The developing rat kidney : the dopamine system and related serine/threonine kinases and phosphatases
Sammanfattning: The kidney originates from the fusion and conversion of two tissues: the metanephric mesenchyme and the ureter epithelium, an evagination from the Wolffian duct. Although a number of candidate molecules have been shown to be of importance for the differentiation of the embryonic kidney, many of the signalling pathway leading to this development are still obscure. A large proportion of important functions in eukaryotic cells is meticulously regulated by reversible protein phosphorylation, carried out by protein kinases and phosphates. This thesis presents data from five studieson serine/threonine protein phosphorylation during the foetal and postnatal development of the rat kidney. The isoforms of protein phosphatases (PP) 1 and PP2A had differential and age-dependent expression patterns, initially with a general distribution, shifting to restricted distributions later in nephrogenesis. All were present in the peripheral, nephrogenic, zone. Inhibition of PP1 and PP2A with okadaic acid in metanephric culture severely disturbed growth and differentiation and induced apoptosis. Protein kinase C (PKC) isoforms [alpha], [delta], [zeta] and [lambda]were age-dependently expressed with individual distribution patterns. Inhibition of PKC in primary culture of proximal tubule cells and in metanephric organ culture resulted in disrupted morphogenesis, increased apoptosis and age-dependent growth inhibition. Inhibitor-1, an endogenous and specific inhibitor of PP1, was specifically expressed in uninduced metanephric mesenchyme at embryonic day (E) 15. Inhibitor-1 is the first known marker of these cells. DARPP-32, another specific PP1 inibitor with a central position in dopaminergic cell signalling, was not detected until E 18. From that age, DARPP-32 was highly concentrated in ureter bud tips, the inducing tissue. The same distribution of DARPP-32 was observed in foetal mouse kidneys. Mice deficient in DARPP-32 had a 28% nephron deficit. They were also unable to regulate natriuresis in response to atrial natriuretic peptide. Furthermore, these mice were hypertensive. Dopamine and its synthesizing (AADC) and metabolizing (COMT) enzymes, as well as the dopamine D1A receptor, were all detected in foetal rat kidneys, each with specific distribution. Conclusion: Reversible protein phosphorylation is an important intracellular modulating system for kidney morphogenesis. This study also presents, for the first time, data indicating important roles for dopamine and dopamine-related signalling during nephrogenesis. The DARPP-32 deficient mice constitute a new model for limited nephron deficit and may be valuable in future research on renal insufficiency and hypertension.
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