the lungs of 48 infants dying of either bronchopulmonary dysplasia (BPD) or hyaline membrane disease (HMD) to obtain a better idea of the disease process. The infants ranged from 24 weeks of gestation to 1 1/2 postnatal years. The lungs of BPD and HMD infants had higher contents of DNA, alkalisoluble protein, hydroxyproline, and desmosine, as well as increased concentrations of DNA, hydroxyproline, and desmosine when compared with the lungs of 72 control infants. BPD was classified histologically into 4 groups: Group I was a phase of acute lung injury, Group II the proliferative phase; Group III the phase of early repair, and Group IV the phase of late repair. We saw a significant increase in hydroxyproline concentration in Groups II and III. The ratio of type I/III collagen decreased in BPD Groups II to IV. Desmosine was significantly higher only in Group III than in controls. When the pathological classification was related to biochemical and clinical features of BPD, the classification showed dependence on the number of days the infant survived postnatally and not on the gestational age of the infant. The number of days on assisted ventilation was a slightly better predictor of the disease classification than days on > 60% oxygen. A statistical model correctly predicted the pathologic classification 83% of the time.

Time course of lung growth following exposure to hypobaria and/or hypoxia in rats.

Four-week-old rats were divided into five groups: general controls, weight-matched controls (weight matched to hypobaric hypoxia), hypobaric hypoxia, normobaric hypoxia, and hypobaric normoxia. Lung growth impairment in weight-matched animals occurred by reduction in cell number and size. In both hypoxic groups, lung weight, RNA and protein were significantly higher on day 3, and DNA on day 5 and remained higher thereafter. Maximum 3H-TdR incorporation occurred on day 3 in both hypoxic groups. Hypoxia increased RNA/DNA ratio on day 1 and protein/DNA on day 3. Following 3 days of recovery, DNA synthesis and RNA/DNA ratio of hypoxic groups and controls were identical. DNA synthesis also doubled on day 5 in hypobaric normoxia compared to general controls. Hypoxia up regulates lung growth despite down regulation by undernutrition. Maximum lung growth stimulation occurs during early exposure by cellular hypertrophy followed by hyperplasia. Low pressure by itself also stimulates lung growth. Cellular activity returns immediately to normal levels after removal of hypoxic stimulus.

Development of the pulmonary airways in the fetal rat and its relation to the prenatal environment.

We studied the left lung using multi-focus microphotography in 378 rat fetuses, assessing airway branching from day 13 to day 19 of gestation, and lung growth variables from day 13 to day 21. Longitudinal growth, and monopodial and dichotomous branching brought about a consistent airway pattern with variations within each day of gestation and a small overlap between adjacent days. Amniotic fluid weight and pole to pole (PTP) distance of the lung increased quadratically with age, while fetal weight and the peripheral airway count (PAC) increased exponentially. The location of the fetus within the uterus had no effect on fetal variables, but correlations were found between maternal weight gain and both fetal weight and PTP. Fetal weight was the best predictor of PAC from gestational ages 15 to 19 days (P < 0.008). The method described allows for observations that are reproducible within the environmental variations present in normal gestation and can be used to study the effect of external factors on lung development.

Cytodynamics of in vitro developing airways and interaction with extracellular matrix proteins.

We studied epithelial and mesenchymal cell kinetics during lung branching and the possible importance of proteoglycan and collagen in rat fetal organ culture for 5 days, starting from 15 days gestation. The lungs were randomly divided into three groups: (1) basal medium (controls); (2) basal medium + beta-D-xyloside (beta-XYL); and (3) basal medium+cis-4-hydroxy-L-proline (cis-HYP). The labeling index of incorporation of 5-bromo-2-deoxyuridine (BrdU) was used as an indicator of DNA synthesis and assessed in the parent airways and in the tips and sides of peripheral buds. In controls, BrdU incorporation was highest in bud tips, followed by bud sides and then parent airway for both epithelium and mesenchyme. These data suggest that preferential cell proliferation occurs in normal airway branching. In parent airways, labeling of epithelium and mesenchyme was consistently diminished throughout the culture period in beta-XYL, whereas in cis-HYP this was not the case. In the peripheral buds, the DNA synthetic activity in both epithelium and mesenchyme was impaired in beta-XYL and cis-HYP, but the effect was more marked in beta-XYL. We conclude that although collagen may be required in peripheral bud morphogenesis, the proteoglycans appear to be essential in growth and development of both parent airways and peripheral buds in the lung.

Lung cytokinetics after exposure to hypobaria and/or hypoxia and undernutrition in growing rats.

Lung cellular dynamics were examined in growing rats from 4 to 7 wk of age after exposure to 1) room air (general controls), 2) hypobaric normoxia, 3) normobaric hypoxia, 4) hypobaric hypoxia, and 5) room air and restricted food intake (weight-matched controls). Tritiated thymidine ([3H]TdR) incorporation diminished in weight-matched controls. In both hypoxic groups, maximum [3H]TdR incorporation occurred on day 3 in all cells of peripheral alveoli, capillary endothelium of central alveoli, airway walls and epithelium, and arterial wall and endothelium, but maximum [3H]TdR labeling of interstitial and unidentifiable cells of central alveoli occurred on day 5. The percent labeling with [3H]TdR was higher in cells of peripheral alveolar walls than in cells of central alveolar walls. Labeling of the interstitium was higher in hypobaric hypoxic than normobaric hypoxic rats. In hypobaric normoxia, DNA synthetic activity increased in alveolar wall cells, except for capillary endothelium. In hypobaric hypoxia, DNA synthesis occurred primarily because of low O2, but low pressure may also affect cytokinetics. [3H]TdR incorporation is greater and earlier in the alveoli of the peripheral part of the lung than central alveoli, and the cellular response of the various cells types is not synchronous.

Effects of streptozotocin-induced diabetes on postpneumonectomy lung growth and connective tissue levels.

Diabetes is generally accompanied by abnormal levels of growth hormone and adrenal steroids, hormones known to modulate postpneumonectomy (post-PNX) compensatory lung growth. Thus, we examined the possibility that diabetes may affect post-PNX lung growth processes. Left PNX was performed in young diabetic rats (streptozotocin-induced; 75 mg/kg body weight) (DM-PNX) and in control rats (C-PNX), for comparison with sham-operated control rats (C-SHAM). The rats were permitted free access to food and water. Examination at day 7 after surgery showed that right lung absolute dry weight and absolute DNA, collagen and elastin contents were increased in C-PNX and DM-PNX rats (but only C-PNX values reached those of both lungs in C-SHAM rats). Body weights (BW) of DM-PNX rats were lower than those of C-PNX and C-SHAM rats. Lung DNA/BW in C-PNX and DM-PNX rats were comparable, and matched values for both lungs in C-SHAM rats. Lung dry weight/BW, collagen/BW, and elastin/BW in DM-PNX rats exceeded values in C-PNX rats and even more for values of both lungs in C-SHAM rats. Data of another experiment, comparing DM-PNX rats with body weight-matched (by food limitation) PNX and control rats (WMC-PNX and WMC-SHAM rats, respectively) indicated comparable lung absolute DNA contents in DM-PNX and WMC-PNX rats (which matched values for both lungs in WMC-SHAM rats), however, lung absolute collagen and elastic contents were greater in the DM-PNX rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Pulmonary function alterations after 3 wk of exposure to hypobaria and/or hypoxia in growing rats.

We studied lung growth in rats between 4 and 7 wk of age under different conditions. There were five groups, seven animals in each: 1) general controls (ambient pressure and room air, food ad libitum); 2) hypobaric normoxic [barometric pressure (PB) 410 mmHg, PO2 153 Torr]; 3) normobaric hypoxic (ambient pressure, PO2 80 Torr); 4) hypobaric hypoxic (PB 410 mmHg, PO2 80 Torr); and 5) weight-matched controls to hypobaric hypoxic. Residual volume, functional residual capacity, vital capacity, and total lung capacity grew 10-20% more in both hypoxic groups than in weight-matched and general controls. Expiratory flow rates corrected for forced vital capacity decreased, and specific airway resistance increased significantly. In addition, the ratio of forced expiratory volume in 0.1 s to %forced vital capacity, peak expiratory flow rate, and forced maximal midexpiratory flow were also lower in normobaric hypoxic animals compared with weight-matched controls. Above a transpulmonary pressure of 6 cmH2O, flows were reduced in both hypoxic groups. No differences were observed between hypobaric normoxic and general control groups for lung volume and lung function. In weight-matched animals, total lung capacity decreased but lung function remained unchanged. We conclude that accelerated lung growth in hypobaric hypoxia and normobaric hypoxia is dysanaptic. Lung growth in hypobaric hypoxia is primarily induced by low oxygen, but differences between hypobaric hypoxia and normobaric hypoxia suggest a beneficial effect of low pressure.

Measurement of emphysema in autopsy lungs, with emphasis on interlobar differences.

This study clarifies interlobar differences in the severity of emphysema and examines the feasibility of assessing emphysema in a whole lung using either the upper or lower lobes. Midsagittal, paper-mounted whole-lung sections from 79 consecutive autopsies were used. The whole-lung sections were scored by comparing them with the panel of standard pictures from Grades 0 to 100, after which the upper or lower lobes were scored in the same way but with the other lobe covered. Scores made on lobes and whole lungs correlated significantly, but while projections to whole-lung scores were quite accurate from the upper lobe, the projections from the lower lobe were less precise. This was particularly true in patients with mild emphysema. Individual lobes of the same lung sections were assessed by the Ryder grid method. This analysis showed that emphysema was more severe in the upper lobe of the 79 consecutive cases. The panel grading method showed more severe emphysema in the upper lobe than in the lower lobe to a panel score of 40; after this, the severity of emphysema in the two lobes was similar. Scores made from the Ryder grid were very similar to those made from the panel. Grading emphysema from one lobe, particularly the lower lobe, is not appropriate if precise clinicopathologic correlations are required.

Correlation of structure and function in idiopathic pulmonary fibrosis.

The early stage of idiopathic pulmonary fibrosis (IPF) is thought to involve a smaller number of alveoli and to be characterized predominantly by cellularity and minimal fibrosis, whereas advanced disease involves a large number of alveoli and is characterized predominantly by fibrosis with minimal cellularity. In addition, correlative studies have indicated that prognosis and response to therapy is determined in part by the extent of fibrosis and cellularity. This study was undertaken to determine whether pulmonary function assessment would help distinguish between the cellular and fibrotic phases of this disorder, as determined by a semiquantitative pathology scoring system that comprised four factor scores: fibrosis, cellularity, granulation/connective tissue, and desquamation. Ninety-six untreated patients with biopsy-confirmed IPF (27 never smokers, 32 current smokers, and 37 ex-smokers) were evaluated. In the group as a whole, there was no significant relationship between the fibrosis or the connective/granulation tissue factor scores and any of the physiologic parameters. The DLCO correlated with the "desquamation" and the total pathology scores, whereas the TLC and FVC correlated with the cellularity factor score. In the current smokers, the coefficient of elastic retraction, DLCO/VA, and FEV1/FVC ratio were significantly lower than in never smokers and ex-smokers, and TLC and FVC were higher than in never smokers. Also, the mean cellularity and granulation/connective tissue factor scores were significantly lower, and the desquamation factor score was significantly higher than those in never smokers and ex-smokers. Both age and smoking status were significant for the cellularity factor score, whereas for the connective/granulation tissue factor score, age was not significant but smoking status was.(ABSTRACT TRUNCATED AT 250 WORDS)

Responsiveness and variability of airflow obstruction in chronic obstructive pulmonary disease. Clinicopathologic correlative studies.

We have studied the relationships between pulmonary lesions and bronchodilator response and variability of FEV1 in 41 patients enrolled in the National Institutes of Health Intermittent Positive Pressure Breathing Trial who died, came to autopsy, and provided adequate tissue to quantitate lesions. The patients had moderate to severe chronic airflow obstruction and various degrees of response to 250 micrograms isoproterenol inhalation. Airway responsiveness was positively correlated with bronchial eosinophilia, bronchial inflammation, and bronchiolar fibrosis, and it was negatively correlated with bronchiolar goblet cell metaplasia and emphysema. Patients with an increase of 190 ml or more in FEV1 after bronchodilator had less bronchial cartilage and less goblet metaplasia in bronchioles. Airway smooth muscle was not related to airway responsiveness and variability. Flow rates were adversely affected by bronchial eosinophilia for given emphysema scores. This study shows the importance of the eosinophil as part of chronic nonspecific lung disease. Lack of airway responsiveness was associated with lesions such as emphysema and goblet cell metaplasia, which by themselves cause severe chronic airflow obstruction. The better-preserved lung function in patients with increased airway responsiveness is attributed to negative correlations with emphysema and positive correlations with bronchial eosinophilia.

Effect of indomethacin on lung development in postnatal rats: possible role of prostaglandin in alveolar formation.

We administered 1.3 mg ip of indomethacin, a prostaglandin synthetase inhibitor, per 100 g body wt to male rat pups daily on postnatal days 4-13 and examined their lungs on day 14. Indomethacin administration produced abnormal lung structure with diminished alveolar air, increased alveolar duct air, increased mean linear intercept (gas-exchanging wall distance), diminished gas-exchanging surface area and surface-to-volume ratio, increased septal wall thickness, diminished the number of alveolar crests, and increased the number of lamellar bodies in alveolar type II cells. However, this procedure did not alter quantitative lung growth (normal lung weights, volumes, and DNA and protein contents). Tissue prostaglandin content was decreased. The total amount of lung collagen or elastin was unchanged, but when collagen was analyzed into soluble and insoluble components, soluble collagen was increased. Supplementation with 1.0 g of prostaglandin E2 per 100 g body wt to animals treated with indomethacin reduced the abnormalities in pulmonary architecture. We conclude that indomethacin affects lung structure in growing rats and that it is an unusual model in that lung growth is normal, but lung development is abnormal. We also suggest that prostaglandins may play a significant role in alveolar formation in postnatal lung development in rats.

Lung growth in hypobaric normoxia, normobaric hypoxia, and hypobaric hypoxia in growing rats. I. Biochemistry.

Adaptive changes in cellular and connective tissue components of the lung after chronic exposure to reduced ambient oxygen and/or pressure were studied. Four-week-old male Sprague-Dawley rats were randomly divided into five groups (n = 12 each): 1) general control, room air (GC); 2) hypobaric normoxic; 3) normobaric hypoxic; 4) hypobaric hypoxic; and 5) weight-matched control, restricted food intake (WMC; weight matched to hypobaric hypoxic animals). Lung growth (lung weight and DNA, RNA, protein, hydroxyproline, and desmosine contents) diminished in WMC compared with GC. Somatic growth decreased in hypobaric and normobaric hypoxic rats compared with GC. Lung weight; DNA, RNA, protein, hydroxyproline, and desmosine contents; and RNA/DNA, protein/DNA, and desmosine/DNA ratios increased in both hypobaric and normobaric hypoxic rats compared with WMC. Hydroxyproline and desmosine contents and the hydroxyproline/DNA ratio were significantly higher in hypobaric than normobaric hypoxic rats. Hypobaric normoxia caused a slight somatic growth reduction, but biochemical parameters of lung growth remained unaffected. In conclusion, in growing animals, despite inhibition of lung growth due to reduced food consumption, accelerated lung growth in hypobaric or normobaric hypoxia occurs by hyperplastic and hypertrophic changes. Hypobaric normoxia does not affect lung growth, but connective tissue proteins accumulate slightly more in hypobaric hypoxia than in hypoxia alone.

Scanning electronmicroscopic morphometry of emphysema in humans.

We quantitated the holes in alveolar walls in 11 nonemphysematous lungs and in 11 lungs with mild emphysema, all of which were removed at surgery. We found that in the nonemphysematous lungs, 94.1% of the holes were smaller than 10 microns in diameter and only 0.2% were larger than 20 microns. In the lung parenchyma distant from emphysema, both the maximum diameter of the holes and the diameter of alveoli increased. In the parenchyma between emphysema, the areas of alveolar walls represented by holes also increased, as did the average hole area and number of holes per alveolus. We found that alveolar holes in the regions between emphysema correlated better with pulmonary function tests than did those in regions distant from emphysema. The maximum diameter of holes and the number of holes per alveolus correlated with functional residual capacity, residual volume, closing capacity expressed as a proportion of total lung capacity (CC/TLC), and static recoil pressure of the lung at TLC. Emphysema correlated with CC/TLC and with the transpulmonary pressure at 90% TLC. Bronchiolar lesions were not related to pulmonary function tests. Our data provide support for the hypothesis that the tissue surrounding emphysematous lesions contributes to loss of recoil.