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Biomedical subjects

J Pfeilschifter

Publications and source records attributed to J Pfeilschifter.

At least 271 records · Page 15Linked to original sources
PubMed1988

Transforming growth factor beta inhibits bone resorption in fetal rat long bone cultures.

TGF-beta 1 is a polypeptide that is abundant in bone matrix, is produced by bone cells, and modulates proliferation and differentiated functions of osteoblastic cells in vitro. TGF-beta 2 is a closely related polypeptide that was originally isolated from bone matrix. TGF-beta 1 has been shown previously to stimulate prostaglandin production in cultures of neonatal mouse calvariae, which causes these bones to resorb. We found similar effects with TGF-beta 2. In comparison, TGF-beta 1 and TGF-beta 2 failed to stimulate bone resorption in fetal rat long bone cultures during a 3-d incubation period in concentrations up to 50-100 times greater than those capable of inducing bone resorption in calvariae. Incubation with TGF-beta 1 for a further 3 d decreased bone resorption up to 30%. Moreover, bone resorption induced by the bone-resorbing agents IL 1 and 1,25-dihydroxyvitamin D3 was partially or completely inhibited by TGF-beta 1 and TGF-beta 2 during the second half of the 6-d incubation period. Inhibition of DNA synthesis with hydroxyurea inhibited bone resorption in long bones in a similar pattern to that seen with TGF-beta 1. The inhibitory effects of TGF-beta 1 and TGF-beta 2 on bone resorption in long bone cultures may therefore be due to inhibition of osteoclast precursor proliferation.

Animals↗
PubMed1987

Cyclosporin A augments angiotensin II-stimulated rise in intracellular free calcium in vascular smooth muscle cells.

Pretreatment of rat vascular smooth muscle cells with the immunosuppressive drug cyclosporin A caused concentration- and time-dependent increases in both the amplitude and duration of the angiotensin II-induced rise in cytosolic free calcium, as measured with quin 2. Cyclosporin A had no significant effect on basal quin 2 fluorescence. However, cyclosporin A increased the basal 45Ca2+ influx. This stimulation of 45Ca2+ influx was not blocked by nifedipine (10(-6) M). Cyclosporin A also augmented the angiotensin II-stimulated influx and efflux of 45Ca2+. These results demonstrate that cyclosporin A increases the permeability of the plasma membrane for Ca2+ and also augments the angiotensin II-induced increases in cytosolic free calcium.

Angiotensin II↗
PubMed1987

Different effects of phorbol ester on angiotensin II- and stable GTP analogue-induced activation of polyphosphoinositide phosphodiesterase in membranes isolated from rat renal mesangial cells.

Pretreatment with pertussis toxin inhibits angiotensin II-induced activation of polyphosphoinositide phosphodiesterase in rat renal mesangial cells [Pfeilschifter & Bauer (1986) Biochem. J. 236, 289-294]. Furthermore, activation of protein kinase C by the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) and by 1-oleoyl-2-acetylglycerol (OAG) abolishes angiotensin II-induced formation of inositol trisphosphate (IP3) in mesangial cells [Pfeilschifter (1986) FEBS Lett. 203, 262-266]. Using membrane preparations of [3H]inositol-labelled mesangial cells we tried to obtain further insight as to the step at which protein kinase C might interfere with the signal transduction mechanism in mesangial cells. Angiotensin II (100 nM) stimulates IP3 formation from membrane preparations of [3H]inositol-labelled mesangial cells with a half-maximal potency of 1.1 nM. The angiotensin II-induced formation of IP3 is enhanced by GTP. This effect of angiotensin II is completely blocked by the competitive antagonist [Sar1,Ala8]angiotensin II. Guanosine 5'-[gamma-thio]triphosphate (GTP gamma S) and guanosine 5'-[beta gamma-imido]triphosphate (Gpp[NH]p), non-hydrolysable analogues of GTP, stimulate IP3 production in the absence of angiotensin II with Kd values of 0.19 microM and 2.4 microM, respectively. Angiotensin II augments the increase in IP3 formation induced by GTP gamma S. However, when mesangial cells were pretreated with TPA there was a dose-dependent inhibition of the synergistic action of angiotensin II on GTP gamma S-induced IP3 production. Comparable results are obtained with OAG, while the non-tumour-promoting phorbol ester 4 alpha-phorbol 12,13-didecanoate is without effect. These results suggest that activation of protein kinase C in mesangial cells does not impair phosphoinositide hydrolysis by stable GTP analogues but somehow seems to interfere with the stimulatory interaction of the occupied angiotensin II receptor with the transducing G-protein.

Angiotensin II↗
PubMed1987

Cyclosporin A inhibits PGE2 release from vascular smooth muscle cells.

The influence of the fungoid undecapeptide cyclosporin A (CyA) on PGE2 release from cultured rat aortic smooth muscle cells was investigated in this study. We found that CyA time and concentration dependently (ED50:500 ng/ml) inhibited PGE2 release from the cells. CyA attenuated both basal and PGE2 release evoked by angiotensin II (10(-10)-10(-6) M), arginine vasopressin (10(-10)-10(-6) M) and ionomycin (10(-9)-10(-6) M). CyA (1 microgram/ml) did not affect the conversion of exogenous arachidonic acid (1 microM) into PGE2. The inhibitory effect of CyA was neutralized by high concentrations of the calcium ionophore ionomycin (greater than 3 X 10(-6) M). Taken together our results indicate that CyA inhibits both basal and vasoconstrictor evoked PGE2 release from vascular smooth muscle by impairing the availability of free arachidonic acid rather than by inhibiting the conversion of arachidonic acid into PGE2.

Angiotensin II↗
PubMed1987

Inflammation-mediated osteopenia (IMO) during acute inflammation in rats is due to a transient inhibition of bone formation.

Local inflammation was induced in rats through the subcutaneous injection of magnesium silicate. Trabecular bone volume of the tibia decreased progressively during a 3 week observation period following the inflammatory stimulus. The trabecular bone surface covered with osteoblasts was strikingly reduced during the first week but had normalized by the end of the third week. Calcification rate in the cortical bone of the tibia was reduced with a parallel reduction in endosteal osteoid seam width. Both calcification rate and tetracycline double-labeled surface of vertebral trabecular bone were reduced during the first 2 weeks. Neither total bone resorption surface nor active bone resorption surface were increased. There was a decrease in osteoclast numbers/mm2 bone tissue associated with decreasing bone volume. Our data demonstrate a transient inhibition of bone formation during acute inflammation in the rat and indicate that changes in osteoblast function are part of the acute phase response following local inflammation.

Acute-Phase Reaction↗
PubMed1987

Modulation of type beta transforming growth factor activity in bone cultures by osteotropic hormones.

Type beta transforming growth factor (TGF-beta) activity was found in conditioned medium harvested from fetal rat and neonatal mouse calvariae by using the anchorage-independent growth of normal rat kidney fibroblasts as an indicator system. Calvariae incubated with parathyroid hormone, 1,25-dihydroxyvitamin D3, and interleukin 1, all factors that stimulate bone resorption showed a concentration-dependent increase in TGF-beta activity in the culture medium. Increases in TGF-beta activity persisted undiminished for 48 hr after removal of these factors. The increases in TGF-beta activity from the resorbing bone cultures were relatively greater than the increases in bone resorption. Calcitonin inhibition of bone resorption correlated with a decrease in TGF-beta activity. Thus, agents that modulate bone resorption also affect TGF-beta activity in the bone culture medium. Changes in local concentrations of TGF-beta activity by osteotropic hormones may be important in the regulation of normal bone remodeling.

Animals↗
PubMed1987

Mechanism of NaCl transport-stimulated prostaglandin formation in MDCK cells.

Recently we have found that stimulation of NaCl transport in high-resistance MDCK cells enhances their prostaglandin formation. In the present study, we investigated the mechanisms by which prostaglandin formation could be linked to the ion transport in these cells. We found that stimulation of transport caused a transient stimulation of prostaglandin formation lasting 5-10 min. The rise in prostaglandin formation was paralleled by a rise of free intracellular arachidonic acid. Analysis of membrane lipids revealed that the rise of free arachidonic acid was paralleled by a loss of arachidonic acid from polyphosphoinositides. We failed to obtain indications for the stimulation of calcium-dependent phospholipase A2. However, we did obtain evidence that the incorporation of arachidonic acid into phospholipids was diminished during stimulation of ion transport, indicating a decreased rate of reesterification. Despite the fact that there was no significant fall in total cellular ATP on stimulation of ion transport, we found a high and transient rise of lactate production of the cells on stimulation of the ion transport indicating an alteration of the ADP/ATP ratio. Moreover, prostaglandin formation and lactate formation were linearly correlated in this situation. When glucose utilization was inhibited by mannoheptulose, the rise in lactate formation was abolished, whereas that of PG formation was unaltered, indicating that lactate formation and prostaglandin formation were not causally linked on stimulation of ion transport. Our results suggest that an increase in the rate of sodium chloride transport by MDCK cells stimulates formation by an inhibition of reesterification of free arachidonic acid.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenosine Triphosphate↗
PubMed1987

Effects of transforming growth factor-beta on osteoblastic osteosarcoma cells.

Transforming growth factor-beta (TGF beta), a polypeptide that controls growth and differentiation in many cell types and has recently been found in abundant amounts in bone, was examined for its effects on cells with the osteoblast phenotype using the clonal osteoblastic osteosarcoma cell line ROS 17/2.8. TGF beta increased alkaline phosphatase (AP) activity and the rate of collagen synthesis per cell. Cell proliferation was inhibited, and the morphological appearance of the cells was markedly changed. All effects were observed at concentrations as low as 0.1 ng/ml TGF beta. Increases in AP activity were detectable after 24 h and increased progressively with time. TGF beta increased AP activity under serum-free conditions and during thymidine-induced inhibition of DNA synthesis. The increase in AP activity mediated by TGF beta could be completely inhibited with actinomycin D and cycloheximide. 1,25-Dihydroxyvitamin D3 at 10(-7) M slightly increased AP activity in ROS 17/2.8 cells, but strongly inhibited AP activity when the cells were pretreated with TGF beta. The data suggest that TGF beta stimulates expression of the osteoblastic phenotype in ROS 17/2.8 cells and that TGF beta may be an important regulator of local bone remodeling.

Alkaline Phosphatase↗
PubMed1986

Tumour promotor 12-O-tetradecanoylphorbol 13-acetate inhibits angiotensin II-induced inositol phosphate production and cytosolic Ca2+ rise in rat renal mesangial cells.

Preincubation of rat renal mesangial cells with 12-O-tetradecanoylphorbol 13-acetate (TPA) strongly inhibited the increases of inositol phosphates and of free cytosolic Ca2+ induced by angiotensin II (10(-7) M). TPA had no significant effect on the basal values of inositol phosphates and of free cytosolic Ca2+. Inhibition appeared already after 1 min and was maximal after 5 min. These effects occur without significant changes on angiotensin II binding in intact cells. The concentration of TPA needed (10(-9)-10(-7) M) was in the range believed to cause specifically an activation of protein kinase C. Furthermore the biologically inactive phorbol ester 4 alpha-phorbol 12,13-didecanoate was without effect. From the entirety of these results it is likely that protein kinase C inhibits angiotensin II activation of phospholipase C at a stage distal to receptor occupancy.

Angiotensin II↗
PubMed1986

Pertussis toxin abolishes angiotensin II-induced phosphoinositide hydrolysis and prostaglandin synthesis in rat renal mesangial cells.

Incubation of rat renal mesangial cells with angiotensin II (0.1 microM) resulted in transient breakdown of phosphatidylinositol 4,5-bisphosphate, rapid generation of diacylglycerol and phosphatidic acid, increased 45Ca2+ influx, increased intracellular [Ca2+] as measured by quin 2, and increased prostaglandin E2 synthesis. All of these processes were markedly inhibited time- and dose-dependently by prior exposure of cells to pertussis toxin. In contrast, the effects of the ionophore A23187 on 45Ca2+ influx and prostaglandin E2 synthesis were not altered by the exposure of the cells to pertussis toxin. The action of the toxin was not associated with alterations in cellular concentrations of cyclic AMP. Incubation of membrane fraction of mesangial cells with pertussis toxin resulted in ADP-ribosylation of Mr-42,000 protein. From all these results, it is likely that a G protein is involved in receptor-mediated signal transduction in renal mesangial cells.

Angiotensin II↗
PubMed1986

Role of phospholipase C and protein kinase C in vasoconstrictor-induced prostaglandin synthesis in cultured rat renal mesangial cells.

It was the aim of the present study to find out if a common mechanism exists by which the vasoconstrictive hormones angiotension II, noradrenaline and 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine (AGEPC) increase prostaglandin E2 (PGE2) synthesis in cultures of rat renal mesangial cells. Angiotension II, noradrenaline and AGEPC stimulated PGE2 synthesis and uptake of 45Ca2+ in cultured mesangial cells. Both of these effects could be completely suppressed by the calcium channel blocker verapamil. Angiotensin II, noradrenaline and AGEPC caused a rapid breakdown of phosphatidylinositol 4,5-bisphosphate with a concomitant increase of 1,2-diacylglycerol and inositol trisphosphate, indicating an activation of phospholipase C by these hormones. Addition of verapamil had no effect on the hormone-induced stimulation of phospholipase C. The synthetic analogue of diacylglycerol, 1-oleoyl-2-acetylglycerol, and the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA), both of which are known to stimulate protein kinase C, enhanced PGE2 synthesis. Chelation of extracellular calcium with EDTA or addition of verapamil abolished the effect of 1-oleoyl-2-acetylglycerol and phorbol ester on PGE2 synthesis. 1-Oleoyl-2-acetylglycerol and phorbol ester increased the uptake of 45Ca2+ by the cells in a dose-dependent manner and this effect could be blocked by verapamil. The entirety of these data leads us to suggest that vasoconstrictor-evoked synthesis of PGE2 in rat mesangial cells is mediated by the subsequent activation of phospholipase C and protein kinase C. The activation of protein kinase C by diacylglycerol is likely to be involved in the increase of the calcium permeability of the plasma membrane which is a prerequisite for PGE2 synthesis induced by vasoconstrictive hormones.

Animals↗
PubMed1986

Atrial natriuretic peptide inhibits renin release from juxtaglomerular cells by a cGMP-mediated process.

We have examined the effect of a synthetic analogue of human alpha-atrial natriuretic peptide (ANP), APII, on renin release in cultured renal juxtaglomerular cells (JGA cells). Using cell cultures containing 80-90% renal juxtaglomerular cells, we found that ANP (10(-13)-10(-9) M) strongly inhibited renin release from the cells in a dose-dependent fashion (ki, 10 pM) to about 10% of control. Inhibition of renin release by ANP was paralleled by an increase in cellular cGMP levels; while in the presence of the cGMP-phosphodiesterase inhibitor M&B 22948 (1 mM), concentrations of ANP lower by a factor of 100 were required to obtain the same effects on renin release and cGMP levels. The guanylate cyclase inhibitor methylene blue (10 microM), on the other hand, shifted the dose-response curves for renin release and cGMP levels to 100-fold higher concentrations of ANP. Neither the influx of 45Ca into the cells nor the intracellular quin-2 signal, which is a measure for changes of intracellular Ca concentration, was in any way altered by ANP. Our results suggest that ANP inhibits renin release from juxtaglomerular cells by a cGMP-dependent process that does not involve changes in intracellular calcium.

Aminoquinolines↗
PubMed1986

Role of protein kinase C in inhibition of renin release caused by vasoconstrictors.

It was the aim of the present study to get insight into some of the intracellular mechanisms by which the vasoconstrictor hormones angiotensin II (ANG II), arginine vasopressin (AVP), and norepinephrine (NE) inhibit renin release from renal juxtaglomerular cells. To this end a primary cell culture from rat renal cortex was established that consisted of 50% juxtaglomerular cells. The cultured juxtaglomerular cells contained prominent renin granules closely resembling those in the intact kidney and responded to a number of stimuli of renin release. By using these cultures, we found that ANG II (10(-7) M), AVP (10(-6) M), and NE (10(-5) M) inhibited renin release and increased the calcium permeability of the plasma membrane of the cultured cells. Both the effects on renin release and on calcium permeability could be diminished or even be abolished by the calcium channel blocker verapamil (Vp) (10(-5) M). ANG II, AVP, and NE led to an increased formation of diacylglycerol (DAG), a well-known stimulator of protein kinase C (PKC). Moreover, a direct stimulation of PKC by 12-O-tetradecanoylphorbol-13-acetate (TPA) (10(-8)-10(-6) M) also inhibited renin release and increased the calcium permeability of the cell membrane. Similar to ANG II, AVP, and NE, the effects of TPA on calcium permeability and renin release could be diminished by Vp. In conclusion, these results point toward a common mechanism by which vasoconstrictors inhibit renin release from renal juxtaglomerular cells: ANG II, AVP, and NE activate a phospholipase C, which generates DAG.(ABSTRACT TRUNCATED AT 250 WORDS)

Angiotensin II↗
PubMed1986

NaCl transport stimulates prostaglandin release in cultured renal epithelial (MDCK) cells.

Prostaglandins (PGs) can modulate a variety of renal functions, including Na+ and Cl- reabsorption. However, it is not known if a direct interdependence exists between PG synthesis and transport activity. The present study was done to find out whether or not the rate of NaCl transport has an influence on PG synthesis in renal tubular cells. For our studies we used cultures of so-called high-resistance MDCK cells, which were originally derived from canine kidney. This cell type has a loop diuretic- and ouabain-sensitive NaCl transport that can be enhanced by activation of the adenylate cyclase (AC). In MDCK cell cultures we found that each state of increased NaCl transport during stimulation of AC by either epinephrine (10(-6) M), isoprenaline (10(-5) M), or forskolin (10(-5) M) was accompanied by a twofold increase in PG release. During inhibition of NaCl transport by furosemide (10(-4) M) or ouabain (2 X 10(-4) M), stimulation of AC failed to increase PGE2 release, whereas basal PG production was not inhibited by either furosemide or ouabain. Furthermore, PG formation during activation of AC was dependent on the concentration of extracellular Na+, whereas PG formation in the absence of activators of AC was independent of extracellular Na+. These results suggest that increased NaCl transport stimulates PG formation in cultures of high-resistance MDCK cells.

Adenylyl Cyclases↗
PubMed1986

Effect of synthetic atrial natriuretic peptide on rat renal juxtaglomerular cells.

We examined the effect of synthetic atrial natriuretic peptide (ANP) on unstimulated renin release from rat renal juxtaglomerular cells. Using renal cortical cell cultures containing 80-90% juxtaglomerular cells, we found that ANP strongly inhibited renin release from the cells in a dose-dependent fashion. Half-maximal inhibition was observed at 10(-11) mol/l ANP. Furthermore, ANP produced a rise in the intracellular concentration of cyclic guanosine monophosphate (cGMP) and a fall in the concentration of cyclic adenosine monophosphate (cAMP). Data indicated that the inhibition of renin release by ANP was related to the rise in cGMP and not to the fall of cAMP. Atrial natriuretic peptide (10(-10) mol/l) had no influence on the transmembrane calcium influx nor did it alter the intracellular calcium concentration of the juxtaglomerular cell. We found, however, that the inhibitory effect of ANP on renin release could be attenuated by the calcium channel blocker verapamil. Our results suggest that ANP inhibits renin release from juxtaglomerular cells by a mechanism that is mediated by cGMP; the mechanism is not linked to any alteration in intracellular calcium concentration but requires a normal level of calcium.

Animals↗
PubMed1985

Inhibition of renin secretion by platelet activating factor (acetylglyceryl ether phosphorylcholine) in cultured rat renal juxtaglomerular cells.

Acetylglyceryl ether phosphorylcholine (AGEPC), commonly known as platelet activating factor, was found to strongly inhibit renin secretion in cultures rich in juxtaglomerular cells. This inhibitory action of AGEPC was accompanied by an enhanced calcium permeability of the cell membrane as evaluated from measurements of the uptake of 45Ca. Simultaneous addition of the calcium channel blocker verapamil abolished the effects of AGEPC on both renin secretion and calcium permeability. Furthermore, addition of AGEPC to the cell cultures led to a decrease of 32P-labeled phosphatidylinositol 4,5-bisphosphate and to an increase in 3H-labeled diacylglycerol, indicating an activation of phospholipase C by AGEPC.

Animals↗