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Exosome-mediated repair of spinal cord injury: a promising therapeutic strategy.

Spinal cord injury (SCI) is a catastrophic injury to the central nervous system (CNS) that can lead to sensory and motor dysfunction, which seriously affects patients' quality of life and imposes a major economic burden on society. The pathological process of SCI is divided into primary and secondary injury, and secondary injury is a cascade of amplified responses triggered by the primary injury. Due to the complexity of the pathological mechanisms of SCI, there is no clear and effective treatment strategy in clinical practice. Exosomes, which are extracellular vesicles of endoplasmic origin w

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… To date, ultracentrifugation is the most commonly used method of exosome isolation; however, the exosomes acquired by ultracentrifugation may contain other impurities, such as extracellular vesicles with diameters close to those of the exosomes [ 188 ] (Fig. 6 ). …

Role of Exosomal Non-Coding RNAs in Bone-Related Diseases.

Bone-related diseases seriously affect the lives of patients and carry a heavy economic burden on society. Treatment methods cannot meet the diverse clinical needs of affected patients. Exosomes participate in the occurrence and development of many diseases through intercellular communication, including bone-related diseases. Studies have shown that exosomes can take-up and "package" non-coding RNAs and "deliver" them to recipient cells, thereby regulating the function of recipient cells. The exosomal non-coding RNAs secreted by osteoblasts, osteoclasts, chondrocytes, and other cells are invol

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… Ultracentrifugation is used as the “gold standard” for separation and purification of exosomes, among which density gradient ultracentrifugation is the best method. The advantages are simple pretreatment, non-requirement of specialist knowledge, and economic affordability ( Li et al., 2017 ). …

Cinobufagin Suppresses Lipid Peroxidation and Inflammation in Osteoporotic Mice by Promoting the Delivery of miR-3102-5p by Macrophage-Derived Exosomes.

BackgroundCinobufagin, the primary active compound in toad venom, is commonly used for anti-tumor, anti-inflammatory, and analgesic purposes. However, its specific bone-protective effects remain uncertain. This research aims to ascertain the bone-protective properties of cinobufagin and investigate underlying mechanisms.MethodsMice were ovariectomized to establish an osteoporosis model, followed by intraperitoneal injections of cinobufagin and cinobufagin-treated RAW.264.7-derived exosomes for therapy. MicroCT, HE staining, and TRAP staining were employed to evaluate bone mass and therapeutic

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… Exosomes were then isolated by ultracentrifugation at 118,000×g for 70 minutes. The exosomes, labeled with DiO-green (C1038, Beyotime Biotechnology) or DID-red (c1039, Beyotime Biotechnology), were resuspended in PBS and underwent final centrifugation at 118,000×g for 70 minutes. …

m6A reader hnRNPA2B1 drives multiple myeloma osteolytic bone disease.

Rationale: Bone destruction is a hallmark of multiple myeloma (MM) and affects more than 80% of patients. Although previous works revealed the roles of N6-methyladenosine (m6A) reader hnRNPA2B1 in the development of tumors, whether hnRNPA2B1 regulates bone destruction in MM is still unknown. Methods: Alizarin red S staining, TRAP staining, ELISA and quantitative real-time PCR assays were used to evaluate osteogenesis and osteoclastogenesis in vitro. X ray and bone histomorphometric analysis were preformed to identify bone resorption and bone formation in vivo. Exosome isolation and characteriz

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… We isolated exosomes from cell culture medium using ultracentrifugation and confirmed their identity by electron microscopy and dynamic light scattering analysis ( Figure 4 A ). This was further confirmed by the expression of exosome markers ( Figure 4 B-D ). …

Discovery and validation of surface N-glycoproteins in MM cell lines and patient samples uncovers immunotherapy targets.

BackgroundMultiple myeloma (MM) is characterized by clonal expansion of malignant plasma cells in the bone marrow. While recent advances in treatment for MM have improved patient outcomes, the 5-year survival rate remains ~50%. A better understanding of the MM cell surface proteome could facilitate development of new directed therapies and assist in stratification and monitoring of patient outcomes.MethodsIn this study, we first used a mass spectrometry (MS)-based discovery-driven cell surface capture (CSC) approach to map the cell surface N-glycoproteome of MM cell lines. Next, we developed t

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… A membrane-enriched fraction was prepared by ultracentrifugation at 210,000 × g for 18 hours. The resulting membrane pellet was digested with trypsin and biotinylated glycopeptides were captured by immobilized streptavidin resin and stringently washed to remove non-specifically bound peptides. …

The Role of Extracellular Vesicles in Bone Metastasis.

Multiple types of cancer have the specific ability to home to the bone microenvironment and cause metastatic lesions. Despite being the focus of intense investigation, the molecular and cellular mechanisms that regulate the metastasis of disseminated tumor cells still remain largely unknown. Bone metastases severely impact quality of life since they are associated with pain, fractures, and bone marrow aplasia. In this review, we will summarize the recent discoveries on the role of extracellular vesicles (EV) in the regulation of bone remodeling activity and bone metastasis occurrence. Indeed,

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… The procedure for their isolation should be optimized, since many protocols are used, including differential ultracentrifugation, flotation on density gradients, polyethylene glycol precipitation, polymer-based precipitation, and immunoprecipitation. …

Synthesis and biological evaluation of a new polymeric conjugate and nanocarrier with osteotropic properties.

Bone-seeking (osteotropic) drug delivery systems (ODDS) represent an interesting solution for targeting different types of drugs to the bones. In particular, anticancer and antibacterial agents could take advantage of such therapeutic strategy. We have recently developed an innovative approach to this aim: a new osteotropic biomaterial was prepared, based on the conjugation of a poly(lactide-co-glycolide) (PLGA) with the bisphosphonate drug alendronate (PLGA-ALE); its hemo- and cytocompatibility were verified. Starting with this copolymer, an osteotropic nanoparticle system (NP) was produced f

Демиелинция производит сдвиг в населении кортикальных нейронов, которые синапсируют с каллосальными олигодендроцитными клетками.

Клетки потомства олигодендроцитов (ОРК) получают синаптическое воздействие от различных нейронов в мозгу развивающихся и взрослых. Понимание того, меняются ли нейронные популяции, которые синапируют с OPC в здоровом мозге в результате обезвоживания и/или ремиэлирования, может способствовать совершенствованию стратегий нейрозащиты или миелинового ремонта, разрабатываемых для демиелинизирующих заболеваний, таких, как множественный склероз. Чтобы изучить эту возможность, мы использовали трансгенную технологию крелокса, чтобы облегчить инфекцию ОПС модифицированным вирусом бешенства, что позволяет

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… The pseudotyped SADΔG-GFP-(EnvA) viral supernatant was collected and filtered, and the SADΔG-GFP-(EnvA) virus concentrated by ultracentrifugation at 24,000 rpm (∼72,000 ×  g ) for 2 h at 4°C using a Sorvall WX+ Ultracentrifuge (Thermo Fisher Scientific, 75000100) with a TH-641 Swinging Bucket Rotor ( …

Изображение миэломы экзосома0002724 поощрение остеокластогенеза через miR-4753-3p/IFIT1 и ось RANK в болезни костей миэломы.

КОЛИЧЕСТВО: Болезнь кости миеломы (МБД) вызвана чрезмерной активацией остеокласта и постепенным разрушением костей, однако роль экзосомных круговых РНК (КРН) из клеток миэломы остается неясной. МЕТОДЫ: Экзосомы из клеток U266 и MM1S были изолированы и использовались для лечения THP-1 клетки. Остеокластная дифференциация была оценена с помощью анализа пятен и ресорбции костей. Для выявления совпадающих пятен вокруг РНК была проведена последовательность последовательности экзосомы U266 и крови пациента MBD. Сеть ceRNA была построена с помощью циркбанка и миРТарбазы в сочетании с одноклеточной RN

Техническое обслуживание дисмиелинических небольших диаметровых аксонов 14-3-3s в центральной нервной системе.

В центральной нервной системе миелин, образующийся вокруг нервных аксонов олигодендроцитом, позволяет эффективно проводить потенциалы действий и поддерживает аксонный гомеостаз. Таким образом, в случае демиелинизирующих заболеваний, таких, как множественный склероз (MS), наблюдается аксональное дегенерация и потеря, особенно в аксонах малых диаметров. Teneurin-4 Недостатки (недостатки)Ten-4 -) Мыши демонстрируют сильное дисмиэлирование микродиаметровых аксонов, но выживают больше года. Здесь мы хотели прояснить механизм выживания аксонов малого диаметра без миелина с помощью Ten-4 - Мыши. Имму

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… Next, we prepared a barrel with a density gradient for ultracentrifugation and pipetted the lightest gradient first, and then, pipetted the heavier gradient at the bottom with a 10 mL syringe connected to a long needle, to build up the density gradient. …

Создание блокирующей ЭЛИСА, использующей VP1-specific моноклонные антитела для обнаружения антител DHAV3.

Вирус гепатита Утки А (DHAV) вызывает острую и серьезную инфекционную болезнь, характеризующуюся опухолью печени и кровоизлиянием в организм утят. three неделями в возрасте. Эта болезнь приводит к значительным экономическим потерям в уточной промышленности. В Китае широко распространены как DHAV1, так и DHAV3, при этом более доминирующим является DHAV3. В числе three Структурные белки DHAV, белки VP1 являются наиболее критическими, поскольку они вызывают нейтрализующее производство антител, служат связующим белком для вирусной адсорбции рецепторов для конкретных клеток и определяют вирусную ан

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… The suspensions were first centrifuged at 500 g for 5 minutes to remove cell debris, followed by ultracentrifugation at 100,000g for 1 hour to collect the VLP pellet. …

Новые клеточные модели для изучения взаимодействия Мьелин и Микроглиа.

Множественный склероз (MS) характеризуется обезвоживанием и нейроинфламмацией, при этом окислительный стресс играет ключевую роль в патологии поражения. Цель этого исследования заключалась в изучении дифференциальной реакции клеток на миелиновый мусор в различных окислительных состояниях. Окисление Myelin было вызвано с помощью системы Cuperoxid, что подтверждается повышенными уровнями TBARS и автофлюоресценцией. Из-за воздействия миелина на жизнеспособность микроглии BV-2 так и не была затронута. Однако окисленный миелин значительно изменил маркеры окислительного стресса, автофагию и метаболи

Exosomes as a Novel Approach to Reverse Osteoporosis: A Review of the Literature.

Osteoporosis is a chronic disease requiring long-term, sometimes lifelong, management. With the aging population, the prevalence of osteoporosis is increasing, and with it so is the risk of hip fracture and subsequent poor quality of life and higher mortality. Current therapies for osteoporosis have various significant side effects limiting patient compliance and use. Recent evidence has demonstrated the significant role of exosomes in osteoporosis both in vivo and in vitro. In this review, we summarize the pathogenesis of senile osteoporosis, highlight the properties and advantages of exosome

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… In addition, current isolation methods of exosomes include mainly repeated ultracentrifugation and ultrafiltration ( Lobb et al., 2015 ), techniques which are too time-consuming and remain somewhat controversial. …

Exogenous Heat Shock Proteins in Oncology: Biological Roles and Clinical Implications.

Heat shock proteins (HSPs), particularly HSP70 and HSP90, are highly conserved molecular chaperones that protect cells from a wide range of stressors and maintain proteome homeostasis. In cancer, tumor cells frequently overexpress and actively release HSPs into the extracellular space and circulation in response to metabolic alterations, hypoxia, oxidative stress, and therapeutic interventions. Consequently, circulating HSP levels are often elevated in patients with malignancies compared with healthy individuals. This review summarizes current evidence on the diagnostic, prognostic, and predic

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… We recommend that future validation protocols employ dual isolation methods (combining affinity capture or ultracentrifugation for the vesicular fraction with whole-serum ELISA) to formally distinguish these biological sources [ 11 , 33 ]. … Therefore, future studies and clinical validation protocols should provide for the mandatory use of dual isolation methods—a combination of ultracentrifugation or affinity capture for the vesicular fraction with standard total pool ELISA—to distinguish the biological source of the circulating signal. …

Стратегия FACS-Proteomics, направленная на экстраклелектальную эмоциональную характеристику однофенотипов в биологических жидкостях: изучение роли лейкоцитных ЭМ в множественном склерозе.

СПРАВОЧНАЯ ИНФОРМАЦИЯ: Изоляция и протеомика внеклеточной везиклетки (ЭМ) от жидкостей организма сопряжены с трудностями из-за их большой неоднородности. Недавно мы продемонстрировали, что срабатывание ячеек (FACS) с флюоресцентной активацией эффективно изолирует весь ЭМ-диапазон непосредственно от нетронутых жидкостей тела, что позволяет провести всеобъемлющий протеомикальный анализ ЭМ. Здесь мы впервые описали однофенотипный ЭВ, сортируя лейкоцитные ЭВ (Leuko EVs) из периферической крови и слез здоровых добровольцев.Используя оптимизированный и запатентованный протокол пятна всего ЭМ-отсека,

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… Other studies employed the use of chemical precipitation kits [ 13 , 14 ], or several series of ultrafiltration combined with ultracentrifugation or size-exclusion chromatography (SEC) [ 15 , 16 ], or immuno-isolation with magnetic beads like ExoChip [ 17 ], and microfluidic separations with ciliated …

Tumour-Derived Extracellular Vesicles (EVs): A Dangerous "Message in A Bottle" for Bone.

Several studies have shown the importance of Extracellular Vesicles (EVs) in the intercellular communication between tumour and resident cells. Through EVs, tumour cells can trigger cell-signalling molecules and shuttle exogenous information to target cells, thus promoting spread of the disease. In fact, many processes are fuelled by EVs, such as tumour invasion and dormancy, drug-resistance, immune-surveillance escape, extravasation, extracellular matrix remodelling and metastasis. A key element is certainly the molecular profile of the shed cargo. Understanding the biochemical basis of EVs w

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… So far, the most employed method is ultracentrifugation, which, however, is time-consuming, and the obtained EVs are often a mix of different subtypes of EVs, being a limitation for those researchers interested to a specific subset of EVs. …

Triple‑negative breast cancer cell‑derived piR‑31115 promotes the proliferation and migration of endothelial cells via METTL3‑mediated m6A modification of YAP1.

Triple‑negative breast cancer (TNBC), a highly malignant breast cancer subtype with a pronounced metastatic propensity, forms the focus of the present investigation. MDA‑MB‑231, a prevalently utilized TNBC cell line in cancer research, was employed. In accordance with the tumour angiogenesis theory, cancer cells are capable of instigating angiogenesis and the formation of a novel vascular system within the tumour microenvironment, which subsequently sustains malignant proliferation and metastasis. Consequently, impeding the growth of tumour blood vessels holds substantial significance in suppr

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… After 48 h, the cell supernatant was collected and the lentiviral particles were then collected through ultracentrifugation. To infect the target cells, a multiplicity of infection of 50 was applied. The transduction of the cells of interest lasted ~6 h. …

Exosomes in oncofertility: emerging roles in chemotherapy-induced reproductive damage and fertility preservation.

ABSTRACT: Oncofertility has emerged as a critical interdisciplinary field addressing the reproductive challenges faced by cancer patients, particularly those undergoing chemotherapy. While chemotherapeutic agents remain indispensable in cancer therapy, their gonadotoxic effects frequently result in diminished ovarian reserve, impaired spermatogenesis, and long-term infertility. Exosomes - small extracellular vesicles enriched with nucleic acids, proteins, and lipids - are increasingly recognized as key mediators of intercellular communication in both pathological and regenerative contexts. Rec

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… Differential ultracentrifugation remains the gold standard, separating vesicles by size and density ( Huang et al. 2022 ). Ultrafiltration, using molecular weight cutoff membranes, offers faster processing of large volumes ( Diaz et al. 2018 , Abady et al. 2023 ). … Ultracentrifugation yields high-purity vesicles but is time-consuming, costly, and requires specialized equipment, while other methods often result in contamination from lipoproteins or ribonucleoprotein complexes, complicating downstream analysis – especially when RNA yields are low ( Hosseini et al …

Advanced biomaterial-based approaches for medication-related osteonecrosis of the jaw.

Medication-related osteonecrosis of the jaw (MRONJ) is a debilitating adverse effect linked to anti-resorptive and anti-angiogenic therapies, with an incidence of 4.34 % (6.22 % in oncology patients and 0.59 % in osteoporosis patients) based on a pooled analysis of 92 observational studies (2015-2020), characterized by persistent bone exposure and poor healing. Conventional treatment modalities often yield suboptimal outcomes, with recurrence and procedural morbidity. In recent years, biomaterial-based strategies have emerged as promising alternatives by targeting MRONJ's complex pathophysiolo

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… In addition, technical challenges persist in scaling up exosome production while maintaining consistency in purity, stability, and yield across manufacturing batches, which are mainly issues largely tied to limitations in purification methods (e.g., ultracentrifugation efficiency), bioreactor design … Current lab-scale isolation methods, such as ultracentrifugation, are low-yield, time-consuming, and prone to contamination, making them unsuitable for clinical-grade manufacturing. …

Modeling and addressing on-target/off-tumor toxicity of claudin 18.2 targeted immunotherapies.

Successfully extending immunotherapies to solid tumors involves addressing several key challenges, importantly the "antigen dilemma", the expression of a solid tumor target antigen on the normal tissue of tumor origin. Claudin 18.2 (CLDN18.2) has emerged as an important target for upper gastrointestinal (GI) cancer therapies (such as Zolbetuximab, a naked antibody, recently approved; or CT041, a second-generation chimeric antigen receptor (CAR) T cell therapy with promising clinical data). However, GI toxicities are reported from clinical use of both Zolbetuximab and CT041. Here, we describe c

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… Viral supernatants were used directly for transduction or concentrated by ultracentrifugation (110,000 × g , 2 h, 4 °C), aliquoted, and stored at −80 °C. Deidentified, discarded blood from donors was used for the isolation of immune cells for the production of CAR T cells. …