Characterization of group Ⅰ metabotropic glutamate receptors in rat superior cervical ganglion and their changes following chronic intermittent hypoxia / 解剖学报

Objective To observe the expression and localization of group Ⅰ metabotropic glutamate receptors (mGluR1/ 5) in rat superior cervical ganglion (SCG) and the effect of chronic intermittent hypoxia (CIH) on mGluR1/ 5 protein level. Methods Twelve male SD rats were randomly divided into control group(Ctrl)and CIH group(CIH), 6 rats in each group. After 6 weeks of modeling, the effect of CIH on mGluR1/ 5 protein level was detected by Western blotting, the expression and distribution of mGluR1/ 5 in SCG were detected by immunohistochemistry and double-immunofluorescent staining. Results mGluR1/ 5 was expressed in rat SCG. mGluR1 was distributed in neurons and small intensely fluorescent (SIF) cells, but not in satellite glial cells (SGCs), nerve fibers and blood vessels, whereas mGluR5 was mainly distributed in nerve fibers and a little in neurons, but not in SGCs, SIF cells and blood vessels. CIH increased the protein levels of mGluR1/ 5 (P<0. 01) in rat SCG. Conclusion Both mGluR1 and mGluR5 are expressed in the rat SCG, but their distribution are different, and the increased protein levels of both may be involved in CIH-induced hypertension.

Activation of metabotropic glutamate receptor 1 inhibits chronic intermittent hypoxia-induced carotid body plasticity in rats / 生理学报

The purpose of the present study was to explore the role of carotid body metabotropic glutamate receptor 1 (mGluR1) in chronic intermittent hypoxia (CIH)-induced carotid body plasticity. Sprague Dawley (SD) rats were exposed to CIH (6%-21% O2, 4 min/cycle, 8 h/day) for 4 weeks. The blood pressure of rats was monitored non-invasively by tail-cuff method under consciousness. RT-qPCR was used to examine the mRNA expression level of mGluR1 in rat carotid body. Western blot was used to detect the protein expression level of mGluR1 in rat carotid body. The role of mGluR1 in CIH-induced carotid body sensory long-term facilitation (sLTF) was investigated by ex vivo carotid sinus nerve discharge recording, and the carotid body sLTF was evoked by a 10-episode of repetitive acute intermittent hypoxia (AIH: 1 min of 5% O2 interspersed with 5 min of 95% O2). The results showed that: 1) CIH increased the systolic blood pressure (P < 0.001), diastolic blood pressure (P < 0.005) and mean arterial blood pressure (P < 0.001) of rats; 2) CIH decreased the mRNA and protein levels of mGluR1 in the rat carotid body (P < 0.01); 3) 4 weeks of CIH induced carotid body sLTF significantly, exhibiting as an increasing baseline sensory activity during post-AIH, which was inhibited by application of an agonist of group I metabotropic glutamate receptors, (S)-3,5-dihydroxyphenylglycine (DHPG), during sLTF induction (P < 0.005). In summary, these results suggest that activation of mGluR1 inhibits CIH-induced carotid body plasticity in rats.

Glutamate and its ionotropic receptor agonists inhibit the response to acute hypoxia in carotid body of rats / 生理学报

The purpose of this study was to investigate the effect of glutamate and its ionotropic receptor agonists on the response to acute hypoxia in rat carotid body in vitro. Briefly, after SD rats were anesthetized and decapitated, the bilateral carotid bifurcations were rapidly isolated. Then bifurcation was placed into a recording chamber perfused with 95% O2-5% CO2 saturated Kreb's solution. The carotid body-sinus nerve complex was dissected, and the carotid sinus nerve discharge was recorded using a suction electrode. To detect the response of carotid body to acute hypoxia, the chamber was perfused with 5% O2-5% CO2-90% N2 saturated Kreb's solution for a period of 100 s at an interval of 15 min. To observe the effect of glutamate, ionotropic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor agonist AMPA or N-methyl-D-aspartate (NMDA) receptor agonist NMDA on the response to acute hypoxia in rat carotid body, the chamber was perfused with 5% O2-5% CO2-90% N2 saturated Kreb's solution containing the corresponding reagent. The results showed that glutamate (20 μmol/L), AMPA (5 μmol/L) or NMDA (10 μmol/L) inhibited the acute hypoxia-induced enhancement of carotid sinus nerve activity, and these inhibitory effects were dose-dependent. In summary, the activation of glutamate ionotropic receptors appears to exert an inhibitory effect on the response to acute hypoxia in carotid body of rats.

Role of group II and III mGluRs in carotid body plasticity induced by chronic intermittent hypoxia in rats / 生理学报

The aim of the present study was to explore the role of group II and III metabotropic glutamate receptors (mGluRs) in carotid body plasticity induced by chronic intermittent hypoxia (CIH) in rats. Sprague Dawley (SD) rats were treated with CIH in Oxycycler A84 hypoxic chamber for 4 weeks, and the tail artery blood pressure was measured at the end of model preparation. RT-qPCR was performed to examine the mRNA expression levels of mGluR2/3/8 in rat carotid body. Carotid sinus nerve activity was detected by ex vivo carotid sinus nerve discharge recording technique, and acute intermittent hypoxia (AIH) was administered to induce carotid body sensory long-term facilitation (sLTF), in order to observe the role of group II and group III mGluRs in carotid body plasticity induced by CIH. The results showed that: 1) After 4 weeks of CIH exposure, the blood pressure of rats increased significantly; 2) CIH down-regulated the mRNA levels of mGluR2/3, and up-regulated the mRNA level of mGluR8 in the carotid body; 3) AIH induced sLTF in carotid body of CIH group. In the CIH group, activation of group II mGluRs had no effect on sLTF of carotid body, while activation of group III mGluRs completely inhibited sLTF. These results suggest that CIH increases blood pressure in rats, and group III mGluRs play an inhibitory role in CIH-induced carotid body plasticity in rats.

Effects of acetyl-CoA carboxylase 1 on proliferation, migration and invasion of glioma cell line U87 / 解剖学报

Objective To explore the effect of acetyl-CoA carboxylase 1(ACC1) on cell proliferation, migration and invasion of human glioma cell line U87. Methods Western blotting was performed to examine endogenous ACC1 expression in human glioma cell lines U87, U251 and U373. ACC1 overexpression plasmid and the plasmid vector were transiently transfected into U87 cells. The level of ACC1 in control and ACC1 overexpression cells was examined by Western blotting. The effect of ACC1 on U87 cells migration and invasion was detected by Transwell assay. The effect of ACC1 on U87 cells scratch healing ability was detected by scratch test. The effect of ACC1 on U87 cells proliferation was investigated by MTT assay. Western blotting was conducted to detect the level changes of proteins. Results Among three human glioma cell lines U87, U251 and U373, endogenous ACC1 level in U87 cells was lower than that in other two cell lines. ACC1 overexpression inhibited U87 cell proliferation, as well as cell migration, invasion and scratch healing ability (P < 0.05). Vimentin, fibronectin, urokinase type plasminogen activator (uPA), Bcl-2, cyclin B, cyclin D and p-STAT3 were down-regulated (P< 0.05), P21 was up-regulated (P < 0.05) after ACC1 overexpression. Conclusion These results suggest that ACC1 suppresses the proliferation, migration and invasion of human glioma cells, probably by inhibiting STAT3 activity.

Expression of β-site amyloid precursor protein cleaving enzyme 1 in rat superior cervical ganglion tissues / 解剖学报

Objective To observe the expression and localization of β-site amyloid precursor protein cleaving enzyme 1 (BACEl) in rat superior cervical ganglion and the effect of chronic intermittent frypoxia (CIH) on BACEl level. Methods The expression and distribution of BACEl in superior cervical ganglion were detected by RT-PCR, Western blotting and immunohistochemistry. Totally 16 male SD rats were randomly divided into control group and CIH group, 8 rats in each group. After 2 weeks of modeling, the effect of CIH on BACEl and peroxisome proliferators activated receptor gamma coactivator 1 alpha (PGC-la) mRNA level was detected by RT-PCR. Results BACEl was expressed in rat superior cervical ganglion, and mainly distributed in satellite glial cells and nerve fibers, but not in blood vessels, neurons and small intesely fluorecent(SIF) cells. CIH down-regulated BACEl mRNA level, but up-regulated PGC-la mRNA level ( P < 0.01). Conclusion BACEl is located in satellite glial cells and nerve fibers in the superior cervical ganglion of rats. The decreased level of BACEl ma)' be involved in the regulation of CIH-induced synaptic plasticity of superior cervical ganglion.

Berberine Inhibits Mechanical Stretch Stress induced MAPK Phosphorylation，Proliferation and Migration of Vascular Smooth Muscle Cells / 中山大学学报(医学科学版)

@#【Objective】To observe whether berberine can inhibit vascular smooth muscle cells（VSMC）proliferation induced by mechanical strength stress and to investigate the role of MAPK pathway in it.【Methods】The cultured VSMC were divided into 4 groups：negative control group（NC group），stretch stress group（SS group），berberine pretreated and stretch stress stimulation group（BBR+SS group），and berberine group. In NC group，phosphate buffer saline was used as a negative control；in SS group，stretch stress was given to VSMC；in BBR+SS group，VSMC were pretreated with berberine for 1 hour and then exposed to stretch stress；in BBR group，VSMC were treated only with berberine for 1 hour and cultured in serum- free DMEM afterwards. We collected VSMC in each group ，detected and analyzed their MAPK phosphorylation，proliferation and migration by using Western blotting，immunofluorescence and wound-healing assay respectively. 【Results】 Compare with NC group，stretch stress markedly induced VSMC proliferation and migration ，which could be inhibited significantly by berberine. Stretch stress obviously increased phosphorylation of MAPK （ERK，JNK，p38），which could be inhibited by berberine in a concentration dependent manner. 【Conclusion】 Berberine inhibits hypertension-induced proliferation and migration of VSMC through MAPK pathway. The results revealed the new use and mechanism of berberine，and provided important data for further study on the prevention and treatment of vascular remodeling caused by abnormal increase of mechanical stress in hypertension.

Effect of cyclic intermittent hypoxia on the expression of β-site amyloid precursor protein cleaving enzyme 1 in rat adrenal gland / 解剖学报

Objective To observe the expression and localization of β-site amyloid precursor protein cleaving enzyme 1 (BACE1) in rat adrenal gland and to detect the effect of cyclic intermittent hypoxia (CIH) on the expression of BACE1. Methods The expression and localization of BACE1 in rat adrenal gland were detected by Western blotting and immunohistochemistry. Sixteen male Sprague-Dawley (SD) rats were randomly divided into two groups: control group and CIH group, 8 rats in each group. The protein levels of BACE1 and tyrosine hydroxylase (TH) in rat adrenal medulla were detected by Western blotting after CIH 2 weeks treatment. Results BACE1 was mainly localized in rat adrenal medullary nerve fibers. Compared with the control group, BACE1 protein level decreased and TH protein level increased in the adrenal medulla in the CIH group. Conclusion BACE1 is located in rat adrenal medullary nerve fibers. The decreased level of BACE1 may participate in slowing down the excessive enhancement of sympathetic activity induced by CIH.

Study on control of graft-versus-host disease by blocking CD137-CD137L ligand costimulatory pathway in mice / 中华血液学杂志

<p><b>OBJECTIVE</b>To explore the in vitro effect on control of graft-versus-host disease (GVHD) and its mechanism in mice by blockade of CD137-CD137L pathway.</p><p><b>METHODS</b>Responder spleen cells from BALB/c donor mice (H-2(d)) were incubated with stimulator spleen cells from C57BL/6 ( H-2(b)) recipient mice, with or without anti-CD137L mAb. Lethally irradiated C57BL/6 mice were transplanted with donor bone marrow cells plus primary MLC spleen T cells. Group A (Allo-BMT control group): allo-BMT mice not receiving any prevention measures for GVHD. Group B (CsA + MTX control group): CsA and MTX given to C57BL/6 mice after transplantation. Group C (experimental group): donor spleen cells from BALB/c mice treated with anti-CD137L mAb. The percentages of CD3+ CD8+ T and CD3+ CD4+ T cells in the three groups were detected by flow cytometry, and the level of cytokines (IFN-gamma, IL-2, IL-10, IL-4) by RT-PCR.</p><p><b>RESULTS</b>The incidence of GVHD in group C was 70%, while in group A and group B were 100%. The survival rate was higher and the median survival time was longer of group C than that of group A and B (P < 0.01). All mice in group A died of aGVHD within 15 ds, while 30% of mice in group C survived more than 30 ds. Symptoms and histological signs of GVHD in group C were the mildest among the three groups. The percentage of CD3+ CD8+ T cells and the levels of IFN-gamma were significantly lower (P < 0.01), and the levels of IL-10 were significantly higher in group C than those in group A and B (P < 0.01).</p><p><b>CONCLUSION</b>Treatment of donor T cells with anti-CD137L mAb in vitro may relieve GVHD, thereby improve the survival time and survival rate, which maybe related to increasing Th1 cytokine (IFN-gamma) and decreasing Th2 cytokine (IL-10) as well as reducing CD3+ CD8+ T cells.</p>