Histochem Cell Biol (2009) 132:199–210 DOI 10.1007/s00418-009-0588-y 123 ORIGINAL PAPER Doxycycline accelerates renal cyst growth and Wbrosis in the pcy/pcy mouse model of type 3 nephronophthisis, a form of recessive polycystic kidney disease Larissa Osten · Marion Kubitza · Anna Rachel Gallagher · Jürgen Kastner · Heike Olbrich · Uwe de Vries · Frieder Kees · Brigitte Lelongt · Stefan Somlo · Heymut Omran · Ralph Witzgall Accepted: 18 March 2009 / Published online: 21 April 2009  Springer-Verlag 2009 Abstract Nephronophthisis belongs to a family of reces- sive cystic kidney diseases and may arise from mutations in multiple genes. In this report we have used a spontaneous mouse mutant of type 3 nephronophthisis to examine whether the doxycycline-inducible synthesis of Timp-2, a natural inhibitor of matrix metalloproteinases, can inXuence renal cyst growth in transgenic mice. Metalloproteinases may exert either a negative or a positive eVect on the pro- gression of cystic kidney disease, and we reasoned that this may be most eVectively examined by using a natural inhibitor. Surprisingly, already the application of doxycycline, which also inhibits matrix metalloproteinases, accelerated renal cyst growth and led to increased renal Wbrosis, an additional eVect of Timp-2 was not detected. The positive eVect of doxycycline on kidney size was not due to a non-speciWc “anabolic eVect” but was speciWc for cystic kidneys because it was not observed in non-cystic kidneys. When looking for potential metabolic changes we noticed that the urine of control animals led to an increase in the calcium response of LLC-PK 1 cells, whereas the urine of doxycy- cline-treated mice showed the opposite eVect and even antagonized the urine of control animals. Further experi- ments demonstrated that the urine of control animals con- tained a heat-labile, proteinase K-resistant substance which appears to be responsible for the induction of a calcium response in LLC-PK 1 cells. We conclude that doxycycline accelerates cyst growth possibly by the induction of a sub- stance which lowers the intracellular calcium concentra- tion. Our data also add a note of caution when interpreting phenotypes of animal models based upon the tet system. Keywords Doxycycline · Polycystic kidney disease · Nephronophthisis · Intracellular calcium · Primary cilia Introduction Although many genes mutated in various forms of cystic kidney diseases have been identiWed, it is still not clear how cysts are formed. Evidence has been presented that primary cilia, extensions of the apical plasma membrane, transmit mechanical stimuli to tubular epithelial cells and that fail- ure to do so initiates a cystogenic program (Nauli et al. 2003; Praetorius and Spring 2001). What such a cystogenic program looks like has been an intense subject of specula- tion. Since the tubules and cysts are surrounded by a base- ment membrane and additional extracellular matrix which forms a mechanical barrier to expansion, it is reasonable to assume that the extracellular matrix has to be remodeled for Larissa Osten and Marion Kubitza have contributed equally. L. Osten · M. Kubitza · J. Kastner · U. de Vries · R. Witzgall (&) Institute for Molecular and Cellular Anatomy, University of Regensburg, Universitätsstrasse 31, 93053 Regensburg, Germany e-mail: ralph.witzgall@vkl.uni-regensburg.de A. R. Gallagher · S. Somlo Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA H. Olbrich · H. Omran Department of Pediatrics, University of Freiburg, Freiburg, Germany F. Kees Department of Pharmacology and Toxicology, University of Regensburg, Regensburg, Germany B. Lelongt Inserm U702, Hôpital Tenon, Université Pierre Et Marie Curie (Paris 6), Paris, France