Identification of potential biomarkers related to mesenchymal stem cell response in patients with Alzheimer's disease.

BACKGROUND: Preclinical studies showed that mesenchymal stem cells (MSCs) ameliorate tau phosphorylation, amyloid-beta accumulation, and inflammation in Alzheimer's disease (AD) mouse models via secretion of neurotrophic factors and cytokines. We aimed to identify CSF biomarkers that can be used to predict or monitor the response to MSCs in patients with AD. METHODS: AD patients were injected with human umbilical cord blood-MSCs (n = 22) or placebo (n = 12). The cerebrospinal fluid (CSF) samples were collected at baseline, one day after the first injection, and one day after the third injection. The patients injected with MSCs were classified into good responder (GR) or poor responder (PR) groups based on the rate of changes in the ratio of total-tau and phosphorylated-tau in the CSF. We selected three typical participants in each group, and their CSF protein levels were analyzed using liquid chromatography/tandem mass spectrometry (LC-MS/MS). RESULTS: In the LC-MS/MS analysis, 1,667 proteins were identified. Eleven proteins showed significant differences between the typical GR and PR at baseline. Based on their significance level and known functions, two proteins, reticulocalbin-3 (RCN3) and follistatin-related protein 3 (FSTL3), were selected as potential biomarkers to predict MSC response. A total of 173 proteins showed significant change one day after the third injection compared to the baseline in typical GR. We excluded 45 proteins that showed significant change after the third injection compared to the baseline in the typical PR. Based on their significance level and known function, four proteins, scrapie-responsive protein 1 (SCRG1), neural proliferation differentiation and control protein (NPDC1), apolipoprotein E (ApoE), and cystatin C (CysC), were selected as potential biomarker to monitor MSC response. Additionally, functional analysis revealed that the increased CSF proteins after the third injection compared to the baseline in the typical GR were associated with synaptogenesis. CONCLUSIONS: This study identified two proteins (RCN3 and FSTL3) that may be potential biomarkers for predicting MSC response and four proteins (SCRG1, NPDC1, ApoE, CysC) that may be potential biomarkers for monitoring MSC response in patients with AD. Further studies are needed to validate our results. Trial registration Clinical Trials.gov, NCT02054208. Registered on 4 February 2014. Samsung Medical Center IRB File No.2017-04-025. Registered on 20 June 2017.

Immunosuppressant Drugs Mitigate Immune Responses Generated by Human Mesenchymal Stem Cells Transplanted into the Mouse Parenchyma.

It has been widely accepted that mesenchymal stem cells (MSCs) can evade the immune surveillance of the recipient. However, emerging research cast doubt on whether MSCs are intrinsically immune-privileged. Previously, we observed that the transplantation of human MSCs (hMSCs) into the mouse parenchyma attracted a high infiltration of leukocytes into the injection tract. Thus, in order to reduce the immune responses generated by hMSCs, the aim of this study was to assess which immunosuppressant condition (dexamethasone only, tacrolimus only, or dexamethasone and tacrolimus together) would not only reduce the overall immune response but also enhance the persistence of MSCs engrafted into the caudate putamen of wild-type C57BL/6 mice. According to immunohistochemical analysis, compared to the hMSC only group, the administration of immunosuppressants (for all three conditions) reduced the infiltration of CD45-positive leukocytes and neutrophils at the site of injection. The highest hMSC persistence was detected from the group that received combinatorial administrations of dexamethasone and tacrolimus. Moreover, compared to the immunocompetent WT mouse, higher MSC engraftment was observed from the immunodeficient BALB/c mice. The results of this study support the use of immunosuppressants to tackle MSC-mediated immune responses and to possibly prolong the engraftment of transplanted MSCs.

Ethionamide Preconditioning Enhances the Proliferation and Migration of Human Wharton's Jelly-Derived Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) are a useful source for cell-based therapy of a variety of immune-mediated diseases, including neurodegenerative disorders. However, poor migration ability and survival rate of MSCs after brain transplantation hinder the therapeutic effects in the disease microenvironment. Therefore, we attempted to use a preconditioning strategy with pharmacological agents to improve the cell proliferation and migration of MSCs. In this study, we identified ethionamide via the screening of a drug library, which enhanced the proliferation of MSCs. Preconditioning with ethionamide promoted the proliferation of Wharton's jelly-derived MSCs (WJ-MSCs) by activating phosphatidylinositol 3-kinase (PI3K)/Akt and mitogen-activated protein kinase/extracellular signal-regulated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK)1/2 signaling. Preconditioning with ethionamide also enhanced the migration ability of MSCs by upregulating expression of genes associated with migration, such as C-X-C motif chemokine receptor 4 (CXCR4) and C-X-C motif chemokine ligand 12 (CXCL12). Furthermore, preconditioning with ethionamide stimulated the secretion of paracrine factors, including neurotrophic and growth factors in MSCs. Compared to naïve MSCs, ethionamide-preconditioned MSCs (ETH-MSCs) were found to survive longer in the brain after transplantation. These results suggested that enhancing the biological process of MSCs induced by ethionamide preconditioning presents itself as a promising strategy for enhancing the effectiveness of MSCs-based therapies.

SERPINB2 Is a Novel Indicator of Cancer Stem Cell Tumorigenicity in Multiple Cancer Types.

: Drug resistance is one of the major characteristics of cancer stem cells (CSCs) and a mechanism of tumor recurrence. Therefore, selectively targeting CSCs may be an effective therapeutic strategy to overcome cancer recurrence. In the present study, we found that exposure to tumorigenic compounds significantly increased the growth potential and stem-cell-like properties of various CSCs. Early-response genes involved in tumorigenesis can be used as specific markers to predict potential tumorigenicity. Importantly, for the first time we identified, a labile tumorigenic response gene-SERPINB2-and showed that tumorigenic compound exposure more profoundly affected its expression in CSCs than in non-stem cancer cells, although both cells exhibit basal expression of SERPINB2 in multiple cancer types. Our data also revealed a strong relationship between the significantly enhanced expression of SERPINB2 and metastatic progression in multiple cancer types. To the best of our knowledge, this is the first study to focus on the functions of SERPINB2 in the tumorigenicity of various CSCs and these findings will facilitate the development of promising tumorigenicity test platforms.

SERPINB2 is a novel indicator of stem cell toxicity.

The toxicological evaluation of potential drug candidates is very important in the preclinical phase of drug development. Toxic materials may cause serious decline in stem cell function and loss of stemness. Indeed, we found that toxic exposure more profoundly suppressed the growth of stem cells than terminally differentiated fibroblasts. Importantly, toxic exposure suppressed stem cell migration and multi-lineage differentiation potential in vitro and in vivo. Moreover, early-response genes involved in stem cell properties such as self-renewal and differentiation capabilities can be used as specific markers to predict toxicity. In the present study, we also identified a labile toxic response gene, SERPINB2, which is significantly increased in response to various toxic agents in human stem cells in vitro and in vivo. Consistently, self-renewal, migration, and multi-lineage differentiation potential were markedly decreased following SERPINB2 overexpression. To the best of our knowledge, this is the first study to focus on the functions of SERPINB2 on the regenerative potential of stem cells in response to various existing chemicals, and the findings will facilitate the development of promising toxicity test platforms for newly developed chemicals.

HIF2α/EFEMP1 cascade mediates hypoxic effects on breast cancer stem cell hierarchy.

Breast cancer stem cells (BCSCs) have been shown to contribute to tumor growth, metastasis, and recurrence. They are also markedly resistant to conventional cancer treatments, such as chemotherapy and radiation. Recent studies have suggested that hypoxia is one of the prominent micro-environmental factors that increase the self-renewal ability of BCSCs, partially by enhancing CSC phenotypes. Thus, the identification and development of new therapeutic approaches based on targeting the hypoxia-dependent responses in BCSCs is urgent. Through various in vitro studies, we found that hypoxia specifically up-regulates BCSC sphere formation and a subset of CD44+/CD24-/low CSCs. Hypoxia inducible factors 2α (HIF2α) depletion suppressed CSC-like phenotypes and CSC-mediated drug resistance in breast cancer. Furthermore, the stimulatory effects of hypoxia-induced HIF2α on BCSC sphere formation were successfully attenuated by epidermal growth factor-containing fibulin-like extracellular matrix protein 1 (EFEMP1) knockdown. Taken together, these data suggest that HIF2α mediates hypoxia-induced cancer growth/metastasis and that EFEMP1 is a downstream effector of hypoxia-induced HIF2α during breast tumorigenesis.

Haemodynamic alterations in cerebral blood vessels after carotid artery revascularisation: quantitative analysis using 2D phase-contrast MRI.

OBJECTIVES: This study was conducted to evaluate the effect of revascularisation, whether revascularisation improves total cerebral blood flow volume (FVTCBF), and how cerebral veins would respond to altered FVTCBF. METHODS: The 39 carotid artery stenoses in 37 patients who underwent revascularisation including 32 stentings and 7 endarterectomies were included in this prospective study. From the two-dimensional phase-contrast (2D-PC) MRI acquired before and after revascularisation, the flow volumes (FVs) of the arteries and veins were compared using paired t-test. The relationships between these parameters were correlated using Pearson's correlation coefficient. RESULTS: The mean FV in the treated carotid artery (proportion of treated artery among total FV) increased from 162.06 ml/min (25.80 %) to 267.71 ml/min (37.21 %; P < 0.001). Revascularisation increased the FVTCBF of patients from 638.66 ml/min to 716.72 ml/min (P < 0.001). The FV of the internal jugular veins, superior sagittal and straight sinuses (FVSS + SSS), and transverse sinuses increased after revascularisation (P < 0.05). Positive relationships were shown between the FVTCBF and the FVSS + SSS (r = 0.584-0.741, P < 0.001). CONCLUSIONS: Revascularisation improves the FVTCBF by increasing the FV in the treated carotid artery. The venous drainages are closely linked to FVTCBF. 2D-PC-MRI is a feasible method for evaluating comprehensively the haemodynamic improvement after revascularisation. KEY POINTS: • Revascularisation may be beneficial in ischaemic strokes due to carotid artery stenosis. • Revascularisation of the affected artery increases total cerebral blood flow volume ( FV TCBF). • Cerebral venous drainage, closely linked to FV TCBF, is also improved. • Two-dimensional phase-contrast MRI can comprehensively assess these haemodynamic improvements after carotid revascularisation.

Human intestinal epithelial cells are broadly unresponsive to Toll-like receptor 2-dependent bacterial ligands: implications for host-microbial interactions in the gut.

Intestinal epithelial cells (IEC) interact with a high density of Gram-positive bacteria and are active participants in mucosal immune responses. Recognition of Gram-positive organisms by Toll-like receptor (TLR)2 induces proinflammatory gene expression by diverse cells. We hypothesized that IEC are unresponsive to Gram-positive pathogen-associated molecular patterns and sought to characterize the functional responses of IEC to TLR2-specific ligands. Human colonic epithelial cells isolated by laser capture microscopy and IEC lines (Caco-2, T84, HT-29) were analyzed for expression of TLR2, TLR6, TLR1, and Toll inhibitory protein (Tollip) mRNA by RT-PCR and quantitative real-time PCR. Response to Gram-positive bacterial ligands was measured by NF-kappa B reporter gene activation and IL-8 secretion. TLR2 protein expression was analyzed by immunofluorescence and flow cytometry. Colonic epithelial cells and lamina propria cells from both uninflamed and inflamed tissue demonstrate low expression of TLR2 mRNA compared with THP-1 monocytes. IECs were unresponsive to TLR2 ligands including the staphylococcal-derived Ags phenol soluble modulin, peptidoglycan, and lipotechoic acid and the mycobacterial-derived Ag soluble tuberculosis factor. Transgenic expression of TLR2 and TLR6 restored responsiveness to phenol soluble modulin and peptidoglycan in IEC. In addition to low levels of TLR2 protein expression, IEC also express high levels of the inhibitory molecule Tollip. We conclude that IEC are broadly unresponsive to TLR2 ligands secondary to deficient expression of TLR2 and TLR6. The relative absence of TLR2 protein expression by IEC and high level of Tollip expression may be important in preventing chronic proinflammatory cytokine secretion in response to commensal Gram-positive bacteria in the gut.