Improvement of Culture Conditions and Plant Growth Regulators for In Vitro Callus Induction and Plant Regeneration in Paeonia lactiflora Pall.

Owing to its high ornamental, medicinal and horticultural values, herbaceous peony (Paeonia lactiflora Pall.) has been widely used as a landscaping and economical plant around the world. However, the lack of an efficient and stable regeneration system in P. lactiflora restricts its rapid propagation and large-scale production. By testing the key factors affecting callus formation, proliferation, adventitious bud induction and rooting, here, we developed an in vitro system for callus induction and regeneration in P. lactiflora. Our results show that callus formation was affected by explant types, culture environment, basal medium and plant growth regulators. Using cotyledons as explants, we established good conditions for P. lactiflora callus induction and callus proliferation. We effectively obtained adventitious buds differentiated from callus in Murashige and Skoog (MS) medium containing kinetin (KT) and thidiazuron (TDZ). Adventitious bud growth can be further promoted by adding gibberellin 3 (GA3), 1-naphthaleneacetic acid (NAA) and 6-benzyleaminopurine (6-BA) into the MS medium. A high percentage of rooting can be achieved by adding indolebutyric acid (IBA) and activated carbon (AC) to ½ MS medium. Overall, our system promotes callus induction and adventitious bud regeneration for P. lactiflora through improved culture conditions and plant growth regulators in the culture media, and lays a foundation for subsequent genetic engineering research.

Re-Du-Ning injection ameliorates radiation-induced pneumonitis and fibrosis by inhibiting AIM2 inflammasome and epithelial-mesenchymal transition.

BACKGROUND: Radiation-induced lung injury (RILI) is a common side effect in chest radiotherapy patients, and there is no good medicine to treat it. Re-Du-Ning (RDN) injection is a traditional Chinese medicine that is clinically used to treat upper respiratory tract infections and acute bronchitis. RDN has the advantage of high safety and mild side effects. The mechanism of most traditional Chinese medicine preparations is unknown. PURPOSE: To illustrate the mechanisms of RDN for the treatment of RILI. METHODS: Female C57BL/6 mice were used to establish a RILI model via irradiation, and RDN injection was intraperitoneally administered at doses of 5, 10, and 20 ml/kg. The cytokines were measured by ELISA and qPCR. The data related to Absent in melanoma 2 (AIM2) inflammasome were analyzed via ELISA and a network pharmacological approach. In addition, the data related to epithelial-mesenchymal transition (EMT) were analyzed via immunofluorescence, Western blotting, and a network pharmacological approach. RESULTS: RDN robustly alleviated RILI. Meanwhile, RDN downregulated inflammatory cells' infiltration and the expression of pro-inflammatory cytokines, such as IL-1ß, IL-6, and TNF-α. Next, the potential molecular mechanisms of RDN were predicted through network pharmacology analysis. RDN may ameliorate radiation pneumonitis (RP) by inhibiting AIM2-mediated pyroptosis. Moreover, RDN treatment inhibited EMT and phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT) pathway. The active compounds from Lonicera japonica Thunb. decreased the phosphorylation of Akt. CONCLUSION: These findings demonstrate that RDN, as a traditional Chinese medicine preparation, will be a candidate drug for treating RILI.

Treatment of urinary incontinence after total hysterectomy with acupuncture: A case report.

RATIONALE: Acupuncture is a significant feature of traditional Chinese medicine, which can dredge the channels, harmonize qi and blood, replenish deficiency and relieve excess, strengthen the body and remove pathogens to treat urinary incontinence after hysterectomy, and improve the quality of life, which is simple, convenient, inexpensive, and practical. PATIENT CONCERNS: After a total hysterectomy, the catheter was retained every day, causing urinary incontinence and elderly urine wetness for 30 days. DIAGNOSES: Postoperative urinary incontinence for 1 month; type 2 diabetes for 4 years. Hypertension for 2 years. INTERVENTIONS: From the first month after operation, acupuncture on bilateral, Ciliao (BL32), Zhongliao (BL33), and Xialiao (BL34). OUTCOMES: The patient experienced bladder fullness on the 2nd day. On the 3rd day, the patient could arose from bed and urinated on her own. On the 4th day, she could urinate freely. The time and frequency of urination were normal. LESSONS: acupuncture is safe and effective mode for the treatment of urinary incontinence issues after total hysterectomy. It greatly improves the quality of life and daily wellbeing.

Cycloastragenol ameliorates experimental heart damage in rats by promoting myocardial autophagy via inhibition of AKT1-RPS6KB1 signaling.

Cycloastragenol, a naturally occurring compound in Astragali Radix, has been demonstrated to possess various pharmacological actions including anti-aging, anti-inflammation, anti-fibrosis, antibacterial, liver and endothelium protection. However, whether cycloastragenol ameliorates heart failure remains unclear. Isoproterenol administration to rats triggered classic cardiac damage, as demonstrated by objective parameters of cardiac dysfunction. The treatment of cycloastragenol improved deranged cardiac parameters in the isoproterenol-induced heart damage model in a dose-dependent manner. At the same time, cycloastragenol markedly ameliorated cardiac histological changes and down-regulated serum levels of various neuroendocrine factors including norepinephrine, aldosterone, brain natriuretic peptide, endothelin 1, angiotensin II and so on. Moreover, the expressions of matrix metalloproteinase-2 (MMP-2) and MMP-9 in rat heart were also inhibited by cycloastragenol. Mechanistically, augmenting autophagy of myocardial cells via the inhibition of AKT1-RPS6KB1 signaling contributed to the improvement of isoproterenol-induced rat heart failure by cycloastragenol. These results suggest that cycloastragenol ameliorates cardiac dysfunction and remodeling through promoting autophagy in myocardial cells and suppressing MMP-2 and MMP-9 expressions, indicating that it could be a drug candidate for patients with congestive heart failure.

Icariin attenuated oxidative stress induced-cardiac apoptosis by mitochondria protection and ERK activation.

Our previous study showed that Icariin (ICA) has anti-cardiac hypertrophy effect in rats with an unknown mechanism. In the present study, we aimed to clarify the cardiac protective effect and mechanism of ICA in vitro. H9C2 cardiac myocytes were incubated with H2O2 to build up the oxidative stress injury model. The results showed that pre-treatment of ICA protected cells against the toxicity induced by H2O2. H2O2 treatment significantly reduced H2O2-induced apoptosis, evidenced by lower Annexin V/PI stained cells and less PARP and caspase-3/9 activation. Mitochondria membrane potential (MMP) dissipation occurred following the exposure of H2O2, which could be prevented by ICA treatment. Moreover, Ca2+ homeostasis was preserved by ICA and ROS generation was significantly suppressed by ICA incubation. Interestingly, ICA treatment increased the phosphorylation of upstream ERK mitogen-activated protein kinase (MAPK) while ERK inhibitor U1026 could reverse the protective effect of ICA. Overall, ICA seems to protect the cardiac cells from oxidative stress injury through ROS scavenge and stimulation of ERK pathway which may explain its effects in vivo.

Icariin attenuates cardiac remodelling through down-regulating myocardial apoptosis and matrix metalloproteinase activity in rats with congestive heart failure.

OBJECTIVES: In this study, the anti-heart failure effect of icariin, a natural flavonol glycoside, and the underlying mechanisms were investigated. METHODS: Heart failure was induced by isoproterenol in male Sprague-Dawley rats. Matrix metalloproteinase activity was determined by gelatin zymography assay. The mRNA expression was determined by real-time PCR. The protein expression was determined by Western bolt. Mitochondria structure was examined by transmission electron microscopy. KEY FINDINGS: Isoproterenol administration resulted in a severe heart failure, as shown by the increased levels of left ventricular weight index, heart rate, left ventricular end diastolic pressure, maximal rate of left ventricular pressure decline (dp/dt(min) ), decreased levels of left ventricular systolic pressure and maximal rate of left ventricular pressure rise (dp/dt(max) ). Against these, icariin dose-dependently reversed the changes of these cardiac morphometric and haemodynamic parameters. In addition, icariin significantly inhibited serum levels of tumour necrosis factor-α, noradrenaline, angiotensin II and brain natriuretic peptide in rats with congestive hear failure and improved the histological changes, including cardiocyte hypertrophy, cardiocyte degeneration, inflammatory infiltration and cardiac desmoplasia. Furthermore, the expression and activity of matrix metalloproteinase (MMP)-2 and MMP-9, which regulate collagen production, were also blocked by icariin. Moreover, myocardial apoptosis was remarkably attenuated by icariin through regulating Bcl-2/Bax axle. CONCLUSIONS: Icariin ameliorates left ventricular dysfunction and cardiac remodelling through down-regulating matrix metalloproteinase-2 and 9 activity and myocardial apoptosis in rats with congestive heart failure.

[Levels of homocysteine and polymorphisms of homocysteine metabolism-related enzymes in patients with type 2 diabetes mellitus and coronary heart disease].

OBJECTIVE: To explore the significance of Hey, the gene polymorphisms of methylenetetrahydrofolate reductase (MTHFR) C677 T and cystathionine beta- synthase (CBS844) ins68 in type 2 diabetes mellitus (DM) with coronary heart disease (CHD) in China. Methods We selected 70 patients with type 2 DM and CHD, 71 type 2 diabetes patients, and 85 controls in Han nationality from northern China. Hey levels were measured by fluorescence polarization immunoassay (FPIA) and the plasma folate and vitamin B12 levels by microparticle enzyme immunoassay (MEIA). The gene polymorphisms of the MTHFR C677 T were determined by PCR- RFLP assay and the gene polymorphisms of the CBS 844ins68 were determined by PCR assay. RESULTS: The plasma Hey levels in DM with CHD group (14.8 micromol/L) were significantly higher than in DM group (11.1 micromol/L) and control group (11.2 micromol/L), (P < 0.01). The levels of plasma folate and Vitamin B12 in DM with CHD group were significantly lower than in DM group and control group, (P < 0.05). The T allelic frequency of MTHFR in DM and CHD group was significantly higher than that in DM group and controls (45% vs 26.8%, 31.2%, P < 0.01). There were no significant differences in the frequencies of CBS 844ins68 polymorphism among 3 groups (P > 0.05). Logistic-regression analysis indicated that the OR of HHcy was 4.547 (95% CI 1.97-10.496) (P < 0.01), the OR of MTHFR 677 with T (including MTHFR CT genotype and Tr genotype)was 2.369 (95% CI 1.160-4.841), (P = 0.018), and the OR of CBS 844ins68 was 0.384 (95% CI 0.033-4.423), (P = 0.443). CONCLUSION: Hyperhomocysteinemia and MTHFR with T allele might be the risk factors for DM with CHD in northern Chinese Han population.

Ethanol extract from Epimedium brevicornum attenuates left ventricular dysfunction and cardiac remodeling through down-regulating matrix metalloproteinase-2 and -9 activity and myocardial apoptosis in rats with congestive heart failure.

Epimedium, a traditional Chinese herb, has been used for the remedy of coronary heart disease, impotence and osteoporosis in traditional oriental medicine. However, despite extensive pharmacological studies, the molecular mechanism of the anti-heart failure effect of epimedium is little known. In the present study, we investigated the pharmacological action mechanism of ethanol extract of epimedium (EPI-ext) on isoproterenol-induced congestive heart failure (CHF) in rats. Isoproterenol administration resulted in severe heart failure, as shown by the increased levels of left ventricular (LV) weight index and heart rate, as well as LV end diastolic pressure, and by the decreased levels of LV systolic pressure, maximal rate of LV pressure rise, and maximal rate of LV pressure decline. EPI-ext dose-dependently reversed the changes of these cardiac morphometric and hemodynamic parameters. In addition, EPI-ext significantly inhibited the serum levels of tumor necrosis factor alpha, norepinephrine, angiotensin II and brain natriuretic peptide in rats with CHF and improved the histological changes including cadiocyte hypertrophy, cadiocyte degeneration, inflammatory infiltration, and cardiac desmoplasia. Furthermore, the expression and activities of matrix metalloproteinase-2 and -9, which regulate collagen production, were also blocked by EPI-ext. Moreover, myocardial apoptosis was remarkably attenuated by EPI-ext through the regulating Bcl-2/Bax axle. In conclusion, EPI-ext ameliorates LV dysfunction and cardiac remodeling through down-regulating matrix metalloproteinase-2 and -9 activity and myocardial apoptosis in rats with CHF.

Apolipoprotein B gene 3'VNTR polymorphism: association with plasma lipids and coronary heart disease in Han Chinese.

BACKGROUND: Studies that considered polymorphisms within the apolipoprotein B (APOB) gene as risk factors for coronary heart disease (CHD) have reported conflicting results. METHODS: The phenotypic effects of the 3'VNTR polymorphism of the APOB gene on the susceptibility to CHD were investigated in 120 unrelated healthy individuals and 137 CHD patients. The internal structure of APOB gene 3'VNTR alleles was also analyzed by the methods of SspI restriction mapping and DNA sequencing of the allele fragments. RESULTS: In total, 14 segregating alleles and 32 genotypes of APOB gene 3'VNTR were characterized in the pooled total of 257 subjects. The frequency of 3'VNTR-B alleles [hypervariable element (HVE) > or =38)] in the CHD cases was higher than that of the controls (10.95% vs. 5.00%, p<0.05). 3'VNTR-B allele was dependently related to total cholesterol levels (p<0.05). Compared with SS homozygotes, 3'VNTR-B allele carriers were associated with an increased risk of CHD (OR=2.137, 95% CI=1.055-4.328, p=0.0349). No significant differences in the internal structure and sequences of APOB gene 3'VNTR alleles were found between cases and controls. CONCLUSIONS: APOB gene 3'VNTR polymorphism exerts an impact on lipid metabolism and may contribute to the susceptibility to the development of CHD in Han Chinese.

Aqueous extract of Yin-Chen-Hao decoction, a traditional Chinese prescription, exerts protective effects on concanavalin A-induced hepatitis in mice through inhibition of NF-kappaB.

In traditional oriental medicine, Yin-Chen-Hao decoction is used for the remedy of liver diseases such as hepatitis, fatty liver, hepatocirrhosis and jaundice. However, despite extensive pharmacological study, the molecular mechanism of the anti-inflammatory effect of Yin-Chen-Hao decoction is poorly understood. In this study, we have investigated the pharmacological action on the mechanism of concanavalin A-induced T cell-dependent hepatitis in mice. Concanavalin A administration resulted in a severe liver injury. This was shown through increased levels of serum transaminase and lactic dehydrogenase, and increased liver DNA fragmentation and caspase-3 activity. Pretreatment with the aqueous extract from Yin-Chen-Hao decoction dose-dependently inhibited the elevation in transaminase and lactic dehydrogenase activity, and reduced liver DNA fragmentation and caspase-3 levels. There was an improvement in histological changes including inflammatory infiltration, hepatocyte necrosis and degeneration, and Kupffer cell hyperplasia. In addition, Yin-Chen-Hao decoction significantly inhibited tumour necrosis factor-alpha (TNF-alpha) production in-vitro and in-vivo. Moreover, the activation of nuclear factor kappa B (NF-kappaB), which regulates TNF-alpha production, was blocked by Yin-Chen-Hao decoction in-vitro and in-vivo. In conclusion, Yin-Chen-Hao decoction was capable of regulating T-cell-mediated liver injury in-vivo. This event may have depended on the decrease of TNF-alpha production through the inhibition of NF-kappaB activation.

Apolipoprotein A5 gene polymorphism -1131T-->C: association with plasma lipids and type 2 diabetes mellitus with coronary heart disease in Chinese.

Type 2 diabetes mellitus (DM) is associated with significant abnormalities of lipoprotein metabolism and coronary heart disease (CHD). The most commonly recognized lipid abnormality in type 2 DM is hypertriglyceridemia, which is known to be an independent risk factor for CHD in diabetics. The -1131T-->C polymorphism found in the newly identified apolipoprotein A5 ( APOA5 ) gene has been found to be associated with elevated plasma triglyceride (TG) concentrations in different racial groups. In this study, DNA samples from 155 control subjects, 172 type 2 diabetics and 113 type 2 DM patients with CHD were analyzed to examine the influence of APOA5 1131T-->C polymorphism on plasma lipids and the susceptibility to CHD in type 2 diabetics. The frequency of the APOA5 -1131C allele in the DM+CHD group was significantly higher than that of control subjects (37.2% vs. 27.7%, p=0.021). The distribution of the APOA5 -1131T-->C genotypes (TT, TC and CC) was 36.3%, 53.1% and 10.6% in type 2 DM patients with CHD, and 53.6%, 37.4% and 9.0% in controls, respectively (p=0.018). The frequencies of alleles and genotypes in type 2 diabetics were not significant compared to controls. In controls, plasma TG concentrations in subjects with the TT genotype were significantly lower than in those with TC/CC (0.92, 1.28 and 1.35 mmol/L for TT, TC and CC, respectively; p = 0.003 by ANOVA). These data suggest that the APOA5 -1131T-->C polymorphism might play a role in elevated plasma TG levels in type 2 diabetic patients in the Chinese population.

Clinical study of qingluo tongbi granules in treating 63 patients with rheumatoid arthritis of the type of yin-deficiency and heat in collaterals.

The study is to observe the therapeutic effects of qingluo tongbi granules (QTG) in patients with rheumatoid arthritis (RA) and the changes of immune indexes. In this series there are 63 patients with RA of the type of yin-deficiency and heat in collaterals treated with QTG as the treated group and 55 patients of the same type treated with Tripterygium glycosides as the control group. As a result, in the treated group, the curative rate is 9.52% and markedly effective rate 38.10%, with a total effective rate of 90.48%, while the corresponding rates in the control group are 0, 20.00% and 83.64%, respectively. The curative effect in the treated group is better than that in the control group (P<0.05). Besides, no obvious adverse reactions are found in the treated group. Therefore it is concluded that as a new medicinal preparation QTG is safe and effective in the treatment of RA.

[Study on serum inhibin concentrations in women during menopausal transition].

OBJECTIVE: To investigate the changes of serum inhibin (Inh)-A, Inh-B concentrations during menopausal transition and the time relationship between changes of serum Inh-A, Inh-B and other reproductive hormone levels. METHODS: Serum Inh-A, Inh-B concentrations were measured by Serotec modified two-site enzyme immunoassay during different phases of normal menstrual cycle in 10 healthy reproductive women, any time of 10 postmenopausal women. So did the serum Inh-A on 5 - 9 days prior to next period (premenstrual phase) and Inh-B determinations on the 3rd day of menstrual cycle in 40 women of age 43 - 52 (menopausal transition group). In addition, serum follicle stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E(2)), progesterone (P) levels were also determined when necessary for the purpose of analysis. RESULTS: The fluctuation patterns of serum Inh-A, Inh-B concentrations during normal menstrual cycle were completely different. About 48% of women during menopausal transition had normal luteal function as shown by the P levels during the premenstrual phase. The only significant change in these women as compared with the young was decrease of Inh-A concentration [(24.7 +/- 13.0) ng/L Vs (42.9 +/- 12.1) ng/L, P = 0.017] in the same phase. Further significant declines of serum Inh-A levels were seen in the luteal phase defect (LPD) and anovulatory (AOV) groups [(12.4 +/- 10.2) ng/L and (5.3 +/- 3.8) ng/L, P = 0.033, P < 0.000 1 respectively], until undetectable in the postmenopausal group. The day 3 Inh-B levels tended to decrease in the normal luteal and LPD groups, became undetectable in the AOV and postmenopausal groups (P = 0.001). Day 3 Inh-B levels was significantly lower in women with day 3 FSH > or = 10 IU/L than those with < 10 IU/L [(16.2 +/- 4.0) ng/L Vs (62.0 +/- 43.8) ng/L]. The elevation of day 3 FSH, LH levels was not significant until the AOV group (P = 0.009, P = 0.027 respectively), and the drop of E(2) levels until the postmenopausal group (P < 0.001). CONCLUSIONS: It is suggested that serum reproductive hormones should be measured in women of menopausal transition in order to know the stage of menopausal transition and to guide the clinical management. The decrease of serum Inh-A levels during the premenstrual phase is the earliest change of menopausal transition, and decrement of day 3 Inh-B levels a marker of decreased ovarian reserve.

[Hyperhomocysteinemia and deep-vein thrombosis].

OBJECTIVE: To study the relationship between plasma homocysteine (Hcy) level and deep-vein thrombosis (DVT), and analyze the interaction of Hcy, folate and methylenetetrahydrofolate reductase (MTHFR) gene polymorphism in patients with DVT. METHODS: Totally 69 patients with DVT and 111 healthy controls were included in our case-control study. We determined the MTHFR C677T genotypes by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP), measured the serum folate and vitamin B12 by radioimmunoassay (RIA), and measured the plasma homocysteine level by fluorescence polarization immunoassay (FPIA). RESULTS: The frequency of the MTHFR C677T TT genotype had no significant difference between DVT group and control group (P > 0.05). The plasma Hcy level was significantly higher in DVT group than in control group (13.03 +/- 8.74 mumol/L vs 10.14 +/- 4.30 mumol/L, P < 0.05). Both serum folate and VitB12 of patients with DVT were not significantly different from those of controls. The odds radios (OR) of hyperhomocysteinemia for DVT was 2.53 (95% CI 1.08-5.92). The interaction of low folate level and TT genotype increased the risk of DVT (OR = 3.12, 95% CI 1.17-8.38). CONCLUSION: Hyperhomocysteinemia may be an independent risk factor for DVT in Han nationality, while serum folate level and MTHRF C677T genotype are not. An interaction between serum folate level and MTHFR genotype that affect the Hcy level is an important risk factor for DVT.

[Clinical evaluation of two kinds homogenous assays for determination of high-density lipoprotein cholesterol].

OBJECTIVE: To evaluate the clinical efficacy of two kinds homogenous assays for direct determination of high-density lipoprotein cholesterol (HDL-C) based on the principle of polyanion polymer/detergent (PPD method) and polyethylene glycol-modified enzyme (PEGME) method. METHODS: The two homogenous methods were compared with the precipitation method (PTA-Mg2+ method), their precision, accuracy, specificity and interference were also analyzed. RESULTS: Both homogenous HDL-C assays were precise, having a within-run CV < 3%, day-to-day CV < 3% and total CV < 4%. The HDL-C values measured by the two homogenous methods correlated well with those by PTA-Mg2+ method (X): Y = 0.9316 X + 0.1063, r = 0.9762 for PPD method (Y); and Y = 0.9106 X + 0.1368, r = 0.9894 for PEGME method (Y). The linearity studies showed the two homogenous methods to be linear up to 4.14 mmol/L. The lowest detectable concentration of the two methods was apparently 0.08 mmol/L. Recoveries of the two methods were 94.1%-106.2%. Hemoglobin did not interfere with the HDL-C results in the two homogenous methods, whereas icteric samples with total bilirubin > 200 mg/L showed discrepancies. Lipemia up to triglyceride concentration of 17.0 mmol/L did not interfere with the two homogenous HDL-C assays. CONCLUSIONS: The two new homogenous HDL-C assays meet the requirements for accuracy, precision, ease of handling with massive sample, allow full automation, and are clinically useful.