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J Ostrowski

Publications and source records attributed to J Ostrowski.

At least 55 records · Page 3Linked to original sources
PubMed1996

Two distinct actions of retinoid-receptor ligands.

Signalling by all-trans retinoic acid is mediated through RXR-RAR retinoid receptor heterodimers, in which RXR has been considered to act as a transcriptionally silent partner. However, we show here that in cultured NB4 (ref. 6) human acute promyelocytic leukaemia cells treated with either an RAR-alpha-selective agonist alone, or certain RAR-alpha antagonists in combination with an RXR agonist, receptor-DNA binding is induced in vivo, resulting in expression of the target genes of retinoic acid as well as acute promyelocytic leukaemia protein (PML) relocation to nuclear bodies and differentiation before apoptosis. These results indicate that RAR-alpha ligands can induce two separate events: one enables RXR-RAR-alpha heterodimers to bind to DNA in vivo and allows RXR agonists to act; the other induces transcriptional activity of RAR-alpha. The availability of receptor-specific synthetic retinoids that can induce distinct receptor functions has potential in extending the therapeutic repertoire of retinoids.

Animals↗
PubMed1996

Cell-type and promoter-context dependent retinoic acid receptor (RAR) redundancies for RAR beta 2 and Hoxa-1 activation in F9 and P19 cells can be artefactually generated by gene knockouts.

By using RAR type (alpha, beta, or gamma)-specific synthetic retinoids and a pan-retinoic X receptor (RXR)-specific ligand, we have investigated the contribution of RARs and RXRs in the activation of RA target genes and the differentiation of embryonal carcinoma cells. We demonstrate cell-type- and promoter context-dependent functional redundancies that differ between the three RAR types for mediating the induction of RARbeta2 and Hoxa-1 in wild-type, RARgamma-/- and RARalpha-/- F9 cells and in P19 cells. The extent of redundancy between RARs is further modulated by the synergistic activation of RXRs with a pan-RXR agonist. We also demonstrate that the expression of RARbeta2 is auto-inducible in RARgamma-/- but not in wild-type F9 cells, indicating that the functional redundancies observed between RARs in gene disruption studies can be artefactually generated. Thus, even though all three RARs can functionally substitute each other for inducing the expression of RA target genes and cell differentiation, one RAR can cell-specifically override the activity of the other RARs. Interestingly, only RARgamma can mediate the retinoic acid-induced differentiation of wild-type F9 cells, whereas the differentiation of P19 cells can be mediated by either RARalpha or RARgamma.

Cell Differentiation↗
PubMed1996

Retinoic acid receptor beta,gamma-selective ligands: synthesis and biological activity of 6-substituted 2-naphthoic acid retinoids.

In search for retinoic acid receptor (RAR) selective ligands, a series of 6-substituted 2-naphthoic acid retinoids were synthesized and evaluated in vitro in a transactivation assay and a competition binding assay for all RARs. These derivatives, in general, showed RAR beta,gamma selectivity. Among these naphthoic acids, oxime derivative 12 was identified as a potent RAR gamma-selective retinoid, while olefinic derivative 11 was found to be comparable to retinoic acid and slightly RAR beta,gamma selective. For the bioassays, a general correlation was observed between the binding affinity of the ligand to the receptors and the potency of the compounds in the transactivation assay. The structure-activity relationship of these naphthoic acids will be discussed.

Binding, Competitive↗
PubMed1996

Phorbol ester (12-O-tetradecanoylphorbol 13-acetate) prevents ornithine decarboxylase inhibition and apoptosis and L1210 leukemic cells exposed to TGF-beta 1.

Previous studies have shown that growth suppression and apoptosis of leukemic cells exposed to TGF-beta 1 is associated with the inhibition of ornithine decarboxylase (ODC)--the key enzyme of polyamine pathway. The aim of the present study was to evaluate the influence of 12-O-tetradecanoylphorbol 13-acetate (TPA)--a potent ODC inducer on antiproliferative and apoptotic effects of TGF-beta 1 in L1210 leukemic cells. Cells were incubated in 2% FCS/RPMI-1640 medium, supplemented with TGF-beta 1 (2 ng/ml). TPA (100 ng/ml) or alpha-difluoromethylornithine (DFMO) (5 mM). Cell proliferation, apoptosis and necrosis were evaluated using [methyl-3H] thymidine, electron microscopy, electrophoresis of DNA and trypan blue exclusion. Expression and activity of ODC were determined by RT-PCR and measurement of 14CO2 release from L-1-14C ornithine, respectively. TGF-beta 1 inhibited proliferation and induced apoptotic and necrotic cell death in L1210 leukemic cells. The above effects were associated with the inhibition of ODC expression and activity, measured 2 and 4 hr after TGF-beta 1 administration, respectively. The presence of DFMO, an irreversible inhibitor of ODC, led to apoptotic fragmentation of DNA, similar to that observed in TGF-beta 1-treated cultures. Administration of TPA simultaneously with TGF-beta 1 significantly reduced antiproliferative, apoptotic and necrotic effects of TGF-beta 1, and prevented its inhibitory action of ODC expression and activity. It is concluded that: down-regulation of ODC expression may be one of the early events associated with TGF-beta 1-evoked suppression of growth and apoptosis; ODC is involved in the mechanism of protective action of TPA on TGF-beta 1-related growth inhibition of L1210 leukemic cells.

Animals↗
PubMed1995

The K protein domain that recruits the interleukin 1-responsive K protein kinase lies adjacent to a cluster of c-Src and Vav SH3-binding sites. Implications that K protein acts as a docking platform.

The heterogeneous ribonucleoprotein particle (hnRNP) K protein interacts with multiple molecular partners including DNA, RNA, serine/threonine, and tyrosine kinases and the product of the proto-oncogene, Vav. The K protein is phosphorylated in vivo and in vitro on serine/threonine residues by an interleukin 1 (IL-1)-responsive kinase with which it forms a complex. In this study we set out to map the K protein domains that bind kinases. We demonstrate that the K protein contains a cluster of at least three SH3-binding sites (P1, PPGRGGRPMPPSRR, amino acids 265-278; P2, PRRGPPPPPPGRG, 285-297; and P3, RARNLPLPPPPPPRGG, 303-318) and that each one of these sites is capable of selectively engaging c-Src and Vav SH3 domains but not SH3 domains of Abl, p85 phosphatidylinositol 3-kinase, Grb-2, and Csk. We demonstrate that the K protein domain that recruits and is phosphorylated in an RNA-dependent manner by the IL-1-responsive kinase, designated KPK for K protein kinase, is contained within the 338-425-amino acid stretch and thus is contiguous but does not include the cluster of the SH3-binding sites. K protein and KPK co-immunoprecipitate from cell extracts with either c-Src or Vav, suggesting that K protein-KPK-c-Src and K protein-KPK-Vav complexes exist in vivo. Furthermore, in the context of K protein, c-Src can reactivate KPK in vitro. The succession of kinase-binding sites contained within the K protein that allow it to form multienzyme complexes and facilitate kinase cross-talk suggest that K protein may serve as a docking platform that promotes molecular interactions occurring during signal transduction.

Amino Acid Sequence↗
PubMed1995

Chromosomal localization and catalytic properties of the recombinant alpha subunit of human lymphocyte methionine adenosyltransferase.

Human lymphocyte methionine adenosyltransferase (HuLy MAT) consists of heterologous subunits alpha and beta. The cDNA sequence of the alpha subunit of HuLy MAT from Jurkat leukemic T cells was identical to that of the human kidney alpha subunit and highly homologous to the sequence of the extrahepatic MAT from other sources. The 3'-untranslated sequence was found to be highly conserved, suggesting that it may be important in regulating the expression of MAT. The extrahepatic alpha subunit unit of MAT was found to be expressed also in human liver, and no differences were found in the sequence of the alpha subunit from normal and malignant T cells. The sequence of two unspliced introns found in the cDNA clones from the Jurkat library enabled us to isolate genomic clones harboring the human extrahepatic alpha subunit gene and to localize it to the centromere on chromosome arm 2p, an area that corresponds to band 2p11.2. Expression of the alpha subunit cDNA in Escherichia coli yielded two peptides with the immunoreactivity and mobilities of authentic alpha/alpha' subunits from HuLy. The Km of the recombinant alpha subunit was 80 microM, which is 20-fold higher than found for the (alpha alpha')x beta y holoenzyme purified from leukemic lymphocytes and 4-10-fold higher than found for the normal lymphocyte enzyme. The data suggest that the alpha/alpha' subunits mediate the enzyme catalytic activity and that the beta subunit may be a regulatory subunit of extrahepatic MAT.

Adult↗
PubMed1995

Description of an IL-1-responsive kinase that phosphorylates the K protein. Enhancement of phosphorylation by selective DNA and RNA motifs.

The K protein was first identified in the heterogeneous ribonucleoprotein particle (hnRNP). Subsequently, K protein was shown to bind sequence-specific single-and double-stranded DNA, stimulate transcription, and bind Src, Fyn, Lyn, and Vav via SH3 interactions. The K protein also binds to the kappa B enhancer motif which stimulates its phosphorylation in vitro by an associated serine/threonine kinase. To gain more insight into this unique nucleic acid-dependent phosphorylation process, we set out to examine the regulation of this kinase. We demonstrate that the K protein exists in a complex with an IL-1-responsive kinase and that phosphorylation of the K protein by this kinase is augmented by either cognate DNA or RNA sequences. The IL-1-responsive kinase activity associated with the K protein is reduced by phosphatase treatment, suggesting that the K protein kinase activity is regulated by phosphorylation. The observation that phosphorylation of the K protein is DNA- or RNA-dependent and IL-1-responsive suggests that the function of the K protein is tightly regulated.

Amino Acid Sequence↗
PubMed1995

RAR-specific agonist/antagonists which dissociate transactivation and AP1 transrepression inhibit anchorage-independent cell proliferation.

Using retinoic acid receptor (RAR) reporter cells specific for either RAR alpha, beta or gamma, we have identified synthetic retinoids which specifically induce transactivation by RAR beta, while antagonizing RA-induced transactivation by RAR alpha and RAR gamma. Like RA, these synthetic retinoids allow all three RAR types to repress AP1 (c-Jun/c-Fos) activity, demonstrating that the transactivation and transrepression functions of RARs can be dissociated by properly designed ligands. Using AP1 reporter cells, we also show that glucocorticoids or vitamin D3, together with either RA or these 'dissociating' synthetic retinoids, can synergistically repress phorbol ester-induced AP1 activity. RA, but not these 'dissociating' retinoids, induced transcription of an interleukin-6 promoter-based reporter gene transiently transfected into HeLa cells together with RARs. Using Ki-ras-transformed 3T3 cells as a model system, we show that both RA and the 'dissociating' retinoids inhibit anchorage-independent cell proliferation, suggesting that retinoid-induced growth inhibition may be related to AP1 transrepression.

Cell Division↗
PubMed1995

The N-terminal portion of domain E of retinoic acid receptors alpha and beta is essential for the recognition of retinoic acid and various analogs.

Utilizing a strategy involving domain exchange between retinoic acid receptors alpha and beta (RAR alpha and RAR beta) and monitoring the transcriptional activity of the resulting chimeric receptors with receptor-selective retinoids, we identified a 70-aa region within the N-terminal portion of the RAR alpha and -beta domain E which is important for an RAR alpha- or RAR beta-specific response. Two amino acid residues within this region, serine-232 (S232) and threonine-239 (T239) in RAR alpha and the corresponding alanine-225 (A225) and isoleucine-232 (I232) in RAR beta, were found to be essential for this effect. In addition, binding studies using the chimeric receptors expressed in Escherichia coli showed that the N-terminal portion of domain E was also important for the characteristic binding profile of t-RA and various retinoids with RAR alpha or RAR beta. Structural predictions of the primary amino acid sequence in this region indicate the presence of an amphipathic helix-turn-helix structure with five hydrophobic amino acids that resemble a leucine zipper motif. The amino acid residues identified by domain swapping, S232 and T239 in RAR alpha and A225 and I232 in RAR beta, were found within the hydrophobic face of an alpha-helix in close proximity to this zipper motif, suggesting that the ligand may interact with the receptor in the region adjacent to a surface involved in protein-protein interactions. This finding may link ligand binding to other processes important for transcriptional activation.

Amino Acid Sequence↗
PubMed1995

Inhibition of acid secretory response and induction of ornithine decarboxylase and its mRNA by TGF alpha and EGF in isolated rat gastric glands.

TGF alpha shows structural and functional homology to EGF, but TGF alpha's mitogenic potency is greater. Our previous study showed that EGF may inhibit parietal cell secretory response through the induction of ornithine decarboxylase (ODC). The aim of this study was to determine parietal cell acid production in vitro in response to stimulation by TGF alpha and EGF and to compare the effect of these two growth factors on ODC activity and ODC mRNA in isolated rat gastric glands. 45 min treatment with TGF alpha and EGF had no effect on basal acid production but did inhibit histamine-stimulated acid production in a dose dependent manner. The two growth factors did not inhibit histamine-stimulated aminopyrine (AP) uptake from incubation medium with concentration of KCl increased from 5 to 70 mM. In the presence of specific ODC inhibitor, alpha-difluoromethylornithine (DFMO), both EGF and TGF alpha failed to inhibit histamine-stimulated AP accumulation. Polyamine spermine also inhibited AP accumulation but this inhibitory effect was not affected by DFMO. After 1 h treatment with TGF alpha and EGF, ODC activities increased to an average 283 +/- 78% and 227 +/- 64% above the basal activity, respectively. 30 min treatment of gastric glands with TGF alpha and EGF resulted in, respectively, 2.9 +/- 0.4- and 2.7 +/- 0.5-fold increases of ODC mRNA level, as assessed by RT-PCR.(ABSTRACT TRUNCATED AT 250 WORDS)

Aminopyrine↗
PubMed1995

Effect of orotic acid on TGF-beta 1-induced growth inhibition of L1210 leukemic cells.

UNLABELLED: Transforming growth factor-beta 1 (TGF-beta 1) exerted growth-inhibitory effect on L1210 leukemic cell line, manifested by the decrease in viable and increase in dead cells. The cell death evoked by TGF-beta 1 was both necrotic and apoptotic, quantified by the trypan blue exclusion method and apoptotic index, respectively. The induction of programmed cell death by TGF-beta 1 was confirmed by gel electrophoresis of DNA, where the characteristic 'DNA ladder' resulting from the internucleosomal DNA cleavage was visualized. The enhancement of cell mortality by TGF-beta 1 was associated with the inhibition of ornithine decarboxylase (ODC) expression (measured by the reverse transcriptase-polymerase chain reaction method) and impaired activity of this key enzyme in polyamine synthesis. Orotic acid (OA)--a known tumor promoter--stimulated proliferation of L1210 leukemic cells and diminished the necrotic effect of TGF-beta 1, but it did not change the extent of apoptosis evoked by TGF-beta 1. OA increased the expression of ODC and diminished depressional influence of TGF-beta 1 on transcription and activity of ODC in leukemic cells. IN CONCLUSION: OA is a bioactive compound stimulating the growth of leukemic cells and diminishing the growth-inhibitory effect of TGF-beta 1. ODC gene is probably one of the targets for both OA and TGF-beta 1 influences in L1210 leukemic cells.

Animals↗
PubMed1995

The effect of OA on proliferation and polyamine metabolism of K 562 leukemic cells and their responsiveness to natural killer cell activity.

Orotic acid (OA), a known promoter of carcinogenesis, significantly stimulated proliferation of K 562 leukemic cells even at as high a concentration as 0.1 mM. This effect was accompanied by a significant increase of the activity of two key enzymes of the polyamine pathway, i.e. ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase (SAMDC). The induction of ODC activity was associated with increased expression of the ODC gene. The participation of ODC in early events evoked by OA in leukemic cells was confirmed by the decrease of the stimulatory effect of OA on cell proliferation in the presence of alpha-difluoromethylornithine (DFMO)--an irreversible inhibitor of ODC. The involvement of protein kinase C (PK-C) and cyclic nucleotide-dependent kinases in OA action on K 562 leukemic cells was demonstrated by a significant reduction of cell proliferation by addition of H-7 (1-(5-isoquinolinesulphonyl)-2-methylpiperazine). Since PK-C is involved both in induction of ODC activity and in membrane transport of polyamines, H-7 significantly inhibited the proliferation of K 562 leukemic cells even in the presence of OA and exogenous putrescine. The importance of extracellular sources of polyamines for leukemic cell growth was shown by supplementation of the incubation medium with putrescine. Exogenous putrescine significantly enhanced the concentration of spermidine and spermine within the cell and increased the number of cells. The effect of OA on natural killer (NK) cell cytotoxicity was also examined. Rat peripheral blood mononuclear cells were used as effector cells and K 562 cells as targets. OA, progressively with dose, significantly decreased specific lysis when targets were preincubated with it. On the other hand, pretreatment of PBMC effector cells with OA, regardless of the applied concentration, did not affect the amount of 51Cr released from lysed cells. OA as a promoter of carcinogenesis stimulates proliferation of leukemic cells and impairs their responsiveness to NK activity. ODC/polyamine system and PK-C appear to be involved in OA action on K 562 cells. The presented observations are important from a practical point of view, since an elevated blood concentration of OA resulting from the impaired kidney function in hematological proliferative diseases may accelerate their progression.

Base Sequence↗
PubMed1995

Retinoic acid receptor gamma mediates topical retinoid efficacy and irritation in animal models.

Among retinoic acid receptors (RARs) alpha, beta, and gamma, the messenger RNA level of RAR-gamma is the most readily detectable by Northern blotting in human and mouse skin. This observation suggests that RAR-gamma may play a critical role in the modulation of the therapeutic benefits and side effects of retinoids in skin. To test this hypothesis, 11 RAR-gamma selective retinoids were synthesized based on three related structures. Each compound was found to prefer RAR-gamma when assessed by retinoid-induced transcriptional activity (RAR-gamma > RAR-beta > RAR-alpha). The apparent Kd for binding to recombinant receptor protein was found to follow a similar trend. To correlate this receptor selectivity with in vivo activity, the compounds were tested topically in the Rhino mouse utriculi reduction and rabbit irritation models, two assays widely used to screen retinoids for efficacy and side effects, respectively. The results indicated that for these compounds, both efficacy in the utriculi reduction assay and irritation potential in rabbits correlated positively with the RAR-gamma transactivation activity, with r2 of 0.9 and 0.8, respectively. These data suggest that RAR-gamma is an important regulator of retinoic acid efficacy in skin and further, that the irritation associated with the use of retinoids is most likely a receptor-mediated process.

Animals↗
PubMed1995

Role of retinoic acid receptor gamma in the Rhino mouse and rabbit irritation models of retinoid activity.

The three retinoic acid receptors (RAR alpha, RAR beta and RAR gamma) are known to modulate the transcription of target genes through interaction of the individual receptors with their naturally occurring ligand, retinoic acid (RA). Since RA has multiple effects in vivo, considerable effort has recently been devoted to finding selective compounds to elucidate the functions of individual receptors and to relate these functions to specific in vivo effects. The racemic synthetic retinoid 6-[(5,5,8,8-tetramethyl-5,6,7,8-tetrahydro-2-naphthyl)hydroxy-methyl]-2- naphthalene carboxylic acid has recently been identified as an RAR gamma-selective agonist. A synthetic method involving lipase-mediated transformation has been developed to prepare the individual enantiomers. Discrimination between the two enantiomers is seen in both transcriptional activity and binding to recombinant receptors with the (S)-enantiomer being the more active. Differences between the two compounds are also seen in the Rhino mouse utriculi reduction assay and the rabbit irritation model. In both animal models, the (S)-enantiomer consistently gave a greater response. Taken together, these results suggest that the activity and irritation seen with RA and related compounds is receptor mediated. Further, the strong selectivity of the compounds reported here for RAR gamma suggests that this receptor plays an important role in these in vivo biological activities. The discrimination between these enantiomers may be useful in the design of novel retinoids with uniquely defined biological properties.

Animals↗
PubMed1995

[Evaluation of acceptance rate and outcome of renal replacement therapy in patients with diabetic nephropathy--multicenter study].

Development of dialysis methods and progress in kidney and pancreas transplantation allowed to treat an increasing number of patients suffering from diabetic nephropathy (D.N.). This report evaluates availability and results of treatment in these patients. 31.12.93 in Gdansk and Bydgoszcz area there were treated 519 patients, including 43 (8.2%) with D.N. It is impossible to evaluate the demand for renal replacement therapy in patients with D.N., because there is no exact data concerning diabetic patients with progressing renal failure. Up to now 88 patients with D.M. (68 with IDDM, 20 with NIDDM) were treat in this area. Most of them (92%) were treated with hemodialysis is and only a few with CAPD, 13 patients received a kidney graft. The average patient survival on dialysis treatment in NIDDM patients was 15 months and in IDDM patients was 11 months. Deaths were mainly caused by cardiovascular complications. The results of renal replacement therapy in these patients cannot be compared with data from other re ports, because the treatment was introduced at advanced stage of D.N. in patients with systemic complications (serum creatinine in IDDM was 9.7 md% and in NIDDM was 6.2% mg%). Following conclusions can be drawn from our observations: 1. There is a need for close cooperation between diabetologist and nephrologist in repeat of evaluation of the demand for renal replacement therapy and time for its institution in a particular patient. 2. The choice of method of renal replacement therapy depends on clinical findings in a particular patient but also on methods available in a particular center. 3. Improvement of therapy outcome can be achieved primarily by earlier institution of dialysis (serum creatinine below 5 m5%).

Adolescent↗
PubMed1994

Purification, cloning, and expression of a murine phosphoprotein that binds the kappa B motif in vitro identifies it as the homolog of the human heterogeneous nuclear ribonucleoprotein K protein. Description of a novel DNA-dependent phosphorylation process.

The kappa B enhancer element regulates expression of many genes involved in immune responses and other processes. kappa B motif binds a number of proteins, some but not all, are related to the NF-kappa B family of transcription factors. We have previously identified a 65-kDa phosphoprotein that is specifically recognized by the kappa B motif (Ostrowski, J., Sims, J. E., Sibley, C. H., Valentine, M. A., Dower, S. K., Meier, K. E., and Bomsztyk, K. (1991) J. Biol. Chem. 266, 12722-12733). This protein is closely associated with a serine/threonine kinase that is responsive to treatment of cells with interleukin-1 and other agents. We report here purification, cloning, and expression of this kappa B motif-binding phosphoprotein. The primary structure deduced from the isolated murine cDNA, identifies the protein as the homolog of the human heterogeneous nuclear ribonucleoprotein K protein. Antipeptide antibodies and expression of the cloned cDNA in Escherichia coli, demonstrated that the K protein is the authentic phosphoprotein that binds the kappa B motif in vitro. We also demonstrate that the in vitro phosphorylation of the natural and the recombinant K proteins by the associated kinase is stimulated by the kappa B motif.

Amino Acid Sequence↗
PubMed1994

Functional and structural changes of isolated rat parietal cells during membrane potential modulation.

The present experiments were undertaken to extent our earlier observations (J Physiol Pharmacol 1991, 42, 367-79) relating membrane potential with membrane recycling of parietal cells. Studies were performed in vitro using gastric glands that were isolated through the use of rat stomachs transformed into "everted sacs" and filled with hyperosmolar NaCl-EDTA solution. Acid production was indirectly determined by accumulation of 14C-aminopyrine (AP) and its translocation by measurement of acridine orange fluorescence. H+/K(+)-ATPase activity was assayed by measurement of K(+)-stimulated p-nitrophenylphosphatase (pNPPase) of the proton pump. Morphologic state of parietal cells in relation to their functional activity was observed using electron microscopy. Changes in the membrane potential were obtained by the treatment of gastric glands with protonophore carbonyl cyanide m-chlorophenylhydrazone (CCCP) in the incubation media of different pH. CCCP caused time-dependent decrease in AP accumulation by parietal cells from the medium of pH 6.6 but not that of pH 7.8. pNPPase activity increased in aplical and decreased in tubulovesical membrances prepared from CCCP treated glands which were incubated in the medium being more acidic than cell cytoplasm. Electron microscopic assessment showed morphological transformation of resting parietal cells treated with CCCP in pH 6.6 from nonsecreting to secreting state. CCCP acting in acidic incubation medium also caused the decrease in acridine orange fluorescence in the cytoplasm of parietal cells with some temporary increase of its fluorescence in the lumen o gastric glands. These findings support our hypothesis that changes in parietal cell membrane potential by protonophore CCCP may translocate HCl from tubulovesicles to secretory canaliculi. While the above explanation is suggestive, the exact mechanisms controlling a membrane recycling during the secretory response of parietal cells in vitro remain to be elucidated.

4-Nitrophenylphosphatase↗