Pathology underrates colon cancer extranodal and nodal metastases; ex vivo radioimmunodetection helps staging.

PURPOSE: Colorectal carcinoma is frequently accompanied by small lymph nodes metastases that often escape pathologic examination. We evaluated whether ex vivo radioimmunodetection with the Affinity Enhancement System (AES) could improve detection of mesocolonic metastases. EXPERIMENTAL DESIGN: A bivalent 111In-labeled hapten was injected (16 patients) 4 days after a bispecific antibody (anticarcinoembryonic antigen, antihapten). Surgery was done 1 to 3 days later, and radioactive uptake in the mesocolon was recorded. Extensive pathologic examination of the mesocolon (reference method) was done after fat dissolution. This method visualizes all lymph nodes but is not in routine use. RESULTS: The reference method disclosed 705 nodes. There was no significant difference between the number of node metastases detected by AES or by the reference method (16 versus 17). Better detection would have been obtained by AES than by routine pathology (P<0.01). In addition 12 extranodal metastases were found in this study of which eight were detected by AES. The prognostic importance of such extranodal metastases has been underlined in the literature. Routine pathology combined with AES would have disclosed all node metastases and 86% of total metastases versus 35% by routine pathology alone. CONCLUSIONS: Ex vivo radioimmunodetection could improve nodal and extranodal metastases detection in patients with colorectal cancer. Its value for improving pathologic analysis, together with the effect of these small metastases on prognosis, should be further evaluated. The benefit of adjuvant chemotherapy for patients upstaged with radioimmunodection should also be assessed because adjuvant chemotherapy improves the 5-year survival of stage III patients.

Ventilator-induced lung injury.

During mechanical ventilation, high end-inspiratory lung volume (whether it be because of large tidal volume (VT) and/or high levels of positive end-expiratory pressure) results in a permeability type pulmonary oedema, called ventilator-induced lung injury (VILI). Previous injury sensitises lung to mechanical ventilation. This experimental concept has recently received a resounding clinical illustration after a 22% reduction of mortality was observed in acute respiratory distress syndrome patients whose VT had been reduced. In addition, it has been suggested that repetitive opening and closing of distal units at low lung volume could induce lung injury but this notion has been challenged both conceptually and clinically after the negative results of the Acute Respiratory Distress Syndrome clinical Network Assessment of Low tidal Volume and Elevated end-expiratory volume to Obviate Lung Injury (ARDSNet ALVEOLI) study. Experimentally and clinically, involvement of inflammatory cytokines in VILI has not been unequivocally demonstrated. Cellular response to mechanical stretch has been increasingly investigated, both on the epithelial and the endothelial side. Lipid membrane trafficking has been thought to be a means by which cells respond to stress failure. Alterations in the respiratory system pressure/volume curve during ventilator-induced lung injury that include decrease in compliance and position of the upper inflection point are due to distal obstruction of airways that reduce aerated lung volume. Information from this curve could help avoid potentially harmful excessive tidal volume reduction.

Glucose transporter gene expression in freshly isolated and cultured rat pneumocytes.

Alveolar epithelium in situ takes up luminal glucose by cotransport with sodium. Cultured alveolar type II pneumocytes have only sodium-independent glucose uptake. It is unclear which isoforms are responsible for glucose transport in these cells and why sodium-glucose cotransport activity disappears during culture. GLUT1, GLUT4, GLUT5 and SGLT1 mRNA were detected in freshly isolated rat alveolar type II cells by reverse transcriptase-polymerase chain reaction. We show that SGLT1 mRNA was 90% lower in cells cultured in plastic wells for 2 or 4 days than in freshly isolated cells. mRNAs coding for the facilitated transporters were reduced from 40% (GLUT1) and 75% (GLUT4 and GLUT5) in cultured cells. Cells cultured at the air-liquid interface better preserved their phenotype as attested by significantly higher surfactant-associated protein mRNA levels. However, these cells had no higher GLUT1 and SGLT1 gene expression. Thus, alveolar type II cells lose sodium-glucose cotransport activity in part because of a decrease in mRNA levels. These changes in gene expression and/or mRNA stability may be an additional consequence of the shift towards the type I cell phenotype observed in cultured type II pneumocytes.

Significance of the changes in the respiratory system pressure-volume curve during acute lung injury in rats.

UNLABELLED: The hypothesis that the changes in the respiratory system pressure- volume (PV) curve during pulmonary edema mainly reflect distal airway obstruction was investigated in rats. Normal rats had a well-defined upper inflection point (UIP) at low airway pressure. Airway occlusion by liquid instillation decreased compliance (Crs) and the volume (Vuip) of the UIP, and increased end-inspiratory pressure. The same changes were observed during the progression of edema produced by high volume ventilation (HV). Changes in Vuip and in Crs produced by HV were correlated with edema severity in normal rats or rats with lungs preinjured with alpha-naphthylthiourea. Vuip and Crs changes were proportional, reflecting compression of the PV curve on the volume axis and suggesting reduction of the amount of ventilatable lung at low airway pressure. In keeping with this explanation, the lower Vuip and Crs were before HV, the more severe HV-induced edema was in alpha-naphthylthiourea-injected rats. When edema was profuse, PV curves displayed a marked lower inflection point (LIP), the UIP at low pressure disappeared but another was seen at high volume above the LIP, and the correlation between Vuip changes and edema severity was lost. These observations may have clinical relevance in the context of the "open lung" strategy. KEYWORDS: ventilator-induced lung injury; respiratory mechanics; acute respiratory distress syndrome

Production of inflammatory cytokines in ventilator-induced lung injury: a reappraisal.

We investigated the production of proinflammatory cytokines by the lung during high mechanical stretch in vivo. To do this, we subjected rats to high-volume (42 ml/kg tidal volume [VT]) ventilation for 2 h. The animals developed severe pulmonary edema and alveolar flooding, with a high protein concentration in bronchoalveolar lavage fluid (BALF). The animals' BALF contained no tumor necrosis factor (TNF)-alpha, negligible amounts of interleukin (IL)-1beta, and less than 300 pg/ml of the chemokine macrophage inflammatory protein (MIP)-2, an amount similar to that found in rats ventilated with 7 ml/kg VT. Systemic cytokine levels were below the detection threshold. Because isolated lungs have been shown to produce high levels of proinflammatory cytokines when ventilated with a similarly high VT for the same duration (Tremblay, et al. J Clin Invest 1997;99:944-952), we reconsidered this specific issue. We ventilated isolated, unperfused rat lungs for 2 h with 7 ml/kg or 42 ml/kg VT, or maintained them in a statically inflated state. Negligible amounts of TNF-alpha were found in the BALF whatever the ventilatory condition applied. The BALF IL-1beta concentration was slightly elevated and higher in lungs ventilated with 42 ml/kg VT than in those ventilated with 7 ml/kg VT or in statically inflated lungs (p < 0.05). The BALF MIP-2 concentration was moderately elevated in all isolated lungs (200 to 300 pg/ml), and was slightly higher (p < 0.05) in lungs ventilated with 42 ml/kg VT. After lipopolysaccharide (LPS) challenge, high levels of TNF-alpha, IL-1beta, and MIP-2 were found in the animals' plasma before the lungs were removed. Negligible amounts of TNF-alpha and IL-1beta were retrieved from the BALF of statically inflated lungs. The concentrations of TNF-alpha and IL-1beta were higher in the BALF of ventilated lungs (p < 0.001). The TNF-alpha level did not differ with the magnitude of VT, whereas the level of IL-1beta was significantly higher in BALF of lungs ventilated with 42 ml/kg VT (p < 0.01). The MIP-2 concentrations were similar for the two ventilatory conditions. These results suggest that ventilation that severely injures lungs does not lead to the release of significant amounts of TNF-alpha or IL-1beta by the lung in the absence of LPS challenge but may increase lung MIP-2 production.

Spermine increases the active and passive transport across the alveolar epithelium in situ: effect of thiol reagents.

An intact alveolar epithelial barrier is thought to be important for alveolar liquid absorption. However, polycations increase alveolar permeability without affecting alveolar liquid absorption (Saumon et al., Am J Physiol 1995: 269:L185-L194). We have reconsidered this issue using polyamines. The polyamine spermine (10(-3) mol/l) produced a large (up to 20-fold), sustained increase in the permeability of the alveolar barrier to mannitol (PAMan) and in alveolar liquid absorption (Jw, twofold) in isolated rat lungs. These increases were inhibited by 5 x 10(-3) mol/l putrescine and 2 x 10(-3) mol/l spermidine. Because spermine is known to affect the phosphoinositide/Ca2+ signalling pathway, we evaluated the effects of thiol reagents known to interfere with this pathway in different ways. Thimerosal, a thiol reagent which sensitizes the inositol 1,4,5-trisphosphate (IP3) receptor, inhibited the spermine-induced increase in PA(Man) and, to a lesser extent, that of Jw. Mersalyl, a thiol reagent which blocks IP3-gated Ca2+ channels, enhanced spermine's effect, whereas N-ethylmaleimide, a non-specific thiol reagent, had no effect. These observations show that large increases in permeability may coexist with increases in Jw. They also suggest that the phosphoinositide/Ca2+ second messenger pathway is involved in modulating the tightness of the alveolar barrier and alveolar liquid absorption.

Chloride fluxes during cAMP stimulation of liquid absorption across the native rat alveolar epithelium.

cAMP activates Cl- channels in the alveolar epithelium of rabbits (Nielsen et al. Am J Physiol. 1998; 275:L1127-L1133), resulting in Cl- secretion; and of rats (Jiang et al. Am J Physiol. 1998; 275:C1610-C1620), resulting in Cl- absorption. The relationship between Cl- fluxes and liquid absorption across the alveolar barrier was examined using isolated perfused rat lungs. Unidirectional 36Cl- fluxes (apparent PACl products) in the apical-to-basal (absorption, PAClab) and reverse (secretion, PAClba) directions were identical under control conditions. Both increased about 2-fold during stimulation of liquid absorption by dibutyrylcAMP + isobutylmethylxanthine (P < .001). Inhibiting Na+ and liquid absorption during cAMP stimulation by adding amiloride to the alveolar instillate decreased PAClab to control level (P < .01), but did not affect PAClba. Neither alveolar liquid absorption nor PAClba was affected by apical N-phenylanthranilic acid or basolateral bumetanide during cAMP stimulation. Mannitol permeability--surface area products did not differ with the experimental condition. These observations indicate that cAMP stimulation results in enhanced Cl- and liquid absorption from rat airspaces and Cl- secretion into them. They suggest that Cl- absorption follows Na+ transport, but fail to demonstrate any significant participation of cAMP-activated Cl- channels in these changes.

A method for measuring the oxygen consumption of intact cell monolayers.

This report describes an open-air method for measuring the O(2) consumption (QO(2)) of intact monolayers of cultured cells. This method is based on Fick's second law of diffusion. It requires only a micromanipulator and a miniature O(2) electrode to measure the PO(2) gradient in the culture medium in the well. It was compared with the conventional oxygraph chamber method. Both methods gave the same value for QO(2) in freshly isolated rat type II cells: 166 +/- 15.3 nmol. h(-1). 10(6) cells(-1) for the open-air method and 151 +/- 11.6 nmol. h(-1). 10(6) cells(-1) for the oxygraph chamber method (n = 11 experiments). But the open-air method gave significantly larger values for QO(2) in cells cultured for 2 days (236 +/- 8.8 nmol. h(-1). 10(6) cells(-1)) than the oxygraph method (71 +/- 15.2 nmol. h(-1). 10(6) cells(-1); P < 0.001; n = 12 experiments). This suggests that the way cells are detached from their substratum to be placed in the oxygraph chamber affects their QO(2). The open-air method may be useful for studies on the metabolic properties of monolayers because the cells do not risk being damaged.

Alveolar permeability and liquid absorption during partial liquid ventilation of rats with perflubron.

We examined the effect of instilled perflubron (LiquiVent) on the transport properties of alveolar epithelium in anesthetized rats. Krebs-Ringer bicarbonate (1 to 4 ml) containing (125)I-albumin, [(3)H]mannitol and [(14)C] sucrose was instilled into airspaces either alone (n = 29), or with 1 (n = 21) or 2 (n = 12) ml perflubron and sampled 30 min later. Absorption was deduced from the changes in (125)I-albumin activity per unit volume in the airspace instillate, and changes in [(3)H]mannitol and [(14)C]sucrose activity per unit volume were used to evaluate the passive permeability of the alveolar-airway barrier. The rate of Ringer absorption depended on the volume instilled [0.38 (ml/h)/ml Ringer]. Perflubron (1 or 2 ml) increased Ringer absorption by 0.26 (p < 0. 001) and 0.19 ml/h (p < 0.05), respectively. However, 2 ml perflubron increased absorption less than did the same additional volume of Ringer (p < 0.001). The passive permeability of the alveolar-airway barrier increased exponentially with instilled Ringer volume. Sucrose/mannitol size selectivity was lost when Ringer volume was > 2 ml and albumin leaked from airspaces when it was 4 ml. Instillation of 2 ml perflubron prevented this increase in permeability, but 1 ml did not. No albumin leaked with perflubron even when the total volume of liquid in airspaces (Ringer + perflubron) was > 4 ml. These results suggest that perflubron can be beneficial in pulmonary edema by redistributing the alveolar liquid over a larger surface area, thus accelerating resorption. In addition, larger doses of perflubron may better preserve epithelial permeability during alveolar flooding.

Efficiency and safety of mechanical ventilation with a heat and moisture exchanger changed only once a week.

The cost of mechanical ventilation (MV) is high. Efforts to reduce this cost, as long as they are not detrimental for the patients, are needed. MV with heat and moisture exchangers (HME) changed every 48 h is safe, efficient, and cost-effective. Preliminary reports suggest that the life span of these filters may be prolonged. We determined prospectively whether a hygroscopic and hydrophobic HME (Hygrobac-Dar; Mallinckrodt) provided safe and efficient heating and humidification of the inspired gases when changed only once a week. Patients who were considered to require mechanical ventilation for more than 48 h were included in the study. HMEs were initially set for 7 d. Efficient airway heating and humidification were assessed by clinical parameters (number of tracheal suctionings and instillations required, peak airway pressures) and hygrometric measurements performed by psychrometry. Resistance was measured from Day 0 to Day 7. Bacterial colonization of circuits and HMEs was studied. A total of 377 days of mechanical ventilation with 60 HMEs was studied. Clinical parameters and hygrometric measurements did not change between Day 0 and Day 7. Mean absolute humidity was 30.3 +/- 1.3 mg H(2)O/L on Day 0 and 30.8 +/- 1.5 mg H(2)O/L on Day 7 (p = 0.7). Endotracheal tube occlusion never occurred. Three HMEs were replaced prematurely because of insufficient absolute humidity. This rare event occurred only in patients with COPD and after the third day of use. In addition, the absolute humidity delivered by the HMEs was significantly lower in patients with COPD than in the rest of the population. Resistance did not change from Day 0 to Day 7 (2.4 +/- 0.3 versus 2.7 +/- 0.3 cm H(2)O/L/s; p = 0.4). Bacterial samples of both circuits and ventilator sides of HMEs were sterile in most cases. We conclude that mechanical ventilation can be safely conducted in non-COPD patients using an HME changed only once a week, leading to substantial cost savings (about $110,000 per year if these findings were applied to the university-affiliated hospitals in Paris).

Alveolar sodium and liquid transport in mice.

We have developed a simple isolated lung preparation for measurement of liquid and solute fluxes across mouse alveolar epithelium. Liquid instilled into air spaces was absorbed at the rate (J(w)) of 3.7 +/- 0.32 ml x h(-1) x g dry lung wt(-1) x J(w) was significantly depressed by ouabain (P < 0.001) and amiloride (P < 0.001). Omission of glucose from the instillate or addition of the Na(+)-glucose cotransport inhibitor phloridzin did not affect J(w). However, the low epithelial lining fluid glucose concentration (one-third that of plasma), the larger-than-mannitol permeability of methyl-alpha-D-glucopyranoside, and the presence of Na(+)-glucose cotransporter SGLT1 mRNA in mouse lung tissue suggest that there is a Na(+)-glucose cotransporter in the mouse alveolar-airway barrier. Isoproterenol stimulated J(w) (6.5 +/- 0.45 ml x h(-1) x g dry lung wt(-1); P < 0.001), and this effect was blocked by amiloride, benzamil, ouabain, and the specific beta(2)-adrenergic antagonist ICI-118551 but not by atenolol. Similar stimulation was obtained with terbutaline (6.4 +/- 0.46 ml x h(-1) x g dry lung wt(-1)). Na(+) unidirectional fluxes out of air spaces varied in agreement with J(w) changes. Thus alveolar liquid absorption in mice follows Na(+) transport via the amiloride-sensitive pathway, with little contribution from Na(+)-glucose cotransport, and is stimulated by beta(2)-adrenergic agonists.

Hyperinflation-induced lung injury during alveolar flooding in rats: effect of perfluorocarbon instillation.

Mechanical nonuniformity of diseased lungs may predispose them to ventilator-induced lung injury (VILI) by overinflation of the more compliant, aerated zones. Perfluorocarbon (PFC) may reduce this nonuniformity by suppressing air-liquid interfaces. Saline (6.8 ml/kg) was instilled into the trachea to mimic alveolar edema and reduce aerated lung volume before mechanical ventilation (6, 16, 24, or 32 ml/kg tidal volume [VT]) for 10 min in rats. Flooding significantly aggravated VILI when VT was 24 or 32 ml/kg, with an increase in the distribution space of albumin in lungs (p < 0.001). Tracheal instillation of a low dose (3.3 ml/kg) of PFC (Liquivent) either before or after the instillation of saline considerably reduced VILI (p < 0.001). Saline instillation raised the lower inflection point of the respiratory system pressure-volume curve to values as high as 25 cm H2O, and produced a significant increase in end-inspiratory pressure (from 38 +/- 2.0 cm H2O to 61 +/- 2.4 cm H2O, for a VT of 32 ml/kg; p < 0.001). PFC significantly reduced the pressure at the lower inflection point and normalized end-inspiratory pressure. These decreases were correlated with a smaller albumin distribution space (p < 0.001). Animals in which PFC instillation failed to reduce the albumin space had pressures similar to those of animals given saline alone. In conclusion, the effectiveness of PFC instillation in reducing VILI may be predicted by the shape of the pressure-volume curve. These findings may help in designing safer clinical studies of mechanical ventilation and in reducing the cost of partial liquid ventilation by reducing doses of PFC.

[Experimental changes in the alveolo-capillary barrier induced by artificial ventilation]. / Altérations expérimentales de la barrière alvéolo-capillaire induites par la ventilation artificielle.

Mechanical ventilation can have deleterious effects on the lungs. Extra-alveolar escape of air, such as pneumothorax or subcutaneous emphysema, are complications which have long been known. New experimental studies have clearly shown that mechanical ventilation can also result in pathologic changes to the air/blood barrier. Mechanical ventilation with high end-inspiratory pressure and high volume causes lung edema in whose origin abnormal permeability of the alveolo-capillary barrier plays a major role. The abnormalities are in fact the result of pulmonary distention and not of elevated air pressure; this justifies the term "volume traumatism". The presence of a previous acute pulmonary injury considerably increases the deleterious effects of mechanical ventilation on the lungs. Although the clinical implications of these experimental studies are difficult to assess, they have nevertheless resulted in major changes in ventilation strategy for acute lung diseases such as the acute respiratory distress syndrome of the adult.

Pulmonary distribution of iodoantipyrine: temperature and lipid solubility effects.

In multiple indicator-dilution studies in rat and dog lungs, we have found that the distribution of iodoantipyrine (IAP) is not limited by the endothelium at a temperature > 7 degrees C but is barrier limited at the epithelium at a temperature < 15 degrees C (permeability coefficient of 6.3 x 10(-5) cm/s at 8 degrees C). IAP extraction from the vascular surface to the tissues is greater than those of antipyrine (AP) and tritiated water (THO). IAP transmittance from the alveolar surface to the vascular compartment is smaller than those of AP and THO: a lung lipid compartment, probably in the lamellar bodies of the type II cells, is more accessible to IAP than to AP or THO because IAP has a higher oil-to-water distribution coefficient. Our mathematical model takes into account these matters and also the low surface density of the type II cells: some of the IAP may bypass the lipid compartment. Lipid may affect the transit of solutes with high oil-to-water distribution coefficients in the lungs and across the alveolar-capillary barrier.

Comparison of the effects of heat and moisture exchangers and heated humidifiers on ventilation and gas exchange during weaning trials from mechanical ventilation.

Heat and moisture exchangers (HME) are increasingly used to warm and humidify inspired gases in intubated ventilated patients. But these devices add dead space that may alter the alveolar ventilation. This could impair the efficiency of spontaneous ventilation (SV) during weaning trials from mechanical ventilation. Fifteen patients were tested with an HME (Hygrobac-DAR) and a heated humidifier (HH) (Fischer-Paykel MR 450) in a random order during weaning trials in SV with inspiratory pressure support. Minute ventilation VE, tidal volume), and respiratory rate were recorded and arterial blood was sampled for blood gas analysis with each device. The HME gave a significantly greater VE than the HH (9.3 +/- 0.8 L/min vs 8.1 +/- 0.8 L/min; p < 0.005), because of increased respiratory rate (21 +/- 2/min vs 19 +/- 2/min; p < 0.05). Tidal volume was unchanged for HME and HH (470 +/- 32 mL vs 458 +/- 39 mL). The higher PaCO2 with HME than with HH (44 +/- 2 mm Hg vs 42 +/- 2 mm Hg; p < 0.005) revealed an insufficient alveolar ventilation response to the increase in dead space. Arterial Po2 rose with the HME, but not significantly above the HH values (103 +/- 6 mm Hg vs 97 +/- 6 mm Hg; p = 0.055), possibly because of a positive end-expiratory pressure effect of the HME. The need to increase VE in SV when an HME is used should be taken into account during difficult weaning from mechanical ventilation.

Glucose transport and equilibrium across alveolar-airway barrier of rat.

The glucose concentration in the epithelial lining fluid (ELF) results from a balance between cellular uptake and paracellular leakage. The present study examines whether the ELF glucose concentration can be predicted from the kinetics of glucose transport obtained in fluid-filled lungs. Isolated rat lungs were filled via the trachea with instillate containing 0-10 mM glucose; the perfusate glucose concentration was 10 mM. The rate of glucose removal from airspaces depended on luminal glucose concentration and was saturable [maximum uptake rate = 101 +/- 8.6 mumol.h-1.g dry lung wt-1; apparent Michaelis constant K(m) = 1.5 +/- 0.43 mM; R2 = 0.79]. Glucose removal was inhibited by phloridzin but not by phloretin or by inhibiting glycolysis. The steady-state concentration in fluid-filled lungs was estimated to be 0.15 +/- 0.034 mM. It agreed with that (< 1/20 plasma) calculated using glucose transport kinetics and paracellular permeability. The ELF glucose concentration obtained by bronchoalveolar lavage was 0.39 +/- 0.012 plasma in vivo and 0.39 +/- 0.021 perfusate in air-filled isolated lungs. The equilibrium ELF/perfusate distribution ratio of alpha-methyl-glucose was similar to that of glucose. Thus there is a major difference between the alveolar steady-state glucose concentration in air- and fluid-filled lungs despite similar mechanisms of airspace glucose removal. This suggests that glucose kinetics or access to uptake sites differ in air- and fluid-filled lungs.