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AIM: To investigate the role of hydrogen sulfide (H_2S) in the cholecystokinin octapeptide (CCK-8) attenuating lipopolysaccharide (LPS)-induced lung injury. METHODS: A rat model of lung injury induced by intravenous injection of LPS was developed. Male Wistar rats were divided into normal control group, LPS group, LPS+CCK-8 group and CCK-8 group. Six hours after LPS injection, partial pressure of oxygen in the arterial blood (PaO_2), H_2S content and cystathionine-γ-lyase (CSE) activity in lung tissue were detected. The mRNA expression of CSE in lung tissue was determined by RT-PCR;the structure of lung tissues was observed under optical microscope. RESULTS: Compared to normal control rats, the LPS-treated rats had significantly decreased PaO_2 level, increased index of quantitative assessment (IQA) score, while H_2S content, CSE activity and the mRNA expression of CSE in lung tissue were significantly increased (all P<0.05). Administration of CCK-8 into LPS-treated rats increased the PaO_2 level and alleviated the degree of lung injury (measured by IQA score). In addition, CCK-8 decreased H_2S content, CSE activity, and the mRNA expression of CSE (all P<0.05). No significant difference of the above-mentioned parameters between CCK-8 group and normal control group was observed. CONCLUSION: CCK-8 reduces LPS-induced lung injury through inhibiting the generation of endogenous H_2S.
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Objective To investigate the changes of neuroglobin (Ngb) expression in the CA1 hippocampus after cerebral ischemia and the effect of limb ischemic preconditioning (LIP) on it in young and aged rats. Methods SD rats aged 3 months and 21-23 months with permanently occluding bilateral vertebral arteries were randomly divided into cerebral ischemic group and LIP + cerebral ischemic group, respectively. The expression of Ngb mRNA and protein in the hippocampus were investigated by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot methods. The profile of delayed neuronal death (DND) of pyramidal neurons in the hippocampus CA1 was evaluated by using thionin staining under light microscope by determining the neuronal density (ND) and histological grade (HG). Results Ngb mRNA and protein expressions were 0.16±0.02 and 0.32±0.07, 0.52±0.04 and 0.91±0.06, 0.09±0.01 and 0.22±0.08, 0.21±0.01 and 0.66± 0. 06 in young cerebral ischemia group, LIP + young cerebral ischemia group, aged cerebral ischemia group and LIP + aged cerebral ischemia group, respectively. The expressions of Ngb mRNA and protein after cerebral ischemia for 8 minutes in aged rats were decreased compared with those in the young rats which suffered an identical cerebral ischemia with the aged rats (P<0.05). LIP up-regulated Ngb mRNA and protein expressions in both young and aged rats which suffered cerebral ischemia (P<0.05). However, the up-regulation of Ngb expression in aged rats was significantly less than that in young rats (P<0.05). Neuropathological evaluation showed that ND was 38.8±10.9, 171.5±16.9, 21.2±12.2 and 102.7±15.4 in young cerebral ischemic group, LIP + young cerebral ischemic group, aged cerebral ischemic group and LIP + aged cerebral ischemic group, respectively. It showed that obvious DND of pyramidal neurons was found in young and aged rats after cerebral ischemia. Although LIP effectively protected the pyramidal neurons in the CA1 hippocampus against DND normally induced by ischemic insult, the neuroprotection of LIP for aged rats was less effective than that for young rats. Conclusions The expression of Ngb and the up-regulation effect of LIP on the expression in aged rats are significantly decreased compared to those in young rats when the rats suffer cerebral ischemia. These differences may be one of underlying reasons why the aged rats exhibit severe DND after cerebral ischemia and why the neuroprotective effect of LIP is less in the aged rats than that in the young rats.
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AIM: To study the signal pathway involved in up-regulation of LPS-induced HO-1 expression by CCK-8. METHODS: Forty-two SD rats were divided into 7 groups (six rats each) randomly as follows: control group, LPS group, LPS+SP600125 (JNK-specific inhibitor) group, CCK-8+LPS group, CCK-8+LPS+SP600125 group, CCK-8 group and CCK-8 +SP600125 group. Lungs from the rats in these 7 groups were excised 6 h after the agents were administered. HO-1 mRNA expression was examined by RT-PCR. The protein expression of HO-1 was detected by Western blotting and immunofluorescence flow cytometry (FCM). RESULTS: There were significant positive expression of HO-1 mRNA in LPS group compared to control group. CCK-8 enhanced LPS-induced HO-1 mRNA expression and CCK-8 alone induced HO-1 mRNA expression as well. The mRNA expressions of HO-1 in LPS group, CCK-8+LPS group and CCK-8 group were 3.01 (P<0.01), 5.88 (P<0.01) and 3.45 (P<0.01) times as many as that in control group, respectively. SP600125 inhibited the mRNA expression of HO-1 induced by CCK-8 and (or) LPS. The change of HO-1 protein expression was in accordance with that of HO-1 mRNA expression by Western blotting and immunofluorescence FCM. CONCLUSION: These results suggest that JNK/c-Jun signal pathway plays an important role in the up-regulation of LPS-induced HO-1 expression by CCK-8.
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AIM: To explore the role of NO/ inducible nitric oxide synthase (iNOS) in the metabotromi glutamate receptor 2/3C (mGluR2/3) mediated-brain ischemic tolerance induced by cerebral ischemic preconditioning (CIP), and to observe the influences of α-methyl- (4-tetrazolyl- phenyl) glycine (MTPG), an antagonist of mGluR2/3, on the expression of iNOS during the induction of brain ischemic tolerance. METHODS: Thirty-six Sprague-Dawley rats were subjected to four vessel occluding global brain ischemic model. Thionin staining and immunohistochemistry were used for neuropathological evaluation and assay of iNOS expression in the hippocampal CA1 subregion of the rats. RESULTS: In the sham group, weak expression of iNOS was detected. The expression of iNOS in the CIP and CIP+ischemic insult groups were increased significantly compared with that in the sham group. Administration of MTPG via lateral cerebral ventricle 20 min before CIP blocked the up-regulation of iNOS induced by CIP, but had no influence on the pyramidal neuron survival. However, in the MTPG+CIP+ischemic insult group, the expression of iNOS was extremely intensive compared to that in CIP and MTPG+CIP groups. Importantly, this up-regulation was accompanied with obvious delayed neuronal death. CONCLUSION: NO/iNOS pathway plays an important role in the process of mGluR2/3 mediated-brain ischemic tolerance induced by CIP.
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AIM To observe whether limb ischemic preconditioning (LIP) could attenuate pyramidal neuronal apoptosis of the CA1 hippocampus and brain edema evoked by brain ischemia in rats. METHODSSeventy-two rats whose bilateral vertebral arteries occluded permanently were randomly assigned into 6 groups: sham, LIP(bilateral femoral arteries were clamped for 10 min, 3 times, in a 10-min interval), brain ischemic insult, LIP+brain ischemic insult, DMSO+LIP+brain ischemic insult and SB 203580+LIP+brain ischemic insult groups. Assays for neuronal apoptosis were performed using TUNEL staining. The percentage of wet over dry tissue weight of the brain was measured by weighing method. RESULTS There were almost no TUNEL-positive cells in the CA1 hippocampus in either sham or LIP group. Clear TUNEL-positive pyramidal neurons of the CA1 hippocampus and increase in brain water content were detected in rats subjected to brain ischemic insult. But the number of TUNEL-positive cells and the increase in brain water content were significantly decreased in LIP+brain ischemic insult group compared with that in brain ischemic insult group, indicated that LIP prevented the occurrence of apoptosis of pyramidal neurons of the CA1 hippocampus and brain edema induced by brain ischemic insult. Pretreatment with SB 203580, an inhibitor of mitogen activated protein kinase p38(p38 MAPK), significantly increased the number of TUNEL-positive cells and brain water in SB 203580+LIP+brain ischemic insult group compared with that in DMSO+LIP+brain ischemic insult group, indicated that SB 203580 blocked the protection of LIP against neuronal apoptosis in the CA1 hippocampus and brain edema. CONCLUSION LIP could attenuate pyramidal neurons apoptosis of the CA1 hippocampus and brain edema evoked by brain ischemia, which maybe related to the activation of p38 MAPK.
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AIM To explore the role of superoxide dismutase (SOD) in the p38 mitogen-activated protein kinase (MAPK) mediated brain ischemic tolerance induced by limb ischemic preconditioning (LIP). METHODS The Wistar rats with permanent occlusion of the bilateral vertebral arteries were subjected to occlude the bilateral femoral arteries for 10 min, 3 times, at an interval of 10 min to get the LIP, then global brain ischemia was induced immediately by occluding the bilateral common carotid arteries for 8 min. SB 203580 (100 μmol·L-1, in a volume of 25 μL), an inhibitor of p38 MAPK, was intraventricularly injected 30 min before LIP in SB 203580+LIP+brain ischemia group. Xanthinoxidase and thiomalonylurea methods were used to determine SOD activity and malondialdehyde (MDA) content of the hippocampus, respectively. Thionin staining was used for observing histological changes of the hippocampus. RESULTS LIP significantly prevented the decrease of SOD activity, the increase of MDA content and the delayed neuronal death in the CA1 hippocampus induced by the brain ischemia. SB 203580 pretreatment evidently blocked the protective effect of LIP against the delayed neuronal death and the modulation on SOD activity and MDA content. CONCLUSIONSOD may play an important role served as a downstream molecule of p38 MAPK in the induction of brain ischemic tolerance by LIP.
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AIM: The present study was designed to observe the effect of [D- Arg1, D- Trp7,9, Leu11] - substance P (spantide), a non- selective antagonist of NK receptors, on the up- regulation of nitric oxide synthase (NOS) induced by formalin test. METHODS: Formalin (5%, 0.2 mL) was subcutaneously injected into the plantar side of the right hind paw to produce persistent pain and hyperalgesia. The pain response was determined by spontaneous flinch reflex test. NOS expression was examined using NADPH- d histochemical staining. Spantide was intrathecally injected via L5 - L6 intervertebral space 5 min prior to the formalin injection. RESULTS: Injection of formalin resulted in a characteristic behavioral response consisting of vigorous scratching, biting, licking and lifting of the injected hind paw from the box' s bottom. Following these behavioral responses, the NOS expression was up- regulated in the pericentral canal region of the L5 segment of the spinal cord. Pre- treatment with spantide depressed the spontaneous flinches of the injected paw in the second phase of the formalin test. At the same time, the upregulation of NOS was substantially inhibited. CONCLUSION: It might be concluded that substance P played an important role in the up - regulation of NOS in the pericentral canal region of the spinal cord in the formalin test.