Functional and histological effects of Anthurium schlechtendalii Kunth extracts on adenine-induced kidney damage of adult Wistar rats.

Anthurium schlechtendalii Kunth is used by the Zoque group in southeastern Mexico for kidney and urinary diseases, but its safety and effectiveness are unproven, therefore a model of adenine-induced renal failure in rats was performed. The rats were fed with solid and aqueous plant extracts for 4 weeks to study its effects on kidney histological morphology. Kidneys were examined, and statistical analysis was performed. The adenine-containing diet caused renal failure, characterized by crystal deposits, cystic dilatation of tubules, and micro-abscesses. Both extracts caused tubular damage and collagen increase without inflammation. However, when combined with adenine, the extracts showed some protective effects, although cystic dilatation and granulomatous inflammation were observed. The extracts at the tested doses resulted in glomerular and tubular damage, aggravating cystic degeneration, therefore, its indiscriminate use in Humans is not safe. Additionally, the extracts can serve as a model for studying renal damage without crystal deposits.

Vascular, valvular and kidney calcification manifested in mouse models of adenine-induced chronic kidney disease.

BACKGROUND: Ectopic calcification (EC) involves multiple organ systems in chronic kidney disease (CKD). Previous CKD-animal models primarily focused on a certain histological abnormality but did not show the correlation with calcified development among various tissues. This study compared calcified deposition in various tissues during CKD progression in mice. METHODS: Male 8-week-old C57BL/6J mice were randomly allocated to the seven groups: a basic, adenine, high-phosphorus, or adenine and high-phosphorus diet for 12-16 weeks (Ctl16, A12, P16, or AP16, respectively); an adenine diet for 4-6 weeks; and a high-phosphorus or adenine and high-phosphorus diet for 10-12 weeks (A6 + P10, A4 + P12, or A4 + AP12, respectively). RESULTS: Compared to the Ctl16 mice, the P16 mice only displayed a slight abnormality in serum calcium and phosphorus; the A12 mice had the most serious kidney impairment; the A4 + P12 and A6 + P10 mice had similar conditions of CKD, mineral abnormalities, and mild calcification in the kidney and aortic valves; the A4 + AP12 and AP16 groups had severe kidney impairment, mineral abnormalities and calcification in the kidneys, aortic valves and aortas. Furthermore, calcium-phosphate particles were deposited not only in the tubulointerstitial compartment but in the glomerular and tubular basement membrane. The elemental composition of EC in various tissues matched the calcification of human cardiovascular tissue as determined by energy dispersive spectroscopy. CONCLUSIONS: The severity of CKD was unparalleled with the progression of mineral metabolism disorder and EC. Calcification was closely related in different tissues and observed in the glomerular and tubular basement membranes.

Previous CKD-animal models primarily focused on a certain histological abnormality but lacked investigations of the interplay of EC in various tissues. This study compared calcified deposition in several tissues during CKD progression in mice, which was closely related. The severity of CKD was unparalleled with the development of ectopic calcification. Glomerular and tubular basement membrane calcification was detected in CKD mice, which has been considered extremely rare in clinical.

Integration of network pharmacology and intestinal flora to investigate the mechanism of action of Chinese herbal Cichorium intybus formula in attenuating adenine and ethambutol hydrochloride-induced hyperuricemic nephropathy in rats.

CONTEXT: Cichorium intybus L. (Asteraceae) formula (CF) has been applied as a folk medicine to treat hyperuricemic nephropathy (HN). However, the exact mechanism remains unclear. OBJECTIVE: To explore the therapeutic effect and mechanism of CF on HN. MATERIALS AND METHODS: Through network pharmacological methods, the targets of the active component of CF against HN were obtained. Subsequently, Male Wistar rats were divided into control, HN, allopurinol (50 mg/kg), CF high-dose (8.64 g/kg) and CF low-dose (2.16 g/kg) groups. The HN model was induced via intragastric administration of adenine (100 mg/kg) and ethambutol hydrochloride (250 mg/kg) for 3 weeks. After CF treatment, biochemical indicators including UA, UREA and CREA were measured. Then, HE staining, qRT-PCR and gut microbiota analysis were conducted to further explore the mechanism. RESULTS: The network pharmacology identified 83 key targets, 6 core genes and 200 signalling pathways involved in the treatment of HN. Compared to the HN group, CF (8.64 g/kg) significantly reduced the levels of UA, UREA and CREA (from 2.4 to 1.57 µMol/L, from 15.87 to 11.05 mMol/L and from 64.83 to 54.83 µMol/L, respectively), and mitigated renal damage. Furthermore, CF inhibited the expression of IL-6, TP53, TNF and JUN. It also altered the composition of gut microbiota, and ameliorated HN by increasing the relative abundance of some probiotics. CONCLUSIONS: This work elucidated the therapeutic effect and underlying mechanism by which CF protects against HN from the view of the biodiversity of the intestinal flora, thus providing a scientific basis for the usage of CF.

Total flavonoids in Epimedium koreanum Nakai alleviated chronic renal failure via promoting AMPK activation.

Chronic renal failure (CRF) is a result of the progression of chronic kidney diseases (CKD), a global health problem with a high cost of treatment and no ideal therapy. The aim of this study is to evaluate the pharmacological efficacy of the total flavonoids in Epimedium koreanum Nakai (TFE), a dietary supplement, against CRF and to determine the mechanism of actions. An adenine-induced CRF rat model and a TGF-ß1 induced human kidney proximal tubule epithelial (HK-2) cell based in vitro renal fibrosis model were established and used to evaluate TFE's efficacy. Renal hemodynamics, biochemical indexes, inflammatory cytokines, histopathology and the reactive oxygen species (ROS) levels were determined to evaluate the efficacy of TFE on CRF. NMR-based metabolomics, immunohistochemical (IHC) staining, immunofluorescence (IF) staining, quantitative real time-PCR (qRT-PCR) and western blotting were conducted to determine the mechanism. The results showed that TFE had a significant effect on CRF at 150 mg kg-1 d-1 and could significantly alleviate renal fibrosis in the animal model. Twelve potential biomarkers, which mainly involve energy metabolism pathways, for CRF were identified using the metabolomics approach. The mechanism study suggested that TFE regulated AMP-activated protein kinase (AMPK)/acetyl-CoA carboxylase (ACC) and AMPK/silent information regulator 1 (SIRT1)/nuclear factor kappa-B (NF-κB) signaling pathways. Furthermore, the effect of TFE was inhibited by compound C in the in vitro experiment, which also confirmed the above conclusion.

Huangjinsan ameliorates adenine-induced chronic kidney disease by regulating metabolic profiling.

Chronic kidney disease is an increasingly serious public health problem worldwide. Our recent studies have shown that Huangjinsan has a renal protective effect on chronic kidney disease, but the specific mechanism by which this effect occurs is not clear. To study the therapeutic effect of Huangjinsan on chronic kidney disease and to explore its possible mechanism of action through nontargeted metabolomics methods, a chronic kidney disease rat model was induced by adenine, and the Huangjinsan extract was given by oral gavage. Body weight, the kidney index, pathological sections, and a series of biochemical indicators were measured. High-performance liquid chromatography quadrupole time-of-flight mass spectrometry was used to analyze the changes in the plasma metabolome. Huangjinsan significantly reduced indicators of kidney damage, including total protein, albumin, the total protein to creatinine ratio, and the albumin to creatinine ratio in urine, as well as IL-2, MCP-1α, and blood urea levels in plasma. Based on nontargeted metabolomics, 13 metabolites related to chronic kidney disease were discovered. These metabolites are closely related to glycerophospholipid metabolism, arginine and proline metabolism, and sphingolipid metabolism. We found that Huangjinsan can restore the renal function of adenine-induced chronic kidney disease by regulating the metabolic profile.

Effects of prebiotic oligofructose-enriched inulin on gut-derived uremic toxins and disease progression in rats with adenine-induced chronic kidney disease.

Recent studies suggest that dysbiosis in chronic kidney disease (CKD) increases gut-derived uremic toxins (GDUT) generation, leads to systemic inflammation, reactive oxygen species generation, and poor prognosis. This study aimed to investigate the effect of oligofructose-enriched inulin supplementation on GDUT levels, inflammatory and antioxidant parameters, renal damage, and intestinal barrier function in adenine-induced CKD rats. Male Sprague-Dawley rats were divided into control group (CTL, n = 12) fed with standard diet; and CKD group (n = 16) given adenine (200 mg/kg/day) by oral gavage for 3-weeks to induce CKD. At the 4th week, CKD rats were subdivided into prebiotic supplementation (5g/kg/day) for four consecutive weeks (CKD-Pre, n = 8). Also, the control group was subdivided into two subgroups; prebiotic supplemented (CTL-Pre, n = 6) and non-supplemented group (CTL, n = 6). Results showed that prebiotic oligofructose-enriched inulin supplementation did not significantly reduce serum indoxyl sulfate (IS) but did significantly reduce serum p-Cresyl sulfate (PCS) (p = 0.002) in CKD rats. Prebiotic supplementation also reduced serum urea (p = 0.008) and interleukin (IL)-6 levels (p = 0.001), ameliorated renal injury, and enhanced antioxidant enzyme activity of glutathione peroxidase (GPx) (p = 0.002) and superoxide dismutase (SOD) (p = 0.001) in renal tissues of CKD rats. No significant changes were observed in colonic epithelial tight junction proteins claudin-1 and occludin in the CKD-Pre group. In adenine-induced CKD rats, oligofructose-enriched inulin supplementation resulted in a reduction in serum urea and PCS levels, enhancement of the antioxidant activity in the renal tissues, and retardation of the disease progression.

Lack of Endothelial Nitric Oxide Synthase Accelerates Ectopic Calcification in Uremic Mice Fed an Adenine and High Phosphorus Diet.

Ectopic calcification is a risk of cardiovascular disease in chronic kidney disease (CKD) patients, and impaired endothelial nitric oxide synthase (eNOS) is involved in the CKD complications. However, whether eNOS dysfunction is a cause of ectopic calcification in CKD remains to be elucidated. To address this issue, we investigated the role of eNOS in ectopic calcification in mice with renal injury caused by an adenine and high-phosphorus (Ade + HP) diet. DBA/2J mice, a calcification-sensitive strain, were fed Ade + HP for 3 weeks. Expression levels of eNOS-related genes were reduced significantly in their calcified aorta. C57BL/6J is a calcification-resistant strain, and wild-type mice showed mild calcified lesions in the aorta and kidney when given an Ade + HP diet for 4 weeks. In contrast, a lack of eNOS led to the development of severe aortic calcification accompanied by an increase in runt-related transcription factor 2, an osteochondrogenic marker. Increased renal calcium deposition and the tubular injury score were remarkable in mice lacking eNOS-fed Ade + HP. Exacerbation of ectopic calcification by a lack of eNOS is associated with increased oxidative stress markers such as nicotinamide adenine dinucleotide phosphate oxidases. In conclusion, eNOS is critically important in preventing ectopic calcification. Therefore, the maintenance of eNOS is useful to reduce cardiovascular disease events and to improve prognosis in CKD patients.

AT2 receptor stimulation inhibits phosphate-induced vascular calcification.

Vascular calcification is a common finding in atherosclerosis and in patients with chronic kidney disease. The renin-angiotensin system plays a role in the pathogenesis of cardiovascular remodeling. Here, we examined the hypothesis that angiotensin II type 2 receptor (AT2) stimulation has inhibitory effects on phosphate-induced vascular calcification. In vivo, calcification of the thoracic aorta induced by an adenine and high-phosphate diet was markedly attenuated in smooth muscle cell-specific AT2-overexpressing mice (smAT2-Tg) compared with wild-type and AT2-knockout mice (AT2KO). Similarly, mRNA levels of relevant osteogenic and vascular smooth muscle cell marker genes were unchanged in smAT2-Tg mice, while their expression was significantly altered in wild-type mice in response to high dietary phosphate. Ex vivo, sections of thoracic aorta were cultured in media supplemented with inorganic phosphate. Aortic rings from smAT2-Tg mice showed less vascular calcification compared with those from wild-type mice. In vitro, calcium deposition induced by high-phosphate media was markedly attenuated in primary vascular smooth muscle cells derived from smAT2-Tg mice compared with the two other mouse groups. To assess the underlying mechanism, we investigated the effect of PPAR-γ, which we previously reported as one of the possible downstream effectors of AT2 stimulation. Treatment with a PPAR-γ antagonist attenuated the inhibitory effects on vascular calcification observed in smAT2-Tg mice fed an adenine and high-phosphate diet. Our results suggest that AT2 activation represents an endogenous protective pathway against vascular calcification. Its stimulation may efficiently reduce adverse cardiovascular events in patients with chronic kidney disease.

Production Inhibition and Excretion Promotion of Urate by Fucoidan from Laminaria japonica in Adenine-Induced Hyperuricemic Mice.

This work aims to explore the amelioration of fucoidan on adenine-induced hyperuricemia and hepatorental damage. Adenine-induced hyperuricemic mice were administered with fucoidan, allopurinol and vehicle control respectively to compare the effects of the drugs. Serum uric acid, urea nitrogen, hepatorenal functions, activities of hepatic adenosine deaminase (ADA), xanthine oxidase (XOD), renal urate transporter 1 (URAT1) and NF-κB p65 were assessed. As the serum uric acid, urea nitrogen, creatinine, glutamic oxalacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), superoxide dismutase (SOD), catalase (CAT) and malondialdehyde (MDA) data demonstrated, the adenine not only mediated hepatorenal function disorders, but also induced hyperuricemia in mice. Meanwhile, activities of hepatic ADA and XOD were markedly augmented by adenine, and the expression of URAT1 was promoted, which was conducive to the reabsorption of urate. However, exposure to fucoidan completely reversed those adenine-induced negative alternations in mice, and the activities of hepatic ADA and XOD were recovered to the normal level. It was obvious that hepatic and renal functions were protected by fucoidan treatment. The expression of URAT1 was returned to normal, resulting in an increase of renal urate excretion and consequent healing of adenine-induced hyperuricemia in mice. Expression and activation of NF-κB p65 was promoted in kidneys of adenine treated mice, but suppressed in kidneys of mice exposed to fucoidan from Laminaria japonica or allopurinol. In conclusion, the fucoidan is a potential therapeutic agent for the treatment of hyperuricemia through dual regulatory roles on inhibition of hepatic metabolism and promotion of renal excretion of urate.

Omega-3 polyunsaturated fatty acids alleviate adenine-induced chronic renal failure via regulating ROS production and TGF-ß/SMAD pathway.

OBJECTIVE: To explore the role of omega-3 polyunsaturated fatty acids (ω-3 PUFAs) in adenine-induced rat chronic renal failure and its underlying mechanism. MATERIALS AND METHODS: 30 Sprague Dawley (SD) rats were randomly assigned into three groups, namely sham group, adenine induction group (adenine group) and adenine induction + ω-3 PUFAs treatment group (ω-3 PUFAs group), with 10 rats in each group. Serum and kidney samples were collected after rats were sacrificed. Serum levels of Cr (creatinine) and BUN (urea nitrogen) were detected using commercial kits. HE (hematoxylin and eosin) staining was performed to evaluate the pathological changes of kidneys. Levels of oxidative stress indicators in rat kidney homogenate were detected by relative commercial kits, including SOD (superoxide dismutase), GSH (reduced glutathione), CAT (catalase), and T-AOC (total antioxidant capacity). Reactive oxygen species (ROS) production was also detected by immunofluorescence. Protein expressions of nuclear factor E2 related factor 2 (Nrf2) and transforming growth factor-beta (TGF-ß)/SMAD pathway-related genes were detected by Western blot. RESULTS: Serum levels of Cr and BUN in ω-3 PUFAs group were remarkably decreased compared with those of adenine group. Higher contents of SOD, GSH, CAT and T-AOC were observed in ω-3 PUFAs group compared with those of adenine group. Besides, MAD content and ROS production were lower in ω-3 PUFAs group than those of adenine group. Pathological changes of kidneys were alleviated after ω-3 PUFAs treatment. Western blot results demonstrated that ω-3 PUFAs treatment remarkably upregulates Nrf2, HO-1, NQO1, but downregulates relative genes in TGF-ß/SMAD pathway. CONCLUSIONS: ω-3 PUFAs alleviated adenine-induced chronic renal failure through enhancing antioxidant stress and inhibiting inflammatory response via regulating Nrf2 and TGF-ß/SMAD pathway.

Chitosan-Fe (III) Complex as a Phosphate Chelator in Uraemic Rats: A Novel Treatment Option.

Phosphate retention and hyperphosphataemia are associated with increased mortality in patients with chronic kidney disease (CKD). We tested the use of cross-linked iron chitosan III (CH-FeCl) as a potential phosphate chelator in rats with CKD. We evaluated 96 animals, divided equally into four groups (control, CKD, CH-FeCl and CKD/CH-FeCl), over 7 weeks. We induced CKD by feeding animals an adenine-enriched diet (0.75% in the first 4 weeks and 0.1% in the following 3 weeks). We administered 30 mg/kg daily of the test polymer, by gavage, from the third week until the end of the study. All animals received a diet supplemented with 1% phosphorus. Uraemia was confirmed by the increase in serum creatinine in week 4 (36.24 ± 18.56 versus 144.98 ± 22.1 µmol/L; p = 0.0001) and week 7 (41.55 ± 22.1 versus 83.98 ± 18.56 µmol/L; p = 0.001) in CKD animals. Rats from the CKD group treated with CH-FeCl had a 54.5% reduction in serum phosphate (6.10 ± 2.23 versus 2.78 ± 0.55 mmol/L) compared to a reduction of 25.6% in the untreated CKD group (4.75 ± 1.45 versus 3.52 ± 0.74 mmol/L, p = 0.021), between week 4 and week 7. At week 7, renal function in both CKD groups was similar (serum creatinine: 83.98 ± 18.56 versus 83.10 ± 23.87 µmol/L, p = 0.888); however, the CH-FeCl-treated rats had a reduction in phosphate overload measured by fractional phosphate excretion (FEPi) (0.71 ± 0.2 versus 0.4 ± 0.16, p = 0.006) compared to the untreated CKD group. Our study demonstrated that CH-FeCl had an efficient chelating action on phosphate.

Protective effects of Salvia miltiorrhiza on adenine-induced chronic renal failure by regulating the metabolic profiling and modulating the NADPH oxidase/ROS/ERK and TGF-ß/Smad signaling pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Chronic renal failure (CRF) is defined as a progressive and irreversible loss of renal function and associated with inflammation and oxidative stress. Salvia miltiorrhiza (SM) is an important Chinese herb used in traditional Chinese medicine for treating cardiovascular diseases. The previous studies showed the SM exhibited significant protective effects on CRF. In this present study, the metabolic profiling changes and action mechanism of SM on CRF were explored. AIMS OF THE STUDY: The aims of this study were to illustrate the metabolic profiling changes of adenine induced CRF and analyze the protective effects and action mechanisms of SM ethanol extract (SMEE) and water extract (SMWE). MATERIALS AND METHODS: The animals were divided into normal group, CRF model group, Huangkui capsule-treated group, SMEE-treated group and SMWE-treated group. The UPLC-QTOFMS coupled with multivariate statistical methods were used to explore the changes of metabolic profile in plasma, urine and renal tissue from CRF rats simultaneously after treatment with SMEE and SMWE. Hematoxylin eosin (HE) staining and Masson staining were applied to observe pathological changes in renal tissue. Biochemical indicators including serum urea nitrogen (BUN), urine protein (UP) and serum creatinine (Scr) were measured according to the manufacturer's instructions of kits. Furthermore, HK-2 cell damaged model induced by ISF was established to access the protective effects and action mechanism. The dichlorodihydrofluorescein diacetate (DCFH-DA) assay was used to determine the reactive oxygen species (ROS) and Western blot was applied to analyze the expression of pathogenesis-related proteins in different groups. RESULTS: The results showed that the ethanol extract (SMEE) and water extract (SMWE) of SM significantly inhibited the elevation of serum creatinine (Scr), blood urea nitrogen (BUN), urine protein (UP) and indoxyl sulfate (ISF) in adenine-induced CRF rats, especially SMEE exhibited more significant effects. Moreover, SM extracts obviously improved the symptoms of glomerular and tubular atrophy, focal calcium deposits, interstitial fibrosis, interstitial inflammation, and renal tissues. By metabolomics analysis, fifty-nine metabolites (thirteen in plasma, twenty-seven in urine and nineteen in kidney tissue) were up-regulated or down-regulated and contributed to CRF progress. After treatment of SM extracts, the altered metabolites were restored back to normal level. These potential biomarkers underpinning the metabolic pathways are including phenylalanine metabolism, pyrimidine metabolism, purine metabolism and tryptophan metabolism. Furthermore, SM extracts prevent epithelial-mesenchymal transition (EMT) of human renal tubular epithelial (HK-2) cell by inhibiting NADPH oxidase/ROS/ERK and TGF-ß/Smad signaling pathways. CONCLUSIONS: SMEE and SMWE can significantly alleviate adenine-induced CRF via regulation of the metabolic profiling and modulation of NADPH oxidase/ROS/ERK and TGF-ß/Smad signaling pathways, which provided important supports for the development of protective agent of SM for CRF.

Renoprotective effect of Chinese chive polysaccharides in adenine-induced chronic renal failure.

In this work, we investigated the effect of Chinese chive polysaccharides (CCP) on renal function in mice with adenine-induced chronic renal failure (CRF). Results exhibited that adenine treatment caused serious renal pathological damages and elevation of serum creatinine and blood urea nitrogen of mice. However, these changes could be significantly reversed by the administration of CCP in a dose-dependent manner. When CCP dosage reached 200mg/kg/day, the area of renal pathological damage was decreased by 59.2%, and the levels of serum creatinine and blood urea nitrogen were decreased by 23.9% and 34.7% compared to those of model group. Moreover, it was found that renal oxidative damage, inflammation and fibrosis of adenine-induced CRF mice could also be significantly inhibited by CCP. These results suggested that CCP could improve the kidney functions of adenine-induced CRF mice and the renoprotective effect might be associated with its antioxidant, anti-inflammatory and anti-fibrosis activities.

Apigenin, a novel candidate involving herb-drug interaction (HDI), interacts with organic anion transporter 1 (OAT1).

BACKGROUND: Apigenin is a flavonoid compound, widely distributed in natural plants. Various studies have suggested that apigenin has inhibitory effects towards several drug transporters, such as the organic anion transporting (OAT) polypeptides, 1B1 and 1B3 (OATP1B1 and OATP1B3). However, the mechanism by which apigenin interacts with OAT1 has not been well studied. METHODS: MDCK cells stably-expressing OAT1 were used to examine the inhibitory effects of apigenin on OAT1. UPLC-MS/MS was used to evaluate the in vitro and in vivo effects of apigenin on the uptake of acyclovir by OAT1. Cytotoxicity was determined by the cell viability, MTT assays. RESULTS: Apigenin effectively inhibited the activity of OAT1 in a dose-dependent manner with an IC50 value of 0.737µM. Pre-incubation of cells with apigenin caused a time-dependent inhibition (TDI) of OAT1. Additionally, we examined the interactions between apigenin and acyclovir or adefovir. Data showed that apigenin (1µM) significantly blocked the uptake of acyclovir by OAT1 in vitro with an inhibition rate of 55%. In vivo, apigenin could increase the concentration of acyclovir in plasma when co-administered with acyclovir. Importantly, the MTT assays showed that, at a dose of 50µM, apigenin significantly reduced the cytotoxicity of adefovir and substantially increased cell viability from 50.6% to 112.62%. CONCLUSION: Our results demonstrate that apigenin regulates OAT1, and can cause TDI or herb-drug interaction (HDI) when used in combination with acyclovir or adefovir. Therefore, apigenin could be used as a nephroprotective agent when used in combination with the substrates of OAT1.

Effect of aqueous extract and anthocyanins of calyces of Hibiscus sabdariffa (Malvaceae) in rats with adenine-induced chronic kidney disease.

OBJECTIVES: The aim of this work was to assess the possible beneficial effects of aqueous extracts of Hibiscus sabdariffa L. calyces and anthocyanins isolated therefrom in an adenine-induced chronic kidney disease (CKD) model. METHODS: Rats were orally given, for 28 consecutive days, either adenine alone or together with either aqueous extract of H. sabdariffa calyces (5 and 10%) or anthocyanins (50, 100 and 200 mg/kg of anthocyanin concentrate). For comparative purposes, two groups of rats were given lisinopril (10 mg/kg). KEY FINDINGS: When either H. sabdariffa aqueous extract or the anthocyanins isolated from it was administered along with adenine, the adverse effects of adenine-induced CKD were significantly lessened, mostly in a dose-dependent manner. The positive effects were similar to those obtained by administration of lisinopril. CONCLUSIONS: The results obtained show that both H. sabdariffa and its anthocyanins could be considered as possible promising safe dietary agents that could be used to attenuate the progression of human CKD. This could have added significance as H. sabdariffa tea is widely consumed in many parts of Africa and Asia and is thus readily available.

Near-infrared photothermal therapy using EGFR-targeted gold nanoparticles increases autophagic cell death in breast cancer.

Although triple negative breast cancer (TNBC) is a small percentage of all breast cancers, to date, TNBC is one of the most challenging types of breast cancer for basic and clinic research because TNBC patients display a high risk of relapse, shorter overall survival and limited therapeutic options after completion of conventional chemotherapy compared with patients with other breast cancer subtypes. The epidermal growth factor receptor (EGFR) is a promising target for TNBC treatment. Although near infrared-photothermal therapy (NIR-PTT) using anti-EGFR antibody-conjugated gold nanorods (anti-EGFR-GNs), has attracted considerable interest for non-invasive and targeted TNBC treatment through an activation of apoptotic pathway, it is unclear whether anti-EGFR-GNs-combined NIR-PTT modulates the induction of autophagy contributing to cell death. Therefore, we investigated the autophagic cell death in cultured TNBC cells and mouse xenograft tumors during anti-EGFR-GNs-combined NIR-PTT. We here found that the cytotoxicity induced by anti-EGFR-GNs-combined NIR-PTT was rescued by treatment with autophagy inhibitor, 3-methyladenine (3-MA). Anti-EGFR-GNs-combined NIR-PTT induced remarkable levels of autophagy activity as evidenced by a large number of autophagic vesicles and a significant increase in autophagy-specific proteins; microtubule-associated protein light chain 3 (LC3), p62, beclin-1, and autophagy-related gene5 (Atg5), accompanying the inhibition of AKT-mTOR signaling pathway responsible for inducing autophagy. Moreover, in mouse xenograft tumors, anti-EGFR-GNs-combined NIR-PTT also increased LC3 and beclin-1 levels. Our findings, for the first time, demonstrate that anti-EGFR-GNs-combined NIR-PTT remarkably induces autophagy leading to EGFR-targeted cancer cell death.

Rhein reverses Klotho repression via promoter demethylation and protects against kidney and bone injuries in mice with chronic kidney disease.

Rhein is an anthraquinone compound isolated from the medicinal plant rhubarb and mainly used in the clinical treatment of diabetic nephropathy. Rhein exhibits various renoprotective functions, but the underlying mechanisms are not fully determined. However, its renoprotective properties recapitulate the role of Klotho, a renal-specific antiaging protein critical for maintaining kidney homeostasis. Here we explored the connections between rhein renoprotection and Klotho in a mouse model of adenine-induced chronic kidney disease. In addition to being an impressive Klotho upregulator, rhein remarkably reversed renal Klotho deficiency in adenine-treated mice. This effect was associated with significant improvement in disturbed serum biochemistry, profibrogenic protein expression, and kidney and bone damage. Further investigation of the molecular basis of Klotho loss revealed that these kidneys displayed marked inductions of DNA methyltransferase DNMT1/DNMT3a and Klotho promoter hypermethylation, whereas rhein treatment effectively corrected these alterations. The renal protective effects of rhein were largely abolished when Klotho was knocked-down by RNA interferences, suggesting that rhein reversal of Klotho deficiency is essential for its renoprotective actions. Thus, our study clarifies how rhein regulation of Klotho expression contributes to its renoprotection and brings new insights into Klotho-targeted strategy for the treatment of kidney diseases of various etiologies.

Activation of AMP-Activated Protein Kinase by Adenine Alleviates TNF-Alpha-Induced Inflammation in Human Umbilical Vein Endothelial Cells.

The AMP-activated protein kinase (AMPK) signaling system plays a key role in cellular stress by repressing the inflammatory responses induced by the nuclear factor-kappa B (NF-κB) system. Previous studies suggest that the anti-inflammatory role of AMPK involves activation by adenine, but the mechanism that allows adenine to produce these effects has not yet been elucidated. In human umbilical vein endothelial cells (HUVECs), adenine was observed to induce the phosphorylation of AMPK in both a time- and dose-dependent manner as well as its downstream target acetyl Co-A carboxylase (ACC). Adenine also attenuated NF-κB targeting of gene expression in a dose-dependent manner and decreased monocyte adhesion to HUVECs following tumor necrosis factor (TNF-α) treatment. The short hairpin RNA (shRNA) against AMPK α1 in HUVECs attenuated the adenine-induced inhibition of NF-κB activation in response to TNF-α, thereby suggesting that the anti-inflammatory role of adenine is mediated by AMPK. Following the knockdown of adenosyl phosphoribosyl transferase (APRT) in HUVECs, adenine supplementation failed to induce the phosphorylation of AMPK and ACC. Similarly, the expression of a shRNA against APRT nullified the anti-inflammatory effects of adenine in HUVECs. These results suggested that the role of adenine as an AMPK activator is related to catabolism by APRT, which increases the cellular AMP levels to activate AMPK.

Chinese Herbal Formulas Si-Wu-Tang and Er-Miao-San Synergistically Ameliorated Hyperuricemia and Renal Impairment in Rats Induced by Adenine and Potassium Oxonate.

BACKGROUND/AIMS: Hyperuricemia is an independent risk factor for chronic kidney disease and cardiovascular disease. Here, we examined the combined protective effects of Chinese herbal formula Si-Wu-Tang and Er-Miao-San on hyperuricemia and renal impairment in rats. METHODS: Rats were randomly divided into normal rats, hyperuricemic rats, and hyperuricemic rats orally administrated with benzbromarone (4.5 mg·kg⁻¹·d⁻¹), Si-Wu-Tang (3.78 g·kg⁻¹·d⁻¹) and Si-Wu-Tang plus Er-Miao-San (6.48 g·kg⁻¹·d⁻¹) for 4 weeks. Hyperuricemic rats were orally gavaged with adenine (0.1 g·kg⁻¹·d⁻¹) and potassium oxonate (1.5 g·kg⁻¹·d⁻¹) daily for 4 weeks. Serum uric acid, creatinine, total cholesterol (TCH), triglyceride and blood urea nitrogen (BUN) concentrations, as well as urinary uric acid and microalbuminuria were measured weekly. Serum xanthine oxidase (XOD) activity and renal histopathology were also evaluated. The renal expression of organic anion transporter 1 (OAT1) and organic anion transporter 3 (OAT3) was detected by western blot. RESULTS: Si-Wu-Tang plus Er-Miao-San lowered serum uric acid, creatinine, triglyceride and BUN levels to a greater degree than did Si-Wu-Tang alone. Si-Wu-Tang plus Er-Miao-San ameliorated microalbuminuria and renal histopathology, as well as decreased serum TCH concentration and XOD activity in hyperuricemic rats. Combination of Si-Wu-Tang and Er-Miao-San also led to a greater increase in OAT1 and OAT3 expression than did Siwutang alone. CONCLUSION: Si-Wu-Tang and Er-Miao-San synergistically ameliorated hyperuricemia and renal impairment in rats through upregulation of OAT1 and OAT3.

Lycium barbarum polysaccharide improves bipolar pulse current-induced microglia cell injury through modulating autophagy.

Blindness and visual impairments are heavy loads for modern society. Visual prosthesis is a promising therapy to treat these diseases. However, electric stimulation (ES)-induced damage of the optic nerve and adjacent cells are problems that must not be overlooked. In the current study, we aimed to investigate the effects of ES on cultured microglia cells and the potential protective mechanisms from a natural compound Lycium barbarum polysaccharide (LBP). Cellular injuries were induced by 9 mA bipolar pulse current in BV-2 cells for 15 min. Treatment with LBP alone or in association with either autophagic inhibitor 3-MA or autophagic agonist rapamycin was preadded for 2 h before the ES challenge. After that, morphological and molecular changes of the cells were measured at 2 h or 6 h postchallenges. We found that ES induced evident morphological and pathological changes of BV-2 cells, including oxidative stress, inflammation, and apoptosis. Pretreatment with LBP significantly attenuated these injuries with enhanced endogenous autophagy. When cellular autophagy was inhibited or enhanced by corresponding drug, the protective properties of LBP were partly inhibited or maintained, respectively. In addition, we demonstrated that ERK and p38 MAPK exerted diversified roles in the protection of LBP against ES-induced cellular damages. In conclusion, LBP improves bipolar pulse current-induced microglia cell injury through modulating autophagy and MAPK pathway.