Biomedical subjects
Publications and source records attributed to S Stein.
Up to now around 170 different homeobox genes have been cloned from vertebrate genomes. A compilation of the various isolates from mouse, chick, frog, fish and man is presented in the form of a concise checklist, including the designations from the original publications. Putative homologs from different species are aligned, and key characteristics of embryonic or adult expression domains, as well as mutant phenotypes are briefly indicated.
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An increase in melting temperature for DNA:DNA duplexes had been observed previously (Zhu et al. Antisense Res. Dev. 3:349-356, 1993) when an oligo(delta)ornithine moiety was covalently appended to a short oligodeoxynucleotide. We now report the analysis of duplex formation by electrophoretic gel shift analysis. In the particular example studied, an increase in Tm of 4 degrees C was found to correspond to about a fivefold increase in binding constant. A similar enhancement by the appended cationic peptide was observed when the target strand was RNA. The use of a competitive assay format for avoidance of adsorptive loss at low concentrations (< 10(-7)M) of the oligonucleotide-oligo(delta)ornithine conjugate is presented.
PURPOSE: The presenting signs, treatment, and outcome of an epidural hematoma of the cauda equina in a child with severe hemophilia are reported for the first time. PATIENTS AND METHODS: A 20-month-old boy with severe hemophilia A (factor VIII <0.01 U/ml) presented with a 12-day history of refusal to stand and constipation of 5-7 days duration. He had normal deep tendon reflexes with normal sensation and withdrawal to pinprick of his lower extremities bilaterally. He stood on his right leg, but had inversion of his left foot and refused to bear weight on his left leg. MRI revealed an epidural hematoma of the cauda equina and a distended bladder. Factor VIII replacement therapy and lumbosacral laminectomy with evacuation of the hematoma resulted in recovery of a normal gait, but bladder dysfunction persisted for 11 months. Clean intermittent catheterization (CIC) was required until bladder function returned. RESULTS: Complete neurologic recovery occurred 11 months after presentation CONCLUSION: This case demonstrates the following points: (a) an epidural hematoma of the cauda equina in a child with severe hemophilia can present with neurologic findings that are as subtle as those seen in normal children; (b) CIC can be performed safely over an extended period without factor VIII replacement; and (c) complete recovery is possible, despite prolonged bladder dysfunction and a 12-day interval between the onset of symptoms and treatment.
The IIIa and IIIb electrophoretic variants of arylsulfatase A (EC 3.1.6.8) are 12 times more prevalent in alcoholic than in nonalcoholic populations. These variant enzymes, found in a subset of alcoholics, possess the pseudodeficient Asn350-Ser mutation of arylsulfatase A and, consequently, lack an N-linked glycan unit. These genetically determined variants of arylsulfatase A show reduced intracellular half-life, and cells from such individuals possess reduced enzymic activity. We propose that this polymorphism is an underlying genetic and biochemical factor contributing to the neuropathology and/or addiction pathway of this disease.
Several electrophoretic forms of human platelet arylsulfatase A (ASA), including variant type IIIa and normal type IV(a), have been identified by nondenaturing polyacrylamide gel electrophoresis. An alcoholic population that we have analyzed is enriched in variant type IIIa compared with nonalcoholic psychiatric and normal controls. Individuals with the IIIa enzyme possess greatly reduced levels of ASA activity. To understand further the structural basis for the differences and their potential biological consequences, the nature of the ASA variant expressed by fibroblasts from different individuals was explored. The electrophoretic patterns of fibroblast ASA from the IIIa and IV(a) individuals differ in degree of phosphorylation. Furthermore, fibroblast ASA from IIIa individuals lacks an N-linked glycan found in ASA from IV(a) individuals. In addition, differences in peptide and/or posttranslational modification unrelated to the N-linked carbohydrate or phosphorylation exist between the fibroblast ASA from IIIa and IV(a) individuals. The finding that both fibroblasts and platelets exhibit related electrophoretic isoform patterns characteristic of the donor's ASA type allows for the use of fibroblasts to study the impact of ethanol on the metabolism of cells possessing different ASA types.
Interference of binding of Tat protein to TAR RNA in HIV-1-infected cells may be a useful therapeutic strategy for AIDS. An electrophoretic assay to screen potential low-molecular-weight (< 2 kDa) Tat antagonists has been established. A radiolabeled TAR RNA fragment (delta TAR) is retarded in mobility when bound by a Tat peptide-polyethylene glycol conjugate (Tat-PEG), which is used in place of the Tat protein. The assay determines the ability of a potential antagonist to compete with Tat-PEG for binding to delta TAR, as measured by interference with the gel shift of delta TAR. To discriminate between specific and nonspecific interactions, the assay is done in the absence or the presence of a 250-fold molar excess of tRNA.
Patients with the acquired immunodeficiency syndrome (AIDS) are characteristically hypouricemic. Therefore, the occurrence of gouty arthritis in association with AIDS would be expected to be a rare phenomenon. We describe a patient with AIDS in whom gouty arthritis developed. Features of both diseases in relation to their coexistence in this patient are discussed.
We have isolated a chicken cDNA clone, Cnot, resembling in sequence and expression pattern the Xenopus homeobox gene Xnot. The major, early transcription domains of Cnot are the node, the notochord and prenodal and postnodal neural plate caudal from the prospective hindbrain level. All these cell populations appear to be descendants of the Cnot-expressing cells of the node, suggesting a cell lineage relationship. After the onset of somitogenesis, a second, independent expression domain appears in the neural folds at the prospective mid- and forebrain levels, and further transcripts are found in the epiphysis, the ventral diencephalon, the preoral gut and the limb buds. Transplantation of nodes from extended streak embryos leads to the formation of ectopic notochords, which express Cnot in the typical, cranially decreasing gradient. Transplantation of young nodes to young hosts has previously been described to induce secondary embryos. We observed that secondary chick embryos express Cnot in node derived, notochord-like structures and in the anterior neural plate, similar to the domains seen in primary embryos. However, expression was absent from the posterior neural plate, which in the induction experiments is excluded from the node lineage. This finding corroborates our initial conclusion about a cell lineage relationship between node, notochord, and neural plate defined by Cnot expression. The midline mesoderm of vertebrate embryos consists of two tissues, the prechordal mesoderm and the notochord. The anterior notochord, the head process, may represent an intermediate form. The transition from prechordal to chordal mesoderm can be followed by the expression of the two marker homeobox genes goosecoid and Cnot, first in the primitive streak, and then in the head process. We suggest that expression of goosecoid or Cnot is involved in the specification of a prechordal or notochordal identity, respectively. A transition from goosecoid to Cnot expression may proceed, while cells are still in the epiblast, but not after becoming mesodermal. A molecular coding of axial positions in the midline mesoderm may occur by specific homeobox genes, similar to the situation in the neural tube and the somitic mesoderm.
It is well known that Down syndrome (DS) affects multiple organ systems, including cardiovascular, gastrointestinal, hematological, and musculoskeletal. Involvement of the lungs by a characteristic form of subpleural cystic disease is well described in the pathology literature. The imaging characteristics of changes caused by this disease have not been previously described in detail in the radiology literature, possibly because they are not well appreciated on plain chest radiographs. We present a child with DS in whom extensive cystic lung disease was discovered incidentally at CT, and review the current literature regarding the pathogenesis of cystic pulmonary abnormalities in children with DS.
The authors report the first excision of a herniated lumbar disk using the combination of laparoscopic exposure and contact laser application. A 27-year-old patient experienced immediate complete relief of back and leg pain. Laser-assisted laparoscopic lumbar diskectomy offers unique advantages.
The concept of antigene therapy of disease is based on the ability of an oligonucleotide (the therapeutic agent) to bind to double-stranded genomic DNA (the target associated with the disease). Examples are herein given of the linkage of a series of polyamines to a 21-mer homopyrimidine oligonucleotide. These conjugated 21-mers can each form a triple helix with an appropriate double-stranded homopurine-homopyrimidine DNA according to Hoogsteen base-pairing rules. No triple helix was found when unmodified third strand was used at 10 mM sodium phosphate, pH 6.5, 100 mM sodium chloride solution. In contrast, the spermine-conjugated oligonucleotide had a melting temperature of 42 degrees C. According to the melting profile, the appended spermine moiety was found to affect the Tm only of the triple helix, but not of the subsequent melting of the underlying double helix. The Tm enhancing ability of the spermine-conjugate was found to be better than that of other polyamine-conjugates.
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A monoclonal antibody (H9724), specific for the 41-kDa flagellar protein of the Lyme disease pathogen Borrelia burgdorferi, cross-reacts with human axons and detects one major protein in human neuroblastoma cell extracts. The homologous cross-reacting protein has now been isolated from calf adrenal and identified as chaperonin-HSP60 by N-terminal sequencing.
Noncovalent binding of proteins to membranes is often employed for dot-blot analysis with various visualization techniques. These techniques are usually not applicable to peptide dot-blot analysis due to peptide wash-off during the staining procedure. As exemplified with a synthetic peptide and peptides produced by proteolysis of a protein, it is possible to achieve efficient covalent attachment to Immobilon-AV membranes. The utility of this membrane has been demonstrated with immunostaining and carbohydrate staining procedures.
We show that automated external defibrillation training of emergency medical technicians (EMTs) is less time consuming than manual defibrillation training, and hypothesize that both improve survival from sudden cardiac death. Data on 91 cardiac arrests over 27 months among five basic life support services was collected before EMT-defibrillation (EMT-D) training. Subsequently, seven BLS services were trained in EMT-D using either manual difibrillation or automated external defibrillation technology, and 55 sudden cardiac death patients were entered after training. Manual defibrillation required 11 more hours per student in initial training. Survival to hospital discharge improved from two of 91 patients (2.2%) in the series before EMT-D training to nine of 55 patients (16.4%) after EMT-D training (P = .001). Improved survival was correlated with shorter prehospital defibrillation times, 8.84 minutes, when EMTs performed defibrillation versus 16.3 minutes before training when EMTs awaited advanced life support defibrillation (P < .001). To enhance equipment familiarity we allowed EMTs to apply three-lead electrode monitors to all medical/cardiac patients during transport (surveillance). There were six emergency medical service-witnessed "surveillance" arrests and three arrests survived to hospital discharge (50% survival). This group represented 33% of all survivors in the series. We recommend automated external defibrillation training for EMTs. Improved survival in sudden cardiac death cases in well-run emergency medical service systems should result from EMT-D training. Finally, we recommend that routine "surveillance" of high-risk patients during transport by defibrillation-capable EMTs be considered in EMT-D programs, rather than limiting EMT-D only to units capable of rapid "man-down" response.
On the basis of the reported enhanced antisense activity of polylysine-oligonucleotide conjugates, a synthetic 12-mer oligodeoxyribonucleotide has been coupled at its 5' terminus to a series of positively charged (delta-ornithine)n cysteine peptides. Binding between the nucleic acid-peptide conjugate and its complementary DNA target sequence was detected by the impact of complexation on the melting temperature (Tm). It was found that the Tm for the nucleic acid-peptide gradually increased with increasing net charge on the conjugated peptide. Site-directed cleavage with RNase H demonstrates that the peptide-modified oligomer also hybridizes with its RNA target sequence. Increased affinity for target mRNA with net charge was shown by a cell-free translation arrest assay.
Oligodeoxynucleotides (12-mers) having either a 3' aminolinker or both 3' and 5' aminolinker groups were synthesized and then reacted with N-iodoacetoxysuccinimide. Separately, a series of peptides, consisting of cysteine carboxyamide and a varying number of residues of ornithine coupled through their side-chain amino groups, was prepared, leaving on the final Fmoc protecting group. Reaction of each Fmoc-peptide with an activated oligodeoxynucleotide yielded the desired conjugates, which were purified by an Fmoc-on/Fmoc-off two-step reversed-phase HPLC procedure. On hybridization with a complementary unmodified 12-mer oligodeoxynucleotide, it was found that there is a gradual increase in melting temperature with the number of ornithine residues, whereas the appended peptide did not perturb the B form of the duplex.