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Protodioscin ameliorates fructose-induced renal injury via inhibition of the mitogen activated protein kinase pathway.


Background: High dietary fructose can cause metabolic syndrome and renal injury.

Purpose: The effects of protodioscin on metabolic syndrome and renal injury were investigated in mice receiving high-dose fructose.

Methods: Mice received 30% (w/v) fructose in water and standard chow for 6 weeks to induce metabolic syndrome and were divided into four groups to receive carboxymethylcellulose sodium, allopurinol (5 mg/kg) and protodioscin (5 and 10 mg/kg) continuously for 6 weeks, respectively. The glucose intolerance. serum uric acid (UA), blood urea nitrogen (BUN), creatinine (Cr), total cholesterol (TC), triglyceride (TG), interleukin-1[beta] (IL-1[beta]), interleukin-6 (IL-6) and tumor necrosis factor-[alpha] (TNF-[alpha]) were determined. Results: Protodioscin significantly improved glucose intolerance and reduced the levels of serum UA, BUN, Cr, TC and TG. Histological examinations showed that protodioscin ameliorated glomerular and tubular pathological changes. Protodioscin significantly reduced renal concentrations of IL-1[beta], IL-6 and TNF-[alpha] by inhibiting the activation of nuclear factor-[kappa]B, c-Jun N-terminal kinase, p38 mitogen-activated protein kinase and extracellular signal-regulated kinase. In addition, the effect of protodioscin on the mitogen activated protein kinases (MAPK) pathway was examined.

Conclusion: Taken together, protodioscin is a potential drug candidate for high dietary fructose-induced metabolic syndrome and renal injury.



High fructose feeding

Renal injury


MAPK pathway


High-fructose com syrup was introduced in the late 1960s, and since then, fructose consumption has increased markedly (Malik et al., 2006). Unlike other sugars, approximately 70% of fructose is taken up by the liver and the kidney (Aoyama et al., 2012). Epidemiological, clinical and experimental studies have proved that high dietary fructose may cause metabolic syndrome, such as dyslipidemia, insulin resistance, hyperglycemia, hypertension, hyperuricaemia and cardiovascular disease (Aoyama et al., 2012; Stanhope and Havel, 2010; Zawiasa and Nowicki, 2013). Metabolic syndrome is a constellation of risk factors of chronic kidney disease (CKD) and is a serious public health problem. Increasing research supports the view that insulin resistance induced by high dietary fructose, can cause hyperuricaemia and hyperglycaemia. The increase of both serum uric acid (UA) and blood glucose levels leads to renal injury through multiple avenues (Ma et al., 2015). UA has been suggested as an independent predictor of CKD, which can cause renal endothelial dysfunction, glomerular hypertension and cortical vasoconstriction (Chen et al., 2013). Previous evidence indicates that hyperglycaemia can activate transforming growth factor-[beta]1 (TGF-[beta]1) and lead to overexpression of inflammatory cytokines, subsequently inducing changes in the renal structure and function (Kajitani et al., 2010; Qi et al., 2008). Currently, only a few drugs are effective against CKD; therefore, there is a need to search for safe and effective drugs against CKD.

Dioscoreae rhizome (DR, Shan-yao in Chinese), the rhizome of Dioscorea oppositifolia L. (Dioscoreaceae) is an important food in China. DR has also been used in traditional Chinese medicine for centuries for invigorating the spleen, stomach and kidney (Shujun et al., 2006). Pharmacological studies have demonstrated the therapeutic effects of DR in numerous diseases, such as cardiovascular disorders, cancers, diabetes, neurodegenerative disorders, allergic diseases and amelioration of menopausal symptoms (Huang et al., 2011; Lee et al., 2002; McAnuff-Harding et al., 2006; Zhao et al., 2005). Protodioscin (Fig. 1), a major steroidal saponin in DR, has been shown to exhibit multiple biological actions, such as anti-hyperlipidemia, anti-cancer, sexual effects and cardiovascular properties (Gauthaman et al., 2003; He et al., 2006; Hu and Yao, 2002; Wang et al., 2010; Zhang et al., 2016).

In regards to the pathogenesis of renal injury induced by high dietary fructose and the pharmacological actions of protodioscin, we hypothesized that protodioscin has a beneficial effect in renal injury. The present study investigated the effects of protodioscin on metabolic syndrome and renal injury in mice treated with highdose fructose. To elucidate its potential mechanism of action, the effect of protodioscin on the mitogen activated protein kinases (MAPK) pathway was examined.

Materials and methods

Chemicals and reagents

Protodioscin (96.2% purity, batch number: 2,015,602) was supplied by Spring Autumn Biological Engineering, Co., Ltd, Nanjing, China. Allopurinol, the positive control used in the experiments, was obtained from Xinyi Pharmaceutical Ltd, Shanghai, China. Protodioscin and allopurinol were suspended in 0.4% carboxymethylcellulose sodium (CMC-Na) solution for the animal experiments. Fructose was purchased from Aldrich Chemical Company (St Louis, MO, USA), Inc. Kits for blood glucose, blood urea nitrogen (BUN), creatinine (Cr), total cholesterol (TC), triglyceride (TG), UA and protein concentration were purchased from Nanjing Jiancheng Institute, Nanjing, China. Enzyme-linked immunosorbent assay kits for tumor necrosis factor-[alpha] (TNF-[alpha]), interleukin-1[beta] (IL-1[beta]), interleukin-6 (IL-6), leptin and insulin were obtained from Cusabio Biotech Co., Ltd, Wuhan, China. Antibodies of c-Jun N-terminal kinases QNK), phosphorylated JNK (p-JNK), extracellular signalregulated kinase (ERK), phosphorylated ERK (p-ERK), protein-38 mitogen activated protein kinases (p38 MAPK) and phosphorylated p38 (p-p38 MAPK) were purchased from Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA. Antibodies against nuclear transcription factor-[kappa]B P65 (NF-[kappa] BP65), phosphorylated NF-[kappa] BP65 (p-NF-[kappa] BP65), inhibitor of NF-[kappa]B-[alpha] (I[kappa]B[alpha]) and phosphorylated I[kappa]B[alpha] (p-I[kappa]B[alpha]) were purchased from Signalway Antibody Co., Ltd, College Park, MD, USA. Horseradish peroxidase-conjugated antibody against glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was purchased from Santa Cruz Biotechnology Inc., Santa Cruz, CA, USA. Whole cell lysis assay kits and nuclei isolation lysis assay kits were purchased from Keyge Biotech Co., Ltd, Nanjing, China.


Adult male ICR mice (18-22 g) were purchased from Jiangsu University Laboratory Animal Center and kept at a maintained environment (23 [+ or -] 2 [degrees]C, humidity of 50% [+ or -]10%. and 12 h light/dark cycle) with access to normal laboratory food and water. All procedures were carried out in accordance with the Guide for the Care and Use of Laboratory Animals, and were approved by the Ethics Committee of China Pharmaceutical University.

Experimental design and treatment protocol

After habituation for 7 days, the animals were randomly divided into either the control (n= 10) or experimental (n=40) group. Mice in the control group received drinking water and standard chow, while the experimental group received 30% (w/v) fructose in drinking water and standard chow for 12 weeks (Yang et al., 2015). After 6 weeks, mice receiving high-dose fructose were divided into four subgroups: fructose group (treated with CMC-Na in a matched volume), allopurinol (Alio) group (administered 5 mg/kg allopurinol hydrochloride), protodioscin-5 (Pdio-5) group (administered 5 mg/kg protodioscin) and protodioscin-10 (Pdio-10) group (administered 10 mg/kg protodioscin). Dose of protodioscin was selected according to other reports and the clinical adult dose of DR (Wang et al., 2010). According to the pharmacopoeia of China, the dose of DR for human is 30g/day. Equivalently, the calculated dose of DR based on respective body surface areas for rats is 2.6 g/kg/day. The average content of protodioscin in DR is 0.183% (Liu et al., 2006a), and so the dose of protodioscin for rats is 4.76 mg /kg/day. Therefore, we chose 5 mg/kg/day as low dose, and 10 mg/kg/day as high dose in this study. All drugs were administered orally once daily between 9:00 and 11:00 a.m., continuously for 6 weeks.

Glucose tolerance

Glucose tolerance was estimated by an oral glucose tolerance test (OGTT) at the end of the treatment. After 12 h of fasting, mice were administered 2g/kg glucose orally and blood samples were collected from the caudal vein at 0, 30, 60, 90 and 120 min after glucose administration to measure the blood glucose levels. The results were expressed as an integrated area under glucose concentration time curve (AUC).

Urine, blood and tissue processing

Three days after the OGTT, urine samples were collected from mice, who were housed in individual metabolic cages for 24 h, for analysis of urine volume and biochemical parameters. The blood was collected via the abdominal aorta and centrifuged (3000 g) for 10 min to obtain serum for TC, TG, insulin, leptin, Cr, BUN and UA analysis. After being weighed, the right kidneys were collected for histology examination and the left kidneys were stored in liquid nitrogen for western blot analyses and biochemical estimations.

Histological examination of the mice kidneys

The kidneys from the mice were fixed in 10% neutral formalin for 24 h at room temperature, dehydrated through a graded alcohol series, embedded in paraffin, and cut into 4/xm-thick sections. The sections were stained with hematoxylin-eosin and the related indicators were examined under light microscope. Histological assessment was examined by two experienced morphologists, who were blinded to the origin of the slides. The surface area of glomeruli were measured by computer image analysis system, and glomerular volume was calculated by the formula (Honore et al., 2012): glomerular volume = [[mean glomerular tuft profile area].sup.1.5] x 1.01/1.382. The index of mesangial expansion was scored by a quantitative estimate of the mesangial zones width in glomerulus: 0, normal glomeruli; 1 = matrix expansion occurred in up to 25% of a glomerulus; 2 = matrix expansion occurred in 25-50% of a glomerulus; 3 = matrix expansion occurred in 50-75% of a glomerulus; 4- matrix expansion occurred in 75-100% of aglomerulus (Cai et al., 2010).

Analysis of TNF-[alpha], IL-1[beta] and IL-6 levels in the renal tissues

The levels of TNF-[alpha], IL-1[beta] and IL-6 in the renal homogenates were determined using commercial ELISA kits according to the manufacturer's instructions. The levels of these cytokines in the renal tissues were normalized to the protein content.

Analysis of serum levels of TC. TG, Cr, BUN, UA, insulin and leptin

The levels of TC, TG, Cr, BUN, UA, insulin and leptin were determined using commercial kits. Homeostatic model assessment (HOMA), an estimation of insulin resistance, was calculated by the HOMA2 Calculator according to previous reports.

Western blot analysis

NF-[kappa]b p65 expression was measured in nuclear fraction, while I[kappa]B[alpha], JNK, ERK, and p38 MAPK expression was measured in total proteins. Nuclear extract proteins and total proteins were extracted from renal tissues using commercial nuclear extract kits and following the manufacturer's instructions. The protein concentration was determined using a protein assay kit. All samples were separated on 10% sodium dodecyl sulphate-polyacrylamide gel electrophoresis gels, and western blotting was performed as previously described (Yang et al., 2015). Relative quantitation for western blot analysis was calculated after normalization to the amount of GAPDH protein levels. Antibodies included mouse monoclonal antibodies against GAPDH (1:500), I[kappa]B[alpha] (1:1000), p- I[kappa]B[alpha] (1:1000), JNK (1:1000), p-JNK (1:1000), ERK (1:1000), p-ERK (1:1000), p38 MAPK (1:1000), p-p38 MAPK (1:1000), NF-[kappa] BP65 (1:1000) and p-NF-[kappa] BP65 (1:1000).

Statistical analysis

All results were expressed as mean [+ or -] standard deviation for ten animals in each group. Data sets with multiple comparisons were evaluated with one-way analysis of variance (ANOVA), followed by a post hoc test (Dunnett's test). In all cases, probability values of P < 0.05 were considered statistically significant.


Protodioscin ameliorated dyslipidaemia, insulin resistance, hyperleptinemia and hyperuricaemia in high fructose fed mice

As shown in Tables 1 and 2, high dietary fructose induced metabolic syndrome in mice, characterized by dyslipidaemia, insulin resistance and hyperuricaemia. Compared with the normal control group, the fructose group significantly increased serum levels of TC, TG, insulin and UA (p < 0.001). Treatment with protodioscin at both 5 and 10mg/kg markedly reduced the serum levels of TC (5 mg/kg, p < 0.05; 10 mg/kg, p < 0.01) and TG (5 mg/kg, p<0.01; 10 mg/kg, p< 0.001) in fructose-fed mice. Furthermore, protodioscin significantly attenuated the increase of serum insulin levels induced by high fructose feeding (p<0.05), and improved glucose intolerance in the OGTT (Fig. 2A and B). Protodioscin also significantly reduced HOMA values compared with high fructose-fed group (5 mg/kg, p<0.05; 10 mg/kg, p<0.01). As shown in Table 1, hyperleptinemia was detected in fructose fed mice (p<0.01), which was reversed by the treatment of protodioscin (5 mg/kg, p < 0.05; 10 mg/kg, p < 0.01). At doses of 5 and 10 mg/kg, protodioscin significantly reduced serum UA levels in high fructose-fed mice (5 mg/kg, p<0.01; 10 mg/kg, p< 0.001). These results indicate that protodioscin treatment could effectively attenuate high dietary fructose-induced metabolic syndrome in mice.

Protodioscin improved renal pathomorphology changes and function in high fructose-fed mice

The kidney sections from control mice showed a normal histology (Fig. 3A). Fig. 3B shows the pathological changes of the glomeruli and podocytes of high fructose-fed mice. In high fructose-fed mice, the brush borders of epithelial cells disappeared and the inflammatory cells infiltrated into the interstitium. Protodioscin treatment for 6 weeks significantly ameliorated these changes in the kidneys (Fig. 3D and E). Allopurinol also ameliorated the pathological changes of kidneys (Fig. 3C). As shown in Fig. 3F, the mean glomerular volume of high fructose-fed mice was significantly greater than that of the normal control mice (p < 0.05). However, treatment with protodioscin significantly lowered the glomerular volume (10 mg/kg, p < 0.05). Further, morphometry analysis indicated that protodioscin significantly inhibited (10 mg/kg, p < 0.01) the increase of mesangial expansion index (Fig. 3G). Those results suggested that protodioscin improved glomerular hypertrophy and mesangial expansion.

As shown in Table 2, the kidney index of high fructose-fed mice was 24% higher than the index of the normal control mice (p < 0.01), while treatment with protodioscin significantly reduced the increase of the kidney index (p<0.05). Likewise, BUN and serum Cr levels in high fructose-fed mice were significantly higher than those in normal mice (p< 0.001). Both 5 and 10 mg/kg of protodioscin significantly reduced BUN (p< 0.001) and serum Cr (p< 0.001) levels, indicating improvement of kidney function. Excretion of urinary albumin is a marker of glomerular dysfunction and tubular impairment. In this study, the excretion of urinary albumin was markedly elevated in high fructose-fed mice (p< 0.001). However, treatment with protodioscin for 6 weeks significantly decreased the elevation of urinary albumin excretion (5 mg/kg, p< 0.05; 10 mg/kg, p < 0.01).

Protodioscin reduced the level of renal inflammatory factors in high fructose-fed mice

As expected, high fructose feeding induced renal inflammatory reaction, characterized by significant elevation of IL-1 [beta] (p < 0.001), IL-6 (p < 0.001) and TNF-[alpha] levels (p < 0.01). Protodioscin at both 5 and 10mg/kg significantly reduced IL-1[beta] (5mg/kg, p<0.01; 10 mg/kg, p < 0.001), IL-6 (p < 0.01) and TNF-[alpha] levels (p < 0.01). Allopurinol also demonstrated a similar reduction effect (Fig. 4A. B, and C).

Protodioscin suppresses the activation of NF-[kappa]B p65 by upregulating I[kappa]B[alpha]

Previous studies have shown that NF-[kappa]B activation is important in the pathogenesis of renal inflammation (Xie et al., 2013). To investigate whether protodioscin reduced renal inflammation via the NF-[kappa]B pathway, we examined the activation of NF-[kappa]B p65 (the major subunit of NF-[kappa]B) and I[kappa]B[alpha] (upstream mediator of NF-[kappa]B) in high fructose-fed mice. As shown in Fig. 5A, NF-[kappa]B p65 expression in the kidney did not differ between control and high fructose-fed mice: however, high fructose feeding significantly increased NF-[kappa]B p65 phosphorylation (p < 0.001) and significantly decreased I[kappa]B[alpha] expression (p < 0.001) in the kidney. As shown in Fig. 5B and C, protodioscin at both 5 and 10mg/kg significantly increased renal I[kappa]B[alpha] expression (5mg/kg, p<0.05; 10mg/kg, p < 0.01) and decreased I[kappa]B[alpha] phosphorylation (5mg/kg, p<0.01; 10mg/kg, p < 0.001). Moreover, protodioscin significantly restored high fructose-induced upregulation of renal p-NF-[kappa]B p65 without significant changes in NF-[kappa]B p65 expression (5mg/kg, p<0.01; 10 mg/kg, p< 0.001).

Protodioscin suppresses the MAPK signaling pathway in high fructose-fed mice

The MAPK family comprises p38 MAPK, ERK and JNK (Lakshmanan et al., 2012). Recent studies have demonstrated the potential link between the MAPK family members and renal hypertrophy (Fujita et al., 2004). Therefore, we investigated whether protodioscin would influence p38 MAPK, ERK and JNK protein levels and phosphorylation in the kidneys of high fructose-fed mice. As shown in Fig. 6A, B, and C, high fructose feeding did not affect p38 MAPK, ERK and JNK protein levels in the kidneys of mice, but it markedly elevated p-p38 MAPK (p< 0.001), p-ERK (p < 0.001) and p-JNK (p < 0.001) protein levels. Treatment with protodioscin significantly restored high fructose-induced upregulation of renal p-p38 MAPK (p <0.001), p-ERK (5 mg/kg, p <0.05; 10 mg/kg, p <0.01) and p-JNK (p <0.001) levels in the kidney of high fructose-fed mice.


DR is a main staple of the tropical and subtropical countries (McAnuff-Harding et al., 2006). Protodioscin, one of the primary furostanol saponin of Dioscorea species (including Dioscoreae Rhizome, Dioscoreae Nipponicae Rhizome, Dioscoreae Spongiosae Rhizome and Dioscoreae Hypoglaucae Rhizome), is a precursor of steroid hormones (Liu et al., 2006b). In this study, we evaluated the effect of protodioscin on high fructose-induced metabolic syndrome and renal injury.

It has been suggested that a high fructose diet is closely associated with the development of metabolic syndrome, including hyperleptinemia, insulin resistance and lipid metabolism abnormalities (Aoyama et al., 2012; Malagrino et al., 2016). Since it is most likely that foods that have abundant fructose will continue to be widely available (Stanhope and Havel, 2010), it is necessary to search for alternative compounds against metabolic syndrome induced by high dietary fructose. To the best of our knowledge, this is the first study that demonstrated the protective effects of protodioscin against metabolic syndrome induced by high dietary fructose, as evidenced by the improvement of glucose intolerance and reduced serum levels of insulin, leptin, TG and TC.

High fructose consumption causes chronic inflammation, which has been suggested to contribute to the process of reduced renal function (Dungey et al., 2013). TNF-[alpha], IL-6 and IL-1[beta] are widely used as markers of inflammation, and the increase or persistent elevations in TNF-[alpha], IL-6 and IL-1[beta] are strongly correlated with predictive mortality of chronic renal diseases (Lee et al., 2015a; Meuwese et al., 2011). TNF-[alpha] is cytotoxic to glomerular and mesangial cells and could directly cause renal injury (Kolati et al., 2015). Studies have reported that mice deficient in TNF receptor-1 were resistant to lipopolysaccharide-induced renal failure (Cunningham et al., 2002). In addition, TNF-[alpha] antibodies significantly decreased sepsis-induced renal failure (Cohen and Carlet, 1996). In experimental models of nephropathy, the renal expression of IL-6 increased, which elevated endothelial permeability, mesangial cell proliferation and fibronectin expression (Navarro-Gonzalez and Mora-Fernandez, 2008). IL-1[beta] is also involved in renal injury. IL-1[beta] directly initiates glomerular hypercellularity and increases vascular endothelial cell permeability (Jones et al., 2001; Royall et al., 1989). Consistent with previous research, we observed that high dietary fructose induced renal inflammatory response (as evidenced by increased levels of TNF-[alpha], IL-6 and IL-1[beta]) and caused kidney injury (characterized by increased levels of BUN and serum Cr). However, protodioscin treatment successfully reduced the levels of TNF-[alpha], IL-6 and IL-1[beta], which prevented potential kidney injury.

Numerous clinical and experimental studies have shown that NF-[kappa]B activation plays a critical role in the development of renal injury (Impellizzeri et al., 2014; Xie et al., 2013). NF-[kappa]B is expressed in glomerular endothelial and epithelial cells as well as in tubular epithelial cells of patients with nephropathy and this correlated with the progression of tissue injury (Ashizawa et al., 2003). Xie et al. (2013) have demonstrated that renal NF-[kappa]B is markedly activated in diabetic nephropathy rats. NF-[kappa]B is a transcription factor that regulates numerous cytokines (including IL-1, IL-2, IL-6, IL-8 and TNF-or), correlating with the pathophysiology of renal inflammation and fibrosis (Tomita et al., 2001). In addition, NF-[kappa]B affects the activity of TGF-[beta]1, which plays a central role in the progressive scarring of CKD (Kim et al., 2009; Xie et al., 2013). IkB suppresses DNA binding and NF-[kappa]B transport. Under normal circumstances, NF-[kappa]B combines with I[kappa]B protein as inactive dimers in the cytoplasm. Upon stimulation, I[kappa]B is degraded, and the active NF-[kappa]B is released and translocated to the nucleus (Wang et al., 2011). The results showed that protodioscin markedly increased I[kappa]B[alpha] expression, and attenuated the increase of NF-[kappa]B p65 phosphorylation. It is suggested that protodioscin suppresses NF-[kappa]B activation through stabilization of I[kappa]B[alpha].

Increasing number of studies have reported that the MAPKs family, including p38 MAPK, ERK and JNK is important in the development of renal disease. Adhikary et al. (2004) reported that p38 MAPK regulates the activation and expression of NF-[kappa]B, and leads to the release of pro-inflammatory cytokines (IL-1, IL-6 and TNF-[alpha]) in renal cells (Ahad et al., 2015). Some p38 MAPK inhibitors have demonstrated effectiveness in preclinical disease models including renal disease, such as diabetic nephropathy (Adhikary et al., 2004). It has been reported that ERK is crucial for cell proliferation and differentiation. In addition, ERK is responsible for the overexpression of fibronectin and fibrosis in human mesangial cells (Lakshmanan et al., 2012). Activation of JNK is closely associated with numerous renal diseases, and in addition, pharmacological inhibitors of JNK were shown to attenuate renal inflammation, fibrosis and apoptosis (Lee et al., 2015b). In this study, high fructose stimulated increase phosphorylation of p38 MAPK, ERK and JNK. We further demonstrated that protodioscin treatment significantly prevented the over activation of p38 MAPK, ERK and JNK in high fructose-fed mice.

Allopurinol is known to inhibit XOD activity and reduce the formation of UA. It also prevents liver lipid accumulation and ameliorates renal disease (Hu et al., 2012). A previous study has shown that allopurinol improved renal lipid accumulation and injury under hyperuricemia and hyperlipidemia (Hu et al., 2012). Allopurinol also improved podocyte injury and albuminuria in fructose-induced metabolic syndrome in rats (Wang et al., 2015). The potential mechanism maybe that allopurinol inhibit p38 MAPK/ NOD-like receptor pyrin domain-containing 3 inflammasome pathway activation (Wang et al., 2015). In this paper, allopurinol significantly prevented the over activation of p38 MAPK, ERK1/2, JNK and NF-[kappa]B, and downregulated the renal levels of TNF-[alpha], IL-6 and IL-1 [beta] in high fructose-fed mice. It is also a potential mechanism of allopurinol against metabolic syndrome-associated renal injury.


In summary, the present study demonstrated for the first time that protodioscin attenuates metabolic syndrome and renal injury induced by high dietary fructose in mice. Moreover, protodioscin significantly prevented the overactivation of p38 MAPK, ERK1/2, JNK and NF-[kappa]B, and downregulated the renal levels of TNF-[alpha], IL-6 and IL-1[beta] in high fructose-fed mice. Taken together, the renoprotective activity of protodioscin may be mediated by inhibition of the MAPK pathway to reduce inflammation. These results suggest that protodioscin could be a therapeutic candidate for the treatment of metabolic syndrome and renal injury induced by high dietary fructose. Further study is necessary to elucidate protodioscin's molecular mechanism of action and establish its beneficial activity in humans.


Article history:

Received 3 May 2016

Revised 18 July 2016

Accepted 27 August 2016

Conflict of interest

The authors have declared that there is no conflict of interest.


This work was supported by a project funded by the College Students Innovation Project for the R8D of Novel Drugs (NO. J1030830). The assistance of the staff is gratefully acknowledged.


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Jinyang Shena (1), Xiaolin Yang (b,1), Zhaoqing Meng (c), Changrun Guo (a), *

(a) State Key Laboratory of Natural Medicines. China Pharmaceutical University. No. 24 Tongjia Lane Nanjing 210009, PR China

(b) Jiangsu Key Laboratory of Research and Development in Marine Bio-resource Pharmaceutics. College of Pharmacy. Nanjing University of Chinese Medicine. Nanjing 210023. PR China

(c) Jiangsu Kanion Pharmaceutical Co., Ltd, Lianyungang 222001, PR China

Abbreviations: Alio, allopurinol; BUN. blood urea nitrogen; CKD. chronic kidney disease; CMC-Na, carboxymethylcellulose sodium; Cr. creatinine; Dio. diosgenin; DR. Dioscoreae rhizome; ERK. extracellular signal-regulated kinase; CAPDH. glyceraldehyde-3-phosphate dehydrogenase; HOMA. homeostatic model assessment; I[kappa]B[alpha], inhibitor of NF-[kappa] B-[alpha]; IL-1[beta], interleukin-1[beta]; IL-6, interleukin-6; JNK, c-Jun N-terminal kinases; NF-[kappa] BP65. nuclear transcription factor-[kappa] B P65; OGTT, oral glucose tolerance test; p38 MAPK, protein-38 mitogen activated protein kinases: TC. total cholesterol; TC. triglyceride; TGF-[beta]1, transforming growth factor-[beta]1; UA. uric acid.

* Corresponding author.

E-mail address: (C. Guo).

(1) These authors contributed equally to this work.

Table 1
Effects of protodioscin on insulin resistance, hyperleptinemia.
dyslipidemia and hyperuricemia in high fructose-fed mice.

Groups            Serum insulin (ng/dl)   HOMA Value

Control           4.50 [+ or -] 0.51        1.31 [+ or -] 0.21
Fructose          6.62 [+ or -] 034 ***     2.27 [+ or -] 0.14 ***
Allopurinol       5.36 [+ or -] 0.49 (##)   2.28 [+ or -] 0.19
Protodioscin-5    5.88 [+ or -] 0.62 (#)    2.16 [+ or -] 0.12 (#)
Protodioscin-10   5.54 [+ or -] 0.63 (#)    1.80 [+ or -] 0.13 (##)

Groups            Serum leptin (ng/dl)    Serum TC (mg/dl)

Control           0.46 [+ or -] 0.05        75.7 [+ or -] 14.6
Fructose          0.62 [+ or -] 0.04 **     124.3 [+ or -] 6.5 ***
Allopurinol       0.47 [+ or -] 0.09 (##)   95.5 [+ or -] 16.8 (##)
Protodioscin-5    0.53 [+ or -] 0.06 (#)    107.9 [+ or -] 9.1 (#)
Protodioscin-10   0.49 [+ or -] 0.07 (##)   102.1 [+ or -] 9.0 (##)

Groups            Serum TG (mg/dl)        Serum UA (mg/dl)

Control           134 [+ or -] 24           2.11 [+ or -] 0.22
Fructose          359 [+ or -] 40 ***       3.76 [+ or -] 0.26 ***
Allopurinol       243 [+ or -] 28 (###)     2.68 [+ or -] 0.24 (##)
Protodioscin-5    274 [+ or -] 23 (##)      3.18 [+ or -] 0.23 (#)
Protodioscin-10   250 [+ or -] 24 (###)     2.72 [+ or -] 0.26 (##)

*** p < 0.001 compared with the control group: (#) p < 0.05,
(##) p < 0.01. (###) p < 0.001 compared with the fructose group.

Table 2
Effects of protodioscin on renal dysfunction in high fructose-fed

Groups            Kidney index (g/lOg)      Serum Cr (mg/dl)

Control           0.578 [+ or -] 0.061      1.20 [+ or -] 0.121
Fructose          0.716 [+ or -] 0.073 **   3.50 [+ or -] 0.38 ***
Allopurinol       0.604 [+ or -] 0.056 (#)  1.59 [+ or -] 0.13 (###)
Protodioscin-5    0.646 [+ or -] 0.052 (#)  2.02 [+ or -] 0.26 (###)
Protodioscin-10   0.601 [+ or -] 0.063 (#)  1.89 [+ or -] 0.27 (###)

Groups            BUN (mg/dl)               Urinary albumin (Mg/ml)

Control           10.2 [+ or -] 1.5         21.7 [+ or -] 1.5
Fructose          18.1 [+ or -] 1.7 ***     31.2 [+ or -] 1.2 ***
Allopurinol       12.9 [+ or -] 1.3 (##)    23.4 [+ or -] 1.1 (##)
Protodioscin-5    14.1 [+ or -] 1.2 (#)     26.8 [+ or -] 1.9 (#)
Protodioscin-10   12.4 [+ or -] 1.4 (##)    24.2 [+ or -] 1.4 (##)

*** p < 0.001 compared with the control group: (#) p < 0.05,
(##) p < 0.01. (###) p < 0.001 compared with the fructose group.


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Title Annotation:Original article
Author:Shen, Jinyang; Yang, Xiaolin; Meng, Zhaoqing; Guo, Changrun
Publication:Phytomedicine: International Journal of Phytotherapy & Phytopharmacology
Article Type:Report
Date:Nov 15, 2016
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