Thorax 1986;41:176-189 Pathogenesis of pleurisy, pleural fibrosis, and mesothelial proliferation AMANDA HERBERT From the Department of Histopathology, Southampton General Hospital, Southampton Systemic and pulmonary diseases that affect the pleura are usually characterised by accumulation of fluid in the pleural cavity or by fibrous healing of damaged mesothelium. Some of the reactive changes in the mesothelial cells and fibroblasts concerned in these processes may closely mimic neoplasia and must be distinguished from metastatic carcinoma and malignant mesothelioma. Before we consider the pathogenesis of these conditions it is necessary to understand the development and structure of pleura, the unique mechanism for regeneration of meso- thelium, and the factors responsible for breakdown of this mechanism and for the resulting fibrous repair. Development and structure of the pleura The pleura and other serous cavities develop from the extraembryonic coelom, which appears in the blasto- cyst as early as the second week of embryonic life. The parietal pleura is derived from the somatopleura, which also covers the amnion and lines the tro- phoblast; whereas the visceral pleura is derived from the splanchnopleura, which also surrounds the yolk sac.1 The pleural connective tissue and its highly spe- cialised mesothelial lining are thus derived entirely from mesoderm: parietal and visceral pleurae develop separately in the early embryo, preserving certain structural and functional differences in the adult. The anatomical layers of the pleura are shown dia- gramatically in the figure. The mesothelium and a thin layer of submesothelial connective tissue cover a well developed network of fibres that form the exter- nal elastic lamina. This is separated from the internal elastic lamina by the interstitial layer, which contains lymphatics and blood vessels and is continuous with the interlobular connective tissue. The internal elastic lamina is present in both parietal and visceral pleura, although in the latter it is indistinguisable from the elastic of the peripheral alveoli.2 Address for reprint requests: Dr Amanda Herbert, Department of Histopathology, Southampton General Hospital, Southampton S09 4XY. PLEURAL LYMPHATICS The lymphatic flow in the adult lung is directed by valves.2 3 Subpleural lymphatic vessels, situated in the deep aspect of the interstitial layer, drain along the surface of the lung, as well as through intra- pulmonary lymphatic vessels, to the hilar lymph nodes.2 Intralobular pulmonary lymphatics drain outwards to the subpleural network.3 The anterior parietal and diaphragmatic lymphatics drain to the internal mammary chain. The posterior parietal and diaphragmatic lymphatics drain mainly to the inter- costal and paravertebral nodes, but also through the diaphragm to the retroperitoneal nodes.2 The controversy about the existence of stomata between the pleural space and lymphatics has been resolved largely by scanning electron microscopy and is reviewed by Whitaker etal.4 The existence of sub- diaphragmatic stomata, as described by Allen in 1936,5 has been confirmed.6 Stomata have also been shown to be present in the parietal pleura of the cau- dal and ventral mediastinum and the lower part of the costal pleura, in both experimental animals and man.7 The distribution of the stomata is similar to that of the aggregates of macrophages and specialised mesothelial cells refered to as "Kampmeier foci"8 9: these structures are not found in the visceral pleura.7 Large particles and cells pass through the stomata,7 while protein is absorbed exclusively through the pleural lymphatics.'0 PLEURAL BLOOD SUPPLY There has been disagreement about the origin of the blood vessels in the interstitial layer of the pleura. Although von Hayekt2 stated that the pulmonary arteries supplied the visceral pleura, others have shown that in man the pleura is supplied by branches of the bronchial arteries.2 3 According to Miller, the subpleural veins drain to the pulmonary vein, whereas Nagaishi has shown that some drain to the pulmonary veins and others via the extrapulmonary bronchial vein to the right atrium.2 3 THE MESOTHELIUM The mesothelial cells form a complete layer of cells 176 on June 17, 2020 by guest. Protected by copyright. http://thorax.bmj.com/ Thorax: first published as 10.1136/thx.41.3.176 on 1 March 1986. Downloaded from
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Thorax 1986;41:176-189
Pathogenesis of pleurisy, pleural fibrosis, andmesothelial proliferationAMANDA HERBERT
From the Department of Histopathology, Southampton General Hospital, Southampton
Systemic and pulmonary diseases that affect thepleura are usually characterised by accumulation offluid in the pleural cavity or by fibrous healing ofdamaged mesothelium. Some of the reactive changesin the mesothelial cells and fibroblasts concerned inthese processes may closely mimic neoplasia and mustbe distinguished from metastatic carcinoma andmalignant mesothelioma. Before we consider thepathogenesis of these conditions it is necessary tounderstand the development and structure of pleura,the unique mechanism for regeneration of meso-thelium, and the factors responsible for breakdown ofthis mechanism and for the resulting fibrous repair.
Development and structure of the pleura
The pleura and other serous cavities develop from theextraembryonic coelom, which appears in the blasto-cyst as early as the second week ofembryonic life. Theparietal pleura is derived from the somatopleura,which also covers the amnion and lines the tro-phoblast; whereas the visceral pleura is derived fromthe splanchnopleura, which also surrounds the yolksac.1 The pleural connective tissue and its highly spe-cialised mesothelial lining are thus derived entirelyfrom mesoderm: parietal and visceral pleurae developseparately in the early embryo, preserving certainstructural and functional differences in the adult.The anatomical layers of the pleura are shown dia-
gramatically in the figure. The mesothelium and athin layer of submesothelial connective tissue cover awell developed network of fibres that form the exter-nal elastic lamina. This is separated from the internalelastic lamina by the interstitial layer, which containslymphatics and blood vessels and is continuous withthe interlobular connective tissue. The internal elasticlamina is present in both parietal and visceral pleura,although in the latter it is indistinguisable from theelastic of the peripheral alveoli.2
Address for reprint requests: Dr Amanda Herbert, Department ofHistopathology, Southampton General Hospital, SouthamptonS09 4XY.
PLEURAL LYMPHATICSThe lymphatic flow in the adult lung is directed byvalves.2 3 Subpleural lymphatic vessels, situated in thedeep aspect of the interstitial layer, drain along thesurface of the lung, as well as through intra-pulmonary lymphatic vessels, to the hilar lymphnodes.2 Intralobular pulmonary lymphatics drainoutwards to the subpleural network.3 The anteriorparietal and diaphragmatic lymphatics drain to theinternal mammary chain. The posterior parietal anddiaphragmatic lymphatics drain mainly to the inter-costal and paravertebral nodes, but also through thediaphragm to the retroperitoneal nodes.2The controversy about the existence of stomata
between the pleural space and lymphatics has beenresolved largely by scanning electron microscopy andis reviewed by Whitaker etal.4 The existence of sub-diaphragmatic stomata, as described by Allen in1936,5 has been confirmed.6 Stomata have also beenshown to be present in the parietal pleura of the cau-dal and ventral mediastinum and the lower part of thecostal pleura, in both experimental animals andman.7 The distribution of the stomata is similar tothat of the aggregates of macrophages and specialisedmesothelial cells refered to as "Kampmeier foci"8 9:these structures are not found in the visceral pleura.7Large particles and cells pass through the stomata,7while protein is absorbed exclusively through thepleural lymphatics.'0
PLEURAL BLOOD SUPPLYThere has been disagreement about the origin of theblood vessels in the interstitial layer of the pleura.Although von Hayekt2 stated that the pulmonaryarteries supplied the visceral pleura, others haveshown that in man the pleura is supplied by branchesof the bronchial arteries.2 3 According to Miller, thesubpleural veins drain to the pulmonary vein,whereas Nagaishi has shown that some drain to thepulmonary veins and others via the extrapulmonarybronchial vein to the right atrium.2 3
THE MESOTHELIUMThe mesothelial cells form a complete layer of cells
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connected by overlapping cytoplasmic processes withvariable tight and gap junctions, many of whichappear to be in a state of formation and dis-mantlement.13 The cells are flattened, with the cyto-plasm raised over a central nucleus. Surface microvilliare more numerous on visceral than parietal meso-thelial cells, possibly because of the greater absorptivepotential of the visceral pleura.'4 The micrcovilli areclosely associated with pinocytotic vesicles and vacu-oles that communicate with the luminal surface, inter-cellular channels, and the basal lamina.4 15 Experi-mental studies, reviewed by Whitaker eta,4 includingcytochemical analysis of membrane associatedenzymes, provide evidence that mesothelial cells are
engaged in active transport rather than passivediffusion, supporting observations made many years
ago by Starling and Tubby.'6 Fluid and small par-
ticles are transported through pinocytotic vesicles or
intercellular channels,'5 depending on size.4 Themicrovilli have a cell coat or glycocalyx, with a strongaffinity for acid mucopolysaccharide; and it has beensuggested that the slippery villous surface protects thepleura from frictional damage.'7 The role of themicrovilli and glycocalyx in lubricating the pleuralcavity is, however, uncertain and recent research hasindicated that the pleura may be lubricated by phos-pholipid surfactants, producing a dry, hydrophobicsurface analogous to an empty polythene bag.'8 19
Mesothelial cells in culture synthesise hyaluronicacid, but in far smaller quantities than do fibroblasts,possibly reflecting their divergent differentiation at anearly stage of embryogensis.20 In culture they alsoproduce small or even large amounts of collagen.20 21
There is evidence that mesothelial cells are capableof phagocytosis.22 Although they can usually be dis-tinguished from serosal macrophages, which are are
derived from the bone marrow,23 intermediate formsare sometimes seen,24 25 raising the possibility thatmesothelial cells in effusions may develop into facul-tative macrophages. Furthermore, mesothelial cellsmay contain non-specific esterase, acid phosphatase,alpha naphthol acetate esterase, cxl antitrypsin and a,antichymotrypsin, all of which are present in macro-
phages but in greater amounts.25-28
Injury and repair
INJURY TO MESOTHELIAL CELLSMesothelial cells are highly susceptible to damagefrom agents that they do not normally encounter,such as air,2930 water,31 asbestos,22 foreign pro-
tein,32 silica,33 and even saline.30 All these cause
swelling of the cells, clubbing of microvilli, and sepa-ration of the cells from each other and from the basallamina, resulting in exfoliation. The denuded surface
Interstitiallayercontinuouswithinterlobularsepta
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then shows an exudative inflammatory reaction, fre-quently accompanied by an effusion.
REGENERATION OF MESOTHELIUMThe exact mechanism of mesothelial regeneration iscontroversial, although there has been general agree-ment with Hertzler's observation that "the entire sur-face becomes endothelialised simultaneously, and notgradually, from the border, as in epidermitisation ofskin wounds.34 This process begins within 24 hoursby the appearance of macrophages on the wound sur-face and is generally complete by 8-10 days.4 Themain controversy has revolved around the origin ofthe new mesothelial cells. Initial observations indi-cated that they may be derived from submesothelialfibroblasts35 36; but Johnson and Whitting37 pro-posed that serosal macrophages might differentiateinto mesothelial cells, a theory supported by light andtransmission electron microscopic studies38 39 andlater by scanning electron microscopy.40 Recentexperiments, combining several investigative tech-niques, have indicated that mesothelial cells adjacentto the wound and on the opposing serous surfaceexfoliate, proliferate, and repopulate the area,replacing the macrophages that initially seal thewound.4' This would confirm that mesothelial regen-eration can occur without the participation of theunderlying connective tissue,40 and suggests thatthere is not transformation of either macrophages orfibroblasts into mesothelial cells. Many years agoCameron eta!42 showed that detached mesothelialcells proliferate to repair peritoneal defects and,although it was subsequently accepted that this mech-anism may play a part in mesothelial repair,3740 itwas regarded as minor and relatively unim-portant.43
FIBROUS REPAIR OF SEROSAL SURFACESAlthough extensive areas of bare submesothelial con-nective tissue can be replaced by mesothelium, evenminor surgical procedures sometimes give rise todense peritoneal adhesions.35 Similarly, fibrous oblit-eration of the pleura may follow both pleurisy andeffusion-although, on the other hand, large amountsof fluid and even blood may be reabsorbed withoutany resultant fibrosis.45 There has been extensiveresearch into the pathogenesis of fibrous adhesionsand the important factors in their development arethe presence of blood,46 extensive and persistentcrushing or abrasion of the mesothelium,47 the pres-ence of foreign material,48 and local ischaemia.49 Thecritical factor appears to be the presence of a fibrinousexudate and whether or not this is absorbed.47 49 Thisis closely related to the fibrinolytic power of serosalsurfaces, which was recognised for many years beforeit was shown to be a specific property of mesothelial
Herbertcells.50 - 53 Newly regenerated mesothelium hasgreatly enhanced fibrinolytic activity, but fibrinolyticactivity is depressed by damage to mesothelial cellsand by dilution with serosal fluid.30 52 Persistentdepression of fibrinolysis is associated with the devel-opment of fibrosis.47
Despite their common origin from mesoderm,mesothelial cells, macrophages, and fibroblastsappear to be separate specialised populations of cells,which, although they overlap in their functional char-acteristics, play different parts in the response toinjury. The mesothelium has great power of regen-eration, but when this breaks down the pleural spacebecomes obliterated by fibrous connective tissue.
Manifestations of benign pleural disease
PLEURAL EFFUSIONPleural effusions occur in human disease when alter-ations in the hydrostatic and colloid osmotic pres-sures result in transudation, or when alterations incapillary permeability associated with the inflam-matory response result in exudation of protein richfluid. Once an effusion has formed, whatever itscause, it is in a dynamic state, with a turnover rate of30-75% per hour.54 Protein is removed exclusively bythe lymphatics, and may be blocked by ligating thethoracic duct.-l Lymphatic flow has been measuredin man as an average of 0.37 ml/kg per hour duringthe day, falling to 0.2 ml/kg per hour during the night,possibly because of the reduction in movement of thediaphragm and intercostal muscles. 1' The removal ofpara-aminohippurate (a relatively small molecule)through the pleural capillaries was estimated to be4.5 mg/kg per hour, 13 times higher than the rate oflymphatic drainage and roughly correlating with ablood flow of 300 ml/h in an average adult."1
It has been accepted that fluid is secreted by theparietal pleura and reabsorbed by the visceral pleura,largely because of visceral capillary pressure has beenthought to be that of the pulmonary circula-tion'9 55 56_although recently some doubt has beencast on this.57 58 In 1957 Agostoni et al demonstrateda visceral pleural absorptive force of 15mm Hg inthoracotomised dogs.59 Later Black,60 using Ago-stoni's work as a basis, calculated that the parietalpleural secretory pressure is 6cm H20, and the vis-ceral pleural resorptive force 13 cm H20. Pulmonaryand systemic capillary pressure and the osmotic pres-sure of pleural fluid and plasma were taken intoaccount. It was assumed that the visceral pleural cap-illary pressure was the same as the pulmonary arterialpressure.
Black's calculations result in a net pressure of 7 cmH20 favouring absorption, which would be evengreater in the absence of fluid in the pleural space.
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Pathogenesis ofpleurisy, pleuralfibrosis, and mesothelial proliferation
The presence of fluid in the normal pleural space hasbeen doubted'9 and is difficult to explain if Black'sfigures are accepted. These calculations have recentlybeen recognised as an oversimplification,57 58 andthey are not consistent with the anatomy of thehuman lung as described by Miller and Nagaishi, whoshowed that although subpleural veins drain, at leastin part, into the pulmonary vein, the visceral pleuralarteries in man are branches of the bronchial arte-ries.' 3 The net hydrostatic pressure in the visceralpleural capillaries must therefore be higher than in thepulmonary circulation but lower than in the systemiccirculation. A modification of the theory is necessaryto explain how effusions can form with relatively lowprotein concentrations, and why pleural effusion doesnot result in pulmonary oedema." The classical the-ory also tends to overlook the role of the mesotheliumin active transport of fluid and the anatomical sepa-ration of the visceral and parietal capillaries by meso-
thelium, submesothelial connective tissue, and theexternal elastic lamina. Nevertheless, the visceralpleura would appear to have greater absorptivepotential than the parietal pleura and probably a con-
siderably lower hydrostatic capillary pressure, whichwould tend to limit the development of effusions.
TransudatesTransudates occur when the plasma colloid osmoticpressure is reduced, as in hypoalbuminaemia, or whenthe systemic or pulmonary venous pressure isincreased; an additional factor is reduced renal excre-tion of sodium, which occurs in both cirrhosis andcongestive cardiac failure. Transudates are not usu-ally associated with primary pathological conditionsof the pleura. The protein concentration is less than3.0g/100ml, corresponding to a specific gravity of1.016, and the cell count is low. When cells arepresent, typically they are degenerate mesothelial cellsand macrophages with signet ring forms.6' Poly-morphs are absent but lymphocytes may be present,which may be explained by reduced lymphatic flowthrough the thoracic duct when there is systemicvenous hypertension.55Congestive cardiacfailure In man a combination ofsystemic and pulmonary venous hypertension is morelikely to cause pleural effusion than is systemic venoushypertension alone. It is therefore common in thecongestive cardiac failure of heart disease but rare inright ventricular failure due to chronic lung disease.55Effusions in congestive cardiac failure may containmany proliferating mesothelial cells in addition todegenerating forms with vacuolated cytoplasm.6' 63
This would imply damage to the mesothelium, andSpriggs and Boddington suggest that the presence ofan appreciable number of mesothelial cells indicates apulmonary complication such as infarction.6'
Nephrotic syndrome This is a classic cause of aneffusion, which is invariably a transudate, and iscaused by severe reduction in plasma colloid osmoticpressure. Pleural effusion develops at a stage whenoedema and ascites are already present,56 indicatingthat the resorptive potential of the pleura is reduced.Cirrhosis Pleural effusion in cirrhosis is usuallyassociated with ascites64 but may occur in itsabsence.6566 Ten per cent of patients with cirrhosisdevelop a pleural effusion,64 most frequently on theright side. Studies with radiolabelled albumin haveshown that fluid is transferred directly across thediaphragm, presumably in response to the negativeintrapleural pressure.6467 Lieberman et al showeddiaphragmatic defects in some of the patients in theirstudy, and suggested that these rather than the trans-diaphragmatic lymphatics are the route of the fluid;this is supported by recent reports.6566 Althougheffusions are typically transudates, occasionally pro-liferating and often atypical mesothelial cells arepresent in large numbers.6263 These cells may appearvery similar to malignant cells, and a cytogeneticstudy of ascitic fluid from patients with alcoholic cir-rhosis has shown karyotypic abnormalities indistin-guishable from malignancy.68 The authors suggestedthat a mutagenic effect of alcohol might be impli-cated, but abnormal karyotypes have been demon-strated on rare occasions in mesothelial cells fromother types of reactive effusions.69
ExudatesExudative pleural effusions form when an acuteinflammatory response causes increased permeabilityof pleural capillaries to protein and cells. The meso-thelial cells round up and separate, allowing the pas-sage of cells and protein into the pleural space. Bothleucocytes70 and fibrin32 can be shown to passbetween cells in experimental peritonitis. A furtherfactor, which is often not emphasised, is the obstruc-tion of lymphatic drainage by hilar lymph-adenopathy. Exudates may or may not be purulent;the latter type includes eosinophilic and lymphocyticexudates.
NON-PURULENTEXUDATES Non-purulent exudatescontain neutrophils, lymphocytes, macrophages,exfoliated mesothelial cells, eosinophils, andbasophils, roughly in that order of frequency. Thepercentage of the cell types varies. Lymphocytes oreosinophils may predominate, for example, in thelater stages of a pneumonic effusion.6'Pneumonia The incidence of effusion in pneumoniavaries with the type of organism and is reviewed indetail by Light.57 It is much higher with f haemolyticstreptococcus (90%) than pneumococcus (40-60%),and occurs in about 20% of cases of viral pneumonia.
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Organisms are grown from the fluid in less than halfsuch effusions, with the exception of staphylococcusand Haemophilus influenzae in children andEscherichia coli and anaerobic organisms in adults.Anaerobic lung infections frequently cause effusionsand are often associated with alcoholism and factorspredisposing to inhalation. The cell counts are veryhigh, usually over 10 x 106/1, 90% being neutrophils.Exfoliated mesothelial cells are present but are notprominent, possibly reflecting the lack of cell damageand the frequency of complete resolution, whichoccurs in about 90% of cases. Large, particularlyblood stained, effusions may fail to resolve, and ifthey are not drained lead to fibrous pleurisy andrespiratory impairment.56Pulmonary infarction An assessment of theincidence of pleural effusion in pulmonarythromboembolism is complicated by the fact thatunrelated causes of effusion may also be present.Bynum and Wilson reported that 40% of patients hadeffusions directly attributable to thromboembolismwhen other causes had been excluded, and that onlyhalf of these had radiological evidence of pulmonaryinfarction. Two thirds of those with radiologicalevidence of infarction had effusions: these were larger,more often blood stained, and more likely topersist.7" The fluid is blood stained in up to 65% ofcases of effusion related to thromboembolism,72suggesting that not all infarcts are radiologicallyvisible. Effusions in the absence of infarction may beattributed to transient pulmonary ischaemia,atelectasis, or right sided heart failure. The cell counttends to be high, with a composition that varies withthe time from the onset of the disease.73 There may bea high percentage of neutrophils, in which case thepicture is similar to that of a pneumonic effusion. Oneimportant difference is the frequent presence ofexfoliated mesothelial cells in large numbers, whichmay be atypical and closely mimic malignantcells,63 73 probably reflecting ischaemic damage.Most of these effusions resolve, but those associatedwith radiological evidence of infarction may persistand give rise to localised pleural fibrosis.7'Malignancy Malignant infiltration is a commoncause of pleural effusion, and the histological andcytological examination of pleural biopsy materialand fluid is largely directed towards the identificationof malignant cells. Not all such effusions containmalignant cells, and they may show the features of anon-purulent inflammatory exudate, a lymphocyticeffusion, or, very occasionally, a transudate.6Effusions in which malignant cells cannot beidentified may be caused by concomitant pneumonia,congestive heart failure, and hypoproteinaemia,74 orby the obstructive pneumonitis or bronchiectasis oflung cancer. This is relevant to whether a patient with
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lung cancer meets the criteria for operability, sinceeven blood stained effusions may be produced byinflammation rather than malignant infiltration.75There is also evidence that effusion in lung cancer,76malignant lymphoma,77 and Hodgkin's disease78 ismore likely to be caused by disease of mediastinallymph nodes than by pleural infiltration by tumour,and this may also be a factor in the development ofeffusions in other forms of metastic cancer,79particularly breast cancer. Goldsmith et al reportedthat effusions in breast cancer were present in 63% ofpatients with pulmonary lymphangitic spread, butalso in 40% of patients without such metastases.Particularly in the latter group, the effusion was moreoften on the same side as the primary tumour.80Seventy five per cent of malignant effusions arecaused by lung cancer, breast cancer, or malignantlymphoma8" and in all three cases the effusions are atsites that share lymphatic drainage with the pleura.Cytological examination gives positive results in50-60% of clinically malignant effusions. Thisproportion is increased by 10-20% by the use ofcytogenetic investigations69 or immunocytochemicalmethods.82 The remaining patients may well not havedirect infiltration of the pleura.
Eosinophilic effusionAn eosinophilic effusion is defined as one in whichthere are more than 10% of eosinophils and it mayoccur in pneumonic effusion, particularly those asso-ciated with viral and pneumococcal infection; in thelatter it tends to develop two to three weeks after theonset of the disease.83 It is said to be rare in tuber-culosis and malignancy, with the exception of Hodg-kin's disease.57 A high percentage of eosinophils hasbeen reported in collagen disease,83 and is a particu-lar feature of the effusions that occur as a response topneumothorax. Surgical specimens from patientshaving a pleurectomy for spontaneous pneumothoraxshow appreciable swelling and activation of the meso-thelial cells, with an infiltrate of eosinophils andhistiocytic giant cells. The picture may bear a
superficial resemblance to the infiltrate of eosinophilicgranuloma (histiocytosis X), and probably representsa direct response of mesothelial cells to the irritanteffect of air.85 Other causes include hypersensitivityto drugs and parasitic infections.57
Lymphocytic effusionA lymphocytic effusion is usually defined as anexudative effusion containing more than 50% of lym-phocytes, but much higher percentages are seen insome effusions, particularly in tuberculosis and malig-nancy. Lymphocytes from malignant effusions cul-tured with phytohaemagglutinin undergo blast
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Pathogenesis ofpleurisy, pleuralfibrosis, and mesothelial proliferation
transformation86 and form rosettes with sheep eryth-rocytes.87 The ratio of T to B lymphocytes is greaterin pleural fluid than peripheral blood, suggestingmigration ofT lymphocytes into effusions in the pres-ence of cancer.87 A recent study using monoclonalantibodies to T cell subsets has shown that thesechanges occur in all types of lymphocytic effusions,not only those associated with cancer.88 The predom-inant subset in all types of effusion was helper-inducer(OKT4 positive). Tuberculous and malignanteffusions also showed a relative increase in the Erosetting, OKT3 negative subset recently described asnatural killer cells.89 Lymphocytes in reactiveeffusions may be so numerous that they mimic malig-nant lymphoma or chronic lymphatic leukaemia.6'Since the latter are most commonly B cell lines, anincrease in the ratio of B to T cells in the fluid may beuseful in differentiating malignant from benign lym-phoid cells.90Tuberculosis Tuberculous pleurisy remains one ofthe major non-malignant causes of a pleural effusion.Classically, it occurs as a complication of primaryinfection, and is regarded as an expression of delayedhypersensitivity to bacilli entering the pleural space.The experimental work of Paterson is accepted as themodel for the development of tuberculous effusions inman.91 Tubercle bacilli introduced into the pleura oftuberculin sensitised guinea pigs induce a vigorouspleurisy with effusion, whereas tuberculin negativecontrols do not have effusions but develop dissem-inated tuberculosis. This experimental model cor-relates well with clinical tuberculous pleurisy, as wasshown in- a clinical and pathological study by Steadetal, in which a focal pulmonary tuberculosis lesionwas found in direct continuity with the pleura in 13out of 15 men undergoing thoracotomy for this con-dition.92 The pleura showed extensive replacement bya layer of tuberculous granulation tissue, which wasdecorticated. Effusions occur less frequently in post-primary tuberculosis, but the pathology is similar.
Despite the extensive and severe pleural reaction,most tuberculosis effusions resolve spontaneouslyeven in the absence of treatment, although 65% of thepatients who recover initially develop pulmonary orextrapulmonary tuberculosis within the following fiveyears.93 Although tuberculous pleurisy presents as anacute illness in young people, the onset is moreinsidious in older patients94; and now that primarytuberculosis occurs more frequently in an older agegroup a tuberculous effusion is more likely to be con-fused with other forms.95At the very earliest stage there are neutrophils in
the fluid, but usually there is a high proportion oflymphocytes. Mesothelial cells are characteristicallyabsent, which may suggest the diagnosis.96 Abram'sneedle biopsy yields 70-80% positive results in
tuberculosis95 `9 because pleural disease is usuallywidespread. The lack of mesothelial cells can bepartly explained by destruction of mesothelium, butthis is not entirely consistent with the clinical resolu-tion that often occurs. Spriggs and Boddington96quote an early, unsubstantiated histological study bySaltykow,"'0 in which an intact mesothelium wasseen under a fibrinous exudate; and this may explainthe lack of mesothelial cells in the pleural fluid.83 Oth-ers have shown that mesothelial cells may be presentin tuberculous effusions, particularly where there isactive disease of the pleural surface.62 63 Sometimesthey show atypical proliferative changes. Possibly thehilar lymph node disease, which is prominent in pri-mary but far less so in postprimary tuberculosis,10' isimportant either as an additional source of bacillientering the pleural space'02 or as a cause of obstruc-tion to lymphatic drainage. A reduced clearance ofprotein has been found in both experimental103 andclinical tuberculous pleurisy.'04Yellow nail syndrome Lymphocytic effusion is a fea-ture of the yellow nail syndrome, in which there ishypoplasia of lymphatics.'05 It must be distinguishedfrom chylothorax, which occurs when the thoracicduct ruptures or is obstructed.'06 56 Chyle is rich inlymphocytes, but also contains triglycerides in agreater quantitity than is ever found in the blood. Themost common causes are trauma, thoracic oper-ations, and malignant lymphoma. It is also a featureof lymphangioleiomatosis. 107
PURULENT EFFUSION In empyema fluid there arelarge numbers of dead and dying neutrophils, withmany smeared nuclei. The appearance is quitedifferent from that of a non-purulent exudate, andsignifies bacterial infection of the pleural membraneitself, which can be demonstrated by culture of thefluid. Tuberculous empyema produces a very similarpicture.6 Mesothelial cells are seldom observedexcept in the very early stages. The widespreaddestruction of the mesothelial lining accounts for thefibrous scarring that invariably accompanies thiscondition.
Other causes ofeffusionOther causes of effusion are reviewed extensivelyelsewhere,55 57 and will not be described in detail.Effusions occur in rheumatoid disease, 108 systemiclupus erythematosus,'09 myxoedema,10 uraemia,"'postmyocardial infarction syndrome, 1 12sarcoidosis,'13 mediastinal irradiation,"4 andasbestos exposure. Peritoneal effusions of many typesmay be associated with pleural effusions, includingpancreatitis,l1 subphrenic abcess,l16 peritonealdialysis,"7 viral hepatitis,"18 and the Meigs-Salmonsyndrome. "19
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A variable proportion of cases of pleural effusionremains unexplained. In the past idiopathic effusions,particularly in young people, would have been con-sidered tuberculous unless proved otherwise.93Recently there has been emphasis on asbestosexposure as a possible cause,120 121 but an extensivesurvey of patients with pleural thickening on radio-graphs failed to show a significant association withasbestos exposure.122 A proportion of patients withunexplained pleural effusions subsequently developmalignant lymphoma, mesothelioma, or car-cinoma. 123
PLEURAL FIBROSISThe mechanisms concerned in mesothelial injury andrepair and the circumstances in which pleural effusionmay undergo fibrous organisation are important inthe pathogenesis of pleural fibrosis. This may takeseveral forms, which may or may not be associatedwith mesothelial regeneration.
Pleural adhesionsLocalised damage to the pleura, from whatever cause,may result in minor degrees of pleural fibrosis oradhesions between the lung and the chest wall, whichbecome relined by mesothelium derived from sur-rounding mesothelial cells.41 Pleural adhesions haveless clinical importance than those that form in theperitoneum, but they may result in the developmentof abnormal communications between the pulmonaryand chest wall vasculature and lymphatic drainage.These may play a part in the spread of disease and thepathogenesis of air embolism and metastatic cerebralabscess.45
Diffuse fibrosing pleurisyAt its most extensive, fibrosing pleurisy totally oblit-erates the pleural cavity, with limitation of chestmovement and consequent respiratory disability. Itmay follow any prolonged exudative or blood stainedeffusion, but is particularly liable to occur in tuber-culous pleurisy, empyema, and asbestos exposure.Rarely, it is associated with rheumatoid effusions,124uraemia, 1 1 1 pancreatitis, 1 15 or traumatic hae-mothorax.45Whatever the cause of the fibrosis, the pleural
lining is replaced by a layer of dense collagenous tis-sue, which may be several centimetres thick andextends from the interstitial layer deep to the externalelastic lamina, often affecting the interlobular septa.It can be stripped easily from the lung (unless there isunderlying pulmonary fibrosis) and from thediaphragm, but with much more difficulty from thechest wall, where it may infiltrate the intercostalmuscle. The mediastinal pleura is usually not affectedin reactive pleurisy, in contrast to malignant meso-
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thelioma.45 The superficial layer is composed oforganising purulent exudate, tuberculous granulationtissue, rheumatoid nodules, or fibrinosanguinousexudate, depending on the aetiology. Histologically,benign fibrous pleurisy may be difficult to distinguishfrom mesenchymal malignant mesothelioma. Theintact elastic lamina may be helpful,125 and the orien-tation of the granulation tissue is less haphazard thanin malignant mesothelioma. 126 Fibrous pleurisyappears to be entirely mesenchymal, whereas thefibroblastic component of mesothelioma often looksmore epithelial and may include cells that express epi-thelial cell markers.127 And although mesothelialcells may form clefts in fibrous pleurisy,'28 they arenot prominent,92 126 presumably because they havebeen largely destroyed in the original inflammatoryprocess.
Benign asbestos pleural effusion andfibrosisPleural fibrosis is a feature of pulmonary asbes-tosis, l29 130 but has only recently been recognised as acause of restrictive lung disease in the absence ofappreciable pulmonary fibrosis.131 132 Diffusefibrosis may follow acute asbestos pleurisy witheffusion,128 and is probably caused by direct damageto the mesothelium. Asbestos is known to have adirect cytotoxic effect on mesothelial cells,22 but themechanism whereby the fibres reach the pleural spaceis unknown. The association of asbestos exposurewith benign pleural effusion was first reported byEisenstadt'33 and further cases were described byGaensler and Kaplan' 34 and Mattson, who suggestedthat asbestos is a frequent cause of apparently idio-pathic effusions.'20 Such recurrent and often blood-stained effusions frequently lead to fibrosis. Althoughthis condition is relatively benign (in one seriespatients were followed for up to nine years), deco-rtication may be required and a considerable numberof patients later develop mesothelioma or lungcancer. 135
Pleural plaquesHyaline pleural plaques must be distinguished frompleural fibrosis, and they probably arise by a com-pletely different mechanism. They consist of raised,sharply defined, serpiginous, shiny yellow whitelesions, which are not associated with adhesions andcan be readily stripped from the chest wall. 136 Micro-scopially they are composed of avascular, poorly cel-lular collagen, which shows hyaline degeneration andoften calcification. They are most frequently found onthe posterior costal parietal pleura, where they tend tofollow the lines of the ribs, and the diaphragmaticpleura, where they are often firmly adherent to thecentral tendon.' 37 They also occur on the mediastinalpleura and cardiac fold but rarely on the visceral
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Pathogenesis ofpleurisy, pleural fibrosis, and mesothelial proliferationpleura. The incidence of hyaline plaques in routinenecropsies has been found to be 11-12% in urbancommunities'37 -139 and almost 40% in a miningcommunity in Finland.'40 In these series they werestrongly associated with an occupational and envi-ronmental exposure to all types of asbestos and withthe presence of asbestos bodies in lung tissue. Anal-ysis of pleural plaques has revealed fine asbestosfibres, less than 2 pm long, mainly in the calcified cen-tral zones. '' Some plaques occur in the apparentabsence of asbestos exposure,'42 143 but may berelated to fibres in the environment such as theamphibole asbestos tremolite. 130 and also non-asbestos minerals. Asbestos fibre counts of the lungsof patients with pleural plaques show a relative pre-dominance of commercial amphiboles (crocidoliteand amosite), contrasting with the predominance ofchrysotile fibres found in controls. 144 The presence ofplaques does not correlate with the degree of pul-monary asbestosis.130The sites of formation of pleural plaques coincide
with pathways of lympathic drainage of the pleuraand in particular with the Kampmeier foci, which arethe sites of uptake of particulate matter into the pari-etal pleural lymphatics. The pathogenesis of pleuralplaques is incompletely understood but Thompsonhas shown that they develop deep to an intact meso-thelium, which suggests that direct mesothelial dam-age does not play a part in their development.145 Thisis supported by the absence of surface adhesions146and of mesothelial hyperplasia.147 Fibres less than5 um are readily taken up by macrophages22 and arecarried in lymphatics to hilar and mediastinal lymphnodes. It has been postulated that they might alsoreach the parietal pleura via lymphatics'138; but thisseems unlikely as it would require retrograde flowagainst valves, and there is no reason for there to belymphatic obstruction. It seems more likely that thefibres are held up at the site of uptake into the lym-phatics; but how they reach the pleural space, in theabsence of pleural or pulmonary scarring, remainsunexplained-although Thompson has suggesteddirect penetration through the visceral pleura.145 Arecent experimental study has shown that asbestosfibres introduced directly into the pleural space inrabbits produced extensive plaques; but these did notform if the animals had previously been treated withnitrogen mustard, when a severe pleurisy with fibrosisdeveloped.This study suggests that the formation ofplaques depends on mobilisation of pleural macro-phages, which is prevented in the animals treated withnitrogen mustard as a secondary effect of inhibitingpolymorphs from entering the pleural space.148 Thisexperimental work may be relevant to clinical condi-tions in that the immunological abnormalities thathave been described in asbestosis'49 have not been
observed in people with pleural plaques.150
Shrinking pleuritis with rounded atelectasis (foldedlung)The association of a visceral pleural plaque or an areaof localised fibrosis with an underlying area ofatelectasis can give a highly characteristic radiologicalappearance of a rounded pleural based mass with cur-vilinear shadows extending towards the hilum,s5'which can simulate a peripheral tumour.152 Thislesion was first described by Blesovsky in threepatients, all of whom had some degree ofoccupational exposure to asbestos.'53 Thoracotomyrevealed an area of pleural fibrosis overlying theatelectatic segment, which readily re-expanded afterremoval of the plaque of fibrosis. Long term followup in a recent series has shown recurrence aftersurgery in one patient. 154 Subsequent casesconfirmed the association with pleural plaques,'51 155but not all patients had a history of asbestosexposure. 5 Dernevik and coworkers have reportedimmunological abnormalities in patients with thissyndrome who had been exposed to asbestos and alsoto quartz. 56 The visceral lesion has usually beendescribed as a glistening fibrous plaque, typically notadherent to the overlying parietal pleura.'5' Probablythe atelectasis results from the contraction of collagenthat occurs during the development of the plaque,157and it is unlikely to result from pleural effusion as thishas seldom been present in reported cases.'55 158 Thisview of the pathogenesis of folded lung is consistentwith the hypothesis that pleural plaques are sub-mesothelial structures, not associated with meso-thelial damage.
Localisedfibrous mesotheliomaThe term localised fibrous mesothelioma is commonlyused to describe a rare group of primary tumoursunrelated to asbestos exposure, which arise in the vis-ceral or parietal pleura, often as pedunculated masses.They are composed of immature mesenchymal spin-dle cells admixed with mature hyalinised collagen andsometimes areas resembling haemangiopericytomas.Large, thin walled blood vessels are often present.Although usually covered by normal mesothelium,they frequently contain clefts of bronchiolar epi-thelium or mesothelium towards the periphery.'59Associated systemic manifestations include fingerclubbing160 and hypoglycaemia. 16' Although mostare benign, a small proportion show histological evi-dence of malignancy and may recur; but they veryrarely metastasise.162 Most authors now hold theview that these tumours are derived from sub-mesothelial mesenchymal cells'25 159 163 and shouldbe clearly distinguished from true mesotheliomas, asoriginally proposed by Klemperer and Rabin.'64
183
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Some ultrastructural studies, however, favour amesothelial origin.165 - 167 This view was supportedby the belief that regenerating mesothelial cells werederived from submesothelial fibroblasts,4445 whichhas now been shown to be highly unlikely.4142
MESOTHELIAL PROLIFERATIONThe ability of mesothelial cells to proliferate andmimic primary or secondary neoplasia is a majorproblem in the interpretation of cytological prepara-tions and biopsy material. It can be understood farbetter on the basis of the functional changes that havealready been discussed with reference to mesothelialdamage, exfoliation, and regeneration. A distinctionmust be made between the changes seen in exfoliatedcells and those seen in the intact lining of the pleuralsurface.
Proliferation of exfoliated mesothelial cellsMesothelial cells are not evident in washing from nor-mal serous surfaces6' but may be abundant in manyforms of effusion. Their morphology changes whenthey exfoliate, and nuclei become active, often withone or more prominent nucleoli. Mitoses may bepresent, as the fluid acts as a tissue culture medium.63Surface microvilli and pinocytotic vesicles tend to dis-appear, presumably because their function is altered;and surfaces either are smooth or have characteristicsurface blebs.24 The nuclei may become hyper-chromatic and the cytoplasm vacuolated.6' Althoughsome of these changes are compatible with a responseto injury, others are more suggestive of active regen-eration. Exfoliated mesothelial cells may also have alimited phagocytic function.
Metastatic malignant cells can usually be dis-tinguished from mesothelial cells as a separate popu-lation with malignant nuclear characteristics and fea-tures that sometimes identify specific tumour types.63These cells are shown by cytogenetic analysis to haveclonal karyotypic abnormalities-a reliable thoughtime consuming way of identifying malignant cells ineffusions, which can, however, be used to supplementthe cytological diagnosis.69
It must be emphasised that mesothelial cells mayshow extreme nuclear atypia, possibly as a result ofcell damage, and occasionally isolated karyotypicabnormalities similar to those seen in malignantcells,68 69 resulting in an occasional "false positive"diagnosis of malignancy. These atypical mesothelialcells may show obvious signs of response to injury,and have homogenous, hyperchromatic, or multiplenuclei; but there may be a more coarse and activechromatin pattern with multiple nucleoli, closelyresembling malignancy. They are typically seen inpulmonary infarction, cirrhosis, and uraemia, and asa result of chemotherapy or irradiation.63 They can
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be distinguished from metastatic carcinoma becausethey do not form a separate population, but are partof a range of appearances that includes normal meso-thelial cells. Typical mesothelial cells are recognisedby the characteristic, sharply defined cytoplasm, con-densed around the nucleus, and often separated fromadjacent cells by intercellular gaps.63 This type ofmesothelial atypia is less often encountered in histo-logical sections because the mesothelial cells will haveexfoliated. When these cells line the inflamed surfacein biopsy specimens they do not have the infiltrativepattern of a tumour and are less likely to be confusedwith malignancy. Hyperchromatic mesothelial cellsmay also be associated with infiltration by metastaticcarcinoma,126 and the presence of atypical reactivemesothelial cells in effusions does not exclude car-cinoma.
Benign mesothelial cells may also mimic carcinomaor mesothelioma when small papillary aggregates arepresent. These are a feature of regeneration, and areseen in many forms of effusion, including thoseresulting from tuberculosis and infarction.62 63 73 Theaggregates are usually smaller than those of papillarycarcinoma or mesothelioma. The latter, with whichthey are most likely to be confused, are three dimen-sional and form tightly packed "morulae" of cellswith nuclei bulging from the surface,168 169 whereasbenign cell aggregates are usually in flat squamoidsheets.'70 Although cells with malignant character-istics can usually be identified in malignant meso-thelioma, particularly in cells separated from theclumps, one of the difficulties in making a cytologicaldiagnosis of this tumour is that the nuclei sometimesappear deceptively benign.63 169
Papillary clusters of mesothelial cells also occur inthe benign proliferative mesothelial lesions that areoccasionally associated with effusion17' (see below).There is no doubt that the cytological appearance ofmesothelioma overlaps with that of some of thesebenign processes as well as with adenocarcinoma. It istherefore unwise to give a certain cytological diagno-sis of mesothelioma without close clinical cor-relation,170 although a probable diagnosis can oftenbe made and is an indication for definitive biopsy.
Although mesothelial cells are mesenchymal in ori-gin they contain cytokeratins,172 but do not usuallyexpress epithelial cells markers, such as carcino-embryonic antigen, human milk fat globule, or Ca1.82 173 Identification of these markers by immu-nocytochemical techniques has proved useful in iden-tifying malignant cells not recognisable by mor-phology alone. None of the antisera used in thesetechniques are entirely specific, and agreementbetween immunocytochemical and morphologicalfindings is necessary for avoiding false positive diag-noses.
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Pathogenesis ofpleurisy, pleural.fibrosis, and mesothelial proliferation
Proliferation ofsurface mesotheliumBenign proliferative mesothelial lesions occur in theperitoneum,174 pericardium,'75 hernial sacs,'76 andpleura.'77 178 A series of five cases has been reportedrecently, with effusions in one or more body cavities,including the pleura, in three.17' They consist offronds of mature connective tissue lined by benignmesothelium. There is no biphasic pattern, and theyare histologically benign, although they can give riseto false positive cytological diagnoses of malig-nancy. '71 175 They may be manifestations of regen-eration and fibrosis rather than true tumours.176Although true papillary structures with a loose con-nective tissue core are seldom seen in small biopsyspecimens in the absence of mesothelioma,'26 theyshould not be regarded as mesotheliomatous in theabsence of unequivocal invasion.A combination of closely applied fibrous bands and
mesothelial regeneration may appear similar to thesebenign nodular lesions, and may be difficult to dis-tinguish from malignant mesothelioma, particularlyin patients with a history of exposure to asbestos. Inbiopsy material the regenerating mesothelium isheaped up and multilayered on the serosal surface,'26but it becomes single layered as it relines the fibrousbands. Entrapment of islands of surviving meso-thelium beneath the fibrous connective tissue can bedistinguished from true invasion by the benignappearance of the cells.170 The key to appreciatingthe benign nature of the process again lies in recog-nising that fibrosis and the mesothelial regenerationare separate processes, whereas malignant meso-thelioma shows simultaneous differentiation towardsmesenchymal and epithelial structures.179
Conclusion
The pleura is a remarkable structure, adapted to forma lubricated surface that allows the lungs to expandand contract without resistance, and to resist theaccumulation of fluid despite negative intrapleuralpressure. Partly because of its extreme sensitivity to awide variety of agents, Barrett has described thepleura as "an anatomical luxury and a pathologicalhazard."45 Elephants and other members of the Pro-boscideae, which develop high intrathoracic negativepressures, lose their pleural spaces during fetal lifeand apparently show no respiratory disability. Manyof the enigmas of the pleural reponse to injury resultfrom the divergent functions of mesothelium, whichshows partial differentiation towards epitheliumthough derived entirely from mesenchyme. A dis-tinction must be drawn between the mesothelialresponse to injury and the process of fibrous repairthat takes over when the mesothelium is irreversiblydestroyed.
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