Kaempferol treatment ameliorates memory impairments in STZ­induced neurodegeneration by acting on reelin signaling.

Many treatment initiatives, like herbal products and their active ingredients, aim to alleviate neurodegeneration to increase cognitive functions. Kaempferol may be a candidate molecule for treating neurodegeneration because of its antioxidant effects. In the present study, we examined the molecular changes associated with kaempferol's memory­enhancing effects on streptozotocin (STZ)­induced neurodegeneration. After intracerebroventricular STZ injection in Long­Evans male rats, intraperitoneal kaempferol was administered for 12 days. The Morris water maze (MWM) was used to measure learning and memory performance in the rats, and proteins related to memory formation were investigated in the hippocampi with western blotting. Kaempferol improved learning performance and memory decline in STZ­treated rats. At the molecular level, STZ­induced neurodegeneration resulted in a decrease in the expression of GAD67, reelin, and phosphorylated­NMDAR. However, kaempferol treatment ameliorated these changes by enhancing their levels similar to the controls. While neither STZ injection nor kaempferol treatment produced any significant change in phosphorylated­CAMKII levels, they increased the expression of klotho and prealbumin. These results show that kaempferol has positive effects on memory loss, affecting synaptic plasticity by ameliorating both the levels and activity of memory­relevant molecules through reelin signaling. In summary, this study provides a guide to future studies by examining in detail the healing effect of kaempferol as a candidate molecule in the treatment of neurodegeneration, such as that observed in Alzheimer's disease.

Kaempferol attenuates diabetic nephropathy in streptozotocin-induced diabetic rats by a hypoglycaemic effect and concomitant activation of the Nrf-2/Ho-1/antioxidants axis.

This study examined the protective effect of Kaempferol against streptozotocin-induced diabetic nephropathy (DN) in rats and studies the underlying mechanisms. Rats were divided into 4 groups as control, control + Kaempferol, STZ, and STZ + Kaempferol. All treatments were conducted for 8 weeks daily after the induction of diabetes. Kaempferol prevented STZ-induced weight and food loss and attenuated renal damage and the alterations in all biochemical related parameters. Concomitantly, Kaempferol reduced renal levels of TNF-α and IL-6, cleaved caspase-3, p38, and Bax, suppressing JNK phosphorylation and NF-κB p65 transactivation, and upregulation of Bcl-2. In both control and STZ-diabetic rats, Kaempferol reduced fasting glucose levels, increased fasting insulin levels and HOMA-ß, reduced the levels of ROS and MDA, stimulated SOD and GSH levels, and increased the expression of Nrf2 and HO-1. In conclusion, Kaempferol prevents STZ-induced diabetic nephropathy, mainly, by antioxidant potential, mediated by the upregulation of the Nrf-2/HO-1 axis.

Network pharmacology strategy to investigate the pharmacological effects of Suanzaoren Decoction on insomnia

Abstract Suanzaoren Decoction (SZRD) is an ancient prescription used in the treatment of insomnia. This study aimed to investigate the components and targets of SZRD in treating insomnia. First, the compounds of five herbs in SZRD were collected from the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (TCMSP), and the putative targets for treating insomnia were obtained from DrugBank to construct the herb-compound-target- disease network. A protein-protein interaction (PPI) network was constructed in the STRING database, and then Gene Ontology functional enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were performed to predict the mechanism of action of intersection target. Finally, 30 mice were divided into five groups: control, model, and quercetin groups (100, 50, 25 mg/kg). The sleep latency and duration of pentobarbital-induced sleeping were measured. The production of interleukin-6 (IL-6) and γ-aminobutyric acid (γ-GABA) was detected by using an enzyme-linked immunosorbent assay kit (ELISA), and Gamma-aminobutyric acid type a receptor subunit alpha1 (GABRA1) was tested by Reverse Transcription-Polymerase Chain Reaction (RT-PCR). A total of 152 active ingredients, including 80 putative targets of SZRD, were obtained. The main active compounds included quercetin and kaempferol, and the key targets involved IL-6 and nitric oxide synthase 3 (NOS3). The results of pathway enrichment analysis indicated that the putative targets of SZRD mainly participated in Neuroactive ligand-receptor interaction. The experiment of P-chlorophenylalanine (PCPA)-induced insomnia model showed that quercetin obviously shortened the sleep latency and prolonged the sleep duration of the insomnia model. The production of IL-6, γ-GABA, and GABRA1 mRNA was significantly increased in mice treated with quercetin. This study predicted the active ingredients and potential targets of SZRD on insomnia on the basis of a systematic network pharmacology approach and illustrated that SZRD might exert hypnotic effects via regulating IL-6, γ-GABA, and GABRA1

Astragalin induced selective kidney cancer cell death and these effects are mediated via mitochondrial mediated cell apoptosis, cell cycle arrest, and modulation of key tumor-suppressive miRNAs.

PURPOSE: Kidney cancer is responsible for a significant number of deaths worldwide. This cancer is often diagnosed at advanced stages and there are frequent relapses following chemotherapy. Target therapies are used now for kidney cancer, while the use of chemotherapy declines. The currently used chemotherapeutic drugs have a number of adverse effects. Herein, we examined the anticancer effects of Astragalin against a panel of kidney cancer cells. METHODS: CellTiter-Glo Luminescent Cell Viability Assay Kit was used to examine the anti-proliferative effects of Astragalin. Acridine orange (AO)/ethidium bromide (EB), DAPI and annexin V/promidium iodide (PI) staining assays were used to examine the apoptotic cell death. Cell cycle analysis was performed by flow cytometry. The mRNA expression was checked by qRT-PCR and protein expression was examined by western blotting. RESULTS: Astragalin inhibited the growth of the all kidney cancer cell lines with IC50 ranging between 20 to 50 µM. Of note, Astragalin had low cytotoxic effects on the normal kidney cells with an IC50 of 110 µM. The experiments have shown that Astragalin exerts antiproliferative effects on the A498 kidney cancer cells by apoptotic cell death. This effect was concomitant with upregulation of apoptotic proteins such as caspase 3 and 9 and Bax. Astragalin also induced arrest of the A498 cells at the G2/M checkpoint of the cell cycle. Also, Astragalin could upregulate the expression of tumor-suppressive microRNAs. CONCLUSIONS: These results suggest that Astragalin exerts potent anticancer effects on kidney cancer cells and could pave the way in the management of kidney cancer provided clinical studies are carried out.

Antidiabetic potential of polyherbal formulation DB14201: Preclinical development, safety and efficacy studies.

ETHNOPHARMACOLOGICAL RELEVANCE: The poly-herbal formulation DB14201 is a new combination of ayurvedic ingredients for treatment of diabetes. The aim of present study was to investigate safety and in vivo efficacy of DB14201 extract. Further this work was aimed to develop, characterize and standardize DB14201 extract and develop it as a botanical drug. MATERIALS AND METHODS: The polyherbal extract was standardized using four chemical markers. The LC-MS/MS method was developed for identification and quantification of mangiferin, berberine, kaempferol and curcumin. The extract was standardized for heavy metal content, aflotoxins, and microbial tests. The mechanism of action of DB14201 extract was explored through glucose uptake by adipocytes, TNF-α production and free fatty acid release, in vitro, was studied using murine adipocytes (3T3-L1). The effect of extract on insulin release was evaluated using murine pancreatic beta cell (ß TC-6). The safety and in vivo efficacy of extract was studied using suitable animal model. Hematology and blood biochemistry parameters were also assessed. RESULTS: In vitro studies of DB14201 in murine adipocytes and murine pancreatic beta cells demonstrated the plausible mechanism of action of DB14201 could be through increase in glucose uptake and by stimulation of insulin release by RIN-5f cells. The microbial load, heavy metals were found to be within the AYUSH permissible limits and aflotoxins were absent. Preclinical efficacy studies in animal models proved the anti-diabetic potential of the extract. The preclinical acute dose toxicity study and 90-days repeated dose toxicity study of DB14201 extract in wistar rats by oral route indicated that the extract is safe up to 1000mg/kg dose. Hematology and blood biochemistry parameters were within the normal range. CONCLUSIONS: The data presented herein demonstrated anti-diabetic potential of developed DB14201 extract and this study will serve as the benchmark for the further research on this polyherbal formulation.

Anti-Japanese-encephalitis-viral effects of kaempferol and daidzin and their RNA-binding characteristics.

BACKGROUND: New therapeutic tools and molecular targets are needed for treatment of Japanese encephalitis virus (JEV) infections. JEV requires an α-1 translational frameshift to synthesize the NS1' protein required for viral neuroinvasiveness. Several flavonoids have been shown to possess antiviral activity in vitro against a wide spectrum of viruses. To date, the antiviral activities of flavonol kaempferol (Kae) and isoflavonoid daidzin (Dai) against JEV have not been described. METHODOLOGY/PRINCIPAL FINDINGS: The 50% cytotoxic concentration (CC(50)) and 50% effective concentration (EC(50)) against JEV were investigated in BHK21 cells by MTS reduction. Activity against viral genomic RNA and proteins was measured by real-time RT-PCR and western blotting. The frameshift site RNA-binding characterization was also determined by electrospray ionization mass spectrometry, isothermal titration calorimetry and autodocking analysis. EC(50) values of Kae and Dai were 12.6 and 25.9 µM against JEV in cells pretreated before infection, whereas in cells infected before treatment, EC(50) was 21.5 and 40.4 µM, respectively. Kae exhibited more potent activity against JEV and RNA binding in cells following internalization through direct inhibition of viral replication and protein expression, indicating that its antiviral activity was principally due to direct virucidal effects. The JEV frameshift site RNA (fsRNA) was selected as a target for assaying Kae and Dai. ITC of fsRNA revealed an apparent K(b) value for Kae that was nine fold stronger than that for Dai. This binding was confirmed and localized to the RNA using ESI-MS and autodock analysis. Kae could form non-covalent complexes with fsRNA more easily than Dai could. CONCLUSIONS/SIGNIFICANCE: Kae demonstrates more potent antiviral activity against JEV than does Dai. The mode of action of Kae as an anti-JEV agent seems to be related to its ability to inactivate virus by binding with JEV fsRNA.

Effects in rats of maternal exposure to raspberry leaf and its constituents on the activity of cytochrome p450 enzymes in the offspring.

The goal of our study was to determine whether maternal exposure to red raspberry leaf (RRL) and its constituents can permanently alter biotransformation of fluorogenic substrates by cytochrome P450 (CYP) in the livers of male and female offspring. Nulliparous female rats received vehicle, raspberry leaf, kaempferol, quercetin, or ellagic acid orally once breeding had been confirmed until parturition. Hepatic microsomes were prepared from animals at birth (postnatal day 1 [PND1]), weaning (PND21), PND65, and PND120 to determine the biotransformation of 8 fluorogenic substrates. The pattern of biotransformation of all but 2 of the substrates was gender specific. Maternal consumption of RRL increased biotransformation of 3 substrates by female offspring at PND120 resulting in a more masculine profile. Kaempferol and quercetin had a similar effect to RRL. These results suggest that maternal consumption of either RRL or some of its constituents leads to long-term alterations of CYP activity in female offspring.

Dietary flavonoids induce MLL translocations in primary human CD34+ cells.

Genetic abnormalities leading to infant leukemias already occur during fetal development and often involve rearrangements of the mixed-lineage leukemia (MLL) gene. These rearrangements resemble the aberrations observed in therapy-related leukemias following treatment with topoisomerase II (topoII)-inhibiting agents such as etoposide. Since flavonoids are potent topoII inhibitors, we examined the role of three widely consumed dietary flavonoids (quercetin, genistein and kaempferol) on the development of MLL rearrangements in primary human CD34(+) cells. Using the neutral Comet assay, we demonstrated a dose-dependent double-strand break (DSB) formation after exposure to flavonoids. An incorrect repair of these DSBs resulted in chromosomal translocations that co-localized with those identified in infant leukemias. Most of these translocations were formed by microhomology-mediated end joining. Moreover, in all but one translocation, SINE/Alu or LINE/L1 repetitive elements were present in at least one side of the breakpoint junction. Beside MLL translocations, fluorescence in situ hybridization analysis demonstrated monosomy or trisomy of MLL in 8-10% of the quercetin-exposed CD34(+) cells. Our study demonstrates that biologically relevant concentrations of flavonoids can induce MLL abnormalities in primary hematopoietic progenitor cells. This is particularly alarming knowing that the differences in metabolism and excretion rate between mother and fetus can lead to a higher flavonoid concentration on the fetal side. Therefore, it is important to raise public awareness and set guidelines for marketing flavonoid supplements to reduce the risk of infant leukemias.