Slide 1

Mr Chairman, Ladies and Gentlemen, first of all thank you very much for the kind invitation and the kind introduction. I’ve  really enjoyed the debates so far, both presentations before have given rise to a heated discussion and one of the gentlemen in the audience asked about what about the genetic forms of hypertension and I will go through that in a few slides if you don’t mind and I will tell you my opinion which is a bit different from the one of the previous speaker.

Slide 2

Now, I was given a challenging topic by Carmine, I don’t know if he’s in the audience and I nearly said well, do you really want me to talk about that? Then I said well, let’s take the challenge and let’s see what the evidence is about a role of salt-sensitivity in predisposing hypertensive nephrosclerosis and diabetic nephropathy.
Now, I assume that we accept the concept of salt-sensitivity that means that if you take subjects and you change their salt content in the diet in a standardised fashion, that some of them will respond with a certain amount of blood pressure increase, whereas others will not have a significant blood pressure increase. We don’t have to think of salt sensitivity as a bimodel distribution it’s a continuum but some will have a significant increase, whereas some will not have a significant increase.

Now what we know from the epidemiological studies is that salt-sensitivity is more prevalent in hypertensive subjects than in normotensive ones and it is more prevalent also in the black population compared to the white population.

Slide 3

Now, this last observation is actually very interesting, it points to an interplay between salt content in the diet and some genetic aspects. Now, we will have to ask the question, well, if salt sensitivity plays a role in hypertensive and diabetic renal disease, then we would expect that subjects with a high prevalence of salt sensitivity have actually a higher incidence of these diseases. There is an indirect indication that may be the case. These are data from the USRDS data registry from last year and on the left hand side of the cartoon you see the incident rates of diabetes or diabetic ESRD in the black population, the blue lines and the white population, the brown lines. Now there is a significantly higher incident rate in black diabetics aged 50-59. Now, you could argue they have a higher prevalence of diabetes full stop but the difference is significantly higher for the incident rate of ESRD compared to the prevalence of diabetes.
The same applies to hypertensive kidney disease in the black population both in the younger age group 40-49 and the older age group 50-59. The prevalence is vastly different than in the white population.

Slide 4

Now, I’ll go back to the story of the genetic hypertension and I have to maybe explain a few issues. In the last ten years there has been a wide development based on molecular biology advances and we have been able to characterise a whole series of genetic forms of hypertension. Now there are two interesting observations. First of all, all the genetic forms, monogenetic forms of hypertension characterised to date involve an abnormality in sodium transport in the kidney with an increased sodium reabsorption. The second fact is that these are forms that are associated with hypokalemia, metabolic alkalosis and usually suppressed renin which is what you have when you have volume overload. This is a cartoon that shows some data that I had the privilege to investigate and that’s about a syndrome that’s called Apparent Mineralocorticoid Excess. Now this syndrome is caused by the mutation of a gene which is called 11βHSD2 it stands for 11β hydroxysteroid dehydrogenase type 2 which is an enzyme that inactivates cortisol into the ketoform cortisone and the interesting thing is that cortisol and aldosterone have the same affinity for the mineralocorticoid receptor but in organs like the kidney cortisol is inactivated and therefore, the mineralocorticoid receptor is protected by over activation, by the reactive glucocorticoid. In patients with a mutation in this gene cortisol is present intracellularly in very high concentrations and stimulates permanently the mineralocorticoid receptors and enhances the sodium and water retention and hypertension. What happens in this situation is that patients develop hypertension and what I’ve got here at the bottom is different phenotypes of these patients, different degrees of hypokalemia, different severities of hypertension and here I have the ratio of the urinary metabolite of active cortisol F to inactive cortisol which is a parameter of 11βHSD2 activity. If the activity is completely abolished, this ratio will be very high, if the activity is partially there, the ratio is very low. These are in vitro expressions, you can take the gene expressed in the mammalian cell, measure the conversion of radio labelled cortisol into radio labelled cortisone and thereby determining whether the activity is partially blunted or completely blunted.
Now regardless of the different results and the different kindreds that we have examined here with Xavier Jeunemaitre and Gilles Morineau in Paris an interesting observation is that with these forms of salt-sensitive hypertension where salt plays an important role, the prevalence of kidney damage being either proteinuria, increase in creatinine or in two of these cases, index cases, end stage renal disease was very high. So you have a form of salt-dependent hypertension, whereas the presence of kidney damage is almost always invariably there.

Slide 5

Now this is something that we use to learn, it doesn’t mean that all patients with salt-sensitivity who develop nephrosclerosis will have this. An interesting observation comes from Watson who looked at microsatellite flanking the 11βHSD2 gene and he took this marker as an indication of possible variable expression of the gene and he looked at that in a population of black hypertensives with end stage renal disease, black normotensive controls and white population. He made two observations, the first one is that there was a difference in the allele distribution between white and black population and the second one was that there was allele heterogeneity for the allele length 216bp in the black hypertensive population with ESRD compared to the control subjects. Now, this would also suggest that genetically altered 11βHSD2 activity could lead to the salt sensitivity in this sub-group of African-Americans that have a high prevalence of ESRD.

Slide 6

The final interesting observation from clinical data comes from a study that nowadays would not be possible because we treat kidney transplant patients with cyclosporine and therefore, we induce hypertension but at this time when Curtis published the data in the New England Journal of Medicine in 1983 the standard therapy was prednisolone and azathioprine and what happened in this observation was that patients who had ESRD due to essential hypertension and were salt-sensitive as we usually know about the black population, when they received a kidney from a normotensive donor and in most instances probably from the white background, their hypertension was cured. This suggests that the kidney actually determines the blood pressure and the blood pressure goes with the kidney. They did also some studies looking at Na loading and Na deprivation and the response of blood pressure was normal. Now, this would suggest that salt-sensitivity accompanies the kidney and is not determined by extrarenal factors.

Slide 7

Now, the question is well is there any evidence that we have histological damage suggesting nephrosclerosis in salt-sensitivity? We have to resort to animal experiments. In this study by Siegel there were three groups of rats; spontaneous hypertensive rats, SHR, the Dahl salt-sensitive rat and the spontaneous hypertensive rat stroke prone strain who were first analysed on normal or low salt diets and then another group that was treated with 4% of sodium chloride in the diet and this for all three strains.
Now, in the salt-sensitive Dahl model and the stroke prone SHR group blood pressure went up significantly on a high salt diet. What also happened is that there was proteinuria that was vastly increased in both the stroke prone strain and the salt-sensitive Dahl rats. The salt-sensitive Dahl rats already had some degree of microalbuminuria on a low salt diet.

Slide 8

If you look at the histology in these rats and these are just some examples in micrographs of some of the rats, the first two panels here on this side of the graph show glomeruli from SHR rats and SHRSP strain on a low salt diet which are virtually normal and these are the corresponding glomeruli of these rat strains on a high salt diet and you see that there is some hyaline thrombosis and there is glomerular collapse in these two glomeruli. This is then something that’s a little bit different, the Dahl salt-sensitive show actually glomerular hypertrophy and some mesangial expansion. --- from salt-sensitive Dahl rats and SHR on a high salt, they show even more significant hypertrophy, they show some significant mesangial damage and also some degree of monocellular infiltration.

Slide 9

Another study that looked at Dahl salt-sensitive rats on a standard chow and kidney damage at 4 months and 12 months and looking at markers of profibrotic markers in the kidney such as collagen type IV, laminin and TGFβ, well here is at 4 months, the glomerulus is virtually normal, whereas at 12 months there is glomerular hypertrophy and then there is a higher expression of collagen type IV, laminin and TGFβ suggesting the progression of glomerulosclerosis.

Slide 10

The question is, is there an indication of podocyte damage? This has been shown very elegantly by Nagase looking at Dahl-resistant and Dahl-sensitive rats on a low salt diet and a high salt diet what happened is that only the Dahl salt-sensitive rats on a high salt diet had an increase in blood pressure, they developed proteinuria and at 6 weeks they had also an increase in serum creatinine and these are for the 4 groups of rats. Immunostaining for desmin, B7-1 and nephrin all markers of podocyte activity and you see that there is a significant increase in the Dahl salt-sensitive rats on a high salt diet of desmin and B7-1 and a decrease in neprhin expression.

Slide 11

Now the other part of the question was, is there any indication that salt-sensitivity predisposes to diabetic nephropathy? There is a little bit less literature here and we have to speculate a little bit more but I think that what we can agree on is that obesity is a significant risk factor for diabetes and the two share something in common that the reason enhanced tubular sodium reabsorption that this enhanced tubular sodium reabsorption causes a rightward shift of the pressure natriuresis curve and that the steady state in-out sodium homeostasis functions at a higher level meaning a salt-sensitivity is the consequence.

Slide 12

Now, to show this we have to go back many years and I just put up this slide because it was done by Peter Weidman in Bern in 1979 and Peter was my mentor and he’s the one that gave me the back to do nephrology. What he did he measured exchangeable sodium in normal subjects and then he measured exchangeable sodium with radio labelled sodium in 17 diabetics on placebo and then he treated them with chlorthalidone. What you see is that diabetics on placebo have a significant increase in exchangeable sodium meaning that the salt sensitivity, this rightward shift of the curve actually causes an increase in total body sodium and that this is accompanied by hypertension and it is a correction by chlorthalidone reduction of the total exchangeable sodium is accompanied by a correction of the hypertension.

Slide 13

Now, to support the fact that salt-sensitivity is important in diabetics I just want to recall this interesting review by Curb a few years ago and we’re all well aware of the metabolic side effects of diuretics. They increase impaired glucose tolerance, they increase serum cholesterol, they decrease potassium and these side effects are more important in patients with diabetes and therefore, we shouldn’t treat patients with diabetes and hypertension with diuretics. But if you look at the effect on total mortality or myocardial infarction, actually diuretics did a better job in diabetic patients than they did in non-diabetic patients and maybe this suggests it’s because the mechanism there is salt-sensitivity and it’s more rational to give diuretics in this group than in the non-diabetic population. Certainly these metabolic side effects are actually overrun by the evidence that stringent end points, and significant end points are clearly improved.

Slide 14

But let’s go back to the story whether salt-sensitivity might predispose to diabetic nephropathy and again, is a study looking at exchangeable body sodium that was done by Feldt-Rasmussen in Denmark about 20 years ago and here in yellow are the normal subjects, in orange are the diabetic patients with type I diabetes this time but no signs of nephropathy and the two in red are with incipient nephropathy or overt nephropathy. What you see is that there is an increase in total body sodium already in the stage of incipient nephropathy and this is of higher magnitude in patients with overt nephropathy. You might tell me, well that’s not an explanation of salt-sensitivity as a cause of nephropathy, it could be the other way round that the more severe the degree of nephropathy, the more salt-sensitive the patient will become. Together with Professor Ritz and with Professor Strojek in Poland we then had a look at offspring of patients with diabetes and other non-nephropathy or parents with diabetes and diabetic nephropathy and we looked at whether these subjects had salt-sensitivity or not and what could be the potential mechanism. One of the mechanisms we looked at was 11β HSD2.

Slide 15

Now what you see here is that offspring of diabetic parents with nephropathy they were more often salt-sensitive these are the crosses in red than the non-nephropathic parents and the control group. What you see here the dots and the crosses, they characterise the level of 11β HSD2 activity measured in vivo as the ratio of these tetrahydro-metabolites for cortisol and cortisone. This is the normal range when the ratio is high, there is a decreased activity of this enzyme. So this could suggest that salt-sensitive blood pressure could be an intermediate phenotype predisposing to diabetic nephropathy.

Slide 16

I just want to conclude reminding you that hypertensive patients with salt-sensitivity have a greater propensity to nephrosclerosis-associated ESRD.
Glomerular abnormalities in animal experiments have been shown and there is an increased expression of profibrotic mediators that are seen in these animals like Dahl salt-sensitive or stroke prone hypertensive rats. Patients with diabetes mellitus have increased total body sodium which is associated with hypertension and particularly nephropathy and finally, like the study that we did in offspring of diabetic patients with and without nephropathy the salt-sensitive blood pressure appears to be an intermediate phenotype predisposing to diabetic nephropathy. Thank you for your attention.

Slide 17

Chairman: Thank you very much Doctor Ferrari. Your presentation is open to questions and comments. Maybe I could start with the first question? In the salt-resistant Dahl rat you showed that high salt intake was not associated with renal lesions and not associated with high blood pressure. So do you think high blood pressure is the essential link or as the subsequent speaker will probably tell us there maybe blood pressure independent salt effects also of the kidney maybe?

Prof Ferrari: I think it’s always difficult when you look at inbred rat strain to know whether the effect that you see by selecting the rats in this case dependent or independent of blood pressure is not also associated with another gene that has been selected that predisposes to some sort of damage but the fact is that if you keep the salt-sensitive rat on a low salt diet for a very long time, they will not develop the glomerulosclerosis and therefore, it will be an argument against the fact that there is a genetic component or some other factors rather than hypertension causing the damage.

Chairman: Any other questions? May I ask you about the Milan strain the --- abnormality, is this strain also sensitive to salt in terms of development of renal lesions?

Prof Ferrari: There have been a couple of studies where they altered the salt content in the Milan hypertensive rats and there doesn’t seem to be a significant effect of low or high salt. I’m not arguing that salt-sensitive is the only effect but I think salt-sensitive may play a role in a subset of patients but as it is clearly known once you develop a certain degree of renal failure everybody becomes salt-sensitive. So what’s the chick and what’s the egg?

Chairman: Very difficult to say indeed we know that haemodialysis patients are extremely sensitive to salt. Ok thank you very much we’ll stop here.