Resistant hypertension (RH) is a condition characterized by elevated blood pressure (BP) that persists despite the use of a diuretic and at least two other antihypertensive medications. Patients with RH are at an increased risk of a future cardiovascular events compared to those whose BP is controlled (Bangalore et al., 2014; Irvin et al., 2014). Whilst there are many classes of antihypertensive drugs available today, successful reduction in BP remains difficult. To combat the problem of treatment resistance to medications, renal sympathetic denervation (RDN) has been explored clinically as an alternative approach.

While epidemiological studies have clearly established that high BP is linked to cardiovascular disease, it is also known that the risk is further increased by the addition of obesity. Visceral obesity is highly prevalent among RH patients. The pathophysiology of obesity-associated hypertension is complex and characterized by activation of the sympathetic nervous system (Esler et al., 2006), renin-angiotensin system (Boustany et al., 2004), endothelial impairment (de Jongh et al., 2004) and other mechanisms.

The view that adipose tissue is merely a repository of stored lipids has been revised over recent years. As many as 50 different substances have been discovered to be synthesized by adipocytes, many of which play a role in more than lipid metabolism. In fact, adiponectin, resistin and leptin, mainly, but not exclusively, synthesized by adipocytes, have been implicated in BP control. For example, it has been shown in experimental models of obesity that increase in leptin, associated with body weight gain, is followed by an elevation in systolic and diastolic BP after 12 weeks of high fat diet (Simonds et al., 2014). The administration of leptin antibody to the same mice resulted in a significant reduction in BP (Simonds et al., 2014). We have also demonstrated in our earlier studies that leptin levels are associated with renal sympathetic nerve activity (Eikelis et al., 2003), which can lead to increase in BP. On the other hand, both clinical and experimental studies have demonstrated that adiponectin levels are inversely related to BP: adiponectin levels fall with increasing BP and vice versa (Imatoh et al., 2008; Baden et al., 2013).

While the effect of renal denervation (RDN) on the reduction of BP, which is, at least partially, achieved via the reduction of sympathetic stimulation to the kidney, has been examined in the last couple of years, not much is known in relation to the effect of RDN on adipokines. The aim of the current study was to determine the effect of renal ablation on adipokine profile in obese patients with RH. In particular whether the potential effect of RDN on the adipokine profile is associated with the fall in BP in patients with RH.

This study was undertaken to determine whether RDN has any effect on the adipokine profile in mostly overweight/obese patients with RH. Here we report that RDN was associated with a significant increase in adiponectin levels at 3 months post-treatment. Adiponectin is almost solely synthesized by adipocytes. Its effects are known to be cardioprotective due largely to its anti-inflammatory and anti-atherogenic properties (Ohashi et al., 2012). Adiponectin is distinct from other adipokines as it is negatively correlated with body fat stores. Plasma adiponectin levels are reduced in people with obesity (Ryan et al., 2003). In the current study there were no changes in BMI between the two visits nor did we observe any significant changes in plasma leptin, which is a marker of fat stores. Therefore, we can conclude that the observed changes in adiponectin levels are not related to adipose stores.

Clinical studies suggest that adiponectin levels strongly correlate with BP, with hypoadiponectinaemia (low adiponectin levels) being a risk factor for hypertension. Baden and colleagues have demonstrated a significant negative correlation between adiponectin levels and BP in a group of healthy people over a wide adiponectin range (Baden et al., 2013). Adiponectin has been shown to exert its effects by stimulating the enzymatic activity of eNOS in endothelial cells. Such direct action of adiponectin on endothelial cells may represent the association of this adipokine and BP. Previous studies have demonstrated that adiponectin can decrease BP not only in hypertensive population but also in those with normal BP (Baden et al., 2013).

There is some evidence that sympathetic nervous activity can modulate adiponectin concentration. For example, Nowak et al demonstrated that 6 months treatment with the centrally acting sympatholytic agent, rilmenidine, in patients with essential hypertension was associated with increased adiponectin levels without changes in total body fat content (Nowak et al., 2005). On the other hand, cold exposure, which leads to heightened sympathetic activity, significantly reduced both adiponectin serum levels and adiponectin mRNA expression in subcutaneous, mesenteric and epididymal fat tissues in mice (Imai et al., 2006). It is well established that increased sympathetic drive is a hallmark of hypertension. Sympathetic activation, in its turn, has been shown to inhibit both synthesis and secretion of adiponectin. For example, administration of β-adrenergic agonists resulted in a reduction in adiponectin gene expression in human visceral adipose tissue (Delporte et al., 2002). We have demonstrated previously that RDN was associated with a significant reduction in renal noradrenaline spillover (indicative of renal sympathetic activity) (Krum et al., 2009) and muscle sympathetic nerve activity (Hering et al., 2013). Although sympathetic activity to adipose tissue was not examined in this study, we can only speculate that there was a reduction in sympathetic drive to adipose tissue after RDN which consequently resulted in higher circulating adiponectin.

On the other hand, there is also evidence that adiponectin might be involved in sympathetic regulation. Adiponectin was detected to be present in cerebrospinal fluid in humans which suggests that it may be able to act centrally to influence cardiovascular function (Kusminski et al., 2007). In support of this, both direct central and peripheral injections of adiponectin in anesthetized rats were associated with reduction of renal sympathetic nerve activity and BP (Tanida et al., 2007), therefore suggesting that BP lowering effects of adiponectin are at least partially mediated via suppression of sympathetic activity. While the nature of this study does not allow to state with certainty whether reduction in sympathetic activity after RDN caused adiponectin levels to increase or vice versa, it is probably the former, given that there was no change in weight at follow up visit. However, what might be the precise mechanism of adiponectin action in the central nervous system that would lead to changes in sympathetic outflow is not clear, but may be perhaps via leptin signaling.

In this study we also observed a significant increase in fasting insulin levels at 3 months after renal ablation. This finding was somewhat unexpected given Mahfoud et al. reported a significant reduction of fasting insulin levels in a similar group of resistant hypertensive patients at 3 months follow up accompanied by improvement in glucose metabolism (Mahfoud et al., 2011). Others have reported an improved glucose metabolism post-RDN procedure (Witkowski et al., 2011). It is possible that reported improvement is due to sympathetic inhibition which follows RDN (Hering et al., 2013). On the other hand, Miroslawska et al. reported no changes in either whole-body (peripheral) or hepatic insulin sensitivity in a group of resistant hypertensive patients at 6 month follow up after denervation despite a significant drop in BP (Miroslawska et al., 2016). It is important to note that in the current study we measured fasting insulin levels but did not evaluate insulin sensitivity and as such we cannot speculate whether there were changes in insulin or glucose status. It is possible; however, that renal ablation had a favorable effect on the recovery of the islet cells, which in turn resulted in an increase in insulin levels. Different medication regime and adherence, which was not assessed in the current study, might account for differences between data from different studies. The mechanisms by which RDN can affect insulin metabolism remain unknown and further studies should be designed to clarify the role of RDN on insulin sensitivity.

Resistin, while classified as an adipokine, is a protein produced primarily by macrophages in humans (Yang et al., 2003) and, when first discovered, was thought to provide a link between obesity and insulin resistance (Steppan et al., 2001). Later studies pointed to a role of resistin in the pathogenesis of hypertension (Zhang et al., 2010) possibly via inflammatory processes (Bokarewa et al., 2005). We did not see any changes in resistin concentration at 3 months after the procedure. Other studies have shown that hypertensive patients have higher resistin levels compared to normotensive individuals. We have not measured resistin levels in healthy individuals, however, compared to previously published reports, (Papadopoulos et al., 2005; Pantsulaia et al., 2007; Jamaluddin et al., 2012), resistin levels in the current cohort were not different from what is seen in healthy young people.

One of the explanations as to why resistin levels in our cohort were within the normal range could be due to the fact that resistin is affected by cholesterol lowering drugs such as statins, and that the 65% of patients in the current study were on statin therapy. For instance, treatment with atorvastatin for 6 months at a dose of 10 mg/day resulted in reduced resistin levels in patients with type 2 diabetes (Ichida et al., 2006). In the same study, the authors demonstrated that incubation of monocyte/macrophages with atorvastatin for 24/48 h led to a significant reduction in resistin expression as measured by qRT-PCR. Interestingly, no such effect of statin has been reported for adiponectin in hypertensive or type 2 diabetes patients (Kumada et al., 2003; Chu et al., 2008; Al-Azzam et al., 2013).

In this study we demonstrated a significant reduction in plasma NEFA levels post-RDN. NEFA is predominately derived from adipose tissue; and as such, NEFA concentration is increased in overweight/obese people (for review see Karpe et al., 2011). Circulating NEFAs are known to mediate adverse metabolic effects; with elevated NEFA at baseline serving a predictive value for type 2 diabetes. However, it should be mentioned that the increase in NEFA levels in obese individuals is not proportional to their BMI. In our study, the weight of the patients did not differ between the baseline and follow up visits. As mentioned above, leptin levels also remained unchanged. Therefore, the observation of reduced NEFA concentration post-denervation is intriguing. NEFA levels were reduced on average by half in 48 patients (out of 55 examined in this study).

Based on the results of clinical studies it would appear that there is a close association between NEFA and BP. For example, Steinberg et al. demonstrated that acute elevation in free fatty acids leads to a rise in systolic BP in lean insulin-sensitive patients (Steinberg et al., 1997). Furthermore, intravenous infusion of intralipid to healthy volunteers resulted in short term rise in BP and heart rate, suggesting altered central autonomic control (Stojiljkovic et al., 2001). NEFA can cross the blood-brain barrier where it can exert direct effects on sympathetic outflow to the muscle bed or other organs (Lam et al., 2005; Florian and Pawelczyk, 2009).

Reduction in NEFA could be due to reduction in the rate of lipolysis. One of the regulators of lipolysis is the sympathetic nervous system. In fact, about 40% of total free fatty acids released from adipocytes was shown to be attributed to sympathetic stimulation of adipose tissue (Hucking et al., 2003). Assessment of muscle sympathetic nerve activity and renal sympathetic nerve activity (using renal noradrenaline spillover technique) provided evidence of the reduction of the sympathetic drive to these organs after renal ablation (Krum et al., 2009; Hering et al., 2013, 2014). Therefore, it can be speculated that there may be a similar reduction of sympathetic activity to adipose tissue after RDN procedure. Further studies will need to address this issue directly.

It has been reported that adiponectin levels can affect NEFA concentration (Lavoie et al., 2009). Lavoie et al. demonstrated that there was an inverse association between circulating total and high molecular weight (HMW) adiponectin and the appearance of plasma NEFA during elevation of intravascular triacylglycerol lipolysis in healthy males (Lavoie et al., 2009). Due to the design of the study it cannot be concluded what was the nature of causal relationship between adiponectin and NEFA levels.

In conclusion, to our knowledge, this is the first study to demonstrate a favorable effect of RDN on adiponectin and NEFA levels, which may, at least in part, contribute to the reduction in BP. Whether adiponectin and NEFA changes subsequent to RDN are independent of the reduction of the sympathetic activation remain to be elucidated. While the clear-cut reductions in NEFA and increase in adiponectin after RDN in the absence of changes in weight, medication or other factors are suggestive of a potential causal association, in this observational study, we cannot establish the causation; nevertheless, the results are of interest and merit further investigation.

NE, conception of the manuscript/data acquisition/data analysis/drafting of the manuscript; DH, data acquisition/critical editing of the manuscript; PM, data acquisition/critical editing of the manuscript; JD, data acquisition/critical editing of the manuscript; LH, data acquisition/critical editing of the manuscript; AW, renal denervation/critical editing of the manuscript; EL, data analysis/critical editing of the manuscript; ME, critical editing of the manuscript; GL, data analysis/critical editing of the manuscript; MS, conception of the manuscript/data analysis/critical editing of the manuscript.