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B9 Dietary calcium attenuates platelet aggregation and intracellular Ca2+-mobilization in spontaneously hypertensive rats
AJH-APRIL 1997-VOL. 10, NO. 4, PART 2
ASH XII ABSTRACTS
B9
B1O
DIETARY CALCIUM ATTENUATES PLATELET AGGREGATION AND INTRACELLULAR Ca2+MOBILIZATION fN SPONTANEOUSLY HYPERTENSIVE RATS Keiichi Otsuka, M Watanabe, Q Yue, DA McCarron*, D Hatton*. Division of Nephrology and Hypertension, Department of Medicine, Oregon Health Sciences University, Portland, OR. Spontaneously hypertensive rats (SHR) are known to he blood pressure-sensitive to dietary calcium. The effects ofdietay calcium on platelet aggregation (AG) and intracellular Ca’+ ([Ca’+]i) mobilization were assessed by tttrbidimetric methods and fura-2 methods respectively in washed platelets of SHR. As shown in Table, 6 weeks of dietary calcium supplementation attenuated the increase of systolic blood pressure (SBP) and thromhin-induced platelet aggregation (Representative experiments are shown in Fig.) at 9 weeks of age. The ionomycin-induced [Ca’+]i peak 100
ABNORMAL VASCULAR REACTIVITY rN EXPERIMENTAL DIET-INDUCED DrABETES (C57BU6J MICE), J.-B. Roullet’, H. Xue, J.C. Chapman, C.M. Roullet and D.A. McCarron, O.H.ELI., Portland, OR, 97201. When fed a high-fat, high-carbohydrate diet (HFC), C57BU6J mice become obese and hypertensive, and exhibita several of the metabolic disturbances associated with non-insulin dependent (type 11)diabetes: elevated glucose and insulin levels and dyslipidemia. However, the functional properties of their arteries have not been characterized. Therefore, we studied the reactivity of smalr arteriea w isolated from HFC-fad (18 ... weeks) C57BU6J mice by —m m comparison with arteries ,, 0 ‘=~ isolated from normal-chow m ... 9 MJ fed (NFC) animara (n = 12/ ● Non-susceptible , ‘~.. ‘ 1 a 8 ....@ #;;p/~J) micewere arso s .. atudied in order to m determine the interaction k
~
whichreflects[Ca’+] istorag~~ize, ‘n*eabsen’eOfex’ema’ ICa’+]i ca’ release andthmmbin-evoked from [Ca’+]istorage were decreased (Table). In addition, SBP was positively correlated with platelet aggregation (r= .703, P = .0088), a thrombin-evoked [Ca’+]i (r = .739, 01 P = .0044) and ionomycin-induced 0 time (rein) 5 [Ca’+]i (r = .591, P= .0415), respectively. These results suggest that dietary calcium supplementation had a beneficial effect on platelets of SHR by attenuating [Ca’+]i mobilization from [Ca’+]i storage. The hypotensive effect of dietary calcium might be associated with attenuated [Ca’+]i mobilization in SHR. ;;,
0“2;=:
SBP (mmHg)
AG at 5 min [%]
Aeonist-induced rca’+li (mzloriL) ‘ ‘ Tbrombin [onomycin
Diet Ca 9 weeks 0.2% 199 + 16 84.5+ 3.7 339 + 29 472 + 55 2.0% 170 i 9* 73.7 + 7.47 278 + 33j 370 + 23* “ = 7 m 8, Values are means+ SD. * P <.001, ‘1P <.004, I P <.002 Vs O.z%ca Key Words: spontaneously hypertensive rats, dietarycalcium, plateletaggregation,intracellularcalcium
0 .04
44
between diet and atrsin. Arteriesweremountedina
wire myograph and the GOw% norepiresponse to nephrine (NE), serotonin (5-HT), acetylcholine (Ach) and Na nitroprusside (SNP) was determined. The data show that HFC diet decreased significantly Achdependent relaxation (P
Bll
B12
DrETARYCALCIUM IMPROVES BONE MINERAL DENsrn (BMD) AND LoWER.5 BLOODpRE55URE (Bp) IN SPONTANEOUSLYHYPERTENSIVE RATS (SHR). Q&l. Bcsu!W’,E. orwoll, S. Orvmll,J.B. Roullet”,Q. Yue”, D.A. McCarmn’, D.C. Hatton*,OregonHealthSciencesUniversity, Portland,OR. An increasein calciumintakehas been proposedas a non-pharmacological treatmentof highBP. We previously reparteda decreasedintestinalC@ transpotlin SHR. Moreover,an inverserelationshiphas been observed betweenbonemineralcontentand BP. We postulatedthat dietaryCa’+ improvesCa** homeostasis,restoringBMD and BP to normal. We thereforemeasuredBP (mmHg), BMD (mg/area),calcitriol(D, pg/ml),PTH (pg/ml),ionized Ca’+(iCa, mM) and CSBP-9K(pg/mg, a markerof intestinal Ca2’transpoti)in SHR fed low(0.1, 0.2%), normal(1.0%) or high(2,00A)Cafi dietsfor 6 weeks (n=16/group,half SHR providedbyTaconic(T), and half by Charles River (CR). Resultswere:
KCI rNCREASES THE SEVERITY OF HYPERTENSION (HT), RENAL LESIONS AND THE fNCIDENCE OF STROKE fN THE NaClLOADED STROKE-PRONE SPONTANEOUSLY HYPERTENSIVE RAT (SHRSP). M Tanaka*, O Schmidlin, JL Olson, RC Morris Jr.* UCSF, Dept. of Medicine & Pathology, GCRC, San Francisco, CA In SHRSP (bred at University of Iowa) fed normal NaCl (0.4%), we previously reported that supplemental KC I exacerbates the severity of HT and renal lesions, whereas supplemental KHCO, attenuates both, suggesting that both are Cr.-sensitive. In the present study of the SHRSP fed high NaCl (2.6%), we investigated the effect of supplemental KCI (2%K, .=15) by comparing its effectswith those of supplemental KHCO, (2%K, n=16) and nothing (CTL, .=15) over 5 wks starting at 10 wks of age. Aortic BP was measured telemetrically every 10 minutes. The incidence of stroke was determined by clinical signs and brain lesions. UrinaW protein excretion (UP) and plasma renin activity (PRA) were measured at 14 wks. The severity of renal lesions was ranked tlom I (the least), to 46 (tbe most). We found that in alr groups NaCl bading induced greater increases in BP than those occurring at comparable times in our previous study with normal NaC1. KCI loading exacerbated the NaC1-induced increase in BP, whereas KHCO, had nO effect On Bp. At 14 wks mean SBP was significantly higher in KCI than either CTL or KHC03 (246 vs. 221 or 217mmHg, p<.01), as was mean renal histology rank (34 vs. 22 or 19, P<.01), median UP (497 vs. 94 or 117mg/day,
elssEfd QaMa
212.&t4.O BP” 1.26Kt.02 iCa” 503,%7.0 D“ 34,6il.7 CaBP’ s% BMD” PTH r 211.5-5.6 CR 134.0M6.O (+,p
Q2KQslQws
211.%3.S 1.33W.01 ls2.2&.2 26.1* 1.5 Ioais 164.= 1,8 llS.6ti4.2
2W.1*3.S 1.35i’o.ol 92.Si4.7 20.4*1. I 133W 106.*1O.2 159,7s?0.1
-
17e.&4.7 1.34M,01 56.2*3.3 17.4* 1.1 135i2 s3.4*lf.4 105.1M2.5
The followingcorrelationswere found:iCafD (PcO.0001, F-O.54); iCafPTH(p
Words:hypertension,bone density, calcium,hormones
P<.05) and the incidence of stroke (53 vs. 7 or 12.5%, p<.01). For all rats combined, mean SBP correlated strongiy with the ranked severity of renal lesions (R’=.79, P<.0001) and PRA (R2=.73, p<.0001). Stroke occurred only in rats with SBP >250 mmHg and PRA >23ng/ml/h. These results demonstrate that in the NaC1-loaded SHRSP 1) the Cl- component of KCI amplifies the pressor effect of NaCl; 2) the presser effect of Cl” overrides any antipressor effect of K’. The results suggest that in the SHRSP large increases in dietary Cl”, severity of HT and PRA are all major determinants of the severity of hypertensive nephropathy and the incidence of stroke. Key Words: SHRSP, Cl, HT, nephropathy, stroke
ASH XII ABSTRACTS
B9
B1O
DIETARY CALCIUM ATTENUATES PLATELET AGGREGATION AND INTRACELLULAR Ca2+MOBILIZATION fN SPONTANEOUSLY HYPERTENSIVE RATS Keiichi Otsuka, M Watanabe, Q Yue, DA McCarron*, D Hatton*. Division of Nephrology and Hypertension, Department of Medicine, Oregon Health Sciences University, Portland, OR. Spontaneously hypertensive rats (SHR) are known to he blood pressure-sensitive to dietary calcium. The effects ofdietay calcium on platelet aggregation (AG) and intracellular Ca’+ ([Ca’+]i) mobilization were assessed by tttrbidimetric methods and fura-2 methods respectively in washed platelets of SHR. As shown in Table, 6 weeks of dietary calcium supplementation attenuated the increase of systolic blood pressure (SBP) and thromhin-induced platelet aggregation (Representative experiments are shown in Fig.) at 9 weeks of age. The ionomycin-induced [Ca’+]i peak 100
ABNORMAL VASCULAR REACTIVITY rN EXPERIMENTAL DIET-INDUCED DrABETES (C57BU6J MICE), J.-B. Roullet’, H. Xue, J.C. Chapman, C.M. Roullet and D.A. McCarron, O.H.ELI., Portland, OR, 97201. When fed a high-fat, high-carbohydrate diet (HFC), C57BU6J mice become obese and hypertensive, and exhibita several of the metabolic disturbances associated with non-insulin dependent (type 11)diabetes: elevated glucose and insulin levels and dyslipidemia. However, the functional properties of their arteries have not been characterized. Therefore, we studied the reactivity of smalr arteriea w isolated from HFC-fad (18 ... weeks) C57BU6J mice by —m m comparison with arteries ,, 0 ‘=~ isolated from normal-chow m ... 9 MJ fed (NFC) animara (n = 12/ ● Non-susceptible , ‘~.. ‘ 1 a 8 ....@ #;;p/~J) micewere arso s .. atudied in order to m determine the interaction k
~
whichreflects[Ca’+] istorag~~ize, ‘n*eabsen’eOfex’ema’ ICa’+]i ca’ release andthmmbin-evoked from [Ca’+]istorage were decreased (Table). In addition, SBP was positively correlated with platelet aggregation (r= .703, P = .0088), a thrombin-evoked [Ca’+]i (r = .739, 01 P = .0044) and ionomycin-induced 0 time (rein) 5 [Ca’+]i (r = .591, P= .0415), respectively. These results suggest that dietary calcium supplementation had a beneficial effect on platelets of SHR by attenuating [Ca’+]i mobilization from [Ca’+]i storage. The hypotensive effect of dietary calcium might be associated with attenuated [Ca’+]i mobilization in SHR. ;;,
0“2;=:
SBP (mmHg)
AG at 5 min [%]
Aeonist-induced rca’+li (mzloriL) ‘ ‘ Tbrombin [onomycin
Diet Ca 9 weeks 0.2% 199 + 16 84.5+ 3.7 339 + 29 472 + 55 2.0% 170 i 9* 73.7 + 7.47 278 + 33j 370 + 23* “ = 7 m 8, Values are means+ SD. * P <.001, ‘1P <.004, I P <.002 Vs O.z%ca Key Words: spontaneously hypertensive rats, dietarycalcium, plateletaggregation,intracellularcalcium
0 .04
44
between diet and atrsin. Arteriesweremountedina
wire myograph and the GOw% norepiresponse to nephrine (NE), serotonin (5-HT), acetylcholine (Ach) and Na nitroprusside (SNP) was determined. The data show that HFC diet decreased significantly Achdependent relaxation (P
Bll
B12
DrETARYCALCIUM IMPROVES BONE MINERAL DENsrn (BMD) AND LoWER.5 BLOODpRE55URE (Bp) IN SPONTANEOUSLYHYPERTENSIVE RATS (SHR). Q&l. Bcsu!W’,E. orwoll, S. Orvmll,J.B. Roullet”,Q. Yue”, D.A. McCarmn’, D.C. Hatton*,OregonHealthSciencesUniversity, Portland,OR. An increasein calciumintakehas been proposedas a non-pharmacological treatmentof highBP. We previously reparteda decreasedintestinalC@ transpotlin SHR. Moreover,an inverserelationshiphas been observed betweenbonemineralcontentand BP. We postulatedthat dietaryCa’+ improvesCa** homeostasis,restoringBMD and BP to normal. We thereforemeasuredBP (mmHg), BMD (mg/area),calcitriol(D, pg/ml),PTH (pg/ml),ionized Ca’+(iCa, mM) and CSBP-9K(pg/mg, a markerof intestinal Ca2’transpoti)in SHR fed low(0.1, 0.2%), normal(1.0%) or high(2,00A)Cafi dietsfor 6 weeks (n=16/group,half SHR providedbyTaconic(T), and half by Charles River (CR). Resultswere:
KCI rNCREASES THE SEVERITY OF HYPERTENSION (HT), RENAL LESIONS AND THE fNCIDENCE OF STROKE fN THE NaClLOADED STROKE-PRONE SPONTANEOUSLY HYPERTENSIVE RAT (SHRSP). M Tanaka*, O Schmidlin, JL Olson, RC Morris Jr.* UCSF, Dept. of Medicine & Pathology, GCRC, San Francisco, CA In SHRSP (bred at University of Iowa) fed normal NaCl (0.4%), we previously reported that supplemental KC I exacerbates the severity of HT and renal lesions, whereas supplemental KHCO, attenuates both, suggesting that both are Cr.-sensitive. In the present study of the SHRSP fed high NaCl (2.6%), we investigated the effect of supplemental KCI (2%K, .=15) by comparing its effectswith those of supplemental KHCO, (2%K, n=16) and nothing (CTL, .=15) over 5 wks starting at 10 wks of age. Aortic BP was measured telemetrically every 10 minutes. The incidence of stroke was determined by clinical signs and brain lesions. UrinaW protein excretion (UP) and plasma renin activity (PRA) were measured at 14 wks. The severity of renal lesions was ranked tlom I (the least), to 46 (tbe most). We found that in alr groups NaCl bading induced greater increases in BP than those occurring at comparable times in our previous study with normal NaC1. KCI loading exacerbated the NaC1-induced increase in BP, whereas KHCO, had nO effect On Bp. At 14 wks mean SBP was significantly higher in KCI than either CTL or KHC03 (246 vs. 221 or 217mmHg, p<.01), as was mean renal histology rank (34 vs. 22 or 19, P<.01), median UP (497 vs. 94 or 117mg/day,
elssEfd QaMa
212.&t4.O BP” 1.26Kt.02 iCa” 503,%7.0 D“ 34,6il.7 CaBP’ s% BMD” PTH r 211.5-5.6 CR 134.0M6.O (+,p
Q2KQslQws
211.%3.S 1.33W.01 ls2.2&.2 26.1* 1.5 Ioais 164.= 1,8 llS.6ti4.2
2W.1*3.S 1.35i’o.ol 92.Si4.7 20.4*1. I 133W 106.*1O.2 159,7s?0.1
-
17e.&4.7 1.34M,01 56.2*3.3 17.4* 1.1 135i2 s3.4*lf.4 105.1M2.5
The followingcorrelationswere found:iCafD (PcO.0001, F-O.54); iCafPTH(p
Words:hypertension,bone density, calcium,hormones
P<.05) and the incidence of stroke (53 vs. 7 or 12.5%, p<.01). For all rats combined, mean SBP correlated strongiy with the ranked severity of renal lesions (R’=.79, P<.0001) and PRA (R2=.73, p<.0001). Stroke occurred only in rats with SBP >250 mmHg and PRA >23ng/ml/h. These results demonstrate that in the NaC1-loaded SHRSP 1) the Cl- component of KCI amplifies the pressor effect of NaCl; 2) the presser effect of Cl” overrides any antipressor effect of K’. The results suggest that in the SHRSP large increases in dietary Cl”, severity of HT and PRA are all major determinants of the severity of hypertensive nephropathy and the incidence of stroke. Key Words: SHRSP, Cl, HT, nephropathy, stroke