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1 J. Nutr. Sci. Vitaminol., 33, 31-36, 1987 Effect of Milk Protein and Fat Intake on Blood Pressure and the Incidence of Cerebrovascular Diseases in Stroke-Prone Spontaneously Hypertensive Rats (SHRSP) Katsumi IKEDA,1 Shinichi MOCHIZUKI,2 Yasuo NARA,1 Ryoichi HORIE,1 and Yukio YAMORI1 1 Department of Pathology, Shimane Medical University and Japan Stroke Prevention Center, Enya-cho, Izumo 693, Japan 2 Research Institute of Life Science, Snow Brand Milk Products, Co., Ltd., Shimotsuga-gun, Tochigi , Japan (Received June 2, 1986) Summary The intake of two milk protein-rich diets containing casein and whey protein attenuated the development of severe hypertension in stroke-prone spontaneously hypertensive rats (SHRSP), and extended their life span in comparison with SHRSP on a regular stock diet. Milk fat-rich diet intake reduced the incidence of cerebrovascular disease in SHRSP without a significant fall in blood pressure. These results suggest that certain milk components have a preventive effect on hypertension and cerebrovascular disease in SHRSP. Key Words SHRSP, blood pressure, stroke, whey protein, casein, milk fat The importance of the interaction between genetic and environmental factors in the pathogenesis of stroke in stroke-prone spontaneously hypertensive rats (SHRSP) has been well documented (1,2). We have previously shown that certain dietary factors can prevent the development of severe hypertension and cerebrovas cular diseases in SHRSP (3,4). The intake of fish and soybean was found to be particularly effective in protecting against such diseases in this rat strain (5). Given that increased longevity is associated with not only an increased fish and soybean intake but also increased milk consumption in certain regions in Japan and in the world, in the present study we have examined the effect of certain cow's milk components on the development of hypertension and cerebrovascular diseases in SHRSP. MATERIALS AND METHODS Three types of diets (casein-rich, whey protein-rich and milk fat-rich) were 31

2 32 K. IKEDA et al. Table 1. Components of experimental diets.(%) prepared for this study. The components of these diets were as follows: the casein rich diet (CD) contained 14.6% milk casein in the Funahashi SP diet (SP) which is the regular stock diet for SHRSP in our colony (6); the whey protein-rich diet (WPD) contained 17.0% whey protein in the SP diet; and the milk fat-rich diet (MFD) contained 20% milk fat in the SP diet. Total calories of SP, CD, WPD and MFD were 4.14, 4.09, 4.10, and 5.11 kcal/g, respectively (Table 1). In Experiment 1, sixteen 7-week-old SHRSP were divided into 3 groups that were fed on CD, WPD or SP diet, respectively. In Experiment 2, SHRSP were divided into 2 groups consisting of more than 6 rats in each group at the age of 12 weeks; the diet for one group was changed from SP to MFD while the other group remained on SP. Body weight and tail cuff blood pressure were measured monthly in each group until natural death. The rats were autopsied after death for routine histopathologi cal examination to determine the incidence of cerebrovascular diseases. Statistical differences in blood pressure and the incidence of cerebrovascular diseases were analyzed by Student's t-test and ƒô2 test, respectively. RESULTS Experiment 1: In the groups fed the CD or WPD diet the development of severe hypertension was attenuated and blood pressure was significantly decreased compared with the control group on the SP diet. There were no significant differences in body weight among these groups until the age of 150 days when the body weight of control rats was significantly greater than that of the CD and WPD groups (Fig. 1). Life span in the CD and WPD groups was significantly extended when compared to that in the control group (Fig. 2). Experiment 2: The incidence of cerebrovascular diseases in the groups fed on the MFD and SP diets were 1/7 and 5/6, respectively, with a significant difference between the 2 groups (p<0.05) (Table 2). In the group on the SP diet, two rats showed cerebral hemorrhage and softening, and three other rats had cerebral softening with or without cyst formation. However, blood pressure and life span in J. Nutr. Sci. Vitaminol.

3 EFFECT OF MILK PROTEIN AND FAT 33 Fig. 1. Blood pressure and body weight in SHRSP on casein-rich (CD) or whey protein diet (WPD). Upper panel, blood pressure; lower panel, body weight. Fig. 2. Life span of SHRSP on casein-rich or whey protein-rich diet. the MFD group were not significantly different from the age-matched control group on the SP diet and body weight in the former was greater than in the latter (Table 2, Fig. 3). Vol. 33, No. 1, 1987

4 34 K. IKEDA et al. Table 2. Life span and the incidence of cerebrovascular diseases in SHRSP fed on milk fat-rich diet (MFD). a Significant difference from control group on SP diet (p<0.05). Fig. 3. Blood pressure and body weight in SHRSP on milk fat diet. Upper panel pressure; lower panel, body weight., blood The mean intakes of the SP, CD, WPD, and MRD diets were 22.4 }0.46, 21.4 }0.81, 22.4 }0.93, and 21.4 }0.87g/day/rat, respectively significant difference among the dietary groups., and there was no DISCUSSION Both casein and whey proteins derived from milk effectively suppressed the J. Nutr. Sci. Vitaminol.

5 EFFECT OF MILK PROTEIN AND FAT 35 development of severe hypertension in SHRSP and extended their life span, although these diets contained a lower concentration of protein than the fish protein diet. In our previous reports, we demonstrated that the preventive effect of fish or soybean protein diets on stroke could be ascribed to certain amino acids in the protein, such as taurine and methionine in fish protein and lysine in soybean protein (1-5). These sulfur containing amino acids and lysine are contained in casein and whey proteins, and the methionine concentration in the SP, WPC, CD and the fish protein diets were as follows: 0.30, 0.54, 0.60, 0.84g/100g, respectively. Thus the amino acid compositions in casein and whey protein may have partly contributed to the attenuation of severe hypertension and longevity in SHRSP fed on casein and whey proteins in this study. The preventive effect of protein-rich or amino acid-rich diets in SHRSP may be attributed to the diuretic or saliuretic effect of urea (7), central suppression of sympathetic nervous system activity by sulfur amino acids (8) and the preservation of vascular elasticity due to long-term protein feeding (3). The interaction of high protein, the source of protein and the content of methionine in the WPC and CD diets may have resulted in the attenuation of the development of severe hypertension in SHRSP. The reduced body weight observed in protein-fed SHRSP over 250 days was not the cause of blood pressure reduction because reduced blood pressure was observed from the age of 160 days and thereafter. While milk fat did not affect blood pressure, it significantly reduced the incidence of cerebrovascular disease in SHRSP. This finding suggests that the intake of milk fat may change vascular wall characteristics in small cerebral arteries in SHRSP, as previously demonstrated in SHRSP on soybean protein-rich diets which reduced the incidence of cerebrovascular diseases without a significant fall in blood pressure (3). We have recently reported that palmitoleic acid (POA, C16:1), which is relatively rich in milk fat, reduced the incidence of stroke in SHRSP even while on high-salt diets (9). The mechanism by which POA reduces the incidence of stroke appears to be independent of the antithrombotic activity of eicosapentaenoic acid and other fatty acids which decrease the incidence of stroke by reducing vascular reactivity and blood pressure (10). REFERENCES 1) Yamori, Y. (1981): Environmental influences on the development of hypertensive vascular diseases in SHR and related models, and their relation to human diseases, in New Trends in Arterial Hypertension, ed. by Worcel, M., Bonvalet, J. P., Langer, S. Z., Menard, J., and Sassard, J., Elsevier, Amsterdam, pp ) Yamori, Y. (1983): Physiopathology of the various strains of spontaneously hyper tensive rats, in Hypertension, ed. by Genest, J., Kuchel, O., Hamet, P., and Cantin, M., McGraw-Hill, Montreal, pp ) Yamori, Y., Hone, R., Ikeda, K., Nara, Y., and Lovenberg, W. (1979): Prophylactic effect of dietary protein on stroke and its mechanisms, in Prophylactic Approach to Hypertensive Diseases, ed. by Yamori, Y., Lovenberg, W., and Freis, E. D., Raven Vol. 33, No. 1, 1987

6 36 K. IKEDA et al. Press, New York, pp ) Yamori, Y., Hone, R., Ohtaka, M., Nara, Y., Ooshima, A., and Fukase, M. (1979): From scientific detection of predisposition to prevention of hypertensive diseases recent progress in animal models, in Prophylactic Approach to Hypertensive Diseases, ed. by Yamori, Y., Lovenberg, W., and Freis, E. D., Raven Press, New York, pp ) Yamori, Y., Hone, R., Nara, Y., Kihara, M., Fujiwara, K., Nabika, T., Mano, M., and Ikeda, K. (1984): Nutritional causation and prevention of cardiovascular diseases -experimental evidence in animal models and man, in Nutritional Prevention of Cardiovascular Disease, ed. by Lovenberg, W., and Yamori, Y., Academic Press, Orlando, pp ) Yamori, Y., Hone, R., Tanase, H., Fujiwara, K., Nara, Y., and Lovenberg, W. (1984): Possible role of nutritional factors in the incidence of cerebral lesions in stroke-prone spontaneously hypertensive rats. Hypertension, 6, ) Wang, H., Ikeda, K., Kihara, M., Nara, Y., Hone, R., and Yamori, Y. (1984): Effect of. dietary urea on blood pressure in spontaneously hypertensive rats. Clin. Exp. Pharmacol. Physiol., 11, ) Yamori, Y., Nara, Y., Hone, R., Ooshima, A., and Lovenberg, W. (1981): Pathophysiological role of taurine in blood pressure regulation in stroke-prone spontaneously hypertensive rats (SHR), in The Effects of Taurine on Excitable Tissues, ed. by Schaffer, S. W., Baskin, S. L., and Kocsis, J. J., Chapter 27, Spectrum Publications, Inc., New York, pp ) Yamori, Y., Nara, Y., Tsubouchi, T., Sogawa, Y., Ikeda, K., and Hone, R. (1986): Dietary prevention of stroke and its mechanisms in stroke-prone SHR (SHRSP) - preventive effect of dietary fiber and palmitoleic acid- J. Hypertension, 4 (Suppl.), pp ) Yamori, Y., and Hone, R. (1977): Vascular reactivity in pathological states, in Factors Influencing Vascular Reactivity, ed. by Shibata, S., and Carvier, P., Igaku-Shoin, Tokyo, pp J. Nutr. Sci. Vitaminol.

Dietary Protein, but Not Carbohydrate, Is a Primary Determinant of the Onset of Stroke in Stroke-Prone Spontaneously Hypertensive Rats

Dietary Protein, but Not Carbohydrate, Is a Primary Determinant of the Onset of Stroke in Stroke-Prone Spontaneously Hypertensive s Tsuyoshi Chiba, PhD; Tatsuki Itoh, PhD; Masaki Tabuchi, PhD; Takao Satou,