Role of heme oxygenase in cholecystokinin octopeptide ameliorating acute lung injury induced by lipopolysaccharide

AIM: To study the role of heme oxygenase (HO)-1 in the mechanism of cholecystokinin-octapeptide (CCK-8) for attenuation of acute lung injury (ALI) induced by lipopolysaccharide (LPS).METHODS: Adult male rats were randomly divided into five groups: control group,LPS group,CCK-8+LPS group,LPS+ Hm (hemin,HO-1 donor) group and LPS+ZnPP (zinc protoporphyrin,specific inhibitor of HO-1) group.PMN number in bronchoalveolar lavage fluid (BALF),the structure of the lung,MDA content,HO-1 activity,the expressions of HO-1 mRNA and protein in the lung were detected respectively.RESULTS: The lung injury in LPS group was observed,at the same time the numbers of PMN,the content of MDA,the activity and the expression of HO-1 were all higher than those in control group (all P<0.05).The degree of lung injury,PMN numbers and MDA content were lower,while the activity and the expression of HO-1 in CCK-8+LPS and LPS+Hm group were higher than those in LPS group (all P<0.05).However,the degree of lung injury,PMN numbers and MDA content were higher,the activity and the expression of HO-1 were lower in LPS+ZnPP than those in LPS group respectively (all P<0.05).CONCLUSION: CCK-8 attenuates the LPS-induced ALI by means of anti-oxidation and inhibits PMN aggregation,which are both mediated by HO-1 partly.     

Role of CD163/TWEAK pathway in atherosclerosis

AIM:To investigate the effect of CD163/tumor necrosis factor-like weak inducer of apoptosis (TWEAK) pathway on atherosclerosis in mice. METHODS:APOE^-/- mice and wild-type (WT) C57BL/6 mice were divided into 4 groups (8~10 weeks, n=10):APOE^-/- +normal diet (ND) group, APOE^-/- +western diet (WD) group, WT+ND group, and WT+WD group. Detection of blood lipid levels and oil red O staining of aorta artery were performed to confirm whether the atherosclerotic model was well established in 16 weeks after feeding. The aortic tissues were harvested to measure CD163 and TWEAK protein levels by Western blot, and immunohistochemical staining was also performed to localize CD163 and TWEAK protein expression on atherosclerotic plaque in each group. The cell experiments were conducted to study whether CD163 regulated TWEAK expression in M1 macrophages and foam cells, and the possible downstream pathway was investigated. RESULTS:The blood lipid levels and aorta oil red O staining showed that the animal model of atherosclerosis was successfully established in APOE^-/- +ND group and APOE^-/- +WD group. The protein level of CD163 was significantly increased in the aortic tissue in APOE^-/- mice (P<0.05) as compared with C57BL/6 mice (P<0.05). Consistently, the protein level of TWEAK was also markedly higher in APOE^-/- +ND group and APOE^-/- +WD group than that in WT+ND group and WT+WD group (P<0.05). Immunohistochemical staining showed that CD163 was mainly expressed in the parts away from the lipid core, and TWEAK was found in all parts of the atherosclerotic plaque. CD163 significantly inhibited the protein expression of TEWAK in the M1 macrophages, and also significantly down-regulated the level of nuclear factor-κΒ (NF-κB) in the M1 macrophages and foam cells (P<0.05). CONCLUSION:The protein levels of CD163 and its ligand TWEAK are significantly increased in atherosclerotic mice. The CD163 positive macrophages are mainly located at the site far away from the lipid core, and CD163 may play an anti-atherosclerotic effect by inhibiting TWEAK/NF-κB pathway.     

Delayed protection of HO-1 in the exercise preconditioning from the myocardial relative ischemic reperfusion injury

AIM:To explore the delayed protection of heme oxygenase-1 (HO-1) in the exercise preconditioning (EP) from the myocardial relative ischemia reperfusion injury (rI/R). METHODS:40 Wistar Rats were divided into 5 groups randomly:control group (CN), rI/R group (IR), EP+rI/R group (EI), HO-1 inductor hemin+rI/R group (HE) and HO-1 inhibitor ZnPP+EP+rI/R group (EZ). The following indexes were detected, including the HO-1activity in myocardium, the cardiac function parameter-pressure-rate product (heart rate × left ventricular developed pressure, PRP) and the content of MDA in coronary effluent. RESULTS:After myocardial rI/R, HO-1 activity increased significantly. Moreover, EP or HO-1 inductor could enhance this effect manifestly. Nevertheless, when the HO-1 inhibitor was administered before EP，HO-1 activity decreased. In addition, there was no distinct difference in the HO-1 activity between EI group and HE group. At the 30 min point of reperfusion, the PRP recovery rate of EI group was higher clearly than that of IR group. However, there was reverse effect between the EZ group and the EI group. The MDA in coronary effluent of EI group, EZ group and HE group were lower obviously than that of IR group and there was significant difference between EI group and EZ group. CONCLUSION:EP could protect the heart from the rI/R injury occurring 24 hours later, which might be performed through activating the HO-1.     

Probucol inhibits proliferation of rat aortic smooth muscle cells stimulated with oxidized low-density lipoprotein

AIM: To investigate the relationships between antiproliferative mechanisms of probucol and protein expressions of signaling molecules ERK1/2, MKP-1, HO-1 and Trx-1 in rat aortic smooth muscle cells (RASMCs) stimulated with ox-LDL. METHODS: The effects of probucol on cell cycle, cell proliferation and the expressions of ERK1/2, MKP-1, HO-1 and Trx-1 in the presence of ox-LDL were observed by means of MTT test, FCM and Western blotting. RESULTS: (1) Probucol significantly inhibited the proliferation of RASMCs stimulated with ox-LDL. A value in 100 μmol/L probucol+35 mg/L ox-LDL group was reduced by 34.9% as compared to ox-LDL group (P<0.01). (2) Probucol protected against ox-LDL-induced RASMCs proliferation through inducing cell growth arrest at G0/G1 phase and cell apoptosis. (3) ox-LDL increased the expression of p-ERK1/2 by 34.7% (P<0.01) and decreased MKP-1 by 60.0% (P<0.01), respectively, as compared to control. Probucol attenuated the increase in ox-LDL-stimulated p-ERK1/2 level by 15.7%, but increased MKP-1 expression by 2 times (P<0.01). (4)ox-LDL at concentration of 35 mg/L decreased the intracellular Trx-1 expression by 28.9% (P<0.05), and slightly increased the level of HO-1 expression as compared to control (P<0.05). Probucol enhanced the expression of Trx-1 by 91.6% (P<0.01) and HO-1 by 31.9% (P<0.01), respectively as compared to ox-LDL group. CONCLUSION: Probucol inhibits ox-LDL-stimulated the proliferation of RASMCs through increases in MKP-1/HO-1 expression, suppression of cell cycle progression and induction of cell apoptosis.     

CCK-8 up-regulats signal pathway of LPS-induced HO-1 expression in rat lungs

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.     

Role of heme oxygenase-1 in the ischemic preconditioning of isolated rat heart

AIM: To investigate the influence of ischemic preconditioning on heart function, the activities of lactate dehydrogenase (LDH), malondialdehyde (MDA), and heme oxygenase-1 (HO-1) after ischemia/reperfusion in isolated rat heart. METHODS: The model of Langendorff was used in isolated rat heart perfusion. Ischemic preconditioning protocol: stopping perfusion for 5 minutes and reperfusion for 5 minutes, repeating three times. Ischemia protocol: stopping perfusion for 40 minutes and reperfusion for 20 minutes. Indexes of heart function were recorded in control M8, ischemia and reperfusion group (IR), and ischemic preconditioning group (IPC). The content of LDH of coronary effluent was measured. Moreover, the content of MDA and activity of HO-1 in myocardium were also measured. RESULTS: The recovery percentage of heart function in IPC group was significantly higher than that in IR group (P<0.01) and the activity of heme oxygenase-1 also increased significantly (P<0.05). CONCLUSION: The contents of LDH and MDA significantly decreased in IPC group compared with IR group. The increase in heme oxygenase-1 activity might be involved in the protective effect of ischemic preconditioning on ischemic/reperfused rat heart.     

Relationship between expression of heme oxygenase-1 mRNA and pulmonary hypertention induced by hypoxic hypercapnia

AIM: To study the effect of chronic hypoxic hypercapnia on expression of heme oxygenase-1 (HO-1). METHODS: Sprague-Dawley rats were randomly divided into three groups: control group(A),hypoxic hypercapnic group(B), hypoxic hypercapnia+hemin group(C). HO-1 and HO-1 mRNA were observed in pulmonary arterioles by the technique of immunohistochemistry and in situ hybridization. RESULTS: ① mPAP and weight ratio of right ventricle (RV) to left ventricle plus septum (LV+S) were significantly higher in rats of B group than those of A and C group (P<0.01). Differences of mCAP were not significant in three groups(P>0.05). ② Blood CO concentration was significantly higher in rats of B group than that of A group (P<0.01), it was much higher in C group than that of B group(P<0.01). ③ Light microscopy showed that vessel well area/total area (WA/TA), density of medial smooth muscle cell (SMC) and media thickness of pulmonary arterioles were much higher in rats of B group than those of A and C group (P<0.01). ④ The observation by electron microscopy showed proliferation of medial smooth muscle cells and collageous fibers of pulmonary arterioles in rats of B group, hemin could reverse the changes mentioned above. ⑤ HO-1 and HO-1 mRNA in pulmonary arterioles was significantly higher in rats of B group than those of A group(P<0.01), and they were significantly higher in rats of C group than those of B group (P<0.01). CONCLUSION: Expression of HO-1 mRNA and HO-1 in pulmonary arterioles was enhanced by hypoxic hypercapnia. Hemin partly inhibited pulmonary hypertension and pulmonary vessel remodeling by enhancing the expression of HO-1 mRNA and HO-1.     

Effect of combination of Penthorum chinense Pursh and Puerariae flos on L-02 liver cell injury induced by alcohol

AIM: To explore the effect of Penthorum chinense Pursh and Puerariae flos-containing serum on L-02 liver cell injury induced by alcohol and its possible mechanism. METHODS: After preparing drug-containing serum, the L-02 cells cultured in vitro were divided into 6 groups:blank control group, model group, 1:1 group, 2:1 group and 1:2 group of combination of Penthorum chinense Pursh and Puerariae flos, and tiopronin group. The viability of the L-02 cells was measured by MTT assay. The activity of alanine aminotransferase (ALT), aspartate aminotransferase (AST) and superoxide dismutase (SOD), and the content of malondialdehyde (MDA) were detected by enzyme label methods. The expression of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) at mRNA and protein levels was determined by real-time PCR and Western blot, respectively. RESULTS: Compared with control group, the levels of ALT, AST and MDA were increased significantly, and SOD was decreased in model group (P<0.01). Compared with model group, these indexes in all treatment groups were opposite (P<0.01). Compared with control group, the expression of TNF-α and IL-6 at mRNA and protein levels was significantly increased, the mRNA and protein expression of Nrf2 and HO-1 was decreased (P<0.01). Compared with model group, these indexes in combination groups were opposite (P<0.01). CONCLUSION: The therapeutic effects of Pentehorum chinensa Pursh and Puerariae flos-containing serum may affect the expression levels of TNF-α, IL-6, Nrf2 and HO-1, and reduce the inflammatory reaction and oxidative stress in alcohol-induced L-02 liver cells, which plays a role in attenuating alcoholic liver injury.     

Dexmedetomidine attenuates acute kidney injury induced by hemorrhagic shock/resuscitation in rats

AIM: To investigate the effects of dexmedetomidine on hemorrhagic shock/resuscitation (HS/R)-induced acute kidney injury (AKI) in rats, and to explore the possible mechanisms. METHODS: Wistar rats (n=32) were randomly divided into 4 groups (n=8):normal saline control group (NS group), dexmedetomidine group (D group), HS/R group and HS/R+D group. The animals were sacrificed at 6 h after resuscitation. The levels of serum creatinine (Cr) and blood urine nitrogen (BUN) were examined. The kidneys of all rats were removed for evaluation of histological characteristics, and the levels of malondialdehyde (MDA), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and superoxide dismutase (SOD) were measured. The expression of nuclear factor-κB (NF-κB) and hemeoxygenase-1 (HO-1) was determined by Western blot. RESULTS: Compared with NS group, the levels of Cr, BUN, MDA, TNF-α and IL-1β were obviously increased in HS/R group, which were obviously decreased in HS/R+D group (P<0.05). Compared with NS group, the SOD activity was obviously decreased in HS/R group, which was obviously increased in HS/R+D group (P<0.05). Compared with NS group, the protein expression of NF-κB was obviously increased in HS/R group, which was obviously decreased in HS/R+D group (P<0.05). Compared with NS group, the protein expression of HO-1 was increased in HS/R group. Compared with HS/R group, the protein expression of HO-1 was obviously increased in HS/R+D group. Compared with NS group, HS/R induced marked kidney histological injury, which was less pronounced in HS/R+D group.CONCLUSION: Dexmedetomidine effectively protects rats against AKI caused by HS/R, and its mechanism may be associated with the increase in HO-1 expression and the inhibition of NF-κB expression.     

Effects of pituitary adenylate cyclase-activating polypeptide on cardiac remodeling in coronary atherosclerotic rabbits

AIM:To study the intervention effects of pituitary adenylate cyclase-activating polypeptide (PACAP) on cardiac remodeling during the development of rabbit coronary atherosclerosis. METHODS:60 male New Zealand rabbits were equally divided into 3 groups randomly: control group (C group), atherosclerosis model group (A group) and PACAP intervention group (P group). At the 4th, 8th and 12th week, 5-6 cases of rabbits in each group were sacrificed, cardiac tissue with coronary arteries were harvested to make paraffin sections. The sections were stained with hematoxylin-eosin and van Gieson separately. The qualitative observation and/or quantitative analysis were made by light microscope. RESULTS:（1）There was no lesion in C group. For A group and P group, there were plaques in large epicardial coronary arteries and small coronary arteries; an impressive accumulation of collagen was also observed in myocardium. In P group, the lesions of small coronary arteries were less serious, and the degrees of perivascular and myocardial fibrosis also appeared to be less.（2）For A group, the wall-to-lumen ratios in small coronary arteries were significantly greater at the 12th week (2.58±1.54) than C group (1.34±0.58) and P group (1.39±0.48) （P<0.05）； and the width of cardiomyocyte (13.85 μm±2.27 μm) was already remarkably narrower than C group (14.68 μm±2.40 μm) at the 8th week (P<0.05) and narrower significantly than C group and P group at the 12th week. （3）There were not difference significantly between the above-related parameters of P group and C group (P>0.05). CONCLUSION:Structure changes exist in coronary arteries and myocardium during the development of rabbit coronary atherosclerosis, PACAP can inhibit the cardiac remodeling.     

Effect of mesenchymal stem cells transfected with human heme oxygenase-1(HO-1)gene on myocardial apoptosis and angiogenesis

AIM: To investigate the effects of mesenchymal stem cells(MSCs)transfected with human heme oxygenase-1(HO-1)gene on myocardial apoptosis and angiogenesis. METHODS: MSCs were acquired from the bone marrow of adult rats. The cells were isolated, purified, cultured, and transfected with Adv-HO-1 in vitro before transplantation. At 1 h after left coronary artery ligation, Adv-HO-1-MSCs or MSCs were directly injected into the border of cardiac infarction in rats. Western blotting analysis was used to measure HO-1, and Bax protein expression in the border of cardiac infarction. ELISA was used to measure the expressions of VEGF and bFGF in the border of cardiac infarction. At 4 weeks after transplantation, the heart functions in survival rats were examined by the Buxco system. The rats were killed, then the myocardial infarct size was measured with Masson’s trichrome, and the expression of CD34 in myocardial infarction area was detected by immunohistochemical method. RESULTS: HO-1-MSCs exhibited increased HO-1 expression. The expression of HO-1, VEGF and bFGF in the border of cardiac infarction in the rats treated with HO-1-MSCs were higher than those in the rats treated with MSCs and PBS(P<0.01). However, the expression of apoptotic protein Bax was significantly lower than that in the rats treated with MSCs and PBS(P<0.01). The number of capillary vessels in the border of cardiac infarction in the rats treated with HO-1-MSCs was significantly higher than that in the rats treated with MSCs and PBS. The cardiac function in the rats treated with HO-1-MSCs was better than that in the rats treated with MSCs and PBS(P<0.01). CONCLUSION: The favorable effect on heart function appears to be a combined outcome of HO-1 and paracrine factors released by MSCs.     

Changes of hydrogen sulfide and heme oxygenase-1 in blood of children with congenital heart disease and pulmonary hypertension before and after surgery

AIM: To evaluate the changes and clinical significance of hydrogen sulfide (H2S) and heme oxygenase-1 (HO-1) in patients with congenital heart disease (CHD) and pulmonary hypertension (PH). METHODS: Forty-eight patients with CHD and left-to-right shunt were selected randomly from cardiac surgery in the Fourth Hospital of Hebei Medical University. Forty-eight patients were divided into 3 groups according to the pulmonary artery systolic pressure (PASP) by Doppler echocardiography before operation: group A was normal (PASP<30 mmHg, n=15); group B was mild PH (PASP 30-49 mmHg, n=15); group C was moderate-severe PH (PASP≥50 mmHg, n=18). The radical operation was performed successfully under complex intravenous anesthesia in all 48 patients. The radial arterial blood was collected immediately before operation, 1 h and 24 h after operation. The concentration of plasma H2S was measured by optical absorbance method at 670 nm and HO-1 activity in serum was detected by dual wavelength spectrophotometer at different time points. RESULTS: Plasma level of H2S in all groups after operation was higher than that in the same group before operation. Significant difference of the plasma H2S level was observed between pre-operation and 24 h post-operation in the same group (P<0.05). The plasma level of H2S in group C and group B at time points of pre-operation, 1 h post-operation or 24 h post-operation was significantly lower than that in group A, the plasma level of H2S in group C was significantly lower than that in group B. Compared to the values in each group before operation, the activity of serum HO-1 among groups after operation was not obviously and statistically different (P>0.05). No obvious difference of HO-1 activity in blood among 3 groups was observed (P>0.05). There was a negative correlation between PASP and the level of H2S in 3 groups at time points of pre-operation, 1 h and 24 h post-operation (pre-operation, r=-0.66, P<0.01; 1 h post-operation, r=-0.458, P<0.01; 24 h post-operation, r=-0.730, P<0.01). No correlation between PASP and HO-1 activity in blood serum was found. CONCLUSION: H2S plays an important role in the formation of PH and remodeling of pulmonary vasculature. There is no correlation between PASP and HO-1 activities in blood, but HO-1 may play an indirect action in the formation of PH and remodeling of pulmonary vasculature. Measuring the level of H2S may be a reliable method to follow the change of pulmonary pressure and worsened PH.     

Changes of ceramide and apoptosis during myocardial ischemia/reperfusion

AIM: To study the change of myocardial ceramide during myocardial ischemia/reperfusion and the relationship between ceramide and apoptosis and oxidative stress. METHODS: After inducing myocardial ischemia/reperfusion (I/R) injury in mice with pituitrin (Pit), myocardial SOD activity and MDA content were measured. DNA agarose gel electrophoresis and fluorescent staining of DAPI were done to check up apoptosis. The content of myocardial ceramide (μg/kg) was measured through HPTLC and scan of thin plate. RESULTS: The myocardium of I/R model group had the phenomenon of DNA ladder. Apoptosis index and ceramide content in I/R model group were higher than those in normal control group (P<0.01). SOD activity in I/R modal group was lower than that in normal control group (P<0.01). The apoptosis index and ceramide content in I/R model group were positive correlative (r=0.970，P<0.01). The myocardial content of ceramide and MDA were positively correlative too (r=0.974, P<0.01). CONCLUSION: The results indicate that there are apoptosis, oxidative stress and increase in ceramide content in ischemia/reperfusion myocardium.     

Erianin induces apoptosis of human lung cancer A549 cells via ROS/p38 MAPK pathway

AIM:To investigate the effect of erianin on the viability and apoptosis of human lung cancer A549 cells and its possible mechanism. METHODS:A549 cells and BEAS-2B cells were cultured in vitro and treated with erianin at 10, 20, 40, 80 and 160 nmol/L for 48 h. The cell viability was measured by CCK-8 assay. The apoptosis and reactive oxygen species (ROS) were analyzed by flow cytometry. The activity of superoxide dismutase (SOD) was detected by WST-8 method, and the content of malondialdehyde (MDA) was detected by barbituric acid method. The protein levels of nuclear factor E2-related factor 2 (Nrf2), NAD(P)H:quinone oxidoreductase-1 (NQO1), heme oxygenase-1 (HO-1), p38 MAPK, p-p38 MAPK, caspase-3 and cleaved caspase-3 were determined by Western blot.RESULTS:Erianin remarkably reduced the viability of A549 cells in a dose-dependent manner (P<0.05) with IC₅₀ at 52.64 nmol/L. Erianin also induced apoptosis (P<0.05), increased ROS level and MDA content (P<0.05), diminished SOD activity (P<0.05), and down-regulated the protein levels of Nrf2, NQO1 and HO-1 (P<0.05), in a dose-dependent manner. Meanwhile, erianin up-regulated the levels of p-p38 MAPK and cleaved caspase-3 (P<0.05), and these effects were inhibited by N-acetyl-L-cysteine and SB203580 (P<0.05).CONCLUSION:Erianin may induce apoptosis of human lung cancer A549 cells most likely via inhibiting SOD activity and down-regulating the protein levels of Nrf2, NQO1 and HO-1, thus resulting in an increase in ROS and activation of p38 MAPK.     

Premature response to luteinizing hormone of granulosa cells from women with polycystic ovary syndrome

AIM:To demonstrate the relationship between hormones in follicular fluid and the expression of LH receptor in granulosa cells (GC) in anovulatory women with polycystic ovary syndrome (PCOS). METHODS:Follicles were obtained from 12 women with PCOS and 15 women with normal menstrual period through surgery at time between day 7 and day 10 of menstrual cycle. The accumulations of estrogen (E2), progesterone (P), luteinizing hormone (LH), follicle stimulating hormone (FSH) and insulin in follicular fluid were determined by a automatism chemiluminescent microparticle immunoassay (CMIA) for the quantitative determination. The accumulation of androstenediol (A) was determined by ELISA. The amounts of the mRNA expressions of LH receptors from GC and theca cells (TC) respectively were measured by RT-PCR using β-actin as intra-control simultaneously. RESULTS:The levels of LH ［(3.8±2.1 vs 1.7±0.8)IU/L, P<0.01］, A ［(600.0±373.4 vs 212.4±205.4)μg/L, P<0.05］ and expressions of LH receptor mRNA of GC (0.29±0.16 vs 0.12±0.13, P<0.01) and TC (0.46±0.14 vs 0.34±0.09，P<0.05) in the women of PCOS group were statically higher than those in control group. The expression of LH receptor mRNA was not detected by RT-PCR in control group when the diameter of an follicle was less than 7 mm, while it was detected in women with PCOS when it remained as small as 4 mm. Expression of LH receptor mRNA in granulose cells was positive related to the concentration of LH (r=0.67, P<0.01) and insulin (r=0.51, P<0.05) in follicular fluid, and that in theca cells (r=0.60, P<0.01). CONCLUSION:The high level of LH in follicular fluid occurs and GC responds to LH prematurely and more intensively in anovulatory women with PCOS. Larger amount of A and P was produced as a result. All of above may contribute to the mechanism of anovulatory.     

Pulmonary function in patients with type 2 diabetes mellitus

AIM: To evaluate the pulmonary function in patients with type 2 diabetes mellitus in order to identify whether the lung is a target organ of chronic pathologic changes in diabetes mellitus. METHODS: Pulmonary ventilation function and diffusion capacity were studied in 107 patients with type 2 diabetes mellitus and 61 healthy subjects matched for age and sex. Glycosylated hemoglobin (HbA1c), urine albumin excretion rate (AER), fundus examination and nerve conduction velocity were included as parameters of glycemic control and diabetic microangiopathies. RESULTS: Pulmonary ventilation function was similar in type 2 diabetic group and the control. Compared with the control, carbon monoxide diffusion capacity (DLCO) and DLCO corrected by alveolar volume (DLCO/VA) were significantly lower in type 2 diabetic group (P<0.05). DLCO and DLCO/VA were inversely correlated with microangiopathy score (r: -0.291, -0.324， respectively, P<0.01). Furthermore, DLCO/VA was negatively correlated with age and duration of diabetes mellitus (r: -0.269, -0.236， respectively, P<0.05). CONCLUSIONS: Pulmonary ventilation function is normal in patients with type 2 diabetes mellitus, but their diffusion capacity is impaired. It suggests that the lung may also be the target organ of the chronic pathologic changes of diabetes mellitus.     

Plasma levels of endothelin-1, nitric oxide,malondialdehyde and superoxide dismutase in rats exposed to infrasound

AIM: To observe the changes of NO, ET-1, SOD and MDA levels in plasma of rats exposed to infrasound. METHODS: Using infrasound (frequency: 8 Hz; sound pressure level:130 dB), the rats were exposed for 1 d， 7 d， 14 d， 21 d and 28 d, 2 h daily, then the levels of NO, ET-1, SOD and MDA were measured after exposure. RESULTS: The changes of NO levels in plasma significantly declined at 7 d and 14 d （P<0.01）, then 1, 21 and 28 d normally （P>0.05）. The changes of ET-1 levels in all groups in plasma were significantly increased （P<0.01）, mostly at 7 d, least at 14 d. The changes of SOD activity in all groups in plasma were significantly declined （P<0.01）. The changes of MDA levels in all groups in plasma were significantly increased （P<0.01）. CONCLUSIONS: Infrasouns induces changes of NO, ET-1, SOD and MDA in rat plasma, and it depends on infrasound exposure time.     

Naringenin attenuates myocardial injury by regulating AMPK/Nrf2/HO-1 signaling pathways in diabetic mice

AIM: To investigate the effects of naringenin (NAR) on the myocardium as well as its effects on adenosine monophosphate-activated protein kinase (AMPK)/nuclear factor (NF)-E2-related factor 2 (Nrf2)/heme oxygenase 1 (HO-1) signaling pathways in diabetic mice. METHODS: C57BL/6 mice (n=50) were randomly divided into normal group (N group) and model group. The mouse model of type 1 diabetes mellitus was established by intraperitoneal injection of streptozotocin (STZ), then the mice were divided into diabetes group (D group), diabetes+low dose of NAR intervention group (D+L-NAR group), diabetes+middle dose of NAR intervention group (D+M-NAR group) and diabetes+high dose of NAR intervention group (D+H-NAR group). The mice in intervention groups were received NAR at low, middle and high doses respectively by gavage, and the mice in N group and D group were received equal volume of normal saline. After 6 weeks, the mice were sacrificed to observe the effects of NAR at different doses on the body weight and blood glucose. The histopathological changes of the cardiac tissues were observed with HE staining. The myocardial collagen volume fraction (CVF) was calculated by Masson staining. Immumohistochemical staining was used to test the protein levels of interleukin-6 (IL-6) and IL-10, and the TUNEL was used to observe the apoptosis of myocardial tissues. The production of reactive oxygen species (ROS) in the myocardial cells was analyzed by fluorescence probe of DHE, and superoxide dismutase (SOD) activity and malondiodehyde (MDA) content in the myocardial cells were measured by SOD and MDA kits. Western blot was applied to determine the protein levels of p-AMPKα, AMPKα, Nrf2, HO-1, NAD(P)H:quinone oxidoreductase 1 (NQO1) and cleaved caspase-3 in the myocardial tissues. RESULTS: Compared with N group, the blood glucose of the mice in D groups was increased and the body weight was decreased significantly. Compared with D group, the blood glucose of the mice in NAR intervention groups was decreased and the body weight was increased. Compared with N group, the CVF, apoptotic rate and the protein levels of IL-6, cleaved caspase-3 were increased, while the protein levels of IL-10, p-AMPKα, Nrf2, HO-1, NQO1 and SOD activity were decreased, the ROS production rate and MDA content was increased significantly in D group (P < 0.05). Compared with D group, the CVF, apoptotic rate and the protein levels of IL-6, cleaved caspase-3 were relatively decreased, conversely the protein levels of IL-10, p-AMPKα, Nrf2, HO-1, NQO1 were increased in NAR intervention groups(P < 0.05). No significantly difference of the ROS production rate, SOD activity and MDA content between D group and D+L-NAR group was observed. However, the ROS production rate and MDA content was decreased,SOD activity were increased in D+M-NAR group and D+H-NAR group as compared with D group. CONCLUSIONS: NAR attenuates myocardial injury in diabetic mice, and its mechanism may be related to regulation of AMPK/Nrf2/HO-1 signaling pathway, enhancement of the antioxidant reaction, reduction of myocardial fibrosis, apoptosis and inflammation.     

Transfection of mesenchymal stem cells with human heme oxygenase-1 gene improves survival under conditions of serum-free and hypoxia

AIM: To investigate the effects of human heme oxygenase-1 (HO-1) gene transfection on mesenchymal stem cells (MSCs) survival under the conditions of serum-free and hypoxia. METHODS: MSCs were acquired from the bone marrow of adult rats. The cells were isolated, purified, cultured, and transfected with Adv-HO-1. The expression of GFP was detected by immunofluorescence. Cell apoptosis was detected by nuclear DAPI staining and FACS. The concentrations of VEGF, HGF and b-FGF in the culture supernatant were measured by ELISA. The caspase-3 protein level and activity of cardiomyocytes cultured with the supernatants from different MSCs under the condition of serum-free and hypoxia were assayed by Western blotting and fluorimetry, respectively. RESULTS: HO-1-MSCs exhibited strong expression of GFP. The fragmented or condensed chromatin diminished in HO-1-MSCs compared with the MSCs lacking exogenous transfection of HO-1 gene. A lower proportion of apoptosis was observed in HO-1-MSCs compared with MSCs under the conditions of serum-free and hypoxia (P<0.01). The expressions of VEGF, HGF and b-FGF in the supernatants of HO-1-MSCs were higher than those in MSCs (P<0.01). A significant reduction of caspase-3 level and activity in the cardiomyocytes treated with the supernatants from HO-1-MSCs was observed, compared to that treated with supernatants from MSCs (P<0.01). CONCLUSION: HO-1 improves the MSCs survival under the conditions of serum-free and hypoxia. Several cytokines released by HO-1-MSCs may protect the cardiomyocytes against apoptosis.