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H Chao

Publications and source records attributed to H Chao.

At least 37 records · Page 2Linked to original sources
PubMed1999

Persistent expression of canine factor IX in hemophilia B canines.

We previously demonstrated that direct intramuscular injection of recombinant adeno-associated virus (rAAV) carrying the human FIX (hFIX) cDNA can safely be administered to hemophilic B canines and express human factor IX protein; however, the functional activity of the hFIX protein could not be assessed due to anti-human FIX antibody (inhibitor) formation. To test the therapeutic efficacy of rAAV in hemophilic dogs, rAAV type 2 (rAAV2) carrying canine FIX (cFIX) cDNA was injected into the skeletal muscle of two dogs at doses of 1012-13particles. Circulating cFIX protein levels were maintained for 1 year at levels of 1-2% of normal. Hemostatic correction (WBCT and APTT) paralleled plasma FIX antigen levels. Both dogs still required plasma infusion for spontaneous and traumatic bleeding events. Inhibitors to cFIX protein were not detected in either animal by Bethesda assay. Neutralizing antibodies directed against AAV-2 capsid were pronounced and persistent. Vector DNA and mRNA transcripts were detected only at the injected skeletal muscle tissue. Analysis of both high and low molecular weight DNA identified both replicative episomal and integrated AAV species. These results demonstrate that persistent secretion of the FIX transgene protein, necessary for successful gene therapy of hemophilia B, can be achieved using the parvovirus-based rAAV vector

Animals↗
PubMed1999

Malignant mixed mesodermal tumors of the ovary: preoperative diagnosis.

Early diagnosis of malignant mixed mesodermal tumors of the ovary is very difficult because of the rarity and the insidious onset. The purpose of this report is to review the magnetic resonance imaging features of an ovarian malignant mixed mesodermal tumor, which occurred in a 52-year-old woman, aiding in the differential diagnosis.

Chemotherapy, Adjuvant↗
PubMed1999

[Metastatic tumors of the vulva: a report of 78 cases].

OBJECTIVE: To study the clinical characteristics, diagnosis and reasonable therapy for metastatic tumors of vulva. METHODS: Seventy eight cases with metastatic tumors of the vulva were retrospectively reviewed. The cervical carcinoma was the most frequent primary site (61 cases), followed by the urethra (5 cases), vagina (4 cases), endometrium (3 cases), breast (2 cases), ovary (1 cases), rectum carcinoma (1 cases) and malignant lymphoma (1 cases). RESULTS: The median age of this series was 55. 75.6%(59/78) of the patients subsequently developed vulvar metastasis from 15 days to 18 years after the primary tumors diagnosed, the median interval was 2.5 years. 61.5%(48/78) patients combined with primary site recurrence or other place metastasis, primary site recurrence were 35.4%, inguinal lymph node metastasis 45.8%, supraclavicular lymph node metastasis 8.3%, pelvic metastasis 8.3% and hematogenous metastasis 12.5% (liver, lung, skeleton). Radiotherapy was the main therapeutic approch, some cases combined with surgery and/or chemotherapy. The overall 1 year, 3 years and 5 years survival rates after vulvar metastatic occurrence were 65.4%, 29.5% and 16.7% respectively. CONCLUSIONS: The prognosis of the vulva metastatic tumors was poor. Radiotherapy combine with surgery and/or chemotherapy can improve the survival.

Adult↗
PubMed1999

Evi-1 and MDS1-Evi-1 genes in pathogenesis of myelodysplastic syndromes and post-MDS acute myeloid leukemia.

OBJECTIVE: To investigate expression of Evi-1 and MDS1-Evi-1 genes in myelodysplastic syndromes (MDS) and post-MDS acute myeloid leukemia (post-MDS AML), and its role in pathogenesis or progression of MDS and post-MDS AML. METHODS: Expression of Evi-1 and MDS1-Evi-1 genes was examined in 31 MDS, 11 post-MDS AML, and 34 de novo AML patients by a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR). RESULTS: Evi-1 expression was not detected in bone marrow samples of 8 normal controls, but low MDS1-Evi-1 expression levels (MDS1-Evi-1/GAPDH < 0.1) were detected in 3 of the 8 controls. Evi-1 RNA was expressed in 1 of 8 RA, 8 of 13 RAEB and 6 of 9 RAEB-T patients, and the percentage of Evi-1 expression in RAEB(T) patients was higher than that in RA (P < 0.05). MDS1-Evi-1 expression was detected in 5 of 8 RA, 9 of 13 RAEB and 5 of 9 RAEB-T patients, and MDS1-Evi-1 expression levels (MDS1-Evi-1/GAPDH > 0.1) were markedly higher than those in the controls. Evi-1 expression was gradually increased in 4 of 5 RAEB-T patients with transformation from MDS to AML. The percentages of Evi-1 and MDS1-Evi-1 expression in post-MDS AML patients were significantly (P < 0.01 and P < 0.05 respectively) higher than those in de novo AML. The colonies of hematopoietic progenitor cells were decreased in Evi-1 and MDS1-Evi-1-positive MDS patients as compared with those in Evi-1 and MDS1-Evi-1-negative patients. CONCLUSION: Abnormal expression of the Evi-1 gene and overexpression of MDS1-Evi-1 gene may play a role in the pathogenesis or progression of MDS and post-MDS AML.

Adult↗
PubMed1998

Use of a heterodimeric coiled-coil system for biosensor application and affinity purification.

The two-stranded alpha-helical coiled-coil is now recognized as one of nature's favorite ways of creating a dimerization motif. Based on the knowledge of protein folding studies and de novo design model systems, a novel heterodimeric coiled-coil protein was synthesized. The heterodimeric E/K coiled-coil was constructed with two distinct peptides (E and K) that will spontaneously associate into a full helical coiled-coil structure in solution. Equilibrium CD, NMR and real time biosensor kinetics experiments showed that the E/K coiled-coil is both structurally (deltaG(unfold)=11.3 kcal/mol) and kinetically (Kd approximately 1 nM) stable in solution at neutral pH. The engineered coiled-coil had been applied as a dimerization and capture domain for biosensor based applications and used in an expression/detection/affinity chromatography system. Specific test examples demonstrated the usefulness of the E/K heterodimeric system in these applications. The universality of coiled-coil as a dimerization motif in nature and our ability to design and synthesize these proteins suggest a wide variety of applications.

Amino Acid Sequence↗
PubMed1998

Binding of an oligopeptide to a specific plane of ice.

The alpha-helical antifreeze protein (AFP) from winter flounder inhibits ice growth by binding to a specific set of pyramidal surface planes that are not otherwise macroscopically expressed. The 37-residue AFP contains three 11-amino acid repeats that make a stereo-specific fit to the ice lattice along the <01-12> direction of the (20-21) and equivalent binding planes. When the AFP was shortened to delete two of the three 11-amino acid ice-binding repeats, the resulting 15-residue peptide and its variants were less helical and showed no antifreeze activity. However, when the helicity of the peptide was reinforced by an internal lactam bridge between Glu-7 and Lys-11, the minimized AFP was able to stably express the pyramidal plane (20-21) on the surface of growing ice crystals. This dynamic shaping of the ice surface by a single ice-binding repeat provides evidence that AFP adsorption to the ice lattice is not an "all-or-nothing" interaction. Instead, a partial interaction can help develop the binding site on ice to which the remainder of the AFP (or other AFP molecules) can orient and bind.

Antifreeze Proteins↗
PubMed1997

Effect of mobile phase on the oligomerization state of alpha-helical coiled-coil peptides during high-performance size-exclusion chromatography.

Important structural motifs involving amphipathic helices include two-stranded and multiple-stranded coiled-coils. High-performance size-exclusion chromatography (HPSEC) is a useful tool to examine both the oligomerization state of coiled-coils as well as the stability of such motifs, due to the facile manipulation of the mobile phase and the lack of interaction of the peptide solutes with the stationary phase. In the present study, HPSEC was applied to two series of de novo designed model amphipathic alpha-helical peptides with the sequences (1) Ac-(E-A-L-K-A-E-I)n-E-A-C-K-A-amide, where n = 1 or 3, Ac-E-I-(E-A-L-K-A-E-I)4-E-A-C-K-A-amide and (2) Ac-(K-L-E-A-L-E-A)n-amide, where n = 1, 2 or 4. Observation of the retention behaviour of Series 1 under both denaturing and non-denaturing conditions at pH 7.0 offered insights into the effect of polypeptide chain length and disulphide bridge formation on the stability of alpha-helical coiled-coils. In contrast, the Series 2 peptides showed promise as peptide standards to monitor the effect of environment on the multi-strandedness of coiled-coils, since the 28-residue peptide of this series was eluted as a monomer, dimer or trimer depending on mobile phase conditions.

Amino Acid Sequence↗
PubMed1997

A diminished role for hydrogen bonds in antifreeze protein binding to ice.

The most abundant isoform (HPLC-6) of type I antifreeze protein (AFP1) in winter flounder is a 37-amino-acid-long, alanine-rich, alpha-helical peptide, containing four Thr spaced 11 amino acids apart. It is generally assumed that HPLC-6 binds ice through a hydrogen-bonding match between the Thr and neighboring Asx residues to oxygens atoms on the {2021} plane of the ice lattice. The result is a lowering of the nonequilibrium freezing point below the melting point (thermal hysteresis). HPLC-6, and two variants in which the central two Thr were replaced with either Ser or Val, were synthesized. The Ser variant was virtually inactive, while only a minor loss of activity was observed in the Val variant. CD, ultracentrifugation, and NMR studies indicated no significant structural changes or aggregation of the variants compared to HPLC-6. These results call into question the role of hydrogen bonds and suggest a much more significant role for entropic effects and van der Waals interactions in binding AFP to ice.

Amino Acid Sequence↗
PubMed1997

Multiecho approaches to spectroscopic imaging of the brain.

Spectroscopic imaging (SI) with nuclear magnetic resonance (NMR) is one of the most powerful tools available for studying brain chemistry in vivo. Both proton (1H) and phosphorus (31P) NMR offer valuable biochemical information that can in principle be mapped throughout the entire brain, thereby enhancing our understanding of brain function. With the exception of protons from tissue water and the triglycerides of adipose tissue, however, nuclei contributing to the NMR signals of living tissue are in relatively small (millimolar) concentrations. The low concentration of metabolite nuclei reduces the overall sensitivity of conventional SI techniques, making high-quality metabolite mapping a lengthy procedure. This problem has led to the development and testing of nonconventional methods for reducing SI scan times, including techniques based on the collection of multiple spin-echoes. The extent to which multiecho methods can be used to decrease SI scan times and maintain high-quality metabolite mapping depends on several factors. These include the spectral transverse relaxation times, the spectral resolution required, and J-coupling interactions. We have discussed these various technical aspects of multiecho SI methods as applied to 1H and 31P spectroscopic imaging of the living brain.

Animals↗
PubMed1997

Two novel mutations in phenylalanine hydroxylase gene and in vitro expression analysis on mutation Arg252Gln.

We report novel mutations in exon 7 of human phenylalanine hydroxylase (PAH) gene of phenylketonuria (PKU) in southern Chinese, analysed by using PCR-DGGE (denaturing gradient gel electrophoresis), solid phase DNA sequencing and in vitro expression. One of the 2 novel mutations, IVS6nt-1, is an intron-exon junctional mutation which results a splicing defect in mRNA. Arg252Gln is another novel mutation with residual PAH activity only 24% compared to wild type in in vitro mutagenesis and expression in Cos-1 cell. Other 3 known mutations and polymorphism including Arg241Cys, Arg243Gln and Val245Val (GTG to GTA) together with these novel mutations composed the mutational profile of exon 7 in the PAH gene of PKUs in this populations.

Exons↗
PubMed1997

[Preliminary study on oncogene MDM2 in acute leukemias].

OBJECTIVE: To explore the causes of MDM2 gene overexpression. METHODS: The amplification and overexpression of MDM2 gene in 41 acute leukemia patients and two leukemic cell lines (K051 and HL60) were studied by using Southern blot, dot hybridyzation and RT-PCR techniques. RESULTS: Overexpression of MDM2 gene was found in 51.2% (21/41) of the patients and HL60 cell line, but no amplification of the gene was found in all the patients and the two cell lines. CONCLUSION: The level of MDM2 gene expression was related to the prognoses of the patients but not to FAB subtypes of acute leukemias.

Actins↗
PubMed1997

[Clonal analysis of blood cells using X-linked HUMARA gene polymorphism].

OBJECTIVE: To explore the method for clonal analysis of blood cells. METHODS: Thirty female AML patients and 20 normal female subjects were analysed by the analysis of X-linked HUMARA gene polymorphism and methylation pattern. RESULTS: The heterozygote rate of HUMARA gene in Chinese women was 88% and the gene had a stable methylation pattern. Out of 20 control females, 17 were heterozygotes and among them 15 were polyclonal, 2 (11.8%) showed skewed X-inactivation. All heterozygote AML were clonal hematopoiesis. CONCLUSION: HUMARA gene polymorphism is a good marker for clonality analysis of blood cells.

Adult↗
PubMed1997

[A clinical analysis of 153 uterine sarcomas].

OBJECTIVE: To evaluate the prognostic factors and treatment methods of 153 uterine sarcomas. METHODS: 153 cases of the uterine sarcoma were eligible for this retrospective study. Of the 153 cases, 48 were leiomyosarcomas, 47 mixed mesodermal sarcomas, 37 endometrial stromal sarcomas, 8 carcinosarcomas, 4 sarcoma botryoides, 1 fibrosarcoma, and 8 malignant lymphomas. 81 cases were in stage I, 11 stage II, 33 stage III and 11 stage IV. 38 cases were treated by surgery alone, 24 by surgery combined with radiation therapy, 50 by surgery plus chemotherapy, 23 by surgery plus radiation therapy and chemotherapy, 4 by radiation therapy alone, 3 by chemotherapy alone, and 11 by radiation therapy plus chemotherapy. RESULTS: The overall 5-year survival rate was 49.0%, and that of leiomyosarcomas, mixed mesodermal sarcomas and endometrial stromal sarcomas was 46.9%, 34.1% and 69.3% respectively (P < 0.01). The 5-year survival rate of lesions limited to the uterus (stage I + II), and that of pelvic cavity invasion (stage III) and distant metastases was 59.6%, 25.6% and 10.0% respectively (P < 0.01. When the uterus was smaller than a 3 months pregnant uterus, the 5-year survival rate was 49.9%. When the uterus size larger than a 3 months pregnant uterus, the survival rate was 18.8% (P < 0.05). Premenopausals surviving 5-year accounted for 56.3% and post-menopausal 28.9% (P < 0.01). CONCLUSIONS: The prognosis of uterine sarcoma is significantly associated with histologic type, clinical and surgico-pathological stage, uterine size and pre- or post-menopausal status. Radiation or chemotherapy alone is palliative. Postsurgical adjuvant radiotherapy significantly decreased vaginal and pelvic recurrences rates. A combination of surgery, radiotherapy and chemotherapy can reduce pelvic recurrence as well as enhance survivals.

Adolescent↗
PubMed1996

NMR characterization of side chain flexibility and backbone structure in the type I antifreeze protein at near freezing temperatures.

The flexibility of the polar side chains in the alpha-helical Type I antifreeze protein (AFP) near the solution freezing temperature was investigated by two-dimensional nuclear magnetic resonance spectroscopy. These experiments were conducted to define the rotameric conformations of the proposed ice-binding groups, threonines and asparagines, in order to probe the molecular mechanism for ice binding. On the basis of the 3J alpha beta 2 NMR coupling constant values of 7.1, 8.5, 8.5, and 6.8 Hz for residues T2, T13, T24, and T35, respectively, it can be calculated that the regularly spaced ice-binding threonines sample many possible rotameric states prior to ice binding. The lack of a dominant side chain rotamer is further corroborated by nuclear Overhauser distance measurements for T13 and T24. N16 and N27, both with 3J alpha beta 2 and 3J alpha beta 3 coupling constants of 8.4 and 4.5 Hz, respectively, show a slight preference for the side chain conformation with a chi 1 of -60 degrees. These data suggest that prior to ice binding the threonine and asparagine side chains are free to rotate and that a unique preformed ice-binding structure in solution is not apparent. These observations do not support the rigid side chain model proposed recently by an X-ray study [Sicheri, F., & Yang, D. S. C. (1995) Nature 375, 427-431].

Amino Acid Sequence↗
PubMed1996

Structural basis for the binding of a globular antifreeze protein to ice.

Antifreeze proteins (AFPs) have the unique ability to adsorb to ice and inhibit its growth. Many organisms ranging from fish to bacteria use AFPs to retard freezing or lessen the damage incurred upon freezing and thawing. The ice-binding mechanism of the long linear alpha-helical type I AFPs has been attributed to their regularly spaced polar residues matching the ice lattice along a pyramidal plane. In contrast, it is not known how globular antifreeze proteins such as type III AFP that lack repeating ice-binding residues bind to ice. Here we report the 1.25 A crystal structure of recombinant type III AFP (QAE isoform) from eel pout (Macrozoarces americanus), which reveals a remarkably flat amphipathic ice-binding site where five hydrogen-bonding atoms match two ranks of oxygens on the [1010] ice prism plane in the <0001> direction, giving high ice-binding affinity and specificity. This binding site, substantiated by the structures and properties of several ice-binding site mutants, suggests that the AFP occupies a niche in the ice surface in which it covers the basal plane while binding to the prism face.

Animals↗
PubMed1996

Kinetic study on the formation of a de novo designed heterodimeric coiled-coil: use of surface plasmon resonance to monitor the association and dissociation of polypeptide chains.

The surface plasmon resonance (SPR) technique was used to study the formation kinetics of a de novo designed coiled-coil (E/K coil). The E/K coil is made up of two distinct peptides (E and K) each with five heptad (g-a-b-c-d-e-f) repeats. The E peptide's heptad sequence is E-V-S-A-L-E-K, and the K peptide's heptad sequence is K-V-S-A-L-K-E. A linker C-nL-G-G-G (nL = norleucine) is present at the C-terminus of the E peptide and at the N-terminus of the K peptide for the SPR studies. Heterodimer formation involves both electrostatic and hydrophobic interactions at the dimer interface. Under conditions that favor the heterodimer formation, the CD signal ([theta]222) varied as a function of peptide concentration. The estimated dissociation constant (Kd) was 2.45 +/- 0.71 nM. Denaturation studies with guanidine-HCI (GdnHC11/2 = 3.9 M) suggested a value of 3.53 +/- 0.48 nM. For the SPR investigation, the peptides were biotinylated and linked to streptavidin in order to increase their effective molecular weight and consequently enhance the signal intensity. Biotinylation in itself did not impede coiled-coil formation based on CD measurements. The biosensor study revealed a slow dissociation rate constant for the heterodimer (kd approximately 2 x 10(-4) s-1) and a moderately fast association rate constant [ka approximately (4.27-4.53) x 10(5) M-1 s-1). This gives a calculated Kd of 0.47-0.50 nM, which agrees reasonably well with the equilibrium CD studies. Therefore, based on the SPR data, the preference for heterodimer formation is due to a combination of moderately fast association and slow dissociation rates.

Amino Acid Sequence↗
PubMed1996

A natural variant of type I antifreeze protein with four ice-binding repeats is a particularly potent antifreeze.

A 4.3-kDa variant of Type I antifreeze protein (AFP9) was purified from winter flounder serum by size exclusion chromatography and reversed-phase HPLC. By the criteria of mass, amino acid composition, and N-terminal sequences of tryptic peptides, this variant is the posttranslationally modified product of the previously characterized AFP gene 21a. It has 52 amino acids and contains four 11-amino acid repeats, one more than the major serum AFP components. The larger protein is completely alpha-helical at 0 degree C, with a melting temperature of 18 degrees C. It is considerably more active as an antifreeze than the three-repeat winter flounder AFP and the four-repeat yellowtail flounder AFP, both on a molar and a mg/mL basis. Several structural features of the four-repeat winter flounder AFP, including its larger size, additional ice-binding residues, and differences in ice-binding motifs might contribute to its greater activity. Its abundance in flounder serum, together with its potency as an antifreeze, suggest that AFP9 makes a significant contribution to the overall freezing point depression of the host.

Amino Acid Sequence↗