[Tianma Gouteng Decoction regulates MFN2 expression to delay vascular aging in spontaneously hypertensive rats].

We studied the effect of Tianma Gouteng Decoction on vascular aging in spontaneously hypertensive rats(SHRs) to explore the molecular mechanism of the decoction in improving arterial vascular aging by regulating the expression of mitofusin 2(MFN2).Twenty 64-weeks-old SHRs were randomly assigned into the aging group and the treatment group(Tianma Gouteng Decoction 5.48 mg·kg~(-1)).Wistar-Kyoto(WKY) rats of 64 weeks old were taken as the normal group and SHR rats of 14 weeks old as the young group.The intervention with Tianma Gouteng Decoction lasted for 12 weeks.We then employed HE staining and Masson staining to observe the morphology of thoracic aorta under an electron microscope and measured the malondialdehyde(MDA) content, superoxide dismutase(SOD) activity, respiratory chain complex Ⅲ level, and thioredoxin peroxidase(TPX) activity.The vascular aging was detected via the biomarker senescence-associated beta-galactosidase(SA-ß-Gal).The expression levels of MFN2 and aging marker proteins silent information regulator 1(SIRT1), Klotho, p21, and p53 in thoracic aorta were detected by immunohistochemistry/fluorescence, qRT-PCR, and Western blot.Compared with the young group and the normal group, the aging group had risen blood pressure, lumen stenosis caused by thickened intima of blood vessels, decreased SOD and TPX activities, increased MDA and mitochondrial respiratory chain complex Ⅲ levels, down-regulated expression of MFN2, SIRT1, and Klotho, and up-regulated expression of p21 and p53(P<0.01 or P<0.05).The treatment with Tianma Gouteng Decoction significantly lowered blood pressure, mitigated vascular intimal thickening, increased SOD and TPX activities, and decreased MDA and mitochondrial respiratory chain complex Ⅲ levels, thus alleviating vascular aging.At the same time, the decoction up-regulated the expression of MFN2, SIRT1, and Klotho, while down-regulated that of p21 and p53(P<0.01 or P<0.05).In summary, Tianma Gouteng Decoction can significantly delay the vascular aging in hypertension.Specifically, it may up-regulate the expression of MFN2 and regulate the expression of aging-related proteins to alleviate oxidative stress.

Mitofusin 2 Downregulation Triggers Pulmonary Artery Smooth Muscle Cell Proliferation and Apoptosis Imbalance in Rats With Hypoxic Pulmonary Hypertension Via the PI3K/Akt and Mitochondrial Apoptosis Pathways.

During hypoxia-induced pulmonary hypertension (HPH), pulmonary artery smooth muscle cells (PASMCs) proliferate as part of the characteristic pulmonary vascular remodeling. We investigated the expression of mitofusin 2(Mfn2) and its role in maintaining the balance between PASMC proliferation and apoptosis during hypoxia. In an experimental model of HPH, we exposed rats to hypoxia (10% ± 0.5% O2) or room air for 4 weeks. We found that Mfn2 messenger RNA and protein levels were reduced and that proliferating cell nuclear antigen protein expression was upregulated in HPH rat lung tissues. We also exposed primary cultured PASMCs from rat pulmonary arterioles to normoxia (21% O2/5% CO2) or hypoxia (2.5% O2/5% CO2) for 24 hours. We found that PASMC proliferation increased under hypoxic conditions and that more hypoxic cells than normoxic cells entered the S + G2/M phase. Additionally, phosphorylated Akt and proliferating cell nuclear antigen expression increased, whereas Mfn2 expression, cleaved caspase 9 expression, and the ratio of mitochondrial to cytosolic cytochrome C expression each decreased. These hypoxia-induced effects were reversed in PASMCs by Mfn2 overexpression and by phosphatidylinositide 3-kinases (PI3K) inhibition. Our results indicate that downregulation of Mfn2 in HPH may activate the PI3K/Akt pathway, thereby causing more cells to enter the S + G2/M phase of the cell cycle and inhibiting the mitochondrial apoptosis pathway.

[Effect of Pinggan Qianyang Recipe Containing Serum on Angiotensin II Induced Vascular Smooth Muscle Cell Proliferation and Migration and DNA Methylation].

OBJECTIVE: To observe the effect of Pinggan Qianyang Recipe (PQR) on inhibiting angiotensin II (Ang II) induced proliferation and migration of vascular smooth muscle cells (VSMCs) and changes of DNA methylation. METHODS: VSMCs were cultured using tissue explant method, and PQR containing serum was prepared. Primarily cultured VSMCs were divided into four groups, the normal group, the model group, the folate group (folic acid intervention) , and the PQR group. The proliferation and migration of VSMCs was duplicated by Ang II. After 24-h Ang II induced culture, 40 microg/mL folic acid was added to the folate group for 48 h, while 5% PQR containing serum was added to the PQR group for 48 h. The cell growth curve of VSMCs was drawn by using Cell Counting Kit (CCK-8). The proliferative activity of VSMC was determined by MTT assay. The migration of VSMCs was measured by Millicell chamber. The general level of cytosine methylation in cell nucleus was detected via 5-mC antibodies immunofluorescence, and mRNA expression levels of DNA methyltransferase 1 (DNMT1) were measured by Real-time q-polymerase chain reaction (q-PCR). RESULTS: VSMCs were promoted by Ang II at 10(-6) mol/L for 24 h. Compared with the normal group, the proliferative activity and migration quantity of VSMCs obviously increased, and DNA methylation level obviously decreased (P < 0.05, P < 0.01). Compared with the model group, the cell growth, proliferative activity and migration quantity of VSMCs obviously decreased and the general DNA methylation level increased in the folate group and the PQR group (P < 0.05, P < 0.01). Compared with the normal group, the mRNA expression of DNMT1 decreased in the model group (P < 0.01). Compared with the model group, mRNA expression of DNMT1 in Ang II induced VSMCs was obviously enhanced in the folate group and the PQR group (P < 0.01). CONCLUSIONS: PQR could inhibit Ang II induced proliferation and migration of VSMCs, and cause high genomic DNA methylation level. Changes of DNA methylation might be associated with DNMT1 expression.

[Association between gene polymorphism of calcium/calmodulin-dependent kinase 4 and efficacy of amlodipine in the treatment of hypertension in Chinese Han nationality].

OBJECTIVE: To evaluate the association between single nucleotide polymorphisms of calcium/calmodulin-dependent kinase 4 (CAMK4) and the therapeutic effect of amlodipine in essential hypertensive patients in Chinese Han nationality.
 METHODS: A total of 108 mild-to-moderate essential hypertension patients in Chinese Han nationality were treated with amlodipine for 8 weeks at a dosage of 5 mg/d. Polymerase chain reaction-restriction fragment length polymorphism was performed to detect the genotypes (rs10491334). Blood pressure was measured and analyzed.
 RESULTS: The result of rs10491334 polymorphism of CAMK4 was consistent with Hardy-Weinberg equilibrium distribution and the frequencies of C allele and T allele were 88.89% and 11.11%, respectively. The systolic blood pressure and diastolic blood pressure before amlodipine treatment were not statistically different among different genotype carriers (P>0.05). The blood pressure was significantly reduced in all patients after amlodipine treatment (P<0.05). Systolic blood pressure was significantly decreased in patients with rs10491334 CC genotype and CT genotype compared with those patients with rs10491334 TT genotype. Total effective rates of CT and TT carriers were higher than those of the CC genotype carriers (P<0.01).
 CONCLUSION: The CAMK4 gene polymorphism might be associated with the efficacy of calcium channel blocker in treating mild-to-moderate essential hypertension patients.

[Effect of Pinggan Qianyang recipe on the expression of Tpx II HSP27 and ANXA1 in the hypothalamus of spontaneously hypertensive rats with hyperactivity of liver-YANG syndrome].

OBJECTIVE: To investigate the effect Pinggan Qianyang recipe on expression of Tpx, HSP27 and ANXA1 in the hypothalamus of spontaneously hypertensive rats (SHRs) with the hyperactivity of liver-YANG syndrome. METHODS: A total of 30 SHRs were subjected to administration of Aconiti Praeparatae Decoction to establish the model of SHR with liver-YANG hyperactivity first, then they were randomly divided into three groups: the control group, the model group and the treatment group (n=10 per group). A total of 10 SD rats were served as the normal group. The rats in control group and treatment group were given Enalapril plus Pinggan Qianyang recipe for four weeks. The change of behavior and blood pressure of rats were monitored. RT-PCR and Western-blot were performed to detect the expression of Tpx II, HSP27 and ANXA1 mRNA and protein in the hypothalamus, respectively. RESULTS: Compared with the normal SD rats, the heart rate, blood pressure and grade of irritability were significantly increased while rotation endurance time was dramatically reduced in the SHR model with liver-YANG hyperactivity (P<0.01), these changes were reversed by the application of Enalapril plus Pinggan Qianyang recipe. Compared with the normal SD rats, the protein and mRNA expression of Tpx II and ANXA1 in the model group were significantly upregulated (P<0.01) while the HSP27 was significantly downregulated (P<0.01). Compared with the model group, the protein and mRNA expression of Tpx II and ANXA1 in the control group or treatment group were significantly decreased (P<0.05 or P<0.01) while HSP27 was significantly increased (P<0.05 or P<0.01). Compared with the control group, the expression of Tpx II and ANXA1 protein in treatment group were significantly reduced (P<0.05 or P<0.01). CONCLUSION: Pinggan Qianyang recipe can improve the blood pressure and behavior in SHRs with hyperactivity of Liver-YANG syndrome, which might be related to the regulation of Tpx, HSP27 and ANXA1 expression in hypothalamuses.

[Study of dahuangzhechong pills on anti-arterial thrombosis with the orthogonal design].

OBJECTIVE: To screen the main component of Dahuangzhechong pill's anti-arterial thrombosis with the orthogonal design and refine Dahuangzhechong pills. METHODS: In accordance with the orthogonal design table (L(16)2(15)), divided herbs into 16 groups and made the appropriate liquid. The liquid was gave to SD rats by intragastric administration,the model group, normal control group received the same volume of physiological saline. Isolated rats' carotid artery after intragastric administration a week,modeled according to ferric chloride inducement the carotid artery thrombosis method, then collected blood, detected content of platelet, thromboxane B2 (TXB2) and 6-keto-prostaglandin F1alpha (6-keto-PGF1alpha), sheared and measured dry weight of the modeling artery, then placed arteries in 10% formalin fixation, observed morphological changes in vascular tissue by HE staining. RESULTS: Pathological examination revealed: each experimental group had thrombosis, softening, dissolution, absorption, and intimal injury, but the severity of thrombosis were diferent. Orthogonal analysis showed: 1, influence on dry weight of thrombus: rhubarb, ground beetle, leeches, peach seed, dry paint, except dry paint P<0.05, the others P<0.01.2, influence on plasma 6- keto-PGF1alpha level: peach seed, dry paint, ground beetle, gadfly, grubs, leeches, rhubarb, except rhubarb P<0.05, the others P<0.01.3, influence on plasma TXB2: ground beetle, peach seed, dried paint, rhubarb, leeches, except leech P<0.05, the others P<0.01.4, influence on platelet count: peach seed, dry paint, rhubarb, ground beetle, gadfly, leeches, except gadfly, leeches P<0.05, the others P<0.01. CONCLUSION: Anti-artery thrombosis of Dahuangzhechong Pill is most closely related with rhubarb, ground beetle, leeches, peach seed, dry paint and gadfly.

[Changes of neuroglobin in the pathologic process of contusion and laceration of brain in children].

OBJECTIVE: To study the role of neuroglobin (Ngb) in the pathologic process of contusion and laceration of brain in children. METHODS: The proteins in the brain tissue were extracted by two-dimensional gel electrophoresis in 3 children undergoing brain ventricular neoplasms resection (normal brain tissue) and in 8 children with contusion and laceration of brain. The image analysis was done using the PDQuest 7.0 software. The differential protein spots were detected and analyzed with Applied Biosystems Voyager System 4307 MALDI-TOF Mass Spectrometer and bioinformatical skills. Ngb expression in the brain tissue was measured using immunohistochemisty. Ngb expression in plasma was measured using ELISA in 15 children with contusion and laceration of brain and 10 healthy children. RESULTS: Expression maps of the brain tissue were established by two-dimensional gel electrophoresis in children with contusion and laceration of brain and healthy children. Six differential protein spots were found and 5 of them were identified by mass spectrum. Immunohistochemisty assay showed that Ngb expression in the brain tissue in children with contusion and laceration of brain was significantly higher than in normal controls (P<0.05). ELISA results showed that Ngb expression in the plasma increased significantly 6, 12, 18, 24 and 48 hours after trauma in children with contusion and laceration of brain compared with healthy children (P<0.01). CONCLUSIONS: Ngb may play an important role in the pathologic process of contusion and laceration of brain in children.

[Corrigendum] Astragaloside IV attenuates hypoxia­induced pulmonary vascular remodeling via the Notch signaling pathway.

Subsequently to the publication of the above article, an interested reader drew to the authors' attention that, in Fig. 4A on p. 6, the 'T' and 'DAPT' data panels appeared to contain some of the same data, although one of the panels appeared to have been rotated through 180° relative to the other. The authors have re­examined their original data, and have realized that this figure was assembled incorrectly. Subsequently, the authors have reperformed the experiments that were concerned with the immunohistochemical detection of PCNA in rat lung tissue samples, and the new results for Fig. 4A are shown in a corrected version of Fig. 4 on the next page. Note that the errors made in assembling the original version of this figure did not quantitatively affect either the results or the overall conclusions reported in this paper. The authors are grateful to the Editor of Molecular Medicine Reports for allowing them this opportunity to publish a Corrigendum; furthermore, they apologize to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 23: Article no. 89, 2021; DOI: 10.3892/mmr.2020.11726].

[Herbs for calming liver and suppressing yang in treatment of hyperthyroidism with hyperactive liver yang: herbal effects on lymphocyte protein expression].

OBJECTIVE: To observe the herbal effects on hyperthyroidism patients with syndrome of hyperactivity of liver-Yang by method for calming the liver and suppressing Yang and investigate its effects on the lymphocyte protein expression. This approach may lay a foundation for the further investigation of the curative mechanisms of calming the liver and suppressing Yang treatment. METHOD: A total of 48 hyperthyroidism patients with syndrome of hyperactivity of liver-Yang were randomly divided into treatment group and control group. The treatment group was treated by method for calming the liver and suppressing Yang in accordance with traditional Chinese medicine (TCM) and the control group with thiamazole tablets for three periods of treatment The therapeutic effects, the score of TCM symptom, electrocardiogram (P wave), thyroid hormones and ultrasound were observed in both groups before and after the treatment. The side effects in the treatment course were observed in both groups. The level of differential protein expression was analyzed by two-dimensional electrphoresis and matrix assisted laser desorption/ionizaton time-of-flight mass spectrometry. RESULT: The treatment group has the effect on stepping down the heart rate, cutting down the P wave amplitude changes, regulating the level of thyroid hormones and decreasing the volume of thyromegaly. There are not statistically significant between the treatment group and control group. However, the treatment group has obviously better effect on regulating TCM symptom and decreasing the side reaction than the control group (P<0.05). There are not statistically significant on the total effective between the treatment group and control group. The average spots in lymphocyte for normal people, before and after treating hyperthyroidism patients with syndrome of hyperactivity of liver-Yang were (429 +/- 31), (452 +/- 28) and (437 +/- 36) spots respectively. Eight down-regulated protein expressions and 11 up-regulated protein expressions were obtained in the hyperthyroidism patients with syndrome of hyperactivity of liver-Yang and normal people. Five strengthened expressions of protein were also obtained in 8 down-regulated expressions of protein and 8 lower expressions of protein in 11 up-regulated expressions of protein before and after treating the migraine patients with syndrome of hyperactivity of liver-Yang. Ten of the total 8 differential protein spots were successfully identified by MALDI-TOF-MS. The functions of these proteins were involved in metabolism associated, transportation, antioxidation, sigal transduction and immume associated protein, etc. according to information provided by NCBI and MSDB database. CONCLUSION: In this study, the TCM complex prescription with herbs for calming the liver and suppressing Yang can regulate the thyroid hormones, improves TCM symptoms, and decrease the adverse reaction. It can possibly regulate lymphocyte protein expression.

[Changes of protein expression profile in vascular tissues of spontaneously hypertensive rats treated by a compound Chinese herbal medicine].

OBJECTIVE: To investigate the effects of a Chinese herbal formula for calming liver and suppressing yang on the protein expressions of vascular tissues in spontaneously hypertensive rats (SHRs), and to explore the mechanism of efficacy. METHODS: Twenty SHRs were randomly divided into model group and treatment group. Another 10 Wistar-Kyoto rats were selected as a normal control. SHRs in the treatment group were administered with the formula for calming liver and suppressing Yang for 4 weeks. During the course of treatment, blood pressure and heart rates were monitored every week and the ethology of rats, including irritability and rotation endurance was also evaluated. After treatment, thoracic aorta was obtained and its proteins were separated by 2-dimensional gel electrophoresis (2-DE). The differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and database query. RESULTS: The formula for calming liver and suppressing yang not only decreased the systolic blood pressure and heart rate, but also improved irritability degree and rotation endurance time of SHRs. This experiment had established the 2-DE graph of protein expressions of vascular tissues in SHRs. Compared with the normal group, the expressions of 15 proteins were down-regulated, and 12 proteins were up-regulated in vascular tissues of the model group. The formula for calming liver and suppressing yang treatment up-regulated expressions of 10 proteins in the 15 down-regulated proteins, and down-regulated 8 proteins in the 12 up-regulated proteins in vascular tissues of SHRs. After analysis, 16 obviously differentially expressed proteins were found, and 13 of them were identified. CONCLUSION: The formula for calming liver and suppressing yang can improve the ethology of SHRs. The mechanism is probably concerned with regulating the protein expressions of vascular tissues.

Comparative transcriptomic analysis revealed novel potential therapeutic targets of traditional Chinese medicine (Pinggan-Qianyang decoction) on vascular remodeling in spontaneously hypertensive rats.

BACKGROUND: Both experimental and clinical studies have revealed satisfactory effects with the traditional Chinese formula Pinggan Qianyang decoction (PGQYD) for improving vascular remodeling caused by essential hypertension. The present study explored various therapeutic targets of PGQYD using mRNA transcriptomics. METHODS: In this study, rats were randomly divided into three groups: Wistar-Kyoto (WKY; normal control), spontaneously hypertensive (SHR), and PGQYD-treated rat groups. After 12 weeks of PGQYD treatment, behavioral tests were employed and the morphology of thoracic aortas were examined with hematoxylin-eosin (HE) and Masson staining and electron microscopy. The mRNA expression profiles were identified with RNA-Seq and quantitative real-time PCR to validate changes in gene expression observed with microarray analysis. The gene ontology and pathway enrichment analyses were carried out to predict gene function and gene co-expressions. Pathway networks were constructed to identify the hub biomarkers, which were further validated by western blotting and immunofluorescence analysis. RESULTS: After PGQYD treatment, the behavioral tests and histological and morphological findings of vascular remodeling were obviously meliorated compared with the SHR group. In the rat thoracic aorta tissues, 626 mRNAs with an exact match were identified. A total of 129 of mRNAs (fold change > 1.3 and P-value < 0.05) were significantly changed in the SHR group compared to the WKY group. Among them, 16 mRNAs were markedly regulated by PGQYD treatment and validated with quantitative real-time PCR. Additionally, target prediction and bioinformatics analyses revealed that these mRNAs could play therapeutic roles through biological processes for regulating cell metabolic processes (such as glycation biology), biological adhesions, rhythmic processes, and cell autophagy. The cellular signaling pathways involved in autophagy may be AGE-RAGE/PI3K/Akt/mTOR signaling pathway. CONCLUSION: The present study provides novel insights for future investigations to explore the mechanisms by which PGQYD may effectively inhibit vascular remodeling by activating the AGE-RAGE/PI3K/Akt/mTOR signal pathway in cell autophagy biology.

Astragaloside IV attenuates hypoxia­induced pulmonary vascular remodeling via the Notch signaling pathway.

The Notch signaling pathway participates in pulmonary artery smooth muscle cell (PASMC) proliferation and apoptosis. Astragaloside IV (AS­IV) is an effective antiproliferative treatment for vascular diseases. The present study aimed to investigate the protective effects and mechanisms underlying AS­IV on hypoxia­induced PASMC proliferation and pulmonary vascular remodeling in pulmonary arterial hypertension (PAH) model rats. Rats were divided into the following four groups: i) normoxia; ii) hypoxia (10% O2); iii) treatment, hypoxia + intragastrical administration of AS­IV (2 mg/kg) daily for 28 days; and iv) DAPT, hypoxia + AS­IV treatment + subcutaneous administration of DAPT (10 mg/kg) three times daily. The effects of AS­IV treatment on the development of hypoxia­induced PAH, right ventricle (RV) hypertrophy and pulmonary vascular remodeling were examined. Furthermore, PASMCs were treated with 20 µmol/l AS­IV under hypoxic conditions for 48 h. To determine the effect of Notch signaling in vascular remodeling and the potential mechanisms underlying AS­IV treatment, 5 mmol/l γ­secretase inhibitor [N­[N­(3,5­difluorophenacetyl)­L­alanyl]­S­phenylglycine t­butyl ester (DAPT)] was used. Cell viability and apoptosis were determined by performing the MTT assay and flow cytometry, respectively. Immunohistochemistry was conducted to detect the expression of proliferating cell nuclear antigen (PCNA). Moreover, the mRNA and protein expression levels of Notch­3, Jagged­1, hes family bHLH transcription factor 5 (Hes­5) and PCNA were measured via reverse transcription­quantitative PCR and western blotting, respectively. Compared with the normoxic group, hypoxia­induced PAH model rats displayed characteristics of PAH and RV hypertrophy, whereas AS­IV treatment alleviated PAH and prevented RV hypertrophy. AS­IV also inhibited hypoxia­induced pulmonary vascular remodeling, as indicated by reduced wall thickness and increased lumen diameter of pulmonary arterioles, and decreased muscularization of distal pulmonary vasculature in hypoxia­induced PAH model rats. Compared with normoxia, hypoxia promoted PASMC proliferation in vitro, whereas AS­IV treatment inhibited hypoxia­induced PASMC proliferation by downregulating PCNA expression in vitro and in vivo. In hypoxia­treated PAH model rats and cultured PASMCs, AS­IV treatment reduced the expression levels of Jagged­1, Notch­3 and Hes­5. Furthermore, Notch signaling inhibition via DAPT significantly inhibited the pulmonary vascular remodeling effect of AS­IV in vitro and in vivo. Collectively, the results indicated that AS­IV effectively reversed hypoxia­induced pulmonary vascular remodeling and PASMC proliferation via the Notch signaling pathway. Therefore, the present study provided novel insights into the mechanism underlying the use of AS­IV for treatment of vascular diseases, such as PAH.

[Clinical efficacy study on calming liver and restraining Yang formula in treating patients with mild or moderate degree of essential hypertension].

OBJECTIVE: To observe the therapeutic effect of calming the liver and restraining the Yang formula in treating patients with mild or moderate degree of essential hypertension (syndrome of hyperactivity of liver-Yang), and to explore its mechanism in lowering blood pressure. METHOD: The 348 patients with EH of stage I , II were randomly divided into two groups, the 174 patients in the treated group were treated with the calming the liver and restraining the Yang formula, and the 174 patients in the control group were treated with amlodipine. The treatment course for them all was 12 weeks. The related clincial symptoms score and quality of life score estimated before and after treatment at 4th week, 8th week and 12th week were observed. Before and after treatment, the ambulatory blood pressure (AMBP), heart rate, blood lipid, serum livels of high-sensitivity C-reactive protein (Hs-CRP), Angiotensin-II (Ang II) and calcitonin gene-related peptide (CGRP) were measured respectively in 40 patients of the treared group and 40 patients of the control group. RESULT: After treatment, the treatment in the treated group showed an effect better than that in the control group in terms of nigh-time blood pressure reducing rate (P < 0.05). The reducing blood pressure variability and total effective rate in the treated group were no significant than that in the control group. In respect of reducing symptomatic scores on dizzy, soreness and weakness of the waist and knees, disturbed and dry and bitter of mouth, ameliorating quality of life score, decreasing the levels of heart rate, total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol(HDL-C) in the treated group were showing marked improvement as compared with that in the control group (P < 0.05 or P < 0.1). The improvement in the level of Ang II , Hs-CRP and CGRP before treatment in two groups were more significant than that after treatment (P < 0.05). However There were no difference in after treatment between the treated group and the control group. CONCLUSION: The calming the liver and restraining the Yang formula shows favorable efficacy in lowering blood pressure on the patients with mild or moderate degree of essential hypertension. It can reduce the clincial symptoms, improve the quality of life, regulate blood lipid metabolism. Its mechanism may be related to the functional relieving inflammatory reaction and inhibition the activity of renin-angiotensin-aldosterone system (RAAS).

[Effect of the formulae for calming the liver and suppressing YANG on lymphocyte proteome in migraine rats with syndrome of hyperactivity of liver-YANG].

OBJECTIVE: To explore the molecular mechanism of the formulae for calming the liver and suppressing YANG in migraine rat model with syndrome of hyperactivity of the liver-YANG. METHODS: A rat model of migraine with hyperactivity of liver-YANG was established through electrical trigeminal ganglion stimulation and syndrome of oral administration of Fuzi decoction. The total proteins of the lymphocyte in the rats were separated by immobilized pH gradient-based 2-dimensional gel electrophoresis (2-DE), and the 2-DE image was analyzed by PDQuest 7.0 software. Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS) and the SWISS-PORT and MSDB database were used to identify differential proteins. RESULTS: The formulae for calming the liver and suppressing YANG could also improve headache. Well-resolution and reproducible 2-DE patterns of rat lymphocyte from normal, model, and therapy tissues were obtained. Eleven of the total 13 differential protein spots were successfully identified by MALDI-TOF-MS. These proteins were alcohol dehydrogenase 3 (ADH3), glycogen phosphorylase, ATP synthase D chain, annexin-3, ubiquitin, neutrophil defensin 4 precursor, melanoma-associated antigen E2, heat shock protein-27, annexin-A1, peroxirdoxin-II, MU class glutathione S-transferase (Fragment)(GSH). CONCLUSION: Differences occur in the expression of lymphocyte proteins in migraine rats with syndrome of hyperactivity of liver-YANG after treatment with the formulae for calming the liver and suppressing YANG, and the 11 identified protein spots may be associated with its mechanism.

[Differential proteomic analysis of thalamus in rats with diffuse axonal injury].

OBJECTIVE: To examine the differential expression of protein of thalamus in rats with diffuse axonal injury. METHODS: Twenty-five rats were randomly divided into a normal group (n=10) and a trauma group (n=15). Total proteins of brain trauma tissue and normal brain tissue were extracted separately, and then proteins were separated by two dimensional gel electrophoresis and stained with Coomassie brilliant blue. The differentially expressed protein spots were identified with biospectrometry. Images were analyzed by PDQuest 7.0. RESULTS: The distribution of protein spots in the trauma group was similar to that of the normal group, the matching rate was 95%, and the repeatability was good. Proteins were mainly displayed at pI 3-8, with relative molecular mass 14.4-75.0 kD. Compared with the normal group, 16 spots of proteins increased and 18 spots of proteins decreased in the trauma group. CONCLUSION: There is some difference in protein expression between the normal group and the trauma group. Brain trauma may lead to changes of proteins in the thalamus.

[Herbs for calming liver and suppressing liver-yang in treatment of migraine with hyperactive liver-yang syndrome and its effects on lymphocyte protein expression: a randomized controlled trial].

OBJECTIVE: To observe the efficacy of herbs for calming liver and suppressing liver-yang in treatment of migraine patients with hyperactivity of liver-yang syndrome and to investigate its effects on the lymphocyte protein expression. This approach may lay a foundation for the further investigation of pathogenic mechanisms in migraine with hyperactive liver-yang syndrome and the curative mechanisms of calming liver and suppressing liver-yang treatment. METHODS: A total of 32 migraine patients with hyperactivity of liver-yang syndrome were randomly divided into treatment group (16 cases) and control group (16 cases). The patients in the treatment group were treated with herbs for calming liver and suppressing liver-yang in accordance with traditional Chinese medicine (TCM) theory and the patients in the control group were treated with Flunarizine Capsules for two courses of treatment. The therapeutic effects, the score of TCM symptom and the changes of headache attack were observed in both groups before and after the treatment. The side effects were also observed in both groups. The level of differential protein expression was analyzed by two-dimensional electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). RESULTS: The herbs for calming liver and suppressing liver-yang had better effects on headache improvement than the Flunarizine Capsules (P<0.01). The cure rates in the treatment and control groups were 87.5% and 75.0% respectively. Vertigo, restlessness and tantrum, and prosopo-heat in the treatment group were also improved as compared with those in the control group (P<0.05). After treatment, the score of TCM symptom in the two groups were all decreased (P<0.01), and there was a significant difference between the treatment group and the control group (P<0.01). The herbs for calming liver and suppressing liver-yang had no side effects. The average protein spots in the blood lymphocyte of normal people, migraine patients with hyperactivity of liver-yang syndrome in the treatment group before and after the treatment were (534+/-42), (552+/-54) and (529+/-55) spots respectively. Six down-regulated protein expressions and 14 up-regulated protein expressions were obtained in the treatment group. Four strengthened protein expressions in the six down-regulated proteins and 11 low protein expressions in the 14 up-regulated proteins were also obtained after treatment. Ten of the total 12 differential protein spots were successfully identified by MALDI-TOF-MS. The functions of these proteins were involved in metabolism, energy generation, transportation, antioxidation, signal transduction and immune, etc. According to information provided by NCBI and MSDB database, there were some proteins closely related to migraine with hyperactivity of liver-yang syndrome, such as peroxiredoxin 2, heat shock protein 27 and annexin A1. CONCLUSION: Herbs for calming liver and suppressing liver-yang is effective in treating migraine, and can improve TCM symptoms. The effects on migraine patients with hyperactivity of liver-yang syndrome may be related to regulating the blood lymphocyte protein expression.

Hypoxia promotes pulmonary vascular remodeling via HIF-1α to regulate mitochondrial dynamics.

BACKGROUND: Increasing research suggests that mitochondrial defect plays a major role in pulmonary hypertension (PH) pathogenesis. Mitochondrial dynamics and quality control have a central role in the maintenance of the cell proliferation and apoptosis balance. However, the molecular mechanism underlying of this balance is still unknown. METHODS: To clarify the biological effects of hypoxic air exposure and hypoxia-inducible factor-1α (HIF-1α) on pulmonary arterial smooth muscle cell (PASMC) and pulmonary arterial hypertension rats, the cells were cultured in a hypoxic chamber under oxygen concentrations. Cell viability, reactive oxygen species level, cell death, mitochondrial morphology, mitochondrial membrane potential, mitochondrial function and mitochondrial biosynthesis, as well as fission-and fusion-related proteins, were measured under hypoxic conditions. In addition, rats were maintained under hypoxic conditions, and the right ventricular systolic pressure, right ventricular hypertrophy index and right ventricular weight/body weight ratio were examined and recorded. Further, we assessed the role of HIF-1α in the development and progression of PH using HIF-1α gene knockdown using small interfering RNA transfection. Mdivi-1 treatment was performed before hypoxia to inhibit dynamin-related protein 1 (Drp1). RESULTS: We found that HIF-1α expression was increased during hypoxia, which was crucial for hypoxia-induced mitochondrial dysfunction and hypoxia-stimulated PASMCs proliferation and apoptosis. We also found that targeting mitochondrial fission Drp1 by mitochondrial division inhibitor Mdivi-1 was effective in PH model rats. The results showed that mitochondrial dynamics were involved in the pulmonary vascular remodeling under hypoxia in vivo and in vitro. Furthermore, HIF-1α also modulated mitochondrial dynamics in pulmonary vascular remodeling under hypoxia through directly regulating the expression of Drp1. CONCLUSIONS: In conclusion, our data suggests that abnormal mitochondrial dynamics could be a marker for the early diagnosis of PH and monitoring disease progression. Further research is needed to study the signaling pathways that govern mitochondrial fission/fusion in PH.

[Adrenal protein expressions after Pinggan Qianyang Formula treatment in hypertensive rats with liver-yang hyperactivity: a comparative proteomic analysis].

OBJECTIVE: To explore the pathogenic mechanism of liver-yang hyperactivity type of hypertension and to observe the effects of Pinggan Qianyang Formula (PGQYF), a compound of traditional Chinese herbals for calming the liver and suppressing yang, so as to provide experimental evidence for new marker proteins of drug therapy. METHODS: A rat model of liver-yang hyperactivity was prepared with spontaneous hypertensive rats (SHRs) by administration of Aconiti Praeparatae Decoction. Adrenal proteins were separated by 2D gel electrophoresis (2-DE). The differentially expressed proteins were identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and database analysis. RESULTS: The rat model of liver-yang hyperactivity was successfully reproduced, and the PGQYF could decrease the grades of irritability, conjunctival congestion and systolic blood pressure of the rats (P<0.05, P<0.01). After analysis, twelve obviously differentially expressed proteins were found, eight of which were identified. The expression levels of isocitrate dehydrogenase and steroidogenic acute regulatory protein in the untreated group were up-regulated as compared with those in the normal control group, and down-regulated in the treatment group. The expression levels of ferritin light chain, elongation factor Tu, Rho GDP disassociation inhibitor 1, flavin reductase and basic transcription factor 3 in the untreated group were down-regulated as compared with those in the normal control group, and up-regulated in the treatment group. CONCLUSION: Differentially expressed adrenal proteins in SHRs with live-yang hyperactivity are successfully identified. This approach may lay a foundation for the further investigation of pathogenic mechanisms in hypertension with liver-yang hyperactivity and the mechanisms of PGQYF treatment.

[Effect of pinggan qianyang on hypothalamic proteome in the hyperthyroid rats with hyperactivity of liver-yang].

OBJECTIVE: To investigate the effect of pinggan-qianyang (PGQY), a Chinese medicine, on hypothalamic proteome in the hyperthyroid rats with hyperactivity of liver-yang, and to explore its mechanism. METHODS: The rat model was established by intraperitoneal injection of levo-thyroxine (L-T4) and fuzi decotion. All the quantitative and qualitative changes of the protein expressions were compared among the normal group,the model group and the treatment group by proteomic techniques. RESULTS: The protein spots in the 3 groups were mainly displayed at the isoelectric point (pI) 3 approximately 10, and the molecular weights were 13.8 approximately 98.8 kD.Compared with the normal group, 6 spots of protein expression increased and 10 decreased in the model group. All the changed protein in the model group returned to normal level after PGQY treatment. Mass-spectrometer and bio-informatics indicated that these proteins were Prohibitin, Peroxiredoxin-6, histidine triad nucleotide-binding protein 1, protein-tyrosine-phosphatase, predicted protein, profilin-2, peroxir doxin-II, heat shock protein-27, and annexin-A1. CONCLUSION: There are differences in the expression of hypothalamus proteins in the hyperthyroid rats with hyperactivity of liver-yang after the treatment with PGQY, and the 9 identified protein spots may be associated with the mechanism of PGQY.

[Effect of calming the liver and suppressing the hyperactive YANG drugs on lymphocyte protein in hypertension patients with hyperactivity of liver-YANG].

OBJECTIVE: To explore the effect of calming the liver and suppressing the hyperactive YANG drugs on the lymphocyte protein and clinical efficacy in the hypertension patients with hyperactivity of liver-YANG, and to identify the therapy. METHODS: Twenty-six hypertension patients with hyperactivity of liver-YANG were treated by calming the liver and suppressing the hyperactive YANG drugs for 2 courses. Symptoms of Chinese medicine and blood pressure were observed, and the separated lymphocyte total protein of normal and hypertensions before and after the treatment were examined by the solid-state pH gradient 2-dimensional gel electrophoresis. The differences of the protein expression were analyzed by ImageMaster 2DE analysis software with two-way patterns. RESULTS: The total efficiency rate of calming the liver and suppressing the hyperactive YANG drugs was 88.5%, and the drugs could significantly relieve the symptoms, such as headache, dizziness, dry mouth, irritability, etc. Calming the liver and suppressing the hyperactive YANG drugs could also remarkably reduce the blood pressure,with significant different between pre-treatment and post-treatment (P<0.05). The average spots of lymphocyte gel proteins in the normal and the hypertension patients with syndrome of hyperactivity of liver-YANG before and after the treatment were 527+/-41,559+/-62, and 543+/-59, respectively. Compared with normal people, the expression of 15 proteins was down-regulated, and 10 up-regulated in the hypertension patients with syndrome of hyperactivity of liver-YANG. Compared with the pre-treatment, the expression of 12 proteins was increased in the 15 down-regulated proteins, and 6 decreased in the 10 up-regulated proteins after the treatment in the hypertension patients with syndrome of the hyperactivity of liver-YANG. CONCLUSION: Calming the liver and suppressing the hyperactive yang drugs may mildly depress the blood pressure and improve the symptoms of Chinese medicine. The effect of drugs in treating hypertension may probably be associated with regulating the expression of some proteins in lymphocytes.