1.The change of cellular repressor of E1A-stimulated genes during adventitial fibroblast phenotypic switching
Yang LI ; Yaling HAN ; Chenghui YAN
Chinese Journal of Interventional Cardiology 2014;(8):509-514
Objective To explore possible role of cellular repressor of E1A-stimulated genes(CREG) in the process of phenotypic switching of adventitial fibroblasts(AFs). Methods Immunofluorescent staining was performed with tissue sections from mouse carotid arteries to evaluate the relationship between the expression of CREG and smooth muscle actin-α(α-SMA) in injured arteries, especially in the adventitia. Tissue block pasted culture method was used to isolate and culture AFs. RT-PCR and Western-blot were used to detect the change of CREG andα-SMA mRNA and protein expression in AFs in the presence of different concentrations of AngⅡfor 12 h/24 h or in the presence of 100 nmol/L Ang Ⅱ for different times. Results Normal mouse carotid arteries had little α-SMA expression throughout the tunica adventitia. Arteries at day 1 and day 3 post-injury exhibited significantly higher immunofluorescence of α-SMA compared with non-injured arteries. Alpha-SMA expression began to decrease on day 7 and progressively declined on day 14. In contrast, immunofluorescent staining revealed that CREG was expressed in the adventitia of normal arteries. Expression of CREG in the adventitia of injured arteries was decreased on the 1st day, reached its lowest value on the 3rd day, and increased gradually from the 7th day, and was higher compared with that in non-injured arteries on the 14th day after injury. Similarly, the expression of CREG in AFs was very high, and AngⅡremarkably decreased mRNA and protein expression levels of CREG in a dose-dependent and time-dependent manner. Conclusions The changes in CREG expression correlate with AF phenotypic modulation, and CREG down-regulation may facilitate AF phenotypic switching into myofibroblasts (MFs).
2.Bioinformatic analysis of the cellular repressor of E1A-stimulated genes proximal promoter and primary identification of transcriptional regulation function
Yaling HAN ; Xin ZHAO ; Chenghui YAN
Chinese Journal of Practical Internal Medicine 2000;0(12):-
Objective To analyze the human cellular repressor of E1A-stimulated genes (hCREG)using bioinformatics tools and predict the promoter position and transciption factor binding sites.Methods Complete coding hCREG sequences were obtained from Genbank.hCREG was analyzed with First EF software in order to obtain the promoter sequence of hCREG.Moreover,transcription factor binding sites of hCREG was convinced by Motif software.Subsequently,the transcription factor of hCREG by bioinformatics tools was related between hCREG and smooth muscle cells differentiation observed by both Western blot and immunoflourescence.Results The promoter of hCREG was located in -109~-359 bp of up-stream of transcriptional site,which was predicted with 945bp length.Transciption factor binding sites of hCREG was predicted by Motif software which was found with 80 transciption factors.The expression of hCREG and smooth muscle ?-actin(SM ?-actin)increased in HITASY after 72 hours with serum deprivation detected by both Western blot and immunoflourescence.Meanwhile,the results showed that the expression of wtp53 increased significantly.These results suggested that transcription factor wtp53 might regulate the expression of hCREG and promoted dedifferentiated phenotype of VSMCs.Conclusion The transcriptional information of the proximal promoter of hCREG obtained.As up-stream regulational factor of hCREG,wtp53 can up-regulate the expression of hCREG and may play a vital role in the process of VSMCs differentiation and phenotype modulation.
3.Effects of Moldavica Total Flavone on Vascular Endothelial Growth Factor, Intercellular Adhesinon Molecule-1 and Other Cytokines in Bronchial Asthmatic Rats
Xiaolong KANG ; Chenghui HE ; Jianguo XING ; Lili YAN
Chinese Journal of Information on Traditional Chinese Medicine 2013;(9):34-36
Objective To investigate the effects of moldavica total flavone on vascular endothelial growth factor (VEGF), intercellular adhesion molecule-1 (ICAM-1), nitric oxide (NO) and endothelin-1 (ET-1) in ovalbumin-induced bronchial asthmatic rats, and explore its mechanism. Methods A total of 60 SD rats was randomly divided into control group, model group, dexamethasone (1 mg/kg) group and moldavica total flavone low (90 mg/kg), medium (180 mg/kg) and high (360 mg/kg) dose group. Rats asthma model was established by ovalbumin challenge methods except the control group. After modeling, rats in control and model groups were intragastrically given distilled water, rats in medicine groups were intragastrically given moldavica total flavone and dexamethasone for 30 d. VEGF in lung tissue and ICAM-1 in bronchoalveolar lavage fluid (BALF) were detected respectively by ELISA. The content of ET-1 and NO in BALF were analyzed with radioimmunity method and nitric acid reductase method. Results The VEGF in lung tissue and ICAM-1, NO and ET-1 in BALF were decreased by moldavica total flavone at 180 and 360 mg/kg (P<0.05, P<0.01) in ovalbumin-induced bronchial asthmatic rats. Conclusion Moldavica total flavone is effective in adjusting abnormal increase of VEGF, ICAM-1, ET-1 and NO of bronchial asthma rats.
4.A MORPHOLOGICAL STUDY OF THE EFFECT OF ACUTE EXPERIMENTAL ISCHEMIA ON THE PYRAMIDAL CELLS OF CANINE CEREBRAL CORTEX
Shuyuan LIU ; Xikai XIONG ; Yan ZHANG ; Chenghui LI ;
Acta Anatomica Sinica 1989;0(S1):-
In the present study, 10 healthy adult dogs were used,8 for the experimental group and 2 for the normal group. We observed the histochemical and ultrastrac- tural changes of the enzymes in the pyramidal cells in the cerebral cortex under the condition of acute ischemia. The result demonstrated that the activity of LDH, AcP and AchE was increased, while the activity of SDH, MAO, ATP ase was decreased. The ultrastructural chnges showed that in the acute ischemia gromp, there were enlargement perinucleus spaces of neurone, brisement, disappearance and vasicularization of the mitochondrial crista. This indicated that acute ischemia had an obvious effect on the histochcmistry of enzymes and the ultra- structure of the neurone in the cerebral cortex.
5.Interactions between the recombinant human CREG protein and cathepsins and M6P/IGFIIR
Mingyu SUN ; Chenghui YAN ; Xiaoxiang TIAN ; Yang LI ; Yaling HAN
Chinese Journal of Tissue Engineering Research 2015;(37):5961-5965
BACKGROUND:It has been found that cel ular repressor of E1A-stimulated genes (CREG) is a lysosomal protein binding directly to the mannose-6-phosphate (M6P)/insulin-like growth factor II receptor (IGFIIR) and depends on the interaction with M6P receptors for efficient delivery to lysosomes OBJECTIVE:To study the interactions between the exogenous CREG protein and cathepsins and M6P/IGFIIR and to confirm the effect of CREG protein on expression and distribution of M6P/IGFIIR. METHODS:Double-stained immunofluorescence and coimmunoprecipitation were applied to observe the interactions between the exogenous CREG protein and cathepsin B, cathepsin L and M6P/IGFIIR. Using gain-of-function and loss-of-function approaches, the effect of CREG on expression and distribution of M6P/IGFIIR were studied by western blot assay and immunofluorescence staining. RESULTS AND CONCLUSION:Double-stained immunofluorescence and coimmunoprecipitation analyses confirmed the direct interactions between the exogenous CREG protein and cathepsin B, cathepsin L and M6P/IGFIIR. It was verified that CREG plays a critical role not in the expression but in the distribution of M6P/IGFIIR using gain-of-function and loss-of-function approaches. These findings provide evidence that exogenous CREG protein is located in lysosomes and has interactions with cathepsins and M6P/IGFIIR, also CREG plays a critical role in the distribution of M6P/IGFIIR.
6.Influence of Different Dietary Salt Concentration on the Renal Fibrosis and Macrophages Infiltration in Salt Sensitive Hypertension
Meili LIU ; Dan LIU ; Yanxia LIU ; Xiaoxiang TIAN ; Chenghui YAN
Progress in Modern Biomedicine 2017;17(27):5214-5218
Objective:To investigate the influence of different salt concentration on the renal fibrosis and macrophages infiltration in salt sensitive hypertension.Methods:Dahl salt sensitive rats were randomly divided into the normal salt (0.3 % nacl) group,4 % high salt,8 % high salt groups at six weeks continuously feeding for 8 weeks,each group contained 15 rats.Tail-cuffmethod was used to value rat blood pressure at 8 weeks,Masson trichromatic method was used to detect renal fibrosis of the three groups at 8 week.Immunohistochemistry and Western blot method were used to depict the renal macrophage infiltration at 8 week.Results:1) The blood pressure of 4 % salt and 8% high salt group rats were significantly higher than those of the normal salt group at 8week,meanwhile the blood pressure of 8 % high salt was further increased than that of 4 % high salt group at 8 week.2) The relative kidney weight and renal fibrosis of 4 % salt and 8 % high salt group rats were obviously higher than that of normal salt group at 8week,meanwhile the relative kidney weight and renal fibrosis of 8 % high salt were further increased than those of 4 % high salt group at 8 week.3) The macrophage infiltration of 4 % salt and 8% high salt group rats were higher than that of the normal salt group at 8week,meanwhile the macrophage infiltration of 8 % high salt was further increased than that of 4 % high salt group at 8 week.Conclusion:Different high salt concentrations had different effect on the renal fibrosis and macrophage infiltration in the salt sensitive hypertension,high salt concentration could exacerbate the renal fibrosis and macrophage infiltration.
7.Effect of Moldavica Total Flavone on Pulmonary Function in Ovalbumin-induced Bronchial Asthmatic Rats
Xiaolong KANG ; Mingda YAO ; Chenghui HE ; Lili YAN
Chinese Journal of Information on Traditional Chinese Medicine 2013;(12):24-25,28
Objective To investigate the effect of moldavica total flavone on pulmonary function and collagen type Ⅲ in lung tissue of ovalbumin-induced bronchial asthmatic rats. Methods A total of 60 SD rats were randomly divided into control group, model group, dexamethasone group, and moldavica total flavone low, medium and high dose group. Rat asthma model was established by ovalbumin challenge methods except the control group. The control and model group were intragastrically given distilled water, medicine groups were intragastrically given moldavica total flavone and dexamethasone for 30 d, once per day. After the last administration, the pulmonary function and airway responsiveness including breathing frequency (F), minute ventilation (MVb), peak inspiratory flow (PIFb), peak expiratory flow (PEFb) and enhanced pause (Penh) were measured by using non-invasive measurement system (Penh system, Buxco, USA). The collagen type Ⅲ in lung tissue were detected by ELISA. Results The moldavica total flavone at 180 and 360 mg/kg decreased F and Penh (P<0.05, P<0.01), increased MVb, PIFb and PEFb (P<0.05), and decreased collagen type Ⅲ in lung tissue (P<0.05, P<0.01). Conclusion Moldavica total flavone can relieve the airway hyperresponsiveness and remodeling, improve respiratory function of ovalbumin-induced bronchial asthmatic rats.
8.Potassium antagonizes damage of coronary artery induced by high salt in-take
Yujie ZHANG ; Chenghui YAN ; Nan ZHU ; Xiaolin ZHANG ; Xin ZHAO ; Yaling HAN
Chinese Journal of Pathophysiology 2014;(6):1010-1016
AIM:To investigate the effect of potassium treatment on coronary arterial impairment induced by high salt intake.METHODS:Sprague-Dawley rats (4-week-old, n=10 in each group) received distilled water (NS), water containing 1.5%NaCl (HS), or 1.5% NaCl and 0.5% KCl (HS+HP) for 16 weeks.Systolic blood pressure (SBP) was determined by tail plethysmography every 2 weeks.After 16 weeks of treatment, vascular remodeling, superox-ide production, malondialdehyde (MDA) content, and endothelial nitric oxide synthase (eNOS) and gp91 expression in the coronary arteries were detected .RESULTS:After 16 weeks of salt loading , the rats in HS group was divided into salt sensitive subgroup and salt resistance subgroup according to the tail-cuff blood pressure .In this experiment , the salt-sensi-tive rats were selected as HS group .In HS group, salt loading significantly increased SBP , serum MDA and gp91 expres-sion, decreased serum NO and eNOS expression in the coronary arteries , and induced the coronary artery remodeling com-pared with NS group .In salt-loaded SD rats , 16-week potassium treatment abrogated the effects induced by salt loading . CONCLUSION:High salt may affect structural and functional changes in coronary arteries by activating oxidative stress . Potassium treatment antagonizes the effect of high salt intake .
9.Effects of chemokine CCL2 on the p38MAPK-HSP27 pathway in the platelets
Yu CAO ; Xiaolin ZHANG ; Dan LIU ; Xiaoxiang TIAN ; Yi LI ; Chenghui YAN ; Yaling HAN
Medical Journal of Chinese People's Liberation Army 2017;42(5):407-412
Objective To investigate whether chemokine CC motif 2 (CCL2) is involved in the high residual platelet response,and the mechanism of CCL2 being involved in the regulation of platelets.Methods Forty patients with ST elevation myocardial infarction (STEMI) were admitted.P2Y12 reaction unit (PRU) was detected by VerifyNow.Forty patients were divided into high platelet reactivity group (high reactivity group,n=24) and normal platelet reactivity group (normal reactivity group,n=16) according to the results of PRU detection.Plasma CCL2 concentration of the STEMI patients was examined by ELISA.The expressions of CCL2 and CCR2 in the platelets were detected by Western blotting.After CCL2 stimulation,the kinases of which phosphorylation was changed in the platelets were screened by ARY003B protein chips.The phosphorylation of p38MAPK and HSP27 in the platelets was tested by Western blotting after CCL2 stimulation in the presence or absence of CCR2 antagonist (RS 102895) or p38MAPK signal pathway inhibitor (SB 203580).Results The plasma CCL2 concentration of high reactivity group was markedly higher than that of normal reactivity group.Moreover,compared with normal reactivity group,the expressions of CCL2 and CCR2 in the platelets of high reactivity group significantly increased.After the platelets were stimulated by CCL2,the phosphorylation of p38α and HSP27 enhanced in the platelets by protein chips screening.When RS 102895 or SB 203580 was treated before CCL2 stimulation,the phosphorylation of p38MAPK and HSP27 decreased.Conclusions CCL2 participates in high residual platelet response in an autocrine/paracrine way.CCL2/CCR2 might affect the function ofplatelets through p38MAPKHSP27 signal pathway.
10.Retrovirus-mediated cellular repressor of E1A-stimulated genes inhibits neointima formation in the rat carotid artery after balloon injury
Yaling HAN ; Jie DENG ; Ming LIANG ; Jian KANG ; Haiwei LIU ; Hongmei XU ; Chenghui YAN
Chinese Journal of Pathophysiology 2000;0(10):-
AIM: To evaluate the effects of over-expression of cellular repressor of E1A-stimulated genes(CREG) mediated by retrovirus on neointima formation in injured rat carotid.METHODS: The pluronic F127 containing pLNCX/CREG or pLNCX/GFP retroviral vectors was placed around the injured rat carotid.The neointima,media areas and the intima to media ratio were calculated.Expressions of CREG,SM ?-actin and Ki-67 were detected.RESULTS: The GFP expression was observed at day 2 in pLNCX/GFP groups.The expression of exogenous CREG was also significantly increased in arteries at day 2 after pLNCX-CREG infection.Over-expression of CREG significantly suppressed neointima formation,attenuated the expression of Ki-67 and up-regulated SM ?-actin expression.CONCLUSION: Over-expression of CREG inhibits VSMCs proliferation and promotes VSMCs differentiation after vascular injury.It suggests that modulation of CREG expression or activity may be a viable approach to treat neointimal restenosis after percutaneous coronary intervention.