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Biomedical subjects

X He

Publications and source records attributed to X He.

At least 127 records · Page 7Linked to original sources
PubMed2001

[The mutation of anti-CD3 antibody (HIT3a) gene and its expression].

OBJECTIVE: To improve the expression of anti-CD(3) single chain Fv (scFv) by site mutation and identify its biological activity. METHODS: Anti-CD(3) scFv gene was mutated by PCR, the target clones were screened by both the fingerprints of DNA restriction endonuclease digestion and Western blot, the antigen-binding activity of scFv was examined by FACS, competitive inhibition was performed with (125)I-labeled HIT3a and the cytotoxic effect mediated by the anti-CD(3) scFv-activated T lymphocytes was analyzed by (51)Cr-released assays. RESULTS: The DNA sequencing showed that the 6th amino acid of the anti-CD(3) antibody (HIT3a) heavy chain gene was mutated from E (GAG) to Q (CAG). The expression of mutated anti-CD(3) scFv (m2) was increased by 100 times higher than that of the parent scFv, and there was no difference in the Jurkat cell (CD(3)(+))-binding activity between the (m2) and parent scFv. The preliminary results of competitive assays showed that m2 could partially block the sites of CD(3)(+) Jurkat cells where the parent antibody bound to. Cytotoxicity assays demonstrated that CD(3)AK cells induced by IL-2 and m2 showed stronger cytotoxic effect than that of LAK cells induced by IL-2 alone in vitro. CONCLUSION: By site mutation, a high expression fragment m2 of anti-CD(3) scFv antibody was obtained. The results of some experiments indicated that m2 could bind to CD(3)(+) Jurkat cells, furthermore, by co-stimulated with IL-2, it could activate peripheral T lymphocytes and induce CD(3)AK cytotoxic effect.

CD3 Complex↗
PubMed2001

[Detection of aflatoxin-albumin adducts in human serum and its application].

Exposure to aflatoxin B1 (AFB1) is one of the major risk factors in the etiology of human hepatocellular carcinoma (HCC). The development of highly sensitive and specific methods for detecting AFB1 and its macromolecular biomarkers are critical for identifying individuals at risk for hepatocellular carcinoma. The detection of aflatoxin-albumin adducts (AFB-ALB) in serum has been used to investigate the relationship between aflatoxin exposure and HCC. A more sensitive and reliable method to detect AFB-ALB in serum was modified by separating serum albumin with Microcon-50 microconcentrators in stead of using precipitation. The method could separate serum albumin quickly and make protein digested in the same container. Thus, the step for transferring samples was reduced and recovery rate was improved. The AFB-ALB adducts were then determined by radioimmunoassay. The AFB-ALB adducts of human serum samples from Qidong, Jiangsu Province, and Fusui, Guangxi Zhuang Autonomous Region of the People's Republic of China were detected. The results indicated that the exposure of AFB1 is common and the positive rate for AFB-ALB adducts in serum of these populations is high.

Aflatoxin B1↗
PubMed2001

[Cloning and expression of D-arabitol dehydrogenase gene from Acetobacter suboxydans in Escherichia coli].

The partial genomic library of Acetobacter suboxydans was constructed using Yeast-E. coli shuttle plasmid YEp352 as vector. Two positive transformants, designated as DH5 alpha(pAD91) and DH5 alpha(pAD98), were obtained by screening the growth of transformants on the agar plate in which D-arabitol was used as the sole carbon source. The results of Southern blot and restriction endonuclease analysis showed that the two recombinants are identical. The insert is about 2.3 kb. Arabitol dehydrogenase activity assay indicated that the transformants could produce D-xylulose-forming D-arabitol dehydrogenase. Hence, the gene encoding D-arabitol dehydrogenase exists in the cloned DNA fragment.

Acetobacter↗
PubMed2001

[Expression of alpha-acetolactate decarboxylase gene from Bacillus subtilis in Brewer's yeast].

Yeast YIp-type expression recombinant plasmid(pCMA2-1) was constructed. The expression of alpha-acetolactate decarboxylase gene from Bacillus subtilis was controled by CUP1 promoter and its own terminator. The recombinant plasmid pCMA2-1 was introduced into the brewer's yeast PJ3-5. Transformants were selected using copper resistance as selected marker. The results of activity assay showed that PJ3-5 didn't produce alpha-acetolactate decarboxylase(ALDC), where as the activity of alpha-ALDC in transformants were induced by copper sulfate. The laboratory scale fermentation test confirmed that the total diacetyl concentration was reduced effectively by alpha-ALDC in transformant.

Bacillus subtilis↗
PubMed2001

[Effect of manganese exposure on brain development in postnatal mice].

Effects of manganese (Mn) exposure on brain development in postnatal mice were studied. The results were as follows: (1) The gain of body weight, brain weight and the ratio of brain weight to body weight in the high dose Mn-exposed group was significantly decreased. (2) In the Morris Water Maze Test, the average latency discovering the hidden platform was obviously increased every day during the first 5 days in the high Mn-exposed group, but the latency probing the quadrant in which the hidden platform was located previously was reduced at the 6th day. (3) The protein content of brain tissue was significantly decreased in the high Mn-exposed mice, but the activity of AChE in brain was decreased. (4) The immunoreactivity of glial fibrillary acid protein (GFAP) and the average relative density of GFAP-positive products in hippocampus (area CA3) of both low and high Mn-exposed group, especially the high dose ones, was significantly higher than that of the control group.

Acetylcholinesterase↗
PubMed2001

Clinical characteristics and basic research development of Peutz-Jeghers syndrome.

OBJECTIVE: To improve clinical knowledge of Peutz-Jeghers syndrome. METHODS: Eight patients with Peutz-Jeghers syndrome from 1984 to 1998 in our hospital were retrospectively reviewed and analyzed in the present study. RESULT: The result of this analysis showed that there were 4 patients appeared with family histories of Peutz-Jeghers syndrome. All of the included patients admitted to the hospital with various complications, and eventually received surgical interventions for these complications, among which, 6 of them had intestinal obstructions mostly (5/6) due to small bowel intussusception, and 2 of them suffered with hemafecia. Post-operative recoveries were generally satisfactory with zero mortality. CONCLUSION: Peutz-Jeghers syndrome is an uncommon digestive dominant hereditary disease. The diagnosis of it with history, symptoms, signs, and proper examinations usually is not difficult Surgical interventions are necessary once complications occur.

Adolescent↗
PubMed2001

[Spectrophotometric determination of trace copper in electroplating wastewater].

In the presence of CPB and OP, the chromogenic reaction of DBH-PF with Cu(II) was studied. The results show that the purple micelle reaction has high sensitivity in the medium of (CH2)6N4-HCl(pH = 5.4) buffered solution, the molar absorptivity is 1.8 x 10(5) L.mol-1.cm-1, and Beer's law is obeyed for Cu(II) in the range of 0.0-9.0 micrograms.25 mL-1 at 540 nm. Combined with solvent extraction separation, the method has been applied to the determination of trace Cu in electroplating wastewater with satisfactory results.

Copper↗
PubMed2001

[Determination of trace manganese in coal gangue by catalytic spectrophotometry].

In HAc-NaAc solution trace Mn(II) catalyzes strongly decolorization reaction of bright green SF(BGSF) by oxidizing with potassium periodate and its catalytic extent is linear with the contents of Mn(II) in the certain range. Based on this study, a catalysis spectrophotometric method for determining trace Mn(II) was developed. The results show that the maximum absorption of the complex is at 651 nm and the detection limits of the method is 0.060 microgram.L-1 for Mn(II) and Beer's law is obeyed for Mn(II) in the range of 0.03-0.3 microgram.50 mL-1. The method has been applied to the determination of trace Mn in coal gangue with satisfactory results.

English Abstract↗
PubMed2001

[T-cell immunity against autologous leukemic cell mediated by in vitro bone marrow-derived dendritic cell from patients with acute myeloid leukemia in complete remission].

OBJECTIVE: To explore if a specific cytolytic T lymphocyte (CTL) response could be in vitro generated by using autogenous-leukemic cells lysates (ACL) pulsed dendritic cells ( DCs ) from AML-CR patients. METHODS: T-cell depleted bone marrow mononuclear cells (TD-BMNC) were isolated from AML-CR patients with E-rosetting procedure, and then were cultured in vitro in the presence of combined cytokines ( GM-CSF, IL-4, SCF/TNF-alpha) for generation of mature DCs, and pulsed with ACL on day 5. After 7-day culture, the cells were harvested and the expression of mature DC marker was determined by FACS. For generation of a specific CTL to the respective AML cells, the cells were co-cultured with pre-activated auto-T lymphocytes by McAb anti-CD3 in the presence of low concentration of IL-2 for 7 days. Cytotoxicity was determined with LDH release. RESULTS: Cultured TD-BMNCs from 12 AML-CR patients developed morphologic and phenotypic characteristics of mature DCs. CTL assay was performed in 6 out of the 12 samples. At effector/target ratio of 20:1, auto-T lymphocytes primed with ACL pulsed DC exhibited significant killing activity to auto-AML cells but not to K562 cells as compared with that stimulated by IL-2 alone or primed by non-pulsed DC (P = 0.001). CONCLUSION: AML cell associated antigen specific CTL responses can be in vitro generated by priming auto-T lymphocytes with ACL pulsed DCs. These findings might prove useful for immunotherapy of AML.

Adolescent↗
PubMed2001

Liver transplantation for patients with hepatitis B virus-related diseases.

OBJECTIVES: To summarize the results of the orthotopic liver transplantation (OLT) for HBV-related diseases and assess the efficiency of lamivudine on preventing HBV recurrence. METHODS: From April 1993 to December 2000, 54 patients with HBV-related liver disease were treated by OLT at the Transplantation Center of Sun Yet-sen University of Medical Sciences (SUMS). Seventeen patients who had postnecrotic cirrhosis (PNC) as a result of HBV infection were designated as group 1; 25 patients coexisting hepatocellular carcinoma (HCC) served as group 2; the remaining 12 patients who had no liver disease but fulminant hepatic failure (FHF) due to HBV infection served as group 3. The survival rate, the causes of early death and the HBV recurrence rate under lamivudine monotherapy were retrospectively analyzed. RESULTS: The overall survival rate within 2 months of transplantation was 75.9% and the morbidity in group 3 was obviously higher than in group 1 and group 2; the outcome patients with small HCC was superior to that of patients with large HCC; lamivudine monotherapy can effectively prevent and treat HBV recurrence while no side effect attributable to it had been identified. CONCLUSION: Combined with the use of lamivudine, liver transplantation can be justified in patients with fulminant hepatitis B, hepatitis B-related liver cirrhosis and coexisting small HCC or even selected large HCC.

Antiviral Agents↗
PubMed2000

Insulin suppresses transactivation by CAAT/enhancer-binding proteins beta (C/EBPbeta). Signaling to p300/CREB-binding protein by protein kinase B disrupts interaction with the major activation domain of C/EBPbeta.

CAAT/enhancer-binding proteins (C/EBPs) play an important role in the regulation of gene expression in insulin-responsive tissues. We have found that a complex containing C/EBPbeta interacts with an insulin response sequence in the insulin-like growth factor-binding protein-1 (IGFBP-1) gene and that a C/EBP-binding site can mediate effects of insulin on promoter activity. Here, we examined mechanisms mediating this effect of insulin. The ability of insulin to suppress promoter activity via a C/EBP-binding site is blocked by LY294002, a phosphatidylinositol 3-kinase inhibitor, but not by rapamycin, which blocks activation of p70(S6 kinase). Dominant negative phosphatidylinositol 3-kinase and protein kinase B (PKB) block the effect of insulin, while activated PKB suppresses promoter function via a C/EBP-binding site, mimicking the effect of insulin. Coexpression studies indicate that insulin and PKB suppress transactivation by C/EBPbeta, but not C/EBPalpha, and that N-terminal transactivation domains in C/EBPbeta are required. Studies with Gal4 fusion proteins reveal that insulin and PKB suppress transactivation by the major activation domain in C/EBPbeta (AD II), located between amino acids 31 and 83. Studies with E1A protein indicate that interaction with p300/CBP is required for transactivation by AD II and the effect of insulin and PKB. Based on a consensus sequence, we identified a PKB phosphorylation site (Ser(1834)) within the region of p300/CBP known to bind C/EBPbeta. Mammalian two-hybrid studies indicate that insulin and PKB disrupt interactions between this region of p300 and AD II and that Ser(1834) is critical for this effect. Signaling by PKB and phosphorylation of Ser(1834) may play an important role in modulating interactions between p300/CBP and transcription factors and mediate effects of insulin and related growth factors on gene expression.

Amino Acid Sequence↗
PubMed2000

On the self-referential nature of naive MHC class II-restricted T cells.

The use of mutant mice expressing a normal MHC class II molecule surface level but a severely restricted self-peptide diversity (H-2Malpha(-/-)) previously revealed that T cells carrying the Ealpha(52-68)-I-A(b) complex-specific 1H3.1 TCR rely on self-peptide(s) recognition for both their peripheral persistence in irradiated hosts and their intrathymic positive selection. Here, we identify Ealpha(52-68) structurally related self-peptide(s) as a major contributor to in vivo positive selection of 1H3.1 TCR-transgenic thymocytes in I-A(b+)/I-Ealpha(-) mice. This is demonstrated by the drastic and specific reduction of the TCR high thymocyte population in 1H3.1 TCR-transgenic (Tg) mice treated with the Ealpha(52-68)-I-A(b) complex-specific Y-Ae mAb. Self-peptide(s) recognition is also driving the maturation of T cells carrying a distinct MHC class II-restricted specificity (the Ealpha(6) alphass TCR), since positive selection was also deficient in Ealpha(6) TCR Tg H-2Malpha(-/-) thymi. Such a requirement for recognition of self-determinants was mirrored in the periphery; Ealpha(6) TCR Tg naive T cells showed an impaired persistence in both H-2Malpha(-/-) and I-A(b)ss(-/-) irradiated hosts, whereas they persisted and slowly cycled in wild-type recipients. This moderate self-peptide(s)-dependent proliferation was associated with a surface phenotype intermediate between those of naive and activated/memory T cells; CD44 expression was up-regulated, but surface expression of other markers such as CD62L remained unaltered. Collectively, these observations indicate that maturation and maintenance of naive MHC class II-restricted T cells are self-oriented processes.

Amino Acid Sequence↗
PubMed2000

Sequence-specific and methylation-dependent and -independent binding of rice nuclear proteins to a rice tungro bacilliform virus vascular bundle expression element.

Nuclear proteins from rice (Oryza sativa) were identified that bind specifically to a rice tungro bacilliform virus promoter region containing a vascular bundle expression element (VBE). One set of proteins of 29, 33, and 37 kDa, present in shoot and cell suspension extracts but hardly detectable in root extracts, bound to a site containing the sequence AGAAGGACCAGA within the VBE, which also contains two CpG and one CpNpG potential methylation motifs. Binding by these proteins was determined to be cytosine methylation-independent. However, a novel protein present in all analyzed extracts bound specifically to the methylated VBE. A region of at least 49 nucleotides overlapping the VBE and complete cytosine methylation of the three Cp(Np)G motifs was required for efficient binding of this methylated VBE-binding protein (MVBP).

5-Methylcytosine↗
PubMed2000

LDL-receptor-related proteins in Wnt signal transduction.

The Wnt family of secreted signalling molecules are essential in embryo development and tumour formation. The Frizzled (Fz) family of serpentine receptors function as Wnt receptors, but how Fz proteins transduce signalling is not understood. In Drosophila, arrow phenocopies the wingless (DWnt-1) phenotype, and encodes a transmembrane protein that is homologous to two members of the mammalian low-density lipoprotein receptor (LDLR)-related protein (LRP) family, LRP5 and LRP6 (refs 12-15). Here we report that LRP6 functions as a co-receptor for Wnt signal transduction. In Xenopus embryos, LRP6 activated Wnt-Fz signalling, and induced Wnt responsive genes, dorsal axis duplication and neural crest formation. An LRP6 mutant lacking the carboxyl intracellular domain blocked signalling by Wnt or Wnt-Fz, but not by Dishevelled or beta-catenin, and inhibited neural crest development. The extracellular domain of LRP6 bound Wnt-1 and associated with Fz in a Wnt-dependent manner. Our results indicate that LRP6 may be a component of the Wnt receptor complex.

Animals↗
PubMed2000

A NK1.1+ thymocyte-derived TCR beta-chain transgene promotes positive selection of thymic NK1.1+ alpha beta T cells.

As a consequence of the peptide specificity of intrathymic positive selection, mice transgenic for a rearranged TCR beta-chain derived from conventional alphabeta T lymphocytes frequently carry mature T cells with significant skewing in the repertoire of the companion alpha-chain. To assess the generality of such an influence, we generated transgenic (Tg) mice expressing a beta-chain derived from nonclassical, NK1.1+ alphabeta T cells, the thymus-derived, CD1. 1-specific DN32H6 T cell hybridoma. Results of the sequence analysis of genomic DNA from developing DN32H6 beta Tg thymocytes revealed that the frequency of the parental alpha-chain sequence, in this instance the Valpha14-Jalpha281 canonical alpha-chain, is specifically and in a CD1.1-dependent manner, increased in the postselection thymocyte population. In accordance, we found phenotypic and functional evidence for an increased frequency of thymic, but interestingly not peripheral, NK1.1+ alphabeta T cells in DN32H6 beta Tg mice, possibly indicating a thymic determinant-dependent maintenance. Thus, in vivo expression of the rearranged TCR beta-chain from a thymus-derived NK1.1+ Valpha14+ T cell hybridoma promotes positive selection of thymic NK1.1+ alphabeta T cells. These observations indicate that the strong influence of productive beta-chain rearrangements on the TCR sequence and specificity of developing thymocytes, which operates through positive selection on self-determinants, applies to both classical and nonclassical alphabeta T cells and therefore represents a general phenomenon in intrathymic alphabeta T lymphocyte development.

Animals↗
PubMed2000

Determinants of recognition of ligands binding to benzodiazepine receptor/GABA(A) receptors initiating sedation.

Complementary behavioral and computational studies of 21 structurally diverse, gamma-amino butyric acid (GABA)(A) benzodiazepine receptor ligands that influence spontaneous locomotor activity have been performed in this work. This behavioral endpoint is a well-accepted indicator of sedation particularly for GABA(A)/benzodiazepine receptor ligands. The goal of the work presented here is the identification and assessment of the minimum requirements for ligand recognition of GABA(A)/benzodiazepine receptors leading to activity at the sedation endpoint embedded in a common 3D pharmacophore for recognition. Using the experimental results, together with a systematic computational procedure developed in our laboratory, a five-component 3D pharmacophore for recognition of the GABA(A) receptor subtypes associated with the sedative behavioral response has been developed consisting of: two proton-accepting moieties, a hydrophobic region, a ring with polar moieties and an aromatic ring in a common geometric arrangement in all ligands having an effect at the sedation endpoint. To provide further evidence that the 3D pharmacophore developed embodied common requirements for receptor recognition, a pharmacophore analysis was performed for agonists, inverse agonists and antagonists separately. Each of the resulting pharmacophores contained the same five moieties at comparable distances to those found for the pharmacophore generated using all of them together. This result confirms that this pharmacophore constitutes a recognition pharmacophore representing required features in the overlapping portion of their binding sites. The reliability of this 3D pharmacophore was then assessed in several ways. First, it was determined that ligands that had no effect at the sedation endpoint did not comply with the pharmacophore requirements. Second, four benzodiazepine receptor ligands known to have an effect at the sedation endpoint, but not used in the pharmacophore development were found to satisfy the requirements of this pharmacophore. Third, the geometric and chemical requirements embedded in this pharmacophore were used to search 3D databases resulting in the identification of benzodiazepine receptor ligands known to affect sedation, but not included in the pharmacophore development. Finally, a 3D-quantitative structure analysis procedure (QSAR) model was developed based upon the ligands in the training set superimposed at their sedation pharmacophore points. The 3D-QSAR model shows good predictivity for binding of these ligands to receptor subtypes containing alpha1 but not alpha5 subunits. The pharmacophore developed for the sedation endpoint thus provides a predictive binding model for diverse ligand binding to alpha1 containing receptor subtypes.

Animals↗
PubMed2000

Transient sister chromatid separation and elastic deformation of chromosomes during mitosis in budding yeast.

The accurate segregation of chromosomes at mitosis requires that all pairs of chromatids bind correctly to microtubules prior to the dissolution of sister cohesion and the initiation of anaphase. By analyzing the motion of GFP-tagged S. cerevisiae chromosomes, we show that kinetochore-microtubule attachments impose sufficient tension on sisters during prometaphase to transiently separate centromeric chromatin toward opposite sides of the spindle. Transient separations of 2-10 min duration occur in the absence of cohesin proteolysis, are characterized by independent motion of the sisters along the spindle, and are followed by the apparent reestablishment of sister linkages. The existence of transient sister separation in yeast explains the unusual bilobed localization of kinetochore proteins and supports an alternative model for spindle structure. By analogy with animal cells, we propose that yeast centromeric chromatin acts as a tensiometer.

Chromatids↗