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Biomedical subjects

R Rosales

Publications and source records attributed to R Rosales.

At least 37 records · Page 2Linked to original sources
PubMed1995

Cerebral microembolism during carotid endarterectomy.

BACKGROUND: This study was designed to assess the intraoperative risk of cerebral microembolism, as detected by transcranial Doppler ultrasonography, during carotid endarterectomy. PATIENTS AND METHODS: Thirty-six patients (37 procedures) with symptomatic (n = 35) or asymptomatic (n = 2) internal carotid artery origin stenosis(> 50%) were monitored continuously during carotid endarterectomy. Special instrumentation was used to detect high-intensity transient signals (HITS) in the middle cerebral artery on the carotid endarterectomy side. All HITS satisfied a priori established criteria. RESULTS: The incidence of carotid endarterectomies with formed-element HITS increased at clamp release (23/37, P < 0.001) and shunt opening (7/11, P = 0.014), and during wound closure (13/22, P< 0.005) and shunting (5/11, P = 0.046). HITS with air microbubble characteristics were detected at clamp release (22/37, P < 0.001) and shunt opening (5/11, P = 0.025). CONCLUSIONS: HITS do not occur randomly during carotid endarterectomy. Shunting, unclamping, and wound closure are high-risk periods.

Aged↗
PubMed1994

Purification and identification of a vaccinia virus-encoded intermediate stage promoter-specific transcription factor that has homology to eukaryotic transcription factor SII (TFIIS) and an additional role as a viral RNA polymerase subunit.

Enzymes and factors, required for in vitro transcription of templates regulated by vaccinia virus intermediate stage promoters, are present in HeLa cells infected with vaccinia virus in the presence of an inhibitor of DNA replication. Previous studies indicated that in vitro transcription could be reconstituted by adding a partially purified transcription factor to the viral RNA polymerase and capping enzyme. By using an independent purification procedure, we isolated two vaccinia virus intermediate were necessary for transcription of several different intermediate stage promoter templates but not for early or late stage promoter templates. VITF-1 was purified to homogeneity, and the sequences of two tryptic peptides were mapped to the fourth open reading frame within the HindIII E fragment (E4L) of the vaccinia virus genome, which had previously been shown to encode an RNA polymerase subunit of 30 kDa (RPO30) with homology to eukaryotic transcription elongation factor SII. Co-chromatography of VITF-1 with the E4L-derived protein was demonstrated using specific antiserum. In addition, transcriptionally active recombinant VITF-1 was made by expressing the E4L open reading frame in Escherichia coli. Thus, E4L encodes a multifunctional protein, serving as a RNA polymerase subunit and a stage-specific transcription factor. The stepwise binding of capping enzyme, VITF-1, and VITF-2 to a DNA/viral RNA polymerase complex was demonstrated.

Amino Acid Sequence↗
PubMed1994

A cellular factor is required for transcription of vaccinia viral intermediate-stage genes.

The cytoplasmic location of vaccinia virus replication and evidence that the multisubunit DNA-dependent RNA polymerase, early and late stage transcription factors, capping and methylating enzymes, and poly(A) polymerase are virus encoded raised the possibility that all of the proteins needed for viral mRNA synthesis are of viral origin. Previous studies showed that four components from infected cells, the viral RNA polymerase and capping enzyme and two factors called vaccinia virus intermediate transcription factors (VITFs) 1 and 2, can reconstitute transcription of vaccinia virus intermediate-stage genes in vitro. Here, we demonstrate that VITF-2 can be isolated from the nuclei of uninfected HeLa cells as well as from the cytoplasm of infected cells. The proteins with VITF-2 activity from uninfected and infected cells cochromatographed and cosedimented, suggesting that they are identical. VITF-2 activity was found in extracts of other uninfected human and monkey cells but not in nonpermissive Trichoplusia ni insect cells or in conditionally permissive rabbit kidney 13 cells. VITF-2 activity was present, however, in a permissive line of rabbit kidney 13 cells that had been stably transfected with the vaccinia virus K1L host range gene. We suggest that the VITF-2 level acts as a gauge of the permissive state of the cell and thereby regulates the length of the early prereplicative phase of the infection.

Cell Cycle Proteins↗
PubMed1994

Modification of the cascade model for regulation of vaccinia virus gene expression: purification of a prereplicative, late-stage-specific transcription factor.

In vivo and in vitro studies have provided evidence that vaccinia virus late gene transcription factors are intermediate gene products synthesized exclusively after DNA replication. Here, we describe an additional transcription factor (P3 factor) that stimulates late gene transcription between 10- and 40-fold but is made in the absence of viral DNA replication. P3 factor activity was not detected either in uninfected cells or in purified virions. A > 1,500-fold purification of P3 factor was achieved by column chromatography of cytoplasmic extracts prepared from cells infected with vaccinia virus in the presence of a DNA replication inhibitor. P3 factor was stage specific, since it could not substitute for early or intermediate transcription factors. Evidence that late stage-specific transcription factors are made both before and after DNA replication necessitates a modification of the cascade model for vaccinia virus gene regulation.

Gene Expression Regulation, Viral↗
PubMed1994

Stable expression of the vaccinia virus K1L gene in rabbit cells complements the host range defect of a vaccinia virus mutant.

Modified vaccinia virus Ankara (MVA), having acquired genomic deletions during passage in chicken embryo fibroblasts, is highly attenuated and unable to productively infect most mammalian cell lines. Multiplication in rabbit kidney-derived RK13 cells, but not other nonpermissive cells, can be restored by insertion of the vaccinia virus K1L gene into the MVA genome. During nonproductive infection of RK13 cells by MVA, transcription of representative viral early genes was revealed by Northern (RNA) blotting, whereas synthesis of an intermediate mRNA and replication of viral DNA could not be detected. Despite the persistence of viral early mRNA for at least several hours, synthesis of virus-induced polypeptides occurred only during the first hour and was followed by abrupt inhibition of all protein synthesis. Transfection of RK13 cells with a eukaryotic expression plasmid that contained the K1L gene allowed MVA infection to proceed to late stages of viral protein synthesis. Moreover, RK13 cell lines that stably expressed the K1L gene were permissive for MVA as well as a K1E deletion mutant of the WR strain of vaccinia virus. This is the first description of the complementation of a poxvirus mutant by cells that stably express a viral gene.

Animals↗
PubMed1994

[Importance of the deficit of crystallization inhibitors in the etiopathogenesis of urolithiasis].

BACKGROUND: five percent of consultations at the Emergency Room of Catholic University Hospital are due to nephrolithiasis. The causes of this high frequency remain unknown. AIM: to know the main metabolic and anatomic factors involved in the genesis of neprholithiasis. PATIENTS AND METHODS: forty one patients (31 male) were studied presenting with a renal colic were studied as soon as the acute episode subsided and without diet modifications. Fasting blood calcium and creatinine and 24 h urine calcium, uric acid, citrate, magnesium and pH were measured and an intravenous pyelogram was performed. Twenty one subjects without a history of nephrolithiasis were used as controls. RESULTS: Patients with nephrolithiasis did not differ from controls in urinary calcium (159 +/- 67 and 172 +/- 67 mg/24 h respectively), uricosuria (417 +/- 171 and 431 +/- 121 mg/24 h respectively) or urinary magnesium (55 +/- 19 and 62 +/- 21 mg/24 h respectively, whereas urinary citrate was lower (219 +/- 172 vs 319 +/- 179 mg/24 h in controls p < 0.05). All patients had normal renal functions, urinary acidification and intravenous pyelogram. Seven percent of patients with nephrolithiasis had hypercalciuria, 2.4% had hyperuricosuria. 68.3% had a low urinary citrate and 44.4% had low urinary magnesium. CONCLUSIONS: in this sample, there is a strong association of nephrolithiasis with low levels of crystallization inhibitors in special with urinary citrate, a crystallization inhibitor.

Adult↗
PubMed1993

Transcription initiation factor activity of vaccinia virus capping enzyme is independent of mRNA guanylylation.

Cytoplasmic extracts of vaccinia virus-infected HeLa cells blocked in DNA replication were capable of transcribing templates containing the minimal promoter sequences derived from three viral intermediate-stage genes (A1L, A2L, and G8R) but not promoters from early or late genes. One of three isolated components required for transcription copurified with the viral capping enzyme, a heterodimeric protein responsible for forming the 7-methyl-guanosine(5')triphospho(5')nucleoside [m7G(5')ppp(5')N-] structure at the 5' end of mRNAs, as had been reported using a template with another intermediate promoter [Vos, J. C., Sasker, M. & Stunnenberg, H. G. (1991) EMBO J. 10, 2553-2558]. Transcription factor activity was associated with partially, purified capping enzyme from infected cell extracts, homogeneous enzyme from purified virions, and recombinant viral enzyme from Escherichia coli. By transcribing truncated templates of different sizes, we determined that RNA chains of 35 nt were capped whereas those of 15 nt were not. Nevertheless, the capping enzyme was required for formation of short uncapped transcripts, indicating that capping and transcription initiation factor activities are independent functions.

Base Sequence↗
PubMed1990

Identification of an estrogen response element upstream of the human c-fos gene that binds the estrogen receptor and the AP-1 transcription factor.

Transcription of the proto-oncogene c-fos is stimulated by 17 beta-estradiol in estrogen responsive human and rat cells. To understand the molecular mechanisms of estrogen regulation of c-fos gene transcription, the human c-fos gene promoter, with 2.25 Kb of 5'-flanking DNA, was cloned upstream of the bacterial CAT gene and tested for estrogen regulation by transient transfection in HeLa cells. When an expression vector coding for the human estrogen receptor was co-transfected with the fos -CAT reporter, the promoter was found to respond to 17 beta-estradiol. An element responsible for estrogen induction was mapped in a 240 bp region localized 1060 to 1300 bases upstream of the startsite of transcription of the gene. Sequence analysis revealed, clustered in a 19 bp sub-region, a sequence corresponding to an imperfectly palindromic ERE: CGGCAGCGTGACC and two sequences: CTGAG and GTGAC, homologous to the core sequence of AP-1 transcription factor binding sites. A synthetic oligonucleotide reproducing this sub-region binds 'in vitro' both the estrogen receptor and AP-1 factor(s) and confers estrogen-responsivity to the HSV-tk gene promoter. Transcriptional activation by the estrogen receptor is prevented by mutations in the fos ERE that hamper binding of the receptor in vitro. Activation of the c-fos gene promoter in HeLa cells requires the DNA binding domain of the estrogen receptor, and can be achieved independently by the TAF-1 and the TAF-2 transcriptional activation functions of this molecule. A receptor mutant lacking the hormone binding domain can activate the c-fos gene promoter in the absence of estrogen.

Base Sequence↗
PubMed1989

The SV40 TC-II(kappa B) enhanson binds ubiquitous and cell type specifically inducible nuclear proteins from lymphoid and non-lymphoid cell lines.

We have characterized the complexes resulting from the specific binding in vitro of proteins present in nuclear extracts of several lymphoid and non-lymphoid cell lines to the TC-I and TC-II sequences of the simian virus 40 (SV40) enhancer. No proteins could be detected, binding selectively to the TC-I sequence, but two proteins TC-IIA and TC-IIB were identified interacting specifically with both the TC-II/kappa B enhanson, 5'-GGAAAGTCCCC-3' (important for the activity of the SV40 enhancer in vivo), and with the related H-2Kb enhanson, 5'-TGGGGATTCCCCA-3'. The binding of these two proteins to mutated TC-II enhansons correlates with the effect of these mutations in vivo, suggesting that both proteins may be important for SV40 enhancer activity. The TC-IIA binding activity was present in nuclear extracts of mature lymphoid B cells and was increased in pre-B cell nuclear extracts by lipopolysaccharide (LPS) and cycloheximide treatment. Furthermore, complex formation between the TC-IIA protein and the TC-II enhanson was efficiently competed by the kappa B motif from the kappa chain enhancer, indicating that TC-IIA is the NF-kappa B factor or a closely related protein. However, in contrast to previous reports, a TC-IIA/NF-kappa B-like protein whose properties could not be distinguished from those of the TC-IIA protein present in lymphoid B cells, was found in nuclear extracts of several untreated non-lymphoid cell lines, notably of HeLa cells, but not of undifferentiated F9 embryonal carcinoma (EC) cells [F9(ND)]. The TC-IIA binding activity which was moderately increased in HeLa cell nuclear extracts by 12-O-tetradecanoylphorbol-13-acetate (TPA) and/or cycloheximide treatment could be induced in nuclear extracts of F9(ND) cells by cycloheximide, but not by TPA. Moreover, the TC-IIA binding activity could be induced in cytosolic fractions from F9(ND) cells by treatment with deoxycholate, indicating that these cells contain an inhibitor protein similar to the previously described NF-kappa B inhibitor, I kappa B. The second TC-II enhanson binding protein, TC-IIB, which could be clearly distinguished from the TC-IIA/NF-kappa B-like protein, by a number of differential properties, resembles the previously described KBF1/H2TF1 protein as it binds with a higher affinity to the H-2Kb enhanson than to the TC-II/kappa B enhanson, and its pattern of methylation interference on the H-2Kb and TC-II/kappa B enhansons is identical to that reported for the KBF1/H2TF1 protein.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

The HeLa cell protein TEF-1 binds specifically and cooperatively to two SV40 enhancer motifs of unrelated sequence.

We have purified a protein (TEF-1) that specifically binds to two sequence unrelated motifs (GT-IIC and Sph) of the simian virus 40 (SV40) enhancer. TEF-1 binds cooperatively to templates containing tandem but not inverted or spaced repeats of its cognate motifs. This cooperative binding correlates with the ability of the tandem repeats to generate enhancer activity in vivo. In contrast, TEF-1 and a second SV40 enhancer binding protein, TEF-2, bind independently to templates containing the cognate motifs of both proteins (GT-I and either GT-IIC or Sph motifs) even though these motifs cooperate in enhancer activity in vivo. These results allow us to distinguish different classes of enhancer factors.

Allosteric Regulation↗
PubMed1987

Clinical electrophysiologic studies of HTLV-I-associated myelopathy.

Six patients with human T-cell lymphotropic virus type I (HTLV-I)-associated myelopathy (HAM) were studied by electrophysiologic methods. Upper-limb short-latency somatosensory evoked potentials showed slight delay of the N9-N20 interpeak latency in one of 12 limbs, while the lower-limb short-latency somatosensory evoked potentials showed prolonged N20-P40 interpeak latency in eight of 12 limbs. Frequent polyphasic potentials and occasional giant spikes were observed in the distal extremities. F-wave conduction velocity was delayed in some patients. Results of the other nerve conduction studies were unremarkable. Our data provide a valuable extension of the clinical examination of HAM and offer encouragement for a more extensive electrophysiologic study of this entity, especially in the spinal cord.

Action Potentials↗
PubMed1987

One cell-specific and three ubiquitous nuclear proteins bind in vitro to overlapping motifs in the domain B1 of the SV40 enhancer.

We have used the gel retardation assay to investigate the binding of nuclear proteins to the domain B1 of the SV40 enhancer, which contains the GT-II motif. Four proteins (GT-IIA, GT-IIB alpha, GT-IIB beta and GT-IIC) were detected, three of which were present in nuclear extracts from several cell lines. The fourth protein (GT-IIC) showed a clear cell-specificity, being absent from the lymphoid cell extracts tested. The results of methylation interference assays and of the binding of the proteins to mutated templates indicate that the domain B1 contains three distinct, but overlapping, protein-binding motifs (GT-IIA, B and C). The cell-specific binding of protein GT-IIC in vitro correlates with the in vivo enhancer activity of its cognate motif, strongly suggesting that this protein acts as a positive trans-acting enhancer factor. Two of the proteins also recognize other enhancer motifs; protein GT-IIB alpha binds to the microE3 motif present in the immunoglobulin heavy chain enhancer; protein GT-IIC binds to an enhancer motif of the polyomavirus mutant PyEC9.1 adapted to growth in F9 embryonal carcinoma cells, but not to the corresponding wild-type sequence.

Base Sequence↗
PubMed1987

In vitro binding of cell-specific and ubiquitous nuclear proteins to the octamer motif of the SV40 enhancer and related motifs present in other promoters and enhancers.

We have used the gel retardation and DNase I footprinting assays to investigate the in vitro binding of nuclear proteins to the octamer motif present in domain A of the SV40 enhancer and in other enhancer and promoter elements. Three apparently cell-specific (oct-B1A, oct-B1B and oct-B2) and one ubiquitous (oct-B3) proteins were detected in various lymphoid and non-lymphoid cell extracts. We show that the previously described 'ubiquitous' NF-A1 factor may correspond in fact to two proteins, oct-B1A in HeLa cells and oct-B1B in lymphoid cells. Interestingly, the HeLa cell protein oct-B1A formed a complex with the SV40 octamer, which could be detected in gel retardation, but not in DNase I footprinting assays. This absence of protection from DNase I digestion correlates with the inactivity of the SV40 octamer in HeLa cells in vivo. We have also found that the in vitro interaction between the SV40 octamer motif and the lymphoid cell-specific protein oct-B2 was negatively modulated by a component present in the nuclear extracts from several lymphoid cell lines. The interactions between the multiple octamer-binding proteins and the related octamer motifs present in other promoter and enhancer elements were systematically compared and the possible role of these proteins in the control of transcription is discussed.

Base Sequence↗
PubMed1987

In vitro binding of several cell-specific and ubiquitous nuclear proteins to the GT-I motif of the SV40 enhancer.

We have investigated the specific in vitro binding of nuclear proteins from several cell lines to the GT-I motif of the SV40 enhancer which overlaps with the canonical enhancer "core" homology. The binding of three proteins (GT-IA, GT-IB, and GT-IC), one of which (GT-IC) exhibits cell specificity, was detected. Competition and direct binding experiments demonstrated that the two ubiquitous proteins also bind to the GC-rich motif III from the 21-bp repeat upstream element of the SV40 early promoter and that protein GT-IA is most probably the transcription factor Sp1. The third, cell-specific protein GT-IC exhibited a high affinity for both the GT-I motif and an upstream element in the promoter of the mouse beta-major-globin gene, suggesting that this protein can act both as an enhancer and an upstream element trans-acting factor. The good correlation between the known cell-specific in vivo activity of the wild-type and mutated GT-I motif and the cell-specific binding of protein GT-IC in vitro strongly supports the conclusion that this protein is an enhancer factor. Interestingly, its cognate recognition sequence does not coincide with the core homology.

Animals↗
PubMed1984

Cholecystoduodenocolic fistula with recurrent gallstone ileus.

The combination of cholecystoduodenocolic fistula with gallstone ileus is rarely seen. To our knowledge, there have only been six previous reports with these findings. A 66-year-old woman's condition was diagnosed preoperatively as small-bowel obstruction and communications between the gallbladder, duodenum, and colon. The small-bowel obstruction was successfully relieved by removing a large gallstone from the midileum, leaving the inflammatory mass in the right upper quadrant undisturbed. Three weeks later the small-bowel obstruction recurred. At reoperation two gallstones were found obstructing the midileum and were removed. The cholecystoduodenocolic fistula was dissected and the duodenum and colon were repaired. A cholecystostomy was done. The patient recovered and has been well four years later.

Aged↗
PubMedDate not supplied

Incidence of Trypanosoma cruzi infection in two Guatemalan communities.

The prevalence of human infection by Trypanosoma cruzi was assessed using an enzyme-linked immunosorbent assay (ELISA) in a serological survey in 1998 of 2 rural communities (SMH and PS) in Guatemala. In SMH (Department of Zacapa), where Rhodnius prolixus was the principal vector, the seroprevalence amongst 373 people tested was 38.8%. In PS (Department of Santa Rosa), where the main vector was Triatoma dimidiata, 8.9% of the 428 people tested were seropositive. The overall prevalence of seropositivity was higher in females than in males in both SMH (40% vs 36%) and PS (11.9% vs 4.9%), although this difference was significant only in PS. Historical seroconversion rates, estimated retrospectively by fitting a transmission model to the age-prevalence curves, were 3.8% per year in SMH and 0.5% per year in PS. There was some indication of a recent reduction in incidence in both villages. In PS, but not in SMH, both the observed prevalence and the estimated incidence rates were significantly higher in females than in males.

Adolescent↗