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Biomedical subjects

L Navarro

Publications and source records attributed to L Navarro.

51 records · Page 3Linked to original sources
PubMed1993

Diagnosis and monitoring of Clostridium difficile infections with the polymerase chain reaction.

Toxigenic Clostridium difficile is the etiologic agent of pseudomembranous colitis. We have developed an assay system for the rapid direct detection of toxigenic C. difficile in human stool samples. After DNA extraction, polymerase chain reaction (PCR) amplification is undertaken with primers targeting specific sequences in the C. difficile 16S rRNA gene. Next, toxigenic strains of C. difficile are distinguished from nontoxigenic strains by PCR amplification of toxin A and/or B gene sequences. This study included 12 patients with C. difficile colitis, seven of whom had clinical relapses after discontinuation of vancomycin therapy. We detected toxigenic C. difficile in stools from four (57%) of these seven patients before relapse--at a time when no toxin B was detectable in stools and results of anaerobic culture were negative. The PCR assay is 100-fold more sensitive than anaerobic culture methods. The course of the infection in one patient (both during and after therapy) was monitored by the PCR technique. The multigene analysis approach permitted the detection of colonization with a nontoxigenic strain when this patient's relapses had cleared. This clinically applicable assay allows earlier detection of infection with toxigenic C. difficile. The result is more timely therapeutic intervention.

Aged↗
PubMed1993

Characterization of field strains of group A bovine rotaviruses by using polymerase chain reaction-generated G and P type-specific cDNA probes.

Dot and Northern blot hybridization assays were used to analyze field strains of group A bovine rotaviruses (BRVs) by using nucleic acid probes representing P and G type specificities. The probes were prepared by polymerase chain reaction amplification of hyperdivergent regions of the cloned VP4 (nucleotides 211 to 686) and VP7 (nucleotides 51 to 392) genes from four serotypically distinct (in P or G types) strains of rotaviruses: NCDV (G6, P1), IND (G6, P5), 69M (G8, P10), and Cr (G10, P11). The P and G type cDNA probes were radiolabeled with [32P]dCTP and hybridized with RNA extracted from reference cell culture-passaged rotavirus strains or the field samples. The field samples were obtained from young diarrheic calves from Ohio, Nebraska, Washington State, and Canada. The cDNA probes were specific for their respective G or P types on the basis of analysis of known P and G type reference strains. The G typing analysis of 102 field samples revealed that 36.3% (37 of 102) were G6, 2.9% (3 of 102) were G8, 12.7% (13 of 102) were G10, and 23.5% (24 of 102) were untypeable. The P typing results for 93 samples indicated that 2.2% (2 of 93) were P1 (NCDV-like), 20.4% (19 of 93) were P5 (UK-like), 9.3% (10 of 93) were P11 (B223-like), and 40.8% (38 of 93) were untypeable. This is the first report of the identification among BRV strains in North America of a G type other than G6 or G10. Our report further confirms that G6, P5 rotaviruses are predominant among the BRV field strains that we examined, and the P types of these strains differ from that of the BRV vaccine strain used in the United States (G6, P1). The large number of untypeable G (23.5%) and P (40.8%) types suggests that other or new P and G types exist among BRV field strains.

Animals↗
PubMed1992

Isolation of the plasma membrane and organelles from Chinese hamster ovary cells.

Two methods are described enabling the plasma membrane from Chinese hamster ovary (CHO) cells to be obtained rapidly, relatively pure and with a good yield. In both cases, cells were disrupted by nitrogen cavitation in an isoosmotic buffer either at pH 5.4 or at pH 7.4. In the first approach, cells were lysed at pH 7.4 and the plasma membrane and cell organelles were isolated on a self-generated gradient of Percoll, at neutral pH. Mitochondria and endoplasmic reticulum were recovered in the denser fractions, plasma membrane fragments were found in the lighter fractions, but always contaminated by lysosomes. Because lysosomes were found to sediment in acidic conditions, cells were lysed at pH 5.4 and presedimentation (1500 x g) of the cell homogenate at the same pH enabled more than 80% of the lysosomes to be removed. Then, ultracentrifugation of the supernatant over a Percoll gradient at neutral pH yielded plasma membrane fractions practically free of lysosomes with an enrichment ratio of 3 and fractions of mitochondria and endoplasmic reticulum with enrichment ratios of 17 and 6, respectively. A major problem was encountered in the final step of elimination of Percoll from the purified plasma membrane fractions. Whatever the technique used for eliminating Percoll, plasma membranes were observed to be contaminated by a Percoll constituent which prevented further purification and biochemical identification of the lipids extracted from these membrane fractions to be carried out. A second method of plasma membrane preparation was tested consisting first in the coating of the cell surface with positive colloidal silica which was stabilized by an anionic polymer. Then, and through differential centrifugations, plasma membrane fractions were easily obtained within less than 1 h, with a yield of 65% and an enrichment ratio of 7. The coating pellicle was quantitatively removed thus enabling any biochemical manipulation of the plasma membrane to be carried out. The lipids present in the plasma membrane of CHO cells were analyzed and are described, both in terms of headgroup and acyl chain composition.

Animals↗
PubMed1991

Neuroendocrine regulation of adrenal 5'-monodeiodination during acute cold exposure in the rat. I. Effects of hypophysectomy.

Circulating levels of T4, T3, corticosterone, noradrenaline, and adrenaline, as well as 5'-monodeiodinase activity (5'-MA) were measured in control and hypophysectomized rats acutely exposed to cold environment (15-120 min, 4 C). In addition to the well known activation of the sympathoadrenomedullary system and the hypothalamic-pituitary-adrenal and-thyroid axes, cold exposure was followed by a rapid and sustained increase of 5'-MA in the hypothalamus, and a byphasic course of activation in the adrenal gland in control rats. The adrenal rapid activation (30 min) corresponded to the medulla and the slower activation (120 min) to the cortex. Both, the basal adrenal 5'-MA and the response to cold in adrenal and hypothalamus were 2-fold higher in hypophysectomized rats compared to control. The time course of enzyme activation in these structures suggests that: 1) organ-specific increases in 5'-MA may be associated to a simultaneous rise in their metabolic and/or functional activity, 2) the triggering mechanisms involves an immediate sympathetic signal activating the hypothalamic-adrenal medulla response and a pituitary signal eliciting a slower adrenocortical response, and 3) the compensatory sympathetic hyperactivity after panhypopituitarism contribute to enhance both the adrenal enzyme basal activity and the hypothalamic and adrenal hyperresponse to cold stress.

Adrenal Cortex↗
PubMed1989

Inverse correlation of expiratory lung flows and sputum eosinophils in status asthmaticus.

Seventy-six consecutive patients admitted to Los Angeles County General Hospital with acute asthma were studied. Blood and sputum smears for cell counts were obtained on all patients within 12 hours of admission. Fifty-one (67%) patients were able or willing to perform spirometry and flow/volume curves in the first 24 hours of hospitalization. The severity of airway obstruction as assessed by forced expiratory volume in one second (FEV1), maximum mid-expiratory flow rate (MMFR), and forced vital capacity (FVC) was compared with blood and sputum eosinophil counts. Although there was no relation between the blood eosinophilia and airway obstruction, an inverse relationship between the number of eosinophils in the sputum and airway flow rates was observed. Higher percentages of sputum eosinophils were associated with diminished flow rates. We believe that sputum eosinophils may be helpful in the initial assessment of severe bronchial asthma.

Adolescent↗
PubMed1988

Effect of seeding density and time in culture on vascular smooth muscle cell proteins.

The purpose of this work was to determine the effect of seeding density and time in culture on quantitative and qualitative characteristics of myosin in primary cultures of vascular smooth muscle cells (VSMCs). Enzymatically dispersed VSMCs from femoral arteries and saphenous veins of adult dogs were seeded at a density of 10(3)-10(5) cells/cm2 and assayed after 7 days or at 10(5) cells/cm2 and assayed between 1 and 10 days. Myosin, actin, and total protein contents as well as electrophoretic and immunoreactive characteristics of myosin heavy chains (MHCs) were determined. Total and contractile protein contents were independent of seeding density and increased with time in culture. Freshly dispersed cells exhibited two MHCs (MHC-1 and MHC-2) but, within 24 h after culturing, only cells attached to the dish expressed a third protein band (MHC-3), which had electrophoretic mobility and immunoreactivity similar to purified platelet MHC. MHC-3 appeared before onset of cell division and, by 4 days in culture when cells were proliferating, became the dominant MHC form. Loss of MHC-1 and MHC-2 could be prevented by growing cells in a serum-free, defined media that prevented proliferation. These data indicate that seeding density does not affect myosin content, but that with time in culture expression of a MHC with characteristics similar to nonmuscle myosin occurs. Expression of MHC-3 is associated with cell attachment, whereas loss of MHC-1 and MHC-2 requires proliferation.

Actins↗
PubMed1986

Incidence and mortality of adult respiratory distress syndrome: a prospective analysis from a large metropolitan hospital.

We examined the incidence and mortality of adult respiratory distress syndrome (ARDS) in patients receiving emergency medical care at a large metropolitan medical center. The patients were classified into eight high-risk categories and monitored prospectively until discharge or death. Over a period of 12 months, 11,112 such patients entered the emergency room. Of 4926 who were admitted to the hospital acutely ill, 90 (2%) developed ARDS. Thirty-six percent of these survived. ARDS occurred in 25% of patients admitted with acquired immune deficiency syndrome and Pneumocystis carinii pneumonia, and mortality in these patients was 86%. Aspiration pneumonia was the primary cause of ARDS in 37% of the patients, who also had a similar mortality rate (85%). Mortality was also high in patients with multisystem organ failure or with ARDS and disseminated intravascular coagulation. These results indicate that there probably is a relatively low incidence of ARDS among hospital patients admitted with high-risk diagnoses.

Acquired Immunodeficiency Syndrome↗
PubMedDate not supplied

Cryopreservation of ovules and somatic embryos of citrus using the encapsulation-dehydration technique.

Cryopreservation of ovules and somatic embryos from several genotypes of citrus was achieved using the encapsulation-dehydration technique. Survival of cryopreserved ovules was occasional and erratic after different pregrowth conditions in liquid medium with 0.75M, 1M or up to 1.25M sucrose. An efficient cryopreservation protocol was established for somatic embryos derived from two embryogenic sources (ovules and cut thin layer explants from stigma, style and ovaries). High survival rates (75-100%) were consistently obtained after 1 day pregrowth in 0.75M sucrose, desiccation down to 20-25% moisture content in the beads and direct immersion in liquid nitrogen. The histological study showed that embryos subjected to the encapsulation-dehydration, accumulated high sucrose levels which appear to ensure the recovery of the whole embryo after cryopreservation.

Cell Survival↗