Transforming growth factor β (TGF-β ) has been shown to participate in the pathophysiology of diabetic complications. As shown most recently, TGF-β stimulates the expression of a distinct ...serine/threonine kinase (hSGK) which had previously been cloned as an early gene transcriptionally regulated by cell volume alterations. The present study was performed to elucidate transcription and function of hSGK in diabetic nephropathy. As shown by Northern blotting, an increase of extracellular glucose concentration increased hSGK mRNA levels in cultured cells, an effect qualitatively mimicked by osmotic cell shrinkage or treatment with TGF-β (2 μ g/liter), phorbol 12,13-didecanoate (1 μ M), or the Ca2+ionophore ionomycin (1 μ M) and blunted by high concentrations of nifedipine (10 and 100 μ M). In situ hybridization revealed that hSGK transcription was markedly enhanced in diabetic nephropathy, with particularly high expression in mesangial cells, interstitial cells, and cells in thick ascending limbs of Henle's loop and distal tubules. According to voltage clamp and tracer flux studies in Xenopus oocytes expressing the renal epithelial Na+channel ENaC or the mouse thick ascending limb Na+,K+,2Cl-cotransporter BSC-1, coexpression with hSGK stimulated ENaC and BSC-1 11-fold and 6-fold, respectively, effects reversed by kinase inhibitors staurosporine (1 μ M) and chelerythrine (1 μ M) and not elicited by inactive hSGK. In conclusion, excessive extracellular glucose concentrations enhance hSGK transcription, which in turn stimulates renal tubular Na+transport. These observations disclose an additional element in the pathophysiology of diabetic nephropathy.
Earlier, we have introduced the spectral index (SI), which was derived from the harmonic content of the blood flow velocity envelope of the ophthalmic artery. SI changed in dependency on the baseline ...blood pressure (bBP). We now examined SI during sympathetic activation by cold stimulation for 300 s in dependency on bBP to investigate the response to sympathetic neural activity in arterial hypertension. Ten men and 12 women with normal bBP (age, 60.5+/-4.6 years and 61.9+/-7.2 years) and age-adjusted men and women with increased bBP underwent the cold pressor test, including a periodical measurement of blood pressure and blood flow velocity in the ophthalmic artery, the latter by pulsed Doppler sonography. From this, the course of the SI was calculated. During cold stimulation men with increased bBP achieved their SI peak and their systolic blood pressure peak earlier than those with normal bBP (P=0.002 and P=0.035, respectively) and their SI slope was steeper than in normotensive men (P=0.002). Multiple testing showed that the difference of SI decrease between men with normal and increased bBP occurs on average 60 s after the beginning of cold stimulation (P=0.018). These differences were not found between female blood pressure groups, but the results in women may be influenced by antihypertensive treatment of some of the hypertensive women. In conclusion, the SI is useful to evaluate the response to sympathetic activation in hypertensive men but a larger study population should confirm the study results in women.
Ample pharmacological evidence points to a role of kinases in the regulation of cell volume. Given the limited selectivity of most inhibitors, however, the specific molecules involved have remained ...largely elusive. The search for cell volume regulated genes in liver HepG2 cells led to the discovery of the human serum- and glucocorticoid-dependent serine/threonine kinase hsgk1. Transcription and expression of hsgk1 is markedly and rapidly upregulated by osmotic and isotonic cell shrinkage. The effect of osmotic cell shrinkage on hsgk1 is mediated by p38 kinase. Further stimuli of hsgk1 transcription include glucocorticoids, aldosterone, TGF-β1, serum, increase of intracellular Ca
2+ and phorbolesters, whereas cAMP downregulates hsgk1 transcription. The hsgk1 protein is expressed in several epithelial tissues including human pancreas, intestine, kidney, and shark rectal gland. Co-expression of hsgk1 with the renal epithelial Na
+-channel ENaC or the Na
+/K
+/2Cl
−-cotransporter NKCC2 (BSC1) in
Xenopus oocytes, accelerates insertion of the transport proteins into the cell membrane and thus, stimulates channel or transport activity. Thus, hsgk1 participates in the regulation of transport by steroids and secretagogues increasing intracellular Ca
2+-activity. The stimulation of hsgk1 transcription by TGF-β1 may further bear pathophysiological relevance.
The serum- and glucocorticoid-dependent kinase SGK1 is regulated by alterations of cell volume, whereby cell shrinkage increases and cell swelling decreases the transcription, expression and activity ...of SGK1. The kinase is expressed in all human tissues studied including the brain. The present study was performed to localize the sites of SGK1 transcription in the brain, to elucidate the influence of the hydration status on SGK1 transcription and to explore the functional significance of altered SGK1 expression. Northern blot analysis of human brain showed SGK1 to be expressed in all cerebral structures examined: amygdala, caudate nucleus, corpus callosum, hippocampus, substantia nigra, subthalamic nucleus and thalamus. In situ hybridization and immunohistochemistry in the rat revealed increased expression of SGK1 in neurons of the hippocampal area CA3 after dehydration, compared with similar slices from brains of euvolaemic rats. Additionally, several oligodendrocytes, a few microglial cells, but no astrocytes, were positive for SGK1. The abundance of SGK1 mRNA in the temporal lobe, including hippocampus, was increased by dehydration and SGK1 transcription in neuroblastoma cells was stimulated by an increase of extracellular osmolarity. Co-expression studies in Xenopus laevis oocytes revealed that SGK1 markedly increased the activity of the neuronal K+ channel Kv1.3. As activation of K+ channels modifies excitation of neuronal cells, SGK1 may participate in the regulation of neuronal excitability.
Transcript levels of the human serine/threonine kinase h-sgk have been found to be highest in pancreas. In the present study, localization and regulation of h-sgk transcription in pancreatic tissue ...were elucidated. As was apparent from radioactive in situ hybridization, most pancreatic acinar cells expressed high levels of h-sgk mRNA. h-sgk mRNA-positive cells were also found in ductal epithelia but not in pancreatic islets. In biopsy specimens from patients with pancreatitis, h-sgk mRNA levels were decreased in acinar cells but abundant in numerous mononuclear interstitial cells within areas of pancreatic necrosis and fibrosis. As shown by Northern blotting, h-sgk transcription in DAN-G pancreatic tumor cells is upregulated by osmotic cell shrinkage, serum, phorbol esters (phorbol 12,13-didecanoate), and Ca(2+) ionophore A-23187 and decreased by staurosporine and cAMP. In conclusion, h-sgk transcription is regulated not only by cell volume but also by serum, protein kinase C stimulation, cAMP, and increase of intracellular Ca(2+) activity. The kinase may participate not only in normal function of exocrine pancreas but also in fibrosing pancreatitis.
Fragestellung: Das Glaukom ist eine Erkrankung mit steigender Pravalenz. Dabei weisen ein Drittel der Patienten ein Normaldruckglaukom (NDG) auf, das nicht durch erhohten Augeninnendruck wie das ...primare Offenwinkelglaukom (POWG) verursacht wird. Die Unterscheidung dieser Formen ist bislang mittels ophthalmologischer Untersuchungen allein nicht sicher moglich, vor dem Hintergrund einer grundsatzlich unterschiedlichen Therapie aber von grosstem Interesse. Ziel dieser studie war es zu untersuchen, ob sich die beiden Glaukomformen anhand unterschiedlicher Veranderungsmuster der fraktionalen Anisotropie (FA) innerhalb der sehbahnen mittels DTI unterscheiden lassen. Methoden: 19 NDG- und 22 POWG-Patienten im fortgeschrittenen Krankheitsstadium sowie 25 sehgesunde Personen im Alter zwischen 24 und 85 Jahren wurden bei 3 T mit einer hochaufgelosten DTI-Sequenz (Matrix = 320 x 256) untersucht. Mit Hilfe einer automatisierten Matlab-Routine wurde die FA im 3. Neuron der sehbahn (sehnerv bis zum corpus geniculatum laterale) und im 4. Neuron (sehstrahlung bis zum optischen Kortex) berechnet und die Mittelwerte innerhalb der einzelnen Gruppen miteinander verglichen. Ergebnisse: Gegenuber den sehgesunden Personen zeigten NDG- und POWG-Patienten eine signifikante Reduktion der durchschnittlichen FA um 11% bzw. 14% entlang der sehbahn. Innerhalb der NDG- und POWG-Patienten fand sich zudem eine unterschiedliche Auspragung der FA-Reduktion im 3. bzw. 4. Neuron: 4% bzw. 14% bei NDG und 19% bzw. 5% bei POWG. Im Gegensatz zu POWG-Patienten fand sich bei NDG-Patienten eine signifikant hohere FAReduktion im 4. Neuron (P < 0,01). Schlussfolgerung: Erste Ergebnisse deuten darauf hin, dass DTI anhand der unterschiedlichen FA-Anderungsmusterinnerhalb der sehbahnen eine sichere Differenzierung zwischen Normaldruck- und Hochdruckglaukom ermoglicht.
One of the striking morphological features of renal failure is an increase of cell volume. This review explores the role of cell volume regulatory mechanisms in the pathophysiology of progressive ...renal disease. The case is made that TGF-beta, a major cytokine involved in the development of progressive renal failure, upregulates the transcription of the serum and glucocorticoid-dependent kinase hSGK1, involved in cell volume regulation. Excessive extracellular glucose concentrations stimulate TGF-beta1 expression and thus similarly enhance hSGK1-transcription. The kinase stimulates two mechanisms important for cell volume regulation, i.e. the renal epithelial Na+ channel ENaC and the thick ascending limb Na+,K+,2Cl- cotransporter BSC1. On the one hand, stimulation of renal tubular transport leads to renal retention of Na+, which favours the development of hypertension. On the other, the increase of cell volume stimulates protein synthesis and inhibits protein degradation, contributing to the enhanced net formation and deposition of matrix proteins. At later stages, the increase of cell volume may be reversed to atrophy, and cell death may lead to loss of functional tissue. In conclusion, progressive renal disease is paralleled by deranged cell volume regulatory mechanisms.
Glomerulonephritis is paralleled by excessive formation of transforming growth factor-beta (TGF-beta), which participates in the pathophysiology of the disease. Recently, a novel downstream target of ...TGF-beta has been identified, i.e. the human serum and glucocorticoid-dependent kinase 1 (hSGK1), a serine/threonine kinase participating in the regulation of Na(+) transport. The present study was performed to elucidate transcriptional regulation of hSGK1 in glomerulonephritis. To this end, in situ hybridization was performed in biopsies from patients with clinical diagnosis of glomerulonephritis. hSGK1 transcript levels were moderately enhanced in 5 out of 9 patients and strongly enhanced in 4 out of 9 patients. Distal nephron epithelial cell hSGK1 transcript levels were low or absent in 7 of the 9 patients but markedly enhanced in 2 of the 9 patients. In conclusion, glomerulonephritis leads to glomerular and in some cases to epithelial up-regulation of hSGK1 transcription.
The excessive matrix deposition in lung fibrosis is thought to be due to enhanced formation and activity of TGFbeta1, which stimulates synthesis and inhibits degradation of matrix proteins. The ...cellular mechanisms triggered by TGFbeta1 are still incompletely understood. Recently, a novel transcriptional target of TGFbeta1 has been identified, i.e. the human serum and glucocorticoid dependent kinase hSGK1. The present study has been performed to explore whether TGFbeta1 stimulates hSGK1 transcription in lung fibroblasts and whether lung fibrosis is associated with enhanced hSGK1 expression. As evident from Northern Blotting, TGFbeta1 strongly upregulates hSGK1 in human lung fibroblasts, an effect partially reversed by p38-kinase inhibitor SB203580. In situ hybridization experiments reveal that in intact lung tissue hSGK1 is expressed in single type II alveolar pneumocytes and macrophages. In contrast, in fibrotic lung tissue a dramatic upregulation of hSGK1 mRNA as well as a strong expression of hSGK1 protein is observed in epithelial cells and interstitial cells comprising macrophages and fibroblasts. In conclusion, in lung fibrosis, the serine/threonine kinase hSGK1 is upregulated, an effect at least partially accounted for by TGFbeta1. The full effect of TGFbeta1 requires the activation of p38 kinase.
The human serine/threonine kinase hSGK1 is expressed ubiquitously with highest transcript levels in pancreas and liver. This study has been performed to determine the hSGK1 distribution in normal ...liver and its putative role in fibrosing liver disease. HSGK1-localization was determined by in situ hybridization, regulation of hSGK1-transcription by Northern blotting, fibronectin synthesis and hSGK1 phosphorylation by Western blotting. In normal liver hSGK1 was mainly transcribed by Kupffer cells. In liver tissue from patients with chronic viral hepatitis, hSGK1 transcript levels were excessively high in numerous activated Kupffer cells and inflammatory cells localized within fibrous septum formations. HSGK1 transcripts were also detected in activated hepatic stellate cells. Accordingly, Western blotting revealed that tissue from fibrotic liver expresses excessive hSGK1 protein as compared to normal liver. TGF-beta1 (2 ng/ml) increases hSGK1 transcription in both human U937 macro-phages and HepG2 hepatoma cells. H(2)O(2) (0.3 mM) activated hSGK1 and increased fibronectin formation in HepG2 cells overexpressing hSGK1 but not in HepG2 cells expressing the inactive mutant hSGK1(K127R). In conclusion hSGK1 is upregulated by TGF-beta1 during hepatitis and may contribute to enhanced matrix formation during fibrosing liver disease.
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