In order to affirm the cardioactive components in Fuzi, we identified a group of aminoalcohol- diterpenoid alkaloids in Fuzi using ultra high-performance liquid chromatography coupled with electrospray ionization mass spectrometer (UPLC-ESI-MS) method. Among a total of forty-one isolated ingredients, thirteen major aminoalcohol-diterpenoid alkaloids were identified by comparing their retention times and MS spectra with those of the reference substances. Moreover, Fuzi samples from different places of origin and with different processing methods were examined and their components displayed a pattern of high similarity, though the relative abundance varies probably due to their different processing methods. Furthermore, the cardiac effect of each identified alkaloid was individually evaluated using the isolated bullfrog heart perfusion experiment. Among the thirteen aminoalcohol diterpenoid alkaloids tested, six of them significantly enhanced the amplitude rates. Taken together, we affirm that the cardioactive components in Fuzi are aminoalcohol-diterpenoid alkaloids, shedding light on future studies of the mechanisms and development of these cardioactive compounds.
Aconitum ; chemistry ; Alkaloids ; chemistry ; Amino Alcohols ; chemistry ; Animals ; Cardiotonic Agents ; chemistry ; Chromatography, High Pressure Liquid ; Drugs, Chinese Herbal ; chemistry ; Heart ; drug effects ; In Vitro Techniques ; Plant Extracts ; chemistry ; Rana catesbeiana ; Spectrometry, Mass, Electrospray Ionization

Objective To observe the different effects of iodine excess on inducing two strain mice thyroiditis. Methods NOD and Balb/c mice, each having 14 mice, were divided into NaI and control group. The mice were given 0.05% NaI water for 8 weeks in NaI group. RIA and ELISA were used respectively to detect TT4, TgAb, TPOAb and TSH level in serum. Morphology changes of thyroid and apoptosis of thyrocytes stained by immunohistochemistry were observed under light microscope. Lymphocytic proliferation of cervical lymph node and spleen to responding to Tg were detected by MTr method. Results After intake of iodine water for 8 weeks, NOD and Balb/c mice showed relative quality of thyroid in Nal group[(104.83±14.52), (155.79±20.77)mg/kg]obviously increased compared with control group[(71.80±20.42), (105.15±21.98)mg/kg, t values:-3.293,-4.429, all P< 0.01)], enlarged follicular lumen with colloid accumulation were observed in thyroid. Serum level of TT4 in Nal group [(29.52±4.42), (19.53± 2.35)nmol/L]to control group[33.40±5.38), (23.47±6.22)nmol/L]of NOD and Balb/c mice showed a decreasing tendency(t values: 1.374,1.567, all P > 0.05). TSH of Nal group showed an increasing tendency in Balb/c mice[(4.14±1.71)μg/L, compared with control [(3.55±1.41)μg/L, t values:-0.705, P > 0.05]and obviously increased in NOD mice [(6.98±0.66)μg/L, compared with control[(555±056)μg/L, t values:-3.562, P< 0.01], but no change of TgAb and TPOAb level in Nal group(1281,1364 cpm, 2.50×103, 0.14×103U/L were observed, compared with control(1297,1220 cpm, 3.17×103,0.03×103 U/L; Zvalues:-0.081,-0.703, -0.244,-1.293, all P > 0.05). In NOD mice NaI group, apoptosis of thyrocytes was more intense than Balb/c mice, obvious infiltration of lymphoeytes, disorganization and focus fibrosis was seen in thyroid. The cell amount of NaI group increased in NOD mice lymph node and spleen cells[(1.100±0.014), (1.076±0.033)]were more than that in the control group [(0.993±0.011), (1.005±0.003), t value:-11.672,-4.314, P < 0.01). Conclusions Iodine leads to enlargement of thyroid and malfunction of thyroid in Balb/c mice. Besides, NOD mice have generate inflammatory reaction in thyroid and produced sensitized lymphocytes to Tg. Iodine excess can induce NOD mice to occur autoimmune thyroiditis.

Objective: To observe the clinical efficacy of spleen-invigorating and qi-benefiting pediatric massage (tuina) for treating recurrent respiratory tract infection in children with cerebral palsy due to qi deficiency of spleen and lung. Methods: A total of 70 children with cerebral palsy who suffered from recurrent respiratory tract infection due to qi deficiency of spleen and lung were randomized into an observation group and a control group by the random number table method, with 35 cases in each group. Both groups were treated with conventional rehabilitation training, while the observation group was given additional spleen-invigorating and qi-benefiting pediatric massage, and the control group additionally took oral Yu Ping Feng granule. The clinical efficacy was evaluated by the total effective rate, traditional Chinese medicine (TCM) symptom score, and serum levels of immunoglobulin (Ig) A, IgM and IgG. Results: The difference in total effective rate between the two groups after treatment was statistically significant (P<0.05). After treatment, the TCM symptom scores in both groups decreased to varying degrees than those before treatment, and the intra-group differences were statistically significant (all P<0.01); the differences in the scores of various TCM symptoms between the two groups were statistically significant (all P<0.05). After treatment, the levels of IgA, IgM and IgG of the children in both groups increased to varying degrees, and the intra-group differences were statistically significant (all P<0.05). The between-group differences in the IgA, IgM and IgG levels after treatment were statistically significant (all P<0.05). Conclusion: Spleen-invigorating and qi-benefiting pediatric massage can effectively treat recurrent respiratory tract infection due to qi deficiency of spleen and lung in children with cerebral palsy, relieve the clinical symptoms and improve immune function, and thus is worthy of clinical promotion and application.

OBJECTIVETo analyze the relationship between the pulse pressure (PP) and atherosclerotic renal artery stenosis (ARAS) in middle-age and eldery paients with hyperensio, so as to provide the basis for the diagnosis and treatment of ARAS.

METHODSWe tetro spectively analyzed the data of 257 patients with hypertension undergoing renal arteriography at General Hospital of PLA between 2009 and 2014. Their mean age was (64.09 +/- 7.81) years. According the results of arteriography, they were divided into two groups: ARAS group (n = 145), and non-ARAS group (n = 112). Their PP and multinomial clinical indexes with ARAS were statistically analyzed.

RESULTSSingle factor correlation and Logisitic analysis showed that age, PP, serum creatinine (SCr) levels, and new onset of hypertension after 50 years of age were strongly positively related to ARAS. The age and PP and SCr was a team of continuous variable.

CONCLUSIONPP was the first new risk factor of ARAS in middile-aged and elderly patients with hypertension. It is suggested that PP may be a early predictive indicator and a new therapeutic target for ARAS.

Angiography ; Arteriosclerosis ; physiopathology ; Blood Pressure ; Humans ; Hypertension ; Middle Aged ; Renal Artery ; physiopathology ; Renal Artery Obstruction ; Risk Factors

OBJECTIVETo explore the effect of Xinfeng Capsule (XC) on lipoprotein metabolism of rheumatoid arthritis (RA) patients.

METHODSTotally 180 RA patients were assigned to the experimental group and the control group by random digit table, 90 in each group. Patients in the experimental group took XC (three pills each time, three times daily), while those in the control group took Methotrexate Tablet (four tablets each time, once per week). One month consisted of one therapeutic course and all patients were treated for two therapeutic courses. A healthy control group consisting of 60 patients was also set up. Changes of lipoprotein indices, clinical efficacy, lipid metabolism, joint symptoms and signs, activity indicators were observed, and correlation analyses were performed.

RESULTSCompared with the healthy control group, expression levels of prealbumin (PA), globulin (GLO), high-density lipoprotein (HDL), apolipoprotein Al (Apo-A1) were lowered in RA patients (P <0. 05, P <0. 01). Correlation analyses showed that PA was negatively correlated with joint tenderness, morning stiffness time, disease activity score (DAS-28), C-reactive protein (CRP), interleukin (IL)-6, respectively. Total protein (TP) was negatively correlated with joint tenderness. GLO was negatively correlated with joint tenderness and DAS-28. HDL was negatively correlated with erythrocyte sedimentation rate (ESR) and endothelin (ET)-1. Apo-Al was negatively correlated with joint pain; Apo-B was negatively correlated with CRP; LDL was negatively correlated with morning stiffness time (P <0. 05, P <0. 01). Compared with before treatment, expression levels of PA, HDL, Apo-A1 , Apo-B, and serum IL-10 contents increased, and expression levels of ESR, CRP, IL-6, ET-1 , joint pain, joint swelling, morning stiffness time, and DAS-28 decreased in the experimental group (P <0. 05, P <0. 01). PA increased more after treatment than before treatment in the control group (P <0. 01). There was statistical difference in joint symptoms (except joint tenderness) and activity indices (except ET-1) in the control group (P <0. 05, P <0. 01). Compared with the control group after treatment, PA and HDL increased, ET-1 and duration of morning stiffness decreased in the experimental group (all P <0. 05).

CONCLUSIONSLipoprotein metabolic disorder exists in RA patients, and it is associated with disease activity. XC could obviously improve lipoprotein metabolism and joint symptoms.

Arthritis, Rheumatoid ; drug therapy ; metabolism ; Blood Sedimentation ; C-Reactive Protein ; Capsules ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Humans ; Interleukin-10 ; Interleukin-6 ; Lipoproteins ; Lipoproteins, HDL ; metabolism ; Methotrexate

Nerve growth factor is one of the most important factors playing an important role in regulating the growth, development and survival of the neuron. The purified NGF from human placenta has been reported, the tissue from which can be isolated the NGF is very limited. It is important for basic research and clinic application to expression hNGF by genetic engineering. By polymerase chain reaction,gene fragment encoding the mature part of ?-NGF was amplified using the DNA of human placenta as template. The fragment was sequenced and inserted into expression vector pET-15b, and the recombinant expression vector pET15b-NGF was transformed into E.coli BL21(DE3)pLysS. After inducing with IPTG the NGF was higher expressed up to 25% of the total cell proteins. The expression product was purified with metal chelate affinity chromatography on Ni-IDA agarose under denaturing condition. The purity of rNGF was higher than 90% and yield of rNGF was 4.56mg/L expressing bacteria. SDS-PAGE revealed the NGF expression product had a Mr 16kDa. Western-blot displayed the recombinant product had strong immunological activity with rabbit anti-human ?-NGF polyclonic antibodies. The expression products were dealed with solubilizing inclusion bodies and refolding protein. The test of nerve fiber growth of chicken embryo DRG neurons displayed rNGF had biological activity.

Pseudorabies virus (PRV) is a swine herpesvirus of the Alphaherpesvirinae subfamily and a pathogen of swine resulting in devastating disease and economic losses worldwide. Cre/loxP site-specific system has the character of site specific, time specific, tissue specific and high efficiency in recombination, which makes this system universal in vivo and in vitro recombination of bacteria, fungus, plants, insects and mammals. A recombinant PRV which contain a loxP site in TK locus by using Cre/LoxP recombinant system was construsted. A pair of primers were synthesized according to the pEGFP-C1 sequence published on GenBank, and were used to amplify the EGFP gene expression cassette with two loxP sites flanking each side. This target gene was cloned into pSKLR, the resulting transfer vector pSKLR-GFP-loxP was then cotransfected into 293T cells with PRV SH strain genomic DNA. The recombinant virus rPRV1 was selected and purified in TK-143 cells by choosing fluorescent expressing plaques. Cre expression vector pOG231 was cotransfected into 293T cells with rPRV1 genomic DNA. The second recombinant virus rPRV2 was obtained, which contains only one loxP site in TK locus. Sequencing results of rPRV2 TK gene indicated that 34bp loxP site was inserted into rPRV2 genome and there were 270bp deletion in TK gene. PCR amplifying different generations of rPRV2 TK gene showed that the mutant was stable when passages in RK-13 cells. TCID_ 50 assay indicated that rPRV2 grows well on RK-13 cells. The LD_ 50 test results on BALB/C mice suggested that the virulence of rPRV2 was reduced. As a conclusion, the report gene GFP expression cassette was removed successfully from rPRV1 genome and only one LoxP site was leaved in rPRV2 genome by using Cre/LoxP recombinant system.

Objective To investigate the clinical effects of the modified Kock method of bladder re- construction with ileum,and to provide the objective basis for wide application of this technique.Methods A total of 51 patients with bladder cancer(T_2N_0M_0 stage tumor in 37 cases,T_3N_0M_0 stage tumor in 14; and pathologic gradeⅡ-Ⅲin all)were included.After radical cystectomy,modified Kock reconstruction of bladder with ileum was performed in them.The procedure consisted of preparing the segment of the ileum for reconstruction of the reservoir,anastomosing the low part of the neobladder with the urethra and regaining the original urinary conduit.Results The mean operative time was 6.5h(range,5.5-8.5h);mean in- traoperative blood loss was 650ml(range,300-1200ml),with blood transfusion in 31 cases.Perioperative complications included stress ulcer in 6 cases and urinary leakage in 1.The other 44 cases had no severe complications.Four cases died of cancer metastasis at 6-18 months after operation.During a follow-up of 8 -32 months,the other 47 cases recovered well and have been alive till now.Two cases had ureteral urine reflux with no impairment of renal function.The daytime urinary continent rate was 100%;and nocturnal uri- nary incontinence occurred in 6 cases.Conclusions The modified Koek reconstruction of bladder with il- eum can improve the patients'quality of life with fewer complications,therefore is a better treatment choice for infiltrative bladder cancer after radical eystectomy.

Objective Imatinib mesylate (IM) is the most active agent in treating chronic myeloid leukemia (CML). 5-Aza-2-deoxycytidine (DAC) is a cytosine analogue that inhibits DNA methylation and the activity in myeloid leukemia. Therefore,we investigated combining these two drugs in human leukemia cell line K562. Methods The effects of IM and DAC was examined in K562 cells including cell viability using MTT method,cell cycle phase and cell death using flow cytometric (FCM),and the expression of bcr-abl mRNA by RT-PCR. Results Both DAC and IM resulted in time and concentration-dependent induction of cell death. DAC and IM in combination produced a greater inhibition of growth against K562 cells (F =43.947,165.580,321.193,296.101,P<0.05). The main effect and interaction between two drugs was statistically significant (F = 202.759,168.457,417.538,P <0.001) after 24 h,48 h,72 h and a greater reduction in expression of bcr-abl mRNA than either agent alone. The difference was statistically significant (F =71.981,P <0.05). The number of G1 phase cells were increased significantly when induced by single agent. 48 h incubation with IM 0.2 μmol/L alone or combined with DAC 4 μmol/L showed 6.7 %,8.4 % pre-apoptosis cells,respectively. After incubation for 48 h with DAC 4 μmol/L,the expression of mRNA were decreased by 14 %,IM 0.2 μmol/L showed 40 % reduction,and combination group were significantly depressed for the mRNA expression by 60 %. Conclusion The combination of DAC and IM showed synergistic effects on cell death in K562 cells. These data suggested that DAC used in combination with IM has clinical potential in the treatment of chronic myeloid leukemia.

Tuberculosis (TB) is a global burden disease and is being resurrected as a major worldwide public health problem after two decades of neglect.In 1993,the World Health Organization (WHO) declared that TB had been a global emergency because of the scale of the epidemic and the urgent need to improve global tuberculosis control.China is one of the countries with the largest population,and also the top of the 22 TB high-burden countries in the world.In the United States,the longstanding downward trend in TB incidence was interrupted in the mid-to-late 1980s,where the national TB incidence peaked in 1992.Sub-Saharan Africa is one of the three regions to dominate the worldwide distribution of notified TB cases.Of the 15 countries with the highest estimated tuberculosis incidence rates in the world,13 are in sub-Saharan Africa,where HIV is the most important single predictor of tuberculosis incidence.The largest share of the global burden of HIV-related tuberculosis falls on this region.The reasons for the persisting global tuberculosis burden include increased poverty in some regions,immigration from countries with high tuberculosis prevalence,the impact of HIV,and most importantly,the failure to maintain the necessary public health infrastructure under the mistaken belief that tuberculosis was a problem of the past.Relying on currently available methods of diagnosis and treatment,the DOT strategy promoted by the WHO for global tuberculosis control is effective,affordable,and adaptable in different settings.