Alpha T3 Research Articles

SF-1 Expression Is Mostly Directed by Distal Regulatory Elements.

Steroidogenic factor 1 (SF-1/Ad4BP/NR5A1) is a nuclear hormone receptor that has a pivotal role in endocrine regulation and development of the adrenal glands and gonads. Considering the critical nature of SF-1, many studies have focused on uncovering transcriptional mechanisms that drive SF-1 expression. Despite these efforts, little has been revealed regarding the mechanism that direct SF-1 to its appointed cell types, a result largely due to its requirement for distal regulatory sequences in directing tissue-specific expression. Previous transgenic studies identified a 153kb region, containing Nr5a1, that was sufficient for proper SF-1 expression, which provided an important initiation point for the identification of essential regulatory elements. Comparative genomics through the web-based tool ECR browser (http://ecrbrowser.dcode.org/) was performed and identified within the Nr5a1 locus several evolutionarily conserved non-coding regions called "ECR"s. Four of the most conserved ECRs were examined for transcriptional regulatory activity. The ECRs were cloned into a vector that contained SF-1 promoter sequence from -734 to +60 upstream of a luciferase reporter. Transcriptional activity of each ECR was tested individually using transient transfection analysis in different SF-1-expressing cell types. Our results identified one ECR (ECR3) with activity that increased transcription in alpha T3(gonadotrope) and Y-1(adrenal) cells and decreased it in MA-10(Leydig), Myoid, MSC-1(Sertoli), and primary Sertoli cells. The data indicate that ECR3 contains control elements necessary for SF-1 transcriptional induction in the pituitary and adrenal cells and repression in the testis cells. Mutagenesis and DNA/protein interaction studies identified sequences within ECR3 that were important for activity in alpha T3 cells. One such sequence contains an E-box that bound the class A basic-helix-loop-helix (bHLH) protein E2A, suggesting studies will reveal a class B bHLH, which exhibit tissue-specific expression, as its dimeric partner. This Research was supported by a Center Grant in Reproductive Sciences (U54 HD33994 from NICHD to LLH) and a KUMC Biomedical Research Training Grant to R.K.

Induction of Cells Expressing Vascular Endothelium Markers from Undifferentiated Xenopus Presumptive Ectoderm by Co-treatment with Activin and Angiopoietin-2

Activin is a potent inducer of mesoderm in amphibian embryos. We previously reported that low concentrations of activin could induce the formation of blood cells from Xenopus explants (animal caps). Both hematopoietic and vascular endothelial cell lineages are believed to share a common precursor, termed hemangioblasts. In this study, we tried to induce differentiation of vascular endothelial cells in aggregates derived from Xenopus animal caps. Aggregates formed from cells that were co-treated with activin and angiopoietin-2 expressed the vascular endothelial markers, X-msr, Xtie2 and Xegfl7. However, none of these aggregates expressed the hematopoietic marker genes, globin alpha T3, alpha T5, alpha A or GATA-1. We used microarray analysis to compare the gene expression profiles of aggregates treated with activin alone or with activin and angiopoietin. The combination, but not activin alone, induced expression of vascular-related genes such as Xl-fli and VEGF. These results demonstrate that treatment of dissociated animal cap cells with activin and angiopoietin-2 can induce differentiation of endothelial cells, and provides a promising model system for the in vitro study of blood vessel induction in vertebrates.

Testosterone Stimulates Follicle-Stimulating Hormone β Transcription via Activation of Extracellular Signal-Regulated Kinase: Evidence in Rat Pituitary Cells1

This study investigated whether estradiol (E2) or testosterone (T) activate extracellular signal-regulated kinase (ERK) and calcium/calmodulin-dependent kinase II (Ca/CaMK II), as indicated by enzyme phosphorylation in rat pituitaries. In vivo studies used adult female rats given E2, T, or empty silastic capsules (vehicle controls). Twenty-four hours later, the rats were given a single pulse of GnRH (300 ng) or BSA-saline (to controls) and killed 5 min later. GnRH stimulated a two- to three-fold rise in activated Ca/CaMK II, and E2 and T had no effect on Ca/CaMK II activation. In contrast, both GnRH and T stimulated threefold increases in ERK activity, with additive effects seen following the combination of GnRH+T. E2 had no effect on ERK activity. In alpha T3 clonal gonadotrope cells, dihydrotestosterone did not activate ERK alone but enhanced and prolonged the ERK responses to GnRH, demonstrating direct effects on the gonadotrope. Thus, the ERK response to GnRH plus androgen was enhanced in both rat pituitary and alpha T3 cells. In vitro studies with cultured rat pituitary cells examined the effect of GnRH+/-T in the presence of the mitogen-activated protein (MAP) kinase kinase inhibitor, PD-098059 (PD). Results showed that PD suppressed ERK activational and FSH beta transcriptional responses to T. These findings suggest that one site of T regulation of FSH beta transcription is through the selective stimulation of the ERK pathway.

Different Composite Regulatory Elements Direct Expression of the Human α Subunit Gene to Pituitary and Placenta

To identify elements of the human alpha subunit gene necessary for cell-specific expression, we generated an array of block mutations spanning approximately 400 base pairs (bp) of promoter proximal region and examined them using transient transfection analysis in pituitary (alpha T3) and placental (BeWo) cell lines. Comparison of promoter activity in the two cell types revealed both common and unique elements required for transcription in pituitary and placenta. Two strong elements, the cyclic AMP response element (CRE) and the upstream regulatory element (URE), regulate expression of the alpha subunit gene in BeWo cells. In contrast, promoter activity in alpha T3 cells requires an array of weaker elements. These include the CREs, the URE, as well as two previously described elements, pituitary glycoprotein hormone basal element (PGBE) and gonadotrope-specific element (GSE), and two new elements we designated as the alpha basal elements 1 and 2 (alpha BE1 and alpha BE2). These new elements reside between -316 and -302 bp (alpha BE1) and -296 and -285 bp (alpha BE2) of the human alpha subunit promoter and bind distinct proteins designated alpha BP1 and alpha BP2, respectively. Southwestern blot analysis revealed that alpha BE1 specifically binds 54- and 56-kDa proteins. Additional studies disclosed several potential interactions between proteins that bind the CRE and proteins that occupy PGBE, alpha BE1, and alpha BE2, suggesting that gonadotrope-specific expression occurs through a unique composite regulatory element that includes components of the placenta-specific enhancer.

Absence of rapid desensitization of the mouse gonadotropin-releasing hormone receptor

Desensitization of gonadotropin release by the pituitary gland in response to gonadotropin-releasing hormone (GnRH) agonists has clinical applications in the treatment of gonadal-hormone-dependent disorders. We therefore investigated possible desensitization of inositol phosphate (IP) responses of GNRH receptors. No short-term homologous desensitization of the IP response to GnRH was observed in either alpha T3 gonadotrope cells line or GH3 cells transfected with GnRH receptor cDNA. The absence of homologous desensitization is unusual among G-protein-coupled receptors, and may be due to the absence of a C-terminal cytoplasmic tail, a unique feature of the GnRH receptor. Several potential protein kinase C phosphorylation sites which might mediate heterologous desensitization are present on the GnRH receptor. In both alpha T3 cells and GnRH-receptor-transfected Cos-1 cells, activation of protein kinase C by pretreatment with phorbol ester caused a 35-53% decrease in the IP response to GnRH. However, phorbol ester also inhibited guanosine 5'-[gamma-thio]triphosphate-stimulated IP production in permeabilized Cos-1 cells, suggesting that this inhibition is mediated at a post-receptor site.

Gonadotropin-releasing hormone stimulates glycoprotein hormone alpha- subunit messenger ribonucleic acid (mRNA) levels in alpha T3 cells by increasing transcription and mRNA stability

Pulsatile GnRH stimulates gonadotropin secretion, whereas continuous exposure to GnRH causes pituitary desensitization and suppressed levels of LH and FSH. At the level of gene expression, continuous GnRH also causes partial or complete suppression of the LH beta and FSH beta genes, but expression of the alpha-subunit gene is stimulated without evidence of desensitization. In this report, we examined the transcriptional and posttranscriptional mechanisms by which GnRH controls alpha-gene expression using the gonadotrope-derived alpha T3 cell line. Continuous GnRH caused a 4- to 5-fold accumulation of alpha mRNA over 72 h, without evidence of a decline. In contrast, measurements of alpha-gene transcription, either by nuclear run-on assays or using a stably integrated alpha LUC reporter gene, revealed that GnRH caused a transient increase in alpha-promoter activity, followed by a decline after 4-6 h. The prolonged accumulation of alpha mRNA at a time when transcriptional activity had abated was accounted for by independent effects of GnRH on alpha mRNA stability. After prior treatment with GnRH, its removal either by washout or using a GnRH receptor antagonist caused an abrupt decline in steady state alpha mRNA levels (t1/2, < 2 h). Readdition of GnRH prevented the decay in alpha mRNA, and experiments using the transcriptional inhibitor actinomycin-D confirmed that this effect of GnRH did not require transcription. Consistent with these results, pulse-chase analyses of mRNA stability demonstrated that GnRH increased the alpha mRNA half-life 6.7-fold, from 1.2 h in the absence of GnRH to 8.0 h in the presence of GnRH. We conclude that GnRH induces a transient burst of alpha-gene transcription that is accompanied by marked induction of mRNA stability.

Gonadotropin-releasing hormone stimulates glycoprotein hormone alpha-subunit messenger ribonucleic acid (mRNA) levels in alpha T3 cells by increasing transcription and mRNA stability.

Pulsatile GnRH stimulates gonadotropin secretion, whereas continuous exposure to GnRH causes pituitary desensitization and suppressed levels of LH and FSH. At the level of gene expression, continuous GnRH also causes partial or complete suppression of the LH beta and FSH beta genes, but expression of the alpha-subunit gene is stimulated without evidence of desensitization. In this report, we examined the transcriptional and posttranscriptional mechanisms by which GnRH controls alpha-gene expression using the gonadotrope-derived alpha T3 cell line. Continuous GnRH caused a 4- to 5-fold accumulation of alpha mRNA over 72 h, without evidence of a decline. In contrast, measurements of alpha-gene transcription, either by nuclear run-on assays or using a stably integrated alpha LUC reporter gene, revealed that GnRH caused a transient increase in alpha-promoter activity, followed by a decline after 4-6 h. The prolonged accumulation of alpha mRNA at a time when transcriptional activity had abated was accounted for by independent effects of GnRH on alpha mRNA stability. After prior treatment with GnRH, its removal either by washout or using a GnRH receptor antagonist caused an abrupt decline in steady state alpha mRNA levels (t1/2, < 2 h). Readdition of GnRH prevented the decay in alpha mRNA, and experiments using the transcriptional inhibitor actinomycin-D confirmed that this effect of GnRH did not require transcription. Consistent with these results, pulse-chase analyses of mRNA stability demonstrated that GnRH increased the alpha mRNA half-life 6.7-fold, from 1.2 h in the absence of GnRH to 8.0 h in the presence of GnRH. We conclude that GnRH induces a transient burst of alpha-gene transcription that is accompanied by marked induction of mRNA stability.

Gonadotropin-releasing hormone causes transcriptional stimulation followed by desensitization of the glycoprotein hormone alpha promoter in transfected alpha T3 gonadotrope cells.

Pulsatile GnRH regulates the biosynthesis and secretion of gonadotropins. Continuous administration of GnRH is known to desensitize gonadotropin secretion, but its effects on gonadotropin gene expression are less well characterized. We used a cell line of gonadotrope lineage (alpha T3 cells) to examine GnRH regulation of glycoprotein hormone alpha-subunit gene transcription. The alpha-subunit promoter, linked to a luciferase reporter gene (alpha LUC), was stably transfected into alpha T3 cells. Treatment with GnRH stimulated alpha LUC activity 3-fold. Stimulation of alpha LUC by GnRH was transient, with maximal activity after 6 h of treatment, followed by a return to baseline after 24 h. Stimulation of alpha-promoter activity by GnRH was inhibited entirely by a 10-fold molar excess of antide, a GnRH antagonist. Antide partially blocked GnRH stimulation even when added 4 h after GnRH, suggesting that a brief exposure to GnRH is not sufficient for maximal transcriptional stimulation. alpha LUC activity was also stimulated by treatment with 8-bromo-cAMP (3.5-fold), phorbol 12-myristate 13-acetate (TPA; 2.6-fold), or Bay K 8644 (3.3-fold). To assess whether the transient nature of GnRH stimulation was due to transcriptional desensitization, cells were pretreated with GnRH, followed by a second treatment with GnRH, cAMP, TPA, or Bay K. After pretreatment with GnRH, no further stimulation was seen after the addition of GnRH or TPA, but alpha LUC activity was further stimulated after the addition of either cAMP or Bay K. These findings indicate that the pathway for transcriptional activation by GnRH is desensitized and suggest that GnRH also desensitizes TPA-mediated stimulation. Similarly, pretreatment with TPA, but not cAMP or Bay K, prevented subsequent stimulation by GnRH. We conclude that GnRH transiently stimulates alpha gene transcription and that desensitization occurs with continuous exposure to GnRH, probably because of down-regulation of the protein kinase-C pathway.

Gonadotropin-releasing hormone causes transcriptional stimulation followed by desensitization of the glycoprotein hormone alpha promoter in transfected alpha T3 gonadotrope cells

Gonadotropin-releasing hormone causes transcriptional stimulation followed by desensitization of the glycoprotein hormone alpha promoter in transfected alpha T3 gonadotrope cells

Co-ordinate and restricted expression of the ProXaaArg/Lys-specific GalNAc-transferase and the GalNAc beta 1,4GlcNAc beta 1,2Man alpha-4-sulfotransferase

Asn-linked oligosaccharides terminating with the sequence SO4-4GalNAc beta 1,4GlcNAc beta 1,2Man alpha (S4GGnM) are present on the glycoprotein hormones lutropin and thyrotropin, pro-opiomelanocortin, and tissue factor pathway inhibitor. The peptide motif ProXaaArg/Lys (PXR/K), which is recognized by a PXR/K-specific GalNAc-transferase, is present in each of these glycoproteins 6-9 residues NH2-terminal to an Asn glycosylation site. Both the PXR/K-specific GalNAc-transferase and a GalNAc beta 1,4GlcNAc beta 1,2Man alpha (GGnM)-4-sulfotransferase are required for synthesis of the S4GGnM sequence. Glycoproteins which do not contain the PXR/K motif but bear Asn-linked oligosaccharides terminating with GGnM or sialic acid alpha 2,3/6GGnM have also been described, suggesting a distinct GalNAc-transferase may be responsible for their synthesis. We have examined a number of tissues and cultured cell lines for the transfer of sulfate to the trisaccharide acceptor GGnM and transfer of GalNAc to oligosaccharide acceptors on protein which do, human chorionic gonadotropin (hCG), and do not, transferrin (Trf), contain the PXR/K motif. The PXR/K-specific GalNAc-transferase and the GGnM-4-sulfo-transferase are expressed in salivary gland, pituitary, lacrimal gland, kidney, and brain, and in the cell lines AtT-20, 293, SHSY5Y, and alpha T3. In contrast Bowes, EL-4, and B16L6 cell extracts transferred GalNAc to oligosaccharides acceptors on Trf but not on hCG. A number of tissues and cell lines displayed transfer of GalNAc to both hCG and to Trf suggesting that two distinct GalNAc-transferases were present. The GGnM-4-sulfotransferase was expressed in tissues and cell lines which expressed the PXR/K-specific GalNAc-transferase but not in cell lines expressing exclusively the Trf-specific GalNAc-transferase. Thus, the PXR/K-specific GalNAc-transferase and the GGnM-4-sulfotransferase are coordinately expressed in a number of tissues other than pituitary. The Trf-specific GalNAc-transferase may account for the presence of beta 1,4-linked GalNAc on glycoproteins which do not contain the PXR/K motif.

Studies on the transfer of DNA into cells through use of avidin-polylysine conjugates complexed to biotinylated transferrin and DNA.

A simple procedure has been developed for studying the transfer of DNA into cells using the process of receptor-mediated endocytosis. Poly-L-lysine 460 was covalently attached to the carbohydrate chains of avidin via periodate oxidation and NaBH4 reduction to give avidin-pLys460. Following purification through Sephacryl S-300, the conjugate was reacted with biotinylated transferrin. The resulting avidin-pLys460-[bio-transferrin] could be either fractionated by Superose 12 gel chromatography or used directly in experiments with DNA. Determination of the interaction between avidin-pLys460-]bio-transferrin] and DNA was carried out by nitro cellulose filter binding and agarose gel retardation assays. The conjugate was shown to bind DNA strongly, giving stable complexes soluble in 0.15-0.2 M salt solutions. Gene transfer using avidin-pLys460-[bio-transferrin] and the luciferase plasmid pRSVL was accomplished with Hela cells, alpha T3 pituitary cells and a human melanoma cell line used in the present study. Transfection was dependent on bio-transferrin and stimulated by the lysosomotropic agent chloroquine. The results are consistent with a receptor-mediated endocytosis mechanism of DNA delivery for Hela cells and a combination of receptor and adsorptive endocytosis for the alpha T3 pituitary and melanoma T-5 cell lines.

Molecular cloning and expression of cDNA encoding the murine gonadotropin-releasing hormone receptor.

The primary structure of the gonadotropin-releasing hormone (GnRH) receptor was determined by sequencing a functional receptor cDNA isolated by expression cloning from an immortalized murine gonadotroph (alpha T3) cell line. Positive clone pools from a cDNA library were detected by screening expressed RNA in aequorin-injected Xenopus laevis oocytes, in which receptor-mediated calcium responses were monitored as light emission during stimulation by GnRH. The isolated receptor cDNA encodes a 327-amino acid protein that has seven putative transmembrane regions and is unique among G protein-coupled receptors in that the predicted sequence lacks a carboxyl-terminal cytoplasmic domain. COS-7 cells transfected with the receptor cDNA expressed high affinity binding sites for GnRH and its agonist and antagonist analogs and exhibited calcium responses to GnRH stimulation. These, and the prominent calcium responses of Xenopus oocytes injected with receptor RNA, were inhibited by GnRH antagonists. Northern blot analysis revealed two mRNAs (1.6 and 3.5 kilobases) in alpha T3 cells and in the mouse pituitary gland, and both transcripts were shown to encode functional GnRH receptors when expressed in Xenopus oocytes. In contrast, a single 4.6-kilobase receptor mRNA was present in rat anterior pituitary gland, ovary, and Leydig cells. The absence of a carboxyl-terminal cytoplasmic domain indicates the importance of other regions of the GnRH receptor in agonist-induced signal transduction, and possibly in receptor desensitization and sequestration.

The C'-terminal interaction domain of the thyroid hormone receptor confers the ability of the DNA site to dictate positive or negative transcriptional activity.

To investigate mechanisms responsible for positive and negative transcriptional control, we have utilized two types of promoters that are differentially regulated by thyroid hormone (T3) receptors. Promoters containing the palindromic T3 response element TCAGGTCA TGACCTGA are positively regulated by the T3 receptor after the administration of T3, whereas otherwise identical promoters containing the estrogen response element TCAGGTCA CTG TGACCTGA can be regulated negatively; converse effects are observed with the estrogen receptor. We describe evidence that the transcriptional inhibitory effects of the T3 or estrogen receptors on the estrogen or T3 response elements, respectively, are imposed by amino acid sequences in the C'-terminal region that colocalize with dimerization and hormone-binding domains and that these sequences can transfer inhibitory functions to other classes of transcription factors. Removal of the C'-terminal dimerization and hormone-binding domains of either the alpha T3 or estrogen receptors permits each receptor to act constitutively to enhance transcription on both T3 and estrogen response elements. It is, therefore, suggested that protein-protein interactions between receptor C' termini limit the subset of DNA binding sites on which transcriptional activation occurs.

Gonadotropin-Releasing Hormone Receptor Expression inXenopusOocytes

The rodent GnRH receptor was characterized in Xenopus oocytes injected with RNA isolated from rat pituitary and from a gonadotrope cell line, alpha T3, derived from a transgenic mouse. Three to 4 days after 150-200 ng RNA injection, 93% of the oocytes, which were recorded by voltage clamp, responded to 10(-7) M GnRH. The mean inward currents obtained after RNA injection were 620 +/- 88 nA (n = 22) with pituitary RNA and 1415 +/- 598 (n = 4) with alpha T3 RNA. The threshold GnRH concentration able to evoke the dose dependent current after pituitary RNA injection was 3 x 10(-9) M GnRH. The GnRH receptor response of the oocyte was antagonized by [D-Phe2,6,Pro3] GnRH and [N-Ac-D-Na](2)1, D-alpha D-Me, pCl-Phe2, D-Arg6, D-Ala10-NH2]GnRH and could be elicited by D-Ser(But)6,Pro9-N-ethylamide GnRH (buserelin). The reversal potential of the GnRH generated current as determined by voltage-ramp was -22.5 +/- 1.0 mV (n = 7) and -25.6 +/- 3.3 mV (n = 3) in pituitary and cell line RNA-injected oocytes respectively, consistent with the chloride reversal potential. The GnRH receptor response was virtually eliminated by intracellular EGTA injection but was unaffected by ligand application in calcium-free perfusate. The GnRH-evoked response is mimicked by intracellular injection of inositol 1,4,5-trisphosphate. To determine the size of the GnRH receptor mRNA, alpha T3 RNA was size fractionated through a sucrose gradient. The maximal GnRH response was induced by a fraction larger than the 28S ribosomal peak. Thus we find that oocytes injected with RNA from an appropriate source develop an electrophysiological response to GnRH which is dependent on intracellular calcium mobilization, is independent of extracellular calcium, and may be mediated by inositol 1,4,5-trisphosphate.

Heterogeneity of long-term cultured activated killer cells induced by anti-T3 antibody.

The present study has characterized the short term and long term cultured murine-activated killer (AK) cells that are induced by antibody directed against the epsilon-chain of T3 complex. The conventional lymphokine AK (LAK) cells were generated by culturing normal B6 spleen cells with purified human rIL-2. The alpha T3-induced AK cells (T3AK) were induced by culturing normal B6 spleen cells with alpha T3 and were then maintained in culture medium supplemented with human rIL-2 and/or alpha T3. After initial activation with alpha T3, lymphocyte proliferation and generation of cytotoxic effectors (T3AK) were noted, and these events were related to the endogenous production of IL-2 and IL-4. Addition of alpha IL-2 and/or alpha IL-4 suppressed both the proliferative response and the cytotoxic response induced by alpha T3. In comparing the T3AK cells with the conventional LAK cells, there were many similarities as well as some distinct differences. Both cells displayed a similar cytotoxic spectrum against a variety of tumor targets. The T3AK cells usually gave much higher levels of cytotoxic activity against susceptible targets. However, the susceptibility of different tumor targets to conventional LAK cells and T3AK cells varied. The time course for the generation of lytic activity also differed between the conventional LAK and T3AK cells. One distinct difference was their ability to survive in vitro. The conventional LAK cells survived in culture for only 1 wk. The T3AK cells could survive for at least 4 to 5 wk with active growth. The serologic phenotype of the LAK precursors was asialo GM1 (AsGM1+) cells, but the T3AK precursors could be either AsGM1+ or AsGM1-, depending on the target cell. The LAK effectors were both Lyt-2+ and Lyt-2-, but the short-term T3AK effectors were exclusively Lyt-2+. The long term T3AK cells (cultured for more than 2 wk) were found to consist of Lyt-2+ and Lyt-2- cells, and these subsets of T3AK cells showed different target specificities. These findings demonstrate the heterogeneity of LAK and T3AK cells, and this heterogeneous property may contribute to their diversity in specificity against different tumor targets and thus may affect their effectiveness in the immunotherapy of cancer.

Augmentation by anti-T3 antibody of the lymphokine-activated killer cell-mediated cytotoxicity.

This study showed that a mAb (145-2C11) against the T3 epsilon-chain of the TCR complex augmented the cytotoxic activity of the lymphokine-activated killer (LAK) effectors. The LAK cells were induced by culturing normal spleen cells with purified human rIL-2. Adding alpha T3 at the effector phase of the cytotoxic reactions augmented the LAK-mediated cytotoxicity. The alpha T3-augmented LAK killing was seen only with tumor targets, and there was no increase of killing against Con A-induced lymphoblasts. The augmentation effect was dose dependent on both the amounts of alpha T3 and the number of LAK cells added. A very low concentration of alpha T3 (1/10,000 dilution of culture supernatants) was sufficient to induce alpha T3-augmented LAK-mediated cytotoxicity. Human rIL-2 at 10 to 30 U/ml was sufficient to generate LAK cells for maximal alpha T3 augmentation, whereas 300 to 1000 U/ml of IL-2 were needed to generate maximal LAK activity when tested in the absence of alpha T3. LAK cells generated for longer periods of time showed a progressive increase of alpha T3-augmented cytotoxicity. For some targets, the alpha T3-augmented LAK killing was FcR dependent as evidenced by the ability of alpha FcR mAb 2.4G2 to inhibit, and for others it was not inhibited. The alpha T3-augmented killing did not correlate with the FcR expression on target cells as defined by 2.4G2. The LAK cells were both Lyt-2+ and Lyt-2-, but the LAK cells involved in alpha T3-augmented killing were exclusively Lyt-2+. Preincubation of LAK cells with alpha T3, but not preincubation of targets with alpha T3, resulted in augmented killing suggesting that the alpha T3 effect was unrelated to an antibody-dependent cell-mediated cytotoxicity. Our findings indicate that alpha T3 is a potent reagent to augment the cytotoxic reaction of LAK cells. These results suggested that a relationship might exist between the T3 complex and the cytotoxic activity of a subpopulation of Lyt-2+ LAK cells.

Specific heat of the Anderson lattice at low temperatures

The authors extend previous functional integral approaches to the Anderson lattice in the boson representation by taking into account gaussian fluctuations. They extract a contribution to the specific heat proportional to alpha T3 ln T at low T. The sign of alpha is positive in the Kondo limit and mod alpha mod decreases on approaching the mixed-valence regime.

Low-temperature specific heats of pseudobinary (Ni1-xCux)80P20and (Ni1-xCux)77B13Si10metallic glasses

The low-temperature specific heats in the temperature range 1.5-6K have been measured on four (Ni1-xCux)80P20 (x=0, 0.18, 0.67 and 0.73) and two (Ni1-xCux)77B13Si10 (x=0 and 0.1) pseudobinary metallic glasses. The results can be well fitted to the equation C= gamma T+ alpha T3+A for all alloys studied except for (Ni0.27Cu0.73)80P20, for which the equation C= gamma T+ alpha T3+ delta T5 is better fitted. The band structure change associated with the progressive filling of the Ni d band is discussed on the basis of the plot of the gamma value against the electron concentration. The d band is found to be essentially filled at the Cu content midway between x=0.18 and 0.67 in the Ni-Cu-P alloy system.

Low-temperature specific heats of pseudobinary (a1-xbx)77Si10B13(a, b=Fe, Co, Ni) metallic glasses

Low-temperature specific heats were measured in the temperature range 1.5-6K on 20 metallic glasses in total in the three alloy systems (Fe1-xCox)77Si10B13, (Fe1-xNix)77Si10B13 and (Co1-xNix)77Si10B13. The specific heat is decomposed into three contributions: the electronic specific heat gamma T, the lattice specific heat alpha T3 and the magnetic specific heat Cm. The band structure, as deduced from the concentration dependence of the value of gamma , is found to be quite structureless in comparison with the corresponding behaviour of FCC alloys. The magnetic specific heat depends strongly on the magnetic state involved. The ferromagnetic alloys containing Co exhibit the nuclear specific heat due to the hyperfine interaction of 59Co nuclei with the internal magnetic field. The magnetic specific heat in the spin-glass state is large and its unique temperature dependence can be accounted for in terms of a computer-simulated 'amorphous' spin-glass model.

Low-temperature specific-heat study of Fe-rich Fe-Zr amorphous alloys

Low-temperature specific heats for Fe-rich Fe100-xZrx amorphous alloys with compositions of x=8, 9, 10 and 11 were measured in the temperature range between 1.5 and 6K. The total specific heat is found to follow the linear relation C= gamma T+ beta with a negative coefficient beta , in sharp contrast with the ordinary relation C= gamma T+ alpha T3. The linearly temperature-dependent coefficient is extremely high and well exceeds 20 mJ mol-1 K-2. It increases further with decreasing Zr content. The large coefficient gamma can be ascribed to the sum of electronic and magnetic contributions. The magnetic specific heat can be expressed in the form A0+A1T+A3T3 in the temperature range studied.