Bioengineered ethosomes encapsulating AgNPs and Tasar silk sericin proteins for non melanoma skin carcinoma (NMSC) as an alternative therapeutics.

Rising cases of Non melanoma skin carcinoma (NMSC) and escalating levels of ultraviolet radiations have underlined a profound correlation with the elevating levels of environmental detoriation and increasing health issues. However, the availability of therapeutics has not aided in controlling the recurrence rates of skin carcinoma. Frequent administration of therapeutics with higher chances of facial deformity escalates the patient's treatment expenses. Thus, this study initiates a low cost effective and biodegradable therapy by exploring four formulations with combinations of silver nanoparticles (AgNPs), sericin (isolated from cocoons of Antherea mylitta) and chitosan. Subsequently, various ethosomal formulations were evaluated as a platform for transdermal delivery vehicle for efficient skin intervention therapeutics. Characterization using UV visible spectroscopy, Dynamic light scattering, Fourier Infrared spectroscopy, X-ray dispersion, Transmission electron microscopy, Fluorescence assisted cell sorting and in vitro studies were done and it was inferenced that equal combination of AgNPs and sericin facilitated to combat the morphological and cellular deformation of the epidermoid A431skin carcinoma cells. The overproduction of superoxide (O2.) and nitric oxide (NO) radicals consequently depolarized the mitochondrial membrane potential triggering apoptosis and necrosis. The in vivo experiments exhibited the stimulation of IgM secretion with T cell-mediated immune response. Therefore, this study proposes a novel approach for treatment of NMSC using biocompatible formulations delivered through ethosomes.

Gelatin carrageenan sericin hydrogel composites improves cell viability of cryopreserved SaOS-2 cells.

Cryopreservation and the low revival rate of cryopreserved cells remains a major challenge in cell based bone regeneration therapies. In our current study we aimed to develop a sericin based hydrogel composite incorporating various drugs and growth factors to enhance cell attachment, cryopreservation to increase the cellular viability upon revival. Sericin, gelatin and carrageenan blended hydrogel composites were prepared and explored for their physicochemical properties. The hydrogels prepared were porous and showed higher biocompatibility. Further, silver nanoparticles, alendronate and insulin like growth factor (IGF-1) were incorporated into the hybrid hydrogels individually and checked for sustained drug release profile. IGF-1 incorporated hydrogels composites showed better osteogenic cell attachment, proliferation and cell revival upon cryopreservation. The clonogenic potential of seeded cells upon 30 days of cryopreservation was also evaluated which was 55% in IGF-1 incorporated scaffold cells. A flow cytometry based staining protocol using Annexin V was developed which showed a live cell population up to 80% even after 30 days of crypreservation. These results validate the potential of our formulated hydrogels as cell based systems aimed for increasing cell survival upon cryopreservation and thus has a great potential for bone repair and regeneration.

Stavudine loaded gelatin liposomes for HIV therapy: Preparation, characterization and in vitro cytotoxic evaluation.

Despite continuous research and availability of 25 different active compounds for treating chronic HIV-1 infection, there is no absolute cure for this deadly disease. Primarily, the residual viremia remains hidden in latently infected reservoir sites and persistently release the viral RNA into the blood stream. The study proposes the dual utilization of the prepared stavudine-containing nanoformulations to control the residual viremia as well as target the reservoir sites. Gelatin nanoformulations containing very low dosage of stavudine were prepared through classical desolvation process and were later loaded in soya lecithin-liposomes. The nanoformulations were characterized through dynamic light scattering (DLS), Transmission electron microscopy (TEM), X-ray diffraction (XRD) and ATR-FTIR. All the formulations were in nano regime with high hemocompatibility and exhibited dose-dependent cytotoxicity towards Raw 264.7 macrophages. Among the various formulations, SG-3 (Stavudine-Gelatin Nanoformulation sample 3) and SG-LP-3 (Stavudine-Gelatin Nano-Liposome formulation sample 3) showed the best results in terms of yield, size, charge, encapsulation efficiency, hemocompatibility and % cell viability. For the first time, liposomal delivery of antiretroviral drugs using nanocarriers has been demonstrated using very low dosage (lower than the recommended WHO dosage) showing the prominent linear release of stavudine for up to 12h which would reduce the circulatory viremia as well as reach the sanctuary reservoir sites due to their nanosize. This method of liposomal delivery of antiretroviral drugs in very low concentrations using nanocarriers could provide a novel therapeutic alternative to target HIV reservoir sites.

Biofilm Impeding AgNPs Target Skin Carcinoma by Inducing Mitochondrial Membrane Depolarization Mediated through ROS Production.

Reactive oxygen species (ROS) are a double-edged sword that possesses both beneficial and harmful effects. Although basic research on skin cancer prevention has undergone a huge transformation, cases of recurrence with higher rates of drug resistance are some of its drawbacks. Therefore, targeting mitochondria by ROS overproduction provides an alternate approach for anticancer therapy. In the present study, green-synthesized silver nanoparticles (AgNPs) were explored for triggering the ROS production in A431 skin carcinoma cells. The synthesized AgNPs were characterized for size, charge, morphology, and phase through high-throughput DLS, Fe-SEM, XRD, and ATR-FTIR techniques. Their physiochemical properties with hemoglobin and blood plasma were screened through hemolysis, hemagglutination assay, and circular dichroism spectroscopy confirmed their nontoxic nature. The AgNPs also exhibited additional efficacy in inhibiting biofilm produced by V. cholerae and B. subtilis, thereby facilitating better applicability in wound-healing biomaterials. The depolarization of mitochondrial membrane potential ΔΨm through excess ROS production was deduced to be the triggering force behind the apoptotic cell death mechanism of the skin carcinoma. Subsequent experimentation through DNA fragmentation, comet tail formation, cell membrane blebbing, and reduced invasiveness potentials through scratch assay confirmed the physiological hallmarks of apoptosis. Thus, depolarizing mitochondrial membrane potential through green-synthesized AgNPs provides an economic, nontoxic, specific approach for targeting skin carcinoma with additional benefits of antibacterial activities.

In silico identification of outer membrane protein (Omp) and subunit vaccine design against pathogenic Vibrio cholerae.

Virulence-related outer membrane proteins (Omps) are expressed in bacteria (Gram-negative) such as V. cholerae and are vital to bacterial invasion in to eukaryotic cell and survival within macrophages that could be best candidate for development of vaccine against V. cholerae. Applying in silico approaches, the 3-D model of the Omp was developed using Swiss model server and validated byProSA and Procheck web server. The continuous stretch of amino acid sequences 26mer: RTRSNSGLLTWGDKQTITLEYGDPAL and 31mer: FFAGGDNNLRGYGYKSISPQDASGALTGAKY having B-cell binding sites were selected from sequence alignment after B cell epitopes prediction by BCPred and AAP prediction modules of BCPreds. Further, the selected antigenic sequences (having B-cell epitopes) were analyzed for T-cell epitopes (MHC I and MHC II alleles binding sequence) by using ProPred 1 and ProPred respectively. The epitope (9mer: YKSISPQDA) that binds to both the MHC classes (MHC I and MHC II) and covers maximum MHC alleles were identified. The identified epitopes can be useful in designing comprehensive peptide vaccine development against V. cholerae by inducing optimal immune response.

Ameliorating Effects of Green Synthesized Silver Nanoparticles on Glycated End Product Induced Reactive Oxygen Species Production and Cellular Toxicity in Osteogenic Saos-2 Cells.

Advanced glycation end-products (AGEs) that result from nonenzymatic glycation are one of the major factors involved in diabetes and its secondary complications and diseases. This necessitates our urge to discover new compounds that may be used as potential AGEs inhibitors without affecting the normal structure and function of biomolecules. In the present study, we investigated the inhibitory effects of AgNP (silver nanoparticles) on AGEs formation as well as their inhibitory effects on glycation mediated cell toxicity via reactive oxygen species (ROS) production and DNA damage. The excitation-emission fluorescence spectroscopy was employed to investigate the interaction of AgNP during glycation. The values of conditional stability constant (log Ka = 4.44) derived from the Stern-Volmer equation indicate that AgNP have strong binding capacity for glycated protein. UV-vis, fluorescence, and Fourier transform infrared spectral data reveal complexation of AgNP with glycated bovine serum albumin, which significantly inhibits AGEs formation in a concentration-dependent manner. Cytotoxic evaluations suggest that simultaneous administration of AgNP and glycated product reduces cell death (42.82% ± 3.54) as compared to the glycated product alone. Similarly, ROS production in AgNP treated cells is significantly less compared to only glycated product treated cells. Although DNA damage studies show DNA damage in both GP and GP-AgNP treated cells, fluorescence activated cell sorting analysis demonstrates that glycated products induce cell death by necrosis, while AgNP cause cell death via apoptotic pathways. AgNP have a positive effect on restoring native protein structure deduced from spectral studies, and hence, inferences can be drawn that AgNP have ameliorating effects on glycated induced cytotoxicity observed in osteogenic Saos-2 cells.

Development of highly biocompatible Gelatin & i-Carrageenan based composite hydrogels: In depth physiochemical analysis for biomedical applications.

In present investigation, gelatin and iota-carrageenan (CG) were used for the fabrication of composite hydrogels in different formulations (G1-G6). The hydrogels were characterized through optical, scanning electron and confocal microscopy to visualize their internal morphology. X-ray diffraction and ATR- FTIR spectroscopy were used for analyzing the chemical interaction between gelatin and CG. The hydrogels were found to be hemocompatible with high mucoadhesive and swelling properties. The standard drug ciprofloxacin was incorporated within the hydrogels and its interaction with the polymers was monitored through XRD and ATR-FTIR spectroscopy. In physiological pH 7.4, the G4 formulation showed a linear release profile. The antimicrobial activity was tested against nosocomial strains of Bacillus sp, Vibrio sp, Pseudomonas sp and Escherichia coli which showed a zone of inhibition between 8.5-20.7mm against the marketed Ciplox ointment. The cytocompatibility of the prepared hydrogels were investigated using normal HaCaT and HEK293 cell lines which showed their negligible cytotoxicity.

Enhanced efficacy of clindamycin hydrochloride encapsulated in PLA/PLGA based nanoparticle system for oral delivery.

Clindamycin hydrochloride (CLH) is a clinically important oral antibiotic with wide spectrum of antimicrobial activity that includes gram-positive aerobes (staphylococci, streptococci etc.), most anaerobic bacteria, Chlamydia and certain protozoa. The current study was focused to develop a stabilised clindamycin encapsulated poly lactic acid (PLA)/poly (D,L-lactide-co-glycolide) (PLGA) nano-formulation with better drug bioavailability at molecular level. Various nanoparticle (NPs) formulations of PLA and PLGA loaded with CLH were prepared by solvent evaporation method varying drug: polymer concentration (1:20, 1:10 and 1:5) and characterised (size, encapsulation efficiency, drug loading, scanning electron microscope, differential scanning calorimetry [DSC] and Fourier transform infrared [FTIR] studies). The ratio 1:10 was found to be optimal for a monodispersed and stable nano formulation for both the polymers. NP formulations demonstrated a significant controlled release profile extended up to 144 h (both CLH-PLA and CLH-PLGA). The thermal behaviour (DSC) studies confirmed the molecular dispersion of the drug within the system. The FTIR studies revealed the intactness as well as unaltered structure of drug. The CLH-PLA NPs showed enhanced antimicrobial activity against two pathogenic bacteria Streptococcus faecalis and Bacillus cereus. The results notably suggest that encapsulation of CLH into PLA/PLGA significantly increases the bioavailability of the drug and due to this enhanced drug activity; it can be widely applied for number of therapies.

Fabrication of poly(vinyl alcohol)-Carrageenan scaffolds for cryopreservation: Effect of composition on cell viability.

The present investigation reports the fabrication of three dimensional (3D), interconnected, highly porous, biodegradable scaffolds using freeze-gelation technique. The hydrogels prepared with different ratios (5:5, 6:4, 7:3, 8:2 and 9:1) of poly(vinyl alcohol) (PVA) and Carrageenan (Car) was lyophilized to obtain their respective scaffolds. The PVA-Car scaffolds were further characterized by scanning electron microscopy (SEM), X-ray diffraction (XRD) and attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR). The prepared scaffolds were found to be biodegradable and highly compatible with hemoglobin. Further, normal keratinocyte (HaCaT) and osteosarcoma (Saos-2) cells seeded on PVA-Car scaffolds were cryopreserved for 15days and their viability was checked at regular interval of 3days (0, 3, 6, 9, 12, 15 days) through MTT assay and fluorescence microscopy. Overall, the collective results indicate the scaffold constructs with 7:3 and 8:2 PVA-Car ratios possess ideal characteristics for tissue engineering applications and for long term cryopreservation of cells.

Synergistic combination of antioxidants, silver nanoparticles and chitosan in a nanoparticle based formulation: Characterization and cytotoxic effect on MCF-7 breast cancer cell lines.

HYPOTHESIS: Chitosan (Cs) is a biocompatible, biodegradable cationic polymer having the ability of targeted drug delivery. Vitamin E and C are not synthesized in our body thus, when encapsulated within a carrier system these vitamins in combination with/alone can be utilized for their anti-cancer potentials. EXPERIMENT: The present investigation was conducted to develop a stable nanoparticle based formulation encapsulating antioxidants (Vitamin E, catechol) and silver nanoparticles synthesized from Hibiscus rosa-sinensis (HRS) petal extracts within a chitosan matrix. The prepared nanoformulations were characterized using Field emission scanning electron microscopy (Fe-SEM), X-ray diffraction (XRD) and Attenuated Total Reflection Fourier Transform Infrared spectroscopy (ATR-FTIR). They were further tested for their antioxidant potentials using DPPH assay, hydrogen peroxide scavenging assay, nitric oxide scavenging assay and ferrous antioxidant reducing potential assay. FINDINGS: The nanoformulations were found to be highly hemocompatible and showed high encapsulation efficiency up to 76%. They also showed higher antioxidant activity than their base materials. Further, their anti-cancer efficacy was observed against MCF-7 breast cancer cells having IC50 values of 53.36±0.36µg/mL (chitosan-ascorbic acid-glucose), 55.28±0.85µg/mL (chitosan-Vitamin E), 63.72±0.27µg/mL (Chitosan-catechol) and 58.53±0.55µg/mL (chitosan-silver nanoparticles). Thus, the prepared formulations can be therapeutically applied for effective and targeted delivery in breast cancer treatment.

Bark extract mediated green synthesis of silver nanoparticles: Evaluation of antimicrobial activity and antiproliferative response against osteosarcoma.

In the current investigation we report the biosynthesis potentials of bark extracts of Ficus benghalensis and Azadirachta indica for production of silver nanoparticle without use of any external reducing or capping agent. The appearance of dark brown color indicated the complete nanoparticle synthesis which was further validated by absorbance peak by UV-vis spectroscopy. The morphology of the synthesized particles was characterized by Field emission- scanning electron microscopy (Fe-SEM) and atomic force microscopy (AFM). The X-ray diffraction (XRD) patterns clearly illustrated the crystalline phase of the synthesized nanoparticles. ATR-Fourier Transform Infrared (ATR-FTIR) spectroscopy was performed to identify the role of various functional groups in the nanoparticle synthesis. The synthesized nanoparticles showed promising antimicrobial activity against Gram negative (Escherichia coli, Pseudomonas aeruginosa and Vibrio cholerae) and Gram positive (Bacillus subtilis) bacteria. The synthesized nano Ag also showed antiproliferative activity against MG-63 osteosarcoma cell line in a dose dependent manner. Thus, these synthesized Ag nanoparticles can be used as a broad spectrum therapeutic agent against osteosarcoma and microorganisms.

Biosynthesis, characterisation and antimicrobial activity of silver nanoparticles using Hibiscus rosa-sinensis petals extracts.

Green synthesis of metallic nanoparticles has lured the world from the chemical and physical approaches owing to its rapid, non-hazardous and economic aspect of production mechanism. In this study, silver nanoparticles (AgNPs) were synthesised using petal extracts of Hibiscus rosa-sinensis. The AgNPs displayed characteristic surface plasmon resonance peak at around 421 nm having a mean particle size of 76.25±0.17 nm and carried a charge of -41±0.2 mV. The X-ray diffraction patterns displayed typical peaks of face centred cubic crystalline silver. The surface morphology was characterised by scanning electron microscopy and atomic force microscopy. Fourier transform infrared spectroscopy studies confirmed the surface modifications of the functional groups for the synthesis of AgNPs. Furthermore, the synthesised AgNPs displayed proficient antimicrobial activity against pathogenic strains of Vibrio cholerae, Escherichia coli, Klebsiella pneumoniae and Staphylococcus aureus.

Biologically synthesised silver nanoparticles from three diverse family of plant extracts and their anticancer activity against epidermoid A431 carcinoma.

HYPOTHESIS: Biological synthesis of silver nanoparticles is a cost effective natural process where the phytochemicals specifically phenols, flavonoids and terpenoids present in the plant extracts act as capping and reducing agent. Due to their nano size regime the silver nanoparticles may directly bind to the DNA of the pathogenic bacterial strains leading to higher antimicrobial activity. EXPERIMENT: In the current study silver nanoparticles were synthesised using plant extracts from different origin Cucurbita maxima (petals), Moringa oleifera (leaves) and Acorus calamus (rhizome). The synthesised nanoparticles were characterized by UV-visible spectroscopy, dynamic light scattering (DLS), X-ray diffraction (XRD), field emission scanning electron microscopy (Fe-SEM) and Fourier transform infrared spectroscopy (FTIR). FINDING: Highly crystalline, roughly spherical and cuboidal silver nanoparticles of 30-70 nm in size were synthesised. The nanoparticles provided strong antimicrobial activity against pathogenic strains. The effect of the synthesised nanoparticles against A431 skin cancer cell line was tested for their toxicity by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) dye. The IC50 values of 82.39±3.1, 83.57±3.9 and 78.58±2.7 µg/ml were calculated for silver nanoparticles synthesised by C. maxima, M. oleifera and A. calamus respectively.