Effects of mesenchymal stem cells combined with iron-quercetin complex on relieving brain injury after intracerebral hemorrhage and stem cell tracking by magnetic resonance imaging

Abstract

Background Intracerebral hemorrhage (ICH) is a severe stroke type with various pathological changes, such as cerebral edema and neuroinflammation, which subsequently induces neurological deficits. The transplantation of mesenchymal stem cells (MSCs) as a neuroprotective therapy has been widely utilized in various brain diseases due to its anti-inflammatory effect. Nevertheless, following ICH onset, the severe inflammatory response affects and restricts MSCs’ survival rate, viability, and effectiveness. Therefore, improving their biological characteristics will be a promising therapeutic strategy in ICH. Interestingly, previous studies have demonstrated that the extraordinary dual capabilities of iron-quercetin complex (IronQ) as a stimulating agent for cell growth and an imaging probe by magnetic resonance imaging (MRI) were verified. Therefore, this study hypothesized that IronQ could improve the survival and viability of MSCs in treating ICH while labeling MSCs for their tracking by MRI. This study aimed to investigate the effects of MSCs with IronQ in regulating inflammation and further clarify their potential mechanisms in the ICH animal model. Methods MSCs were extracted from the femurs and tibias of C57BL/6 mice and identified by flow cytometry. Then, MSCs incubated with IronQ for 24h, the labeled efficiency of which was determined by Prussian blue staining, and cells were imaged through MRI in the ICH mice model. Conditioned medium (CM) from the MSCs cultured with IronQ was utilized to examine antioxidant activity in hydrogen peroxide (H2O2) activated-SH-SY5Y cells, and reactive oxygen species (ROS) was checked by flow cytometry. In this thesis, male C57BL/6 mice were induced with collagenase I to induce ICH and were independently divided into subgroups for therapeutic testing: (1) a control model group, Groups receiving (2) oral quercetin, (3) MSCs implanted (MSCs), and (4) those receiving IronQ-labeled MSCs cells for 24 h (MSCs+IronQ). Then the neurological deficits score, brain water content (BWC), inflammatory factors (TNF-α, IL-6), M1/M2 phenotypic protein expressions (CD80, CD86, CD206, Arg-1), neuroprotective related protein expressions (NeuN, MBP, and GFAP), in brain tissues were investigated. In addition, Mincle protein and its downstream targets were measured. Furthermore, the lipopolysaccharide (LPS)-induced BV2 cells were utilized to examine the neuroprotection of CM of MSCs co-cultured with IronQ in vitro. Results MSCs incubated with IronQ for 24 h showed increased Prussian blue staining when observed under a microscope. IronQ-labeled MSCs were seen as white spots in the hemorrhagic regions of ICH mice on MRI images using the T1 weighting technique. In vitro, results showed that the CM obtained from incubating MSCs with IronQ inhibited intracellular free radicals of SH-SY5Y induced by H2O2 and could prevent LPS-induced inflammation of BV2 cells. It has an anti-inflammatory mechanism by inhibiting the Miincle/syk signaling pathway. Studies in the ICH mice model showed that administering IronQ-labeled MSCs improved inflammation-induced neurodegenerative disorders and reduced cerebral water content better than using MSC stem cells alone and better than quercetin, respectively. It was found that inflammation in the brain tissue decreased. Pro-inflammatory agents (TNF-α, IL-6) and Mincle/syk protein expression were reduced in M1 macrophages, while the expression of the anti-inflammatory protein (NeuN, MBP, and GFAP) soared. Conclusions This dissertation suggested that the implanted IronQ-labeled MSCs could be tracked at the lesion of the ICH using T1-weighted magnetic resonance imaging. Further, IronQ combined with MSCs could alleviate ICH-induced inflammation by downregulating the Mincle/syk signaling pathway, further improving the neurologic deficits and brain edema.