P2X7 receptor antagonist improves gastrointestinal disorders in spontaneously hypertensive rats

The purinergic system participates in the control of blood pressure. Hypertension promotes the occurrence of gastrointestinal disorders such as intestinal inflammation and gastric emptying delay. This study aimed i) to investigate the participation of the P2X7 receptor blocker Brilliant Blue G (BBG) on gastric emptying of solids and changes in oxidative stress in the gastric fundus, duodenum, and colon of spontaneously hypertensive rats (SHR) and ii) to study the putative relationship of this effect with the renin-angiotensin system. Rats were divided into five groups: Control, SHR, SHR+BBG, SHR+BBG+ATP, and SHR+BBG+ANG II. In the gastrointestinal tract, we assessed gastric emptying (GE) and oxidative stress markers (NOx, MPO, GSH, SOD). We observed a decrease in the GE rate (P<0.05) in SHR vs control rats (21.8±2.0% vs 42.8±3.5%). The decrease in GE was returned (P<0.05) to control levels by BBG in SHR rats (21.8±2.0% vs 41.6±3.2%). Co-administration of ATP or ANG II together with BBG bypassed the effect of the P2X7 antagonist on GE in SHR (P<0.05) (21.9±5.0% vs 25.6±3.0% vs 41.6±3.2%). The MPO activity increased (P<0.05) in the gastric fundus of SHR compared to control rats (6.12±2.26 vs 0.077±0.02 UMPO/mg tissue); this effect was prevented (P<0.05) by BBG (0.55±0.15 vs 6.12±2.26 UMPO/mg tissue). Data demonstrated that blockage of P2X7 receptors with BBG can improve the GE delay and oxidative stress biomarkers in SHR animals. This preventive effect of BBG on GE delay was abrogated by ANG II and ATP, thus prompting crosstalk between renin-angiotensin and the purinergic signaling systems underlying this phenomenon.

Liquid chromatography-mass spectrometry method for the quantification of an anti-sclerostin monoclonal antibody in cynomolgus monkey serum / 药物分析学报

Liquid chromatography tandem mass spectrometry (LC-MS/MS) has gradually become a promising alternative to ligand binding assay for the bioanalysis of biotherapeutic molecules,due to its rapid method development and high accuracy.In this study,we established a new LC-MS/MS method for the determination of the anti-sclerostin monoclonal antibody (SHR-1222) in cynomolgus monkey serum,and compared it to a previous electrochemiluminescence method.The antibody was quantified by detecting the surrogate peptide obtained by trypsin digestion.The surrogate peptide was carefully selected by investigating its uniqueness,stability and MS response.The quantitative range of the pro-posed method was 2.00-500 μg/mL,and this verified method was successfully applied to the tox-icokinetic assessment of SHR-1222 in cynomolgus monkey serum.It was found that the concentrations of SHR-1222 in cynomolgus monkeys displayed an excellent agreement between the LC-MS/MS and electrochemiluminescence methods (ratios of drug exposure,0.8-1.0).Notably,two monkeys in the 60 mg/kg dose group had abnormal profiles with a low detection value of SHR-1222 in their individual sample.Combining the high-level anti-drug antibodies (ADAs) in these samples and the consistent quantitative results of the two methods,we found that the decreased concentration of SHR-1222 was due to the accelerated clearance mediated by ADAs rather than the interference of ADAs to the detection platform.Taken together,we successfully developed an accurate,efficient and cost-effective LC-MS/MS method for the quantification of SHR-1222 in serum samples,which could serve as a powerful tool to improve the preclinical development of antibody drugs.

Caracterização do tecido ósseo peri-implantar em ratas hipertensas e com deficiência de estrógeno, após ação sinérgica de losartan e alendronato de sódio / Characterization of peri-implant bone tissue in hypertensive and estrogen-deficient rats after the synergistic action of losartan and alendronate sodium

Introdução: Este estudo visou caracterizar morfológica e estruturalmente o tecido produzido na interface osso-implante em ratas espontaneamente hipertensas ovariectomizadas com implantes instalados em suas tíbias, e analisou como o tratamento associado de losartan sistêmico e alendronato de sódio local influenciou no reparo ósseo peri-implantar. Material e Métodos: Foram utilizadas ratas espontaneamente hipertensas (SHR) que receberam losartan (30 mg/kg, p.o.). Após uma semana, implantes de titânio tratados (Medens, Ribeirão Preto, São Paulo, Brazil) ou não com alendronato de sódio (ALE) foram instalados nas tíbias. Sessenta dias após a implantação, a estabilidade do implante foi avaliada pela medição de torque reverso, considerado como desfecho primário. A microtomografia computadorizada e a análise por confocal foram parâmetros secundários. Resultados: A ação sinérgica do losartan e do alendronato de sódio na superfície do implante aumentou o torque reverso no grupo SHR SHAM ALE. Enquanto a microtomografia também revelou maior extensão de contato entre osso e implante, volume ósseo e espessura trabecular nos animais SHR SHAM ALE. Por último, o losartan e o alendronato de sódio não alterou significativamente os parâmetros de osseointegração nas ratas ovariectomizadas. Conclusões: Os resultados apresentados sugerem que a ação sistêmica do losartan somada à atuação local do alendronato de sódio na superfície dos implantes melhoram os parâmetros de osseointegração em tíbias de ratas hipertensas e não ovariectomizadas(AU)

Background: This study aims to characterize morphologically and structurally the tissue produced at the bone-implant interface in spontaneously hypertensive ovariectomized rats that will have implants placed in their tibiae, and to analyze how the associated treatment of systemic losartan and local sodium alendronate might influence the peri-implant bone healing. Methods: They are used spontaneously hypertensive (SHR) rats that received losartan (30 mg/kg, p.o.). After one week, titanium implants treated (Medens, Ribeirão, São Paulo, Brazil) or not with sodium alendronate (ALE) were installed in the tibiae. Sixty days after implantation, implant stability was assessed by measuring the removal torque considered the primary end point. Computed tomography and confocal analysis were secondary parameters. Results: The synergistic action of losartan and sodium alendronate on the implant surface increased the reverse torque in the SHR SHAM ALE group. While microtomography also revealed a greater extent of contact between bone and implant, bone volume and trabecular thickness in SHR SHAM ALE animals. Finally, losartan and sodium alendronate did not significantly alter osseointegration parameters in ovariectomized rats. Conclusions: The results presented suggest that systemic losartan plus the local action of sodium alendronate on implants surface improves osseointegration parameters in tibias of hypertensives and non-ovariectomized rats(AU)

Electrocardiographic Abnormalities in Hypertension Models

Abstract Background: Hypertensive condition can lead to abnormalities in heart structure and electrical activity. The electrocardiogram (ECG) is a recording of the electrical activity of the heart and widely used to diagnose and detect heart problem. Objective: We conducted a comparative ECG analysis between two hypertension models (L-NAME and SHR) and their controls (Wistar and Wistar-Kyoto) at six and 15 th week of age. Methods: Blood pressure was measured at the end of the 15 th week, and electrocardiography was performed at six and 15 weeks of age in anaesthetized rats. Data normality was confirmed by Kolmogorov-Smirnov test followed by unpaired Student's t-test and the Mann-Whitney for parametric and non-parametric data, respectively. Results are expressed as mean ± SD. The accepted level of significance was set at p < 0.05. Results: L-NAME exhibited prolongation of JT and QT intervals and SHR showed a decrease in heart rate when compared to Wistar-Kyoto and L-NAME. Wistar-Kyoto exhibited short PR interval with increased QRS complex, and only QT prolongation at 15 weeks compared to Wistar. Conclusions: All the hypertension models used in this study featured an increase in blood pressure. However, while SHR showed cardiac dysfunction, L-NAME exhibited changes in ventricular performance. These results may guide future studies on different types and models of hypertension.

A via Nrf2 participa da maior modulação endotelial induzida pela prenhez sobre reatividade de aortas à fenilefrina / The Nrf2 pathway participates in the greater endothelial modulation induced by pregnancy on reactivity of aortas to phenylephrine

Alterações em diferentes vias de sinalização levam a disfunção vascular e endotelial e consequentemente a hipertensão. A hipertensão está relacionada diretamente com o aumento da produção de espécies reativas de oxigênio (ROS) e diminuição da biodisponibilidade de óxido nítrico (NO) nos vasos sanguíneos. A via do Nrf2 (fator nuclear eritróide 2) está envolvida nos mecanismos que levam ao aumento da biodisponibilidade vascular de NO, pois controla a expressão de enzimas antioxidantes. A ativação do Nrf2 é modulada por sua ligação com a Keap-1 (Kelch-like ECH-associated protein 1) e sua atividade é modulada pelo fator de transcrição Bach-1, que compete pelo mesmo sitio ativo no DNA com o Nrf2. Em ratas normotensas e em ratas espontaneamente hipertensas (SHR) é observada uma redução da pressão arterial ao final da prenhez, que tem sido associada à redução do estresse oxidativo e maior biodisponibilidade de NO. Com o aumento da biodisponibilidade de NO, é aumentada a modulação do endotélio sobre a reatividade vascular à agonistas vasoconstritores, como à fenilefrina (PE). Levantamos a hipótese que a prenhez altera a expressão e ou a atividade do Nrf2 e de seus inibidores Keap-1 e Bach-1 e que estas possíveis alterações estariam associadas à maior modulação endotelial sobre contração de aortas à PE. Para testarmos esta hipótese, a expressão de Nrf2, Keap-1 e Bach-1 e também das enzimas antioxidantes transcritas pelo Nrf2, como a NADP(H) quinona oxirredutase-1 (NQO1), SOD-1 e SOD-2 foram avaliadas em aortas de ratas prenhes e comparadas as aortas de ratas não-prenhes. A fim de identificarmos outros possíveis mecanismos alterados pela prenhez em ratas Wistar e SHR, avaliamos a expressão de NOXO-1, subunidade regulatória da NOX1 e de p47phox, subunidade regulatória de NOX2. A participação do Nrf2 na produção de NO endotelial em aortas de ratas prenhes, foi avaliada pela utilização de Brusatol, uma droga inibidora do Nrf2. Avaliamos também a participação do Nrf2 na reatividade de aortas de ratas prenhes à fenilefrina e à acetilcolina, utilizando o Brusatol. Todos os resultados foram comparados entre ratas não-prenhes normotensas (Wistar) e hipertensas (SHR) e entre ratas não-prenhes e prenhes nos grupos (análise de multivariância, post-test Tukey, p< 0,05). Os resultados mostraram que a expressão de Nrf2 está aumentada em aortas de ratas prenhes Wistar, apesar da expressão de Keap-1 e de Bach1 não estar alterada. Associado a expressão aumentada de Nrf2 observamos maior expressão de SOD-2, mas não de SOD-1 ou NQO1, em aortas de ratas prenhes. Em aortas de ratas SHR não prenhes, observamos entre todas as proteínas avaliadas, menor expressão de Bach-1 e de NQO1 quando comparadas às aortas de ratas normotensas. A prenhez reduziu ainda mais a expressão apenas de NQO1 em aortas de SHR. A prenhez reduziu a expressão de NOXO-1 e de p47phox em aortas de SHR, enquanto que em aortas de ratas Wistar reduziu apenas a expressão de NOXO-1. Os resultados obtidos neste estudo mostraram também que a incubação de HUVEC com Brusatol aumentou as concentrações intracelulares de ERO, mas não alterou as concentrações de NO, no entanto, reduziu significativamente a concentração de NOx estimulada pela ACh em aortas de ratas prenhes, Wistar ou SHR. Além disto, o Brusatol aumentou a reatividade à PE em aortas de ratas normotensas não prenhes e prenhes, igualando a reatividade de aortas de ratas prenhes as aortas de ratas não prenhes. No entanto, o Brusatol não alterou a reatividade de aortas de SHR, prenhes ou não-prenhes. Nenhum efeito significativo do Brusatol foi observado na reatividade à Acetilcolina em aortas de ratas Wistar ou SHR, prenhes ou não prenhes. Em conclusão, nossos resultados sugerem que a atividade da via de sinalização do Nrf2 está aumentada, favorecendo a maior atividade de enzimas antioxidantes como a SOD-2, que contribuiria para maior biodisponibilidade de NO e maior modulação endotélio-dependente da contração vascular à PE em aortas de ratas normotensas prenhes. No entanto, em aortas de SHR prenhes, este mecanismo parecer não ser mais importante que a redução da atividade de isoformas NOX e de suas subunidades regulatórias que contribuiria para menor geração de O2•- e consequentemente, maior biodisponibilidade de NO(AU)

Modifications in different signaling pathways lead to vascular and endothelial dysfunction and, consequently, hypertension. Hypertension is directly related to an increase in the production of reactive oxygen species (ROS) and a decrease in the bioavailability of nitric oxide (NO) in blood vessels. The Nrf2 (erythroid nuclear factor 2) pathway is involved in the mechanisms that lead to the increased vascular bioavailability of NO, as it controls the expression of antioxidant enzymes. The activation of Nrf2 is modulated by Keap-1 (Kelch-like ECH-associated protein 1) and its activity is modulated by the transcription factor Bach-1, which competes for the same active site in DNA with Nrf2. In normotensive rats and spontaneously hypertensive rats (SHR), a reduction in blood pressure at the end of pregnancy is observed, which has been associated with a reduction in oxidative stress and greater bioavailability of NO. This increased NO bioavailability has been associated with the higher endothelium modulation over blood vessel reactivity to vasoconstrictor agonists, such as phenylephrine (PE), observed in pregnant rats. We hypothesized that pregnancy alters the expression and/or activity of Nrf2 and its inhibitors Keap-1 and Bach-1 and that these changes are associated with greater endothelial modulation on the contraction of aortas to PE. To test this hypothesis, the expression of Nrf2, Keap-1 and Bach-1 and the antioxidant enzymes transcribed by Nrf2, such as NADP (H) quinone oxidoreductase-1 (NQO1), SOD-1 and SOD-2 were evaluated in aortas of pregnant rats and compared to aortas of non-pregnant rats. In order to identify other possible mechanisms altered by pregnancy in Wistar rats and SHR, we evaluated the expression of NOXO-1, NOX1 regulatory subunit and p47phox, NOX2 regulatory subunit. The role of Nrf2 in the production of endothelial NO in aortas of pregnant rats was evaluated by use of Brusatol, an inhibitor of Nrf2. We also evaluated the role of Nrf2 in the reactivity of aortas of pregnant rats to phenylephrine and acetylcholine, using Brusatol. All results were compared between normotensive (Wistar) and hypertensive (SHR) non-pregnant rats and between pregnant and non-pregnant rats in the groups (multivariate analysis, Tukey post-test, p< 0.05). The results showed that the expression of Nrf2 is increased in aortas of pregnant Wistar rats, although the expression of Keap-1 and Bach-1 is not altered. The increased expression of Nrf2 was associated with the greater expression of SOD-2, but not of SOD-1 or NQO1, in aortas of pregnant rats. In aortas of non-pregnant SHR rats, we observed among all evaluated proteins, lower expression of Bach-1 and NQO1 when compared to the aortas of normotensive rats. Pregnancy reduced even more the expression of NQO1 in SHR aortas. Pregnancy reduced the expression of NOXO-1 and p47phox in SHR aortas, whereas in aorta of Wistar rats it reduced only the expression of NOXO-1. The results obtained in this study also showed that the incubation of HUVEC with Brusatol increased the intracellular concentrations of ROS, but did not alter the concentrations of NO, however, Brusatol significantly reduced the concentration of NOx stimulated by ACh in aortas of pregnant rats, Wistar or SHR. Moreover, Brusatol increased the reactivity to PE in aortas of pregnant normotensive rats and pregnant, matching the reactivity of aortas of pregnant rats to aortas of non-pregnant rats. However, Brusatol did not alter the reactivity of pregnant or non-pregnant SHR aortas. No significant effect of Brusatol was observed in the reactivity to Acetylcholine in aortas of Wistar rats or SHR, pregnant or non-pregnant rats. In conclusion, our results suggest that the activity of the Nrf2 signaling pathway is increased, favoring the greater activity of antioxidant enzymes such as SOD-2, which would contribute to greater bioavailability of NO and greater endothelium-dependent modulation of vascular contraction to PE in aortas of pregnant normotensive rats. However, in pregnant SHR aortas, this mechanism appears to be no more important than the lower the activity of NOX isoforms and their regulatory subunits that would contribute to lower O2•- generation and, consequently, greater NO bioavailability(AU)

Relationship between microangiogenesis and astrocytes after cerebral infarct in SHR-SP rats / 西安交通大学学报(医学版)

Objective: To investigate the possible role of astrocytes after brain infarction in stroke-prone, spontaneously hypertensive (SHR-SP) rats and the association with angiogenesis and the architecture. Methods: We maintained SHR-SP rats on high sodium water starting to accelerate the stroke onset. The 3D quantification of microvasculatures (diameter, branch number) by cofocal microscope after FITC-dextran was injected into the rats via the left femoral vein. Glial fibrillary acidic protein (GFAP) expression and microvessel density (MVD) using counting the number of factor -positive endothelial cells were evaluated by immunofluorescence and immunohistochemistry, respectively. Results: Cerebral infarction occurred at week 7 after high sodium water intake (13 g/L NaCl) in SHR-SP group. When compared with the non-infarcted contralateral hemisphere and SHR-SP on normal sodium intake and WKY rats, GFAP expression and MVD were significantly increased, respectively, and the diameter and the branch number of vessels were decreased, respectively, in cerebral infarcts with boundary zones of SHR-SP rats (P<0.01). Linear correlation analysis showed that GFAP expression was positively correlated with MVD and the diameter and the branch number of vessels in cerebral infarcts in SHR-SP (P<0.01). Conclusion: Astrocytes hyperplasia may be associated with increased regional angiogenesis and the changes of architecture in SHR-SP rats with high sodium water (13 g/L NaCl) that induces focal cerebral infarcts.

Value of stress hyperglycemia ratio in predicting the prognosis of patients with acute myocardial infarction / 上海交通大学学报（医学版）

Objective • To explore the value of stress hyperglycemia ratio (SHR) in predicting the prognosis of patients with acute myocardial infarction(AMI). Methods • A total of 434 patients diagnosed as having AMI were enrolled from October 2014 to October 2015. Immediate blood glucose, glycosylated hemoglobin (HbA1c) and SHR of each subjects were collected and calculated. Patients with and without diabetes were divided into several groups according to the occurrence of in-hospital and 2-year major adverse cardiovascular and cerebrovascular events (MACCEs). Indicators above were evaluated to predict the prognosis of patients with AMI. Results • Immediate blood glucose (P=0.003) and SHR (P=0.005) were significantly higher in MACCEs group than those in 2-year MACCEs-free group. Among AMI patients with diabetes mellitus, SHR in hospital MACCEs group was significantly higher than that in MACCEs-free group (P=0.001). Among AMI patients without diabetes, values of immediate blood glucose (P=0.001) and SHR (P=0.001) were higher in 2-year MACCEs group than those in MACCEs-free group. All-cause mortality was the highest (P=0.047, P=0.007) in the highest SHR group (three-digit relationship), no matter with or without diabetes mellitus. AMI patients with and without diabetes were further divided into two groups according to the median of SHR. High SHR could better predict in hospital MACCEs rate in AMI patients with diabetes as well as 2-year MACCEs rate without diabetes (P=0.023, P=0.000). Conclusion • Abnormal stress hyperglycemia is a risk factor of long-term poor prognosis in AMI patients. The value of SHR is more valuable in predicting the poor prognosis of AMI than immediate blood glucose, no matter the patients with or without diabetes.

The clinical reports on adrenal insufficiency of patients with advanced solid tumors accepting anti-PD-1 antibody, SHR-1210 therapy / 中华肿瘤杂志

Objective@#To investigate the adrenocortical function changes of patients with advanced solid tumors who received the anti- programmed cell death protein-1 (PD-1) antibody, SHR-1210 therapy.@*Methods@#The clinical data of 98 patients with advanced solid tumors who were enrolled in a prospective phase I trial of SHR-1210 therapy at our institution between April 27, 2016 and June 8, 2017 were collected. The levels of plasma adrenocorticotropic hormone (ACTH) and cortisol were evaluated in 96 patients. The clinical manifestations, laboratory tests and radiologic data were collected to define the immune-related adrenal insufficiency.@*Results@#Until December 14th, 2018, no SHR-1210 related primary adrenal insufficiency occurred, and the incidence of immune-related secondary adrenal insufficiency was 1.0% among the 96 patients, which was identified as grade 2. No patient developed grade 3-4 adrenal insufficiency. The main clinical manifestations of the patient who was diagnosed as secondary adrenal insufficiency were grade 2 fatigue, anorexia and headache.The patient developed fatigue and anorexia at the 267th day after receiving the first dose of SHR-1210, the hypocortisolism occurred on the 279th day, and the headache emerged on the 291th day. The anorexia of patient who treated by physiological replacement doses of glucocorticoid since the 457th day was attenuated.The patient whose cortisol level was still below the normal limit continued to accept the hormone replacement therapy up to 776 days after the initial administration of SHR-1210.@*Conclusions@#The incidence of SHR-1210 related adrenal insufficiency of patients with advanced solid tumors is low, and the symptoms can be effectively ameliorated by hormone replacement therapy. The potential adverse outcome of adrenal insufficiency following immunotherapy should be noticed by clinicians to avoid the occurrence of adrenal crisis.

The anti-proliferative and anti-inflammatory mechanisms of JAK1 inhibitor SHR0302 versus Ruxolitinib in SET2 cell line and primary cells / 中华血液学杂志

Objective@#To explore the effects and molecular mechanism of the selective JAK1inhibitor SHR0302 and Ruxolitinib on myeloproliterative neoplasms (MPN) cell line SET2 and primary cells in vitro.@*Methods@#Cell proliferation was detected by CCK8 kit. Colony forming experiment was conducted to evaluate erythroid burst colony formation unit (BFU-E) of primary cells from MPN patients. Multi-factor kits were used to detect six inflammatory cytokines. Phosphorylated proteins of Jak-Stat signaling pathway were tested by Western blot.@*Results@#At different time points after treated with SHR0302 and Ruxolitinib, the inhibition of cell proliferation was dose dependent by both drugs (P<0.01) . The inhibitory rates of 2.5 μmol/L SHR0302 and 0.1 μmol/L Ruxolitinib on SET2 cells for 72 h were comparable, i.e. (59.94±0.60) % and (64.00±0.66) %, respectively, suggesting that the inhibitory effect of SHR0302 was weaker than that of Ruxolitinib. Similarly, both SHR0302 and Ruxolitinib inhibited BFU-E in primary marrow cells from MPN patients in a dose-dependent manner. SHR0302 1.0 μmol/L produced similar degree of inhibition compared to Ruxolitinib 0.2 μmol/L. Except IL-12, the expression of other 5 cytokines (IL-6, TNF-α, IL-1β, IL-2, IL-8) was significantly inhibited by 1.6 μmol/L SHR0302 in SET2 cells at 24 h (P<0.01) , while Ruxolitinib 1.0 μmol/L had the same effect. Several phosphorylated molecules of Jak-Stat signaling pathway were significantly inhibited by SHR0302 in SET2 cells only for 3 h. P-stat1 (Tyr701) , p-stat3 (Tyr705) were down-regulated when treated with SHR0302 1.0 μmol/L (P<0.05) , p-jak1 (tyr1022/1023) and p-stat5 (Tyr694) were inhibited at 5.0 μmol/L (P<0.05) . Ruxolitinib significantly inhibited the downstream STAT protein at 0.1 μmol/L. Again, the inhibitory effect of SHR0302 on protein expression was weaker than that of Ruxolitinib.@*Conclusion@#SHR0302 can effectively inhibit the proliferation of MPN cell line and patients' primary cells, as well as the expression of inflammatory factors. The molecular mechanism is possibly related to the down-regulation of phosphorylated proteins of Jak-Stat signaling pathway. Overall, the anti-proliferative and anti-inflammatory effects of SHR0302 are weaker than those of Ruxolitinib.

Effect of baicalin on growth state of attention deficit hyperactivity disorder animal model and its regulation on Ca MKⅡ and ERK1/2 / 中国中药杂志

The aim of this study was to observe the effect of baicalin on the growth state of attention deficit hyperactivity disorder animal model and its regulation on Ca MKⅡand ERK1/2.In the present study,a total of 40 SHR rats were randomly divided into model group,methylphenidate hydrochloride group,and low,medium,and high dose baicalin groups,with 8 rats in each group.Eight WKYrats were selected as a normal control group.The methylphenidate hydrochloride group(0.07 g·L~(-1))and the low(3.33 g·L~(-1)),medium(6.67 g·L~(-1)),and high dose(10 g·L~(-1))baicalin groups received corresponding drugs by gavage administration according to the body weight(0.015 m L·g~(-1)),while the normal group and the model group received the same volume of normal saline by gavage.Thegavage administration lasted for 4 weeks,twice a day.The body weight of the rats and the amount of remaining feed were weighed daily,and the growth state of the rats was statistically evaluated weekly.Percoll density gradient centrifugation was used to prepare brain synaptosomes and an electron microscope was used to observe their structures.The Ca MKⅡand ERK1/2 protein and mRNA expression levels were detected with Western blot and Real-time PCR methods,respectively.RESULTS: showed that baicalin did not affect the normal eating and weight gain of rats,and the weight gain of rats was even more significant than that in the normal group(P<0.05).In the study of its effects on Ca MKⅡand ERK1/2 protein expression in rat synaptosomes,the expression of both proteins in each drug-administered group was higher than that in the model group(P<0.05);besides,the expression levels of Ca MKⅡand ERK1/2 protein were significantly increased in both baicalin high dose group and the methylphenidate hydrochloride group(P<0.05).The relative expression of Ca MKⅡand ERK1/2 mRNA in synaptosome was detected by PCR.The results showed that medium and high doses of baicalin and methylphenidate hydrochloride significantly increased the relative expression of Ca MKⅡand ERK1/2 mRNA in synaptosomes of SHR rats(P<0.05).In conclusion,baicalin does not affect the normal growth and development of SHR rats,so it is safe for administration.Both baicalin and methylphenidate hydrochloride could up-regulate the relative expression of Ca MKⅡand ERK1/2 in mRNA and protein,and the pharmacodynamic stability of baicalin is in a dose-dependent manner to certain extent.

Moderate-intensity exercise allows enhanced protection against oxidative stress-induced cardiac dysfunction in spontaneously hypertensive rats

The progression of myocardial injury secondary to hypertension is a complex process related to a series of physiological and molecular factors including oxidative stress. This study aimed to investigate whether moderate-intensity exercise (MIE) could improve cardiac function and oxidative stress in spontaneously hypertensive rats (SHRs). Eight-week-old male SHRs and age-matched male Wistar-Kyoto rats were randomly assigned to exercise training (treadmill running at a speed of 20 m/min for 1 h continuously) or kept sedentary for 16 weeks. Cardiac function was monitored by polygraph; cardiac mitochondrial structure was observed by scanning electron microscope; tissue free radical production was measured using dihydroethidium staining. Expression levels of SIRT3 and SOD2 protein were measured by western blot, and cardiac antioxidants were assessed by assay kits. MIE improved the cardiac function of SHRs by decreasing left ventricular systolic pressure (LVSP), and first derivation of LVP (+LVdP/dtmax and −LVdP/dtmax). In addition, exercise-induced beneficial effects in SHRs were mediated by decreasing damage to myocardial mitochondrial morphology, decreasing production of reactive oxygen species, increasing glutathione level, decreasing oxidized glutathione level, increasing expression of SIRT3/SOD2, and increasing activity of superoxide dismutase. Exercise training in SHRs improved cardiac function by inhibiting hypertension-induced myocardial mitochondrial damage and attenuating oxidative stresses, offering new insights into prevention and treatment of hypertension.

Diferencias intra-especie en respuesta al estrés: Hacia enfoques traslacionales / Intra-specie differences in response to stress: Looking for translational view

Resumen: El estrés produce la sobreactivación del eje HPA y sistema neuroendocrino. Se ha mostrado que existe daño en estructuras relacionadas con el procesamiento emocional (amígdala) aprendizaje (hipocampo), toma de decisiones y prospección (corteza prefrontal). Sin embargo, se generalizan los efectos del estrés sin ponderar el tipo de estrés (crónico o agudo), duración, especie, etc. Esto permite que hallazgos se contrapongan a nivel cortical, neuroquímico, hormonal y conductual. El objetivo fue evaluar los efectos del estrés crónico impredecible (ECI) en diferentes cepas de ratas y sus efectos inmediatos o a largo plazo. Se utilizaron ratas macho Wistar, Wistar Kyoto y SHR en condiciones estándar de laboratorio. Se aplicó una batería de ECI y una batería de evaluación conductual para evaluar efectos previos, agudos y crónicos. La cepa Wistar Kyoto muestra deficiencias previas a la exposición. La cepa SHR muestra mayor movilidad y sesgos atencionales, lo que produce un efecto que perdura a largo plazo. La cepa Wistar muestra una gran capacidad de adaptación ya que aunque se observaron deficiencias inmediatamente después de la exposición al estrés, éstas se recuperan e largo plazo. Se infiere que las precondiciones de los sujetos podrían funcionar como biomarcadores y poder prevenir padecimientos relacionados al estrés.

Abstract: Stress produces the over activation of the Hypothalamus Pituitary Adrenal axis (HPA) and the neuroendocrine system. It has been shown that it could damage structures related with the emotional processing (amygdala), learning and memory (Hippocampus), decision making and prospection (prefrontal cortex). However, the stress affects are generalized without weighting all the elements related with this conditions, for example the kind of stress stimuli (acute or chronic), duration, species, etc. This allowed that some findings it will go against each other in relation to cerebral cortex function, neurochemical, hormonal and behavioral. The main porpoise of this research was to evaluate the effects of the unpredictable chronic stress on several rat strains (Wistar, Wistar Kyoto and SHR) and its immediate effects or in long term so. Wistar, Wistar Kyoto and SHR rats were used. All animals were housed in standard laboratory conditions and we follow the international guide for use and care of laboratory animals. The subjects were exposed to the Chronic Unpredictable Stress Battery (CUSB) and to evaluate the stress effects all the subjects were evaluated with a Battery of Behavioral Evaluation to find the previous, immediate or the long-term effects of CUSB exposition. The Wistar Kyoto strain showed deficits before the stress exposure. Whereas the SHR rats showed more mobility and poor attention which produces a long-term effect. The Wistar strain show a high adaptation to the adverse conditions because until the animals showed strong effects immediately after the stress exposure they showed a good recovery in the long term. In conclusion we can asseverate that the preconditions in every strain plays a major role in the stress response and that preconditions it could be used as biomarkers and in that way infer if the subjects are more susceptible to suffer high stress or some other related disease.

The influence of benazepril and amlodipine on the expression of secretin and somatostatin in spontaneously hypertensive rats / 中国应用生理学杂志

OBJECTIVES@#Investigate the influence of benazepril and amlodipine on the expression of secretin (PZ) and somatostatin (SS) in spontaneously hypertensive rats (SHR).@*METHODS@#Forty-five SHRs (14 weeks old, male) were randomly assigned into 3 groups (=15):SHR group, Benazepril group (which was given benazepril 0.90 mg·kg·d) and Amlodipine group (SHRs were given amlodipine 0.45 mg· kg·d), taking WistarKyoto(WKY) as normal control (=15), meanwhile, rats in SHR group and WKY group were given the same volume of distilled water. After 8 weeks of intervention, the expression of protein and mRNA of PZ in duodenum and SS in sinuses ventriculi was detected by enzyme-linked immunoassay and RT-PCR.@*RESULTS@#After 8 weeks of intervention, compared with the WKY group, the expression of protein and mRNA of PZ in duodenum and SS in sinuses ventriculi was increased significantly in SHR group (<0. 05). Compared with SHR group, the expression of PZ in duodenum and SS in sinuses ventriculi was decreased significantly in Benazepril group and Amlodipine group (<0.05). Compared with Benazepril group, in Amlodipine group the expression of PZ mRNA in duodenum and SS mRNA in sinuses ventriculi was decreased more significantly (<0.05).@*CONCLUSIONS@#The regulation disorder of PZ in duodenum and SS in sinuses ventriculi exists in SHR. The antihypertensive effect of benazepril and amlodipine may be realized by regulating the expression of PZ and SS, while the regulation of amlodipine is more obvious than benazepril.

Effects of Taxilli Herba from different hosts on lowering blood pressure of spontaneously hypertensive rats / 中成药

AIM To observe the effects of Taxilli Herba from six different hosts (Morus alba L.,Salix babylonica L.,Camellia oleifera Abel.,Castanea mollissima B1.,Liquidambar formosana Hance and Nerium indicum Mill.) on lowering blood pressure of spontaneously hypertensive rats (SHR).METHODS SHR were randomly divided into 14 groups,captopril positive group (20 mg/kg),model group,and Taxilli Herba groups of 6 different hosts,and each Taxilli Herba group was further divided into high-dose group (5.9 g/kg) and low-dose group (1.48 g/kg);WKY,in addition,was the blank control.And the 20-day consective correspondence medication was applied to the groups,each with eight rats.The caudal arterial systolic blood pressure (SBP) was measured by tail-cuff method before the administration,and on the 10th day and 20th day of the administration.Anaesthesia was performed at the blood collection 12 h after the last administration;and thus final serum contents of nitric oxide (NO) and changes of superoxide dismutase (SOD) activity,plasmatic contents of angiotensin Ⅱ (Ang Ⅱ) and endothelin-1 (ET-1) were determined.RESULTS From the data before and after administration,an SBP drop among all SHR groups was observed on the 10th day of administration,among which the blood pressure lowering effect by high-dose Taxilli Herba from Morus alba L.was very obvious (P ＜ 0.01);remarkable SBP decrease on the 20th day of administration induced by Taxilli Herba from Salix babylonica L,Liquidambarformosana Hance and Camellia oleifera Abel,and high-dose Taxilli Herba from Morus alba L,low-dose Taxilli Herba from Castanea mollissima B1 were detected (P ＜ 0.01).No significant SBP variation was available between the model group and Taxilli Herba groups after10-day administration;all the Taxilli Herba groups exhibited obvious effect in lowering SBP except Taxilli Herba from Nerium indicum Mill,low-dose Taxilli Herba from Morus alba L.and high-dose Taxilli Herba from Castanea mollissimaBl.after 20-day administration,compared to the model group (P ＜ 0.05).Highdose Taxilli Herba from Morus alba L.and Salix babylonica L significantly decreased plasmatic Ang Ⅱ and ET-1 contents of SHR groups in comparison with the model group (P ＜0.01).High-dose Taxilli Herba from Morus alba L.,Salix babylonica L.and Liquidambarformosana Hance significantly increased serum NO release and superoxide dismutase (SOD) activity (P ＜ 0.01).CONCLUSION Taxilli Herba from the five different hosts,except Nerium indicum Mill,can lower blood pressure,and there exists an effect difference due to the host variation.

Regulatory effect of Shudihuang on expressions of BDNF/TrkB and NRG-3 in prefrontal cortex and striatum of ADHD model rats / 中国中药杂志

To observe the effect of Shudihuang on behaviors and expression of BDNF/TrkB and NRG-3 in prefrontal cortex and striatum of attention deficit hyperactivity disorder (ADHD) model rats. Thirty 4-week-old spontaneous hypertension rats (SHR) were randomly divided into model group, methylphenidate hydrochloride (MPH, 2 mg·kg⁻¹·d⁻¹) and Shudihuang group (2.4 g·kg⁻¹·d⁻¹). Another 10 Wistar-Kyoto (WKY) rats were selected as normal control group. The 0.5% CMC-Na solution was administered to model group and WKY rats (2 mL·kg⁻¹·d⁻¹). All of the rats were treated for 4 weeks. The open field test was performed at the 14th and 28th days after gavage, in order to evaluate the spontaneous and impulsive behaviors. Subsequently, gene and protein expressions of BDNF/TrkB and NRG-3 were tested by RT-qPCR and Western blot. Compared with model group, MPH and Shudihuang groups showed significant reduction in total distance, mean velocity and central distance in the open field test (<0.05), and Shudihuang group displayed a shorter central distance than MPH group (<0.05). RT-qPCR and Western blot analysis indicated that expressions of BDNF/TrkB and NRG-3 were lower in prefrontal cortex and striatum of SHR compared with WKY rats. Four weeks later after administration, both Shudihuang and MPH significantly elevated mRNA and protein expressions of BDNF/TrkB and NRG-3 (<0.05).In conclusion, neurodevelopmental disorder mediated by BDNF/TrkB and NRG-3 was closely related with SHR rats' behaviors. Shudihuang may ameliorate the spontaneous and impulsive behaviors by up-regulating the expressions of BDNF/TrkB and NRG-3 and improving growth and maturation of neurons in SHR.

Impact on NO, iNOS, IL-1 and IL-6 levels of isosorbide mononitrate in lung tissue of spontaneously hypertensive rats (SHR) / 实用医学杂志

Objective To investigate the effect of isosorbide mononitrate on the levels of NO,iNOS,IL-1 and IL-6 in lung tissue of spontaneously hypertensive rats (SHR).Methods Fourteen-week-old Wistar and SHR male rats were randomly divided into the W0,W1,S0 and S1 group,with 10 rats ineach group.Rats in the W0 and S0 group were fed with the normal saline and the ordinary food,rats in the W1 and S1 group were fed with isosorbide mononitrate and the ordinary food.Twelve weeks later,levels of NO,iNOS,IL-1 and IL6 in rat lung tissue were detected.Results Compared with the W0 group,levels of NO,iNOS,IL-1 and IL6 were significantly increased in the W1 groups (P ＜ 0.05,respectively).Compared with the SO group,levels of NO,iNOS,IL-1 and IL6 were significantly increased in the S1 group (P ＜ 0.05,respectively).In the W1 and S1 group,levels of iNOS and NO were positively correlated with IL-1 and IL-6.Conclusion 1.Isosorbide mononitrate may lead to increases of NO,iNOS,IL-1 and IL6 in lung tissue of Wistar rats,which indicates the presence of chronic inflammation.2.Longterm feeding of isosorbide mononitrate may lead to increases of inflammatory factors in SHR rats,contributing to the inflammatory state in rats.

Nf1 and BDNF Dysregulation in the Prefrontal Cortex Involved in the Pathogenesis of Attention Deficit Hyperactivity / 医学研究杂志

Objective To investigate the interaction between the Nf1 (Neurofibromin 1)and the BDNF (brain derived neurotrophic factor) in the mechanism of attention deficit hyperactivity disorder (ADHD).Methods Spontaneously hypertensive rats were chosen as the experimental group and WKY servesd as control.RT-PCR,Western-blot were used to detect the mRNA and protein expression levels of Nf1 and BDNF.To explore their relationship,BDNF expression levels were detected after Nf1 expression plasmid transferred and transfection in PC12H and CBRH-7919 cells.Results were quantified by Image-pro plus software.Results In the PFC of ADHD model rats SHR,Nf1 expression was abnormally degressive(P =0.000),in common with the expression of BDNF(P =0.000).Ectopic expression of Nf1 further encouraged the expression of BDNF (PC12H:P =0.000,7919:P ＜0.05).Conclusion Nf1 expression was significantly lower in PFC of SHR than control,and positively correlated with the levels of BDNF.These findings show that in the prefrontal cortex,Nf1-BDNF dysregulation may be involved in the pathomechanism of ADHD.

Imunomarcação para TRAP em tecido de reparação óssea de ratos espontaneamente hipertensos (SHR) tratados com Atenolol / Immunostaining for TRAP in bone healing tissue of Spontaneously Hypertensive Rats (SHR) treated with Atenolol

Introdução: A hipertensão arterial tem sido um dos maiores problemas de saúde no mundo, com grandes alterações para as doenças cardiovasculares e renais. O tecido ósseo tem função importante no suporte, proteção e locomoção e está sob o controle de fatores sistêmicos como hormônios e fatores locais, entre eles os fatores de crescimento e citocinas. A Fosfatase Ácida Tartarato Resistente (TRAP) é uma enzima que faz parte da família das fosfatases ácidas e apresenta localização intracelular; mais especificamente dentro do compartimento lisossomal de osteoclasto, macrófagos e células dendríticas, tem sido utilizada como um marcador histoquímico da atividade osteoclástica. Objetivos: Avaliar a expressão da proteína TRAP em alvéolos dentários de ratos hipertensos (SHR) e normotensos tratados ou não com atenolol. Métodos: Neste estudo foram utilizados 4 grupos de ratos sendo: 1) W (wistar sem tratamento), 2) WT (wistar tratado com atenolol), 3) S (SHR sem tratamento) e 4) ST (SHR tratado com atenolol), submetidos a exodontia do incisivo superior direito, com eutanásia no 7º, 14º, 21 e 28º dia pós-operatório. A análise dos mecanismos biológicos envolvidos no processo de reparo alveolar foi obtida pela análise da expressão de proteínas TRAP por meio da técnica de imunoistoquímica. Os resultados foram analisados pela média e erro padrão da média e aplicado o teste paramétrico ANOVA, com pos-test de Tukey para avaliar os períodos dentro de cada grupo e entre os grupos, sendo consideradas as diferenças significativas quando p<0,05. Resultados: Os resultados mostraram que a marcação TRAP aumenta em alvéolo dentais de ratos Wistar durante todos os períodos pós ­ operatórios. A marcação TRAP aumenta apenas ao 14o nos dias de reparação alveolar em alvéolo dental de SHR não tratados. O atenolol não altera o processo de reparo alveolar em ratos Wistar, porém o atenolol promoveu a redução da marcação de TRAP em SHR ao 14º dia. Conclusão: A hipertensão aumenta a expressão da proteína TRAP no 14o dia pós-cirúrgico de reparação alveolar e o atenolol promove redução da marcação aumentada de TRAP ao 14º dia pós-cirúrgico em alvéolos de SHR(AU)

Introduction: Arterial hypertension has been one of the world's biggest health problems, with considerable alterations for cardiovascular and renal diseases. The bone tissue has an important role in support, protection and locomotion and is controlled by systemic factors like hormones and local factors, such as growth factors and cytokines. The Tartrate-resistant Acid Phosphatase (TRAP) is an enzyme that belongs to the Acid Phosphatases family and has an intracellular location, more specifically inside the lysosomal compartment of osteoclasts, macrophages and dendritic cells. It has been used as a histochemical marker of the osteoclast activity. Objectives: Evaluate TRAP protein's expression in the dental alveoli of normotensive and hypertensive rats (SHR) treated or not treated with Atenolol. Methods: In this study, four groups of rats were used: 1) W (with no treatment), 2) WT (wistar treated with Atenolol), 3) S (SHR without treatment) and 4) ST (SHR treated with Atenolol), all of which underwent exodontia of the upper right incisor with euthanasia on the 7th, 14th, 21st and 28th day after the operation. The analysis of the biological mechanisms involved in the process of alveolar repair was obtained by the expression of TRAP proteins in the alveolar process through an immunohistochemistry technique. The results were analyzed through the average and its standard error. The parametric test ANOVA was applied with Tukey's posttest were applied to evaluate the periods within each group and between the groups, considering the significant differences when p< 0,05. Results: The results demonstrated that TRAP staining increases in the dental alveoli of Wistar rats during all the post-surgical periods. TRAP staining increases only on the 14th day of alveolar recovery in the dental alveoli of non-treated SHR. Atenolol does not change the process of alveolar repair in Wistar rats, but Atenolol promoted the reduction of TRAP staining among SHR on the 14th day. Conclusion: Hypertension increases the expression of TRAP proteins on the 14th alveolar recovery postsurgical day and Atenolol promotes the reduction of the increased TRAP staining on the 14th postsurgical day in SHR's alveoli(AU)

Microarray Analysis in Spontaneously Hypertensive Rat Heart after Losartan Treatment / 이화의대지

OBJECTIVES: Spontaneously hypertensive rats (SHR) are frequently used as rat models of essential hypertension. The mechanism for the development of hypertension is complicated and it is unknown. The renin-angiotensin system (RAS) plays a key role in the control of blood pressure. Microarrays are a powerful tool for studying genetics. The purpose of this study was to investigate changes of gene expression in the heart tissues of SHR after losartan treatment to provide basic data that is useful in the early diagnosis of hypertension and gene treatment. METHODS: Rats were divided into three groups: the control (C) group; the hypertension (H) group (SHR), and the losartan (L) group; treated with losartan (10 mg/kg/day) in SHR. Rats were sacrificed at week 5 and microarray analysis was performed. RESULTS: 102 gene expressions including the genes associated with cell proliferation such as Raf1, Uchl1, Btla, Spock1 were increased. The other 139 gene expressions, including the genes related to the regulation of metabolism such as TFIID, Auf1, Bmp, Hub, Taf51 showed decreases in gene expression. A total of 31 genes were differentially expressed in the L group compared to the H group. Of these, 16 genes including the genes associated with macromolecule metabolism such as MGC105766, Ppp1r1a, Rpl3l showed increased expression. The other 15 genes including the genes associated with primary metabolism such as Mcpt4, Ngn3, Tdo, Ak2 Hyal2 showed decreased expressions. CONCLUSION: According to microarray analysis, there was significant gene expression change in SHR compared with normal rats as well as significant gene expression changes after losartan treatment in SHR.