Enterotropic mouse hepatitis virus infection in nude mice

The cause of emaciation and diarrhea in athymic nude mice was found to be hyperplastic typhlocolitis resulting from infection with enterotropic mouse hepatitis virus (MHV). The disease was reproduced in experimentally-inoculated nude mice using intestinal homogenates from affected mice and cell culture-derived virus. Material derived from an experimental mouse was passed into neonatal Swiss mice and caused acute typhlocolitis. Virus failed to grow in NCTC-1469 cells and 17Cl-1 cells, which are normally permissive for MHV, but grew to low titer in a mouse rectal carcinoma cell line, CMT 93. These results show that an enterotropic strain of MHV can cause chronic enteric disease in athymic nude mice. The pattern of infection differs markedly from the more common MHV wasting syndrome in nude mice caused by non-enteric strains of MHV.     

Tumorigenicity of persistently infected tumors in nude mice is a function of both virus and host cell type.

Although BHK-21 cells persistently infected with wild-type vesicular stomatitis virus (VSV) are sensitive to natural killer (NK) cells and do not form tumors in athymic nude mice, BHK-21 cells persistently infected with a previously isolated mutant virus (VSV-P) are resistant to NK cells and form tumors in nude mice. We used this VSV-P mutant to persistently infect HeLa cells and mouse tumor cell lines. A mouse mastocytoma line (P815) persistently infected with VSV-P was similar to BHK-21 cells in that it was resistant to NK cell lysis and formed tumors in nude mice. However, neither HeLa cells nor mouse myeloma lines persistently infected with VSV-P were resistant to NK cell lysis in vitro, and neither formed tumors in nude mice. Rejection by nude mice of HeLa cells and mouse myeloma cell lines persistently infected with VSV-P could be ablated by rabbit antiserum to asialo-GM1, implicating NK cells in the in vivo rejection of these persistently infected tumors. These results suggest that NK cell recognition and killing of virus-infected cells in vivo and in vitro depend upon genetic contributions from both the virus and the host cell.     

Establishment of a primary human sarcoma model in athymic nude mice

Improvement of soft tissue sarcoma patient outcome requires well-characterized animal models in which to evaluate novel therapeutic options. Xenograft sarcoma models are frequently used, but commonly with established cell lines rather than with primary human sarcoma cells. The objective of the present study was to establish a reproducible xenograft model of primary human soft tissue sarcoma in athymic nude mice. Primary soft tissue sarcoma cells from four resected human sarcomas were isolated, cultured until the third passage and injected subcutaneously into athymic nude mice. The sarcoma xenograft was further analyzed by histological and immunohistochemical staining. In two out of four sarcomas tumor growth could successfully be established leading to solid tumors of up to 540 mm³ volume. Histological and immunohistochemical staining confirmed the mouse xenograft as identical sarcoma compared with the original patient’s tissue. In the present study a reproducible xenograft model of primary human soft tissue sarcoma in athymic nude mice was established. This animal model is of great interest for the study of sarcomogenesis and therapy.     

Intracranial growth of pulmonary small cell carcinoma cells in nude athymic mice

Pulmonary small cell carcinoma of the lung cells were inoculated intracranially and subdermally in nude athymic mice (Balb/c strain). There was a 100% intracranial tumor incidence in response to 1/10th the cell dose required to produce subdermal tumors in immunosuppressed nude mice. Tumors did not grow in the brains of Balb/c normal (haired) mice.     

Neoplasms in NIH type II athymic (nude) mice

Necropsy and histopathology were performed over an 18-month period on 173 NIH type II athymic (nude) mice and 53 NIH type II mice heterozygous at the nu locus. A total of 149 mice were used in studies of tumor transplantation while 77 mice were screened as part of the quality assurance program for the colony. Twenty-nine neoplasms were found in 173 nu/nu mice. Only one neoplasm, an ovarian granulosa cell tumor, was found in 53 nu/+ mice. In nu/nu mice, there were nineteen lymphosarcomas, nine ovarian granulosa cell tumors, and one transitional cell carcinoma of the urinary bladder. A greater number of lymphosarcomas occurred in mice greater than 6 months old. A greater number of tumors, particularly lymphosarcomas, were found in nu/nu mice than in nu/+ mice.     

Changes in histological differentiation of human tumors transplanted to athymic nude mice: a morphometric study

Differing conclusions have been reached regarding the phenotypic stability of human tumors transplanted to athymic nude mice. Since previous conflicting studies of tumor histology have been largely subjective, quantitative methodology was applied to an analysis of 13 human adenocarcinoma tumor lines that were originally derived directly from surgical specimens. Glandular differentiation was quantitated, both in the original human tumor (OHT) and in a minimum of 6 serial passages of the nude mouse-grown tumors (MGT), by means of point counting. A significant change in differentiation was noted in 12 lines, with 9 showing a decrease. Variance from the OHT was most commonly noted in the initial MGT, but additional changes were also noted in 8 lines during subsequent passages. Most of the lines also showed increased necrosis in the MGTs. Since histological differentiation and necrosis are related to tumor aggressiveness, it would appear that the predominant tendency was to evolve toward a more malignant phenotype. These changes may mimic those seen in human tumor metastases.     

Human immunodeficiency virus (HIV)-infected tumor xenografts as an in vivo model for antiviral therapy: role of alpha/beta interferon in restriction of tumor growth in nude mice injected with HIV-infected U937 tumor cells.

The host factors involved in the restriction of tumor growth were studied in nude mice transplanted with a cloned line of chronically human immunodeficiency virus (HIV)-infected U937 cells. HIV-infected and uninfected U937 cells exhibited the same growth patterns in culture. However, HIV-infected cells were not tumorigenic when injected subcutaneously in nude mice, whereas large solid tumors were observed in mice injected with uninfected U937 cells. Injection of nude mice with antibody to alpha/beta interferon (IFN-alpha/beta) enabled HIV-infected U937 cells to grow progressively in approximately 90 to 100% of mice. HIV-infected U937 cells formed solid tumors in the majority (60 to 90%) of either immunosuppressed (splenectomized, irradiated, and anti-asialo-GM1-treated) or genetically immunodeficient (bg/nu/xid) nude mice. In mice treated with antibodies to IFN-alpha/beta with established HIV-positive tumors, a direct correlation was found between p24 antigenemia and tumor size. Treatment of established HIV-positive U937 cell tumors with human IFN-alpha or mouse IFN-alpha/beta resulted in a clear-cut inhibition of both tumor growth and p24 HIV antigenemia. In contrast, treatment with tumor necrosis factor alpha markedly inhibited tumor growth but did not significantly decrease serum p24 levels. 3'-Azido-3'-deoxythymidine treatment did not affect either tumor growth or the levels of serum p24 antigen. These data indicate that endogenous IFN-alpha/beta is a crucial factor in the restriction of both tumor growth and p24 antigenemia in mice injected with HIV-infected tumor cells. Moreover, the results suggest that the development of HIV-1 p24 antigenemia in athymic immunosuppressed mice may represent an interesting in vivo model for anti-HIV therapy.     

Cortisone-sensitive, innate resistance to Hymenolepis nana infection in congenitally athymic nude rats

The innate resistance of the unnatural rat host to the mouse tapeworm Hymenolepis nana is cortisone sensitive but thymus independent. When congenitally athymic nude rats were orally given eggs, cysticercoids, or adult worms of H. nana, no lumenal adults were established except when they were treated with cortisone acetate during the expected lumenal development. The effect of cortisone to promote adult maturation in the rats was compared in nude and normal rats given eggs of H. nana. The fecundity of the worms (assessed by the fresh worm biomass and the number of infective eggs produced) was much higher in cortisone-treated nude rats than in cortisone-treated normal rats. When the nude rats reconstituted with thymocytes were given eggs and treated with cortisone, the fecundity of H. nana dropped to the same level as in cortisone-treated normal rats. It is strongly suggested that the unnatural rat host has thymus-independent cortisone sensitive resistance to an initial infection (which is the main component of the innate resistance and blocks the lumenal establishment of this parasite) and thymus-dependent resistance (which suppresses the established worms' fecundity and may be ascribed to acquired resistance to the ongoing infection).     

Human lymphomas transplanted in nude mice

This study was carried out to determine the take rate of malignant lymphomas transplanted subcutaneously in nude mice. Lymphoid tissue from patients with a variety of lymphoproliferative and lymphomatous disorders was employed, but only four (8.3% of total) non-Hodgkin's lymphomas of various types grew in the nude mice. These transplants were from patients with lymphoplasmacytic, lymphoblastic, immunoblastic and centroblastic lymphomas; two of them were serially transferred. High grade malignant lymphomas seem easier to transplant than low grade ones. In each case, the tumor growing in nude mice was similar to the patient's tumor. In one case, metastases were found in mesenteric lymph nodes and lung. Electron microscopic study showed that neoplastic cells were associated with peculiar dark cells the nature of which is discussed. Furthermore, in two transplanted lymphomas, neoplastic cells contained C type virus particles, indicating contamination by the murine xenotropic type C virus.     

Characterization of implants from Dupuytren's contracture tissue into the nude (athymic) mouse

Dupuytren's contracture tissues were obtained from six patients as excess surgical material. Pieces of these tissues (a total of 38 implants) were placed into subcutaneous pockets in the suprascapular area of nude (athymic) mice. The objective was to determine whether the implant tissues would be maintained in the mouse with the characteristics of Dupuytren's tissue. The implants were removed for study at 14-179 days after implantation. Microvascular anastomosis between implant and host skin was established within the first 14 days. Histologic character and electron microscopic structure of the implants did not change during the course of the study. The implants became reduced in size with time. However, neither the spatial pattern of collagen nor the appearance of fibroblast cells changed. The original high levels of chondroitin-4-sulfate were significantly decreased in the 66- to 179-day postimplantation group, but were not significantly different from the values for normal fascial bands. The hyaluronic acid of the implants increased significantly with time of implantation, but never reached the level found in the normal fascial bands. The use of implants into nude mice may be useful for further experimental studies of Dupuytren's contracture.     

Immunological surveillance against DNA-virus-transformed cells: correlations between natural killer cell cytolytic competence and tumor susceptibility of athymic rodents.

Adenovirus type 2 (Ad2)-transformed hamster and rat cells are susceptible to lysis by natural killer (NK) cells from the host of origin and are nontumorigenic in immunocompetent hamsters and rats, respectively. These NK-cell-susceptible, virus-transformed cells are, however, highly tumorigenic in athymic (nude) mice--animals with intact NK-cell responses. In vitro lysis of these xenogeneic, Ad2-transformed cells by nude-mouse NK cells was found to be defective. In contrast, Ad2-transformed hamster and rat cells were highly susceptible to lysis by nude-rat NK cells. Furthermore, xenogeneic, Ad2-transformed hamster cells were nontumorigenic in nude rats unless the NK-cell responses of the challenged animals were compromised. The results of the nude-rat studies show that thymus-dependent, cytotoxic T-lymphocyte-mediated, host cellular immune responses are not essential for rejection of xenogeneic cells transformed by nononcogenic Ad2. The data suggest instead that immunologically nonspecific host cellular immune responses, such as those mediated by NK cells, are sufficient for rejection of Ad2-transformed cells. These results indicate that biologically important differences exist in the NK-cell-mediated defenses mounted by nude mice and nude rats against transformed cells that may account for the different patterns of tumor induction by various neoplastic cell types in these athymic animals.     

Use of athymic nude mice for in vivo studies of human growth-hormone-secreting pituitary adenomas.

Human growth hormone (hGH)-secreting pituitary adenoma tissue of 31 acromegalic patients was transplanted subcutaneously onto 291 athymic nude mice. 37% of the transplanted adenoma fragments could be maintained vital up to 46 days. Histological examinations of the transplants revealed neither alterations in their morphological characteristics nor signs of growth. A maintenance or linear decline of hGH secretion of the transplants related to their vitality was observed by hGH radioimmunoassay. Estimation of graft vitality was improved by GH-releasing hormone (GHRH) stimulation in regular intervals. The rate of pituitary adenomas responding to GHRH was as high as in a major collective of acromegalic patients. Our method of positive selection of vital xenotransplanted hGH-secreting pituitary adenomas via hGH detection at regular intervals in combination with GHRH stimulation gives the opportunity of reliable in vivo research with these tumors.     

Generation of cells with phenotypes of both intercalated duct-type and myoepithelial cells in human parotid gland adenocarcinoma clonal cells grown in athymic nude mice

A neoplastic epithelial cell line was established from nude mice tumors grown after transplantation of surgical specimens from a human parotid gland adenocarcinoma. This cell line, which had ultrastructure similarities to salivary intercalated duct cells, was found by immunohistochemical techniques to contain amylase, but myosin was not detected. Ultrastructurally, cells of an intermediate type between intercalated ductal and myoepithelial cells were found in the transplanted tumors. Moreover, the expression of myosin in addition to the presence of amylase was detected in the tumors. These findings indicate that some transplanted tumor cells appear to be differentiating towards myoepithelial cells.     

Papovaviral sialoadenitis in athymic nude rats

A wasting disease was found in 32 athymic nude rats. The rats had parotid sialoadenitis with intranuclear inclusion bodies in ductal and acinar epithelial cells. Other common lesions included bronchitis, bronchiolitis and secondary bacterial pneumonia. Less commonly, rhinitis and Harderian adenitis were seen. Intranuclear inclusions were also seen in bronchial epithelium of 1 rat, Harderian gland acini of 1 rat and laryngeal glands of 2 rats. Viral particles, averaging 45 nm in diameter, sometimes in crystalline arrays, were found in the nucleus of parotid epithelial cells. By the use of the avidin-biotin-peroxidase complex (ABC) immunoperoxidase technique, antibodies to disrupted SV40 virus (the group specific antigen of the polyomavirus (miopapovavirus) genus of the papovavirus family) reacted with intranuclear inclusions and cytoplasm of parotid epithelium and inclusions in lung and Harderian gland. The viral antigen did not cross react with antibodies to mouse polyoma, mouse K or disrupted bovine papilloma viruses.     

Impaired clonal expansion in athymic nude CD8+CD4- T cells

A comparative study of the phenotype and immune functions of highly purified CD8+CD4- T cells obtained from the spleen and thymus of normal mice and from the spleen of athymic nude mice was conducted. Of seven individual normal and nude mice examined, the range of V beta 8+ cells among CD8+ T cells was a heterogeneous 4.3 to 30.5% for athymic nude mice and a much more uniform spread from 14.7 to 18.5% for normal mice. In six of the seven nude mice examined, the fraction of V beta 8+ cells was below the lower limit of the V beta 8 distribution in normal mice. However, one of the seven nude mice contained nearly twice the percentage of normal V beta 8+ cells. A reduction in the density of V beta 8 as well as CD3 Ag expression was also observed in athymic CD8+CD4- cells although an Ly-6-linked Ag, B4B2 displayed a highly increased expression. Considering the battery of Ag analyzed in entirety, athymic CD8+CD4- T cells were clearly distinct from their "counterpart" CD8+CD4- T cells isolated from either thymus or spleen of normal (euthymic) mice. Anti-CD3-mediated triggering of the TCR:CD3 complex caused extensive clonal proliferation in cultures to which single responding CD8+ T cells had been deposited. Under identical conditions, however, anti-CD3 caused little, if any clonal expansion in CD8+ cells from athymic nude mice. Highly purified athymic CD8+CD4- cells produced readily detectable IL-2R expression and IL-2 synthesis and secretion upon stimulation by anti-CD3 and by Con A. Production of IL-2 by purified athymic CD8+CD4- cells was due to CD8+CD4- cells and not due to a minor population of contaminating CD8- cells as anti-CD8 + C treatment completely abrogated the ability of athymic CD8+CD4- cells to produce IL-2. Despite IL-2 production and IL-2R expression by athymic nude CD8+CD4- T cells in response to anti-CD3 and to Con A, an impaired proliferative response followed.     

Differentiation of human testicular embryonal carcinoma and teratocarcinoma grown in nude mice and soft-agar cultures

The differentiation pattern of two human germ cell tumors, grown in nude mice and in vitro is described. Tumor A was an embryonal carcinoma (EC) of borderline histology with characteristics of yolk sac tumor and of seminoma; tumor B was a teratocarcinoma with yolk sac elements and syncytiotrophoblastic giant cells. The morphology of an EC as well as cytogenetic characteristics were maintained during 20 passages in nude mice from tumor A and over 11 passages from tumor B. Tumor A did not grow in vitro. Cell suspensions prepared from xenografted tumor B grew into cystic embryoid bodies in semi-solid tissue culture medium. These embryoid bodies showed cuboidal and flattened cells with microvilli, junctional complexes, peripheral microfilaments, and annulated lamellae, reminiscent of the 'inner cell mass' of a blastula and of endoderm, respectively. When such colonies were transplanted into nude mice, however, only tumors with the morphology found in the transplants appeared.     

Tumor formation by SV40-transformed human cells in nude mice: the role of SV40 T antigens

Human cells transformed in vitro by SV40 rarely form tumors in nude mice. We examined whether these cells as a group are inherently nontumorigenic or whether they are potentially tumorigenic but rejected by the athymic host, possibly by nonspecific immune mechanisms. SV80 and NG8 are SV40-transformed human cell lines that express all of the transformed properties, including anchorage-independent growth, but do not form tumors in adult nude mice after injection of as many as 10(8) cells. Both the SV80 and NG8 cell lines have SV40-specific transplantation antigens which crossreact with those present on SV40-transformed (but tumorigenic) rodent cells. We found that SV80 cells, though not NG8 cells, induced progressively growing lethal tumors if the cells are injected repeatedly into neonatal nude mice. Somatic cell hybrids between SV80 or NG8 cells and a highly tumorigenic cell line derived from a human tumor continue to express the virus-induced antigens and fail to form tumors in adult nude mice. These results strongly suggest that at least for some SV40-transformed human cells, the failure to form tumors in nude mice may be due to their expression of virus-induced transplantation antigens rather than the absence of tumorigenic potential.     

Eradication of a large MOPC-315 tumor in athymic nude mice by chemoimmunotherapy with Lyt2+ splenic T cells from melphalan-treated BALB/c mice bearing a large MOPC-315 tumor

Summary We have previously shown that spleen cells from BALB/c mice that are in the process of eradicating a large MOPC-315 tumor following low-dose (2.5 mg/kg) melphalan (l-phenylalanine mustard) therapy are effective in preventing tumor progression upon adoptive transfer into BALB/c mice bearing a barely palpable tumor that had been treated with a subcurative dose of melphalan [Mokyr et al. (1989) Cancer Res 49: 4597]. Here we show that such spleen cells in conjunction with a subcurative dose of drug (adoptive chemoimmunotherapy, ACIT) can cause the complete regression of a large (15–20 mm) s.c. MOPC-315 tumor in a large percentage of T-cell-deficient (athymic nude) tumor-bearing mice. Spleen cells that were effective in ACIT of athymic nude mice displayed in vitro a substantial direct lytic activity against MOPC-315 tumor cells, and the lytic activity was greatly enhanced when the spleen cells were cultured for 5 days with or without mitomycin-C-treated MOPC-315 stimulator tumor cells. The cells responsible for the therapeutic effectiveness of the spleen cells in ACIT of athymic nude mice, as well as the cells responsible for the direct in vitro anti-MOPC-315 lytic activity of the spleen cells, were of the Lyt 2 and not the L3T4 phenotype. Most of the athymic nude mice that completely eradicated a large MOPC-315 tumor as a consequence of ACIT were capable of rejecting a challenge with 30–100 times the minimal lethal tumor dose for 100% of normal BALB/c mice administered more than 1 month after the ACIT. The ability of these athymic nude mice to resist the tumor challenge was associated with the presence of a greatly elevated percentage of cells expressing T cell surface markers in their spleens. Thus, it is conceivable that splenic Lyt 2⁺ T cells from melphalan-treated BALB/c mice bearing a large MOPC-315 tumor mediate their therapeutic effectiveness in ACIT of athymic nude mice bearing a large MOPC-315 tumor, at least in part, through direct cytotoxicity for MOPC-315 tumor cells. In addition, eradication of a large MOPC-315 tumor through cooperation between antitumor immunity and melphalan toxicity endues the athymic nude mice with an elevated percentage of T cells in their secondary lymphoid organs, and these T cells are probably responsible for the long-lasting protective antitumor immunity exhibited by these mice.Supported by research grant IM-435A from the American Cancer Society and research grant B-8806 from the Bane EstateIn partial fulfillment of the requirements for the Doctor of Philosophy DegreeRecipient of Career Development Award CA-01350 from the National Cancer Institute     

Dying and regeneration of human tumor cells after heterotransplantation to athymic mice

The histologic phenomena occurring immediately after heterotransplantation of two human colon adenocarcinomas to athymic mice have been studied. The tumors differed with respect to velocity of growth and passage age. Three phases were discernible in both cases. (1) During the first phase, most inoculated tumor cells died. (2) The second phase was characterized by removal of the necrotic tumor cells by immigrated inflammatory cells and by penetration of the connective tissue of host animals from peripheral into central areas of the implants. The first mitoses occurred within tumor cells in close proximity to these connective tissue septa. (3) During the third phase, signs of regeneration and proliferation of tumor cells resulted in the macroscopic enlargement of xenografts. Only in this phase, the typical histologic characteristics of the tumors were formed. These observations point to the host connective tissue invading into implants to be of great importance for the stimulation of tumor cell proliferation and, therefore, for the growth of xenografts. Thus, successful heterotransplantation is obviously based on mutual events between the transplanted tumor cells and host connective tissue.     

Collagenase inhibitors retarding invasion of a human tumor in nude mice

Tumor invasion has been correlated with the ability of tumor cells to produce collagenolytic enzymes which are capable of degrading normal host tissues. However, the human small cell carcinoma implanted subcutanouesly and growing progressively in athymic (nude) mice produced large quantities of collagenase but did not appear to significantly infultrate adjacent host tissue. In comparison, subcutaneously implanted murine Lewis lung tumors produced similar quantities of collagenase and were locally invasive. The human tumors were surrounded by a compact layer of fibroblast cells in a fibrous matrix. This fibrous sheath exhibited anticollagenase activity and indicated a mechanism of host tissue resistance to invasion via the formation of inhibitors to degradative enzymes produced by tumor cells.