Immunomodulatory Mechanism of Kangxian Yixin Prescriptions on Autoimmune Injury Mice Based on Th17/Treg Cell Balance / 中国实验方剂学杂志

ObjectiveTo investigate the immunomodulatory mechanism of Kangxian Yixin prescription （KYP） on autoimmune injury mice and its relationship with the T helper 17 （Th17）/regulatory T cell （Treg） balance. MethodSixty healthy 8-week-old male BALBc mice were randomly divided into a normal group and an experimental group at a ratio of 1∶5. On the 0th， 7th， and 28th days， 0.2 mL of porcine cardiac myosin emulsion （containing 0.1 mg of porcine cardiac myosin） was subcutaneously injected into the groin， armpit， and back of the mice in the experimental group to induce an animal model of myocardial immune injury. Mice with myocardial immune injury were randomly divided into a model group （Model）， a KYP group （20.4 g·kg-1·d-1， ig）， and a valsartan group （12 mg·kg-1·d-1， ig）. Mice in the control group and the model group received the same amount of normal saline by gavage. After four weeks of intervention， the heart tissues were collected. Hematoxylin-eosin （HE） staining and Masson staining were used to detect pathological damage in heart tissues. Enzyme-linked immunosorbent assay （ELISA） was used to detect the expression of type B-type natriuretic peptide （BNP）， anti-cardiac antibody， interleukin-17 （IL-17）， and interleukin-10 （IL-10） in the serum of mice， and the expression levels of Th17 cells and Tregs in the spleen were detected by flow cytometry. The protein expression of B-cell lymphoma-2 （Bcl-2） and Bcl-2-associated X （Bax） in heart tissues was detected by Western blot， and the mRNA expression of retinoid-related orphan receptor gamma t （RORγt） and forkhead box P3 （FoxP3） in the spleen was detected by quantitative real-time polymerase chain reaction （Real-time PCR）. ResultCompared with the control group， the model group showed worsened pathological damage in heart tissues， elevated serum levels of BNP， anti-myocardial antibody， and IL-17， decreased serum expression of IL-10 （P<0.05）， increased expression of Th17 cells and reduced expression of Tregs in spleen tissues （P<0.05）， increased protein expression of Bax， diminished Bcl-2 protein expression， elevated Bax/Bcl-2 ratio， up-regulated mRNA expression of RORγt， dwindled mRNA expression of FoxP3， and elevated ratio of RORγt/FoxP3 （P<0.05）. Compared with the model group， the KYP group and the valsartan group displayed relieved pathological damage in heart tissues， decreased serum expression of BNP， anti-myocardial antibody， and IL-17， increased serum expression of IL-10 （P<0.05）， reduced expression of Th17 cells and increased Tregs in spleen tissues （P<0.05）， dwindled protein expression of Bax and elevated protein expression of Bcl-2 in heart tissues （P<0.05）， diminished Bax/Bcl-2 ratio， reduced mRNA expression of RORγt， up-regulated FoxP3， and down-regulated ratio of RORγt/FoxP3 （P<0.05）. ConclusionKYP may improve myocardial immune damage by regulating the Th17/Treg cell balance.

Kangxian Yixin Prescription Interferes with Mitochondrial Permeability Transition Pore to Inhibit Cardiomyocyte Apoptosis / 中国实验方剂学杂志

Objective:To explore the mechanism of Kangxian Yixin prescription in regulating mitochondrial permeability transition pore（mPTP）and inhibiting cardiomyocyte apoptosis. Method:H9c2 cardiomyocytes were cultured routinely. After 8 h of starvation，the cells were divided into the normal group，model group，Kangxian Yixin prescription（0.25 g·L^-1） group，and cyclosporin A（CsA，10 μmol·L^-1） group and treated with the corresponding drugs for 24 h for follow-up experiments. The H9c2 cardiomyocyte hypertrophy model was induced by norepinephrine（NE），whose optimal concentration was determined by real-time polymerase chain reaction （Real-time PCR）. The degree of mPTP opening was detected by flow cytometry， followed by the measurement of mRNA and protein expression levels of apoptosis-related factors cyclophilin D（Cyp-D），cytochrome C（Cyt-C），and cysteine aspartate-specific protease-3（Caspase-3） after mPTP opening and the quantification of mitochondrial membrane potential. Result:When the concentration of NE was 200 μmol·L^-1， the mRNA expression levels of atrial natriuretic peptide（ANP） and brain natriuretic peptide（BNP） were the highest， implying that it was the optimal concentration to induce H9c2 cell hypertrophy. Compared with the normal group，the model group exhibited excessive opening of mPTP，weakened relative fluorescence intensity in mitochondria， decreased mitochondrial membrane potential（<italic>P</italic><0.05，<italic>P</italic><0.01），and elevated mRNA and protein expression of Cyp-D，Cyt-C，and Caspase-3（<italic>P</italic><0.05）. Compared with the model group，both Kangxian Yixin prescription and CsA inhibited mPTP opening，enhanced the relative fluorescence intensity of mitochondria， increased mitochondrial membrane potential（<italic>P</italic><0.05，<italic>P</italic><0.01），and lowered the mRNA and protein expression of Cyp-D，Cyt-C，and Caspase-3 （<italic>P</italic><0.05）. Conclusion:Kangxian Yixin prescription inhibits cardiomyocyte apoptosis possibly by regulating mPTP opening and inhibiting the expression of apoptosis-related factors Cyp-D，Cyt-C， and Caspase-3.

Study on establishing animal model of myocardial infarction of blood-stasis and Xin-qi deficiency syndrome or blood-stasis and Xin-yang deficiency syndrome type / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To investigate the method for establishing animal model of integrative medical disease/ syndrome and its evaluation.</p><p><b>METHODS</b>Rat myocardial (heart failure) model was established by anterio-descending coronary arterial ligation, and treated by nitric oxide synthase inhibitor 9-12 weeks after operation to induce hypertension and aggravate heart failure. The model rat was observed 12 weeks to collect the information of four-diagnosis, for visceral qi-blood and excess-deficiency syndrome differentiation in combining with the eight-principal syndrome differentiation according to the standard of deficiency syndrome and blood-stasis syndrom issued by Chinese Association of integrative medicine.</p><p><b>RESULTS</b>After modeling, the model rats revealed ascended ST segment and abnormal Q-wave in ECG, with the visceral ratio, left ventricular area and myocardial collagen content significantly more than those in the sham-operative group (P < 0.01), showing a blood-stasis syndrome. Besides, Xin-qi deficiency syndrome, exhibiting as quickened heart rate, shortened swimming time and lowered cardiac function, appeared in the model rats (P < 0.01), which was aggravated in the late stage due to the increased blood pressure and deteriorated cardiac function, even revealed the manifestation of Yang-deficiency syndrome as low body temperature and polyuria.</p><p><b>CONCLUSION</b>The basic thinking path of TCM animal disease/syndrome model establishment and evaluation should adopt the normative, mature and unified standard and methods formed in clinical and experimental study of integrative medicine.</p>

Comparative study on early application of the recipe for activating blood circulation and the recipe for supplementing qi for inhibiting left ventricular remodeling and apoptosis in rats with heart failure / 中国中药杂志

<p><b>OBJECTIVE</b>To compare the characteristics of early application of the recipe for activating blood circulation and the recipe for supplementing qi for inhibiting left ventricular remodeling and apoptosis in rats with heart failure.</p><p><b>METHOD</b>The left coronary artery occlusion was conducted to establish the rat model of heart failure after myocardial infarction. The model rats were randomly divided into 5 groups, including model group, activating blood circulation group (8 g x kg(-1)), supplementing qi group (8 g x kg(-1)), activating blood circulation plus supplementing qi group (16 g x kg(-1)), and captopril group (10.125 g x kg(-1)), and a sham operation group was set up as negative control group. The drugs were administrated on the second day after myocardial infarction with a therapeutic course of 4 weeks or 8 weeks. The heart function was detected by impedance method; Pathological staining and image analysis were used to determine the perimeter and the area of left ventricular cavity, and myocardial nuclei number and collagen content per unit area; Apoptosis percentage of the myocardial cell was detected by TUNEL and the content of Ang II in the cardiac muscle was measured by radioimmunoassay.</p><p><b>RESULT</b>In comparison with the model group, the function of left ventricular contraction function improved, the area of left ventricular cavity diminished, and proliferation of collagen, content of Ang II and apoptosis percentage of the myocardial cell reduced in all of the treatment groups (P < 0.05 or P < 0.01). After treatment of 8 weeks, the activating blood circulation group was similar to the sham operation group in improvement of cardiac index, and decreases of the area of left ventricular cavity and the content of Ang II; Apoptosis percentage of the myocardial cell in the activating blood circulation group was significantly lower than that in the supplementing qi group.</p><p><b>CONCLUSION</b>Both the recipes for activating blood circulation and for supplementing qi can inhibit left ventricular remodeling and myocardial apoptosis, and delay development of heart failure, with the best effect in the activating blood circulation group after treatment of 8 weeks.</p>

Comparative study on effect of recipe for activating blood circulation and replenishing Qi on left ventricular remodeling in rats with left heart failure after myocardial infarction / 中国中西医结合杂志

<p><b>OBJECTIVE</b>Comparative study to the effect of Chinese herbal medicine on left ventricular remodeling in rats with left heart failure after myocardial infarction (MI).</p><p><b>METHODS</b>Rat's model of left heart failure after myocardial infarction was treated with injection for activating blood circulation (ABCI, consisted of R. Salviae miltiorrhizea; Rh. Ligusticum wallichii and F1. Carthamus tinctorius) and injection for replenishing Qi (RQI, consisted of R. Codonopsis Pilosulae and R. Astragalus membranaceus) respectively. The effect of treatment were evaluated by observing and comparing the changes of heart morphological structure, collagen element, heart weight/body weight ratio (HW/BW), left intraventricular area (LVA), ratio of ventricular wall thinning in MI area and myocardial nuclei number (MNN) per square area.</p><p><b>RESULTS</b>In comparison with the model group, the reduction of collagen tissue around myocardial cells in living area of MI, HW/BW and LVA of ABCI and RQI group were lower, and MNN per square area was higher significantly (all P < 0.05).</p><p><b>CONCLUSION</b>Both ABCI and RQI, though without positive myodynamia, showed certain inhibitory effect of left ventricular remodeling in rats with left heart failure after MI.</p>