Retrovirus-mediated immunosuppression. II. FeLV-UV alters in vitro murine T lymphocyte behavior by reversibly impairing lymphokine secretion

Murine splenocytes and cloned murine T cells were used to study the in vitro immunosuppressive effects of UV-inactivated feline leukemia virus (FeLV-UV) on lymphokine secretion. FeLV-UV can significantly depress the accumulation of IL 2 in cultures of Con A-stimulated C57BL/6 splenocytes and in cultures containing the alloreactive helper T cell clone B6D/2-2m plus Con A. Inhibition of lymphokine accumulation in these cultures could not be attributed to absorption or inactivation of IL 2 by the FeLV-UV or to the FeLV-UV-induced production of substances which interfere with the IL 2 bioassay. Thus, FeLV-UV appears to block production and/or secretion of IL 2 by a direct inhibitory effect on IL 2-secreting murine T lymphocytes. Additional studies indicate that FeLV-UV impairs IL 2 production only if added very soon after lymphocyte contact with lymphokine-inducing agents and that IL 2 secretion resumes when FeLV-UV is removed from the culture. FeLV-UV also impairs accumulation of MAF (interferon-gamma?) in cultures of Con A-stimulated C57BL/6 splenocytes and in cultures containing the alloreactive cytotoxic T lymphocyte clone B6D/2-7c plus Con A. The latter observation again suggests that FeLV-UV impairs lymphokine secretion by a direct effect on lymphokine-producing T lymphocytes. Furthermore, it suggests that FeLV-UV does not selectively impair production of IL 2 nor does it have selective inhibitory effects on helper T cells. Rather, FeLV-UV appears to have a general inhibitory effect on lymphokine production by T lymphocytes. Finally, concentrations of FeLV-UV which suppress MAF production by the CTL clone have little influence on the cytolysis mediated by the same cloned T cell population. Thus, the immunosuppressive influence of FeLV-UV is selective for phenomena associated with induction of new T lymphocyte functions, such as lymphokine secretion, and spares other immune functions already expressed by the same cells.     

Differential distribution of antigen-specific helper T cells and cytotoxic T cells after antigenic stimulation in vivo. A functional study using limiting dilution analysis

We have developed modified limiting dilution analysis (LDA) techniques that distinguish in vivo Ag-stimulated murine helper T lymphocytes (HTL) and CTL from unstimulated precursor T cells, even those with the same Ag specificity. We refer to these cells that are detectable in the modified LDA as "Ag-conditioned" T cells (cHTL and cCTL). We have used the modified LDA techniques in conjunction with conventional LDA techniques (which enumerate all Ag-specific T cells) to evaluate the in vivo distribution of Ag-conditioned cHTL and cCTL following in vivo sensitization to alloantigens via sponge matrix or skin allografts. In general, we observed the following regarding the distribution of cHTL and cCTL: 1) Ag-conditioned HTL and CTL were detectable only after in vivo sensitization with alloantigen: 2) not all Ag-reactive T cells became conditioned T cells after in vivo Ag deposition; 3) the percentage of Ag-reactive T cells that converted to conditioned T cells after Ag deposition varied among different lymphoid compartments; 4) a high percentage of cHTL, but a low percentage of cCTL, accumulated in regional lymph nodes and spleen; 5) cHTL accumulated in peripheral blood, whereas cCTL did not; 6) Ag-conditioned cHTL were detectable in various lymphoid tissues for greater than 60 days following Ag deposition, whereas cCTL were detectable for only 14 to 20 days; and 7) unlike the other lymphoid sites, the site of Ag deposition accumulated a high percentage of both Ag-stimulated cHTL and cCTL. Furthermore, cHTL and cCTL appeared to reside in phenotypically distinct T cell subsets in that in vivo treatment with anti-L3T4 mAb abrogated the accumulation of HTL, but not CTL, at the site of Ag deposition. These data demonstrate differential compartmentalization of Ag-conditioned cHTL and cCTL subsequent to in vivo Ag deposition. The implications of these findings regarding the monitoring of in vivo immune responses are discussed.     

Lymphocyte entry into inflammatory tissues in vivo. Qualitative differences of high endothelial venule-like vessels in sponge matrix allografts vs isografts

Sponge matrix allografts and isografts become extensively encapsulated and neovascularized after s.c. implantation. Sponge allografts acquire alloantigen-reactive T lymphocytes, whereas sponge isografts fail to do so, even though these T cells are continuously circulating in the peripheral blood. We have investigated the possibility that the vascular endothelia regulates lymphocytic accumulation in sponge matrix implants. In normal lymph nodes, specialized high endothelial venules (HEV) regulate lymphocyte extravasation from the blood. We have now identified HEV-like vessels in sponge matrix allografts. These vessels are operationally defined as "HEV-like" in that they react with mAb MECA 325 which identifies murine HEV, and bind lymphocytes in ex vivo adhesion assays. In contrast, sponge isografts contain MECA 325 reactive vessels that are significantly smaller than those found in allografts. Further, vessels of sponge isografts do not readily bind lymphocytes in ex vivo adhesion assays. Immunohistologic analysis also revealed that the small MECA 325+ vessels present in sponge isografts are consistently found in close proximity to nerve bundles. Although this MECA 325 reactive vessel-nerve bundle association is also observed in sponge allografts, large MECA 325 reactive vessels are widely distributed in allografts. Our data suggest that small, poorly adhesive MECA 325 reactive vessels develop in sponge isografts and allografts, possibly under the influence of local nerve tissue. These vessels respond to regional alloimmune responses by developing into the larger HEV-like vessels capable of binding lymphocytes in sponge allografts. The value of this experimental system as an in vivo model to evaluate mechanisms involved in neovascularization and endothelial differentiation is discussed.     

Characterization of aarA, a pleiotrophic negative regulator of the 2'-N-acetyltransferase in Providencia stuartii.

We have utilized transposon mutagenesis to obtain insertional mutations in Providencia stuartii that activate the chromosomal aac(2')-la gene. Two closely linked mini-Tn5Cm insertions were obtained in a locus designated aarA, and a single insertion was obtained in a separate locus, aarC. Nucleotide sequence analysis, complementation studies, and localization of the sites of mini-Tn5Cm insertion have allowed the identification of the aarA coding region. The deduced AarA protein had a molecular mass of 31,086 kDa and displayed characteristics of an integral membrane protein. A strain deleted for the aarA gene by allelic exchange showed at least a fourfold increase in the accumulation of aac(2')-la mRNA and an eightfold increase in aminoglycoside resistance. Mutations in aarA were pleiotrophic and also resulted in loss of pigmentation and a deficiency in cell separation during division.     

Corticotroph,somatotroph and mammotroph cell kinetics in the postnatal infant female rat

The aim of this work was to detect if hypothalamic-pituitary maturation was accompanied by significant proliferation changes in differentiated pituitary cell pools. For this purpose, pituitary corticotroph (Ct), mammotroph (Mt) and somatotroph (St) proliferation activities were scanned in intact female rats during the postnatal (P) period (1–35 postnatal days). The techniques of tritiated thymidine labelling, immunostaining and autoradiography were combined to visualize DNA synthesis of hormone containing cells. Immunoreactive cell densities were measured using image analysis, and double labelled cells were counted. Corticotroph proliferation activity increased significantly on day P12, followed by an increase in the Ct proportion on days P13–14. This is the first observation of a spontaneous change of corticotroph proliferation at the end of the stress nonresponsive period. The mammotroph density and proliferation rate increased gradually during postnatal maturation, until the Mt pool overran other cell types of the female hypophysis on day 35. The somatotroph pool was the most numerous until day P20; the proliferation rate remained constant while St proportions increased reaching a plateau between days P13 and 20, then decreased to the adult level. Each cell type examined showed a characteristic, individual density and proliferation pattern.     

Stimulation of folliculogenesis in domestic cats with human FSH and LH

Folliculogenesis in response to exogenous stimulation by human urinary follicle stimulating hormone (huFSH) and human menopausal gonadotropin (hMG) was evaluated in the domestic queen (Felis catus). The role of LH and/or FSH in folliculogenesis was examined by measuring concentrations of estradiol 17beta (E(2)) and progesterone (P) in the serum. Additionally, changes in the number and size of follicles from before the administration of exogenous hormones to surgical oocyte collection were monitored. Findings indicated that in queens receiving huFSH or hMG followed by human chorionic gonadotropin (hCG) to induce ovulation, the numbers of follicles from 1 to 3 mm increase with statistical significance (P<0.005) from before the initiation of treatment to surgical collection of oocytes. Although E(2) concentrations in cats receiving hMG increased above baseline by the third exogenous hormone injection, mean E(2) concentrations did not increase in the groups that received both huFSH and hCG, or hCG only, until after the administration of hCG. This suggests that the exogenous administration of LH contained in both hMG and hCG was necessary for E(2) to rise to levels associated with estrus.