Nanovaccines: An effective therapeutic approach for cancer therapy.

Cancer vaccines hold considerable promise for the immunotherapy of solid tumors. Nanomedicine offers several strategies for enhancing vaccine effectiveness. In particular, molecular or (sub) cellular vaccines can be delivered to the target lymphoid tissues and cells by nanocarriers and nanoplatforms to increase the potency and durability of antitumor immunity and minimize negative side effects. Nanovaccines use nanoparticles (NPs) as carriers and/or adjuvants, offering the advantages of optimal nanoscale size, high stability, ample antigen loading, high immunogenicity, tunable antigen presentation, increased retention in lymph nodes, and immunity promotion. To induce antitumor immunity, cancer vaccines rely on tumor antigens, which are administered in the form of entire cells, peptides, nucleic acids, extracellular vesicles (EVs), or cell membrane-encapsulated NPs. Ideal cancer vaccines stimulate both humoral and cellular immunity while overcoming tumor-induced immune suppression. Herein, we review the key properties of nanovaccines for cancer immunotherapy and highlight the recent advances in their development based on the structure and composition of various (including synthetic and semi (biogenic) nanocarriers. Moreover, we discuss tumor cell-derived vaccines (including those based on whole-tumor-cell components, EVs, cell membrane-encapsulated NPs, and hybrid membrane-coated NPs), nanovaccine action mechanisms, and the challenges of immunocancer therapy and their translation to clinical applications.

Antibacterial and antibiofilm effects of Pseudomonas aeruginosa derived outer membrane vesicles against Streptococcus mutans.

Antimicrobial resistance (AMR) is a serious and most urgent global threat to human health. AMR is one of today's biggest difficulties in the health system and has the potential to harm people at any stage of life, making it a severe public health issue. There must be fewer antimicrobial medicines available to treat diseases given the rise in antibiotic-resistant organisms. If no new drugs are created or discovered, it is predicted that there won't be any effective antibiotics accessible by 2050. In most cases, Streptococcus increased antibiotic resistance by forming biofilms, which account for around 80 % of all microbial infections in humans. This highlights the need to look for new strategies to manage diseases that are resistant to antibiotics. Therefore, development alternative, biocompatible and high efficacy new strategies are essential to overcome drug resistance. Recently, bacterial derived extracellular vesicles have been applied to tackle infection and reduce the emergence of drug resistance. Therefore, the objective of the current study was designed to assess the antibacterial and antibiofilm potential of outer membrane vesicles (OMVs) derived from Pseudomonas aeruginosa againstStreptococcus mutans. According to the findings of this investigation, the pure P. aeruginosa outer membrane vesicles (PAOMVs) display a size of 100 nm. S. mutans treated with PAOMVs showed significant antibacterial and antibiofilm activity. The mechanistic studies revealed that PAOMVs induce cell death through excessive generation of reactive oxygen species and imbalance of redox leads to lipid peroxidation, decreased level of antioxidant markers including glutathione, superoxide dismutase and catalase. Further this study confirmed that PAOMVs significantly impairs metabolic activity through inhibiting lactate dehydrogenase activity (LDH), adenosine triphosphate (ATP) production, leakage of proteins and sugars. Interestingly, combination of sub-lethal concentrations of PAOMVs and antibiotics enhances cell death and biofilm formation of S. mutans. Altogether, this work, may serve as an important basis for further evaluation of PAOMVs as novel therapeutic agents against bacterial infections.

Extracellular nanovesicles produced by Bacillus licheniformis: A potential anticancer agent for breast and lung cancer.

Cancer is a major public burden and leading cause of death worldwide; furthermore, it is a significant barrier to increasing life expectancy in most countries of the world. Among various types of cancers, breast and lung cancers lead to significant mortality in both males and females annually. Bacteria-derived products have been explored for their use in cancer therapy. Although bacteria contain significant amounts of anticancer substances, attenuated bacteria may still pose a potential risk for infection owing to the variety of immunomodulatory molecules present in the parental bacteria; therefore, non-cellular bacterial extracellular vesicles (BEVs), which are naturally non-replicating, safer, and are considered to be potential anticancer agents, are preferred for cancer therapy. Gram-positive bacteria actively secrete cytoplasmic membrane vesicles that are spherical and vary between 10 and 400 nm in size. However, no studies have considered cytoplasmic membrane vesicles derived from Bacillus licheniformisin cancer treatment. In this study, we investigated the potential use of B. licheniformis extracellular nanovesicles (BENVs) as therapeutic agents to treat cancer. Purified BENVs from the culture supernatant of B. licheniformis using ultracentrifugation and ExoQuick were characterized using a series of analytical techniques. Human breast cancer cells (MDA-MB-231) and lung cancer cells (A549) were treated with different concentrations of purified BENVs, which inhibited the cell viability and proliferation, and increased cytotoxicity in a dose-dependent manner. To elucidate the mechanism underlying the anticancer activity of BENVs, the oxidative stress markers such as reactive oxygen species (ROS) and glutathione (GSH) levels were measured. The ROS levels were significantly higher in BENV-treated cells, whereas the GSH levels were markedly reduced. Cells treated with BENVs, doxorubicin (DOX), or a combination of BENVs and DOX showed significantly increased expression of p53, p21, caspase-9/3, and Bax, and concomitantly decreased expression of Bcl-2. The combination of BENVs and doxorubicin enhanced mitochondrial dysfunction, DNA damage, and apoptosis. To our knowledge, this is the first study to determine the anticancer properties of BENVs derived from industrially significant probacteria on breast and lung cancer cells.

Bacterial extracellular vesicles: Emerging nanoplatforms for biomedical applications.

Bacterial extracellular vesicles (BEVs) are nanosized lipid bilayers generated from membranes that are filled with components derived from bacteria. BEVs are important for the physiology, pathogenicity, and interactions between bacteria and their hosts as well. BEVs represent an important mechanism of transport and interaction between cells. Recent advances in biomolecular nanotechnology have enabled the desired properties to be engineered on the surface of BEVs and decoration with desired and diverse biomolecules and nanoparticles, which have potential biomedical applications. BEVs have been the focus of various fields, including nanovaccines, therapeutic agents, and drug delivery vehicles. In this review, we delineate the fundamental aspects of BEVs, including their biogenesis, cargo composition, function, and interactions with host cells. We comprehensively summarize the factors influencing the biogenesis of BEVs. We further highlight the importance of the isolation, purification, and characterization of BEVs because they are essential processes for potential benefits related to host-microbe interactions. In addition, we address recent advancements in BEVs in biomedical applications. Finally, we provide conclusions and future perspectives as well as highlight the remaining challenges of BEVs for different biomedical applications.

Postbiotics: Functional Food Materials and Therapeutic Agents for Cancer, Diabetes, and Inflammatory Diseases.

Postbiotics are (i) "soluble factors secreted by live bacteria, or released after bacterial lysis, such as enzymes, peptides, teichoic acids, peptidoglycan-derived muropeptides, polysaccharides, cell-surface proteins and organic acids"; (ii) "non-viable metabolites produced by microorganisms that exert biological effects on the hosts"; and (iii) "compounds produced by microorganisms, released from food components or microbial constituents, including non-viable cells that, when administered in adequate amounts, promote health and wellbeing". A probiotic- and prebiotic-rich diet ensures an adequate supply of these vital nutrients. During the anaerobic fermentation of organic nutrients, such as prebiotics, postbiotics act as a benevolent bioactive molecule matrix. Postbiotics can be used as functional components in the food industry by offering a number of advantages, such as being added to foods that are harmful to probiotic survival. Postbiotic supplements have grown in popularity in the food, cosmetic, and healthcare industries because of their numerous health advantages. Their classification depends on various factors, including the type of microorganism, structural composition, and physiological functions. This review offers a succinct introduction to postbiotics while discussing their salient features and classification, production, purification, characterization, biological functions, and applications in the food industry. Furthermore, their therapeutic mechanisms as antibacterial, antiviral, antioxidant, anticancer, anti-diabetic, and anti-inflammatory agents are elucidated.

Graphene Oxide Enhances Biogenesis and Release of Exosomes in Human Ovarian Cancer Cells.

Background: Exosomes, which are nanovesicles secreted by almost all the cells, mediate intercellular communication and are involved in various physiological and pathological processes. We aimed to investigate the effects of graphene oxide (GO) on the biogenesis and release of exosomes in human ovarian cancer (SKOV3) cells. Methods: Exosomes were isolated using ultracentrifugation and ExoQuick and characterized by various analytical techniques. The expression levels of exosome markers were analyzed via quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. Results: Graphene oxide (10-50 µg/mL), cisplatin (2-10 µg/mL), and C6-ceramide (5-25 µM) inhibited the cell viability, proliferation, and cytotoxicity in a dose-dependent manner. We observed that graphene oxide (GO), cisplatin (CIS), and C6-Ceramide (C6-Cer) stimulated acetylcholine esterase and neutral sphingomyelinase activity, total exosome protein concentration, and exosome counts associated with increased level of apoptosis, oxidative stress and endoplasmic reticulum stress. In contrast, GW4869 treatment inhibits biogenesis and release of exosomes. We observed that the human ovarian cancer cells secreted exosomes with typical cup-shaped morphology and surface protein biomarkers. The expression levels of TSG101, CD9, CD63, and CD81 were significantly higher in GO-treated cells than in control cells. Further, cytokine and chemokine levels were significantly higher in exosomes isolated from GO-treated SKOV3 cells than in those isolated from control cells. SKOV3 cells pre-treated with N-acetylcysteine or GW4869 displayed a significant reduction in GO-induced exosome biogenesis and release. Furthermore, endocytic inhibitors decrease exosome biogenesis and release by impairing endocytic pathways. Conclusion: This study identifies GO as a potential tool for targeting the exosome pathway and stimulating exosome biogenesis and release. We believe that the knowledge acquired in this study can be potentially extended to other exosome-dominated pathologies and model systems. Furthermore, these nanoparticles can provide a promising means to enhance exosome production in SKOV3 cells.

Antifungal Effect of Nanoparticles against COVID-19 Linked Black Fungus: A Perspective on Biomedical Applications.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a highly transmissible and pathogenic coronavirus that has caused a 'coronavirus disease 2019' (COVID-19) pandemic in multiple waves, which threatens human health and public safety. During this pandemic, some patients with COVID-19 acquired secondary infections, such as mucormycosis, also known as black fungus disease. Mucormycosis is a serious, acute, and deadly fungal infection caused by Mucorales-related fungal species, and it spreads rapidly. Hence, prompt diagnosis and treatment are necessary to avoid high mortality and morbidity rates. Major risk factors for this disease include uncontrolled diabetes mellitus and immunosuppression that can also facilitate increases in mucormycosis infections. The extensive use of steroids to prevent the worsening of COVID-19 can lead to black fungus infection. Generally, antifungal agents dedicated to medical applications must be biocompatible, non-toxic, easily soluble, efficient, and hypoallergenic. They should also provide long-term protection against fungal growth. COVID-19-related black fungus infection causes a severe increase in fatalities. Therefore, there is a strong need for the development of novel and efficient antimicrobial agents. Recently, nanoparticle-containing products available in the market have been used as antimicrobial agents to prevent bacterial growth, but little is known about their efficacy with respect to preventing fungal growth, especially black fungus. The present review focuses on the effect of various types of metal nanoparticles, specifically those containing silver, zinc oxide, gold, copper, titanium, magnetic, iron, and carbon, on the growth of various types of fungi. We particularly focused on how these nanoparticles can impact the growth of black fungus. We also discussed black fungus co-infection in the context of the global COVID-19 outbreak, and management and guidelines to help control COVID-19-associated black fungus infection. Finally, this review aimed to elucidate the relationship between COVID-19 and mucormycosis.

Epigenetic regulator Cfp1 safeguards male meiotic progression by regulating meiotic gene expression.

Meiosis occurs specifically in germ cells to produce sperm and oocytes that are competent for sexual reproduction. Multiple factors are required for successful meiotic entry, progression, and termination. Among them, trimethylation of histone H3 on lysine 4 (H3K4me3), a mark of active transcription, has been implicated in spermatogenesis by forming double-strand breaks (DSBs). However, the role of H3K4me in transcriptional regulation during meiosis remains poorly understood. Here, we reveal that mouse CXXC finger protein 1 (Cfp1), a component of the H3K4 methyltransferase Setd1a/b, is dynamically expressed in differentiating male germ cells and safeguards meiosis by controlling gene expression. Genetic ablation of mouse CFP1 in male germ cells caused complete infertility with failure in prophase I of the 1st meiosis. Mechanistically, CFP1 binds to genes essential for spermatogenesis, and its loss leads to a reduction in H3K4me3 levels and gene expression. Importantly, CFP1 is highly enriched within the promoter/TSS of target genes to elevate H3K4me3 levels and gene expression at the pachytene stage of meiotic prophase I. The most enriched genes were associated with meiosis and homologous recombination during the differentiation of spermatocytes to round spermatids. Therefore, our study establishes a mechanistic link between CFP1-mediated transcriptional control and meiotic progression and might provide an unprecedented genetic basis for understanding human sterility.

The role of extracellular vesicles in animal reproduction and diseases.

Extracellular vesicles (EVs) are nanosized membrane-enclosed compartments that serve as messengers in cell-to-cell communication, both in normal physiology and in pathological conditions. EVs can transfer functional proteins and genetic information to alter the phenotype and function of recipient cells, which undergo different changes that positively affect their structural and functional integrity. Biological fluids are enriched with several subpopulations of EVs, including exosomes, microvesicles (MVs), and apoptotic bodies carrying several cargoes, such as lipids, proteins, and nucleic acids. EVs associated with the reproductive system are actively involved in the regulation of different physiological events, including gamete maturation, fertilization, and embryo and fetal development. EVs can influence follicle development, oocyte maturation, embryo production, and endometrial-conceptus communication. EVs loaded with cargoes are used to diagnose various diseases, including pregnancy disorders; however, these are dependent on the type of cell of origin and pathological characteristics. EV-derived microRNAs (miRNAs) and proteins in the placenta regulate inflammatory responses and trophoblast invasion through intercellular delivery in the placental microenvironment. This review presents evidence regarding the types of extracellular vesicles, and general aspects of isolation, purification, and characterization of EVs, particularly from various types of embryos. Further, we discuss EVs as mediators and messengers in reproductive biology, the effects of EVs on placentation and pregnancy disorders, the role of EVs in animal reproduction, in the male reproductive system, and mother and embryo cross-communication. In addition, we emphasize the role of microRNAs in embryo implantation and the role of EVs in reproductive and therapeutic medicine. Finally, we discuss the future perspectives of EVs in reproductive biology.

Diverse Effects of Exosomes on COVID-19: A Perspective of Progress From Transmission to Therapeutic Developments.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a new strain of coronavirus and the causative agent of the current global pandemic of coronavirus disease 2019 (COVID-19). There are currently no FDA-approved antiviral drugs for COVID-19 and there is an urgent need to develop treatment strategies that can effectively suppress SARS-CoV-2 infection. Numerous approaches have been researched so far, with one of them being the emerging exosome-based therapies. Exosomes are nano-sized, lipid bilayer-enclosed structures, share structural similarities with viruses secreted from all types of cells, including those lining the respiratory tract. Importantly, the interplay between exosomes and viruses could be potentially exploited for antiviral drug and vaccine development. Exosomes are produced by virus-infected cells and play crucial roles in mediating communication between infected and uninfected cells. SARS-CoV-2 modulates the production and composition of exosomes, and can exploit exosome formation, secretion, and release pathways to promote infection, transmission, and intercellular spread. Exosomes have been exploited for therapeutic benefits in patients afflicted with various diseases including COVID-19. Furthermore, the administration of exosomes loaded with immunomodulatory cargo in combination with antiviral drugs represents a novel intervention for the treatment of diseases such as COVID-19. In particular, exosomes derived from mesenchymal stem cells (MSCs) are used as cell-free therapeutic agents. Mesenchymal stem cell derived exosomes reduces the cytokine storm and reverse the inhibition of host anti-viral defenses associated with COVID-19 and also enhances mitochondrial function repair lung injuries. We discuss the role of exosomes in relation to transmission, infection, diagnosis, treatment, therapeutics, drug delivery, and vaccines, and present some future perspectives regarding their use for combating COVID-19.

Biogenesis, Membrane Trafficking, Functions, and Next Generation Nanotherapeutics Medicine of Extracellular Vesicles.

Extracellular vesicles (EVs) are a heterogeneous group of membrane-limited vesicles and multi-signal messengers loaded with biomolecules. Exosomes and ectosomes are two different types of EVs generated by all cell types. Their formation depends on local microdomains assembled in endocytic membranes for exosomes and in the plasma membrane for ectosomes. Further, EV release is a fundamental process required for intercellular communication in both normal physiology and pathological conditions to transmit/exchange bioactive molecules to recipient cells and the extracellular environment. The unique structure and composition of EVs enable them to serve as natural nanocarriers, and their physicochemical properties and biological functions can be used to develop next-generation nano and precision medicine. Knowledge of the cellular processes that govern EVs biology and membrane trafficking is essential for their clinical applications. However, in this rapidly expanding field, much remains unknown regarding EV origin, biogenesis, cargo sorting, and secretion, as well as EV-based theranostic platform generation. Hence, we present a comprehensive overview of the recent advances in biogenesis, membrane trafficking, and functions of EVs, highlighting the impact of nanoparticles and oxidative stress on EVs biogenesis and release and finally emphasizing the role of EVs as nanotherapeutic agents.

High-Quality Nucleic Acid Isolation from Hard-to-Lyse Bacterial Strains Using PMAP-36, a Broad-Spectrum Antimicrobial Peptide.

The efficiency of existing cell lysis methods to isolate nucleic acids from diverse bacteria varies depending on cell wall structures. This study tested a novel idea of using broad-spectrum antimicrobial peptides to improve the lytic efficiency of hard-to-lyse bacteria and characterized their differences. The lysis conditions of Staphylococcus aureus using recombinant porcine myeloid antimicrobial peptide 36 (PMAP-36), a broad-spectrum pig cathelicidin, was optimized, and RNA isolation was performed with cultured pellets of ten bacterial species using various membranolytic proteins. Additionally, three other antimicrobial peptides, protegrin-1 (PG-1), melittin, and nisin, were evaluated for their suitability as the membranolytic agents of bacteria. However, PMAP-36 use resulted in the most successful outcomes in RNA isolation from diverse bacterial species. The amount of total RNA obtained using PMAP-36 increased by ~2-fold compared to lysozyme in Salmonella typhimurium. Streptococci species were refractory to all lytic proteins tested, although the RNA yield from PMAP-36 treatment was slightly higher than that from other methods. PMAP-36 use produced high-quality RNA, and reverse transcription PCR showed the efficient amplification of the 16S rRNA gene from all tested strains. Additionally, the results of genomic DNA isolation were similar to those of RNA isolation. Thus, our findings present an additional option for high quality and unbiased nucleic acid isolation from microbiomes or challenging bacterial strains.

Palladium Nanoparticle-Induced Oxidative Stress, Endoplasmic Reticulum Stress, Apoptosis, and Immunomodulation Enhance the Biogenesis and Release of Exosome in Human Leukemia Monocytic Cells (THP-1).

BACKGROUND: Exosomes are endosome-derived nano-sized vesicles that have emerged as important mediators of intercellular communication and play significant roles in various diseases. However, their applications are rigorously restricted by the limited secretion competence of cells. Therefore, strategies to enhance the production and functions of exosomes are warranted. Studies have shown that nanomaterials can significantly enhance the effects of cells and exosomes in intercellular communication; however, how palladium nanoparticles (PdNPs) enhance exosome release in human leukemia monocytic cells (THP-1) remains unclear. Therefore, this study aimed to address the effect of PdNPs on exosome biogenesis and release in THP-1 cells. METHODS: Exosomes were isolated by ultracentrifugation and ExoQuickTM and characterized by dynamic light scattering, nanoparticle tracking analysis system, scanning electron microscopy, transmission electron microscopy, EXOCETTM assay, and fluorescence polarization. The expression levels of exosome markers were analyzed via quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay. RESULTS: PdNP treatment enhanced the biogenesis and release of exosomes by inducing oxidative stress, endoplasmic reticulum stress, apoptosis, and immunomodulation. The exosomes were spherical in shape and had an average diameter of 50-80 nm. Exosome production was confirmed via total protein concentration, exosome counts, acetylcholinesterase activity, and neutral sphingomyelinase activity. The expression levels of TSG101, CD9, CD63, and CD81 were significantly higher in PdNP-treated cells than in control cells. Further, cytokine and chemokine levels were significantly higher in exosomes isolated from PdNP-treated THP-1 cells than in those isolated from control cells. THP-1 cells pre-treated with N-acetylcysteine or GW4869 showed significant decreases in PdNP-induced exosome biogenesis and release. CONCLUSION: To our knowledge, this is the first study showing that PdNPs stimulate exosome biogenesis and release and simultaneously increase the levels of cytokines and chemokines by modulating various physiological processes. Our findings suggest a reasonable approach to improve the production of exosomes for various therapeutic applications.

Role and Therapeutic Potential of Melatonin in Various Type of Cancers.

Cancer is a large group of diseases and the second leading cause of death worldwide. Lung, prostate, colorectal, stomach, and liver cancers are the most common types of cancer in men, whereas breast, colorectal, lung, cervical, and thyroid cancers are the most common among women. Presently, various treatment strategies, including surgical resection combined with chemotherapy, radiotherapy, nanotherapy, and immunotherapy, have been used as conventional treatments for patients with cancer. However, the clinical outcomes of advanced-stage disease remain relatively unfavorable owing to the emergence of chemoresistance, toxicity, and other undesired detrimental side effects. Therefore, new therapies to overcome these limitations are indispensable. Recently, there has been considerable evidence from experimental and clinical studies suggesting that melatonin can be used to prevent and treat cancer. Studies have confirmed that melatonin mitigates the pathogenesis of cancer by directly affecting carcinogenesis and indirectly disrupting the circadian cycle. Melatonin (MLT) is nontoxic and exhibits a range of beneficial effects against cancer via apoptotic, antiangiogenic, antiproliferative, and metastasis-inhibitory pathways. The combination of melatonin with conventional drugs improves the drug sensitivity of cancers, including solid and liquid tumors. In this manuscript, we will comprehensively review some of the cellular, animal, and human studies from the literature that provide evidence that melatonin has oncostatic and anticancer properties. Further, this comprehensive review compiles the available experimental and clinical data analyzing the history, epidemiology, risk factors, therapeutic effect, clinical significance, of melatonin alone or in combination with chemotherapeutic agents or radiotherapy, as well as the underlying molecular mechanisms of its anticancer effect against lung, breast, prostate, colorectal, skin, liver, cervical, and ovarian cancers. Nonetheless, in the interest of readership clarity and ease of reading, we have discussed the overall mechanism of the anticancer activity of melatonin against different types of cancer. We have ended this report with general conclusions and future perspectives.

Human endogenous retrovirus-enveloped baculoviral DNA vaccines against MERS-CoV and SARS-CoV2.

Here we report a recombinant baculoviral vector-based DNA vaccine system against Middle East respiratory syndrome coronavirus (MERS-CoV) and the severe acute respiratory syndrome coronavirus-2 (SARS-CoV2). A non-replicating recombinant baculovirus expressing the human endogenous retrovirus envelope gene (AcHERV) was constructed as a DNA vaccine vector for gene delivery into human cells. For MERS-CoV vaccine construction, DNA encoding MERS-CoV S-full, S1 subunit, or receptor-binding domain (RBD) was inserted into the genome of AcHERV. For COVID19 vaccine construction, DNA encoding SARS-CoV2 S-full or S1 or a MERS-CoV NTD domain-fused SARS-CoV2 RBD was inserted into the genome of AcHERV. AcHERV-DNA vaccines induce high humoral and cell-mediated immunity in animal models. In challenge tests, twice immunized AcHERV-MERS-S1 and AcHERV-COVID19-S showed complete protection against MERS-CoV and SARS-CoV2, respectively. Unlike AcHERV-MERS vaccines, AcHERV-COVID19-S provided the greatest protection against SARS-CoV2 challenge. These results support the feasibility of AcHERV-MERS or AcHERV-COVID19 vaccines in preventing pandemic spreads of viral infections.

Correction: Gurunathan, S. et al. Review of the Isolation, Characterization, Biological Function, and Multifarious Therapeutic Approaches of Exosomes. Cells 2019, 8, 307.

In the original review [...].

A Comprehensive Review on Factors Influences Biogenesis, Functions, Therapeutic and Clinical Implications of Exosomes.

Exosomes are nanoscale-sized membrane vesicles secreted by almost all cell types into the extracellular environment upon fusion of multivesicular bodies and plasma membrane. Biogenesis of exosomes is a protein quality control mechanism, and once released, exosomes transmit signals to other cells. The applications of exosomes have increased immensely in biomedical fields owing to their cell-specific cargos that facilitate intercellular communications with neighboring cells through the transfer of biologically active compounds. The diverse constituents of exosomes reflect their cell of origin and their detection in biological fluids represents a diagnostic marker for various diseases. Exosome research is expanding rapidly due to the potential for clinical application to therapeutics and diagnosis. However, several aspects of exosome biology remain elusive. To discover the use of exosomes in the biomedical applications, we must better understand the basic molecular mechanisms underlying their biogenesis and function. In this comprehensive review, we describe factors involved in exosomes biogenesis and the role of exosomes in intercellular signaling and cell-cell communications, immune responses, cellular homeostasis, autophagy, and infectious diseases. In addition, we discuss the role of exosomes as diagnostic markers, and their therapeutic and clinical implications. Furthermore, we addressed the challenges and outstanding developments in exosome research, and discuss future perspectives.

Platinum Nanoparticles Enhance Exosome Release in Human Lung Epithelial Adenocarcinoma Cancer Cells (A549): Oxidative Stress and the Ceramide Pathway are Key Players.

BACKGROUND: Several studies have demonstrated various molecular mechanisms involved in the biogenesis and release of exosomes. However, how external stimuli, such as platinum nanoparticles (PtNPs), induces the biogenesis and release of exosomes remains unclear. To address this, PtNPs were synthesized using lutein to examine their effect on the biogenesis and release of exosomes in human lung epithelial adenocarcinoma cancer cells (A549). METHODS: The size and concentration of isolated exosomes were characterized by dynamic light scattering (DLS) and nanoparticle tracking analysis system (NTA). Morphology and structure of exosomes were examined using scanning electron microscopy and transmission electron microscopy (TEM), respectively. Quantification of exosomes were analyzed by EXOCETTM assay and fluorescence polarization (FP). The expression of typical markers of exosomes were analyzed by quantitative reverse transcription-polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). RESULTS: A549 cells cultured with PtNPs enhance exosome secretion by altering various physiological processes. Interestingly, A549 cells treated with PtNPs increases total protein concentration, biogenesis and release of exosomes associated with PtNPs-induced oxidative stress. GW4869 inhibits PtNPs induced biogenesis and release of exosomes and also acetylcholinesterase (AChE), neutral sphingomyelinase activity (n-SMase), and exosome counts. A549 cells pre-treated with N-acetylcysteine (NAC) significantly inhibited PtNPs induced exosome biogenesis and release. These findings confirmed that PtNPs-induced exosome release was due to the induction of oxidative stress and the ceramide pathway. These factors enhanced exosome biogenesis and release and may be useful in understanding the mechanism of exosome formation, release, and function. CONCLUSION: PtNPs provide a promising agent to increase exosome production in A549 cells. These findings offer novel strategies for enhancing exosome release, which can be applied in the treatment and prevention of cancer. Importantly, this is the first study, to our knowledge, showing that PtNPs stimulate exosome biogenesis by inducing oxidative stress and the ceramide pathway.

Transgenic Mice Overexpressing PG1 Display Corneal Opacity and Severe Inflammation in the Eye.

Antimicrobial peptides (AMPs) are of interest as alternatives to antibiotics or immunomodulators. We generated and characterized the phenotypes of transgenic mice overexpressing protegrin 1 (PG1), a potent porcine cathelicidin. No obvious differences were observed between PG1 transgenic and wild-type mice in terms of growth, development, general behaviour, and the major immune cell population. However, PG1 transgenic mice intranasally infected with Staphylococcus aureus resulted in a reduction in microscopic pulmonary injury, improved clearance of bacteria, and lower proinflammatory cytokine secretion, compared to those of wild-type mice. On the other hand, approximately 25% of PG1 transgenic mice (n = 54/215) showed corneal opacity and developed inflammation in the eye, resulting ultimately in phthisis bulbi. Immunohistochemical analyses revealed that PG1 and its activator, neutrophil elastase, localized to the basal cells of the cornea and glands in eyelids, respectively. In addition, apoptosis indicated by a Terminal deoxynucleotidyl transferase dUTP nick end labelling (TUNEL)-positive signal was detected from flat cells of the cornea. Our study suggests that the expression regulation or localization of AMPs such as PG1 is important to prevent their adverse effects. However, our results also showed that the cytotoxic effects of PG1 on cells could be tolerated in animals, except for the eyes.

Cathelicidin ΔPb-CATH4 derived from Python bivittatus accelerates the healing of Staphylococcus aureus-infected wounds in mice.

The effects of ΔPb-CATH4, a cathelicidin derived from Python bivittatus, were evaluated against Staphylococcus aureus-infected wounds in mice. These effects were comparable to those of classical antibiotics. ΔPb-CATH4 was resistant to bacterial protease but not to porcine trypsin. A reduction in the level of inflammatory cytokines and an increase in the migration of immune cells was observed in vitro. Thus, ΔPb-CATH4 can promote wound healing by controlling infections including those caused by multidrug-resistant bacteria via its immunomodulatory effects.