Robert G. McKinnell

Temperature-dependent metastasis of the Lucke renal carcinoma and its significance for studies on mechanisms of metastasis

SummaryMetastasis is temperature dependent in the renal adenocarcinoma of the North American leopard frog, Rana pipiens. Widespread, multiple, metastatic colonies occur in tumor-bearing frogs kept at 28°C for 50 days while tumor-bearing frogs kept at 7°C for 98 days or more have either no secondary deposits or they have only an occasional small metastatic nodule. An attractive aspect of the frog tumor is that invasion and metastasis can be permitted or inhibited by the manipulation of temperature alone-no exogenous chemicals or drugs are required for the effect. Because of this, biological variables which reproducibly and specifically associate with metastasis permissive conditions when ambient temperature is cycled between permissive and inhibitory values are strong candidates for being causal elements in the multistep process leading to metastasis.Intravascularly injected labelled renal tumor cells reached all organs studied in as little as 15 minutes at both metastasis restrictive and permissive temperature. The results with tumor cell inoculation dispose of the possibility that failure of metastasis in chilled animals is due to cold-induced changes in blood flow. Histologically typical metastatic colonies developed in frogs, kept at the permissive temperature, after injection with disaggregated tumor cells which were previously cryopreserved.Frog tumors elaborate type I collagenase in a temperature dependent manner. Type IV collagenase has been demonstrated as well. Tumor cell detachment in vitro, assembly and disassembly of tumor cell cytoplasmic microtubules, and invasion in vitro, are all temperature dependent.

Genomic studies of the Lucké tumor herpesvirus (RaHV-1)

Abstract Ranid herpesvirus 1 (RaHV-1) is the etiological agent of the Lucké renal adenocarcinoma of the North American leopard frog Rana pipiens. Construction of cosmid libraries containing RaHV-1 DNA inserts allowed the derivation of a BamHI map for the viral genome. Summation of fragment sizes indicates that the genome is 217 kbp in size, a value in accordance with the most recent published estimate (220 kbp) obtained by field-inversion gel electrophoresis. The DNA sequence of the 39,757-bp insert in 1 cosmid (cos54) was determined and was predicted to contain 21 complete and 3 partial genes. In all, 12 genes have distant counterparts in a fish herpesvirus (ictalurid herpesvirus 1) and are present in 2 blocks, 1 of which is relatively inverted. This indicates that RaHV-1 belongs to the fish virus lineage of the herpesvirus family rather than to the lineage populated by mammalian and avian viruses. The remainding 12 genes in cos54 lack counterparts in any other herpesvirus. One of these encodes a putative DNA (cytosine-5) methyltransferase. This raises the possibility that biological processes induced in the host by RaHV-1 might involve methylation of cellular DNA by the viral enzyme.

Luckeé renal adenocarcinoma, an anuran neoplasm: Studies at the interface of pathology, virology, and differentiation competence

The northern leopard frog, Rana pipiens, is vulnerable to a herpesvirus‐induced renal tumor. The Lucké renal adenocarcinoma is metastatic as a function of temperature. The cloning procedure of nuclear transplantation has been used to study the differentiation potential of the tumor genome. This paper summarizes current studies of the pathology, virology, and differentiation competence of the Lucké tumor. J. Cell. Physiol. 173:115–118, 1997.

The influence of glucocorticoids on survival and growth of allografted tumors in the anterior eye chamber of leopard frogs.

Abstract The Lucke renal adenocarcinoma of northern leopard frogs, Rana pipiens , can be propagated by culturing it in the anterior eye chamber (AEC). In this so-called immunologically privileged site, however, only about 22 percent of first-set tumor alloimplants and 5 percent of second-set alloimplants survived for more than 40 days. The mean time of onset of first- and second-set rejections was about 20 and 13 days. Thus while tumor rejection in the AEC occurred more slowly than rejection of allografted tumor or skin at other sites, implantation in the AEC does not promote permanent survival. The glucocorticoids, corticosterone (500 ug/20 g) and aldosterone (50 ug/20 g) enhanced survival and growth of tumor alloimplants in the AEC. Seventy-three percent and 78 percent of tumor implants survived more than 40 days in frogs treated with aldosterone and corticosterone, respectively. Enhanced growth and survival of tumor implants in steroid-treated animals was thought to be due to a lympholytic action of the hormones. However, the number of circulating small lymphocytes in steroid-treated frogs was not significantly different from that of uninjected or solvent-injected frogs after six weeks of continuous treatment.

Invasion in vitro by explants of Lucke renal carcinoma cocultured with normal tissue is temperature dependent.

Fragments of renal carcinoma of the northern leopard frog, Rana pipiens, were cocultured in vitro with small pieces of tadpole heart, frog heart and frog kidney with gyrotory shaking for up to 14 days at 21°C and 28°C. No invasion by renal carcinoma occurred in confrontation cultures at 21°. However, the three normal tissues were invaded by renal carcinoma in confrontation cultures incubated at 28°C. Invasion in vitro by histologically typical renal carcinoma is thus similar to temperature-dependent invasion by the renal carcinoma-derived cell line PNKT-4B and affords an opportunity for the identification of cell or biochemical events which may be activated at invasion-permissive temperature. Cell or biochemical events which are selectively activated and subsequently repressed as the renal tumor is incubated at invasion-permissive and invasion-restrictive temperatures become significant candidates as events involved in, or causal for, malignant invasion.

Lucke Tumor of Frogs

The Lucke renal adenocarcinoma of the Northern leopard frog, Rana pipiens, has been studied more than any other neoplasm that afflicts amphibians. Abundant scientific literature relates, in part, to the biology of the species and is, in part, due to the biology of the tumor.

Continued diminished prevalence of the Lucke renal adenocarcinoma in Minnesota leopard frogs

No Luck? renal adenocarcinomas were detected at autopsy of 1216 northern leopard frogs (Rana pipiens) collected in Minnesota during the spring and autumn of 1978 and the spring of 1979, seasons in which the tumor was formerly obtained with a high frequency. The prevalence of renal tumors has been declining in Minnesota since at least 1966, and R. pipiens has become progressively scarce in Minnesota since that time. We suggest a causal relationship between reduced frog populations and decreased tumor prevalence.

Temperature-dependent malignant invasion in vitro by frog renal carcinoma-derived PNKT-4B cells.

The northern leopard frog,Rana pipiens, may be afflicted with a herpesvirustransmitted renal carcinoma which has the interesting property that its metastatic behavior is temperature-related. PNKT-4B is a cell line derived from a pronephric carcinoma arising in a tadpole. We sought to ascertain if invasion of normal tissue by PNKT-4B cells in three-dimensional confrontation culturein vitro is similarly temperature-dependent. Normal fragments of tadpole and frog organs are invaded by PNKT-4B cells at 28°C but not at 20°C or 21°C. PNKT-4B cells failed to invade tadpole tissues at 7°C. A temperature critical for invasion was sought. Temperatures of 21°C and cooler are invasion-restrictive and 23°C and warmer are invasion-permissive under the conditions of this study. Identification of a critical permissive temperature allows for the characterization of biochemical events which may be activated at the same temperature. The biochemical changes, which are selectively activated and subsequently repressed as tumor cells are cycled through invasion-permissive and invasion-restrictive temperatures, become compelling candidates as reactions involved in. or causal for, malignant invasion.

Cytogenetic analysis of triploid renal carcinoma in Rana pipiens

To provide a cytogenetic marker for nuclear transplantation experiments, triploid Rana pipiens embryos were produced. These embryos were injected with Lucké tumor herpesvirus. The chromosome profile of a renal carcinoma that developed in one of these triploid embryos was compared to the chromosomal profiles of a naturally occurring diploid renal carcinoma and a diploid renal tumor maintained as serial anterior eye chamber allografts for over 7 yr. Examination of Ag-NOR-stained chromosome spreads from the putative triploid and naturally occurring putative diploid tumor revealed the expected results. The vast majority of the chromosome spreads, 54/57 (95%) and 6/7 (86%), respectively, displayed euploid chromosome and Ag-NOR profiles: 3N = 39 with three Ag-NORs at the secondary constrictions in the long arm of chromosome 10 (10q) and 2N = 26 with two Ag-NORs in 10q. Chromosome profiles from the long-term allografted tumor were highly aneuploid (82%) and, based on their Ag-NOR content, displayed variations in their 2N, 3N, and 4N numbers. These data indicate that the majority of recently transformed triploid Lucké tumor cells can provide donor nuclei suitable for the characterization of developmental potential.

Are Renal Carcinomas Increasing in Rana pipiens after a Decade of Reduced Prevalence

-We report finding 12 Lucke renal carcinomas from autopsy of 409 frogs (2.9%) since 30 January 1987 to the present date (6 April 1989). While tumor prevalence remains low, tumors appear more abundant now than they have been in the recent past. Between 1977 and 30 January 1987, only one tumor was found at autopsy of 4532 frogs.