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Biomedical subjects

C Castro

Publications and source records attributed to C Castro.

At least 37 records · Page 2Linked to original sources
PubMed2001

Hepatitis C virus RNA-dependent RNA polymerase (NS5B) as a mediator of the antiviral activity of ribavirin.

Ribavirin is administered in combination with interferon-alpha for treatment of hepatitis C virus (HCV) infection. Recently, we demonstrated that the antiviral activity of ribavirin can result from the ability of a viral RNA polymerase to utilize ribavirin triphosphate and to incorporate this nucleotide with reduced specificity, thereby mutagenizing the genome and decreasing the yield of infectious virus (Crotty, S., Maag, D., Arnold, J. J., Zhong, W., Lau, J. Y., Hong, Z., Andino, R., and Cameron, C. E. (2000) Nat. Med. 6, 1375-1379). In this study, we performed a quantitative analysis of a novel HCV RNA polymerase derivative that is capable of utilizing stably annealed primer-template substrates and exploited this derivative to evaluate whether lethal mutagenesis of the HCV genome is a possible mechanism for the anti-HCV activity of ribavirin. These studies demonstrate HCV RNA polymerase-catalyzed incorporation of ribavirin opposite cytidine and uridine. In addition, we demonstrate that templates containing ribavirin support CMP and UMP incorporation with equivalent efficiency. Surprisingly, templates containing ribavirin can also cause a significant block to RNA elongation. Together, these data suggest that ribavirin can exert a direct effect on HCV replication, which is mediated by the HCV RNA polymerase. We discuss the implications of this work on the development of nucleoside analogs for treatment of HCV infection.

Antiviral Agents↗
PubMed2001

A novel mechanism to ensure terminal initiation by hepatitis C virus NS5B polymerase.

Hepatitis C virus (HCV) nonstructural protein 5B (NS5B) RNA-dependent RNA polymerase (RdRp) has acquired a unique beta-hairpin in the thumb subdomain which protrudes toward the active site. We report here that this beta-hairpin plays an important role in positioning the 3' terminus of the viral RNA genome for correct initiation of replication. The presence of this beta-hairpin interferes with polymerase binding to preannealed double-stranded RNA (dsRNA) molecules and allows only the single-stranded 3' terminus of an RNA template to bind productively to the active site. We propose that this beta-hairpin may serve as a "gate" which prevents the 3' terminus of the template RNA from slipping through the active site and ensures initiation of replication from the terminus of the genome. This hypothesis is supported by the ability of a beta-hairpin deletion mutant that utilizes dsRNA substrates and initiates RNA synthesis internally. The proposed terminal initiation mechanism may represent a novel replication strategy adopted by HCV and related viruses.

3' Untranslated Regions↗
PubMed2001

Asymptomatic lingual tonsillar hypertrophy and difficult airway management: a report of three cases.

PURPOSE: To report on the airway management of three cases of asymptomatic lingual tonsillar hypertrophy (LTH). MATERIAL: On three separate occasions, patients presenting for elective surgery were subsequently found to have asymptomatic LTH. In all cases preoperative airway examination was essentially unremarkable and no unusual difficulties were anticipated. In the first case, despite an inability to visualize the glottic opening, the patient was intubated successfully on the initial attempt and had no further problems in the perioperative period. In the second case, neither direct laryngoscopy, utilizing the MacIntosh and McCoy blades, nor fibreoptic visualization enabled successful intubation. Ventilation was maintained with a laryngeal mask airway (LMA) until the anesthetic was reversible. Upon awakening and removal of the LMA, the patient totally obstructed and could not be ventilated, necessitating emergency cricothyroidotomy. The third patient was an elderly gentleman in whom successful intubation was eventually achieved, with considerable difficulty, by the otorhinolaryngologist (ENT surgeon) utilizing a straight blade. On a second occasion, he was again intubated by the same ENT surgeon, this time utilizing the anterior commissure blade. All three patients were subsequently discharged without further sequelae. CONCLUSION: Asymptomatic LTH can cause varying degrees of unexpected difficulty in securing the airway and, at present, no single method will necessarily improve the chances of successful intubation. Therefore, strategies to manage unanticipated difficult intubation secondary to supraglottic airway pathology need to be performed and practiced, including the establishment of a transtracheal airway.

Adult↗
PubMed2001

Aortic smooth muscle cell proliferation and endothelial nitric oxide synthase activity in fructose-fed rats.

The aim of this study was to evaluate the proliferative behavior of vascular smooth muscle cells in primary culture (pC-SMC) and the endothelial nitric oxide synthase (eNOS) activity in the endothelial lining of the aorta of fructose-fed rats (FFR). This is an experimental model of syndrome X, a cluster of cardiovascular risk factors including hyperinsulinemia, insulin resistance, and hypertension that has been suggested to be of pathophysiologic importance for the development of atherosclerosis. Male Wistar rats were used: Control (n = 12) and FFR (n = 12). After receiving fructose in drinking water (10% w/v) during 8 weeks, biochemical parameters, systolic blood pressure (SBP) and relative heart weight (RHW) were determined. The proliferative effect of 10% fetal calf serum (FCS) was examined in aortic pC-SMC by [3H]thymidine incorporation and by cell counting. Ca2+/calmodulin-dependent NOS activity was estimated in aortic endothelial lining and in heart tissue homogenates by conversion of [3H]arginine into [3H]citrulline. Fructose-fed rats showed hyperinsulinemia (P = .0263), altered glucose tolerance test (P < .001), higher SBP (P < .0001), and RHW (P = .0145), compared to control rats. These animals also showed an increase of 10% FCS-induced [3H]thymidine incorporation (P < .0001) and cell number of aortic pC-SMC (P = .0049) and decreased eNOS activity in both aortic endothelium (P = .0147) and cardiac tissue (P < .0001). These data support the hypothesis that syndrome X is associated to changes in SMC proliferation and endothelial dysfunction, which could be involved in the onset or progression of the atherogenic process.

Animals↗
PubMed2001

Binding thermodynamics of the transition state analogue coformycin and of the ground state analogue 1-deazaadenosine to bovine adenosine deaminase.

Binding of the transition state analogue coformycin and the ground state analogue 1-deaazadenosine to bovine adenosine deaminase have been thermodynamically characterized. The heat capacity changes for coformycin and 1-deazaadenosine binding are -4.7 +/- 0.8 kJ/mole-K and -1.2 +/- 0.1 kJ/mole-K, respectively. Since the predominant source of heat capacity change in enzyme interactions are changes in the extent of exposure of nonpolar amino acid side chains to the aqueous environment and the hydrophobic effect is the predominant factor in native structure stabilization, we propose that the binding of either class of ligand is associated with a stabilizing enzyme conformational change with coformycin producing the far greater effect. Analysis of the T dependence of the second order rate constant for formation of the enzyme/coformycin complex further reveals that the conformational change is not rate limiting. We propose that the enzyme may facilitate catalysis via the formation of a stabilizing conformation at the reaction transition state.

Adenosine Deaminase↗
PubMed2001

The mechanism of action of ribavirin: lethal mutagenesis of RNA virus genomes mediated by the viral RNA-dependent RNA polymerase.

Ribavirin (1-beta-D-ribofuranosyl-1,2,4-triazole) is a broad-spectrum antiviral nucleoside that is currently used in combination with interferon-alpha to treat hepatitis C virus infection and as a monotherapy to treat severe cases of respiratory syncytial virus infection and Lassa fever virus infection. The mechanism of action of ribavirin has been studied for decades. These studies have suggested that the antiviral activity of ribavirin may be related to its ability to cause a decrease in intracellular guanosine triphosphate pools, to inhibit capping of viral transcripts or to suppress humoral and cellular immune responses. Last year, another possibility was added to this list. The new proposition is that ribavirin, when converted to the triphosphate, is utilized by the viral RNA-dependent RNA polymerase and causes lethal mutagenesis of the viral genome. In this article, the data supporting this new hypothesis are reviewed. We discuss the implications of these data on alternative explanations for the apparent failure of ribavirin monotherapy in the treatment of hepatitis C virus infection, connections between developmental defects induced by ribavirin and posttranscriptional gene silencing/RNA interference, and the use of lethal mutagenesis and related concepts as strategies for antiviral therapy.

Antiviral Agents↗
PubMed2000

Characterization of the chitin biosynthesis process as a compensatory mechanism in the fks1 mutant of Saccharomyces cerevisiae.

Deletion of the 1,3-beta-D-glucan synthase gene FKS1 in Saccharomyces cerevisiae induces a compensatory mechanism that is reflected in a significant increase in chitin synthase III (CSIII) activity, leading to high rates of chitin synthesis. Deregulation of CSIII activity is mainly due to the intracellular delocalization of Chs3p and Chs4p, the two main components of the CSIII active complex.

Carrier Proteins↗
PubMed2000

Opsonic potential, protective capacity, and sequence conservation of the Treponema pallidum subspecies pallidum Tp92.

By means of a differential screening technique, a 92-kDa antigen, designated Tp92, was identified from Treponema pallidum subspecies pallidum. This protein is similar in sequence to the protective surface antigens D15 from Haemophilus influenzae and Oma87 from Pasteurella multocida. Amino acid sequence analyses revealed a cleavable N-terminal signal sequence and predicted the outer membrane location for Tp92. In support of this, antiserum raised against recombinant Tp92 promotes opsonization and phagocytosis of T. pallidum by rabbit macrophages, and anti-Tp92 reactivity is absent from washed treponemal preparations presumed to be lacking outer membranes. The Tp92 amino acid sequence is 95.5%-100% conserved among 11 strains representing 4 pathogenic treponemes, and immunization with recombinant Tp92 partially protected rabbits from subsequent T. pallidum challenge. These results demonstrate that Tp92 is an invariant, immunoprotective antigen that may be present on the surface of T. pallidum and may represent a potential vaccine candidate for syphilis.

Amino Acid Sequence↗
PubMed2000

Singlet-triplet energy separation of cyclobutylidene

Ab initio (MP2, CCSD(T)) and density functional theory (BLYP, B3LYP) calculations provide insight concerning novel aspects of structure and bonding in cyclobutylidene (1). Singlet cyclobutylidene ((1)1) adopts a bicyclobutane-like structure (C(s) symmetry) that includes a weak, transannular bonding interaction between the carbene carbon and the opposing CH(2) group. Conformational ring inversion in (1)1 occurs through a transition state of C(2)(v)() symmetry (TS(1)1) with an enthalpy barrier of approximately 3 kcal/mol. Stabilization afforded the singlet state by the transannular interaction appears to be largely offset by a loss of hyperconjugative stabilization from the adjacent C-H bonds. Triplet cyclobutylidene ((3)1) exhibits a C(2)(v)() structure and conventional bonding. The triplet state lies 5.9 kcal/mol above the singlet ground state at the CCSD(T)/TZP//CCSD(T)/DZP level of theory. The singlet-triplet energy gap of cyclobutylidene (-5.9 kcal/mol) lies between that of an acyclic analogue, dimethylcarbene (-1.6 kcal/mol), and a highly strained analogue, cyclopropylidene (-13.8 kcal/mol). The magnitude of the energy gap suggests that triplet cyclobutylidene ((3)1) will be thermally accessible under a variety of experimental conditions.

Journal Article↗
PubMed2000

Eleven novel APC mutations identified in Portuguese FAP families.

Germline mutations of the adenomatous polyposis coli (APC) gene are responsible for familial adenomatous polyposis (FAP), an autosomal dominant predisposition to colorectal cancer. In the present study we screened all of the exons of the APC gene in individuals belonging to 85 Portuguese FAP families. We here report eleven novel mutations which are predominantly frameshifts or single base substitutions, resulting in premature stop codons. Hum Mutat 16:178, 2000.

Adenomatous Polyposis Coli↗
PubMed2000

Compartment syndrome following intravenous regional anesthesia.

PURPOSE: To present two cases of upper extremity compartment syndrome following intravenous regional anesthesia. CLINICAL FEATURES: Case 1: A 57-yr-old man presented for surgical release of a left-hand Dupuytren's contracture. The procedure was performed under iv regional anesthesia with 360 mg lidocaine and sedation with 150 microg fentanyl and 1.5 mg midazolam. Tourniquet time was 107 min at a pressure of 260 mmHg using three different tourniquet sites. Within minutes of tourniquet release, increased forearm muscle tension, hand anesthesia, pallor, and limited motor function developed. Serum CK and myoglobin levels rose. Myoglobinuria was present. Several fasciotomies and aggressive fluid therapy were performed. Patient made almost full recovery. Case 2: A 73-yr-old woman with controlled hypertension had Dupuytren fasciotomy of her right hand under iv regional anesthesia with 200 mg lidocaine and sedation using 75 microg fentanyl and 1.5 mg midazolam. Tourniquet time was 64 min at a pressure of 250 mmHg using three different tourniquet sites. The patient complained of pain at the iv site during injection of local anesthetic, third tourniquet inflation and after deflation of tourniquet. Thirty minutes after arrival in PACU, her fingers were bluish. She complained of pain and swelling of the forearm. Under general anesthesia, fasciectomy was performed. Myoglobin and CPK levels rose. CPK MB was high but troponin was negative. Three days later she developed pulmonary embolism. She was heparinized and subsequently discharged home. She recovered completely. CONCLUSION: Compartment syndrome may have a rapid and severe onset. Etiology of our cases is still not established. We postulate that increased tissue pressure may be the cause. The anesthesiologists must be aware of compartment syndrome during regional anesthesia.

Aged↗
PubMed2000

Do changes in physical activity lead to dietary changes in middle and old age?

INTRODUCTION: Research examining the relationship between dietary factors and physical activity patterns has yielded conflicting findings. Few prospective studies have examined whether individuals who change their level of physical activity make corresponding changes in their diet. METHODS: The current study examined, in two randomized clinical trials of physical activity, whether self-report of dietary factors changed over a 1-year period, whether these changes differed by gender or exercise-intervention group, and whether changes in physical activity led to changes in dietary factors. Participants included 268 men and women aged 50-65 years who participated in the Stanford-Sunnyvale Health Improvement Project I (SSHIP-I) and 103 men and women aged 65-82 years who participated in SSHIP-II. In both trials we recruited participants through random-digit telephone dialing and citywide promotion. RESULTS: Overall, participants adopted healthier diets over the 1-year period. The SSHIP-I participants decreased their consumption of total calories, total fat, saturated fat, protein, and cholesterol. The SSHIP-II participants decreased their consumption of high-fat, high-cholesterol foods. The SSHIP-I women were more likely than men to decrease their intake of total fat and protein (p < 0.01), as well as total calories, saturated fat, and cholesterol (p < 0.05). However, we found that changes in physical activity were not associated with changes in self-report measures of dietary habits. CONCLUSIONS: We found no evidence to support the idea that changes in physical activity are a gateway for changes in dietary factors in middle-aged and older adults.

Aged↗
PubMed2000

Personal and environmental factors associated with physical inactivity among different racial-ethnic groups of U.S. middle-aged and older-aged women.

Personal, program-based, and environmental barriers to physical activity were explored among a U.S. population-derived sample of 2,912 women 40 years of age and older. Factors significantly associated with inactivity included American Indian ethnicity, older age, less education, lack of energy, lack of hills in one's neighborhood, absence of enjoyable scenery, and infrequent observation of others exercising in one's neighborhood. For all ethnic subgroups, caregiving duties and lacking energy to exercise ranked among the top 4 most frequently reported barriers. Approximately 62% of respondents rated exercise on one's own with instruction as more appealing than undertaking exercise in an instructor-led group, regardless of ethnicity or current physical activity levels. The results underscore the importance of a multifaceted approach to understanding physical activity determinants in this understudied, high-risk population segment.

Activities of Daily Living↗
PubMed2000

Cutaneous primary paracoccidioidomycosis.

Paracoccidioidomycosis is a common fungal infection in Latin America. Few cases have been described in non-endemic countries and their diagnosis without the correct suspicion, bearing in mind that the latency period might be up to 60 years, may not be easy. We report the case of a 59-year-old man who worked as a taxi driver for 25 years in Venezuela. Cutaneous paracoccidioidomycosis was diagnosed one year after he returned to Spain. No internal signs or symptoms of the disease were found and cell-mediated immunity showed no depression. Our case suggested us to consider direct cutaneous infection, although respiratory tract is believed to be the most frequent portal of entry for this infection.

Humans↗
PubMed2000

Molecular characterization and identification of biocontrol isolates of Trichoderma spp.

The most common biological control agents (BCAs) of the genus Trichoderma have been reported to be strains of Trichoderma virens, T. harzianum, and T. viride. Since Trichoderma BCAs use different mechanisms of biocontrol, it is very important to explore the synergistic effects expressed by different genotypes for their practical use in agriculture. Characterization of 16 biocontrol strains, previously identified as "Trichoderma harzianum" Rifai and one biocontrol strain recognized as T. viride, was carried out using several molecular techniques. A certain degree of polymorphism was detected in hybridizations using a probe of mitochondrial DNA. Sequencing of internal transcribed spacers 1 and 2 (ITS1 and ITS2) revealed three different ITS lengths and four different sequence types. Phylogenetic analysis based on ITS1 sequences, including type strains of different species, clustered the 17 biocontrol strains into four groups: T. harzianum-T. inhamatum complex, T. longibrachiatum, T. asperellum, and T. atroviride-T. koningii complex. ITS2 sequences were also useful for locating the biocontrol strains in T. atroviride within the complex T. atroviride-T. koningii. None of the biocontrol strains studied corresponded to biotypes Th2 or Th4 of T. harzianum, which cause mushroom green mold. Correlation between different genotypes and potential biocontrol activity was studied under dual culturing of 17 BCAs in the presence of the phytopathogenic fungi Phoma betae, Rosellinia necatrix, Botrytis cinerea, and Fusarium oxysporum f. sp. dianthi in three different media.

DNA, Mitochondrial↗
PubMed2000

The tprK gene is heterogeneous among Treponema pallidum strains and has multiple alleles.

We have previously shown that the TprK antigen of T. pallidum, Nichols strain, is predominantly expressed in treponemes obtained 10 days after infection and that the hydrophilic domain of TprK is a target of opsonic antibodies and confers significant protection against homologous challenge. The T. pallidum genome sequence reported the presence of a single copy of the tprK gene in the Nichols strain. In the present study we demonstrate size heterogeneity in the central portions of the TprK hydrophilic domains of 14 treponemal isolates. Sequence analysis of the central domains and the complete open reading frames (ORFs) of the tprK genes confirms this heterogeneity. Further, multiple tprK sequences were found in the Nichols-defined tprK locus in three isolates (Sea 81-4, Bal 7, and Bal 73-1). In contrast, only a single tprK sequence could be identified in this locus in the Nichols strain. Alignment of the DNA and deduced amino acid sequences of the whole tprK ORFs shows the presence of seven discrete variable domains flanked by highly conserved regions. We hypothesize that these heterogeneous regions may be involved in antigenic heterogeneity and, in particular, evasion of the immune response. The presence of different tprK alleles in the tprK locus strongly suggests the existence of genetically different subpopulations within treponemal isolates.

Alleles↗
PubMed2000

Multiple alleles of Treponema pallidum repeat gene D in Treponema pallidum isolates.

Two new tprD alleles have been identified in Treponema pallidum: tprD2 is found in 7 of 12 T. pallidum subsp. pallidum isolates and 7 of 8 non-pallidum isolates, and tprD3 is found in one T. pallidum subsp. pertenue isolate. Antibodies against TprD2 are found in persons with syphilis, demonstrating that tprD2 is expressed during infection.

Alleles↗