Pharmaceutical characterization and exploration of Arkeshwara rasa in MDA-MB-231 cells.

BACKGROUND: The diverse specificity mode of cancer treatment targets and chemo resistance demands the necessity of drug entities which can address the devastating dynamicity of the disease. OBJECTIVES: To check the anti-tumour potential of traditional medicine rich in polyherbal components and metal nanoparticle namely Arkeshwara rasa (AR). MATERIAL METHODS: The AR was prepared in a modified version with reference from Rasaratna Samuchaya and characterized using sophisticated instrumental analysis including XRD, SEM-EDAX, TEM, TGA-DSC, and LC-MS and tested against the MDA-MB-231 cell line to screen cell viability and the cytotoxicity with MTT, SRB and the AO assay. RESULTS: XRD pattern shows cubic tetrahedrite structure with Sb, Cu, S peaks and trace elements like Fe, Mg, etc. The particle size of AR ranges between 20 and 30 nm. The TGA points thermal decomposition at 210 °C and the metal sulphide peaks in DSC. LC-MS analysis reveals the components of the formulation more on the flavonoid portion. The IC50 value of MTT and SRB are 25.28 µg/mL and 31.7 µg/mL respectively. The AO colorimeter substantiated the cell viability and the apoptosis figures of the same cell line. The AR exhibits cytotoxicity and reaffirms the apoptosis fraction with SRB assay. CONCLUSIONS: The Hesperidine, Neohesperidin, Rutin components in the phytochemical pool can synergize the anti-tumour potential with either influencing cellular pathways or decreasing chemo resistance to conventional treatment. AR need to be further experimented with reverse transcription, flow cytometry, western blotting, etc.

Ocimum sanctum Alters the Lipid Landscape of the Brain Cortex and Plasma to Ameliorate the Effect of Photothrombotic Stroke in a Mouse Model.

Stroke-like injuries in the brain result in not only cell death at the site of the injury but also other detrimental structural and molecular changes in regions around the stroke. A stroke-induced alteration in the lipid profile interferes with neuronal functions such as neurotransmission. Preventing these unfavorable changes is important for recovery. Ocimum sanctum (Tulsi extract) is known to have anti-inflammatory and neuroprotective properties. It is possible that Tulsi imparts a neuroprotective effect through the lipophilic transfer of active ingredients into the brain. Hence, we examined alterations in the lipid profile in the cerebral cortex as well as the plasma of mice with a photothrombotic-ischemic-stroke-like injury following the administration of a Tulsi extract. It is also possible that the lipids present in the Tulsi extract could contribute to the lipophilic transfer of active ingredients into the brain. Therefore, to identify the major lipid species in the Tulsi extract, we performed metabolomic and untargeted lipidomic analyses on the Tulsi extract. The presence of 39 molecular lipid species was detected in the Tulsi extract. We then examined the effect of a treatment using the Tulsi extract on the untargeted lipidomic profile of the brain and plasma following photothrombotic ischemic stroke in a mouse model. Mice of the C57Bl/6j strain, aged 2-3 months, were randomly divided into four groups: (i) Sham, (ii) Lesion, (iii) Lesion plus Tulsi, and (iv) Lesion plus Ibuprofen. The cerebral cortex of the lesioned hemisphere of the brain and plasma samples were collected for untargeted lipidomic profiling using a Q-Exactive Mass Spectrometer. Our results documented significant alterations in major lipid groups, including PE, PC, neutral glycerolipids, PS, and P-glycerol, in the brain and plasma samples from the photothrombotic stroke mice following their treatment with Tulsi. Upon further comparison between the different study groups of mice, levels of MGDG (36:4), which may assist in recovery, were found to be increased in the brain cortexes of the mice treated with Tulsi when compared to the other groups (p < 0.05). Lipid species such as PS, PE, LPG, and PI were commonly altered in the Sham and Lesion plus Tulsi groups. The brain samples from the Sham group were specifically enriched in many species of glycerol lipids and had reduced PE species, while their plasma samples showed altered PE and PS species when compared to the Lesion group. LPC (16:1) was found in the Tulsi extract and was significantly increased in the brains of the PTL-plus-Tulsi-treated group. Our results suggest that the neuroprotective effect of Tulsi on cerebral ischemia may be partially associated with its ability to regulate brain and plasma lipids, and these results may help provide critical insights into therapeutic options for cerebral ischemia or brain lesions.

Quantitative analysis of the bioenergetics of Mycobacterium tuberculosis along with Glyoxylate cycle as a drug target under inhibition of enzymes using Petri net.

The bacteria Mycobacterium tuberculosis is responsible for the infectious disease Tuberculosis. Targeting the tubercule bacteria is an important challenge in developing the antimycobacterials. The glyoxylate cycle is considered as a potential target for the development of anti-tuberculosis agents, due to its absence in the humans. Humans only possess tricarboxylic acid cycle, while this cycle gets connected to glyoxylate cycle in microbes. Glyoxylate cycle is essential to the Mycobacterium for its growth and survival. Due to this reason, it is considered as a potential therapeutic target for the development of anti-tuberculosis agents. Here, we explore the effect on the behavior of the tricarboxylic acid cycle, glyoxylate cycle and their integrated pathway with the bioenergetics of the Mycobacterium, under the inhibition of key glyoxylate cycle enzymes using Continuous Petri net. Continuous Petri net is a special Petri net used to perform the quantitative analysis of the networks. We first study the tricarboxylic acid cycle and glyoxylate cycle of the tubercule bacteria by simulating its Continuous Petri net model under different scenarios. Both the cycles are then integrated with the bioenergetics of the bacteria and the integrated pathway is again simulated under different conditions. The simulation graphs show the metabolic consequences of inhibiting the key glyoxylate cycle enzymes and adding the uncouplers on the individual as well as integrated pathway. The uncouplers that inhibit the synthesis of adenosine triphosphate, play an important role as anti-mycobacterials. The simulation study done here validates the proposed Continuous Petri net model as compared with the experimental outcomes and also explains the consequences of the enzyme inhibition on the biochemical reactions involved in the metabolic pathways of the mycobacterium.

Ocimum sanctum [Tulsi] as a Potential Immunomodulator for the Treatment of Ischemic Injury in the Brain.

Stroke causes brain damage and is one of the main reasons for death. Most survivors of stroke face long-term physical disabilities and cognitive dysfunctions. In addition, they also have persistent emotional and behavioral changes. The two main treatments that are effective are reperfusion with recombinant tissue plasminogen activator and recanalization of penumbra using mechanical thrombectomy. However, these treatments are suitable only for a few patients due to limitations such as susceptibility to hemorrhage and the requirement for administering tissue plasminogen activators within the short therapeutic window during the early hours following a stroke. The paucity of interventions and treatments could be because of the multiple pathological mechanisms induced in the brain by stroke. The ongoing immune response following stroke has been attributed to the worsening brain injury. Hence, novel compounds with immunomodulatory properties that could improve the outcome of stroke patients are required. Natural compounds and medicinal herbs with anti-inflammatory activities and having minimal or no adverse systemic effect could be beneficial in treating stroke. Ocimum sanctum is a medicinal herb that can be considered an effective therapeutic option for ischemic brain injury. Ocimum sanctum, commonly known as holy basil or "Tulsi," is mentioned as the "Elixir of Life" for its healing powers. Since antiquity, Tulsi has been used in the Ayurvedic and Siddha medical systems to treat several diseases; it possesses immuno-modulatory activity, which can alter cellular and humoral immune responses. Tulsi can be considered a potential option as an immuno-modulator for treating various diseases, including brain stroke. In this review, we will focus on the immunomodulatory properties of Tulsi, specifically its effect on both innate and adaptive immunity, as well as its antioxidant and anti-inflammatory properties, which could potentially be effective in treating ongoing immune reactions following ischemic brain injury.

Phytochemical: a treatment option for heavy metal induced neurotoxicity.

Heavy metals are known to be carcinogenic, mutagenic, and teratogenic. Some heavy metals are necessary while present in the growing medium in moderate concentrations known to be essential heavy metals as they required for the body functioning as a nutrient. But there are some unwanted metals and are also toxic to the environment and create a harmful impact on the body, which termed to be non-essential heavy metals. Upon exposure, the heavy metals decrease the major antioxidants of cells and enzymes with the thiol group and affect cell division, proliferation, and apoptosis. It interacts with the DNA repair mechanism and initiates the production of reactive oxygen species (ROS). It subsequently binds to the mitochondria and may inhibit respiratory and oxidative phosphorylation in even low concentrations. This mechanism leads to damage antioxidant repair mechanism of neuronal cells and turns into neurotoxicity. Now, phytochemicals have led to good practices in the health system. Phytochemicals that are present in the fruits and herbs can preserve upon free radical damage. Thus, this review paper summarized various phytochemicals which can be utilized as a treatment option to reverse the effect of the toxicity caused by the ingestion of heavy metals in our body through various environmental or lifestyles ways.

Mechanistic insights into the antimycobacterial action of unani formulation, Qurs Sartan Kafoori.

Background and aim: Tuberculosis (TBC) is a deadly disease and major health issue in the world. Emergence of drug resistant strains further worsens the efficiency of available anti-TBC drugs. Natural compounds and particularly traditional medicines such as Unani drugs are one of the promising alternatives that have been widely used nowadays. This study aims to evaluate the efficacy of unani drug Qurs-e-Sartan Kafoori (QSK) on Mycobacterium tuberculosis (MTB). Experimental procedures: Drug susceptibilities were estimated by broth microdilution assay. Cell surface integrity was assessed by ZN staining, colony morphology and nitrocefin hydrolysis. Biofilms were visualized by crystal violet staining and measurement of metabolic activity and biomass. Lipidomics analysis was performed using mass spectrometry. Host pathogen interaction studies were accomplished using THP-1 cell lines to estimate cytokines by ELISA kit, apoptosis and ROS by flow cytometry. Results: QSK enhanced the susceptibilities of isoniazid and rifampicin and impaired membrane homeostasis as depicted by altered cell surface properties and enhanced membrane permeability. In addition, virulence factor, biofilm formation was considerably reduced in presence of QSK. Lipidomic analysis revealed extensive lipid remodeling. Furthermore, we used a THP-1 cell line model, and investigated the immunomodulatory effect by estimating cytokine profile and found change in expressions of TNF-α, IL-6 and IL-10. Additionally, we uncover reduced THP-1 apoptosis and enhanced ROS production in presence of QSK. Conclusion: Together, this study validates the potential of unani formulation (QSK) with its mechanism of action and attempts to highlight its significance in MDR reversal.

Study of the bioenergetics to identify the novel pathways as a drug target against Mycobacterium tuberculosis using Petri net.

Tuberculosis is one of the life-threatening diseases globally, caused by the bacteria Mycobacterium tuberculosis. In order to control this epidemic globally, there is an urgent need to discover new drugs with novel mechanism of action that can help in shortening the duration of treatment for both drug resistant and drug sensitive tuberculosis. Mycobacterium essentially depends on oxidative phosphorylation for its growth and establishment of pathogenesis. This pathway is unique in Mycobacterium tuberculosis as compared to host due to the differences in some of the enzyme complexes carrying electron transfer. Hence, it serves as an important drug target area. The uncouplers which inhibit adenosine triphosphate synthesis, could play a vital role in serving as antimycobacterial agents and thus could help in eradicating this deadly disease. In this article, the bioenergetics of Mycobacterium tuberculosis are studied with and without uncouplers using Petri net. Petri net is among the most widely used mathematical and computational tools to model and study the complex biochemical networks. We first represented the bioenergetic pathway as a Petri net which is then validated and analyzed using invariant analysis techniques of Petri net. The valid mathematical models presented here are capable to explain the molecular mechanism of uncouplers and the processes occurring within the electron transport chain of Mycobacterium tuberculosis. The results explained the net behavior in agreement with the biological results and also suggested some possible processes and pathways to be studied as a drug target for developing antimycobacterials.

Understanding Human Microbiota Offers Novel and Promising Therapeutic Options against Candida Infections.

Human fungal pathogens particularly of Candida species are one of the major causes of hospital acquired infections in immunocompromised patients. The limited arsenal of antifungal drugs to treat Candida infections with concomitant evolution of multidrug resistant strains further complicates the management of these infections. Therefore, deployment of novel strategies to surmount the Candida infections requires immediate attention. The human body is a dynamic ecosystem having microbiota usually involving symbionts that benefit from the host, but in turn may act as commensal organisms or affect positively (mutualism) or negatively (pathogenic) the physiology and nourishment of the host. The composition of human microbiota has garnered a lot of recent attention, and despite the common occurrence of Candida spp. within the microbiota, there is still an incomplete picture of relationships between Candida spp. and other microorganism, as well as how such associations are governed. These relationships could be important to have a more holistic understanding of the human microbiota and its connection to Candida infections. Understanding the mechanisms behind commensalism and pathogenesis is vital for the development of efficient therapeutic strategies for these Candida infections. The concept of host-microbiota crosstalk plays critical roles in human health and microbiota dysbiosis and is responsible for various pathologies. Through this review, we attempted to analyze the types of human microbiota and provide an update on the current understanding in the context of health and Candida infections. The information in this article will help as a resource for development of targeted microbial therapies such as pre-/pro-biotics and microbiota transplant that has gained advantage in recent times over antibiotics and established as novel therapeutic strategy.

Protein kinases as potential anticandidal drug targets.

Candidal infections are increasing at an alarming rate due to hospital acquired infections causing high mortality rates worldwide. Moreover, the emergence of drug resistant Candida strains is the major impediment against effective therapeutics. Thus, there is an imperious need to search for novel antifungal drug targets. Among various fungi, Candida albicans is one of  the most prevalent human fungal pathogen. Protein kinases modify other signaling molecules through phosphorylation and transduce extracellular stimuli for adaptation ensuing C. albicans growth, persistence and pathogenesis. In C. albicans, there are various kinds of kinases such as MAP (Mitogen Activated Protein) kinase cascade involving Hog1 (High-osmolarity glycerol) and Cek1 (C. albicans ERK-like Kinase1) mediated pathways, cyclin dependent pathway, cAMP (cyclic adenosine monophosphate) -dependent protein kinase pathway and TOR signaling pathway. Herein we have reviewed the variety of functions served by protein kinases in C. albicans.  Additionally, we have discussed the inhibitors for targeting these kinases. Together, we explore the potential of these kinases as effective drug target and discuss the progress made in the development of inhibitors against these targets.

Antifungal Action of Methylene Blue Involves Mitochondrial Dysfunction and Disruption of Redox and Membrane Homeostasis in C. albicans.

Candida albicans is known to cause infections ranging from superficial and systemic in immunocompromised person. In this study, we explored that the antifungal action of Methylene blue (MB) is mediated through mitochondrial dysfunction and disruption of redox and membrane homeostasis against C. albicans. We demonstrated that MB displayed its antifungal potential against C. albicans and two clinical isolates tested. We also showed that MB is effective against two non- albicans species as well. Notably, the antifungal effect of MB seems to be independent of the major drug efflux pumps transporter activity. We explored that MB treated Candida cells were sensitive on non-fermentable carbon source leading us to propose that MB inhibits mitochondria. This sensitive phenotype was reinforced with the fact that sensitivity of Candida cells to MB could be rescued upon the supplementation of ascorbic acid, an antioxidant. This clearly suggests that disturbances in redox status are linked with MB action. We further demonstrated that Candida cells were susceptible to membrane perturbing agent viz. SDS which was additionally confirmed by transmission electron micrographs showing disruption of membrane integrity. Moreover, the ergosterol levels were significantly decreased by 66% suggesting lipid compositional changes due to MB. Furthermore, we could demonstrate that MB inhibits the yeast to hyphal transition in C. albicans which is one of the major virulence attribute in most of the hyphal inducing conditions. Taken together, the data generated from present study clearly establishes MB as promising antifungal agent that could be efficiently employed in strategies to treat Candida infections.

Insights into the mode of action of anticandidal herbal monoterpenoid geraniol reveal disruption of multiple MDR mechanisms and virulence attributes in Candida albicans.

The anticandidal potential of Geraniol (Ger) against Candida albicans has already been established. The present study reveals deeper insights into the mechanisms of action of Ger. We observed that the repertoire of antifungal activity was not only limited to C. albicans and its clinical isolates but also against non-albicans species of Candida. The membrane tampering effect was visualized through transmission electron micrographs, depleted ergosterol levels and altered plasma membrane ATPase activity. Ger also affects cell wall as revealed by spot assays with cell wall-perturbing agents and scanning electron micrographs. Functional calcineurin pathway seems to be indispensable for the antifungal effect of Ger as calcineurin signaling mutant was hypersensitive to Ger while calcineurin overexpressing strain remained resistant. Ger also causes mitochondrial dysfunction, impaired iron homeostasis and genotoxicity. Furthermore, Ger inhibits both virulence attributes of hyphal morphogenesis and biofilm formation. Taken together, our results suggest that Ger is potential antifungal agent that warrants further investigation in clinical applications so that it could be competently employed in therapeutic strategies to treat Candida infections.

Therapeutic Potential of Dietary Phenolic Acids.

Although modern lifestyle has eased the quality of human life, this lifestyle's related patterns have imparted negative effects on health to acquire multiple diseases. Many synthetic drugs are invented during the last millennium but most if not all of them possess several side effects and proved to be costly. Convincing evidences have established the premise that the phytotherapeutic potential of natural compounds and need of search for novel drugs from natural sources are of high priority. Phenolic acids (PAs) are a class of secondary metabolites spread throughout the plant kingdom and generally involved in plethora of cellular processes involved in plant growth and reproduction and also produced as defense mechanism to sustain various environmental stresses. Extensive research on PAs strongly suggests that consumption of these compounds hold promise to offer protection against various ailments in humans. This paper focuses on the naturally derived PAs and summarizes the action mechanisms of these compounds during disease conditions. Based on the available information in the literature, it is suggested that use of PAs as drugs is very promising; however more research and clinical trials are necessary before these bioactive molecules can be made for treatment. Finally this review provides greater awareness of the promise that natural PAs hold for use in the disease prevention and therapy.

Iron Deprivation Affects Drug Susceptibilities of Mycobacteria Targeting Membrane Integrity.

Multidrug resistance (MDR) acquired by Mycobacterium tuberculosis (MTB) through continuous deployment of antitubercular drugs warrants immediate search for novel targets and mechanisms. The ability of MTB to sense and become accustomed to changes in the host is essential for survival and confers the basis of infection. A crucial condition that MTB must surmount is iron limitation, during the establishment of infection, since iron is required by both bacteria and humans. This study focuses on how iron deprivation affects drug susceptibilities of known anti-TB drugs in Mycobacterium smegmatis, a "surrogate of MTB." We showed that iron deprivation leads to enhanced potency of most commonly used first line anti-TB drugs that could be reverted upon iron supplementation. We explored that membrane homeostasis is disrupted upon iron deprivation as revealed by enhanced membrane permeability and hypersensitivity to membrane perturbing agent leading to increased passive diffusion of drug and TEM images showing detectable differences in cell envelope thickness. Furthermore, iron seems to be indispensable to sustain genotoxic stress suggesting its possible role in DNA repair machinery. Taken together, we for the first time established a link between cellular iron and drug susceptibility of mycobacteria suggesting iron as novel determinant to combat MDR.