Construction of chrysophanol loaded nanoparticles with N-octyl-O-sulfate chitosan for enhanced nephroprotective effect.

Natural occurring anthraquinone like chrysophanol has been studied because of its anti-diabetic, anti-tumor, anti-inflammatory, hepatoprotective and neuroprotective properties. Nonetheless, its poor water solubility and unstable nature are big concerns in achieving efficient delivery and associated pharmacokinetic and pharmacodynamic effects. Herein, this study sought to solve the above-mentioned problem through development of chrysophanol-loaded nanoparticles to enhance the bioavailability of chrysophanol and to evaluate its anti-renal fibrosis effect in rats. After synthesis of a safe N-octyl-O-sulfate chitosan, we used it to prepare chrysophanol-loaded nanoparticles through dialysis technique before we performed and physical characterization. Also, we tested the stability of the nanoparticles for 21 days at 4 °C and room temperature (25 °C) and evaluated their pharmacokinetics and anti-renal fibrosis effect in rat model of chronic kidney disease (CKD). In terms of results, the nano-preparation demonstrated an acceptable narrow size distribution, wherein the encapsulation rate, size, polydispersed index (PDI) and electrokinetic potential at room temperature were respectively 83.41±0.89 %, 364.88±13.62 nm, 0.192±0.015 and 23.78±1.39 mV. During 21 days of storage, we observed that size of particles and electrokinetic potential altered slightly but the difference was statistically insignificant (p > 0.05). Also, in vitro release studies showed that the formulation reached 84.74 % at 24 h. Chrysophanol nanoparticles showed a 2.57-fold increase in bioavailability compared to unformulated chrysophanol. More importantly, chrysophanol nanoparticles demonstrated certain renal internalization properties and anti-renal fibrosis effects, which could ultimately result in reduced blood-urea nitrogen (BUN), kidney-injury molecule-1 (KIM-1) and serum creatinine (SCr) levels in model rats. In conclusion, the prepared chrysophanol-loaded nanoparticles potentially increased bioavailability and enhanced nephroprotective effects of chrysophanol.

Chrysophanol, a main anthraquinone from Rheum palmatum L. (rhubarb), protects against renal fibrosis by suppressing NKD2/NF-κB pathway.

PURPOSE: Chronic kidney disease (CKD), characterized as renal dysfunction and multi-system damage, has become a serious public health problem with high prevalence and mortality. Rheum palmatum L. (rhubarb) is one of the most widely used Chinese herb with renal protective activity. However, the active components and underlying mechanisms of rhubarb remain unknown. In this work, we tried to explore the pharmacological mechanism of chrysophanol, a main anthraquinone from rhubarb, against CKD by in vivo and in vitro models. STUDY DESIGN: The therapeutic effect of chrysophanol and its underlying mechanism were investigated using CKD mouse model induced by unilateral ureteral occlusion (UUO), and human kidney 2 (HK-2) cells stimulated by TGF-ß1 in vivo. METHODS: The impact of chrysophanol on renal function, inflammation, fibrosis of CKD mice were evaluated. Then, the protein expressions of FN1, collagen ɑI, α-SMA, NF-κB and naked keratinocyte homolog 2 (NKD2) were investigated. In vitro studies, the inhibition on inflammation and fibrogenesis by chrysophanol was further validated in TGF-ß1-stimulated HK2 cells, and the regulation of chrysophanol on NKD2/NF-κB pathway was analyzed. Moreover, NKD2 was overexpressed in HK-2 cells to confirm the role of NKD2/NF-κB pathway in chrysophanol-mediated efficacy. Finally, the binding mode of chrysophanol with NKD2 was studied using in silico molecular docking and microscale thermophoresis (MST) assay. RESULTS: Chrysophanol could significantly improve the kidney dysfunction, alleviate renal pathology, and reverse the elevated levels of renal fibrosis markers such as FN1, collagen ɑI and α-SMA. Furthermore, chrysophanol effectively inhibited TNF-α, IL-6, and IL-1ß production, and suppressed NF-κB activation and NKD2 expression. The findings of in vitro study were consistent with those of animal expriment. Using NKD2-overexpressing HK-2 cells, we also demonstrated that overexpression of NKD2 significantly compromised the anti-fibrotic effects of chrysophanol. In addition, molecular docking and MST analysis revealed that NKD2 was a direct target of chrysophanol. CONCLUSION: Together, our work demonstrated for the first time that chrysophanol could effectively ameliorate renal fibrosis by inhibiting NKD2/NF-κB pathway. Chrysophanol can potentially prevent CKD by suppressing renal NKD2 expression directly.

Glycyrrhetinic Acid Protects Renal Tubular Cells against Oxidative Injury via Reciprocal Regulation of JNK-Connexin 43-Thioredoxin 1 Signaling.

Background and Objective: The incidence of chronic kidney disease (CKD) is steadily increasing. Although renal tubular epithelium injury is closely correlated with the prognosis of CKD, the underlying mechanism is not fully understood and therapeutic strategies are limited. The main bioactive component of the Chinese medicine herb, glycyrrhiza, is 18α-glycyrrhetinic acid (Ga), which is also a pharmacological inhibitor of gap junctions. Our previous studies indicated that Ga is able to ameliorate renal cell injury. The present study explored the regulatory role of Ga in redox signaling in renal tubular epithelial cells with oxidative injury. Methods: Rat renal tubular epithelial cells, NRK-52E, were incubated with Px-12, a thioredoxin inhibitor, to mimic thioredoxin deficiency and induce oxidative injury in vitro. A Cell Counting Kit-8 was used to analyze cell viability while a reactive oxygen species (ROS)/superoxide (O2 -) fluorescence probe was employed to determine oxidative stress. Apoptosis was evaluated using DT-mediated dUTP nick end labeling/4,6-diamidino-2-phenylindole staining and cleaved caspase 3 protein analysis. Western blot analysis was used to analyze the expression of specific proteins while siRNA transfection was performed to downregulate targeted proteins. Results: Inhibition of thioredoxin 1 by Px-12 triggered renal tubular cell oxidative injury as evidenced by morphological change, loss of cellular viability, over production of ROS and O2 -, and appearance of cleaved caspase-3. Ga significantly attenuated cell oxidative injury, as indicated by the parameters mentioned above. Px-12 induced phosphorylation of c-Jun N-terminal kinase (JNK) and subsequently the expression of connexin 43 (Cx43) in NRK-52E cells. Ga and the JNK inhibitor, sp600125, markedly suppressed Px-12-induced generation of intracellular ROS and O2 -. Inhibition of JNK improved Px-12-elicited NRK-52E cell injury. Moreover, sp600125 inhibited Cx43 expression. After downregulation of Cx43 via Cx43 siRNA transfection, the phosphorylation of JNK was markedly reduced. Furthermore, Ga restored the expression of thioredoxin 1 inhibited by Px-12. Conclusion: ROS-JNK-Cx43-thioredoxin 1 signaling plays a crucial role in renal tubular cell injury. JNK is involved in the regulation of thioredoxin 1 and Cx43, and Cx43 reciprocally regulates thioredoxin 1. Inhibition of gap junctions by Ga alleviated renal tubular oxidative injury via improvement of thioredoxin 1-mediated redox signaling.

Improved oral bioavailability and anti-chronic renal failure activity of chrysophanol via mixed polymeric micelles.

AIMS: This study was designed to prepare chrysophanol-loaded micelles (CLM) to improve the oral bioavailability, targetability and anti-chronic renal failure (CRF) activity of chrysophanol (CH). METHODS: The preparation of CLM was achieved via thin-film dispersion technique. The in vitro release of CLM compared with free CH was measured in phosphate buffer solution (PBS) containing 0.5%w/v sodium dodecyl sulphate (pH 6.8) while the pharmacokinetic and anti-CRF activity study was also conducted in rats. Moreover, the tissue distribution of CLM was investigated in the mice. RESULTS: The CLM had particle size (PS) of 29.64 ± 0.71 nm, and encapsulation efficiency (EE) of 90.48 ± 1.22%w/w. The cumulative release rate of CH from the micellar system was significantly higher than that of the free CH (86%m/m vs. 15%m/m, p < 0.01). In vivo pharmacokinetic studies showed that the bioavailability of CLM after oral administration was substantially improved (about 3.4 times) compared with free drugs (p < 0.01). Also, it was observed that CLM accumulated well in the liver and brain. Moreover, in vitro renal podocytes study showed that CLM had better protection against renal podocyte damage than the free CH. In addition, CLM significantly (p < 0.01) reduced levels of blood urea nitrogen (BUN), kidney injury molecule-1 (Kim-1), and serum creatinine (SCr), which obviously improved kidney damage in rats with CRF. CONCLUSIONS: Collectively, these findings suggest that mixed micelles may be used as a promising drug delivery system for oral bioavailability improvement and concomitantly enhance the anti-CRF activity of CH, as well as provide a basis for the clinical application of CH.

Ureic clearance granule, alleviates renal dysfunction and tubulointerstitial fibrosis by promoting extracellular matrix degradation in renal failure rats, compared with enalapril.

ETHNOPHARMACOLOGICAL RELEVANCE: Chinese herbal compound prescription has a unique therapeutic action on chronic kidney disease (CKD) in China. In clinics, Uremic Clearance Granules (UCG), a compounded Chinese patent medicine, has been frequently used to treat chronic renal failure (CRF) patients for nearly 30 years, however, the deep therapeutic mechanisms involved in vivo remain a challenge. This study aims to demonstrate the effects and mechanisms of UCG on renal dysfunction and tubulointerstitial fibrosis by regulating extracellular matrix (ECM) degradation and transforming growth factor (TGF)-beta1/Smad signaling activity in vivo, compared with enalapril. MATERIALS AND METHODS: Twenty-six rats were randomly divided into 4 groups, a sham-operated group (Sham group), a vehicle-intervened group (Vehicle group), a UCG-treated group (UCG group) (5g/kg/day) and an enalapril-treated group (Enalapril group) (20mg/kg/day). The rats with renal failure were induced by adenine (150 mg/kg/day) and unilateral ureteral obstruction (UUO), and killed on day 35 after the administration. Proteinuria, urinary N-acetyl-beta-D-glucosaminidase (UNAG), blood biochemical parameters, renal morphological changes, collagen type IV (CIV), matrix metalloproteinase (MMP)-2, MMP-9 and tissue inhibitors of metalloproteinase (TIMP)-1, as well as the key molecular protein expressions in TGF-beta1/Smad signaling pathway were observed, respectively. RESULTS: Adenine administration and UUO induced severe renal damages, as indicated by renal dysfunction, proteinuria and the marked histopathological injuries in the tubules and interstitium, which were associated with MMP-2/TIMP-1 imbalance and TGF-beta1/Smad signaling activity, as shown by up-regulation of the protein expressions of TGF-beta1, TGF-beta receptor type I (RI), TGF-beta receptor type II (RII), Smad2/3, phosphorylated-Smad2/3 (p-Smad2/3) and Smad4, as well as down-regulation of the protein expression of Smad7 in the kidney. UCG treatment, however, significantly not only attenuated renal dysfunction and tubulointerstitial fibrosis, but also improved the protein expressions of MMP-2, TIMP-1, TGF-beta1, TGF-beta RI, p-Smad2/3, Smad4 and Smad7 in the kidney. Besides, the effects of UCG were stronger than those of enalapril partly. CONCLUSION: UCG similar to enalapril, is renoprotective via ameliorating renal dysfunction and tubulointerstitial fibrosis in the renal failure model. The potential mechanisms by which UCG exerts its therapeutical effects in vivo are through promoting ECM degradation and regulating MMP-2/TIMP-1 balance or signaling molecular activity in TGF-beta1/Smad pathway in the kidney. These findings suggest that UCG treatment is undoubtedly useful in preventing the progression of CRF.

[Molecular mechanisms of renal extracellular matrix degradation and interventional effects of Chinese herbal medicine].

The reduction of extracellular matrix (ECM) degradation in kidney is taken as the morphological features and pathological base in renal injury in chronic kidney disease (CKD). ECM degradation is controlled by the catabolic enzyme systems in glomerulus and renal interstitium, in which matrix metalloproteinases (MMPs) play a key role. The expression and activity of MMPs are regulated by the classical pathway, such as the genic transcription, the activation of zymogen, and the specific inhibitor. The previous studies showed that, Uremic Clearance granule, as a representation, and other prescriptions of Chinese herbal medicine, as well as some extracts from Chinese herbal medicine could intervene the pathway of ECM degradation through promoting the degradation of ECM components, affecting the expression of catabolic enzymes, regulating the genetic transcription of MMPs, and inhibiting the relative signaling transduction of MMPs.

[Overview research of uremic ulearance granule treating chronic renal failure].

The effective bioactivity compositions of uremic clearance granul (UCG) include isoflavonoids, emodin, astragaloside, paeoniflorin, salvianolic acid A, and so on. The effects of UCG treating chronic renal failure (CRF) in clinical pharmacodynamics mainly refer to improve renal function and the complications of CRF. The mechanisms involved in vivo basically include depressing transforming growth factor (TGF)-beta1 over-expression, lessening podocyte injury,inhibiting tubular epithelial myofibroblast transdifferentiation, ameliorating microinflammation status, retarding oxidative stress, and alleviating insulin resistance.

[Effects of huangkui capsule on renal inflammatory injury by intervening p38MAPK signaling pathway in rats with adriamycin-induced nephropathy].

OBJECTIVE: To explore the potential mechanisms of huangkui capsule (HKC), an extract from Abelmoschus manihot (AM), for ameliorating renal inflammatory injury by regulating p38 mitogen-activated protein kinase (MAPK) signaling pathway in rats with adriamycin-induced nephropathy (ADRN). METHOD: Nineteen Sprague-Dawley (SD) rats were randomly divided into three groups, the sham-operation group, the untreated model group,and the HKC-treated group. Rats in the untreated model group and the HKC-treated group were made into ADRN model by right nephrectomy and twice intravenous injections of adriamycin( ADR, 0.4 mL and 0.2 mL respectively within 4 weeks). After the model successfully established, rats in the HKC-treated group were orally given HKC (2 mg x kg(-1) per day), while rats in the untreated model group and the sham-operation group were intervened with distilled water respectively. The intervention for all rats was 4 weeks. Rats' body weight were weighted and 24 h urinary protein excretion (Upro) was detected at the end of the 1st, 2nd, 3rd, and 4th week after the intervention of HKC or distilled water. All rats were sacrificed at the end of the 8th week after nephrectomy, and then, to withdraw blood and kidney to examine the blood biochemical parameters, the glomerular morphological changes, alpha-smooth muscle actin (alpha-SMA) and collagen type I expressions,and the glomerular macrophages infiltration. Besides, the protein expression of transforming growth factor (TGF)-beta1, p38MAPK, as well as phosphorylated p38MAPK (p-p38MAPK) in renal tissues were detected by Western blotting. RESULT: As compared with rats in the untreated model group, in the HKC-treated group,the HKC treatment significantly improved Upro, serum albumin, mesangial cell proliferation, extracellular matrix (ECM) and collagen deposition,and decreased the expression of alpha-SMA and collagen type I and the infiltration of ED1+ and ED3+ cells in glomeruli. In addition, it significantly down-regulated the protein expression of TGF-beta1 and p-p38MAPK in renal tissues. CONCLUSION: HKC had the effects on ameliorating renal inflammatory injury in vivo. It could reduce the expression of TGF-beta1 and improve the infiltration and activation of inflammatory cells in glomeruli by way of intervening p38MAPK signaling pathway in kidney through down-regulating the protein expression of p-p38MAPK, as the key signal molecule.

[Mechanisms and effects of Abelmoschus manihot preparations in treating chronic kidney disease].

Abelmoschus manihot (AM) is a medicinal plant rich in twenty kinds of separated active bio-components including flavones, polysaccharides, trannic acid, and long chain hydrocarbons. Among these, total flavones of A. manihot (TFA) are the major active component. In this review, the mechanisms of Huangkui capsule will be discussed as a preparation of AM to treat chronic kidney disease (CKD) by improving immunological reaction, inflammation, renal fibrosis, and renal tubular epithelial injury. Additionally, it has been reported that Huangkui capsule can ameliorate some clinical symptoms, proteinuria, hematuria, and renal function in patients with common CKD, such as nephrotic syndrome, diabetic nephropathy, Henoch-Schönlein purpura nephritis, IgA nephropathy, and membranous nephropathy.

[Regulatory mechanism of p38MAPK signaling pathway on renal tissue inflammation in chronic kidney disease and interventional effect of traditional Chinese medicine].

The inflammatory reaction of renal tissues and its relevant tissue damages (such as glomerulosclerosis and renal interstitial fibrosis) are important factors for the development of chronic kidney diseases (CKD) to end-state renal diseases. Of them, p38 mitogen-activated protein kinase (MAPK) signaling pathway plays an important role in regulating expression and bioactivity of multiple nuclear transcription factors, impacting synthesis of downstream inflammatory mediators and activating inflammatory cells. Some monomer traditional Chinese medicines and their extracts (such as emodin and berberine) and some traditional Chinese medicine compound prescriptions (such as Yishen Huoxue decoction) can affect inflammatory reaction of renal tissues by regulating p38MAPK signaling pathway, thas improving reduce glomerulus and renal interstitial inflammatory injury.

Contrasting dose-effects of multi-glycoside of Tripterygium wilfordii HOOK. f. on glomerular inflammation and hepatic damage in two types of anti-Thy1.1 glomerulonephritis.

Multi-glycoside, one of the extracted compounds from Tripterygium wilfordii HOOK. f. (GTW), has been shown to be clinically effective in suppressing glomerular inflammation in chronic kidney disease. However, its clinical application is often limited by its cytotoxic actions on the liver. This study was performed to contrast the dose-effects of GTW on glomerular inflammation and hepatic damage in two types of anti-Thy1.1 glomerulonephritis (GN). Rats with acute or chronic anti-Thy1.1 GN were either left untreated (the Vehicle group) or treated with a high or low dose of GTW and sacrificed on day 7 or day 45. GTW was administrated 3 days before or at the same time as the antibody injection and lasted until sacrifice. GTW at high dose ameliorated glomerular macrophage accumulation, mesangial proliferation, proteinuria, and interleukin-2 expression in the acute anti-Thy1.1 GN model, but caused structural and functional lesions in the liver. In contrast, GTW at low dose improved activated macrophage and T lymphocyte infiltration, mesangial injury, proteinuria, and interleukin-2 and interferon-γ expressions without hepatic toxicity in the chronic model of GN induced by anti-Thy1.1 antibody. In conclusion, GTW at low dose not only effectively inhibits glomerular inflammation but also avoids severe injuries to the liver.

[Mechanisms and effects of chinese herbal medicine delaying progression of chronic renal failure].

In this review,firstly,it has been discussed the mechanisms of Chinese herbal medicine ameliorating glomerulosclerosis and renal interstitial fibrosis during the progression of chronic renal failure (CRF) by improving glomerular hemodynamics turbulence, podocyte injury, transforming growth factor (TGF)-beta over-expression, hyperlipidemia, macrophage infiltration, tubular epithelial myofibroblast transdifferentiation, and nephrotoxicity of proteinuria. Secondly,it has been reported the clinical effects of Chinese herbal medicine improving renal function and some clinical complications in the patients with progressive CRF through various treatments including oral administration or coloclysis of Chinese herbal medicine, oral administration combined with coloclysis of Chinese herbal medicine, and colonic dialysis combined with coloclysis of Chinese herbal medicine. Finally,it has been reviewed the beneficial influences of Chinese herbal medicine on metabolic dysequilibrium of calcium and phosphonium, microinflammatory state, and uremic toxins in patients with uremia.