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Biomedical subjects

E Canalis

Publications and source records attributed to E Canalis.

At least 55 records · Page 3Linked to original sources
PubMed1998

Basic fibroblast growth factor destabilizes osteonectin mRNA in osteoblasts.

Osteonectin (secreted protein acidic and rich in cysteine, 40-kDa basement membrane) is a glycoprotein abundantly expressed in bone and in other tissues undergoing active remodeling. Fibroblast growth factors (FGFs) are important in skeletal development and fracture repair, events associated with extracellular matrix remodeling. We used the murine osteoblastic cell line MC3T3 to determine whether basic FGF (bFGF) regulates osteonectin expression in bone. Northern blot analysis showed that bFGF decreased osteonectin transcripts in a dose- and time-dependent manner. This regulation was independent of the mitogenic effect of bFGF but was dependent on new protein synthesis. Immunoprecipitation of [35S]methionine-cysteine osteoblast-conditioned medium and cell layer proteins showed that bFGF decreased osteonectin synthesis. Nuclear runoff assays failed to reveal regulation of osteonectin gene transcription by bFGF. However, bFGF dramatically decreased the stability of osteonectin mRNA in transcriptionally arrested osteoblasts. This destabilization of osteonectin mRNA may be one means by which bFGF regulates extracellular matrix remodeling.

Animals↗
PubMed1998

Insulin-like growth factors sustain insulin-like growth factor-binding protein-5 expression in osteoblasts.

Insulin-like growth factors (IGFs) I and II are considered to be autocrine regulators of bone cell function. Recently, we demonstrated that IGF-I induces IGF-binding protein-5 (IGFBP-5) expression in cultures of osteoblast-enriched cells from 22-day fetal rat calvariae (Ob cells). In the present study, we postulated that IGFs play an autocrine role in the maintenance of IGFBP-5 basal expression in Ob cells. IGFBP-2 and -3, at concentrations that bind endogenous IGFs, decreased IGFBP-5 mRNA levels, as determined by Northern blot analysis, and protein levels, as determined by Western immunoblots of extracellular matrix extracts of Ob cells. IGFBP-2 and -3 in excess inhibited IGFBP-5 heterogeneous nuclear RNA levels, as determined by RT-PCR, and did not alter the half-life of IGFBP-5 mRNA in transcriptionally arrested Ob cells. In conclusion, blocking endogenous IGFs in Ob cells represses IGFBP-5 expression, suggesting that IGFs are autocrine inducers of IGFBP-5 synthesis in osteoblasts.

Animals↗
PubMed1998

Insulin-like growth factor II promoter expression in cultured rodent osteoblasts and adult rat bone.

Insulin-like growth factor (IGF)-II stimulates bone formation by increasing the replication of cells of the osteoblastic lineage and by enhancing the differentiated function of the osteoblast. Although IGF-II is synthesized by skeletal cells, little is known about the mechanisms involved and its regulation by growth factors. IGF-II expression is tissue specific and is developmentally regulated. In the present study, we examined the expression of IGF-II in fetal rat, newborn mouse and MC3T3-E1 osteoblastic (Ob) cells, and in adult rat calvariae. We also determined mechanisms involved in the regulation of IGF-II by platelet-derived growth factor (PDGF) BB, fibroblast growth factor-2 (FGF-2), and transforming growth factor (TGF) beta1. Northern analysis revealed IGF-II transcripts of 3.6 and 1.2 kb in osteoblastic cells and adult rat calvariae. Ribonuclease (RNase) protection assay using probes specific to the three known IGF-II promoters, P1, P2, and P3, demonstrated messenger RNA (mRNA) expression driven by P3 in osteoblasts and adult rat calvariae, but no expression of P1 or P2 transcripts. PDGF BB, FGF-2, and TGF beta1 inhibited the expression of IGF-II P3 mRNA by 50%. PDGF BB, FGF-2, and TGF beta1 also decreased the rates of IGF-II transcription in rat Ob cells as determined by nuclear run-on assays and did not modify the decay of IGF-II in transcriptionally arrested rat Ob cells. In conclusion, the synthesis of IGF-II in osteoblastic cells and in adult rat calvariae is driven by IGF-II P3 and is regulated by skeletal growth factors acting at the transcriptional level using the IGF-II P3.

Animals↗
PubMed1997

Insulin-like growth factor I inhibits the transcription of collagenase 3 in osteoblast cultures.

Insulin-like growth factor (IGF) I is an autocrine regulator of bone remodeling which inhibits bone collagen degradation and interstitial collagenase 3 mRNA levels. The mechanism of this inhibitory effect on collagenase 3 expression is not known. We tested the effects of IGF I on collagenase 3 gene expression in cultures of osteoblast-enriched cells from 22 day fetal rat calvariae (Ob cells) to determine whether transcriptional or posttranscriptional mechanisms were involved in the regulation of the collagenase 3 gene. IGF I at 10-100 nM caused a dose-dependent decrease in collagenase mRNA and protein levels. IGF I did not modify the half-life of collagenase 3 mRNA in transcriptionally arrested Ob cells, whereas it decreased the levels of interstitial collagenase 3 heterogeneous nuclear RNA. In addition, IGF I decreased the rates of transcription of the collagenase gene and the activity of a 2.1 kilobase collagenase 3 promoter construct transiently transfected into Ob cells. In conclusion, IGF I decreases the expression of collagenase 3 mRNA by transcriptional mechanisms.

Animals↗
PubMed1997

Interleukin 6 is autoregulated by transcriptional mechanisms in cultures of rat osteoblastic cells.

Interleukin 6 (IL-6), a cytokine produced by skeletal cells, stimulates osteoclast recruitment. The IL-6 soluble receptor (sIL-6R) increases IL-6 activity, and IL-6 and sIL-6R levels are increased in conditions of increased bone resorption. We examined the production of IL-6 by primary rat osteoblasts (Ob cells) cultured in the presence of IL-6 and sIL-6R. IL-6 alone did not induce IL-6 transcripts, but IL-6 was stimulatory in the presence of sIL-6R. Furthermore, sIL-6R by itself increased IL-6 transcripts. Cycloheximide superinduced IL-6 transcripts and did not prevent the effect of IL-6 and sIL-6R. IL-6 in the presence of sIL-6R stimulated IL-6 rates of transcription and the activity of IL-6 promoter fragments in transiently transfected Ob cells. 5' deletions of the IL-6 promoter and targeted mutations of the multiple response element (MRE)/cAMP responsive element (CRE), the nuclear factor for IL-6 (NF-IL-6), and the nuclear factor-kappaB (NF-kappaB) binding sites indicated that NF-IL-6 and NF-kappaB, in combination with MRE/CRE, binding sites are required for the induction of the IL-6 promoter by IL-6. In conclusion, IL-6 induces its own synthesis in osteoblasts by transcriptional mechanisms. This positive feedback may be important in conditions of increased bone resorption.

Animals↗
PubMed1997

Growth factor regulation of insulin-like growth factor binding protein-6 expression in osteoblasts.

Previously we have shown that transforming growth factor beta (TGF beta) 1, basic fibroblast growth factor (FGF), and platelet-derived growth factor (PDGF) BB inhibit the synthesis of insulin-like growth factor (IGF) II, but their effects on IGF binding protein (IGFBP)-6 in osteoblast cultures are not known. IGFBP-6 binds IGF II with high affinity and prevents IGF II-mediated effects, so that a possible mode of regulating the IGF II available to bone cells would be by changing the levels of IGFBP-6. To enhance our understanding of the actions of growth factors on the IGF II axis in bone, we tested the effects of TGF beta 1, basic FGF, PDGF BB, IGF I, and IGF II on the expression of IGFBP-6 in cultures of osteoblast-enriched cells from 22 day fetal rat calvariae (Ob cells). Treatment of Ob cells with TGF beta 1 caused a time- and dose-dependent decrease in IGFBP-6 mRNA levels, as determined by Northern blot analysis. The effect was maximal after 48 h and observed with TGF beta 1 concentrations of 0.04 nM and higher. TGF beta 1 also decreased IGFBP-6 polypeptide levels in the medium, as determined by Western immunoblot analysis. Cycloheximide at 3.6 microM decreased IGFBP-6 transcripts and prevented the effect of TGF beta 1. The decay of IGFBP-6 mRNA in transcriptionally arrested Ob cells was not modified by TGF beta 1. In addition, TGF beta 1 decreased the rates of IGFBP-6 transcription as determined by a nuclear run-on assay. In contrast, basic FGF, PDGF BB, IGF I, and IGF II did not change IGFBP-6 mRNA levels in Ob cells. In conclusion, TGF beta 1 inhibits IGFBP-6 expression in Ob cells by transcriptional mechanisms. Since IGFBP-6 binds IGF II and prevents its effects on bone cells, decreased synthesis of IGFBP-6 induced by TGF beta 1 could be a local feedback mechanism to increase the amount of IGF II available in the bone microenvironment.

Animals↗
PubMed1997

Interleukin-6 and its soluble receptor cause a marked induction of collagenase 3 expression in rat osteoblast cultures.

Interleukin-6 (IL-6), a cytokine produced by skeletal cells, increases bone resorption, but its effects on collagenase expression are unknown. We tested the effects of IL-6 and its soluble receptor on collagenase 3 expression in osteoblast-enriched cells from fetal rat calvariae (Ob cells). IL-6 caused a small increase in collagenase mRNA levels, but in the presence of IL-6-soluble receptor (IL-6sR), IL-6 caused a marked increase in collagenase transcripts after 2-24 h. In addition, IL-6sR increased collagenase mRNA when tested alone. IL-6 and IL-6sR increased immunoreactive collagenase levels. Cycloheximide and indomethacin did not prevent the effect of IL-6 and IL-6sR on collagenase mRNA levels. IL-6 and IL-6sR did not alter the decay of collagenase mRNA in transcriptionally arrested Ob cells and increased the levels of collagenase heterogeneous nuclear RNA and the rate of collagenase gene transcription in Ob cells. IL-6 and IL-6sR increased collagenase 3 mRNA in MC3T3 cells but only modestly in skin fibroblasts. IL-6 and IL-6sR enhanced the expression of tissue inhibitor of metalloproteinases 1. In conclusion, IL-6, in the presence of IL-6sR, increases collagenase 3 synthesis in osteoblasts by transcriptional mechanisms. This effect may contribute to the action of IL-6 on bone matrix degradation and bone resorption.

Animals↗
PubMed1997

Transforming growth factor beta1 inhibits collagenase 3 expression by transcriptional and post-transcriptional mechanisms in osteoblast cultures.

Transforming growth factor (TGF) beta1 is an autocrine regulator of bone cell function. We demonstrated that TGF beta1 enhances bone collagen synthesis, but its effects on collagen degradation are not well characterized. We tested the effects of TGF beta1 on rat collagenase 3 expression in cultures of osteoblast-enriched cells from fetal rat calvariae (Ob cells). Treatment with TGF beta1 at 0.4 nM decreased steady state collagenase mRNA levels after 2 to 24 h. This dose-dependent effect was observed at TGF beta1 concentrations of 4 pM to 1.2 nM, and was accompanied by decreased levels of immunoreactive procollagenase. The protein synthesis inhibitor cycloheximide increased collagenase transcripts, but did not prevent the effect of TGF beta1 on collagenase mRNA levels. TGF beta1 accelerated the decay of collagenase mRNA in transcriptionally arrested Ob cells. In addition, TGF beta1 decreased the levels of collagenase heterogeneous nuclear RNA and the rate of collagenase gene transcription in Ob cells. TGF beta1 enhanced the expression of tissue inhibitors of metalloproteinases (TIMP) 1 and 3 and caused a modest decrease of TIMP 2 mRNA levels. In conclusion, TGF beta1 decreases interstitial collagenase transcripts and protease levels in Ob cells by transcriptional and post-transcriptional mechanisms, and this effect may contribute to its actions on bone matrix.

Animals↗
PubMed1997

Retinoic acid regulates the expression of insulin-like growth factors I and II in osteoblasts.

Insulin-like growth factors (IGF) I and II are the most abundant growth factors secreted by skeletal cells, and retinoic acid has many important action on cell differentiation and osteoblastic function. Some of these actions may be mediated by changes in the expression of IGF I and II since IGFs are known to enhance the differentiated function of the osteoblast. We examined the effects of all-transretinoic acid on IGF I and IGF II expression in cultures of osteoblast-enriched cells from 22 day fetal rat calvariae (Ob cells). Retinoic acid caused a transient increase in IGF I and IGF II mRNA levels after 6 h, but after 24 and 48 h of treatment a dose-dependent decrease was observed. Cycloheximide prevented the inhibitory effect of retinoic acid. Retinoic acid treatment for 48 h decreased IGF I polypeptide levels in the culture medium. In contrast, 48 h exposure to retinoic acid increased IGF II polypeptide levels, possible due to increased levels of IGF binding protein-6. The decay of IGF I and II mRNA in transcriptionally arrested Ob cells was similar in control and retinoic acid-treated cells. After 2 h, retinoic acid increased the rates of IGF I and II transcription, as determined by a nuclear run-on assay and heterogeneous nuclear RNA levels, but after 24 h retinoic acid was inhibitory. Retinoic acid had opposite effects to IGFs in osteoblasts and inhibited DNA and collagen synthesis. In conclusion, following a small transient increase, retinoic acid causes a pronounced decrease in IGF I and IGF II mRNA expression in Ob cells. However, treatment with retinoic acid causes a decrease in IGF I and an increase in IGF II polypeptide levels. These changes in the IGF/IGFBP axis may be relevant to the mechanism of action of retinoic acid in bone.

Animals↗
PubMed1997

Surgical treatment of primary spontaneous pneumothorax with video-assisted thoracic surgery.

We report the results of our experience using video-assisted thoracoscopic surgery (VATS) to treat primary spontaneous pneumothorax (PSP) from January 1992 until December 1994 in a multicentered co-operative study. A total of 132 patients (110 males and 22 females, aged 13-38 yrs, mean age 26 yrs) were treated by VATS to deal with the PSP that they presented with. A standard VATS technique was used. Apical bullae were always removed, and mechanical pleural abrasion was performed, leaving a pleural drainage tube. In two cases (1.5%), a switch to thoracotomy was necessary. In eight cases (6%), air leakage persisted for 5 days after surgery, which resolved with pleural drainage. There were eight postoperative relapses (6%), which were treated with pleural drainage (n = 4), VATS (n = 3) or axillar thoracotomy (n = 1). The average postoperative stay was 5.6 days (range 2-15 days). We conclude that video-assisted thoracoscopic surgery is a viable alternative for the treatment of primary spontaneous pneumothorax. There is, however, a high relapse rate, and in a number of cases air leakage persists in the postoperative period.

Adolescent↗
PubMed1997

Interleukin-6 with its soluble receptor enhances the expression of insulin-like growth factor-I in osteoblasts.

Interleukin (IL)-6, a cytokine produced by skeletal cells and known to increase bone resorption, has mitogenic effects for bone cells, possibly by regulating the synthesis of other local factors. We tested the effects of IL-6 and its soluble receptor (IL-6sR) on the expression of insulin-like growth factor (IGF)-I and IGF-II in cultured osteoblast-enriched cells from fetal rat calvariae (Ob cells). IL-6 did not modify IGF-I messenger RNA (mRNA) levels, but when tested in the presence of IL-6sR, IL-6 at 1 to 100 ng/ml increased IGF-I transcripts by up to 3.2-fold after 24 h. IL-6sR caused a small increase in IGF-I mRNA levels when tested alone. IL-6 and IL-6sR increased immunoreactive IGF-I levels by 2.4-fold after 24 h and 6.4-fold after 48 h. Cycloheximide prevented, and indomethacin markedly decreased, the effect of IL-6 and IL-6sR on IGF-I mRNA levels, but hydroxyurea did not. IL-6 and IL-6sR did not alter the decay of IGF-I mRNA in transcriptionally arrested Ob cells, and the half-life of the predominant 6.5-kb IGF-I transcript was about 11 h in control and treated cells. In addition, IL-6 and IL-6sR increased the levels of IGF-I heterogeneous nuclear RNA. IL-11 also increased IGF-I mRNA levels, whereas oncostatin M and leukemia-inhibitory factor did not. In contrast to their effects on IGF-I, IL-6 and IL-6sR caused only a modest increase in IGF-II mRNA and polypeptide levels. In conclusion, IL-6, in the presence of IL-6sR, increases IGF-I synthesis in Ob cells; this effect may lead to a secondary increase in bone formation.

Animals↗
PubMed1997

Regulation of collagenase-3 by bone morphogenetic protein-2 in bone cell cultures.

Bone morphogenetic protein-2 (BMP-2), a member of the transforming growth factor superfamily of peptides, induces ectopic bone formation in vivo. The actions of BMP-2 on osteoblastic cells include stimulation of collagen synthesis, but the role of BMP-2 on collagen degradation is not known. We examined whether BMP-2 affects the expression of collagenase-3, an enzyme that degrades type I collagen at neutral pH, and that of tissue inhibitors of matrix metalloproteinases (TIMPs) in primary osteoblast-enriched cells from 22-day-old fetal rat calvariae. BMP-2 suppressed collagenase messenger RNA (mRNA) and immunoreactive protein levels. BMP-2 did not affect collagenase mRNA stability, but it reduced collagenase heterogeneous nuclear RNA levels and decreased the rate of transcription of the collagenase gene. BMP-2 also stimulated TIMP 1 and TIMP 3 mRNA levels, but failed to alter TIMP 2 expression. In conclusion, our studies indicate that BMP-2 suppresses collagenase-3 gene transcription and stimulates TIMP 1 and TIMP 3 expression in osteoblasts. The regulation of collagenase and TIMPs by BMP-2 in osteoblasts may play a role in osteoinduction.

Animals↗
PubMed1997

Interleukin-6 and its soluble receptor regulate the expression of insulin-like growth factor binding protein-5 in osteoblast cultures.

Interleukin-6 (IL-6), a cytokine produced by bone cells, is known to influence bone resorption by stimulating the development of osteoclasts from precursor cells and to have mitogenic actions on osteoblastic cells. Insulin-like growth factors (IGFs) are important local regulators of bone formation, and IGF binding protein (IGFBP)-5 stimulates bone cell growth and enhances the effects of IGF-I. We tested the effects of IL-6 in the presence and absence of its soluble receptor (sIL-6R) on IGFBP-5 expression in cultures of osteoblast-enriched cells from 22-day-old fetal rat calvariae (Ob cells). When tested individually, IL-6 and sIL-6R had a modest stimulatory effect on IGFBP-5 messenger RNA (mRNA) levels. In contrast, when IL-6 and sIL-6R were tested in combination, they caused a considerable increase in IGFBP-5 mRNA levels, and IL-6 at 100 ng/ml and sIL-6R at 125 ng/ml increased IGFBP-5 transcripts by 5- to 7-fold after 24 h. The effect of IL-6 and sIL-6R on IGFBP-5 transcripts was not blocked by indomethacin, but cycloheximide markedly inhibited IGFBP-5 mRNA levels in control and treated cultures. IL-6 and sIL-6R did not modify the decay of IGFBP-5 mRNA in transcriptionally arrested Ob cells, and stimulated the rate of IGFBP-5 transcription as demonstrated by a nuclear run-on assay. IL-6 and sIL-6R did not increase intact IGFBP-5 levels in the extracellular matrix and increased IGFBP-5 fragments in the culture medium. Conditioned medium from Ob cells induced the proteolytic fragmentation of an IGFBP-5 standard, an effect that was accelerated and enhanced by conditioned medium from IL-6/sIL-6R-treated cultures and prevented by metalloprotease inhibitors. In conclusion, IL-6, in the presence of sIL-6R, stimulates IGFBP-5 mRNA expression in Ob cells by transcriptional mechanisms, and accelerates the fragmentation of the protein.

Animals↗
PubMed1996

Cortisol inhibits the synthesis of insulin-like growth factor-binding protein-5 in bone cell cultures by transcriptional mechanisms.

Glucocorticoids inhibit the synthesis of insulin-like growth factor-binding protein-5 (IGFBP-5) in osteoblasts, but the mechanisms involved are unknown. IGFBP-5 stimulates bone cell growth, and its inhibition by glucocorticoids may be relevant to the action of this binding protein on bone formation. We tested the effects of cortisol on IGFBP-5 expression in cultures of osteoblast-enriched cells from fetal rat calvariae (Ob cells). Cortisol decreased IGFBP-5 polypeptide levels in the extracellular matrix and caused a time- and dose-dependent decrease in IGFBP-5 mRNA. IGFBP-5 transcripts were markedly decreased by cycloheximide, and further suppressive effects of cortisol could not be determined. Cortisol did not modify the decay of IGFBP-5 mRNA in transcriptionally arrested Ob cells. Cortisol decreased IGFBP-5 hnRNA, the rate of IGFBP-5 transcription, and the activity of the murine IGFBP-5 promoter by 35% in transient transfection experiments. Deletion analysis showed that the region responsive to cortisol is from base pairs -70 to +22, and E-box-binding proteins or c-Myb-related nuclear factors may be involved in its regulation. In conclusion, cortisol inhibits IGFBP-5 transcription in Ob cells through the Myb-binding domain. This effect may be partly responsible for the effect of glucocorticoids on bone formation.

Animals↗
PubMed1996

Retinoic acid stimulates the transcription of insulin-like growth factor binding protein-6 in skeletal cells.

Retinoic acid has important actions on cell differentiation and osteoblastic function, and some of these actions may be mediated by changes in the insulin-like growth factor (IGF) axis. Skeletal cells synthesize IGF I and II and the six known IGF binding proteins (IGFBP). IGFBP-6 binds IGF II with high affinity and prevents IGF II-mediated effects. In fibroblasts, IGFBP-6 levels are regulated by retinoic acid, and we postulated that retinoic acid may regulate IGF II in bone by altering IGFBP-6 synthesis. We examined the effect of retinoic acid on IGFBP-6 expression in cultures of osteoblast-enriched cells from 22-day fetal rat calvariae (Ob cells). Retinoic acid caused a time- and dose-dependent increase in IGFBP-6 mRNA levels, as determined by Northern blot analysis. The effect was maximal after 48 h of treatment and observed with retinoic acid at concentrations of 10 nM to 1 microM. Retinoic acid increased IGFBP-6 polypeptide levels in the culture medium, as determined by Western immunoblot analysis. Cycloheximide at 3.6 microM slightly decreased IGFBP-6 transcripts but did not prevent the stimulatory effect of retinoic acid. The decay of IGFBP-6 mRNA in transcriptionally arrested Ob cells was similar in control and retinoic acid-treated cells, and retinoic acid increased the rates of IGFBP-6 transcription, as determined by nuclear run on assays. In conclusion, retinoic acid enhances IGFBP-6 expression in Ob cells by transcriptional mechanisms. Since IGFBP-6 prevents the effects of IGF II, increased synthesis of IGFBP-6 could mediate selected actions of retinoic acid in bone.

Animals↗