Galactosylceramidase deficiency causes sperm abnormalities in the mouse model of globoid cell leukodystrophy A. Luddi a , M. Strazza a , M. Carbone b , E. Moretti a , E. Costantino-Ceccarini b, * a Department of Pediatrics, Obstetrics and Reproductive Medicine, Section of Biology, University of Siena, Italy b Istituto fisiologia Clinica, Sezione di Siena, CNR, Via Pendola 62, 53100 Siena, Italy Received 18 May 2004, revised version received 27 September 2004 Available online 8 December 2004 Abstract The classical recessive mouse mutant, bthe twitcher,Q is one of the several animal models of the human globoid cell leukodystrophy (Krabbe disease) caused by a deficiency in the gene encoding the lysosomal enzyme galactosylceramidase (GALC). The failure to hydrolyze galactosylceramide (gal-cer) and galactosylsphingosine (psychosine) leads to degeneration of oligodendrocytes and severe demyelination. Substrate for GALC is also the galactosyl-alkyl-acyl-glycerol (GalAAG), precursor of the seminolipid, the most abundant glycolipid in spermatozoa of mammals. In this paper, we report the pathobiology of the testis and sperm in the twitcher mouse and demonstrate the importance of GALC for normal sperm maturation and function. The GALC deficit results in accumulation of GalAAG in the testis of the twitcher mouse. Morphological studies revealed that affected spermatozoa have abnormally swollen acrosomes and angulation of the flagellum mainly at midpiece–principal piece junction. Multiple folding of the principal piece was also observed. Electron microscopy analysis showed that in the twitcher sperm, acrosomal membrane is redundant, detached from the nucleus and folded over. Disorganization and abnormal arrangements of the axoneme components were also detected. These results provide in vivo evidence that GALC plays a critical role in spermiogenesis. D 2004 Elsevier Inc. All rights reserved. Keywords: Galactosylceramidase; Sperm abnormalities; Globoid cell leukodystrophy Introduction In humans and mice, the relationship between the development of cells of the nervous system and the male reproductive system is progressively understood. This relationship becomes apparent by the evaluation of several unlinked autosomal mutations, which affect both systems. This interrelation can be easily investigated in the mouse mutants, taking advantage of several unlinked autosomal mutations causing defects in both systems. Some of these mutants are generated by spontaneous mutations while others are by gene targeting. In both instances, there are some that are authentic models of human diseases, while others do not have a known corresponding human disorder. Examples are Purkinje cell degeneration [1], quaking [2], wobbler [3], weaver [4], and Osp/Claudin-11 [5] mouse. The general importance of lysosomal hydrolases in male fertility is well documented. Recent studies have indicated that mutations affecting lysosomal enzymes exert pleio- tropic effects specifically on spermiogenesis. In fact, in the knockout mice for the lysosomal enzymes sphingomyeli- nase, a, or h hexosaminidase, both nervous system and reproductive system are affected [6, 7]. Evidence exists to suggest that the sperm acrosome is a modified lysosome containing several acidic hydrolases, including h-hexosa- minidase, h-glucuronidase, a-mannosidase, and h-galacto- sidase [8]. In the present study, the twitcher mouse with a mutation in the galactosylceramidase (GALC) gene, was used to investigate the role of this hydrolase in normal sperm 0014-4827/$ - see front matter D 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.yexcr.2004.10.034 * Corresponding author. Fax: +39 0577 233509. E-mail address: costantino@unisi.it (E. Costantino-Ceccarini). Experimental Cell Research 304 (2005) 59 – 68 www.elsevier.com/locate/yexcr