Search PubMed⌕ Search

Biomedical subjects

S Demczuk

Publications and source records attributed to S Demczuk.

31 records · Page 2Linked to original sources
PubMed1992

Thyroid hormone alters the DNA binding properties of chicken thyroid hormone receptors alpha and beta.

The effects of thyroid hormone agonists on thyroid hormone receptor (TR)/DNA complex formation was investigated to elucidate the mechanism by which TRs transactivate genes in response to ligand. The data, obtained from gel shift experiments, indicate that thyroid hormones alter the conformation of TRs bound to DNA, irrespective of if the element is occupied by monomeric TR, homodimeric TR/TR, or heterodimeric complexes with the retinoid receptors RAR or RXR. Furthermore, triiodo-thyronine (T3) prevents 2 TR molecules from binding to oligonucleotides containing direct repeats or inverted palindromes of the consensus AGGTCA motif, an effect that was not detected with palindromic elements. Heterodimers bound to direct repeats were less affected: RXR/TR were fully and RAR/TR complexes partially resistant to thyroid hormone. The data suggest that a ligand-induced conformational change in TR prevents double TR occupancy of a response element containing 2 direct repeats of the consensus binding motif, possibly by steric hindrance, whereas such an event does not prevent TR/RXR heterodimers from binding to DNA. Finally, our data show that a monomeric, liganded TR bound preferentially to the second half site in a AGGTCActcaAGGTCA element, and therefore indicate that nucleotides adjacent to the consensus half site contribute to binding specificity.

Animals↗
PubMed1991

Involvement of a calpain-like protease in the processing of the murine interleukin 1 alpha precursor.

Interleukin (IL) 1 alpha is synthesized as a 33-kDa precursor that is enzymatically cleaved to the 15-17-kDa forms that are found in the culture supernatants of activated macrophages. We have explored the possibility that calcium might enhance IL-1 processing and secretion via the stimulation of a calcium-dependent protease. We have found that lysates prepared from human peripheral blood monocytes, the human histiocytic lymphoma cell line U937, and the murine macrophage cell line P388D1 contain a calcium-dependent IL-1 alpha processing activity that cleaves the IL-1 alpha precursor to its mature form. Although NIH 3T3 mouse fibroblast cell lysates also contain IL-1 processing activity, lysates from the murine thymoma EL-4, the human epidermoid cell line HEp-2, and the human foreskin fibroblast line FS-4 lack this activity. IL-1 processing activity is inhibited by leupeptin and exhibits a molecular mass of 80-110 kDa. The processing activity is also inhibited by a monoclonal antibody directed against calpain type I. These results indicate that the processing of the IL-1 alpha precursor is mediated, at least in part, by a member of the calpain family of proteases. Mixing experiments revealed that lysates from EL-4 or HEp-2 cells contain an inhibitor(s) of the calpain-like protease in macrophage extracts. It is, therefore, likely that many non-macrophage cell types are unable to process the IL-1 alpha precursor because the calpain present in these cells is only weakly active due to the presence of a specific inhibitor(s) such as calpastatin.

Animals↗
PubMed1991

Oncogenes: a review of their clinical application.

One objective of this review is to sort through and collate the recent data that suggest that human cellular oncogenes, which have been implicated as the etiologic agents in both animal and human malignancies, have also the potential to be employed as clinical tools in the struggle against cancer. For nearly 10 years, reports have been suggesting that advantage can be taken of cellular oncogenes as to their use as diagnostic and prognostic indicators of cancer and eventually as therapeutic cancer agents. It is also the purpose of this review to give an objective evaluation of these predictions. Moreover, this review will try to highlight some of the significant advances in this most rapidly evolving field of biology. Although the enormity of what has been learned about cellular oncogenes is nothing less than impressive, it is the view here that the routine implementation of oncogenes into the clinical setting will not become evident as early as the many predictions had purported.

Gene Expression Regulation, Neoplastic↗
PubMed1988

IL1/ETAF activity and undetectable IL1 beta mRNA and minimal IL1 alpha mRNA levels in normal adult heat-separated epidermis.

Interleukin 1 (IL 1), a cytokine expressed from a number of cell types, plays a vital part in immune and inflammatory processes. Epidermal IL1-like activity, also known as epidermal cell thymocyte-activating factor (ETAF), is associated with whole skin, epidermis, Langerhans cells and cultured keratinocytes and keratinocyte cell lines. We show that preparations of heat-separated epidermis possess abundant IL1/ETAF activity as indicated by PGE/collagenase induction in cultured synovial cells and proliferation of mouse thymocytes. However, using cDNA probes to interleukin-1 alpha and beta we show that freshly isolated heat-separated epidermis from healthy adult donors contains only a slight detectable level of IL1 alpha mRNA by dot blot analysis and no IL1 beta and IL1 alpha mRNA by Northern blot analysis. The implications of these observations will be discussed.

Blotting, Northern↗
PubMed1988

Differential effects of in vitro mycoplasma infection on interleukin-1 alpha and beta mRNA expression in U937 and A431 cells.

Cultured cells depend on cytokine mediators for sustained growth and maintenance and are routinely employed in bioassays to detect and measure minute changes in biological mediators, e.g. the interferons and interleukins. We evaluated the effects of mycoplasma infection on the steady-state mRNA levels of two cytokines IL-1 alpha and beta. Noninfected human squamous carcinoma cell line A431 expressed constitutively IL-1 alpha and beta mRNA. In contrast freshly isolated peripheral blood mononuclear cells and the monocytic cell line U937 expressed abundant IL-1 mRNA only after the appropriate stimulation. Peripheral blood mononuclear cells and U937 steady-state IL-1 beta mRNA levels were considerably greater than IL-1 alpha mRNA levels, whereas nearly equivalent high levels of IL-1 alpha and beta mRNA were detected in A431 cells. Mycoplasma infection of cultured A431 cells reduced the steady-state levels of IL-1 alpha and beta mRNA. This effect was nonspecific for A431 cells as actin mRNA steady-state levels showed similar decreases to mycoplasma contamination. However, this response was cell specific since mycoplasma-free and contaminated U937 cells differed little in IL-1 mRNA expression. These results show that the response to mycoplasma infection is at least partly cell-type dependent.

Cell Line↗
PubMed1987

Limited effects of recombinant human and murine interleukin 1 and tumour necrosis factor on production of acute phase proteins by cultured rat hepatocytes.

Albumin, fibrinogen, alpha 1-acid glycoprotein and cysteine proteinase inhibitor were determined by electroimmunoassay in the media of primary cultures of rat hepatocytes exposed to dialysed supernatants of rat, mouse and human macrophages or to recombinant human and murine interleukin 1 and tumour necrosis factor. Recombinant cytokines in the range of 1 to 1000 ng/ml caused only reduction of albumin synthesis and slight stimulation of alpha 1 acid glycoprotein production while crude preparations of macrophage cytokines elicited typical acute phase response. The results suggest that interleukin 1 or tumour necrosis factor are not likely the principal mediators responsible for the direct stimulation of normal rat hepatocytes to acute phase protein synthesis.

Animals↗
PubMed1987

Expression of human IL 1 alpha and beta messenger RNAs and IL 1 activity in human peripheral blood mononuclear cells.

The macrophage-derived lymphokine interleukin 1 (IL 1) plays a critical role in modulating immune (cellular and humoral) and nonimmune responses. For example, the relative expression of IL 1 alpha and beta under various states may be crucial to the success of the immune system in response to infection. Until recently, a comparative study of IL 1 mRNA expression and IL 1 biological activity was not possible. We have cloned both IL 1 alpha and beta cDNAs and employed them as probes in Northern blot analysis to determine in mitogen-stimulated peripheral blood mononuclear cells the steady-state expression of their cognate mRNAs with respect to IL 1 activity. IL 1 was determined by the lymphocyte-activating factor (IL 1/LAF) and the mononuclear cell factor (IL 1/MCF) activities. In lectin-stimulated PBMC, maximum cell-associated activities whereas detected at 12 and 24 hr after stimulation whereas maximum extracellular activities appeared between 24-48 hr. In the same cultures, the kinetics of IL 1 mRNA steady-state expression were determined by Northern gel blot analysis with IL 1 alpha and beta cDNA probes. IL 1 mRNAs were undetectable in noncultured freshly isolated PBMC (time zero). Both IL 1 mRNAs appeared as early as 4 hr after lectin stimulation as did IL 1 beta mRNA in unstimulated cultures. Both IL 1 alpha and beta mRNA steady-state levels were barely detectable by 48 hr. At all time points, IL 1 mRNA levels were considerably lower in unstimulated cultures. IL 1 beta mRNA was always considerably more abundant than IL 1 alpha mRNA. The less abundant IL 1 alpha mRNA showed a decrease in its stead-state levels prior to the reduction in the levels of IL 1 beta mRNA. TNF alpha activity and mRNA were not detected under these culture conditions. Poly(A) + RNA injected into Xenopus oocytes revealed that the Northern blot detected IL 1 mRNAs were biologically active. To understand the precise nature of IL 1 in immune and nonimmune events, we felt it necessary to first study the kinetics of IL 1 mRNA steady-state levels with respect to its cell-associated and extracellular biological activities. The data presented here may allow for a better understanding of the etiology of various immune and nonimmune responses that are modulated through the expression of IL 1.

Animals↗
PubMed1986

Purification and characterization of human interleukin-1 beta expressed in recombinant Escherichia coli.

The high-level expression of human interleukin-1 beta in Escherichia coli is described. The protein contributes about 12% of the total cell protein and is associated with the soluble cytoplasmic fraction of the cell. A method for the purification of the protein is given which is based on anion- and cation-exchange chromatographies. The isolated protein, shown to be homogeneous by several analytical methods, has been characterized by amino acid analysis, N- and C-terminal sequence analysis and analytical centrifugation. The protein is biologically active as demonstrated by two different in vitro assays, namely, the mononuclear cell factor (IL-1/MCF) assay and lymphocyte activating factor (IL-1/LAF) assay. The specific activities determined with the IL-1/MCF and IL-1/LAF assays, are 2 X 10(7) and 4 X 10(7) units mg-1, respectively.

Amino Acid Sequence↗
PubMed1986

Human recombinant interleukin 1 stimulates collagenase and prostaglandin E2 production by human synovial cells.

The pathogenesis and progression of rheumatoid arthritis involves the production of biologically active lymphokines and monokines. Of these, interleukin 1 (IL-1) has been somewhat of a controversial molecule because it seems to evoke various biological responses in several different tissues. In these studies we demonstrate that three biological properties of human monocyte-derived IL-1 (T-lymphocyte activation and human synovial cell prostaglandin E2 and collagenase production) co-purify. The complementary DNA for the prominent pI 7 form of human IL-1 was expressed, purified, and tested. Any controversy now appears resolved since homogeneous recombinant human IL-1 stimulates prostaglandin E2 and collagenase from human synovial cells as well as activates T cells in vitro.

Animals↗
PubMed1986

Changes in hepatic RNA, poly(A)+RNA, and poly(A)-RNA during the acute phase response to inflammation.

The steady state changes in total rat hepatic cytoplasmic RNA, poly(A)+ RNA and poly(A)-RNA were assessed in response to turpentine induced inflammation. From 18 to 24 h after injury, cytoplasmic RNA doubled, while poly(A)+ RNA peaked at 24 h, 3.5 times over control animals. Cell-free translation showed significant increases in messenger RNA levels beginning at 18 h. Gel electrophoresis of translation products revealed significant increases in several polypeptides and a decrease in others. Poly(A)-RNA from control and injured rats translated to an insignificant level and the electrophoretic gel patterns of their proteins were similar. Furthermore, no change had occurred in the 3' poly(A)-sequences during the course of inflammation.

Acute Disease↗