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The aim of this study was to observe the electrophysiological characteristics of the isolated rat muscle spindle. The muscle spindle was isolated from rat soleus and the afferent discharge of the isolated muscle spindle was recorded by air-gap technique. In the basic physiological salt solution, the spontaneous impulses of muscle spindle were at a lower level with irregular intervals. The mean frequency of afferents was (51.78 ± 25.63) impulses/1 000 s (n = 13). The muscle spindle afferents were significantly increased and maintained over time by the addition of certain amino acids during the observation. The number of the action potential recorded per 1 000 s was 200-1 000 [mean: (687.62 ± 312.56) impulses/1 000 s, n = 17]. In addition to the typical propagated action potential, a large number of abortive spikes were observed. The results indicate that the activities of isolated muscle spindles in rats can be well maintained by the addition of certain amino acids. The results initially establish and provide the possibility for further research conducted in isolated rat muscle spindles.

Objective: To study the diagnostic value of anti-cyclic citrullinated peptide antibody (anti-CCP) and magnetic resonance imaging (MRI) of metacarpophalangeal joints (MCP) and wrist in early rheumatoid arthritis (RA)., Methods: MRI of MCP and wrist joint, laboratory indices of anti-CCP and rheumatic factor (RF) were performed and recorded in the 94 early-stage RA patients, 24 non-typical monoarthritis and 35 other arthritis. The MRI findings and OMERACT (outcome measures in rheumatoid arthritis clinical trials) score were analyzed in comparison with their clinical and laboratory indices., Results: The sensitivity of anti-CCP, synovitis, bone erosion and bone erosion was 55.3%, 100%, 25.5% and 88.3% respectively in early-stage RA patients. The specificity was 88.6%, 71.4%, 94.3% and 65.7% respectively. There was significant difference between early-stage RA group and other arthritis group (P < 0.05). Bone erosion was found to be the most specific among MRI findings. And bone erosion of wrist joint had a positive correlation with anti-CCP. MRI was a more efficient supplemental modality in diagnosing early-stage RA as compared with conventional radiological films. Among 94 early-stage RA, their MRI findings and OMERACT scores of wrist joint appeared more obvious., Conclusion: Anti-CCP has a better specificity for early-stage RA than RF. But MRI may visualize the disorder of RA earlier while it is often missed by radiological films. Both have prominent diagnostic values in early RA patients. Bone erosion of wrist joint has a positive correlation with anti-CCP. MRI may help to differentiate those RA patients with negative anti-CCP.

In an attempt to find new types of anti-hypoxic agents from herbs, we identified 5-hydroxymethyl-2-furfural (5-HMF) as a natural agent that fulfills the criterion. 5-HMF, the final product of carbohydrate metabolism, has favorable biological effects such as anti-oxidant activity and inhibiting sickling of red blood cells. The role of 5-HMF in hypoxia, however, is not yet. Our pilot results showed that pretreatment with 5-HMF markedly increased both the survival time and the survival rate of mice under hypoxic stress. The present study was aimed to further investigate the protective role of 5-HMF and the underlying mechanisms in hypoxic injury using ECV304 cells as an in vitro model. ECV304 cells pretreated with or without 5-HMF for 1 h were exposed to hypoxic condition (0.3% O(2)) for 24 h and then cell apoptosis, necrosis, the changes of mitochondrial membrane potential (MMP) and the expressions of phosphorylation- extracellular signal-regulated kinase (p-ERK) were investigated. Pretreatment with 5-HMF markedly attenuated hypoxia-induced cell necrosis and apoptosis at late stage (p < 0.01). Furthermore, pretreatment with 5-HMF rescued both the decline of the MMP and the increase of p-ERK protein under hypoxia. In a word, these results indicated that 5-HMF had protective effects against hypoxic injury in ECV304 cells, and its effects on MMP and p-ERK may be involved in the mechanism.

Objective: To study the learning and memory ability, expressions of hippocampal N-methyl D-aspartate (NMDA) subunit NR2A and NR2B, and EphB2 receptor in fatigue rats, and to observe effects of Sini Powder, Shenghui Decoction, and Sihui Mixture on them., Methods: The central nervous system fatigue model was duplicated by paradoxical sleep deprivation for 168 h using multiple platform method. Experimental rats were randomly divided into the normal control group, the model group, the Sini Powder group, and the Shenghui Decoction group, ten in each. Corresponding medicines and distilled water were given to them by gastrogavage at 6, 30, 54, 78, 102, 126, and 150 h after sleep deprivation. Changes of the learning and memory ability were observed using Y maze. mRNA expressions of NMDA subunit NR2A and NR2B, and EphB2 receptor in fatigue rats were quantitatively analyzed using Real-time PCR., Results: Compared with the normal control group, the Y maze correct percentage in the model group obviously decreased (P<0.05), mRNA expressions of NR2B and EphB2 obviously decreased (P<0.901), with no obvious change in NR2A. Compared with the model group, Sihui Mixture could obviously improve Y maze results and mRNA expressions of NR2A and NR2B, and EphB2 (P<0 01). No statistical difference was found between the Sini Powder group and the Shenghui Decoction group. Compared with the Sini Powder group, mRNA expressions of EphB2 obviously increased in the Sihui Mixture group (P<0 01). mRNA expression of NR2A could be more obviously increased in the Shenghui Decoction group than in the model group (P <0 01)., Conclusion: The central nervous system fatigue could result in decreased Y maze results and gene expressions of hippocampal NR2B and EphB2. Sihui Mixture could improve rats' learning and memory ability, which might be possibly achieved through up-regulating mRNA expressions of hippocampal EphB2 and NR2B.

Using numerical simulation, we have studied in detail vacuum electron acceleration driven by two crossed Airy beams with identical characteristics except for opposite accelerating directions. An electron injected along the longitudinal central axis is only affected by the combined longitudinal electric field. In addition, a suitable crossed Airy beams scheme is more beneficial to the energy gain of an electron than the single Airy beam acceleration scheme [Opt. Lett. 35, 3258 (2010)]. Meanwhile, the cross angle, the injection energy of the electron, and the initial phase of the Airy beams play significant roles in the energy gain of the electron.

The present study aimed to study the changes of neurotrophin-3 (NT-3) expression of intrafusal muscle fibers in rat soleus muscles under simulated weightlessness. The tail-suspension (SUS) rat model was used to simulate weightlessness. Forty mature female Sprague-Dawley rats were randomly assigned to ambulatory control (CON), 3-day SUS, 7-day SUS, 14-day SUS and 21-day SUS groups. Immunohistochemistry ABC staining method and enzyme linked immunosorbent assay (ELISA) were used to detect the NT-3 expression of intrafusal muscle fibers in rat soleus muscles. The results from the immunohistochemistry staining technique showed that the extrafusal muscle fibers did not exhibit the NT-3-like immunoreactivity, and NT-3-like immunoreactivity was mainly expressed in nuclear bag 1 and nuclear bag 2 fibers of the muscle spindles. The ELISA results showed that the expression quantity of NT-3 in rat soleus muscles in control, 3-day SUS, 7-day SUS, 14-day SUS and 21-day SUS groups were (14.23±1.65), (14.11±1.53), (13.09±1.47), (12.45±1.51) and (9.85±1.52) pg/mg of tissue respectively. Compared to the control group, the expression quantity of NT-3 was significantly decreased after 14 days of SUS (P<0.05). After 21 days of SUS, the NT-3 expression was further reduced (P<0.01). These results suggest that simulated weightlessness induces an obvious decrease in the NT-3 expression level of intrafusal fibers in rat soleus muscles. Accompanying the simulated weightlessness extension, NT-3 expression in rat soleus muscle spindles is progressively decreased. These changes may contribute to the proprioceptive adaptations to microgravity.

Objectives: To study the effect of Houxue Zhitong decoration on the expression of the mitochondria-initiated apoptosis pathway and transmembrane protein I an II of the epidural scar tissue., Methods: A total of 60 New Zealand rabbits (weight: 2.5-3.0 kg) were randomly divided into four groups, sham operation group (D, n=15), control group (B, n=14), sodium hyaluronate group (C, n=15), Houxue Zhitong decoration group (D, n=15). Except for group A, 1.0 cm x 1.0 cm dura mater uncovered area laminectomy was performed at I (4) and I(5), covered with 0.5 ml sodium hyaluronate in group C, covered with same amount of saline in group B and D. First 2 weeks after operation, animals in group D were lavaged with 2.5 ml/kg Houxue Zhitong decoction by one a day for 14 days. Five rabbits of each group selected randomly were killed in the 2,4,8 weeks after laminectomy. The specimens were prepared for determination of the expression of Fas and FasL, at scar tissue by semiquantitative reserve transcription-polymerase chain reaction (RT-PCR). The degree of scar adhesion was evaluated according by Rydell method., Results: The adhesion area in group B was larger than of group C and D in the 4th and 8th week. However, the number of fibroblasts and inflammantory cells in group D was the least among the three groups in the 8th week. At 2, 4, 8 weeks after operation, as compared with group B the expression of Fas, FasL of group C and D were decreased (P < 0.05). Especially, at 2 weeks, as compared with group B the expression of this two cytokines of group D was significant decreased (P < 0.05), too. In group C and D the duramater adhesion was decreased (P < 0.05). The proliferation of fibroblast and fibroblastic function were inhibited (P < 0.05)., Conclusion: Huoxue Zhitong is able to down-regulated the expression of Fas, FasL, which inhibited the proliferation of fibroblast, the fibroblastic function and the synthesis of extracellular matrix in the epidural scar tissue. It is an effective way of reducing peridural scar formation and prevent the failed back surgery syndrome.

Objective: To study the effect of Huoxuezhitong recipe on the pathway of mitochondrial apoptosis and the expression of related gene (Caspase-3, -9, Bax, Bcl-2) in epidural scar tissue., Methods: A total of 60 New Zealand rabbits, ranging in weight from 2.5 to 3.0 kg, were randomly divided into sham operative group (group A), control group (group B), sodium hyaluronate group (group C), Huoxuezhitong recipe group (group D, included Danggui 20 g, Chishao 20 g, Honghua 20 g, Ruxiang 15 g, Myrrh 15 g etc.) with 15 rabbits in each group. Except for group A, L4,5 vertebral plate were resected leading to 1.0 cm x 1.0 cm dura mater exposed area, which were covered with 0.5 ml sodium hyaluronate in group C, covered with same amount of saline in group B and D. At 2 weeks after operation, the rabbits of group D were administered with Huoxuezhitong recipe (2.5 ml/kg, once a day, continuous 14 d). Five rabbits of each group were selected randomly and were killed at 2, 4 and 8 weeks after operation; and the expression of Caspase-3, -9, Bax, Bcl-2 in scar tissue were surveyed by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR)., Results: The results by RT-PCR showed significant different in expressions of Caspase-3, -9, Bax, Bcl-2 of group A and others groups at each period (P < 0.01). At 2, 4, 8 weeks after operation, compared with group B, the expressions of Caspase-3, -9, Bcl-2 in group C and D were decreased (P < 0.05), but the expression of Bax in group C and D were increased (P < 0.05). At 8 weeks after operation, the expressions of Caspase-3, -9, Bax, Bcl-2 in group D was significant decreased compared with group C (P < 0.05)., Conclusion: Huoxuezhitong recipe can regulate the levels of expression of Caspase-3, -9, Bax, Bcl-2 mRNA and further induce the pathway of mitochondrial apoptosis, thereby, achieve the effect of prophylaxis for the proliferation and conglutination of epidural scar tissue.

During hind limb unloading (HU), the soleus is often in a shortened position and the natural physiological stimulus of muscle spindles is altered, such that muscle spindle activity also changes. Using isolated spindle conditions, the present study investigates the electrophysiological activity and ultrastructure of muscle spindles following HU. Results show that muscle spindle discharges fall into either of two main patterns, single spikes or spike clusters in shortened positions, with a steady frequency of 18-38 spikes/s (mean 29.08 +/- 2.45) in an extended position. Following 14-day HU, afferent discharge activity was significantly altered in soleus muscle spindles. Duration of individual spikes was significantly prolonged, from 0.54 +/- 0.05 ms for control rats to 1.53 +/- 0.25 ms for rats in the HU group. In a shortened position, regular rhythm afferent discharges were obviously depressed, and the majority of muscle spindles became silent, while in an extended position, the discharges remained continuous but with decreased frequency. Results also show that the ultrastructure of muscle spindles experience degenerative changes during HU. Altered muscle spindle afference could possibly modify the activity of motor neurons and further affect the activity of extrafusal fibers.

Background: Stathmin was identified as an endometriosis-related protein by comparative proteomics in our previous study. As a microtubule-destabilizing factor, stathmin was shown to participate in the relay and integration of diverse intracellular signaling pathways involved in cell proliferation, differentiation, and many other cellular activities. To investigate whether stathmin is involved in the pathogenesis of endometriosis, we examined the expression of stathmin in eutopic endometrium of women with or without endometriosis., Methods: Eutopic endometrium samples were collected from thirty-six patients who were diagnosed as endometriosis and the nineteen age-matched patients who were confirmed to be free of endometriosis surgically and histologically. The expression of stathmin mRNA was detected by real-time PCR, and its protein was detected by Western blotting and immunohistochemistry., Results: Stathmin was overexpressed in eutopic endometrium of women with endometriosis detected by real-time PCR in mRNA levels and by Western blotting in protein levels, without significant difference between proliferative and secretory phase. Immunohistochemistry showed that stathmin protein was localized in both endometrial glandular and stromal cells throughout the menstrual cycle., Conclusions: Stathmin is overexpressed in endometrium of patients with endometriosis and may play a role in the pathogenesis of endometriosis.

Human leukocyte antigen-G (HLA-G) expression in 55 cervical intraepithelial neoplasia (CIN) patients with or without human papillomavirus (HPV) infection and 116 patients with squamous cell cervical cancer were examined using immunohistochemistry. Host immune response was assessed by estimating the number of intratumoral lymphocyte infiltration (TIL) in all lesions and counting CD57-expressing cells in the neoplasm lesions. The means of HLA-G immunoreactive scores were compared by the Mann-Whitney test and 1-way analysis of variance (ANOVA). The association of HLA-G expression with disease progression, HPV infection and host immune response was calculated using the Pearson Chi-square test. It was found that HLA-G expression increasingly progressed from patients with CIN 1 to CIN 2/3 and was highest in patients with cervical cancer. Human leukocyte antigen-G expression was also significantly higher in CIN and cancer patients with HPV 16/18 than in CIN patients without HPV. A significant correlation between HLA-G expression and TIL score or the counting of CD57-expressing cells was also evident in CIN patients with HPV infection and cervical cancer cases. These results suggest that HLA-G expression in cervical lesions is associated with carcinogenesis, HPV infection, and host immune response.

It has been demonstrated that hypoxic preconditioning (HP) enhances the survival ability of the organism against the subsequent acute anoxia (AA). However, it is not yet clear whether necrosis induced by AA can be prevented by HP, and what are the underlying mechanisms. In this study, we examined the effect of HP (10% O(2), 48 h) on necrosis induced by AA (0% O(2), 24 h) in PC12 cells. We found that HP delayed the regulatory volume decrease and reduced cell swelling after 24 h of exposure to AA. Since aldose reductase (AR) is involved in cell volume regulation, we detected AR mRNA expression with reverse transcription-polymerase chain reaction (RT-PCR) techniques. The AR mRNA level was dramatically elevated by HP. Furthermore, an HP-induced decrease in cell injury was reversed by berberine chloride (BB), the inhibitor of AR. In addition, sorbitol synthesized from glucose catalyzed by AR is directly related to cell volume regulation. Subsequently, we tested sorbitol content in the cytoplasm. HP clearly elevated sorbitol content, while BB inhibited the elevation induced by HP. Further study showed that a strong inhibitor of sorbitol permease, quinidine, completely reversed the protection induced by HP after AA. These data provide evidence that HP prevents necrosis induced by AA and is mediated by AR and sorbitol pathway.

It is important to identify pigments of painting and colored drawing in ancient architectures in order to restore and conserve them. The components of green pigments were detected with X-ray diffraction (XRD), X-ray fluorescence (XRF) and scanning electron microscopy-energy dispersive X-ray (SEM-EDX). Twenty-seven samples were collected from painting and colored drawing in northern Chinese ancient architectures in Beijing, Shanxi province and Gansu province. The experiment results showed that emerald green [CuCH3COO]2 x Cu(AsO2)2], a complex of copper aceto-arsenite pigment, had been used as the colored component in fifteen samples, whereas organic materials synthesized in the rest. However, in all samples there were no malachite and atacamite, green pigments commonly used in ancient time a long time ago. These two pigments have been found in Qin Shihuang's Terracotta Army and the wall paintings at Mogao Grettoes, Dunhuang, and some other famous wall paintings and color pottery figurines. However, emerald green was used many years later. It was reported that emerald green was synthesized by Germany in 1814 and had been widely used in China as watercolor on pith paper works and on scroll paintings since the 1850s. Because painting and colored drawing in ancient architectures stands outside, under sunlight and rain, it must be repaired and repainted in less than fifty years. Therefore, it is not surprising that emerald green was used in them. In recent years, artificial organic materials are increasingly used in painting and colored drawing in ancient architectures. From experiments it was also showed that in the same recolored painting and colored drawing, organic materials are usually in the later layers, but emerald green is in the earlier layers. This work supplies a lot of data for the purpose of selecting restoration materials and identifying painting and colored drawing in ancient architectures with a new method.

The flame temperature was measured by the double line of atomic emission spectroscopy according to the spectra of K (766.5 and 769.9 nm) whose relative intensity was obtained by fiber spectrometer. The principles, methods and experiment system were described. The temperature measured by the double line of atomic emission spectroscopy was compared to the results measured by thermal couple under the condition of thermal equilibrium of blackbody furnace. The comparison indicated a good coherence between these two measurement methods. The method was demonstrated on coal powder and timber, and the temperature measured corresponded to reality.

To date, the medium and long-term space flight is urgent in need and has become a major task of our manned space flight program. There is no doubt that medium and long-term space flight has serious damaging impact upon human physiological systems. For instance, atrophy of the lower limb anti-gravity muscle can be induced during the space flight. Muscle atrophy significantly affects the flight of astronauts in space. Most importantly, it influences the precise manipulation of the astronauts and their response capacity to emergencies on returning to the atmosphere from space. Muscle atrophy caused by weightlessness may also seriously disrupt the normal life and work of the astronauts during the re-adaptation period. Here we summarize the corresponding research concentrating on weightlessness-induced changes of muscular structure and function. By combining research on muscle pain, which is a common clinical pain disease, we further provide a hypothesis concerning a dynamic feedback model of "weightlessness condition right triple arrow muscular atrophy <--> muscle pain". This may be useful to explore the neural mechanisms underlying the occurrence and development of muscular atrophy and muscle pain, through the key study of muscle spindle, and furthermore provide more effective therapy for clinical treatment.

Objective: To detect the specific mutations in rpoB gene of Mycobacterium tuberculosis by oligonucleotide microarray., Methods: Four wild-type and 8 mutant probes were used to detect rifampin resistant strains. Target DNA of M. tuberculosis was amplified by PCR, hybridized and scanned. Direct sequencing was performed to verify the results of oligonucleotide microarray., Results: Of the 102 rifampin-resistant strains 98 (96.1%) had mutations in the rpoB genes., Conclusion: Oligonucleotide microarray with mutation-specific probes is a reliable and useful tool for the rapid and accurate diagnosis of rifampin resistance in M. tuberculosis isolates.

Bone transplantation is widely used in orthopaedics, and allograft bone transplantation is being more and more emphasized. In this article, the allograft bone combined with growth factors transplantation for repairing bone defects were reviewed. Moreover, the way to compound many growth factors and other material is the tendency of allogenic bone grafting, which enhance the opportunity of success in bone transplanting.

A series of novel 12-(aryloxyacyloxyimino)-1,15-pentadecanlactone derivatives (3) were synthesized, and their structures including configuration of C=N bond were confirmed by (1)H NMR, elemental analysis and X-ray diffraction analysis. The bioassay showed that some of them exhibited excellent herbicidal activity against Amaranthus tricolor L. The activity of compounds 3 except compounds 3A1-2 was much higher than the commercial herbicide 2,4-D and the activity of about half of compounds 3 was comparable to the commercial herbicide tribenuron-methyl. The further bioassay showed that the representative of compounds 3, 3A1-12, exhibited excellent herbicidal activity not only against dicotyledon, such as Amaranthus tricolor L., Cucumis sativus L., Glycine max L., and Phaseolus radiatus L., but also against monocotyledon, such as Zea mays L. and Oryza sativa L.

The present study investigates changes in red nucleus (RN) neuronal activity and the role of glutamate receptors (GluRs) after simulated microgravity (tail-suspension) in the rat using single-unit recording and microinjection. The results showed that tail-suspension for 3, 7, and 14 days could induce a significant decrease in spontaneous firing rate of RN neurons in a time-dependent manner. Unilateral microinjection of glutamate into the RN significantly increased the firing rate of RN neurons, but the increased firing rate was significantly reduced following tail-suspension time. Microinjection of the NMDA receptor antagonist MK-801 or the non-NMDA receptor antagonist DNQX into the RN blocked this excitatory effect induced by glutamate. However, microinjection of the metabotropic glutamate receptor (mGluR) antagonist (+/-)-MCPG into the RN had no effect. These results suggest that simulated microgravity can reduce excitability of RN neurons following a functional impairment of glutamate receptors. NMDA and non-NMDA receptors, but not mGluRs, are involved in the mediation of glutamate-evoked excitation of RN neurons. The decrease in excitability of RN neurons may be involved in simulated microgravity-induced muscle atrophy.

4-nitrobenzoic acid and hydraxine sulfate were cyclized with H3PO4/P2O5 to afford 2,5-bis(4-nitrophenyl)-1, 3,4-oxadiazole(1), then(1)was reduced to 2,5-bis(4-aminophenyl)-1,3,4-oxadiazole using Zn/CaCl2 as reduction reagent, and at last, a series of novel di-schiff base derivatives containing oxadiazole ring were obtained by the reaction of 2,5-bis(4-aminophenyl)-1, 3,4-oxadiazole with aromatic aldehydes (yield: 65%-81%). The structures of these compounds were confirmed by 1H NMR, FTIR and MS. Their UV-Vis spectra as well as fluorescence spectra were studied and the electrochemical properties were tested with cyclic voltammetry. The UV-Vis spectroscopy results show that the characteristic absorption peaks of 1, 3, 4-oxadiazole unit and di-schiff base unit disappeared. The maximum absorption wavelength was detected at 345-357 nm because conjugation effect made the two energy bands partially hybrid and form a new energy band structure. Fluorescence spectroscopy results show that they emit strong blue and purple fluorescence, and the emission maximum wavelengths of di-schiff base are from 390 to 407 nm. Electrochemical properties results show that they have higher affinity energy (2.36-3.04 eV) and ionic potential (5.35-6.06 eV), which indicated that they have better electron-transporting properties than PBD except 3 a and 3 h. The results give a reference to further application to organic electroluminescent devices of the target compound.

Objective: To analyze LC-MS fingerprints of Aristolochia manshuriensis for quality assessment with two different chemical pattern recognition models., Method: LC-MS fingerprints of A. manshuriensis were established from 24 batches of samples from different habitats. SIMCA and Clustering analysis were used to compare the parameters of the 29 common peaks., Result: Two methods had good consistency, while they reflected the inherent sample information from different perspectives, respectively., Conclusion: Modern equipment analysis technology and multivariable chemical pattern recognition would be an efficient way for quality control and variety identification of A. manshuriensis.

To determine whether the neuronal activity of the spindle cord in muscle spindle afferent pathways is altered after a period of hindlimb unloading (Hu), and after recovery, we focused on c-Fos-immunoreactivity in the spinal cord evoked by excitation of the muscle spindle in normal gravity and after 3, 7, and 14 days of Hu, and after 14 days of Hu with an additional 2, 5, and 9 days of unrestricted cage activity. High frequency sinusoidal vibration (HFV) applied to the Achilles tendon was used for activation of the muscle spindle. Results showed that c-Fos-immunoreactive neurons evoked by HFV were mainly concentrated in lamina IV-VII of the ipsilateral spinal cord. After 14 days of Hu, the total number of labeled neurons in the spinal cord was significantly increased (P<0.05). The number of c-Fos-immunoreactive neurons in lamina IV and VI-VII were also significantly higher than that in normal rats (P<0.05). After 9 days of reloading, the total number of c-Fos-immunoreactive neurons and the pattern of their lamellar distribution were both recovered to pre-experimental levels. Our findings suggest that simulated weightlessness induces a large increase in neuronal excitation of the spinal cord in muscle spindle afferent pathways. Weight bearing immediately after Hu could induce completed recovery of excitability of the spinal cord in muscle spindle afferent pathways.

Objective: By means of the detection of the numbers of CD34(+) cells and eosinophils (EOS), and the level of IL-5 in peripheral blood from normal controls and patients with allergic rhinitis pre- or post-treatment, the role of EOS-stem cells paths for treatment effect in allergic rhinitis (AR) was studied so as to find the convenient and quick indicators which could be used to evaluate the clinical therapeutic effect and adjust the methods of the hormone therapy., Methods: There were 2 groups. (1) experimental group: 44 patients, including 24 males and 20 females with a age range of 7 to 68 years old. The patients received the treatment of fluticasone spurt for four weeks. (2) CONTROL GROUP: 30 cases, from normal health examination. The EOS numbers, CD4(+) cell numbers and the IL-5 level were examined in control group as well as before and after therapy in experimental group., Results: The IL-5 level and CD numbers before therapy in experimental group were (88.25 +/- 33.47) ng/L, (9.24 +/- 2.15)/10(5), significantly higher than that after therapy and experimental group which were (44.34 +/- 16.32) ng/L, (6.31 +/- 1.83)/10(5) and (31.24 +/- 8.43) ng/L, (3.47 +/- 1.32)/10(5) respectively. The IL-5 level was positively correlated with the CD34(+) number pre-and post treatment in experimental group (r = 0.64, P <0.01; (r = 0.61, P <0.01). The EOS number was positively correlated with the level of IL-5 (r = 0.64, P <0.01)., Conclusions: IL-5 and CD34(+) cells can be regarded as indicators to evaluate the therapeutic effect. The EOS, CD34(+) cells and the level of IL-5 in the peripheral blood are correlated with the pathogenesis of AR, suggesting that there is a related path between the local nasal tissue of AR patients and the marrow.

Objective: To clone pIA and pIB genes of Neisseria gonorrhoeae,and to construct pIA-pIB fusion gene and its prokaryotic expression system, and to establish enzyme linked immunosorbent assay (ELISA) based on rPIA-PIB for detecting serum and pus samples from gonorrhea patients and to evaluate the sensitivity and specificity of the ELISA., Methods: pIA-pIB fusion gene was constructed by polymerase chain reaction (PCR) using linking primers and a prokaryotic expression system of the fusion gene was constructed by using routine molecular biological methods. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) plus BioRad Gel Image Analyzer was used to measure the expression of the target recombinant protein rPIA-PIB. Ni-NTA affinity chromatography was performed to extract and purify rPIA-PIB. An ELISA by using rPIA-PIB as the coated antigen for detecting the specific IgG against rPIA and/or rPIB in gonorrhea patients' sera as well as another ELISA by using rPIA-PIB antiserum as the first antibody for detecting the rPIA and/or rPIB in gonorrhea patients' pus samples were established. In these experiments, ELISAs associated with rPIA, rPIB and their antisera were applied as the controls., Results: 100% similarities of the nucleotide and putative amino acid sequences of the pIA-pIB fusion gene were confirmed when compared with the original sequences. The output of rPIA-PIB was 29.8% of the total bacterial proteins. The purified rPIA-PIB only showed a single target protein segment in gel after SDS-PAGE. Using a positive rate (98.3%) of rPIA-PIB-IgG-ELISA to detect 119 cases of gonorrhea patients' serum samples was remarkably higher than that of rPIA-IgG-ELISA (30.3%) or rPIB-IgG-ELISA (66.4%) (P<0.01). The positive rate (91.6%) of rPIA-PIB-ELISA to detect 119 cases of gonorrhea patients' pus samples was also significantly higher than that of rPIA-IgG-ELISA (27.7%) or rPIB-IgG- ELISA (62.2%) (P<0.01)., Conclusion: In this study we successfully constructed pIA-pIB fusion gene of N. gonorrhoeae and its prokaryotic expression system while rPIA-PIB showed obvious superiority used as the antigen in gonorrhea associated detection kits compared to both the rPIA and rPIB.

Objective: To investigate the protective action of Epimedium against chemotherapy-induced damage to rat epididymides., Methods: Fifty 60-day-old male rats were divided into a control, a model and a treatment group. Procarbazine was injected into the abdominal cavity of the model rats at the dose of 30 mg/(kg x d). In addition to procarbazine, Epimedium was given intragastrically to the treatment group. The changes in the ultrastructure of the epididymis were observed after 10 and 20 days., Results: Electron microscopy showed that the chemotherapy-induced damages to the epididymal epithelia mainly included cell swelling, local cavitation of mitochondria, tumor-like change in nucleoli, agglutination of marginal translocation of heterochromatin and cell apoptosis. The damage to the epithelial ultrastructure was slight in the treatment group as compared with the model rats. Chemotherapy significantly affected sperm concentration, sperm viability and sialic acid (SA), which were (15.59 +/- 4.01) x 10(6)/ml, (76.71 +/- 10.11)% and (19.38 +/- 9.34) g/mg prot in the model group in comparison with (10.63 +/- 3.82) x 10(6)/ml (P < 0.01), (60.03 +/- 7.54)% (P < 0.01) and (13.62 +/- 7.81) g/g prot (P < 0.05) in the control. Epimedium significantly increased sperm viability in the treatment group (60.03 +/- 7.54)% as compared with the model rats (69.90 +/- 12.58)% (P < 0.05)., Conclusion: Epimedium can lessen chemotherapy-induced damage to the epididymis and protect the reproductive function of rats.

The aim of this study was to investigate the efficacy of diacetyl hexamethylene diamine (CAHB) for patients with high risk myelodysplastic syndrome (MDS), and to explore the effect of CAHB on HL-60 cells in vitro and its possible mechanism. 8 patients with high risk MDS were treated with CAHB by continuous intravenous infusion for 10 days, and repeated once after an interval of 28 days. The count of the granulo- and mono-blasts in bone marrow (BM) aspirate was measured before and after treatment. HL-60 cells were treated with different concentrations of CAHB for 72 hours in vitro. The inhibitory effect of CAHB on proliferation of HL-60 cells in vitro was measured by MTT assay. Differentiation of HL-60 cells was detected by the changes of CD11b and CD14 expression on cell surface. Apoptosis of HL-60 cells was detected by double staining of Annexin V and PI. The cell cycle distribution change of HL-60 cells was analyzed by flow-cytometry. The results indicated that the granulo- and mono-blasts in BM decreased in all the 8 patients after CAHB treatment. The main side effect of CAHB on hematological system was thrombocytopenia. After being treated with 1, 2, 3, 4 mmol/L CAHB for 72 hours in vitro, the result of MTT assay showed the inhibitory effect of CAHB on the proliferation of HL-60 cells in dose-dependent manner. After being treated manner 1, 2, 3, 4 mmol/L CAHB for 72 hours, the CD11b positive HL-60 cells were 22.39+/-3.97%, 33.12+/-4.46%, 49.25+/-5.27%, 78.05+/-5.66%, respectively, which were significantly different from the control group (CD11b positive HL-60 cells was 5.89+/-2.94%) (p<0.01). The CD14 expression was negative in all the 5 groups. These results suggested that CAHB could induce HL-60 cells to differentiate into mature granulocytes, and the effect of CAHB appeared in dose-dependent manner. After being treated for 72 hours by 1, 2, 3, 4 mmol/L CAHB, the apoptotic cells (Annexin V(+)/PI(-) cells) increased mildly, which suggested that CAHB only weakly induces HL-60 cells to apoptosis at the concentration of 1 to 4 mmol/L. Along with the concentration increase of CAHB, the ratio of cells in G(0)/G(1) phase increased, and ratio of cells in S phase and G(2)/M phase decreased correspondingly, it indicated that CAHB could arrest HL-60 cells in G(0)/G(1) phase in a dose-dependent manner. It is concluded that induction of cell differentiation may be the primary effect of CAHB on MDS. Cell cycle arrest may be essential to the effect of CAHB as well. Side effect of CAHB on platelet count may correlated with its inhibitory effect on hematopoiesis.

Objective: To clone tpn17 and tpn47 genes of Treponema pallidum and then construct their prokaryotic expression systems,to establish ELISAs based on rTpN17 and rTpN47 as antigens and to evaluate the sensitivity and specificity of the ELISAs for detection of serological diagnosis of syphilis., Methods: The whole length of tpn17 and tpn47 genes was amplified by PCR and then their prokaryotic expression systems were constructed. SDS-PAGE was used to measure the expression of the target recombinant proteins rTpN17 and rTpN47. Ni-NTA affinity chromatography was applied to extract rTpN17 and rTpN47, while Western blot was performed to determine the specific immunoreactivity of rTpN17 and rTpN47. By using rTpN17 and rTpN47 as the coated antigen, respectively, ELISAs (rTpN17-ELISA and rTpN47-ELISA) were established to detect serum samples from 200 healthy individuals, 25 RA patients, 17 SLE patients and 211 syphilis patients. The detection effects of the ELISAs were compared to those of TRUST and TPHA., Result: The sequence similarity of the cloned tpn17 and tpn47 genes was 100 % compared with the corresponding sequences in GenBank. The expression outputs of rTpN17 and rTpN47 were approximately 37.2 % and 26.8 % of the total bacterial proteins, respectively. Both the extracted rTpN17 and rTpN47 could take place remarkable conjugation reactions to the sera with positive antibody against Treponema pallidum.The positive detection rate of TPHA (99.1%) was the highest (P<0.001). The positive detection rates of rTpN17-ELISA (85.3 %) and rTpN47-ELISA (84.3 %) were similar (P>0.05). The positive detection rates of TRUST (72.5 %) was lower than that of rTpN17-ELISA (P=0.001) but similar to that of rTpN47-ELISA (P=0.014). The detection results of all the serum samples from healthy individuals, RA patients and SLE patients were negative, whereas 7.1 % (3/42) of the samples from RA or SLE patients were positive., Conclusion: rTpN17 and rTpN47 are still maintaining their original immunoreactivity. The ELISAs using rTpN17 or rTpN47 as the antigen are rapid, simple and convenient, higher sensitivity and specificity methods for serological screening and detection of syphilis.

Chlorophyll a and chlorophyll b play a significant role in the plant growth process. Precise determination of its content could provide the scientific basis for the corps growth state, the plant pathology diagnosis and so on, and is the key point of implementing accurate agriculture. The authors transfered the absorption spectrum experiment data of chlorophyll a and chlorophyll b determined by predecessor into a transmission spectrum. Observing the transmission spectrum using human eyes, the color of chlorophyll a and chlorophyll b was obtained. Thus, according to the predecessor's experiment curve of absorption spectrum of chlorophyll a and chlorophyll b, combining the three primary color principles of colorimetry, and choosing a standard light source with spectrum power distribution similar to D65 as the illumination, the authors determined their chromaticity coordinates in the 1931CIE-x, y chromaticity diagram: Chlorophyll a is (0.198 1, 0.334 1), which falls in the cyan color region; Chlorophyll b is (0. 270 4, 0. 566 3), which falls in the yellowish green region. The main point of above processing is: reducing the spectrum curves of chlorophyll a and the chlorophyll b to coordinates on the chromaticity diagram, and the result could offer the essential theoretical support for a new non-contact, long-distance and non-damage technology to determine the pigment content in single leaf or mass.

Objective: To explore the role of connective tissue growth factor (CTGF) on pulmonary artery remodeling induced by smoke exposure in rats., Methods: Thirty-five male Wistar rats were randomly assigned into a control group (A group), a smoke exposure one month group (B group), a smoke exposure and high dose CTGF antisense oligonucleotide (ASON) one month group (C group), a smoke exposure and low dose CTGF ASON one month group (D group), a smoke exposure two month group (E group), a smoke exposure and high dose CTGF ASON two month group (F group), and a smoke exposure and low dose CTGF ASON two month group (G group). Pulmonary artery remodeling was observed by hematoxylin-eosin staining, and the CTGF mRNA expressions of pulmonary arteries were evaluated by RT-PCR. Immunohistochemistry methods were performed to determine CTGF protein expression in pulmonary artery smooth muscle. The difference between the groups was analyzed., Results: (1) The pulmonary artery WA% of the seven groups were respectively (28.6 +/- 1.2)%, (42.5 +/- 2.3)%, (33.7 +/- 1.8)%, (42.1 +/- 2.4)%, (49.6 +/- 2.1)%, (34.3 +/- 1.9)% and (38.4 +/- 2.0)%. There was significant difference between B group and C group (q = 5.09, P < 0.01). Compared to E group, there were significant decreases in F group and G group (q = 8.15, 3.75, all P < 0.05). (2) The CTGF protein expressions (A value) of pulmonary artery smooth muscle were respectively 0.098 +/- 0.015, 0.159 +/- 0.023, 0.118 +/- 0.017, 0.153 +/- 0.022, 0.406 +/- 0.036, 0.109 +/- 0.012 and 0.146 +/- 0.024. There was significant difference between B group and C group (q = 3.26, P < 0.05). Compared to E group, there were significant decreases in F group and G group (q = 67.08, 18.09, all P < 0.01). (3) The CTGF mRNA expressions (A(CTGF)/A(beta-actin)) of pulmonary artery were respectively 0.051 +/- 0.010, 0.823 +/- 0.096, 0.216 +/- 0.056, 0.810 +/- 0.085, 2.452 +/- 0.267, 0.207 +/- 0.062 and 0.509 +/- 0.067. There was significant difference between B group and C group (q = 53.50, P < 0.01). Compared to E group, there were significant decreases in F group and G group (q = 132.22, 93.70, all P < 0.01)., Conclusion: Administration of CTGF ASON in vivo could significantly reduce pulmonary artery remodeling induced by smoke exposure in rats, indicating that CTGF may play an important role in pulmonary hypertension.

Aim: To clone HPV58 E6 gene and analyze the HLA-DQB1 03-restricted T cell epitopes on E6 protein., Methods: Total genome DNA was isolated from a cervical cancer sample. The HPV58 E6 gene was amplified by PCR and inserted into pGEM-T Easy vector. The recombinant plasmid was identified by restriction endonuclease analysis and sequencing. HLA-DQB1 03-restricted T cell epitopes on E6 protein were predicted and analyzed by the position-specific scoring matrix, support vector machine theory and prediction algorithm for proteasomal cleavages., Results: A HPV58 E6 gene was successfully cloned and submitted to GenBank (EF060239). Epitope 47(FADLRIAYRDGNPFA) and Epitope 102(RCIICQRPLCPQEKK) were theoretic HLA-DQB1 03-restricted T cell epitopes on E6 protein., Conclusion: These two epitopes could serve as candidates for screening and identification of the vaccine against HPV58 infection in the further study.

Objective: To establish a method to analyze HPLC fingerprint characteristics of 10 plants from Selaginella., Method: HPLC was applied for establishment of fingerprints, which were used to evaluate and distinguish the different species of Selaginella., Result: The different species from Selaginella showed different HPLC fingerprint characteristic. The samples of the same species but collected in different period, different environment or different locations showed certain difference in fingerprints, Conclusion: 2 important mutual fingerprint peaks were found in the 10 plants of Selaginella species and 5 peaks can be used as "main fingerprint peaks". The dates of these peaks can used for assessment of phylogenetic relation among species and evaluation of quality.

A series of star-shaped oligofluorenes end-capped with carboxylic acid groups were synthesized. Different numbers of carboxyl groups that can form hydrogen bonds, and long alkane chains that have stabilizing effects, were intentionally introduced. The resulting molecular architectures of the so-prepared star-shaped oligofluorenes at the liquid-solid interface were investigated by scanning tunneling microscopy. It is found that the number of hydrogen-bonding groups and the symmetry of the target molecules have crucial influences on the structures of the ordered assemblies.

Objective: To study the occurrence and predictive factors of hepatitis B virus DAN negative conversion among patients with chronic hepatitis B and liver cirrhosis during lamivudine therapy., Methods: Clinical data of 202 patients with chronic hepatitis B and 55 patients with post-hepatitis B cirrhosis treated with lamivudine therapy were analyzed and PCR as well as mispairing PCR-restricted fragment length polymorphism (RFLP) assay were applied. The data including ALT, AST, total bilirubin (TBIL), albumin, etc. were analyzed by SPSS software., Results: The HBV DNA negative conversion occurred in 87.62%, 95.04%, 86.63%, 82.65%, 75.43%, and 77.27% of the chronic hepatitis B patients and in 94.54%, 98.18%, 81.81%, 74.35%, 73.91%, and 63.63% of liver cirrhosis patients, respectively, after 3, 6, 12, 18, 24, 30 months. The results of log-rank test and Cox proportional hazard model analysis indicated that combination of lamivudine and interferon, high ALT level,low HBV DNA level, and low albumin level at baseline were significantly related to HBV DNA negative conversion., Conclusion: Combination treatment with lamivudine and interferon, high ALT level, low HBV DNA level, and low albumin level at baseline were significant predicting predicting factors for HBV DNA negative conversion in chronic hepatitis B patients.

Background: Tuberculosis remains the leading cause of human death. Currently, Bacillus Calmette-Guérin (BCG) is the only available vaccine against tuberculosis but its efficacy is highly variable. Thus, developing new tuberculosis vaccines becomes an urgent task. In this study, we evaluated in BALB/c mice the humoral and cellular immune responses of recombinant BCG expressing the antigen ESAT-6 from Mycobacterium tuberculosis., Methods: Escherichia coli-BCG shuttle plasmid named pDE22-esat-6 was constructed by inserting the BamHI/EcoRI digested esat-6 gene PCR product into the similarly digested parental plasmid pDE22. BCG cells were transformed with pDE22-esat-6, which was named recombinant BCG (rBCG). BALB/c mice were immunized subcutaneously on the back with 100 microl normal saline containing 10(6) CFU of BCG or rBCG. They were sacrificed after 4 weeks to detect their humoral and cellular responses., Results: There was no any significant differences in the growth characteristics between the conventional BCG and rBCG. In immunized mice, the IgG antibody titres of rBCG group were as high as 1:8000, which was significantly higher than that in BCG group (1:1400, P < 0.05). The elicited IFN-gamma level of rBCG group was (1993 +/- 106) pg/ml, which was also significantly higher than that in BCG group ((1463 +/- 105) pg/ml, P < 0.05). The splenocyte proliferation index of rBCG group reached 4.34 +/- 0.31, which was higher than that of BCG group (3.79 +/- 0.24, P < 0.05)., Conclusion: rBCG secreted expressing antigen ESAT-6 stimulated stronger humoral and cellular immune responses than BCG did, and, therefore may be the better vaccine against mycobacterium tuberculosis.

Application of HPLC-ESI-ITMS in the quality control of carboxyterminal sequence confirmation for insulin and insulin chain B was studied. The solution of intact insulin or insulin chain B was added to the solution of carboxypeptidase P (CPP) and carboxypeptidase Y (CPY). Fractions of appropriate volume were removed at some appointed time points, acidified with the same amount of 1% formic acid to stop the digestion, and then briefly vortexed for HPLC-ESI-ITMS analysis. Mobile phase A consisted of 0.02% TFA in 98% ultra-pure water and 2% acetonitrile. Mobile phase B consisted of 0.02% TFA in 98% acetonitrile and 2% ultra-pure water. The solution used for post-column fix consisted of propionic acid and isopropyl alcohol (20 : 80, v/v). Chromatographic separation was carried out on a reversed-phase column (Zorbax Prosphere C18, 300A, 5 microm, 2.1 mm ID x 150 mm length). The molecular weights of the multiply charged ions representing consecutive truncated losses of carboxyterminal amino acids were determined by the use of HPLC-ESI-ITMS. The differences between the consecutive truncated peptides are the experimental weights of the carboxyterminal amino acid residues. The carboxyterminal amino acid residue Ala, which released from chain B of intact insulin, was confirmed in the nanomolar concentration range by analyzing the molecular weight of the truncated peptides. Another one carboxyterminal amino acid Ala was confirmed in the nanomolar concentration range of insulin chain B. In the quality control for recombinant DNA product or natural protein, the confirmation of 1 - 3 carboxyterminal amino acid residues is regarded to be up to standard. One amino acid residue of insulin or insulin chain B could be confirmed accurately in the nanomolar concentration range. The results showed that intact insulin could be directly sequenced in the quality control without separating chain B from chain A. There would be no need to separate chain A from chain B to identify carboxyterminal of intact insulin. Furthermore, the method saved us a lot of trouble from the preparation and purification of insulin chain A and chain B.

To study the possibility of utilizing genetically engineered Leifsonia xyli subsp. cynodontis (Lxc) as an endophytic bacterium in rice, we constructed an Escherichia coli-Lxc shuttle vector, pLGUS, containing a beta-glucuronidase reporter gene, which was stable both in vitro and in vivo. Lxc grows and expresses the beta-glucuronidase reporter gene in all parts of rice, except for seed. A 2-year field study using three rice varieties from China showed that Lxc inoculation did not have a negative effect on the growth and yield of any of these varieties. Therefore, Lxc has the potential to be used as a benign endophyte for the expression of foreign genes in rice.

Unlabelled: Natural IgM can recognize apoptotic cells, but the molecular structure and the role in macrophage phagocytosis of apoptotic cells remain unclear., Objectives: (1) To examine the binding of previously isolated natural IgM (3B4) to apoptotic cells and its effects on phagocytosis of apoptotic cells. (2) To characterize the molecular structure of 3B4., Methods: 3B4 binding to apoptotic thymocytes was examined by flow cytometry. Polyreactivity of 3B4 was assayed by ELISA. PKH26-labeled Macrophages were incubated with PKH67-stained apoptotic cells in the presence of 3B4. Macrophages phagocytosis of apoptotic cell was evaluated by flow cytometry. The DNA segments of 3B V(H) and V(K) were sequenced and analyzed., Results: 3B4 IgM recognized late apoptotic cells. Polyreactive-recognitions of lysophosphatidylcholine (LPC) as well as some autoantigens were observed in 3B4. Phagocytosis of late apoptotic cells was increased in the presence of 3B4. The V(H) and V(K) genes of 3B4 showed a germline gene context, while N-sequences and nucleotide loss were observed in CDR3., Conclusion: 3B4 promotes macrophage phagocytosis of late apoptotic cells in a complement-independent process. 3B4 has a germline configuration and is possibly ligand-selected. Out experiments suggest an independent role of natural IgM as opsonin in clearance of late apoptotic cells.

In the present study, HPLC-ESI-IT (ion-trap) MS was used for carboxyterminal (C-terminal) amino acid sequence confirmation of intact recombinant Hirudin Variant 3 (HV3) and alkylated HV3. The C-terminal amino acid sequence of HV3 was determined by the use of carboxypeptidase P (CPP), and by the combined use of carboxypeptidase P and carboxypeptidase Y (CPY). The C-terminal amino acid sequence of alkylated HV3 with 1,4-dithiothreitol (DTT) reduction was also confirmed by the combined use of CPP and CPY (abbreviated to CPP-CPY). Up to 19 amino acid residues were confirmed in the nanomolar concentration range by analyzing the molecular weights of the truncated peptides of HV3. Another five amino acids were confirmed in the nanomolar concentration range of alkylated HV3 with DTT reduction. For sequencing alkylated HV3 with DTT reduction, HV3 reduced with DTT followed by alkylation with iodoacetamide. The reaction mixture, which included alkylated HV3, DTT, and iodoacetamide, was then directly sequenced without any further pre-treatment. The reaction was designed in a time-, and concentration-dependent manner to obtain the maximum sequence information. The results showed that HPLC-ESI-ITMS cannot only determine the C-terminal amino acid sequence of HV3, but also gives important information about the enzymatic degradation and subsequent release of the C-terminal amino acids of HV3.

Most adenoviruses bind to the coxsackie- and adenovirus receptor (CAR). Surprisingly, CAR is not expressed apically on polarized cells and is thus not easily available to viruses. Consequently, alternative mechanisms for entry of coxsackievirus and adenovirus into cells have been suggested. We have found that tear fluid promotes adenovirus infection, and we have identified human lactoferrin (HLf) as the tear fluid component responsible for this effect. HLf alone was found to promote binding of adenovirus to epithelial cells in a dose-dependent manner and also infection of epithelial cells by adenovirus. HLf was also found to promote gene delivery from an adenovirus-based vector. The mechanism takes place at the binding stage and functions independently of CAR. Thus, we have identified a novel binding mechanism whereby adenovirus hijacks HLf, a component of the innate immune system, and uses it as a bridge for attachment to host cells.

Objectives: To study the effects of gonadotropin on the expression of growth differentiation factor-9 (GDF-9) and -9B (GDF-9B) in mouse ovary., Methods: We chose follicles of mature mice cultured in vitro and mature BALB/c mice as our animal models. (1) In vivo experiment. Twenty mice were divided into two groups (groups A and B) randomly with ten mice in each group. Each mouse was injected with 10 IU pregnant mare's serum gonadotropin in group A and saline of the same volume was given to the other group. Twenty-four hours later we obtained ovarian tissue and immunohistochemistry and in situ hybridization were performed to detect the expression of GDF-9 and GDF-9B. (2) In vitro experiment. Fifty-eight separated follicles were divided into two groups (groups C and D) randomly and cultured in vitro. Thirty-two follicles in group C were cultured in medium with follicular stimulating hormone (FSH) for 72 hours while 26 were cultured without FSH. Immunohistochemistry was performed to detect the expression of GDF-9 in both groups., Results: We abserved 370 follicles by immunohistochemistry. The weak positive (+ or ++) rate was 22.0% and 46.2% while the strong positive (+++ or ++++) rate was 22.6% and 9.1% in 186 follicles of group A. The weak positive (+ or ++) rate in group B was 42.4% and 42.9% while the strong positive (+++ or ++++) rate was 14.1% and 0.5%. The expression of GDF-9 was higher in group A than that of group B. The weak positive rate, positive rate and strong positive rate in group C was 28.1%, 53.1% and 18.8% while that in group D was 61.5%, 30.8% and 7.7%, respectively. The expression of GDF-9 was higher in group C than that of group D. We observed 362 follicles by in situ hybridization. The weak positive (+ or ++) rate was 31.5% and 46.2% while the strong positive (+++ or ++++) rate was 21.2% and 1.1% in 184 follicles of group A. The weak positive (+ or ++) rate in group B was 39.9% and 41.6% while the strong positive (+++ or ++++) rate was 15.7% and 2.8%. There were no differences in the expression of GDF-9B between groups A and B., Conclusion: Gonadotropin increases the expression of GDF-9 in vivo while FSH increases GDF-9 in vitro. Gonadotropin has no effects on the expression of GDF-9B.

Objective: To compare the treatment results between radical surgery and late course accelerated hyperfractionated radiotherapy (LCAHFR) for patients with resectable esophageal cancer in the chest., Methods: From June 1998 to September 2002, 269 patients with resectable esophageal cancer in the chest were randomized into two groups: 135 in surgery group and 134 in radiotherapy. The surgery group received esophagectomy including resection of the lesion and 5 cm margin at both ends from the lesion as well as surrounding lymph nodes > or = 5 mm and fatty tissue. In the radiotherapy group: irradiation field for the lesion in the upper esophageal cancer included the gross lesion, bilateral supraclavicular nodes and 4 cm of normal esophagus from lower margin of the gross disease; for the esophageal cancer at the middle segment, it included the gross disease with 4 cm normal esophagus from both ends of the lesion; for the lesion in the lower esophageal cancer, it included 4 cm of normal esophagus and the gross lesion as well as the draining gastric lymph nodes. The width of the irradiation field was 5-6 cm. The 90% isodose volume was covered by the entire CTV with 3-5 beams, in a conventionally fractionated RT at 1.8-2.0 Gy/d for the first two thirds of treatment course to a dose of about 50-50.4 Gy followed by LCAHFR using reduced fields (2 cm extended margin at both ends of the lesion) , twice daily at 1.5 Gy per fraction ( with aminimal interval of 6 h between fractions) to a dose of 18-21 Gy. The total dose whole radiotherapy was 68.4-71.0 Gy., Results: The 1-, 3- and 5-year overall survival rate was 93.3%, 61.5% and 36.9% in the surgery group versus 88.6%, 56.2% and 34.7% in the radiotherapy group without statistical difference between the two groups. The 1-, 3- and 5-year progression free survival rate was 75.9%, 43.7% and 23.1% in the surgery group and 73.3%, 39.7% and 20.6%, respectively, in the radiotherapy group without statistical difference between the two groups either., Conclusion: The results treated by late course accelerated hyperfractionated conformal radiotherapy alone may be comparable to that by radical surgery for patient with resectable esophageal cancer in the chest.

The live vaccine Mycobacterium bovis bacillus Calmette-Guérin (BCG) provides variable efficacy against adult pulmonary tuberculosis (TB). Recombinant BCG, expressing either immunodominant antigens or Th1 cytokines, is a promising strategy for developing a new TB vaccine. However, not much is known about whether the introduction of cytokine and specific antigen genes concurrently into the BCG strain could improve the immunogenicity of BCG. In this study, a recombinant BCG strain (rBCG) expressing the fusion protein human interleukin (IL)-2 and ESAT-6 (early secreted antigenic target-6 kDa) antigen of Mycobacterium tuberculosis was constructed. Six weeks after BALB/c mice (H-2d) were immunized with 106 colony forming units (CFUs) BCG or rBCG, splenocyte proliferation was determined with MTT [3-(4,5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide] assay, IL-4 and interferon (IFN)-gamma produced by splenocytes were tested by enzyme linked immunosorbent assay (ELISA,) and the cytotoxicity of splenocytes from immunized mice to P815 cells (H-2d) expressing ESAT-6 protein was measured using CytoTox 96 Non-Radioactive Cytotoxicity Assay. Compared with native BCG-vaccinated mice, rBCG induced stronger Th1 responses that were confirmed by high lymphoproliferative responses and IFN-gamma production to culture filtrate protein (CFP) or ESAT-6 protein. Moreover, rBCG induced significant enhanced CTL responses against P815-ESAT-6 cells. Results from rBCG-immunized mice demonstrated that introducing the il-2 and esat-6 genes into BCG could enhance Th1 type immune responses to ESAT-6. Further investigation is needed by introducing other Th1 cytokines and antigens into BCG to optimize the protective efficacy against TB.

Objective: To investigate the effects of prenatal stress (PNS) on kinetic properties of high-voltage-activated (HVA) Ca(2+) channel in freshly isolated offspring rat hippocampal CA3 pyramidal neurons., Methods: The pregnant rats were exposed to restraint stress, and the hippocampal CA3 pyramidal neurons were freshly isolated from the offspring rats to record voltage-gate Ca(2+) channel currents in these neurons using whole-cell configuration of patch-clamp technique., Results: PNS significantly increased HVA calcium current density and integral current but did not alter the steady-state activation and steady-state inactivation properties of HVA Ca(2+) channel in the offspring rat CA3 neurons. The maximal HVA calcium current density was -40.89-/+0.31 pA/pF in the control group (n=10) and -49.44-/+0.37 pA/pF in PNS group (n=8, P<0.01). The maximal integral current of the HVA Ca(2+) channel was 106.81-/+4.20 nA*ms in the control group (n=10) and 133.49-/+2.59 nA*ms in the PNS group (n=8, P<0.01)., Conclusion: Maternal exposure to stress during the critical phase of pregnancy may result in long-lasting effects on the ion channels of the hippocampal neurons in the offspring rats.

Aim: To record calcium and potassium currents in acutely isolated smooth muscle cells of mesenteric arterial branches in rats., Methods: Smooth muscle cells were freshly isolated by collagenase digest and mechanical trituration with polished pipettes. Patch clamp technique in whole-cell mode was employed to record calcium and potassium currents., Results: The procedure dissociated smooth muscle cells without impairing the electrophysiological characteristics of the cells. The voltage-gated Ca2+ and potassium currents were successfully recorded using whole-cell patch clamp configuration., Conclusion: The method dissociates smooth muscle cells from rat mesenteric arterial branches. Voltage-gated channel currents can be recorded in this preparation.

To investigate the effects of prenatal stress (PNS) on high-voltage-activated (HVA) Ca2+ channel and delayed rectifier potassium currents (I(KD)) of freshly isolated rat offspring hippocampal CA3 pyramidal neurons, the pregnant rats of PNS group were exposed to restraint stress and patch clamp technique of whole-cell mode was employed to record HVA Ca2+ and K+ channel currents in hippocampal CA3 pyramidal neurons. It was observed that PNS increased HVA calcium peak current amplitude. The maximal HVA calcium peak current amplitudes were -576.52 +/- 7.03 pA in the control group and -702.05 +/- 6.82 pA in the PNS group respectively (P < 0.01). The conductance-voltage relationship of HVA Ca2+ channel was not changed. Current-voltage relationship and conductance-voltage relationship of I(KD) in offspring CA3 neurons were also not affected by PNS. The data suggested that exposure of pregnant animals to a period of stress could impose lasting effects on the offspring hippocampus CA3 neurons Ca2+ channel during a critical phase of fetal development. The mechanism may include two ways. One was that high CORT led to enhance level of expression for the HVA Ca2+ channel subunit mRNAs and increased HVA calcium currents. Another was that PNS caused an increase in the production of ROS in hippocampal CA3 region and ROS caused an increase in the phosphorylation of HVA Ca2+ channel of offspring hippocampal CA3 neurons.

Background: Identification of critical autoantigenic T-cell epitopes is key to developing antigen-based therapies for autoimmune diseases, including psoriasis. Our previous work demonstrated that 3 peptides on keratin 17 are able to stimulate peripheral blood lymphocytes of HLA-DRB1*07-positive patients with psoriasis and to serve as immunodominant T-cell epitopes., Objective: We sought to determine antagonistic altered peptide ligands to psoriatic T cells with a down-modulatory effect in inhibiting keratinocyte proliferation., Methods: Psoriatic altered peptide ligands were generated by single alanine residue substitutions at a critical T-cell receptor contact residue position. Antagonistic altered peptide ligands were identified by suppression screening of psoriatic T-cell activation and keratinocyte proliferation., Results: Altered peptide ligands 119R and 355L can inhibit psoriatic T-cell activation more effectively than other altered peptide ligands, especially 355L, with inhibition of T-cell proliferation and the secretion of interferon gamma and interleukin 2 in parallel with the up-regulation of interleukins 4 and 10 as well as transforming growth factor-beta. In coincubation assay, altered peptide ligands 119R and 355L can down-regulate the function of psoriatic T cells more effectively than wild-type epitopes solely, but less effectively than altered peptide ligands solely. In prepulse assay altered peptide ligand 119R can down-regulate the activation of psoriatic T cells more effectively than in coincubation but less effectively as compared with altered peptide ligand 119R only. Altered peptide ligand 355L was also shown to have a similar presentation. T-cell culture supernatants (1:100) from the concentrations (10 microg.mL(-1) and 100 microg.mL(-1) with 119R, 100 microg.mL(-1) with 355L) were more effective than the other ratios in inhibiting keratinocyte proliferation., Limitations: This study had a relatively small sample size (52 patients and 48 healthy controls)., Conclusion: Our findings show that the altered peptide ligands 119R (VAALEEANTELEVKI) and 355L (ENRYCVQASQIQGLI) are capable of inhibiting proliferative responses of psoriatic T cells and keratinocyte proliferation in vitro, at least, with enhanced helper T cell type 2 polarization. Thus, to our knowledge, this article is the first report of the demonstration of therapeutic activity of altered peptide ligands derived from keratin 17.

Aim: To study the effects of inosine on myosin heavy chain (MHC) activity in rats soleus muscle after tail-suspension., Methods: Weightlessness was simulated by tail-suspension of rats. Using immunohistochemistry technique, changes of expression of MHC in intrafusal and extrafusal fibers in soleus muscle were detected., Results: Expression of fast MHC in intrafusal and extrafusal fibers in soleus muscle increased after 14 d of simulated weightlessness, whereas in tail-suspended rats given inosine, the expression of fast MHC was not detected., Conclusion: Inosine could counteract the changes in expression of MHC in intrafusal and extrafusal fibers of rats soleus under simulated weightless situation.

Background: Heat shock protein 70 (HSP70) is expressed highly in epithelial tumours associated closely with human papillomavirus 16 (HPV16) infections. However, evidence about the direct relationship between HSP70 expression and HPVs infections are still lacking. In the present study, we examined the expression of HSP70 in keratinocytes introduced with HPV16 E6/E7 oncogenes., Methods: Stable transfected cells were established by transfection of the plasmids pLXSN16E6/E7 into cultured primary keratinocytes and subsequently selected by plasmid specific selection antibiotic (G418) at the required concentration. The expression of HSP70 in pLXSN16E6/E7 transfected keratinocytes was determined by Western blot. The correlation of HSP70 expression and E6/E7 transfection was further confirmed by doubly labelled immunofluorescent staining., Results: Compared to non-transfected keratinocytes, there was a significant trend for higher levels of HSP70 in pLXSN16E6/E7 transfected keratinocytes. Doubly labelled immunofluorescent staining experiment showed that the co-localization of HPV16 E6/E7 and HSP70 in transfected keratinocytes was observed and increased expression of HSP70 was strongly associated with the transfection of HPV16 E6/E7., Conclusions: Our studies demonstrated increased levels of HSP70 proteins in keratinocytes stably transfected by HPV16 E6/E7 oncogenes. It suggests that the expression of HSP70 is modulated by HPV16 E6/E7 proteins, which may be involved in HPV16 E6/E7 induced immortalization.

The present paper establishes the optimization criteria for achieving the optimum measurements in three-band pyrometry under the assumption of linear emissivity model and the sensor design with Gaussian distribution. By varying the FWHM of Gaussian distribution, the optimization analysis is investigated with the choice of the wavelength bandwidth in three-band pyrometry. For specific cases, the simulation results present the optimum bandwidth distribution. Therefore, the discussions in this paper will provide significant theoretical instructions for the design of radiation pyrometer.