Dataset Information

Proteomics of hypoxic mesenchymal stem cells

ABSTRACT: Background: Neonatal hypoxic-ischemic (HI) brain injury, is one of the leading causes of mortality and long-term neurological morbidity in newborns. Current treatment options for HI brain injury are very limited, but mesenchymal stem cell (MSC) therapy is a promising strategy to boost neuroregeneration after injury. Optimization strategies to further enhance the potential of MSCs are in development. In the current study we aimed to test the potency of hypoxic preconditioning (HP) to enhance the therapeutic efficacy of MSCs in a mouse model for neonatal HI injury. Methods: HI was induced on postnatal day 9 in C57Bl/6 mouse pups. MSCs were cultured at 1% oxygen levels for 24 hours prior to use (HP-MSCs) or under normoxic (21% O2) control conditions (NP-MSCs). At 10 days after induction of HI brain injury, HP-MSCs or NP-MSCs were intranasally administered. Lesion size, sensorimotor outcome, MSC migration and neuroinflammation were assessed by HE staining, cylinder rearing task, gold nanoparticle-labeled MSC tracing and IBA1 staining, respectively. In vitro, the effect of hypoxic preconditioning on MSC migration, potency and proteome profile was studied using assays for transwell-migration, neural stem cell differentiation and neuroinflammation, and LC-MS/MS, respectively. Results: HP-MSCs were superior to NP-MSCs in reducing lesion size and improving sensorimotor outcome after HI. Moreover, hypoxic preconditioning enhanced MSC migration specifically to the HI lesion after intranasal application and in vitro. Additionally, HP-MSCs enhanced neural stem cell (NSC) differentiation into more complex neurons in vitro but did not enhance anti-inflammatory effects compared to NP-MSCs. Lastly, hypoxic preconditioning enriched the expression of pathways mainly related to glucose metabolism and extracellular matrix remodeling in MSCs. Conclusions: Overall, this study showed for the first time that intranasal HP-MSC therapy is a promising optimization strategy to superiorly reduce lesion size and improve neurodevelopmental outcome in a mouse model of neonatal HI brain injury.

INSTRUMENT(S): Orbitrap Eclipse

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Stem Cell

DISEASE(S): Wounds And Injuries

SUBMITTER: Tatiana Shamorkina

LAB HEAD: Albert JR Heck

PROVIDER: PXD053838 | Pride | 2025-06-30

REPOSITORIES: Pride

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			Action	DRS
	20231122_E1_UM1_shamo002_SA_EXT00_MSCs_18plexPro_HpH_F1.raw	Raw
	20231122_E1_UM1_shamo002_SA_EXT00_MSCs_18plexPro_HpH_F10.raw	Raw
	20231122_E1_UM1_shamo002_SA_EXT00_MSCs_18plexPro_HpH_F11.raw	Raw
	20231122_E1_UM1_shamo002_SA_EXT00_MSCs_18plexPro_HpH_F12.raw	Raw
	20231122_E1_UM1_shamo002_SA_EXT00_MSCs_18plexPro_HpH_F13.raw	Raw

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Publications

Hypoxic Preconditioning Enhances the Potential of Mesenchymal Stem Cells to Treat Neonatal Hypoxic-Ischemic Brain Injury.

De Palma Sara T ST Hermans Eva C EC Shamorkina Tatiana M TM Trayford Chloe C van Rijt Sabine S Heck Albert J R AJR Nijboer Cora H A CHA de Theije Caroline G M CGM

Stroke 20250418 7

<h4>Background</h4>Neonatal hypoxic-ischemic (HI) brain injury is one of the leading causes of long-term neurological morbidity in newborns. Current treatment options for HI brain injury are limited, but mesenchymal stem cell (MSC) therapy is a promising strategy to boost neuroregeneration after injury. Optimization strategies to further enhance the potential of MSCs are under development. The current study aimed to test the potency of hypoxic preconditioning of MSCs to enhance the therapeutic e ...[more]

PMID: 40248869

Similar Datasets

Project description:Hypoxic-ischemic (HI) injury in the developing brain is a common cause of disability in children, and there are no effective treatments at this time. Exposure to sublethal hypoxic conditions (hypoxic preconditioning) 24 hours prior to hypoxic-ischemic insult is protective in the developing rat model. We have observed protective effects on brain histopathology and on long-term sensory-motor behavioral tasks. Changes in gene expression are thought to underlie this protective effect. By comparing gene expression in rats subjected to hypoxic preconditioning or sham conditioning at several time points from 0 to 24 hrs after preconditioning, we should gain insight into the mechanisms underlying these neuroprotective effects and may identify targets for therapeutic intervention. The aim of this study is to determine the effect of hypoxic preconditioning on global gene expression, and, in littermates, to examine the effect of hypoxic preconditioning 24 h prior to hypoxic-ischemic insult on brain histopathology. We hypothesize that changes in gene expression underlie the protective effect of hypoxic preconditioning against subsequent hypoxic-ischemic insult. Gene expression will be examined in two groups, 1) preconditioned and 2) sham controls, at 4 time points. On postnatal day 6, preconditioned animals are exposed to normothermic hypoxia for 3 hrs (8.0% oxygen, 36 degrees C), and sham animals are simultaneouosly exposed to normoxia at 36 degrees C. Animals are then returned to their dams until euthanized at 4 time points (0h, 2h, 8h, and 24h later). Five brains/group/timepoint will be used, with an equal number of males and females in each group. Brains are removed and dissected on ice. Cerebral cortex is dissected from both hemispheres and rapidly frozen on dry ice. Total RNA is isolated using the QIAGEN RNeasy Protect Maxi Kit. Littermates of these animals will be exposed to hypoxic preconditioning or sham preconditioning and subjected to hypoxic-ischemic injury 24 h later. These animals are euthanized at postnatal day 14 for histopathologic evaluation of injury.

Project description:Background Extracellular vesicles (EVs) secreted by mesenchymal stromal cells (MSCs) provide significant protection against renal ischemia-reperfusion injury (IRI). Hypoxia is considered an important method for enhancing the tissue repair capabilities of MSCs. However, the specific effects of hypoxia on MSCs and MSC-EVs, as well as their therapeutic potential for renal IRI, remain unclear. In this study, we investigated the alterations in MSCs and the production of MSC-EVs following hypoxia pre-treatment, and further explored the key intrinsic mechanisms by which hypoxic MSC-EVs treat renal IRI. Methods Human umbilical cord MSCs were cultured under normoxic and hypoxic conditions. Proliferation and related pathways were measured, and RNA sequencing was used to detect changes in the transcription profile. MSC-EVs from both normoxic and hypoxic conditions were isolated and characterized. In vivo, the localization and therapeutic effects of MSC-EVs were assessed in a rat renal IRI model. Histological examinations were employed to assess the structure, proliferation, and apoptosis of IRI kidney tissue respectively. Renal function was measured by analyzing serum creatinine and blood urea nitrogen levels. In vitro, the therapeutic potential of MSC-EVs were measured in renal tubular epithelial cells injured by antimycin A. Protein sequencing analysis of hypoxic MSC-EVs was conducted, and the depletion of Glutathione S-Transferase Omega 1 (GSTO1) in hypoxic MSC-EVs was performed to verify its key role in alleviating renal injury. Results Hypoxia alters MSCs transcription, promotes their proliferation, and increases the production of EVs. Hypoxia-pretreated MSC-EVs exhibited a superior ability to mitigate renal IRI, enhancing proliferation and reducing apoptosis of renal tubular epithelial cells both in vivo and in vitro. Protein profiling of the EVs revealed an accumulation of numerous anti-oxidative stress proteins, with GSTO1 being particularly prominent. GSTO1 knock down was significantly reduced the antioxidant and therapeutic effects in renal IRI of hypoxic MSC-EVs. Conclusions Hypoxia significantly promotes MSC-EVs generation and enhances the therapeutic effect of EVs on renal IRI. The effect of antioxidant stress induced by GSTO1 is one of the most important underlying mechanisms. Our findings underscore that hypoxia-pretreated MSC-EVs represent a novel and promising therapeutic intervention for renal IRI.

Project description:Intervertebral disc (IVD) degeneration is a common pathological condition associated with low back pain. Recent evidence suggests that MSCs promote IVD regeneration, but underlying mechanisms remain poorly defined. One postulated mechanism is via modulation of macrophage phenotypes. MSCs encounter drastic declines in oxygen tension when injected into the avascular IVD; thus, understanding how oxygen tension affects interactions between MSCs and macrophages may offer clues as to how MSCs act therapeutically. We investigated how conditioned medium (CM) derived from MSCs cultured in hypoxia altered macrophage subsets. We cultured human, bone marrow-derived STRO3+ MSCs in hypoxia or normoxia, and collected CM. We then cultured human bone marrow-derived macrophages with IFN-y, followed by either hypoxic or normoxic STRO3+ MSC CM. We also cultured macrophages in hypoxic MSC CM in the presence of IL-4 and collected cells for scRNA-seq analyses. Bioinformatic analyses confirmed multiple macrophage subpopulations. We first performed comparative analyses between macrophages cultured in hypoxic and normoxic MSC CM. Integration of these data sets showed large overlap between macrophage subsets; however, we identified a unique hypoxic MSC CM-induced macrophage cluster. Gene Ontogeny (GO) analyses revealed enrichment in IL-10, Ccl5/Ccr5, G alpha (i) Signaling, and Rho GTPases within this unique cluster. To determine if factors from MSC CM simulated effects of the anti-inflammatory cytokine IL-4, we integrated the data from macrophages cultured in hypoxic MSC CM with and without IL-4 addition. Integration of these data sets showed considerable overlap, demonstrating that hypoxic MSC CM simulates the effects of IL-4. Interestingly, macrophages cultured in normoxic MSC CM in the absence of IL-4 did not significantly contribute to the unique cluster within our comparison analyses and showed differential TGF-β signaling; thus, normoxic conditions did not approximate IL-4. In addition, TGF-β neutralization partially limited the effects of MSC CM. Our study identifies a unique macrophage subset induced by MSCs within hypoxic conditions, and supports that MSCs alter macrophage phenotypes through TGF-β-dependent mechanisms