Nanoquercetin and Extracellular Vesicles as Potential Anticancer Therapeutics in Hepatocellular Carcinoma.

Despite world-class sophisticated technologies, robotics, artificial intelligence, and machine learning approaches, cancer-associated mortalities and morbidities have shown continuous increments posing a healthcare burden. Drug-based interventions were associated with systemic toxicities and several limitations. Natural bioactive compounds derived nanoformulations, especially nanoquercetin (nQ), are alternative options to overcome drug-associated limitations. Moreover, the EVs-based cargo targeted delivery of nQ can have enormous potential in treating hepatocellular carcinoma (HCC). EVs-based nQ delivery synergistically regulates and dysregulates several pathways, including NF-κB, p53, JAK/STAT, MAPK, Wnt/ß-catenin, and PI3K/AKT, along with PBX3/ERK1/2/CDK2, and miRNAs intonation. Furthermore, discoveries on possible checkpoints of anticancer signaling pathways were studied, which might lead to the development of modified EVs infused with nQ for the development of innovative treatments for HCC. In this work, we abridged the control of such signaling systems using a synergetic strategy with EVs and nQ. The governing roles of extracellular vesicles controlling the expression of miRNAs were investigated, particularly in relation to HCC.

Extracellular vesicles in neurodegenerative diseases: A systematic review.

Introduction: Extracellular vesicles (EVs) are known to have a significant role in the central nervous system (CNS) and neurodegenerative disease. Methods: PubMed, Scopus, ISI Web of Science, EMBASE, and Google Scholar were used to identify published articles about EV modifications (2012 to Feb 2022). Results: In total, 1,435 published papers were identified among the searched articles, with 1,128 non-duplicate publications being identified. Following the screening of titles and abstracts, 214 publications were excluded; following the full-text screening of 93 published articles, another 33 publications were excluded. The remaining 60 studies were considered. The kappa statistic of 0.868 indicated that the raters were highly reliable. Furthermore, the inter-reliability and intra-reliability coefficients were found to be 0.931 and 0.908, respectively, indicating strong reliability and consistency between the eligible studies identified by the raters. A total of 27 relevant studies demonstrated the role of EVs as therapeutic and diagnostic biomarkers in neurodegenerative diseases. Of note, 19 and 14 studies, respectively, found EVs to be pioneering in diagnostic and therapeutic roles. Discussion: EVs play an important role in the central nervous system (CNS), aiding in cell-to-cell communication and serving as a diagnostic marker and therapeutic target in a variety of neurodegenerative diseases. EVs are the home of several proteins [including-synuclein (-syn) and tau proteins], lipids, and genetic materials such as DNA and RNA. The presence of novel miRNAs in EVs suggests biomarkers for the diagnosis and screening of neurodegenerative disorders. Furthermore, EVs play an important role in the pathogenesis of such disorders. This systematic review discussed the current state of EVs' role in neurological diseases, as well as some preclinical studies on the therapeutic and diagnostic potential of EVs.

Engineered Extracellular Vesicles in Treatment of Type 1 Diabetes Mellitus: A Prospective Review.

Insulin replacement is an available treatment for autoimmune type 1 diabetes mellitus (T1DM). There are multiple limitations in the treatment of autoimmune diseases such as T1DM by immunosuppression using drugs and chemicals. The advent of extracellular vesicle (EV)-based therapies for the treatment of various diseases has attracted much attention to the field of bio-nanomedicine. Tolerogenic nanoparticles can induce immune tolerance, especially in autoimmune diseases. EVs can deliver cargo to specific cells without restrictions. Accordingly, EVs can be used to deliver tolerogenic nanoparticles, including iron oxide-peptide-major histocompatibility complex, polyethylene glycol-silver-2-(1'H-indole-3'-carbonyl)-thiazole-4-carboxylic acid methyl ester, and carboxylated poly (lactic-co-glycolic acid) nanoparticles coupled with or encapsulating an antigen, to effectively treat autoimmune T1DM. The present work highlights the advances in exosome-based delivery of tolerogenic nanoparticles for the treatment of autoimmune T1DM.

Protective role of engineered extracellular vesicles loaded quercetin nanoparticles as anti-viral therapy against SARS-CoV-2 infection: A prospective review.

Quercetin (QCT) is a naturally occurring phenolic flavonoid compound with inbuilt characteristics of antioxidant, anti-inflammatory, and immune protection. Several recent studies have shown that QCT and QCTits nanoparticles have therapeutic potential against severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. Novel therapeutics also include the implication of extracellular vesicles (EVs) to protect from SARS-CoV-2 viral infection. This article highlighted the therapeutic/prophylactic potential of engineered EVs loaded with QCT against SARS-CoV-2 infection. Several biotechnological engineering approaches are available to deliver EVs loaded with QCT nanoparticles. Among these biotechnological advances, a specific approach with significantly higher efficiency and yield has to be opted to fabricate such drug delivery of nano molecules, especially to combat SARS-CoV-2 infection. The current treatment regime protects the human body from virus infection but has some limitations including drugs and long-term steroid side effects. However, the vaccine strategy is somehow effective in inhibiting the spread of coronavirus disease-19 (COVID-19) infection. Moreover, the proposed exosomal therapy met the current need to repair the damaged tissue along with inhibition of COVID-19-associated complications at the tissue level. These scientific findings expand the possibilities and predictability of developing a novel and cost-effective therapeutic approach that combines the dual molecule, EVs and QCT nanoparticles, to treat SARS-CoV-2 infection. Therefore, the most suitable engineering method to fabricate such a drug delivery system should be better understood before developing novel therapeutics for clinical purposes.

New horizons of biomaterials in treatment of nerve damage in diabetes mellitus: A translational prospective review.

Background: Peripheral nerve injury is a serious concern that leads to loss of neuronal communication that impairs the quality of life and, in adverse conditions, causes permanent disability. The limited availability of autografts with associated demerits shifts the paradigm of researchers to use biomaterials as an alternative treatment approach to recover nerve damage. Purpose: The purpose of this study is to explore the role of biomaterials in translational treatment approaches in diabetic neuropathy. Study design: The present study is a prospective review study. Methods: Published literature on the role of biomaterials in therapeutics was searched for. Results: Biomaterials can be implemented with desired characteristics to overcome the problem of nerve regeneration. Biomaterials can be further exploited in the treatment of nerve damage especially associated with PDN. These can be modified, customized, and engineered as scaffolds with the potential of mimicking the extracellular matrix of nerve tissue along with axonal regeneration. Due to their beneficial biological deeds, they can expedite tissue repair and serve as carriers of cellular and pharmacological treatments. Therefore, the emerging research area of biomaterials-mediated treatment of nerve damage provides opportunities to explore them as translational biomedical treatment approaches. Conclusions: Pre-clinical and clinical trials in this direction are needed to establish the effective role of several biomaterials in the treatment of other human diseases.

A systematic review on the modifications of extracellular vesicles: a revolutionized tool of nano-biotechnology.

BACKGROUND: Tailoring extracellular vesicles (EVs) can bequeath them with diverse functions and efficient performance in nano-biotechnology. Engineering and modification of EVs improves the targeted drug delivery efficiency. Here, we performed systematic review of various methods for EVs modifications. METHODS: PubMed, Scopus, ISI Web of Science, EMBASE, and Google Scholar were searched for available articles on EVs modifications (up to March 2021). In total, 1208 articles were identified and assessed, and then only 36 articles were found eligible and included. RESULTS: Six studies demonstrate the application of click chemistry, seven studies used co-incubation, two studies used chemical transfection, four studies implicated electroporation and sonication approach for modification of EVs. Moreover, two studies utilized microfluidics as suitable approach for loading cargo into EVs, while eight studies showed freeze-thaw method as feasible for these biological nanoparticles. CONCLUSION: Freeze-thaw approach is found to be convenient and popular among researchers for performing modifications in EVs for the purpose of targeted drug delivery loading. Clinical-grade EVs production with good clinical practices (GCPs) is challenging in the current scenario. More studies are needed to determine the best suitable approach for cargo loading of EVs that may be exploited for research and therapeutic use.

Newer Horizon of Mesenchymal Stem Cell-Based Therapy in the Management of SARS-CoV-2-Associated Mucormycosis: A Safe Hope for Future Medicine.

SARS-CoV-2-infected patients are reported to show immunocompromised behavior that gives rise to a wide variety of complications due to impaired innate immune response, cytokine storm, and thrombo-inflammation. Prolonged use of steroids, diabetes mellitus, and diabetic ketoacidosis (DKA) are some of the factors responsible for the growth of Mucorales in such immunocompromised patients and, thus, can lead to a life-threatening condition referred to as mucormycosis. Therefore, an early diagnosis and cell-based management cosis is the need of the hour to help affected patients overcome this severe condition. In addition, extended exposure to antifungal drugs/therapeutics is found to initiate hormonal and neurological complications. More recently, mesenchymal stem cells (MSCs) have been used to exhibit immunomodulatory function and proven to be beneficial in a clinical cell-based regenerative approach. The immunomodulation ability of MSCs in mucormycosis patient boosts the immunity by the release of chemotactic proteins. MSC-based therapy in mucormycosis along with the combination of short-term antifungal drugs can be utilized as a prospective approach for mucormycosis treatment with promising outcomes. However, preclinical and in mucormyIn mucormycosis, the hyphae of clinical trials are needed to establish the precise mechanism of MSCs in mucormycosis treatment.

Mesenchymal Stromal Cell-Derived Tailored Exosomes Treat Bacteria-Associated Diabetes Foot Ulcers: A Customized Approach From Bench to Bed.

Exosomes are nano-vesicles of endosomal origin inherited with characteristics of drug delivery and cargo loading. Exosomes offer a diverse range of opportunities that can be exploited in the treatment of various diseases post-functionalization. This membrane engineering is recently being used in the management of bacteria-associated diabetic foot ulcers (DFUs). Diabetes mellitus (DM) is among the most crippling disease of society with a large share of its imposing economic burden. DM in a chronic state is associated with the development of micro- and macrovascular complications. DFU is among the diabetic microvascular complications with the consequent occurrence of diabetic peripheral neuropathy. Mesenchymal stromal cell (MSC)-derived exosomes post-tailoring hold promise to accelerate the diabetic wound repair in DFU associated with bacterial inhabitant. These exosomes promote the antibacterial properties with regenerative activity by loading bioactive molecules like growth factors, nucleic acids, and proteins, and non-bioactive substances like antibiotics. Functionalization of MSC-derived exosomes is mediated by various physical, chemical, and biological processes that effectively load the desired cargo into the exosomes for targeted delivery at specific bacterial DFUs and wound. The present study focused on the application of the cargo-loaded exosomes in the treatment of DFU and also emphasizes the different approaches for loading the desired cargo/drug inside exosomes. However, more studies and clinical trials are needed in the domain to explore this membrane engineering.

Mesenchymal Stem Cell-Derived Exosomes Exhibit Promising Potential for Treating SARS-CoV-2-Infected Patients.

The novel coronavirus severe acute respiratory syndrome-CoV-2 (SARS-CoV-2) is responsible for COVID-19 infection. The COVID-19 pandemic represents one of the worst global threats in the 21st century since World War II. This pandemic has led to a worldwide economic recession and crisis due to lockdown. Biomedical researchers, pharmaceutical companies, and premier institutes throughout the world are claiming that new clinical trials are in progress. During the severe phase of this disease, mechanical ventilators are used to assist in the management of outcomes; however, their use can lead to the development of pneumonia. In this context, mesenchymal stem cell (MSC)-derived exosomes can serve as an immunomodulation treatment for COVID-19 patients. Exosomes possess anti-inflammatory, pro-angiogenic, and immunomodulatory properties that can be explored in an effort to improve the outcomes of SARS-CoV-2-infected patients. Currently, only one ongoing clinical trial (NCT04276987) is specifically exploring the use of MSC-derived exosomes as a therapy to treat SARS-CoV-2-associated pneumonia. The purpose of this review is to provide insights of using exosomes derived from mesenchymal stem cells in management of the co-morbidities associated with SARS-CoV-2-infected persons in direction of improving their health outcome. There is limited knowledge of using exosomes in SARS-CoV-2; the clinicians and researchers should exploit exosomes as therapeutic regime.

CRISPR/Cas9 Edited sRAGE-MSCs Protect Neuronal Death in Parkinson’s Disease Model.

BACKGROUND AND OBJECTIVES: Parkinson’s disease (PD) is a fatal and progressive degenerative disease of the nervous system. Until recently, its promising treatment and underlying mechanisms for neuronal death are poorly understood. This study was investigated to identify the molecular mechanism of neuronal death in the substantia nigra and corpus striatum of PD. METHODS: The soluble RAGE (sRAGE) secreting Umbilical Cord Blood-derived Mesenchymal Stem Cell (UCB-MSC) was generated by gene editing method using clustered regularly interspaced short palindromic repeats/CRISPR associated protein 9 (CRISPR/Cas9). These cells were transplanted into Corpus Striatum of rotenone-induced PD animal models then behavioral test, morphological analysis, and immunohistochemical experiments were performed to determine the neuronal cell death and recovery of movement. RESULTS: The neuronal cell death in Corpus Striatum and Substantia Nigra was dramatically reduced and the movement was improved after sRAGE secreting UCB-MSC treatment in PD mice by inhibition of RAGE in neuronal cells. CONCLUSIONS: We suggest that sRAGE secreting UCB-MSC based therapeutic approach could be a potential treatment strategy for neurodegenerative disease including PD.

Advanced glycation end-products produced systemically and by macrophages: A common contributor to inflammation and degenerative diseases.

Advanced glycation end products (AGEs) and their receptor have been implicated in the progressions of many intractable diseases, such as diabetes and atherosclerosis, and are also critical for pathologic changes in chronic degenerative diseases, such as Alzheimer's disease, Parkinson's disease, and alcoholic brain damage. Recently activated macrophages were found to be a source of AGEs, and the most abundant form of AGEs, AGE-albumin excreted by macrophages has been implicated in these diseases and to act through common pathways. AGEs inhibition has been shown to prevent the pathogenesis of AGEs-related diseases in human, and therapeutic advances have resulted in several agents that prevent their adverse effects. Recently, anti-inflammatory molecules that inhibit AGEs have been shown to be good candidates for ameliorating diabetic complications as well as degenerative diseases. This review was undertaken to present, discuss, and clarify current understanding regarding AGEs formation in association with macrophages, different diseases, therapeutic and diagnostic strategy and links with RAGE inhibition.

Intravenously Infused F3.Olig2 Improves Memory Deficits via Restoring Myelination in the Aged Hippocampus Following Experimental Ischemic Stroke.

Oligodendrocytes play a crucial role in creating the myelin sheath that is an important component in neural transmission. In an animal model of transient cerebral ischemia, application of oligodendrocyte progenitor cells (OPCs) has not yet been reported. In this study, the effects of F3.Olig2 transplantation on memory and cognitive dysfunction were investigated in the aged gerbil in which ischemic stroke was induced. To investigate the possible mechanisms underlying repair, changes in the expression of myelin basic protein (MBP), oligodendrocyte-specific protein (OSP), and brain-derived neurotrophic factor (BDNF) were examined. Experimental ischemic stroke was induced by occlusion of bilateral common carotid arteries in aged gerbils. Gerbils (n=31 per group) were randomly divided into three groups: (1) vehicle sham group, (2) vehicle ischemia group, and (3) F3.Olig2 ischemia group. After 1, 3, and 7 days of ischemiareperfusion (I-R), saline or F3.Olig2 cells (1106 cells in 100 l) were injected into the gerbils intravenously. The gerbils were sacrificed 10 days after I-R for identification of grafted F3.Olig2 cells, and 15 and 30 days after I-R for tissue analysis after conducting passive avoidance and novel object recognition test. Injected F3.Olig2 cells and MBP, OSP, and BDNF were detected by specific antibodies using immunohistochemistry and/or Western blots. Memory and cognition were significantly increased in the F3.Olig2 ischemia group compared with the vehicle ischemia group. In the F3.Olig2 ischemia group, the neurons were not protected from ischemic damage; however, MBP, OSP, and BDNF expressions were significantly increased. Our results show that injection of F3.Olig2 cells significantly improved impaired memory and cognition, which might be related to increased MBP expression via increasing OSP and BDNF expression in the aged gerbil hippocampus following transient cerebral ischemia.

3D texture analysis in renal cell carcinoma tissue image grading.

One of the most significant processes in cancer cell and tissue image analysis is the efficient extraction of features for grading purposes. This research applied two types of three-dimensional texture analysis methods to the extraction of feature values from renal cell carcinoma tissue images, and then evaluated the validity of the methods statistically through grade classification. First, we used a confocal laser scanning microscope to obtain image slices of four grades of renal cell carcinoma, which were then reconstructed into 3D volumes. Next, we extracted quantitative values using a 3D gray level cooccurrence matrix (GLCM) and a 3D wavelet based on two types of basis functions. To evaluate their validity, we predefined 6 different statistical classifiers and applied these to the extracted feature sets. In the grade classification results, 3D Haar wavelet texture features combined with principal component analysis showed the best discrimination results. Classification using 3D wavelet texture features was significantly better than 3D GLCM, suggesting that the former has potential for use in a computer-based grading system.

Changes of calcium binding proteins, c-Fos and COX in hippocampal formation and cerebellum of Niemann-Pick, type C mouse.

Niemann-Pick disease, type C (NPC) is an intractable disease that is accompanied by ataxia, dystonia, neurodegeneration, and dementia due to an NPC gene defect. Disruption of calcium homeostasis in neurons is important in patients with NPC. Thus, we used immunohistochemistry to assess the expression levels of calcium binding proteins (calbindin D28K, parvalbumin, and calretinin), c-Fos and cyclooxygenase-1,2 (COX-1,2) in the hippocampal formation and cerebellum of 4 and 8 week old NPC+/+, NPC+/-, and NPC-/- mice. General expression of these proteins decreased in the hippocampus and cerebellum of NPC-/- compared to that in both young and adult NPC+/+ or NPC+/- mice. Parvalbumin, COX-1,2 or c-Fos-immunoreactive neurons were widely detected in the CA1, CA3, and DG of the hippocampus, but the immunoreactivities were decreased sharply in all areas of hippocampus of NPC-/- compared to NPC+/+ and NPC+/- mice. Taken together, reduction of these proteins may be one of the strong phenotypes related to the neuronal degeneration in NPC-/- mice.

Caffeine-induced endothelial cell death and the inhibition of angiogenesis.

Numerous studies have shown that adenosine or adenosine agonists can stimulate angiogenesis. However, the effect of caffeine (a known adenosine receptor antagonist) on angiogenesis has not been previously studied. Accordingly, this study was undertaken to examine the effect of caffeine on angiogenesis and to clarify the mechanism involved. Chick chorioallantoic membrane assays were used to investigate the effect of caffeine on angiogenesis and proliferation assays using human umbilical vein endothelial cells (HUVECs), were used to study its effects on specific aspects of angiogenesis. The expressions of caspase-3 and Bcl-2 were examined by western blotting, immunofluorescence staining was used to identify HUVEC morphological changes, and fluorescence activated cell sorting (FACS) and DAPI staining were used to detect HUVEC apoptosis. Caffeine was found to inhibit blood vessel formation dose-dependently and to inhibit the proliferation of HUVECs time- and dose-dependently. FACS analysis and DAPI staining showed that inhibitory effect of caffeine on HUVEC proliferation was the result of apoptosis and the up-regulation of thrombospondin-1 (TSP-1). Furthermore, TSP-1 levels were down-regulated by NECA but were unaffected by CGS21680, indicating that caffeine regulated TSP-1 expression via adenosine A2B receptor. In addition, caffeine up-regulated caspase-3 and down-regulated Bcl-2 at the protein level. These results suggest that the inhibitory effect of caffeine on angiogenesis is associated, at least in part, with its induction of endothelial cell apoptosis, probably mediated by a caspase-3 dependent mechanism.

Quantitative proteomic analysis reveals that lipopolysaccharide induces mitogen-activated protein kinase-dependent activation in human microglial cells.

Microglial cells act as the first and main form of active immune defense in the central nervous system related to inflammation and neurodegenerative disease. Lipopolysaccharide (LPS) induces many genes encoding inflammatory mediators, including cytokines such as tumor necrosis factor-α, interleukin-1ß, (IL-1ß), and IL-6, chemokines, and prostaglandins in microglial cells. Quantitative proteomics methods with isobaric chemical labeling using tandem mass tags and 2D-nano LC-ESI-MS/MS were used to systematically analyze proteomic changes in microglia responding to LPS stimulation. As a result, we found that the expression level of 21 proteins in human microglial cells changed after activation. Among those, one of the strong mitogen-activated protein kinase (MAPK) regulator proteins, CMPK1 was highly upregulated after LPS stimulation in human microglial cells. We detected and validated upregulation of MAPK including ERK1/2, p38, and SAPK/JNK by immunohistochemistry and Western blotting. NFκB, strong transcription factor of CMPK1, was translocated to the nucleus from the cytosol by high contents screening after LPS stimulation. Taken together, we conclude that MAPK signaling plays an important role in LPS-induced human microglial activation related to inflammatory response.

Activated microglial cells synthesize and secrete AGE-albumin.

A holy grail of curing neurodegenerative diseases is to identify the main causes and mechanisms underlying neuronal death. Many studies have sought to identify these targets in a wide variety of ways, but a more important task is to identify critical molecular targets and their origins. Potential molecular targets include advanced glycation end products (AGEs) that can promote neuronal cell death, thereby contributing to neurodegenerative disorders such as Alzheimer disease or Parkinson disease. In this study, we showed that AGE-albumin (glycated albumin) is synthesized in microglial cells and secreted in the human brain. Our results provide new insight into which microglial cells can promote the receptor for AGE-mediated neuronal cell death, eventually leading to neurodegenerative diseases.

Recovery of CNS pathway innervating the sciatic nerve following transplantation of human neural stem cells in rat spinal cord injury.

Stem cell research has been attained a greater attention in most fields of medicine due to its potential for many incurable diseases through replacing or helping the regeneration of damaged cells or tissues. Here, we demonstrated the functional recovery and structural connection of the central nervous system pathway innervating the sciatic nerve after total transection of the spinal cord followed by the transplantation of human neural stem cells (hNSC) in the injured rat spinal cord site. The limb function of hNSC-treated group recovered dramatically compared with that in the sham group by Basso-Beattie-Bresnahan (BBB) scores. Transplanted hNSC differentiated into astrocytes and neurons in the injured site. In addition, immunohistochemistry for growth-associated protein 43 showed axonal regeneration in the injured spinal cord site. The pseudorabies viral-Ba (PRV-Ba) tracing method revealed that transplanted hNSC and their differentiated neurons showed positive labeling after sciatic nerve injection. In addition, the PRV-Ba labeling was also observed in several nuclei in the brain innervating the sciatic nerve. This result implies that the rat CNS motor pathway could be reconstructed by hNSC transplantation, and it may contribute to the functional recovery of the limb.

Alteration of the CNS pathway to the hippocampus in a mouse model of Niemann-Pick, type C disease.

Niemann-Pick type C disease (NPC) is an autosomal recessive disorder that results in premature death due to progressive neurodegeneration including dementia. To understand neuronal pathways connecting to the hippocampus, retrograde transneuronal labeling method with Bartha strain of pseudorabies virus (PRV) was employed in 40 NPC+/+, NPC+/- and NPC-/- mice. Immunohistochemistry using polyclonal antibody against PRV and streological counting were used. The number of neurons and synapse in CA2&3 regions of the hippocampus decreased dramatically in the NPC-/- mouse compared to the NPC+/+ or +/- mouse. The number of PRV positive cell was significantly decreased in several regions including the entorhinal and piriform cortex in the NPC-/- mouse. More severely, lateral septal dorsal nucleus, dorsal entorhinal cortex and medial geniculate body showed no positive labeling in the NPC-/- mouse. However, the hippocampus, medial septal and supramammilary nuclei showed increased immunoreactivity in the NPC-/- mouse. Our data suggest that the synaptic loss and discontinuity of the CNS hippocampal pathway may contribute to understanding the mechanism of symptoms and functional disabilities such as memory and learning disturbance in NPC patients.