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Biomedical subjects

L M Hoffman

Publications and source records attributed to L M Hoffman.

At least 19 recordsLinked to original sources
PubMed1999

Alcohol promotes in vitro chondrogenesis in embryonic facial mesenchyme.

Ethanol is a well-recognized teratogen in vertebrates that can perturb the development of the facial primordia and various other embryonic structures. However,the mechanisms underlying alcohol's effects on embryogenesis are currently unclear. Recent evidence suggests that the cranial neural crest, which forms the entire facial skeleton, may be a particularly sensitive target of ethanol teratogenicity. In the present study we have examined the influence of in vitro ethanol exposure on cartilage differentiation in micromass cultures of mesenchymal cells isolated from the various facial primordia (maxillary, mandibular, frontonasal, and hyoid processes) of the stage 24 chick embryo. In all four populations of facial mesenchyme, exposure to 1-1.5% ethanol promoted marked increases in Alcian blue-positive cartilage matrix formation, a rise in 35SO4 accumulation into matrix glycosaminoglycans, and enhanced expression of cartilage-characteristic type II collagen and aggrecan gene transcripts. In frontonasal and mandibular mesenchyme cultures, which undergo extensive spontaneous cartilage formation, ethanol treatment quantitatively elevated both matrix production and cartilage-specific gene transcript expression. In cultures of maxillary process and hyoid arch mesenchyme, which form little or no cartilage spontaneously, ethanol exposure induced the formation of chondrogenic cell aggregates and the appearance of aggrecan and type II collagen mRNAs. These actions were not restricted to ethanol, since tertiary butanol treatment also enhanced cartilage differentiation in facial mesenchyme cultures. Our findings demonstrate a potent stimulatory effect of alcohol on the differentiation of prechondrogenic mesenchyme of the facial primordia. Further analysis of this phenomenon might yield insight into the developmental mechanisms underlying the facial dysmorphologies associated with embryonic ethanol exposure.

Aggrecans↗
PubMed1997

Cell-mediated immune response and stability of intraocular transgene expression after adenovirus-mediated delivery.

PURPOSE: To evaluate the role of cell-mediated immunity in the stability of ocular adenovirus-mediated transgene expression. METHODS: Adenovirus (4 x 10(6) pfu) containing lacZ (Ad.CMVlacZ) was injected intravitreally or subretinally into one or both eyes of immunocompetent (+/+) and immunocompromised (nu/nu) CD-1 mice. Control eyes received injections of saline. Additional +/+ mice received subretinal injections of Ad.CMVlacZ with coadministration of 200 microg of human immunoglobulin (Ig) G or CTLA4Ig by intraperitoneal, intravitreal, or subretinal injection. The mice were killed at various times after injection, and their eyes were examined histologically and immunohistochemically. RESULTS: LacZ expression was extended from 1 week to more than 5 weeks in the corneal endothelium, iris, and trabecular meshwork of nu/nu mice compared with time of expression in +/+ mice when adenovirus was administered intravitreally. In contrast, subretinal injection resulted in only a minimal increase in transgene stability in nu/nu mice compared with that in +/+ mice. Neither systemic nor intraocular administration of IgG or CTLA4Ig affected the stability of lacZ expression in the retina or retinal pigment epithelium after subretinal injection in +/+ mice. Immunoglobulin G and CTLA4Ig enhanced the stability of transgene expression in the trabecular meshwork. CONCLUSIONS: A T-cell-mediated immune response appears to play a role in the loss of adenovirus-mediated lacZ expression after intravitreal but not after subretinal injection. This result implies that the subretinal space is an immune-privileged site and a favorable site for gene transfer.

Abatacept↗
PubMed1996

Ethanol exposure stimulates cartilage differentiation by embryonic limb mesenchyme cells.

Studies of neural, hepatic, and other cells have demonstrated that in vitro ethanol exposure can influence a variety of membrane-associated signaling mechanisms. These include processes such as receptor-kinase phosphorylation, adenylate cyclase and protein kinase C activation, and prostaglandin production that have been implicated as critical regulators of chondrocyte differentiation during embryonic limb development. The potential for ethanol to affect signaling mechanisms controlling chondrogenesis in the developing limb, together with its known ability to promote congenital skeletal deformities in vivo, prompted us to examine whether chronic alcohol exposure could influence cartilage differentiation in cultures of prechondrogenic mesenchyme cells isolated from limb buds of stage 23-25 chick embryos. We have made the novel and surprising finding that ethanol is a potent stimulant of in vitro chondrogenesis at both pre- and posttranslational levels. In high-density cultures of embryonic limb mesenchyme cells, which spontaneously undergo extensive cartilage differentiation, the presence of ethanol in the culture medium promoted increased Alcian-blue-positive cartilage matrix production, a quantitative rise in 35SO4 incorporation into matrix glycosaminoglycans (GAG), and the precocious accumulation of mRNAs for cartilage-characteristic type II collagen and aggrecan (cartilage proteoglycan). Stimulation of matrix GAG accumulation was maximal at a concentration of 2% ethanol (v/v), although a significant increase was elicited by as little as 0.5% ethanol (approximately 85 mM). The alcohol appears to directly influence differentiation of the chondrogenic progenitor cells of the limb, since ethanol elevated cartilage formation even in cultures prepared from distal subridge mesenchyme of stage 24/25 chick embryo wing buds, which is free of myogenic precursor cells. When limb mesenchyme cells were cultured at low density, which suppresses spontaneous chondrogenesis, ethanol exposure induced the expression of high levels of type II collagen and aggrecan mRNAs and promoted abundant cartilage matrix formation. These stimulatory effects were not specific to ethanol, since methanol, propanol, and tertiary butanol treatments also enhanced cartilage differentiation in embryonic limb mesenchyme cultures. Further investigations of the stimulatory effects of ethanol on in vitro chondrogenesis may provide insights into the mechanisms regulating chondrocyte differentiation during embryogenesis and the molecular basis of alcohol's teratogenic effects on skeletal morphogenesis.

Aggrecans↗
PubMed1996

The reovirus nonstructural protein sigma1NS is recognized by murine cytotoxic T lymphocytes.

The cytotoxic T-lymphocyte (CTL) response in reovirus-infected C3H mice was investigated by using reovirus-vaccinia virus recombinants. Results of cytotoxicity assays indicated that the nonstructural protein sigma1NS elicited a significant CTL response. Experiments with sigma1NS-specific CTL lines showed that both strain-specific and cross-reactive epitopes exist in the sigma1NS protein.

Animals↗
PubMed1994

Cyclic AMP causes differentiation and decreases the expression of neutral glycosphingolipids in cell cultures derived from a malignant glioma.

Cultures derived from a malignant glioma (U-87 MG) were treated with 3 mM dibutyryl cAMP. The treatment resulted in morphological differentiation of the cultures and a decrease in cell proliferation. Biochemically, dibutyryl cAMP treatment caused a general reduction in the concentration of neutral glycosphingolipids in the U-87 MG cells. The concentration of individual neutral glycosphingolipids in the untreated cells was 1.8- to 3.0-fold higher than in cells treated for 72 h with 3 mM dibutyryl cAMP. Cells were labeled with [3H]galactose to monitor synthesis of the neutral glycosphingolipids. Decreased synthesis was noted in cells treated with dibutyryl cAMP as compared with untreated cells as indicated by decreased uptake of [3H]galactose label. The ganglioside composition of the cells was essentially unchanged after dibutyryl cAMP treatment.

Bucladesine↗
PubMed1994

Enzymatic synthesis of milligram quantities of ribozymes in small volumes.

Milligram quantities of ribozymes (Rzs) can be synthesized in vitro in reaction volumes of 1 mL or less using AmpliScribe T7 RNA polymerase kits to transcribe either linear plasmids or oligodeoxynucleotide templates. Model hammerhead Rzs were synthesized that specifically cleave RNA-encoding chloramphenicol acetyl transferase sequences. Methods are presented for the transcription of Rzs of virtually any length and sequence composition at a fraction of the cost of chemical synthesis.

Base Sequence↗
PubMed1993

Migraine.

Explore the source record for details and available documents.

Acetaminophen↗
PubMed1991

SV-40 transformation: effect on GM2 ganglioside in cultured cell lines.

In previous work, we observed the presence of substantially elevated levels of GM2 after Simian Virus 40 (SV-40) transformation of human fetal brain cells. This elevated level of GM2 contrasted with the reports of many other investigators who had often observed decreased levels of GM2 and a simplification of ganglioside pattern in various non-neural rodent cell lines. In order to determine if the increase in GM2 in the transformed human brain cells would also be found in transformed rodent brain cells, we analyzed ganglioside changes after transformation in mouse brain cell lines and observed the increase in GM3 and low levels or lack of GM2 usually noted in rodent SV-40 transformed cell lines. In addition, we analyzed changes after SV-40 transformation in three human fibroblast lines and found that all three lines contained substantially elevated levels of GM2 after SV-40 transformation. As a result of this study, our earlier work on SV-40 transformed human brain cells, and occasional other reports of high levels of GM2 in human SV-40 transformed cell lines, elevated levels of GM2 may be considered a marker for SV-40 transformed human cells of both fibroblastic and neural origin.

Animals↗
PubMed1989

Gangliosides in SV-40-transformed cells derived from Tay-Sachs disease fetal brain.

A human glial brain cell line derived from a Tay-Sachs disease fetal cerebellum was transformed with SV-40 virus in order to obtain a transformed brain cell line which reflected the characteristics of the disease. It was shown that the transformed TSD cell line maintained an elevated level of GM2 which was similar to that shown by the nontransformed precursor. In addition, the TSD transformed line lacked hexosaminidase A.

Cerebellum↗
PubMed1988

Vectors for in vitro synthesis of poly(A)+RNA transcripts.

An RNA expression plasmid containing a maize zein storage protein gene was modified by the addition of an oligodeoxynucleotide duplex containing oligo(A30) in the RNA sense strand to the 3' flank of the gene. By linearizing the plasmid at either side of the oligo(A) region, transcripts with or without a poly(A) tail may be synthesized. Poly(A) addition dramatically increases the translation of SP6-transcribed zein transcripts in Xenopus laevis oocytes. We describe a derivative of pSP64, pSP64A, containing the oligo(A) region, which may be useful for the transcription of polyadenylated RNAs from any template DNA.

DNA Restriction Enzymes↗
PubMed1988

Induction of lymphomas and fibrosarcomas in nude mice after implantation of simian virus 40-transformed human meningioma.

Human cells transformed by Simian virus 40 (SV40) usually show little or no tumor formation after implantation in nude mice. In long-term studies, however, we have observed that 127 to 366 days after subcutaneous injection of SV40-transformed human meningioma cells (KJ-M2-T) into nude mice (Nu/Cox), 6 of 15 animals developed lymphomas or fibrosarcomas, usually at the site of inoculation. The induced tumors were of murine origin and were positive for SV40-T antigen. Chromosome analysis and G11 staining revealed no evidence of hybridization between human and mouse cells. No spontaneous shedding of SV40 was noted with KJ-M2-T cells in vitro; however, SV40 could be rescued after fusion of KJ-M2-T with BS-C-1 monkey kidney cells, but not with L929 mouse fibroblasts. A parallel study using SV40-transformed human fetal brain cells failed to induce tumors in nude mice despite the demonstration that infectious SV40 could also be rescued from this line after fusion with BS-C-1 but not with L-929. Subcutaneous injection of 5 X 10(3) TCID's of SV40 (strain J436) into nude mice resulted in the induction of fibrosarcomas at the injection site in 6 of 15 mice after 273 to 396 days. The induction of malignant lymphomas after implantation of SV40 transformed cells contrasted with the development of fibrosarcomas after injection of free virus. This study suggests that after subcutaneous implantation into nude mice, some SV40-transformed human tumor cell lines can serve as vectors for transmitting SV40 to murine cells causing transformation and tumor development in the host animal.

Animals↗
PubMed1985

Nucleotide sequences from phaseolin cDNA clones: the major storage proteins from Phaseolus vulgaris are encoded by two unique gene families.

The nucleotide sequences of eight partial and five full-length phaseolin cDNA clones show that phaseolin polypeptides are encoded by two distinct gene families which differ in their coding regions by the presence or absence of two different size direct repeats. The alpha-type phaseolin polypeptides are encoded by genes containing direct repeats which encode 14 additional amino acids. Aside from these differences, the alpha-and beta-type phaseolin genes show a high degree of homology (98%) which is consistent with these genes being derived from a common ancestral gene. Much of the heterogeneity found in the phaseolin polypeptides appears to be due to post-translational processing. Nucleotide sequence analysis demonstrates that the alpha-type genes contain only a few amino acid replacement substitutions and that the beta-type genes appear to contain no amino acid replacement substitutions. S1 nuclease mapping shows a complex pattern for transcriptional initiation of phaseolin mRNA. Hydropathy analysis shows that phaseolin polypeptides are predominately hydrophilic, and that the two N-glycosyl recognition sites are located in different hydropathic environments.

Amino Acid Sequence↗
PubMed1985

Characterization of two Phaseolus vulgaris phytohemagglutinin genes closely linked on the chromosome.

A lambda 1059 library of Phaseolus vulgaris cv. 'Tendergreen' DNA was screened with a cloned lectin-like cDNA. Among the phages selected was clone lambda B10 containing two complete lectin genes in the same orientation approximately 4 kb apart. The DNA sequences of the lectin genes and their flanking regions have been determined and their transcriptional initiation sites were located by S1 nuclease mapping. On the basis of the deduced amino acid sequences and compositions and the mol. wts. of their encoded glycoproteins, the genes, dlec1 and dlec2, are predicted to encode erythro- and leucoagglutinating phytohemagglutinins (PHA-E and PHA-L), respectively. The mRNA coding regions of dlec1 and dlec2 are 90% homologous, suggesting an origin involving duplication of an ancestral gene. Both lectin genes are intronless and have at least two ATG codons in a short (11-14 bp) 5'-untranslated region. Most of their 5'-untranslated regions consist of alternating pyrimidines and purines (RY repeats). Upstream sequences are also highly conserved between dlec1 and dlec2, including stretches of nine or more alternating R and Y residues. RY repeats of such length are not found within the protein coding portion of dlec1, dlec2 or a Phaseolus lectinlike gene previously described. Overlapping double (dlec1) or triple (dlec2) polyadenylation addition signals are found and there is an unusually high degree of homology (84%) between their 3'-untranslated regions.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1984

Structure of a chromosomal Phaseolus vulgaris lectin gene and its transcript.

A genomic library of Phaseolus vulgaris DNA was screened using a lectin cDNA clone that includes the entire coding region to obtain a lambda clone containing the corresponding nuclear lectin gene. Comparison of the cDNA and genomic nucleotide sequences showed that the gene has no intervening sequences. The 5' untranslated region was found to be only 10-16 base pairs long by S1 nuclease mapping. The lectin transcriptional unit is flanked by sequences which are exceptionally high (72-75%) in A + T content and its 5' flanking region contains potential transcription control elements.

Amino Acid Sequence↗
PubMed1982

Molecular cloning of Phaseolus vulgaris lectin mRNA and use of cDNA as a probe to estimate lectin transcript levels in various tissues.

The complete amino acid sequence of a Phaseolus vulgaris cv. Tendergreen lectin was deduced from the nucleotide sequence of a cDNA plasmid. The lectin appears to be synthesized as a prepeptide consisting of a signal sequence of at least 20 residues and a mature protein of 223 amino acids. The 5' region of the lectin cDNA contains four AUG codons, one of which may initiate a 58 amino acid peptide out of phase with lectin translation. Lectin transcripts 1.1 kb long begin accumulating in cotyledons by the 9 mm stage, increasing in amount until about the 16 mm stage, after which their levels dramatically decrease. In addition, larger lectin-specific transcripts are detected in mid-maturation phase cotyledons. A 1.1 kb RNA species in leaves from plants filling their pods hybridizes with lectin cDNA, but no hybridizing RNA is found in primary leaves or roots.

Amino Acid Sequence↗