Inhibition of aquaporin-4 expression in astrocytes by RNAi determines alteration in cell morphology, growth, and water transport and induces changes in ischemia-related genes.
Recent studies indicate a key role of aquaporin (AQP) 4 in astrocyte swelling and brain edema and suggest that AQP4 inhibition may be a new therapeutic way for reducing cerebral water accumulation. To understand the physiological role of AQP4-mediated astroglial swelling, we used 21-nucleotide small interfering RNA duplexes (siRNA) to specifically suppress AQP4 expression in astrocyte primary cultures. Semiquantitative RT-PCR experiments and Western blot analysis showed that AQP4 silencing determined a progressive and parallel reduction in AQP4 mRNA and protein. AQP4 gene suppression determined the appearance of a new morphological cell phenotype associated with a strong reduction in cell growth. Water transport measurements showed that the rate of shrinkage of AQP4 knockdown astrocytes was one-half of that of controls. Finally, cDNA microarray analysis revealed that the gene expression pattern perturbed by AQP4 gene silencing concerned ischemia-related genes, such as GLUT1 and hexokinase. Taken together, these results indicate that 1) AQP4 seems to be the major factor responsible for the fast water transport of cultured astrocytes; 2) as in skeletal muscle, AQP4 is a protein involved in cell plasticity; 3) AQP4 alteration may be a primary factor in ischemia-induced cerebral edema; and 4) RNA interference could be a new potent tool for studying AQP pathophysiology in those organs and tissues where they are expressed.
Nicchia
Grazia Paola
GP
Department of General and Environmental Physiology and Center of Excellence in Comparative Genomics (CEGBA), University of Bari, I-70126 Bari, Italy.
Frigeri
Antonio
A
Liuzzi
Grazia Maria
GM
Svelto
Maria
M
D000818:Animals; D051401:Aquaporin 4; D020346:Aquaporins / Q000096:biosynthesis / Q000235:genetics* / Q000502:physiology*; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D001692:Biological Transport; D001921:Brain / Q000166:cytology*; D001929:Brain Edema / Q000235:genetics / Q000378:metabolism; D002545:Brain Ischemia / Q000235:genetics / Q000378:metabolism; D002455:Cell Division; D002478:Cells, Cultured; D020869:Gene Expression Profiling; D034622:RNA Interference; D014867:Water / Q000378:metabolism*
D051401:Aquaporin 4; D020346:Aquaporins; D014867:Water