BACKGROUNDNowadays it is now a focus topic in schistosomiasis research to find ideal vaccine candidates and new drug targets for developing anti-schistosomiasis vaccine. We cloned a new gene, casein kinase II beta subunit, of Schistosoma japonicum (S. japonicum) and express it in Escherichia coli (E. coli).

METHODSThe ESTs obtained in our laboratory were analyzed by homologous searching, and a new gene was recognized. The full-length cDNA of the new gene was obtained by joining the 3'RACE PCR fragment and the EST clone. To express the new gene, the cDNA was cloned into pGEX-4T-1 vector and then transformed into E. coli JM109. The recombinant protein was analyzed by SDS-PAGE and Western-blot.

RESULTSA 908 bp cDNA was isolated from S. japonicum and identified to be casein kinase II beta subunit gene by sequence analysis. The open reading frame of the gene encodes a protein of 217 amino acids exhibiting 75.8%, 75.8%, 73.9%, 68.2%, 51.6% identity to the amino acids sequence of the corresponding genes of Homo sapiens (H. sapiens), Xenopus laevi (X. laevi), Drosophila melanogaster (D. melanogaster), Caenorhabditis elegan (C. elegan), and Schizosaccharomyces pombe (S. promber) respectively. The predicted molecular weight of the protein was 24.921 kDa. The new cDNA sequence had been submitted to GenBank, and its accession number is AY241391. This cDNA was subcloned into the pGEX-4T-1 vector and expressed in E. coli JM109. The recombinant protein could be recognized by the S. japonicum infected rabbit serum.

CONCLUSIONThe full-length cDNA sequences encoding S. japonicum casein kinase II beta subunit were firstly sequenced, cloned, and expressed in E. coli.

Amino Acid Sequence ; Animals ; Base Sequence ; Blotting, Western ; Casein Kinase II ; chemistry ; genetics ; Cloning, Molecular ; DNA, Complementary ; chemistry ; isolation & purification ; Escherichia coli ; genetics ; Molecular Sequence Data ; Rabbits ; Schistosoma japonicum ; enzymology ; genetics

OBJECTIVETo compare changes of hypothalamus-pituitary-adrenal axis (HPAA) in different rat models of Gan stagnation (GS), Pi deficiency (PD), Gan stagnation Pi deficiency (GSPD) syndromes, and to observe interventional effect of Chaishu Sijun Decoction (CSD, capable of soothing Gan-qi invigorating Pi) on them.

METHODSSeventy Wistar rats were divided into the normal control group (group 1), the GS group (group 2), the PD group (group 3), the GSPD group (group 4), the GS intervention group (group 5), the PD intervention group (group 6), and the GSPD intervention group (group 7) according to random digit table, 10 in each group. Rats in group 1 received no treatment. Rats in group 2 and 5 were modeled by chronic restraint method. Rats in group 3 and 6 were modeled by excess fatigue plus alimentary abstinence method. Rats in group 4 and 7 were modeled by chronic restraint, excess fatigue, and alimentary abstinence method. At the 2nd weekend of modeling, CSD at 2.86 g/kg was fed to rats in group 5, 6, and 7 by gastrogavage for 2 successive weeks. Equal volume of distilled water was given to rats in the rest 4 groups. On the 29th day, rats were killed, adrenal weight weighed, and adrenal index calculated. Levels of plasma and hypothalamus corticotropin-releasing hormone (CRH), plasma and pituitary adrenocorticotrophic hormone (ACTH), and plasma corticosterone (CORT) were determined using radioimmunity.

RESULTSCompared with group 1, adrenal index significantly decreased in group 2, 3, and 4 (P < 0.05). Of them, plasma and hypothalamus CRH, plasma CORT increased significantly in group 2 and 4 (P < 0.05). Besides, plasma and pituitary ACTH increased in group 4 (P < 0.05). Plasma and pituitary ACTH, as well as plasma CORT decreased significantly in group 3 (P < 0.05). Compared with group 2, 3, and 4, adrenal index increased significantly in group 5, 6, and 7 (P < 0.05). Compared with group 2, plasma CORT, hypothalamus CRH, and pituitary ACTH decreased significantly in group 5 (P < 0.05). Compared with group 3, plasma ACTH and CORT increased significantly in group 6 (P < 0.05). Compared with group 4, plasma CRH, ACTH, CORT, hypothalamus CRH, and pituitary ACTH decreased in group 7 (P < 0.05).

CONCLUSIONSThe function of HPA .axis was damaged to varying degrees in rats of the three models in this experiment. Hyperactivity of HPA axis existed in GS syndrome and GSPD syndrome. Impairment of feedback regulation in hypothalamus and pituitary was accompanied in GSPD syndrome. Hypofunction of HPA axis existed in PDS. CSD, capable of soothing Gan-qi invigorating'Pi, showed improvement on disarranged HPAA, but with optimal effect on GSPD syndrome. CSD had higher correlation with GSPD syndrome.

Adrenocorticotropic Hormone ; metabolism ; Animals ; Corticosterone ; Corticotropin-Releasing Hormone ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Hypothalamo-Hypophyseal System ; metabolism ; Hypothalamus ; metabolism ; Medicine, Chinese Traditional ; Models, Animal ; Pituitary Gland ; metabolism ; Pituitary-Adrenal System ; metabolism ; Rats ; Rats, Wistar

Contraction of smooth muscle cells is triggered by an increase in cytosolic Ca(2+) upon agonist stimulation. Ca(2+) influx across the plasma membrane constitutes a major component of the agonist-induced response in smooth muscle cells. Traditionally, voltage-operated Ca(2+) channel (VOCC) is considered as the channel mediating the Ca(2+) entry. However, this view has been challenged by recent discoveries, which demonstrated that other types of ion channels, such as store-operated and/or receptor-operated Ca(2+) channels (SOCC and/or ROCC), also participate in Ca(2+) response induced by agonists in smooth muscle cells. SOCC is defined as the channel activated in response to the depletion of the internal Ca(2+) stores, an event secondary to G protein coupled receptor or receptor tyrosine kinase stimulation. The Ca(2+) flow mediated by SOCC is termed as capacitative Ca(2+) entry (CCE). Previous study from other group has demonstrated that VOCC played a predominant role in ACh-induced contraction of distal colon smooth muscle in guinea pig. However, whether SOCC participates in the agonist-induced contractile response in this particular tissue is unknown. The present study was performed to investigate the role of CCE in ACh-induced mechanical activity of distal colon smooth muscle in rats. The contractile function of the smooth muscle was assessed by measuring isometric force of isolated rat distal colon rings. We showed that both high extracellular K(+) (40 mmol/L) and ACh (5 mumol/L) evoked striking contraction of the smooth muscle. The contractile responses were almost abolished by removal of extracellular Ca(2+) with ethylene glycol-bis(2-aminoethylether)-N,N,N',N' tetraacetic acid (EGTA), suggesting a critical contribution of extracellular source of Ca(2+) to the contraction. Verapamil (5 mumol/L), an L-type VOCC blocker, significantly attenuated, but didn't completely eliminate the high K(+)- and ACh-induced contraction (74% and 41% for high K(+) and ACh, respectively), indicating that additional channels might be involved in the contractile mechanism. Furthermore, ACh only induced transient contractions in the absence of extracellular Ca(2+). Readmission of Ca(2+) into the extracellular compartment resulted in a significant and sustained increase in the tension of the smooth muscle. This response was not affected by verapamil (5 mumol/L) and Cd(2+) (5 mumol/L), both of which efficiently block VOCC at the doses. However, La(3+), a known inhibitor of SOCC, significantly suppressed the Ca(2+) readdition-induced contraction in a dose-dependent manner. On the basis of these results, we conclude that contraction of smooth muscle in the distal colon is regulated by multiple Ca(2+) channels. In addition to VOCC-mediated Ca(2+) influx, SOCC-mediated CCE participates in agonist-induced contractile response of distal colon smooth muscle in rats.
Acetylcholine ; physiology ; Animals ; Calcium ; metabolism ; Calcium Channels ; physiology ; Colon ; physiology ; Female ; Male ; Muscle Contraction ; physiology ; Muscle, Smooth ; physiology ; Myocytes, Smooth Muscle ; physiology ; Rats ; Rats, Sprague-Dawley ; Verapamil ; pharmacology

OBJECTIVEThe aim of this paper was to investigate the factors associated with viral response and HBeAg seroconversion and the relationship between them at different stages of interferon treatment in HBeAg-positive chronic hepatitis B patients.

METHODSPEG-IFN alfa-2a was injected subcutaneously in doses of 180 microg once a week for 48 weeks to HBeAg-positive chronic hepatitis B patients, and the patients were followed for another 24 weeks after the treatment. The serum HBV DNA load was measured by real-time quantitative PCR assay. Microparticle enzyme immunoassay analysis (MEIA) was then carried out by an automatic enzyme immunoassay analysis instrument to measure HBeAg and anti-HBe. Virological response and HBeAg seroconversion rates, and the factors associated with them were analyzed.

RESULTSThe differences in ALT baselines between viral responding and non-responding groups were significant at treatment time and at the end of the follow-up period. These differences were also significant in patients with HBeAg seroconversion at 12 weeks and at the end of the follow-up period compared with the non-conversion group. No significant difference of HBV DNA baseline was observed between the HBeAg seroconversion and non-conversion group. At 12, 24 and 48 weeks, in patients with viral response during the treatment, their HBeAg seroconversion rates were 43.8%, 21.4% and 18.9% respectively; their respective HBeAg seroconversion rates remaining at 72 weeks were 42.9%, 33.3% and 27.6%. HBeAg seroconversion was related to HBV DNA negativity at 48 weeks treatment in the multivariate analysis (OR=2.15, 95.0% CI=1.744-2.664, P less than 0.01).

CONCLUSIONSViral response and early and sustained HBeAg seroconversion were associated with pretreatment ALT levels. HBeAg seroconversion was related to viral response during IFN treatment, but not to the baseline HBV DNA load.

Adolescent ; Adult ; Antiviral Agents ; therapeutic use ; Female ; Hepatitis B e Antigens ; blood ; Hepatitis B virus ; drug effects ; Hepatitis B, Chronic ; blood ; drug therapy ; virology ; Humans ; Interferon-alpha ; therapeutic use ; Male ; Middle Aged ; Polyethylene Glycols ; therapeutic use ; Recombinant Proteins ; Young Adult

Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) is the most abundant kinase within excitatory synapses in the mammalian brain. It interacts with and phosphorylates a large number of synaptic proteins, including major ionotropic glutamate receptors (iGluRs) and group I metabotropic glutamate receptors (mGluRs), to constitutively and/or activity-dependently regulate trafficking, subsynaptic localization, and function of the receptors. Among iGluRs, the N-methyl-D-aspartate receptor (NMDAR) is a direct target of CaMKII. By directly binding to an intracellular C-terminal (CT) region of NMDAR GluN2B subunits, CaMKII phosphorylates a serine residue (S1303) in the GluN2B CT. CaMKII also phosphorylates a serine site (S831) in the CT of α-amino-3-hydroxy-5- methylisoxazole-4-propionic acid receptors. This phosphorylation enhances channel conductance and is critical for synaptic plasticity. In addition to iGluRs, CaMKII binds to the proximal CT region of mGluR1a, which enables the kinase to phosphorylate threonine 871. Agonist stimulation of mGluR1a triggers a CaMKII-mediated negative feedback to facilitate endocytosis and desensitization of the receptor. CaMKII also binds to the mGluR5 CT. This binding seems to anchor and accumulate inactive CaMKII at synaptic sites. Active CaMKII dissociates from mGluR5 and may then bind to adjacent GluN2B to mediate the mGluR5-NMDAR coupling. Together, glutamate receptors serve as direct substrates of CaMKII. By phosphorylating these receptors, CaMKII plays a central role in controlling the number and activity of the modified receptors and determining the strength of excitatory synaptic transmission.
Calcium-Calmodulin-Dependent Protein Kinase Type 2 ; metabolism ; Neuronal Plasticity ; Phosphorylation ; Receptor, Metabotropic Glutamate 5 ; metabolism ; Receptors, Metabotropic Glutamate ; metabolism ; Receptors, N-Methyl-D-Aspartate ; metabolism ; Serine ; metabolism ; Synapses ; Synaptic Transmission

Animals ; Female ; Liver ; blood supply ; Male ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase Type III ; metabolism ; Rats ; Rats, Wistar ; Reperfusion Injury ; metabolism ; Sex Factors

OBJECTIVETo study the diagnostic value of color Doppler ultrasonography(CDUS) and nocturnal electrobioimpedance volumetric assessment (NEVA) in the assessment of erectile dysfunction (ED) and in differentiating the causes of ED.

METHODSCDUS and NEVA were performed in the 45 patients with ED. The patients were classified into 3 groups according to their results of CDUS, and compared all parameters of NEVA between each two groups, and then studied the correlation between CDUS and NEVA in the assessment of ED.

RESULTSIn the non-vasculogenic ED group, 17 (94.4%) patients had normal nocturnal penile tumescence (NPT); and in contrast, there were 9(75.0%) and 8(72.7%) patients with abnormal NPT in the arteriogenic and venogenic ED groups, respectively. Except that the blood volume change of penis in the venogenic ED group was significantly lower than that in the non-vasculogenic ED group (P = 0.033), there were no significant difference in the other parameters of NEVA between each two groups.

CONCLUSIONThe results of NEVA are well correlated with the functions of artery and venous which were indicated by CDUS. NEVA can indicate the causes of ED to some extent.

Adolescent ; Adult ; Electric Impedance ; Erectile Dysfunction ; diagnosis ; diagnostic imaging ; physiopathology ; Humans ; Male ; Middle Aged ; Ultrasonography, Doppler, Color

A headspace solid-phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) method was developed and optimized for qualitative and semi quantitative analysis of volatile components from Schisandra propinqua, a kind of Yi Nationality herb medicine. This method was used for analysis and evaluation of volatile components from S.propinqua from four different geometrical origins (Yunnan Wuding, Yunnan Luoping, Guizhou Qingzhen and Hubei Shennongjia). 51, 53, 52， 50 compounds were identified from the above four kinds of samples respectively, in which 46 volatile components were contained in all of these four origins. The volatile components in these samples from different geographical origins were then compared by using principal component analysis, cluster analysis and other chemometrics methods. The results suggested that the analysis of volatile components can be used to distinguish the S.propinqua from four origins, and those samples from close geographical origins were more similar in volatile components. The developed method was stable, reliable, and suitable for rapid analysis of volatile components from S.propinqua, providing reference for quality control, drug development and scientific utilization of the herb.

BACKGROUNDChromosomal abnormalities have been shown to play an important prognostic role in multiple myeloma (MM). Interphase fluorescence in situ hybridization (i-FISH) has been much more effective to identify cytogenetic aberrations in MM than conventional cytogenetic technique (CC). To clearly determine the cytogenetic features of Chinese MM patients and identify their prognostic implications, we designed a multicenter study based on i-FISH including 672 patients from 52 hospitals in China.

METHODSAll 672 patients were systematically screened for the following genomic aberrations: del(13q), IgH rearrangement, del(p53) and 1q21 amplifications.

RESULTSThe analysis showed that the chromosomal changes were detected in 22.1% patients by CC and in 82.3% patients by i-FISH. The most common abnormalities by CC were chromosome 1 aberrations (48.4%), -13/13q- (37.6%), hyperdiploidy (36.6%), hypodiploidy (30.1%) and IgH rearrangements (23.7%). The most frequent abnormalities by FISH was del(13q), which was found in 60.4% patients, whereas IgH rearrangement, 1q21 amplification and p53 deletions were detected in 57.6%, 49.0% and 34.7% cases, respectively. By statistical analysis, -13/13q- by CC was associated with low level of platelet (P = 0.015), hyperdiploidy was associated with low level of serum albumin (P = 0.028), and IgH rearrangement by FISH was associated with high level of β2 microglobulin (P = 0.019). Moreover, 1q21 amplification and del(p53) by FISH conferred a high incidence of progressive disease (PD) after initial therapy. Metaphase detection of IgH rearrangements and chromosome 1 aberrations concurrently was associated with a short progression free survival (PFS) (P = 0.036). No significant prognostic implications of other cytogenetic abnormalities were found associated with overall survival and PFS.

CONCLUSIONSChinese MM patients had similar cytogenetic abnormalities compared with the previous reported studies. However, the prognostic significance of FISH aberrations were not clearly determined and further study is required.

Adult ; China ; Chromosome Aberrations ; Chromosomes, Human, Pair 1 ; genetics ; Cytogenetic Analysis ; Female ; Humans ; In Situ Hybridization, Fluorescence ; Karyotyping ; Male ; Middle Aged ; Multiple Myeloma ; genetics ; pathology

Objective To observe the morphological changes in the testes and ovaries of adult 12th-generation Oncomelania hupensis bred for 12 winters in Weishan Lake areas. Methods The offspring of the adult O. hupensis snails bred in the Weishan Lake that were originated from the Yangzhou section of the Yangtze River was defined as the experiment group, while uninfected, adult O. hupensis snails captured from the marshland of the Yangzhou section of the Yangtze River served as the control group. Snails were dissected and intact testicular and ovarian specimens were sampled, routinely fixed, dehydrated, embedded, polymerized in an oven and sliced on an ultramicrotome. The sections were visualized under a transmission electron microscope, and the ultrastructure of the snail gonad was compared between the experiment and control groups. Results Transmission electron microscopy showed “9 + 2” microtubules on the transverse sections of the tails of sperm cells in the testes of male snails in the control group, with triangular acrosomes and spiral, dense nuclei seen in the tip, while in the experiment group, the “9 + 2” microtubules disappeared on the transverse sections of the tails of sperm cells in the testes of male snails, with low chromatin density found in the tip. Transmission electron microscopy revealed clear nucleolus and nuclear membranes in the ova of female snail ovaries, and displayed yolk body, liposomes and endoplasmic reticulum in the cytoplasm, bilayer twists of nuclear membrane and a uniform nucleolus in the control group, while in the experiment group, smooth nuclear membrane and unclear nucleolus were observed in the ova of female snail ovaries, with few contents seen within cells. Conclusions Following breeding for 12 winters in the Weishan Lake, the 12th-generation O. hupensis snails fail to fully adapt to the natural environment in northern China, and the remarkable changes in the ultrastructure of the snail gonad may be a cause of gradual decline and even extinction of O. hupensis in the Weishan Lake areas.