scholarly journals Human recombinant interleukin-1 regulates cellular mRNA levels of dermatan sulphate proteoglycan core protein

1988 ◽  
Vol 252 (1) ◽  
pp. 309-312 ◽  
Author(s):  
J Heino ◽  
V M Kähäri ◽  
A Mauviel ◽  
T Krusius
Keyword(s):  
Culture Medium ◽  
Core Protein ◽  
Interleukin 1 ◽  
Mrna Levels ◽  
Interleukin 1 Beta ◽  
Cell Culture Medium ◽  
Human Skin Fibroblasts ◽  
Dermatan Sulphate ◽  
Maximal Increase ◽  
Sulphate Proteoglycan

Human skin fibroblasts were exposed to various concentrations (from 0.01 to 5.0 units/ml) of human recombinant interleukin-1 alpha and interleukin-1 beta (IL-1 alpha and IL-1 beta). Both IL-1 alpha and IL-1 beta were found to increase dermatan-sulphate-proteoglycan (DSPG) core-protein mRNA levels. Maximal increase (3.0-fold) was seen at 48 h after addition of 1 unit of IL-1 beta/ml. In spite of the elevated DSPG-core-protein mRNA only a slight increase (from 10 to 18%) could be seen in the production of DSPG to cell-culture medium. No changes in the molecular mass of DSPG could be detected.

Thyroid hormone excess stimulates the synthesis of proteoglycan in human skin fibroblasts in culture

1990 ◽  
Vol 123 (5) ◽  
pp. 541-549 ◽  
Author(s):  
Yoshimasa Shishiba ◽  
Yasuhiro Takeuchi ◽  
Noriko Yokoi ◽  
Yasunori Ozawa ◽  
Taeko Shimizu
Keyword(s):  
Thyroid Hormone ◽  
Human Skin ◽  
Specific Activity ◽  
Heparan Sulphate ◽  
Skin Fibroblasts ◽  
Proteoglycan Synthesis ◽  
Heparan Sulphate Proteoglycan ◽  
Human Skin Fibroblasts ◽  
Dermatan Sulphate ◽  
Sulphate Proteoglycan

Abstract We previously demonstrated that proteoglycan accumulated in the affected skin of circumscribed pretibial myxedema of Graves' disease. As an underlying mechanism responsible for the accumulation, we sought to determine whether excess thyroid hormone was partially responsible for the increase in proteoglycan synthesis. Human skin fibroblasts were cultured in Ham's F-10 medium containing 1% Nutridoma with graded doses of T3 (0.184 × 10−9 to 46 × 10−9 mol/l) and were labelled with [35S]sulphate and [3H]glucosamine. Proteoglycans were purified by Sephadex G-50, Q-Sepharose chromatography with NaCl-gradient and Sepharose CL-6B chromatography. 35S and 3H incorporated into dermatan sulphate proteoglycan and heparan sulphate proteoglycan and 3H incorporated into hyaluronan were measured. 35S and 3H incorporation into dermatan sulphate proteoglycan was minimum at a T3 concentration of 0.184 × 10−9 mol/l, and increased with increasing doses of T3 up to 46 × 10−9 mol/l. 35S and 3H incorporation into heparan sulphate proteoglycan also increased with increasingdoses of T3. 3H incorporation into hyaluronan was not influenced at all by T3. The increased incorporation of 35S into proteoglycan in high-T3 culture reflects the increased synthesis of proteoglycan because 1. the extent of sulphation of disaccharides examined by thin-layer chromatography was not altered by T3; 2. the specific activity of [35S]sulphate was not influenced by T3, and 3. T3 did not decrease the degradation rate of cell-associated proteoglycan.

scholarly journals Endocytosis of a small dermatan sulphate proteoglycan. Identification of binding proteins

1989 ◽  
Vol 263 (1) ◽  
pp. 137-142 ◽  
Author(s):  
H Hausser ◽  
W Hoppe ◽  
U Rauch ◽  
H Kresse
Keyword(s):  
Binding Proteins ◽  
Core Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Scatchard Analysis ◽  
Dermatan Sulphate ◽  
Intact Cells ◽  
Sulphate Proteoglycan ◽  
Human Osteosarcoma Cells ◽  
Antibody Complex ◽  
Triton X

Endosomal preparations from human osteosarcoma cells and from fibroblasts contain 51,000- and 26,000-Mr proteins which bind a small dermatan sulphate proteoglycan after SDS/polyacrylamide-gel electrophoresis and Western blotting. Binding can be inhibited by unlabelled proteoglycan core protein. The proteins co-precipitate with a proteoglycan core protein-antibody complex. Scatchard analysis of immobilized endosomal proteins yielded a KD of about 37 nM for the proteoglycan. In intact cells proteins of the same size can be found. They are sensitive to trypsinization. A 51,000-Mr protein is the predominant membrane protein with strong binding to immobilized dermatan sulphate proteoglycan. There are additional proteoglycan-binding proteins with Mr values of around 30,000 and 14,000 which are insensitive to trypsin treatment. In contrast with the 51,000- and 26,000-Mr proteins, they resist deoxycholate/Triton X-100 extraction several days after subcultivation.

Colchicine has opposite effects on interleukin-1 beta and tumor necrosis factor-alpha production

1991 ◽  
Vol 261 (4) ◽  
pp. L315-L321 ◽  
Author(s):  
J. N. Allen ◽  
D. J. Herzyk ◽  
M. D. Wewers
Keyword(s):  
Tumor Necrosis Factor ◽  
Tumor Necrosis ◽  
Cytokine Production ◽  
Interleukin 1 ◽  
Tnf Alpha ◽  
Mrna Levels ◽  
Interleukin 1 Beta ◽  
Necrosis Factor Alpha ◽  
Factor Alpha ◽  
Necrosis Factor

To study the role of microtubules in cytokine production, the effect of the microtubule depolymerizing agent colchicine on lipopolysaccharide endotoxin (LPS)-induced interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha) release by blood monocytes and alveolar macrophages were examined. Immunofluorescence microscopy demonstrated that LPS resulted in the appearance of microtubule-containing cytoplasmic appendages and that colchicine, which resulted in microtubule disruption in monocytes, blocked appendage formation. Colchicine resulted in approximately 50% increase in LPS-induced IL-1 beta release and a 50% decrease in LPS-induced TNF-alpha release by human monocytes at all doses of LPS tested. Although colchicine resulted in a statistically significant increase in LPS-stimulated human alveolar macrophage IL-1 beta release, the increase was not as great as that observed with monocytes. Northern blot analysis suggested that the colchicine effect occurs pretranslationally because colchicine caused an increase in LPS-stimulated IL-1 beta mRNA levels and a decrease in TNF-alpha mRNA levels. These results suggest that microtubules contribute to the regulation of endotoxin-stimulated mononuclear phagocyte cytokine production and that this regulation differs significantly between IL-1 beta and TNF-alpha.

The Histone Deacetylase-8 (HDAC8) Selective Inhibitor PCI-34051 Decreases Interleukin-1 Beta Secretion in Vitro and Reduces Inflammation in Vivo

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 2581-2581
Author(s):  
Sriram Balasubramanian ◽  
Susanne Steggerda ◽  
Mint Sirisawad ◽  
Marshall Schreeder ◽  
Luke Doiron ◽  
...  
Keyword(s):  
T Cell ◽  
Secretory Pathway ◽  
Interleukin 1 ◽  
Contact Hypersensitivity ◽  
Secretory Process ◽  
Mrna Levels ◽  
Interleukin 1 Beta ◽  
Caspase 1 ◽  
T Cell Lymphomas

Abstract Inhibitors of histone deacetylases (HDACs) which are currently in clinical testing for treating various cancers typically inhibit multiple isoforms of the 11-member HDAC family. We have developed an isoform-selective HDAC inhibitor, PCI-34051, that inhibits HDAC8 with a Ki of 10 nM and greater than 200-fold selectivity over other HDAC isoforms (Balasubramanian et al. (2008) Leukemia,22:1026–34). We have shown that PCI-34051 selectively induced apoptosis in cell lines derived from T-cell lymphomas and leukemias, but not in other tumor or normal cell types. Here we show that it potently inhibits the secretion of the pro-inflammatory cytokine interleukin-1 beta (IL-1b) in lipopolysaccharide (LPS)-stimulated peripheral mononuclear blood cells (PBMC) and isolated monocytes. PCI-34051 inhibited IL-1b secretion (by 80% compared to control) from LPS-stimulated human PBMC with an IC50 of 0.6 uM, which is much lower than the growth inhibitory concentrations of 2.4–4 uM required in T-cell lymphomas. We found that PCI-34051 also inhibited the secretion of interleukin-18 (IL-18) to a similar extent as IL-1b, but secretion of other pro-inflammatory cytokines, including MIP-1b, MCP-1, TNFa, and IL-6, was inhibited to a smaller extent. Interestingly, IL-18, like IL-1b, is synthesized without a signal peptide, and also utilizes the same non-classical endosomal secretory pathway as IL-1b including cleavage of the pro-form by caspase-1. Thus, we theorized that the modulatory effect of PCI-34051 is likely to involve modulation of the post-translational secretory process. In accordance, we found that the IL-1b mRNA levels were reduced by only 20% compared to control, but the intracellular protein levels of the pro-form was increased by >50% in primary monocytes after treatment with PCI-34051, indicating that the mechanism was due to inhibition of the processing from the pro- to the mature form of the cytokine. We showed that this was not due to a direct inhibition of caspase-1 or TACE (TNF-alpha converting enzyme), but is likely due to an as-yet unidentified substrate of HDAC8. In vivo, PCI-34051 inhibited ear swelling induced by oxazolone in a model of contact hypersensitivity in mice, and we showed that this was accompanied by a reduction in IL-1b at both the protein and mRNA levels. Based on this result, we examined the effect of PCI-34051 on IL-1b secretion in human keratinocytes, as well as in PBMC from psoriasis patients, and found that it could reduce IL-1b secretion in both. We found that PCI-34051 decreased IL-1b by 60% in LPS-stimulated PBMC from rheumatoid arthritis (RA) patients, but the pan-HDAC inhibitors which were only weakly inhibitory to HDAC8 did not have this effect, indicating a specific role for HDAC8 in the secretory process. Finally, we found that in unstimulated PBMC from RA patients that had basal production of IL-1b that this could be decreased by 90% by treatment with PCI-34051. Taken together, these findings indicate that PCI-34051 is an active drug that could be useful for the treatment of T-cell lymphoma as well as for autoinflammatory diseases such as RA and psoriasis.

scholarly journals Dermatan sulphate proteoglycans of human articular cartilage. The properties of dermatan sulphate proteoglycans I and II

1989 ◽  
Vol 262 (3) ◽  
pp. 823-827 ◽  
Author(s):  
P J Roughley ◽  
R J White
Keyword(s):  
Articular Cartilage ◽  
Gel Electrophoresis ◽  
Core Protein ◽  
Polyacrylamide Gel Electrophoresis ◽  
Polyacrylamide Gel ◽  
Human Articular Cartilage ◽  
Amino Acid Residues ◽  
Dermatan Sulphate ◽  
Sulphate Proteoglycan ◽  
Core Proteins

Dermatan sulphate proteoglycans were purified from juvenile human articular cartilage, with a yield of about 2 mg/g wet wt. of cartilage. Both dermatan sulphate proteoglycan I (DS-PGI) and dermatan sulphate proteoglycan II (DS-PGII) were identified and the former was present in greater abundance. The two proteoglycans could not be resolved by agarose/polyacrylamide-gel electrophoresis, but could be resolved by SDS/polyacrylamide-gel electrophoresis, which indicated average Mr values of 200,000 and 98,000 for DS-PGI and DS-PGII respectively. After digestion with chondroitin ABC lyase the Mr values of the core proteins were 44,000 for DS-PGI and 43,000 and 47,000 for DS-PGII, with the smaller core protein being predominant in DS-PGII. Sequence analysis of the N-terminal 20 amino acid residues reveals the presence of a single site for the potential substitution of dermatan sulphate at residue 4 of DS-PGII and two such sites at residues 5 and 10 for DS-PGI.

Regulation of prostaglandin endoperoxide synthase gene expression in rat mesangial cells by interleukin-1 beta

1994 ◽  
Vol 266 (1) ◽  
pp. F39-F45 ◽  
Author(s):  
D. Rzymkiewicz ◽  
K. Leingang ◽  
N. Baird ◽  
A. R. Morrison
Keyword(s):  
Steady State ◽  
Mesangial Cells ◽  
Interleukin 1 ◽  
Primary Cultures ◽  
Northern Analysis ◽  
Gene Transcript ◽  
Mrna Levels ◽  
Interleukin 1 Beta ◽  
Extracellular Medium ◽  
Prostaglandin Endoperoxide

In primary cultures of rat mesangial cells from passage 3 to 6, interleukin-1 beta (IL-1) induced a time-dependent increase in prostaglandin E2 (PGE2) formation and release into the extracellular medium. This increase was associated with a dramatic upregulation of the steady-state levels of mRNA for the prostaglandin endoperoxide synthetase (PES)-2 gene transcript as demonstrated by Northern analysis. In contrast, there did not appear to be a significant increase in the mRNA levels for a 2.8-kb transcript for the PES-1 gene. At 18 h of exposure to IL-1, the steady-state level of message for PES-2 remained elevated at 50% of the 2-h time point. Culturing the cells in the presence of cycloheximide and IL-1 demonstrated a superinduction of the PES-2 message without any change in PES-1 message. The tumor-promoting phorbol ester, phorbol myristate acetate (PMA), was also associated with an upregulation of the message for the PES-2 gene and did not influence the levels of the message for the PES-1 gene as demonstrated by Northern analysis. Dexamethasone (Dex) inhibited to control levels the induction by PMA, but the induction of the message by IL-1 was only inhibited 30%. Despite 70% of the message being present by 2 h of induction, Dex was capable of totally inhibiting the inductive effect of IL-1 with respect to PGE2 biosynthesis. Immunocytochemical studies demonstrated a dramatic induction of PES-2 protein by IL-1, which was inhibited by Dex. The data suggest that Dex inhibits the translation of the PES-2 protein.

Effects of intracellular ions on interleukin-1 beta production by lipopolysaccharide-activated human monocytes

1992 ◽  
Vol 263 (5) ◽  
pp. C1073-C1080 ◽  
Author(s):  
U. Orlinska ◽  
R. C. Newton
Keyword(s):  
Culture Medium ◽  
Choline Chloride ◽  
Interleukin 1 ◽  
Disulfonic Acid ◽  
Interleukin 1 Beta ◽  
Activated Monocytes ◽  
Lps Activation ◽  
Intracellular Ions ◽  
Ionophore Monensin

Following the observation that interleukin 1 beta (IL-1 beta) production in lipopolysaccharide (LPS)activated monocytes increases in concert with a rise in intracellular pH (pHi), the role of ion transport in IL-1 beta production was investigated. The amiloride analogue 5-(N-ethyl-N-isopropyl)amiloride (EIPA), an inhibitor of the Na(+)-H+ antiporter, inhibited extracellular IL-1 beta. The replacement of Na+ in the culture medium with sucrose or choline chloride also prevented monocyte activation. The sodium ionophore monensin, in doses from 100 pM to 1 microM, potentiated LPS-stimulated extracellular IL-1 beta when compared with LPS alone. In the absence of LPS activation, monensin by itself at 10 nM stimulated IL-1 beta production to 63%. EIPA at 10 microM inhibited the Na+ influx, the rise in pHi, and intra- and extracellular IL-1 beta production in activated monocytes; this inhibition was reversed by 10 nM monensin. In the absence of bicarbonate, or in the presence of 10 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, the pHi of activated monocytes and the total protein synthesis did not change, but the production of IL-1 beta was inhibited. The data suggest that the stimulated influx of Na+ via the Na(+)-H+ antiporter regulates both pHi and IL-1 beta production in LPS-activated monocytes. The requirement for bicarbonate indicates an additional mechanism(s), separate from the modulation of pHi and intracellular Na+.

Posttranscriptional modulation of gene expression in cultured rat hepatocytes

1984 ◽  
Vol 4 (9) ◽  
pp. 1929-1934
Author(s):  
D M Jefferson ◽  
D F Clayton ◽  
J E Darnell ◽  
L M Reid
Keyword(s):  
Mrna Stability ◽  
Culture Medium ◽  
Rat Hepatocytes ◽  
Mrna Levels ◽  
Cell Culture Medium ◽  
Mrna Stabilization ◽  
Specific Mrnas ◽  
Cultured Rat Hepatocytes ◽  
Modulation Of Gene Expression ◽  
Specific Mrna

The maintenance of high levels of two liver-specific mRNAs in cultured hepatocytes was achieved in a serum-free hormonally defined cell culture medium. However, this maintenance of liver-specific mRNA levels did not correlate with the level of transcription of the genes but was apparently due to increased stabilization of the tissue-specific mRNAs. The mRNA stabilization did not occur in serum-supplemented medium. In both defined and serum-supplemented medium, actin and tubulin mRNAs were also greatly increased, in both cases predominantly if not entirely due to increased mRNA stability.

scholarly journals Characterization of proteoglycans synthesized by human adult glomerular mesangial cells in culture

1991 ◽  
Vol 277 (1) ◽  
pp. 81-88 ◽  
Author(s):  
G J Thomas ◽  
R M Mason ◽  
M Davies
Keyword(s):  
Culture Medium ◽  
Mesangial Cells ◽  
Cell Layer ◽  
Heparan Sulphate ◽  
Glomerular Mesangial Cells ◽  
Heparan Sulphate Proteoglycan ◽  
Dermatan Sulphate ◽  
Sulphate Proteoglycan ◽  
Human Adult ◽  
Core Proteins

1. The newly synthesized proteoglycans from human adult glomerular mesangial cells labelled in vitro for 24 h with [35S]sulphate have been characterized using biochemical and immunological techniques. 2. The following proteoglycans were identified (% of total synthesized). (i) A large chondroitin sulphate proteoglycan, CSPG-I, Mr approximately 1 x 10(6) (10.6%). This proteoglycan consisted of a protein core of Mr approximately 4 x 10(5) and glycosaminoglycan chains of Mr 2.5 x 10(4), and was present in both the cell layer and the culture medium. (ii) A major small dermatan sulphate proteoglycan, DSPG-I, Mr 3.5 x 10(5) (46%), which was mainly located in the culture medium. (iii) A second minor small dermatan sulphate, DSPG-II, Mr approximately 2 x 10(5) (9.8%). This molecule was exclusively located in the culture medium. (iv) A large heparan sulphate proteoglycan, HSPG-I, Mr 8 x 10(5) (3.3%). (v) A second large heparan sulphate proteoglycan HSPG-II, Mr approximately 6 x 10(5) (23%). HSPG-I and HSPG-II were extracted from both the culture medium and the cell layer. 3. Western blot analysis of the core proteins released by chondroitin ABC lyase treatment of DSPG-I and DSPG-II identified these dermatan sulphate proteoglycans as biglycan and decorin respectively. Both DSPG-I and DSPG-II had core proteins of Mr 45,000. 4. The cell-layer-associated forms of CSPG-I, HSPG-I and HSPG-II were accessible to limited trypsin treatment, bound to octyl-Sepharose and could be inserted into liposomes, indicating a possible cell membrane location. 5. Pulse-chase experiments indicated that the cell-layer-associated [35S]proteoglycans undergo limited metabolism to inorganic [35S]sulphate, the majority of which is accounted for by the degradation of HSPG-II and to a lesser extent DSPG-I.

scholarly journals Inhibition of in vitro maturation of equine oocytes by interleukin 1 beta via specific IL-1 receptors

Reproduction ◽  
2003 ◽  
pp. 509-515 ◽  
Author(s):  
A Martoriati ◽  
M Caillaud ◽  
G Goudet ◽  
N Gerard
Keyword(s):  
Oocyte Maturation ◽  
In Vitro Maturation ◽  
Culture Medium ◽  
Interleukin 1 ◽  
Inhibitory Effect ◽  
Interleukin 1 Beta ◽  
Nuclear Maturation ◽  
Epidermal Growth ◽  
Dose Dependent

Interleukin 1 beta (IL-1 beta) inhibits the LH-induced resumption of meiosis of equine oocytes in vitro. The present study was performed to clarify this inhibitory effect of IL-1 beta by testing increasing concentrations of IL-1 beta, and by measuring the effect of addition of IL-1 receptor antagonist (IL-1RA) to the culture medium. The effect of IL-1 beta on epidermal growth factor (EGF)-induced resumption of meiosis was also studied. Cumulus-oocyte complexes (COCs) were collected from subordinate follicles on ovaries obtained from an abattoir. In five distinct experiments, COCs were cultured for 30 h and nuclear maturation of oocytes was evaluated by DNA staining. In Expt 1, seven different media were tested: medium 1 (TCM199+BSA); medium 2 (medium 1+50 ng IL-1 beta ml(-1)); medium 3 (medium 1+eLH); and media 4, 5, 6 and 7 (medium 3 containing 0.1, 1.0, 10.0 and 50.0 ng IL-1 beta ml(-1), respectively). In Expt 2, four different media were tested: medium 1 (TCM199+BSA+eLH); medium 2 (medium 1+50 ng IL-1 beta ml(-1)); and media 3 and 4 (medium 2+IL-1RA at 50 and 100 ng ml(-1), respectively). In Expt 3, three different media were tested: medium 1 (TCM199+BSA+eLH); medium 2 (medium 1+50 ng IL-1RA ml(-1)); and medium 3 (medium 2+50 ng IL-1 beta ml(-1)). In Expt 4, four different media were tested: medium 1 (TCM199+BSA+eLH); and media 2, 3 and 4 (medium 1+IL-1RA at 50, 100 and 150 ng ml(-1), respectively). In Expt 5, three different media were tested: medium 1 (TCM199+BSA+EGF); medium 2 (medium 1+50 ng IL-1 beta ml(-1)); and medium 3 (medium 2+50 ng IL-1RA ml(-1)). In Expt 1, LH alone induced an increase in the rate of in vitro maturation (IVM) of equine oocytes (P<0.05), whereas IL-1 beta alone did not have any effect compared with medium 1. IL-1 beta (50 ng ml(-1)) significantly inhibited the eLH-induced IVM of oocytes (P<0.05) compared with medium 3. A decrease in rate of maturation was observed from a concentration of 10 ng IL-1 beta ml(-1) onwards. In Expt 2, the presence of IL-1RA in the culture medium inhibited the effect of IL-1 beta and restored the rate of oocyte maturation (P<0.05) observed in the presence of LH alone. In Expts 3 and 4 it was demonstrated that IL-1RA alone had no positive effect on the eLH-induced rate of maturation. In Expt 5, IL-1 beta inhibited the EGF-induced resumption of meiosis (P<0.05). The addition of IL-1RA inhibited this effect and restored the rate of oocyte maturation (P<0.05) observed with EGF alone. In conclusion, the present data confirm the inhibitory effect of IL-1 beta on IVM of equine oocytes induced by eLH and demonstrate its inhibitory effect on EGF-induced oocyte maturation. The rate of maturation decreased in a dose-dependent way and the lowest rate of maturation was observed at 50 ng IL-1 beta ml(-1) (P<0.05). The use of IL-1RA inhibited these effects, demonstrating that the action of IL-1 beta is receptor-mediated. Moreover, the results clearly show that, in equine species, IL-1 beta is involved in the physiology of COCs by regulating resumption of meiosis.

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