Molecular interaction of quercetin and its derivatives against nucleolin in breast cancer: in-silico and in-vitro study.

Nucleolin, a multifaceted RNA binding domain protein is overexpressed in various cancers leading to dysfunction of several cellular signaling pathways. Quercetin, a distinctive bioactive molecule, along with its derivatives have shown exclusive physio-chemical properties which makes them appealing choices for drug development, yet their role in targeted cancer therapy is limited. Here, the RBD domain structure of Nucleolin was modeled and stabilized by MD simulations for a period of 1000 ns. Molecular docking was performed to determine the binding capability of ligands with the target. To determine the stability of the ligand inside the binding pocket of the protein, MD simulation was performed for a period of 250 ns each for Quercetin-4'-o'-Glucoside, Quercetin_9 and Quercetin complexes. Further, in-vitro studies including cytotoxicity and RT-PCR assays were performed to validate quercetin against Nucleolin. Molecular docking and MD Simulation studies suggested a potential mechanism of interaction of Quercetin-4'-o'-Glucoside, Querectin_9 and Quercetin with Nucleolin with the binding free energy of -63.653, -58.86 and -46.9 kcal/mol, respectively. Moreover, Lys 348 and Glu379 were identified as important amino acids in ligand interaction located at the RRM2 motif of Nucleolin. In-vitro studies showed significant downregulation in Nucleolin expression by 15.18 and 2.51-fold at 48h and 72h respectively in MCF-7 cells with Quercetin (IC50 = 160 µM). Our findings suggested the potential role of specific RRM motifs in interaction with natural compounds targeting Nucleolin. This could be an effective strategy in the identification of potential molecules in targeting Nucleolin which can be further explored for developing targeted therapies for breast cancer.Communicated by Ramaswamy H. Sarma.

Trolox aids coenzyme Q10 in neuroprotection against NMDA induced damage via upregulation of VEGF in rat model of glutamate excitotoxicity.

Glutamate induced damage to retinal ganglion cells (RGCs) requires tight physiological regulation of the N-methyl-D-aspartate (NMDA) receptors. Previously, studies have demonstrated the neuroprotective abilities of antioxidants like coenzyme Q10 (CoQ10) and vitamin E analogs like α-tocopherol against neuropathies resulting from NMDA insult, but have failed to shed light on the effect of CoQ10 and trolox, a hydrophilic analog of vitamin E, on glaucomatous neurodegeneration. In the current study, we wanted to investigate whether the combined effect of trolox with CoQ10 could alleviate NMDA-induced death of retinal cells while also trying to elucidate the underlying mechanism in relation to the yet unexplained role of vascular endothelial growth factor (VEGF) in NMDA-mediated excitotoxicity. After successful NMDA-induced degeneration, we followed it up with the treatment of combination of Trolox and CoQ10. The structural damage by NMDA was repaired significantly and retina retained structural integrity comparable to levels of control in the treatment group of Trolox and CoQ10. Detection of ROS generation after NMDA insult showed that together, Trolox and CoQ10 could significantly bring down the high levels of free radicals while also rescuing mitochondrial membrane potential (MMP). A significant increase in NMDA receptor Grin2A by CoQ10 alone as well as by CoQ10 and trolox was accompanied by a lowered Grin2B receptor expression, suggesting neuroprotective action of Trolox and CoQ10. Subsequently, lowered VEGFR1 and VEGFR2 receptor expression by NMDA treatment also recovered when subjected to combined treatment of Trolox and CoQ10. Western blot analyses also indicated the same whereby Trolox and CoQ10 could increase the diminished levels of phosphorylated VEGFR2. Immunofluorescence studies also indicated a positive correlation between recovered VEGFR2 and NMDAR2A levels and diminished levels of NMDAR2D, confirming the results obtained by RT-PCR analysis. This is the first report in our knowledge that demonstrates the efficacy of trolox in combination with CoQ10 highlighting the importance of maintaining VEGF levels that are lowered in ocular diseases due to NMDA-related toxicities.

In-vitro antiproliferative efficacy of Abrus precatorius seed extracts on cervical carcinoma.

Abrus precatorius is a tropical medicinal plant with multiple medicinal benefits whose seeds have not yet been studied against cervical cancer. Herein, we have assessed the antioxidant and antiproliferative properties of seed extracts (ethyl acetate and 70% ethanol) prepared from Soxhlet and Maceration extraction methods against Hep2C and HeLa Cells. We observed that the APE (Sox) extract had a significantly higher total flavonoid content, APA (Mac) extract had a high total phenolic content, and APA (Sox) extract had a high total tannin content. Further, HPLC analysis of extracts revealed the presence of tannic acid and rutin. Moreover, APA (Sox) exhibited the highest free radical scavenging activity. APE (Mac) had the best antiproliferative activity against Hep2C cells, while APA (Sox) had the best antiproliferative activity against HeLa cells. In Hep2C cells, APE (Mac) extract revealed the highest SOD, catalase activity, GSH content, and the lowest MDA content, whereas APA (Mac) extract demonstrated the highest GST activity. In HeLa cells, APA (Sox) extract showed the highest SOD, GST activity, GSH content, and the least MDA content, whereas APA (Mac) extract showed the highest catalase activity. Lastly, docking results suggested maximum binding affinity of tannic acid with HER2 and GCR receptors. This study provides evidence that A. precatorius seed extracts possess promising bioactive compounds with probable anticancer and antioxidant properties against cervical cancer for restricting tumor growth.

Insulin like growth factor-1 works synergistically with dopamine to attenuate diabetic retinopathy by downregulating vascular endothelial growth factor.

AIM: Levels of Insulin-like growth factor-1 (IGF-1), a proangiogenic growth factor is elevated and dopamine downregulated in proliferative diabetic retinopathy (PDR). This study aims to investigate whether IGF-1 with dopamine can together modulate vascular endothelial growth factor (VEGF) to prevent proliferative diabetic retinopathy while also attenuating angiogenic effects of IGF-1. METHODS: Effect of combination of levodopa L-Dopa with IGF-1 was tested on normal retinal pigment epithelium cells (ARPE-19) and human umbilical vein endothelial cells (HUVEC), followed by tube formation. Invivo analysis of anti-angiogenic potential assessed by chick chorioallantoic membrane (CAM) assay. Diabetes induction in wistar rats at two time points, 12 and 16 weeks, treated with L-Dopa+IGF-1 and analysed for morphological variations, serum and tissue dopamine levels, gene expression by real-time PCR and western blot assay. RESULTS: L-Dopa+IGF-1 on ARPE-19 cells caused no toxicity and worked synergistically. Reduced number of vessels observed. Significant improvement in inner retina thickness (*p < 0.05) was observed when L-Dopa was given alone and/or with IGF-1. Dopamine levels improved significantly in both serum and tissue (*p < 0.05). Levels of VEGF and IGF-1 receptors reduced significantly in 12 weeks. Western studies suggest that L-Dopa+IGF-1 modulates its effects via Akt/ERK dependent pathway. CONCLUSION: First ever report on synergistic effect of L-Dopa+IGF-1 in a rat model of diabetic retinopathy. Even though the effect of L-Dopa in combination with IGF-1 is comparable to levels of L-Dopa alone, this study presents an interesting finding of neuroprotective function of IGF-1, which has been studied in disease models of Parkinson's but not diabetes.

Lack of fiber cell induction stops normal growth of rat lenses in organ culture.

PURPOSE: Lens organ culture has been widely used as a model system for studying cataract induction and prevention. While rat lenses remain transparent and viable for a week or longer in culture, they do not increase in weight. This study was undertaken to determine what accounts for the lack of weight increase. METHODS: Lenses from 4-week-old Sprague-Dawley rats were cultured using standard methods. Histological analysis was performed on sections from methacrylate embedded tissue. 35S-labeled amino acids were used to metabolically label lenses in culture for the purpose of analyzing protein synthesis. BrdU labeling was used to assess synthesis of DNA in vivo and in vitro. RESULTS: Lenses from young, rapidly growing rats do not increase in weight after being put into organ culture. Protein synthesis continues in the cultured lenses although at decreased levels as time in culture increases. Lens epithelial cells continue to synthesize DNA as indicated by BrdU labeling, however, the normal migration of epithelial cells from the proliferative zone to the equator does not occur in culture. In the cultured lens, the shape of the lens bow gradually changes, becoming compressed towards the capsule. CONCLUSIONS: The differentiation of lens epithelial cells into fibers is arrested in the cultured lens; consequently lenses in organ culture do not grow normally.