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Biomedical subjects

H Ji

Publications and source records attributed to H Ji.

At least 109 records · Page 6Linked to original sources
PubMed1997

Identification of a breast cancer-specific gene, BCSG1, by direct differential cDNA sequencing.

A high-throughput direct-differential cDNA sequencing approach was employed to identify genes differentially expressed in normal breast as compared with breast cancer. Approximately 6000 expressed sequence tags (ESTs) from cDNA libraries of normal breast and breast carcinoma were selected randomly and subjected to EST-sequencing analysis. The relative expression levels of more than 2000 unique EST groups were quantitatively compared in normal versus cancerous breast. Of many putative differentially expressed genes, a breast cancer-specific gene, BCSGC1, which was expressed in high abundance in a breast cancer cDNA library but scarcely in a normal breast cDNA library, was identified as a putative breast cancer marker. In situ hybridization analysis demonstrated stage-specific BCSG1 expression as follows: BCSG1 was undetectable in normal or benign breast lesions, showed partial expression in ductal carcinoma in situ, but was expressed at an extremely high level in advanced infiltrating breast cancer. The predicted amino acid sequence of BCSG1 gene has a significant sequence homology to the non-amyloid beta protein fragment of the Alzheimer's disease amyloid protein. BCSG1 overexpression may indicate breast cancer malignant progression from benign breast or in situ carcinoma to the highly infiltrating carcinoma.

Amino Acid Sequence↗
PubMed1997

The human A33 antigen is a transmembrane glycoprotein and a novel member of the immunoglobulin superfamily.

The mAb A33 detects a membrane antigen that is expressed in normal human colonic and small bowel epithelium and > 95% of human colon cancers. It is absent from most other human tissues and tumor types. The murine A33 mAb has been shown to target colon cancer in clinical trials, and the therapeutic potential of a humanized antibody is currently being evaluated. Using detergent extracts of the human colon carcinoma cell lines LIM1215 and SW1222, in which the antigen is highly expressed, the molecule was purified, yielding a 43-kDa protein. The N-terminal sequence was determined and further internal peptide sequence obtained following enzymatic cleavage. Degenerate primers were used in PCRs to produce a probe to screen a LIM1215 cDNA library, yielding clones that enabled us to deduce the complete amino acid sequence of the A33 antigen and express the protein. The available data bases have been searched and reveal no overall sequence similarities with known proteins. Based on a hydrophilicity plot, the A33 protein has three distinct structural domains: an extracellular region of 213 amino acids (which, by sequence alignment of conserved residues, contains two putative immunoglobulin-like domains), a single hydrophobic transmembrane domain, and a highly polar intracellular tail containing four consecutive cysteine residues. These data indicate that the A33 antigen is a novel cell surface receptor or cell adhesion molecule in the immunoglobulin superfamily.

Amino Acid Sequence↗
PubMed1997

Inherited mutations in PTEN that are associated with breast cancer, cowden disease, and juvenile polyposis.

PTEN, a protein tyrosine phosphatase with homology to tensin, is a tumor-suppressor gene on chromosome 10q23. Somatic mutations in PTEN occur in multiple tumors, most markedly glioblastomas. Germ-line mutations in PTEN are responsible for Cowden disease (CD), a rare autosomal dominant multiple-hamartoma syndrome. PTEN was sequenced from constitutional DNA from 25 families. Germ-line PTEN mutations were detected in all of five families with both breast cancer and CD, in one family with juvenile polyposis syndrome, and in one of four families with breast and thyroid tumors. In this last case, signs of CD were subtle and were diagnosed only in the context of mutation analysis. PTEN mutations were not detected in 13 families at high risk of breast and/or ovarian cancer. No PTEN-coding-sequence polymorphisms were detected in 70 independent chromosomes. Seven PTEN germ-line mutations occurred, five nonsense and two missense mutations, in six of nine PTEN exons. The wild-type PTEN allele was lost from renal, uterine, breast, and thyroid tumors from a single patient. Loss of PTEN expression was an early event, reflected in loss of the wild-type allele in DNA from normal tissue adjacent to the breast and thyroid tumors. In RNA from normal tissues from three families, mutant transcripts appeared unstable. Germ-line PTEN mutations predispose to breast cancer in association with CD, although the signs of CD may be subtle.

Adenoma↗
PubMed1997

Two-site immunofluorometric assay of intact salmon calcitonin with improved sensitivity.

We recently developed a two-site immunofluorometric assay (IFMA) of salmon calcitonin (SCT) by DELFIA (dissociation enhancement lanthanide fluoroimmunoassay) technique using the same polyclonal antibodies both for "catching" the antigen and for signaling. In the present study we used a monoclonal antibody to SCT 1-11 as the capture antibody. This antibody was biotinylated before use in streptavidin-coated microtitration plates. The polyclonal antibody labeled with Eu chelate was used as a signaling marker. This combination of antibodies resulted in an assay that was three to four times more sensitive than the previous IFMA, with a detection limit of 0.3 pmol/L serum. Intact SCT 1-32 was detected by the assay (recoveries 94-96%), but not the fragments SCT 1-11 and SCT 10-32 or human calcitonin. Dilutions of plasma samples containing SCT were parallel to the calibration curve of SCT 1-32. Pharmacokinetic studies of SCT, 100 IU administered intramuscularly to 10 men, indicated peak serum concentrations of 32-128 pmol/L within 10-20 min with apparent half-life of 56+/-18 min (mean+/-SD). This new assay will allow study of the pharmacokinetics of new calcitonin preparations that do not require injection.

Adult↗
PubMed1997

Quantification of parathyroid hormone-related protein mRNA by competitive PCR and time-resolved lanthanide fluorometry.

Using dissociation and enhancement time-resolved lanthanide fluorometry, we have developed a quantitative competitive (QC)-PCR for measuring parathyroid hormone-related protein (PTHrP) mRNA after reverse transcription. A cloned PTHrP cDNA target was also modified by deletion of 10 bp and insertion of 21 bp in the midregion of the fragment and cloned for use as a competitor (i.e., internal standard). Two primers spanning 362 bp of target and 373 bp of competitor were designed and one of the primers was biotinylated. Two oligonucleotide probes, one recognizing the target and the other hybridizing to the competitor, were labeled with Eu chelate. Two equal aliquots of PCR products were assayed with each probe separately in streptavidin-coated wells. After 35 PCR cycles, the competitor signal decreased exponentially (y = e(3.74 - 0.624x); r2 = 0.965) and the target signal increased exponentially (y = e(1.14 + 0.497x); r2 = 0.984) when 1000 copies/tube of the competitor and 0-100,000 copies/tube of the target DNA were added. Log-transformed data for the ratio of target to competitor signals (y) and the copies of the target DNA added (x) were used for plotting the linear calibration curve (y = 2.79 + 2.76x; r2 = 0.976). This QC-PCR enables analysis of multiple samples simultaneously and can be used to study PTHrP gene expression in malignancy and physiology.

Europium↗
PubMed1997

Microlamellar keratectomy for correction of high myopia.

OBJECTIVE: To evaluate the preliminary results of high myopia correction by microlamellar keratectomy (MLK). METHODS: Seventy-three eyes of 43 patients with high myopia were treated by MLK. The preoperative refraction ranged from -7.00D to -32.00D (mean -18.45D +/- 6.18D). All the operations were performed with the special apparatus for MLK. RESULTS: At 6 months after surgery, in the 73 eyes except 2 with new other symptoms, all the uncorrected visual acuity and corrected visual acuity were better than or equal to their preoperative ones respectively. Uncorrected visual acuity was better than or equal to the preoperative corrected visual acuity in 51 eyes (71.83%). No severe complications occurred during and after operation. The postoperative refractive status was stable. CONCLUSIONS: MLK is effective to reduce high myopia and has some special advantages as compared with radial keratotomy (RK), photorefractive keratectomy (PRK) and laser in-situ keratomileusis (LASIK). But influenced by several factors, the predictability of this procedure is merely fair. It is suggested that this kind of procedure must be done with excellent microsurgery equipments, microsurgical skills and much more clinical experience of managing corneal disorders.

Adolescent↗
PubMed1996

Purification and characterization of a novel restricted antigen expressed by normal and transformed human colonic epithelium.

A cell surface antigen that is expressed by normal and 95% of transformed colonic epithelium and is recognized by the monoclonal antibody A33 (Welt, S., Divgi, C. R., Real, F. X., Yeh, S. D., Garin-Chesa, P., Finstad, C. L., Sakamoto, J., Cohen, A., Sigurdson, E. R., Kemeny, N., Carswell, E. A., Oettgen, H. F., and Old, L. J. (1990) J. Clin. Oncol. 8, 1894-1906) has been purified to homogeneity from the human colonic carcinoma cell line LIM1215. The A33 protein was purified from Triton X-114 extracts of LIM1215 cells under nondenaturing conditions. These extracts were applied sequentially to Green-Sepharose HE-4BD, Mono-Q HR 10/10, Superose 12 HR 10/30, and micropreparative Brownlee Aquapore RP 300. The purification was monitored by biosensor analysis using surface plasmon resonance detection with a F(ab')2 fragment of the humanized A33 monoclonal antibody immobilized on the sensor surface and Western blot analysis following SDS-polyacrylamide gel electrophoresis (PAGE) under nonreducing conditions using humanized A33 monoclonal antibody. The purified A33 antigen has a Mr on SDS-PAGE of 43,000 under nonreducing conditions. By contrast, the purified protein displayed a Mr of approximately 180,000 under native conditions on both size exclusion chromatography and native PAGE, possibly due to the formation of a homotetramer. N-terminal amino acid sequence analysis of the purified protein identified 34 amino acid residues of a unique sequence: ISVETPQDVLRASQGKSVTLPXTYHTSXXXREGLIQWD. A polyclonal antibody was raised against a synthetic peptide corresponding to residues 2-20 of this sequence. The antipeptide serum recognized the purified protein using Western blot analysis under both nonreducing (Mr 43,000) and reducing (Mr 49,000) conditions.

Amino Acid Sequence↗
PubMed1996

Purification of single-stranded M13 DNA by cooperative triple-helix-mediated affinity capture.

A solid-phase triple-helix-mediated affinity capture method is described for the purification of single-stranded M13 DNA for use as template in fluorescence-based DNA sequencing reactions. In this method, a biotinylated polypyrimidine oligonucleotide "loop" bound to streptavidin-coated magnetic beads is used to selectively capture single-stranded M13 DNA from high-titer phage supernatant through the formation of a cooperative triple helix (CTH) complex between the oligonucleotide and a polypurine site previously cloned into the M13 vector. Capture is accomplished at acidic pH to encourage triple-helix formation, while elution is performed at alkaline pH with heating to destroy the CTH complex. The beads can be reused up to three times without probe replenishment. Yields of M13 ssDNA in excess of 1 microgram per milliliter of culture are obtained, sufficient for use as template in fluorescence-based DNA sequencing reactions.

Bacteriophage M13↗
PubMed1996

Rapid restriction mapping of cosmids by sequence-specific triple-helix-mediated affinity capture.

A simple and rapid strategy for restriction mapping based on sequence-specific triple-helix affinity capture (TAC) was developed. The strategy was applied to the analysis of cosmid clones by the construction of a new cosmid vector, ScosTriplex-II, containing two different triple-helix-forming sequences flanking the cloning site of the original SuperCos-1 cosmid vector. For restriction mapping, the recombinant cosmid DNA is digested with NotI restriction enzyme or with one of four intron-encoded endonucleases for excision of intact inserts followed by controlled partial digestion with a mapping enzyme used in conjunction with the corresponding methyltransferase. The partial digestion products are combined with biotinylated triple-helix-forming oligonucleotides to form a triple-helical complex. The triple-helix complexes are immobilized on streptavidin-coated magnetic beads, washed, and eluted with pH 9 buffer solution. The fragments are separated and directly sized by agarose gel electrophoresis. Bidirectional maps are obtained simultaneously by binding to the two different triple-helix-forming oligonucleotides. No probe labeling, gel drying, blotting to membranes, hybridization, or autoradiography is necessary. Also, TAC conditions that permit gel-free isolation of the terminal restriction fragments from cosmid inserts were found. These advantages afforded by ScosTriplex-II should facilitate the automation of cosmid restriction site fingerprinting needed for large-scale mapping and sequencing projects.

Base Sequence↗
PubMed1996

Management of stage-I borderline ovarian tumors.

OBJECTIVE: To evaluate the surgical management and outcome of stage-I borderline ovarian tumors. METHODS: A series of 95 cases of FIGO stage-I borderline ovarian tumors was retrospectively reviewed. RESULTS: The age of the patients ranged from 17.6 to 88.9 years (mean 52.7 years). Serous and mucinous tumors were equally represented (47.5% and 47.4%, respectively), endometrioid and clear cell tumors comprising the remaining three (3.2%) and two (2.1% cases. Extirpative primary surgery was performed on all patients. Three patients received postoperative chemotherapy. Nineteen of the 28 patients (67.9%) who were younger than 40 years underwent conservative surgery in which ovarian tissue and the uterus were preserved in order to maintain reproductive potential, and nine of them had successful pregnancies later. Ninety patients were followed regularly for 0.75-21.3 years (average 7.3 years). Seven patients suffered a tumor relapse (7.8%) within 0.6-4.3 years, and four (57.1%) of them (three with mucinous tumors, one with clear cell tumor) died of the disease within 1-10.8 years. The 5- and 10-year overall survival rates were 97.0% and 86.3%, respectively, and the disease-free survival rates were 90.8% in both 5- and 10-year follow-ups. CONCLUSIONS: Even though the prognosis of surgically treated stage-I borderline ovarian tumors is excellent, mucinous and clear cell subtypes are associated with potential mortality.

Adolescent↗
PubMed1996

Atlantic salmon (Salmo salar) fed L-carnitine exhibit altered intermediary metabolism and reduced tissue lipid, but no change in growth rate.

Metabolic evidence was sought to explain the reduced body fat and increased body protein observed in Atlantic salmon fed diets supplemented with L-carnitine. By stimulating fatty acid oxidation, dietary carnitine might increase flux through pyruvate carboxylase and decrease flux through the branched-chain alpha-keto acid dehydrogenase complex, by increasing regulatory ratios of acetyl CoA:free enzyme A (CoA-SH) and ATP:ADP. Such changes could conserve nitrogen by providing more carbon for amino acid biosynthesis and by blocking oxidative loss of the branched-chain amino acids. Consistent with this hypothesis, salmon fed carnitine (23 mmol/kg diet) for 9 wk exhibited greater metabolic rates than cohorts fed a carnitine-free diet (P < 0.05) for the following: 1) 1-[14C] palmitate oxidation by liver cubes (48%) and by isolated hepatocytes (151%), 2) pyruvate-dependent [14 CO2]-fixation by isolated mitochondria (81%), 3) incorporation of 1-[14C] lactate into glucose by liver cubes (120%) and by isolated hepatocytes (210%), and 4) incorporation of [35S]-methionine into the acid-insoluble fraction of liver cubes (59%) and isolated hepatocytes (89%). Hepatic concentrations of seven amino acids, including the branched-chain amino acids, were greater (7-112%), as were the plasma concentrations of three of these (45-130%). However, 230% more enzyme in the mitochondria of carnitine-fed fish, and not a difference in the ratios of acetyl CoA:CoA-SH or ATP:ADP, appeared to account for accelerated flux through pyruvate carboxylase; flux through the dehydrogenase complex was unchanged. These results implicate induction of pyruvate carboxylase (or a reduction in turnover) and enhanced protein synthesis in the mechanism for carnitine-induced changes in gluconeogenesis and nitrogen metabolism.

Amino Acids↗
PubMed1996

Solid-phase synthesis, metal binding and folding properties of caulimovirus-related 'zinc finger'.

A 17-residue peptide containing the caulimovirus-related "zinc finger' was prepared by solid-phase peptide synthesis. Fluorescence measurements showed that the tryptophan quantum yield was Zn(2+)-dependent, allowing a 1:1 a stoichiometry for the complex to be determined. The structure of the peptide was characterized using circular dichroic spectroscopy, which indicates that the peptide exhibits a random coiled conformation in the absence of zinc but appears to form an ordered structure in the presence of zinc.

Amino Acid Sequence↗
PubMed1996

A long-term culture system for the expansion of hematopoietic stem cells from embryonic yolk sac with the capacity to seed erythroid and lymphoid development in vitro and to reconstitute the lymphoid compartment in severe combined immunodeficient mice.

We have established a long-term culture system under which hematopoietic stem cells derived from the embryonic yolk sac may be maintained for long periods. Evidence for the persistence of stem cells in this culture is provided by three experimental observations. First, erythroid progenitors, as evidenced by morphological, functional, and phenotypical analysis, may be generated from these yolk sac cultures after more than 7 months in culture. The yolk sac derived erythroid progenitors are distinct from those of bone marrow derived cells both qualitatively and quantitatively. This is evidenced by the high colony plating efficiency, large colony size, different growth factor requirements, increased sensitivity to Epo and other cytokines, as well as significant and prolonged expansion capability, suggesting that the yolk sac derived progenitors are both more proliferative and more primitive than their bone marrow derived analogs. Second, under different conditions, lymphoid progenitors may also be derived from these long term yolk sac cultures in the presence of the appropriate cytokines. Third, preliminary data suggest the engraftment of these yolk sac cells and reconstitution of at least some compartments of the hematopoietic system of host animals. This long-term culture system will provide a useful model for the study of early embryonic hematopoiesis, and the cells derived from this culture system may also have the potential of serving as donor cells for hematopoietic cell transplantation.

Animals↗
PubMed1996

[Establishment of anti-HCV NS5 alpha McAb hybridoma cell lines and its application on clinical immunohistochemical diagnosis].

OBJECTIVE: To prepare hybridoma cell lines which secrete anti-HCV recombinant NS5a protein monoclonal antibodies, and to evaluate their usage in investigating the distribution of HCV NS5a antigen in liver tissue. METHODS: The hybridoma cell lines were raised using the conventional protocols. The immunohistochemistry of paraffin embedded liver tissue was performed to determine the distribution of HCV NS5a antigen as well as NS3 antigen in liver tissue. RESULTS: Four hybridoma cell lines, which secrete monoclonal antibodies against HCV NS5a protein, were raised and named as 8B2, 6F11, 4C6 and 7D9. They had specific reaction with HCV NS5a protein and had no reaction with HCV core and NS3 proteins. Immunohistochemistry results indicated that 54.8% of anti-HCV positive patient's liver specimens were positive and 70.6% of active cirrhosis positive. CONCLUSION: The monoclonal antibodies we prepared here are specific to the recombinant HCV NS5 alpha protein and they can be used in the clinical immunohistochemistry diagnosis.

Animals↗
PubMed1996

[The role of substance P in the spinal dorsal horn in the pathogenesis of autoimmune diseases].

The aim of the present study is to explore the role of immunosuppression mediated by substance P (SP) in spinal dorsal horn (SDH) in the pathogenesis of the autoimmune diseases. The experimental allergic neuritis (EAN), experimental allergic encephalomyelitis (EAE) and adjuvant arthritis (AA) animal models were established in the guinea pigs and Wistar rats, respectively. The effects of alteration of SP activity in SDH on immune responses and the pathogenesis of the autoimmune diseases were observed. The results showed that decreasing activity of SP in SDH by pretreatment of capsaicin or intrathecal SP antagonist could enhance cellular and humoral immune responses and aggravate the autoimmune diseases, while intrathecal SP agonist could suppress the immunity and alleviate clinical signs. The contents of SP in SDH was elevated dramatically at the peak of immune responses. These results suggest that SDH SP might participate in the pathogenesis of the autoimmune diseases. The increase of SP contents in SDH may inhibit the immune system via unknown pathway and ease clinical severity of the autoimmune disease, where SP might act as neurotransmitter in the immunoregulation of the negative feedback. To elevate SP content in SDH might be beneficial to the autoimmune diseases.

Animals↗