Experimental study on anaphylaxis of qingkailing injection and its components on Beagle dogs.

OBJECTIVE: To study the anaphylaxis of Qingkailing injection (QI) and its components. METHODS: Experimental anaphylactoid and allergic reactions were used. Changes in the behaviors of Beagles and serum levels of histamine, immunoglobulin (Ig)E, IgG, IgM, eosinophil cationic protein (ECP), and interleukin (IL)-4, as well as blood pressure, after injecting QI and its components on the forelimb veins of Beagles were observed. RESULTS: According to comprehensive determination of abnormal behavior scores and changes in serum levels of histamine, IgE, IgG, IgM, ECP, and IL-4, as well as in blood pressure, radix isatidis and hyodeoxycholic acid caused anaphylactoid reactions, and honeysuckle, radix isatidis, hydrolysate, cholic acid and Gardenia jasminoides caused allergic reactions. The anaphylaxis of QI involved anaphylactoid and allergic reactions. CONCLUSION: QI and its components need to be refined further to improve the safety, efficacy, and quality of its use in clinical settings.

Seasonal variations in metabolite profiling of the fruits of Ligustrum lucidum Ait.

The metabolite profiling of fruits of the herb Ligustrum lucidum Ait collected during different months has been performed using ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC/QTOFMS) and multivariate statistical analysis techniques. The markers such as oleuropein acid, neonuezhenide, specnuezhenide, oleuropein and ligustrosidic acid accountable for such variations were identified through the loadings plot of principal component analysis (PCA), and the tentative identification of the markers is completed by comparing the mass spectra and retention times with those of reference compounds and/or tentatively assigned by matching empirical molecular formulae and MS/MS data with those of the known compounds published. Furthermore, one of the chemical markers, such as specnuezhenide, which is water-soluble, biologically active and also the predominant compound in this crude drug, was quantified by ultra-performance liquid chromatography coupled with a tunable UV detector (UPLC-TUV). The developed UPLC method provides good linearity (r(2)=0.9991), repeatability (RSD=2.96%), intra- and inter-day precisions (RSD=0.21%, 0.96%), with accuracies of 99.18-100.26% and a recovery of specnuezhenide of 97.57%. The fruits of L. lucidum Ait collected from August to December were tested. The results clearly show that the fruits of L. lucidum Ait harvested in October have the highest yields of specnuezhenide. It is also noted that the variations of content of specnuezhenide obtained by both methods have a strong correlation. This suggests that the newly proposed strategy is a reliable and simple method for the rapid discrimination of subtle variations, within the same plant species or strains, due to different seasonal collection times.

To compare the efficacy of two kinds of Zhizhu pills in the treatment of functional dyspepsia of spleen-deficiency and qi-stagnation syndrome: a randomized group sequential comparative trial.

BACKGROUND: In Traditional Chinese Medicine (TCM) theory, functional dyspepsia (FD) can be divided into different syndromes according to different clinical symptoms and signs, and the most common one is spleen-deficiency and qi-stagnation syndrome that can be treated by Chinese traditional patent medicine--two kinds of Zhizhu pills, between which the primary difference in ingredients is that one contains immature orange fruit of Citrus aurantium L.(IFCA) and the other contains that of Citrus sinensis Osbeck (IFCS). The trial's objective was to compare the efficacy of two kinds of Zhizhu pills on symptom changes in patients with FD of spleen-deficiency and qi-stagnation syndrome. METHODS: A randomized, group sequential, double-blinded, multicenter trial was conducted in patients with FD of spleen-deficiency and qi-stagnation syndrome at 3 hospitals in Beijing between June 2003 and May 2005. Participants were randomly allocated into two groups (IFCA group and IFCS group) in a 1:1 ratio, and respectively took one of the two kinds of Zhizhu pills orally, 6 g each time, 3 times a day, for 4 weeks. Statistical analysis was performed with use of a group sequential method, the triangular test (TT). RESULTS: A total of 163 patients were randomized, and 3 patients were excluded from analysis because of early dropouts, leaving 160 patients (IFCA group: n = 82; IFCS group: n = 78) for statistical analysis. Three interim analyses were done after 62, 116, and 160 patients had completed their 4-week treatment, respectively. At the third interim analysis, the sample path crossed the upper boundary and the trial was stopped, the cure-markedly effective rates were 45% for IFCS group and 67% for IFCA group, respectively, the one-sided p-value was 0.0036, the median unbiased estimate of the odds ratio (OR) for the benefit of IFCA relative to IFCS was 2.91 with 95%CI: 1.40 to 6.06.No adverse events were observed in the two groups. CONCLUSIONS: Zhizhu pills containing IFCA was superior to Zhizhu pills containing IFCS in the treatment of FD of spleen-deficiency and qi-stagnation syndrome. The application of group sequential analysis in clinical trials of TCM may offer some financial and ethical benefits. TRIAL REGISTRATION: Chinese Clinical Trial Registry (ChiCTR): ChiCTR-TRC-00000485.

Role of TL1A in the pathogenesis of rheumatoid arthritis.

TNF-like ligand 1A (TL1A), a member of the TNF superfamily, is the ligand of DR3 and DcR3. Several types of cells, such as endothelial cells, monocytes/macrophages, dendritic cells, and CD4 and CD8 T cells, are capable of producing this cytokine. In present study, we demonstrated that TL1A aggravated collagen-induced arthritis in mice. It increased collagen-induced arthritis penetrance and clinical scores as well as the severity of the pathological findings. TL1A administration led to the occurrence of multiple enlarged germinal centers in the spleen, and it boosted serum anti-collagen Ab titers in vivo. In vitro, TL1A augmented TNF-alpha production by T cells upon TCR ligation, and it greatly enhanced Th17 differentiation and IL-17 production. We further showed that human rheumatoid arthritis (RA) synovial fluids had elevated TL1A titers, and human chrondrocytes and synovial fibroblasts were capable of secreting TL1A upon TNF-alpha or IL-1beta stimulation. Taken together, these data suggest that TL1A secretion in lymphoid organs might contribute to RA initiation by promoting autoantibody production, and TL1A secretion stimulated by inflammatory cytokines in RA joints might be a part of a vicious circle that aggravates RA pathogenesis.

[Experimental model of histamine-induced anaphylactoid reaction on beagle dogs].

OBJECTIVE: To establish animal model of histamine-induced anaphylactoid reactions on Beagle dogs through intravenous injection of histamine phosphate injection. METHOD: Anaphylactoid reactions was determined according to the changes of praxiology and blood pressure of Beagle dogs after one intravenous injection of histamine phosphate injection. RESULT: It showed that typical anaphylactoid reactions be caused by histamine phosphate injection on Beagle dogs, and the response of the high-dose group was more obvious than that of the low-dose group. CONCLUSION: Intravenous injection of histamine phosphate injection could simulate Beagle dogs to bring typical clinical anaphylactoid reaction, that could be used as a sensitive animal model to evaluate anaphylactoid reaction of traditional Chinese medicine injections (TCMIs) in the pre-clinical experiments of both TCMIs and their composition.

[Study on allergenicity of chlorogenic acid in Qingkailing injection].

OBJECTIVE: To observe the allergenicity of chlorogenic acid (CA) in Qingkailing injection. METHOD: CA was administrated to Beagle dogs through intravenous injection, and on experimental allergic sensitization of guinea pigs, it was through intraperitoneal and intravenous injection. The behavioral changes of Beagle dogs and guinea pigs were observed, and changes of the content of histamine, IgE, IgG, IgM, ECP and IL-4 in blood were detected. Then the allergenicity of CA was determined by experimental anaphylactoid and allergic methods. RESULT: There were no typical behavioral changes and increasement of the content of histamine, IgE, IgG, IgM, ECP and IL-4 in blood. CONCLUSION: CA can not provoke anaphylactoid and allergic reactions.

[Pharmacokinetics of three alkaloids in Huanglianjiedu decoction in rat serum by LC-MS-MS].

OBJECTIVE: To develop a sensitive and reliable high-performance liquid chromatography-tandem mass spectrometry (LC-MS-MS) method for simultaneous determination of jatrorrhizine, palmatine and berberine in rat serum, and study the pharmacokinetics of the three alkaloids in rat serum after an intragastrical (ig) administration of Huanglianjiedu decoction to rat. METHOD: The optimal ionization and fragmentation conditions, as well as HPLC ones, to detect jatrorrhozine, palmatine and berberine were developed and validated. After the Huanglianjiedu decoction were administered to rats through ig route, LC-MS-MS method has been applied to the pharmacokinetic study of the three alkaloids in rat serum. DAS procedure was used to process concentration-time data. RESULT: The detection was performed by MRM mode via electrospray ionization (ESI) source operating in the positive ionization mode. The precursor-to-product ion transitions were at m/z 338-322 for jatrorrhozine, m/z 351.9-308 for palmatine, m/z 336-319.8 for berberine and m/z 356.1-192.1 for tetrahydropalmatine (IS) respectively. The method was linear over the concentration range of 0.2-25 microg x L(-1) for jatrorrhozine, 0.4-50 microg x L(-1) for palmatine and berberine. The method was validated according to the requirements. The pharmacokinetic process of the three alkaloids after oral administration of Huanglianjiedu decoction was fitted to be a one-compartment model. CONCLUSION: The fully validated LC-MS-MS method has been successfully applied to the pharmacokinetic study of the three alkaloids in rat serum after oral administration of Huanglianjiedu decoction.

Simultaneous determination of seven ginsenosides in Du Shen Tang decoction by rapid resolution liquid chromatography (RRLC) coupled with tandem mass spectrometry.

A rapid quantification of seven ginsenosides in Du Shen Tang decoction, the decoction of the root of Panax ginseng was investigated by rapid resolution liquid chromatography coupled with tandem mass spectrometry (RRLC-MS/MS). All analytes including seven major constituents, namely ginsenoside-Re, Rg1, Rb1, Rc, Rb2, Rd and Rf were analysed simultaneously in short time and provide good linearity (r>0.9976), repeatability (RSD<5.67%), intra- and inter-day precisions (RSD<5.06%) with accuracies (94.65-105.31%) and recovery (93.76-106.82%) by using optimised methods. In addition, the content of seven ginsenosides in the decoction prepared by the root of 2-year-old, 4-year-old and 6-year-old Panax ginseng was also analysed. The results showed the concentration of ginsenosides detected in Du Shen Tang decoction was affected by the difference in collected times of the root of Panax ginseng. The established method could be used as a rapid and reliable approach for assessment of the quantity of Du Shen Tang decoction.

TRAF6 activation in multiple myeloma: a potential therapeutic target.

Multiple myeloma (MM) is an incurable B-lymphocyte malignancy. New therapeutic options have become available during the past several years; however nearly all patients acquire resistance to currently available therapeutic agents. Mechanisms contributing to the pathogenesis and chemoresistance of MM include genetic abnormalities, chromosomal translocations, gene mutations, the interaction between MM cells and the bone marrow microenvironment, and defects in the apoptotic signaling pathways. Survival signaling pathways associated with the pathogenesis of MM and bone marrow stromal cells play crucial roles in promoting growth, survival, adhesion, immortalization, angiogenesis, and drug resistance. The receptor activator of nuclear factor-kappa B/receptor activator of nuclear factor-kappa B ligand/tumor necrosis factor receptor-associated factor (RANK/RANKL-TRAF6) signal pathway mediates osteolytic bone lesions through the activation of the NF-κB and Janus kinase/signal transducer and activator of transcription (JNK) pathways in osteoclast precursor cells and thus contributes to the main clinical manifestations of bone disease. TRAF6 has also been identified as a ligase for Akt ubiquitination and membrane recruitment and its phosphorylation on growth factor stimulation. The inhibition of TRAF6 by silencing RNA or by decoy peptides decreases MM tumor cell proliferation and increases apoptosis as well as bone resorption. Some proteasome inhibitors and benzoxadiazole derivatives showed inhibitory effects on the activity and function of TRAF6. Overall, we propose that TRAF6 may be considered as a potential therapeutic target for the treatment of MM.

Antiarrhythmic effect of acupuncture pretreatment in rats subjected to simulative global ischemia and reperfusion--involvement of adenylate cyclase, protein kinase A, and L-type Ca2+ channel.

Our previous study showed that electro-acupuncture (EA) pretreatment protects the heart from injury of ischemia. The present study explored further whether adenylate cyclase (AC), protein kinase A (PKA), and L-type Ca(2+) channel, the beta(1)-AR signaling components modulating intracellular Ca(2+) ([Ca(2+)](i)), are involved in the mediation of the antiarrhythmic effect of EA pretreatment in the rats from which the hearts were subsequently isolated and subjected to simulative global ischemia and reperfusion (SGIR). SGIR was performed by perfusing the isolated heart at a low flow followed by normal perfusion. Adult rats were randomized into four groups, namely, normal control (NC), SGIR, EA, and NC plus EA (NCEA) groups. The rats in the EA and NCEA groups were given EA pretreatment at bilateral Neiguan points (PC6) for 30 min once a day in 3 consecutive days before the hearts were isolated and perfused. The arrhythmia score and the response of [Ca(2+)](i) to the activators of AC, PKA, and L-type Ca(2+) channel in single ventricular myocyte isolated from the hearts subjected to SGIR were compared among the groups. The results showed that the arrhythmia score was significantly higher in the SGIR group as compared with the NC and NCEA groups. The SGIR-enhanced arrhythmia score was significantly attenuated in the EA group. More interesting, EA pretreatment also attenuated the SGIR-enhanced response of [Ca(2+)](i) to the activators of AC, PKA, and the L-type Ca(2+) channel in the myocytes isolated from the hearts subjected to SGIR. In conclusion, EA pretreatment can produce an antiarrhythmic effect in the rat of SGIR. AC, PKA and the L-type Ca(2+) channel are involved in the mediation of the antiarrhythmic effect of EA pretreatment.