Biomedical subjects
Publications and source records attributed to C Liu.
Two novel chelate resins containing hydroxy-alkylamine group were prepared. The IR spectra of the above mentioned resins and their middle products were measured, and the major IR bands were assigned by such chemical methods as hydrolysis, getling salts, determination of hydroxyl value, etc., the structure of the resins and their middle products were proved.
We describe the use of an HPLC/MS technique for the characterization of nicked fragments of hCG beta-subunit. After reductive alkylation of the nicked hCG beta-subunit with vinylpyridine, endoproteinase Glu-C or trypsin was used to digest the protein to produce peptides that could be analyzed by HPLC/electrospray ionization MS. Human leukocyte elastase digestion was used to produce an experimentally nicked hCG. Two nicking sites were observed, between amino acids 42Thr and 43Arg and between 44Val and 45Leu. The former site has not been previously reported for elastase digestion. The structures of the fragments were confirmed by HPLC/MS after removal of the oligosaccharide by direct mass measurement and by mass determination of their proteolytic digests. Without the glycopeptidase treatment, the microheterogeneity of the two N-linked oligosaccharides could be deduced from the spectra of the proteolytic fragments. Nicking with elastase was found to alter the oligosaccharide structures. Nicked beta-subunit samples isolated from the urine of choriocarcinoma patients were also analyzed and the location of the nicking site(s) agreed with that determined by classical techniques. Important differences in the oligosaccharide structures were also observed in these samples, including the presence of triantennary oligosaccharides not found in hCG from healthy subjects. These findings demonstrate the potential of HPLC/MS for characterization of glycoprotein standard preparations.
OBJECTIVE: To examine the role of otoacoustic emissions and auditory efferent system function in evaluating auditory functional status in both normal and diseased conditions. METHODS: Spontaneous and transient evoked otoacoustic emissions and efferent function in terms of contralateral white noise induced transient evoked otoacoustic emission suppressions were tested and cross compared in 32 normal ears, 24 ears with cochlear hearing loss and 16 ears with retrocochlear hearing loss. RESULTS: Transient evoked otoacoustic emission amplitude and contralateral suppression, spontaneous otoacoustic emission incidence, peaks and frequency range were significantly reduced in cochlear hearing loss while in retrocochlear hearing loss transient evoked otoacoustic emission and spontaneous otoacoustic emission levels were significantly higher than in cochlear hearing loss group and showed no suppression. CONCLUSIONS: Contralateral acoustic stimulation induced transient evoked otoacoustic emission suppression as an index of efferent function is clinically feasible. Combined otoacoustic emission and efferent tests are of great significance in evaluating cochlea status and auditory central mechanisms. Auditory efferent function is weakened in cochlear lesions and severely damaged in retrocochlear lesions.
OBJECTIVE: To observe the effects of methyl carprost suppository on postpartum bleeding in normal parturients. METHODS: Three hundred and sixty normal parturients were randomized into study group (n = 260) and the control group (n = 100). Parturients in study group were administered methyl carprost suppository tents either vaginally (n = 130) or via the rectum (n = 130). The controls were given 10IU oxytocin immediately after the delivery of anterior shoulder of the fetus either intramuscularly (n = 50) or intravenously (n = 50). The amount of bleeding with in 2 hours postpartum was measured. RESULTS: The mean amount of bleeding for parturients who received vaginal or rectal methyl carprost was 146.4 ml and 134.2 ml, respectively. In contrast, the mean amount of bleeding for parturients who received oxytocin intramuscularly or intravenously was 267.9 ml and 210.6 ml, respectively. The differences in the amount of bleeding between the two groups were statistically significant (P < 0.01), whereas differences between the routes of drug administration within each group were not significant (P > 0.05). CONCLUSION: Methyl carprost suppository is more effective than oxytocin in controlling postpartum bleeding. Drug administration through the rectum is easy to perform and, therefore, suitable for use in rural areas.
OBJECTIVE: To evaluate a new scanning method designed for the diagnosis of early transtentorial hernia as compared with the routine scanning method (parallel to orbitomeatal line). METHODS: 102 patients with unilateral high intracranial pressure and 100 volunteers without any cranial symptoms and signs were scanned on (1) the plane perpendicular to the plane of clivus and (2) the plane parallel to orbitomeatal line, a line joining the apex of the dorsum sellae and the lowest point of the clivus. RESULTS: Vertical-to-clivus scanning method directly demonstrated the temporal transtentorial hernia in 48 of 102 patients with unilateral high intracranial pressure, but the orbitomeatal line scanning method was not able to show this direct finding. The projected tissue of the temporal lobe could be obviously shown and the degree of the hernia could be judged directly on the CT imaging with the vertical-to-clivus scanning method. Therefore, the CT diagnosis criterion of the hernia was suggested as follows: the hernia would be determined if the downward shift of the temporal lobe was more than 3 mm and suspected if less than 3 mm with ipsilateral high intracranial pressure. CONCLUSION: This new method presented a superiority in showing the temporal transtentorial hernia for the patients with a cranial emergency.
Explore the source record for details and available documents.
This retrospective study involved 34 patients with multiple mandibular fractures. Monocortical miniplates were used for fixation of fractures followed by a short term intermaxillary fixation. All the patients were evaluated radiographically and clinically. The results indicated that all fractures healed uneventfully. No malocclusion, delayed union, injury of the nerves or teeth due to the procedure were found. We concluded that monocortical miniplate osteosynthesis is effective and reliable for multiple mandibular fractures, and a short term intermaxillary fixation is helpful for the healing of fracture.
Mycoplasmas were examined by cultured in the urogenital secreta and the midpieceurine from 400 cases (88 men and 312 women) of genitourinary infection and 119 cases (positive rate 29.75%) of genitourinary infection in Wuhan area. The positive rates of ureaplasma urealyticum (Uu) and mycoplasma hominis (Mh) were 24.75% and 8.75% respectively. The authors also studied the infection rates of Neisseria gonorrheae, fungus and trichomonad by cultures and microscopic findings. The positive rates of Neisseria gonorrheae, fungus and trichomonad were 14.50%, 13.50% and 4.50% respectively. Results showed that mycoplasma was one of the common pathogens causing genitourinary infection in Wuhan area.
Comparative electrophoretic, thermal denaturation, and spectroscopic studies with dodecamers of the form d(ATTA-XGCX-TAAT) and their self-complementary counterparts suggest that actinomycin D (ACTD) binds strongly to a 5'GC3' site with flanking T/T mismatches and moderately to that with C/C mismatches but weakly to those with other G/G or A/A mismatches. The relative binding order is found to be T/T > C/C > G/G > A/A. The ACTD binding affinity for the GC site with T/T mismatches is comparable to the strong binding of self-complementary-XGCY-sequences. Both the ACTD association and dissociation kinetics at the GC site with flanking T/T mismatches require two-exponential fits. The slow component of the association rates is slower than those of the self-complementary sequences, whereas that of the dissociation is only slightly faster than that of the -TGCA- sequence. Interestingly, the slow component of dissociation is decidedly slower than those of -AGCT- and -CGCG- sites and is more than an order of magnitude slower than those with C/C, G/G, and A/A mismatches. These kinetic results are further corroborated by fluorescence measurements using 7-amino-ACTD, a fluorescent analog of ACTD. In addition, fluorescence and absorbance spectral characteristics indicate that the binding mode at the GC site with flanking T/T mismatches resembles those of strong-binding self-complementary -XGCY- sites which are known to be intercalative in nature. The observed slow ACTD dissociation at the T/T-mismatched site suggests that the minor-groove environment near the T/T-mismatched pairs provides favorable interactions with the pentapeptide rings of the drug, whereas the others, especially those of bulkier purine/purine mismatches, result in less favorable interactions.
Explore the source record for details and available documents.
A new Clostridium cellulovorans (strain ATCC 35296) endoglucanase gene engF has been isolated and sequenced. The gene contains 1671 bp and codes for a protein containing 557 amino acids and a mass of 60.1 kDa. A putative signal peptide of 29 amino acids is present and the mature protein has a mass of 57.1 kDa. EngF does not have amino acid sequence homology to previously isolated EngB and EngD, but does show sequence homology to family 5 glycosyl hydrolases from Bacillus, Erwinia carotovora, and C. acetobutylicum species. EngF is not a component of the cellulosome and does not contain a duplicated sequence (DS) at its C-terminal region. EngF is capable of binding to cellulose and hydrolyzing carboxymethylcellulose but not xylan. The cellulose binding domain (CBD) differs from types I, II and III CBDs and no obvious homology has been found to other CBD types. The maximum activity of EngF occurs at pH 5.5 and at 47 degrees C. Its properties suggest that EngF plays an ancillary role in the degradation of cellulosic materials.
A procedure for rapid purification of polymerase chain reaction (PCR) products allowing precise molecular weight determination using electrospray ionization-Fourier transform ion cyclotron resonance (ESI-FTICR) mass spectrometry is described. PCR amplification utilized the DNA polymerase from Pyrococcus furiosus (Pfu) which, unlike Taq, does not incorporate a nontemplated terminal deoxyadenosine phosphate. An 89-base pair nucleotide portion of the spacer region between the 16S and 23S ribosomal rRNA genes was amplified from the genome of three members of Bacillus cereus group and a 114 nucleotide region from the Bacillus subtilis. PCR involves polymerization of nucleotide precursors using two oligonucleotide primers and an amplification enzyme, as well as the presence of metal ions. Mass spectrometric analysis greatly benefits from removal of the oligonucleotide primers (15- and 17-mers in this instance) and nucleotide precursors since they adversely affect sensitivity and metal ion adduction results in an inaccurate molecular weight determination. In the presence of guanidinium hydrochloride the PCR products bind preferentially to a silica resin, allowing removal of other components (i.e., dNTP's primers, and salts). Further removal of metal ions was accomplished using a microdialysis device, allowing samples to be pumped through a hollow cellulose fiber with external countercurrent flow of 2.5 mM ammonium acetate. Prior to injection into the mass spectrometer, the sample buffer was adjusted to 50 vol % acetronitrile, 25 mM piperidine, and 25 mM imidazole, which enhanced signal intensity. The molecular weights of the PCR products determined by nucleotide sequence and MS analysis were in excellent agreement, and several PCR products were analyzed where mass differences corresponding to single base substitutions could be accurately assigned. These assignments were possible due to the high mass precision, accuracy, and resolution FTICR inherently affords. This constitutes the first report demonstrating the ionization and detection of PCR products by mass spectrometry with mass precision and accuracy for assignment of such modifications or substitutions.
A major challenge for using native or modified T cell epitopes to induce or suppress immunity relates to poor localization of peptides to antigen presenting cells (APCs) in vivo. In this study, we demonstrate enhanced presentation of antigenic and antagonistic peptides by targeting them to the type I Fc receptor for IgG (F(c)gammaRI, CD64) on human monocytes. A Th epitope of tetanus toxoid, TT830, and the antagonistic peptide for TT830, TT833S, were genetically grafted into the constant region of the heavy chain of the humanized anti-CD64 mAb 22 and expressed as monovalent fusion proteins, Fab22-TT830 and Fab22-TT833S. These CD64-targeted peptides were up to 1,000- and 100-fold more efficient than the parent peptides for T cell stimulation and antagonism, respectively, suggesting that such fusion proteins could effectively increase the delivery of peptides to APCs in vivo. Moreover, the F(c)gammaRI-targeted antagonistic peptide inhibited proliferation of TT830-specific T cells even when APCs were first pulsed with native peptide, a situation comparable with that which would be encountered in vivo when attempting to ameliorate an autoimmune response. These data suggest that targeted presentation of antagonistic peptides could lead to promising Ag-specific therapies for T cell-mediated autoimmune diseases.
The biosynthesis of retinoic acid from beta-apocarotenoic acids was examined for a beta-oxidation-like process using both rabbit liver mitochondrial fractions with various beta-apocarotenoic acids (beta-apo-14'-, beta-apo-12'-, and beta-apo-8'-carotenoic acid) and perfusion in ferret liver through the portal vein with beta-apo-8'-carotenoic acid. The in vitro incubation of beta-apo-8', beta-apo-12'-, and beta-apo-14'-carotenoic acids gave rise to shorter chain beta-apocarotenoic acids as well as retinoic acid. The rate of retinoic acid synthesis from 10 microM beta-apo-8', beta-apo-12'-, and beta-apo-14'-carotenoic acids was 11 +/- 2, 18 +/- 3, and 30 +/- 7 pmol/h/mg of protein, respectively. The stepwise oxidation of beta-apocarotenoic acid in mitochondria was dose-related to both protein concentration and substrate concentration. beta-Apocarotenoic acid oxidation was inhibited in a dose-dependent manner when it was co-incubated with oleoyl-CoA. The in vivo perfusion of ferret liver with beta-apo-8'-carotenoic acid resulted in a linear increase in the retinoic acid concentration of bile, which was completely abolished by co-perfusion of 3-mercaptopropionic acid, an inhibitor of long chain acyl-CoA dehydrogenase, and partially inhibited by 2-tetradecylglycidic acid, an inhibitor of carnitine-palmitoyl-CoA transferase I. However, the formation of retinoic acid from the beta-apocarotenoic acids was not inhibited, either in vitro or in vivo, by citral, an inhibitor of retinal oxidase. Thus, the formation of retinoic acid was not occurring by the central cleavage pathway. These data suggest that the oxidation of intermediate compounds between beta-carotene and retinoic acid may undergo a type of beta-oxidative process to form retinoic acid, which is reminiscent of mitochondrial fatty acid beta-oxidation. This pathway may play an important role in the biosynthesis of retinoic acid from beta-carotene.
The early growth response 1 (EGR-1) gene product is a transcription factor with role in differentiation and growth. We have previously shown that expression of exogenous EGR-1 in various human tumor cells unexpectedly and markedly reduces growth and tumorigenicity and, conversely, that suppression of endogenous Egr-1 expression by antisense RNA eliminates protein expression, enhances growth, and promotes phenotypic transformation. However, the mechanism of these effects remained unknown. The promoter of human transforming growth factor beta 1 (TGF-beta 1) contains two GC-rich EGR-1 binding sites. We show that expression of EGR-1 in human HT-1080 fibrosarcoma cells uses increased secretion of biologically active TGF-beta 1 in direct proportion (rPearson = 0.96) to the amount of EGR-1 expressed and addition of recombinant human TGF-beta 1 is strongly growth-suppressive for these cells. Addition of monoclonal anti-TGF-beta 1 antibodies to EGR-1-expressing HT-1080 cells completely reverses the growth inhibitory effects of EGR-1. Reporter constructs bearing the EGR-1 binding segment of the TGF-beta 1 promoter was activated 4- to 6-fold relative to a control reporter in either HT-1080 cells that stably expressed or parental cells cotransfected with an EGR-1 expression vector. Expression of delta EGR-1, a mutant that cannot interact with the corepressors, nerve growth factor-activated factor binding proteins NAB1 and NAB2, due to deletion of the repressor domain, exhibited enhanced transactivation of 2- to 3.5-fold over that of wild-type EGR-1 showing that the reporter construct reflected the appropriate in vivo regulatory context. The EGR-1-stimulated transactivation was inhibited by expression of the Wilms tumor suppressor, a known specific DNA-binding competitor. These results indicate that EGR-1 suppresses growth of human HT-1080 fibrosarcoma cells by induction of TGF-beta 1.
A major limitation of electrospray ionization mass spectrometry (ESI-MS) for oligonucleotide analysis arises due to sodium adduction, a problem that increases with molecular weight. Sodium adduction can preclude useful measurements when limited sample sizes prevent off-line cleanup. A novel and generally useful on-line microdialysis technique is described for the rapid (approximately 1-5 min) DNA sample cleanup for ESI-MS. Mass spectra of oligonucleotides of different size and sequence showing no significant sodium adduct peaks were obtained using the on-line microdialysis system with sodium chloride concentrations as high as 250 mM. Signal-to-noise ratios were also greatly enhanced compared to direct infusion of the original samples. By using ammonium acetate as the dialysis buffer, it was also found that the noncovalent association of double-stranded oligonucleotides could be preserved during the microdialysis process, allowing analysis by ESI-MS.
Two types of interleukin (IL)-1 receptors with three extracellular immunoglobulin-like domains, limited homology (28%), and different pharmacological characteristics termed type I and type II have been cloned from mouse and human cell lines. Both receptors exist in transmembrane and soluble forms; the soluble IL-1 receptor is thought to be post-translationally derived from cleavage of the extracellular portion of the membrane receptors. In preliminary cross-linking studies with radiolabeled IL-1, we found that monkey kidney COS1 cells express a soluble receptor with molecular mass of approximately 55-60 kDa, which is different from previously reported soluble IL-1 receptors. This soluble IL-1 receptor protein from COS1 cells was purified to homogeneity by affinity chromatography using recombinant IL-1beta as the ligand and shown to have an affinity for human 125I-IL-1beta (KD approximately 2-3 nM) comparable to the human type II IL-1 receptor (IL-1RII). The purified protein was microsequenced, and the sequence information was used to design primers to clone the COS1 IL-1RII using reverse transcription-coupled polymerase chain reaction; the DNA comparison with monkey COS1 and human IL-1RII indicate that they are 95% identical at the nucleic acid and amino acid levels. In addition, another cDNA, which represents an alternatively processed mRNA of the IL-1RII gene, was also cloned both from monkey COS1 and human Raji cells and was shown to have approximately 95% sequence identity between these species. While the cDNA of the novel alternatively processed gene has a 5' end identical to the IL-1RII, the 200 base pairs at the 3' end are different and the sequence predicts a soluble IL-1 receptor protein of 296 amino acids. Radioligand binding studies of the alternatively processed IL-1RII mRNA demonstrated kinetic and pharmacological characteristics similar to the known type II IL-1 receptor. COS7 cells (which lack IL-1 receptor) transfected with the transmembrane form of the human IL-1RII cDNA showed 125I-IL-1beta binding in both the membrane fractions and supernatant. In contrast, COS7 cells transfected with the alternatively processed human IL-1RII cDNA showed high affinity 125I-IL-1beta binding (Ki approximately 1.2 nM) predominantly in the supernatant; a very small amount of detectable membrane IL-1 binding activity was also observed presumably due to association of the soluble IL-1 receptor and membrane-integrated proteins. In cross-linking and ligand blot studies, the alternatively processed human IL-1RII cDNA-transfected COS7 cells expressed a soluble IL-1 receptor with molecular masses ranging from 60 to 160 kDa, further indicating the association between this soluble IL-1 receptor and other soluble proteins. In summary, we report the purification and characterization of a soluble IL-1 receptor expressed by COS1 cells and the cloning of an alternatively processed type II IL-1 receptor mRNA from both human and COS1 cells. The alternative splicing of a primary transcript leading to a secreted protein provides a potentially important mechanism by which soluble IL-1RII can be produced.
The maytansinoid drug DM1 is 100- to 1000-fold more cytotoxic than anticancer drugs that are currently in clinical use. The immunoconjugate C242-DM1 was prepared by conjugating DM1 to the monoclonal antibody C242, which recognizes a mucin-type glycoprotein expressed to various extents by human colorectal cancers. C242-DM1 was found to be highly cytotoxic toward cultured colon cancer cells in an antigen-specific manner and showed remarkable antitumor efficacy in vivo. C242-DM1 cured mice bearing subcutaneous COLO 205 human colon tumor xenografts (tumor size at time of treatment 65-130 mm3), at doses that showed very little toxicity and were well below the maximum tolerated dose. C242-DM1 could even effect complete regressions or cures in animals with large (260- to 500-mm3) COLO 205 tumor xenografts. Further, C242-DM1 induced complete regressions of subcutaneous LoVo and HT-29 colon tumor xenografts that express the target antigen in a heterogeneous manner. C242-DM1 represents a new generation of immunoconjugates that may yet fulfill the promise of effective cancer therapy through antibody targeting of cytotoxic agents.