Clinicopathological features of narrow-band imaging endoscopy and immunohistochemistry in ultraminute esophageal squamous neoplasms.

To reveal clinicopathological features of narrow-band imaging (NBI) endoscopy and immunohistochemistry in ultraminute esophageal squamous neoplasms. If a lesion diameter was smaller or same compared with a width of closed biopsy forceps, a lesion was defined to be an ultraminute lesion. Twenty-five consecutive patients with 33 ultraminute esophageal lesions that were removed by endoscopic mucosal resection were included in the present study. We conducted two questionnaire surveys of six endoscopists by their retrospective review of endoscopic still images. The six endoscopists evaluated the endoscopic findings of the ultraminute lesions on still images taken by conventional white-light imaging endoscopy and non-magnified NBI endoscopy in the first questionnaire, and taken by magnified NBI endoscopy in the second questionnaire. An experienced pathologist who was unaware of any endoscopic findings made histological diagnosis and evaluated immunoexpression of p53 and Ki67. The 33 ultraminute lesions were all determined to be either 11 high-grade intraepithelial neoplasias (HGIENs) or 22 low-grade intraepithelial neoplasias (LGIENs). The tumor diameters were histologically confirmed to be <3 mm. All of the ultraminute tumors were visualized as unstained areas and brownish areas by real-time endoscopy with Lugol dye staining and non-magnified NBI endoscopy, respectively. All of the ultraminute IENs were visualized as brownish areas by real-time non-magnified NBI endoscopy. Three of the 25 patients with the ultraminute IENs (12%) had multiple brownish areas (more than several areas) in the esophagus on real-time non-magnified NBI endoscopy. All of the ultraminute IENs were visualized as unstained areas by real-time Lugol chromoendoscopy. Twenty of the 25 patients (80%) had multiple unstained areas (more than several areas) in the esophagus on real-time Lugol chromoendoscopy. The first questionnaire survey revealed that a significantly higher detection rate of the ultraminute IENs on non-magnified NBI endoscopy images compared with conventional white-light imaging endoscopy ones (100% vs. 72%, respectively: P < 0.0001). The second questionnaire survey revealed that presence rates of any magnified NBI endoscopy findings were not significantly different between HGIENs and LGIENs. Proliferation, dilation, and various shapes of intrapapillary capillary loops indicated remarkably high presence rates of more than 90% in both HGIENs and LGIENs. Six of 22 LGIENs (27%) and 3 of 11 HGIENs (27%) show a positive expression for p53. None of peri-IEN epithelia was positive for p53. A mean of Ki67 labeling index of LGIENs was 33% and that of HGIENs 36%. Ki67 labeling index was significantly greater in the LGIENs and HGIENs compared with that in the peri-IEN epithelia. There were no significant differences in p53 expression and Ki67 labeling index between the HGIENs and LGIENs. Non-magnified/magnified NBI endoscopy could facilitate visualization and characterization of ultraminute esophageal squamous IENs. The ultraminute HGIENs and LGIENs might have comparable features of magnified NBI endoscopy and immunohistochemistry.

Olanzapine increases hepatic glucose production through the activation of hypothalamic adenosine 5'-monophosphate-activated protein kinase.

AIMS: To investigate the mechanism of the metabolic disturbance induced by the atypical antipsychotic olanzapine, we examined whether adenosine 5'-monophosphate-activated protein kinase (AMPK) in the hypothalamus and hepatic glucose production are involved in the effect of olanzapine. METHODS: Male 6-week-old ICR mice were used. Blood glucose levels were determined by the glucose oxidase method. The mRNA levels of gluconeogenic or glycolytic enzymes were measured by reverse transcription polymerase chain reaction (RT-PCR). AMPK expression was measured by Western blotting. RESULTS: Systemic injection of olanzapine increased blood glucose levels in both unfasted and fasted mice. However, the increase in fasted mice was less than that in unfasted mice. Central administration of olanzapine also increased the blood glucose levels in unfasted mice, but not in fasted mice. In a pyruvate tolerance test, olanzapine significantly increased blood glucose levels. In addition, olanzapine increased the mRNA levels of glucose-6-phosphatase (G6Pase), a gluconeogenic enzyme, in the liver. Furthermore, olanzapine increased phosphorylated AMPK in the hypothalamus of unfasted mice, and olanzapine-induced hyperglycaemia was inhibited by the AMPK inhibitor compound C. Central administration of the AMPK activator AICAR significantly increased G6Pase mRNA levels in the liver and blood glucose levels. Moreover, both olanzapine- and AICAR-induced hyperglycaemia were attenuated by the ß-adrenergic receptor antagonist propranolol, suggesting that olanzapine and AICAR induce hepatic glucose production through the sympathetic nervous system. CONCLUSIONS: Our results indicate that olanzapine activates AMPK in the hypothalamus, which increases hepatic glucose production via the sympathetic nervous system.

Molecular analysis of new isolates of Tomato leaf curl Philippines virus and an associated betasatellite occurring in the Philippines.

Three new begomovirus isolates and one betasatellite were obtained from a tomato plant exhibiting leaf curl symptom in Laguna, the Philippines. Typical begomovirus DNA components representing the three isolates (PH01, PH02 and PH03) were cloned, and their full-length sequences were determined to be 2754 to 2746 nucleotides. The genome organizations of these isolates were similar to those of other Old World monopartite begomoviruses. The sequence data indicated that PH01 and PH02 were variants of strain B of the species Tomato leaf curl Philippines virus, while PH03 was a variant of strain A of the species Tomato leaf curl Philippines virus. These isolates were designated ToLCPV-B[PH:Lag1:06], ToLCPV-B[PH:Lag2:06], and ToLCPV-A[PH:Lag3:06], respectively. Phylogenetic analysis revealed that the present isolates form a separate monophyletic cluster with indigenous begomoviruses reported earlier in the Philippines. A betasatellite isolated from same sample belongs to the betasatellite species Tomato leaf curl Philippines betasatellite and designated Tomato leaf curl Philippines betasatellite-[Philippines:Laguna1:2006], ToLCPHB-[PH:Lag1:06]. When co-inoculated with this betasatellite, tomato leaf curl Philippines virus induced severe symptoms in N. benthamiana and Solanum lycopersicum plants. Using a PVX-mediated transient assay, we found that the C4 and C2 proteins of tomato leaf curl Philippines virus and the ßC1 protein of ToLCPHB-[PH:Lag1:06] function as a suppressor of RNA silencing.

First Report of Tomato leaf curl New Delhi virus Infecting Cucumber in Central Java, Indonesia.

Cucumber (Cucumis sativus L.) is an important vegetable in Indonesia. Cucumber plants showing yellowy green mosaic symptoms on leaves were observed in Klaten, Central Java, Indonesia in August 2008. Total DNAs were extracted from symptomatic leaves, and the putative viral genomes were amplified by PCR with the Deng A and B primers (2). The PCR-amplified viral genomic DNA was sequenced. The remaining part of DNA-A was amplified with two primers sets (ToLCNDV-A1F 5'-ACCAACAGGCCGATGAACA-3' and ToLCNDV-A1R 5'-TTCCCACTATCTTCCTGTGCA-3'; ToLCNDV-A2F 5'-TCGAGTGTGATRAAGAYTGCA-3' and ToLCNDV-A2R 5'-ACTAACTAAGCATTGCAGCGTC-3' [R = A and G, Y = C and T]) and sequenced. The remaining part of DNA-B was amplified with two primers sets (ToLCNDV-B1F 5'-ARGAGTTYMCRYYTGTGGA-3' and ToLCNDV-B1R 5'-TKCWGTYGGTCATGTCGT-3'; ToLCNDV-B2F 5'-TCYGTCAATCKCATGTCGYGT-3' and ToLCNDV-B2R 5'-CCTTACGCGTATAYTGTYTRGA-3' [K = G and T, M = A and C, W = A and T]) and sequenced. Full-length DNA-A (2,739 nt; GenBank Accession No. AB613825) and DNA-B (2,690 nt; GenBank Accession No. AB613826) sequences of a bipartite Tomato leaf curl New Delhi virus (ToLCNDV) from Central Java were obtained and they were most similar to the corresponding sequences of both DNA-A and DNA-B of ToLCNDV-[cucumber:Thailand] (DNA-A, GenBank Accession No. AB330079; DNA-B, GenBank Accession No. AB330080) at 95.5 and 91.0% nucleotide identities, respectively. On the basis of high nucleotide sequence identity with ToLCNDV-[cucumber:Thailand] and the demarcation criteria in species identification (3), the virus isolate from the diseased cucumber in Central Java is considered as a variant of ToLCNDV and was accordingly named ToLCNDV-Indonesia[Indonesia:Java:Cucumber:2008] (ToLCNDV-ID[ID:Jav:Cuc:08]). Although the importance of begomovirus diseases on chili pepper (Solanaceae) is currently highly noticed in Indonesia (1), ToLCNDV was newly isolated from cucumber (Cucurbitaceae) in this study. Therefore, farmers in Indonesia should pay more attention to controlling begomovirus vectors, white flies, on Cucurbitaceae. To our knowledge, this is the first report of the natural occurrence of ToLCNDV in Indonesia. References: (1) P. J. D. Barro et al. Biol. Invas.10:411, 2008. (2) D. Deng et al. Ann. Appl. Biol. 125:327, 1994. (3) C. M. Fauquet et al. Arch. Virol. 153:783, 2008.

Magnifying endoscopy with narrow band imaging for predicting the invasion depth of superficial esophageal squamous cell carcinoma.

The invasion depth of superficial esophageal squamous cell carcinoma is important in determining therapeutic strategy. The aim of this study was to prospectively investigate the clinical utility of magnifying endoscopy with narrow band imaging compared with that of non-magnifying high-resolution endoscopy or high-frequency endoscopic ultrasonography in predicting the depth of superficial esophageal squamous cell carcinoma. The techniques were carried out in 72 patients with 101 superficial esophageal squamous cell carcinomas, which were then resected by either endoscopic mucosal resection or esophagectomy. The histological invasion depth was divided into two: mucosal or submucosal carcinoma. We investigated the relationship between endoscopic staging and histology of tumor depth. Non-magnifying high-resolution endoscopy, magnifying endoscopy with narrow band imaging, and high-frequency endoscopic ultrasonography had overestimation/underestimation rates of 7/5, 4/4 and 8/3%, respectively. The sensitivity rates for the three techniques were 72, 78, and 83%, respectively, and the specificity rates were 92, 95, and 89%, respectively. There were no statistically significant differences among the three endoscopic techniques. Clinical utility of magnifying endoscopy with narrow band imaging does not seem to be significantly different from that of non-magnifying high-resolution endoscopy or high-frequency endoscopic ultrasonography in predicting the depth of superficial esophageal squamous cell carcinoma. Magnifying endoscopy with narrow band imaging may have potential to reduce overestimation risks of non-magnifying high-resolution endoscopy or high-frequency endoscopic ultrasonography.

EFFECT ON MICROBIAL PRODUCTS ON CAESIUM ELUTION BEHAVIOUR FROM CLAY MINERALS.

Some microorganisms in the environment make siderophores, which are low molecular chelators, to take up minerals from soil. Eleven bacteria were separated from the root of white clover by chlome azrol S (CAS) assay. Each bacterium was incubated in casamino acid (CAA) culture, and siderophores in CAA culture were purified. These extractions were applied to biotite or vermiculite spiked with Cs. From each clay mineral, 57.1-72.8% (5100 ppm), 55.6-63.8% (920 ppm) and 48.6-54.3% (2300 ppm), 31.6-34.4% (520 ppm) was eluted, respectively. To understand elution behaviour, Cs desorption ratio of each clay was measured every 30 min. The results indicate Cs elution was occurred quickly.

The begomoviruses Honeysuckle yellow vein mosaic virus and Tobacco leaf curl Japan virus with DNAbeta satellites cause yellow dwarf disease of tomato.

The complete nucleotide sequences of two begomoviruses (Nara virus-1 and Nara virus-2), a satellite DNA (DNAbeta-Nara) and defective DNAs were obtained from honeysuckle (Lonicera japonica) showing characteristic yellow vein mosaic symptoms in Nara Prefecture, Japan. One begomovirus (Ibaraki virus) and a satellite DNA (DNAbeta-Ibaraki) was isolated and cloned from honeysuckle plants exhibited typical yellowing of veins and small elliptical shaped enations along veins on the under side of the leaves in Ibaraki Prefecture, Japan. The genome organization of the three viruses is the same as those of other Old World monopartite begomoviruses. Nara virus-1 had overall nucleotide sequence identity with Nara virus-2 of 94% and Ibaraki virus of 90%. DNAbeta-Nara had overall nucleotide sequence identity with DNAbeta-Ibaraki of 83%. Comparison of the nucleotide sequences with other begomoviruses revealed that Nara virus-1 and Nara virus-2 are strains of Honeysuckle yellow vein mosaic virus (HYVMV), hence named as HYVMV-Nara1 and HYVMV-Nara2, whereas Ibaraki virus was a strain of Tobacco leaf curl Japan virus (TbLCJV), designated as TbLCJV-Hs[Iba]. HYVMV-Nara1 and HYVMV-Nara2 have hybrid genomes, which are likely to have formed recombination between HYVMV and TbLCJV. TbLCJV-Hs[Iba] or HYVMV-Nara2 could infect and cause yellowing, leaf crinkling and stunting symptoms when partial tandem dimeric constructs were agroinoculated on tomato plants. However, in the presence of DNAbeta, both TbLCJV-Hs[Iba] or HYVMV-Nara2 produced more severe stunting symptoms in tomato plants. Therefore, these viruses along with their satellites are causal agents of tomato yellow dwarf disease in Japan, and honeysuckle acts as a potential reservoir host. Previously available evidence indicated that DNAbeta elements do not contain iteron sequences of their helper viruses; hence this is the first evidence that DNAbeta satellites have the iteron of their helper virus.

Surfactant and pulmonary blood flow distributions following treatment of premature lambs with natural surfactant.

Prematurely delivered lambs were treated with radiolabeled natural surfactant by either tracheal instillation at birth and before the onset of mechanical ventilation, or after 23 +/- 1 (+/- SE) min of mechanical ventilation. Right ventricular blood flow distributions, left ventricular outputs, and left-to-right ductal shunts were measured with radiolabeled microspheres. After sacrifice, the lungs of lambs receiving surfactant at birth inflated uniformly with constant distending pressure while the lungs of lambs treated after a period of ventilation had aerated, partially aerated, and atelectatic areas. All lungs were divided into pieces which were weighed and catalogued as to location. The amount of radiolabeled surfactant and microsphere-associated radioactivity in each piece of lung was quantified. Surfactant was relatively homogenously distributed to pieces of lung from lambs that were treated with surfactant at birth; 48% of lung pieces received amounts of surfactant within +/- 25% of the mean value. Surfactant was preferentially recovered from the aerated pieces of lungs of lambs treated after a period of mechanical ventilation, and the distribution of surfactant to these lungs was very nonhomogeneous. Right ventricular blood flow distributions to the lungs were quite homogeneous in both groups of lambs. However, in 8 of 12 lambs, pulmonary blood flow was preferentially directed away from those pieces of lung that received relatively large amounts of surfactant and toward pieces of lung that received less surfactant. This acute redirection of pulmonary blood flow distribution may result from the local changes in compliances within the lung following surfactant instillation.

Sequential treatments of premature lambs with an artificial surfactant and natural surfactant.

To test an artificial surfactant in vivo, six 120-d gestational age lambs were treated at birth with a mixture of a 9:1 M ratio of [14C]dipalmitoyl phosphatidylcholine (DPC) and phosphatidylglycerol at a dose of 100 mg DPC/kg. Nine other lambs were not treated. The mean PO2 values of the lambs treated with artificial surfactant were 65.7 +/- 11 mm Hg vs. 24.8 +/- 1.6 mm Hg for the untreated lambs (P less than 0.001). All lambs then were treated with 50 mg/natural surfactant lipid per kg, which promptly improved PO2 in all lambs. The PO2 values of those lambs previously treated with artificial surfactant remained greater than 100 mm Hg for 2.5 +/- 0.5 h vs. 0.9 +/- 0.3 h for lambs untreated with artificial surfactant (P less than 0.01). The pH and PCO2 values were not strikingly different between the two groups of lambs. Airway samples taken from lambs treated with artificial surfactant before treatment with natural surfactant had minimal surface tensions of 32 +/- 2.9 dyn/cm, whereas the artificial surfactant reisolated from these samples by centrifugation had minimum surface tension of 0 dyn/cm. The minimum surface tension of artificial surfactant was inhibited by fetal lung fluid from the premature lambs, whereas the minimum surface tension of natural surfactant was much less sensitive to inhibition. Artificial surfactant did not improve the pressure-volume characteristics of unventilated premature lung, whereas natural surfactant did. The change in specific activity of [14C]DPC following treatment with natural surfactant indicated that approximately 50% of the DPC initially administered was no longer associated with the airways.

Corticosteroids and surfactant change lung function and protein leaks in the lungs of ventilated premature rabbits.

Fetal rabbits were treated with corticosteroids by maternal administration for 48 h before delivery at 27 d gestational age. The treated and control rabbits were placed on ventilator-plethysmographs so that ventilation could be adjusted by regulation of tidal volumes to 10-13 ml/kg body wt. [125I]albumin was mixed with fetal lung fluid at birth, alternate rabbits from each litter were treated with Surfactant-TA, and [131I]albumin was injected intravascularly. The movement of the labeled albumins into and out of the alveolar wash and lung tissue was measured after 30 min of ventilation. Corticosteroid treatment (total dose, 0.2 mg/kg betamethasone) significantly decreased the protein leak across the endothelium (P less than 0.001) but increased the protein leak across the epithelium (P less than 0.001). Surfactant treatment decreased both the endothelial and epithelial leaks, and the combination of surfactant and corticosteroid treatments decreased endothelial leaks to 29% of control values and increased compliance more than either treatment alone. The 48-h corticosteroid treatment did not increase alveolar surfactant pool sizes. Corticosteroids significantly changed lung protein leaks independently of surfactant, and improved the response of the preterm lung to surfactant treatments.

Effect of fetal adrenalectomy on catecholamine release and physiologic adaptation at birth in sheep.

Plasma catecholamine levels increase dramatically at birth. To determine the contribution of adrenal catecholamine secretion to the surge in catecholamines at birth and the role in newborn adaptation, we performed surgical adrenalectomy or sham operation on near-term ovine fetuses. After recovery in utero, the animals were delivered and supported by mechanical ventilation. Plasma catecholamine levels, heart rate, blood pressure, cardiac output, pulmonary function, surfactant secretion, and release of free fatty acids (FFA) and glucose were compared in control and adrenalectomized animals. Plasma epinephrine increased rapidly at birth in controls but was undetectable in adrenalectomized animals. Norepinephrine levels were not statistically different. Heart rate, blood pressure, cardiac output and contractility increased abruptly after cord cutting in controls but did not increase in adrenalectomized animals. Lung compliance, pulmonary function, surfactant pool size, glucose and FFA levels were significantly decreased in adrenalectomized animals. These results suggest that adrenal epinephrine secretion is vital to many of the adaptive events at birth.

Duration and characteristics of treatment of premature lambs with natural surfactant.

Premature lambs were treated with 50 mg/kg of natural surfactant lipid by tracheal instillation either at birth or shortly thereafter when respiratory failure was documented. All lambs were delivered by cesarean section and supported on infant ventilators with 100% oxygen under conditions to mimic the care of human infants with the respiratory distress syndrome. The natural surfactant used for therapy was recovered by lavage from sheep lung. Six 120-d gestational age lambs treated at birth had an initial mean oxygen pressure (pO2) value of 270 +/- 35 mm Hg; this fell within 3 h to less than 100 mm Hg. By 8.3 +/- 0.3 h after birth the lambs were in severe respiratory failure with a mean pH less than 7.1 and a mean pCO2 greater than 70 mm Hg. Six untreated lambs had pH values below 7.0 within 40 min of life despite more intensive respiratory support than was given the treated animals. Treatment with natural surfactant of 17 lambs of 120 and 130 d gestational age after early respiratory failure resulted in a prompt increase in pO2 values from about 35 mm Hg to values over 200 mm Hg and a fall in pCO2 values to normal levels in the majority of animals. This response lasted only approximately 3 h, and a second treatment was less predictably effective.

First Report of Pepper yellow leaf curl Indonesia virus in Ageratum conyzoides in Indonesia.

Ageratum conyzoides L. plants affected with yellow vein disease were collected from Magelang, Bandung, and Purwokerto locations in Indonesia during 2001. A. conyzoides is a naturally occurring weed that is found in and around fields of cultivated pepper (Capsicum annuum L.) and tomato (Lycopersicon esculentum L.). It is frequently found with symptoms of yellow vein disease and the abundance of whiteflies on the affected plants suggested the possible involvement of a geminivirus. Total nucleic acids were extracted from nine samples collected from these locations of A. conyzoides-affected plants exhibiting yellow vein disease and amplified using PCR with geminivirus DNA-A-specific designed primers (virion-sense primer 5'-GAGCTCTTAGCCGCCTGAATGTTC-3'; complementary-sense primer 5'-GAGCTCGTCAGATGTTAAGACCTAC-3') (1). A PCR-amplified product of approximately 2.7 kbp was obtained from each sample. Five independent sequences were cloned and sequenced from each sample. Sequence analysis showed that five of nine samples were Ageratum yellow vein virus (one each from Bandung and Purwokerto and three from Magelang) and the remaining four samples (two samples each from Bandung and Purwokerto) were a strain of Pepper yellow leaf curl Indonesia virus (PepYLCIDV). Full-length DNA-A of PepYLCIDV from systemic A. coniziodes was amplified using PCR with additional primers designed at only one restriction site (BamHI) (5'-GGATCCGCTTGTTCATCCTTTTCCAG-3'/5'-GGATCCCACATCTTTGGTTAGTGGAGGGTG-3') and cloned. Three independent clones obtained were sequenced and analyzed. The sequence of a full-length DNA-A component was determined (2,760 bases, GenBank Accession No. AB267838). PCR using degenerate primers (DNABLC1: 5'-GTVAATGGRGTDCACTTCTG-3'; DNABLC2: 5'-RGTDCACTTCTGYARGATGC-3', DNABLV2: 5'-GAGTAGTAGTGBAKGTTGCA-3') of begomovirus DNA-B component (2), five independent clones were obtained and sequenced. Primers designed to amplify a full-length B component were constructed around a unique restriction site (BamHI) (5'-GGATCCCCTCATTCCTTTTGCGGAG-3'/5'-GGATCCACAGAGGAAAACTCGCAAGGC-3'). A PCR product was obtained from A. conyzoides samples and three independent clones were sequenced and analyzed. A full-length sequence of a begomovirus B component was determined (2,746 bases, GenBank Accession No. AB267839). Five open reading frames (ORF) were found in DNA-A and two in DNA-B. The DNA-A and DNA-B had a common region (CR) (74% nucleotide sequence identity) that comprised approximately 160 nucleotides. The DNA-A and DNA-B had an identical 31-base stem loop region in the CR. In addition, DNA-A and DNA-B had the highest nucleotide sequence identity (93%) with those of PepYLCIDV (GenBank Accession Nos. AB267834 and AB267835), suggesting it is a strain of PepYLCIDV, which is widely prevalent in Indonesia. To our knowledge, this is the first report of PepYLCIDV isolated from A. conyzoides plants affected with yellow vein disease. References: (1) R. W. Briddon and P. G. Markham. Mol. Biotechnol. 1:202, 1994. (2) S. K. Green et al. Plant Dis. 85:1286, 2001.

Three-Dimensional Study of the Terminal Portion in Sprague-Dawley Rat Ejaculatory Ducts.

In mammals, a pair of ejaculatory ducts exists in the urethra at the seminal colliculus. The detailed anatomical structures of the distal end of the ejaculatory ducts of Sprague-Dawley rats were investigated by the computer-assisted three-dimensional reconstruction analysis using light-microscopic serial sections. A three-dimensional reconstruction revealed that in adult rats, the ejaculatory sinus pair consists of two parts: the cranial section - a compartment region composed of a fusion of the ampullary gland duct and the seminal vesicle duct, and the caudal section - a grooved region composed of a long slitlike ejaculatory ostium that extends into the urethra on both sides of the seminal colliculus. But the sphincter structure was not observed. The long axis of the compartment region was approximately 58 µm in length, and that of the groove region was approximately 495 µm. Although many epithelial glands ducts were distributed throughout the ejaculatory sinuses, the prostate and coagulation gland ducts did not open in these sinuses. The urethra was composed of transitional epithelium, while the ejaculatory sinuses were composed of single to stratified cuboidal epithelium. The ejaculatory ducts continued to the ejaculatory ostium in male adult Sprague-Dawley rat were composed of the seminal vesicle ducts received the ampullary gland ducts.

Surfactant protein metabolism in vivo.

The surfactant components saturated phosphatidylcholine, SP-B and SP-C, are secreted together in lamellar bodies, and at least a part of the de novo synthesized SP-A is secreted independently. The surface film forms from tubular myelin and loose lipid arrays, and it generates unilamellar vesicles that lack surfactant proteins and are thought to represent catabolic forms. The half-life values for the clearance of surfactant proteins from lungs range from 6.5 to 28 h and vary with species. There is minimal information about the associations of the surfactant proteins with lipids or with each other after film formation, although all surfactant components seem to be recycled back into lamellar bodies in type II cells. The relative importance of type II cells or macrophages to the catabolism of the protein components of surfactant remains to be characterized, as do regulators of surfactant homeostasis.

Changes in the amount of lung and airway phosphatidylcholine in 0.5-12.5-day-old rabbits.

Newborn rabbits delivered spontaneously at term and cared for by the mothers were studied from 0.5 to 12.5 days of age. Curves are constructed to describe the changes in weight, lung and alveolar wash phosphatidylcholine and saturated phosphatidylcholine, and lung protein. The curves are complex and non-linear. However. expressing the increases in lung and alveolar wash phosphatidylcholine and saturated phosphatidylcholine pool sizes relative to animals weight results in a decreasing linear relationship from 0.5 to 12.5 days of age. By 12.5 days the ratios of lung phosphatidylcholine and saturated phosphatidylcholine to weight approximate the ratios measured for adult rabbits. The ratios of saturated to total phosphatidylcholine in the alveolar washes and lungs remained invarient throughout the study period.

Liposomes of dipalmitoylphosphatidylcholine associate with natural surfactant.

Unilamellar liposomes of an average diameter of 0.05 micron formed by sonication of dipalmitoylphosphatidylcholine associate in vitro with the large aggregate forms of natural surfactant. The liposomal-surfactant aggregates are stable and previously associated liposomes are not released from the aggregates by the addition of more liposomes. Radiolabeled liposomes, surfactant, and preformed liposomal-surfactant aggregates were injected at a dose of 8-10 mg lipid (about 2-times the endogenous surfactant pool size) into the airways of 3-day-old rabbits. Following airway injection, labeled phosphatidylcholine from the liposomal-surfactant aggregates were recovered in approximately equal amounts by alveolar wash and in the residual lung tissue fractions. This recovery pattern and the clearance kinetics were equivalent for 48 h after airway injection to those measured with radiolabeled surfactant alone. In contrast, following the injection of liposomes alone, labeled phosphatidylcholine from the liposomes was recovered primarily by alveolar wash at 3 and 24 h. The overall clearance of the liposomal-derived phosphatidylcholine from the lung was more rapid than was the clearance of the phosphatidylcholine from the surfactant or liposome-surfactant complexes. Liposomes can interact with surfactant in vitro, and the liposomes associated with the surfactant aggregate have a metabolic fate in vivo similar to surfactant and different from liposomes alone.

Surfactant protein B metabolism in newborn rabbits.

Surfactant protein B (SP-B) is critical to the biophysical function of surfactant. To characterize its metabolism in vivo in the newborn, we administered [35S]methionine and [3H]palmitate to newborn rabbits intravascularly. Three groups of 4 rabbits per group were killed at each of 4 time points followed by isolation of SP-B from alveolar wash and lamellar bodies. The labeling kinetics for alveolar wash associated SP-B and saturated phosphatidylcholine (Sat PC) had similar patterns. To characterize SP-B clearance from the airspace, rabbit SP-B was iodinated, mixed with [14C]dipalmitoylphosphatidylcholine and given by intratracheal injection. Alveolar washes and lamellar bodies were recovered from 4 animals at each of 7 time points. Both SP-B and Sat PC were cleared slowly from the total lung (half-life values approximately 25 h). However, SP-B was cleared more rapidly from the airspaces than was Sat PC. The ratio of [125I]SP-B to [14C]Sat PC in lamellar bodies increased 2-fold by 8 h. These results support the concept of linked secretion and clearance pathways for SP-B and Sat PC, although small differences in reuptake were detected.

The in vivo labeling with acetate and palmitate of lung phospholipids from developing and adult rabbits.

The labeling with radiolabeled acetate and palmitate of lung, microsomes isolated from lung, and surfactant phospholipids from adult, 3-day-old, and newborn rabbits was studied. The half-life of phosphatidylcholine from lung and microsomal fractions was shorter when labeled with acetate than when labeled with palmitate. Half-time values similarly measured for phosphatidylglycerol, phosphatidylinositol or phosphatidylethanolamine were not different for the two labels. Acetate and palmitate-labeled phospholipids appeared in the surfactant fraction with similar accumulation curves. The relative specific activities of acetate-labeled phosphatidylcholine from adult, 3-day-old, and newborn rabbits, respectively, were 1.30, 1.86 and 1.77 times those measured for those measured for the palmitate label. Surfactant phosphatidylinositol and phosphatidylethanolamine from 3-day-old animals similarly were labeled preferentially with acetate. However, phosphatidylglycerol purified from the surfactant fraction contained equivalent relative amounts of the acetate and palmitate labels in 3-day-old and adult rabbits. These results suggest that the type II pneumocyte may use acetate preferentially for the synthesis of palmitic acid which then is incorporated into surfactant phospholipids.