Biomedical subjects
Publications and source records attributed to G Chan.
In this paper, the contents of total flavonoids, essential oil, tannin, trace elements along with bacteriostatic action in Sophora Powder have been measured. These studies present a scientific evaluation on the quality of Sophora Powder.
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Selective immunoglobulin deficiencies have been shown to be associated with atopic disease. In this study, serum immunoglobulin (Ig) G, A, M, E and IgG subclasses of 92 Asian asthmatic children were studied and compared with those of age-matched controls. The children, aged between 0.7 and 17.4 years (mean age 7.5 years), were recruited from the National University Hospital, Singapore. The serum Ig levels were measured by enzyme-linked immunosorbent assay, except for IgE which was measured by the fluorescent allergosorbent test. As expected, serum total IgE levels were markedly higher in the asthmatic children than in the controls (geometric mean = 513 units/ml and 164 units/ml, respectively; p < 0.0001). Serum IgM levels were also slightly higher in the asthmatic patients than in the controls (geometric mean = 1.74 and 1.51 milligrams, respectively; p < 0.04). Mean serum IgG and A and IgG subclasses (1-4) levels in the asthmatics did not differ significantly from those in the controls. However, four asthmatic children were found to have selective IgA deficiency (serum IgA < 0.08 milligrams). None of the patients was found to be IgG subclass-deficient.
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Rabbit cytochrome P450 2C3 was expressed from its cDNA in Escherichia coli as a chimeric enzyme in which a portion of the N-terminal membrane anchor sequence of 2C3 was replaced with a modified sequence derived from P450 17 alpha. The nucleotide sequence encoding the N-terminus of P450 17 alpha was modified previously to achieve a high level of expression of P450 17 alpha in E. coli by altering the first eight codons of P450 17 alpha to reflect second codon preferences for high expression and to minimize the potential for the formation of a stable secondary structure of the corresponding RNA transcript. The modified P450 2C3 was expressed at > 400 nmol/liter of culture. P450 2C3 was isolated to apparent electrophoretic homogeneity and a specific content > 14 nmol P450/mg protein. When reconstituted with P450 reductase and dilauroyl-L-alpha-lecithin, the purified E. coli-expressed P450 2C3 catalyzed 16 alpha, but not 6 beta-hydroxylation of progesterone. Expression of unmodified 2C3 from its cDNA in COS-1 cells confirmed the absence of detectable 6 beta-hydroxylase activity. In addition, the enzyme expressed in E. coli is activated by the allosteric effector 5 beta-pregnane-3 beta,20 alpha-diol, with a resultant Vmax = 10 min-1 and Km = 20 microM and is not inhibited by 16 alpha-methylprogesterone. These results indicate that the 2C3 cDNA encodes an enzymatic form characteristic of IIIvo/J and B/J inbred rabbits rather than a second enzymatic form expressed in most outbred and some inbred strains that catalyzes both high efficiency 16 alpha- and 6 beta-hydroxylation of progesterone. Our results have identified the enzyme variant encoded by the 2C3 cDNA and have demonstrated the utility of E. coli for the expression of recombinant P450 enzymes.
Eighteen patients with presumed childhood acquisition of chronic hepatitis B virus infection were initially entered into this randomized controlled trial. Twelve were treated with prednisolone for 4 weeks followed, after a 2-week gap, by thrice weekly lymphoblastoid alpha-interferon for 12 weeks. Two of these had previously acted as untreated controls. Three of the 12 patients (25%) [who were initially hepatitis B virus (HBV) surface antigen (HBsAg), 'e' antigen (HBeAg) and HBV-DNA positive] became HBeAg and HBV-DNA negative during therapy and remained so after 12 months post-therapy follow-up. One of these also lost HBsAg. A further two patients lost HBeAg and HBV-DNA during therapy but relapsed 6 and 9 months later. Two additional patients were HBV-DNA negative but HBeAg positive at the end of follow-up. None of the eight untreated control patients seroconverted during an identical follow-up period. Two further patients were HBsAg and HBeAg positive but HBV-DNA negative at the start of therapy. These were omitted from the final analysis: both subsequently lost HBeAg. The treatment response was associated with a rise in aspartate aminotransferase, peaking 2-6 weeks after prednisolone withdrawal, loss of HBV-DNA 0-8 weeks later and subsequent normalization of liver function tests. Treatment was well tolerated.
OBJECTIVE: Human T-cell leukemia virus types I (HTLV-I) and II (HTLV-II) are closely related human retroviruses. HTLV-I has been implicated in a chronic progressive myelopathy, known as tropical spastic paraparesis (TSP) or HTLV-I-associated myelopathy (HAM). We sought to determine whether autoantibodies to brain antigens were present in the cerebrospinal fluid (CSF) of a patient with chronic progressive spastic myelopathy with evidence of both HIV-1 infection and HTLV-I/II seropositivity. DESIGN: A 54-year-old bisexual man with clinical features of HAM/TSP of over 20 years' duration was followed. METHODS: We applied discriminatory DNA amplification (polymerase chain reaction) to distinguish HTLV-I from HTLV-II and to verify co-infection with HIV-1. The patient's CSF was used to screen a human brain cDNA expression library to identify antibodies directed against brain antigens. Autoreactive bacteriophage clones were isolated and sequenced. RESULTS: The patient was found to be co-infected with both HIV-1 and HTLV-II, but not with HTLV-I. HTLV-II proviral levels in the peripheral blood remained relatively constant, despite therapy with zidovudine. Prominent oligoclonal banding of immunoglobulins was present in the patient's CSF. A single repeatedly reactive cDNA clone was identified, by screening with CSF antibody, sequenced, and found to be the human homologue of the rat insulinoma gene, rig. CONCLUSIONS: HTLV-II infection may predispose to development of a HAM/TSP-like illness. Autoimmune mechanisms, such as autoantibody formation, may play a role in pathogenesis.
22 cases with convergence insufficiency (CI) were presented here. It is found that there was an important relationship between CI and asthenopia. The causes clinical findings, diagnostic criteria, treatment of CI and relation with asthenopia were discussed.
To determine the genetic and molecular basis for rheumatoid factor (RF) autoantibody reactivity in patients with destructive, erosive arthritis, we established a human lymphoblastoid cell line (hRF-1) from a patient with polyarthritis that produced an IgG RF mAb, mAb hRF-1. Studies of isolated H and L chains showed that the specificity of RF reactivity is conferred by mAb hRF-1 L chains. The L chain gene was cloned from a cDNA library prepared from hRF-1 cells. The nucleotide sequence was similar to known V kappa II L chains except for a two nucleotide change corresponding to a change of two amino acids in an invariable region of FR3. A germ-line gene with one of the nucleotide changes was identified by polymerase chain reaction in multiple cell lines, including K562 that does not rearrange Ig genes, but the other nucleotide change appeared to be due to mutation. Either or both of these amino acid changes may contribute to the RF reactivity, because an antibody with the same V kappa II L chain except for these two amino acid changes in FR3 did not have RF reactivity. The RF reactivity of isolated L chains from mAb hRF-1 was confirmed by transfecting COS cells with an expression vector encoding the hRF-1 kappa-chain and showing that the secreted k-chains had RF reactivity. Expression of this variant V kappa II L chain gene may form the basis for RF autoantibody reactivity in some patients.
Fragments of rabbit DNA have been cloned which encompass the gene for rabbit cytochrome P450IIC3. Cytochrome P450IIC3 is a constitutive member of the cytochrome P450IIC subfamily which contains both constitutive and phenobarbital-responsive genes. The cytochrome P450IIC3 gene spans at least 25 kbp and contains 8 introns which have 5'GT's and 3'AG's and are located in the same positions as in other family II genes. The 5' flanking region contains a consensus TATA site about 25 bp from the RNA initiation site which was mapped by the primer extension method. Other potential regulatory sequences include a CCAAT sequence and sequences similar to binding sites for the liver-specific factor HNF-1 and the general transcription factors AP-1 and OCT. The gene sequence differs from the cytochrome P450IIC3 cloned cDNA sequence in only 2 of 1400 nucleotides, 1 of which results in an amino acid change. Since sequences coding for exon 1 and part of exon 2 were not present in the cloned cytochrome P450IIC3 cDNA, a 475-nucleotide fragment of the 5' end of the cDNA was amplified by the polymerase chain reaction, cloned, and sequenced. The sequence of this cDNA was identical with the gene exon sequence. The protein sequence derived from the gene differs in nine positions from that determined directly for the protein. A comparison of the amino acid sequence by exons with other cytochrome P450IIC proteins revealed that, in general, the similarity was greater in C-terminal exons than in N-terminal ones.(ABSTRACT TRUNCATED AT 250 WORDS)
In order to identify unique structural features of pathogenic autoantibodies to DNA in SLE, a murine anti-anti-DNA (anti-Id) mAb (mAb 1C7) was produced in response to immunization of lupus mice with a syngeneic anti-DNA mAb (mAb 3E10). Immunization of lupus mice with mAb 3E10 inhibited production of native anti-DNA antibodies, suppressed development of lupus kidney disease (nephritis), and induced production of anti-anti-DNA (anti-Id) antibodies. mAb 1C7 bound F(ab')2 fragments of mAb 3E10, and it bound other murine anti-DNA mAb, but not murine mAb or polyclonal serum antibodies unreactive with DNA. Moreover, binding of mAb 1C7 anti-Id to mAb 3E10 was inhibited by DNA, suggesting anti-Id binding within or near the binding site for DNA. Furthermore, mAb 1C7 bound serum IgG immunoglobulins from 9/12 patients with lupus nephritis and serum anti-DNA antibodies compared to only 3/12 SLE patients with comparable serum levels of anti-DNA antibodies, but without nephritis (p = 0.04), and only 1/53 SLE patients without serum anti-DNA antibodies, 0/49 patients with rheumatoid arthritis, and 1/47 healthy subjects (p less than 0.001). These results provide evidence that mAb 1C7 identifies a conserved Id associated with anti-DNA antibodies in murine and human SLE and may be useful as a structural probe to characterize pathogenic anti-DNA antibodies in SLE.
To measure rheumatoid factors specific for patients with rheumatoid arthritis an enzyme linked immunosorbent assay (ELISA) was developed to measure rheumatoid factors in human serum that bind a cross-reactive determinant shared on human and other mammalian IgG. Rheumatoid factors that cross link human IgG and sheep IgG in a double binding ELISA were almost completely specific (greater than 99%) for rheumatoid arthritis in assays of 108 sera from patients with rheumatoid arthritis compared with 231 sera from patients with other connective tissue diseases and 365 sera from healthy subjects and patients without these diseases. Moreover, positive tests occurred primarily in patients with active arthritis (r = 0.68). In contrast, these rheumatoid factor autoantibodies were not detected in sera from most of the patients with other autoimmune diseases, including patients with systemic lupus erythematosus. These results show that rheumatoid factors identified in human sera by the double binding test are specific for active rheumatoid arthritis.
OBJECTIVE: To confirm the findings of pilot studies that interferon alfa is an effective treatment of Europid men with chronic hepatitis B virus infection. DESIGN: Randomised controlled trial of three months treatment with interferon alfa followed by 12 months of observation. SETTING: Outpatient clinic of a tertiary referral centre. PATIENTS: 37 Treated men (six anti-HIV positive) and 34 untreated men (nine anti-HIV positive) who met the criteria for the trial. Four controls failed to complete follow up. INTERVENTIONS: The treated group received subcutaneous injections of 5-10 MU interferon alfa/m2 daily for five days, then 10 MU/m2 thrice weekly for 11 weeks. Follow up continued at monthly intervals for 12 months. Untreated controls were monitored over the same period. MAIN OUTCOME MEASURE: Hepatitis B e antigen and hepatitis B virus DNA state after 15 months of observation. RESULTS: 12 Of the 37 treated patients cleared hepatitis B e antigen and hepatitis B virus DNA, whereas only one of 30 untreated controls seroconverted over the same period--an increased response rate of 29% (95% confidence interval 13% to 45%). The life table estimate of response at 15 months was 35% in treated patients, an increase of 32% above controls (95% confidence interval 16% to 48%). The response rates in groups by predictive pretreatment variables were 12 of 31 anti-HIV negative patients (excess response 34%; 95% confidence interval 14% to 54%), 12 of 26 with chronic active hepatitis before treatment (excess response 46%; 27% to 65%), and 12 of 21 with a pretreatment serum aspartate aminotransferase activity greater than 70 IU/l (excess response 46%; 16% to 76%). The combination of these factors predicted response with a sensitivity of 100% and a specificity of 80%. Four of the 12 responders, who had all been infected for less than two years, also lost hepatitis B surface antigen. Treatment was well tolerated. CONCLUSIONS: Interferon alfa is effective in the treatment of a proportion of Europid men with chronic hepatitis B virus infection, who might be identified before treatment. Additional strategies are required to improve the rate of response.
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Though bilirubin is reported to affect a variety of cellular functions, the primary target of its toxic effect is still not known. A major problem in understanding this is the wide variation in results reported by different groups. This is probably due to the differences in stability of bilirubin solutions arising from large differences in bilirubin:albumin molar ratios used in experiments. Hence in studying the toxic effects of bilirubin in tissue culture systems, it is important to be certain that the bilirubin is maintained in solution throughout the time of the exposure to bilirubin. Spectrophotometric measurements have shown that bilirubin is stable in Dulbecco's modified Eagle medium solution at bilirubin:albumin molar ratios up to 3. Under these defined conditions, bilirubin was found to affect Na+K+ ATPase, [3H]-thymidine uptake, L-[35S]methionine incorporation into protein and mitochondrial function at bilirubin concentrations up to 125 microM and bilirubin:albumin molar ratio of 1.5. Toxic effects on all parameters measured were evident at bilirubin:albumin molar ratio of 1.5 after a minimum of 2 h of exposure. No effect was evident at a bilirubin:albumin molar ratio below 1. Although it is not possible to identify with certainty the primary target, the effect on mitochondrial function appeared earlier and was more profound than that seen with the other assessed functions.
A set of four cerebrosides was isolated from a Pachybasium species and purified by preparative reversed-phase HPLC. All four products displayed activity in a natural product screen aimed at detecting novel cell wall-active antifungal agents based on synergy with the known glucan synthetase inhibitor, aculeacin. Based on degradation studies, fast atom bombardment mass spectrometry and 13C and high field 1H NMR techniques, the structure of the major cerebroside was determined to be (4E,8E)-N-D-2'-hydroxy-(E)-3'- hexadecenoyl-1-O-beta-D-glucopyranosyl-9-methyl-4,8-sphingadiene. The other components were found to be the corresponding 2'-hydroxypalmitic acid analog with one less double bond and an analogous pair containing 2'-hydroxystearic acid with and without the 3' double bond.