Expression of lectin receptors during hepatocarcinogenesis induced by chemical carcinogen in rats*
1Department of Pathology, Qingdao Medical College, Qingdao 266021, Shandong Province, China
2Department of Foreign Languages, Qingdao Institute of Architecture and Engineering, Qingdao 266033, Shandong Province, China
3Institute of Dermatology, Qingdao Medical College, Qingdao 266012, Shandong Province, China
NIU Zhao-Shan, male, born on 1964-03-25 in Qingdao City, Shandong Province and graduated from Binzhou Medical College in 1986, lecturer, engaged in the basic research of hepatocellular carcinoma; having 8 papers published.
Tel: +86·532·3838480-3758
*Supported by the foundation of Shandong Provincial Science and Technology Committee (No. 1992-18); this subject has been awarded the third prize for Shandong Provincial Science and Technology Progress (No. 95-3-89-1).
Correspondence to NIU Zhao-Shan.
Received 1997-03-16
Subject headings carcinogens; liver neoplasms, experimental/metabolism; receptors, mitogen/metabolism
Abstract
AIM To study the relationship between oval cells, transitional cells derived from oval cells and hepatocarcinogenesis.
METHODS The expressions of peanut agglutinin (PNA), phasolus vulgaris agglutinin (PHA), ulex europaeus agglutinin (UEA) and soybean agglutinin (SBA) receptors during hepatocarcinogenesis induced by chemical carcinogens were studied with affinitive histochemistry.
RESULTS Some of the oval and transitional cells were positive for PNA, PHA and UEA; normal liver cells, hyperplastic foci and nodules were negative for PNA, PHA, UEA and SBA; and hepatocellular carcinoma (HCC) was positive for PNA, PHA, UEA and SBA.
CONCLUSION Carcinogen-induced oval cells and transitional cells derived from oval cells share some common glycoproteins with HCC. Oval cells and transitional cells are the early cells of hepatocarcinogenesis. PNA, PHA and UEA might be used as an index of early lesions during experimental hepatocarcinogenesis.
INTRODUCTION
Lectin is a type of protein or glycoprotein which can specifically identify different oligosaccharide architectures. Lectin receptors in tissues may reflect some features of complex carbohydrate architecture. Therefore, lectins have become important tools in studying complex carbohydrate in tumor tissues[1]. In order to investigate the relationship between oval cells, transitional cells derived from oval cells and hepatocarcinogenesis, the expressions of 4 types of lectin receptors in oval cells, transitional cells and hepatocellular carcinoma (HCC) tissues during hepatocarcinogenesis induced by chemical carcinogens were studied with affinitive histochemistry in the present study.
MATERIALS AND METHODS
MaterialsSixty Fischer 344 (F344) male rats with the body weight of 130g-150g were chosen, which were fed with a diet containing 0.06% 3′-methyl-4-dimethylaminoazobenzene (3′-Me-DAB) for 98 days, then they were given a standard diet. Three to 5 rats were killed at an interval of 21 days.
MethodsThe rat liver tissues were fixed in 95% alcohol, embedded in paraffin, and consectutive sections cut at 4μm, the expressions of peanut agglutinin (PNA), phasolus vulgaris agglutinin (PHA), ulex europaeus agglutinin (UEA) and soybean agglutinin (SBA) receptors were carried out by avidin-biotin-peroxidase complex (ABC) method. Biotin and ABC reagent were purchased from the Vector Company USA, PNA, SBA and PHA had a same dilution of 1∶500, and the dilutions for UEA and ABC were 1∶400 and 1∶200.
RESULTS
Pathological changesGrossly, on the 14th day after the 3′-Me-DAB administration, the size of the liver was apparently reduced and the weight was lost. Between 14-35 days, round and grey hyperplastic nodules of 1mm diameter were seen on the surface and sectioning surface of the liver. After 140 days, the number of the hyperplastic nodules gradually decreased and the surface of the liver became smoother.
Histologically, there were three phases according to different pathological changes in the course of 3′-Me-DAB administration. ① Early stage: the proliferation stage of oval cells, transitional cells and small hepatocytes. On the 14th day after 3′-Me-DAB administration, the toxic effects of carcinogen brought degeneration and necrosis to hepatocytes and the architectures of hepatic lobules were destroyed with foci of necrosis. Subsequently, oval cells proliferated and extended from the portal tracts into lobules. Oval cells appeared oval in shape, small in size, scanty pale stained cytoplasm and often arranged in two separate ranks with small cracks but not glandular structures between them. After 28 days, oval cells proliferated greatly, and more transitional cells appeared which were slightly larger in size than oval cells. These transitional cells had somewhat abundant cytoplasm and piled up and showed acinar structure. On the 42nd day, the proliferation of the oval and transitional cells reached their peak; and on the 70th day, they gradually decreased. ② Middle stage: the formation stage of hyperplastic foci and nodules. On the 21st day after 3′-Me-DAB administration, hyperplastic foci were formed. On the 28th day, hyperplastic nodules appeared, with more clear cell nodules and acidophilic cell nodules and few basophilic cell nodules. Liver cell dysplasia occurred on the 70th day. ③Late stage: the formation stage of hepatocellular carcinoma. 140 days after 3′-Me-DAB administration, the hyperplastic nodules gradually decreased and liver architecture tended to be normal. There existed the proliferation of basophilic large cells in basophilic cells nodules. Hepatocellular carcinoma was the main histological pattern of liver carcinoma. The carcinoma cell varied in shape which could be arranged in cordal, lamellar, trabecular, papillary and glandular shapes, etc.
Expressions of lectin receptors by ABC methodThe expressions of lectin receptors were variable, some of the oval and transitional cells were positive for PNA, PHA and UEA receptors; the bile ductular cells positive for PNA, PHA, SBA and UEA receptors; the hepatic sinusoidal endothelial cells positive for PHA and UEA receptors; normal liver cells, hyperplastic foci and nodules were negative for PNA, PHA, UEA and SBA receptors; and the HCC positive for PNA, PHA, UEA and SBA receptors. The positive expressions might be divided into cytoplasmic, nuclear, membranous, lacunar and secretive patterns.
DISCUSSION
Carbohydrate plays an important role in vital activities, which is associated with the occurrence, recurrence and metastasis of tumors[2]. Recent findings suggest that the structure of complex carbohydrate has greatly altered when tumor occurs and the emergence of certain complex carbohydrates is related with some biological behaviors of tumors[2]. Abnormal expressions of surface complex carbohydrate of tumor cells are: ①certain normal oligosaccharide residues lost; ②the synthesis of normal oligosaccharide residues increased; ③the distribution of normal carbohydrates altered; ④ some immature or new oligosaccharides appeared and became the determiner of the tumor associated antigens[3].
Makino[4] found different expressions of lectin receptors in the liver lesions induced by administration of the carcinogens 3′-Me-DAB, 2-acetylaminofluorene (2-AAF) and non-carcinogene alpha-nephthylisothiocyanate (ANIT). Two different binding patterns were observed in oval and proliferated bile ductular cells. One group showed increased binding of PNA, while the other showed intense binding of UEA. These results suggested that oval and proliferated bile ductular cells had their own specific carbohydrate residues. Lemire[5] also noticed that oval cells were positive for PNA receptors. Among the 4 types of lectin receptors in this study, the expressions of the HCC were all positive, those of the normal liver cells were all negative, some of the oval and transitional cells were positive for PNA, PHA and UEA receptors, which suggested that oval and transitional cells share some common glycoproteins with HCC. Our results suggest that oval and transitional cells derived from oval cells are pro-cells of hepatocarcinogenesis, which gradually gained PNA, PHA and UEA receptors in the process of their proliferation. So, PNA, PHA and UEA receptors might be used as one of the indicators of early lesions during experimental hepatocarcinogenesis.
The relationship between oval cells, transitional cells derived from oval cells and hepatocarcinogenesis has always been one of the key points in studying experimental HCC. To study the differentiation of oval cells during hepatocarcinogenesis in F344 rats by means of double staining for gamma-glutamyl transpeptidase (GGT) and alpha fetoprotein (AFP), Zhang ZC et al[6] confirmed that oval cells staining was positive for GGT and not for AFP; transitional cells staining was positive for GGT and AFP; and the HCC was positive for both GGT and AFP. This result further suggests that oval and transitional cells are related to hepatocarcinogenesis. The results of morphological measurement indicated that there were significant difference between the surface marks on oval and transitional cells showed by morphological measurement (P<0.01), suggesting that they are different cell populations. We think that transitional cells induced by the carcinogen may be a new cell population. Arising from the oval cells but different from oval cells, transitional cells are the pro-cells of hepatocarcinogenesis.
The results of the present study indicated that hyperplastic foci and nodules were negative for PNA, PHA, UEA and SBA receptors, but some oval and transitional cells were positive for PNA, PHA and UEA receptors before hyperplastic foci and nodules formed. It was because oval and transitional cells derived from oval cells, proliferated in the early stage after 3′-Me-DAB administration, were still puerile, while hyperplastic foci and nodules formed in the mid stage, were more mature. We think that oval and transitional cells bear an inner and inevitable relationship with hyperplastic foci and nodules. With carcinogens, the liver cell necrosis is extensive and on this basis, the oval cells proliferate markedly and differentiate into transitional cells, then into small hepatocytes which proliferate into hyperplastic foci and nodules and finally into HCC.
REFERENCES
1 Allison RT. Lectins in diagnostic histopathology: a review. Med Lab Sci, 1986;43(4):369-376
2 Nicolson GL. Cell surface molecules and tumor metastasis. Regulation of metastatic phenotypic diversity. Exp Cell Res,
1984;150(1):3-22
3 Damjanov I. Lectin cytochemistry and histochemistry. Lab Invest, 1987;57(1):5-20
4 Makino Y, Yamamoto K, Tsuji T. Specific lectin binding of oval cells and proliferated bile ductules. Gastroenterology (Jpn),1988;23(6):658-665
5 Lemire JM, Shiojiri N, Fausto N. Oval cell proliferation and the origin of small hepatocytes in livers injury induced by
D-galactosamine. Am J Pathol, 1991;139(3):535-552
6 Zhang ZC, Xu YD, Ying YY. The study of oval cell differentiation during hepatocarcinogenesis in rats with the help of
double staining. Acta Acad Med Qingdao, 1989;25(4):257-261, 百拇医药(NIU Zhao-Shan1, ZHANG Zhao-Cheng1,ZOU Wei1, WANG Mei2 and WANG Guo-Ying3)
2Department of Foreign Languages, Qingdao Institute of Architecture and Engineering, Qingdao 266033, Shandong Province, China
3Institute of Dermatology, Qingdao Medical College, Qingdao 266012, Shandong Province, China
NIU Zhao-Shan, male, born on 1964-03-25 in Qingdao City, Shandong Province and graduated from Binzhou Medical College in 1986, lecturer, engaged in the basic research of hepatocellular carcinoma; having 8 papers published.
Tel: +86·532·3838480-3758
*Supported by the foundation of Shandong Provincial Science and Technology Committee (No. 1992-18); this subject has been awarded the third prize for Shandong Provincial Science and Technology Progress (No. 95-3-89-1).
Correspondence to NIU Zhao-Shan.
Received 1997-03-16
Subject headings carcinogens; liver neoplasms, experimental/metabolism; receptors, mitogen/metabolism
Abstract
AIM To study the relationship between oval cells, transitional cells derived from oval cells and hepatocarcinogenesis.
METHODS The expressions of peanut agglutinin (PNA), phasolus vulgaris agglutinin (PHA), ulex europaeus agglutinin (UEA) and soybean agglutinin (SBA) receptors during hepatocarcinogenesis induced by chemical carcinogens were studied with affinitive histochemistry.
RESULTS Some of the oval and transitional cells were positive for PNA, PHA and UEA; normal liver cells, hyperplastic foci and nodules were negative for PNA, PHA, UEA and SBA; and hepatocellular carcinoma (HCC) was positive for PNA, PHA, UEA and SBA.
CONCLUSION Carcinogen-induced oval cells and transitional cells derived from oval cells share some common glycoproteins with HCC. Oval cells and transitional cells are the early cells of hepatocarcinogenesis. PNA, PHA and UEA might be used as an index of early lesions during experimental hepatocarcinogenesis.
INTRODUCTION
Lectin is a type of protein or glycoprotein which can specifically identify different oligosaccharide architectures. Lectin receptors in tissues may reflect some features of complex carbohydrate architecture. Therefore, lectins have become important tools in studying complex carbohydrate in tumor tissues[1]. In order to investigate the relationship between oval cells, transitional cells derived from oval cells and hepatocarcinogenesis, the expressions of 4 types of lectin receptors in oval cells, transitional cells and hepatocellular carcinoma (HCC) tissues during hepatocarcinogenesis induced by chemical carcinogens were studied with affinitive histochemistry in the present study.
MATERIALS AND METHODS
MaterialsSixty Fischer 344 (F344) male rats with the body weight of 130g-150g were chosen, which were fed with a diet containing 0.06% 3′-methyl-4-dimethylaminoazobenzene (3′-Me-DAB) for 98 days, then they were given a standard diet. Three to 5 rats were killed at an interval of 21 days.
MethodsThe rat liver tissues were fixed in 95% alcohol, embedded in paraffin, and consectutive sections cut at 4μm, the expressions of peanut agglutinin (PNA), phasolus vulgaris agglutinin (PHA), ulex europaeus agglutinin (UEA) and soybean agglutinin (SBA) receptors were carried out by avidin-biotin-peroxidase complex (ABC) method. Biotin and ABC reagent were purchased from the Vector Company USA, PNA, SBA and PHA had a same dilution of 1∶500, and the dilutions for UEA and ABC were 1∶400 and 1∶200.
RESULTS
Pathological changesGrossly, on the 14th day after the 3′-Me-DAB administration, the size of the liver was apparently reduced and the weight was lost. Between 14-35 days, round and grey hyperplastic nodules of 1mm diameter were seen on the surface and sectioning surface of the liver. After 140 days, the number of the hyperplastic nodules gradually decreased and the surface of the liver became smoother.
Histologically, there were three phases according to different pathological changes in the course of 3′-Me-DAB administration. ① Early stage: the proliferation stage of oval cells, transitional cells and small hepatocytes. On the 14th day after 3′-Me-DAB administration, the toxic effects of carcinogen brought degeneration and necrosis to hepatocytes and the architectures of hepatic lobules were destroyed with foci of necrosis. Subsequently, oval cells proliferated and extended from the portal tracts into lobules. Oval cells appeared oval in shape, small in size, scanty pale stained cytoplasm and often arranged in two separate ranks with small cracks but not glandular structures between them. After 28 days, oval cells proliferated greatly, and more transitional cells appeared which were slightly larger in size than oval cells. These transitional cells had somewhat abundant cytoplasm and piled up and showed acinar structure. On the 42nd day, the proliferation of the oval and transitional cells reached their peak; and on the 70th day, they gradually decreased. ② Middle stage: the formation stage of hyperplastic foci and nodules. On the 21st day after 3′-Me-DAB administration, hyperplastic foci were formed. On the 28th day, hyperplastic nodules appeared, with more clear cell nodules and acidophilic cell nodules and few basophilic cell nodules. Liver cell dysplasia occurred on the 70th day. ③Late stage: the formation stage of hepatocellular carcinoma. 140 days after 3′-Me-DAB administration, the hyperplastic nodules gradually decreased and liver architecture tended to be normal. There existed the proliferation of basophilic large cells in basophilic cells nodules. Hepatocellular carcinoma was the main histological pattern of liver carcinoma. The carcinoma cell varied in shape which could be arranged in cordal, lamellar, trabecular, papillary and glandular shapes, etc.
Expressions of lectin receptors by ABC methodThe expressions of lectin receptors were variable, some of the oval and transitional cells were positive for PNA, PHA and UEA receptors; the bile ductular cells positive for PNA, PHA, SBA and UEA receptors; the hepatic sinusoidal endothelial cells positive for PHA and UEA receptors; normal liver cells, hyperplastic foci and nodules were negative for PNA, PHA, UEA and SBA receptors; and the HCC positive for PNA, PHA, UEA and SBA receptors. The positive expressions might be divided into cytoplasmic, nuclear, membranous, lacunar and secretive patterns.
DISCUSSION
Carbohydrate plays an important role in vital activities, which is associated with the occurrence, recurrence and metastasis of tumors[2]. Recent findings suggest that the structure of complex carbohydrate has greatly altered when tumor occurs and the emergence of certain complex carbohydrates is related with some biological behaviors of tumors[2]. Abnormal expressions of surface complex carbohydrate of tumor cells are: ①certain normal oligosaccharide residues lost; ②the synthesis of normal oligosaccharide residues increased; ③the distribution of normal carbohydrates altered; ④ some immature or new oligosaccharides appeared and became the determiner of the tumor associated antigens[3].
Makino[4] found different expressions of lectin receptors in the liver lesions induced by administration of the carcinogens 3′-Me-DAB, 2-acetylaminofluorene (2-AAF) and non-carcinogene alpha-nephthylisothiocyanate (ANIT). Two different binding patterns were observed in oval and proliferated bile ductular cells. One group showed increased binding of PNA, while the other showed intense binding of UEA. These results suggested that oval and proliferated bile ductular cells had their own specific carbohydrate residues. Lemire[5] also noticed that oval cells were positive for PNA receptors. Among the 4 types of lectin receptors in this study, the expressions of the HCC were all positive, those of the normal liver cells were all negative, some of the oval and transitional cells were positive for PNA, PHA and UEA receptors, which suggested that oval and transitional cells share some common glycoproteins with HCC. Our results suggest that oval and transitional cells derived from oval cells are pro-cells of hepatocarcinogenesis, which gradually gained PNA, PHA and UEA receptors in the process of their proliferation. So, PNA, PHA and UEA receptors might be used as one of the indicators of early lesions during experimental hepatocarcinogenesis.
The relationship between oval cells, transitional cells derived from oval cells and hepatocarcinogenesis has always been one of the key points in studying experimental HCC. To study the differentiation of oval cells during hepatocarcinogenesis in F344 rats by means of double staining for gamma-glutamyl transpeptidase (GGT) and alpha fetoprotein (AFP), Zhang ZC et al[6] confirmed that oval cells staining was positive for GGT and not for AFP; transitional cells staining was positive for GGT and AFP; and the HCC was positive for both GGT and AFP. This result further suggests that oval and transitional cells are related to hepatocarcinogenesis. The results of morphological measurement indicated that there were significant difference between the surface marks on oval and transitional cells showed by morphological measurement (P<0.01), suggesting that they are different cell populations. We think that transitional cells induced by the carcinogen may be a new cell population. Arising from the oval cells but different from oval cells, transitional cells are the pro-cells of hepatocarcinogenesis.
The results of the present study indicated that hyperplastic foci and nodules were negative for PNA, PHA, UEA and SBA receptors, but some oval and transitional cells were positive for PNA, PHA and UEA receptors before hyperplastic foci and nodules formed. It was because oval and transitional cells derived from oval cells, proliferated in the early stage after 3′-Me-DAB administration, were still puerile, while hyperplastic foci and nodules formed in the mid stage, were more mature. We think that oval and transitional cells bear an inner and inevitable relationship with hyperplastic foci and nodules. With carcinogens, the liver cell necrosis is extensive and on this basis, the oval cells proliferate markedly and differentiate into transitional cells, then into small hepatocytes which proliferate into hyperplastic foci and nodules and finally into HCC.
REFERENCES
1 Allison RT. Lectins in diagnostic histopathology: a review. Med Lab Sci, 1986;43(4):369-376
2 Nicolson GL. Cell surface molecules and tumor metastasis. Regulation of metastatic phenotypic diversity. Exp Cell Res,
1984;150(1):3-22
3 Damjanov I. Lectin cytochemistry and histochemistry. Lab Invest, 1987;57(1):5-20
4 Makino Y, Yamamoto K, Tsuji T. Specific lectin binding of oval cells and proliferated bile ductules. Gastroenterology (Jpn),1988;23(6):658-665
5 Lemire JM, Shiojiri N, Fausto N. Oval cell proliferation and the origin of small hepatocytes in livers injury induced by
D-galactosamine. Am J Pathol, 1991;139(3):535-552
6 Zhang ZC, Xu YD, Ying YY. The study of oval cell differentiation during hepatocarcinogenesis in rats with the help of
double staining. Acta Acad Med Qingdao, 1989;25(4):257-261, 百拇医药(NIU Zhao-Shan1, ZHANG Zhao-Cheng1,ZOU Wei1, WANG Mei2 and WANG Guo-Ying3)