Abstract The knowledge of alterations in regulation of autophagy during tumorigenesis may also help our under- standing of its normal control. We established an experi- mental system and reported recently that autophagic ca- pacity, measured as the cell's capability of increasing seg- regation (formation of autophagosomes) and subsequent degradation of cytoplasmic quanta were highly increased in premalignant nodule cells 6 months after initiation by azaserine in the rat pancreas in vivo. In the present study, we followed changes of these autophagic functions throughout the tumour progression. We carried out elec- tron-microscopic morphometrical analysis of the expan- sion of autophagic vacuole compartment and subcompart- ments induced by vinblastine (an in vivo segregation en- hancer), as well as their regression upon segregation-in- hibitor cycloheximide post-treatment. Premalignant tu- mour samples were taken at month 5, month 8 (nodules), month 10 and month 15 (adenomas) after initiation. In all these stages, a highly increased and varying autophagic capacity was found compared with the host tissue. The basal (non-stimulated) autophagic compartment was mea- surable only at month 5 and month 15, and its regression upon cycloheximide was consistent with increased basal autophagic activity. Compared with the host tissue, au- tophagic capacity profoundly decreased in the differenti- ated and anaplastic adenocarcinomas at month 20, when, surprisingly, cycloheximide was unable to inhibit segre- gation. Our conclusion is that down-regulation of the cy- cloheximide sensitive segregation and a partly compensa- tory up-regulation of an alternative pathway of segrega- tion might occur along with malignant transformation. Keywords Macroautophagy · Vinblastine · Nodule · Adenoma · Adenocarcinoma · Morphometry · Rat (Wistar) Introduction Autophagy, i.e. lysosomal degradation of endogenous cell constituents, is a complex cell function that has gained growing interest in recent years. One ubiquitous pathway of eukaryotic lysosomal degradation is macro- autophagy, which is at least a three-step process, starting with segregation of large cytoplasmic quanta by specific prelysosomal membranes (Réz and Meldolesi 1980; Kovács et al. 2000). The resulting autophagosomes will gain lytic enzymes later by fusion with other members of the lysosomal supercompartment, after which the segre- gated material undergoes autolysosomal degradation (re- views: Pfeifer 1987; Mortimore et al. 1989; Seglen and Bohley 1992; Dunn 1994; Blommaart et al. 1997; Kim and Klionsky 2000). Being an important pathway of the catabolic side of cellular steady state and growth regulation, autophagy is expected to change along with metabolic rearrangement during tumour progression. In general, autolysosomal proteolysis was found down-regulated in malignant (Gunn et al. 1977; Rønning et al. 1981a, 1981b; Baccino et al. 1984; Gronotajski and Pardee 1984; Tessitore et al. 1988; Houri et al. 1993; Kisen et al. 1993; Seglen 1997) and some premalignant (Ahlberg et al. 1987; Schwarze and Seglen 1985; Seglen 1997) cells in vitro. However, the nature of this regulational change and its timing in the tumour progression in vivo remained unclear. Eluci- dation of carcinogenesis-associated alterations of au- tophagy is expected to promote understanding of its nor- mal regulation, too. To this end, we analysed autophagic capacity and activity of rat pancreatic tumour cells at dif- ferent stages of chemical carcinogenesis. In our terms autophagic capacity can be characterised on the one hand This study was partly supported by a Copernicus contract (CIP- ACT 930210) from the European Union. S. Tóth · K. Nagy · Z. Pálfia · G. Réz ( ✉ ) Department of General Zoology, Loránd Eötvös University, Budapest, Pf 120, 1518, Hungary e-mail: grez@cerberus.elte.hu Tel.: +36-1-2090555 ext. 8640/8639, Fax: +36-1-3812184 Cell Tissue Res (2002) 309:409–416 DOI 10.1007/s00441-001-0506-7 REGULAR ARTICLE Szilveszter Tóth · Krisztina Nagy · Zsolt Pálfia Gábor Réz Cellular autophagic capacity changes during azaserine-induced tumour progression in the rat pancreas Up-regulation in all premalignant stages and down-regulation with loss of cycloheximide sensitivity of segregation along with malignant transformation Received: 19 March 2001 / Accepted: 4 December 2001 / Published online: 10 July 2002 © Springer-Verlag 2002