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K Li

Publications and source records attributed to K Li.

At least 109 records · Page 6Linked to original sources
PubMed2001

Replication competition between lactate dehydrogenase-elevating virus quasispecies in mice. Implications for quasispecies selection and evolution.

The common quasispecies of lactate dehydrogenase-elevating virus (LDV), LDV-P and LDV-vx, are highly resistant to the humoral host immune response because the single neutralization epitope on the ectodomain of the primary envelope glycoprotein, VP-3P, carries three large N-glycans. Two laboratory mutants, LDV-C and LDV-v, have lost two of the N-glycans on the VP-3P ectodomain, thereby gaining neuropathogenicity for AKR/C58 mice but at the same time, becoming susceptible to the humoral immune response of the host. In attempts to further assess the origins and evolution of these LDVs we have determined their competitiveness by monitoring their fate in mixed infections of wild type, SCID, nude, and cyclophosphamide-treated mice by reverse transcription/polymerase chain reaction assays that distinguish between them. In mixed infections with LDV-P and LDV-vx, LDV-C and LDV-v became rapidly lost even when present initially in large excess over the former. In mixed infections of mice unable to generate neutralizing antibodies, LDV-C and LDV-v also became replaced by LDV-P and LDV-vx as predominant quasispecies but more slowly than in immunocompetent mice. The results indicate that the humoral immune response plays an important role in the displacement of LDV-C and LDV-v by LDV-P and LDV-vx but that in addition, LDV-C and LDV-v possess an impaired ability to compete with LDV-P and LDV-vx in the productive infection of the subpopulation of macrophages that represents the host for all these LDVs. In addition, LDV-v outcompeted LDV-C in mixed infections and the same was the case for neutralization escape mutants of LDV-v and LDV-C which had regained all three N-glycosylation sites on the VP-3P ectodomain. Thus a hierarchy exists in replication fitness: LDV-P/LDV-vx>LDV-v>LDV-C, which is unrelated to the number of N-glycans on the VP-3P ectodomain. The implications of the results in relation to the evolution and selection of the LDV-quasispecies is discussed. LDV-P and LDV-vx are genetically highly stable and thus seem to have achieved evolutionary stasis with optimum ability to establish viremic persistent infections of mice that are unimpeded by the host immune responses.

Amino Acid Sequence↗
PubMed2001

Inhibitory effects of rosa roxburghii tratt juice on in vitro oxidative modification of low density lipoprotein and on the macrophage growth and cellular cholesteryl ester accumulation induced by oxidized low density lipoprotein.

BACKGROUND: Rosa roxburghii tratt juice (RRTJ) administration has been shown to significantly ameliorate atherosclerotic diseases in cholesterol-fed animals. However, the mechanism for the antiatherogenic effect of RRTJ is not clear. METHODS: We investigated the effects of RRTJ on in vitro oxidative modification of LDL and on LDL-induced macrophage growth and cellular cholesteryl ester (CE) accumulation. The effects of RRTJ on LDL oxidative modification were assessed by relative electrophoretic migration, thiobarbituric acid-reactive substance (TBARS) content, and the formation of conjugated dienes. The inhibition of RRTJ on oxidized LDL (Ox-LDL)-induced murine peritoneal macrophage growth was evaluated by a cell-counting assay and an MTT assay. The effect of RRTJ on Ox-LDL-induced cellular CE accumulation was examined after macrophages were incubated with Ox-LDL in the presence of RRTJ. To clarify the mechanism of the inhibitory effect of RRTJ on Ox-LDL-induced CE accumulation in macrophages, its capacity for cholesterol efflux from macrophage-derived foam cells were examined. RESULTS: We showed that RRTJ significantly reduced LDL oxidative susceptibility. In addition, RRTJ effectively suppressed Ox-LDL-induced macrophage growth and especially Ox-LDL-induced CE accumulation in murine peritoneal macrophages by promoting cellular cholesterol efflux. CONCLUSION: These results indicated that RRTJ exerted its antiatherogenic effects largely due to its ability to inhibit the oxidative modification of LDL and to suppress the formation of foam cells.

Animals↗
PubMed2001

Toxicity evaluation of sewage sludges in Hong Kong.

Anaerobically digested sewage sludges collected from four wastewater treatment plants located in Sha Tin, Tai Po, Yuen Long, and Shek Wu Hui in Hong Kong were subjected to chemical characterization and toxicity testing to provide preliminary assessment of their suitability for land application. All sewage sludges were slightly alkaline with pH range of 8.3-8.7. Electrical conductivity (EC; 0.69 dS m(-1)) and soluble NH4-N content (996 mg kg(-1)) of sewage sludge from Yuen Long were lower than that of other plants. Concentrations of heavy metals were determined as total, extractable, and water-soluble fraction using mixed acid digestion, DTPA (pH 7.3), and distilled water, respectively. Yuen Long sludge was most polluted with Zn and Cr higher than the pollutant concentration limits listed in Part 503 of USEPA, owing to the effluent coming from the tannery industry. High concentration of Ni was found in sludge from Sha Tin that originated mainly from the electroplating industry. DTPA-extractable Zn contents were high in sludges from Yuen Long (1247 mg kg(-1)) and Shek Wu Hui (892 mg kg(-1)), while 3.7 mg kg(-1) of DTPA-extractable Cr was found in Yuen Long sludge. Metal speciation of sludges showed that Pb was major in residual phase while Cu, Cr, and Ni in organic and residual phases, and Zn did not show any dominant chemical phase. The sludge extracts did not exert significant adverse effect on seed germination of Brassica chinensis (Chinese cabbage), but Yuen Long sludge caused a reduction in root growth. In view of its lower EC and soluble ammonia contents, the high concentration of Zn and Cr in Yuen Long sludge would likely be responsible for this adverse effect on root growth. Therefore, Yuen Long sludge would likely have a more serious impact on soil quality and plant growth as compared to other sludges. This would require further verification from greenhouse and field experiments.

Ammonia↗
PubMed2001

Histological reaction to hydroxyapatite in the middle ear of rats.

OBJECTIVE: Present study was performed to evaluate the histological response of rat middle ear mucosa following implantation of Apaceram granules, a synthetic dense hydroxyapatite [Ca10(PO4)6(OH)2], prepared from commercially available synthetic auditory ossicle, and to assess the precise histological response of the rat middle ear to implantation of Apaceram granules, by microscopic examination of mucosal tissue at various time points after implantation. METHODS: Apaceram granules were implanted in the temporal bulla of 32 rats. As control, sham surgery was performed in a group of ten rats. Bulla specimens were removed at 1, 3, 7, 14, and 30 days after surgery in the implant and control groups, and at 90, 180 and 300 days in the implant group. Specimens were decalcified, sectioned at a thickness of 6 microm, and stained with haematoxylin and eosin, and Mallory's azan for histological examination of mucosal tissue. RESULTS: Evidence of inflammatory reaction was slightly greater in the implant group than in controls. Lymphocyte and macrophage counts were higher in the implant group 1 day after surgery, but decreased to similar levels by day 3, and continued to decrease thereafter, and few were observed in the implant group at 300 days. Neutrophils observed at 1 day after surgery were not evident in either group at 3 days. Gradual fibrosis development continued in both groups over all time points studied. Foreign body giant cells were never observed in either group. No bony reaction was observed in any specimen. CONCLUSION: The results of this study suggest that Apaceram is biocompatible and suitable for reconstructive ear surgery.

Animals↗
PubMed2001

Inhibition of the HIV-1 rev-RRE complex formation by unfused aromatic cations.

RNA viruses cause a wide range of human diseases. Development of new agents to target such viruses is an active area of research. Towards this goal, a series of diphenylfuran cations as potential inhibitors of the Rev-RRE complex have been designed and synthesized. Analysis of the interaction of the diphenylfurans with RRE and TAR RNA model systems by gel shift assays indicates that they exhibit both sequence and structure-dependent binding modes. Our results show a strong interaction between the diphenylfuran ring system and RRE bases, while the TAR interactions are much weaker with the compounds that are the best inhibitors of Rev-RRE.

Aminoglycosides↗
PubMed2001

Cord blood collection before and after placental delivery: levels of nucleated cells, haematopoietic progenitor cells, leukocyte subpopulations and macroscopic clots.

The number of nucleated cells infused into the recipient of a cord blood (CB) transplant has emerged as the most important factor affecting the probability and speed of engraftment. At present, there is no international consensus on the procedure of CB collection in the maternity ward. In order to maximise the yield of viable cells in a CB unit, we aimed to investigate the efficiency of CB collection, with respect to the time of delivery of the placenta. We analysed stem and progenitor cells in terms of CD34+ cell content and colony-forming activities, lymphocyte subpopulations and the presence of macroscopic clots in 93 paired CB samples, collected before and after the delivery of the placenta. Our results demonstrated that the median concentrations of nucleated cells and total colony-forming unit (CFU) were significantly lower in CB collected after placenta delivery by 9.5% (P < 0.001) and 11.6% (P = 0.015), respectively, when compared to their counterparts collected before placental delivery. A reduction of granulocytes (P < 0.001), monocytes (P < 0.001) and CD19+ B lymphocytes (P = 0.031) was observed, with no significant change in the proportion of T cell subsets (CD4+, CD8+ cells) or activated T cells (CD25+, CD45RO+ cells) in samples collected after placenta delivery. The incidence of macroscopic clots was also higher in these samples (31% vs 1%, P < 0.001). The reduction of stem and progenitor cells correlated significantly with that of major cell populations, indicating a general cell loss, possibly due to clotting activities developed with time. Our study has documented strong evidence for recommending the collection of CB before the delivery of the placenta.

Blood Cell Count↗
PubMed2001

Platelet-derived growth factor enhances ex vivo expansion of megakaryocytic progenitors from human cord blood.

Infusion of ex vivo expanded megakaryocytic (MK) progenitor cells is a strategy for shortening the duration of thrombocytopenia after haematopoietic stem cell transplantation. The cell dose after expansion has emerged as a critical factor for achieving the desired clinical outcomes. This study aimed to establish efficient conditions for the expansion of the MK lineage from enriched CD34(+) cells of umbilical cord blood and to investigate the effect of platelet-derived growth factor (PDGF) in this system. Our results demonstrated that thrombopoietin (TPO) alone produced a high proportion of CD61(+)CD41(+) cells but a low total cell count and high cell death, resulting in an inferior expansion. The addition of interleukin-1 beta (IL-1 beta), Flt-3 ligand (Flt-3L) and to a lesser extent IL-3 improved the expansion outcome. The treatment groups with three to five cytokines produced efficient expansions of CFU-MK up to 400-fold with the highest yield observed in the presence of TPO, IL-1 beta, IL-3, IL-6 and Flt-3L. CD34(+) cells were expanded by five to 22-fold. PDGF improved the expansion of all cell types with CD61(+)CD41(+) cells, CFU-MK and CD34(+) cells increased by 101%, 134% and 70%, respectively. On day 14, the CD61(+) population consisted of diploid (86.5%), tetraploid (11.8%) and polyploid (8N--32N; 1.69%) cells. Their levels were not affected by PDGF. TPO, IL-1 beta, IL-3, IL-6, Flt-3L and PDGF represented an effective cytokine combination for expanding MK progenitors while maintaining a moderate increase of CD34(+) cells. This study showed, for the first time, that PDGF enhanced the ex vivo expansion of the MK lineage, without promoting their in vitro maturation. PDGF might be a suitable growth factor to improve the ex vivo expansion of MK progenitors for clinical applications.

Antigens, CD↗
PubMed2001

Association of clonal T-cell large granular lymphocyte disease and paroxysmal nocturnal haemoglobinuria (PNH): further evidence for a pathogenetic link between T cells, aplastic anaemia and PNH.

There is mounting evidence to suggest that T-cell-mediated suppression of haemopoiesis is a pathogenetic mechanism in three bone marrow failure syndromes: aplastic anaemia (AA), paroxysmal nocturnal haemoglobinuria (PNH) and myelodysplasia (MDS). T-cell microclones can be detected by sensitive polymerase chain reaction (PCR)-based methods in all three disorders. Recently, larger clonal populations of T-cell large granular lymphocytes (T-LGLs) have been observed in some patients with AA and MDS. Here, we report the development of a large clonal T-LGL population in a patient with bona fide PNH. In this patient, we defined part of the sequence of the T-cell receptor (TCR) beta-chain gene, and we have shown that the large T-LGL population emerged from a background of multiple smaller T-cell clones. Thus, T-LGL clones in AA, MDS and PNH probably expand as a result of antigenic stimulation. It is postulated that the antigen driving clonal T-cell proliferations in these disorders exists on haemopoietic stem cells.

Adult↗
PubMed2001

Haematopoietic stem and progenitor cells in human term and preterm neonatal blood.

BACKGROUND AND OBJECTIVES: Whilst cord blood (CB) contains a significant number of haematopoietic stem and progenitor cells suitable for bone marrow transplantation, levels of these cells are very low in the adult circulation. In previous studies, we demonstrated that stem and progenitor cells are present in neonatal blood (NB) and reported the first sibling transplant using a combination of CB and NB for a patient with beta-thalassaemia major. However, our preliminary data showed that the number of CD34+ cells decreased rapidly in the peripheral blood of neonates soon after birth. To further investigate the mechanism of the change of stem and progenitor cells in NB, we measured the steady-state levels of CD34+ cells, early progenitor subsets and the expression of adhesion molecules, in term and preterm neonates. MATERIALS AND METHODS: NB was collected serially from infants at 2, 4, 6, 8, 24 and 48 h after birth and was analysed by three-colour flow cytometry. RESULTS: Our results demonstrated that the number of CD34+ cells rapidly decreased in term NB, particularly during the first 2-6 h of life, by 29.2 +/- 5.55% (P = 0.0003) in absolute counts/ml. A decrease was observed in all subsets of CD34+ cells studied, including the CD33+, CD71+, CD62L+ and CD49d+ populations. In contrast, the CD34+ cell number increased in preterm infants in the first 8 h of life, before starting to decrease. Significant inverse correlations were observed between gestational age and levels of CD34+ cells (P = 0.0065, 4-h collection time-point). CONCLUSION: Our study suggests that changes in the levels of CD34+ stem and progenitor cells in NB might be triggered by parturition and affected by the gestational age. Our findings confirmed that if targeted as a supplementary source of stem and progenitor cells to CB for transplantation, NB should be collected as soon as possible after birth.

Antigens, CD34↗
PubMed2001

Enzymatic and electrochemical oxidation of N-hydroxy compounds. Redox potential, electron-transfer kinetics, and radical stability.

A series of N-hydroxyacetanilide and 1-hydroxybenzotriazole analogues derivatized by various functional substituents were studied with regard to redox potential, oxidation by laccase, oxidative stability, and correlation to the electronic and steric properties of the substituents. It was found that substituents carrying conjugative/pi-electron function influenced the redox potential more than substituents carrying inductive/sigma-electron function, and that the electron-transfer from an N-hydroxy moiety to laccase was significantly affected by the redox potential. Electron-withdrawing substituents tended to reduce the electron density on the N-hydroxy group, leading to higher redox potential and lower oxidation rate. Bulky substitution or absence of N-phenyl tended to increase the Km of the N-hydroxy group, leading to lower oxidation rate. Oxidized N-hydroxy compounds were stabilized by N-phenyl or N-carbonyl group, but not by N-azo or highly strained structure. Potential implication of these effects on laccase-based, N-hydroxy compound-mediated biocatalysis is discussed.

Catalysis↗
PubMed2001

Dextran sedimentation in a semi-closed system for the clinical banking of umbilical cord blood.

BACKGROUND: The results of current processing procedures for reducing volume and recovering HPCs from umbilical cord blood (UCB) before cryopreservation vary. STUDY DESIGN AND METHODS: Dextran was added to bags containing UCB, followed by sedimentation for 30 minutes. The processed UCB was then frozen. RBCs, nucleated cells, MNCs, CD34+ cells, CFUs and long-term culture-initiating cells (LTC-ICs), viability, and sterility were evaluated. Fractionations in ficoll-hypaque and hydroxyethyl starch (HES) were also run in parallel for comparison. RESULTS: The nucleated cell (NC) recovery and RBC depletion were 86.1 percent and 94.3 percent, respectively (n = 50). Sedimentation with dextran also enabled the recovery of 80.7 percent MNCs and 82.6 percent CD34+ cells (n = 30). Postsedimentation samples displayed no impairment of CFU growth (n = 42, 108.7% CFU-C, 104.6% CFU-GEMM, 107% CFU-GM, and 95.7% BFU-E). Long-term cultures on five paired samples before and after sedimentation generated similar numbers of CFU-C each week (p = 0.88). Limiting dilution analysis of 12 paired pre/postsedimentation samples showed comparable median proportions of LTC-ICs (1/6494 vs. 1/5236; p = 0.18). The cell viability of 24 samples of thawed UCB after sedimentation was 90.3 percent (77.5-96%) and the recovery of CFU-C, CFU-GEMM, CFU-GM, and BFU-E of 11 postsedimentation samples was 93.4 percent, 84.9 percent, 92.3 percent, and 83.4 percent, respectively. NC recovery was significantly higher after treatment with dextran than with ficoll-hypaque (n = 30; 88.5% vs. 29.1%; p<0.005) and HES treatment (n = 21; 88.5% vs. 76.4%; p = 0.004). However, MNCs, CD34+ cells, CFUs, LTC-ICs, and RBCs were comparable. Two cycles of dextran sedimentation recovered 93.9 percent of NCs with cell viability of 98.6 percent (96.5-100%), whereas 11.7 percent of RBCs were retained (n = 20). The final yield volume was 33.5 (28-41) mL. CONCLUSION: In a semi-closed system, dextran sedimentation enabled volume reduction of UCB without significant quantitative and qualitative losses of HPCs.

Blood Banks↗
PubMed2001

Preclinical ex vivo expansion of cord blood hematopoietic stem and progenitor cells: duration of culture; the media, serum supplements, and growth factors used; and engraftment in NOD/SCID mice.

BACKGROUND: Ex vivo expansion of cord blood (CB) hematopoietic stem and progenitor cells increases cell dose and may reduce the severity and duration of neutropenia and thrombocytopenia after transplantation. This study's purpose was to establish a clinically applicable culture system by investigating the use of cytokines, serum-free media, and autologous plasma for the expansion of CB cells and the engraftment of expanded product in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice. STUDY DESIGN AND METHODS: Enriched CB CD34+ cells were cultured in four media (Iscove's modified Dulbecco's medium with FCS, Gibco; X-Vivo-10, BioWhittaker; QBSF-60, Quality Biological; and StemSpan SFEM, Stem Cell Technologies) with four cytokine combinations (thrombopoietin [TPO], SCF, Flt-3 ligand [FL] with and without G-CSF, and/or IL-6). The effect of autologous CB plasma was also investigated. The read-out measures were evaluated on Days 8 and 12. After expansion at the optimized condition, cultured cells were transplanted into sublethally irradiated NOD/SCID mice. The engraftment of human CD45+ cells and subsets in the bone marrow, spleen, and peripheral blood was determined. RESULTS: QBSF-60 or StemSpan SFEM supported high yields of early progenitors (CD34+ cells, <or= 64.8-fold; CD34+CD38- cells, 330-fold; CFU-granulocyte erythroid macrophage megakaryocyte [GEMM], 248-fold) and CFUs of the myeloid (CFU-GM, 407-fold) and erythroid (BFU/CFU-E, 144-fold) lineages. The expansion of the megakaryocytic lineage was consistently higher in X-Vivo-10 (CFU-megakaryocyte, 684-fold). Autologous plasma promoted colony formation but reduced CD34+ cells and CFU-GEMM. The addition of G-CSF or IL-6 improved cell yields; G-CSF was more effective for committed progenitors. Expansion products from cultures in QBSF-60 with the cytokines engrafted and differentiated into the myeloid and lymphoid lineages in NOD/SCID mice. CONCLUSION: The data supported the strategy of expansion. The optimized condition may be applicable to clinical expansion for the abrogation or reduction of posttransplant cytopenia.

Animals↗
PubMed2001

The road to danger: the comparative risks of driving while sleepy.

OBJECTIVES/HYPOTHESIS: A large sector of the population of the United States has sleep deprivation directly leading to excessive daytime sleepiness. The prevalence of excessive daytime sleepiness in this population ranges from 0.3% to 13.3%. The consequences of even 1 to 2 hours of sleep loss nightly may result in decrements in daytime functions resulting in human error, accidents, and catastrophic events. The magnitude of risks in the workplace or on the highways resulting from sleepiness is not fully understood or appreciated by the general population. Hence, to more clearly emphasize the magnitude of these risks, we question whether mild sleep deprivation may have the same effect as alcohol on reaction times and driving performance. STUDY DESIGN: Nonrandomized prospective cohort investigation. METHODS: Sixteen healthy matched adult subjects (50% women) were stratified into two groups, sleep deprived and alcohol challenged. The sleep-deprived group was further subdivided into acute (one night without sleep) and chronic (2 h less sleep nightly for 7 d) sleep deprivation. Each group underwent baseline reaction time testing and then drove on a closed course set up to test performance. Seven days later, the group repeated this sequence after either sleep deprivation or alcohol intake. RESULTS: There were no significant between-group differences (sleep deprivation or alcohol challenged) in the changes before and after intervention for all 11 reaction time test metrics. Moreover, with few exceptions, the magnitude of change was nearly identical in the two groups, despite a mean blood alcohol concentration of 0.089 g/dL in the alcohol-challenged group. On-track driving performances were similar (P =.724) when change scores (hits and errors) between groups were compared (baseline minus final driving trial). CONCLUSION: This comparative model suggests that the potential risks of driving while sleepy are at least as dangerous as the risks of driving illegally under the influence of alcohol.

Adult↗
PubMed2001

Radiofrequency treatment of turbinate hypertrophy in subjects using continuous positive airway pressure: a randomized, double-blind, placebo-controlled clinical pilot trial.

OBJECTIVES: To estimate the treatment effect of temperature-controlled radiofrequency (TCRF) reduction of turbinate hypertrophy in patients with sleep-disordered breathing (SDB) treated with nasal continuous positive airway pressure (CPAP), and to assess the impact of study design on this estimate. STUDY DESIGN: Prospective, randomized, double-blind, placebo-controlled clinical pilot trial. METHODS: Twenty-two CPAP-treated patients with SDB with turbinate hypertrophy were randomly assigned to either TCRF turbinate treatment (mean energy 415 +/- 37 J/turbinate; n = 17) or placebo control (n = 5). Changes in nasal obstruction were evaluated between pretreatment and 4 weeks post-treatment. The primary outcome assessed changes in the blinded examiners' findings of nasal obstruction on a visual analogue scale (VAS). Secondary outcomes included blinded patients' and unblinded examiner assessments of nasal obstruction (VAS), nightly CPAP use, adherence, and tolerance, along with sleepiness and general health status scales. The treatment group findings were subtracted from the changes in the placebo group to yield treatment effect. RESULTS: The primary outcome treatment effect by VAS was -0.9 cm (95% confidence interval [CI], -2.4, 0.7), and beyond the placebo effect of -1.5 cm (95% CI: -3.4, 0.3). The secondary treatment effect of the unblinded examiner was -3.0 cm (95% CI, -4.9, -1.1). A beneficial treatment effect was also seen on every secondary outcome except general health status, but only self-reported CPAP adherence (P = .03) was statistically significant. CONCLUSIONS: TCRF turbinate treatment appears to benefit nasal obstruction and CPAP treatment for SDB. Placebo control and double blinding are critical for establishing the true treatment effect. A future definitive trial is feasible to establish statistical significance of these findings.

Adult↗
PubMed2001

Immunocytochemical localization of mitochondrial single-stranded DNA-binding protein in mitochondria-rich cells of normal and neoplastic human tissue.

Mitochondrial single-stranded DNA-binding protein (mtSSB) is necessary for mtDNA replication. So far the protein has been studied mainly in Escherichia coli and in cell cultures of lower mammals. In this investigation, the authors studied the expression of mtSSB in normal and neoplastic human tissue by light and electron immunocytochemistry. mtSSB has been detected in various tissues and particularly in mitochondria-rich tissues such as external eye muscles and parietal cells of the stomach and in mitochondria-rich tumors (oncocytomas) of various origins. Ultraimmunocytochemistry disclosed the specific distribution of immunoreactive mtSSB over the mitochondria. The staining intensity was heterogeneous. Forty-five percent had a labeling index (silver grains/microm2) greater than 1 and less than 3, approximately 20% had an index of 3 or greater, and 15% of mitochondria remained unstained. The mean labeling index was 1.83. Immunolabeling showed a linear correlation with the mitochondrial profile area (r = 0.82). In conclusion, mtSSB is regularly expressed in normal and neoplastic human tissue of different origin, function, and differentiation. The heterogeneous staining pattern most probably reflects the functional heterogeneity of mitochondria.

DNA, Mitochondrial↗
PubMed2001

Recombinant urease and urease DNA of Coccidioides immitis elicit an immunoprotective response against coccidioidomycosis in mice.

Coccidioides immitis antigens which stimulate a T helper cell 1 (Th1) pathway of host immune response are considered to be essential components of a vaccine against coccidioidomycosis. Recombinant urease (rURE) and recombinant heat shock protein 60 (rHSP60) of C. immitis were expressed in Escherichia coli and tested as vaccine candidates in BALB/c mice. A synthetic oligodeoxynucleotide which contained unmethylated CpG dinucleotides and was previously shown to enhance a murine Th1 response was used as an immunoadjuvant. T cells isolated from the spleens and lymph nodes of the rURE- and rHSP60-immune mice showed in vitro proliferative responses to the respective recombinant protein, but only those T lymphocytes from rURE-immunized mice revealed markedly elevated levels of expression of selected Th1-type cytokine genes. BALB/c mice immunized subcutaneously with rURE and subsequently challenged by the intraperitoneal (i.p.) route with a lethal inoculum of C. immitis arthroconidia demonstrated a significant reduction in the level of C. immitis infection compared to control animals. rHSP60 was much less effective as a protective antigen. Evaluation of cytokine gene expression in lung tissue and levels of recombinant urease-specific immunoglobulins (immunoglobulin G1 [IgG1] versus IgG2a) in murine sera at 12 days after challenge provided additional evidence that immunization with rURE stimulated a Th1 response to the pathogen. Urease was further evaluated by expression of the URE gene in a mammalian plasmid vector (pSecTag2A.URE) which was used to immunize mice by the intradermal route. In this case, 82% of the vector construct-immunized animals survived more than 40 days after i.p. infection, compared to only 10% of the mice immunized with the vector alone. In addition, 87% of the pSecTag2A.URE-immunized survivors had sterile lungs and spleens. These data support the need for further evaluation of the C. immitis urease as a candidate vaccine against coccidioidomycosis.

Animals↗