Molecular analysis of T cell receptor (Ti) variable region (V) gene expression. Evidence that a single Ti beta V gene family can be used in formation of V domains on phenotypically and functionally diverse T cell populations.

We examine the rules governing Ti beta variable (V) gene segment usage in the formation of T cell antigen-MHC receptors in diverse regulatory and effector T lymphoid subpopulations. To this end, a single Ti beta V gene family and its products were analyzed. A monoclonal antibody, termed anti-Ti3A, which was shown to be reactive with an epitope encoded by members of the REX cell line Ti beta V gene family, and which was expressed on 2% of human T lymphocytes was used in selection of clones from unprimed peripheral T lymphocytes. Both T4+, as well as T8+ T cell clones with inducer, suppressor, and/or cytotoxic function were defined. Southern analysis, isoelectric focusing and two-dimensional peptide mapping indicated that individual members of the REX V gene family were linked to different Ti beta diversity and/or joining and constant region segments. Moreover, the Ti alpha chains of such clones were distinct. These results imply that Ti beta V gene usage is not restricted to any functionally or phenotypically defined T cell subsets, and there is presumably little, if any, restriction on the mechanisms that generate combinational, junctional or chain association-mediated diversity.

Analysis of T-cell receptor gene rearrangement and expression in human natural killer clones.

A series of clones of human natural killer (NK) cells was characterized with respect to expression of the Ti alpha and Ti beta genes of the T-cell receptor. T11+T3+ NK clones contained Ti alpha and Ti beta RNA transcripts and expressed disulfide-linked heterodimers, demonstrating the presence of a functional T-cell receptor. In contrast, T11+T3- NK clones expressed only 1.0-kilobase truncated Ti beta transcripts, without a Ti alpha transcript and no detectable surface Ti protein. Since previous studies demonstrated that Ti beta gene activation precedes Ti alpha gene activation in thymic ontogeny, the T11+T3- NK cells appear to be derived from T-lineage precursors.

The ontogeny, structure and function of the human T lymphocyte receptor for antigen and major histocompatibility complex.

Recent studies using cloned antigen-specific T lymphocytes and monoclonal antibodies directed at their various surface glycoprotein components have led to identification of the human T cell antigen receptor as a surface complex comprised of a clonotypic 90 kDa Ti heterodimer and the invariant 20 and 25 kDa T3 molecules. Approximately 30,000-40,000 Ti and T3 molecules exist on the surface of human T lymphocytes. These glycoproteins are acquired and expressed during late thymic ontogeny, thus providing the structural basis for immunological competence. The alpha and beta subunits of Ti bear no precursor-product relationship to one another and are encoded by separate germline V, D, J and C segments which rearrange during intrathymic differentiation to form an active gene set. Triggering of the T3-Ti receptor complex induces a rapid increase in free cytoplasmic Ca2+ and gives rise to specific antigen-induced proliferation through an autocrine pathway involving endogenous interleukin-2 production, release and subsequent binding to interleukin-2 receptors. The implications of these findings for understanding of human T cell growth and its regulation in disease states are discussed.

Spironolactone: a glucocorticoid agonist or antagonist?

Spironolactone displaces 3H-dexamethasone from HTC cell nuclei and inhibits the induction of tyrosine aminotransferase (TAT) activity by dexamethasone in HTC cells. Spironolactone alone fails to increase TAT activity in HTC cells or in the liver of adrenalectomized rats. These actions indicate that spironolactone lacks glucocorticoid agonist activity but can be an antagonistic to the actions of glucocorticoid as well as mineralocorticoid steroid hormones.

Mineralo- and glucocorticoid effects on renal excretion of electrolytes.

The acute effects of mineralo- and glucocorticoids on urinary electrolyte excretion were studied in the conscious, acutely potassium deprived, adrenalectomized rat. Sodium, potassium, and creatinine were measured in the urine excreted from 2.5 to 5.5 h after injection of one or more of the following steroids: aldosterone (Aldo), 9-alpha fluorocortisol (FC), deoxycorticosterone (DOC), dexamethasone (Dex), and spironolactone (Spiro). The hierarchy (a) for increasing creatinine excretion was Dex greater than FC greater than Aldo greater than DOC greater than Spiro greater than none, a hierarchy consistent with glucocorticoid potency; and (b) for producing anti-natriuresis was Aldo greater than DOC greater than or equal to FC greater than or equal to none = Spiro greater than Dex, a hierarchy consistent with mineralocorticoid potency. In contrast, the kaliuresis produced by mineralo- and glucocorticoids appears different. A "mineralocorticoid" kaliuresis is 1) elicited by anti-natriuretic doses of Aldo and FC, 2) approximately twice control UKV, 3) unrelated to changes in glomerular filtration rate (GFR), and 4) inhibited by Spiro. A "glucocorticoid" kaliuresis is 1) elicited by Dex and high doses of Aldo and FC, 2) about seven to twenty-fold greater than control UKV, 3) possibly dependent, in part, on changes in GFR, and, 4) not inhibited by Spiro. DOC was not kaliuretic at anti-natriuretic doses. The urinary Na/K ratio was an unreliable index of mineralocorticoid action.

Genes encoding the T-cell receptor alpha and beta subunits are transcribed in an ordered manner during intrathymic ontogeny.

To further characterize sequential events involved in activation of genes encoding the T-cell receptor (a complex of T3 molecules and a disulfide-linked heterodimer designated Ti, T3-Ti) for antigen and the major histocompatibility complex during intrathymic ontogeny, cDNA probes specific for Ti alpha and Ti beta subunits were used for transcriptional analysis. Ti beta transcript levels were minimal in stage I thymocytes, maximal in stage II thymocytes, and intermediate in state III thymocytes. In contrast, Ti alpha transcriptional activity was virtually undetectable in stage I, was low in stage II, and achieved high levels only in the stage III compartment. Analysis of tumor populations derived from individual stages of thymic differentiation confirmed these observations and demonstrated that clonal stage I-II cells often express Ti beta RNA in the absence of Ti alpha RNA. The latter was not a consequence of functional Ti alpha-subunit isotypy because each of 13 interleukin 2-dependent T-cell clones, including inducer, suppressor, and cytotoxic T cells, contain transcripts that hybridize with the Ti alpha probe. From these data it is concluded that Ti beta gene activation precedes Ti alpha gene activation. Moreover, the high level of Ti beta mRNA preceding Ti alpha mRNA expression implies that Ti beta mRNA and/or its protein products may regulate Ti alpha gene transcription.

The structure and function of the T3-Ti molecular complex on human T lymphocytes.

Recent studies using cloned antigen-specific T lymphocytes and monoclonal antibodies directed at their various surface glycoprotein components have led to identification of the human T cell antigen receptor as a surface complex composed of a clonotypic 90-kDa Ti heterodimer and the invariant 20- and 25-kDa T3 molecules. Approximately 30,000-40,000 Ti and T3 molecules exist on the surface of human T lymphocytes. These glycoproteins are acquired and expressed during late thymic ontogeny, thus providing the structural basis for immunologic competence. The Ti alpha and Ti beta subunits bear no precursor-to-product relationship and are encoded by separate germ line V, D, J, and C segments, which rearrange during intrathymic differentiation to form an active gene set. Triggering of the T3-Ti receptor complex induces a rapid increase in free cytoplasmic Ca2+ and gives rise to specific antigen-induced proliferation through an autocrine pathway involving endogenous IL 2 production, release, and subsequent binding to IL 2 receptors.

Intraocular pressure elevation after simple pars plana vitrectomy.

PURPOSE: The purpose of the study is to determine the incidence, timing, and severity of variability in the intraocular pressures (IOPs) from baseline after simple pars plana vitrectomy. METHODS: A prospective study was performed in 25 consecutive patients undergoing simple pars plana vitrectomy. Intraocular pressures were measured before surgery, immediately after surgery, and then at 2, 4, 6, 12, and 24 hours after surgery. RESULTS: The mean IOP was elevated significantly 2 hours after surgery when compared with the mean immediate postoperative IOP (30.3 mmHg +/- 11.0 mmHg vs. 17.4 mmHg +/- 7.0 mmHg, P < 0.001). A steady decline was seen at all succeeding timepoints. The 24-hour mean (17.3 mmHg +/- 4.3 mmHg, P = 0.923) was similar to baseline. Ninety-two percent of eyes had a 2-hour postoperative IOP that was higher than the IOP at the completion of surgery. Forty percent of patients required medical management for IOP greater than or equal to 30 mmHg. CONCLUSIONS: Significant IOP elevation can occur after simple pars plana vitrectomy. The optimal time for detecting the pressure rise during the first 24 hours is 2 hours after surgery.

The human T lymphocyte receptor complex for antigen and MHC.

Recent studies using cloned antigen specific T lymphocytes and monoclonal antibodies directed at their various surface glycoprotein components have led to identification of the human T cell antigen receptor as a surface complex comprised of a clonotypic 90 KD Ti heterodimer and the in-variant 20 and 25 KD T3 molecules. Approximately 30,000-40,000 Ti and T3 molecules exist on the surface of human T lymphocytes. These glycoproteins are acquired and expressed during late thymic ontogeny, thus providing the structural basis for immunologic competence. The alpha and beta subunits of Ti bear no precursor-product relationship to one another and are encoded by separate germline V, D, J and C segments which rearrange during intrathymic differentiation to form an active gene set. Triggering of the T3-Ti receptor complex induces a rapid increase in free cytoplasmic Ca2+ and gives rise to specific antigen-induced proliferation through an autocrine pathway involving endogenous IL-2 production, release, and subsequent binding to IL-2 receptors. The implications of these findings for understanding of human T cell growth and its regulation in disease states are discussed.

The human T-cell receptor.

Recent studies using cloned antigen-specific T lymphocytes and monoclonal antibodies directed at their various surface glycoprotein components have led to the identification of the human T-cell antigen receptor as a surface complex comprised of a clonotypic 90-kD Ti heterodimer and the invariant 20- and 25-kD T3 molecules. Approximately 30,000-40,000 Ti and T3 molecules exist on the surface of human T lymphocytes. These glycoproteins are acquired and expressed during late thymic ontogeny, thus providing the structural basis for immunologic competence. The alpha and beta subunits of Ti bear no precursor-product relationship to one another and are encoded by separate genes. Moreover, the presence of unique peptides following proteolysis of different Ti molecules isolated by non-cross-reactive anticlonotypic monoclonal antibodies supports the notion that variable regions exist within both the alpha and the beta subunits. N-Terminal amino acid sequencing and molecular cloning of the Ti beta subunit further show that it bears an homology to the first V-region framework of immunoglobulin light chains and represents the product of a gene that rearranges specifically in T lymphocytes. Triggering of the T3-Ti receptor complex gives rise to specific antigen-induced proliferation through an autocrine pathway involving endogenous IL-2 production, release, and subsequent binding to IL-2 receptors. The implications of these findings for understanding human T-cell growth and its regulation in disease states are discussed.

Variation among human 28S ribosomal RNA genes.

We report the complete 5025-base sequence of the human 28S rRNA gene. Variability within the species has been demonstrated by sequencing a variable region from six separately cloned genes. This region is one of three large subunit rRNA regions that show extreme sequence and size variation among species. The interspecies differences suggest species-specific functions for these sections, while the intraspecies heterogeneity indicates differences among ribosomes. Comparison of the human gene with a partial sequence from the chimpanzee 28S gene yields divergence rates for the two species: 0.8% for conserved regions of the gene and 3.7% for a variable region. The rapid divergence rates of variable regions in the ribosomal gene may permit answers to the question of time of separation of closely related species.