Path. Res. Pract. 187,931-935 (1991)
Ultrastructure of Mesothelial Regeneration after Intraperitoneal Injection of Asbestos Fibres on Rat Omentum 1 S. Gonzalez2 Department Anatomia Patologica, Escuela de Medicina, Universidad Catolica de Chile, Santiago, Chile
J. Friemann and K.-M. Muller Institut fur Pathologie an den Berufsgenossenschaftlichen Krankenanstalten Bergmannsheil, Universitatsklinik, Bochum, FRG
F. Pott Medizinisches Institut fur Umwelthygiene an der Universitat Dusseldorf, Dusseldorf, FRG
SUMMARY
In order to describe the ultrastructural features of the early phases of regenerating mesothelium in rat peritoneum, 69 cases were examined after intraperitoneal injection of 0.05-15 mg crocidolite, chrysotile B and other mineral and synthetic fibers. The findings show the presence of intermediate or transition cells between proliferating submesothelial connective tissue cells bearing the ultrastructural phenotype of myofibroblasts and mature fully regenerated mesothelium. Our results and data accumulated in the literature provide strong support for the hypothesis of submesothelial cell origin for regenerating mesothelium.
Introduction Under experimental conditions, asbestos fibers induce fibrosis and foreign body granulomas within the submesothelial connective tissue 4,5, 14, 15, but also reactive mesothelial hyperplasia with atypical features 1,5-7, 14, 15. Finally, the development of malignant mesothelioma is observed in a high proportion of the cases 2, 5-8,11. These observations and other recent investigations have suggested an important role for submesothelial cell populations in the regeneration of the injured mesothelium 1,3,5,16. The present report deals with the ultrastructural features of regenera1 With financial support of the Hauptverband der Gewerblichen Berufsgenossenschaften e.V., Bonn, St. Augustin. 2 Fellow of the Alexander von Humboldt Foundation.
© 1991 by Gustav Fischer Veriag, Stuttgart
tion of peritoneal serosa of rat omentum after intraperitoneal injection of asbestos ~ibers and synthetic fibers in order to describe the early phases of regenerating mesothelium. Light microscopic findings and experimental models have been described elsewhere5,8. Material and Methods 64 female Sprague-Dawley rats were sacrificed under anesthesia 8 hours to 15 months after one intraperitoneal injection of 1 to 15 mg crocidolite (32 cases; South African) or chrysotilc B (32 cases; Canadian asbestos), both UICC reference samples, dissolved in 1 ml physiological NaCl solution. In addition, omentum specimens from long-term (25 months) carcinogenicity studies in 5 female Wi star rats with natural and man-made mineral fibers were also included. These experiments have already been referred 0,44-0338/91/0187-0931$3.50/0
932 . S. Gonzalez et al. to elsewhere 8• Representative samples of omentum were immediatly obtained and immersed in 3 % glutaraldehyde with sodium phosphate buffer (pH 7.4) for transmission electron microscopy. Postfixation in a 1% aqueous solution of OS04 was followed by dehydration in graded ethanols and embedding in Epon 812. Semithin sections for preliminary orientation of 0.5-1 11m thickness were stained with toluidine blue. Ultrathin sections were contrasted with uranyl acetate and lead citrate and subsequently examined under a Zeiss 109 electron microscope at 50 and/or 80KV.
Results The ultrastructural findings were similar in both experimental groups, and what follows is a summary of these features. In the early phase, that is 8 hours to 1 week after the injection of asbestos fibers, mesothelial cells were not visible on the omental surface. Cellular debris and a denuded surface zone with fibrin and collagen fibris were focally found. The underlying stroma was loose and inflammatory cells such as lymphocytes, macrophages, neutrophils and scant plasmocytes were also identified. In the intermediate phase, 2 weeks to 6 months after the intraperitoneal injection of asbestos fibres, the omental surface was partly covered by thin, elongated, undifferentiated cells (Fig. 1). These cells neither showed desmosomes nor identifiable organelles. A basal lamina was not visible and they were lying directly on mature collagen fibril bundles. In the underlying stroma fibroblasts and undifferentiated mesenchymal cells were recognized, immersed in a relatively dense fibrous connective tissue. In other areas, especially serosal duplicatures, a stratified layer of some spindle cells already showing ultrastructural
features of immature mesothelium was observed (Fig. 2). The more superficially located cells showed abundant cytoplasm with its largest diameter in horizontal direction; an elongated nucleus with irregular boundaries and heterochromatin clumps was also visible. The cytoplasm showed moderate amounts of rough endoplasmic reticulum cisternae, mitochondria, focal aggregates of tonofilaments and short microvilli. Desmosomes were frequently identified. The cells of deeper layers showed intermediate differentiation features with a rather clear cytoplasm, absence of microvilli, close cellular appositions without recognizable intercellular junctions. Peripheral microfilaments with dense bodies (myofilaments) were also prominent. Within the stroma, but immediately under the superficial stratified layer, myofibroblasts and fibroblasts were also identified. These cells showed abundant elongated cytoplasm with well-developed rough endoplasmic reticulum, delicate and coarser cytoplasmic processes and subplasmalemmal myofilaments. Scattered lymphocytes and macrophages among mature colagen fibrils were also observed. After 6 months and before the full development of mesothelioma, in the proliferative mesothelial phase, the serosal surface was covered by a stratified layer of cells showing mesothelial differentiation and resting on a partially fragmented basal lamina (Fig. 3A). The more superficially located cells showed a cuboidal and/or polygonal contour with numerous long sinous and also shorter microvilli. Desmosomes were numerous. The deeply located cells were elongated, with lesser amounts of superficial microvilli and they were resting on a discontinuous basal lamina. Their cytoplasm showed less organelles, predominantly mitochondria and rough endoplasmic reticulum. The nuclei showed variable amounts of
Fig. 1. 4 months after 15 mg chrysotile injection. The serosal surface is covered only by a thin layer of cytoplasm of an undifferentiated cell. A basal lamina is not recognizable. The underlying stroma is loose with mature collagen fibers and a solitary fibroblast is visible. Uranyl acetate and lead citrate, X 12000.
Mesothelial Regeneration of Asbestos Fibres . 933
Fig. 2. 6 months after 0.05 mg actinolite. The surface layer is made up of mesothelial cells, and underlying transitional cells showing features of myofibroblasts are also visible. Desmosomes and tonofilaments are already identifiable in the more superficially located cells. Uranyl acetate and lead citrate, x 3150.
Fig. 3A. 25 months after 0.05 mg actinolite. The serosa is covered by a well-differentiated mesothelium resting on a partially fragmented basal lamina. The underlying stroma depicts mature collagen fibrils and myofibroblasts in a conspicuous mosaic-like aggregate. Uranyl acetate and lead citrate, x 3150.
934 . S. Gonzalez et al.
f
clumped heterochromatin, and nucleoli were not visible. The cytoplasm showed some dispersed lipid droplets (Fig. 3B). The underlying stroma was composed of abundant bundles of mature collagen fibrils, most of them disorderly arranged, but also around fibroblasts. Occasionally, nests of myofibroblasts and fibroblasts with a conspicuous mosaic-like arrangement were also recognized. Close apposition of cellular membranes was clearly visible, but intercellular junctions were not observed. Asbestos fibres were not found. Discussion The present report shows ultrastructural evidence for the presence of intermediate cells between submesothelial connective tissue cells and mature mesothelium in rat omentum after intraperitoneal injection of asbestos fibers. The origin of regenerated mesothelial cells has been a matter of controversy in the last few years. An origin from undifferentiated mesenchymal cells l , lO, sub mesothelial fibroblasts3, multinuclated giant cells at the margin of mesothelial injury12, macrophageslO, and mesothelial cells of adjacent serosal 7has been postulated. Recent observations of human tissues 1,2 have strongly suggested the existence of a so-called multipotential subserosal cell, this cell representing the proliferative cell after local injury. These authors have found that after the loss of the mesothelial layer the proliferating compartment is made up of spindle-shaped cells that ultrastructurally correspond to myofibroblasts l . RafterylO developed an experimental model in rats with mechanical injury of the visceral and parietal peritoneum and observed that subserosa 1cells proliferated and then suggested that these were capable of mesothelial differentiation. Furthermore, Davis3 injected crocidolite into the peritoneal cavity of rats and observed the formation of spindle-cell nests differentiating eventually towards mesothelial cells. Thus, data accumulated in the literature and our own findings give strong and definitive support to the hypothesis of sub mesothelial cell origin for regenerating mesothelium. In fact, after mesothelial injury and subsequently desquamation of the damaged cells, the
Fig. 3B. Fully regenerated mesothelium showing sur. face microvilli, desmosomes (arrowheads), scant tonofi.: laments, and a continuous basal lamina (arrows). Ura• I • • ':1 nyl acetate and lead citrate, x 7300.
subserosal cells proliferate. These proliferating cells are spindle-shaped and ultrastructurally bear features of myofibroblasts: elongated cell processes, abundant rough endoplasmic reticulum, and peripheral subplasmalemmal myofilaments. The more superficially these cells are located, the more frequent cellular membrane appositions and the more numerous intermediate filaments and microvilli are visible. The cells progressively continue to develop surface microvilli, desmosomes and tonofilaments. Finally, a basal lamina appears. These findings are also supported by immunocytochemical studies. Normal mesothelium expresses cytokeratin, especially high-molecularweight polypeptides l . Subserosa I cells usually express actin and vimentin, but proliferating submesothelial cells that ultrastructurally correspond to myofibroblasts, coexpress actin, vimentin and also low-molecular-weightcytokeratins. These myofibroblasts progressively begin to express high-molecular-weight-cytokeratins and to lose vimentin 1,2. Thus, ultrastructural and immunocytochemical observations confirm the capability of submesothelial myofibroblasts of differentiation towards mesothelium. In our experiments, even 0.05 mg actinolite, crocidolite or chrysotile injected intra peritoneally led to tumor rates of about 30-40%5. Mesothelial proliferation up to malignant mesothelioma was found only in association with submesothelial fibrosis and almost without any topographical relationship to foreign-body granulomas 4,5. The question of which factors and local conditions perpetuate and maintain the proliferation of submesothelial cells and progressively conduce to the development of malignant mesothelioma remains to be answered. In all of our cases asbestos fibers were consistently not found in the vicinity of the proliferating submesothelial and/or mesothelial cells. References 1 Bolen JW, Hammar SP, McNutt MA (1986) Reactive and neoplastic serosal tissue. A light microscopic, ultrastructural and immunocytochemical study. Am] Surg PathollO: 34-47 2 Brockmann M, Brockmann I, Fischer M, Muller K-M (1990) Reactive lesions of the pleura. Immunohistochemical characterization. Path Res Pract 186: 238-246
Mesothelial Regeneration of Asbestos Fibres . 935 3 Davis JMG (1974) Histogenesis and fine structure of peritoneal tumors produced in animals by injection of asbestos. J Nat! Cancer Inst 52: 1823-1837 4 Ellis H, Harrison W, Hugh TB (1965) The healing of peritoneum under normal and pathological conditions. Br J Surg 52:471-476 5 FriemannJ, Voss B, Weller W, Muller K-M (1987) Asbestos induced fibrosis in the omentum of rats. Immunofluorescence microscopical demonstration of collagen types I and III, laminin and fibronectin. Virchows Arch A (Pathol Anat) 411: 403-408 6 Friemann J, Muller K-M, Pott F (1990) Mesothelial proliferation due to asbestos and man-made fibres. Experimental studies on rat omentum. Path Res Pract 186: 117-123 7 Gormley IP, Bolton RE, Brown G, Davis JMG, Donaldson K (1980) Studies on the morphological patterns of asbestos induced mesotheliomas in vivo and in vitro. Carcinogenesis 2: 219-231 8 Moncheaux G, Bignon J, Jaurand MC, Lafuma J, Sebastien P, Masse R, Hirsch A, Goni J (1981) Mesotheliomas in rats following inoculation with acid-leached Chrysotile asbestos and other mineral fibres. Carcinogenesis 2: 232-236 9 Pott F, Ziem U, Reiffer F-J, Huth F, Ernst H, Mohr U (1987) Carcinogenity studies on fibres, metal compounds, and some other dusts in rats. Exp Pathol32: 129-152
10 Raftery AT (1973) Regeneration of partial and visceral peritoneum: An electronmicroscopical study. J Anat 115: 375-392 II Ryan GB, Groberty J,Majno G (1973) Mesothelial injury and recovery. Am J Pathol 71: 93-112 12 Shin ML, Firminger HI (1973) Acute and chronic effects of intraperitoneal injection of two types of asbestos in rats with a study of the histopathogenesis and ultrastructure of resulting mesotheliomas. Am J Pathol 70: 291-314 13 Sinapins D, Vollhaber H (1951) Zur Kenntnis der Pleuradeckzellen. Beitr. Klinik TBK 106: 165-182 14 Wagner JC (1986) Mesothelioma and mineral fibers. Cancer 57: 1905-1911 15 Winkler GC, Ruttner JR (1982) Penetration of asbestos fibers in the visceral peritoneum of mice. A scanning electron microscopic study. Exp Cell Bioi 50: 187-194 16 Winkler GC, Ruttner JR (1983) Early fibrogenicity of asbestos fibers in visceral peritoneum. Exp. Cell Bioi 51: 1-8 17 Whitaker D, Papadimitriou JM (1985) Mesothelial healing: morphological and kinetic investigations. J Pathol 14 5: 159-175
Received November 24, 1990 . Accepted December 7, 1990
Key words: Mesothelium - Asbestos - Mesothelioma - Carcinogenesis - Myofibroblasts S. Gonzalez, Departmento Anatomia Patologica, Escuela de Medicina, Universidad Catolica de Chile, Santiago, Chile
Ultrastructure of Mesothelial Regeneration after Intraperitoneal Injection of Asbestos Fibres on Rat Omentum 1 S. Gonzalez2 Department Anatomia Patologica, Escuela de Medicina, Universidad Catolica de Chile, Santiago, Chile
J. Friemann and K.-M. Muller Institut fur Pathologie an den Berufsgenossenschaftlichen Krankenanstalten Bergmannsheil, Universitatsklinik, Bochum, FRG
F. Pott Medizinisches Institut fur Umwelthygiene an der Universitat Dusseldorf, Dusseldorf, FRG
SUMMARY
In order to describe the ultrastructural features of the early phases of regenerating mesothelium in rat peritoneum, 69 cases were examined after intraperitoneal injection of 0.05-15 mg crocidolite, chrysotile B and other mineral and synthetic fibers. The findings show the presence of intermediate or transition cells between proliferating submesothelial connective tissue cells bearing the ultrastructural phenotype of myofibroblasts and mature fully regenerated mesothelium. Our results and data accumulated in the literature provide strong support for the hypothesis of submesothelial cell origin for regenerating mesothelium.
Introduction Under experimental conditions, asbestos fibers induce fibrosis and foreign body granulomas within the submesothelial connective tissue 4,5, 14, 15, but also reactive mesothelial hyperplasia with atypical features 1,5-7, 14, 15. Finally, the development of malignant mesothelioma is observed in a high proportion of the cases 2, 5-8,11. These observations and other recent investigations have suggested an important role for submesothelial cell populations in the regeneration of the injured mesothelium 1,3,5,16. The present report deals with the ultrastructural features of regenera1 With financial support of the Hauptverband der Gewerblichen Berufsgenossenschaften e.V., Bonn, St. Augustin. 2 Fellow of the Alexander von Humboldt Foundation.
© 1991 by Gustav Fischer Veriag, Stuttgart
tion of peritoneal serosa of rat omentum after intraperitoneal injection of asbestos ~ibers and synthetic fibers in order to describe the early phases of regenerating mesothelium. Light microscopic findings and experimental models have been described elsewhere5,8. Material and Methods 64 female Sprague-Dawley rats were sacrificed under anesthesia 8 hours to 15 months after one intraperitoneal injection of 1 to 15 mg crocidolite (32 cases; South African) or chrysotilc B (32 cases; Canadian asbestos), both UICC reference samples, dissolved in 1 ml physiological NaCl solution. In addition, omentum specimens from long-term (25 months) carcinogenicity studies in 5 female Wi star rats with natural and man-made mineral fibers were also included. These experiments have already been referred 0,44-0338/91/0187-0931$3.50/0
932 . S. Gonzalez et al. to elsewhere 8• Representative samples of omentum were immediatly obtained and immersed in 3 % glutaraldehyde with sodium phosphate buffer (pH 7.4) for transmission electron microscopy. Postfixation in a 1% aqueous solution of OS04 was followed by dehydration in graded ethanols and embedding in Epon 812. Semithin sections for preliminary orientation of 0.5-1 11m thickness were stained with toluidine blue. Ultrathin sections were contrasted with uranyl acetate and lead citrate and subsequently examined under a Zeiss 109 electron microscope at 50 and/or 80KV.
Results The ultrastructural findings were similar in both experimental groups, and what follows is a summary of these features. In the early phase, that is 8 hours to 1 week after the injection of asbestos fibers, mesothelial cells were not visible on the omental surface. Cellular debris and a denuded surface zone with fibrin and collagen fibris were focally found. The underlying stroma was loose and inflammatory cells such as lymphocytes, macrophages, neutrophils and scant plasmocytes were also identified. In the intermediate phase, 2 weeks to 6 months after the intraperitoneal injection of asbestos fibres, the omental surface was partly covered by thin, elongated, undifferentiated cells (Fig. 1). These cells neither showed desmosomes nor identifiable organelles. A basal lamina was not visible and they were lying directly on mature collagen fibril bundles. In the underlying stroma fibroblasts and undifferentiated mesenchymal cells were recognized, immersed in a relatively dense fibrous connective tissue. In other areas, especially serosal duplicatures, a stratified layer of some spindle cells already showing ultrastructural
features of immature mesothelium was observed (Fig. 2). The more superficially located cells showed abundant cytoplasm with its largest diameter in horizontal direction; an elongated nucleus with irregular boundaries and heterochromatin clumps was also visible. The cytoplasm showed moderate amounts of rough endoplasmic reticulum cisternae, mitochondria, focal aggregates of tonofilaments and short microvilli. Desmosomes were frequently identified. The cells of deeper layers showed intermediate differentiation features with a rather clear cytoplasm, absence of microvilli, close cellular appositions without recognizable intercellular junctions. Peripheral microfilaments with dense bodies (myofilaments) were also prominent. Within the stroma, but immediately under the superficial stratified layer, myofibroblasts and fibroblasts were also identified. These cells showed abundant elongated cytoplasm with well-developed rough endoplasmic reticulum, delicate and coarser cytoplasmic processes and subplasmalemmal myofilaments. Scattered lymphocytes and macrophages among mature colagen fibrils were also observed. After 6 months and before the full development of mesothelioma, in the proliferative mesothelial phase, the serosal surface was covered by a stratified layer of cells showing mesothelial differentiation and resting on a partially fragmented basal lamina (Fig. 3A). The more superficially located cells showed a cuboidal and/or polygonal contour with numerous long sinous and also shorter microvilli. Desmosomes were numerous. The deeply located cells were elongated, with lesser amounts of superficial microvilli and they were resting on a discontinuous basal lamina. Their cytoplasm showed less organelles, predominantly mitochondria and rough endoplasmic reticulum. The nuclei showed variable amounts of
Fig. 1. 4 months after 15 mg chrysotile injection. The serosal surface is covered only by a thin layer of cytoplasm of an undifferentiated cell. A basal lamina is not recognizable. The underlying stroma is loose with mature collagen fibers and a solitary fibroblast is visible. Uranyl acetate and lead citrate, X 12000.
Mesothelial Regeneration of Asbestos Fibres . 933
Fig. 2. 6 months after 0.05 mg actinolite. The surface layer is made up of mesothelial cells, and underlying transitional cells showing features of myofibroblasts are also visible. Desmosomes and tonofilaments are already identifiable in the more superficially located cells. Uranyl acetate and lead citrate, x 3150.
Fig. 3A. 25 months after 0.05 mg actinolite. The serosa is covered by a well-differentiated mesothelium resting on a partially fragmented basal lamina. The underlying stroma depicts mature collagen fibrils and myofibroblasts in a conspicuous mosaic-like aggregate. Uranyl acetate and lead citrate, x 3150.
934 . S. Gonzalez et al.
f
clumped heterochromatin, and nucleoli were not visible. The cytoplasm showed some dispersed lipid droplets (Fig. 3B). The underlying stroma was composed of abundant bundles of mature collagen fibrils, most of them disorderly arranged, but also around fibroblasts. Occasionally, nests of myofibroblasts and fibroblasts with a conspicuous mosaic-like arrangement were also recognized. Close apposition of cellular membranes was clearly visible, but intercellular junctions were not observed. Asbestos fibres were not found. Discussion The present report shows ultrastructural evidence for the presence of intermediate cells between submesothelial connective tissue cells and mature mesothelium in rat omentum after intraperitoneal injection of asbestos fibers. The origin of regenerated mesothelial cells has been a matter of controversy in the last few years. An origin from undifferentiated mesenchymal cells l , lO, sub mesothelial fibroblasts3, multinuclated giant cells at the margin of mesothelial injury12, macrophageslO, and mesothelial cells of adjacent serosal 7has been postulated. Recent observations of human tissues 1,2 have strongly suggested the existence of a so-called multipotential subserosal cell, this cell representing the proliferative cell after local injury. These authors have found that after the loss of the mesothelial layer the proliferating compartment is made up of spindle-shaped cells that ultrastructurally correspond to myofibroblasts l . RafterylO developed an experimental model in rats with mechanical injury of the visceral and parietal peritoneum and observed that subserosa 1cells proliferated and then suggested that these were capable of mesothelial differentiation. Furthermore, Davis3 injected crocidolite into the peritoneal cavity of rats and observed the formation of spindle-cell nests differentiating eventually towards mesothelial cells. Thus, data accumulated in the literature and our own findings give strong and definitive support to the hypothesis of sub mesothelial cell origin for regenerating mesothelium. In fact, after mesothelial injury and subsequently desquamation of the damaged cells, the
Fig. 3B. Fully regenerated mesothelium showing sur. face microvilli, desmosomes (arrowheads), scant tonofi.: laments, and a continuous basal lamina (arrows). Ura• I • • ':1 nyl acetate and lead citrate, x 7300.
subserosal cells proliferate. These proliferating cells are spindle-shaped and ultrastructurally bear features of myofibroblasts: elongated cell processes, abundant rough endoplasmic reticulum, and peripheral subplasmalemmal myofilaments. The more superficially these cells are located, the more frequent cellular membrane appositions and the more numerous intermediate filaments and microvilli are visible. The cells progressively continue to develop surface microvilli, desmosomes and tonofilaments. Finally, a basal lamina appears. These findings are also supported by immunocytochemical studies. Normal mesothelium expresses cytokeratin, especially high-molecularweight polypeptides l . Subserosa I cells usually express actin and vimentin, but proliferating submesothelial cells that ultrastructurally correspond to myofibroblasts, coexpress actin, vimentin and also low-molecular-weightcytokeratins. These myofibroblasts progressively begin to express high-molecular-weight-cytokeratins and to lose vimentin 1,2. Thus, ultrastructural and immunocytochemical observations confirm the capability of submesothelial myofibroblasts of differentiation towards mesothelium. In our experiments, even 0.05 mg actinolite, crocidolite or chrysotile injected intra peritoneally led to tumor rates of about 30-40%5. Mesothelial proliferation up to malignant mesothelioma was found only in association with submesothelial fibrosis and almost without any topographical relationship to foreign-body granulomas 4,5. The question of which factors and local conditions perpetuate and maintain the proliferation of submesothelial cells and progressively conduce to the development of malignant mesothelioma remains to be answered. In all of our cases asbestos fibers were consistently not found in the vicinity of the proliferating submesothelial and/or mesothelial cells. References 1 Bolen JW, Hammar SP, McNutt MA (1986) Reactive and neoplastic serosal tissue. A light microscopic, ultrastructural and immunocytochemical study. Am] Surg PathollO: 34-47 2 Brockmann M, Brockmann I, Fischer M, Muller K-M (1990) Reactive lesions of the pleura. Immunohistochemical characterization. Path Res Pract 186: 238-246
Mesothelial Regeneration of Asbestos Fibres . 935 3 Davis JMG (1974) Histogenesis and fine structure of peritoneal tumors produced in animals by injection of asbestos. J Nat! Cancer Inst 52: 1823-1837 4 Ellis H, Harrison W, Hugh TB (1965) The healing of peritoneum under normal and pathological conditions. Br J Surg 52:471-476 5 FriemannJ, Voss B, Weller W, Muller K-M (1987) Asbestos induced fibrosis in the omentum of rats. Immunofluorescence microscopical demonstration of collagen types I and III, laminin and fibronectin. Virchows Arch A (Pathol Anat) 411: 403-408 6 Friemann J, Muller K-M, Pott F (1990) Mesothelial proliferation due to asbestos and man-made fibres. Experimental studies on rat omentum. Path Res Pract 186: 117-123 7 Gormley IP, Bolton RE, Brown G, Davis JMG, Donaldson K (1980) Studies on the morphological patterns of asbestos induced mesotheliomas in vivo and in vitro. Carcinogenesis 2: 219-231 8 Moncheaux G, Bignon J, Jaurand MC, Lafuma J, Sebastien P, Masse R, Hirsch A, Goni J (1981) Mesotheliomas in rats following inoculation with acid-leached Chrysotile asbestos and other mineral fibres. Carcinogenesis 2: 232-236 9 Pott F, Ziem U, Reiffer F-J, Huth F, Ernst H, Mohr U (1987) Carcinogenity studies on fibres, metal compounds, and some other dusts in rats. Exp Pathol32: 129-152
10 Raftery AT (1973) Regeneration of partial and visceral peritoneum: An electronmicroscopical study. J Anat 115: 375-392 II Ryan GB, Groberty J,Majno G (1973) Mesothelial injury and recovery. Am J Pathol 71: 93-112 12 Shin ML, Firminger HI (1973) Acute and chronic effects of intraperitoneal injection of two types of asbestos in rats with a study of the histopathogenesis and ultrastructure of resulting mesotheliomas. Am J Pathol 70: 291-314 13 Sinapins D, Vollhaber H (1951) Zur Kenntnis der Pleuradeckzellen. Beitr. Klinik TBK 106: 165-182 14 Wagner JC (1986) Mesothelioma and mineral fibers. Cancer 57: 1905-1911 15 Winkler GC, Ruttner JR (1982) Penetration of asbestos fibers in the visceral peritoneum of mice. A scanning electron microscopic study. Exp Cell Bioi 50: 187-194 16 Winkler GC, Ruttner JR (1983) Early fibrogenicity of asbestos fibers in visceral peritoneum. Exp. Cell Bioi 51: 1-8 17 Whitaker D, Papadimitriou JM (1985) Mesothelial healing: morphological and kinetic investigations. J Pathol 14 5: 159-175
Received November 24, 1990 . Accepted December 7, 1990
Key words: Mesothelium - Asbestos - Mesothelioma - Carcinogenesis - Myofibroblasts S. Gonzalez, Departmento Anatomia Patologica, Escuela de Medicina, Universidad Catolica de Chile, Santiago, Chile
