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Biomedical subjects

C Luo

Publications and source records attributed to C Luo.

At least 145 records · Page 8Linked to original sources
PubMed1995

A study of biological function of retinoblastoma gene (Rb gene).

The human wild-type Rb gene cDNA has been cis- or trans inserted into the retrovirus vector DOL, resulting in a sense-expression vector DOLRB and an antisense-expression vector DOLRBAS of Rb gene. By eletroporation transfection techniques, the vector DOLRB has been introduced into the human breast carcinoma cell line MDAMB468 and human hepatocellular carcinoma cell line SMMC7721 both of which have an inactivated Rb gene and the vector DOLBAS, into normal human embryonic lung fibroblasts HEL cells. With the expression of Rb protein, the growth rate of the MDAMB468 cells is decreased by about 50%, their colony formation ability in soft agar is repressed completely, and their tumorigenicity in nude mice is repressed partially. Meanwhile, the cell population of G1 phase of Rb+ MDAMB468 cells is increased markedly. About 75% of transfected SMMC7721 cells have been killed by Rb gene product. For HEL cells, with the transient expression of antisense Rb gene, the Rb protein synthesis is reduced and the growth rate of those cells increased, but no colonies of HEL cells are formed in soft agar.

Animals↗
PubMed1994

Human immunodeficiency virus type-1 infection in Zambian children with tuberculosis: changing seroprevalence and evaluation of a thioacetazone-free regimen.

SETTING: This study was conducted at the Department of Paediatrics and Child Health, University Teaching Hospital (UTH), in Lusaka, Zambia. OBJECTIVES: To monitor the seroprevalence of HIV type-1 in children with tuberculosis and to evaluate the response to anti-tuberculosis therapy using a thioacetazone-free treatment regimen. DESIGN: A prospective cross-sectional study of all consecutive newly diagnosed cases of TB in children from 1 month-15 years of age seen at the University Teaching Hospital (UTH) in Lusaka, Zambia between 1 October 1991 and 31 May 1992. RESULTS: 120 children with a clinical diagnosis of tuberculosis and 167 controls were enrolled in the study. The overall HIV type-1 seroprevalence rate in children with tuberculosis was 55.8% (67/120) compared to 9.6% (16/167) amongst the control group (P < 0.0001: odds ratio = 11.50; 95% CI = 5.99-22.7). Common clinical presentations among children with TB were bronchopneumonia (45/162), miliary TB (30/162) and tuberculous lymphadenopathy (21/33). There were no significant differences in clinical presentation of TB between the HIV-negative and HIV-positive groups. The follow-up of those patients with tuberculosis was poor, with only 65 patients (55%) returning to the clinic for scheduled appointments after discharge. All the 16 patients who died did so within 60 days of discharge from hospital; all of them were seropositive for HIV. There were no deaths among the HIV-negative group. Despite the exclusion of thioacetazone from the treatment regimen, cutaneous reactions occurring within 8 weeks of commencing treatment were observed in 7 of the 65 (11%) patients, 2 of whom developed fatal Stevens-Johnson syndrome. All 7 patients were seropositive for HIV-1. CONCLUSIONS: The seroprevalence rate of HIV type-1 among children with tuberculosis in Lusaka continues to rise; careful monitoring of anti-TB therapy (even in regimens excluding thioacetazone) for potentially lethal side effects should be carried out.

Adolescent↗
PubMed1993

Isolation of the cyclosporin-sensitive T cell transcription factor NFATp.

Nuclear factor of activated T cells (NFAT) is a transcription factor that regulates expression of the cytokine interleukin-2 (IL-2) in activated T cells. The DNA-binding specificity of NFAT is conferred by NFATp, a phosphoprotein that is a target for the immunosuppressive compounds cyclosporin A and FK506. Here, the purification of NFATp from murine T cells and the isolation of a complementary DNA clone encoding NFATp are reported. A truncated form of NFATp, expressed as a recombinant protein in bacteria, binds specifically to the NFAT site of the murine IL-2 promoter and forms a transcriptionally active complex with recombinant protein fragment react with T cell NFATp. The molecular cloning of NFATp should allow detailed analysis of a T cell transcription factor that is central to initiation of the immune response.

Amino Acid Sequence↗
PubMed1993

Cloning of a novel surface antigen from the insect stages of Trypanosoma brucei by expression in COS cells.

Trypanosoma brucei cDNA libraries constructed in the vector pCDM8 were screened selectively for insect (procyclic) stage surface antigen cDNAs by transient expression in mammalian COS-7 fibroblasts and "panning" with a rabbit polyclonal antiserum. This strategy yielded two surface antigen cDNAs termed PSSA-1 and PSSA-2. The PSSA-1 cDNA encoded an isotype of procyclin, the major phosphatidylinositol-linked stage-specific glycoprotein antigen of the tsetse fly infective forms of T. brucei. The PSSA-2 cDNA encoded a new and previously unidentified stage-specific surface antigen with the features of a typical transmembrane glycoprotein but with an unusual cytoplasmic tail composed of a proline-rich tandem repeat. Fluorescent antibody staining of PSSA-1 transfected COS cells with a panel of procyclin-specific monoclonal antibodies confirmed that the protein was located on the outer surface of the plasma membrane. Furthermore, the antigen on COS cells was insensitive to treatment with phosphatidylinositol-specific phospholipase C suggesting that the inositol of the glycosylinositol phospholipid-lipid anchor contained the same fatty acyl modification reported recently for the procyclin molecule in cultured procyclic trypanosomes. In contrast the PSSA-2 antigen on COS cells was stained very weakly by whole parasite antisera. Northern blot hybridization revealed that the PSSA-2 antigen was encoded by a single 1.7-kilobase transcript which was present in parasites from the insect procyclic stage of the life cycle but not from the animal bloodstream stage. Southern blot hybridization analysis of DNA from procyclic stage trypanosomes indicated that the gene for PSSA-2 may be present in more than one copy in procyclic trypanosomes.

Amino Acid Sequence↗
PubMed1993

Dissociation of the lipid-containing bacteriophage PRD1: effects of heat, pH, and sodium dodecyl sulfate.

The double-stranded DNA bacteriophage PRD1 replicates in Escherichia coli and Salmonella typhimurium. It has an outer protein coat surrounding a membrane. The phage lipids are derived from the host, but the membrane proteins are of phage origin. In this investigation we studied the effects of heat, pH, and sodium dodecyl sulfate on the integrity of phage particles. Heat and high pH result in the release of the main coat protein, P3, as trimers, whereas treatment of phage particles with detergent results in the solubilization of the membrane. Our results enable a distinction to be made between the phage structural proteins that are embedded in the lipid bilayer and those that appear to be more loosely associated with the membrane.

Capsid↗
PubMed1993

Isolation of a phospholipid-free protein shell of bacteriophage PRD1, an Escherichia coli virus with an internal membrane.

PRD1 is a double-stranded DNA virus infecting Escherichia coli and Salmonella typhimurium. It has an icosahedral outer protein capsid which encloses the viral membrane, inside of which resides the phage genome. In this investigation we demonstrate the detergent resistance of the intact virus particles. The membrane of empty DNA-free particles, however, is very sensitive to detergent action. We assume that their sensitivity is due to the access of detergents through a portal structure to the virus interior. Using the anionic detergent sodium dodecyl sulfate, it is possible to obtain a shell structure composed of the major coat protein P3 alone. The treatment of empty particles with the milder nonionic detergents n-octyl beta-D-glucopyranoside and Triton X-100 yielded P3 particles which retained the membrane-associated proteins P7 and P11. Deoxycholic acid treatment yielded shells of intermediate composition between those obtained with the nonionic detergents and sodium dodecyl sulfate.

Capsid↗
PubMed1993

Bacteriophage PRD1 proteins: cross-linking and scanning transmission electron microscopy analysis.

Bacteriophage PRD1, a double-stranded DNA virus infecting Escherichia coli, has a membrane inside the protein capsid. Chemical cross-linking and scanning transmission electron microscopy showed that the multimeric major coat protein (P3) exists in a trimeric form. Cross-linking revealed, in addition, that protein P11, located between the protein coat and the membrane, exists also as a homotrimer. Minor protein P7 was associated with the major coat protein P3. Under nonreducing conditions the infectivity proteins P16 and P18 formed homomultimeric complexes which were dissociated upon addition of 2-mercaptoethanol.

Capsid↗
PubMed1993

Topology of the major capsid protein P3 of bacteriophage PRD1: analysis using monoclonal antibodies and C-terminally truncated proteins.

Trimeric capsomeres of protein P3 (395 aa) are the main component of the phage PRD1 capsid, which encloses a lipid-protein vesicle containing the viral dsDNA genome. In this study we characterize a panel of monoclonal antibodies (MAb) against P3. The epitopes recognized by the MAbs are analyzed by immunoprecipitation of intact virions or of released P3 trimers, and by Western blotting using a series of C-terminally truncated P3 molecules. Nine of the MAbs recognize epitopes on the virion surface, whereas five require unmasking of epitopes by disruption of the virions. Several of the MAbs are capable of neutralizing the virus; this is most probably due to virus aggregation by the antibodies. Analysis of the C-terminal truncations (the 6 Western blot-positive MAbs were used) delineates three major antigenic regions of the protein. The epitope of MAb 3T74 is included in the 66 N-terminal amino acids, and is not accessible on the virion surface, suggesting that the N-terminus is internally located in the capsid. MAbs 3N81 and 3R2 recognize epitopes in the region of amino acids 159-168, which is part of the first predicted beta-barrel structure of P3. The third antigenic region is in the second predicted beta-barrel, between amino acids 217-242, where the epitopes of 3N180, 3P4, and 3T5 map. The trimerization of P3 was found to be independent of the non-structural assembly factor proteins P10 and P17. Functional studies of the truncated proteins reveal that molecules comprising of 294 or more residues from the P3 N-terminus are capable of trimer formation.

Antibodies, Monoclonal↗
PubMed1993

Angiotensin II and catecholamines interaction in short-term low protein feeding.

Renal and systemic hemodynamic responses to an alpha-adrenergic agonist (norepinephrine, NE) and an alpha-adrenergic antagonist (phentolamine, PHEN) were studied in weanling rats pair-fed isocaloric diets containing either normal (NP, 23%) or low (LP, 6%) protein. Mean arterial pressure (MAP) rose less with NE and fell more with PHEN in LP than in NP. Plasma NE and epinephrine (E; 46 +/- 5 and 51 +/- 4 ng/ml) were higher in LP than in NP (26 +/- 3 and 39 +/- 3 ng/ml). These could not be attributed to changes in red cell mass nor the volumes of plasma, extracellular, or interstitial fluid in LP versus NP. Plasma angiotensin II (Ang II), renin (PRA), and aldosterone (PA) were lower in LP than in NP. An increased number without changes in affinity of glomerular Ang II receptors was found in LP compared to NP, while alpha 1- and alpha 2-adrenergic receptors were down-regulated in LP as compared to NP without changes in affinity for the alpha 1 receptor but with an increase in renal alpha 2 receptor affinity. LP (vs. NP) decreased GFR and RPF, and increased renal vascular resistance (RVR). NE decreased RPF equally in NP versus LP but raised RVR approximately twofold in NP versus LP. PHEN decreased RPF and increased RVR less in LP than in NP. Moreover, PHEN increased renal renin content approximately seven-fold over the basal NP values. Exogenous Ang II increased RVR and lowered RPF more in LP than in NP. Enalapril abolished all the hemodynamic changes of LP and restored the systemic response to NE.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin II↗
PubMed1993

Regulation of renin-angiotensin system in unilateral ureteral obstruction.

The effect of 24-hour unilateral ureteral obstruction (UUO) on the expression and regulation of the renin-angiotensin system (RAS) in rats and of pretreatment with lisinopril (5 mg/kg/day) or the AT1-R inhibitor, losartan, (10 mg/kg/day) on renal hemodynamics was evaluated. Both drugs improved the post-obstructed kidney (POK) renal hemodynamics, lowered MAP, and normalized eicosanoid excretion by the POK. Cortex and medulla POK:CK ratio of relative density R mRNA was approximately 3.5 for both. Sham, POK, and CK showed renin immunoreactivity and R mRNA exclusively in juxtaglomerular position. In addition, in POK renin was expressed in mesangial cells, along greater lengths of afferent arterioles and in dilated distal tubules and loops of Henle. In situ hybridization revealed that approximately 20% more glomeruli in POK than CK overexpressed R mRNA. Blood vessels of POK consistently showed greater ACE and Ao mRNA expression than CK. Overexpression of the genes coding for members of the RAS is possibly responsible for local Ang II production which, in view of the response to CEI and AT1-R inhibitors, is at least partly responsible for the severe hemodynamic changes in UUO.

Animals↗
PubMed1993

Seroprevalence of human immunodeficiency virus type 1 infection in Zambian children with tuberculosis.

Descriptions in the medical literature of human immunodeficiency virus type 1 (HIV-1) in children with tuberculosis (TB) are scanty. This study determined the seroprevalence of HIV-1 in 237 hospitalized children between the ages of 1 month and 14 years with a clinical diagnosis of TB (125 males and 112 females) and in 242 control children (149 males and 93 females). The overall HIV-1 seroprevalence rate in patients with TB was 37% (88 of 237) compared with 10.7% (26 of 242) among the control group (P < 0.00001: odds ratio 5.37, 95% confidence interval = 3.21 < 5.37 < 9.47). HIV-1 seropositivity in children with TB ranged from 53% (31 of 58) in the 12- to 18-month age group to 14% (9 of 61) in the 10- to 14-year-olds. The risk of TB attributable to HIV infection was 29%. The predominant clinical presentation in both seronegative (84.6%) and seropositive (89.7%) groups was that of pulmonary TB and there were no significant differences in clinical presentation between the two groups of patients. Only 54.8% of the patients attended follow-up clinics regularly whereas 32% were lost to follow-up within 3 months. Bacillus Calmette-Guérin vaccination coverage was 87.3% among TB patients and 90.5% in the controls. No significant differences in B. Calmette-Guérin vaccination rates between the seronegative and seropositive children were seen. Coinfection with HIV and TB in children is now one of the major public health problems in Zambian children.

AIDS-Related Opportunistic Infections↗
PubMed1993

Cutaneous hypersensitivity reactions due to thiacetazone in the treatment of tuberculosis in Zambian children infected with HIV-I.

Tuberculosis is one of the most common infections in Zambian adults and children infected with HIV. In Africa, cutaneous hypersensitivity reactions attributed to thiacetazone during treatment of tuberculosis in adults infected with HIV-I have been well documented. This study monitored adverse drug reactions during treatment for tuberculosis over an 18 month period (1 April 1990 to 31 October 1991) in 237 children with a clinical diagnosis of tuberculosis (125 boys and 112 girls; 88/237 (37%) infected with HIV-I) and 242 control children (149 boys and 93 girls; 26/242 (11%) infected with HIV-I). Twenty two (9%) of the 237 children with tuberculosis developed hypersensitivity skin reactions during the course of treatment. Adverse skin reactions were seen more often in children infected with HIV than in those who were not (odds ratio 11.65, 95% confidence interval 3.07 to 34.88). These represented 19 (21%) of 88 children infected with HIV and three (2%) of 149 children not infected with HIV. These skin reactions occurred after a period of treatment ranging between two and four weeks among 14 children receiving the HST (isoniazid, streptomycin, thiacetazone) regimen and eight children receiving the HSTR (isoniazid, streptomycin, thiacetazone, rifampicin) regimen. Twelve (55%) of the 22 children who reacted adversely to treatment developed the Stevens-Johnson syndrome. All 12 of these children with the Stevens-Johnson syndrome were infected with HIV. The mortality among these children who developed the Stevens-Johnson syndrome was 91% (11 of 12 died within three days of the onset of the reaction). No further reactions were observed in the 11 children who recovered from the cutaneous hypersensitivity reactions after thiacetazone was discontinued over a period of six months of further treatment of tuberculosis. The results of this study were in part responsible for the recommendations put forward by the World Health Organization to avoid the use of thiacetazone in the treatment of tuberculosis in children infected with HIV.

AIDS-Related Opportunistic Infections↗
PubMed1993

Renal renin, angiotensinogen, and ANG I-converting-enzyme gene expression: influence of dietary protein.

Low-protein (LP) feeding (6%) of rats results in renal hemodynamic changes that are abolished by converting enzyme and nonpeptide AT1 inhibitors, suggesting a role for intrarenal angiotensin II (ANG II). Dietary protein is a stimulus for the expression of renal renin mRNA in intact and partially nephrectomized rats. In the present study, LP increased renal renin immunoreactivity and renin mRNA as assessed by in situ hybridization and Northern blot analysis; whole kidney expression of angiotensinogen mRNA was unaltered. Whole kidney, cortical, and medullary ANG I-converting-enzyme (ACE) mRNA was also increased in LP vs. normal protein. The changes occurred despite a reduced protein synthetic rate (RNA-to-DNA ratio) in the kidney of LP, which did not change expression of renal glyceraldehyde-3-phosphate dehydrogenase mRNA. These studies show for the first time that LP diet increases the expression of renal renin and ACE mRNA in intact rats; these may be responsible for the renal hemodynamic changes of LP.

Angiotensinogen↗
PubMed1993

Internal cRNA standards for quantitative northern analysis.

We report a simple method using copy RNA (cRNA) internal standards for quantitative Northern hybridization. This was accomplished by synthesis of a full-length or "half"-length cRNA and mixing these RNA internal standards with samples to be tested for the abundance of a given mRNA. Both full-length and truncated cRNAs are detected in Northern analysis by the nucleic acid detection probe (which can be labeled with 32P or biotin), and the known amount of the truncated cRNA is compared to the homologous mRNA present in the specimen being examined. We demonstrate the usefulness of this method by measuring the expression of renin in rat kidney with ureteral obstruction and angiotensin II receptor (AT1-R) mRNA in kidneys from spontaneously hypertensive rats. It was found that this method of preparing cRNA internal standards successfully controlled for variables (such as differences in loading, transfer and hybridization efficiency) that often frustrate efforts to use Northern analysis as a quantitative tool and enabled direct estimation of absolute mRNA amount present in samples. This technique may have wide applicability and permit a more quantitative use of Northern analysis.

Animals↗
PubMed1992

Recombinant human complement subcomponent C1s lacking beta-hydroxyasparagine, sialic acid, and one of its two carbohydrate chains still reassembles with C1q and C1r to form a functional C1 complex.

In contrast to the human serum protein which is approximately one-half erythro-beta-hydroxyasparagine at asparagine 134 [Theilens et al. (1990) Biochemistry 29, 3570-3578], recombinant C1s expressed by insect cells after infection with recombinant baculovirus entirely lacks posttranslational modification at asparagine 134. It is also incompletely glycosylated, lacking, at least, sialic acid. Site-directed mutagenesis of one of the two sites of carbohydrate attachment (Asn 159 to Gln 159) yields a faster migrating recombinant C1s still abundantly secreted. Furthermore, the mutated protein displays good hemolytic activity when reassembled with C1q and either human serum or recombinant C1r, demonstrating that these posttranslational modifications are not critical for any of the multiple interactions between C1s and C1q, C1r, C2, and C4 required for reassembly of the C1 complex, activation, and initiation of the classical complement pathway. The 4.0S recombinant C1s dimerizes to yield 5.6S C1s2 in the presence of Ca2+ and forms the 9.1S C1s-C1r-C1r-C1s tetramer upon the addition of human serum C1r and the 15.6S C1 complex upon the addition of C1q to the tetramer. The recombinant C1s and human serum C1s have identical N-terminal amino acid sequences, indicating proper recognition by the insect signal peptidase. The recombinant C1s is secreted and isolated as the unactivated zymogen, and it may be activated by human serum C1r which cleaves at Arg422-Ile423 to yield the characteristic heavy and light chains. A very tight complex is formed between C1-inhibitor and the light chain of recombinant C1s.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗