Effect of pharmaceutical intervention on AT1R, AT2R, ERK and JNK activity in chronic hibernating myocardium in rabbits

Objective： To investigate in chronic hibernating myocardium in rabbits and the influence and significance of captopril, betaloc, valsartan in angiotensin Ⅱ subtype 1 receptor（AT1R）, angiotensin Ⅱ subtype 2 receptor（AT2R）, extracellular signal regulated kinase 1/2 （ERK1/2）, c-Jun N-terminal kinase（JNK）. Methods： The model of chronic hibernating myocardium（CHM） was established. The changes of AT1R, AT2R, ERK1/2, JNK in different groups were assessed by western blotting and immunohistochemistry. Results： The amount of AT1R decreased while AT2R increased in the CON group compared with in sham group, and both AT1R and AT2R decreased in drug groups compared with the CON group. The content of ERK had no change in each group, while that of ＂expression＂ p-ERK increased in CON group compared with in sham group, and was lower in drug intervention groups than in CON and sham groups. The contents of JNK and p-JNK decreased in CON and drug intervention groups compared with in sham group. The protein levels of JNK, p-JNK in drug intervention groups were lower than in the CON group. Three drugs can inhibit interstitial fibrosis and reduce apoptotic cells. The expression levels in the groups（with different doses） had statistical difference as well as between groups of captopril and other drugs; however the results between betaloc and valsartan had no significant difference. Conclusion： AT1R, AT2R may be the upper stream receptor of ERK and JNK and may participate in generation and evolution of CHM. Captopril, valsartan and betaloc may preserve CHM by inhibiting ATIR, AT2R and JNK activity.     

Effects of Electroacupuncture on the Contents of NO, ET and T-AOC in the Brain Tissues of the Cerebral Hemorrhage Model Rats

In the cerebral hemorrhage model rats established by injection of collagenase and heparin into caudatenucleus,the effects of electroacupuncture (EA) on the contents of nitric oxide (NO) and endothelin (ET),and total anti-oxidation capability (T-AOC) in the brain tissues were investigated.It is found that thecontent of NO in the Shuigou EA group lowered,ET decreased and the T-AOC raised significantly in boththe Fengfu EA group and the Shuigou EA group (all P<0.05) as compared with the model group,indicating that acupuncture can reduce the contents of ET and NO,and increase the T-AOC in the braintissues of the rats with cerebral hemorrhage.     

Effect of Electroacupuncture on Expression of p53 Protein in Cerebral Cortex of Rats with Global Cerebral Ischemia/Reperfusion Injury

Objective: To observe the effect of electroacupuncture (EA) on expression of p53 protein in cerebral cortex of senile rats with global cerebral ischemia/reperfusion (IR) injury and to explore its mechanism. Methods: The cerebral IR injury rat model was established referring to Pulsinelli 4-vessel occlusion method. Thirty-six SD rats were randomly and evenly divided into the control group, the IR group and the IR plus EA (IR-EA) group. The animals in the control group were subjected to electrocauterization of vertebral arteries in bilateral flank orifice alone with the general carotid arteries unoccluded. To rats in the IR-EA group, immediately and 24h, 48h, 72h after cerebral IR, EA treatment on bilateral acupoint "Zusanli" (ST36) was applied once a day, lasting for 60 minutes. After the final treatment, all the rats were sacrificed and their brains were taken to examine p53 protein expression by the immunohistochemical method. Results: Cells with positive p53 immunoreactivity in the cerebral cortex of     

Role of angiotensin Ⅱ and angiotensin Ⅱ receptors in vascular smooth muscle cell migration in vitro

ObjectiveTo determine the biotic effects of angiotensin Ⅱ (Ang Ⅱ) on the migration of rat smooth muscle cells (VSMCs) and investigate the mechanisms involved in the development of vascular injury. Methods VSMCs isolated from aortic media of Wistar rats and cultured by the modified explant method were adopted. In the presence and absence of Ang Ⅱ, the expression of Ang Ⅱ receptor (ATR) and reorganization of the actin cytoskeleton and focal adhesion of VSMCs were studied by an immunocytochemistry technique and fluorocytochemistry technique. Migration assays were performed with a modified Boyden’s chamber. The effects of AT(1)R antagonist (CV- 11974), AT2R antagonist (PD123319) on the aforementioned target were studied. Results VSMCs migration was stimulated by adding Ang Ⅱ. The dynamic reorganization of actin cytoskeleton and focal adhesions may be an important mechanism by which Ang Ⅱ facilitates VSMCs motility. The expression of AT(1)R in VSMCs could be upregulated initially after treatment with Ang Ⅱ, then decreased gradually. The expression of AT(1)R was downregulated by AT(1)R antagonists. The effect of Ang Ⅱ on VSMCs migration was mediated by AT(1)R, while AT2R had no significant effect. Conclusions The dynamic reorganization of focal adhesions and the actin cytoskeleton is required for Ang Ⅱ- induced VSMCs migration. This effect is mediated by AT(1)R.     

Effect of Electro-acupuncture on Rat Joint Pathomorpnology of Chronic Adjuvant Arthritis Model

Objective:To study the effect of electro-acupuncture (EA) on pathomorphological changes of joints in rat model of chronic adjuvant arthritis. Methods: The rat chronic adjuvant arthritis model was established by subcutaneous injection of 0.1 ml of complete Freund's adjuvant to the left hind sole. Forty Wistar rats were randomly divided into the model group, the low frequency (2 Hz) EA group, the high frequency EA (100 Hz) group and the body acupuncture group. After being modeled except the model group, the other three groups were treated with EA or body acupuncture in Yanglingquan points (bilateral) for 3 weeks, the left ankle joints and metatarsal joints of rats were taken for pathological examination by fixing with 10% formalin and embedding in paraffin, sectioning and staining with HE. Results: Obvious inflammatory cell infiltration, loosened synovial tissue, damage of articular cartilage and proliferation of synovial cells and granulation tissue were observed in the sections of joints in model rats.     

Expression of cyclooxygenase-2 and its pathogenic effects in nonalcoholic fatty liver disease

Objective:To investigate the expression of cyclooxygenase-2 and its pathological effect in the experimental nonalcoholic fatty liver of rats,and to explore its possible mechanism.Methods:The rat NAFLD model was established by giving a fat-enriched diet.The blood samples were obtained form abdominal aorta and the levels of serum ALT,AST and IL-1,changes in the hepatic tissue 6-k-PGF1α TXB2 were measured.The expression level of COX-2 in rats livers were assayed by immunohistochemistry,RT-PCR and Western-blot.Results:Light microscope analysis revealed that hepatocytes were injured in the model group and slightly in the treatment group.The levels of serum TXB2 and IL-1 in the fatty liver rats were increased.Compared with the model group,the IL-1 and TXB2 increased significantly(P < 0.05),on the contrary,compared with the normal group,the hepatic tissue 6-Keto-prostagland decreased significantly in the model group(P < 0.05),the treatment group also increased but P > 0.05.There was no positive expression of COX-2 in hepatic tissue of normal rats.In the model group,there was positive expression of COX-2 antigen and the number of COX positive cells progressively increased at 4,8,12 wks.The intensity of expression of COX-2 had significantly increased(P < 0.05)and the intensity of COX-2 expression in the treated group decreased remarkably compared with the model group(P < 0.05).The expression of COX-2 mRNA and the level of COX-2 protein were significantly stronger in the liver of model rats compared with normal rats,and significantly weaker in treated rats,than in 8W and 12W model rats(P < 0.05).Conclusion:The increase of COX-2 expression in NAFLD is closely associated with the severity of liver inflammation and damage.COX-2 may play an important role in the progression of rat NAFLD,and the expression of COX-2 mRNA is downregulated by cyclooxygenase-2 inhibitor,which can depress the oxidative stress and control inflammatory response efficiently.     

Effect of Acupuncture on CXCL8 Receptors in Rats Suffering from Embryo Implantation Failure简

To observe the effect of acupuncture on CXCL8 receptors（CXCR1 and CXCR2） in rat endometrium experiencing embryo implantation failure, 72 pregnant rats were randomly divided into four groups： normal group（N）, embryo implantation failure group（M）, acupuncture treatment group（A）, and progestin treatment group（W）. Then the rats in each group were equally randomized into a day-6（D6） group, a day-8（D8） group, and a day-10（D10） group. The rats in group M, group A, and group W were treated with mifepristone-sesame oil solution on day 1, while the rats in group N were injected with the same amount of sesame oil. Meanwhile, ＂Housanli＂ and ＂Sanyinjiao＂ were selected for acupuncture. From day 1 to the time of death, the rats in group A were fastened up and then acupuncture was administered while the rats in group N and group M were only fixed, and the rats in group W were given progestin. The number of implanted embryos was calculated. The expression of CXCR1 and CXCR2 in rat endometrium was detected by immunohistochemistry, Western blotting and real-time PCR. Compared to group N, the average number of implanted embryos, the protein and mRNA expression of CXCR1（D6, D8 and D10）, and the protein and mRNA expression of CXCR2（D8 and D10） in rat endometrium were significantly decreased in group M. Compared to group M, there was significant elevation in the average number of implanted embryos, the protein expression（D6, D8 and D10） and mRNA expression（D8） of CXCR1 in rat endometrium of group A, and the protein expression（D8 and D10） and mRNA expression（D8） of CXCR2 in rat endometrium of group W. These findings indicated that acupuncture can increase the number of implanted embryos in rats of embryo implantation failure, which may be relevant with up-regulation the expression of CXCR1 and CXCR2 at maternal-fetal interface of rats with embryo implantation failure.     

Protective Effects of Sapindus Saponins in Spontaneously Hypertensive Rats

Objectives:To investigate the protective effects of Sapindus saponins in spontaneously hypertensive rats,and the possible cellular and molecular mechanisms.Methods:Thirty-two 16-week-old spontaneously hypertensive rats were randomly divided into four groups(8 in each group):model group(placebo),positive control group(27 mg/kg of Captopril Tablets),Sapindus saponins groups(27 mg/kg and 108 mg/kg,respectively).Another8 healthy Wistar-Kyoto strain(WKY) rats were used as the normal group.The animals were treated for 8 weeks.Blood pressure of rats was determined by non-invasive blood pressure meter(BP-6).Furthermore,the contents of angiotensin II(Ang Ⅱ) in plasma and myocardial tissue were determined by enzyme-linked immunosorbent assay(ELISA),the gene expression of receptor angiotensin type 1(AT_1R) in aorta was determined by quantitative realtime polymerase chain reaction(qRT-PCR).The protein expression of transforming growth factor- β_1(TGF- β_1)and AT,R in heart was determined by immunohistochemical staining.The protein expression of p-phosphorylation of p38 mitogen-activated protein kinase(p-p38MAPK) was determined by Western blotting.The contents of interleukin(IL)-1,IL-6 and tumor necrosis factor(TNF) in serum were determined by radioimmunoassay.And the histopathological and morphological changes of aorta and heart tissue samples were assessed semi-quantitatively by hematoxylin-eosin(HE) or Masson staining.Results:Thirty minutes after single or continuous treatment,systolic blood pressure(SBP) was reduced significantly in Sapindus saponins groups.And the contents of Ang Ⅱ,IL-1,IL-6 and TNF- α in serum,the expression of AT_1R mRNA,p-p38 MAPK and TGF- β_1 were significantly suppressed dose-dependently(P0.05 or P0.01).With the Sapindus saponins treatment,compared with those of the model group,the cardiac and aortic pathological changes were ameliorated significantly.Conclusions:Our findings suggest that Sapindus saponins might have protective effects in spontaneously hypertensive rats,the cellular and molecular mechanisms of which might be relevant to the regulation of inflammatory responses mediated by p-p38 MAPK signal pathway based on activated Ang II and AT_1R.     

Electrical Signal Propagated across Acupoints along Foot Taiyang Bladder Meridian in Rats

Objective: To investigate the electrical signals propagated along Foot Taiyang Bladder Meridian(BL) in a rat model. Methods: The experiments were performed on Dark-Agouti(DA), DA.1U and Sprague Dawley(SD) rats. The antidromic electrical stimulation was applied on the nerve innervating "Pishu"(BL 20) to mimic the acupoint electro-acupuncture(EA). The activities recording from adjacent nerve innervating acupoint "Danshu"(BL 19) or "Weishu"(BL 21) were recorded as indics for acupoint, including the mechanical threshold and discharge rate. Results: After mimic EA on BL 20, C and Aδ units from adjacent BL 19 or BL 21 were sensitized including the decrease in mechanical threshold and increase in discharge rates in DA, DA.1U and SD rats, especially in DA rats. The average discharge rate increased from 2.40±0.26 to 6.06±0.55 and from 1.92±0.42 to 6.17±1.10 impulse/min(P0.01), and the mechanical threshold decreased from 0.52±0.12 to 0.24±0.05 and from 0.27±0.02 to 0.16±0.01 mmol/L(P0.01) in C(n=15) and Aδ(n=18) units in DA rats. The net change in discharge rates from C units were 152.5%, 144.7% and 42.4% in DA, DA.1U and SD rats, respectively, among which DA rat's was the highest(P0.05). In Aδ units, the net change in DA rats were also the highest(221.5%, 139.2% and 49.2% in DA, DA.1U and SD rats). Conclusions: These results showed that mimic acupoint EA activated adjacent acupoints along BL in three rat strains, which might be related to propagated sensation along meridians(PSM). In addition, DA rats were more sensitive and might be a good model animal for PSM research.     

Effects of Electro-acupuncture at Tongli (HT 5) and Xuanzhong (GB 39) Acupoints from Functional Magnetic Resonance Imaging Evidence

Objective

To explore the specifificity of Tongli (HT 5) and Xuanzhong (GB 39) paired acupionts in aspects of Deqi sensation and brain activation patterns during electroacupuncture.

Methods

In this study, 15 healthy subjects were enrolled. All participants suffered two kinds of functional magnetic resonance imaging (fMRI) examinations randomly: Examination A received electro-acupuncture (EA) at the bilateral Tongli (HT5) and Xuanzhong (GB 39) acupoints (ACU), and examination B received EA at bilateral non-acupoints (NAP). The subjects reported the feeling of Deqi at each examination later respectively. A multi-voxel pattern analysis method and Statistical Program for Social Sciences were used to analyze the data.

Results

The ACU group (Exam A) reported fullness, heaviness, numbness, soreness and throbbing of signifificantly greater intensity than the NAP group (Exam B). In addition, there was no statistical signifificance between two groups in aching, tingling, deep pressure, sharp pain, dull pain, warmness and cold. Meanwhile, fMRI data revealed differences between two groups in discriminating accuracy of brain somatosensory cortex and language-related cortices.

Conclusions

Needling HT 5 and GB 39 may modulate language function through a complex brain network, suggesting that it may be benefificial to the recovery of language function in patients with aphasia.

     

Comparison of Electroacupuncture and Mild-Warm Moxibustion on Brain-Gut Function in Patients with Constipation-Predominant Irritable Bowel Syndrome: A Randomized Controlled Trial

Objective

To compare the effects of electroacupuncture (EA) and mild-warm moxibustion (Mox) therapies for constipation-predominant irritable bowel syndrome (C-IBS) patients.

Methods

Sixty C-IBS patients were assigned to 2 groups by simple randomized method, i.e. EA group (30 cases) and Mox group (30 cases). Both EA and Mox treatments were performed on bilateral Tianshu (ST 25) and Shangjuxu (ST 37) for 30 min each time, 6 times per week, for 4 consecutive weeks. The gastrointestinal symptoms and psychological symptoms of the two groups were scored before and after treatment. The effects on the corresponding functional brain areas, namely the anterior cingulate cortex (ACC), insular cortex (IC) and prefrontal cortex (PFC) were observed by functional magnetic resonance imaging (fMRI) before and after treatment.

Results

Compared with the Mox group, greater improvements in abdominal distension, defecation frequency, diffificulty in defecation and stool features were observed in the EA group (all P<0.01), both Hamilton Anxiety Rating Scale and Hamilton Depression Rating Scale scores were signifificantly decreased in the EA group (all P<0.01). Finally, decreased activated voxel values were observed in the ACC, right IC and PFC brain regions of EA group with 150 mL colorectal distension stimulation (P<0.05 or P<0.01).

Conclusions

Both EA and Mox could signifificantly improve some of the most intrusive symptoms of C-IBS patients, and EA was more effective than Mox. The therapeutic effect of these two therapies might through modulating of the brain-gut axis function. (Registration No. ChiCTRTRC-11001349).

     

Identification of Human Serum Protein Targets of Qianggu Decoction (强骨饮) in Primary Type Ⅰ Osteoporosis Based on Tandem Mass Tag Labeling and Liquid Chromatography-Tandem Mass Spectrometry Technology

Objective: To investigate the serum protein targets of Qianggu Decoction (强骨饮, QGD) on treating osteoporosis by the proteomics analysis using tandem mass tag (TMT) and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Methods: Twenty serum protein samples were recruited (10 patients with primary type Ⅰ osteoporosis before and after QGD treatment) and the high abundance ratios protein was removed, two serum samples were extracted and labeled with TMT reagent. Then, mass spectrometric detection, identification of differentially expressed proteins and bioinformatics analysis of differentially expressed proteins were carried out. Results: A total of 60 proteins were identified, within a 99% confidence interval, to be differentially regulated of which, 34 proteins were up-regulated and 26 proteins were down-regulated. Differentially expressed proteins analyzed by Gene Ontology (GO) annotation mainly get involved in 12 different biological processes, 7 types of cellular components, and 6 kinds of molecular functions. Angiotensinogen (AGT), stromelysin-1 (MMP3), heparanase (HPSE) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) were screened as candidate protein targets of QGD treatment, which were related to metabolic mechanism of bone remodeling and/or bone collagen of osteoporosis. By the utilization of the protein-protein interaction network analysis tool named STRING10.0, it showed that AGT, MMP3, HPSE and GAPDH were located in the key node of the protein-protein interactions network. Furthermore, AGT, MMP3, HPSE and GAPDH were found to be directly related to BMP, MAPK, Wnt, SMAD and tumor necrosis factor ligand superfamily member 11 (TNFSF11) families. Conclusions: The proteomics analysis by using TMT combined with LC-MS/MS was a feasible method for screening the potential therapeutic targets associated with QGD treatment. It suggests that AGT, MMP3, HPSE and GAPDH may be candidate protein targets of QGD treatment which can be used as therapeutic effect monitor and early diagnosis of primary type Ⅰ osteoporosis.     

Chinese Tuina Downregulates the Elevated Levels of Tissue Plasminogen Activator in Sciatic Nerve Injured Sprague-Dawley Rats

Objective

To elucidate the mechanism of Chinese tuina in treating sciatic nerve crush injury, and to detect the levels of tissue plasminogen activator (tPA) and plasminogen activator inhibitor-1 (PAI-1), which is thought to play an important role in nerve regeneration.

Methods

Thirty-two adult male Sprague-Dawley rats were subjected to sciatic nerve crush injury and 16 rats (sham-operated group) went through a sham operation. Control group was given no treatment while tuina group received tuina therapy since day 7 post-surgery. Tuina treatment was performed once a day and lasted for 20 days. The sciatic functional index was examined every 5 days during the treatment session. The rats’ gastrocnemius muscles were evaluated for changes in mass and immunohistochemistry techniques were performed to detect the levels of tPA and PAI-1.

Results

Tuina therapy improved the motor function of sciatic nerve injured rats (P<0.05), however, it did not increase muscle volume (P<0.05). Tuina downregulated the levels of tPA and PAI-1 (P<0.05).

Conclusions

The present study implies that tuina treatment could accelerate rehabilitation of peripheral nerve injury.

     

Three generations of matrilineal excess of birth defects in Irish families with neural tube defects

Background

Neural tube defects (NTDs) and birth defects overall are more likely to occur among maternal compared to paternal relatives in two generations (uncles/aunts and first cousins) of Irish families where an individual has been born with an NTD.

Aims

The aim of this study was to determine if the matrilineal excess persisted into the third generation.

Methods

First cousins were interviewed about their pregnancy outcomes and their offsprings’ health.

Results

Maternal first cousins once removed (FCOR) were more likely to have birth defects than paternal FCOR: 6.7 versus 3.5% (adjusted odds ratio 1.49, 95% CI 0.57, 3.89). No NTDs occurred. Folic acid supplementation significantly reduced the risk of birth defects (P = 0.04).

Conclusions

This study demonstrates an excess of birth defects among maternal relatives in three consecutive generations of NTD families, and supports the hypothesis that an underlying mechanism links distant maternal relatives in at least some NTD families.