Breast arterial calcification on mammography and risk of coronary artery disease: a SCOT-HEART sub-study.

AIM: To assess the prevalence of breast arterial calcification (BAC) in patients who also underwent routine surveillance mammography, and to determine the association with cardiovascular risk factors, coronary artery calcification, and coronary artery disease on coronary computed tomography angiography (CCTA). MATERIALS AND METHODS: Four hundred and five female participants were identified who had undergone CCTA and subsequent mammography in the SCOT-HEART randomised controlled trial of CCTA in patients with suspected stable angina. Mammograms were assessed visually for the presence and severity of BAC. RESULTS: BAC was identified in 93 (23%) patients. Patients with BAC were slightly older (63±7 versus 59±8 years, p<0.001), with a higher cardiovascular risk score (19±11 versus 16±10, p=0.022) and were more likely to be non-smokers (73% versus 49%, p<0.001). In patients with BAC, coronary artery calcification was present in 58 patients (62%; relative risk [RR] 1.26, 95% confidence intervals [CI]: 1.04, 1.53; p=0.02), non-obstructive coronary artery disease in 58 (62%; RR 1.27, 95% CI: 1.04 to 1.54, p=0.02), and obstructive coronary artery disease in 19 (20%; RR 1.62, 95% CI: 0.98, 2.66; p=0.058). Patients without BAC were very unlikely to have severe coronary artery calcification (negative predictive value 95%) but the diagnostic accuracy of BAC to identify coronary artery disease was poor (AUC 0.547). CONCLUSION: Although BAC is associated with the presence and severity of coronary artery calcification, the diagnostic accuracy to identify patients with coronary artery disease or obstructive coronary artery disease is poor.

Coronary artery calcification is associated with mortality independent of pulmonary embolism severity: a retrospective cohort study.

AIM: To assess coronary artery calcification (CAC) and vascular calcification in patients with pulmonary embolism (PE) and correlate this with mortality. MATERIALS AND METHODS: PE severity was quantified using computed tomography pulmonary angiography (CTPA) in 400 consecutive cases using the modified Miller score (1-5, mild; 6-11, moderate; 12-16, severe). Right ventricle strain was assessed using the right/left ventricle diameter (RV/LV) ratio. CAC score (CACS) was assessed using a four-point scale (CACS mild 1-3, moderate 4-8, severe 9-12) for each vessel and summed to give the total CACS. Follow-up for mortality was obtained at 3 years. RESULTS: PE severity was classified as mild in 48%, moderate in 21%, and severe in 32% of cases. The median modified Miller score was 6 (Interquartile range [IQR] 2, 14) and median total CACS was 2 (IQR 0, 7). All-cause mortality occurred in 128 (32%) patients. Patients with CAC were three times more likely to die than patients without CAC (Hazard ratio [HR] 2.96; 95% CI 1.84, 4.77; p<0.001), and patients with severe CAC were at the highest risk (HR 4.62; 95% CI 2.73, 7.83, p<0.001). Gender, modified Miller score and RV/LV ratio were not predictive of mortality. In multivariate analysis both CACS and age were independent predictors of 3-year all-cause mortality. Of the patients with CAC who died, the presence of coronary artery disease was only documented in 34 (32%). CONCLUSION: CACS is an independent predictor of all-cause mortality in patients with PE, and has important implications for subsequent patient management.

Display colour scale effects on diagnostic performance and reader agreement in cardiac CT and prostate apparent diffusion coefficient assessment.

AIM: To investigate the effect of colour scale choice on diagnostic performance in the interpretation of medical images. MATERIALS AND METHODS: Twelve clinicians interpreted 210 myocardial computed tomography (CT) perfusion (CTP) examinations, and nine clinicians interpreted 165 magnetic resonance imaging (MRI) apparent diffusion coefficient (ADC) prostate images. In three separate sessions, each participant read the same image set using greyscale, hot-iron, and rainbow scales, respectively. Participants scored their level of confidence for tumour presence in the ADC study, and for ischaemia in the CTP study, from 0 to 100. The area under the receiver operating characteristic (ROC) curve (AUC) was used as the performance metric. For cases that scored >50, CTP readers' agreement on the ischaemic transmural extent was analysed, and ADC map readers' selected values and coordinates for the lowest ADC within the detected tumour were compared across different colour scales. RESULTS: For CTP detection, the AUC was up to 0.10 higher with greyscale, 0.67±0.02 (standard error), compared to rainbow, 0.56±0.02, and detection with hot-iron was in between (0.61±0.03). For ischaemic transmural lesion categorisation, observed inter-reader agreement was highest with greyscale for category 25-50%. There is a small tendency for rainbow and greyscale to outperform hot-iron in the detection of prostate tumours. The selected lowest ADC value and pixel localisation was similar with all colour scales. CONCLUSIONS: The present findings suggest that colour visualisation has a measurable effect on CTP and ADC performance. Further investigation is necessary to determine the magnitude of the effect in diagnostic tasks.

Genotyping and comparative pathology of Spirocerca in black-backed jackals (Canis mesomelas) in South Africa.

BACKGROUND: The pathology of spirocercosis, a disease caused by the infestation of carnivores with the nematode Spirocerca lupi, has been extensively described in domestic dogs and coyotes. However, it has not been described in wild carnivores in South Africa. The aim of this study was to evaluate whether black-backed jackals are a host for Spirocerca species and to provide a detailed description of the associated pathology. Jackals were also stratified according to age and the Spirocerca species recovered were characterized using molecular techniques. METHODS: Standard necropsies were performed on routinely culled jackals from three of the nine provinces of South Africa during the period June 2012 to February 2013. Jackals were screened for the presence of pathognomonic Spirocerca-induced lesions and for evidence of aberrant migration. Relevant samples were submitted for histopathology and collected larvae were genotyped at nine microsatellite loci. RESULTS: Spirocerca lupi-associated aortic lesions were found in 16 of 93 (17%) black-backed jackals. Of these, four (25%) were associated with S. lupi larvae. Genotyping of the larvae revealed amplification of all nine loci that amplified dog-derived S. lupi, with the same level of polymorphism in the allele size ranges. Only 1 of 93 jackals had an esophageal nodule with concurrent S. lupi-induced aortic aneurysms. The single esophageal nodule found did not contain adult nematodes, nor did it communicate with the esophageal lumen. None of the jackals that were examined had macroscopically evident spondylitis, which is frequently reported in the dog. Histopathology of the S. lupi-induced aortic lesions in the jackal revealed replacement of elastic and smooth muscle fibers by fibrous connective tissue. In cases where inflammation was present, the inflammatory infiltrate consisted predominantly of eosinophils. The single esophageal nodule histologically resembled the early inflammatory nodule described in dogs and consisted of fibrous connective tissue, multifocal accumulation of lymphocytes, plasma cells and rare hemosiderin-laden macrophages. CONCLUSIONS: These lesions suggest that the life cycle of S. lupi may not or only rarely be completed in jackals. A possible explanation might be that jackals are relatively resistant to developing significant pathology associated with S. lupi-infection. However, before any conclusions can be drawn, many more jackals, including those that die naturally will have to be investigated for evidence of S. lupi infection.

CT myocardial perfusion imaging: current status and future directions.

Computed tomography (CT) imaging of the heart has advanced rapidly, and it is now possible to perform a comprehensive assessment at a low radiation dose. CT myocardial perfusion imaging can provide additive information to CT coronary angiography, and is particularly useful in patients with heavily calcified coronary arteries or coronary artery stents. A number of protocols are now available for CT myocardial perfusion including static, dynamic, and dual-energy techniques. This review will discuss the current status of CT myocardial perfusion imaging, its clinical application, and future directions for this technology.

Cardiovascular PET-CT imaging: a new frontier?

Cardiovascular positron-emission tomography combined with computed tomography (PET-CT) has recently emerged as an imaging technology with the potential to simultaneously describe both anatomical structures and physiological processes in vivo. The scope for clinical application of this technique is vast, but to date this promise has not been realised. Nonetheless, significant research activity is underway to explore these possibilities and it is likely that the knowledge gained will have important diagnostic and therapeutic implications in due course. This review provides a brief overview of the current state of cardiovascular PET-CT and the likely direction of future developments.

Aortic stenosis, atherosclerosis, and skeletal bone: is there a common link with calcification and inflammation?

AIMS: The pathophysiology of aortic stenosis shares many similarities with atherosclerosis and skeletal bone formation. Using non-invasive imaging, we compared aortic valve calcification and inflammation activity with that measured in atherosclerosis and bone. METHODS AND RESULTS: Positron emission and computed tomography was performed using 18F-sodium fluoride (18F-NaF, calcification) and 18F-fluorodeoxyglucose (18F-FDG, inflammation) in 101 patients with calcific aortic valve disease (81 aortic stenosis and 20 aortic sclerosis). Calcium scores and positron emission tomography tracer activity (tissue-to-background ratio; TBR) were measured in the aortic valve, coronary arteries, thoracic aorta, and bone. Over 90% of the cohort had coexistent calcific atheroma, yet correlations between calcium scores were weak or absent (valve vs. aorta r(2) = 0.015, P = 0.222; valve vs. coronaries r(2) = 0.039, P = 0.049) as were associations between calcium scores and bone mineral density (BMD vs. valve r(2) = 0.000, P = 0.766; vs. aorta r(2) = 0.052, P = 0.025; vs. coronaries r(2) = 0.016, P = 0.210). 18F-NaF activity in the valve was 28% higher than in the aorta (TBR: 2.66 ± 0.84 vs. 2.11 ± 0.31, respectively, P < 0.001) and correlated more strongly with the severity of aortic stenosis (r(2) = 0.419, P < 0.001) than 18F-NaF activity outwith the valve (valve vs. aorta r(2) = 0.167, P < 0.001; valve vs. coronary arteries r(2) = 0.174, P < 0.001; valve vs. bone r(2) = 0.001, P = 0.806). In contrast, 18F-FDG activity was lower in the aortic valve than the aortic atheroma (TBR: 1.56 ± 0.21 vs. 1.81 ± 0.24, respectively, P < 0.001) and more closely associated with uptake outwith the valve (valve vs. aorta r(2) = 0.327, P < 0.001). CONCLUSION: In patients with aortic stenosis, disease activity appears to be determined by local calcific processes within the valve that are distinct from atherosclerosis and skeletal bone metabolism.

Iterative reconstruction and individualized automatic tube current selection reduce radiation dose while maintaining image quality in 320-multidetector computed tomography coronary angiography.

AIM: To assess the effect of two iterative reconstruction algorithms (AIDR and AIDR3D) and individualized automatic tube current selection on radiation dose and image quality in computed tomography coronary angiography (CTCA). MATERIALS AND METHODS: In a single-centre cohort study, 942 patients underwent electrocardiogram-gated CTCA using a 320-multidetector CT system. Images from group 1 (n = 228) were reconstructed with a filtered back projection algorithm (Quantum Denoising Software, QDS+). Iterative reconstruction was used for group 2 (AIDR, n = 379) and group 3 (AIDR3D, n = 335). Tube current was selected based on body mass index (BMI) for groups 1 and 2, and selected automatically based on scout image attenuation for group 3. Subjective image quality was graded on a four-point scale (1 = excellent, 4 = non-diagnostic). RESULTS: There were no differences in age (p = 0.975), body mass index (p = 0.435), or heart rate (p = 0.746) between the groups. Image quality improved with iterative reconstruction and automatic tube current selection [1.3 (95% confidence intervals (CI): 1.2-1.4), 1.2 (1.1-1.2) and 1.1 (1-1.2) respectively; p < 0.001] and radiation dose decreased [274 (260-290), 242 (230-253) and 168 (156-180) mGy cm, respectively; p < 0.001]. CONCLUSION: The application of the latest iterative reconstruction algorithm and individualized automatic tube current selection can substantially reduce radiation dose whilst improving image quality in CTCA.

Conversion of lamellar body membranes into tubular myelin in alveoli of fetal rat lungs.

Fluid-filled lumina of fetal rat lungs contain lamellar bodies (LBs) as well as tubular myelin (TM), both of which are thought to be stores of phospholipid-rich pulmonary surfactant. The alveolar epithelium is believed to secrete LBs, but neither the origin nor the mechanism of TM formation is entirely certain. The main objective of this study was to determine the relationship between secreted LBs and TM and to define membrane phenomena which occur during TM formation. I examined lung tissues of 20-21 day-old fetuses (day 22 = term) using transmission and high voltage transmission electron microscopy and cytochemistry. My findings indicate that secreted LBs, identified by the presence of an acid-phosphatase reactive core, are the precursor of TM. Secreted LBs are highly organized structures which contain structurally specialized areas, one of which is a "mini-lattice" structure similar to TM. During TM formation, fuzzes or 8.0-nm diameter particles appear on transition membranes, although LB membranes appear to lack both structures. Similar particles are present on TM membranes and are generally associated with membrane intersections. My results provide evidence that TM is formed from LBs within the alveolar lumen by mechanisms which may be complex.

Permeability of muscle capillaries to microperoxidase.

In this study we attempted to identify a morphologic counterpart of the small pore of muscle capillaries. The existence of such a pore has been postulated by physiologists to explain the permeability of muscle capillaries to small macromolecules. We injected mice intravenously with microperoxidase (MP) and fixed specimens of diaphragm at intervals of 0-250 s after the injection to localize the tracer by electron microscopy. The small size of MP (1,900 mol wt and 20 A molecular diameter [MD]) ensures its ready passage through the small pore since the latter is thought to be either a cylindrical channel 90 A in diameter or a slit 55 A wide. MP appears in the pericapillary interstitium within 30 s of initiation of its intravenous injection. The patterns of localization of MP observed within clefts between adjacent capillary endothelial cells indicate that some endothelial junctions are permeable to this tracer. Although small vesicles transfer MP across the endothelium, we do not believe that the vesicles transfer substantial amounts of MP into the pericapillary interstitium. We did not obtain evidence that MP crosses the endothelium of capillaries through channels formed either by a single vesicle or by a series of linked vesicles opening simultaneously at both surfaces of the endothelial cell. From our observations we conclude that some endothelial junctions of capillaries are permeable to MP, and that these permeable junctions are a plausible morphologic counterpart of the small pore.

The permeability of muscle capillaries to horseradish peroxidase.

The main objective of this study was to determine the pathways by which horseradish peroxidase (HRP) can cross the endothelium of muscle capillaries. Specimens of mouse diaphragm were fixed for cytochemical analysis at various intervals after intervenous injection of 0.5 mg HRP, at 4 min after intervenous injection of varied amounts of HRP, and at 4 min after intervenous injections in various volumes of isotonic NaCl. Our findings indicate that endothelial junctions serve as a barrier which may allow passage of very limited amounts of HRP. They also suggest that endothelial vesicles transfer HRP from the capillary lumen to the pericapillary interstitium as well as in the reverse direction. Increasing the volume of solution injected to approximately 30% of total blood volume did not increase the amount of HRP that left the capillary lumen. Our results with HRP do not provide clearcut evidence that endothelial junctions are the site of the small pore.

The pathology of tuberculosis caused by Mycobacterium tuberculosis in a herd of semi-free-ranging springbok (Antidorcas marsupialis).

The first detailed description of the pathology of tuberculosis, caused by Mycobacterium tuberculosis, in springbok is reported. The springbok were part of a semi-free-ranging herd kept on the grounds of iThemba Laboratory for Accelerator Based Science (LABS) in the Kuils River district of the Western Cape Province, South Africa. Mycobacterium tuberculosis was isolated from three animals out of a total of 33 sampled, with two animals showing tuberculosis lesions. The index case was an adult ewe that showed advanced miliary tuberculosis with marked macroscopic and microscopic lesions in the lungs, pleura and respiratory lymph nodes, and numerous acid-fast bacilli. Six healthy rams were sampled nine months later and a pilot study indicated miliary tuberculosis lesions in one ram, which again were macroscopically most prominent in the lungs, pleura and respiratory lymph nodes. Macroscopic lesions were also noted in the sternal, iliac, prefemoral and retropharyngeal lymph nodes. Microscopy in this animal revealed lesions in the macroscopically affected organs as well as numerous other lymph nodes, and suspected lesions occurred in the testicle and colon. Acid-fast bacilli were scarce to moderate in affected organs. Because of the miliary nature of the lesions in both affected animals, the route of infection could not be established conclusively. The lesions in most affected organs of both animals resembled classical tuberculous granulomas. A main study conducted on healthy animals 19 months after the pilot study failed to find any animal with tuberculosis lesions in the group of 25 sampled, and all were negative for mycobacteria via mycobacterial culture.

Multimodality quantitative assessments of myocardial perfusion using dynamic contrast enhanced magnetic resonance and 15O-labelled water positron emission tomography imaging.

Kinetic modelling of myocardial perfusion imaging data allows the absolute quantification of myocardial blood flow (MBF) and can improve the diagnosis and clinical assessment of coronary artery disease (CAD). Positron emission tomography (PET) imaging is considered the reference standard technique for absolute quantification, whilst oxygen-15 (15O)-water has been extensively implemented for MBF quantification. Dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) has also been used for MBF quantification and showed comparable diagnostic performance against (15O)-water PET studies. We investigated for the first time the diagnostic performance of two different PET MBF analysis softwares PMOD and Carimas, for obstructive CAD detection against invasive clinical standard methods in 20 patients with known or suspected CAD. Fermi and distributed parameter modelling-derived MBF quantification from DCE-MRI was also compared against (15O)-water PET, in a subgroup of 6 patients. The sensitivity and specificity for PMOD was significantly superior for obstructive CAD detection in both per vessel (0.83, 0.90) and per patient (0.86, 0.75) analysis, against Carimas (0.75, 0.65), (0.81, 0.70), respectively. We showed strong, significant correlations between MR and PET MBF quantifications (r=0.83-0.92). However, DP and PMOD analysis demonstrated comparable and higher haemodynamic differences between obstructive versus (no, minor or non)-obstructive CAD, against Fermi and Carimas analysis. Our MR method assessments against the optimum PET reference standard technique for perfusion analysis showed promising results in per segment level and can support further multi-modality assessments in larger patient cohorts. Further MR against PET assessments may help to determine their comparative diagnostic performance for obstructive CAD detection.

Discoidal bilayer structure of nascent high density lipoproteins from perfused rat liver.

Rat livers were perfused for 6 h without added plasma proteins using washed erythrocytes and buffer in a recirculating system. An inhibitor to the enzyme lecithin-cholesterol acyltransferase (5,5'-dithionitrobenzoic acid) was added in some experiments to prevent modification of substrate-lipids contained in secreted lipoproteins. The inhibitor did not detectably alter hepatic ultrastructure or gas exchange, but it inhibited the secreted lecithin-cholesterol acyltransferase by more than 85%. Very low density lipoproteins in perfusate were unaltered but the high density lipoproteins obtained from livers perfused with the inhibitor appeared disk-shaped in negative stain by electron microscopy with a mean edge thickness of 46 +/- 5 A and a mean diameter of 190 +/- 25 A. The high density lipoproteins were composed predominantly of polar lipids and protein with only small amounts of cholesteryl esters and triglycerides. The major apoprotein of these discoidal fractions had the same electrophoretic mobility as the arginine-rich apoprotein, whereas plasma high density lipoproteins contained mainly the A-I approtein. In all these respects the discoidal perfusate high density lipoproteins closely resemble those found in human plasma which is deficient in lecithin-cholesterol acyltransferase. Perfusate high density lipoproteins obtained in the absence of the enzyme inhibitor more closely resembled plasma high density lipoproteins in chemical composition (content of cholesteryl esters and apoproteins) and in electron microscopic appearance. Purified lecithin-cholesterol acyltransferase synthesized cholesteryl esters at a substantially faster rate from substrate lipids of perfusate high density lipoproteins than those from plasma. The discoidal high density lipoproteins were the best substrate for this reaction. Thin sections of plasma high density lipoproteins indicated a spherical particle whereas discoidal high density lipoproteins stained with the characteristic trilaminar image of membranes. These observations suggest that the liver secretes disk-shaped lipid bilayer particles which represent both the nascent form of high density lipoproteins and preferred substrate for lecithin-cholesterol acyltransferase.

The FEMA GRAS assessment of aromatic substituted secondary alcohols, ketones, and related esters used as flavor ingredients.

This publication is the 11th in a series of safety evaluations performed by the Expert Panel of the Flavor and Extract Manufacturers Association (FEMA). In 1993, the Panel initiated a comprehensive program to re-evaluate the safety of more than 1700 GRAS flavoring substances under conditions of intended use. The list of GRAS substances has now grown to more than 2100 substances. Elements that are fundamental to the safety evaluation of flavor ingredients include exposure, structural analogy, metabolism, pharmacokinetics and toxicology. Flavor ingredients are evaluated individually and in the context of the available scientific information on the group of structurally related substances. In this monograph, a detailed interpretation is presented on the renal carcinogenic potential of the aromatic secondary alcohol alpha-methylbenzyl alcohol, aromatic ketone benzophenone, and corresponding alcohol benzhydrol. The relevance of these effects to the flavor use of these substances is also discussed. The group of aromatic substituted secondary alcohols, ketones, and related esters was reaffirmed as GRAS (GRASr) based, in part, on their rapid absorption, metabolic detoxication, and excretion in humans and other animals; their low level of flavor use; the wide margins of safety between the conservative estimates of intake and the no-observed-adverse effect levels determined from subchronic and chronic studies and the lack of significant genotoxic and mutagenic potential.

Chronic vanadium poisoning in calves and its treatment with calcium disodium ethylenediaminetetraacetate.

Sixteen Friesland heifer calves aged between 96 and 157 days were removed from a dairy farm that had been polluted with vanadium and randomly allocated into two equal groups (n = 8). The objective of the trial was to determine whether calcium disodium ethylenediaminetetraacetate (CaNa(2)EDTA) could be used as a treatment for cattle running in environments high in background vanadium. The treatment group received 80 mg CaNa(2)EDTA per kg body weight intraperitonealy (i.p.) twice a week over a 10-week period. The control group received normal saline i.p. over the same period. During the trial calves were exposed to a daily intake of vanadium in the form of contaminated tef hay derived from the farm of origin. In addition, the total mixed ration was spiked with a further 20 mg V(2)O(5)/kg feed to compensate for possible on-farm inhalation exposure. A stochastic model was used to estimate daily intake of vanadium as a distribution function. The model estimated that the daily intake of vanadium varied between an absolute minimum of 33 mg/day to an absolute maximum of 124 mg/day. The average intake of vanadium was 71.8 mg per day per calf. Various chemical pathology parameters were measured throughout the trial as well as urine excretion rates of vanadium and lymphocyte stimulation counts. All calves were slaughtered and necropsied in cohorts of 4-6 animals at monthly intervals after completion of the trial and withdrawal of vanadium from the ration. Tissue concentrations of vanadium were determined and necropsy findings were noted. The study found that CaNa(2)EDTA appears to enhance the excretion of vanadium in calves, but could not prove that the treatment had a protective effect against vanadium exposure. Calves were able to tolerate the prolonged treatment with CaNa(2)EDTA without side-effects.

Phospholipid composition and ultrastructure of A549 cells and other cultured pulmonary epithelial cells of presumed type II cell origin.

In order to assess the usefulness of A549, L-2, and AK-D cell lines as model systems for alveolar type II cells, we compared their phospholipid composition to that of fibroblasts grown under similar conditions. The percentage of disaturated phosphatidylcholine and phosphatidylglycerol, key phospholipids of purified surface-active material, was the same in epithelial cells and fibroblasts. When A549 cells were maintained in serum-free media for two days, ultrastructural examination showed an increase in cytoplasmic lamellar inclusions but there was no change in the percentage of disaturated phosphatidylcholine or phosphatidylglycerol. Because the lipid content of these cultured cells was very different from that of freshly isolated rat type II cells, we conclude that their suitability as model cell systems for type II cells is questionable.

Monoclonal antibodies specific to apical surfaces of rat alveolar type I cells bind to surfaces of cultured, but not freshly isolated, type II cells.

The alveolar surface of the lung is lined by two classes of epithelial cells, type I and type II cells. Type I cells cover more than 97% of the alveolar surface. Although this cell type is felt to be essential for normal gas exchange, neither unique identifying characteristics nor functions have been described for the type I cell. We have produced monoclonal antibodies to (a) component(s) of molecular weight 40,000 and 42,000 of the apical surface of rat alveolar type I cells. The antibodies are specific to the lung in Western blots of organ homogenates. In immunocytochemical studies of frozen lung at the level of both light and electron microscopy, the monoclonal antibodies appear to react specifically with the apical plasma membrane of type I cells. Airway, vascular, interstitial cells, type II cells and macrophages are not immunoreactive. Western blots of isolated type I cells (approx. 70% pure) also demonstrate immunoreactivity at molecular weights of 40,000 and 42,000. When the lung is injured, type I cells may be damaged and sloughed from the alveolar surface. Alveolar repair occurs when the second type of alveolar cell, the type II cell, divides. Cell progeny may retain type II cell morphology or may differentiate into type I cells. Western blots of freshly isolated type II cells (approx. 85% pure) do not display immunoreactivity with our monoclonal antibodies. However, type II cells maintained in culture acquire immunoreactivity to monoclonal antibodies, demonstrating that type II cells in vitro have the capacity to develop a characteristic associated with type I cells in situ. The availability of markers for a specific membrane component of type I cells should facilitate the study of many questions on alveolar functions, development and response to injury.

Role of calcium ions the structure and function of pulmonary surfactant.

Pulmonary surfactant isolated by centrifugation in buffers containing ions contains at least three different morphologic structures. The presence of one of these, tubular myelin, is dependent on calcium ions, since chelation of the calcium ions causes disruption of this structure. Addition of EDTA also decreases the ability of the surfactant to absorb rapidly to air-water interfaces and lower surface tension. Titration with calcium ions (2.5 or 5 mM) restores rapid surface adsorption and restores the tubular myelin structural forms. Magnesium ions cannot substitute for calcium ions in these processes. The reversibility of structure and function induced by calcium ions and EDTA is also accompanied by reversible isopycnic density shifts probably related to aggregation and disaggregation of the lipid-protein complex with calcium ions and EDTA, respectively.

Changes in biochemical characteristics and pattern of lectin binding of alveolar type II cells with time in culture.

When cultured on plastic culture dishes for several days, alveolar type II cells gradually lose both their morphologic and biochemical identifying characteristics. Although type II cells cultured on a matrix derived from corneal endothelial cells have previously been reported to retain lamellar bodies for 7-10 days in culture, the ability of type II cells cultured on matrix to synthesize surfactant lipids has not been previously studied. We therefore measured the phospholipid content and the distribution of [14C]acetate into classes of lipids by type II cells maintained in culture. We found no differences between cells cultured on plastic or on matrix. We then studied the binding to type II cells in culture of Maclura pomifera and Ricinus communis I, lectins specific in vivo for type II and type I cells, respectively. We found that the cells progressively bind less M. pomifera and more R. communis I. The change in pattern of lectin binding occurs whether cells are cultured on plastic or matrix, whether lectins are conjugated with fluorescein, rhodamine or ferritin, or whether cells are cultured in the presence or absence of serum. We conclude that type II cells cultured on either tissue culture plastic or matrix derived from corneal endothelial cells lose the ability to synthesize and contain surfactant phospholipids, and, at least in their pattern of lectin binding, become similar to type I cells.