The dynamic changes of heme oxygenase-1 mRNA and protein express at subfornical organ in rats with experimental allergic encephalomyelitis / 中国应用生理学杂志

<p><b>AIM</b>To observe the dynamic changes of heme oxygenase-1 (HO-1) mRNA and protein express in subfornical organ in rats with experimental allergic encephalomyelitis (EAE) to confirm that SFO is one of the sites for blood-bearing signaling molecules entering into brain.</p><p><b>METHODS</b>EAE was induced by CFA-GPSCH on Wistar rats, we observed the levels of HO-1 mRNA and its protein expression with immunohistochemistry and in situ hybridization technology on 1 d, 7 d, 14 d, and 21 d after EAE induction in SFO of rats. The relationship between HO-1 and symptoms of EAE was also investigated.</p><p><b>RESULTS</b>The expression levels of HO-1 mRNA and its protein expression were very low in the brains of the control group, whereas they were enhanced gradually with pathological course in the brain and onsets of symptoms, signs of EAE. On 1 d after induction of EAE, positive cells of HO-1 mRNA and its protein expression were observed at SFO, but the labeled cells were rarely seen in the other brain regions. On 7 d, the positive cells increased markedly. On 14 d the levels of HO-1 mRNA and its protein expression in the brains reached the peak, the positive cells of HO-1 were mainly located at the choroid plexuses and SFO, as well as the regions around "sleeve-like" lesion foci, all of which were coincident with the locations of lesions of EAE. The changes of incidence, symptom, reduction of the body weight, and pathology lesions of EAE in rat brains were the most significant. On 21 d, the levels of HO-1 mRNA and its protein expression reduced gradually, which was in parallel with remitted symptoms of EAE. When a specific inhibitor of HO-1, Snpp9, was applied, the symptoms and pathological lesions of EAE in brains were mitigated markedly.</p><p><b>CONCLUSION</b>SFO may be one of the earliest sites for blood-bearing signaling molecules entering into brain. The dynamic changes of HO-1 mRNA and its protein expression are in parallel with the changes of symptoms and pathological lesions of EAE in the brains. Application of some inhibitors of HO-1 may be one of potential therapeutic methods for prevention and treatment of EAE.</p>

Dynamic changes of heme oxygenase-1 protein and mRNA in the brains of rats with experimental allergic encephalomyelitis / 生理学报

In order to investigate the role of heme oxygenase-1 (HO-1) in the molecular mechanism of experimental allergic encephalomyelitis (EAE), which was induced by guinea pig spinal cord homogenate + complete freund adjuvant on Wistar rats, we observed the gene of HO-1 and its protein expression with reverse transcriptase polymerase chain reaction(RT-PCR) and immunohistochemistry 1, 7, 14, and 21 d after EAE induction in rats. The relationship between HO-1 and the symptoms of EAE was also observed. The results showed that the levels of HO-1 mRNA and its protein expression were very low in the brains of the control group, whereas they were enhanced gradually with pathological course in the brain and onsets of symptoms, signs of EAE. On day 7, the level of HO-1 mRNA reached the peak, but the expression level of HO-1 protein in the brains reached the peak on day 14. The immunoreactive cells of HO-1 were mainly located at the choroid plexuses and subfornical organ (SFO), as well as in regions around the "sleeve-like" lesion foci, all of which were coincident with the locations of lesions of EAE. The levels of HO-1 mRNA and its protein expression were lowered gradually on day 21, which were in parallel with the severities of symptoms and signs of EAE. After a specific inhibitor of HO-1, Snpp-9, was applied, both of the symptoms and pathological lesions of EAE in the rat brains were mitigated markedly. Therefore, these results may suggest that the dynamic changes of HO-1 mRNA and its protein expression are in parallel with the changes of symptoms and pathological lesions of EAE in the brain. In conclusion, the levels of HO-1 mRNA and its protein expression in brains may play an important role in the pathogenesis of EAE, and application of inhibitors of HO-1 may be one of the potential therapeutic ways for the prevention and treatment of EAE.

Cholecystokinin octapeptide increases free intracellular calcium of guinea pig cardiomyocytes through activation of Ca2+ channel and tyrosine kinase / 生理学报

The aim of the present study was to explore the effect of cholecystokinin octapeptide (CCK-8) on [Ca(2+)](i) and its signal transduction mechanism in isolated guinea pig cardiomyocytes. [Ca(2+)](i) was measured by laser scanning confocal microscopy in single ventricular myocytes which were dissociated by enzymatic dissociation method and loaded with Fluo 3-AM. The changes in [Ca(2+)](i) were represented by fluorescent intensity (F(i)) or relative fluorescent intensity (F(i)/F(O)%). The results obtained are as follows. (1) In the normal Tyrode's solution containing 1.0 mmol/ L Ca(2+), CCK-8 (1-10(4) pmol/L) elicited a rapid and marked increase in [Ca(2+)](i). (2) When cardiomyocytes were pretreated with the Ca(2+) chelator EGTA (3 mmol/L) and Ca(2+) channel antagonist nisoldipine (0.5 micromol/L) for 5 min, CCK-8 (10(2)pmol/L) caused a slow and small increase in [Ca(2+)](i) (p< 0.01). (3) Pretreatment with the nonselected CCK- receptor (CCK-R) antagonist proglumide (6 micromol/L) or the tyrosine kinase inhibitor genistein (1 micromol/L) for 5 min could inhibit the increase of [Ca(2+)](i) induced by CCK-8 (10(2) pmol/L) (p<0.01). The results suggest that CCK-8 increases the [Ca(2+)](i) via activating the receptor-operated Ca(2+) channel and eliciting the influx of Ca(2+) in isolated guinea pig cardiomyocytes, in which tyrosine kinase may be involved.

The role of activation of nuclear factor-kappa B of rat brain in the pathogenesis of experimental allergic encephalomyelitis / 生理学报

To investigate the role of activated nuclear factor-kappaB (NF-kappaB) in experimental allergic encephalomyelitis (EAE), the activity and protein expression of NF-kappaB p65 in rat brain tissues, which were extracted from EAE rats at 1, 7, 14 and 21 d respectively after EAE was induced by CFA-GPSCH, were measured with electrophoretic mobility shift assay and immunohistochemistry. The relationship between activated NF-kappaB and symptoms of EAE was also investigated. The results showed that protein expression level and the activity of NF-kappaB were very low in the brain of the control group. After EAE was induced, the activity of NF-kappaB and the level of the protein expression in the brains increased gradually with the development of symptoms and brain pathology of EAE. On d 14, both the activity and the level of protein expression in the brains reached a peak, the positive cells of NF-kappaB were mainly located at the choroid plexuses and subfornical organ, as well as around the regions of sleeve-like lesion foci, which were coincident with the locations of lesions of EAE. The incidence, symptoms, reduction of the body weight and pathology of EAE rats brains at the above locations were most significant. On d 21 the activity of NF-kappaB and level of the protein expression reduced gradually, which was in parallel with a gradual alleviation of the symptoms of EAE rats. After a specific inhibitor of NF-kappaB, PDTC was applied, the symptoms and pathological lesions of EAE rat brain were mitigated markedly. The above results indicate that the dynamic changes in the activity and protein expression of NF-kappaB were in parallel with the changes in symptoms and pathological lesion of EAE rat brains. In conclusion, the activated NF-kappaB in the brain may play a critical role in the pathogenesis of EAE, and application of some inhibitors of NF-kappaB, such as PDTC, may be one of the effective therapeutic methods for prevention and treatment of EAE.

Expression of heme oxygenase-1 induced by lipopolysaccharide in thoracic aortic and its effect on vascular reactivity / 中国应用生理学杂志

<p><b>AIM</b>To explore the effects of heme- heme oxygenase-1 (HO-1)-carbon monoxide(CO)-cyclic GMP (cGMP)on aortic vascular reactivity in endotoxemic rats and its molecular mechanism.</p><p><b>METHODS</b>By using isolated vascular ring tension detecting technique, cumulative responses of thoracic aortic rings (TARs)to phenylephrine (PE) were measured at 6 h after lipopolysaccharide administration. Effects on contractile responses to PE were measured under which the TARs were incubated with hemin (He, donor of CO), zinc-protoporphyrin-IX(ZnPP-IX, selective inhibitor of HO-1), or methylene blue (MB, inhibitor of guanylyl cyclase), respectively. The content of CO and the activity of HO-1 were measured. The protein and the gene expression of HO-1 were examined by Western blot and RT-PCR.</p><p><b>RESULTS</b>Contractile responses of TARs to cumulative doses of PE were depressed by pretreated with LPS. The hyporesponsiveness was partly reversed by incubation with ZnPP-IX and was restored to normal by incubation with MB in endotoxemic rats. Incubation with He could contribute to the vascular hyporeactivity. The content of CO, the activity and the protein and the gene expression of HO-1 were significantly increased in aorta of endotoxemic rats.</p><p><b>CONCLUSION</b>LPS could induce the HO-1 mRNA and the protein expression, the activity of HO-1 increase in aorta, lead to active the pathway of heme-HO-1-CO-cGMP, which is one of the important mechanisms of the vascular hyporeactivity in endotoxemic rats.</p>