Biomedical subjects

J Cai

Publications and source records attributed to J Cai.

At least 253 records · Page 14Linked to original sources

PubMed1995

Measurement of the hemoglobin N-(2-oxoethyl)valine adduct in ethyl carbamate-treated mice.

Ethyl carbamate (EC) is bioactivated by CYP2E1 through vinyl carbamate to its epoxide, a reactive electrophile. This carcinogen reacts with macromolecules, including hemoglobin (Hb). This report defines a method to examine levels of N-(2-oxoethyl) adduct on the N-terminal valine of Hb after EC treatment at carcinogenic doses. Concentrations were determined 24 hr following an oral dose of EC (1 mg/g body wt) to strains A/J and C57BL/6 mice. Globin samples were isolated by precipitation in acidified acetone, washed, dried, and stored frozen at -20 degrees C until analyzed. Weighed aliquots were treated with sodium borohydride to reduce the aldehyde of the 2-oxoethyl group to the N-(2-hydroxyethyl) adduct. The adduct valine was cleaved using phenylisothiocyanate to form a substituted phenylthiohydantoin derivative of N-(2-hydroxyethyl)valine in a modified Edman degradation. After reaction with N,O-bis(trimethylsilyl)trifluoroacetamide, the resultant product, 1-(2'-trimethylsilyloxy)ethyl-5-isopropyl-3-phenyl-2-thiohydantoin , was quantified by GC/MS with selected ion monitoring of the molecular ion using synthetic N-(3-hydroxypropyl)valine as an internal standard. No adducts were detected without NaBH4 reduction. Strain A/J mice treated with EC (1 mg/g, N = 10) yielded mean +/- standard deviation (SD) adduct level values of 13.3 +/- 1.03 nmol/g globin; saline-treated A/J controls (N = 7) gave background levels of 4.43 +/- 0.69 nmol/g globin. Strain C57BL/6 mice treated with EC (1 mg/g, N = 6) exhibited mean +/- SD values of 12.0 +/- 1.92 nmol/g globin, while control mice of this strain (N = 4) had adduct levels of 7.23 +/- 1.19 nmol/g globin. These results are consistent with findings of others that bioactivation of EC produces N-(2-oxoethyl)valine hemoglobin adducts. Although the difference between mouse strains in mean total adduct levels following EC treatment was not significant, the differences evident in comparisons within strains due to treatment, between strains in endogenous background levels, and between strains in estimates of mean increases in adduct concentrations resulting from EC treatment were highly significant (p < 0.01). This assay provides a biomarker system for assessment of production from EC of the electrophilic metabolites which are believed to be genotoxic following metabolic activation in vivo.

Animals↗

PubMed1995

Association of vagal tone with serum insulin, glucose, and diabetes mellitus--The ARIC Study.

Reduced vagal activity assessed by heart rate variability (HRV) has been observed in studies of diabetics, but this association has not been reported at the population level. To investigate the association of HRV with diabetes mellitus, as well as fasting serum insulin, and glucose, we examined a stratified random sample of 1933 individuals (154 diabetics and 1779 non-diabetics), aged 45-65 years from the Atherosclerosis Risk in Communities (ARIC) study cohort. Two-minute, resting, supine beat-to-beat heart rate records were collected. Power spectral density estimation was used to derive HRV high frequency power (HF, 0.15-0.35 Hz) as the conventional marker of vagal function. Age, race, and gender-adjusted geometric means of HF were 0.78 and 1.27 (beat/min)(2) for diabetics and non-diabetics respectively (P for mean difference <0.01), reflecting a reduced vagal activity in diabetics. In individuals not diagnosed as diabetics, a graded, inverse association was observed between fasting serum insulin and HF (P for trend <0.01): the age, race, and gender-adjusted geometric mean values of HF in the lowest and highest quartiles of serum insulin were 1.34 and 1.14 (beat/minute)(2), respectively. A similar association was observed between glucose and HF in a univariate model, but not in the adjusted model. This first population-based study on this subject confirmed that diabetics have significantly lower vagal activity than non-diabetics. In individuals not diagnosed as diabetics, serum insulin, and, to a lesser degree, serum glucose were inversely associated with vagal function, suggesting a role in the pathogenesis of diabetic neuropathy.

Blood Glucose↗

PubMed1995

An Escherichia coli chromosomal ars operon homolog is functional in arsenic detoxification and is conserved in gram-negative bacteria.

Arsenic is a known toxic metalloid, whose trivalent and pentavalent ions can inhibit many biochemical processes. Operons which encode arsenic resistance have been found in multicopy plasmids from both gram-positive and gram-negative bacteria. The resistance mechanism is encoded from a single operon which typically consists of an arsenite ion-inducible repressor that regulates expression of an arsenate reductase and inner membrane-associated arsenite export system. Using a lacZ transcriptional gene fusion library, we have identified an Escherichia coli operon whose expression is induced by cellular exposure to sodium arsenite at concentrations as low as 5 micrograms/liter. This chromosomal operon was cloned, sequenced, and found to consist of three cistrons which we named arsR, arsB, and arsC because of their strong homology to plasmid-borne ars operons. Mutants in the chromosomal ars operon were found to be approximately 10- to 100-fold more sensitive to sodium arsenate and arsenite exposure than wild-type E. coli, while wild-type E. coli that contained the operon cloned on a ColE1-based plasmid was found to be at least 2- to 10-fold more resistant to sodium arsenate and arsenite. Moreover, Southern blotting and high-stringency hybridization of this operon with chromosomal DNAs from a number of bacterial species showed homologous sequences among members of the family Enterobacteriaceae, and hybridization was detectable even in Pseudomonas aeruginosa. These results suggest that the chromosomal ars operon may be the evolutionary precursor of the plasmid-borne operon, as a multicopy plasmid location would allow the operon to be amplified and its products to confer increased resistance to this toxic metalloid.

Adenosine Triphosphatases↗

PubMed1995

[Quantitation of Ras p21 oncogene and DNA content of gastrointestinal smooth muscle tumors and prognostic significance].

Expression of ras p21 oncogene and DNA content of paraffin-embedded tissues from 55 smooth muscle tumors of the gastrointestinal tract were analysed simultaneously by using immunofluorescence and flow cytometry. All of 14 leiomyomas were found to be DNA diploid and lower-expression of ras p21. Four cases of DNA aneuploidy (33%) and 9 cases of ras p21 overexpression (75%) were found in 12 potential malignant smooth muscle tumors, while all 29 cases of leiomyosarcomas were aneuploidy (100%) (P < 0.005), but the rate of ras p21 overexpression was not increased continuously (72%) (P < 0.005). 24 of 33 aneuploid tumors (73%) were found to be ras p21 overexpression, while in 22 diploid tumors, there were only 6 cases (27%) (P < 0.005). The outcome of the patients with ras p21 lower-expressed diploid tumor was excellent, compared to the patients with ras p21 overexpressed aneuploid tumor it was worst (P < 0.005). It was suggested that DNA aneuploidy and ras p21 overexpression could be regarded as the mark of malignancy. The overexpression of ras p21 oncogene was always presented in the early stage of malignant smooth muscle tumors, when the cell proliferation was active but the DNA content was still normal, and stable in the whole procss of the tumor progression. ras p21 expression and DNA content could supply a deficiency each other in the diagnosis and could be used as objective parameters in distinguishing malignancy from benign and predicting the prognosis of the patients with smooth muscle tumors of the gastrointestinal tract.

Adolescent↗

PubMed1995

Hormonal and cell density regulation of hepatic gamma-glutamylcysteine synthetase gene expression.

We previously reported that the activity of gamma-glutamylcysteine synthetase (GCS), the rate-limiting enzyme in GSH synthesis, is under both hormonal and cell density regulation in cultured rat hepatocytes. Specifically, the addition of insulin or hydrocortisone to culture media or the lowering of the initial plating cell density increased cell GSH by increasing the activity of GCS. In the present study, we examined the molecular mechanism of these effects. To determine whether the increase in GCS activity is associated with an increase in GCS heavy subunit (GCS-HS) mRNA expression, the steady state mRNA levels of GCS-HS were examined with the use of Northern blots. After 24-hr treatment of high density (0.6 x 10(5) cells/cm2) cultured rat hepatocytes with insulin (1 micrograms/ml) or hydrocortisone (50 nM), the steady state GCS-HS mRNA level increased by approximately 1-2 fold. When the plating density was decreased to 0.1 x 10(5) cells/cm2, the steady state GCS-HS mRNA level also increased by 1-2 fold 24 hr later. An increase in the steady state GCS-HS mRNA level was found within 4 hr of either hormonal treatment or cell density manipulation. The increase in steady state GCS-HS mRNA level resulted from increased gene transcription, as the transcriptional rates of GCS-HS after hormonal or cell density manipulation were increased by 2-3-fold, whereas the rates of GCS-HS mRNA degradation remained unchanged. Western blotting confirmed the increase in GCS-HS protein level after hormone treatment or lowering of plating cell density. When examined in vivo, the steady state GCS-HS mRNA level decreased by 50% in a rat in which diabetes had been induced with streptozotocin for 1 week; this was prevented with insulin replacement. In summary, GCS-HS gene expression is under both hormonal and cell density regulation.

Amino Acid Sequence↗

PubMed1995

[The origin of midline malignant reticulosis].

On the basis of light microscope examination and clinical pathological retrospective analysis, 32 cases of highly suspected midline malignant reticulosis (MMR) were investigated for immunophenotype expression (UCHL-1 for T lymphocytes and L26 for B cells) and TCR beta or IgH gene rearrangement. Scrapped tissue from stained and unstained formalin-fixed paraffin-embedded tissue sections were used for PCR gene rearrangement analysis. The results showed: Atypical lymphoid cells (ALC) in 24 cases of MMR expressed UCHL-1 marker, only one case was positive for L26. UCHL-1 and L26 positive cells often coexisted in the lesion. TCR and IgH gene rearrangement analysis using scrapped tissue showed 17 cases to be positive for TCR beta gene, which included 12 for UCHL-1 positive and 5 for UCHL-1 negative. The only one which was L26 positive showed IgH gene rearrangement. The gene rearrangement analysis further demonstrated that most MMR are T lymphocyte neoplastic proliferating disorder (extranodal T lymphoma) and could also originate from B lymphocytes.

Adolescent↗

PubMed1995

Chinese squash leaf curl virus: a new whitefly-transmitted geminivirus.

Coat protein (CP) gene of the Chinese squash leaf curl virus (SqLCV-C) was amplified through PCR, cloned and completely sequenced. Based on the comparisons at the levels of both CP gene nucleotide and CP-deduced amino acid sequences with other geminiviruses, SqLCV-C is confirmed to be distinct from the American squash leaf curl virus (SqLCV-E). It is a new geminivirus transmitted by whitefly Bemisia tabaci, which infects dicotyledonous plants and is more closely related to the Indian cassava mosaic virus (ICMV).

Amino Acid Sequence↗

PubMed1995

[Fate of human fetal dopamine neurons transplanted into rhesus monkey model of Parkinson's disease: a tyrosine hydroxylase immunocytochemical study].

To predicate the value of human fetal substantia nigra transplantation in clinical treatment of Parkinson's disease (PD), dissociated cells of substantia nigra from 8-12 week old abortive human fetus were grafted into the neostriatum of 5 adult rhesus monkeys with hemiparkinsonism induced by unilateral injection of MPTP. At 2, 5 and 12 months after transplanting the monkeys were sacrificed for tyrosine hydroxylase (TH) immunocytochemistry to examine the survival and possible synaptic contact of transplanted dopamine (DA) neurons. Transplanted TH immunoreactive cells took a pattern of patches scattered in the neostriatum. Each of the cell patches consisted of 3-10 cells. The TH immunoreactive fiber network was seen in the neostriatum. Electron microscopic survey revealed that TH+ buttons arising from grafted DA neurons formed symmetric or asymmetric synapses with TH- dendritic shafts/spines, and TH+ dendrites were seen to form synapses with TH- axons of the host. Additionally, there were a few synapses formed by TH+ axonal terminals with negative buttons. The results suggest that DA neurons from 8-12 week old abortive human fetus are able to survive grafting into the neostriatum of monkey, a species phylogenetically very close to human, and to establish reciprocal synaptic connectivity with the host even at 2 months post-transplanting. It is, therefore, inferable that embryonic human DA neurons transplanted into human neostriatum may have the same fate as in monkeys.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗

PubMed1995

[Karyotype analysis of Candida species by PFGE and its use in classicification].

Electrophoretic karyotype in 7 medically important Candida species was analyzed by pulsed-field gel electrophoresis. The chromosomal DNAs of the Candida species were separated into 4-10 bamds ranging in size from 0.5-2.8 Mb. These patterns were species-specific, so the electrophoretic karyotype can be used as a genotype basis for classification and identification of Candida species.

Candida↗

PubMed1995

Quantitation of P53 protein expression in gastrointestinal smooth muscle tumors. Clinicopathological correlation and prognostic significance.

Quantitative analysis of P53 protein expression was performed on paraffin-embedded tissues from 55 smooth muscle tumors of the gastrointestinal tract, using immunofluorescence and flow cytometry. No positive expression was found in normal smooth muscle tissues of the gastrointestinal tract. Over-expression of P53 gene was found in a significantly higher proportion in leiomyosarcomas (90%) and potentially malignant smooth muscle tumors (75%) as compared to leiomyomas (14%) (P < 0.005). The quantitation of P53 expression was found to be progressively enhanced in the sequence from leiomyoma through potentially malignant smooth muscle tumor to leiomyosarcoma (P < 0.005). It was markedly over-expressed when the mitotic counts ranged from one to more than one per 10 high power fields (P < 0.005) or the mild cytologic atypia was found (P < 0.005). The five-year survival rate was significantly higher in patients with low-expression of P53 than in those with over-expression of P53 (P < 0.005). It was suggested that P53 over-expression might be associated with the transformation of leiomyoma into leiomyosarcoma and could be used as an objective parameter in distinguishing the malignant from the benign and predicting the prognosis of patients with smooth muscle tumors of the gastrointestinal tract.

Adolescent↗

PubMed1995

[Gastrointestinal smooth muscle tumors: a study of DNA content and the association with clinicopathology].

Quantitative analysis of DNA content and ploidy pattern was performed in 84 cases of smooth muscle tumors of the gastrointestinal tract by flow cytometry, and the results were correlated with histopathologic features. The aneuploid rate was found to be 0, 50%, 75%, 92.8% and 100% respectively in the tumors with mitotic figures of 0,1, 2-4, 5-9, or over 10/10HPF (P < 0.005), as to be 3.2%, 60%, 95.8% and 100% in tumors with zero, mild, moderate or severe cellular atypia (P < 0.005). There were no aneuploidy in the 30 cases of leiomyomas, but all 40 cases of leiomyosarcomas were aneuploidy, while in the 14 potentially malignant smooth muscle tumors, there were 4 cases of aneuploidy (P < 0.01). The aneuploid rate was significantly higher in tumors with contiguous organ inveasion or distant metastasis (79.5%), and in tumors of large size (64.5%) as well as in tumors with center necrosis (68.3%). 5 year survival rate in patients with aneuploid tumors (41.6%) was significantly lower than those with diploid tumors (96%) (P < 0.005). The results suggested that DNA content and ploidy pattern be used as an objective parameter to distinguish malignants from benigns and predict the prognosis of patients with gastrointestinal smooth muscle tumors.

Adolescent↗

PubMed1994

Determination of trace impurities of peptides and alkaloids by capillary electrophoresis-ion spray mass spectrometry.

Two different mixtures have been analyzed by CE-UV-MS using selected ion monitoring (SIM) conditions to evaluate whether this technique can detect trace impurities in such mixtures. The first mixture consisted of two bioactive peptide analogues which included Lys-bradykinin (kallidin) and Met-Lys-bradykinin. The presence of 0.1% Lys-bradykinin was detected by SIM CE-MS but not by CE-UV at the 0.1% level as it migrated from the capillary column prior to the major component, Met-Lys-bradykinin. The second mixture consisted of two antibacterial alkaloids, berberine and palmatine. The presence of 0.15% palmatine was detected by CE-UV and SIM CE-MS at the 0.15% level as it migrated from the capillary column following the major component, berberine. These results suggest that SIM CE-MS offers the necessary separation efficiencies and sensitivity to provide a complementary analytical determination of trace components in such sample mixtures.

Alkaloids↗

PubMed1994

Two independent and interactive DNA-binding subdomains of the Pax6 paired domain are regulated by alternative splicing.

Vertebrate Pax proteins share a conserved 128-amino-acid DNA-binding motif, the paired domain. The PAX6 gene, which is mutated in the murine Small eye and human aniridia developmental defects, also encodes a second protein with a 14-amino-acid insertion in the paired domain. This protein, which arises by alternative mRNA splicing, exhibits unique DNA-binding properties. Unlike other paired domains, which bind DNA predominantly by their amino termini, the extended Pax6 paired domain interacts with DNA exclusively through its carboxyl terminus. This property can be stimulated by deletion of 30 amino-terminal residues from the Pax6 or Pax2 paired domains. Thus, the insertion acts as a molecular toggle to unmask the DNA-binding potential of the carboxyl terminus. The functional nonequivalence of the two Pax6 proteins is underscored by a T-->C mutation at position -3 of the alternative splice acceptor site that changes the ratio of the two isoforms and causes a distinct human ocular syndrome.

Alternative Splicing↗

PubMed1994

Identification of a Pax paired domain recognition sequence and evidence for DNA-dependent conformational changes.

Pax genes encode a family of developmentally regulated transcription factors that have been implicated in a number of human and murine congenital disorders, as well as in tumorigenesis (Gruss, P., and Walther, C. (1992) Cell 69, 719-722; Hill, R., and van Heyningen, V. (1992) Trends Genet. 8, 119-120; Chalepakis, G., Tremblay, P., and Gruss, P. (1992) J. Cell Sci. Suppl. 16, 61-67; Maulbecker, C. C., and Gruss, P. (1993) EMBO J. 12, 2361-2367; Walther, C., Guenet, J. L., Simon, D., Deutsch, U., Jostes, B., Goulding, M. D., Plachov, D., Balling, R., and Gruss, P. (1991) Genomics 11, 424-434; Barr, R. G., Galili, N., Holick, J., Biegel, J. A., Rovera, G., and Emanuel, B. S. (1993) Nature Genet. 3, 113-117). These genes are defined by the presence of an evolutionarily conserved DNA binding domain, termed the paired domain. The structure and the DNA binding characteristics of the paired domain remain largely unknown. We have utilized repetitive rounds of a polymerase chain reaction-based selection method to identify the optimal DNA binding sequences for the Pax-2 and Pax-6 paired domains. The results suggest that the paired domain family of peptides bind similar DNA sequences. Identification of this binding site has revealed an important structural clue regarding the mechanism of paired domain binding to DNA. CD and NMR structural analyses of the purified Pax-6 paired domain reveal it to be largely structureless in solution. Upon binding the recognition sequence, the complex becomes markedly less soluble and displays CD spectroscopic evidence of significant alpha-helical structure.

Animals↗

PubMed1994

Isolation and characterization of the mouse heme oxygenase-1 gene. Distal 5' sequences are required for induction by heme or heavy metals.

Mouse genomic fragments encoding heme oxygenase-1 (HO-1) were isolated from a recombinant lambda library by in situ plaque hybridization. The mouse HO-1 gene, approximately 7 kilobase pairs (kbp) in length, is organized into 5 exons and 4 introns. The primary structure of the exons and 1287 base pairs (bp) of the 5'-flanking region was determined. The deduced amino acid sequence of the mouse HO-1 gene is identical to that of p32, initially identified as a stress-induced protein in mouse BALBc/3T3 cells. A single, major transcription initiation site is utilized for constitutive and heme- or metal-induced expression of the HO-1 gene in mouse hepatoma (Hepa) cells. The transcriptional activity of the 5'-flanking region was examined by transient expression assays using the chloramphenicol acetyltransferase gene as the reporter gene. Basal promoter activity in several cell lines was localized to within 149 bp of the upstream sequence by deletion analysis. This proximal promoter region of the mouse HO-1 gene contains several sequence elements that are not only conserved in both the rat and human HO-1 genes but also resemble consensus binding sites of various transcription factors including AP-1, AP-4, C/EBP and c-Myc:Max/USF. Heavy metals activate HO-1 gene transcription and the rat gene contains a putative metal regulatory element (Müller, R. M., Taguchi, H., and Shibahara, S. (1987) J. Biol. Chem. 262, 6795-6802) that is completely conserved in the mouse gene. Transient expression analyses, however, indicate that this sequence, which contains a core heptanucleotide, TGCACTC, identical to that of the strongest metal regulatory element of the mouse metallothionein-1 gene, is not responsive to Cd2+ or Zn2+. Stable transfection of constructs containing the entire mouse HO-1 gene and various portions of the 5'-flanking region into rat C6 glioma cells and simultaneous, quantitative analysis of the mouse and rat HO-1 mRNAs indicate that distal 5' sequences, between positions -3.5 and -12.5 kbp, are required for induction of mouse HO-1 gene transcription by both heme and heavy metals. A 5-7-fold difference in the levels of induction between stably integrated and transiently expressed mouse HO-1 gene constructs is observed in this cell line.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1994

Generation and characterization of dihydroxycarbene, HO-C-OH, by neutralization/reionization mass spectrometry.

Dihydroxycarbene is produced in the gas phase by neutralization of the HO-C-OH+. radical cation which is formed by dissociative electron ionization of oxalic acid. Reionization approximately 0.3 microseconds later shows that HO-C-OH can survive intact and, thus, exists as a stable species with appreciable barriers for dissociation or rearrangement to formic acid (HCOOH). Within the time scale of the experiment, a small fraction of the carbene decomposes to H2O+CO. Comparison of the experimental results with ab initio theory shows that the dissociating HO-C-OH molecules are generated in the electronically excited triplet state, while the large amount of surviving carbene molecules is formed in the singlet ground state.

Mass Spectrometry↗

PubMed1994

Dissection of the Drosophila paired protein: functional requirements for conserved motifs.

The Drosophila paired gene encodes three conserved motifs: a homeodomain, paired domain and PRD (his/pro) repeat. To investigate the functional importance of the PRD repeat and paired domain, we tested deletion mutants using an ectopic expression assay in embryos. Our results suggest that the PRD repeat is not required for the in vivo regulation of the target genes, engrailed and gooseberry. However, the PRD repeat appears to be embedded within a proline-rich transcriptional activation domain required for the regulation of these genes. Our analysis of the paired domain indicated that its N-terminal half, which is required for DNA binding in vitro, is also required for in vivo function, whereas surprisingly, the C-terminal half is dispensable for the regulation of engrailed and gooseberry.

Amino Acid Sequence↗

PubMed1994

Evidence for a close phylogenetic relationship between Melissococcus pluton, the causative agent of European foulbrood disease, and the genus Enterococcus.

The 16S rRNA gene sequence of Melissococcus pluton, the causative agent of European foulbrood disease, was determined in order to investigate the phylogenetic relationships between this organism and other low-G + C-content gram-positive bacteria. A comparative sequence analysis revealed that M. pluton is a close phylogenetic relative of the genus Enterococcus.

Animals↗