Food Additives & Contaminants
ISSN: 0265-203X (Print) (Online) Journal homepage: http://www.tandfonline.com/loi/tfac19
The effect of dietary retinyl palmitate on the retinyl ester content of human or rat liver tumours A. Periquet , B. Periquet , C. Frayssinet , I. Tomatis , J. Ghisolfi & J. P. Thouvenot To cite this article: A. Periquet , B. Periquet , C. Frayssinet , I. Tomatis , J. Ghisolfi & J. P. Thouvenot (1990) The effect of dietary retinyl palmitate on the retinyl ester content of human or rat liver tumours, Food Additives & Contaminants, 7:S1, S69-S72, DOI: 10.1080/02652039009373851 To link to this article: http://dx.doi.org/10.1080/02652039009373851
Published online: 10 Jan 2009.
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Date: 06 November 2015, At: 17:31
FOOD ADDITIVES AND CONTAMINANTS, 1990, VOL. 7, SUPPLEMENT NO. 1, S 6 9 - S 7 2
The effect of dietary retinyl palmitate on the retinyl ester content of human or rat liver tumours A. PERIQUET, B. PERIQUET†, C. FRAYSSINET‡, I. TOMATIS†, J. GHISOLFI† and J. P. THOUVENOT† INSERM U-87, Université Paul Sabatier, 2 rue François Magendie, 31400 Toulouse, France, † Groupe d'Etudes en Nutrition Infantile CHU Purpan, Toulouse, France, and ‡ CNRS, Institut de Recherches Scientifiques sur le Cancer, Villejuif, France
Downloaded by [Universite Laval] at 17:31 06 November 2015
Introduction
Research and public attention have been increasingly focussed on the hypothesis that vitamins and micronutrients may decrease the incidence of some cancers (Mettlin et al. 1980, Krale et al. 1983, Birt 1986, Woutersen and GarderenHoetmer 1988). The possible role of vitamin A in chemoprevention and chemotherapy of cancer is an issue that is widely discussed but the only physiological function of the vitamin A that has been clearly denned on a molecular basis is its role in vision (Bollag 1979, Lotan 1980, Moon et al. 1983, Sporn and Roberts 1983). The study in our laboratory of two cases of human hepatoblastomas showed different qualitative and quantitative distributions of the vitamin A esters (Tomatis et al. 1988). It seemed important to verify, at an experimental level, the relationship existing between the qualitative and quantitative aspects of vitamin A metabolism during the different stages of carcinogenesis. Materials and methods
Human hepatoblastomas Three fragments drawn by partial hepatectomy were analysed in each case: fragment no. 1: the anatomy pathology showed no sign of neoplastic invasion: 'normal part', fragment no. 2: the anatomy pathology revealed a massive invasion: 'tumoural part', fragment no. 3: partially invaded: 'mixed tissue'. Animals and diets The experiments were carried out with male weanling Sprague-Dawley rats. The rats, housed in separate cages, were assigned to different dietary groups. The basal diet was the same in each group but supplemented with different levels of retinyl palmitate: group 1 (18 rats) with 1 IU/g diet, group 2 (18 rats) with 2 IU/g diet, group 3 (18 rats) with 20 IU/g diet and group 4 (18 rats) with 200 IU/g diet. Twelve rats in each dietary group were treated according to the Solt and Farber protocol (1976) viz.: aflatoxin Bi initiation at six weeks (2 mg/kgb.w., i.p.), selection by acetylamino fluorene from 8 to 10 weeks (0-02% in the diet), partial hepatectomy at 9 weeks and promotion from the 1 lth week by phenobarbital in the 0265-203X/90 $3.00© 1990 Taylor & Francis Ltd.
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drinking water (1 g/1). The six rats left in each group constituted the control animals. At 4 months post initiation, the rats were killed by decapitation. Blood was collected for analysis of plasma retinol according to the HPLC method of De Leenheer et al. (1979). Livers were removed, weighed and kept at 4°C. Homogenates were assayed according to our method published earlier (Periquet etal. 1985). Quantification of retinyl fatty acyl esters 7ra«s-retinol, /ra«s-retinyl palmitate and frans-retinyl acetate were purchased from Sigma Chemicals Co (St Louis, Missouri, USA). The fatty acyl chlorides were obtained from Aldrich Chemical Co Inc (Milwaukee, Wisconsin, USA). The retinyl ester standards (laurate, myristate, palmitoleate, pentadecanoate, linoleate, heptadecanoate, oleate and stearate) (were synthesized by reaction of trans-vttmoX with the corresponding acyl chloride as described by Huang and Goodman (1965) with slight modifications (Periquet etal. 1988). Retinyl esters present in the liver lipid extract were separated and quantified using an HPLC technique as described earlier (Periquet etal. 1988). Results In this article are reported the results obtained with the human hepatoblastomas and the experimental results obtained with the control and treated rats receiving a normal daily dose of vitamin A: 20 IU/g diet. The study of two cases of human hepatoblastomas showed: (1) at a quantitative level: a very high vitamin A level in the liver part considered healthy and especially in the mixed part after anatomopathological examination and, in contrast, a very low vitamin A level in the cancerous part (table 1). (2) at a qualitative level, a difference in the stocking profile with, in particular, an increase in all the minor forms at the expense of the retinyl palmitate (table 2). As for the results for the animals, table 3 shows that for the same daily contribution in vitamin A (20 IU/g diet as retinyl palmitate) there was four times less vitamin A in the treated rats' livers than in the controls' livers after twelve weeks of promotion. On the other hand, circulating vitamin A levels were comparable and stayed within normal limits. At 4 months post-initiation, at a qualitative level, no significant difference was noted between the stocking profile and the healthy human liver. The results illustrated in table 4 show that retinyl palmitate/oleate constituted the most important ester form (78-84%) of the total retinyl fatty acyl esters followed by stearate (11-15%), laurate, palmitoleate/myristate, linoleate, heptadecanoate and pentadecanoate totalling 6%. Table 1. Retinyl palmitate contents of a human liver tumour.
Total homogenate (retinyl palmitate) Hg/g
Normal part
Mixed tissue
Tumourous part
no. 1
no. 3
no. 2
663
1177
195
Effect of dietary retinyl palmitate on tumours
S71
Table 2. Distribution of the esters of retinol (%) in the normal human liver and in the healthy part of a cancerous human liver. Retinyl esters (%)
Normal human liver
Downloaded by [Universite Laval] at 17:31 06 November 2015
Normal part of cancerous human liver
C16:0
C18:0
C12.-0
80-85
10-15
0-4-0-6
65-70
9-12
0-4-0-7
C14:0 and C16:l
C18:2
C15:0
C17:0
1-0-2
0-7-1
0-2-0-6
traces
2-5
2-6
0-2-0-7
5-10
Table 3. Serum concentration of vitamin A and retinyl fatty acyl esters in liver homogenate of rats vitamin A sufficient diets (20 IU/g diet) for up to 4 months post initiation. Serum vit A (mg/1)
C16:0
C18:0
C12:0
C14:0 and C16:l
Control rats
0-42 ±0-087
504-25 ± 30-2
90-64 ± 5-6
4-51 ±0-38
Treated rats n=12
0-56 ±0-055
141-47 ± 16-12
22-38 ± 1-72
1-18 ±0-25
C18:0
C15:0
C17:0
4-70 ±0-59
23-6 ± 1-80
2-53 ±0-42
8-70 ±1-05
2-68 ±0-30
1-92 ±0-08
1-80 ±0-05
2-44 ±0-60
Table 4. Distribution of the esters of retinol (%) in the normal and treated rats' liver (4 months post-initiation). Retinyl esters (%)
C16:0
C18:0
C12:0
C14:0 and C16:l
C18.-2
C15.-0
C17.-0
Control livers
78-84
11-15
0-6-0-8
1-0-1-3
1-2-4
0-3-0-4
1-2-2-4
Treated livers
75-84
11-18
0-4-0-6
1-2-5
0-5-2
0-2-1-9
1-4-2-4
Discussion and conclusion The starting point of this work was the determination of vitamin A and its different esters in some cases of human hepatoblastomas. The results seem to indicate that there is accumulation of vitamin A especially in the mixed zone whereas there is a non-negligible proportion of tumoural cells a priori vitamin A deficient. This 'barrier phenomenon', with a very high vitamin A level separating the cancerous tissue from the healthy one, has not, to our knowledge, been reported. Muto and Omori (1981) made comparable observations concerning the low vitamin A level in the tumour; the hypothesis advanced is a perturbation in the
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A. Periquet et al.
retinol membrane transport from the surrounding healthy tissue and/or for a much faster utilization rate in the tumour cells. De Luca et al. (1984) showed that the nuclear membranes coming from carcinomas are depleted in retinol compared with the nuclear membranes of the healthy hepatic cells; they concluded that the hepatic stocking function for vitamin A has been lost in the tumour. We have tried to obtain similar results in animal experiments by initiating tumours by the Solt and Farber protocol. Our first results obtained in the animals seem to confirm these results and point out an increased vitamin A consumption during a Solt and Farber carcinogenesis process; they consolidate the defined hypothesis: modification of the vitamin A membrane transport in the tumourous tissues, a loss of the stocking characteristic and/or an increased use of this micronutrient by the tumour cells.
Downloaded by [Universite Laval] at 17:31 06 November 2015
References BIRT, D. F., 1986, Update on effects of vitamin A, C. and E and selenium on carcinogenesis. Proceeding of the Society for Experimental Biology and Medicine, 183, 311 -320. BOLLAG, W., 1979, Retinoids and cancer. Cancer Chemotherapy and Pharmacology, 2, 207-215. HUANG, H. S., and GOODMAN, D. S., 1965, Vitamin A and carotenoids I. Intestinal absorption and
metabolism of 14C-labelled vitamin A alcohol and β-carotène in the rat. Journal of Biological Chemistry, 240, 2839-2844. KRALE, G., BJELKE, E., and GART, J. J., 1983, Dietary habits and lung cancer risk. International Journal of Cancer, 31, 397-405. DE LEENHEER, A. P . , DE BEVERE, V., DE RUYTER, M. G. M., and CLAEYS, A. E., 1979, Simultaneous
determination of retinol and tocopherol in human serum by HPLC. Journal of Chromatography, 162, 408-412. LOTAN, R., 1980, Effects of vitamin A and its analogs (retinoids) on normal and neoplastic cells. Biochemistry et Biophysica Acta, 605, 33-91. DE LUCA, L. M., BRUGH, M., and SILVERMAN, J. C. S., 1984, Retinyl palmitate, retinyl phosphate and dolichyl phosphate of post nuclear membrane fraction from hepatoma host liver and regenerating: marginal vitamin A status of hepatoma tissue. Cancer Research, 44, 224-232. METTLIN, C., GRAHAM, S., PRIORE, R., MARSHALL, J., and SWANSON, M., 1980, Diet and cancer of the
esophagus. Nutrition and Cancer, 2, 143-147. MOON, R. C., MCCORMICK, D. L., and MEHTA, R. G., 1983, Chemoprevention of animals tumors by retinoids. In Modulation and Mediation of Cancer by Vitamins, edited by F. L. Meykens and K. N. Prasad (Basel: Karger) pp. 47-57. MUTO, Y., and OMARI, M., 1981, A novel cellular retinoid binding protein, f-type, in hepatocellular carcinoma. Annals of the New York Academy of Science, 359, 91-103. PERIQUET, B., BAILLY, A., GHISOLFI, J., and THOUVENOT, J. P., 1985, Determination of retinyl
palminate in homogenates and subcellular fractions of rat liver by liquid chromatography. Clinica Chimica Acta, 147, 41-49. PERIQUET, B., LAMBERT, W., BAILLY, A., TOMATIS, I., GHISOLFI, J., DE LEENHEER, A. P . , and
THOUVENOT, J. P . , 1988, Fatty acid composition and kinetic behaviour of liver retinyl esters in vitamin A sufficient and deficient rats. Clinica Chimica Ata, 172, 275-290. SOLT, D., and FARBER, E., 1976, New principle for the analysis of chemical carcinogenesis. Nature, London, 263, 701-703. SPORN, M. B., and ROBERTS, A. B., 1983, Role of retinoids in differenciation and carcinogenesis. Cancer Research, 43, 3034-3040. TOMATIS, I., FRAYSSINET, C , PERIQUET, A., PERIQUET, B., GHISOLFI, J., and THOUVENOT, J. P . , 1988,
Etude des différents esters de retinol des hepatoblastomes humains. Compte rendu des Journées du Colloque 'Ailment 2000' Janvier 1989, Ministère de la Recherche et de la Technologie, Paris, dans: Bilan 'Ailment 2000'. WOUTERSEN, R. A., and GARDEREN-HOETMER, A. M., 1988, Inhibition of dietary fat-promoted development of (pre) neoplastic lesions in exocrine pancreas of rats and hamsters by supplemental vitamins A, C and E. Cancer Letters, 41, 179-189.
ISSN: 0265-203X (Print) (Online) Journal homepage: http://www.tandfonline.com/loi/tfac19
The effect of dietary retinyl palmitate on the retinyl ester content of human or rat liver tumours A. Periquet , B. Periquet , C. Frayssinet , I. Tomatis , J. Ghisolfi & J. P. Thouvenot To cite this article: A. Periquet , B. Periquet , C. Frayssinet , I. Tomatis , J. Ghisolfi & J. P. Thouvenot (1990) The effect of dietary retinyl palmitate on the retinyl ester content of human or rat liver tumours, Food Additives & Contaminants, 7:S1, S69-S72, DOI: 10.1080/02652039009373851 To link to this article: http://dx.doi.org/10.1080/02652039009373851
Published online: 10 Jan 2009.
Submit your article to this journal
Article views: 4
View related articles
Full Terms & Conditions of access and use can be found at http://www.tandfonline.com/action/journalInformation?journalCode=tfac19 Download by: [Universite Laval]
Date: 06 November 2015, At: 17:31
FOOD ADDITIVES AND CONTAMINANTS, 1990, VOL. 7, SUPPLEMENT NO. 1, S 6 9 - S 7 2
The effect of dietary retinyl palmitate on the retinyl ester content of human or rat liver tumours A. PERIQUET, B. PERIQUET†, C. FRAYSSINET‡, I. TOMATIS†, J. GHISOLFI† and J. P. THOUVENOT† INSERM U-87, Université Paul Sabatier, 2 rue François Magendie, 31400 Toulouse, France, † Groupe d'Etudes en Nutrition Infantile CHU Purpan, Toulouse, France, and ‡ CNRS, Institut de Recherches Scientifiques sur le Cancer, Villejuif, France
Downloaded by [Universite Laval] at 17:31 06 November 2015
Introduction
Research and public attention have been increasingly focussed on the hypothesis that vitamins and micronutrients may decrease the incidence of some cancers (Mettlin et al. 1980, Krale et al. 1983, Birt 1986, Woutersen and GarderenHoetmer 1988). The possible role of vitamin A in chemoprevention and chemotherapy of cancer is an issue that is widely discussed but the only physiological function of the vitamin A that has been clearly denned on a molecular basis is its role in vision (Bollag 1979, Lotan 1980, Moon et al. 1983, Sporn and Roberts 1983). The study in our laboratory of two cases of human hepatoblastomas showed different qualitative and quantitative distributions of the vitamin A esters (Tomatis et al. 1988). It seemed important to verify, at an experimental level, the relationship existing between the qualitative and quantitative aspects of vitamin A metabolism during the different stages of carcinogenesis. Materials and methods
Human hepatoblastomas Three fragments drawn by partial hepatectomy were analysed in each case: fragment no. 1: the anatomy pathology showed no sign of neoplastic invasion: 'normal part', fragment no. 2: the anatomy pathology revealed a massive invasion: 'tumoural part', fragment no. 3: partially invaded: 'mixed tissue'. Animals and diets The experiments were carried out with male weanling Sprague-Dawley rats. The rats, housed in separate cages, were assigned to different dietary groups. The basal diet was the same in each group but supplemented with different levels of retinyl palmitate: group 1 (18 rats) with 1 IU/g diet, group 2 (18 rats) with 2 IU/g diet, group 3 (18 rats) with 20 IU/g diet and group 4 (18 rats) with 200 IU/g diet. Twelve rats in each dietary group were treated according to the Solt and Farber protocol (1976) viz.: aflatoxin Bi initiation at six weeks (2 mg/kgb.w., i.p.), selection by acetylamino fluorene from 8 to 10 weeks (0-02% in the diet), partial hepatectomy at 9 weeks and promotion from the 1 lth week by phenobarbital in the 0265-203X/90 $3.00© 1990 Taylor & Francis Ltd.
S70
A. Periquet et al.
Downloaded by [Universite Laval] at 17:31 06 November 2015
drinking water (1 g/1). The six rats left in each group constituted the control animals. At 4 months post initiation, the rats were killed by decapitation. Blood was collected for analysis of plasma retinol according to the HPLC method of De Leenheer et al. (1979). Livers were removed, weighed and kept at 4°C. Homogenates were assayed according to our method published earlier (Periquet etal. 1985). Quantification of retinyl fatty acyl esters 7ra«s-retinol, /ra«s-retinyl palmitate and frans-retinyl acetate were purchased from Sigma Chemicals Co (St Louis, Missouri, USA). The fatty acyl chlorides were obtained from Aldrich Chemical Co Inc (Milwaukee, Wisconsin, USA). The retinyl ester standards (laurate, myristate, palmitoleate, pentadecanoate, linoleate, heptadecanoate, oleate and stearate) (were synthesized by reaction of trans-vttmoX with the corresponding acyl chloride as described by Huang and Goodman (1965) with slight modifications (Periquet etal. 1988). Retinyl esters present in the liver lipid extract were separated and quantified using an HPLC technique as described earlier (Periquet etal. 1988). Results In this article are reported the results obtained with the human hepatoblastomas and the experimental results obtained with the control and treated rats receiving a normal daily dose of vitamin A: 20 IU/g diet. The study of two cases of human hepatoblastomas showed: (1) at a quantitative level: a very high vitamin A level in the liver part considered healthy and especially in the mixed part after anatomopathological examination and, in contrast, a very low vitamin A level in the cancerous part (table 1). (2) at a qualitative level, a difference in the stocking profile with, in particular, an increase in all the minor forms at the expense of the retinyl palmitate (table 2). As for the results for the animals, table 3 shows that for the same daily contribution in vitamin A (20 IU/g diet as retinyl palmitate) there was four times less vitamin A in the treated rats' livers than in the controls' livers after twelve weeks of promotion. On the other hand, circulating vitamin A levels were comparable and stayed within normal limits. At 4 months post-initiation, at a qualitative level, no significant difference was noted between the stocking profile and the healthy human liver. The results illustrated in table 4 show that retinyl palmitate/oleate constituted the most important ester form (78-84%) of the total retinyl fatty acyl esters followed by stearate (11-15%), laurate, palmitoleate/myristate, linoleate, heptadecanoate and pentadecanoate totalling 6%. Table 1. Retinyl palmitate contents of a human liver tumour.
Total homogenate (retinyl palmitate) Hg/g
Normal part
Mixed tissue
Tumourous part
no. 1
no. 3
no. 2
663
1177
195
Effect of dietary retinyl palmitate on tumours
S71
Table 2. Distribution of the esters of retinol (%) in the normal human liver and in the healthy part of a cancerous human liver. Retinyl esters (%)
Normal human liver
Downloaded by [Universite Laval] at 17:31 06 November 2015
Normal part of cancerous human liver
C16:0
C18:0
C12.-0
80-85
10-15
0-4-0-6
65-70
9-12
0-4-0-7
C14:0 and C16:l
C18:2
C15:0
C17:0
1-0-2
0-7-1
0-2-0-6
traces
2-5
2-6
0-2-0-7
5-10
Table 3. Serum concentration of vitamin A and retinyl fatty acyl esters in liver homogenate of rats vitamin A sufficient diets (20 IU/g diet) for up to 4 months post initiation. Serum vit A (mg/1)
C16:0
C18:0
C12:0
C14:0 and C16:l
Control rats
0-42 ±0-087
504-25 ± 30-2
90-64 ± 5-6
4-51 ±0-38
Treated rats n=12
0-56 ±0-055
141-47 ± 16-12
22-38 ± 1-72
1-18 ±0-25
C18:0
C15:0
C17:0
4-70 ±0-59
23-6 ± 1-80
2-53 ±0-42
8-70 ±1-05
2-68 ±0-30
1-92 ±0-08
1-80 ±0-05
2-44 ±0-60
Table 4. Distribution of the esters of retinol (%) in the normal and treated rats' liver (4 months post-initiation). Retinyl esters (%)
C16:0
C18:0
C12:0
C14:0 and C16:l
C18.-2
C15.-0
C17.-0
Control livers
78-84
11-15
0-6-0-8
1-0-1-3
1-2-4
0-3-0-4
1-2-2-4
Treated livers
75-84
11-18
0-4-0-6
1-2-5
0-5-2
0-2-1-9
1-4-2-4
Discussion and conclusion The starting point of this work was the determination of vitamin A and its different esters in some cases of human hepatoblastomas. The results seem to indicate that there is accumulation of vitamin A especially in the mixed zone whereas there is a non-negligible proportion of tumoural cells a priori vitamin A deficient. This 'barrier phenomenon', with a very high vitamin A level separating the cancerous tissue from the healthy one, has not, to our knowledge, been reported. Muto and Omori (1981) made comparable observations concerning the low vitamin A level in the tumour; the hypothesis advanced is a perturbation in the
S72
A. Periquet et al.
retinol membrane transport from the surrounding healthy tissue and/or for a much faster utilization rate in the tumour cells. De Luca et al. (1984) showed that the nuclear membranes coming from carcinomas are depleted in retinol compared with the nuclear membranes of the healthy hepatic cells; they concluded that the hepatic stocking function for vitamin A has been lost in the tumour. We have tried to obtain similar results in animal experiments by initiating tumours by the Solt and Farber protocol. Our first results obtained in the animals seem to confirm these results and point out an increased vitamin A consumption during a Solt and Farber carcinogenesis process; they consolidate the defined hypothesis: modification of the vitamin A membrane transport in the tumourous tissues, a loss of the stocking characteristic and/or an increased use of this micronutrient by the tumour cells.
Downloaded by [Universite Laval] at 17:31 06 November 2015
References BIRT, D. F., 1986, Update on effects of vitamin A, C. and E and selenium on carcinogenesis. Proceeding of the Society for Experimental Biology and Medicine, 183, 311 -320. BOLLAG, W., 1979, Retinoids and cancer. Cancer Chemotherapy and Pharmacology, 2, 207-215. HUANG, H. S., and GOODMAN, D. S., 1965, Vitamin A and carotenoids I. Intestinal absorption and
metabolism of 14C-labelled vitamin A alcohol and β-carotène in the rat. Journal of Biological Chemistry, 240, 2839-2844. KRALE, G., BJELKE, E., and GART, J. J., 1983, Dietary habits and lung cancer risk. International Journal of Cancer, 31, 397-405. DE LEENHEER, A. P . , DE BEVERE, V., DE RUYTER, M. G. M., and CLAEYS, A. E., 1979, Simultaneous
determination of retinol and tocopherol in human serum by HPLC. Journal of Chromatography, 162, 408-412. LOTAN, R., 1980, Effects of vitamin A and its analogs (retinoids) on normal and neoplastic cells. Biochemistry et Biophysica Acta, 605, 33-91. DE LUCA, L. M., BRUGH, M., and SILVERMAN, J. C. S., 1984, Retinyl palmitate, retinyl phosphate and dolichyl phosphate of post nuclear membrane fraction from hepatoma host liver and regenerating: marginal vitamin A status of hepatoma tissue. Cancer Research, 44, 224-232. METTLIN, C., GRAHAM, S., PRIORE, R., MARSHALL, J., and SWANSON, M., 1980, Diet and cancer of the
esophagus. Nutrition and Cancer, 2, 143-147. MOON, R. C., MCCORMICK, D. L., and MEHTA, R. G., 1983, Chemoprevention of animals tumors by retinoids. In Modulation and Mediation of Cancer by Vitamins, edited by F. L. Meykens and K. N. Prasad (Basel: Karger) pp. 47-57. MUTO, Y., and OMARI, M., 1981, A novel cellular retinoid binding protein, f-type, in hepatocellular carcinoma. Annals of the New York Academy of Science, 359, 91-103. PERIQUET, B., BAILLY, A., GHISOLFI, J., and THOUVENOT, J. P., 1985, Determination of retinyl
palminate in homogenates and subcellular fractions of rat liver by liquid chromatography. Clinica Chimica Acta, 147, 41-49. PERIQUET, B., LAMBERT, W., BAILLY, A., TOMATIS, I., GHISOLFI, J., DE LEENHEER, A. P . , and
THOUVENOT, J. P . , 1988, Fatty acid composition and kinetic behaviour of liver retinyl esters in vitamin A sufficient and deficient rats. Clinica Chimica Ata, 172, 275-290. SOLT, D., and FARBER, E., 1976, New principle for the analysis of chemical carcinogenesis. Nature, London, 263, 701-703. SPORN, M. B., and ROBERTS, A. B., 1983, Role of retinoids in differenciation and carcinogenesis. Cancer Research, 43, 3034-3040. TOMATIS, I., FRAYSSINET, C , PERIQUET, A., PERIQUET, B., GHISOLFI, J., and THOUVENOT, J. P . , 1988,
Etude des différents esters de retinol des hepatoblastomes humains. Compte rendu des Journées du Colloque 'Ailment 2000' Janvier 1989, Ministère de la Recherche et de la Technologie, Paris, dans: Bilan 'Ailment 2000'. WOUTERSEN, R. A., and GARDEREN-HOETMER, A. M., 1988, Inhibition of dietary fat-promoted development of (pre) neoplastic lesions in exocrine pancreas of rats and hamsters by supplemental vitamins A, C and E. Cancer Letters, 41, 179-189.
