Abstract Spleens obtained from the common snapping turtle, Chelydra serpentina (Linné), have been analyzed for immunological competence in several in vitro systems. Spleen fragments can be maintained in tissue culture for several weeks under conditions permitting antibody formation. Such fragments can produce specific agglutinins to sheep or mouse red blood cells when presented with these antigens in vitro. Cells obtained from adult turtle spleens can elicit splenomegaly or hepatomegaly in vitro in a manner suggestive of a graft‐versus‐host reaction. Using these in vitro procedures, it has been observed that maturation of the immunological system of the turtle occurs several months after hatching, and that hibernating turtles are defective in immunological capacity. Histological studies indicate that the snapping turtle has cloaca‐associated lymphoid tissues similar to the bursa of Fabricius.
Pyrimidinones are low-molecular-weight compounds which are inducers of interferon in several animal species. They have established antiviral, immunomodulatory, and antitumor effects. Four pyrimidinones as well as another potent interferon inducer, polyriboinosinic-polyribocytidylic acid, and beta-interferon were tested for effects on growth of the transplantable mouse bladder tumor (MBT-2). The pyrimidinones 2-amino-5-bromo-6-phenyl-4(3H)pyrimidinone (ABPP) and 2-amino-5-bromo-6-(3-fluorophenyl)-4(3H)pyrimidinone (ABMFPP) significantly inhibited MBT-2 growth in a dose-dependent manner and with equal potency when injected i.p. every 4 days starting 1 day after tumor cell inoculation. Administration of ABPP p.o. was as effective as i.p. injections. Direct intravesical application of ABPP to transplantable tumors growing in the bladder may be more effective in inhibiting MBT-2 growth than the same dose introduced p.o. Although ABPP (100 mg/kg) has an inhibitory effect comparable to 5000 units of beta-interferon, both pyrimidinones even at 500 mg/kg were less inhibitory of tumor growth than 10 mg of polyriboinosinic-polyribocytidylic acid per kg. The pyrimidinones 2-amino-5-bromo-6-(2,5-difluorophenyl)pyrimidine-4(3H)one (ABDFPP) and 2-amino-5-iodo-6-(2,3-difluorophenyl)pyrimidin-4(3H)one (AIDFPP) were also of comparable potency in inhibiting MBT-2 growth and were more effective on mg/kg basis than both ABPP and ABMFPP. Treatment with ABDFPP or AIDFPP also resulted in long-term cures of up to 40% of mice. In this respect these latter two compounds were superior to treatment with 10 mg of polyriboinosinic-polyribocytidylic acid per kg, a treatment which reduced tumor size but had no effects on tumor incidence. The data suggest that tumors of bladder origin may be particularly sensitive to treatment with pyrimidinones.
Interferons (IFNs) have established activities as antivirals and inhibitors of viral and transplantable tumors. To establish whether IFNs or their inducers can affect induction of carcinogenesis in vivo, the bladder-specific carcinogen N-[4-(5-nitro-2-furyl)-2-thiazolyl]formamide (FANFT) was administered in the diet at 0.11 or 0.13% (w/w) to female C3H/He mice beginning at 7 weeks of age. Mice treated with the IFN-inducing bropirimine [2-amino-5-bromo-6-phenyl-4(3H)-pyrimidinone] i.p. twice a week for 14 weeks starting on day 30 of start of FANFT feeding developed fewer transitional cell carcinomas (TCC) than mice treated with the vehicle. Bropirimine (200 mg/kg twice a week) orally resulted in even greater effectiveness: 6 of 43 bladders with TCC for bropirimine-treated mice versus 24 of 39 for control glycine buffer-treated mice (P less than 0.01, x2 test). Mice treated i.p. daily on days 29 through 210 with 5,000 units of beta interferon (specific activity, 2.0 x 10(8) units/mg) had 0 of 15 TCC while control mice had 7 of 13 TCC (P less than 0.001). Bladders of untreated mice were also significantly heavier than those of beta interferon- or bropirimine-treated mice. This dose of IFN treatment was confirmed as effective in a second experiment, in which mice were treated daily on days 30-223 with 5,000 units alpha/beta interferon (specific activity, 1.2 x 10(7) units/mg). This resulted in 4 of 25 bladders with TCC versus 24 of 39 for control mice (P less than 0.001). A higher dose of IFN (50,000 units alpha/beta interferon daily) was toxic; 24 of 30 mice died within 2 months. IFN and an IFN inducer, bropirimine, inhibited development and progression of FANFT-induced bladder TCC in vivo and thus may have roles as chemopreventive modalities.
Abstract The effect of some steroids on the lymphoid differentiation of embryonic mouse thymus glands was studied. Most of the adrenal cortex hormones tested could inhibit lymphoid differentiation when applied in low concentrations to prelymphoid 13‐day glands. The most effective was corticosterone. Careful washing of the affected glands allowed them to develop normally into lymphoid organs. The application of a high dose of corticosterone produced an effect not reversed by thorough washing. Such permanently affected glands were restituted to lymphoid organs by fusion with 13 or 15‐day embryonic mouse liver or with 15‐day spleen. Lymphoid glands were much less sensitive to low concentrations of corticosterone than the prelymphoid glands. The same concentration of corticosterone was more toxic to more advanced fetal glands. The sex hormones tested were practically ineffective at comparatively high doses; very high concentrations were toxic however. Basophilic cells normally found in 12 and 13‐day embryonic thymus glands disappeared after corticosterone treatment even in glands which were still potentially capable of lymphoid differentiation suggesting that these cells are not stem cells.
The presence of a growing tumor can lead to a significant curtailment of a graft-versus-host reaction as measured by the ability of allogeneic spleen cells to induce a host vascular response. This interference with the normal pattern of immunological reactions may be a reason for the survival of tumors in an immunologically alien environment.
Abstract The anatomy of the ventral neck region of the scincid lizards Chalcides ocellatus and Scincus scincus is presented and is found to be similar to that of other lizards as described in the literature. The internal carotid artery arises by 3‐5 roots from the dorsal side of the ascending limb of the carotid arch. During its first part, the internal carotid artery is completely divided into two nearly equal channels. The carotid sinus is more complicated in Chalcides than in Scincus . In lizards, it may be homologous to the carotid labyrinth of fishes and amphibians. Around the origin of the internal carotid artery are two kinds of epithelioid cells scattered in the adventitial connective tissue: a‐ large cells with rounded, faintly stained nuclei, and little, clear cytoplasm; b‐ cells with small darkly stained nuclei. Both kinds of cells appear to represent different levels of secretory activity. The number of the large cells increases with greater complexity of the carotid sinus. The cells also increase in size and number during summer (sexual period); this is especially true in younger animals. The epithelioid cells are considered to be homologous to the carotid body of higher vertebrates. The carotid sinus and epithelioid cells together form a closely interrelated system which may be intermediate between the carotid labyrinth of fishes and amphibians, and the carotid body of birds and mammals.
A new and sensitive assay for the effect of intracutanous administration of immunocompentent lymphocytes into the skin of irradiated unimmunized mice is described. The assay, which we have termed lymphocyte-induced angiogenesis (LIA) involves enumeration of new vascular branches induced by the action of these competent cells. As is the case for the previously described normal lymphocyte transfer reaction, LIA is a manifestation of the graft-vs.-host reaction, as shown by experiments utilizing appropaiate genetic combinations. The reaction is dose-dependent, and within the dose range of 2 times 10 minus 5 -4 times 10-6 cells the mumber of vessels induced correlates with the mumber of immunocompetent cells injected. At these dose levels spleen, lumph node, and hydrocortisone-resistant thymocytes are effective; bone marrow and thymus cells are not. Spleen cells from nude mice are incapable of inducing LIA, while mitomycin-C and irradiated lymphocytes can initiate but not maintain the reaction. The relationship between lymphocyte-induced angiogenesis has been discussed as have the implications of these findings to delayed hypersensitivity, inflammation, and vascular pathology.
A monoclonal antibody has been prepared against rat angiotensin-converting enzyme (ACE). By selection for antibody binding to endothelial cells of bovine rather than rat origin we have obtained a reagent that has broad cross-species binding properties and that can at the same time serve as a useful marker for the surface of endothelial cells. The IgM-producing clone that we have established, alpha-ACE 3.1.1, has been grown in ascites form to yield ascites fluid that binds selectively to immobilized ACE at a greater than 1:10,000 dilution. By use of enzyme-linked immunosorbent assays, immunofluorescence histology, and flow cytometry, we have demonstrated the presence of ACE on endothelial cells of murine, bovine, and human origin. By means of a fluorescence-activated cell sorter (FACS-IV) we have been able to selectively isolate viable endothelial cells from a mixture of endothelial cells and fibroblasts. We believe the antibody will be useful not only for the selection and in vitro cultivation of endothelial cells but also as a tool for the identification and pharmacological study of ACE.
