A simplified workflow with end-point validation of real-time electrical cell-substrate impedance sensing of retinoic acid stimulated neurogenesis in human SH-SY5Y cells in vitro.

OBJECTIVE: Retinoic acid (RA) is known to transition proliferating SH-SY5Y neuroblastoma cells towards functional neurons. However, the activity of RA is restricted due to its photolability where any findings from prolonged time course observations using microscopy may alter outcomes. The aim of the study was to establish a real-time, long-term (9-day) protocol for the screening of differentiation events using Electrical cell-substrate impedance sensing (ECIS). RESULTS AND DISCUSSION: A differentiation baseline for SH-SY5Y cells was established. Cells were seeded and exposed to repeated spikes of RA using the xCELLigence real-time cell analyser single plate (RTCA-SP) for real-time monitoring and identification of differentiation activity over a 9 day period in order to be more representative of differentiation over a prolonged timeline. Specific features associated with differentiation (growth inhibition, neurite outgrowths) were confirmed by end-point analysis. RA-induced growth inhibition and assumed phenotypic changes (i.e. neurite outgrowth) were identified by the xCELLigence analysis and further confirmed by end-point metabolic and phenotypic assays. Change in cellular morphology and neurite outgrowth length was identified by end-point fluorescence detection followed by computational analysis. Based on this it was possible to identify SH-SY5Y phenotypic differentiation with distinct phases observed over 9 days using Electric cell-substrate impedance sensing (ECIS) cell index traces providing a path to application in larger scale neurotrophic factor screening using this scalable technology.

Cytostatic and cytotoxic effects of a hot water and methanol extract of Acokanthera oppositifolia in HepG2 hepatocarcinoma cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Herb-induced liver injury is poorly described for African herbal remedies, such as Acokanthera oppositifolia. Although a commonly used treatment for pain, snake bites and anthrax, it is also a well-known arrow poison, thus toxicity is to be expected. AIM OF THE STUDY: The cytotoxicity and preliminary mechanisms of toxicity in HepG2 hepatocarcinoma cells were assessed. MATERIALS AND METHODS: The effect of hot water and methanol extracts were on cell density, oxidative status, mitochondrial membrane potential, fatty acids, caspase-3/7 activity, adenosine triphosphate levels, cell cycling and viability was assessed. Phytochemicals were tentatively identified using ultra-performance liquid chromatography. RESULTS: The hot water extract displayed an IC50 of 24.26 µg/mL, and reduced proliferation (S- and G2/M-phase arrest) and viability (by 30.71%) as early as 24 h after incubation. The methanol extract had a comparable IC50 of 26.16 µg/mL, and arrested cells in the G2/M-phase (by 18.87%) and induced necrosis (by 13.21%). The hot water and methanol extracts depolarised the mitochondrial membrane (up to 0.84- and 0.74-fold), though did not generate reactive oxygen species. The hot water and methanol extracts decreased glutathione (0.42- and 0.62-fold) and adenosine triphosphate (0.08- and 0.26-fold) levels, while fatty acids (2.00- and 4.61-fold) and caspase-3/7 activity (1.98- and 5.82-fold) were increased. CONCLUSION: Extracts were both cytostatic and cytotoxic in HepG2 cells. Mitochondrial toxicity was evident and contributed to reducing adenosine triphosphate production and fatty acid accumulation. Altered redox status perturbed proliferation and promoted necrosis. Extracts of A. oppositifolia may thus promote necrotic cell death, which poses a risk for inflammatory hepatotoxicity with associated steatosis.

Comparative Evaluation of the Cytotoxicity of Doxorubicin in BT-20 Triple-Negative Breast Carcinoma Monolayer and Spheroid Cultures.

Three-dimensional cell culture models are increasingly adopted as preferred pre-clinical drug testing platforms, as they circumvent limitations associated with traditional monolayer cell cultures. However, many of these models are not fully characterized. This study aimed to characterize a BT-20 triple-negative breast carcinoma spheroid model and assess its susceptibility to doxorubicin in comparison to a monolayer model. Spheroids were developed using the liquid overlay method. Phenotypic attributes were analyzed by characterizing changes in size, gross morphology, protein content, metabolic activity, hypoxic status, and cell-cell junctions. The cytotoxic range of doxorubicin in monolayers was determined using the sulforhodamine B assay, and the comparative effect of toxic and sub-toxic concentrations was assessed in both spheroids and monolayers. Similar to the in vivo microenvironment, spheroids had a heterogeneous spatial cytoarchitecture, inherent hypoxia and strong adherens junctions. Doxorubicin induced dose-dependent cytotoxicity in monolayers (IC25: 130 nM, IC50: 320 nM and IC75: 1580 nM); however, these concentrations did not alter the spheroid size or acid phosphatase activity. Only concentrations ≥6 µM had any effect on spheroid integrity. In comparison to monolayers, the BT-20 spheroid model has decreased sensitivity to doxorubicin and could serve as a better model for susceptibility testing in triple-negative breast cancer.

Utility of Therapeutic Drug Monitoring in Identifying Clinically Significant Interactions Between St. John's Wort and Prescription Drugs.

BACKGROUND: The general population widely uses herbal medicines, as they are regarded as effective and safe. St. John's wort, which is an effective herbal antidepressant, exhibits both pharmacokinetic and pharmacodynamic interactions with several drugs. The aim of this review was to highlight the clinically significant interactions of St. John's wort with drugs that require to be monitored to assess their therapeutic effect. METHODS: Published literature was searched using electronic databases, such as MEDLINE, PubMed, and Elsevier ScienceDirect using terms such as "herbal medicine," "herbal toxicity," "legislation herbal medicine," "drug-herb interactions," "St. John's wort," and "St. John's wort-drug interactions." Searches were limited to the English language, and there was no restriction on the date of publication. RESULTS: St. John's wort exhibits a number of pharmacokinetic and pharmacodynamic interactions with drugs. The most dangerous interactions occurred when used concurrently with the immunosuppressants, cyclosporine, and tacrolimus (treatment failure or organ rejection) or warfarin (treatment failure resulting in thromboembolic events) or antiretroviral agents (treatment failure and the emergence of new viral variants that are resistant to conventional drugs). CONCLUSIONS: Patients should consult their health care providers before consuming herbal supplements, especially St. John's wort, to avoid potentially dangerous drug-herb interactions.

The VEGF/VEGFR Axis Revisited: Implications for Cancer Therapy.

The vascular endothelial growth factor (VEGF)/vascular endothelial growth factor receptor (VEGFR) axis is indispensable in the process of angiogenesis and has been implicated as a key driver of tumor vascularization. Consequently, several strategies that target VEGF and its cognate receptors, VEGFR-1 and VEGFR-2, have been designed to treat cancer. While therapies targeting full-length VEGF have resulted in an improvement in both overall survival and progression-free survival in various cancers, these benefits have been modest. In addition, the inhibition of VEGFRs is associated with undesirable off-target effects. Moreover, VEGF splice variants that modulate sprouting and non-sprouting angiogenesis have been identified in recent years. Cues within the tumor microenvironment determine the expression patterns of these variants. Noteworthy is that the mechanisms of action of these variants challenge the established norm of VEGF signaling. Furthermore, the aberrant expression of some of these variants has been observed in several cancers. Herein, developments in the understanding of the VEGF/VEGFR axis and the splice products of these molecules, as well as the environmental cues that regulate these variants are reviewed. Furthermore, strategies that incorporate the targeting of VEGF variants to enhance the effectiveness of antiangiogenic therapies in the clinical setting are discussed.

Mechanisms facilitating the uptake of carboxyl-polythene glycol-functionalized gold nanoparticles into multicellular spheroids.

OBJECTIVES: Nanomedicines represent theragnostic alternatives to traditional candidate drugs, with increased targeting and delivery potential due to their size and functional tailorability. Biological activity typically relies on nanomaterials permeating into the intracellular environment, necessitating characterization of uptake and intracellular trafficking pathways. Spheroids' three-dimensional architecture and heterogenous cellular distribution offer an in-vivo-representative platform to assess the biological activity of nanoparticles (NPs). This study aimed to develop an A549 alveolar carcinoma spheroid model as a NP uptake assessment platform for carboxyl-polythene glycol-functionalized gold NPs affording further biological characterization opportunities in nanomedicine. METHODS: A549 spheroids were generated via the liquid overlay method, and their morphology and viability were assessed for 21 days. Cytotoxicity was assessed via lactate dehydrogenase release. NP uptake was elucidated using uptake pathway inhibition, combined with CytoViva hyperspectral imaging of sectioned spheroids to count internalized NPs. KEY FINDINGS: Cytotoxicity was absent for all exposure groups. Clathrin-mediated endocytosis was the primary endocytic mechanism (33.5-54.8% of uptake), which may precede lysosomal degradation. Lysosomal membrane permeabilization appears to be a potential downstream application. Low penetration into spheroids (4.5 µm) suggests the failure of NPs to traverse cellular layers in the spheroid. CONCLUSIONS: Although poor uptake was observed, a multicellular spheroid model of A549 alveolar carcinoma cells was established, allowing for similar future uptake assessment of various NPs.

Hybrid Compounds Containing Carvacrol Scaffold: In Vitro Antibacterial and Cytotoxicity Evaluation.

BACKGROUND: The design of hybrid compounds is a distinct approach for developing potent bioactive agents. Carvacrol, an essential oil, exhibits antimicrobial, antifungal, antioxidant, and anticancer activity, making it a good precursor for the development of compounds with potent biological activities. Some patents have reported carvacrol derivatives with promising biological activities. OBJECTIVE: This study aimed to prepare hybrid compounds containing a carvacrol scaffold with significant antibacterial and anticancer activity. METHODS: Esterification reactions between carvacrol and known pharmacophores were performed at room temperature and characterized using 1H-NMR, 13CNMR, and UHPLC-HRMS. In vitro antibacterial study was determined using the microdilution assay and cytotoxicity evaluation using sulforhodamine B staining assay. RESULTS: The FTIR spectra of the carvacrol hybrids revealed prominent bands in the range of 1612-1764 cm-1 and 1014-1280 cm-1 due to (C=O) and (C-O) stretching vibrations, respectively. The structures of the carvacrol hybrids were confirmed by 1H-NMR, 13C-NMR, and UHPLC-HRMS analysis, and compound 5 exhibited superior activity when compared to the hybrid compounds against the strains of bacteria used in the study. The in vitro cytotoxicity evaluation showed that compound 3 induced cytotoxicity in all the cancer cell lines; MDA (16.57 ± 1.14 µM), MCF-7 (0.47 ± 1.14 µM), and DU145 (16.25 ± 1.08 µM), as well as the normal breast cells, MCF-12A (0.75± 1.30 µM). Compound 7 did not induce cytotoxicity in the cell lines tested (IC50 > 200 µM). CONCLUSION: The modification of carvacrol through hybridization is a promising approach to develop compounds with significant antibacterial and anticancer activity.

A Brief Updated Review of Advances to Enhance Resveratrol's Bioavailability.

Resveratrol (RES) has a low bioavailability. This limitation was addressed in an earlier review and several recommendations were offered. A literature search was conducted in order to determine the extent of the research that was conducted in line with these recommendations, along with new developments in this field. Most of the identified studies were pre-clinical and confirmed the heightened activity of RES analogues compared to their parent compound. Although this has provided additional scientific kudos for these compounds and has strengthened their potential to be developed into phytopharmaceutical products, clinical trials designed to confirm this increased activity remain lacking and are warranted.

Synthesis, Antibacterial, and Cytotoxicity Evaluation of Oleanolic Acid-4-aminoquinoline Based Hybrid Compounds.

AIM: The study aims to prepare a class of oleanolic-based compounds. BACKGROUND: Conventional drugs used to treat infectious diseases suffer from limitations such as drug toxicity and drug resistance. The resistance of microbes to antimicrobial agents is a significant challenge in treating microbial infections. Combining two or more drugs with different modes of action to treat microbial infections results in a delay in developing drug resistance by the microbes. However, it is challenging to select the appropriate drugs for combination therapy due to the differences in stability and pharmacokinetic profile of the drugs. Therefore, developing hybrid compounds using the existing drugs is a promising approach to design effective antimicrobial agents. OBJECTIVES: To prepare oleanolic-based hybrid compounds followed by characterization, in vitro antibacterial and cytotoxicity evaluation. METHODS: Oleanolic acid-4-aminoquinoline-based hybrid compounds were synthesized via esterification and amidation. The compounds were characterized using FTIR, NMR, and UHPLC-HRMS. Oleanolic acid (OA) was isolated from the flower buds of Syzygium aromaticum (L.) Merr. & L.M.Perry, a species from Kingdom Plantae, order Mytales in the Myrtaceae family. Antibacterial activity was determined against selected strains of bacteria using the microdilution assay and cytotoxicity activity was assessed using the sulforhodamine B assay against selected cancer cell lines. RESULTS: The synthesized hybrid compounds exhibited antibacterial activity against the Gram-positive bacteria Enterococcus faecalis (ATCC13047), Bacillus subtilis (ATCC19659), Staphylococcus aureus as well as Gram-negative bacteria, Klebsiella oxytoca (ATCC8724), Escherischia coli (ATCC25922), and Proteus vulgaris (ATCC6380) with minimum inhibitory concentrations of 1.25 mg/mL compared to oleanolic acid (2.5 mg/mL). Compounds 13 and 14 displayed cytotoxicity in vitro against the cancer cell lines (MCF-7 and DU 145) compared to the oleanolic acid (IC50 ˃ 200 µM). CONCLUSION: Modification of C28 of OA enhanced its biological activity.

Novel N-benzylpiperidine carboxamide derivatives as potential cholinesterase inhibitors for the treatment of Alzheimer's disease.

A series of fifteen acetylcholinesterase inhibitors were designed and synthesised based upon the previously identified lead compound 5,6-dimethoxy-1-oxo-2,3-dihydro-1H-inden-2-yl 1-benzylpiperidine-4-carboxylate (5) which showed good inhibitory activity (IC50 0.03 ±â€¯0.07 µM) against acetylcholinesterase. A series of compounds were prepared wherein the ester linker in the original lead compound was exchanged for a more metabolically stable amide linker and the indanone moiety was exchanged for a range of aryl and aromatic heterocycles. The two most active analogues 1-benzyl-N-(5,6-dimethoxy-8H-indeno[1,2-d]thiazol-2-yl)piperidine-4-carboxamide (28) and 1-benzyl-N-(1-methyl-3-oxo-2-phenyl-2,3-dihydro-1H-pyrazol-4-yl) piperidine-4-carboxamide (20) afforded in vitro IC50 values of 0.41 ±â€¯1.25 and 5.94 ±â€¯1.08 µM, respectively. In silico screening predicts that 20 will be a blood brain-barrier permeant, and molecular dynamic simulations are indicative of a close correlation between the binding of 20 and the Food and Drug Administration-approved cholinesterase inhibitor donepezil (1).

Synthesis and biological evaluation of 2-chloro-3-[(thiazol-2-yl)amino]-1,4-naphthoquinones.

A series of novel, substituted 2-chloro-3-[(thiazol-2-yl)amino]-1,4-naphthoquinones have been prepared and shown to exhibit promising concentration-dependent activity against human SH-SY5Y cells, Plasmodium falciparum, Mycobacterium tuberculosis and P. aeruginosa. Substituent effects on observed bioactivity have been explored; the para-fluorophenyl derivative 3d exhibited activity across the range of the bioassays employed, indicating the potential of the 2-chloro-3-[(4-arylthiazol-2-yl)amino]-1,4-naphthoquinone scaffold in the development of novel, broad spectrum therapeutics.

Cytotoxic activity of pentachlorophenol and its active metabolites in SH-SY5Y neuroblastoma cells.

As knowledge regarding mechanisms of pentachlorophenol (PCP) toxicity in neuronal cell lines is limited, the aim of the study was to evaluate the effects of PCP and its active metabolites, tetrachloro-1,4-benzoquinone (TCBQ) and tetrachlorohydroquinone (TCHQ) in human neuroblastoma SH-SY5Y cells. All compounds induced cytotoxic effects in time- and dose-dependent manners, and resulted in differential modes of cell death. Reduced mitochondrial membrane potential (Δá´ªM) and oxidative damage lead to apoptosis and necrosis following TCBQ and PCP exposure, respectively. Time-dependent investigations revealed transient Δá´ªM recovery in TCHQ exposed cells, and redox stress. Sufficient Δá´ªM recovery allowed apoptosis completion in TCHQ exposed cells, whereas overwhelming metabolic and oxidative stress saw a conversion from apoptotic to necrotic-like cell death. The onset of mitochondrial dysfunction preceded that of redox damage for all compounds, indicating that oxidative damage is secondary to Δá´ªM insult. Cytotoxic events were further linked to cell cycle. S phase and G2/M blocks were observed after 12 h exposure to TCBQ and TCHQ, respectively, while a G1 block occurred after 24 h exposure to PCP. This study provides new insight regarding time-dependant toxic effects of PCP and its metabolites in human neuronal cells.

Beneficial effects of folic acid on the kidneys and testes of adult albino rats after exposure to methomyl.

The aim of this study was to evaluate the protective effect of folate against methomyl-induced toxicity on the kidneys and testes of male rats. Adult male albino rats were divided into four groups; Group I served as the control (vehicle), Group II received folic acid (1.1 mg per kg b.wt.), Group III methomyl (1 mg per kg b.wt.) and Group IV folic acid and methomyl. Treatments were administered via oral gavage on a daily basis for 14 weeks. Thereafter blood samples were collected and serum creatinine, testosterone and total antioxidant capacity (TAC) were determined. Animals were sacrificed and semen analysis was conducted. The kidneys and testes were excised and malondialdehyde (MDA) levels were determined. Histopathological and immunohistochemical analyses for caspase-3 were also undertaken. Methomyl treatment resulted in a significant (p < 0.001) elevation of creatinine and MDA levels and significant (p < 0.001) reduction in testosterone and TAC levels. Furthermore, methomyl caused a significant (p < 0.001) reduction in sperm quality. Histopathological examination indicated testicular and renal damage with strong immunoreactivity for caspase-3. Functional and tissue damage was prevented in rats treated with a combination of methomyl and folic acid. This is ascribed to the ability of folate to directly scavenge reactive oxygen species and indirectly enhance cellular redox homeostasis. This study identified that folic acid supplementation may have a beneficial effect in preventing or reducing the deleterious effects of methomyl exposure on kidney as well as testis structure and function. Future studies should focus on the fertility outcome/pregnancy index in rats.

Priority areas for cannabis and cannabinoid product research in South Africa.

No abstract available.

Solamargine, a bioactive steroidal alkaloid isolated from Solanum aculeastrum induces non-selective cytotoxicity and P-glycoprotein inhibition.

BACKGROUND: Solanum aculeastrum fruits are used by some cancer sufferers as a form of alternative treatment. Scientific literature is scarce concerning its anticancer activity, and thus the aim of the study was to assess the in vitro anticancer and P-glycoprotein inhibitory potential of extracts of S. aculeastrum fruits. Furthermore, assessment of the combinational effect with doxorubicin was also done. METHODS: The crude extract was prepared by ultrasonic maceration. Liquid-liquid extraction yielded one aqueous and two organic fractions. Bioactive constituents were isolated from the aqueous fraction by means of column chromatography, solid phase extraction and preparative thin-layer chromatography. Confirmation of bioactive constituent identity was done by nuclear magnetic resonance and ultra-performance liquid chromatography mass spectrometry. The crude extract and fractions were assessed for cytotoxicity and P-glycoprotein inhibition in both cancerous and non-cancerous cell lines using the sulforhodamine B and rhodamine-123 assays, respectively. RESULTS: Both the crude extract and aqueous fraction was cytotoxic to all cell lines, with the SH-SY5Y neuroblastoma cell line being most susceptible to exposure (IC50 = 10.72 µg/mL [crude], 17.21 µg/mL [aqueous]). Dose-dependent P-glycoprotein inhibition was observed for the crude extract (5.9 to 18.9-fold at 100 µg/mL) and aqueous fraction (2.9 to 21.2 at 100 µg/mL). The steroidal alkaloids solamargine and solanine were identified. While solanine was not bioactive, solamargine displayed an IC50 of 15.62 µg/mL, and 9.1-fold P-glycoprotein inhibition at 100 µg/mL against the SH-SY5Y cell line. Additive effects were noted for combinations of doxorubicin against the SH-SY5Y cell line. CONCLUSIONS: The crude extract and aqueous fraction displayed potent non-selective cytotoxicity and noteworthy P-glycoprotein inhibition. These effects were attributed to solamargine. P-glycoprotein inhibitory activity was only present at concentrations higher than those inducing cytotoxicity, and thus does not appear to be the likely mechanism for the enhancement of doxorubicin's cytotoxicity. Preliminary results suggest that non-selective cytotoxicity may hinder drug development, however, further assessment of the mode of cell death is necessary to determine the route forward.

African Herbal Remedies with Antioxidant Activity: A Potential Resource Base for Wound Treatment.

The use of traditional herbal remedies as alternative medicine plays an important role in Africa since it forms part of primary health care for treatment of various medical conditions, including wounds. Although physiological levels of free radicals are essential to the healing process, they are known to partly contribute to wound chronicity when in excess. Consequently, antioxidant therapy has been shown to facilitate healing of such wounds. Also, a growing body of evidence suggests that, at least, part of the therapeutic value of herbals may be explained by their antioxidant activity. This paper reviews African herbal remedies with antioxidant activity with the aim of indicating potential resources for wound treatment. Firstly, herbals with identified antioxidant compounds and, secondly, herbals with proven antioxidant activity, but where the compound(s) responsible for the activity has not yet been identified, are listed. In the latter case it has been attempted to ascribe the activity to a compound known to be present in the plant family and/or species, where related activity has previously been documented for another genus of the species. Also, the tests employed to assess antioxidant activity and the potential caveats thereof during assessment are briefly commented on.