(Hypertension. 2001;38:e21.)
© 2001 American Heart Association, Inc.
Letters to the Editor |
Effect of Chronic Blockade of the Kallikrein-Kinin System on the Development of Hypertension in Rats
Professor of Pharmacology, Kitasato University School of Medicine
Emeritus Professor of Pharmacology, Kitasato University School of Medicine
To the Editor:
We found some incorrect statements in an article prepared by Nour-Eddine Rhaleb et al titled "Effect of Chronic Blockade of the Kallikrein-Kinin System on the Development of Hypertension in Rats,"1 and we consider it our duty to point out the errors (and other related irregularities) and to provide the correct information for the readers of Hypertension. It should be mentioned at the outset that Dr Majima was invited to Dr Rhaleb’s laboratory by the director of that laboratory, Dr Oscar A. Carretero, to conduct the experiments in question.
In the Discussion section (pp.126, right column, ll. 28 to 38), appears the following statement": "(1) normal BN ]Brown Norway] rats were given a subcutaneous infusion of Ang II or high salt diet alone or combined with icatibant (given intraperitoneally) and (2) BNK ]Brown Norway Katholiek] rats were given a subcutaneous infusion of a nonpressor dose of Ang II or high salt diet...we were unable to reproduce any of their findings or those of Madeddu et al."1 However, the only 2 experiments that Majima performed with Dr Rhaleb, during a stay of nearly 1.5 months, focused solely on the effects of high salt diet in normal BN rats and on those of the infusion of Ang II in kininogen-deficient BNK rats and normal BN rats. Majima’s work with Dr Rhaleb (as far as it is possible to judge) confirmed our original results. Accordingly, the statement "we were unable to reproduce any of their findings" is not correct, and we can find no justification for it.
In addition, there is no explanation in the text of why the mean blood pressure during week 1 was lower (by 
7 mm Hg) than that in the pretreatment period. According to our data gathered over a period of >10 years, the systolic blood pressure of mutant kininogen-deficient BNK rats (130±2 mm Hg) is not different from that of normal BN Kitasato rats (127±2 mm Hg), and the blood pressure of these rats showed no essential change during vehicle infusion (as originally published in Hypertension, Table 1).2
According to the "Instructions to Authors" of the recent issue of Hypertension, for any acknowledgments, a statement "signed by the corresponding author, stating that those acknowledged in the manuscript have seen and approve of the manner in which their names are mentioned" is required. However, Majima was not asked for permission to include his name in the Acknowledgments section nor was he given an opportunity to see the manuscript before its submission (or acceptance) for publication. His repeated requests to the editor-in-chief to be sent a copy of the statement sent by the authors to the editor-in-chief have not generated any response whatsoever.
References
1.
Rhaleb NE, Yang XP, Nanba M, Shesely EG, Carretero OA. Effect of chronic blockade of the kallikrein-kinin system on the development of hypertension in rats. Hypertension. 2001; 37: 121–128.
2.
Majima M, Mizogami S, Kuribayashi Y, Katori M, Oh-ishi S. Hypertension induced by a nonpressor dose of angiotensin II in kininogen-deficient rats. Hypertension. 1994; 24: 111–119.
Response
Hypertension and Vascular Research Division, Henry Ford Hospital, Detroit, Michigan
We are responding to several points made by Dr Majima in regard to our recent article in Hypertension.1
(1) Dr Majima wanted us to provide an explanation for the 7 mm Hg drop in mean blood pressure (MBP) from the pretreatment period to Week 1 (Figure 1 in our article). This request was surprising, because (a) the difference was not statistically significant, (b) it’s usual to observe such a variation in BP within a few days after surgically installing the telemetry device, as previously reported,2 and (c) by the end of Week 1 and 2 we were expecting instead a 40- to 50-mm Hg increase in BP as reported by Dr. Majima’s group.3
As we stated in our article, MBP was 115±3 mm Hg in normal Brown Norway rats compared with 118±3 mm Hg in Brown Norway Katholiek (BNK) rats. Hence, there is also no difference between the 2 strains in terms of BP, similar to Dr Majima’s data.3 Our BP values were about 12 to 14 mm Hg higher than Dr Majima’s, but that’s normal because we measured BP in the United States with our equipment, whereas Dr Majima’s were done in Japan with their equipment.
(2) As we indicated in the article, Dr Majima visited our laboratory in an attempt to reconcile the different findings between our study and his, but we were not able to do so. We included him in the acknowledgments as a way of expressing our gratitude for his generous assistance. Thus, there is no letter to this effect, as we merely wished to acknowledge his courtesy to us. It does not seem logical to me that it would be necessary for us to first write to Dr Majima and ask for his permission to thank him for helping us. Moreover, the Editorial Board of Hypertension never asked us to provide them with any documentation regarding this matter.
In our article we stated ". . .(1) normal BN rats were given a subcutaneous infusion of Ang II or high salt diet alone or combined with icatibant . . . and (2) BNK rats were given a subcutaneous infusion of a nonpressor dose of Ang II or high salt diet. . . we were unable to reproduce any of their findings or those of Madeddu et al." There was a mistake, and we should have stated "(1) normal BN rats were given high salt diet alone or combined with icatibant . . . and (2) BNK rats were given a subcutaneous infusion of a nonpressor dose of Ang II or high salt diet. . . we were unable to reproduce any of their findings or those of Madeddu et al." Dr Majima did not participate with us in the protocol dealing with the effect of icatibant infusion on the pressor effect of Ang II in normal BN rats. Dr Majima stated that the original results were confirmed while participating with us on a protocol involving BNK rats infused with vehicle or subpressor dose of Ang II. That is wrong, because when we administered a subpressor dose of Ang II, systolic blood pressure was slightly higher in BNK rats compared with rats given vehicle; however, direct arterial MBP in awake and restrained rats was similar in both groups. For that reason, we opted for telemetry and explained the rationale for its use in our paper.
References
1.
Rhaleb N-E, Yang X-P, Nanba M, Shesely EG, Carretero OA. Effect of chronic blockade of the kallikrein-kinin system on the development of hypertension in rats. Hypertension. 2001; 37: 121–128.
2.
Butz GM, Davisson RL. Long-term telemetric measurement of cardiovascular parameters in awake mice: a physiological genomics tool. Physiol Genomics. 2001; 5: 89–97.
3.
Majima M, Mizogami S, Kuribayashi Y, Katori M, Oh-Ishi S. Hypertension induced by a nonpressor dose of angiotensin II in kininogen-deficient rats. Hypertension. 1994; 24: 111–119.
Role of Kinins in Blood Pressure Regulation: Reality or Fiction
Gene Therapy Section, INBB National Laboratory, Osilo, Italy
To the Editor:
We read with interest the article entitled "Effect of Chronic Blockade of the Kallikrein-Kinin System on the Development of Hypertension in Rats" by Rhaleb et al.1 By a pharmacological approach, the authors evaluated if endogenous kinins can mitigate hypertension induced by Ang II or mineralocorticoids. Although unable to confirm the hypothesis under challenge, the authors showed that icatibant prevents the early antihypertensive effect induced by ramipril in uninephrectomized deoxycorticosterone acetate (DOCA)-salt treated rats. Thus, at least under ACE-inhibition, kinins may become important in contrasting mineralocorticoid-induced hypertension.
Recent availability of selective kinin antagonists and genetically modified animals lacking or overexpressing components of the kallikrein-kinin system (KKS) has renewed the interest in the area. However, enthusiasm generated by positive reports2,3 has been criticized by Rhaleb et al,1 unable to confirm the higher BP of kinin B2 knockout mice (B2-KO) and the enhanced pressor response to Ang II or mineralocorticoids in animals with interrupted kinin receptor signaling. Here, we report our concern regarding the sensitivity of these negative studies. In experiments aimed to challenge Ang II action under icatibant, Rhaleb et al used groups of 4 to 5 animals. Considering the small sample size and the large over-time BP variability in controls (20 mm Hg), it can be calculated that the power of the test barely reaches 0.28. Consequently, because of the low sensitivity of measurements, only astonishing BP changes caused by kinin B2 blockade would be significant. Under these conditions, a negative result may configure a type 1 error. Rhaleb et al also refer to a previous work of theirs, affirming that B2-KO are not sensitive to mineralocorticoids.4 The authors showed that BP was increased by 27 mm Hg in DOCA-treated B2KO versus 16 mm Hg in controls (P=0.07). They conclude that the absence of the receptor does not make the difference. However, we argue that with such a level of probability, no strong conclusion can actually be reached. Furthermore, Rhaleb has superimposed high salt and uninephrectomy to DOCA-treatment. This is at variance with our approach, because we preferred to avoid any maneuver that could suppress kallikrein and mask the importance of the system in mineralocorticoid-induced hypertension.5 Thus, we regret to say that Rhaleb’s methodology differs from ours and that his reference to our article is partial and incorrect.
Quite singular is Rhaleb’s tendency of unequally emphasizing the points of agreement and disagreement in the field. The findings from either laboratory are consistent regarding the issue that acute or chronic administration of kinin antagonists does not alter the cardiovascular phenotype of rodents under conditions of salt loading. Furthermore, agreement exists regarding sodium-sensitivity of B2-KO, although our strain was more prone to the toxic effect of high salt.2,6 This prompted us to formulate the hypothesis that genetic drift was causing phenotypic changes in small-size colonies present at various laboratories.7
The variability of B2-KO cardiovascular phenotype seems to confirm our opinion. At variance with Rhaleb’s data, Gavras’ group8 and ourselves2 documented that B2-KO have higher BP and display an enhanced compensatory statement of inducible B1 receptor. Eventually, B2-KO develop heart remodeling, myocardial damage, and failure with aging.2 Sensitivity of B2-KO to Ang II or renin-dependent hypertension was documented by us and Cervenka9 but negated by others.1 The theory of genetic drift driving B2-KO toward fixation of divergent phenotypes was reinforced by the observation that one mutant line developed testicular theratomas at our laboratory (Madeddu P, unpublished observations), a feature typical of some J129-substrains. The absence of the above pathology in other related lines of B2-KO favors the possibility of a nonhomogeneous genetic background among mutants. In line with this theory, Dr Milia, an investigator of my team visiting Max Delbruck Center (Berlin, Germany), was not able to find any alteration in the cardiovascular phenotype of B2-KO (J129 genetic background) derived from a breeding pair that I personally supplied to that institution.10 In our laboratory, we have confirmed these negative results on the progeny of the German stock kindly returned to us by Prof Luft. Interestingly enough, during the same set of measurements, B2-KO mice of a different background (C57/BL6) showed elevated BP and HR (118±3 mm Hg and 628±12 bpm versus 100±3 mm Hg and 531±17 bpm in wild type, P<0.01) and ventricular hypertrophy (Madeddu et al, unpublished observations). Our observation that the hypertensive, hypertrophic phenotype of B2-KO is reproduced in a different genetic background favors a role of B2 receptor signaling in the regulation of cardiovascular function.
Altogether these results indicate that (1) given the redundancy of mechanisms controlling BP, the contribution of kinins may be detected only using high-power experimental strategies; and (2) the lack of phenotypic homogeneity of B2-KO may be attributable to random fixation of alleles caused by genetic drift. Microarray analysis could be extremely useful in elucidating the role of otherwise unappreciated mechanisms responsible for the variability among related substrains.10 Out of these reasons, we hope that apparently conflicting results and queries will not discourage more in depth research in the field of KKS.
References
1.
Rhaleb NE, Yang XP, Nanba M, Shesely EG, Carretero OA. Effect of chronic blockade of the kallikrein-kinin system on the development of hypertension in rats. Hypertension. 2001; 37: 121–128.
2. Emanueli C, Maestri R, Corradi D, Marchione R, Minasi A, Tozzi MG, Salis MB, Straino S, Capogrossi MC, Olivetti G, Madeddu P. Dilated and failing cardiomyopathy in bradykinin B2 receptor knockout mice. Circulation. 1999; 100; 2359–2365.[Medline] [Order article via Infotrieve]
3. Madeddu P, Emanueli C, Maestri R, Salis MB, Minasi A, Capogrossi MC, Olivetti G. Angiotensin II type 1 receptor blockade prevents cardiac remodeling in bradykinin B2 receptor knockout mice. Hypertension. 2000; 35; 391–396.[Medline] [Order article via Infotrieve]
4.
Rhaleb NE Peng H, Alfie ME, Shesely EG, Carretero OA. Effect of ACE inhibitor on DOCA-Salt- and Aortic coartation-induced hypertension in mice. Do kinin B2 receptors play a role? Hypertension. 1999; 33: 329–334.
5. Madeddu P, Anania V, Pinna Parpaglia P, Demontis MP, Varoni MV, Fattaccio MC, Glorioso N. Chronic receptor blockade induces hypertension in deoxycorticoserone-treated rats. Br J Pharmacol. 1993; 108: 651–657.[Medline] [Order article via Infotrieve]
6. Alfie ME, Yang XP, Hess F, Carretero OA. Salt sensitive hypertension in bradykinin B2 receptor knockout mice. Biochem Biophys Res Commun. 1996; 224: 625–630.[Medline] [Order article via Infotrieve]
7. Madeddu P, Emanueli C. Can knockout mice help dissect relevant genes in hypertension? Evidence and confounding factors. Hypertension. 1999; 34; e14–15.[Medline] [Order article via Infotrieve]
8.
Duka I, Kintsurashvili E, Gavras I, Johns C, Bresnahan M, Gavras H. Vasoactive potential of the B1 bradykinin receptor in normotension and hypertension. Circ Res. 2001; 88: 275–281.
9. Cervenka L, Maly J, Karasova L, Simova M, Vitko S, Hellerova S, Heller J, El-Dahr SS. Angiotensin II-induced hypertension in bradykinin B2 receptor knockout mice. Hypertension. 2001; 37; 967–973.[Medline] [Order article via Infotrieve]
10.
Milia AF, Gross V, Plehm R, De Silva JAJr, Bader M, Luft FC. Normal blood pressure and renal function in mice lacking the bradykinin B2 receptor. Hypertension. 2001; 37: 1473–1479.
Response
Hypertension and Vascular Research Division, Henry Ford Hospital, Detroit, Michigan
We have shown that kinins play a role in the antihypertensive effect of ACE inhibitors in mineralocorticoid hypertension in mice1 and rats,2 but only during the early phase of treatment (1 week); they do not participate in the long-term antihypertensive effect of ACE inhibition (4 to 6 weeks) in hypertension induced by mineralocorticoids in mice1 or by aortic coarctation in mice1 and rats.3
In our recent study, we found that chronic blockade of B2 kinin receptors did not enhance the effect of chronic infusion of a subpressor dose of angiotensin II,2 as shown by 2 separate experimental protocols. In one of these protocols, involving 5 to 7 rats per group, variation was minimal, while in the other we only observed marked variation at 2 points by the end of drug infusion.2 There was no tendency toward an enhanced pressor response to angiotensin II in rats chronically infused with a B2 kinin receptor antagonist.
Madeddu suggests that uninephrectomy could suppress kallikrein activity and mask the important role of the kallikrein-kinin system in mineralocorticoid hypertension, which is not supported by the findings of other authors.4,5 Indeed, those authors found that kallikrein activity more than doubled in rats that were uninephrectomized and given high salt plus deoxycorticosterone (DOCA-salt). Therefore, we took those findings into account when we designed our study on the role of kinins in DOCA-salt hypertension.
Madeddu used data recently published by Gavras’s group as a confirmation of his previous finding that kinin B2 knockout (B2-KO) mice are hypertensive.6,7 However, the blood pressures (BP) reported by Gavras’s group were only slightly higher and in some cases no different between B2-KO and control mice.6 In addition, unlike Madeddu’s study,8 2-kidney, 1-clip hypertension resulted in a similar increase in BP and cardiac hypertrophy in both strains. Madeddu’s group reported earlier that B2-KO mice at 
4 months of age were hypertensive compared with wild-type controls,6,9 findings that others, including our laboratory, could not confirm.1,10,11 Recently, Milia et al compared blood pressure of B2-KO mice that originated from Madeddu’s laboratory to controls (129Sv/J).11 They used the telemetry method, a state-of-the-art method for BP determination in mice, to assess BP and heart rate; yet, they were not able to confirm hypertension or fast heart rate in B2-KO mice. Therefore, the "genetic drift" hypothesis postulated by Madeddu may not apply.12 Moreover, we recently compared B2-KO mice to 129X1/SvJ (a control commonly used by Madeddu’s group) and 129/SvEvTac mice (a control used by our laboratory) at 1, 2, 3, and 6 months of age but found no significant difference in tail-cuff systolic pressure or mean blood pressure (MAP) measured directly in awake mice.13 Direct MAP measurement significantly correlates with BP recorded simultaneously via a telemetric device (r2=0.97; P<0.0001) (N.E. Rhaleb and O.A. Carretero, personal observation, 2001). Others have measured MAP in awake mice but found that ablation of B2 receptors does not cause adult hypertension under normal conditions.10 In response to a previous Letter to the Editor in Hypertension by Madeddu’s group,12 we invited him to exchange B2-KO mice with our laboratory and see whether their mice have different phenotypes than ours. However, conveniently Madeddu’s colony is no longer available. On the other hand, our B2-KO colony is still available for any laboratory, including Madeddu’s, that is interested in assessing the role of kinins in the cardiovascular system.
References
1.
Rhaleb N-E, Peng H, Alfie M, Shesely EG, Carretero OA. Effect of ACE inhibitor on DOCA-salt- and aortic coarctation- induced hypertension in mice: do kinin B2 receptors play a role? Hypertension. 1999; 33: 329–334.
2.
Rhaleb N-E, Yang X-P, Nanba M, Shesely EG, Carretero OA. Effect of chronic blockade of the kallikrein-kinin system on the development of hypertension in rats. Hypertension. 2001; 37: 121–128.
3. Nolly HL, Saed G, Scicli G, Carretero OA, Scicli AG. The kallikrein-kinin system in cardiac tissue. Agents Actions Suppl. 1992; 38: 62–72.[Medline] [Order article via Infotrieve]
4.
Majima M, Katori M, Hanazuka M, Mizogami S, Nakano T, Nakao Y, Mikami R, Uryu H, Okamura R, Mohsin SSJ, Oh-Ishi S. Suppression of rat deoxycorticosterone-salt hypertension by kallikrein-kinin system. Hypertension. 1991; 17: 806–813.
5. Katori M, Majima M, Mohsin SSJ, Hanzuka M, Mizogami S, Oh-Ishi S. Essential role of kallikrein-kinin system in suppression of blood pressure rise during the developmental stage of hypertension induced by deoxycorticosterone acetate-salt in rats. Agents Actions. 1992; 38: 235–242.
6.
Duka I, Kintsurashvili E, Gavras I, Johns C, Bresnahan M, Gavras H. Vasoactive potential of the B1 bradykinin receptor in normotension and hypertension. Circ Res. 2001; 88: 275–281.
7.
Madeddu P, Varoni MV, Palomba D, Emanueli C, Demontis MP, Glorioso N, Dessi-Fulgheri P, Sarzani R, Anania V. Cardiovascular phenotype of a mouse strain with disruption of bradykinin B2-receptor gene. Circulation. 1997; 96: 3570–3578.
8.
Madeddu P, Milia AF, Salis MB, Gaspa L, Gross W, Lippoldt A, Emanueli C. Renovascular hypertension in bradykinin B2-receptor knockout mice. Hypertension. 1998; 32: 503–509.
9.
Emanueli C, Maestri R, Corradi D, Marchione R, Minasi A, Tozzi MG, Salis MB, Straino S, Capogrossi MC, Olivetti G, Madeddu P. Dilated and failing cardiomyopathy in bradykinin B2 receptor knockout mice. Circulation. 1999; 100: 2359–2365.
10.
Cervenka L, Harrison-Bernard LM, Dipp S, Primrose G, Imig JD, El-Dahr SS. Early onset salt-sensitive hypertension in bradykinin B2 receptor null mice. Hypertension. 1999; 34: 176–180.
11.
Milia AF, Gross V, Plehm R, De Silva JA,Jr, Bader M, Luft FC. Normal blood pressure and renal function in mice lacking the bradykinin B2 receptor. Hypertension. 2001; 37: 1473–1479.
12. Madeddu P, Emanueli C. Can knockout mice help dissect relevant genes in hypertension?: evidence and confounding factors. Hypertension. 1999; 34: e14–e15.[Medline] [Order article via Infotrieve]
13. Rhaleb N-E, Yang X-P, Peng H, Cavasin MA, Liu Y-H, Yang F, Xu J, Carretero OA. Cardiovascular phenotype of male 129/SvEvTac, 129/SvJ and B2-KO mice. FASEB J. 2001; 15: A101.Abstract.
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