The development of safe and effective immunomodulatory nanoplatforms is critical for advancing next-generation antimicrobial and vaccine strategies. In this study, we investigated the immunomodulatory efficacy and biocompatibility of antigenic PLGA- nanoparticles (aNPs) using zebrafish (Danio rerio) larvae as a vertebrate model of innate immunity. This work extends our previous findings in Caenorhabditis elegans, where Staphylococcus aureus cell wall membrane (CWM) antigens encapsulated within PLGA nanoparticles elicited controlled immune modulation in an invertebrate system relying solely on innate immunity.To evaluate vertebrate translational relevance, zebrafish larvae at early developmental stages, characterized predominantly by innate immune functionality, were exposed to both CWM and aNP formulations. In vivo toxicity analysis revealed no mortality or developmental abnormalities, confirming the biocompatibility of both CWM and aNP formulations. Quantitative gene expression analysis showed that aNP-treated larvae exhibited a regulated upregulation of key innate immune markers, including il-8, tnf-α, nf-κb, and mpo, compared to CWM-treated groups. These findings suggest that nanoparticle encapsulation may influence selected innate immune-related responses while maintaining biocompatibility under the tested conditions. Our findings support the potential of antigenic PLGA nanoparticles as a biocompatible platform for further investigation of immunomodulatory applications in vertebrate systems.

Bacterial derivatives and mimetics are receiving attention as an emerging immunostimulant in animal health, aquaculture, poultry, and immunotherapy due to their ability to enhance innate immune responses in a non-specific manner. Bacterial ghosts (BG) possess an inherent repertoire of surface-associated antigens and pathogen-associated molecular patterns (PAMPs) that confer strong immunostimulatory potential. To determine the immunomodulatory and developmental effect of bacterial ghost cells (GCs) derived from a Gram-positive bacterium, Staphylococcus aureus, prepared using Lugol’s iodine-based protocol, Danio rerio (zebrafish) embryos were exposed to different concentrations of GCs for 6 h, 12 h and 24 h. Time- and dose-dependent changes in zebrafish embryo morphology were assessed microscopically, while the immunostimulatory potential of GCs was evaluated through gene expression analysis using qRT-PCR. Overall, the results demonstrated that even at the highest concentration and longest exposure duration, GCs did not induce harmful effects or developmental abnormalities in the larvae. Gene expression analysis confirmed the immunostimulatory activity of GCs with upregulation of major innate immune signalling genes such as tlr2, nod2, nfkb and cytokines such as tnfa and il1b. These findings highlight the importance of small animal models such as zebrafish for understanding the in vivo behaviour of GCs, where complex interactions among multiple cell types contribute to the systemic response following treatment.

Effective immunomodulation of natural killer (NK) cells holds promise for enhancing innate immune responses in cancer and infectious diseases; however, free immunostimulants often induce only transient activation. In this study, we developed peptidoglycan (PG)-loaded poly(lactic-co-glycolic acid)–polyvinyl alcohol nanoparticles (PGN) as a biocompatible platform for sustained immune signalling. PGN fabricated via nanoprecipitation exhibited a spherical morphology with an average diameter of 150 nm, a zeta potential of − 15 mV, and approximately 18

ABSTRACT Bacterial melanin are macromolecules found in nature that provide a wide range of biological functions, including pigmentation, resistance to radiation, scavenging of free radicals, thermoregulation and protected from oxidative stress and harmful heavy metals. The melanin is crucial for pathogenesis and bacterial survival in a variety of circumstances, and they can also influence how bacteria interact with other organisms. Usually, bacteria produce the melanin is either black or brown colour. The produced melanin has excellent properties like antimicrobial, antioxidant, photoprotective and antibiofilm. This is a report on Corynebacterium amycolatum melanin-producing bacteria isolated from the marine sediment of Thiruvanmiyur beach in Tamil Nadu, India. Corynebacterium amycolatum was screened using tyrosine basal broth (TBB), and UV-visible spectroscopy, FTIR, and SEM were used to analyse the extracted melanin. The non-pathogenic nature of the Cornynebacterium amycolatum strain was verified through antibiotic sensitivity profiling. The cosmetic potential was evaluated using antioxidant and SPF assays. Corynebacterium amycolatum predominantly uses the DOPA pathway for melanin production, was confirmed using kojic acid inhibitor study. The in vitro studies on mouse fibroblast cell line (L929) and in vivo studies on zebra fish embryos shows non-cytotoxicity using this melanin, even in lower concentration confirms its potential to use in cosmetic formulation. This research aims to demonstrate that bacterial melanin is safe for the environment and has qualities that make it safer and more effective in cosmetics.

Immunotherapy is a promising and safer alternative to conventional cancer therapies. It involves adaptive T-cell therapy, cancer vaccines, monoclonal antibodies, immune checkpoint blockade (ICB), and chimeric antigen receptor (CAR) based therapies. However, most of these modalities encounter restrictions in solid tumours owing to a dense, highly hypoxic and immune-suppressive microenvironment as well as the heterogeneity of tumour antigens. The elevated intra-tumoural pressure and mutational rates within fastgrowing solid tumours present challenges in efficient drug targeting and delivery. The tumour microenvironment is a dynamic niche infiltrated by a variety of immune cells, most of which are macrophages. Since they form a part of the innate immune system, targeting macrophages has become a plausible immunotherapeutic approach. In this review, we discuss several versatile approaches (both at pre-clinical and clinical stages) such as the direct killing of tumour-associated macrophages, reprogramming pro-tumour macrophages to anti-tumour phenotypes, inhibition of macrophage recruitment into the tumour microenvironment, novel CAR macrophages, and genetically engineered macrophages that have been devised thus far. These strategies comprise a strong and adaptable macrophage-toolkit in the ongoing fight against cancer and by understanding their significance, we may unlock the full potential of these immune cells in cancer therapy.

In the complex realm of bacterial infections, particularly those caused by Staphylococcus aureus (S. aureus), macrophages play a pivotal role in orchestrating the immune response. During the initial stages of infection, the monocytes give rise to macrophages with a pro-inflammatory (M1 type) behaviour, engulfing and neutralizing the invading pathogens. However, under the sustained influence of S. aureus infection, monocytes can undergo a transition into an anti-inflammatory M2 state (pro-infection) rather than the M1 state (anti-infection), thereby compromising effective infection control. Therefore, it is necessary to develop a strategy that would preserve the pro-inflammatory functions of macrophages, in a safe and controlled manner. For this, we focused on harnessing the potential of S. aureus-derived ghost cells (GCs) which are non-live empty envelopes of bacterial cells, but with the antigenic determinants intact. Through a unique Lugol’s-iodine treatment, we generated GCs and characterization of these GCs using gel electrophoresis, FTIR, flow cytometry, TEM, and SEM confirmed their structural integrity. Following this, we assessed the extend of cellular association of the GCs with RAW267.4 macrophages, and observed an immediate interaction between the two, as evident from the flowcytometry and microscopy studies. We then performed macrophage polarisation on a human monocyte-macrophage model cell line, THP-1. Our findings revealed that GCs effectively activated macrophages, and promoted a pro-inflammatory polarisation with the expression of M1 differentiation markers (CD86, TNFα, IL-1β, IL-6, IL-12) evaluated through both qPCR and ELISA. Interestingly an intermediary expression of M2 markers viz., CD206 and IL-10 was also observed, but was overruled by the enhanced expression of M1 markers at a later time point. Overall, our study introduces a novel approach utilizing GCs to guide naïve macrophages towards M1 subtypes, thereby potentiating immune responses during microbial infections. This innovative strategy can modulate macrophage function, ultimately improving outcomes in S. aureus infections and beyond. A simplistic protocol developed for obtaining ghost cells (GC) from S. aureus. High cellular interaction of GCs demonstrated in macrophages within short incubation time intervals. GCs induced M1 macrophage polarization, upregulating markers like CD86 and cytokines TNF-α, IL-1β, IL-6, and IL-12. Selective M1 polarization demonstrated by GCs evident from the downregulation the anti-inflammatory cytokine IL-10. Novel approach for immunotherapy in S. aureus infections via pro-inflammatory (M1) polarization of macrophages.

This study explores the significance of antigenic nanoformulation in immunomodulation and in the interplay between immune response and nutrition. The work involves the development of a polylactic-co-glycolic acid (PLGA) biopolymer-based nanoparticle with immunogenic inclusions derived from Staphylococcus aureus cell wall and membrane (CWM) through a double emulsion method followed by their physio-chemical characterization and in vivo assessment in Caenorhabditis elegans (C. elegans). The prepared nanoparticles were monodispersed in nature and exhibited a diameter of 25 nm with stable colloidal nature and a zeta potential of − 25 ± 2 mV. The inclusion release and carrier degradation profiling revealed controlled and steady kinetics supporting the sustained availability of the encapsulated payload. The immunomodulatory studies conducted in C. elegans revealed that the expression of the stress indicator gene viz., sodh-1 was significantly upregulated in the CWM-treated worms and was notably reduced in the worms treated with the nanoformulation indicative of the slow release of the antigen which does not trigger untoward stress responses. In contrast, the expression of host defense genes viz., clec-7, ilys-3, igg-1, and cyp-37B1 in response to the CWM treatment was found to be downregulated, while for the nanoformulation treatment, the extent of downregulation was relatively lesser. A notable observation emerged as these genes, previously downregulated, exhibited a significant upsurge when the nutritional supplementation was amplified. This highlighted the profound influence of nutrition in fine-tuning the immune responses. Our data offers insights that could pave the way for further research in designing nutritional strategies to augment immunomodulatory interventions, as well as advocate for nanoparticle-based immunomodulatory approaches to prevent immune stress. Antigenic nanoparticles (aNPs) were precisely synthesized utilizing PLGA as the matrix and CWM as the payload, employing a water-in-oil-in-water emulsion (W/O/W) technique. This nanoformulation exhibited promising capabilities by attenuating the expression of the stress indicator gene and concurrently augmenting the expression of host defense genes under varying nutritional states within C. elegans. Our study stands as an exemplar, emphasizing the pivotal role of nanoformulations in achieving precise immunomodulation while outlining the significance of tailored nutritional strategies for effective immunomodulatory interventions.

The potential of bacteria-based immunotherapy lies in its ability to inherently enhance immune responses. However, the "liveness" of bacteria poses risks of bacterial escape, nonspecific immuno-stimulation, and ethical concerns, limiting their acceptability in immunotherapy. In this scenario, nonliving empty bacterial-cell envelopes, named bacterial ghosts (BGs), have emerged as immuno-stimulants with the potential to side-step the limitations of live bacterial therapies. This study demonstrates the capability of BGs in modulating the functionality of NK-92 cells and Caenorhabditis elegans (C. elegans), as well as perform as cytokine-therapy adjuvants. BGs were obtained through a pH-driven culture method, and were validated for their structural and chemical integrity via electron microscopy and spectroscopy. In NK-92 cells, BGs have shown significant immuno-stimulation by boosting the gene-expression of perforin, granzyme-B, Fas-L, and interferon-gamma by factors of 3.5-, 1.5-, 12.5-, and 8.6-folds, respectively. Combined BG and IL-12 treatment yielded a notable 10.2-fold increase in interferon-gamma protein expression in 24 h. The BGs also significantly influenced the innate immune response in C. elegans through the upregulation of lysozyme genes viz., ilys-3 (8.8-fold) and lys-2 (3.1-fold). Our investigation into the impact of BGs on natural killer cells and C. elegans highlights its potential as a valid alternative approach for new-age immunotherapy and cytokine augmentation.

Melanins are a ubiquitous group of pigments widely acclaimed as potent free-radical scavengers. The present study proposed to harness this property of melanins for anti-inflammatory and anti-cancer applications. Pyomelanin, a potent form of melanin analogous to alkaptomelanin in humans, was derived from Pseudoalteromonas species and fabricated into ultra-small pyomelanin nanogranules (PNGs) by facile routes. These melanin nanogranules were characterized for various physicochemical attributes using DLS, TEM, FTIR, EPR, XRD, and TGDA. Additionally, elemental analysis and long-term particle stability study was also conducted. The ultra-small particles were ~ 5–7 nm in size through TEM with a very strict size distribution. The free-radical scavenging activity of PNG assessed through the DPPH assay was comparable with that of ascorbic acid. Significant inflammatory markers viz., cyclooxygenase, lipoxygenase, myeloperoxidase, and cellular nitrate levels estimated in lipopolysaccharide-triggered RAW264.7 cells were reduced upon PNG treatment. The cancer cell kill effect of PNG was estimated in lung carcinoma cells in comparison with normal fibroblasts, wherein percentage inhibition in cancer cells was ~ 2-fold higher than that observed in normal fibroblasts. Overall, our results demonstrate the proof-of-principle of using pyomelanin nanogranules for therapeutic applications.

Nanomedicines consisting of combinations of cytotoxic drugs and molecular targeted therapeutics which inhibit specific downstream signals are evolving as a novel paradigm for breast cancer therapy. This research addresses one such combination of Paclitaxel (Ptx), having several adversities related to the activation of NF-kappa B pathway, with Epigallocatechin gallate (EGCG), a multiple signaling inhibitor, encapsulated within a targeted core/shell PLGA-Casein nanoparticle. The sequential release of EGCG followed by Ptx from this core/shell nanocarrier sensitized Ptx resistant MDA-MB-231 cells to Ptx, induced their apoptosis, inhibited NF-kappa B activation and downregulated the key genes associated with angiogenesis, tumor metastasis and survival. More importantly, Ptx-induced expression of P-glycoprotein was repressed by the nanocombination both at the protein and gene levels. This combination also offered significant cytotoxic response on breast cancer primary cells, indicating its translational value. (C) 2015 Elsevier Inc. All rights reserved.

The present study reports an engineered poly-L-lactide-co-glycolic acid (PLGA)-casein polymer-protein hybrid nanocarrier 190 +/- 12 nm in size entrapping a combination of chemically distinct (hydrophobic/hydrophilic) model drugs. A simple emulsion-precipitation route was adopted to prepare nearly monodispersed nanoparticles with distinct core/shell morphology entrapping paclitaxel (Ptx) in the core and epigallocatechin gallate (EGCG) in the shell, with the intention of providing a sequential and sustained release of these drugs. The idea was that an early release of EGCG would substantially increase the sensitivity of Ptx to cancer, thereby providing improved therapeutics at lower concentrations, with less toxicity. The hemo- and immunocompatibility of the core/shell nanomedicine was established in this study. The core/shell nanoparticles injected via the tail vein in Sprague-Dawley rats did not reveal any organ toxicity as was evident from histopathological evaluations of the major organs. In vivo pharmacokinetic studies in rats by high-performance liquid chromatography confirmed a sustained and sequential release of both the drugs in plasma, indicating prolonged circulation of the nanomedicine and enhanced availability of the drugs when compared to the bare drugs. Overall, the polymer-protein multilayered nanoparticles proved to be a promising platform for nanopolypharmaceutics. (C) 2013 Acts Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Magnetite/gold (Fe3O4/Au) hybrid nanoparticles were synthesized from a single iron precursor (ferric chloride) through a green chemistry route using grape seed proanthocyanidin as the reducing agent. Structural and physicochemical characterization proved the nanohybrid to be crystalline, with spherical morphology and size ∼35 nm. Magnetic resonance imaging and magnetization studies revealed that the Fe3O4 component of the hybrid provided superparamagnetism, with dark T2 contrast and high relaxivity (124.2 ± 3.02 mM −1 s−1). Phantom computed tomographic imaging demonstrated good X-ray contrast, which can be attributed to the presence of the nanogold component in the hybrid. Considering the potential application of this bimodal nanoconstruct for stem cell tracking and imaging, we have conducted compatibility studies on human Mesenchymal Stem Cells (hMSCs), wherein cell viability, apoptosis, and intracellular reactive oxygen species (ROS) generation due to the particle−cell interaction were asessed. It was noted that the material showed good biocompatibility even for high concentrations of 500 μg/mL and up to 48 h incubation, with no apoptotic signals or ROS generation. Cellular uptake of the nanomaterial was visualized using confocal microscopy and prussian blue staining. The presence of the nanohybrids were clearly visualized in the intracytoplasmic region of the cell, which is desirable for efficient imaging of stem cells in addition to the cytocompatible nature of the hybrids. Our work is a good demonstrative example of the use of green aqueous chemistry through the employment of phytochemicals for the room temperature synthesis of complex hybrid nanomaterials with multimodal functionalities.

Magnetite/gold (Fe(3)O(4)/Au) hybrid nanoparticles were synthesized from a single iron precursor (ferric chloride) through a green chemistry route using grape seed proanthocyanidin as the reducing agent. Structural and physicochemical characterization proved the nanohybrid to be crystalline, with spherical morphology and size ~35 nm. Magnetic resonance imaging and magnetization studies revealed that the Fe(3)O(4) component of the hybrid provided superparamagnetism, with dark T(2) contrast and high relaxivity (124.2 ± 3.02 mM(-1) s(-1)). Phantom computed tomographic imaging demonstrated good X-ray contrast, which can be attributed to the presence of the nanogold component in the hybrid. Considering the potential application of this bimodal nanoconstruct for stem cell tracking and imaging, we have conducted compatibility studies on human Mesenchymal Stem Cells (hMSCs), wherein cell viability, apoptosis, and intracellular reactive oxygen species (ROS) generation due to the particle-cell interaction were asessed. It was noted that the material showed good biocompatibility even for high concentrations of 500 μg/mL and up to 48 h incubation, with no apoptotic signals or ROS generation. Cellular uptake of the nanomaterial was visualized using confocal microscopy and prussian blue staining. The presence of the nanohybrids were clearly visualized in the intracytoplasmic region of the cell, which is desirable for efficient imaging of stem cells in addition to the cytocompatible nature of the hybrids. Our work is a good demonstrative example of the use of green aqueous chemistry through the employment of phytochemicals for the room temperature synthesis of complex hybrid nanomaterials with multimodal functionalities.

Polyoxometalates (POMs) show great molecular diversity and have significant applications in material science as well as in medicine. In this study, nano-complexation of a novel europium containing polyanion [Cs⊂Eu6As6W63O218(H2O)14(OH)4]25− (EuWAs) with biocompatible chitosan was achieved through ionotropic gelation technique without the aid of any cross-linker. Thus obtained chitosan/EuWAs nano-complex was characterized using DLS and Zeta analysis, FT-IR, SEM, AFM, TG/DTA, EDAX and fluorescence spectroscopy. The cross-linking efficiency of EuWAs with chitosan was calculated to be 81% and the release profile recorded at physiological pH was slow and sustained. Cytotoxicity assays performed on a host of cancer cell lines, viz., KB, MCF-7, PC-3 and A549 proved the anticancer activity of the nanocomplex and flow cytometry studies revealed that reactive oxygen species generation can be the plausible mechanism for the apoptosis induced by this material. Our study has thus indicated the feasibility of using chitosan/EuWAs nano-complex for anticancer applications.

The concept of ‘green’ chemotherapy by employing targeted nanoparticle mediated delivery to enhance the efficacy of phytomedicines is reported. Poly (lactide-co-glycolide) (PLGA) nanoparticles encapsulating a well known nutraceutical namely, grape seed extract (GSE)—‘NanoGSE’—was prepared by a nanoprecipitation technique. The drug-loaded nanoparticles of size ∼ 100 nm exhibited high colloidal stability at physiological pH. Molecular receptor targeting of this nanophytomedicine against folate receptor over-expressing cancers was demonstrated in vitro by conjugation with a potential cancer targeting ligand, folic acid (FA). Fluorescence microscopy and flow cytometry data showed highly specific cellular uptake of FA conjugated NanoGSE on folate receptor positive cancer cells. Studies were also conducted to investigate the efficiency of targeted (FA conjugated) versus non-targeted (non-FA conjugated) nanoformulations in causing cancer cell death. The IC50 values were lowered by a factor of ∼ 3 for FA-NanoGSE compared to the free drug, indicating substantially enhanced bioavailability to the tumor cells, sparing the normal ones. Receptor targeting of FA-NanoGSE resulted in a significant increase in apoptotic index, which was also quantified by flow cytometry and fluorescence microscopy. This in vitro study provides a basis for the use of nanoparticle mediated delivery of anticancer nutraceuticals to enhance bioavailability and effectively target cancer by a ‘green’ approach.