Pleural function and lymphatics.

The pleural space plays an important role in respiratory function as the negative intrapleural pressure regimen ensures lung expansion and in the mean time maintains the tight mechanical coupling between the lung and the chest wall. The efficiency of the lung-chest wall coupling depends upon pleural liquid volume, which in turn reflects the balance between the filtration of fluid into and its egress out of the cavity. While filtration occurs through a single mechanism passively driving fluid from the interstitium of the parietal pleura into the cavity, several mechanisms may co-operate to remove pleural fluid. Among these, the pleural lymphatic system emerges as the most important one in quantitative terms and the only one able to cope with variable pleural fluid volume and drainage requirements. In this review, we present a detailed account of the actual knowledge on: (a) the complex morphology of the pleural lymphatic system, (b) the mechanism supporting pleural lymph formation and propulsion, (c) the dependence of pleural lymphatic function upon local tissue mechanics and (d) the effect of lymphatic inefficiency in the development of clinically severe pleural and, more in general, respiratory pathologies.

Impact of respiratory pattern on lung mechanics and interstitial proteoglycans in spontaneously breathing anaesthetized healthy rats.

AIM: The aim of this study was to investigate the effect of different pattern of spontaneous breathing on the respiratory mechanics and on the integrity of the pulmonary extracellular matrix. METHODS: Experiments were performed on adult healthy rats in which different spontaneously breathing pattern was elicited through administration of two commonly used anaesthetic mixtures: pentobarbital/urethane (P/U) and ketamine/medetomidine (K/M). The animals (five per group) were randomized and left to spontaneously breath for 10 min (P/U-sham; K/M-sham) or for 4h (P/U-4h; K/M-4h), targeting the anaesthesia level to obtain a tidal volume of about 8 mL kg(-1) body wt. At the end of the experiment, lung matrix integrity was assessed through determination of the glycosaminoglycans (GAGs) content in the lung parenchyma. RESULTS: Compared with K/M, anaesthesia with P/U cocktail induced: (1) a higher respiratory rate and minute ventilation attained with lower P(a) CO(2) ; (2) a higher pressure-time-product and work of breathing per minute; (3) a lower static lung compliance; (4) an increased activation of lung tissue metalloproteases; and (5) greater extraction of pulmonary interstitial GAGs. CONCLUSIONS: This study suggests that the breathing pattern induced by the different anaesthetic regimen may damage the pulmonary interstitium even during spontaneous breathing at physiological tidal volumes.

International Programme for Resource Use in Critical Care (IPOC)--a methodology and initial results of cost and provision in four European countries.

BACKGROUND: A standardized top-down costing method is not currently available internationally. An internally validated method developed in the UK was modified for use in critical care in different countries. Costs could then be compared using the World Health Organization's Purchasing Power Parities (WHO PPPs). METHODS: This was an observational, retrospective, cross-sectional, multicentre study set in four European countries: France, UK, Germany and Hungary. A total of 329 adult intensive care units (ICUs) participated in the study. RESULTS: The costs are reported in international dollars ($) derived from the WHO PPP programme. The results show significant differences in resource use and costs of ICUs over the four countries. On the basis of the sum of the means for the major components, the average cost per patient day in UK hospitals was $1512, in French hospitals $934, in German hospitals $726 and in Hungarian hospitals $280. CONCLUSIONS: The reasons for such differences are poorly understood but warrant further investigation. This information will allow us to better adjust our measures of international ICU costs.

The cost of a hospital ward in Europe: is there a methodology available to accurately measure the costs?

Costing health care services has become a major requirement due to an increase in demand for health care and technological advances. Several studies have been published describing the computation of the costs of hospital wards. The objective of this article is to examine the methodologies utilised to try to describe the basic components of a standardised method, which could be applied throughout Europe. Cost measurement however is a complex matter and a lack of clarity exists in the terminology and the cost concepts utilised. The methods discussed in this review make it evident that there is a lack of standardized methodologies for the determination of accurate costs of hospital wards. A standardized costing methodology would facilitate comparisons, encourage economic evaluation within the ward and hence assist in the decision-making process with regard to the efficient allocation of resources.

Pulmonary microvascular and perivascular interstitial geometry during development of mild hydraulic edema.

To study pulmonary arteriolar vasomotion in control conditions and in the transition to hydraulic edema, changes in subpleural pulmonary arteriolar diameter and perivascular interstitial volume were evaluated in anesthetized spontaneously breathing rabbits. Images of subpleural pulmonary microvessels were recorded in control conditions and for up to 180 min during a 0.5 ml x kg(-1) x min(-1) intravenous saline infusion through an intact parietal pleural window. Images were digitized and analyzed with a semiautomatic procedure to determine vessel diameter and perivascular interstitial thickness from which interstitial fluid volume was derived. In control vessels, the diameter of approximately 30-, approximately 50-, and approximately 80-microm arterioles and the perivascular interstitial thickness were fairly stable. During infusion, the diameter increased maximally by 20% in approximately 30 microm vessels, was unchanged in approximately 50 microm vessels, and decreased by 25% in approximately 80-microm arterioles; the perivascular interstitial volume increased by 54% only around 30-microm microvessels. In papaverine-treated rabbits, all arterioles dilated and a larger increase in perivascular interstitial thickness was observed. The data suggest that the opposite vasomotor behavior of 30- and 80-microm arterioles during development of mild edema may represent a local specific response of the pulmonary microcirculation to reduce capillary pressure in the face of an increased transendothelial fluid filtration, thus counteracting progression toward severe edema.

Development of lung edema: interstitial fluid dynamics and molecular structure.

Pulmonary interstitium is maintained dehydrated at subatmospheric pressure (-10 cmH(2)O) through low capillary permeability, low tissue compliance, and an efficient lymphatic drainage. Enzymatic degradation of proteoglycans disrupts the endothelial basal membrane and the matrix structure, triggering the development of pulmonary edema.

Pulmonary interstitial pressure and tissue matrix structure in acute hypoxia.

Pulmonary interstitial pressure was measured via micropuncture in anesthetized rabbits in normoxia and after breathing 12% O(2). In normoxia [arterial PO(2) = 88 +/- 2 (SD) mmHg], pulmonary arterial pressure and pulmonary interstitial pressure were 16 +/- 8 and -9.6 +/- 2 cmH(2)O, respectively. After 6 h of hypoxia (arterial PO(2) = 39 +/- 16 mm Hg), the corresponding values were 30+/-8 and 3.5+/-2.5 cm H(2)O (P<0.05). Pulmonary interstitial proteoglycan extractability, evaluated by hexuronate assay after 0.4 M guanidinium hydrochloride extraction, was 12.3, 32.4, and 60.6 microg/g wet tissue in normoxia and after 3 and 6 h of hypoxia, respectively, indicating a weakening of the noncovalent bonds linking proteoglycans to other extracellular matrix components. Gel filtration chromatography showed an increased fragmentation of chondroitin sulfate- and heparan sulfate-proteoglycans during hypoxic exposure, accounting for a loss of extracellular matrix native architecture and basement membrane structure. Gelatin zymography demonstrated increased amounts of the proteolytically activated form of gelatinase B (matrix metalloproteinase-9) after hypoxic exposure, providing evidence that the activation of proteinases may play a role in hypoxia-induced lung injury.

Isolation of pulmonary interstitial fluid in rabbits by a modified wick technique.

Interstitial fluid protein concentration (C(protein)) values in perivascular and peribronchial lung tissues were never simultaneously measured in mammals; in this study, perivascular and peribronchial interstitial fluids were collected from rabbits under control conditions and rabbits with hydraulic edema or lesional edema. Postmortem dry wicks were implanted in the perivascular and peribronchial tissues; after 20 min, the wicks were withdrawn and the interstitial fluid was collected to measure C(protein) and colloid osmotic pressure. Plasma, perivascular, and peribronchial C(protein) values averaged 6.4 +/- 0.7 (SD), 3.7 +/- 0.5, and 2.4 +/- 0.7 g/dl, respectively, in control rabbits; 4.8 +/- 0.7, 2.5 +/- 0.6, and 2.4 +/- 0.4 g/dl, respectively, in rabbits with hydraulic edema; and 5.1 +/- 0.3, 4.3 +/- 0.4 and 3.3 +/- 0.6 g/dl, respectively, in rabbits with lesional edema. Contamination of plasma proteins from microvascular lesions during wick insertion was 14% of plasma C(protein). In control animals, pulmonary interstitial C(protein) was lower than previous estimates from pre- and postnodal pulmonary lymph; furthermore, although the interstitium constitutes a continuum within the lung parenchyma, regional differences in tissue content seem to exist in the rabbit lung.

Subatmospheric pressure in the rabbit pleural lymphatic network.

1. Hydraulic pressure in intercostal and diaphragmatic lymphatic vessels was measured through the micropuncture technique in 23 anaesthetised paralysed rabbits. Pleural lymphatic vessels with diameters ranging from 55 to 950 microm were observed under stereomicroscope view about 3-4 h after intrapleural injection of 20 % fluorescent dextrans. 2. Lymphatic pressure oscillated from a minimum (Pmin) to a maximum (Pmax) value, reflecting oscillations in phase with cardiac activity (cardiogenic oscillations) and lymphatic myogenic activity. With intact pleural space, Pmin in submesothelial diaphragmatic lymphatic vessels of the lateral apposition zone was -9.1 +/- 4.2 mmHg, more subatmospheric than the simultaneously recorded pleural liquid pressure amounting to -3.9 +/- 1.2 mmHg. In extrapleural intercostal lymphatic vessels Pmin averaged -1.3 +/- 2. 7 mmHg. 3. Cardiogenic pressure oscillations (Pmax - Pmin), were observed in all recordings; their mean amplitude was about 5 mmHg and was not dependent upon frequency of cardiac contraction, nor lymphatic vessel diameter, nor the Pmin value. 4. Intrinsic contractions of lymphatic vessel walls caused spontaneous pressure waves of about 7 mmHg in amplitude at a rate of 8 cycles min-1. 5. These results demonstrated the ability of pleural lymphatic vessels to generate pressure oscillations driving fluid from the subatmospheric pleural space into the lymphatic network.

The sensitivity of versican from rabbit lung to gelatinase A (MMP-2) and B (MMP-9) and its involvement in the development of hydraulic lung edema.

Large chondroitinsulphate-containing proteoglycan (versican) isolated from rabbit lung was cleaved by purified gelatinase A (MMP-2) and gelatinase B (MMP-9), as well as by crude enzyme extract from rabbit lung with hydraulic edema. Gelatine zymography, performed after purification of gelatinases by affinity chromatography, demonstrated that the enzyme extract contained two main gelatinolytic bands at about 92 kDa and 72 kDa, identified by specific antisera as the latent proMMP-9 and proMMP-2, respectively. Moreover, enzyme extract from edematous lung showed an increased amount of the proteolytically activated forms of both gelatinases with respect to normal controls. These results suggest that MMP-2 and MMP-9 are involved in the breakdown of versican occurring in rabbit lung during the development of hydraulic edema.

Involvement of lung interstitial proteoglycans in development of hydraulic- and elastase-induced edema.

We extracted and isolated proteoglycans from lung tissue samples obtained from three groups of anesthetized rabbits: 1) control animals (C; n = 8) killed by overdose after 180 min; 2) animals receiving an intravenous saline infusion (S; n = 4, 1.5 ml . kg-1 . min-1) for 180 min; 3) animals receiving an intravenous bolus of 200 microg of pancreatic elastase (E; n = 4), killed after 200 min. The lung dry weight-to-wet weight ratio in the three groups was 5.2 +/- 0.2, 6.0 +/- 0.4, and 5.6 +/- 0.5, respectively. Gel-filtration analysis showed a massive fragmentation of the versican family of the extracellular matrix (ECM) in the S groups and a marked degradation of heparan sulfate-containing proteoglycans, including perlecan of the basement membrane, in the E group. The binding properties of total proteoglycans to other ECM components were lowered in both groups relative to control. The decrease in proteoglycan binding was more pronounced for collagen type IV in the E group relative to C (-93.5%, P < 0.05) and for hyaluronic acid in the S groups (-85.8%, P < 0.05). These findings suggest that elastase treatment produces a major degree of damage to the organization of basement membrane, whereas saline loading affects more markedly the architecture of interstitial ECM. Qualitative zymography performed on lung extracts showed increased gelatinase activities in both S and E groups, providing direct evidence that the activation of tissue proteinases may play a role in acute lung injury.

Proteoglycan involvement during development of lesional pulmonary edema.

We evaluated the effect of pancreatic elastase (7 IU i.v.) on pulmonary interstitial pressure (Pip) in in situ rabbit lungs by a micropuncture technique through the intact parietal pleura. Pip was -10.8 +/- 2.2 (SD) cmH2O in the control condition, increased to +5.1 +/- 1.7 cmH2O at approximately 60 min [condition referred to as mild edema (ME)], and subsequently decreased to -0.15 +/- 0.8 cmH2O, remaining steady from 80 up to 200 min with a marked increase in lung wet-to-dry weight ratio [condition referred to as severe edema (SE)], suggesting an increase in tissue compliance. We functionally correlated the measured Pip to structural modifications of proteoglycans, the major interfibrillar component of the extracellular matrix (ECM). The strength of the noncovalent bonds linking proteoglycans to other ECM components decreased with increasing severity of edema, as indicated by the increased extractability of proteoglycans with guanidine hydrochloride. Total proteoglycan recovery (expressed as microgram hexuronate/g dry tissue) increased from 436.8 +/- 14 in the control condition to 495.3 +/- 23 and 547.0 +/- 10 in ME and SE, respectively. Gel-filtration chromatography showed in ME a fragmentation of heparan sulfate proteoglycans, suggesting that elastase treatment first affected basement membrane integrity, whereas large chondroitin sulfate proteoglycans were degraded only in SE. Elastase caused a fragmentation only of the core protein of proteoglycans, the binding properties of which to collagens, fibronectin, and hyaluronic acid were markedly decreased, as indicated by a solid-phase binding assay. The sequential degradation of heparan sulfate and chondroitin sulfate proteoglycans may account for the initial increase in microvascular permeability, followed by a loss of the native architecture of the ECM, which may be responsible for the increase in tissue compliance.

Respiratory mechanics after 180 days space mission (EUROMIR'95).

The present study reports data on respiratory function of lung and chest wall following the 180 days long European - Russian EuroMir '95 space mission. Data reported refer to two subjects studied before the mission, on day 9 and 175 in flight and on days 1, 10, 12, 27 and 120 after return. In-flight vital capacity (VC) and expiratory reserve volume (ERV) were similar to those in supine posture, namely approximately 5% and approximately 30% less than in sitting posture. On day 1 after return, VC was reduced by approximately 30% in both postures. This reflected a decrease in ERV (approximately 0.5 L) and in IC (inspiratory capacity, approximately 1.7 L) that could be attributed to a marked weakening of the respiratory muscles. Regain of normal preflight values barely occurred 120 days after return. Post-flight pressure-volume curves of the lung, chest wall and total respiratory system are equal to preflight ones. The pressure-volume curve of the lung in supine posture is displaced to the right relative to sitting posture and shows a lower compliance. As far as the lung in-flight condition resembles that occurring in supine posture, this implies a lower compliance, a greater amount of blood in the pulmonary microvascular bed, a more homogeneous lung perfusion and therefore a greater microvascular filtration rate towards lung interstitium.

Pulmonary interstitial pressure and proteoglycans during development of pulmonary edema.

In anesthetized adult rabbits, pulmonary perivascular interstitial pressure (P(ip)), measured by micropuncture technique with intact pleural space, averaged -10.5 +/- 1.9 (SD) cmH2O in control conditions, with a wet-to-dry lung weight ratio (W/D) of 4.8 +/- 0.2. Saline infusion (120 ml i.v. over 120 min) induced interstitial edema, increasing P(ip) to 3.62 +/- 1.6 cmH2O with no significant increase in W/D (5.13 +/- 0.1). For intravenous saline infusion exceeding 140 ml, P(ip) decreased to about atmospheric pressure with development of severe edema that was characterized by an increase of W/D ( > 7) with no further change in P(ip). In a separate set of animals, pulmonary interstitial proteoglycans (PGs) were investigated after sequential extraction of the tissue with 0.4 and 4 M guanidinium chloride (GuHCl) under control conditions and with interstitial (100 ml saline load in 100 min) and severe edema ( > 200 ml total infusion). The extractability of PGs increased constantly with increasing W/D. PG content in total extracts was evaluated by determination of hexuronate content which was 195.4 +/- 1.5 micrograms/g dry tissue in control lungs, 217.9 +/- 1.6 in interstitial edema, and 316.4 +/- 2.7 in severe edema. Moreover, edema development was coupled with an increase in efficiency of PG extraction with 0.4 M GuHCl. These findings suggested a weakening of PG interactions with other components of the extracellular matrix (ECM). Electrophoretic and gel-filtration analyses showed that the relative content of PG populations of large molecular size decreased constantly in 0.4 M GuHCl extract with increasing water loading. We propose relating the inflection of P(ip) in the transition from interstitial to severe edema to PG breakdown, which might greatly affect ECM structural organization, including collagen spreading and/or rupture of epithelial layer.

Pulmonary microvascular pressure profile during development of hydrostatic edema.

OBJECTIVE: To measure, in intact closed chest, the pressure in the pulmonary microvasculature during transition to mild interstitial edema. METHODS: In anesthetized spontaneously breathing rabbits, the pulmonary artery and left atrium were cannulated. Pleural windows were prepared to view the superficial pulmonary microvascular network through the intact parietal pleura. After intravenous infusion of 96.4 +/- 12.3 ml of saline at a rate of 0.5 ml/kg h, the hydraulic pressure in the pulmonary microvessels (15-240 microns in diameter) were measured using glass pipettes driven through the pleural window and connected to a servonull system. RESULTS: After saline, plasma protein concentration decreased from 6 +/- 1 to 4.8 +/- 0.5 g/dl; pulmonary arterial and left atrial pressures averaged 22.3 +/- 6.4 and 2.3 +/- 2 cm H2O in control and 23.1 +/- 4.2 and 4.2 +/- 2 cm H2O after infusion. After saline loading, 16.4% of total pressure drop occurred from pulmonary artery to 80-microns arterioles, 60.3% in 30-80 microns arterioles, 6.9% from 30-microns arterioles to 30-microns venules and 16.4% in the downstream segment. CONCLUSIONS: Mild interstitial edema induced, with respect to control, constriction of small arterioles and capillary recruitment to maintain a low capillary pressure. Hence, in initial edema, pulmonary circulation prevents further fluid filtration, acting like an intrinsic safety factor to delay development of severe edema.

Estimation of in vivo pulmonary microvascular and interstitial geometry using digital image analysis.

OBJECTIVE: To determine microvascular diameter and perivascular interstitium thickness at the lung surface in the in situ, in vivo lung. METHODS: Microscopic images of the lung surface collected through a "pleural window" by a videocamera were digitized with a monochrome frame grabber (512 x 512 pixels, 8 bits per pixels) to be computer analyzed by image processing techniques. RESULTS: We found that the maxima in the distribution of the standard deviations of gray levels in adjacent neighbors 7 x 7 pixels wide identify the edges between the microvessel lumen and the surrounding perivascular interstitium. Furthermore, the maxima in the distribution of the standard deviation of the standard deviations of gray levels identify the edges between the perivascular interstitium and the lung tissue. CONCLUSIONS: This technique can be applied to microvessels ranging in diameter from 30 microns to 200 microns and perivascular interstitial thickness of the order of 10-150 microns. Our approach allows for the definition of microvascular geometry even for noisy images and represents an improvement compared to other edge detection methods. The proposed analytical procedure may provide a useful tool to study lung fluid balance and microvascular reactivity in the in situ lung in the normal state and in response to a variety of functional conditions.

Contribution of lymphatic myogenic activity and respiratory movements to pleural lymph flow.

In 11 anesthetized spontaneously breathing rabbits, we studied the contribution to total pleural lymph flow of myogenic activity of pleural lymphatics ("intrinsic mechanism") and the effect due to mechanical action of respiratory movements ("extrinsic mechanism"). Isoncotic saline solution (5 ml) containing 100 microCi of 125I-lactate dehydrogenase (LDH) was injected into right pleural space; in all but three control rabbits, injectate contained 1 mM amiloride in dimethyl sulfoxide to induce relaxation of smooth muscle tone. At 3 h, rabbits were killed and pleural fluid was collected and its volume measured. LDH radioactivity in pleural liquid and parietal pleural tissue was counted. In control rabbits, net pleural liquid flow (Jnet) at 3 h was -0.17 +/- 0.04 (SD) ml.kg-1.h-1; LDH concentration (C) and quantity (Q) decreased by 40.3 and 51.1% of initial value, respectively; total pleural lymphatic flow (Jl), calculated from LDH clearance, was 0.58 +/- 0.01 ml.kg-1.h-1. In amiloride-treated rabbits, Jnet was 0.01 +/- 0.1 ml.kg-1.h-1, C decreased by 34.4% and Q by 33.1%, and Jl averaged 0.39 +/- 0.02 ml.kg-1.h-1. C in parietal pleura, rich in lymphatics, was 13-fold higher in control than in amiloride-treated animals. The significant decrease of pleural lymphatic flow observed with amiloride (-40% relative to control) resulted from impairment of intrinsic mechanism, whereas, at comparable breathing frequencies, extrinsic mechanism remained unaltered. The direct effect of topical application of 1 mM amiloride was confirmed on exposed mesenteric collecting lymphatic ducts (data from 5 rats): amiloride reduced lymph flow by 40% by decreasing stroke volume without greatly affecting contraction rate of lymphatic walls.

Permeability of parietal pleura to liquid and proteins.

The permselectivity of the parietal pleura was determined in spontaneously breathing anesthetized rabbits and dogs. In rabbits, we injected intrapleurally 5 ml of 1-g/dl albumin solution containing 100 microCi of 131I-labeled albumin plus 100 microCi of either lactate dehydrogenase (LDH) or alpha 2-125I-macroglobulin. Dogs received 100 ml of 1-g/dl albumin solution containing 100 microCi of 131I-albumin plus 100 microCi of alpha 2-125I-macroglobulin. A transpleural pressure gradient was set, lowering the intracapsular pressure to -30 cmH2O. The solvent drag reflection coefficients (sigma f) were calculated as the ratio between tracer concentrations in capsular and pleural liquid collected at 60-180 min. In rabbits sigma f was 0.44 +/- 0.2 (SD) for albumin, 0.84 +/- 0.1 for LDH, and 0.93 +/- 0.05 for alpha 2-macroglobulin. In dogs sigma f was 0.30 +/- 0.19 for albumin and 0.53 +/- 0.15 for alpha 2-macroglobulin. The hydraulic conductivity of the parietal pleura was 2.18 +/- 1.54 microliters.h-1.cmH2O-1.cm-2 in rabbits and 1.22 +/- 1.13 microliters.h-1.cmH2O-1.cm-2 in dogs. The parietal pleura could be modeled by two pore populations with radii of 83-89 and 156-222 A. The permeability coefficient averaged 0.08-0.21 x 10(-6) cm/s for albumin, 0.06-0.09 x 10(-6) cm/s for LDH, and 0.01-0.03 x 10(-6) cm/s for alpha 2-macroglobulin.