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Biomedical subjects

I Schwartz

Publications and source records attributed to I Schwartz.

At least 55 records · Page 3Linked to original sources
PubMed1997

Inhibition of constitutive nitric oxide synthase (NOS) by nitric oxide generated by inducible NOS after lipopolysaccharide administration provokes renal dysfunction in rats.

Excess NO generation plays a major role in the hypotension and systemic vasodilatation characteristic of sepsis. Yet the kidney response to sepsis is characterized by vasoconstriction resulting in renal dysfunction. We have examined the roles of inducible nitric oxide synthase (iNOS) and endothelial NOS (eNOS) on the renal effects of lipopolysaccharide administration by comparing the effects of specific iNOS inhibition, -N6-(1-iminoethyl)lysine (L-NIL), and 2,4-diamino6-hydroxy-pyrimidine vs. nonspecific NOS inhibitors (nitro- -arginine-methylester). cGMP responses to carbamylcholine (CCh) (stimulated, basal) and sodium nitroprusside in isolated glomeruli were used as indices of eNOS and guanylate cyclase (GC) activity, respectively. LPS significantly decreased blood pressure and GFR (112+/-4 vs. 83+/-4 mmHg; 2.66+/-0.29 vs. 0. 96+/-0.22 ml/min, P < 0.05) and inhibited the cGMP response to CCh. GC activity was reciprocally increased. L-NIL and 2, 4-diamino-6-hydroxy-pyrimidine administration prevented the decrease in GFR (2.71+/-0.28 and 3.16+/-0.18 ml/min, respectively), restored the normal response to CCh, and GC activity was normalized. In vitro application of L-NIL also restored CCh responses in LPS glomeruli. Neuronal NOS inhibitors verified that CCh responses reflected eNOS activity. L-NAME, a nonspecific inhibitor, worsened GFR (0.41+/-0.15 ml/min), a reduction that was functional and not related to glomerular thrombosis, and eliminated the CCh response. No differences were observed in eNOS mRNA expression among the experimental groups. Selective iNOS inhibition prevents reductions in GFR, whereas nonselective inhibition of NOS further decreases GFR. These findings suggest that the decrease in GFR after LPS is due to local inhibition of eNOS by iNOS, possibly via NO autoinhibition.

Animals↗
PubMed1997

Cellular changes and induction of apoptosis in human promyelocytic HL-60 cells infected with the agent of human granulocytic ehrlichiosis (HGE).

Human granulocytic ehrlichiosis (HGE) is an emerging and occasionally fatal human infectious disease whose pathogenesis is largely unknown. Goodman et al. (1) recently described the successful cultivation of the HGE infectious agent in human promyelocytic HL-60 leukemic cells. It was reported in the same study that infectivity invariably led to host cell death, although the mechanism by which HGE infection triggers cellular self-destruction is as yet undetermined. In this communication, we show that in vitro passage of HGE pathogen-infected blood elicits a significantly dysfunctional G1-to-S transition. Moreover, we provide evidence that the cytopathic properties of the HGE pathogen are attributed to its ability to induce apoptosis in host HL-60 cells. Determination of specific protein expression changes by Western blot analysis showed that HGE infection resulted in reduced expression of PCNA and pRB, both of which play a role in cell cycling. Moreover, the steady state level of bcl-2, which protects eukaryotic cells against apoptosis, is suppressed by exposure to the HGE agent. These results suggest that this pathogen HGE induces apoptosis in HL-60 cells by a mechanism involving the shut-off of multiple cell cycle and apoptosis regulatory events.

Apoptosis↗
PubMed1997

Molecular cloning and characterization of Borrelia burgdorferi rpoB.

Borrelia burgdorferi rpoB, the gene encoding the beta-subunit of RNA polymerase, has been cloned and sequenced. The full-length gene encodes a protein of 1154 amino acids with a calculated molecular mass of 129.8 kDa. The amino-acid sequence is 49% identical to the corresponding protein from Escherichia coli. B. burgdorferi rpoB is a component of a gene cluster, which includes rplJ, rplL and rpoC. A temperature-sensitive E. coli rpoB mutant could be complemented by introduction of the B. burgdorferi gene, indicating that the B. burgdorferi rpoB is expressed in E. coli and the beta-subunit can be assembled into functional holoenzyme. The wild-type amino-acid sequence of the B. burgdorferi beta-subunit is consistent with those of spontaneously arising rifampicin-resistant mutants of E. coli and Mycobacterium tuberculosis at certain critical residues. This suggests that the natural resistance of B. burgdorferi to rifampicin may be due to the primary amino-acid sequence of its beta-subunit.

Amino Acid Sequence↗
PubMed1997

Positive Lyme disease serology in patients with clinical and laboratory evidence of human granulocytic ehrlichiosis.

In 10 consecutive patients with an acute febrile illness, human granulocytic ehrlichiosis was confirmed with specific polymerase chain reaction studies, serologic conversion, or both. Although no patients had the clinical features most suggestive of early Lyme disease (eg, erythema migrans or cranial nerve palsy), tests for antibody to Borrelia burgdorferi produced a reaction in most patients. In 6 of 7 patients (86%) with evaluable results, enzyme-linked immunosorbent assay yielded positive or equivocal findings, and an immunoblot technique yielded positive findings in 60% to 90% of patients, depending on the criteria used for interpretation. Inasmuch as approximately 25% of nymphal Ixodes scapularis ticks in Westchester County, New York, are infected with B burgdorferi, the probability that at least 9 of these patients were coinfected with B burgdorferi and human granulocytic ehrlichiosis by the same tick bite is estimated to be .00003. These observations suggest that serodiagnosis is insufficient to establish the presence of coinfection with B burgdorferi.

Adult↗
PubMed1997

Inhibition of efficient polymerase chain reaction amplification of Borrelia burgdorferi DNA in blood-fed ticks.

Polymerase chain reaction (PCR) analysis is a widely used method for detection of Borrelia burgdorferi DNA in biological specimens, including ticks. Studies have demonstrated that substances present in mammalian blood can inhibit PCR amplification. This would limit the utility of PCR for determination of B. burgdorferi infection in engorged ticks that have taken a blood meal from a human or other animal host. To systematically assess the potential for such inhibition, nymphal Ixodes scapularis, which had acquired B. burgdorferi as larvae, were fed on rats. These engorged ticks were lysed in standard PCR lysis buffer and aliquots were subjected to PCR analysis; 0 of 56 were PCR positive. An equivalent cohort of unfed (unengorged) ticks had an infection rate of 19% (11 of 57) as determined by identical PCR analysis (P = 0.0006, by Fisher's exact test). When lysates from the engorged ticks were spiked with the 500 prelysed B. burgdorferi, none of the samples yielded a positive PCR signal, indicating the presence of inhibitory substances. Consistent with this observation, PCR amplification of the original engorged tick lysates after extraction with a DNA extraction kit, resulted in detection of B. burgdorferi DNA in 13 specimens (23%). Furthermore, when 500 prelysed B. burgdorferi were added to the treated extracts, all samples (56 of 56) were PCR positive. Thus, extraction resulted in removal of inhibitors of PCR amplification present in unprocessed engorged tick lysates. Furthermore, additional titration experiments showed that some inhibitory substances may also be present in unfed ticks, although this inhibition does not completely prevent detection of B. burgdorferi DNA in unprocessed lysates. This study clearly demonstrates that inhibitors of PCR amplification are present in engorged ticks and prevent accurate determination of B. burgdorferi infection rates by this method unless steps are taken to remove such inhibitors.

Animals↗
PubMed1996

Serologic evidence for bovine immunodeficiency virus infection in France.

We report herein on the first serologic detection of antibodies to bovine immunodeficiency virus (BIV) in France. Serum samples from dairy and beef cattle from southwestern and western France (Landes and Vendée) were tested using a western blot assay with a recombinant 53 kDa gag precursor derived from the Louisiana BIV R29 isolate. We performed our study on the oldest animals from 37 different herds that were under serologic follow up for previous bovine leukemia virus infection. Overall, 398 selected bovine sera were assayed and 15 serum samples from 8 herds reacted with the recombinant 53 kDa BIV R29 gag. Interestingly, reactions obtained with French sera were weaker than with positive Louisiana sera, a finding that may indicate the occurrence of distinct French and Louisiana BIV variants.

Animals↗
PubMed1996

Intraovarian blood flow during spontaneous and stimulated cycles.

This study was undertaken to investigate intraovarian blood flow during the early follicular, periovulatory and mid-luteal phases, during spontaneous and stimulated ovarian cycles. Transvaginal ultrasonography with colour flow imaging was used to measure the pulsatility index in eight patients with spontaneous cycles, 20 patients undergoing induction of ovulation with clomiphene citrate and 11 patients undergoing controlled ovarian hyperstimulation for in-vitro fertilization (IVF)-embryo transfer with gonadotrophin-releasing hormone analogue, human menopausal gonadotrophin, human chorionic gonadotrophin. All patients were studied during the early follicular, periovulatory and mid-luteal phase. Intraovarian blood flow velocity waveforms were found in 20% of cases at the early follicular phase, in 56% during the periovulatory phase and in 85% during the mid-luteal phase (P < 0.001). Pulsatility index at the early follicular phase was found to be 1.05 +/- 0.22, during the periovulatory phase 0.99 +/- 0.22 and during the mid-luteal phase 0.85 +/- 0.22. The appearance of intraovarian flow velocity waveforms may suggest either ovarian neovascularization or final follicular maturation or luteinization, which can be detected or measured by Doppler technology.

Blood Flow Velocity↗
PubMed1996

Molecular typing of Borrelia burgdorferi from Lyme disease patients by PCR-restriction fragment length polymorphism analysis.

Ninety-three Borrelia burgdorferi isolates obtained from erythema migrans lesions or blood of Lyme disease patients in Westchester County, N.Y., between 1991 and 1994 were characterized by PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the 16S-23S rRNA gene spacer. All isolates could be classified into three distinct RFLP types. Among the 82 skin biopsy isolates studied, 21 (25.6%) were type 1, 37 (45.1%) were type 2, and 21 (25.6%) were type 3. Three (3.7%) cultures contained a mixture of two isolates with distinct RFLP types. The 11 isolates cultured from blood showed a similar predominance of RFLP type 2 (6 of 11; 54.5%) relative to types 1 (2 of 11; 18.2%) and 3 (3 of 11; 27.3%). For one patient both skin and blood isolates were cultured, and RFLP analysis revealed that these isolates differed from one another. This study demonstrates that there is genotypic heterogeneity in B. burgdorferi strains infecting Lyme disease patients, and this typing approach may allow differentiation of isolates with various degrees of pathogenic potential.

Bacterial Typing Techniques↗
PubMed1996

Confirmation of Borrelia burgdorferi spirochetes by polymerase chain reaction in placentas of women with reactive serology for Lyme antibodies.

The purpose of our study was to determine whether Borrelia burgdorferi spirochetes were present in placentas of asymptomatic women with reactive Lyme serology using a silver stain, and to confirm the identity of the spirochetes by polymerase chain reaction (PCR). Sixty placentas of asymptomatic women with ELISA-positive or-equivocal serology for Lyme antibodies during pregnancy were examined for spirochetes using a silver stain. The results of the ELISA serology were confirmed by Western blot analysis. PCR amplification for B. burgdorferi was performed on placentas identified to have spirochetes and on a group of placentas negative for spirochetes. Spirochetes were identified by silver staining in 3 (5%) of the 60 placentas. PCR confirmed B. burgdorferi nucleotide sequences in 2 of the placentas. The 5 women had equivocal Lyme ELISA and negative syphilis serology. The results of the Western blot analysis were negative in 2 cases and indeterminate in 1 case. Six controls were negative for spirochetes by silver staining and PCR. A normal perinatal outcome was observed in all cases. Spirochetes identified in placental tissue of pregnancies with reactive Lyme serology were confirmed by PCR to be B. burgdorferi. There was no relationship between the presence of placental spirochetes and the results of Lyme serology or the pregnancy outcome.

Blotting, Western↗
PubMed1995

Two geographically distinct isolates of Borrelia burgdorferi from the United States share a common unique ancestor.

The genetic diversity of Borrelia burgdorferi isolates from several geographic regions was evaluated by nucleotide sequence analysis of the genes encoding 23S ribosomal RNA and outer surface protein A. Comparison of nucleotide sequences spanning 738 bp of the 23S ribosomal DNA from two unusual isolates, DN127 (Del Norte County, California) and 25015 (Millbrook, New York), to homologous sequences from other B. burgdorferi isolates from the United States and Russia identified several nucleotide sequence polymorphisms that are unique to these two isolates. Sequence analysis of a 615 nucleotide segment of the gene encoding outer surface protein A also revealed greater similarity of strains DN127 and 25015 (94.1%) compared to other US and Eurasian isolates. These data were further corroborated by genomic macrorestriction analysis, in which DN127 and 25015 demonstrated unique restriction digestion patterns. Our findings suggest that substantial genetic diversity of B. burgdorferi, rivaling that of European strains, exists among isolates from the United States. Strains DN127 and 25015 are unique among all B. burgdorferi isolates tested to date, and though isolated from opposite longitudinal extremes of the North American continent, are closely related.

Bacterial Outer Membrane Proteins↗
PubMed1995

rRNA operon multiplicity in Escherichia coli and the physiological implications of rrn inactivation.

Here we present evidence that only five of the seven rRNA operons present in Escherichia coli are necessary to support near-optimal growth on complex media. Seven rrn operons are necessary, however, for rapid adaptation to nutrient and temperature changes, suggesting it is the ability to adapt quickly to changing environmental conditions that has provided the selective pressure for the persistence of seven rrn operons in E. coli. We have also found that one consequence of rrn operon inactivation is a miscoordination of the concentrations of initiation factor IF3 and ribosomes.

Adaptation, Physiological↗
PubMed1995

Molecular typing of Borrelia burgdorferi sensu lato by PCR-restriction fragment length polymorphism analysis.

The etiologic agent of Lyme borreliosis, Borrelia burgdorferi sensu lato, has been isolated from many biologic sources in North America and Eurasia, and isolates have been divided into three distinct genospecies (B. burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii). In order to explore the possible association of genospecies with disease manifestation, 60 isolates of B. burgdorferi sensu lato were subjected to 5S rDNA-linked restriction fragment length polymorphism (RFLP) analysis. The results confirmed earlier studies which indicated that virtually all North American isolates are B. burgdorferi sensu stricto, whereas Eurasian strains fall into all three genospecies. Thirty-five isolates were further characterized by PCR amplification of a region of the 16S-23S rDNA spacer and HinfI digestion of the products. This method resulted in the subdivision of B. burgdorferi sensu stricto into two distinct PCR-RFLP types. In contrast, B. garinii isolates all displayed an identical pattern. Additionally, a number of previously unclassified North American isolates (25015, DN127, 19857, 24330) showed distinctively different PCR-RFLP patterns. The application of this method for the typing of uncultured B. burgdorferi directly in biologic samples was demonstrated by analysis of several field-collected Ixodes scapularis tick specimens. The described PCR-RFLP technique should allow for the direct and rapid molecular typing of B. burgdorferi-containing samples and facilitate studies of the relationship between spirochete genotype and clinical disease.

Animals↗
PubMed1995

Identification of novel insertion elements, restriction fragment length polymorphism patterns, and discontinuous 23S rRNA in Lyme disease spirochetes: phylogenetic analyses of rRNA genes and their intergenic spacers in Borrelia japonica sp. nov. and genomic group 21038 (Borrelia andersonii sp. nov.) isolates.

Borrelia spp. associated with Lyme disease possess an rRNA gene organization consisting of a single 16S rRNA gene followed by a spacer of several kilobases and a tandem repeat of a 23S (rrl)-5S (rrf) rRNA gene cluster. The restriction fragment length polymorphism (RFLP) patterns for these genes have been widely used to classify Lyme disease spirochete isolates. We analyzed the rRNA gene organization and sequences for two Ixodes ovatus isolates from Japan (IKA2 and HO14) and two group 21038 isolates associated with Ixodes dentatus ticks or rabbits from North America (isolates 21038 and 19857). This analysis revealed unique polymorphisms not previously described in other Lyme disease spirochete isolates. The molecular basis of these polymorphisms was determined by Southern blotting and PCR analyses. Only one continuous copy of the rrl-rrf gene cluster was identified in isolates IKA2, 19857, and 21038. The second rrl-rrf gene cluster is entirely absent from the IKA2 genome. In isolates 19857 and 21038, an intervening sequence is present, resulting in a fragment rrlB gene. The insertion site of this intervening sequence element differed in each isolate. While isolates 19857 and 21038 were found to carry a fragmented rrlB gene, they lacked rrfB. To determine if these rRNA polymorphisms were indicative of an underlying phylogenetic divergence, sequence analysis of the 16S rRNA (rrs) genes was conducted. The phylogenies inferred from rrs sequence analysis suggest that the polymorphisms resulted from recent mutational events. In addition, the phylogenetic analyses also support the proposed species status of Borrelia japonica sp. nov. and indicate that isolates of genomic group 21038 belong to a previously undescribed species for which we propose the nomenclature Borrelia andersonii sp. nov.

Base Sequence↗