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Experiments are described that characterize the nature of the stimulus leading to lymphocyte-induced angiogenesis (LIA), a reaction previously shown to reflect a local in vivo graft-vs-host reaction. The studies demonstrate that circulating cells of the host animals provide the stimulation essential for activation of donor lymphocytes and that the major allogeneic stimulus in congenic lines of mice is correlated with I-region disparity, primarily associated with IA-controlled determinants. The results are readily compartible with the hypothesis that is proposed that LIA is in large part the consequence of the release of soluble mediators or lymphokines that may act either directly on endothelial cells or indirectly by activating macrophages, which in turn generate the vascular reaction.

We have previously shown that tumor cells inoculated into the thoracic region of the trunk grow much more rapidly and with a higher frequency than do an equivalent number of cells injected more posteriorly. We thought that these results might have been a reflection of an underlying difference in the vascular system supplying these regions of the trunk. The present experiments which were undertaken to characterize various aspects of the vascular system associated with the transplantation sites used in these studies include measurements of rates of diffusion of salt solutions, of protein clearance, and of the retention of radio-labelled cells and cellular debris. We have examined local differences in temperatures and studied the effect of change of ambient temperature on those local differences as well as on tumor growth. More directly, we have measured blood flow to the capillaries by intracardiac administration of radioactive microspheres, and assessed the ability of inoculated tumor cells to induce angiogenesis. We have been unable to detect any major differences in the parameters of the vascular system that we have analyzed. Thus we cannot offer any satisfactory explanation for the observed four-fold differences in tumor growth in anterior vs posterior inoculation sites. The results are discussed in terms of underlying developmental gradients which can be invoked to provide a basis for further experimentation.

SummaryWe have previously shown that immunocompetent lymphocytes can induce angiogenesis following intradermal inoculation into allogeneic or semi-allogeneic host animals. Experiments are described demonstrating that preimmunization of donor animals leads to a specific increase in angiogenesis-inducing capacity of effector spleen cells, while preimmunization of host animals results in a selective, specific reduction in lymphocyte-induced vascular changes.The results are discussed in terms of the increasing evidence that lymphocyte-induced angiogenesis is mediated by soluble factors (lymphokines) released following lymphocyte activation.

Lymphocyte-induced angiogenesis (LIA) is a vascular response observed when allogeneic or semiallogeneic immunocompetent lymphocytes are inoculated intradermally into immunosuppressed or irradiated host mice. The reported experiments were carried out to characterize the effector cell population(s) responsible for causing LIA. Lyt 1.2, Lyt 2.2, and monoclonal Thy 1.2 antisera were used for negative selection with complement (C′) to investigate the ability of selected subsets of lymphocytes to evoke angiogenesis. Treatment of C57BL/6 spleen cells with either anti-Lyt 1.2 or anti-Thy 1.2 and C′ resulted in an almost complete abrogation of the LIA reaction. In contrast, depletion of Lyt 2+ cells, under conditions which fully abrogated their ability to generate cell-mediated cytotoxicity in allogeneic mixed leukocyte cultures, resulted only in a partial (45%) reduction in the induced vascular response. Synergistic interaction between cell preparations treated separately with either anti-Lyt 1.2 or anti-Lyt 2.2 serum was not observed. We conclude that (i) Lyt 1 + 2−T lymphocytes can induce a significant LIA reaction; (ii) lymphocytes resistant to negative selection with anti-Lyt-1.2 serum are incapable of inducing such a reaction; and (iii) Lyt 1 + 2+ cells directly or indirectly play an additional role in generating a maximal LIA response.

Spleens obtained from the common snapping turtle, Chelydra serpentina (Linne), 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.

Parathyroidectomy was fatal in Chalcides ocellatus. Animals died between 3 weeks and more than 3 months; the latent period ranged from 6 to 60 days depending on the temperature. Hyperexcitability, loss of appetite, tremors, tetany, and general weakness were followed by death. In Varanus griseus, the latent period extended from 21 hours to 7 days. Only those younger animals died which presented more severe symptoms. High temperature and loss of blood exaggerated the effect of parathyroid removal. Tremors and tetany were noticed in most skeletal muscles. Injection of 2 ml of 10% calcium gluconate solution intramuscularly almost instantaneously relieved all symptoms. In Varanus, plasma normally contained from 11.0 to 12.8 mg calcium per 100 ml, which decreased by nearly 50% in the parathyroidectomized animals. The plasma calcium level dropped rapidly during the first 24 hours; the decrease then became gradual, and finally insignificant. Unilateral parathyroidectomy had no detectable effect on the animals or on the plasma calcium.

1. Number, position, size, shape, and vascular bed of the parathyroid glands of Chalcides ocellatus and Scincus scincus are described. The collagenous capsule is pigmented in Chalcides and pigment-free in Scincus. The internal “capillary” network is more developed than the subcapsular. 2. Seasonal changes in the activity of the glands are noticed. From April to October, the glands are active having a compact appearance. In the remaining months, “foraminated”, i.e., partially active glands are found. The glandular tissue can rearrange itself according to the degree of activity, either in the form of rounded lobules or cellular cords. In winter, degeneration takes place and nearly half of the parathyroid tissue is destroyed. In Chalcides, this occurs in the centre of the gland while the rest of the cells show limited activity, because the animal is a non-hibernator. In Scincus, mostly the peripheral part degenerates while the rest of the gland appears foraminated, because the animal is a hibernator. The rebuilding of the glandular tissue presumably takes place by amitosis. Partial destruction of the parathyroid may be caused also during summer by starvation. 3. Large acini are rare in the parathyroid gland of Chalcides but are very frequent in that of Scincus. Their size differs greatly and their condition may indicate the degree of activity of the gland. Their presence might represent a phase of storage of the parathormone, or the colloid of the acini may be inert. 4. Glands of Chalcides individuals subjected for a long time to high temperature are large and show abnormally high concentrations of nuclei. Some of the glands show also degeneration in the centre.

WHEN tumour cells are inoculated into syngeneic mice their establishment and growth is subject to regulation by the host animal. We have shown this most effectively for the inoculation of a broad range of tumours into anterior compared with posterior portions of the trunk1. For example, when mammary tumour cells were injected intradermally into thoracic compared with lumbar regions of adult histocompatible mice, tumour growth anteriorly became detectable by more rapid palpation, and tumour growth was three to four times as rapid anteriorly as posteriorly. This difference between anterior and posterior injection sites also became manifest after subcutaneous inoculation; it was apparent after inoculation of a wide variety of tumour types (mastocytoma, melanoma, lymphoma, sarcoma, mammary adenocarcinoma); it was independent of the sex of the host animal; it was found in a variety of mouse strains; and it did not seem to be related to the immunocompetence of the host or to the immunogenicity of the tumour1,7. We have now determined that there are antero-posterior and dorso-ventral differences in the growth of tumour cells inoculated intradermally into adult histocompatible mice. Although we have no explanation for our results we suggest that the observed differences may reflect the existence of morphogenetic gradients reminiscent of those invoked to explain patterns of differentiation during ontogeny.

Interferon-alpha, interferon-beta, and interferon-gamma differ in their antiproliferative effects for several cell lines. Interferons were thus assessed for their activity in inhibiting proliferation of three renal cell carcinoma cell lines. The malignant epithelial phenotype of each of these cell lines was confirmed by electron microscopy, histology, karyotype and tumorigenicity. When compared on an anti-viral unit basis, naturally produced interferon-beta was more effective than natural interferon-alpha for all cell lines and clones. Proliferation of each of the cell lines was inhibited by interferon-gamma. In all cases, removal of interferons from culture media resulted in resumption of the rate of cell growth after a variable delay of 6-10 days. If the antiproliferative effects of interferons predominate in mediating tumor regression, clinical response may depend upon the type of interferon to which the tumor is exposed.

Human malignancies arise commonly after exposure to environmental carcinogens. Bladder cancer is a well-studied and excellent preclinical model for this process. Transitional cell carcinoma (TCC) of the lower urinary tract accounts for approximately 4% of human cancers. Most of the TCC arise in the urinary bladder, with a male:female ratio in the U.S. of about 3:1. Chemical carcinogen-induced mouse and rat tumors in situ and transplantable bladder cancer models, which mimic the pathogenesis of the human disease, have been developed (1–4). Neoplasms can be histologically graded and staged in a manner similar to that for human bladder carcinoma.

Primitive thymus stem cells were studied with regard to in vitro maturation, sensitivity to x radiation, sensitivity to corticosteroids, and development of thymocytes from nude embryos. Studies were also conducted on thymus helper cell function. (HLW)

SummarySpleen explants from hamsters kept at room temperature responded strongly when immunized with sheep red blood cells in vitro. Spleens from cold-adapted animals behaved essentially like those obtained from normal animals. In contrast, spleen explants obtained from hibernating animals tended to be less capable or incapable of giving a detectable response to the same antigen under identical conditons of cultivation. Restoration of competence was obtained by cultivation of spleen explants in combination with a number of other tissues.

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.