ABSTRACT
OBJECTIVE@#To explore the function and mechanism of microRNA-155 to regulate the angiogenesis after the cerebral infarction of rats through the angiotensin II receptor 1 (AT1R)/vascular endothelial growth factor (VEGF) signaling pathway.@*METHODS@#Female SD rats were chosen for the construction of cerebral infarction model of rats using the modified right middle cerebral artery occlusion. The real-time PCR (RT-PCR) method was employed to detect the expression of microRNA-155 in each group at different time points after the cerebral infarction (1 h, l d, 3 d and 7 d). SD rats were randomly divided into four groups (n = 20 rats): sham operation group (Sham group), MACO group, MACO+microRNA-155 mimic group, and MACO+microRNA-155 inhibitor group. Sham group was given the free graft, while MACO+microRNA-155 mimic group and MACO+microRNA-155 inhibitor group were treated with microRNA-155 mimic and microRNA-155 inhibitor respectively. The Zea Longa 5-point scale was used to score the neurologic impairment of rats in each group; 2, 3, 5-triphenyl tetrazolium chloride staining to evaluate the volume of cerebral infarction of rats in each group; the immunohistochemistry to detect the expression of CD31; Western blot and RT-PCR to detect the expression of AT1R and VEGF receptor 2 (VEGFR2).@*RESULTS@#The expression of microRNA-155 was increased in the cerebral ischemia tissue after the cerebral infarction. It was significantly increased at 1 d of ischemia and maintained at the high level for a long time. Rats in the Sham group had no symptom of neurologic impairment, while rats in the MACO group had the obvious neurologic impairment. After being treated with microRNA-155 inhibitor, the neural function of MACO rats had been improved, with the decreased area of cerebral infarction. But after being treated with microRNA-155 mimic, the neural function was further worsened, with the increased area of cerebral infarction. Results of immunohistochemical assay indicated that microRNA-155 inhibitor could up-regulate the expression of CD31, while microRNA-155 mimic could down-regulate the expression of CD31. The RT-PCR found that, after being treated with microRNA-155 inhibitor, MACO rats had the increased expression of AT1R and VEGFR2 messenger RNA (mRNA); but after being treated with microRNA-155 mimic, the expression of AT1R and VEGFR2 mRNA was decreased. Results of Western blot showed that, after being treated with microRNA-155 inhibitor, MACO rats had the increased expression of AT1R and VEGFR2 mRNA; but after being treated with microRNA-155 mimic, the expression of AT1R and VEGFR2 mRNA was decreased.@*CONCLUSIONS@#The inhibition of microRNA-155 can improve the neurologic impairment of rats with the cerebral infarction, reduce the volume of cerebral infarction and effectively promote the angiogenesis in the region of ischemia, which may be mediated through AT1R/VEGFR2 pathway.
ABSTRACT
Objective: To explore the function and mechanism of microRNA-155 to regulate the angiogenesis after the cerebral infarction of rats through the angiotensin II receptor 1 (AT1R)/vascular endothelial growth factor (VEGF) signaling pathway. Methods: Female SD rats were chosen for the construction of cerebral infarction model of rats using the modified right middle cerebral artery occlusion. The real-time PCR (RT-PCR) method was employed to detect the expression of microRNA-155 in each group at different time points after the cerebral infarction (1 h, l d, 3 d and 7 d). SD rats were randomly divided into four groups (n = 20 rats): sham operation group (Sham group), MACO group, MACO+microRNA-155 mimic group, and MACO+microRNA-155 inhibitor group. Sham group was given the free graft, while MACO+microRNA-155 mimic group and MACO+microRNA-155 inhibitor group were treated with microRNA-155 mimic and microRNA-155 inhibitor respectively. The Zea Longa 5-point scale was used to score the neurologic impairment of rats in each group; 2, 3, 5-triphenyl tetrazolium chloride staining to evaluate the volume of cerebral infarction of rats in each group; the immunohistochemistry to detect the expression of CD31; Western blot and RT-PCR to detect the expression of AT1R and VEGF receptor 2 (VEGFR2). Results: The expression of microRNA-155 was increased in the cerebral ischemia tissue after the cerebral infarction. It was significantly increased at 1 d of ischemia and maintained at the high level for a long time. Rats in the Sham group had no symptom of neurologic impairment, while rats in the MACO group had the obvious neurologic impairment. After being treated with microRNA-155 inhibitor, the neural function of MACO rats had been improved, with the decreased area of cerebral infarction. But after being treated with microRNA-155 mimic, the neural function was further worsened, with the increased area of cerebral infarction. Results of immunohistochemical assay indicated that microRNA-155 inhibitor could up-regulate the expression of CD31, while microRNA-155 mimic could down-regulate the expression of CD31. The RT-PCR found that, after being treated with microRNA-155 inhibitor, MACO rats had the increased expression of AT1R and VEGFR2 messenger RNA (mRNA); but after being treated with microRNA-155 mimic, the expression of AT1R and VEGFR2 mRNA was decreased. Results of Western blot showed that, after being treated with microRNA-155 inhibitor, MACO rats had the increased expression of AT1R and VEGFR2 mRNA; but after being treated with microRNA-155 mimic, the expression of AT1R and VEGFR2 mRNA was decreased. Conclusions: The inhibition of microRNA-155 can improve the neurologic impairment of rats with the cerebral infarction, reduce the volume of cerebral infarction and effectively promote the angiogenesis in the region of ischemia, which may be mediated through AT1R/VEGFR2 pathway.
ABSTRACT
Objective To study the characteristics of a new neuron-specific marker in the cerebral cortex and its application method.Methods According to immunofluorescence principles,the neurons in the cerebral cortex cultured in vitro were stained by fluorescein isothiocyanate (FITC)-labeled Tet1; and FITC-Tet1 with NSE or GFAP was employed to determine whether the neurons can be marked and whether they can be distinguished from astrocytes; the conditions of application of FITC-Tet1 were quantified.The HEK and C6 cell lines were performed the above experiments as controls.Results The final concentration was the key factor,which indicated that when the FITC-Tet1 was about 30 μg/mL,the neuron marking ability was the best,and the astrocytes,HEK and C6 were not stained.Neither high (100μg/mL) or low (10μg/mL) levels of FITC-Tet1 were suitableforstainingneurons.Conclusion Tet1 can specifically combine with the neuron in the cerebral cortex,which may make it to be a new staining method or a targeting molecule to neurons in the central nervous system.