GH and bone metabolism: the role of the vitamin D system.
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Animals↗
Biomedical subjects
J Pfeilschifter
Publications and source records attributed to J Pfeilschifter.
Suppression of serum thyrotropin with thyroxine in patients with Graves' disease: effects on recurrence of hyperthyroidism and thyroid volume.
Based on findings that thyroxine may have a beneficial effect on the recurrence of Graves' hyperthyroidism, we prospectively studied the effects of a TSH suppressive treatment with thyroxine on the course of Graves' disease in fifty patients with recent onset of hyperthyroidism. After the normalization of serum tri-iodothyronine (T3) and thyroxine (T4) concentrations, one group of patients was randomly assigned to a combined treatment with carbimazole and a TSH suppressive dose of T4 for 12 months, followed by another 12 months of TSH suppressive therapy alone. The other group of patients also received carbimazole for one year, but T4 was only added as indicated to normalize elevated TSH serum concentrations, and patients received no therapy during the second year. By the end of the second year, a relapse of hyperthyroidism had occurred in 43% of the patients with and in 45% of the patients without suppressive T4 treatment. In those patients without a relapse of hyperthyroidism, initial thyroid size significantly (P = 0.01) decreased with time in both treatment groups. However, patients on suppressive T4 treatment tended to have a greater reduction in thyroid volume than patients with normal TSH serum concentrations (P = 0.05). In conclusion, we were unable to detect a preventive effect of exogenous TSH suppression on the recurrence of hyperthyroidism. However, our data suggest that TSH suppressive treatment may have a beneficial effect on thyroid enlargement in Graves' disease.
Adrenergic beta-Antagonists↗
Apoptosis and expression of inducible nitric oxide synthase are mutually exclusive in renal mesangial cells.
Nitric oxide (NO) is a multipurpose messenger molecule, important for blood vessel relaxation, neuronal communication, and antimicrobial activities. The generation of NO from L-arginine is catalyzed by NO synthase (NOS). An inducible form of NOS, iNOS, was first characterized in macrophages and then in many other tissues and cells, including renal mesangial cells. Mesangial cells play a crucial role in the regulation of the glomerular filtration rate as well as in the pathophysiology of certain forms of glomerulonephritis in which mesangial cells and macrophages produce NO in high amounts. Because reports have associated NO production with apoptotic cell death in macrophages and we recently demonstrated NO-mediated apoptosis in mesangial cells, we searched for the relationship between in situ iNOS induction and apoptosis by iNOS immunocytochemistry and terminal desoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) staining. RAW 264.7 macrophages exhibited homogeneous iNOS expression and apoptotic nuclei in the iNOS-containing cells upon stimulation with interferon-gamma and lipopolysaccharide. In contrast, stimulated rat mesangial cells stained heterogeneously for iNOS, depending on cell passage and iNOS-stimulating pathway. Mesangial cells expressing iNOS did not display signs of apoptosis and, vice versa, cells showing characteristic features of apoptosis did not stain for iNOS. Thus, our study suggests that mesangial cells react to stimulation by interleukin-1 and/or cAMP-elevating compounds with mutually exclusive responses, either by expression of iNOS or by undergoing programmed cell death.
Animals↗
[Impact of pretreatment variables on the outcome of standardized 131I therapy with 150 Gray in Graves' disease].
AIM: We examined the impact of several pretreatment variables on thyroid size and function in 61 patients with Graves' disease one year after a standardized [131]I treatment with 150 Gray. METHODS: FT3, FT4, and TSH serum concentrations were determined before and 1.5, 3, 6, and 12 months after therapy. Thyroid size was measured by ultrasound and scintigraphy before and one year after therapy. RESULTS: One year after therapy, 30% of the patients had latent or manifest hyperthyroidism, 24% were euthyroid, and 46% had developed latent or manifest hypothyroidism. Age and initial thyroid volume were major predictors of post-therapeutical thyroid function. Thus, persistent hyperthyroidism was observed in 70% of the patients age 50 years and older with a thyroid size of more than 50 ml. With few exception, thyroid size markedly decreased after therapy. Initial thyroid size and age were also major predictors of posttherapeutical thyroid volume. Thyroid size normalized in all patients younger than 50 years of age, independent from initial thyroid size. CONCLUSION: Radioiodine treatment with 150 Gray causes a considerable decrease in thyroid size in most patients with Graves' disease. Age and initial thyroid volume are important determinants of thyroid function and size after therapy and should be considered in dose calculation.
Adult↗
The glomerular cell: source and target of various mediators.
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Animals↗
Nitric oxide donors induce apoptosis in glomerular mesangial cells, epithelial cells and endothelial cells.
Renal mesangial cells exposed to inflammatory cytokines produce high concentrations of nitric oxide (NO) which may exert cytotoxic actions. We report here that glomerular mesangial cells, endothelial cells and epithelial cells in culture are themselves targets for NO and undergo apoptotic cell death upon exposure to high concentrations of NO. NO generated from different NO-releasing compounds as well as NO-saturated solution induce apoptosis in all three cell types as demonstrated by internucleosomal DNA fragmentation, an enrichment of cytosolic DNA/histone complexes, an increasing number of cellular 3'-OH-fragmented DNA ends and typical nuclear chromatin condensation. Induction of apoptosis was found to be dependent on protein synthesis and is preceded by expression of the tumour suppressor gene product p53 in mesangial cells. Induction of inducible NO synthase in mesangial cells by interleukin-1 beta leads to excessive formation of NO by the cells as measured by nitrite production. However, there was no evidence for apoptotic changes in mesangial cells triggered by endogenously produced NO. Co-cultures of glomerular endothelial or epithelial cells with interleukin-1 beta-activated mesangial cells expressing inducible NO synthase do not show apoptotic alterations in endothelial or epithelial cells. Moreover, preincubation of mesangial cells with interleukin-1 beta protects the cells from apoptosis induced by subsequent addition of exogenous NO thus suggesting that interleukin-1 beta not only triggers the expression of inducible NO synthase and massive NO formation but simultaneously stimulates a protecting principle in the cells. In summary, these results suggest that exogenous NO can induce apoptosis in all three types of intrinsic glomerular cells. However, whether endogenously produced NO can fulfil this function critically depends on a balance between a yet to be defined protective mechanism and inducible NO synthase expression in mesangial cells in response to interleukin-1 beta and eventually other inflammatory cytokines.
Animals↗
Nitric oxide stimulates stress-activated protein kinases in glomerular endothelial and mesangial cells.
Exposure of rat glomerular mesangial cells and primary cultures of bovine glomerular endothelial cells to compounds releasing nitric oxide (NO), including MAHMA-NONOate, S-nitrosoglutathione, and spermine-NO, results in a time- and concentration-dependent activation of stress-activated protein kinases (SAPK) as measured by the phosphorylation of c-Jun in a solid phase kinase assay. Dibutyryl cGMP had no effect on SAPK activity. Pretreatment of the cells with the tyrosine kinase inhibitor genistein strongly attenuated NO-induced c-Jun phosphorylation. Furthermore, N-acetylcysteine markedly reduced the activation of SAPK in response to NO. These studies identify SAPK as a target for NO which may be critical for the NO-induced apoptosis of glomerular mesangial and endothelial cells.
Animals↗
Interleukin 1 beta and cAMP trigger the expression of GTP cyclohydrolase I in rat renal mesangial cells.
Endogenous synthesis of tetrahydrobiopterin (BH4) is an important requirement for cytokine-stimulated nitric oxide (NO) production in mesangial cells. We have shown that inducible NO synthase is expressed in mesangial cells in response to two principal classes of activating signals, inflammatory cytokines such as interleukin 1 beta (IL-1 beta) and agents that elevate cellular levels of cAMP [Kunz, Mühl, Walker and Pfeilschifter (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 5387-5391]. In the present paper we demonstrate that IL-1 beta and cAMP similarly increase the steady-state mRNA levels of GTP cyclohydrolase I (EC 3,5,4,16), the rate-limiting enzyme in BH4 biosynthesis, as measured by a sensitive and quantitative nuclease protection assay. Stimulation of cells with a combination of IL-1 beta plus cAMP revealed an additive induction profile of GTP cyclohydrolase I mRNA. Message stability studies established that GTP cyclohydrolase I mRNA induced by cAMP has a longer half-life than the IL-1 beta-induced message. Moreover, cAMP exposure markedly prolonged the half-life of GTP cyclohydrolase I mRNA, from 1.5 to 3.4 h. In a next step we generated a rabbit polyclonal antibody against rat GTP cyclohydrolase I expressed in Escherichia coli and demonstrated that IL-1 beta and cAMP elevated GTP cyclohydrolase I protein levels in mesangial cells. Furthermore, IL-1 beta and cAMP led to a marked increase in GTP cyclohydrolase I activity and to increased accumulation of biopterin in mesangial cells. Combinations of IL-1 beta and cAMP resulted in a synergistic stimulation of GTP cyclohydrolase I activity. This may suggest that, in addition to transcriptional and post-transcriptional regulation, there is a prominent post-translational modulation of enzyme activity.
Animals↗
Ceramide-binding and activation defines protein kinase c-Raf as a ceramide-activated protein kinase.
Interleukin 1 is the prototype of an inflammatory cytokine, and evidence suggests that it uses the sphingomyelin pathway and ceramide production to trigger mitogen-activated protein kinase (MAPK) activation and subsequent gene expression required for acute inflammatory processes. To identify downstream signaling targets of ceramide, a radioiodinated photoaffinity labeling analog of ceramide ([125I] 3-trifluoromethyl-3-(m-iodophenyl)diazirine-ceramide) was employed. It is observed that ceramide specifically binds to and activates protein kinase c-Raf, leading to a subsequent activation of the MAPK cascade. Ceramide does not bind to any other member of the MAPK module nor does it bind to protein kinase C-zeta. These data identify protein kinase c-Raf as a specific molecular target for interleukin 1 beta-stimulated ceramide formation and demonstrate that ceramide is a lipid cofactor participating in regulation of c-Raf activity.
Affinity Labels↗
Molecular cloning of the rat inducible nitric oxide synthase gene promoter.
The inducible nitric oxide synthase (iNOS) is not constitutively expressed but is induced in many types of mammalian cells by cytokines and bacterial endotoxins. Previously, we have reported the most interesting feature of rat iNOS gene that is up-regulated by cyclic AMP at transcriptional level as demonstrated by nuclear run-on assay. We now have isolated and sequenced the rat iNOS gene promoter and determined the transcription start site. Moreover, we have constructed a rat iNOS promoter-chloramphenicol acetyltransferase (CAT) fusion gene and analyzed its inducibility by interleukin 1 beta and cyclic AMP in transiently transfected Swiss 3T3 fibroblasts.
3T3 Cells↗
Tetranactin inhibits interleukin 1 beta and cAMP induction of group II phospholipase A2 in rat renal mesangial cells.
Renal mesangial cells express secretory phospholipase A2 in response to two principal classes of activating signals that may interact in a synergistic fashion. These two groups of activators comprise inflammatory cytokines, such as interleukin 1 beta, and agents that elevate cellular levels of cAMP. Treatment of mesangial cells with tetranactin, a cyclic antibiotic produced by Streptomyces aureus with a molecular structure similar to cyclosporin A inhibits interleukin 1 beta- and cAMP-dependent group II phospholipase A2 secretion in a dose-dependent manner with IC50 values of 43 and 33 nM, respectively. However, tetranactin does not directly inhibit group II phospholipase A2 activity. Western blot analyses of mesangial cell supernatants reveal that the inhibition of phospholipase A2 activity is due to suppression of phospholipase A2 protein synthesis. This effect is preceded by the reduction of phospholipase A2 mRNA steady-state levels as shown by Northern blot analyses of total cellular RNA isolated from stimulated mesangial cells. Thus, tetranactin is a potent inhibitor of group II phospholipase A2 expression in cytokine- and cAMP-stimulated mesangial cells and represents a new class of group II phospholipase A2 inhibitors with IC50 values in the low nanomolar range. This compound may be useful in the therapy of diseases associated with increased group II phospholipase A2 secretion.
Analysis of Variance↗
Effects of dexamethasone and transforming growth factor-beta 2 on group II phospholipase A2 mRNA and activity levels in interleukin 1 beta- and forskolin-stimulated mesangial cells.
The expression of 14 kDa group II phospholipase A2 [also referred to as secretory PLA2 (sPLA2)] is induced in rat glomerular mesangial cells by exposure to inflammatory cytokines and forskolin, a cAMP elevating agent. Previously we have shown that dexamethasone and transforming growth factor-beta 2 (TGF-beta 2) suppress sPLA2 protein synthesis and enzyme activity induced by cytokines and forskolin. The regulation of sPLA2 by pro-inflammatory cytokines suggests that the enzyme may play a role in glomerular inflammatory reactions. In order to understand the regulation of sPLA, in more detail, we investigated whether dexamethasone and TGF-beta 2 also suppress sPLA, mRNA after its induction by either interleukin-1 beta (IL-1 beta) or forskolin. We found that IL-1 beta-induced sPLA2 mRNA in rat mesangial cells is not down-regulated by pretreatment of the cells with dexamethasone, even at a concentration of 10 microM, which dramatically decreases sPLA2 protein levels and activity. Metabolic labelling experiments indicated that the decreased sPLA2 levels under these conditions can be explained by inhibition of the rate of sPLA2 synthesis from the elevated mRNA levels. In contrast, the forskolin-induced elevation of sPLA, mRNA is inhibited by dexamethasone in a concentration-dependent manner. Likewise, TGF-beta 2 inhibits the elevation of sPLA, mRNAs induced by either IL-1 beta or forskolin. The decrease in sPLA2 mRNA caused by TGF-beta 2 corresponds with the decrease in sPLA2 enzyme levels and activity. These data suggest that cytokine- and forskolin-induced sPLA2, expression is tightly controlled via both transcriptional and post-transcriptional mechanisms. Furthermore, we show that pretreatment of mesangial cells with epidermal growth factor prior to stimulation with IL-1 beta or forskolin had no suppressing effect on sPLA2 levels or enzyme activity, as has been reported previously for osteoblasts.
Animals↗
Apoptosis is triggered by the cyclic AMP signalling pathway in renal mesangial cells.
Glomerular mesangial cells are regarded as specialized smooth muscle cells located within the renal glomeruli and fulfilling important functions in glomerular physiology and pathophysiology. Here, we report that activation of the cyclic AMP signalling pathway by dibutyryl cyclic AMP, forskolin, or the beta 2-adrenergic receptor agonist salbutamol results in induction of apoptosis in mesangial cells. Activation of the apoptotic programme results in DNA fragmentation which is visible for most forms of apoptosis and is paralleled by enrichment of cytosolic DNA/histone complexes, an increasing number of cellular 3'-OH-fragmented DNA ends and typical nuclear chromatin condensation. Induction of apoptosis was found to be dependent on translation and independent of nitric oxide synthase activity.
Adrenergic beta-Agonists↗
Effect of cyclic AMP and prostaglandin E2 on the induction of nitric oxide- and prostanoid-forming pathways in cultured rat mesangial cells.
Cyclic AMP (cAMP) represents an important cellular signalling molecule. We analysed the effect of dibutyryl cAMP (db-cAMP), a cell-permeable and stable derivative of cAMP, on the regulation and expression of cyclo-oxygenase 2, inducible NO synthase and argininosuccinate synthetase. We observed different transcriptional regulation of these enzymes depending on the db-cAMP concentration used. Low concentrations of db-cAMP in the range 10-50 microM elevated levels of cyclo-oxygenase 2 mRNA, protein and activity, but not the respective mRNA and protein concentrations of the inducible NO synthase or argininosuccinate synthetase. At higher concentrations a massive induction of the latter two enzymes was also apparent. Expression of prostacyclin synthase and argininosuccinate lyase, secondary enzymes of NO- and prostanoid-forming pathways, was not stimulated by db-cAMP. Prostaglandin E2, known to be an intracellular physiological trigger of cAMP formation, stimulated only cyclooxygenase 2 expression and activity at a concentration of 10 microM, and not inducible NO synthase. The induction of the mRNA for the transcription factors JunB and p65, a component of the NF kappa B complex, by prostaglandin treatment of the cells might be a possible mechanistic explanation for this observation.
Animals↗
Molecular mechanisms of dexamethasone inhibition of nitric oxide synthase expression in interleukin 1 beta-stimulated mesangial cells: evidence for the involvement of transcriptional and posttranscriptional regulation.
Inducible nitric oxide synthase (iNOS; EC 1.14.13.39) is expressed in rat glomerular mesangial cells upon exposure to the inflammatory cytokine interleukin 1 beta (IL-1 beta). We have reported that nanomolar concentrations of dexamethasone suppress IL-1 beta-induced iNOS protein expression and production of nitrite, the stable end product of NO formation, without affecting IL-1 beta-triggered increase in iNOS mRNA levels. We now have studied the mechanisms by which dexamethasone suppresses IL-1 beta-stimulated iNOS expression in mesangial cells. Surprisingly, nuclear run-on transcription experiments demonstrate that dexamethasone markedly attenuates IL-1 beta-induced iNOS gene transcription. However, this is counteracted by a prolongation of the half-life of iNOS mRNA from 1 h to 2.5 h by dexamethasone. Moreover, dexamethasone drastically reduces the amount of iNOS protein by reduction of iNOS mRNA translation and increased degradation of iNOS protein. These results indicate that glucocorticoids act at multiple levels to regulate iNOS expression, thus providing important insights into the treatment of inflammatory diseases.
Animals↗
Therapeutic strategies for the inhibition of inducible nitric oxide synthase--potential for a novel class of anti-inflammatory agents.
In recent years, NO, a gas previously considered a potentially toxic chemical, has become established as a diffusible universal messenger mediating cell-cell communication throughout the body. In mammals, NO is a recognized mediator of blood vessel relaxation that helps to maintain blood pressure. In the central nervous system NO acts as a non-conventional neurotransmitter and participates in the establishment of long-term plasticity required for memory formation. In addition, NO is responsible for some parts of the host response to sepsis and inflammation and contributes to certain disease states. A number of strategies have emerged with regard to a pharmacological control of pathological NO overproductions. This review will discuss these novel therapeutic approaches that may provide new means for clinical medicine.
Animals↗
Amplification of nitric oxide synthase expression by nitric oxide in interleukin 1beta-stimulated rat mesangial cells.
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Animals↗
Smoking and endocrine ophthalmopathy: impact of smoking severity and current vs lifetime cigarette consumption.
OBJECTIVE: Smoking has been associated with an increased incidence of endocrine ophthalmopathy (EO). In this study we examined the relation between smoking severity and the incidence of EO symptoms in patients with Graves' hyperthyroidism. DESIGN: Patients were prospectively followed for at least one year after the onset of hyperthyroidism. Smoking and EO status were evaluated at 3-6- months intervals. PATIENTS: Two hundred and fifty-three ambulatory patients with recent onset of Graves' hyperthyroidism were studied. MEASUREMENTS: The incidence of total EO symptoms, proptosis, and diplopia at any time point before and after the occurrence of Graves' hyperthyroidism was assessed by interview and physical examination. RESULTS: Smoking was associated with a 1.3-fold increase in the overall incidence of symptomatic EO, and a 2.6 and 3.1-fold increase in the incidence of proptosis and diplopia, respectively. The relative risk increased in parallel with the current number of cigarettes smoked per day. In contrast, lifetime tobacco use was not an independent risk factor for the development of EO symptoms. Former smokers had a significantly lower risk for the occurrence of proptosis and diplopia than active smokers with a comparable lifetime cigarette consumption. CONCLUSIONS: Our data suggest that current, but not lifetime, tobacco consumption constitutes a risk for the incidence of proptosis and diplopia in patients with Graves' hyperthyroidism, and that this risk increases with smoking severity.
Adult↗