an-enrichment-method-for-small-extracellular-vesicles-derived-from

An Enrichment Method for Small Extracellular Vesicles Derived from Liver Cancer Tissue

Small extracellular vesicles (sEVs) derived from tissue can reflect the functional status of the source cells and the characteristics of the tissue's ...

Gannan Medical University

First Affiliated Hospital of Gannan Medical University

The detection and analysis of methylene tetrahydrofolate reductase (MTHFR) C677T single nucleotide polymorphism (SNP) from blood samples is time-consuming and costly. We aimed to establish a method to detect these SNPs by direct whole blood PCR and without DNA extraction.

Single probe PCR melting curve analysis MTHFR C677T SNP sites.

Aging is a universal phenomenon in all biological organisms, defined by the loss of reproductive capacity and a progressive decline in fitness. In humans, aging is further associated with an increased incidence of disease conditions. The current aging population has become a primary public burden of the 21 century. Therefore, to delay the aging process and maintain fitness in the aging population, the discovery of novel anti-aging drugs remains an urgent need. In recent years, metformin, a widely used hypoglycemic drug, has attracted growing attention in the field of anti-aging research. Reportedly, numerous studies have indicated that metformin regulates aging-related pathways, possibly delaying the aging process by modulating these pathways. The elucidation of these anti-aging effects may provide insights into the age-retarding potential of metformin. The present review focuses on the predominant molecular mechanisms associated with aging, as well as the anti-aging effects of metformin.

Metformin: A Potential Candidate for Targeting Aging Mechanisms.

Small extracellular vesicles are membrane-bound vesicles secreted into extracellular spaces by virtually all types of cells. These carry a large number of membrane proteins on their surface that are incorporated during their biogenesis in cells. The composition of the membrane proteins hence bears the signature of the cells from which they originate. Recent studies have suggested that the proteins on these small extracellular vesicles can serve as biomarkers and target proteins for the diagnosis and treatment of diseases. This article classifies small extracellular vesicle membrane proteins and summarizes their pathophysiological functions in the diagnosis and treatment of diseases.

Advances in Biological Function and Clinical Application of Small Extracellular Vesicle Membrane Proteins.

A potential use of small extracellular vesicles (sEVs) for diagnostic and therapeutic purposes has recently generated a great interest. sEVs, when purified directly from various tissues with proper procedures, can reflect the physiological and pathological state of the organism. However, the quality of sEV is affected by many factors during isolation, including separation of sEV from cell and tissues debris, the use of enzymes for tissue digestion, and the storage state of tissues. In the present study, we established an assay for the isolation and purification of liver cancer tissues-derived sEVs (tdsEVs) and cultured explants-derived sEVs (cedsEVs) by comparing the quality of sEVs derived from different concentration of digestion enzyme and incubation time. The nano-flow cytometry (NanoFCM) showed that the isolated tdsEVs by our method are purer than those obtained from differential ultracentrifugation. Our study thus establishes a simple and effective approach for isolation of high-quality sEVs that can be used for analysis of their constituents.

Comparison of the Variability of Small Extracellular Vesicles Derived from Human Liver Cancer Tissues and Cultured from the Tissue Explants Based on a Simple Enrichment Method.

Current human papillomavirus (HPV) detection methods require complex instruments, skilled staff and have a high cost. Therefore, novel testing approaches are needed which are easy to implement, highly sensitive, and low cost. Loop-mediated isothermal amplification (LAMP) is an isothermal amplification technique. In this study, according to the conditions in China, a novel LAMP method for detecting seven high-risk HPV subtypes (16, 18, 33, 39, 45, 52, and 58) was designed and evaluated. The DNA from plasmid and cervical specimens was extracted using Chelex 100 and measured by qPCR and LAMP assay. LAMP products were observed under ultraviolet light. HPV sequences were successfully amplified and a plateau time of 19-75 min was maintained. The concentration of positive reactions ranged between 20 copies/μL and 200000 copies/μL. Additionally, there was no cross-reactivity between HPV16, 18, 33, 39, 45, 52, 58, 31, 35, 45, 51, 56, 59, 66, or 68. For clinical samples, the LAMP assay had high sensitivity and specificity for HPV16, 18, 33, 39, 45, 52, and 58. However, 5% (72/1447) of the samples tested yielded false-positive results. In conclusion, the novel LAMP assay for HPV16, 18, 33, 39, 45, 52, and 58 has high sensitivity and specificity, a low cost, and is simple and rapid to perform. The LAMP assay can improve HPV detection in resource-limited settings, especially in primary care hospitals and rural areas.

Rapid diagnosis of seven high-risk human papillomavirus subtypes by a novel loop-mediated isothermal amplification method.

Exosomes are tiny vesicles with a double membrane structure that cells produce. They range in diameter from 40 to 150 nm and may contain a variety of biomolecules including proteins and nucleic acids. Exosomes have low toxicity, low immunogenicity, and the ability to encapsulate a wide variety of substances, making them attractive drug delivery vehicles. MSCs secrete large amounts of exosomes and hence serve as an excellent source of exosomes. MSCs-derived exosomes have regenerative and tissue repair functions comparable to MSCs and can circumvent the risks of immune rejection and infection associated with MSC transplantation, indicating that they may be a viable alternative to MSCs' biological functions. In this review, we summarized the drug delivery methods and advantages of exosomes, as well as the advancement of MSC exosomes as drug carriers. The challenges and prospects of using exosomes as drug delivery vectors are presented.

Advances in Mesenchymal Stem Cell-Derived Exosomes as Drug Delivery Vehicles.

Exosomes are a heterogeneous subset of extracellular vesicles (EVs) that biogenesis from endosomes. Besides, exosomes contain a variety of molecular cargoes including proteins, lipids and nucleic acids, which play a key role in the mechanism of exosome formation. Meanwhile, exosomes are involved with physiological and pathological conditions. The molecular profile of exosomes reflects the type and pathophysiological status of the originating cells so could potentially be exploited for diagnostic of cancer. This review aims to describe important molecular cargoes involved in exosome biogenesis. In addition, we highlight exogenous factors, especially autophagy, hypoxia and pharmacology, that regulate the release of exosomes and their corresponding cargoes. Particularly, we also emphasize exosome molecular cargoes as potential biomarkers in liquid biopsy for diagnosis of cancer.

Current perspectives on clinical use of exosomes as novel biomarkers for cancer diagnosis.

Breast cancer (BC) is the most common malignancy and the leading cause of cancer-related deaths in women worldwide. Currently, patients' survival remains a challenge in BC due to the lack of effective targeted therapies and the difficult condition of patients with higher aggressiveness, metastasis and drug resistance. Small extracellular vesicles (sEVs), which are nanoscale vesicles with lipid bilayer envelopes released by various cell types in physiological and pathological conditions, play an important role in biological information transfer between cells. There is growing evidence that BC cell-derived sEVs may contribute to the establishment of a favorable microenvironment that supports cancer cells proliferation, invasion and metastasis. Moreover, sEVs provide a versatile platform not only for the diagnosis but also as a delivery vehicle for drugs. This review provides an overview of current new developments regarding the involvement of sEVs in BC pathogenesis, including tumor proliferation, invasion, metastasis, immune evasion, and drug resistance. In addition, sEVs act as messenger carriers carrying a variety of biomolecules such as proteins, nucleic acids, lipids and metabolites, making them as potential liquid biopsy biomarkers for BC diagnosis and prognosis. We also described the clinical applications of BC derived sEVs associated MiRs in the diagnosis and treatment of BC along with ongoing clinical trials which will assist future scientific endeavors in a more organized direction.

The role and application of small extracellular vesicles in breast cancer.

Early tumor diagnosis is crucial for its treatment and reduction of death, with effective tumor biomarkers being important tools. Extracellular vesicles (EVs) are small vesicles secreted by cells with various biomolecules, including proteins, nucleic acids, and lipids. They harbor a double membrane structure. Previous studies on EVs in cancer diagnosis and therapy focused on miRNAs. Nonetheless, EVs contain proteins that represent physiological and pathological state of their parental cells. EVs proteins can reflect the pathological state of some diseases, which provides a basis for diagnosis and treatment. This study describes the role of EVs in cancer and summarizes the use of EVs proteins as diagnostic markers in different cancer types. Specifically, we discuss the potential and shortcomings of EVs as tumor biomarkers.

Application of extracellular vesicles proteins in cancer diagnosis.

Colorectal cancer (CRC) is a malignancy that seriously threatens human health, and metastasis from CRC is a major cause of death and poor prognosis for patients. Studying the potential mechanisms of small extracellular vesicles (sEVs) in tumor development may provide new options for early and effective diagnosis and treatment of CRC metastasis. In this review, we systematically describe how sEVs mediate epithelial mesenchymal transition (EMT), reconfigure the tumor microenvironment (TME), modulate the immune system, and alter vascular permeability and angiogenesis to promote CRC metastasis. We also discuss the current difficulties in studying sEVs and propose new ideas.

Article	Year
Single probe PCR melting curve analysis MTHFR C677T SNP sites. Analytical biochemistry\| PubMed ID: 33450284	2021
Metformin: A Potential Candidate for Targeting Aging Mechanisms. Aging and disease\| PubMed ID: 33815878	2021
Advances in Biological Function and Clinical Application of Small Extracellular Vesicle Membrane Proteins. Frontiers in oncology\| PubMed ID: 34094979	2021
Comparison of the Variability of Small Extracellular Vesicles Derived from Human Liver Cancer Tissues and Cultured from the Tissue Explants Based on a Simple Enrichment Method. Stem cell reviews and reports\| PubMed ID: 34550537	2022
Rapid diagnosis of seven high-risk human papillomavirus subtypes by a novel loop-mediated isothermal amplification method. Molecular and cellular probes\| PubMed ID: 34875349	2022
Advances in Mesenchymal Stem Cell-Derived Exosomes as Drug Delivery Vehicles. Frontiers in bioengineering and biotechnology\| PubMed ID: 35186913	2021
Current perspectives on clinical use of exosomes as novel biomarkers for cancer diagnosis. Frontiers in oncology\| PubMed ID: 36119470	2022
The role and application of small extracellular vesicles in breast cancer. Frontiers in oncology\| PubMed ID: 36185265	2022
Application of extracellular vesicles proteins in cancer diagnosis. Frontiers in cell and developmental biology\| PubMed ID: 36407096	2022
The Biological Effect of Small Extracellular Vesicles on Colorectal Cancer Metastasis. Cells\| PubMed ID: 36552835	2022

Defa Huang

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