Analysis of the Amine Submetabolome Using Novel Isotope-Coded Pyrylium Salt Derivatization and LC-MS: Herbs and Cancer Tissues as Cases.

The amine submetabolome, including amino acids (AAs) and biogenic amines (BAs), is a class of small molecular compounds exhibiting important physiological activities. Here, a new pyrylium salt named 6,7-dimethoxy-3-methyl isochromenylium tetrafluoroborate ([d0]-DMMIC) with stable isotope-labeled reagents ([d3]-/[d6]-DMMIC) was designed and synthesized for amino compounds. [d0]-/[d3]-/[d6]-DMMIC-derivatized had a charged tag and formed a set of molecular ions with an increase of 3.02 m/z and the characteristic fragment ions of m/z 204.1:207.1:210.1. When DMMIC coupled with liquid chromatography-mass spectrometry (LC-MS), a systematic methodology evaluation for quantitation proved to have good linearity (R2 between 0.9904 and 0.9998), precision (interday: 2.2-21.9%; intraday: 1.0-19.7%), and accuracy (recovery: 71.8-108.8%) through the test AAs. Finally, the methods based on DMMIC and LC-MS demonstrated the advantaged application by the nontargeted screening of BAs in a common medicinal herb Senecio scandens and an analysis of metabolic differences among the amine submetabolomes between the carcinoma and paracarcinoma tissues of esophageal squamous cell carcinoma (ESCC). A total of 20 BA candidates were discovered in S. scandens as well as the finding of 13 amine metabolites might be the highest-potential differential metabolites in ESCC. The results showed the ability of DMMIC coupled with LC-MS to analyze the amine submetabolome in herbs and clinical tissues.

Effects of different drying methods on the chemical constituents of Lilium lancifolium Thunb. based on UHPLC-MS analysis and antidepressant activity of the main chemical component regaloside A.

The Lilium lancifolium Thunb. is a herb with multiple functions in both medicine and food in China, and its extracts have shown antidepressant effects. In this study, fresh bulbs of Lilium lancifolium Thunb. were processed to study the effects of different drying processes on changes in its main chemical components. We found that different drying methods can affect the chemical constituents of the herb. Among these components, Regaloside A has been found as the characteristic component. Here, Cell Counting Kit-8 assay, and Western blotting were used to evaluate the neuroprotective antidepressant effects of Regaloside A. The results showed the cell survival rate was improved, the phosphorylation levels of brain-derived neurotrophic factor, tyrosine kinase receptor B, phosphatidylinositol 3 kinase, protein kinase B, and mammalian target of rapamycin were increased after Regaloside A treatment. In general, different drying methods have a significant influence on the chemical composition of the herb, and Regaloside A may be the main chemical component of the herb. It can alleviate the damage of corticosterone in SH-SY5Y cells, and phosphatidylinositol-3-kinase/protein kinase B/mammalian target of rapamycin signaling mediated by brain-derived neurotrophic factor/tyrosine kinase receptor B may play an important role in the neuroprotective antidepressant effects of Regaloside A.

Salvianolic acid B improves airway hyperresponsiveness by inhibiting MUC5AC overproduction associated with Erk1/2/P38 signaling.

Salvianolic acid B (SalB) is one of the main water-soluble composites from Chinese medicine Dansen (Radix miltiorrhiza). It is used for clinical treatment of various diseases including cardiovascular, lung, Liver, renal and cancers. However, the effects of SalB to allergy induced airway mucin hypersecretion, inflammation and hyperresponsiveness (AHR) remains not clear. Overproduction of airway MUC5AC is a central effector of inflammation that is strongly associated with AHR in asthmatic attack. In this study, we investigated the anti-asthmatic activity and mechanism of SalB in a murine model and human epithelial cells by monitoring changes in mucin expression and secretion, airway inflammation, AHR, and signaling pathways. SalB was administered by intragastric administration (i.g) daily for a week, starting at 21 days after sensitization of ovalbumin (OVA). All examinations were performed 24h after the last antigen challenge. We found that treatments with SalB significantly inhibited increase in the tracheobronchial secretion, glycosaminoglycan levels, interleukin (IL)-13, IL-4, and IL-5 cytokines mRNA and protein expression, and decrease in mucociliary clearance in lung tissues. Histological results demonstrated that SalB attenuated OVA-induced eosinophil infiltration, airway goblet cell hyperplasia, and MUC5AC and MUC5B mRNA and protein expression in lung tissues. SalB exhibited protective effects against AHR in OVA-challenged animals. In vitro, SalB significantly inhibited IL-13-induced MUC5AC and MUC5B mRNA and protein expression in human epithelial cells. These effects were blocked by SalB by downregulating the Erk1/2 and P38 signaling pathways. Taken together, these data indicate that treatment with SalB may improve AHR by inhibiting MUC5AC overproduction.

Ultrasonography for Preoperative Diagnosis of Retroperitoneal Fibrosis.

This study aimed to assess the application of ultrasonography for preoperative diagnosis of retroperitoneal fibrosis (RPF). A total of 51 patients with clinically suspected RPF underwent transabdominal ultrasonography and ultrasound-guided histopathologic biopsy (tAU-UGHB). Retroperitoneal fibrosis was diagnosed in 35 patients; of these, 31 cases (88.6%) received a diagnosis using tAU-UGHB. The ultrasonographic images mainly showed solid, irregular isoechoic masses, which were present behind the retroperitoneum and surrounded the abdominal aorta, inferior vena cava, and ureters and had clear borders with the encapsulated structures. Compared with other retroperitoneal lesions, RPF lesions were mainly located below the initial level of the renal artery, and their internal flow signals were not rich (P < 0.05). Retroperitoneal fibrosis had characteristic ultrasonographic features, and color Doppler ultrasound-guided biopsy could be positioned accurately, safely, and efficiently; therefore, tAU-UGHB can be used as an effective modality for preoperative diagnosis of RPF.

Cryptoporus volvatus polysaccharides attenuate LPS-induced expression of pro-inflammatory factors via the TLR2 signaling pathway in human alveolar epithelial cells.

CONTEXT: Cryptoporus volvatus (Peck) Hubb grows wild in China, and its fruiting bodies have been used traditionally to treat asthma and bronchitis. OBJECTIVES: This study evaluates the anti-inflammatory effect of Cryptoporus polysaccharides (CP) extracted from fruiting bodies of C. volvatus on lipopolysaccharide (LPS)-induced pro-inflammatory factors and the signaling pathways involved in human alveolar epithelial cells. MATERIALS AND METHODS: To evaluate the effects of CP on LPS-induced pro-inflammatory factors, A549 cells were pre-incubated with CP 1, 10, and 100 µg/ml for 1 h and then stimulated with LPS 10 µg/ml for 24 h. The expression of pro-inflammatory factors monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), Toll-like receptor 2 (TLR2), and phosphorylation of ERK1/2, p38, and NF-κB p65 were measured by q-PCR, ELISA, and western blotting. RESULTS: CP decreased LPS-induced mRNA expression of MCP-1, TNF-α, and IL-1ß (IC50 = 83.3, 85.2, and 91.6 µg/ml, respectively) and their correspondent protein expression (IC50 = 88.6, 76.4, and 81.6 µg/ml, respectively). Investigation of potential mechanisms indicated that CP 100 µg/ml reduced LPS-induced expression of TLR2 mRNA (66.9%, p < 0.01) and protein (63.2%, p < 0.01) that was a result of the decreased pro-inflammatory factors. LPS induction increased the expression of TLR2 and the phosphorylation of p38 and ERK1/2, NF-kB p65 concomitantly. CP 100 µg/ml inhibited the LPS-induced phosphorylation of the signaling proteins (p < 0.05). CONCLUSIONS: This suggests that CP pretreatment down-regulates LPS-mediated inflammation in lung epithelial cells. This study further confirmed that CP is a potential anti-inflammatory drug for the treatment of airway inflammatory diseases.

[Effect of intermittent pneumatic compression on coagulation function and deep venous hemodynamics of lower limbs after rectal cancer resection].

OBJECTIVE: To investigate the effects of intermittent pneumatic compression (IPC) on coagulation function, deep venous hemodynamics and prevention of deep venous thrombosis (DVT) of lower limbs in patients after rectal cancer resection. METHODS: A total of 120 patients undergoing rectal cancer resection were randomly divided into non-IPC group (control group, n=60) and IPC group (n=60). The control group received routine treatment after resection and the IPC group received IPC based on the routine treatments. Prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), international normalized ratio (INR) and volume of D-dimer (D-D) were detected before operation and 1-, 3-, 5- and 7-day after operation. Meanwhile, blood flow velocity and caliber of external iliac vein, femoral vein and popliteal vein were examined by color Doppler ultrasound, then the average blood flow velocity and blood flow volume were calculated. RESULTS: Incidence of lower limb DVT was 13.3% (8/60) and 1.7% (1/60) in control group and IPC group respectively with significant difference (P<0.05). The differences in PT, APTT and INR were not significant (P>0.05) at 1-day after operation as compared to the preoperative level, while FIB and D-D both increased (P<0.05), all presented no significant difference among the two groups (P>0.05). PT shortened gradually (P<0.05), APTT and INR did not change significantly (P>0.05), FIB and D-D increased gradually (P<0.05), and no significant differences were found between the two groups at the same time point (all P>0.05). All the above parameters in the control group were significantly lower than those in IPC group (all P<0.05). CONCLUSIONS: IPC can improve hemodynamics indexes of deep veins of lower limb in patients after rectal cancer operation, and prevent the lower limb DVT. IPC is a safe, simple and convenient physical therapy.

Cardiovascular protection and antioxidant activity of the extracts from the mycelia of Cordyceps sinensis act partially via adenosine receptors.

Mycelia of cultured Cordyceps sinensis (CS) is one of the most common substitutes for natural CS and was approved for arrhythmia in China. However, the role of CS in ameliorating injury during ischemia-reperfusion (I/R) is still unclear. We examined effects of extracts from CS on I/R and investigated the possible mechanisms. Post-ischemic coronary perfusion pressure, ventricular function, and coronary flow were measured using the Langendorff mouse heart model. Oxidative stress of cardiac homogenates was performed using an ELISA. Our results indicate that CS affords cardioprotection possibly through enhanced adenosine receptor activation. Cardioprotection was demonstrated by reduced post-ischemic diastolic dysfunction and improved recovery of pressure development and coronary flow. Treatment with CS largely abrogates oxidative stress and damage in glucose- or pyruvate-perfused hearts. Importantly, observed reductions in oxidative stress [glutathione disulfide (GSSG)]/[GSSG + glutathione] and [malondialdehyde (MDA)]/[superoxide dismutase + MDA] ratios as well as the resultant damage upon CS treatment correlate with functional markers of post-ischemic myocardial outcome. These effects of CS were partially blocked by 8-ρ-sulfophenyltheophylline, an adenosine receptor antagonist. Our results demonstrate a suppressive role of CS in ischemic contracture. Meanwhile, the results also suggest pre-ischemic adenosine receptor activation may be involved in reducing contracture in hearts pretreated with CS.

[SUDHL-4 cell culture in vitro and establishment of mouse tumor model].

This study was designed to investigate the biological and immunological characteristics of a human diffuse large B-cell lymphoma (DLBCL) cell line SUDHL-4, and to establish a mouse model for human DLBCL. SUDHL-4 cells were cultured under different conditions. The morphology and in vitro expression of B-cell and tumor-related markers were detected by microscopy and flow cytometry respectively. To establish the transplanted tumor, the cells were injected subcutaneously into SCID mice. Tumor formation and its histomorphology were analyzed. The results showed that the expression of B cell/tumor-related markers was found on cultured SUDHL-4 cells. A stable mouse model of human DLBCL was successfully established in SCID mice by subcutaneous injection of 10(7) SUDHL-4 cells. Tumor tissue from mice exhibited similar histologic manifestation to those of human DLBCL. It is concluded that the SUDHL-4 cells represent a high consistency in immunological characteristics with human DLBCL. Transplantation of SUDHL-4 cells provides a syngeneic mouse model for the study of human DLBCL.

A novel dendrimeric peptide induces high level neutralizing antibodies against classical swine fever virus in rabbits.

The amino acid sequence (TAVSPTTLR, 829-837aa) on the glycoprotein E2 of classical swine fever virus (CSFV) is a conserved and linear neutralizing epitope. In the present study, two peptides were constructed based the core sequence of this neutralizing epitope, the dendrimeric peptide (Th-B(4)) containing four copies of B cell epitope fused to one copy of promiscuous T helper (Th) cell epitope and the peptide Th-B containing a single copy of B cell epitope fused to one copy of Th cell epitope. The dendrimeric peptide Th-B(4) elicited high titers of neutralizing antibodies as detected in an indirect ELISA, blocking ELISA and neutralization test and induced a complete protection against CSFV C strain in rabbits. The Th-B elicited low titers of neutralizing antibodies and did not induce a protection in rabbits. These results suggest that the dendrimeric peptide Th-B(4) may be a promising marker vaccine candidate against CSFV and the multimerization is a requirement for development of a peptide vaccine.

Studies on functional connections between the supraoptic nucleus and the stomach in rats.

The present study was to investigate whether there are functional connections between the hypothalamic supraoptic nucleus (SON) and the stomach, which is the case with the paraventricular nucleus. The rats were divided into four groups. Group I: the neuronal discharge was recorded extracellularly in the NTS, DMV or SON before and after cold physiological saline (4°C) was perfused into the stomach and effused from the duodenum. Group II: the rats were stimulated as for Group I and c-Fos expression in NTS, DMV and SON was examined. Group III: the control to Group II. Group IV: gastric motility was recorded continuously before and after microinjection of L: -Glu into the SON. In Group I, the discharge frequency increased in all the three nuclei, while in Group II, Fos expression in NTS, DMV and SON was, respectively, greater than that of Group III. In Group IV, microinjection of L: -Glu (5 nmol) into SON significantly inhibited gastric motility. These data suggest there are functional connections between SON and stomach.

[Clinical analysis of surgically treated cervical spondylotic myelopathy with erectile dysfunction].

OBJECTIVE: To investigate the recovery of sexual function of surgically treated male patients with cervical spondylotic myelopathy. METHODS: A prospective and a mean 16-month postoperative follow-up were conducted for 22 male patients surgically treated for cervical spondylotic myelopathy complicated by sexual dysfunction. Their neurologic scores were obtained by the Japanese Orthopedic Association (JOA) Scoring System, their sexual function assessed by the International Index of Erectile Function (IIEF), and their pre- and post-operative reflexogenic and psychogenic erection analyzed by comparison. RESULTS: Most of the patients experienced an obvious improvement in neurological function after the surgery, with a significantly higher JOA score than pre-operation ( P < 0.01). Compared with the preoperative rates of abnormal reflexogenic and psychogenic erection, 82% (18/22) and 18% (4/22) , the average IIEF score was elevated from preoperatively (9.90 +/- 2. 22) to postoperatively (20.89 +/- 3.89), with a statistically significant difference (P < 0.01). CONCLUSION: Cervical spondylotic myelopathy induces sexual as well as neurological dysfunction, mostly with abnormal psychogenic but normal reflexogenic erection. With neurological recovery, most of the patients may experience an improvement in their sexual function after surgery.

Protective effects of pravastatin in murine lipopolysaccharide-induced acute lung injury.

1. The present study was designed to determine whether pravastatin, a 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor, could attenuate acute lung injury (ALI) induced by lipopolysaccharide (LPS) in BALB/c mice. 2. Acute lung injury was induced successfully by intratracheal administraiton of LPS (3 microg/g) in BALB/c mice. Pravastatin (3, 10 and 30 mg/kg, i.p.) was administered to mice 24 h prior to and then again concomitant with LPS exposure. 3. Challenge with LPS alone produced a significant increase in lung index and the wet/dry weight ratio compared with control animals. Pulmonary microvascular leakage, as indicated by albumin content in the bronchoalveolar lavage fluid (BALF) and extravasation of Evans blue dye albumin into lung tissue, was apparently increased in LPS-exposed mice. Lipopolysaccharide exposure also produced a significant lung inflammatory response, reflected by myeloperoxidase activity and inflammatory cell counts in BALF. Furthermore, histological examination showed that mice exposed to LPS also exhibited prominent inflammatory cell infiltration and occasional alveolar haemorrhage. 4. Pravastatin (3, 10 or 30 mg/kg, i.p.) produced a significant reduction in multiple indices of LPS-induced pulmonary vascular leak and inflammatory cell infiltration into lung tissue. Elevated tumour necrosis factor (TNF)-alpha levels in lung tissue homogenates of ALI mice were significantly decreased after administration of 10 or 30 mg/kg pravastatin. 5. These findings confirm significant protection by pravastatin against LPS-induced lung vascular leak and inflammation and implicate a potential role for statins in the management of ALI. The inhibitory effect of pravastatin was associated with its effect in decreasing TNF-alpha.

Losartan attenuates bleomycin-induced pulmonary fibrosis in rats.

BACKGROUND: In addition to regulating blood pressure and body fluid homeostasis, the renin-angiotensin system is also involved in lung fibrogenesis. OBJECTIVE: To study the effect of losartan, an angiotensin II antagonist, on bleomycin-induced pulmonary fibrosis in rats and its possible mechanism. METHODS: Pulmonary fibrosis was induced in SD rats by intratracheal instillation of bleomycin (5 mg x kg(-1)). Subsequently, the rats received daily losartan (3, 9 and 27 mg x kg(-1)) or prednisone (20 mg x kg(-1)) orally. Six rats in each group were sacrificed 14 and 21 days after intratracheal instillation. Hydroxyproline, superoxide dismutase (SOD), and malondialdehyde (MDA) levels in lung tissues were determined by spectroscopy. The levels of TGF-beta1 in serum were measured by ELISA. Histological changes in the lungs were evaluated by hematoxylin-eosin stain, and scored. RESULTS: Rat body weight evidently decreased while the indices of lung and hydroxyproline contents in lung tissue were significantly increased 14 and 21 days after intratracheal bleomycin instillation. Inflammatory cell infiltration and fibrotic scores were more prominent in the model group compared to the sham group. Losartan (3, 9 and 27 mg.kg(-1), i.g.) apparently attenuated the degree of pulmonary fibrosis. Further study showed that losartan significantly increased SOD levels while it decreased MDA contents in lung homogenates. Serum TGF-beta1 levels of pulmonary fibrosis rats were also decreased by losartan. CONCLUSIONS: Losartan had an inhibitory effect on bleomycin-induced pulmonary fibrosis, and its effect may be associated with its anti-free radicals and the reduction in TGF-beta1.

Research on history of medicine in China in the last five years.

Since 1999, progress has been made, to varying degrees, in numerous areas of medical history research in China including history of TCM, history of western medicine, history of integrated Chinese and western medicine, history of traditional medicine of Chinese minorities, history of medicine of foreign countries, history of medical exchanges between China and other countries, and history of comparative medical history. Among others, the number of articles on history of diseases, history of specific medical disciplines, modern medical history, medical biographies, medical works, contemporary medical history, and history of medical culture has increased dramatically. In the field of history of diseases, the papers deal with diseases in gynecology and obstetrics, plague, lanhousha (scarlet fever), and nephritis; articles in the field of specific disciplines deal with history of acu-moxibustion, history of prescription-forms, and history of gynecology, endoscopic surgery, and evidence-based medicine. There are even distinguished papers appeared in these aspects. In the aspect of modern medical history, there are papers dealing with the developement of TCM, the introduction of western medicine into China, with some specific researches in these fields. Medical biographies include Tan Yun-xian, Quan Shao-qing, Du Chong-ming etc. Papers on medical works deal with the ancient unearthed literature lost yet spread and extant abroad, medical classics, canons on material medica, cold-pathogenic diseases, and formularies. While papers on history of medical culture discuss basically the influence of Confucianism, Taoism, Buddhism and I-discipline on Chinese medicine. During these five years, 300 original articles have been published in The Chinese journal of Medical History, with another 200 papers published in other Chinese journals. Forty monographs have been published and important ones are A General History of Chinese Medicine, Modern history of TCM, The Historical Development of Acupuncture, A General History of Tiberan Medicine. For the coming years, the stress points should be laid on the research on history of diseases, history of specific medical disciplines, and some specific academic topics. In addition, the weak points in the research fields of theoretical problems in studies of medical history, history of traditional medicine of Chinese minorities, medical history of communication between China and foreign countries, comparative study on medical history, and the study on history of medical also be strengthened continually. In the five years since 1999, progress has been made, more or less, in the research works in medical history in China, including history of TCM, history of western medicine, history of integrated Chinese and western medicine, history of traditional medicine of Chinese minorities, history of medicine of foreign countries, history of medical exchanges between China and other countries, and history of comparative medical history. And the number of articles on history of diseases, history of specific medical disciplines, modern medical history, medical biographies, medical works, contemporary medical history and history of medical culture has increased dramatically. The research work in ancient medical history is still progressing forcefully.

[Liver transplantation for advanced primary hepatocellular carcinoma].

OBJECTIVE: To investigate the value of liver transplantation for late hepatocellular carcinoma. METHODS: Thirty-six patients were treated by liver transplantation from August 2000 to February 2002, of which 15 patients had had advanced hepatocellular carcinoma and thirteen of these 15 patients were evaluated for results. RESULTS: The 1-year survival rate was 86% (6/7). Only one patient died of recurrence within 6 months. The tumor-free survival was 5 to 19 months. Till February 2002, two patients have survived for 10 months and 19 months with recurrence. CONCLUSION: In our country, if the patients can afford liver transplantation, advanced hepatocellular carcinoma without extrahepatic metastasis is still indicated for liver transplantation, since some patients may survive relatively long.