Dataset Information

HTERT mesenchymal stem cells (MSCs), Stem cells EVs LC-MS/MS

ABSTRACT: As the economic challenges associated with vision impairment and ocular injuries continue to escalate, there exists an urgent need to explore innovative remedies for these conditions. Extracellular Vesicles (EVs), containing a diverse array of biological components such as nucleic acids, proteins, and lipids, have garnered substantial scientific support for their healing and immune-regulating attributes in addressing various disorders. Nevertheless, an area that remains relatively uncharted is the potential reparative impact of stem cell-derived EVs within the realm of ocular damage or disease. Furthermore, prevailing literature predominantly focuses on EVs derived from primary stem cells, neglecting a comprehensive assessment of EVs derived from hTERT-immortalized stem cells. Through our extensive EV production platform, we have consistently generated EVs sourced from hTERT mesenchymal stem cells (MSCs) and have employed diverse techniques to characterize and profile their cargo contents. Leveraging established cellular assays, we have conducted a comparative analysis of these EVs' effects on both intact and impaired retinal pigment epithelial cells. To the best of our knowledge, this marks the inaugural study demonstrating the feasibility of consistent, large-scale production of hTERT MSC EVs and exploring their potential regenerative properties against compromised ocular cells. The outcomes of our investigations affirm that hTERT MSCs elicit multiple reparative influences in vitro, underscoring their promise not only as a viable substitute for primary MSC-derived EVs but also as a prospective innovative therapeutic agent for addressing ocular injuries.

INSTRUMENT(S): Orbitrap Exploris 480

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Stem Cell, Cell Culture

DISEASE(S): Retinal Degeneration

SUBMITTER: Fatah Kashanchi

LAB HEAD: Fatah Kashanchi

PROVIDER: PXD044629 | Pride | 2024-06-16

REPOSITORIES: Pride

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Publications

hTERT-Immortalized Mesenchymal Stem Cell-Derived Extracellular Vesicles: Large-Scale Manufacturing, Cargo Profiling, and Functional Effects in Retinal Epithelial Cells.

Hindle Jessica J Williams Anastasia A Kim Yuriy Y Kim Dongsung D Patil Kajal K Khatkar Pooja P Osgood Quinn Q Nelson Collin C Routenberg David A DA Howard Marissa M Liotta Lance A LA Kashanchi Fatah F Branscome Heather H

Cells 20240517 10

As the economic burden associated with vision loss and ocular damage continues to rise, there is a need to explore novel treatment strategies. Extracellular vesicles (EVs) are enriched with various biological cargo, and there is abundant literature supporting the reparative and immunomodulatory properties of stem cell EVs across a broad range of pathologies. However, one area that requires further attention is the reparative effects of stem cell EVs in the context of ocular damage. Additionally, ...[more]

PMID: 38786083

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Project description:Background: Extracellular vesicles (EVs) isolated from mesenchymal stem/stromal cells (MSCs) contribute to recovery of damaged tissue in animals models of human disease. We have previously shown that EVs isolated from porcine MSCs transport mRNA and miRNA capable of modulating several cellular pathways in recipient cells, yet their proteome remained to be profiled. Using a quantitative proteomic strategy, we sought to study the protein cargo of porcine MSC-derived EVs to identify candidate molecules for mediating their therapeutic effect. Methods: Autologous MSCs were collected from abdominal fat of 3 female domestic pigs, and MSC-derived EVs were subsequently isolated, cultured, and characterized by the expression of typical MSC and EV markers. LC-MS/MS proteomic analysis was performed and proteins classified using the Panther Classification System. Functional pathway analysis was performed with DAVID 6.7. Three candidate proteins were selected for validation and their expression in EVs and MSCs confirmed by Western blot. Results: Proteomics analysis identified 5,469 protein groups in MSCs and 4,937 in EVs. Average protein intensity was higher in MSCs compared to EVs (p<0.0001). Differential expression analysis revealed 128 proteins upregulated in EVs vs. MSCs (log2 fold change>10, p<0.05), whereas 563 proteins were excluded from EVs (log2 fold change<-10, p<0.05). Biological functional analysis of proteins enriched in EVs indicated a broad distribution, with the most frequently represented categories being proteins involved in angiogenesis, blood coagulation, apoptosis, extracellular matrix remodeling, and regulation of inflammatory responses. Proteins excluded from EVs were mostly nuclear proteins and proteins involved in nucleotide binding and RNA splicing. Conclusions: The present study provides novel proteomic characterization of the biological signatures of porcine adipose MSC-derived EVs. The selective cargo that EVs shuttle may define the spectrum of their roles in mediating MSC intercellular communication.

			Action	DRS
	03222023_Marissa_MSC.msf	Msf
	03222023_Marissa_MSC.raw	Raw
	03222023_Marissa_PC3.msf	Msf
	03222023_Marissa_PC3.raw	Raw