Resveratrol ameliorates lipopolysaccharide-induced anxiety-like behavior by attenuating YAP-mediated neuro-inflammation and promoting hippocampal autophagy in mice.

Resveratrol, a type of natural polyphenol mainly extracted from the skin of grapes, has been reported to protect against inflammatory responses and exert anxiolytic effect. Yes-associated protein (YAP), a major downstream effector of the Hippo signaling pathway, plays a critical role in inflammation. The present study aimed to explore whether YAP pathway was involved in the anxiolytic effect of resveratrol in lipopolysaccharide (LPS)-treated C57BL/6J male mice. LPS treatment induced anxiety-like behavior and decreased sirtuin 1 while increased YAP expression in the hippocampus. Resveratrol attenuated LPS-induced anxiety-like behavior, which was blocked by EX-527 (a sirtuin 1 inhibitor). Mechanistically, the anxiolytic effects of resveratrol were accompanied by a marked decrease in YAP, interleukin-1ß and ionized calcium binding adaptor molecule 1 (Iba-1) while a significant increase in autophagic protein expression in the hippocampus. Pharmacological study using XMU-MP-1, a YAP activator, showed that activating YAP could induce anxiety-like behavior and neuro-inflammation as well as decrease hippocampal autophagy. Moreover, activation of YAP by XMU-MP-1 treatment attenuated the ameliorative effects of resveratrol on LPS-induced anxiety-like behavior, while blockade of YAP activation with verteporfin, a YAP inhibitor, attenuated LPS-induced anxiety-like behavior and neuro-inflammation as well as hippocampal autophagy. Finally, rapamycin-mediated promotion of autophagy attenuated LPS-induced anxiety-like behavior and decreased interleukin-1ß and Iba-1 expression in the hippocampus. Collectively, these results indicate that amelioration by resveratrol in LPS-induced anxiety-like behavior is through attenuating YAP-mediated neuro-inflammation and promoting hippocampal autophagy, and suggest that inhibition of YAP pathway could be a potential therapeutic target for anxiety-like behavior induced by neuro-inflammation.

Intermedin modulates hypoxic pulmonary vascular remodeling by inhibiting pulmonary artery smooth muscle cell proliferation.

BACKGROUND: Hypoxic pulmonary arterial hypertension (PAH) is a disabling disease with limited treatment options. Hypoxic pulmonary vascular remodeling is a major cause of hypoxic PAH. Pharmacological agents that can inhibit the remodeling process may have great therapeutic value. OBJECTIVE: To examine the effect of intermedin (IMD), a new calcitonin gene-related peptide family of peptide, on hypoxic pulmonary vascular remodeling. METHODS: Rats were exposed to normoxia or hypoxia (∼10% O(2)), or exposed to hypoxia and treated with IMD, administered by an implanted mini-osmotic pump (6.5 µg/rat/day), for 4 weeks. The effects of IMD infusion on the development of hypoxic PAH and right ventricle (RV) hypertrophy, on pulmonary vascular remodeling, on pulmonary artery smooth muscle cell (PASMC) proliferation and apoptosis, and on the activations of l-arginine nitric oxide (NO) pathway and endoplasmic reticulum stress apoptotic pathway were examined. RESULTS: Rats exposed to hypoxia developed PAH and RV hypertrophy. IMD treatment alleviated PAH and prevented RV hypertrophy. IMD inhibited hypoxic pulmonary vascular remodeling as indicated by reduced wall thickness and increased lumen diameter of pulmonary arterioles, and decreased muscularization of distal pulmonary vasculature in hypoxia-exposed rats. IMD treatment inhibited PASMC proliferation and promoted PASMC apoptosis. IMD treatment increased tissue level of constitutive NO synthase activity and tissue NO content in lungs, and enhanced l-arginine uptake into pulmonary vascular tissues. IMD treatment increased cellular levels of glucose-regulated protein (GRP) 78 and GRP94, two major markers of endoplasmic reticulum (ER) stress, and increased caspase-12 expression, the ER stress-specific caspase, in lungs and cultured PASMCs. CONCLUSIONS: These results demonstrate that IMD treatment attenuates hypoxic pulmonary vascular remodeling, and thereby hypoxic PAH mainly by inhibiting PASMC proliferation. Promotion of PASMC apoptosis may also contribute to the inhibitory effect of IMD. Activations l-arginine-NO pathway and of ER stress-specific apoptosis pathway could be the mechanisms mediating the anti-proliferative and pro-apoptotic effects of IMD.

Upregulation of miR-335-3p by NF-κB Transcriptional Regulation Contributes to the Induction of Pulmonary Arterial Hypertension via APJ during Hypoxia.

Pulmonary arterial hypertension (PAH) is a cardiopulmonary disease that can lead to heart failure and eventually death. MicroRNAs (miRs) play essential roles during PAH progression; however, their exact mechanism of action remains unclear. Apelin is a small bioactive peptide with a key protective function in the pathogenesis of PAH mediated by binding to the APJ gene. The aim of the present study was to investigate the role of miR-335-3p in chronic normobaric hypoxia (CNH)-induced PAH in mice and the potential underlying regulatory mechanism. Adult male C57BL/6 mice were exposed to normoxia (~21% O2) or CNH (~10% O2, 23 h/d) for 5 weeks. MiR-335-3p was significantly increased in lung tissue of CNH-induced PAH mice. Blocking miR-335-3p attenuated CNH-induced PAH and alleviated pulmonary vascular remodeling. Bioinformatics analysis and luciferase reporter assay indicated that nuclear factor-kappa beta (NF-κB) acted as a transcriptional regulator upstream of miR-335-3p. Pyrrolidine dithiocarbamate treatment reversed the CNH-induced increase in miR-335-3p expression and diminished CNH-induced PAH. Moreover, p50-/- mice were resistant to CNH-induced PAH. Finally, APJ was identified as a direct targeting gene downstream of miR-335-3p, and pharmacological activation of APJ by its ligand apelin-13 reduced CNH-induced PAH and improved pulmonary vascular remodeling. Our results indicate that NF-κB-mediated transcriptional upregulation of miR-335-3p contributes to the inhibition of APJ and induction of PAH during hypoxia; hence, miR-335-3p could be a potential therapeutic target for hypoxic PAH.

[Effect of apelin on hypoxic pulmonary hypertension in rats: role of the NO pathway].

To investigate the effectiveness and mechanism of apelin against pulmonary hypertension and pulmonary vascular remodeling induced by hypoxia in rats, 24 male Sprague-Dawley rats were randomly divided into normal control (NC) group, 4-week hypoxia (HH) group and 4-week hypoxia with apelin (HA) group (each n=8). The rats of hypoxic group were placed in an isobaric hypoxic chamber, in which O2 and CO2 content was maintained at 9%-11% and <3%, respectively, for 4 weeks (8 h/d, 6 d/week). [pGlu]apelin-13 (10 nmol/kg per day, 28 d) was administered subcutaneously by osmotic mini-pump before hypoxia treatment in HA group. L-arginine (L-Arg) uptake of pulmonary artery was assay by [³H]-L-Arg, while nitric oxide synthase (NOS) activity of pulmonary tissue, and nitrate/nitrite (NO2(-)/NO3(-)) concentrations in pulmonary tissue and plasma were detected by colorimetric technique and nitrate reductase method, respectively. The results showed that mean pulmonary arterial pressure, the ratio value of right ventricle weight to left ventricle plus septum weight, the relative medial thickness, and the relative medial area of pulmonary arterioles were higher in HH group than those in NC group (all P<0.01), while these indices were lower in HA group than those in HH group (P<0.05 or P<0.01). Compared with those in HH group, the uptake of 0.5, 5 and 10 nmol/L [³H]-L-Arg in pulmonary artery in HA group increased by 121.4% (P<0.01), 85.0% (P<0.05) and 61.5% (P<0.05), respectively; cNOS activity of pulmonary tissue increased by 74.3%, while iNOS activity decreased by 25.0% (all P<0.01); and NO2(-)/NO3(-) concentrations in pulmonary tissue and plasma increased by 97.6% and 48.0% (all P<0.05), respectively. Taken together, our results suggest that apelin has a prophylactic effect against hypoxic pulmonary hypertension in rats, and that the mechanism of this effect is possibly associated with activation of the L-Arg/NOS /NO pathway.

Percutaneous biphasic electrical stimulation for treatment of obstructive sleep apnea syndrome.

In this paper, we study the effect of stimulation of the genioglossus with percutaneous biphasic electrical pulses on patients with the obstructive sleep apnea syndrome (OSAS). The experiment was conducted in 22 patients clinically diagnosed with OSAS. The patients were monitored with polysomnography (PSG) in the trial. When the sleep apnea was detected, the genioglossus was stimulated with percutaneous biphasic electrical pulses that were automatically regulated by a microcontroller to achieve the optimal effect. The percutaneous biphasic electrical stimulation caused contraction of the genioglossus, forward movement of the tongue, and relieving of the glossopharyngeal airway obstruction. The SaO2, apnea time, hypoxemia time, and change of respiratory disturbance index (RDI) were compared in patients with treatment and without treatment. With percutaneous biphasic electrical stimulation of the genioglossus, the OSAS patients showed apnea time decreased (P < 0.01), RDI decreased (P < 0.01), and SaO2 increased (P < 0.01). No tissue injury or major discomfort was noticed during the trial. The stimulation of genioglossus with percutaneous biphasic electrical current pulse is an effective method for treating OSAS.

SIRT1 Mediates Apelin-13 in Ameliorating Chronic Normobaric Hypoxia-induced Anxiety-like Behavior by Suppressing NF-κB Pathway in Mice Hippocampus.

We previously showed that apelin-13 ameliorates chronic normobaric hypoxia (CNH)-induced anxiety-like behavior in mice, the mechanism, however, is not well known. This study aims to investigate whether SIRT1 is involved in the anxiolytic effect of apelin-13 in CNH-treated mice, and to illustrate the potential underlying mechanism. We showed that apelin-13 treatment reversed a decrease in SIRT1 and an increase in acetylated p65 (lysine 310) proteins' expression in hippocampus of CNH-treated mice, indicating that apelin-13 inhibited NF-κB signaling pathway by activating SIRT1. Behaviorally, apelin-13 ameliorated CNH-induced anxiety-like behavior, EX-527 blocked the beneficial effect of apelin-13, and the anxiogenic effect of CNH was attenuated by resveratrol pretreatment, suggesting that SIRT1 was involved in the effect of apelin-13 against CNH-induced anxiety-like behavior in mice. We also showed that resveratrol treatment decreased IL-1ß, IL-6, TNF-ɑ, PCNA, Bcl-2, and acetyl-p65 levels, but increased Bax and caspase 3 levels in hippocampus, suggesting a suppressive effect of resveratrol on cellular neuroinflammation and proliferation while a promotive effect on apoptosis of microglia in hippocampus. Finally, blockade of NF-κB activity by PDTC diminished CNH-induced anxiety-like behavior, indicating that NF-κB was involved in CNH-induced anxiety-like behavior in mice. In conclusion, this study provides the first evidence that SIRT1 mediates the anxiolytic effect of apelin-13 in CNH-treated mice through the inhibition of NF-κB pathway. These results imply that dysfunction of the apelin-SIRT1-NF-κB axis in hippocampus represents a potential mechanism that results in the induction of neuroinflammation and reduction in neuroprotection, thus induces anxiety-like behavior in CNH-treated mice.

[Changes of intermedin/adrenomedullin 2 and its receptors in the right ventricle of rats with chronic hypoxic pulmonary hypertension].

The purpose of the present study was to explore the expression changes of intermedin/adrenomedullin 2 (IMD/ADM2), a novel small molecular bioactive peptide, and its receptors, calcitonin receptor-like receptor (CRLR) and receptor activity modifying proteins (RAMP1, RAMP2, RAMP3) in the right ventricle of rats with chronic hypoxia-induced pulmonary hypertension. Twenty male Sprague-Dawley rats were randomly divided into 4-week hypoxia group and normal control group (each n=10). The rats in hypoxia group were placed in an isobaric hypoxic chamber, in which O(2) content was maintained at 9%-11% by delivering N(2), and CO(2) content was maintained at <3% for 4 weeks (8 h/d, 6 d/week). The rats in the control group were housed in room air. The protein levels of IMD/ADM2 and adrenomedullin (ADM) in blood plasma and right ventricular tissue were measured by radioimmunoassay. The mRNA expressions of IMD/ADM2, ADM and their receptors CRLR, RAMP1, RAMP2, RAMP3 in right ventricular tissue were determined by reverse transcription-polymerase chain reaction (RT-PCR). The results showed that the ratio of right ventricle weight to left ventricle plus septum weight [RV/(LV+S)] and mean pulmonary arterial pressure (mPAP) were higher in hypoxia group than those in the control group (all P<0.01), suggesting that the rat model of pulmonary hypertension was successfully established. However, the mean carotid arterial pressure (mCAP) between the two groups had no significant difference. Compared with that in the control group, ADM contents in plasma and right ventricular tissue in hypoxia group increased by 1.26 and 1.68 folds (all P<0.01), respectively. Likewise, IMD/ADM2 contents in blood plasma and right ventricular tissue in hypoxia group increased by 0.90 and 1.19 folds (P<0.01), respectively, compared with that in the control group. The data of RT-PCR showed that mRNA levels of ADM, IMD/ADM2 and RAMP2 in hypoxia group increased by 155.1% (P<0.01), 80.9% (P<0.01) and 52.9% (P<0.05), respectively, compared with those in the control group. There were no significant differences in mRNA expressions of CRLR, RAMP1 and RAMP3 between the two groups (all P>0.05). Taken together, the results show that the level of IMD/ADM2 increases in the rats with chronic hypoxia-induced pulmonary hypertension.

Amelioration of apelin-13 in chronic normobaric hypoxia-induced anxiety-like behavior is associated with an inhibition of NF-κB in the hippocampus.

Apelin, a small bioactive peptide, plays an important role in the pathogenesis of mood disorders through the endogenous ligand APJ. Although the anxiolytic effect of apelin is well established, the mechanisms are poorly understood. In this study, we hypothesized that apelin played an anxiolytic role in chronic normobaric hypoxia (CNH)-induced anxiety like behavior in mice, which might be associated with an inhibition of nuclear factor-κB (NF-κB) activation in the hippocampus. To this end, mice were exposed in a normobaric hypoxic chamber with a fraction of inspired oxygen (FIO2, ∼10%, 23h/d) with or without apelin-13 application (20 nmolkg-1d-1, i.p.), for 4 weeks. The anxiety-like behavior was tested by elevated plus maze and open field. Activities of NF-κB, microglial, and related signaling pathways in the hippocampus during this pathological process were examined. We found that CNH treatment decreased APJ but increased Iba-1 proteins expression, as well as nucleus translocation of p50 and p65 in the hippocampus, which were reversed by apelin-13 treatment. In addition, apelin-13 treatment ameliorated CNH-induced anxiety-like behavior in mice, suggesting anxiogenic effect of apelin-13 might be mediated by an inhibition of NF-κB activation in microglial of the hippocampus. Furthermore, apelin-13 treatment reversed p-CAMKII decrease in the hippocampus under CNH treatment. Apelin-13 treatment did not affect anxiety-like behavior and relative proteins expression in normoxia control mice. Finally, we found that rats with CNH treatment decreased APJ expression while enhanced NF-κB activation in the hippocampus, providing additional evidences that NF-κB activation in hippocampus in CNH-induced anxiety-like behavior in rats we reported previously might be associated with an inhibition of APJ activity. In conclusion, the present results illustrated that inhibition of APJ and promotion of NF-κB activation in the microglial of hippocampus might be involved in anxiogenic effect in CNH-exposed mice, and apelin-13 ameliorates CNH-induced anxiety-like behavior might be associated with an inhibition of NF-κB activation.

[Experimental study of non-invasive percutaneous electrical stimulator for treatment of obstructive sleep apnea syndrome and its clinical effect].

To study the stimulation of the genioglossus with percutaneous biphasic current pulses as a new therapeutical method to treat the obstructive sleep apnea syndrome (OSAS), polysomnography (PSG) was used to synchronously monitor the patient. When OSAS was occurring, the stimulation with the optimal parameter was given in time to make the tongue move forward, the glossopharyngeal airway dilated, the resistance of the upper respiratory tract reduced, the hypoxia at night to be improved and the sleeping structure to be ameliorated because of the function of the dilated muscle of the upper airway. The results of the clinical therapeutic effect indicated that 17 of 22 patients with OSAS had cured effects, 2 of whom improved and 3 of whom were without effect. The effective rate was 77.27%. It is preliminarily proved that this is a new method in the treatment of patients suffered from OSAS.

Blunted inflammation mediated by NF-κB activation in hippocampus alleviates chronic normobaric hypoxia-induced anxiety-like behavior in rats.

This study aims to investigate whether inflammation mediated by NF-κB activation is involved in the induction of anxiety-like behavior in chronic normobaric hypoxia (CNH) exposed rats and to investigate the underlying mechanism. To this end, rats were exposed in a normobaric hypoxic chamber with a fraction of inspired oxygen (FIO2) of ∼ 10%, 23 h/d, continues for 2 weeks. Anxiety-like behavior was tested by elevated plus maze and open field, inflammatory response, nucleus translocation of NF-κB, and signaling pathway in hippocampus were examined. CNH induced a significant increase of anxiety- like behavior and inflammation responses, which were ameliorated by NF-κB inhibitor, PDTC pretreatment, suggesting that the anxiogenic effect induced by inflammation is through NF-κB activation. CNH treatment significantly increased nucleus translocation of p65 and p105 in hippocampus, which was suppressed by PDTC pretreatment. In addition, CNH treatment significantly increased Iba-1, iNOS, COX-2, and p-PKA in hippocampus, which were blocked by PDTC pretreatment, suggesting CNH may activate microglia cells in hippocampus through NF-κB pathway. In conclusion, our results illustrate a mechanism that, activation of NF-κB in hippocampus may trigger the proinflammatory response of microglia cells, and iNOS-PKA pathway may involve in anxiogenic effect in CNH exposed rats.

[Effect of oxygen on L-arginine transport in pulmonary arteries in rats chronically exposed to hypoxia-hypercapnia].

OBJECTIVE: To investigate the effect of chronic hypoxia-hypercapnia and oxygen inhalation on L-arginine transport in rat pulmonary artery. METHODS: Forty male Sprague-Dawley rats were randomly divided into four groups: normal control group (A), the group of 4-week hypoxia-hypercapnia after 4-week hypoxia (B), the group of 4-week oxygen inhalation (D) 4-week exposure to air (C) after 4-week hypoxia and 4-week hypoxia-hypercapnia. Changes in pulmonary artery L-arginine (L-Arg) transport, nitric oxide synthase (NOS) activity, plasma nitrite (NO(2)/NO(3)) concentration and L-Arg level in rats were measured. RESULTS: (1) The mean pulmonary artery pressure (mPAP) and weight ratio of right ventricle to left ventricle plus septum (RV/LV + S) of B group were higher than those of A group (33% and 35%, respectively, P < 0.01), with the D group lower than C group (24% and 24%, respectively, P < 0.05). (2) At low (0.2 mmol/L) or high (5.0 mmol/L) concentration of L-Arg, the velocity of L-Arg transport in B group was lower than that in A group [(3.04 +/- 0.16 vs 4.62 +/- 0.55) micro mol x g(-1) x min(-1) and (8.12 +/- 0.14 vs 11.24 +/- 1.02) micro mol x g(-1) x min(-1), respectively, P < 0.01], and in D group higher than in C group [(4.30 +/- 0.18 vs 3.80 +/- 0.21) micro mol x g(-1) x min(-1) and [(12.31 +/- 0.65 vs 10.04 +/- 1.28) micro mol x g(-1) x min(-1), respectively, P < 0.01]. (3) The activity of pulmonary artery tNOS and iNOS in B group was significantly higher than that in C group [(3.82 +/- 0.42 vs 2.59 +/- 0.22) micro nmol x g(-1) x min(-1) and (3.07 +/- 0.30 vs 1.62 +/- 0.10) nmol x g(-1) x min(-1) respectively, P < 0.01], but cNOS in B group was lower than that in NC group [(0.75 +/- 0.16 vs 0.93 +/- 0.10) nmol x g(-1) x min(-1), P < 0.01]. The activity of tNOS and cNOS was not significantly different between D group and C group (P > 0.05); However, iNOS was lower in D group than in C group [(1.97 +/- 0.18 vs 2.25 +/- 0.17) nmol x g(-1) x min(-1), P < 0.05]. (4) Plasma NO(2)/NO(3) content of B group was decreased by 21%, compared with C group (P < 0.01); and D group was increased by 89% (P < 0.01), compared with C group. (5) Plasma L-Arg level was not significantly different among the four groups (P > 0.05). CONCLUSION: The L-Arg transport injury of the pulmonary artery might be the key cause for the decrease of nitric oxide generation in rats with chronic hypoxia-hypercapnia induced pulmonary hypertension. Oxygen inhalation was found to improve L-Arg transport in the injured arteries.

[Effect of safflower injection on endoplasmic reticulum stress-induced apoptosts in rats with hypoxic pulmonary hypertension].

OBJECTIVE: To explore the effects of safflower injection on prevention and treatment of hypoxic pulmonary hypertension and clarify the function of the endoplasmic reticulum stress apoptosis pathway during the process. METHODS: Thirty male SD rats were randomly grouped as normal control group, hypoxia-hypercapnia group and hypoxia+safflower group. The latter two groups were put in the cabin with oxygen concentration ranged from 9% to 11% and carbon dioxide concentration from 5% to 6%. The pulmonary artery pressure and the index of right ventricular hypertrophy were determined after hypoxia exposure (8 h/dx28 d). Changes in morphology of lung tissue were observed by electron microscopy. To explore the possible mechanisms, we also detected apoptosis and apoptosis-related genes/proteins in lung tissue by TUNEL reactivity and PCR and Western blot. RESULTS: Compared with the normal control group, pulmonary artery pressure and the index of right ventricular hypertrophy in hypoxia group were 45% and 33.4% higher, respectively. Tiny blood vessel wall of lungs was thickened and edema, and proliferation of collagen fibers was obvious under the electron microscope. TUNEL staining of apoptotic cells in lung tissues showed more high brightness green fluorescence (+-++), but less green fluorescence showed in the pulmonary vascular smooth muscle cell layer, and apoptosis index (AI) value was 150% higher; gene and protein expression levels of endoplasmic reticulum stress pathway were increased. Compared with hypoxia-hypercapnia group, pulmonary artery pressure and the index of right ventricular hypertrophy in the hypoxia+safflower group were 18% and 15.6% lower, respectively; collagen fibers were decreased, and smooth muscle cells and epithelial cells were got apoptotic-like changes under the electron microscope. TUNEL staining of apoptotic cells in lung tissues showed brighter green fluorescence (++-+++); the high brightness green fluorescence showed in pulmonary vascular smooth muscle cell layer, and apoptotic index (Al) value was 40% higher; gene and protein expressions of endoplasmic reticulum stress pathway were significantly upregulated. CONCLUSION: Our findings demonstrate that safflower injection could activate endoplasmic reticulum stress-induced apoptosis and especially promote apoptosis in pulmonary vascular smooth muscle cells.

[Changes of endoplasmic reticulum stress-induced apoptosis in pulmonary tissue of rats with hypoxic pulmonary hypertension].

OBJECTIVE: To investigate the changes of endoplasmic reticulum stress-induced apoptosis in pulmonary tissue of rats with hypoxic pulmonary hypertension. METHODS: Twenty two male SD rats were randomly divided into control group and 4-week hypoxia-hypercapnia group (n=11). The mean pulmonary arterial pressure (mPAP) and the mean carotid arterial pressure (mCAP) were monitored, and the weight ratio of right ventricle (RV) to left ventricle plus septum (LV + S) were measured. The rattish pathological model were assessed by mPAP, mCAP, RV/(LV+ S), vessel wall area/total area (WA/TA), vessel cavity area/total area (CA/TA) and media thickness of pulmonary arteriole (PAMT). The pulmonary apoptotic cells were detected by Hoechst staining. RT-PCR was used to study the genetic expression of caspasel2, glucose regulated protein 78 (GRP78) and GRP94 in pulmonary tissue. The expression of GRP94 and GRP78 proteins in pulmonary tissue were determined by using immunohistochemistry. RESULTS: (1) (The mPAP, RV/(LV + S), WA/TA and PAMT were respectively higher by 50.5%, 37.3%, 72.5% and 137% in hypoxic group than those in control group, while CA/TA was lower by 41.9% (all P < 0.01). There was not significant difference of mCAP between the two groups. (2) Hoechst staining showed that the pulmonary apoptotic cells in hypoxic group outnumbered markedly than those in control group, and the apoptotic cells were mainly in pulmonary tissue, while they were rare in pulmonary vascular smooth muscle cell. (3) Compared with control group, the expression of pulmonary caspasel2, GRP78 and GRP94 mRNA in hypoxic group were higher by 144%, 137% and 80.7% (all P < 0.05), respectively. (4) The expression of pulmonary GRP78 and GRP94 proteins were up-regulated in hypoxic group, and these proteins mainly localized in pulmonary vascular endothelial cell. CONCLUSION: The endoplasmic reticulum stress-induced apoptosis may be one of the mechanism of hypoxic pulmonary hypertension and pulmonary vascular wall remodeling.

[Effect of apelin on vasodilatation of isolated pulmonary arteries in rats is concerned with the nitric oxide pathway].

OBJECTIVE: To investigate the effect of apelin on vasodilatation of isolated pulmonary arterial rings in rats and its relationship to the nitric oxide (NO) pathway, and to observe the difference of vasodilatation between hypoxic rats and normoxic rats. METHODS: Thirty-six male Sprague-Dawley (SD) rats were randomly divided into hypoxic group and normoxic group. The effects of accumulated apelin on pulmonary arterial rings preconstricted with norepinephrine (NE) were observed by using tissue organ bath system. After pulmonary arterial rings were pretreated with three methods: removing the endothelium, pretreating with nitric oxide synthase inhibitor L-NAME or soluble guanylatecyclase inhibitor ODQ, the different effect of apelin was observed. In addition, the difference of vasodilatation between hypoxic rats and normal rats were observed. RESULTS: (1) Exposure of intact endothelium pulmonary arterial rings preconstricted by NE to apelin at concentration (0.01 - 100 nmol/L) induced a significant concentration dependent relaxation. The maximal vasorelaxant effect of apelin was 10.62% +/- 2.60%, which was inhibited by removal of the endothelium (P < 0.01), pretreatment with L-NAME (P < 0.01) or ODQ (P < 0.01). (2) Response of pulmonary arterial rings from hypoxic pulmonary hypertension rats was decreased (P < 0.05). Compared to normal rats, at a concentration of 100 nmol/L, the response to apelin on arteries from hypoxic rats decreased 60.45% (P < 0.01). But the values of EC50 were not significantly different (P > 0.05). CONCLUSION: These results indicate that apelin relaxes the pulmonary arterial rings of rats in an endothelium dependent manner, which may have a relationship to NO signaling pathway. The response of vasodilatation is decreased in the pulmonary arterial rings from the hypoxic rats.

[Protective and therapeutic effect of apelin on chronic hypoxic pulmonary hypertension in rats].

OBJECTIVE: To study the role of apelin in the prevention of pulmonary hypertension induced by hypoxia in rats. METHODS: The animal model of hypoxic pulmonary hypertension was established by exposing the rats to isobaric hypoxic chamber for 4 weeks (8 h/d, 6 d/ w). Forty male Sprague-Dawley rats were randomly divided into control group (NC), hypoxic group(HH), hypoxic with low-dose apelin (5 nmol/(kg x d) group(LA) and high-dose apelin (10 nmol/(kg x d) (HA). [pGlu]apelin-13 was administered into the rats of apelin groups by mini-osmotic pump subcutaneously. The mean pulmonary arterial pressure(mPAP) and the mean carotid arterial pressure (mCAP) were measured by either right or left cardiac catheterization, and the weight ratio of right ventricule/left ventricule plus septum (RV/(LV + S)) were calculated. The Masson's trichrome stained lung specimens were examined by light microscope to examine the vessel wall area/total area (WA/TA), vessel cavity area/total area (CA/TA) and media thickness of pulmonary arterioles (PAMT). Meanwhile, the lung homogenates were assayed for the activity of supeeroxide dismutase (SOD) and the content of malondialdehyde (MDA). RESULTS: (1) mPAP and RV/(LV + S) of HH group were significantly higher than those of NC group. mPAP of LA and HA groups were lower than those of HH group. The RV/(LV + S) of HA group was significantly lower than that of HH group, but there was no significant difference between HH group and LA group. (2) Masson's trichrome staining revealed that WA/TA and PAMT of HH group were higher than those of NC group. Administration of apelin significantly eliminated WA/TA and PAMT in LA and HA groups. (3) CA/TA of HH group was lower than that of NC group. Administration of apelin significantly elevated CA/TA in LA and HA groups. (4) The activity of SOD and content of MDA in HH group was, respectively, lower and higher than those in NC group. Apelin treatment increased the activity of SOD in LA and HA groups while decreased the content of MDA. CONCLUSIONS: Apelin could play an important role in treatment of hypoxic pulmonary hypertension of rats and the mechanisms of protection were associated with vasodilation of pulmonary artery and inhibition of oxidative stress.

Chronic intermittent hypoxia exposure induces memory impairment in growing rats.

The study was aimed to examine the effect of chronic intermittent hypoxia (CIH) on spatial memory of growing rats and to explore the possible underlying mechanisms. Sixty two rats were trained to perform the 8-arm radial maze task and were divided into four groups: 2-week-CIH (2IH) 2-week-control (2C) 4-week-CIH (4IH) and 4-week-control (4C). There were more reference memory errors, working memory errors and total memory errors in 2IH and 4IH groups compared to the controls. The levels of 8-iso-Prostaglandin F(2-alpha) [8-ISO-PGF(2-alpha)] an in vivo marker for oxidative stress, in serum, hippocampus and prefrontal cortex were higher in CIH groups than the control groups. There were significant correlations between the levels of 8-ISO-PGF(2-alpha) and numbers of memory errors. The ultrastructural changes were evident in the hippocampal and prefrontal cortical tissues from the CIH groups. These results indicate that CIH can induce oxidative stress in brain tissues involved in spatial memory function.

[Effect of hypoxia on the expressions of intermedin/ adrenomedullin2 in plasma and the tissues of heart and lung in rats].

AIM: To study the effect and significances of two-week hypoxia on the expression of intermedin/adrenomedullin2 (IMD/ADM2) in plasma and the tissues of heart and lung in rats. METHODS: Twenty male SD rats were randomly divided into normal control group and hypoxia group. The concentrations of IMD/ADM2 and adrenomedullin (ADM) in plasma, right ventricle and lung tissue were measured by radioimmunoassay. RT-PCR was used to detect the mRNA levels of IMD/ADM2 and ADM in right ventricle and lung tissue. RESULTS: (1) The mean pulmonary arterial pressure (mPAP) and the weight ratio of right ventricle (RV) to left ventricle plus septum (LV + S) of hypoxia group were significantly higher than those of normal control group (P < 0.01). (2) The concentrations of IMD/ADM2 and ADM in plasma were significantly higher in hypoxia group, compared with normal control group (P < 0.01). (3) The concentration of ADM in right ventricle and lung tissue in hypoxia group was significantly higher than that in normal control group (P < 0.01), while there was no significant difference in IMD/ADM2 between the two groups. (4) The mRNA levels of IMD/ADM2 and ADM in right ventricle and lung tissues were significantly up-regulated in hypoxia group (P < 0.05). CONCLUSION: The expressions of IMD/ADM2 peptides and gene in plasma, right ventricular and pulmonary tissues are different in the early-middle pathological proceeding of pulmonary hypertension induced by two-week hypoxia in rats.

[Changes of adrenomedullin 2/intermedin in the lung of rats with chronic hypoxic pulmonary hypertension].

AIM: To investigate the changes and probable roles of adrenomedullin2/intermedin (AIDM2/IMD), a novel micromolecular bioactive peptide, in the lungs of rats with chronic hypoxic pulmonary hypertension. METHODS: Twenty male SD rats were randomly divided into normal control group (NC) and normobaric hypoxia group (4H). The protein levels of ADM and ADM2/IMD) in the plasma and lung were measured by radioimmunoassay and immunohistochemistry. The mRNA expressions of ADM, ADM2/IMD and their receptors C (RLR, RAMP1, RAMP2 and RAMP3 in the lung tissue were determined by reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: (1) The rat model of chronic pulmonary hypertension was confirmed by the increased mean pulmonary arterial pressure (mPAP) and weight ratio of right ventricle to left ventricle plus septum [RV/(LV + S)] in 4H group compared to NC group. (2) The concentrations of ADM in the plasma and lung homogenate of 4H group were 2.3 and 3.2 folds of NC group, respectively (all P < 0.01). The levels of ADM2/IMD were higher 89.6% and 45.0% in the plasma and lung homogenate of 4H group than those of NC group (respectively, P < 0.01, P < 0.05). (3) The mRNA expressions of ADM2/IMD and ADM in the lung of 4H group were up-regulated (respectively, P < 0.01, P < 0.05 vs. NC group). The expressions of CRLR and RAMP1 mRNAs were down-regulated (all P < 0.01 vs. NC group), while the levels of RAMP2 and RAMP3 mRNAs were no significant difference between the two groups. (4) The strong ADM2/IMD immunostaining was detected in the endothelial and adventitial cells of the rat pulmonary arteriole. CONCLUSION: ADM2/IMD, like its paralog ADM, might be closely related to the chronic hypoxic pulmonary hypertension in rats. The disorders of the gene expression and/or the synthesis and metabolism of ADM2/IMD and its receptor CRLR/RAMP1 possibly take part in the pathogenesis of chronic hypoxic pulmonary hypertension in rats.

[Expression of Urotensin II and its receptor on right ventricle in rats of pulmonary hypertension].

AIM: To observe the expression of Urotensin II (U II) and its receptor (UT) on right ventricle in rats with chronic pulmonary hypertension induced by hypoxia and hypercapnia. METHODS: Twenty male SD rats were randomly divided into normal control group (NC) and hypoxia-hypercapnia 4-week group(HH). Mean pulmonary arterial pressure(MPAP) and the weight ratio of right ventricle (RV) to left ventricle plus septum (LV+ S) were calculated separately. U II in plasma was measured using radioimmunoassay. The expression of U II was observed in right ventricle myocytes and right ventricle arteries by immunohistochemistry. The expression of U II mRNA and UT mRNA were observed in right ventricle myocytes and right ventricle arteries by in situ hybridization. RESULTS: (1) The MPAP and RV/LV + S of HH group were higher respectively than those of NC group (P < 0.01, respectively). (2) The plasma U II content of HH group did not increased obviously than that of NC group. (3) The expression score of U II, U II mRNA, UT mRNA by right ventricle myocytes in HH group were higher significantly than those of NC group (P < 0.01 respectively). (4) The average value of integral light density (LD) of U II, U II mRNA, UT mRNA by right cardial arteries in HH group were higher significantly than those of NC group (P < 0.01, respectively). CONCLUSION: The expression of U II in right ventricle arteries and right ventricle myocytes increase significantly during the formation of pulmonary hypertension and right ventricle hypertrophy in rats chronically exposed to hypoxia-hypercapnia. These changes indicate that U II might be involved in right ventricle remodeling, which promotes proliferation of cardiac muscle cells.

[Effects and relationship between NO and HIF-1alpha in rats with pulmonary hypertension induced by hypoxia].

AIM: To investigate the expression of hypoxia-inducible factor-1 alpha (HIF-1alpha) in rats with chronic pulmonary hypertension induced by hypoxia and hypercapnia and its relationship with nitric oxide(NO). METHODS: Fourty male Sprague-Dawley rats were randomly divided into four groups, normal control group (NC), hypoxia-hypercapnia group (HH), hypoxia - hypercapnia + L-arginine liposome group(HP) and hypoxia-hypercapnia+ N-nitro-L-arginine methylester group (HM). Colorimetric analysis, immunohistochemistry and in situ hybridization were used for detection of NO, HIF-1alpha and constitutive nitric oxide synthase (ecNOS). RESULTS: (1) The mean pulmonary arterial pressure (mPAP) and the weight ratio of right ventricular to left ventricle plus septum (RV/(LV + S)) of HH group were higher than those of NC group (P < 0.05), HP group much lower than HH group (P < 0.01), mPAP of HM higher than HH group ( P < 0.05). 2)0 Contents of NO in plasma and pulmonary tissue homogenates of HH group were much lower than those of NC group (P < 0.01), HP group higher than HH group (P < 0.01). There were no difference between HM group and HH group. (Expression of HIF-1alpha and HIF-1alpha mRNA in pulmonary arterioles of HH group were significantly higher than those of NC group( P < 0.01), HP group lower than HH group (P < 0.01) ,HM group higher than HH group (P < 0.01); Whereas expression of ecNOS and ecNOS mRNA in pulmonary arterioles of HH were lower than those of NC group( P < 0.05, IP group higher than HH group (P < 0.01), HM group lower than HH group (P < 0.05). CONCLUSION: HIF-1alpha is involved in the pathogenesis of chronic pulmonary hypertension induced by hypoxia and hypercapnia. The protective function of NO in the pathogenesis might be partly depended on its effects on the expression/activity of HIF-1alpha in lung.