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1.
Journal of Medical Postgraduates ; (12): 587-591, 2020.
Article in Chinese | WPRIM | ID: wpr-821832

ABSTRACT

ObjectiveNano-graphene oxide quantum dots (GOQDs) can be used to target fluorescent markers. The stem cell labeling is an important method in studying stem cell treatments. Our study aims to explore the possibility of using GOQDs as living cell fluorescent marker materials for human periodontal ligament stem cells (hPDLSCs), and to evaluate the biosecurity and effect as live cell fluorescence markers of GOQDs.Methods GOQDs were testified by TEM, DLS, UV-vis, and PL spectra. hPDLSCs were obtained by tissue cultivation and separated by single cell-derived colony selection. Then the source of the cells was carried out by immunocytochemical staining of anti-vimentin, anti-cytokeratin, and multipotent differentiation was used in the identification of stem cells. hPDLSCs were incubated with different concentrations of GOODs (0, 10, 25, and 50 μg/mL) for 24h and 72 h. Cytotoxicity and proliferation effects were determined using CCK-8, and cell cycles were detected using flow cytometry after the co-culture of GOQDs and hPDLSCs. The fluorescent labeling effect of GOQDs was tested using laser scanning confocal microscopy.ResultsThe characterization of GOQDs showed that the nanoparticles were evenly dispersed in water and showing blue light at 365 nm. TEM and DLS showed GOQDs had good dispersion, and the particle size was (6.36±1.41) nm. Immunocytochemical staining of anti-vimentin was positive while anti-cytokeratin was negative. The results of cytotoxicity showed there were no significant differences in cell activity after incubated with different concentrations of GOODs (0, 5, 10, 25, 50, 100, 200, and 400 μg/mL) (P>0.05), and there was no significant decrease in cell activity between 24h and 72h (P>0.05). There was no significant difference in the proportional distribution of G1, G2, and S phases between the two concentrations of GOQDs (0 μg/mL and 50 μg/mL) (P>0.05). Fluorescent images showed that GOQDs could enter the cell membrane and increase the fluorescence intensity at the concertation of 50 μg/mL.ConclusionGOQDs were confirmed to have good biocompatibility and could be used for live cell labeling of hPDLSCs.

2.
Journal of Medical Postgraduates ; (12): 587-591, 2020.
Article in Chinese | WPRIM | ID: wpr-821813

ABSTRACT

ObjectiveNano-graphene oxide quantum dots (GOQDs) can be used to target fluorescent markers. The stem cell labeling is an important method in studying stem cell treatments. Our study aims to explore the possibility of using GOQDs as living cell fluorescent marker materials for human periodontal ligament stem cells (hPDLSCs), and to evaluate the biosecurity and effect as live cell fluorescence markers of GOQDs.Methods GOQDs were testified by TEM, DLS, UV-vis, and PL spectra. hPDLSCs were obtained by tissue cultivation and separated by single cell-derived colony selection. Then the source of the cells was carried out by immunocytochemical staining of anti-vimentin, anti-cytokeratin, and multipotent differentiation was used in the identification of stem cells. hPDLSCs were incubated with different concentrations of GOODs (0, 10, 25, and 50 μg/mL) for 24h and 72 h. Cytotoxicity and proliferation effects were determined using CCK-8, and cell cycles were detected using flow cytometry after the co-culture of GOQDs and hPDLSCs. The fluorescent labeling effect of GOQDs was tested using laser scanning confocal microscopy.ResultsThe characterization of GOQDs showed that the nanoparticles were evenly dispersed in water and showing blue light at 365 nm. TEM and DLS showed GOQDs had good dispersion, and the particle size was (6.36±1.41) nm. Immunocytochemical staining of anti-vimentin was positive while anti-cytokeratin was negative. The results of cytotoxicity showed there were no significant differences in cell activity after incubated with different concentrations of GOODs (0, 5, 10, 25, 50, 100, 200, and 400 μg/mL) (P>0.05), and there was no significant decrease in cell activity between 24h and 72h (P>0.05). There was no significant difference in the proportional distribution of G1, G2, and S phases between the two concentrations of GOQDs (0 μg/mL and 50 μg/mL) (P>0.05). Fluorescent images showed that GOQDs could enter the cell membrane and increase the fluorescence intensity at the concertation of 50 μg/mL.ConclusionGOQDs were confirmed to have good biocompatibility and could be used for live cell labeling of hPDLSCs.

3.
Article in Chinese | WPRIM | ID: wpr-701073

ABSTRACT

AIM:To investigate the role of hydrogen molecule on apoptosis-related proteins in glomerular me-sangial cells cultured with high glucose and to explore its possible mechanism.METHODS:Mouse glomerular mesangial cells cultured in vitro were divided into 4 groups:normal control group(C group,5.5 mmol/L glucose),mannitol group(G group,5.5 mmol/L glucose+19.5 mmol/L mannitol),high glucose group(H group,25 mmol/L glucose),high glu-cose+hydrogen-rich water group(HH group,25 mmol/L glucose+hydrogen-rich water),and cultured for 48 h.The pro-tein levels of Bax,Bcl-2,cleaved caspase-3,nuclear factor E2-related factor-2(Nrf2),heme oxygenase-1(HO-1)and NAD(P)H:quinone oxidoreductase-1(NQO-1)were determined by Western blot ,and the mRNA expression of HO-1 and NQO-1 was determined by RT-PCR.The level of intracellular reactive oxygen species(ROS)was detected by dihydro-ethidium method,and the activity of superoxide dismutase(SOD)was measured by WST-8 assay.RESULTS:Compared with C group,the protein levels of Bax and cleaved caspase-3 were up-regulated,and Bcl-2 was down-regulated in H group(P <0.05).No significantly difference of the protein levels mentioned above between C and HH group was observed. Compared with H group,the protein levels of Bax and cleaved caspase-3 were down-regulated,and Bcl-2 was up-regulated in HH group(P <0.05).The level of intracellular ROS was higher and the activity of SOD was lower in H group than those in C group(P<0.05).However,there was no difference of the SOD activity between C group and HH group.The level of intracellular ROS decreased and the activity of SOD increased in HH group as compared with H group(P<0.05). Compared with C group,clearly reduced protein expression of Nrf2,HO-1 and NQO-1,and decreased mRNA expression of HO-1 and NQO-1 in H group were observed(P<0.05).Compared with H group,the protein levels of Nrf2,HO-1 and NQO-1 as well as the mRNA levels of HO-1 and NQO-1 were obviously increased in HH group(P<0.05 ).CONCLU-SION:Hydrogen molecule inhibits the expression of pro-apoptotic proteins and induces the expression of anti-apoptotic pro-teins in glomerular mesangial cells cultured with high glucose.The mechanism may be related to activation of Nrf 2 signaling pathway.

4.
Article in Chinese | WPRIM | ID: wpr-773750

ABSTRACT

OBJECTIVE@#To observe the pulmonary vascular remodeling in rats with pulmonary hypertension induced by hypoxia and hypercapnia, and to explore the role of endoplasmic reticulum stress in pulmonary hypertension.@*METHODS@#Forty SD rats were random-ly divided into four groups:normoxic control group (N), hypoxia hypercapnia group (HH), ERS inhibitor 4-phenylbutyric acid group (4-PBA), endoplasmic reticulum stress (ERS) pathway agonist tunicamycin group (TM), ten rats in each group.The mean pulmona-ry artery pressure (mPAP), mean carotid artery pressure (mCAP) and right ventricular hypertrophy index of rats in each group were measured.Pulmonary artery smooth muscle cells were identified by immunofluorescence α-smooth muscle actin (α-SMA).Morphologi-cal changes of lung tissue and pulmonary artery were observed by electron microscope.The apoptotic index of pulmonary artery smooth muscle cells in each group was detected by TUNEL.Reverse transcription polymerase chain reaction (RT-PCR) and Western blot were used to detect the expression of glucose-regulated protein (GRP78), C/EBP homologous protein (CHOP), c-Jun N-terminal kinase (JNK) and cysteinyl aspartate specific proteinase-12 (caspase-12) mRNA and protein in each group.@*RESULTS@#①Compared with the N group, the mPAP, the ratio of right ventricle weight to left ventricle plus ventricular septum weight[RV/(LV+S)]and the ratio of pulmonary artery wall area to total tube area (WA/TA) were increased (<0.01), and the ratio of pulmonary artery luminal area to total tube area (LA/TA) were decreased (<0.01), pulmonary artery smooth muscle cell apoptosis index were decreased (<0.05 or <0.01) in HH group, 4-PBA group and TM group.ERS related protein and mRNA expressions were increased, the differences were statistically significant.②Compared with the HH group, the mPAP, [RV/(LV+S)]and WA/TA of 4-PBA group were decreased ( <0.01), LA/TA and pulmonary artery smooth muscle cell apoptosis index were increased (<0.01, <0.05).The expressions of ERS related protein and mRNA were all decreased (<0.05 or <0.01).③Compared with the HH group, the mPAP, [RV/(LV+S)]and WA/TA of TM group were increased (<0.05 or <0.01), pulmonary artery middle layer thickened, LA/TA and pulmonary artery smooth muscle cell apoptotic index were decreased (<0.01).ERS related protein and mRNA expressions were increased with statistical significance except GRP78 protein.@*CONCLUSIONS@#Pulmonary vascular remodeling in rats with pulmonary hypertension induced by hypoxia and hypercapnia may be related to the excessive proliferation of pulmonary artery smooth muscle cells and too little apopto-sis;ERS related factors (JNK, caspase-12 and CHOP) are involved in the regulation of pulmonary hypertension induced by hypoxia hypercapnia.


Subject(s)
Animals , Endoplasmic Reticulum Stress , Hypercapnia , Hypertension, Pulmonary , Hypoxia , Pulmonary Artery , Rats , Rats, Sprague-Dawley
5.
Chinese Journal of Pathophysiology ; (12): 1945-1950, 2017.
Article in Chinese | WPRIM | ID: wpr-667670

ABSTRACT

AIM:To investigate the protective effect of non-mitogenic fibroblast growth factor 1 (nFGF1) on the aortic vascular function in streptozotocin (STZ)/high-fat diet (HFD)-induced type 2 diabetic rats and its underlying mechanisms. METHODS:Five-week-old male SD rats (n=30) were randomly divided into 3 groups (n=10 in each group),including normal control group,type 2 diabetic group and nFGF1 treatment group(type 2 diabetic rats were intra-peritoneally injected with 0.5 mg/kg nFGF1 every other day for 4 weeks). After the rats were sacrificed, blood glucose, cholesterol and triglyceride levels,aorta diastolic function and superoxide dismutase(SOD) level in the aorta of each group were measured. Besides,the protein levels of cyclooxygenase-2(COX-2),phosphorylated extracellular signal-regulated ki-nase (p-ERK) and endothelial nitric oxide synthase (eNOS) in the aorta were determined by Western blot. RESULTS:nFGF1 markedly lowered blood glucose, cholesterol and triglyceride levels, enhanced aorta SOD activity and upregulated protein level of eNOS in the type 2 diabetic rats. Furthermore,the increased protein levels of COX-2 and p-ERK in the type 2 diabetic rats were largely abrogated by nFGF1. CONCLUSION:nFGF1 effectively attenuates aortic vascular dysfunction in the type 2 diabetic rats,which may be associated with decreasing blood glucose,cholesterol and triglyceride levels,re-ducing inflammation and oxidative stress response,and activating eNOS signaling pathway.

6.
Journal of Clinical Pediatrics ; (12): 355-358, 2017.
Article in Chinese | WPRIM | ID: wpr-608502

ABSTRACT

Objective To investigate the characteristics and essential points of diagnosis and treatment of double trisomy 47,XXX/48,XXX,+8 combined Behcet disease, a rare inherited immunodeficiency disorder. Methods The clinical manifestations, karyotype analysis and gene test results of the patients were analyzed, and relevant literatures were reviewed. Results A 11-year-old girl presented repeated fever for more than 6 years, accompanied with recurrent genital herpes infection and oral apthosis, was clinically diagnosed with Behcet disease. Cytogentic and molecular karyotyping on peripheral lymphocytes demonstrated 47,XXX[12]/48,XXX,+8[18]. Conclusions Conventional karyotype analysis and chromosomal microarray analysis have a complementary role in the diagnosis of the disease. We conclude that patients with constitutional trisomy 8 and those with trisomy 8 confined to the bone marrow are both at increased risk of developing features of Behcet disease. The mechanism may relate to increased gene dosage of candidate genes for Behcet's disease on chromosome 8.

7.
Article in Chinese | WPRIM | ID: wpr-243441

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the expression profile of interleuki-1β (IL-1β) in rat myocardium at different time points during hypoxia/reoxygenation(H/R)transition.</p><p><b>METHODS</b>The isolated Langendorff perfused rat heart model was established.Forty SD rats were randomly divided into sham group (A group) and hypoxia/reoxygenation group (H/R group). The H/R group rats were subdivided into H/R 0.5 h group(B group), H/R 1 h group(C group), H/R 2 h group(D group)according to reoxygenation time. The left ventricular development pressure(LVDP), maximal rates of increase/decrease of the left ventricular pressure(±dp/dtmax) were continuously recorded. The concentration of interleukin-1β(IL-lβ) and creatine kinase-MB (CK-MB) in myocardium was measured by ELISA. The mRNA expression of IL-lβ in myocardium was determined by RT-PCR. Microstructure of myocardium was observed under light microscopy.</p><p><b>RESULTS</b>The value of LVDP and ±dp/dtmax in hypoxia/reoxygenation group rat were significantly lower than that in sham group(P < 0.05). The expression of IL-lβ and CK-MB at protein level and the expression of IL-1β at mRNA level in hypoxia /reoxygenation group were higher than that in sham group(P < 0. 05). There were significant differences of the above parameters among H/R 0.5 h, 1 h, 2 h group(P <0.05). The concentration of IL-1β and CK-MB, the mRNA expression of IL-1β were higher in H/R 2 h group than that of other groups(P < 0.05).</p><p><b>CONCLUSION</b>The high expression of IL-Iβ in myocardium after myocardial hypoxia /reoxygenation in rats might lead to. ischemia/reperfusion injury.</p>


Subject(s)
Animals , Creatine Kinase, MB Form , Metabolism , Disease Models, Animal , Hypoxia , Metabolism , Pathology , Interleukin-1beta , Metabolism , Myocardial Ischemia , Metabolism , Myocardium , Metabolism , Pathology , Rats , Rats, Sprague-Dawley
8.
Chinese Journal of Pathophysiology ; (12): 1645-1650, 2014.
Article in Chinese | WPRIM | ID: wpr-456849

ABSTRACT

AIM:To investigate the effects of voltage-dependent K +channel 1.5 (Kv1.5) on the proliferation and apoptosis of rat pulmonary artery smooth muscle cells (PASMCs) under hypoxia+hypercapnia condition and the relation-ship with mitogen-activated protein kinase(MAPK) signal pathway.METHODS:The PASMCs isolated from the male SD rat were cultured under hypoxia +hypercapnia condition, and randomly divided into normal group (N group), hypoxia+hyper-capnia group (HH group), hypoxia+hypercapnia+DMSO incubation group (HD group), hypoxia+hypercapnia +U0126 (an extracellular signal-regulated kinase 1/2 inhibitor) incubation group (HU group), hypoxia+hypercapnia+SB203580 (a p38 mitogen-activated protein kinase inhibitor ) incubation group (HS group), and hypoxia+hypercapnia+anisomycin (an agonist of MAPK) incubation group (HA group).Cell Counting Kit-8 was used to detect the cell viability.The protein expression of Kv1.5, PCNA and Bax was detected by Western blotting .RESULTS:Compared with N group , the cell via-bility and PCNA protein expression in HH group and HD group were significantly raised (P0.05 ) .Compared with HD group , the cell viability and PCNA protein expression in HU group , HS group and HA group were decreased (P<0.05 or P<0.01), but Kv1.5 protein and Bax protein were raised (P<0.01), with the most significant changes in HA group .CONCLUSION:The regulation of Kv1.5 to the proliferation and apoptosis of PASMCs under hy-poxia+hypercapnia condition might have a relationship with the activation of MAPK signal pathway .

9.
Article in Chinese | WPRIM | ID: wpr-313000

ABSTRACT

<p><b>OBJECTIVE</b>To explore the role of Xuebijing Injection (XBJI) in inhibiting inflammatory factors associated with anoxia/reoxygenation myocardial inflammatory response of rats.</p><p><b>METHODS</b>Totally 36 healthy male Sprague-Dawley rats, 280 ± 30 g were randomly divided into six groups, i.e., the normal control group (N group), the balanced perfusion group (BP group),the model group (M group),the low dose XBJI group (XBJI(L) group), the middle dose XBJI group (XBJI(M) group),and the high dose XBJI group (XBJI(H) group), 6 in each group. The myocardial anoxia/reoxygenation rat model was established by Langendorff isolated heart perfusion. The concentration of TNF-α in the myocardial tissue was detected by ELISA. The expression of nuclear factor kappa B p65 (NF-κB p65) protein and Toll like receptor 4 (TLR4) protein were detected using Western blot. The expression of NF-κB p65 mRNA and TLR4 mRNA was detected by RT-PCR. Ultrastructural changes of anoxia-reoxygenation rats' heart muscle were observed under transmission electron microscope.</p><p><b>RESULTS</b>Compared with the M group,the TNF-α concentration, expression levels of NF-κB p65 protein and mRNA, TLR4 protein and mRNA decreased to various degrees in the XBJI(L) group, the XBJI(M) group, and the XBJI(H) group. The TNF-α expression level decreased most significantly in the XBJI(L), group (P < 0.01), while other indices decreased most obviously in the XBJI(M) group (P < 0.01, P < 0.05). Expression levels of NF-κB p65 and TLR4 protein were obviously lower in the XBJI(M) group than in the XBJI(L) group (P < 0.05). There was no statistical difference in other indices among the three XBJI groups (P > 0.05). Myocardial fibers were loose and broken with disappearance of transverse striation, and mitochondrial cristae was dissolved and severely damaged in the M group. The aforesaid condition was improved after treated by XBJI, with the most obvious effect obtained in the XBJI(M) group.</p><p><b>CONCLUSIONS</b>Different doses of XBJI could attenuate inflammatory reactions after myocardial anoxia/reoxygenation rats' heart muscle through inhibiting TLR4-NF-κB-TNF-α signal transduction pathway. The best effect could be obtained by 4 mL/100 mL XBJI.</p>


Subject(s)
Animals , Drugs, Chinese Herbal , Pharmacology , Therapeutic Uses , Hypoxia , Male , Myocardium , Metabolism , Myocytes, Cardiac , NF-kappa B , Metabolism , Oxygen , Metabolism , RNA, Messenger , Rats , Rats, Sprague-Dawley , Signal Transduction , Toll-Like Receptor 4 , Metabolism , Transcription Factor RelA , Tumor Necrosis Factor-alpha , Metabolism
10.
Article in Chinese | WPRIM | ID: wpr-236386

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the role of Xuebijing injection(XBJI, traditional Chinese medicine), in inhibiting TLR4--NF-kappaB--IL-1beta pathway of myocardial hypoxia/reoxygenation in rats.</p><p><b>METHODS</b>Thirty six male SD rats (280 +/- 30) g were randomly divided into six groups (n = 6): normal group (N group), balanced perfusion group (BP group), model group (M group), low dose XBJI group (XBJI(L) group), middle dose XBJI group (XBJI(M) group), high dose XBJI group (XBJI(H) group). By Langendorff isolated heart perfusion device to establish the model of myocardial hypoxia/reoxygenation in rats. ELISA was used to detect the concentration of interleukin-1beta (IL-1beta); Western blot was used to detect the expression of nuclear factor kappa B p65 (NF-kappaB p65) protein and toll like receptor 4 (TLR4) protein; and RT-PCR to determine the expression of NF-kappaB p65 mRNA and TLR4 mRNA;To observe microstructure changes of hypoxia/reoxygenation myocardial by light microscopy.</p><p><b>RESULTS</b>Compared with M group, the IL-1beta concentration, NF-kappaB p65 and TLR4 protein,NF-kappaB p65 and TLR4 mRNA of XBJIL group, XBJI(M) group, XBJI(H) group expression decreased in varying degrees,and decreased most obviously all in XBJI(M) group (P < 0.05, P < 0.01); Myocardical structural damage was serious in M group, and improved after treatment XBJI, the most obvious was the XBJI(M).</p><p><b>CONCLUSION</b>Different dose of XBJI can inhibit TLR4--NF-kappaB--IL-1beta signal transduction pathway and reduce several inflammatory reaction after myocardial hypoxia/reoxygenation injury, the 4 ml/100 ml of XBJI is the best.</p>


Subject(s)
Animals , Drugs, Chinese Herbal , Pharmacology , Heart , Inflammation , Interleukin-1beta , Metabolism , Male , Myocardium , Pathology , RNA, Messenger , Rats , Rats, Sprague-Dawley , Reperfusion Injury , Drug Therapy , Signal Transduction , Toll-Like Receptor 4 , Metabolism , Transcription Factor RelA , Metabolism
11.
Article in Chinese | WPRIM | ID: wpr-236381

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effect of chloride channel blocker--niflumic acid (NFA) on the pathological process of hypoxia hypercapnia-induced pulmonary vasoconstriction in rats.</p><p><b>METHODS</b>We used the model of hypoxia hypercapnia-induced pulmonary vasoconstriction rats, and divided the second, third branch pulmonary artery rings randomly into four groups (n = 8): control group (N group), hypoxia hypercapnia group (H group), DMSO incubation group (HD group), niflumic acid group (NFA group). Under acute hypoxia hypercapnia conditions, we observed the effects of the three stages of hypoxia hypercapnia-induced pulmonary vasoconstriction (HHPV) incubated by NFA in the second, third brach pulmonary artery rings. At the same time, the values of rings' tension changings were recorded via the method of hypoxia hypercapnia conditions reactivity. And investigated the effect of NFA to HHPV.</p><p><b>RESULTS</b>(1) Under the hypoxia hypercapnia condition, we observed a biphasic pulmonary artery contractile (the phase I rapid contraction and vasodilation; the phase II sustained contraction) response in both the second and the third branch pulmonary artery rings compared with the control group (P < 0.05 , P < 0.01); (2) The second and third pulmonary artery rings incubated by NFA which phase II persistent vasoconstriction were significantly attenuated compared with the H group (P < 0.05 , P < 0.01).</p><p><b>CONCLUSION</b>The blocker of the chloride channels attenuates the second and third branch pulmonary artery rings constriction in rat, especially the phase II persistent vasoconstriction, so then have an antagonistic effect on HHPV.</p>


Subject(s)
Animals , Chloride Channels , Hypercapnia , Hypoxia , Niflumic Acid , Pharmacology , Pulmonary Artery , Pulmonary Circulation , Rats , Vasoconstriction
12.
Article in Chinese | WPRIM | ID: wpr-236333

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the role of p38 MAPK on ischemic postconditioning (IPO) attenuating pneumocyte apoptosis after lung ischemia/reperfusion injury (LIRI).</p><p><b>METHODS</b>Forty adult male SD rats were randomly divided into 5 groups based upon the intervention (n = 8): control group (C), LIR group (I/R), LIR + IPO group (IPO), IPO + solution control group (D), IPO + SB203580 group (SB). Left lung tissue was isolated after the 2 hours of reperfusion, the ratio of wet lung weight to dry lung weight (W/D), and total lung water content (TLW) were measured. The histological structure of the left lung was observed under light and electron transmission microscopes, and scored by alveolar damage index of quantitative assessment (IQA). Apoptosis index (AI) of lung tissue was determined by terminal deoxynuleotidyl transferase mediated dUTP nick end and labeling (TUNEL) method. The mRNA expression and protein levels of and Bax were measured by RT-PCR and quantitative immunohistochemistry (IHC).</p><p><b>RESULTS</b>Compared with C group, W/D, TLW, IQA, AI and the expression of Bax of I/R were significantly increased, the expression of Bcl-2 and Bcl-2/Bax were significantly decreased (P < 0.05, P < 0.01), and was obviously morphological abnormality in lung tissue. Compared with I/R group, all the indexes of IPO except for the expression of Bcl-2 and Bcl-2/ Bax were obviously reduced, the expression of Bcl-2 and Bcl-2/Bax were increased (P < 0.05, P < 0.01). All the indexes between D and IPO were little or not significant( P > 0.05). The expression of Bcl-2 and Bcl-2/Bax of SB were significantly increased and other indexes were reduced than those of IPO (P < 0.05, P < 0.01).</p><p><b>CONCLUSION</b>IPO may attenuate pneumocyte apoptosis in LIRI by inactivation of p38 MAPK, up-regulating expression of Bcl-2/Bax ratio.</p>


Subject(s)
Alveolar Epithelial Cells , Cell Biology , Animals , Apoptosis , Disease Models, Animal , Ischemic Postconditioning , Lung , Pathology , Male , Proto-Oncogene Proteins c-bcl-2 , Metabolism , Rats , Rats, Sprague-Dawley , Reperfusion Injury , Pathology , bcl-2-Associated X Protein , Metabolism , p38 Mitogen-Activated Protein Kinases , Metabolism
13.
Article in Chinese | WPRIM | ID: wpr-236309

ABSTRACT

<p><b>OBJECTIVE</b>To investigate whether resveratrol (RES) plays a protective role in hypothermic preserved isolated rat hearts and whether it is mediated by regulation of silent information regulator protein-1 (Sirt-1) expression.</p><p><b>METHODS</b>The Langendorff model of isolated rat heart was used. After stored in different Celsior solution at 4 degrees C for 9 h, SD rat hearts were randomly divided into 7 groups: blank control group;9 h group (soley hypothermic preservation for 9 h); RES group (3, 10, 30 micromol/L RES treatment plus hypothermic preservation for 9 h ), niacinamide (NAM) group (40 micromol/L NAM added in Celsior solution plus hypothermic preservation for 9 h), RES + NAM group (30 micromol/L RES and 40 micromol/L NAM were added in Celsior solution plus hypothermic preservation for 9 h). The morphological changes of cardiomyocytes were detected by the HE staining with the light microscope. The mRNA and protein expression levels of Sirt-1 were detected by Real-Time PCR and Western blot respectively.</p><p><b>RESULTS</b>(1) Compared with the blank control group, myocardiocytes were injured remarkably in the 9 h group and the Sirt-1 mRNA and protein expression levels were decreased significantly (P < 0.01); (2) Compared with the 9 h group, rat myocardial injury was alleviated gradually in 3, 10, 30 micromol/L RES group and the Sirt-1 mRNA and protein expression levels were increased in a dose-dependent manner (P < 0.05); (3) The above protective effects of RES were attenuated by Sirt-1 inhibitor NAM.</p><p><b>CONCLUSION</b>RES can protect myocardiocytes from injury caused by long range hypothermic preservation and this protective effect maybe mediated by upregulation of Sirt-1 expression.</p>


Subject(s)
Animals , Cryopreservation , Heart , Male , Organ Preservation , Rats , Rats, Sprague-Dawley , Sirtuin 1 , Metabolism , Stilbenes , Pharmacology
14.
Acta Physiologica Sinica ; (6): 203-209, 2014.
Article in Chinese | WPRIM | ID: wpr-297500

ABSTRACT

The aim of the present study was to investigate the roles of calcium-activated chloride channels (Cl(Ca)) in the two-phase hypoxic pulmonary vasoconstriction (HPV). The second pulmonary artery branches were dissected from male Sprague-Dawley rats, and the changes in vascular tone were measured by using routine blood vascular perfusion in vitro. The result showed that, under normoxic conditions, Cl(Ca) inhibitors (NFA and IAA-94) significantly relaxed second pulmonary artery contracted by norepinephrine (P < 0.01), but merely had effects on KCl-induced second pulmonary artery contractions. A biphasic contraction response was induced in second pulmonary artery ring pre-contracted with norepinephrine exposed to hypoxic conditions for at least one hour, but no biphasic contraction was observed in pulmonary rings pre-contracted with KCl. NFA and IAA-94 significantly attenuated phase II sustained hypoxic contraction (P < 0.01), and also attenuated phase I vasodilation, but had little effect on phase I contraction. These results suggest that Cl(Ca) is an important component forming phase II contraction in secondary pulmonary artery, but not involved in phase I contraction.


Subject(s)
Animals , Chloride Channels , Physiology , Glycolates , Pharmacology , Hypoxia , Male , Norepinephrine , Pharmacology , Pulmonary Artery , Rats , Rats, Sprague-Dawley , Vasoconstriction , Vasodilation
15.
Acta Physiologica Sinica ; (6): 283-288, 2014.
Article in Chinese | WPRIM | ID: wpr-297491

ABSTRACT

The aim of the present study is to investigate the expressions of ATP-sensitive K(+) channels (KATP) in pulmonary artery smooth muscle cells (PASMCs) and the relationship with p38 MAPK signal pathway in rats. Male SD rat PASMCs were cultured in vitro, and a model of hypoxia and hypercapnia was reconstructed. PASMCs were divided to normal (N), hypoxia-hypercapnia (H), hypoxia-hypercapnia+DMSO incubation (HD), hypoxia-hypercapnia+SB203580 (inhibitor of p38 MAPK pathway) incubation (HS) and hypoxia-hypercapnia+Anisomycin (agonist of p38 MAPK pathway) incubation (HA) groups. Western blot was used to detect the protein expression of SUR2B and Kir6.1; semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) was used to detect the mRNA expression of SUR2B and Kir6.1. The results demonstrated that: (1) Compared with N, H, HD and HS groups, the expressions of Kir6.1 mRNA and protein in PASMCs of HA group were decreased significantly (P < 0.01), but there were no differences among N, H, HD and HS groups (P > 0.05); (2) Compared with N group, the expressions of SUR2B mRNA and protein in H, HD, HS and HA groups were increased significantly (P < 0.05), but there were no differences among H, HD, HS and HA groups (P > 0.05). The results imply that: (1) Hypoxia-hypercapnia, SB203580 didn't change the expressions of Kir6.1 mRNA and protein in PASMCs, but Anisomycin decreased the expressions of Kir6.1 mRNA and protein, so Kir6.1 may be regulated by the other subfamily of MAPK pathway; (2) Hypoxia-hypercapnia raised SUR2B mRNA and protein expressions in PASMCs, but SB203580 and Anisomycin did not affect the changes, so the increasing of SUR2B mRNA and protein induced by hypoxia-hypercapnia may be not depend on p38 MAPK pathway.


Subject(s)
Animals , Anisomycin , Pharmacology , Cell Hypoxia , Cells, Cultured , Hypercapnia , Imidazoles , Pharmacology , KATP Channels , Metabolism , MAP Kinase Signaling System , Male , Myocytes, Smooth Muscle , Metabolism , Pulmonary Artery , Cell Biology , Pyridines , Pharmacology , Rats , Rats, Sprague-Dawley , Sulfonylurea Receptors , Metabolism , p38 Mitogen-Activated Protein Kinases
16.
Article in Chinese | WPRIM | ID: wpr-294336

ABSTRACT

<p><b>OBJECTIVE</b>To explore the effect of curcumin (CUR) on cycteinyl aspirate specific protease-12 (Caspase-12) and pneumocyte apoptosis in pulmonary ischemia/reperfusion (I/R) injury mice.</p><p><b>METHODS</b>The in vivo unilateral in situ pulmonary I/R injury mouse model was established in C57BL/6J mice. Sixty experimental mice were randomly divided into six groups by random digit table, i. e., the sham-operation group (Sham), the I/R group, the I/R + dimethyl sulfoxide group (I/R + DMSO), the I/R + low dose CUR pre-treated group (I/R + CUR-100), the I/R + middle dose CUR pre-treated group (I/R + CUR-150), the I/R + high dose CUR pre-treated group (I/R + CUR-200), 10 in each group. Mice were euthanized and their left lungs were excised. Wet lung weight to dry lung weight (W/D) and the total lung water content (TLW) were tested. The morphological changes of the lung tissue were observed and index of quantitative evaluation for alveolar damage (IQA) detected under light microscope. The ultra-microstructure of the lung tissue was observed under electron microscope. The mRNA and protein expression levels of Caspase-12 and glucose regulated protein (GRP78) were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot. Apoptosis index (AI) of the lung tissue was determined by terminal-deoxynucleoitidyl transferase mediated nick end labeling (TUNEL) method.</p><p><b>RESULTS</b>Compared with the Sham group, expression levels of Caspase-12, GRP78 mRNA and protein all significantly increased in the I/R group (P < 0.05); W/D, TLW, IQA, and AI were all notably higher (P < 0.05, P < 0.01); the morphological and ultrastructural injury of the lung tissue were notably observed in I/R group. Compared with the I/R + DMSO group, expression levels of GRP78 mRNA and protein were increasingly higher in the I/R + CUR-100 group, the I/R + CUR-150 group, and the I/R +CUR-200 group (P < 0.05), expression levels of Caspase-12 mRNA and protein were lower (P < 0.05); W/D, TLW, IQA, and AI also decreased (P < 0.05, P < 0.01); the morphological and ultrastructural injury of the lung tissue were gradually alleviated in the I/R + CUR groups.</p><p><b>CONCLUSION</b>CUR had better effect on the lung protection against I/R injury, which might be related to inhibition for pneumocyte apoptosis associated with Caspase-12 in excessive unfolded protein response (UPR).</p>


Subject(s)
Animals , Apoptosis , Caspase 12 , Metabolism , Curcumin , Pharmacology , Heat-Shock Proteins , Metabolism , Lung , Male , Mice , Mice, Inbred C57BL , Reperfusion Injury , Metabolism , Pathology
17.
Article in Chinese | WPRIM | ID: wpr-235369

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the effects of curcumin (CUR) on pneumocyte apoptosis and CCAAT/enhancer binding protein homologous protein (CHOP) in pulmonary ischemia/reperfusion injury (PIRI) in mice.</p><p><b>METHODS</b>Sixty C57BL/6J mice were randomly allocated into six groups (n = 10): Sham operation group (Sham group), ischemia/reperfusion group (I/R group), ischemia/reperfusion + dimethyl sulfoxide group (DMSO group), ischemia/reperfusion + curcumin pre-treated with respectively 100 mg/kg, 150 mg/kg and 200 mg/kg groups (CUR-100 group, CUR-150 group and CUR-200 group). Left lung tissue of each group was excised after reperfusion for 3 h. Wet lung weight to dry lung weight (W/D) and total lung water content (TLW) were tested. The morphological and ultrastructural changes of lung tissue were observed under light microscope and electron microscope, and index of quantitative evaluation for alveolar damage (IQA) was calculated. The expression levels of CHOP and glucose regulated protein 78 (GRP78) were detected by RT-PCR and Western Blot. Apoptosis index (AI) of lung tissue was determined by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL) method.</p><p><b>RESULTS</b>Compared with Sham group, the expression levels of CHOP, GRP78 mRNA and protein were all significantly increased (P < 0.05) in I/R group and DMSO group, W/D, TLW, IQA and AI were all notably higher (P < 0.01); morphological and ultrastructural injury in lung tissue were notably observed in I/R group. Compared with DMSO group, the expression levels of GRP78 mRNA and protein were increased higher (P < 0. 05) in CUR-100 group, CUR-150 group, and CUR-200 group, but the expression levels of CHOP mRNA and protein were decreased lower (P < 0.05), W/D, TLW, IQA and AI were also decreased (P < 0.05, P < 0.01); morphological and ultrastructural injury in lung tissue were gradually alleviated in CUR groups.</p><p><b>CONCLUSION</b>I/R induces excessive unfolded protein response (UPR) in lung tissue, in which CHOP participates in pneumocyte apoptosis, leading to lung injury; CUR has notable effects on lung protection against I/R injury, which may be related to inhibition of apoptosis mediated by CHOP in excessive UPR.</p>


Subject(s)
Alveolar Epithelial Cells , Metabolism , Animals , Apoptosis , Curcumin , Pharmacology , Heat-Shock Proteins , Metabolism , Lung , Metabolism , Pathology , Male , Mice , Mice, Inbred C57BL , Reperfusion Injury , Metabolism , Pathology , Transcription Factor CHOP , Metabolism
18.
Chinese Journal of Virology ; (6): 326-330, 2011.
Article in Chinese | WPRIM | ID: wpr-286034

ABSTRACT

To investigate biological characteristics of the IVpi-189 progeny virus derived from the culture of influenza A virus as a live-attenuated vaccine candidate. Persistent infection of a cultured cell line with influenza A virus (MDCK-IVpi) was established by incubating continuously influenza virus-infected cells at a lower temperature. The infectious progeny virus derived from MDCK-IVpi cells at the 189rd subculture was designated as the IVpi-189 strain of influenza virus. The cytopathic effect induced by IVpi-189 virus was observed under different temperature conditions. The production of infectious progeny virus was examined at 38 and 32 degrees C by plaque titration of cell-associated and released virus. IVpi-189 virus showed cytopathic effect as strong as that of IVwt in infected cell line of MDCK at 32 degrees C. However, when culture temperature was raised to 38 degrees C, the cytopathic effect induced by IVpi-189 virus was delayed and less pronounced. Virus growth in IVpi-189 virus-infected cells at 38 degrees C was significantly reduced as compared with that of IVwt virus, although both viruses yielded nearly equivalent high titers of cell-associated and released virus at 32 degrees C. The reasons of the decreased proliferative ability of IVpi-189 virus at high culture temperature were unrelated with virus inactivation or the release of progeny virus, but associated with the decreased replication of infectious progeny virus in the infected cells. IVpi-189 virus derived from MDCK cells infected persistently with influenza A virus showed biological characteristics as a potential live-attenuated vaccine candidate.


Subject(s)
Animals , Cell Line , Cytopathogenic Effect, Viral , Dogs , Humans , Influenza A virus , Genetics , Physiology , Temperature , Virus Cultivation , Methods , Virus Replication
19.
Article in Chinese | WPRIM | ID: wpr-248240

ABSTRACT

<p><b>OBJECTIVE</b>To investigate the optimal dosage and timing for 5-bromodeoxyuridine (BrdU) labeling of endothelial progenitor cells (EPCs) from rat circulating blood.</p><p><b>METHODS</b>The animal model for rat tooth movement was established. EPCs were obtained by density gradient centrifugation. The expressions of specific antigens on cell surface were analysed by immunocytochemistry and fluorescenceochemistry. EPCs were incubated with BrdU at different concentrations (5, 10, 15 micromol/L) for different incubating time (12, 24, 48, 72, 96 h) to identify the optimal BrdU concentration and incubating time for cell labeling. Immunohistochemistry was performed to calculate the labeling index (LI).</p><p><b>RESULTS</b>The culture cell positively expressed CD34, CD133 and could be shown to endocytose DiI-ac-LDL, FITC-UEA-1. Incubation of the EPCs with BrdU at 10 micromol/L and for an optimal length of 72 h appeared to achieve the highest LI (66.8+/-2.9)%, which was significantly higher than group of 5 micromol/L (P<0.05), while there was no significant difference between the group of 15 micromol/L and 10 micromol/L (P>0.05).</p><p><b>CONCLUSION</b>EPCs can be isolated from tooth movement rat circulating blood and cultured. Incubation of the EPCs with BrdU at 10 micromol/L and for an optimal length of 72 h appeared to achieve the optimal LI. This provides a foundation for us to investigate the mechanism of chemiotaxis and differentiation for EPCs.</p>


Subject(s)
Animals , Bromodeoxyuridine , Cell Culture Techniques , Cell Differentiation , Cells, Cultured , Endothelial Cells , Endothelial Progenitor Cells , Rats , Stem Cells , Tooth Movement Techniques
20.
Chinese Journal of Stomatology ; (12): 150-151, 2007.
Article in Chinese | WPRIM | ID: wpr-333381

ABSTRACT

<p><b>OBJECTIVE</b>To study the methods of decalcification for making united slices of tooth and affiliated periodontic tissues.</p><p><b>METHODS</b>Twenty-one samples containing dog molars and affiliated periodontic tissues were divided into seven mean groups. The pH value of solution, time of decalcification, weight and volume of samples, and content of decalcified calcium were detected. The slices were observed by HE, specific, and immunohistochemical stain.</p><p><b>RESULTS</b>The velocity of decalcification increased with decrease of solution pH. The weight of samples lightened by 37.61%, the volume reduced by 25.97% on average, and calcium decalcified was 174.49 mg per gram humid samples. The EDTA decalcification was slowest, but it was best. Decalcification was fast in Plank-Rycho solution while the section was worst, and faster in the formyl solution containing aluminium chloride than in EDTA, and the section was better.</p><p><b>CONCLUSIONS</b>The 50% formyl solution containing aluminium chloride is an ideal decalcifying solution.</p>


Subject(s)
Animals , Decalcification Technique , Methods , Dogs , Edetic Acid , Formates , Microtomy , Molar , Periodontium
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