PC Secretion Research Articles

TANGO1 and SEC12 are copackaged with procollagen I to facilitate the generation of large COPII carriers

Large coat protein complex II (COPII)-coated vesicles serve to convey the large cargo procollagen I (PC1) from the endoplasmic reticulum (ER). The link between large cargo in the lumen of the ER and modulation of the COPII machinery remains unresolved. TANGO1 is required for PC secretion and interacts with PC and COPII on opposite sides of the ER membrane, but evidence suggests that TANGO1 is retained in the ER, and not included in normal size (<100 nm) COPII vesicles. Here we show that TANGO1 is exported out of the ER in large COPII-coated PC1 carriers, and retrieved back to the ER by the retrograde coat, COPI, mediated by the C-terminal RDEL retrieval sequence of HSP47. TANGO1 is known to target the COPII initiation factor SEC12 to ER exit sites through an interacting protein, cTAGE5. SEC12 is important for the growth of COPII vesicles, but it is not sorted into small budded vesicles. We found both cTAGE5 and SEC12 were exported with TANGO1 in large COPII carriers. In contrast to its exclusion from small transport vesicles, SEC12 was particularly enriched around ER membranes and large COPII carriers that contained PC1. We constructed a split GFP system to recapitulate the targeting of SEC12 to PC1 via the luminal domain of TANGO1. The minimal targeting system enriched SEC12 around PC1 and generated large PC1 carriers. We conclude that TANGO1, cTAGE5, and SEC12 are copacked with PC1 into COPII carriers to increase the size of COPII, thus ensuring the capture of large cargo.

Potential Clinical Impact of Choline Supplementation in Cystic Fibrosis Patients

Rationale: In cystic fibrosis (CF) exocrine pancreas insufficiency results in impaired cleavage of bile phosphatidylcholine and faecal choline loss. Choline is essential for phosphatidylcholine synthesis, and in choline deficiency the liver accretes choline/phosphatidylcholine at the expense of the lungs. Hence, plasma choline and phosphatidylcholine concentrations correlate with lung function. Objective: We investigated the effect of choline supplementation on hepatic PC secretion (primary outcome), lung function, liver fat and choline metabolism. Methods: 10 adult male CF patients were recruited from 11/2014-1/2016 for oral supplementation with 3x1g choline chloride for 84(84-91)d. Pre/post supplementation, patients were spiked with 3.6mg/kg [methyl-D9]choline to assess choline/PC metabolism. Mass spectrometry, spirometry and hepatic nuclear resonance spectrometry served for analysis. Main Results: Supplementation decreased plasma D9-PC from 17.7(15.5- 22.4)µmol/L to 12.2(10.5-18.3)µmol/L (t=33h, p<0.01), indicating D9- tracer dilution by higher choline metabolite pools. Whereas plasma PC was unchanged, choline increased from 4.8(4.1-6.2)µmol/L to 14.7(11.8- 16.2)µmol/L at d49, and was 10.5(8.5-15.5)µmol/L at d84 (p<0.01). Supplementation increased forced expiratory volume of 1s percent of predicted (ppFEV1) from 70.0(50.9-74.8)% to 78.3(60.1-83.9)% (p<0.05), and decreased hepatosteatosis from 1.58(0.37-8.82)% to 0.84(0.56-1.17)% liver fat (p<0.01). Post-interventionally, plasma choline returned to baseline concentrations within 60h. Conclusions: Choline supplementation normalized plasma choline concentration, and increased choline-containing PC precursor pools in adult CF patients. Improved lung function, and resolution of hepatosteatosis, suggest that correcting choline deficiency is clinically important in a systemic approach. Choline supplementation may be a therapeutic perspective for CF patients that should be addressed in prospective, randomized and placebo-controlled trials. Clinical Trial Number: Clinical trial registered with clinicaltrials.gov (NCT 03312140). Funding Statement: Margerete-Muller-Bull-Stiftung, Stuttgart, Germany, for project granting. Declaration of Interests: The authors have no conflicts of interest to disclose. Ethics Approval Statement: The protocol was approved by The Institutional Review Board, and registered at ClinicalTrials.gov (Identifier: NCT 03312140). Data collection and management was performed by the Centre for Paediatric Clinical Studies (CPCS) III&IV, University Children’s Hospital, University of Tubingen.

Genetic Mouse Models with Intestinal-Specific Tight Junction Deletion Resemble an Ulcerative Colitis Phenotype.

Background and AimsA key pathogenetic feature of ulcerative colitis [UC] is an intrinsic low mucus phosphatidylcholine[PC] content. Recently, a paracellular transport for PC across tight junctions[TJs] was described, suggesting TJ disturbance as a cause of diminished luminal PC transport. Therefore, we aimed to generate mutant mice with TJ deletion to evaluate whether a UC phenotype developed.MethodsCL57BL/6 control wild-type mice were compared to mutant mice with tamoxifen-induced villin-Cre-dependent intestinal deletion of kindlin 1 and 2.ResultsElectron microscopy of mucosal biopsies obtained from both mutants before overt inflammation following only 2 days of tamoxifen exposure revealed a defective TJ morphology with extended paracellular space and, by light microscopy, expanded mucosal crypt lumina. PC secretion into mucus was reduced by >65% and the mucus PC content dropped by >50%, causing a >50 % decrease of mucus hydrophobicity in both mutants. Consequently, the microbiota was able to penetrate the submucosa. After 3 days of tamoxifen exposure, intestinal inflammation was present in both mutants, with loose bloody stools as well as macroscopic and histological features of colitis. Oral PC supplementation was able to suppress inflammation. By analogy, colonic biopsies obtained from patients with UC in remission also showed a defective epithelium with widened intercellular clefts, and enlarged crypt luminal diameters with functionally impaired luminal PC secretion.ConclusionsGenetic mouse models with intestinal deletion of kindlin 1 and 2 resulted in TJ deletion and revealed pathophysiological features of impaired PC secretion to the mucus leading to mucosal inflammation compatible with human UC.

Functional defect of variants in the adenosine triphosphate-binding sites of ABCB4 and their rescue by the cystic fibrosis transmembrane conductance regulator potentiator, ivacaftor (VX-770).

Disease-causing variations in the ATP-binding sites of ABCB4 cause defects in PC secretion, which can be rescued by ivacaftor. These results provide the first experimental evidence that ivacaftor is a potential therapy for selected patients who harbor mutations in the ATP-binding sites of ABCB4. (Hepatology 2017;65:560-570).

Syntaxin-4 is implicated in the secretion of antibodies by human plasma cells

PCs are responsible for the production and secretion of antibodies, the effector molecules of the humoral immune response. The molecular mechanisms responsible for vesicle docking and secretion implicated in the antibody-secretion process are not well-known, as they have not been studied, but it is known that SNARE proteins are responsible for many membrane-fusion processes in the cell. We show here that freshly isolated human colon LP-PCs and T-PCs from MM-PC patients and the U266 cell line, as a model for PC secretion, contain a set of these proteins. SNAP23, STX3, and STX4 were localized mainly in the plasma membrane of PCs, and interactions of SNAP23 with STX3 and with STX4 were proven by IP. Interaction between SNAP23 and STX4 was also confirmed in situ. With the use of siRNA, as well as shRNA, the functional role of SNAP23, STX3, and STX4 in antibody secretion was also examined. The findings demonstrate that in addition to SNAP23, STX4 is implicated in the antibody secretion by a myeloma cell line and by normal human colon LP-PCs.

Metformin reduces cellular lysophosphatidylcholine and thereby may lower apolipoprotein B secretion in primary human hepatocytes

The biguanide metformin is an oral antihyperglycemic drug for the treatment of type 2 diabetes mellitus. Further, a moderate improvement of dyslipidemia by metformin was reported, and therefore, the effect of metformin on the release of apolipoprotein B (ApoB) and ApoE in primary human hepatocytes was determined. Metformin at 0.5 and 1 mM reduced hepatic ApoB secretion but ApoE was not altered. Metformin is well known to stimulate the AMP kinase that subsequently reduces hepatic nuclear factor 4-α (HNF4-α) and HNF4-α regulated genes like ApoB. However, HNF4-α was only diminished by 1 mM metformin and ApoB mRNA was not suppressed indicating that this pathway may not explain reduced ApoB release. Lower abundance of lysophosphatidylcholine (lysoPC) may also diminish ApoB secretion. Therefore, electrospray ionization tandem mass spectrometry was applied to measure cellular lipids. PC, lysoPC (produced by hydrolysis of PC), phosphatidylserine and sphingomyelin (derived from PC) were lower in metformin-treated hepatocytes whereas phosphatidylethanolamine, an alternative precursor of PC, was not affected. In addition, ABCB4, the canalicular membrane flippase essential for biliary PC secretion, was diminished. Supplementation with lysoPC led to a selective elevation of endogenous lysoPC and rescued ApoB secretion in metformin-treated cells. Therefore, it is concluded that metformin reduces lysoPC in human hepatocytes and this may secondarily lead to a therapeutically beneficial lower release of ApoB.

Abnormal surfactant metabolism and function in preterm ventilated baboons.

We evaluated surfactant metabolism and function and the effects of antenatal glucocorticoids in very preterm baboons. Pregnant baboons were randomized to receive saline (controls) or 6 mg betamethasone (beta) 48 and 24 h before delivery at 125 +/- 2 d gestation (term is 184 d). The newborn baboons were treated with [14C]dipalmitoylphosphatidylcholine-labeled surfactant and ventilated for 6 d. Lung function for six control and six betamethasone-treated animals was similar. Recoveries of 14C-saturated phosphatidylcholine (Sat PC) were similar: 4.8% (control) and 3.6% (beta) in alveolar wash and 15.4% (control) and 17.7% (beta) in total lungs. Alveolar and total lung pool sizes of Sat PC were about 23 and 190 micromol/kg, respectively. The preterm baboons secreted 8.7% (control) and 6.5% (beta) of de novo synthesized Sat PC labeled with 3H-palmitate from Day 5 to Day 6. These preterm baboons had high estimated Sat PC synthetic and net tissue accumulation rates but low secretion of Sat PC. The large aggregate surfactant fractions from the preterm baboons had high minimal surface tensions and were less effective when used to treat surfactant-deficient preterm rabbits than surfactant from newborn or adult baboons. Ventilation of the preterm baboon was associated with surfactant functional and metabolic abnormalities that were not altered by antenatal glucocorticoids.

Osmotic stress induces both secretion and apoptosis in rat alveolar type II cells.

The aim of this study was to analyze the effects of osmotic shock and secretagogues such as ATP and 12-O-tetradecanoylphorbol 13-acetate (TPA) on various intracellular signaling pathways in primary cultures of alveolar type II cells and examine their potential role in regulating events such as secretion and apoptosis in these cells. Sorbitol-induced osmotic stress caused the sustained release of [3H]phosphatidylcholine ([3H]PC) from primary cultures of rat alveolar type II cells prelabeled with [3H]choline chloride. This release was not dependent on protein kinase C because downregulation of the major protein kinase C isoforms (alpha, betaII, delta, and eta) expressed in alveolar type II cells had no effect on [3H]PC secretion. Sorbitol, as well as the known secretagogues TPA and ATP, activated extracellular signal-regulated kinase. Although an inhibitor of the extracellular signal-regulated kinase cascade, PD-98059, blocked this activation, it had no effect on the release of [3H]PC. Sorbitol and ultraviolet C radiation, but not TPA or ATP, were also found to activate both p38 and stress-activated protein kinase/c-Jun NH2-terminal kinase. Furthermore, both sorbitol and ultraviolet C radiation induced apoptosis in alveolar type II cells as demonstrated by Hoechst 33258 staining of the condensed nuclei, the generation of DNA ladders, and the activation of caspases. The data indicate that multiple signaling pathways are activated by traditional secretagogues such as TPA and ATP and by cellular stresses such as osmotic shock and that these may be involved in regulating secretory and apoptotic events in alveolar type II cells.

Clearance of surfactant protein A from rabbit lungs.

Surfactant protein A (SP-A) is a major surfactant protein with multiple biophysical, metabolic, and host defense functions. To further characterize its metabolism in vivo, we measured clearance of SP-A from adult rabbit lungs. Trace amounts of [125I]SP-A radiolabeled by the Bolton-Hunter method and mixed with [3H]dipalmitoylphosphatidylcholine (DPPC) were given intratracheally via a bronchoscope to rabbits. Groups of five to six animals were studied 10 min to 16 h after labeled surfactant administration. After collection of alveolar washes, lamellar bodies were isolated from lung tissue. Macrophages also were isolated from alveolar washes. [125I]SP-A was cleared more rapidly than DPPC from the airspaces. Both [125I]SP-A and [3H]DPPC were lost exponentially from the total lungs, with half-life values of 6.5 h for SP-A and 12 h for DPPC (P < 0.01). In macrophages, the highest radioactivities for SP-A and DPPC were at 10 to 45 min and the radiolabels subsequently disappeared similarly. In lamellar bodies, 125I and 3H radioactivities per mumol saturated phosphatidylcholine (Sat PC) increased in parallel and were highest at 2 h. Whereas radiolabeled lipids were recovered in lamellar bodies for up to 16 h, iodinated SP-A was lost, indicating less recycling of SP-A than DPPC. We previously showed independent pathways of SP-A and Sat PC secretion in rabbits. These results demonstrate the different clearance kinetics of these two principle components of surfactant.

166 DEVELOPMENT CHANGES IN THE RESPONSE OF TYPE II CELLS TO PHOSPHATIDYLCHOLINE (PC) SECRETAGOGUES

PC secretion in adult rat type II cells can be stimulated by several agents acting via different signal-transduction mechanisms. To determine if these mechanisms also operate in developing lung we have compared the response of type II cells isolated from 21 day fetal and 1, 7 and 14 day old rats with those from adults to 36 uM terbutaline, 10 uM N-ethylcarboxamido-adenosine (NECA, adenosine analog), 1 mM ATP, 10 uM tetradecanoylphorbol acetate (TPA) and 25 nM ionomycin. Fetal cells were isolated by trypsin/collagenase digestion and differential adhesion and those from newborns and adults by elastase digestion and panning on IgG-coated dishes. The cells were cultured for 18-20 h with 3H-choline, washed in fresh medium and incubated ± agonists for 90 min after which 3H-PC in cells and medium was measured. The cells were 81-97% type II cells. The rate of basal secretion (PC in medium as % of total in cells + medium) was the same in all 5 groups.However, there was a developmental change in the response to the secretogogues. These data show that different signal-transduction pathways mature at different developmental stages in type II cells. Supported by NIH (HL-31175) and the DFG.

Alveolar Type II Cells Isolated after Silica-induced Lung Injury in Rats Have Increased Surfactant Protein A (SP-A) Receptor Activity

We examined surfactant secretion and its regulation by surfactant protein A (SP-A) in alveolar type II cells isolated from silica-treated rats to determine the role of SP-A-mediated regulatory control of phospholipid secretion in the pathogenesis of silica-induced alveolar proteinosis. Type II cells were isolated at weekly intervals for 28 d after silica or saline instillation. The maximum total binding of [125I]SP-A (internalized and surface-bound SP-A) to type II cells increased with time after silica instillation and, at 21 d, was 4-fold greater than that of type II cells isolated from saline-treated rats (272.8 +/- 42.5 and 65.4 +/- 9.8 ng/10(5) cells, respectively; P less than 0.05). Type II cells isolated from silica-treated rats showed a 2-fold increased surface binding and a 3-fold increased internalization compared to control cells. The receptor affinity for SP-A was the same for type II cells isolated from silica- and saline-treated animals. Type II cells isolated 14 d after silica instillation were separated into normotrophic and hypertrophic populations by centrifugal elutriation. Hypertrophic cells showed significantly elevated maximum total binding compared to normotrophic cells. The secretion of [3H]phosphatidylcholine [( 3H]PC) by type II cells from silica- and saline-treated animals was also compared. Type II cells from silica-treated animals showed lower basal and tetradecanoyl phorbol acetate (TPA)-stimulated [3H]PC secretion than cells from saline-treated animals at each time point after instillation. SP-A inhibited TPA-stimulated [3H]PC secretion similarly in type II cells isolated after either silica or saline instillation.(ABSTRACT TRUNCATED AT 250 WORDS)

Altered surfactant function and metabolism in rabbits with acute lung injury

Lung injury was induced in rabbits with N-nitroso-N-methylurethane (NNNMU), and saturated phosphatidylcholine (Sat PC) pool sizes and phospholipid compositions were measured in alveolar wash subfractions isolated by differential centrifugation (large and small surfactant aggregates). Surfactant metabolism also was studied using intravascular and intratracheal radiolabels. Protein permeability, gas exchange, and compliance were significantly abnormal as lung injury progressed. At peak injury, there was a decrease in the large aggregate Sat PC pool size in alveolar wash accompanied by increased uptake of Sat PC from the air space and increased specific activity of both intravascular and intratracheal radiolabels in lamellar bodies. This was followed by a marked rise in the small aggregate pool size in the alveolar wash and increased secretion of Sat PC into the air spaces. Phospholipid compositions, total phospholipid-to-protein ratios, and in vivo functional studies using a preterm ventilated rabbit model were abnormal for both large and small aggregate surfactant fractions from the lung-injured rabbits. These studies characterize quantitative, qualitative, and functional changes of alveolar wash surfactant subfractions in NNNMU-injured lungs.

Surfactant-associated protein inhibits phospholipid secretion from type II cells.

Secretion of [3H]phosphatidylcholine ([3H]PC) from isolated rat pulmonary type II epithelial cells was inhibited by the surfactant-associated protein of Mr = 35,000 (SAP-35) purified from canine lung surfactant. SAP-35 inhibited [3H]PC secretion in a dose-dependent manner and significantly inhibited basal, phorbol ester, beta-adrenergic, and P2-purinergic agonist-induced [3H]PC secretion. SAP-35 significantly inhibited [3H]PC secretion from 1 to 3 h after treatment. The IC50 for inhibition of [3H]PC secretion by canine SAP-35 was 1-5 X 10(-6) g/ml and was similar for inhibition of both basal and secretagogue-stimulated release. Heat denaturation of SAP-35, addition of monoclonal anti-SAP-35 antibody, reduction and alkylation of SAP-35, or association of SAP-35 with phospholipid vesicles reversed the inhibitory effect on secretagogue-induced secretion. Inhibitory effects of SAP-35 were observed 3 h after cells were washed with buffer that did not contain SAP-35. Although SAP-35 enhanced reassociation of surfactant phospholipid with isolated type II cells, its inhibitory effect on secretion of [3H]PC did not result from stimulation of reuptake of secreted [3H]PC by type II cells. The inhibition of phospholipid secretion by SAP-35 was also not due to inhibition of PC or disaturated PC synthesis by SAP-35. SAP-35, the major phospholipid-associated protein in pulmonary surfactant, is a potent inhibitor of surfactant secretion from type II cells in vitro and may play an important role in homeostasis of surfactant in the alveolar space.

PHOSPHOINOSITIDE CYCLE IN DIFFERENTIATION OF TYPE II ALVEOLAR CELLS

In the present study we have evaluated the role of phospholnositide cycle and protein kinase C in signal transmission because INO accelerates maturation of the surfactant system CPed Res 20:1228, 1986). Rabbit lung explants were grown in serum-free medium, and lung epithelial cells (80% of type II cells) cultured in the presence of serum from adult rabbit. - Lung explants from 22-day-old fetuses did not demonstrate surfactant synthesis in the presence of dexamethasone (1 uM) and thyroxlne (1 uM). Phorbol ester or dioctylglycerol increased the synthesis of surfactant ohosphatidylcholine (PC) by 470%, and increased secretion of PC from type II cells by 57%. In the presence of 1.5 mM INO, the hormones alone increased surfactant synthesis & secretion, whereas the activators of protein kinase C did not have a further effect. Addition of fibroblast-pneumonocyte factor into explants labeled with 1.5 mM 3H-INO, transiently decreased dl- and triphosphoinositide labeling. This was evident only when the medium contained high INO C1.5mM). - We propose that protein kinase C transmits signals into very immature alveolar cells and that high extracellular inositol is required for activation of the phosphoinositide cycle, and of protein kinase C.

PHOSPHATIDYLCHOLINE (PC) SECRETION BY CULTURED FETAL RAT TYPE II PNEUMOCYTES

Purinooeptor and B-adrenergio agonists have been shown to stimulate PC secretion by adult rat type II cells. We have studied the influence of these agents on secretion by fetal type II cells. The cells were isolated by explant culture, cell dissociation, differential adhesion, type II cell aggregation, and monolayer culture. To study secretion the cells were incubated with [3H]-choline for 20h, followed by 90 minutes of exposure to varying concentrations of ATP or terbutaline. Purity of type II cell cultures ranged from 85 to 90%. Post culture viability was 97.3±0.2%. LDH release averaged 1.5%; no difference was observed between control and treated cultures.Baseline secretion of PC from the fetal cells was 0.63±0.11%. We observed dose dependent enhancement of secretion by both ATP and terbutaline. Maximal stimulation by ATP occurred at 10−4 M. At this dose secretion was 1.24±0.28% (97% increase). Maximal stimulation by terbutaline was observed at 10−5 M (75% increase). These data differ from those in adult rat type II cells in which baseline PC secretion ranged from 1% to 3%. ATP and terbutaline produced greater stimulation in adult cells (approximately 400% and 100% respectively) than in fetal cells.We conclude that fetal type II cells secrete PC. Baseline and stimulated secretion is lower in these cells than in adult cells. This suggests that although fetal and adult cells are morphologically similar, they are functionally different. (Supported by HD 07091, HL 19752, HL 31175.)

P2-purinoceptors regulate surfactant secretion from rat isolated alveolar type II cells.

Rat isolated alveolar Type II cells were utilized to examine the effect of purine and pyrimidine analogues on secretion of pulmonary surfactant. ATP potently stimulated [3H]-phosphatidylcholine ([3H]-PC) secretion in a time- and dose-dependent manner. The effect of ATP was noted by one hour of exposure and persisted for three hours. The EC50 (concentration producing 1/2 the maximal response) for ATP-induced [3H]-PC secretion was 100 nM. ADP was also a potent secretagogue for surfactant secretion, but AMP and adenosine had no significant effect on surfactant secretion at concentrations less than or equal to 250 microM. The EC50 for ADP-induced [3H]-PC secretion was 250 nM. Other purine and pyrimidine nucleotides (ITP, GTP, CTP, TTP) were examined for their effect on [3H]-PC secretion. All purine and pyrimidine triphosphates examined significantly augmented [3H]-PC secretion, but were much less potent than ATP. The EC50s were ITP = 10 microM; GTP = 100 microM; CTP = 250 microM; TTP = 100 microM. Neither 8-phenyltheophylline (10 microM, a P1-purinoceptor antagonist), propranolol (100 microM, a beta-adrenoceptor antagonist), nor indomethacin (10 microM, a prostaglandin synthetase inhibitor) inhibited ATP-induced [3H]-PC secretion from isolated Type II cells. These data provide evidence for regulation of surfactant secretion from alveolar Type II cells by a P2-purinoceptor.

264 STIMULATION OF PHOSPHATIDYLCHOLINE (PC) SECRETION BY LEUKOTRIENES C4, D4, AND E4, IN TYPE II CELLS ISOLATED FROM ADULT RAT LUNG

We previously reported that incubation of type II cells with 1-10 μM arachidonic acid (AA) stimulated PC secretion and that this effect was blocked by 0.1 μM nordihydroguaiaretic acid and 10 μM indomethacin (I) but not by 0.1 μM 1 or 10 μM flufenamic acid, ibuprofen or naproxen. Since these data suggested that lipoxygenase products of its metabolism mediated the effect of AA we examined effects of leukotrienes on PC secretion in type 11 cells. Cells isolated from adult rats were cultured for 20 h in medium containing serum, antibiotics and 3H-choline, transferred to fresh serum-free medium and incubated with leukotrienes for 90 min. 3H-PC in cells and medium was then measured. LTC4, LTD4 and LTE4 but not LTB4 stimulated PC secretion and this effect was concentration dependent in the range 10−12-10−6 M. In 7 experiments LTC4, LTD4, and LTE4 (1 μM) increased the rate of PC secretion by 63%, 81% and 100% from 0.7±0.04 (3H-PC in the medium as % of that in cells + medium) in the control cultures to 1.14±0.08 (P<0.001), 1.27±0.12 (P<0.002) and 1.40±0.12 (P<0.001), respectively. The concentration of LTE4, which produced half maximal stimulation (EC50) was 5×10−12 M. Leukotriene receptors have been reported in intact lung tissue. Whether these effects are mediated by specific leukotriene receptors in type II cells remains to be established. (Supported by HL-31175. We are grateful to Dr. J. Rokach, Merck Frosst Canada, who generously supplied the leukotrienes.)

1693 LABOR STIMULATES SURFACTANT SECRETION IN NEWBORN RABBIT LUNG SLICES

We previously reported that labor increased the amount of surfactant phospholipid in newborn rabbit lung lavage but it was unclear if the effect was on synthesis or secretion. We have now examined the effect of labor on secretion in a slice model. Newborn rabbits are injected i.p. with 3H-choline and killed 4 h later. Slices from the blood-free lungs are incubated in buffer for 2.5 h. Initially the lipids secreted are similar to those of lung tissue but after 3 changes of buffer the lipids secreted are very similar to those of lung lavage. We measure the rate of secretion of 3H-phosphatidylcholine (PC) during this period. At 31 days gestation the rate of secretion increased from 198±18 cpm/h/mg tissue protein in 16 litters delivered before labor to 281±21 in 31 litters delivered after oxytocin-induced labor (P<0.02). Spontaneous labor was also stimulatory while oxytocin without labor was not. We attempted to mimic and abolish the effect of labor on PC secretion by incubating the slices with various agents. 10−5M prostaglandin E2 and 10−4M terbutaline stimulated secretion in slices from animals delivered prior to labor by 20% and 33%, respectively. Pilocarpine had no effect. 2×10−4M indomethacin and 10−4M propranolol inhibited secretion in slices from animals delivered after labor by 49% and 42%, respectively. Atropine had no effect. These data show that labor stimulates surfactant secretion and thus may be an important factor in the prevention of RDS. The effect of labor may be mediated, at least in part, by prostaglandins and catecholamines both of which increase during labor and birth. (Supported by HD-11018).

Effect of hydrocortisone on the metabolism of phosphatidylcholine in maternal and fetal rabbit lungs and livers.

Administration of hydrocortisone to pregnant rabbits caused a decrease in weights of fetal body and lung and an increase in the incorporation of choline into fetal lung PC. The authors found no induction of the enzymes related to the incorporation of choline into PC in fetal lung. Also, there was no stimulation of any enzymatic activity of CDP-choline pathway or PC-lysoPC cycle pathway in maternal lung and liver or fetal liver. In addition to the acceleration of choline incorporation into fetal lung PC by the cortisol, hydrocortisone also significantly stimulated the secretion of lung PC affected by glucocorticoids may also be related to apparent fetal lung maturation.

Secretion of hepatic triglycerides into plasma of rats fed retinol.

The effect of feeding 33 mg of retinol daily for two days on liver and plasma lipids of rats has been studied. The secretion of liver TG into plasma of retinol fed rats has been measured by the use of palmitic acid-1-14C and of Triton WR 1339. Liver and plasma lipids, TG, phospholipids and PC were significantly higher in retinol fed rats as compared to control rats. The incorporation of palmitic acid-l-14C into liver TG, PC and PE was significantly higher in retinol fed rats. The labelling pattern with time of liver and plasma TG and PC shows that the secretion of liver TG and PC into plasma was impaired in retinol fed rats.