In-vitro antioxidant and anti-inflammatory potential along with p.o. pharmacokinetic profile of key bioactive phytocompounds of Snow Mountain Garlic: a comparative analysis vis-à-vis normal garlic.

Snow mountain garlic (SMG) is a trans-Himalayan medicinal plant used in the traditional medicine system for several ailments, including inflammatory arthritis. Research studies are insufficient to validate its folk medicinal applications. In the present study, the comparative abundance of its key bioactive phytocompounds, viz., S-allyl-L-cysteine (SAC), alliin, and S-methyl-L-cysteine (SMC) against normal garlic were assessed using the LC-MS/MS-MRM method. In addition, the study also explored the antioxidant and anti-inflammatory potency of crude extract of SMG and purified signature phytocompounds (i.e., SMC, SAC, and alliin) in comparison with normal garlic and dexamethasone in LPS-stimulated RAW264.7 macrophage cells. The LC-MS/MS-MRM study revealed significant differences among SMG and normal garlic, viz., alliin 22.8-fold higher in SMG, and SMC could be detected only in SMG. In the bioassays, SMG extract and purified signature phytocompounds significantly downregulated oxidative damage in activated macrophages, boosting endogenous antioxidants' activity. SMG extract-treated macrophages significantly suppressed NF-κB expression and related inflammatory indicators such as cytokines, COX-2, iNOS, and NO. Notably, the observed anti-inflammatory and antioxidant bioactivities of SMG extract were comparable to signature phytocompounds and dexamethasone. In addition, SAC being uniformly found in SMG and normal garlic, its comparative pharmacokinetics was studied to validate the pharmacodynamic superiority of SMG over normal garlic. Significantly higher plasma concentrations (Cmax), half-life (t1/2), and area under curve (AUC) of SAC following SMG extract administration than normal garlic validated the proposed hypothesis. Thus, the abundance of bioactive phytocompounds and their better pharmacokinetics in SMG extract might be underlying its medicinal merits over normal garlic.

Effect of age of tobacco initiation and number of failed quit attempts on maintenance of tobacco abstinence.

BACKGROUND: The decision to make a quit attempt is the first step toward the tobacco cessation process. It is well established in the literature that if someone does not take tobacco till the age of 21 years then his chances of remaining tobacco-free for life are higher than his counterparts who start tobacco at early developmental ages. METHODOLOGY AND TOOLS: The present study was conducted among 400 university undergraduate students. A cross-sectional survey design was used, multi-stage sampling was done, and four colleges were selected via random sampling. The motivation to quit tobacco, tobacco craving, and maintenance of tobacco abstinence was assessed via contemplation ladder, tobacco craving questionnaire Short Form, and smoking abstinence questionnaire. To validate subjective data, a urine cotinine test was performed. RESULTS: The age of tobacco initiation significantly impacts intentions to quit tobacco and tobacco craving levels. The number of unsuccessful quit attempts was also significantly associated with the maintenance of tobacco abstinence. The failed quit attempts play a vital role in altering tobacco cravings and make the withdrawals more complicated to handle for recent tobacco quitters.

An image analysis approach to characterize micronuclei differences in different subtypes of breast cancer.

BACKGROUND: Micronuclei (MN) have been used as screening, diagnostic and prognostic markers in multiple cancer types, including breast cancer (BC). However, the question that the MN present in all subtypes of BC are similar or different remains unanswered. We thus hypothesized that MN present in different subtypes of BC may differ in their contents which may be visible as differences in their morphologic and morphometric features. This study was thus carried out with the aim to identify the differences between MN morphometry, complexity, and texture in different subtypes of BC, such as estrogen and progesterone receptor-positive (ER+/PR+; MCF-7, T-47D), human epidermal growth factor receptor-positive (Her2 +;SKBR3) and triple-negative BC (TNBC; MDA-MB-231, MDA-MB-468) cell lines (CLs) by ImageJ software. METHODS: For analysis of MN dimensions, MN irregularity, and texture, we used morphometry and two mathematical computer-assisted algorithms, i.e., fractal dimension (FD) and grey level co-occurrence matrix (GLCM) of ImageJ software. RESULTS: MN area and perimeter values showed differences in the size of MN in different subtypes of BC, with the largest MN in TNBC CLs. GLCM parameters (entropy, angular second moment, inverse difference moment, contrast, and correlation) showed highly heterogenous texture in case of TNBC MN as compared to the others. FD analysis also revealed more complexity and irregularity in MN found in TNBC cells. CONCLUSION: The study for the first time showed morphometric, architectural and texture related differences amongst MN present in different subtypes of BC. The above may reflect differences in their composition and contents. Further, these differences may point towards the distinct mechanisms involved in the formation of MN in different subtypes of BC that need to be explored further.

Aspirin as a potential drug repurposing candidate targeting estrogen receptor alpha in breast cancer: a molecular dynamics and in-vitro study.

Estrogen receptor alpha (ERα) is expressed by 70% of breast cancers (BCs). Any deregulation in ERα signaling is crucial for the initiation and progression of BC. Because of development of resistance to anti-estrogenic compounds, repurposing existing drugs is an apt strategy to avoid a long drug-discovery process. Substantial epidemiologic evidence suggests that Aspirin use reduces the risk of different cancers including BC, while its role as an adjuvant or a possible antineoplastic agent in cancer treatment is being investigated. In this study, we attempted to explore possibilities of ERα inhibition by Aspirin which may act through competitive binding to the ligand binding domain (LBD) of ERα. A list of 48 ERα-LBD crystal structures bound with agonists, antagonists, and selective ER modulators (SERMs) was thoroughly analysed to determine interaction patterns specific to each ligand category. Exhaustive docking and 500 ns molecular dynamics (MD) studies were performed on three ERα - Aspirin complexes generated using agonist, antagonist, and SERM-bound crystal structures. Besides, three ERα crystal structures bound to agonist, antagonist, and SERM respectively were also subjected to MD simulations. Aspirin showed good affinity to LBD of ERα. Comparative analyses of binding patterns, conformational changes and molecular interaction profiles from the docking results and MD trajectories suggests that Aspirin was most stable in complex generated using SERM bound crystal structure of ERα and showed interactions with Gly-521, Ala-350, Leu-525 and Thr-347 like SERMs. In addition, in-vitro assays, qPCR, and immunofluorescent assay demonstrated the decline in the expression of ERα in MCF-7 upon treatment with Aspirin. These preliminary bioinformatical and in-vitro findings may form the basis to consider Aspirin as a potential candidate for targeting ERα, especially in tamoxifen-resistant cancers.Communicated by Ramaswamy H. Sarma.

Touch imprint smear: A prerequisite to obtain better quality and "true" tumor RNA in breast tissues.

BACKGROUND: RNA is the primary genetic material required for various molecular studies. RNA derived from breast tissue has low quality and quantity compared to that extracted from other tissues. Therefore, optimization of techniques for breast tissue RNA extraction is a challenging but essential requirement. METHODS: RNA was extracted from 60 samples of breast cancer after dividing them into 2 groups. Each tissue was divided into 2 halves for RNA extraction and histopathology respectively. In group 2 RNA was extracted after taking touch imprints whereas group1 was not subjected to any such procedure. Concentration and purity of RNA was assessed by using spectrophotometer and 1% agarose gel followed by RT-PCR for 18 S rRNA and CCND1 gene. RESULTS: Based on microscopic observations of imprints, group 2 samples were further subdivided into 2 subgroups. Group 2 A (n = 30) showing tumor in imprint smears were found to yield best concentration of pure RNA (1846.50 ng/µl and 1.92) as compared to group 2B (n = 15) with no malignancy in imprints (102.61 ng/µl and 1.53). The correlation of imprint smears with their corresponding H&E-stained slides further leads to grouping of each group in 2 groups. RT-PCR analyses showed better melting peaks and high relative expression of CCND1 in group 2 A. CONCLUSION: Touch imprints may provide valuable information regarding presence or absence of tumor in tissue samples being used for extraction of genetic material. This approach can be used as easy, cheap and fast strategy to resolve the doubts associated with RNA being truly representative of the tumor.

Motivation to quit tobacco; Impact of different types of Anti-tobacco state-sponsored media propaganda messages.

Introduction: Antitobacco media messages can easily reach the mass and play a very positive and significant role in changing the motivational stages among recent quitters. Motivation is the key to changing human behaviour. Motivation can be intrinsic and extrinsic. To modify tobacco-related behaviour, one must have an inherent motivation to quit tobacco. However, the outside factors, for example, protobacco advertisements, antitobacco advertisements, peer pressure, celebrity influence, and family members' influence cannot be ignored. Method: A total of 400 recent tobacco quitters were enrolled from four colleges via a multistage sampling method. Time series research design was used for data collection at three time periods 0, 1, and 3 months. Study participants were divided into four groups: 1) personal testimony group, 2) health warning group, 3) celebrity-influenced public service announcements, and 4) natural exposure group. Media messages containing antitobacco video clippings and pictures were delivered to the participants via phone thrice a week, as per the groups assigned. All four groups were assessed for the motivational stage via contemplation ladder at 0, 1, and 3 months of intervals. Results: Antitobacco personal testimonial media messages are most effective in enhancing the motivation to quit tobacco, followed by the antitobacco health warning messages, which are also proven to be effective in maintaining high motivation levels to remain abstinent from smoking. However, public service announcements are ineffective in keeping the motivation to quit tobacco at higher smoking. Conclusion: Overall, the antitobacco state-sponsored media messages, personal testimonials, and health warnings about tobacco products effectively maintain and enhance motivation to quit tobacco.

Development of nanostructured lipid carriers as a promising tool for methotrexate delivery: physicochemical and in vitro evaluation.

The aim of the present study is to fabricate the stable nanostructured lipid carriers (NLCs) using biocompatible excipients for the encapsulation of Methotrexate (MTX), a chemotherapeutic agent for breast cancer treatment. MTX has restricted clinical applications owing to its low solubility, non-specific targeting and adverse side effects. Glyceryl Monostearate (GMS) and Miglyol 812 (MI1) were chosen as solid and liquid lipids, respectively, for the fabrication of NLCs, and the influence of variation of solid and liquid composition was investigated. The prepared NLCs exhibited long-term stability and spherical shape morphology as characterized by electron microscopy. The internal structure of fabricated NLCs was arranged into cubic crystalline as confirmed by small-angle X-ray scattering (SAXS) analysis. MTX's encapsulation efficiency of ∼85 ± 0.9%. and sustained in vitro release of MTX ∼ 52% ± 3.0 in 24 h was achieved. Classical molecular dynamics (MD) simulations were performed to study the structural stability of the MTX encapsulated NLCs. Hemolysis carried out on the NLCs showcased the biosafety of the formulation under the tolerance limit (<10%). Further, the MTT assay demonstrates that MTX-loaded NLCs exhibited toxicity against HeLa and MCF-7 cell lines as compared to blank NLCs. The finding demonstrates NLCs as promising vehicles for MTX delivery to address cancer.Communicated by Ramaswamy H. Sarma.

Facially Amphiphilic Cholic Acid-Lysine Conjugates as Promising Antimicrobials.

The emergence of multidrug-resistant microbes is a significant health concern posing a constant need for new antimicrobials. Membrane-targeting antibiotics are promising candidates with reduced ability of microbes to develop resistance. In the present investigation, the principal reason behind choosing cholic acid as the crucial scaffold lies in the fact that it has a facially amphiphilic nature, which provides ample opportunity to refine the amphiphilicity by linking the amino acid lysine. A total of 16 novel amphipathic cholic acid derivatives were synthesized by sequentially linking lysine to C3-ß-amino cholic acid methyl ester to maintain the hydrophobic/hydrophilic balance, which could be the essential requirement for the antimicrobial activity. Among the synthesized conjugates, a series with fluorenyl-9-methoxycarbonyl moiety attached to cholic acid via lysine linker showed promising antimicrobial activity against Staphylococcus aureus, Escherichia coli, and Candida albicans. A pronounced effect of increase in lysine residues was noted on the observed activity. The lead compounds were found to be active against drug-resistant bacterial and fungal clinical isolates and also improved the efficacy of antifungal agents amphotericin B and voriconazole. Membrane-permeability studies demonstrated the ability of these compounds to induce membrane damage in the tested microbes. The active conjugates did not show any hemolytic activity and were also found to be nontoxic to the normal cells as well as the examined cancer cell lines. The observed antimicrobial activity was attributed to the facial amphiphilic conformations, hydrophobic/hydrophilic balance, and the overall charge on the molecules.