A mechanosensory defect in a C. elegans amyloid-beta glutamatergic neuron model is reversed following exposure to Salvia species extracts.

Previous research has described promising neuroprotective and/or antioxidant properties for extracts derived from a few Salvia (sage) species. Here, six new Salvia species were isolated during flowering times from plants native to Turkey. Extracts were prepared and then examined for their potential to rescue both anterior and posterior mechanosensory behavioral defects in a transgenic C. elegans Alzheimer's disease model that expresses human amyloid-beta (Aß) peptide (1-42) exclusively in the glutamatergic neurons. Extracts from all six Salvia species rescued anterior touch response defects while only three rescued posterior touch response defects, compared to the Aß controls.

Cellular Membrane Affinity Chromatography Columns to Identify Specialized Plant Metabolites Interacting with Immobilized Tropomyosin Kinase Receptor B.

Chemicals synthesized by plants, fungi, bacteria, and marine invertebrates have been a rich source of new drug hits and leads. Medicines such as statins, penicillin, paclitaxel, rapamycin, or artemisinin, commonly used in medical practice, have been first identified and isolated from natural products. However, the identification and isolation of biologically active specialized metabolites from natural sources is a challenging and time-consuming process. Traditionally, individual metabolites are isolated and purified from complex mixtures, following the extraction of biomass. Subsequently, the isolated molecules are tested in functional assays to verify their biological activity. Here we present the use of cellular membrane affinity chromatography (CMAC) columns to identify biologically active compounds directly from complex mixtures. CMAC columns allow for the identification of compounds interacting with immobilized functional transmembrane proteins (TMPs) embedded in their native phospholipid bilayer environment. This is a targeted approach, which requires knowing the TMP whose activity one intends to modulate with the newly identified small molecule drug candidate. In this protocol, we present an approach to prepare CMAC columns with immobilized tropomyosin kinase receptor B (TrkB), which has emerged as a viable target for drug discovery for numerous nervous system disorders. In this article, we provide a detailed protocol to assemble the CMAC column with immobilized TrkB receptors using neuroblastoma cell lines overexpressing TrkB receptors. We further present the approach to investigate the functionality of the column and its use in the identification of specialized plant metabolites interacting with TrkB receptors.

Identification of TrkB Binders from Complex Matrices Using a Magnetic Drug Screening Nanoplatform.

Brain-derived neurotrophic factor (BDNF) and its receptor tyrosine receptor kinase B (TrkB) have been shown to play an important role in numerous neurological disorders, such as Alzheimer's disease. The identification of biologically active compounds interacting with TrkB serves as a drug discovery strategy to identify drug leads for neurological disorders. Here, we report effective immobilization of functional TrkB on magnetic iron oxide nanoclusters, where TrkB receptors behave as "smart baits" to bind compounds from mixtures and magnetic nanoclusters enable rapid isolation through magnetic separation. The presence of the immobilized TrkB was confirmed by specific antibody labeling. Subsequently, the activity of the TrkB on iron oxide nanoclusters was evaluated with ATP/ADP conversion experiments using a known TrkB agonist. The immobilized TrkB receptors can effectively identify binders from mixtures containing known binders, synthetic small molecule mixtures, and Gotu Kola (Centella asiatica) plant extracts. The identified compounds were analyzed by an ultrahigh-performance liquid chromatography system coupled with a quadrupole time-of-flight mass spectrometer. Importantly, some of the identified TrkB binders from Gotu Kola plant extracts matched with compounds previously linked to neuroprotective effects observed for a Gotu Kola extract approved for use in a clinical trial. Our studies suggest that the possible therapeutic effects of the Gotu Kola plant extract in dementia treatment, at least partially, might be associated with compounds interacting with TrkB. The unique feature of this approach is its ability to fast screen potential drug leads using less explored transmembrane targets. This platform works as a drug-screening funnel at early stages of the drug discovery pipeline. Therefore, our approach will not only greatly benefit drug discovery processes using transmembrane proteins as targets but also allow for evaluation and validation of cellular pathways targeted by drug leads.

Comparison of anticarcinogenic properties of Viburnum opulus and its active compound p-coumaric acid on human colorectal carcinoma.

Resistance to therapeutic agents and the highly toxic side effects of synthetic drugs has spurred new research in the treatment of colon cancer, which has high morbidity and mortality ratios. This study aims to clarify the molecular mechanisms of the anticarcinogenic properties of methanol extract of Viburnum opulus L. (EVO)and its main active compound, trans-p -coumaric acid ( p -CA), on human colon cancer cells (DLD-1, HT-29, SW-620, Caco-2) and healthy colon epithelial cells (CCD-18Co). The effects of EVO on controlled cell death (apoptosis) and the cell division cycle were determined by flow cytometry. Alteration in mRNA and protein expressions of switch genes in colorectal carcinoma (APC, MLH1, TP53, SMAD4, KRAS, and BRAF) were determined by qRT-PCR and Western blot, respectively. Our results show that EVO possesses a strong reducing capacity and free-radical scavenging activity. HPLC analyses prove that p -CAis the main compound of EVO. EVO and p -CA inhibit the proliferation of human colon cancer cells DLD-1 and HT-29 in a dose-dependent manner. EVO increases apoptosis of DLD-1 cells and halts the cell cycle in the G2 stage in HT-29 cells. mRNA and protein expressions of p53 and SMAD-4 are upregulated, while BRAFs are downregulated. The results were directly proportional to p -CA. EVO and p -CA up- and downregulate switch genes and protein expressions of DLD-1 cells, which alter the expression of 186 other genes. This is the first study of pharmacological exploration of V.opulus in human colon cancer. Its antiproliferative effects may be due to the presence of p -CA.