Abstract

Introduction

Chronic kidney disease (CKD) is a slow and progressive decline of kidney function that is often linked to long standing diseases such as diabetes and hypertension (Remuzzi et al., 2002[35]). It is generally characterized by high amount of creatinine, low GFR and proteinuria which predict an accelerated loss of renal function that irreversibly leads to end stage renal disease (Ritz et al., 1999[36]). Oxidative stress and inflammation play important roles in both the progress of CKD and related complications (Massy and Nguyen-Khoa, 2002[30]). Oxidative stress results from an imbalance between ROS overproduction and reduced antioxidant defences within mitochondria, leading to its dysfunction and consequently to renal cell failure (Wallace, 2005[45]).

Grape seed extract (GSE) is a complex polyphenolics mixture containing flavonoïds, non flavonoïds, proanthocyanidins exhibiting multi-organ protection in various experimental settings (Khanal et al., 2009[24]). For instance, GSE protects the heart (Charradi et al., 2011[11]), the liver (Charradi et al., 2014[8]), the brain (Charradi et al., 2012[10]) and the kidney (Charradi et al., 2013[9]) against high fat diet (HFD)-induced obesity and lipotoxicity in rat. Furthermore high dosage GSE was even shown to improve renal injury in type 2 diabetic rats through its anti-oxidant and anti-inflammatory properties (Bao et al., 2015[5]), and also to protect against arsenic (Zhang et al., 2014[47]), cisplatin (Gao et al., 2014[19]), amikacin (Ulusoy et al., 2012[43]), and cyclosporine A-induced nephrotoxicity (Ulusoy et al., 2012[44]).

The present work aims to test the ability of GSE to improve kidney dysfunction of CKD patients from various renal stages i.e stage 2, 3, 4. Data mainly show that although used at a low dosage, GSE improves several kidney function parameters as well as the anti-oxidative status of CKD patients after a 6-month-supplementation period.

Material and Methods

Results

We reported in Figure 1(Fig. 1) the effect of a 6-month-long supplementation with GSE on some renal hemodynamic parameters of patients suffering from kidney deficiency. GSE increased 1/plasma creatinine ratio by 19.42 % (Figure 1A(Fig. 1), p = 0.044), and decreased proteinuria by -32.67 % (Figure 1B(Fig. 1), p = 0.041) but had no significant effect on plasma urea (Figure 1C(Fig. 1)) and uric acid (Figure 1D(Fig. 1)). Moreover GSE slightly improved GFR by +18.70 % (36.74 ± 5.74 versus 43.62 ± 3.68, p = 0.604) (Figure 1E(Fig. 1)).

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Figure 1

Figure 1: Effect of GSE supplementation on renal function biomarkers of CKD patients. 1/plasma creatinine ratio (A), proteinuria (B), plasma urea (C), plasma uric acid (D) and GFR (E). Data are presented as mean ± SEM. GSE (after 6 months treatment) vs. GSE (at 0 month): *p < 0.05. GSE (after 6 months treatment) vs. Placebo (after 6 months treatment): ^#p < 0.05

GSE also affected several plasma biomarkers of the antioxidant status. GSE slightly decreased plasma MDA by -14.96 % (Figure 2A(Fig. 2), p = 0.0307), and protein carbonylation by -38.82 % (Figure 2B(Fig. 2), p = 0.0319) but had no significant effect on hydrogen peroxide (-14 %, Figure 2C(Fig. 2), p = 0.383). Moreover GSE highly increased plasma CAT by +95.94 % (Figure 3A(Fig. 3), p = 0.015) and SOD by +25.99 % (Figure 3C(Fig. 3), p = 0.049) but had no effect on plasma GPx activity (Figure 3B(Fig. 3)).

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Figure 2

Effect of GSE supplementation on plasma pro-oxidant biomarkers of CKD patients. MDA (A), protein carbonylation (B), H₂O₂ (C). Data are presented as mean ±SEM. Placebo (after 6 months treatment) vs. Placebo (at 0 month): ^§p < 0.05. GSE (after 6 months treatment) vs. GSE (at 0 month): *p < 0.05. GSE (after 6 months treatment) vs. Placebo (after 6 months treatment): ^#p < 0.05

Figure 3

Effect of GSE supplementation on plasma antioxidant enzyme activities of CKD patients. CAT (A), GPx (B) and SOD (C) activity. Data are presented as mean ± SEM. GSE (after 6 months treatment) vs. GSE (at 0 month): *p < 0.05. GSE (after 6 months treatment) vs. Placebo (after 6 months treatment): ^#p < 0.05

We reported in Table 1(Tab. 1) the effect of GSE on plasma biochemistry and inflammation of CKD patients. After a six-month-long treatment period, GSE partially alleviated plasma CRP by -12.81 % (+82.83 % for placebo, p = 0.028 against +56.40 % for GSE, p = 0.168), totally counteracted plasma LDH (p = 0.623) and increased free iron by +13.26 % (p = 0.019) versus placebo. Furthermore GSE slightly depressed triglyceridemia by -16.44 % (p = 0.067), increased plasma lipase activity by +29.67 % (p = 0.006), but had no effect on cholesterolemia.

Table 2(Tab. 2) reported the effect of GSE on some haematological parameters of CKD patients. GSE slightly increased white blood cells (WBC) by +23.48 % (p = 0.276) especially the granulocytes. GSE also prevented anemia of CKD patients. A decrease tendency in red blood cells (RBC) from 4.72 ± 0.14 to 4.37 ± 0.19 in control placebo patients (p = 0.127) was noticed which was cleaned by GSE treatment, allowing the stabilization of RBC from 4.22 ± 0.21 ( control 0 month GSE) to 4.23 ± 0.25 (6 months treatment GSE) (p = 0.692). Furthermore GSE did not affect haemoglobin nor did it prevent the decrease in haematocrit of CKD patients from 39.33 ± 0.81 to 35.53 ± 0.96 in control (p = 0.016) and from 37.22 ± 0.84 to 33.30 ± 1.42 in GSE treated patients (p = 0.049). However GSE significantly counteracted thrombocytopenia by +24.63 % (p = 0.030) when compared to placebo which increased platelets by +10.93 % (p = 0.327).

We reported in Table 3(Tab. 3) the effect of a six-month-long GSE treatment on kidney function biomarker as GFR. The initial number of stage 4 and 3 patients tend to decrease from 6 to 2 and from 11 to 10 patients respectively reaching an overall number of 5 patients who swell the ranks of stage 2, rising the cohort of stage 2 patients from 6 to 11 representing + 83.33 %. No GFR change (no improvement nor worsening) occurred in the placebo group.

Discussion

The present work is a preliminary and pilot study dealing with the effect of GSE treatment on CKD patients of various renal failure stages. Data mainly show that even when used at a low dosage, GSE exerts a clear beneficial effect on some renal function biomarkers as well as on the antioxidant status of CKD patients. To our knowledge, our work is the first one to describe such relevant and hopeful clinical trials data for million CKD patients worldwide.

First of all GSE improves some kidney function parameters as it enhances GFR and clearly lowers proteinuria. This result is of utmost importance as reduction in proteinuria to the lowest achievable level is an important predictor of long term renal protection (de Zeeuw et al., 2004[13]), as it is increasingly recognized that proteinuria may actually be pathological and etiological in CKD progress and not just symptomatic (Roscioni et al., 2014[38]). An association between hyperuricemia and CKD in middle-aged population has also been recently described (Zhu et al., 2014[48]). As in our present case GSE affects both plasma, urine urea and uric acid, its clinical use could be envisaged as an uric acid-lowering therapy substitute to allopurinol.

The beneficial effect of GSE on renal function seems to be mediated, at least partially, by its antioxidant and anti-inflammatory properties. After a six-month-period of treatment, GSE counteracts both plasma lipoperoxidation and carbonylation as well as CRP and LDH, and clearly increases plasma CAT and SOD (Rodrigo and Bosco, 2006[37]). Moreover GSE improves some blood parameters that were shown to be linked to CKD as anemia (Tbahriti et al., 2013[41]), thrombocytopenia (Attman et al., 2011[3]), hypertriglyceridemia (Stegmayr, 2014[40]) and low plasma lipase activity (Dorgalaleh et al., 2013[14]). Importantly no patient drop out of the study nor adverse side effects were noted during the entire clinical trial period. Our data corroborate several previous works in the field demonstrating the heightening of oxidative and inflammatory biomarkers along with a decrease in antioxidant biomarkers in CKD patients. As rightly emphasized (Rodrigo and Bosco, 2006[37]), only few studies dealing with the effect of polyphenols in humans are available, and our contribution is the first to show the effectiveness of GSE to improve renal function in CKD patients from various clinical stages. As a support, dietary supplementation with concentrated red grape juice was shown to exert antioxidant and anti-inflammatory effect in haemodialysis patients (Castilla et al., 2008[7]). Moreover our data are in line with recent animal studies demonstrating the usefulness of grape seed polyphenols in the improvement of renal dysfunction (Bao et al., 2015[5]; Zhang et al., 2014[47]; Gao et al., 2014[19]; Ulusoy et al., 2012[43][44]). In particular the use of high dosage GSE (higher than 500 mg/kg) was recently shown to exert relevant beneficial health effects (Jhun et al., 2013[22]; Tsao et al., 2012[42]; Yang et al., 2012[46]) whereas lower dosages were less effective (Goey et al., 2013[20]).

Remarkably that in our present case, GSE improvement in kidney function of CKD patients has been obtained at a low GSE dosage of 35 mg/kg if we consider the weight of the starting grape seed powder, and highly lower if we consider the polyphenol content. Our laboratory has recently demonstrated the efficiency of high dosage GSE (till 4 g/kg) in multi-organ protection against various insults as obesity (Charradi et al., 2011[11], 2012[10], 2013[9]), stroke (Safwen et al., 2015[39]), or chemotherapeutic treatment (Mokni et al., 2012[32]). In fact, our studies were inspired from the pionneering work of Bagchi et al. (2000[4]) and other subsequent studies that led to the elaboration of calibrated and commercial GSE preparations which evidenced the safety of high dosage GSE (till 2 g/kg) in long-term experiments (Bentivegna and Whitney, 2002[6]). In light of these data it appears that the concentration of GSE used in the present study (35 mg/kg) is far less from the No Observed Adverse Effect Level (NOAEL) of IH 636 or oligonol GSE preparation (Fujii et al., 2007[18]). In addition as GSE recently received the Generally Recognized As Safe (GRAS) certification from US FDA (2011[16]), its use opens a new and safe approach for the validation of high dosage GSE to impede CKD. Most importantly and although used at very low level, GSE seems to slow down the progression of CKD and even to reverse the decline in kidney function (Choi et al., 2014[12]) as described in the present pilot study i.e from stages 4 and 3 to stage 2.

Nevertheless the current interventional pilot study suffers from several limitations. First the sample size is too small to be of robust statistical significance. Although GSE was used at low dosage, a significant improvement in the decline of some kidney function biomarkers after a six-month-treatment period was obtained. Duration of the study should be extended to increase blood and urine sampling and a ten fold higher GSE dosage (350 mg/kg) should be tested to reach improvement of statistical significance. Owing to its tremendous safety, GSE is increasingly used at high dosage (more than 500 mg/kg) according to our own data (Charradi et al., 2011[11], 2014[8], 2012[10], 2013[9]) or to current literature (Li et al., 2015[27]; Mansouri et al., 2011[28]; Margalof et al., 2015[29]).

In conclusion, due to the multi-organ safety and beneficial health effects of grape polyphenols, clinical trials using high GSE dosage should be urgently planned to confirm these preliminary and hopeful data.

Conflict of interest

The authors declare no conflict of interest.