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Biomedical subjects

X M Sun

Publications and source records attributed to X M Sun.

At least 55 records · Page 3Linked to original sources
PubMed1996

Cell-specific activation and detoxification of benzene metabolites in mouse and human bone marrow: identification of target cells and a potential role for modulation of apoptosis in benzene toxicity.

The role of cell-specific metabolism in benzene toxicity was examined in both murine and human bone marrow. Hemopoietic progenitor cells and stromal cells are important control points for regulation of hemopoiesis. We show that the selective toxicity of hydroquinone at the level of the macrophage in murine bone marrow stroma may be explained by a high peroxidase/nicotanimide adenine dinucleotide phosphate, reduced [NAD(P)H]:quinone oxidoreductase (NQO1) ratio. Peroxidases metabolize hydroquinone to the reactive 1,4-benzoquinone, whereas NQO1 reduces the quinones formed, resulting in detoxification. Peroxidase and NQO1 activity in human stromal cultures vary as a function of time in culture, with peroxidase activity decreasing and NQO1 activity increasing with time. Peroxidase activity and, more specifically, myeloperoxidase, which had previously been considered to be expressed at the promyelocyte level, was detected in murine lineage-negative and human CD34+ progenitor cells. This provides a metabolic mechanism whereby phenolic metabolites of benzene can be bioactivated in progenitor cells, which are considered initial target cells for the development of leukemias. Consequences of a high peroxidase/NQO1 ratio in HL-60 cells were shown to include hydroquinone-induced apoptosis. Hydroquinone can also inhibit proteases known to play a role in induction of apoptosis, suggesting that it may be able to inhibit apoptosis induced by other stimuli. Modulation of apoptosis may lead to aberrant hemopoiesis and neoplastic progression. This enzyme-directed approach has identified target cells of the phenolic metabolites of benzene in bone marrow and provided a metabolic basis for benzene-induced toxicity at the level of the progenitor cell in both murine and human bone marrow.

Animals↗
PubMed1996

Induction of apoptosis by benzene metabolites in HL60 and CD34+ human bone marrow progenitor cells.

Two cell types, HL60 human promyelocytic leukemia cells and CD34+ human bone marrow progenitor cells, were used as model systems to explore a possible role for apoptosis in the myelotoxicity of the phenolic metabolites of benzene. HL60 cells were treated with either phenol, catechol, hydroquinone, or 1,2,4-benzenetriol and then stained with Hoechst 33342 and propidium iodide and subjected to fluorescent microscopy. Cells with nuclear condensation and fragmentation were scored as apoptotic, and etoposide (40 microM) was used as a positive control. Catechol, 1,2,4-benzenetriol, and hydroquinone induced marked time- (0-24 hr) and concentration- (25-100 microM) dependent apoptosis, whereas phenol (750 microM) did not. Under these conditions, no significant necrosis was observed. The induction of apoptosis was confirmed by internucleosomal cleavage of DNA, assessed by agarose gel electrophoresis. CD34+ cells treated with etoposide (40 microM) or hydroquinone (50 microM) for 18 hr were stained and subjected to fluorescent microscopy as above. The percentage of cells exhibiting nuclear condensation and/or fragmentation as well as high intensity staining significantly increased in both cases. The induction of apoptosis was confirmed using a terminal deoxynucleotidyl transferase assay. These data show that apoptosis can be induced in both HL60 and CD34+ human bone marrow progenitor cells by benzene metabolites. The ability of phenolic metabolites of benzene to induce apoptosis in human bone marrow progenitor cells may contribute to benzene myelotoxicity.

Adult↗
PubMed1996

Characterization of mutations in the low density lipoprotein (LDL)-receptor gene in patients with homozygous familial hypercholesterolemia, and frequency of these mutations in FH patients in the United Kingdom.

Mutations in the gene for the low density lipoprotein (LDL) receptor have been identified in 15 patients with homozygous familial hypercholesterolemia (FH). Five patients are homozygous at the LDL-receptor locus; their mutant alleles include Glu387Lys and Pro664Leu in patients of Asian-Indian descent, Cys292Stop in a Greek Cypriot, Cys281Trp in a Turkish patient, and Gln 540Stop in a West Indian. The other 10 patients (9 of apparently British ancestry) are compound heterozygotes. Mutations have been identified in 18 of 20 possible alleles, including Glu80Lys (2 patients), Pro664Leu (3 patients), Asp69Gly, Cys176Arg, Cys227Tyr, Ser265Arg, Asp280Ala, Asp283Glu, Arg329Pro, Asp461Asn, Leu578Ser, a single bp deletion in exon 15, a 21 bp duplication of codons 200-206 and two large deletions. The seven mutations underlined above have not been described previously. The two uncharacterized mutant alleles fail to produce detectable amounts of mRNA. LDL-receptor activity in cultured cells from 13 of the 15 homozygous patients varied from undetectable to about 30% of normal, but there was no correlation between LDL-receptor activity and the untreated plasma cholesterol concentration in these patients. When genomic DNA from 295 patients with a clinical diagnosis of FH was screened for 29 mutations found in these and other FH patients of British ancestry, most were identified in only one or a few individuals. Four patients heterozygous for the Asp461Asn allele showed a wide range of clinical manifestations. These observations confirm the striking heterogeneity underlying FH in most populations and demonstrate the variability in phenotype between patients with the same mutation.

Adolescent↗
PubMed1995

Discrimination of apoptotic thymocytes by forward light scatter.

The discrimination of apoptotic and normal rat thymocytes by forward-angle light scatter in the UV depended on the angle over which the scattered light was collected. To achieve good separation, the light needed to be collected over a narrow angle. It was also observed that apoptotic thymocytes gave less forward light scatter than normal cells, whereas, under identical conditions, apoptotic cells from a murine cell line gave more light scatter than their normal counterparts. We suggest that the forward light scatter of apoptotic cells is affected by several factors, including changes in cell size and in the internal structure of the cell.

Animals↗
PubMed1995

Endogenous bacterial toxins are required for the injurious action of platelet-activating factor in rats.

BACKGROUND & AIMS: Platelet-activating factor (PAF), an endogenous mediator for experimental sepsis, has been shown to induce shock and intestinal necrosis in vivo. However, it is unclear whether PAF exerts its injurious effects on the intestinal tissue directly or via synergism with other endogenous products. The aim of this study was to examine the role of endogenous bacterial products, such as endotoxin, in PAF-induced intestinal injury. METHODS: PAF (3 micrograms/kg) was injected intravenously into normally colonized rats, germfree rats, and normal rats pretreated with a combination of antibiotics, and the systemic response and intestinal injury were assessed. RESULTS: PAF did not cause prolonged shock, leukopenia, hemoconcentration, and bowel necrosis in germfree rats. When germfree rats were primed with a low dose (0.5 mg/kg) of endotoxin, the protection was lost. Combined treatment of the normally colonized rats with neomycin, polymyxin B, and metronidazole for 7 days largely protected the animal from PAF-induced shock and intestinal necrosis. CONCLUSIONS: PAF does not directly induce prolonged hypotension, hemoconcentration, persistent leukopenia, and gross intestinal necrosis but causes these changes via a synergism with endogenous bacterial toxins, presumably from the gut flora.

Analysis of Variance↗
PubMed1995

Protection of SJL/J mice from demyelinating disease mediated by Theiler's murine encephalomyelitis virus.

Intracerebral infection with the DA strain of Theiler's murine encephalomyelitis virus induces a chronic demyelinating disease in SJL/J mice. Intraperitoneal inoculation with either the wild-type DA virus or an attenuated variant virus of DA, H7A6-2, results in protection from development of chronic demyelinating disease. Protective anti-viral immune responses result in reduced viral titers and decreased inflammation in the central nervous system within the first week following intracerebral challenge with virus. Development of protective immunity requires the presence of B cells and CD4+ T cells but does not require CD8+ T cells. High titers of serum anti-viral IgG and neutralizing antibodies are induced following the intraperitoneal inoculation with the DA virus or H7A6-2 virus prior to challenge. While protection could not be transferred with immune serum from DA virus-infected mice or neutralizing monoclonal antibodies, protection was correlated with increased numbers of DA virus-specific plasma cells in the central nervous system within the first week following intracerebral challenge. Protected mice also had enhanced levels of anti-DA virus IgG and neutralizing antibodies in the cerebral spinal fluid by 1 week following intracerebral challenge with DA virus. Thus, we conclude that vaccination with live virus results in protection from chronic demyelinating disease by inducing immune responses which are manifested in the central nervous system and rapidly clear infection after intracerebral challenge with DA virus.

Animals↗
PubMed1995

A mutation (T-45C) in the promoter region of the low-density-lipoprotein (LDL)-receptor gene is associated with a mild clinical phenotype in a patient with heterozygous familial hypercholesterolaemia (FH).

We have identified a rare mutation (T-45C) in the low density lipoprotein (LDL)-receptor gene in a Welsh patient with a clinical diagnosis of heterozygous familial hypercholesterolaemia (FH). The mutation is in the proximal Sp1 binding site in repeat 3 of the 42 bp region of the promoter required for sterol-dependent regulation of transcription, but the substituted nucleotide is not a strongly conserved base in the consensus sequence for Sp1 binding. Normal and mutant promoter fragments (from base -600 to -5) were linked to a luciferase reporter gene, and transient expression in COS cells showed that the mutation reduced transcriptional activity to approximately 43% of normal in the presence, and 25% in the absence of sterols in the medium. Competitive gel-shift mobility assays showed that the mutation reduced the binding affinity for transcription factor Sp1. Analysis of a neutral polymorphism in the LDL-receptor mRNA from the patient's lymphoblasts showed that expression of one allele was reduced. Since Southern blotting of genomic DNA and sequencing of the entire coding region of the LDL-R gene did not reveal any other potential defects, we infer that the T-45 C mutation is the underlying cause of hypercholesterolaemia in the proband.

Animals↗
PubMed1995

Effect of the StuI polymorphism in the LDL receptor gene (Ala 370 to Thr) on lipid levels in healthy individuals.

We have examined the effect on plasma lipid levels of a single base change in exon 8 of the LDL receptor gene that causes an amino acid change Ala 370 to Thr in a sample of 318 Icelandic individuals selected at random from the general population. The change destroys a StuI restriction site and was detected by digestion of pooled samples in groups of 5. The frequency of the loss of the cutting site was 0.05 (95% CI = 0.014-0.054). In men, those with the Thr allele (n = 18) had 8.3% higher total cholesterol, 11.8% higher LDL cholesterol and 10.3% higher apolipoprotein B than those with the common Ala allele, whereas in women those with the Thr allele (n = 12) had levels lower by 7.4%, 13.3% and 10.1% respectively. These differences reached statistical significance only in the men (p < 0.05). Functional analysis of CHO cells transfected with constructs of the LDL receptor cDNA carrying the Ala370 and Thr370 alleles showed that within the limits of the assays there was no difference in function of the LDL receptor protein as measured by uptake and degradation of LDL. The data raise the possibility that amino acid substitutions that could affect LDL receptor function below the limits of detection by conventional assays, may have an effect on plasma lipid levels in the general population.

Adolescent↗
PubMed1995

B-lymphocyte requirement for vaccine-mediated protection from Theiler's murine encephalomyelitis virus-induced central nervous system disease.

The role of humoral immunity in the protection of vaccinated SJL/J mice from central nervous system disease induced by the DA strain (DAV) of Theiler's murine encephalomyelitis virus was investigated in B-cell-deficient mice. Mice were depleted of B cells by treatment with a mouse monoclonal antibody specific for immunoglobulin M. DAV-vaccinated, B-cell-deficient mice failed to clear viral infection and were no longer protected from Theiler's murine encephalomyelitis virus-mediated central nervous system disease. CD4+ T cells are required in this model of protection to provide help for the development of an antiviral antibody response in the central nervous system.

Animals↗
PubMed1995

Characterization of a splice-site mutation in the gene for the LDL receptor associated with an unpredictably severe clinical phenotype in English patients with heterozygous FH.

We have identified a substitution of G to A in the first base pair of intron 3 in the LDL receptor gene of an English heterozygous familial hypercholesterolemia (FH) patient. Reverse transcription, amplification, and nucleotide sequencing of the LDL receptor mRNA from mononuclear blood cells showed both the normal mRNA and one that lacked the nucleotides encoded by exon 3, which codes for repeat 2 of the ligand-binding domain. The same mutant allele was identified in 2/200 unrelated FH patients from the London area and 4/77 from Manchester. Immunoblotting of cultured lymphoblasts from the index patient revealed the normal receptor protein and smaller amounts of a receptor protein with electrophoretic mobility consistent with a deletion of the 41 amino acid residues encoded by exon 3. Normal amounts of a similar protein were observed when the mutant cDNA was expressed in heterologous cells; this protein showed reduced binding affinity for LDL but bound apoprotein E-containing lipoproteins normally. Despite these and other observations that repeat 2 of the binding domain is relatively unimportant for receptor function in vitro, carriers of this allele exhibit a severe clinical phenotype, typical of FH. Thus, the relationship between genotype and phenotype in heterozygous FH is not always predictable.

Adult↗
PubMed1994

Adult onset immunodeficiency caused by inherited adenosine deaminase deficiency.

Adenosine deaminase (ADA) deficiency is identified here as a cause of adult onset immunodeficiency. Two sisters who noted recurrent, predominantly chest infections in their twenties were found in their thirties to have CD4+ lymphopenia and lymphocyte ADA activity of approximately 5% of the lower limit of normal. Immune function, measured by proliferation of PBMCs in vitro to mitogens and specific Ags, was impaired. Inheritance of a polymorphic marker showed that both patients were heterozygous at the ADA locus. In the paternal allele there was a deletion resulting from homologous recombination between two alu elements that normally flank the first exon and the polymorphic marker. The recombination site was distinct from that in similar deletions described in two infants having severe combined immunodeficiency. This allele is predicted to result in a null phenotype. In the mutant allele inherited from the mother, a C to T transition in a CpG dinucleotide changed the codon for arginine 211, which lies in a conserved sequence close to the active site, to that for cysteine. This mutation has been observed previously in a child in whom the other allele was also a null mutation, but who was diagnosed as having partial ADA deficiency because immune function was apparently normal. The late onset of immunodeficiency in our patients suggests that immune function in children with partial ADA deficiency may deteriorate with time and that ADA deficiency should be regarded as a possible cause of adult onset immune dysfunction of unknown etiology.

Adenosine Deaminase↗
PubMed1994

Formation of large molecular weight fragments of DNA is a key committed step of apoptosis in thymocytes.

Apoptosis is a process in which cells die in a controlled manner and apparently participate in their own demise. It is best characterized morphologically by condensation of chromatin and biochemically by cleavage of chromatin at internucleosomal regions to yield a classical DNA ladder pattern. Apoptosis was induced in thymocytes by exposure to either the glucocorticoid, dexamethasone, or DNA topoisomerase II inhibitor, etoposide. We describe the formation of large m.w. fragments of DNA, 30 to 50 kilobase pairs in length, in a population of these thymocytes at an early stage of apoptosis before internucleosomal cleavage of DNA. These fragments are absent in normal thymocytes and their formation is dependent on protein synthesis. Their appearance is coincident with the commitment of these cells to apoptosis. The formation of these large fragments is associated with the condensation of chromatin, abutting the nuclear membrane, recognized as one of the earliest ultrastructural signs of apoptosis. Subsequent cleavage of these large fragments gives rise to oligonucleosomal fragments and is independent of protein synthesis. We propose that the formation of large fragments of DNA represents a key committed step in apoptosis, and that it is from these fragments that the archetypal DNA ladders associated with apoptosis are derived.

Animals↗
PubMed1994

Mg(2+)-dependent cleavage of DNA into kilobase pair fragments is responsible for the initial degradation of DNA in apoptosis.

Cleavage of DNA into oligonucleosomal fragments, recognizable as a DNA ladder on agarose gel electrophoresis, is usually considered as the biochemical hallmark of apoptosis. Recently, it has been shown that this internucleosomal cleavage is preceded by the formation of large fragments of DNA of > or = 700, 200-250, and 30-50 kilobase pairs (kbp) in length. Using isolated thymocyte nuclei, we now demonstrate that the formation of these large fragments is Mg(2+)- but not Ca(2+)-dependent. Further degradation of > or = 460- and 200-250-kbp fragments to 30-50-kbp fragments but not oligonucleosomal cleavage is also Mg(2+)-dependent but is facilitated by Ca2+. In contrast, formation of oligonucleosomal fragments does not occur in the presence of either Ca2+ or Mg2+ alone but requires the presence of both cations. These results support the hypothesis that the formation of these large fragments and the subsequent internucleosomal cleavage are distinct steps in the degradation of DNA in apoptosis in thymocytes.

Animals↗
PubMed1994

Changes in nuclear chromatin precede internucleosomal DNA cleavage in the induction of apoptosis by etoposide.

Etoposide, a DNA topoisomerase II inhibitor, caused a concentration-dependent induction of apoptosis in immature thymocytes. Using a flow cytometric method to separate and quantify normal and apoptotic cells, etoposide-induced apoptosis was inhibited by cycloheximide and actinomycin D but not by zinc. Etoposide induced a marked cleavage of DNA into nucleosomal length fragments or multiples thereof, which was completely inhibited if the thymocytes were also incubated in the presence of zinc. Etoposide, alone, induced the classical ultrastructural features of apoptosis, but in the presence of zinc, the morphological pattern was markedly different and dominated by discrete clumps of condensed chromatin abutting the nuclear membrane. These latter changes resemble those described as the earliest changes in apoptosis. These results support the hypothesis that, in the induction of apoptosis, critical alterations in nuclear chromatin occur prior to endonuclease cleavage of DNA into nucleosomal fragments.

Apoptosis↗
PubMed1994

The involvement of apoptosis in etoposide-induced thymic atrophy.

A time- and dose-dependent thymic atrophy was observed in young male Fischer 344 rats dosed intraperitoneally with etoposide (10, 30, or 100 mg/kg). Histopathological examination of the thymus revealed that the pattern of cell death in the majority of thymocytes had a characteristic apoptotic morphology typified by nuclear condensation. This observation was supported by the formation of internucleosomal fragments of DNA in thymocytes isolated from animals dosed with etoposide. Administration of the protein synthesis inhibitor, cycloheximide (1.5 mg/kg, ip), 1 hr prior to etoposide inhibited the induction of apoptosis in thymocytes, assessed by both biochemical and histological criteria. Flow cytometric analysis of thymocytes from animals dosed with etoposide in vivo revealed the formation of both apoptotic cells and apoptotic bodies in contrast to previous in vitro studies which showed the formation of only apoptotic cells. Our data indicate that the induction of apoptosis in thymocytes is a major mechanism involved in etoposide-induced thymic atrophy.

Animals↗
PubMed1994

Modeling the fine structure of the 2f1-f2 acoustic distortion product. I. Model development.

The fine structure of the 2f1-f2 acoustic distortion product (ADP) as a function of frequency has been measured in human subjects and shows a series of sharp peaks and valleys (rippling) with peak-to-valley level differences of up to 15-20 dB. In order to delineate the cause of the ADP rippling pattern, a computer model was developed to simulate the behavior of the ADP, specifically the ADP fine structure. The ADP model includes the middle ear and cochlea. The middle ear was treated as a simple signal delivery system in both the forward and reverse directions. The ADP was assumed to be generated within the cochlea by nonlinear elements taken to be the outer hair cells (OHCs), and an array of ADP generators was used to simulate the OHCs along the basilar membrane (BM). The magnitude and phase of the output of each of the ADP generators were functions of the local responses of the two primary traveling waves. The traveling waves were calculated from a passive transmission line model of the BM using the WKB approximation, coupled to a second-order resonance to mimic the contribution from active OHC feedback. The system output of ADP in dB was proportional to the weighted vectorial sum of all the components, arriving at the stapes. Parameters such as lateral coupling and feedback gain were examined.

Acoustics↗
PubMed1994

Modeling the fine structure of the 2f1-f2 acoustic distortion product. II. Model evaluation.

In a previous article, a vector sum model was developed that successfully reproduces the ADP rippling pattern when a nonuniformity is introduced in the active damping factor, a parameter that is inversely related to the energy gain contribution of the outer hair cells [OHCs, Sun et al., J. Acoust. Soc. Am. 96, 2166-2174 (1994)]. Here, the mean of the damping factor is increased nonlinearly with input level, mimicking the saturation of the active feedback of the OHCs. The passive damping factor in the transmission line model was also nonlinearly increased with input level to reproduce the frequency shift of the peak of the traveling wave observed in experimental data. The resulting model simulates an ADP that is compatible with data from human subjects wherein the ADP fine structure does not saturate with level. Moreover, the model suggests that the shifting of the ADP pattern with level is a direct result of the peak shift of the traveling wave, thus implicating the nonlinear damping factors as the underlying basis of this phenomenon. The input/output (I/O) functions of the simulated ADP emissions at specific frequencies were also examined. The resulting functions show a variety of shapes, depending on the pattern of ADP fine structure around the I/O frequency, and the way the fine structure shifts as primary levels increase. These I/O functions are also similar to those observed in human subjects, even with regard to overall slopes which approximate one. Thus the model illustrates how cubic distortion generators coupled with damping on linearities can yield I/O function slopes on the order of one, rather than the expected three.(ABSTRACT TRUNCATED AT 250 WORDS)

Basilar Membrane↗