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Biomedical subjects

M Whitman

Publications and source records attributed to M Whitman.

At least 19 recordsLinked to original sources
PubMed1996

A transcriptional partner for MAD proteins in TGF-beta signalling.

The transforming-growth-factor-beta (TGF-beta) superfamily is critical for establishing mesoderm during early embryogenesis in Xenopus. The transcriptional activation of Mix.2, an immediate-early response gene specific to activin-like members of the TGF-beta superfamily, is associated with the rapid appearance of a site-specific DNA-binding activity that recognizes a fifty-base-pair regulatory element known as ARE in the Mix.2 promoter. Cloning of the site-specific DNA-binding component of this activity revealed it to be a new winged-helix transcription factor and a direct target for signalling by the TGF-beta superfamily. XMAD2, a recently identified TGF-beta signal transducer, forms a complex with the transcription factor in an activin-dependent fashion to generate an activated ARE-binding complex. A model is proposed to explain how TGF-beta superfamily signals might regulate the expression of specific genes in the early embryo.

Activins↗
PubMed1995

Identification of a potential regulator of early transcriptional responses to mesoderm inducers in the frog embryo.

The activin/transforming growth factor-beta (TGF-beta) family of peptide growth factors plays a central role in the induction of mesoderm during early Xenopus embryogenesis. Immediate transcriptional responses to mesoderm-inducing signals have been described, but the signal transduction steps leading to these early responses are unknown. We describe here the first pre-transcriptional response to activin/TGF-beta mesoderm inducers in the early embryo. We have identified a cellular factor which binds to a 50 bp portion of the promoter for an activin/TGF-beta early response gene. This factor is activated within 4 min of treatment of embryonic prospective ectoderm with mesoderm-inducing factors, making it the earliest response to these factors described in early embryos. This factor can be activated throughout early cleavage and blastula stages, is activated by mesoderm inducers of the activin/TGF-beta superfamily but not the fibroblast growth factor (FGF) family, and does not appear to require an endogenous FGF signal for activation. Characterization of this factor provides a powerful tool for studying the early steps in the induction of mesoderm by members of the activin/TGF-beta superfamily.

Activins↗
PubMed1995

Role of MAP kinase in mesoderm induction and axial patterning during Xenopus development.

We have examined the role of MAP kinase during mesoderm induction and axial patterning in Xenopus embryos. MAP Kinase Phosphatase (MKP-1) was used to inactivate endogenous MAP kinase and was found to prevent the induction of early and late mesodermal markers by both FGF and activin. In whole embryos, MKP-1 was found to disrupt posterior axial patterning, generating a phenotype similar to that obtained with a dominant inhibitory FGF receptor. Overexpression of either constitutively active MAP kinase or constitutively active MAP kinase (MEK) was sufficient to induce Xbra expression, while only constitutively active MEK was able to significantly induce expression of muscle actin. When MAP kinase phosphorylation was used as a sensitive marker of FGF receptor activity in vivo, this activity was found to persist at a low and relatively uniform level throughout blastula stage embryos. The finding that a low level of MAP kinase phosphorylation exists in unstimulated animal caps and is absent in caps overexpressing a dominant inhibitory FGF receptor provides a basis for our previous observation that overexpression of this receptor inhibits activin induction. These results indicate that FGF-dependent MAP kinase activity plays a critical role in establishing the responsiveness of embryonic tissues to mesoderm inducers.

Activins↗
PubMed1994

Normal magnetic resonance imaging and abnormal discography in lumbar disc disruption.

STUDY DESIGN: This report describes a clinical series of seven patients who had surgically proven internal disc disruption, normal magnetic resonance imaging, and abnormal discograms morphologically. SUMMARY OF BACKGROUND DATA: Numerous reports in the literature have described the utility of magnetic resonance imaging and discography in diagnosing degenerative disease within lumbar intervertebral discs. RESULTS: Discography may be useful in patients with persistent symptoms despite a normal or equivocal magnetic resonance imaging study.

Adult↗
PubMed1994

Mesoderm induction by activin requires FGF-mediated intracellular signals.

We have examined the role of FGF signaling during activin-mediated mesoderm induction in Xenopus. Using dominant inhibitory mutants of FGF signal transducers to disrupt the FGF-signaling pathway at the plasma membrane or in the cytosol prevents animal cap blastomeres from expressing several mesodermal markers in response to exogenous activin. Dominant inhibitory mutants of the FGF receptor, c-ras or c-raf inhibit the ability of activin to induce molecular markers of both dorsal and ventral mesoderm including Xbra, Mix1 and Xnot. Some transcriptional responses to activin such as goosecoid and Xwnt8 are inhibited less effectively than others, however, suggesting that there may differing requirements for an FGF signal in the responses of mesoderm-specific genes to activin induction. Despite the requirement for this signaling pathway during activin induction, downstream components of this pathway are not activated in response to activin, suggesting that activin does not signal directly through this pathway.

Activins↗
PubMed1992

Involvement of p21ras in Xenopus mesoderm induction.

During early vertebrate embryogenesis, mesoderm is specified by a signal emanating from prospective endoderm. This signal can respecify Xenopus prospective ectoderm as mesoderm, and can be mimicked by members of the fibroblast growth factor and transforming growth factor-beta families. In other systems, the p21c-ras proto-oncogene product has been implicated in signal transduction for various polypeptide growth factors. We report here that a dominant inhibitory ras mutant blocks the mesoderm-inducing activity of fibroblast growth factor and activin, as well as the endogenous inducing activity of prospective endoderm. A constitutively active ras mutant partially mimics these activities. These results indicate that p21ras may have a central role in the transduction of the mesoderm inductive signal. Basic fibroblast growth factor and activin have emerged as candidates for endogenous mesoderm-inducing molecules. The character of the mesoderm induced by these two factors is overlapping but distinct when assessed both by histological and molecular criteria. The signal transduction pathways used during induction by these factors are unknown. We used messenger RNA microinjection of Xenopus eggs to express a dominant inhibitory mutant ras, p21(Asn 17)Ha-ras, in cells competent to respond to inducing factors to examine the role of p21ras in this response. This mutant, which has a reduced affinity for GTP relative to GDP, blocks a variety of mitogenic signals in 3T3 fibroblasts as well as the differentiation of pheochromocytoma cells in response to nerve growth factor.

Activins↗
PubMed1992

PCR detection of Borrelia burgdorferi DNA in cerebrospinal fluid of Lyme neuroborreliosis patients.

We used the polymerase chain reaction (PCR), a method useful in the detection of Borrelia burgdorferi in vitro, to evaluate CSF in patients thought to have neuroborreliosis. Nested pairs of oligonucleotide primers were designed to recognize the C-terminal region of B burgdorferi OspA. CSF samples were obtained from (1) patients with immunologic evidence of systemic B burgdorferi infection and clinical manifestations suggestive of CNS dysfunction, (2) seronegative patients with clinical disorders consistent with Lyme borreliosis, and (3) patient and contamination controls; all were analyzed in a blinded fashion. PCR detected B burgdorferi OspA DNA in CSF of (1) 10 of 11 patients with Lyme encephalopathy, (2) 28 of 37 patients with inflammatory CNS disease, (3) seven of seven seronegative patients with Lyme-compatible disorders, and (4) zero of 23 patient controls. Zero of 83 additional contamination controls were PCR-positive. In eight patients from whom we obtained CSF before and after parenteral antimicrobial therapy, PCR results invariably predicted clinical outcome accurately.

Antibody Formation↗
PubMed1990

Activins are expressed early in Xenopus embryogenesis and can induce axial mesoderm and anterior structures.

We show that mammalian and Xenopus activins induce dorsal axial mesoderm and anterior structures in explants of Xenopus blastula cells that would otherwise form epidermis. The induced explants of animal cap cells can form notochord, muscle, neural tissue, and eyes all arranged in a rudimentary axial pattern. Activin A shares inductive properties and antigenic determinants with PIF, an inducing factor recently isolated from mouse macrophage culture supernatants. Genes encoding Xenopus activin beta A and beta B chains were cloned. Activin beta B transcripts are first detected in Xenopus blastula, whereas activin beta A transcripts do not appear until the late gastrula stage. Recombinant Xenopus activin beta B protein induces mesodermal and neural tissues similar to those induced by mammalian activin A and PIF. Furthermore, ectopic expression of Xenopus activin beta B produces a second body axis in embryos injected with synthetic mRNA. Our results suggest that early induction and axial patterning are accomplished by endogenous activin B, not activin A, in Xenopus.

Activins↗
PubMed1989

Induction of mesoderm by a viral oncogene in early Xenopus embryos.

During frog embryogenesis, mesoderm is specified in the equatorial region of the early embryo by a signal from the vegetal hemisphere. Prospective ectodermal cells dissected from the animal hemisphere can be respecified to form mesodermal tissues by recombination with vegetal tissue or by treatment with any of several polypeptide growth factors or growth factor-like molecules. Together with the discovery that several developmental mutations in Drosophila are in genes with significant homology to mammalian mitogens and oncogenes, these observations suggest that early developmental signals may use similar transduction pathways to mitogenic signals characterized in cultured mammalian cells. Whether mesoderm can be induced by activation of intracellular signal transduction pathways implicated in mitogenesis and oncogenesis has been investigated with the viral oncogene polyoma middle T. Microinjection of middle T messenger RNA into early embryos results in the respecification of isolated prospective ectodermal tissue to form characteristic mesodermal structures. Middle T in frog blastomeres appears to associate with cellular activities similar to those observed in polyoma-transformed mouse cells, and transformation-defective middle T mutants fail to induce mesoderm. These results suggest that early inductive signals and mitogenic and oncogenic stimuli may share common signal transduction pathways.

Animals↗
PubMed1989

Induction of c-fos protein by activation of vasopressin receptors in smooth muscle cells.

Stimulation of vasopressin (V1) receptors of rat aortic smooth muscle cells (A-10, ATCC CRL 1476) results in the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) to inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG) with the mobilization of intracellular calcium. When A-10 cells are exposed to arginine vasopressin (AVP), there is an increase in the level of c-fos oncoprotein. The extent of induction of c-fos oncoprotein depends on both the time of exposure of the cells to AVP, reaching a maximum at 60 min after which there is a slow decline, and the concentration of AVP used, with an approximate EC50 of 1 nM which corresponds well with the Kd of vasopressin binding to these receptors. This vasopressin-mediated increase in c-fos protein level is inhibited by a V1/V2 antagonist (SKF 101498) suggesting that this is a receptor-mediated event. In addition dDAVP, a V2 selective agonist, is much less effective than AVP in inducing c-fos protein suggesting that AVP mediates its effect via V1 receptors. Desensitization of vasopressin receptors by prolonged exposure to AVP resulted in no additional induction of c-fos protein level in response to second challenge of AVP. In addition to AVP, phorbol dibutyrate (PDBu), an activator of protein kinase C (PKC), also stimulates the accumulation of c-fos protein although to a lesser extent than AVP. The above data suggest that c-fos protein levels in smooth muscle cells are regulated by AVP and the hormonal effect may be mediated through PI turnover and DAG, IP3 and Ca2+ signals.

Animals↗
PubMed1989

Homologous and heterologous desensitization mediated by vasopressin in smooth muscle cells.

Prolonged exposure of A-10 cells to Arginine Vasopressin (AVP) resulted in the following responses: (a) loss of vasopressin receptors from the cell surface (30-40%), (b) increased basal levels of inositol and inositol monophosphate, (c) decreased inositol di- and trisphosphate production and decreased intracellular calcium release in response to a second challenge with AVP, (d) attenuation of AVP-mediated inhibition of isoproterenol-stimulated cAMP and ANF-stimulated cGMP accumulation and (e) attenuation of thrombin and ATP-mediated increase in inositol di- and trisphosphate accumulation and intracellular calcium release. All the above responses depended on the time of exposure of the cells to AVP with the responses being attenuated as early as 5-10 min of exposure to AVP. The desensitization also depended on the concentration of AVP used with 50% of maximal desensitization for each response being observed at 5 nM of AVP. This concentration of AVP corresponded well with the Kd of vasopressin for binding to these sites. Desensitization of protein kinase C (PKC) by prolonged exposure of the cells to PDBu or addition of the PKC inhibitor staurosporine during pretreatment with AVP did not prevent AVP-mediated desensitization, suggesting that PKC may not be involved in AVP-mediated desensitization in smooth muscle cells. It is concluded that AVP induced both homologous and heterologous desensitization of phosphatidylinositol turnover and calcium release in smooth muscle cells. The desensitization processes did not appear to be mediated by protein kinase C. The possibility that the locus of the heterologous desensitization may be at the level of substrates such as PI, PIP and PIP2 is discussed.

Adenosine Triphosphate↗