Yonghua Zhao

Sodium Ferulate combined with bone marrow stromal cell treatment ameliorating rat brain ischemic injury after stroke.

Reports suggested that bone marrow stromal cells (BMSCs) could protect brain against ischemic injury. However, the limited number of BMSCs in the brain hampered their application. To explore a way that might improve the migration and differentiation of BMSCs in the brain after stroke, we investigated the additive therapeutic effect of combination Sodium Ferulate (SF) and BMSCs treatment of stroke. In general, BMSCs were primarily cultured from rat bone marrow and were identified by flow cytometry. Permanent middle cerebral artery occlusion (MCAo) was induced and the ischemic brain was observed by 2,3,5-triphenyltetrazolium chloride (TTC) staining. The neurological functional outcome was respectively evaluated at 0 hour, 24 hours, 48 hours, and 72 hours post-operation, and the stromal cell-derived factor-1 alpha (SDF-1α) /chemokine(CXC motif)receptor-4(CXCR4) mRNA and protein expressions at the marginal zone of the ischemic brain were respectively measured by real-time RT-PCR and Western Blot at 3days after stroke; furthermore, Nestin and BrdU double immunostaining was performed for identification of BMSC differentiation. Combination treatment of Stroke significantly improved the neurological functional recovery as early as 48 hours and up-regulated the SDF-1α /CXCR4 axis, enhanced BMSC migration into the ischemic brain and differentiation into Nestin-positive cells. We demonstrate that SF combined with BMSC administration can ameliorate rat brain ischemic injury and has beneficial effect on the rat neurological functional recovery after ischemic stroke.

Kuwanon G Preserves LPS-Induced Disruption of Gut Epithelial Barrier In Vitro

Defects in the gut epithelial barrier have now been recognized to be responsible for diabetic endotoxemia. In everyday life, Mulberry leaf tea is widely used in Asian nations due to its proposed benefits to health and control of diabetes. Evidence indicates the potential role of Kuwanon G (KWG), a component from Morus alba L., on blocking the gut epithelial barrier. In lipopolysaccharides (LPS)-damaged Caco-2 cells, it was found that KWG increased the viability of cells in a concentration-dependent manner. KWG administration significantly elevated the anti-oxidant abilities via increasing ratio of superoxidase dismutase (SOD)/malondialdehyde (MDA) and decreasing reactive oxygen species (ROS) within the cells. During KWG incubation, pro-inflammatory cytokines including interleukin (IL)-1β and tumor necrosis factor (TNF)-α were significantly reduced, tight junction proteins including zonula occludens (ZO)-1, intercellular adhesion molecule (ICAM)-1 and Occludin were dramatically increased as detected by immunofluorescence assay, trans-epithelial electrical resistance was significantly increased and the transmission of albumin-fluorescein isothiocyanate (FITC) across the barrier was decreased. In conclusion, the present study demonstrated that KWG could ameliorate LPS-induced disruption of the gut epithelial barrier by increasing cell viability and tight junction between cells, and decreasing pro-inflammatory cytokines and oxidative damage.

Bone Marrow Stromal Cells Combined with Sodium Ferulate and n-Butylidenephthalide Promote the Effect of Therapeutic Angiogenesis via Advancing Astrocyte-Derived Trophic Factors after Ischemic Stroke

Being a potential candidate for stroke treatment, bone marrow-derived mesenchymal stem/stromal cells (BM-MSCs) have been demonstrated to be able to enhance angiogenesis and proliferation of reactive astrocytes, which subsequently leads to the amelioration of neurological injury. Increasing evidence further indicates that combining BM-MSCs with certain agents, such as simvastatin, may improve therapeutic effects. Sodium ferulate (SF) and n-butylidenephthalide (BP), two main components of Radix Angelica Sinensis, are proven to be important regulators of stem cells in cell migration, differentiation, and pluripotency maintenance. This study aimed to investigate whether combining BM-MSCs with SF and BP had better therapeutic effect in the treatment of stroke, and the underlying molecular basis for the therapeutic effects was also investigated. The results showed that combination treatment notably reduced neurological injury after stroke and increased the expression of astrocyte-derived vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), and von Willebrand factor-positive vascular density in the ischemic boundary zone as evaluated by immuno fluorescence staining. After treatment with BM-MSCs plus SF and BP, astrocytes showed increased expression of VEGF and BDNF by upregulating protein kinase B/mammalian target of rapamycin (AKT/mTOR) expression in an oxygen- and glucose-deprived (OGD) environment. Human umbilical vein endothelial cells (HUVECs) incubated with the conditioned medium (CM) derived from OGD astrocytes treated with BM-MSCs plus SF and BP showed significantly increased migration and tube formation compared with those incubated with the CM derived from OGD astrocytes treated with BM-MSCs alone. These results demonstrate that combination treatment enhances the expression of astrocyte-derived VEGF and BDNF, which contribute to angiogenesis after cerebral ischemia, and the underlying mechanism is associated with activation of the astrocytic AKT/mTOR signaling pathway. Our study provides a potential therapeutic approach for ischemic stroke.

Evolutional Characterization of Photochemically Induced Stroke in Rats: a Multimodality Imaging and Molecular Biological Study.

Photochemically induced cerebral ischemia is an easy-manipulated, reproducible, relatively noninvasive, and lesion controllable model for translational study of ischemic stroke. In order to longitudinally investigate the characterization of the model, magnetic resonance imaging, 18F-2-deoxy-glucose positron emission tomography, fluorescence, and bioluminescence imaging system were performed in correlation with triphenyl tetrazolium chloride (TTC), hematoxylin-eosin staining, and immunohistochemistry examinations of glial fibrillary acidic protein, CD68, NeuN, von willebrand factor, and α-smooth muscle actin in the infarct zone. The results suggested that the number of inflammatory cells, astrocytes, and neovascularization significantly elevated in peri-infarct region from day 7 and a belt of macrophage/microglial and astrocytes was formed surrounding infarct lesion at day 14. Both vasogenic and cytotoxic edema, as well as blood brain-barrier leakage, occurred since day 1 after stroke induction and gradually attenuated with time. Numerous cells other than neuronal cells infiltrated into infarct lesion, which resulted in no visible TTC negative regional existence at day 14. Furthermore, recovery of cerebral blood flow and glucose utilization in peri-infarct zone were noted and more remarkably than that in infarct core following the stroke progression. In conclusion, these characterizations may be highly beneficial to the development of therapeutic strategies for ischemic stroke.

Calycosin Rebalances Advanced Glycation End Products-Induced Glucose Uptake Dysfunction of Hepatocyte In Vitro.

Diabetes mellitus (DM) often accompanies liver dysfunction. Astragali Radix is a traditional Chinese herbal medicine that is widely administrated to ameliorate the symptoms of diabetes as well as liver dysfunction, but its acting mechanism is still not yet fully recognized. Advanced glycation end products (AGEs) play a key role in promoting diabetic organ dysfunction. Both hyperglycemia and AGEs can induce insulin resistance, hepatocyte damage and liver dysfunction. We designed this study to explore the effects of the phytoestrogen Calycosin, a major active component of Astragali Radix, on AGEs-induced glucose uptake dysfunction in the hepatocyte cell line and relevant mechanisms. MTT and BrdU methods were applied to evaluate cell viability. 2-NBDG was used to observe glucose uptake by a live cell imaging system. Immunofluorescence method was carried out to investigate GLUT1, GLUT4, and RAGE protein expressions on cell membrane. cAMP content was determined by an EIA method. We found Calycosin concentration-dependently ameliorated AGEs-induced hepatocyte viability damage. AGEs dramatically reduced basal glucose uptake in hepatocytes, and this reduction could be reversed by Calycosin administration. By immunofluorescence detection, we observed that Calycosin could inhibit AGEs-induced GLUT1 expression down-regulation via estrogen receptor (ER). Furthermore, Calycosin decreased AGEs-promoted RAGE and cAMP elevation in hepatocytes. These findings strongly suggest that Calycosin can ameliorate AGEs-promoted glucose uptake dysfunction in hepatocytes; the protection of cell viability and ER-RAGE and GLUT1 pathways play a significant role in this modulation.

Shengui Sansheng San extraction is an angiogenic switch via regulations of AKT/mTOR, ERK1/2 and Notch1 signal pathways after ischemic stroke

BACKGROUND As a traditional Chinese herbal formula, Shengui Sansheng San (SSS) has been employed for stroke treatment more than 300 years. PURPOSE We hypothesize that SSS extraction is an angiogenic switch in penumbra post-stroke, and corresponding mechanisms are investigated. METHODS In present study, rats were subjected to permanent middle cerebral artery occlusion model (MCAo) and were treated with low, middle and high doses of SSS extraction. We assessed neurological function and survival rate, and measured infarct volume by 2,3,5-triphenyltetrazolium chloride staining on day 7 after ischemia. von Willebrand factor (vWF), stromal cell-derived factor-1 alpha (SDF-1α) /chemokine (C-X-C motif) receptor 4 (CXCR4) axis, vascular endothelial growth factor (VEGF)/VEGF receptor 2 (VEGFR2) as well as protein kinase B (AKT)/mammalian target of rapamycin (mTOR) /hypoxia-inducible factor-1 alpha (HIF-1α), extracellular signal-regulated kinase 1/2 (ERK1/2) and Notch1 signaling pathways were respectively investigated by immunofluorescence assay or western blotting in vivo and oxygen-glucose-deprived (OGD) brain microvascular endothelial cells (BMECs); simultaneously, wound healing of BMECs and tube formation assay were administrated. RESULTS Compared to MCAo group, SSS extraction could significantly improve neurological functional scores, survival rate and cerebral infarct volume, enhance vWF+ vascular density and perimeter, SDF-1α/CXCR4 axis, VEGF expression, as well as activate AKT/mTOR/HIF-1α and ERK1/2 and inhibit Notch1 pathways in penumbra. In vitro, containing SSS extraction serum increased BMEC migration, capillary formation and VEGF expression via up-regulations of AKT/mTOR and ERK1/2 pathways in OGD BMECs, but ERK inhibitor (U0126) reversed the result of VEGF expression in high dose of SSS group. Additionally, VEGFR2 and Notch1 expressions were suppressed by containing SSS extraction serum. All results were in dose dependent manner. CONCLUSION Our study firstly demonstrates that SSS extraction is an angiogenic switch. Due to suppressed VEGFR2/Notch1 cascades and activated AKT/mTOR and ERK1/2 signals in BMECs, a feedback loop of angiogenic homeostasis is established. Furthermore, the comprehensive mediations of SDF-1α/CXCR4 axis, AKT/mTOR/HIF-α, ERK1/2 and Notch1 pathways in penumbra contribute to the improvements of neurological function, survival rate and infarct volume post-stroke.