Hyaluronan and hyaluronidases have been proposed to be involved in tumor angiogenesis and invasion. Three hyaluronidases, HYAL1, HYAL2 and HYAL3, are located at the chromosomal region 3p21. In most small cell lung cancer (SCLC) lines the 3p21 region is part of a homozygote or heterozygote deletion. Gliomas are known to exist in a hyaluronan rich environment and express high levels of the hyaluronan receptor CD44. In a panel of SCLC and glioma cell lines the expression of HYAL1, HYAL2 and HYAL3 mRNA was examined. It was observed that the cell lines differed in their ability to splice out a retained intron in the 5' UTR of HYAL1 mRNA. A correlation seems to exist between the ability to splice out the retained 5' end intron of HYAL1 mRNA and the general hyaluronidase activity. In one cell line a substantial part of the hyaluronidase activity was abolished by immunoprecipitation of Hyal1, which strongly indicates that Hyal1 is the principal hyaluornidase in the examined cell lines. During severe hypoxia a significant reduction in both hyaluronidase mRNA and protein activity was found. These results support the theory of involvement of hyaluronidase in the angiogenic and invasive front of tumors.