Natural killer (NK) cells are 'spontaneously' cytotoxic cells thought to be involved in surveillance against tumour cells, rejection of virally infected cells, and regulation of haematopoietic stem cell differentiation and antibody synthesis. Fetus-derived alpha-fetoprotein (AFP) has been shown to regulate certain T cell-mediated immune reactions in vitro and in vivo. The lack of NK activity in newborn mice with high endogenous levels of AFP, together with the presence of cells expressing NK surface markers, also suggests that AFP may regulate NK activity. In this study we compared the effects of AFP on spontaneous versus activated murine NK activity. The lytic ability of both freshly prepared splenic NK cells and those arising after incubation for 24 h with interferon, Poly I:C, or T-cell growth factor (TCGF) was not affected by AFP if the latter was present only during the killing phase. However, if AFP was added at the beginning and retained for the duration of the 24-h in vitro lymphokine stimulation, the subsequent NK activity induced by interferon, Poly I:C, and TCGF was found to be significantly suppressed. This inhibition is both dose- and time-dependent. Delayed addition experiments showed that when AFP is present during the first 6 h of in vitro stimulation it will suppress interferon and TCGF-boosted NK activity by 50-80%. The AFP-mediated inhibitory effect on lymphokine-stimulated NK activity is not the result of increased death of effector cells nor, in the case of interferon and polyribonucleotides, of non-specific binding of AFP to the enhancing agents. In vivo injections of Poly I:C or TCGF failed to increase neonatal NK function, while administration of interferon did cause slightly higher levels of NK activity. However, spleen cells from newborn animals cultured for 24 h in the presence of lymphokines resulted in markedly elevated NK function and this in vitro activation could be suppressed by purified fetus-derived AFP. Thus, the in vivo pattern of NK activation in newborns with high endogenous levels of AFP was very similar to that of adult NK stimulation in vitro when exogenous AFP was added.

T-cell growth factor (TCGF) is a protein that is required for the continuous proliferation of activated normal T lymphocytes. It is produced by a subset of T lymphocytes upon appropriate stimulation. A human leukemic T-cell line (Jurkat) can be induced with the lectin phytohemagglutinin and the phorbol ester tetradecanoylphorbolacetate (TPA) to produce T-cell growth factor. This production was enhanced by including the lymphokine interleukin 1 in the induction medium. Interleukin 1 alone did not substantially increase T-cell growth factor production by cells treated only with phytohemagglutinin. These effects were preceded by and correlated with the induction of T-cell growth factor mRNA. Northern blot experiments with cloned TCGF DNA as a probe showed that TCGF mRNA was induced rapidly in cells treated with TPA and phytohemagglutinin, and this induction was augmented by interleukin 1. Thus, the production of T-cell growth factor was regulated at the level of its mRNA. Nuclear transcription experiments suggested that the TCGF gene was more actively transcribed in cells treated with TPA and phytohemagglutinin than in cells treated with phytohemagglutinin alone. The transcription of the TCGF gene was further increased when interleukin 1 was included along with TPA and phytohemagglutinin. When continued synthesis of RNA in induced cells was blocked with actinomycin D and cells were subsequently cultured in the presence or absence of inducing agents, the steady-state levels of TCGF mRNA declined in all cultures. This decline was roughly equivalent in cells incubated without the inducers and those incubated with phytohemagglutinin.(ABSTRACT TRUNCATED AT 250 WORDS)

Spleen cells of BALB/c mice that had been inoculated with syngeneic plasmacytoma MOPC 104E were cultured for 11 days in T-cell growth factor (TCGF) and ultrasonicated tumor extract (USE). Cultured lymphocytes (MOPC-CL) possessed three-fold more lytic units than normal spleen cells cultured in TCGF without USE (N-CL). Moreover, the in vivo neutralization assay suggested that MOPC-CL were composed of at least two populations, one possessing tumor-specific and the other nonspecific antitumor activity. When 2 X 10(7) of MOPC-CL were administered IP to mice that had been inoculated IP with 10(5) MOPC 104E cells 5 days previously marginal prolongation of survival was observed. This effect was not augmented by the single injection of a larger number (5 X 10(7] of CL, but was augmented by the repeated daily administration for 4 days (from day 5 to day 8 after the inoculation) of the same total number (5 X 10(7] of CL. In addition, IP injection of the streptococcal preparation OK432 before the transfer of CL significantly enhanced the therapeutic efficacy, and resulted in a cure rate of 20%. The mechanism of this combined effect appears to involve the effect of OK432 on interleukin 2 (IL-2) regulation systems in vivo. Our culture system with TCGF and USE and our therapy system with OK432 and CL allow the clinical application of adoptive immunotherapy for the many types of solid cancers.

Human T cell lymphocyte lines, established from lymphoid tissues from patients with adult T cell malignancies infected with the human T cell lymphoma virus (HTLV), were used in co-culture experiments to infect newborn cord blood lymphocytes (CBL). The infected and non-infected CBL cell lines were typed for HLA alloantigen determinants and tested for cell surface antigens using selected monoclonal antibodies. Infected cord blood lymphocytes showed inappropriate expression of alloantigenic determinants of the HLA-A and -B alleles. The monoclonal antibody 4D12, detecting an antigen common to the HLA-B5 cross-reactive group, was reactive with all infected cultures; this determinant appeared de novo in CBL cells lacking B5 cross-reactive group antigens in the uninfected state, and it increased in density in the infected cultures where the HLA-B5 cross-reactive alloantigens were present in uninfected CBL. HLA-DR was expressed in low levels or not detected on non-infected cultured cord blood lymphocytes and was present on all infected cells. OKT4 positive cells predominated in infected cultures, whereas OKT8 positive cells decreased in number or were absent. The TCGF receptor also increased both in density and in percent positive cells in all infected cell lines. The results suggest that HTLV may be tropic for a subset of T cells expressing mature T cell markers and that viral infections directly affect expression of cell surface antigens controlled by the human major histocompatibility complex (MHC).

We have recently developed a technique for direct expansion of human T lymphocyte clones from cerebrospinal fluid (CSF) of patients with acute infections of the central nervous system (CNS). In the present study, T lymphocyte clones were established directly from the CSF of 4 patients with multiple sclerosis by limiting dilution in the presence of T cell growth factor and irradiated feeder cells. In 3 patients the CSF was obtained during an exacerbation of their disease. Cloning efficiencies ranged between 4 and 6%. About 40 clones per patient were available for surface marker analysis and functional studies. Typing of the clones for membrane antigens revealed the following results: 75-100% had the OKT4+8- and 0-25% the OKT4-8+ phenotype. Only one clone expressed both surface markers. When tested for PHA-dependent cytotoxicity, all OKT8+ clones and about 50% of the OKT4+ clones were found to express cytotoxic activity. Studies on the proliferative response showed that all OKT4+ and the majority of OKT8+ clones were capable of TCGF-independent, mitogen-induced proliferation. Screening of the clones for specific reactivity against a panel of antigens including measles virus, mumps virus, Epstein-Barr virus and myelin basic protein (MBP) did not reveal significant specific reactivity.

Utilizing a 'sandwich' ELISA assay we have been able to demonstrate that mAb W6/32, B1G6 and IL-A19 are reactive with three different monomorphic determinants on bovine class I major histocompatibility complex (MHC) molecules. Sequential immunoprecipitations performed with the mAb revealed that class I molecules on PBM comprise a single population with respect to reactivity with the mAb in that the beta 2m-associated proteins bear all three epitopes. By contrast, TCGF-driven lymphoblasts and cells transformed by Theileria parva (Tp) additionally express molecules of Mr 45000 bound to beta 2m which are recognized by mAb B1G6 and IL-A19 but not by W6/32. These two subclasses of molecules were further distinguished on the basis that, when tunicamycin was added to cultures in the preparation of cells for analysis, mAb W6/32 precipitated class I heavy chains of Mr 39000 while the extra molecules detected only by mAb B1G6 and IL-A19 were of Mr 37000 and 39000. On thymocytes, the mAb W6/32-non-reactive class I molecules are present in low amounts and are expressed by cells in the medulla area, unlike BoT1 (analogous to human CD1) molecules which are expressed by the cortical cells. Our studies also revealed that the supposed beta 2m-specific mAb B1G6 does not recognize the beta 2m-associated molecules (BoT1) precipitated by mAb TH97A and thus the specificity of mAb B1G6 in cattle is for an epitope on bovine beta 2m which is strongly influenced by the nature of the heavy chain with which the beta 2m is associated.

In previous studies, the syngeneic MLR of peripheral T cells was shown to be predominantly an I region-restricted function. In this report we show that adult thymocytes are also capable of responding to syngeneic irradiated stimulator cells in a syngeneic MLR, provided that TCGF is added to the culture system. Using this assay, it was possible for the first time to examine the pattern of I region restriction within the thymus itself. Analysis of the thymocyte syngeneic MLR in thymuses from radiation-induced bone marrow chimeras demonstrated that the MHC preference seen in the peripheral T cell population also existed in cells resident within the thymus. Experiments utilizing congenitally athymic mice transplanted with allogeneic thymic grafts demonstrated that both peripheral T cells and thymocytes from such animals displayed a strong preferential proliferation toward stimulator cells bearing thymic-type MHC determinants. The results in the nude model thus demonstrate that the thymus by itself is sufficient to impart such restriction specificity on a developing T cell repertoire. These results are consistent with the notion that the thymus exerts selective pressure on maturing T cell populations that results in a skewing of the T cell repertoire toward the recognition of thymic-type I region products, and that this MHC preference exists before expansion of T cells in the periphery.

The lymphocyte transformation responses to mitogens (phytohaemagglutinin (PHA), concanavalin A (Con A), and pokeweed mitogen (PWM)), allogeneic cells, and the antigen-purified protein derivative (PPD) were studied in six acquired immunodeficiency syndrome (AIDS) patients and in six healthy controls, each of whom was HLA-DR- and mixed lymphocyte culture (MLC)-identical with one of the AIDS patients. No evidence of suppression was observed when irradiated or non-irradiated AIDS peripheral blood mononuclear cells (PBMC) were added to cultures of HLA-DR-identical PMBC from healthy controls stimulated with the strong mitogens PHA and Con A or with allogeneic cells, but suppression may be involved in the decreased responses in cultures stimulated with PWM or PPD. Addition of supernatants from macrocultures of AIDS cells did not suppress responses of control PBMC. Thus, suppression by any lymphocyte subset or soluble factor alone cannot explain the generally severely depressed transformation responses in AIDS. Addition of heavily irradiated HLA-DR-identical PBMC from healthy controls or supernatants from these cultures led to increased responses in cultures of mitogen-stimulated AIDS PBMC and in some cultures of antigen or allogeneic cell-stimulated AIDS PBMC, which were of the same magnitude as seen after the addition of commercially obtained T-cell growth factor (TCGF). This indicates that AIDS cells are deficient in producing TCGF. Heavily irradiated AIDS PBMC were capable of restoring the transformation responses to mitogens and antigens of purified HLA-DR-identical normal T cells, indicating that AIDS cells have a normal antigen-presenting capacity and interleukin (IL-1) production. However, AIDS PBMC had a very poor capacity to stimulate normal PBMC in MLC. Together, our experiments suggest that the immune deficiency in AIDS cells may be partially due to a decreased capability of T lymphocytes to produce TCGF and that a decreased number and/or function of dendritic cells may also be involved.

The existence and characteristics of bone marrow T-cell progenitors have not yet been established in man. Several pieces of evidence such as the reconstitution of certain immunodeficiencies by bone marrow graft suggest that T-cell precursors are present in the bone marrow. We report the growth of T-cell colonies from bone marrow populations using PHA-stimulated lymphocyte-conditioned medium containing T-cell growth factor (TCGF). Rosetting experiments and complement-dependent cytotoxicity assays with monoclonal antibodies indicate that the bone marrow T colony-forming cells (T-CFC) are E- OKT 3- and la+, i.e., immature progenitors. The colonies derived from these cells have the phenotype of mature T cells: E + OKT 3 + la- with either helper (OKT 4+) and suppressor (OKT 8 +) antigens. These results suggest that a thymic microenvironment may not be necessary for the in vitro proliferation and differentiation of the T-cell lineage in adult humans. These methodologies may permit direct investigation of early phenomena concerning the T-cell lineage, such as the acquisition of self-tolerance, the formation of a repertoire of specificities, and the HLA restriction phenomena that we believe takes place before the thymic maturation.

Human T-cell leukemia-lymphoma virus (HTLV) is a type C retrovirus associated with a subtype of mature T-cell malignancy in humans. HTLV also infects normal human cord blood mature T lymphocytes in vitro and induces a number of phenotypic changes in these cells, including their continuous growth and partial or complete independence of T-cell growth factor (TCGF). As part of our initial study designed to analyze gene(s) specifically activated by HTLV infection, we have isolated a recombinant DNA clone by differential screening of a cDNA library made from mRNA of a human T-cell lymphoma cell line producing HTLV. This cDNA identifies a single-copy gene in all human DNAs and a single mRNA species of 2.3 kilobases expressed at several hundred copies per cell in five HTLV-positive neoplastic T-cell lines. In addition, cord blood T lymphocytes infected with HTLV, but not the uninfected counterparts, express high levels of mRNA from this gene. A survey of different human hematopoietic cell types showed that this gene is expressed at low or undetectable levels (less than 10 copies) in human T, B, myeloid, or erythroid cell lines; in moderate amounts in lymphoid precursor (immature) cell lines; and in high amounts in lectin-activated mature T-cells, comparable to those of HTLV-infected T-cell lines. The precise function of this gene has not yet been determined.

Alloantigen-specific T cell precursors present in spleen cell populations obtained from parental mice that were rendered tolerant by the neonatal injection of F1 spleen cells were quantitated by clonal analysis, i.e., limiting dilution techniques. The results demonstrate that the frequencies for cytolytic as well as TCGF-producing (helper) and MAF-producing T cell precursors were dramatically reduced in tolerant spleen cell populations. The unresponsive state was antigen specific and could not be transferred in vitro in cell mixing experiments. Therefore, clonal deletion (or inactivation) would appear to represent a major mechanism for the observed tolerant state in this model system.

In an attempt to better define the mode of action of TCGF we have investigated the effect of complete removal of TCGF on the cell cycle kinetics of a TCGF-dependent murine CTL cloned line (B6.1). Cells that had been cultured in the presence or absence of TCGF were stained with propidium iodide, a DNA binding dye, and the distribution of DNA content was analyzed by flow cytometry. Results indicated that the proportion of cells entering the S phase of the cell cycle started to decrease 6 hr after removal of TCGF from the culture medium. This fraction continued to decrease as a function of time, and after 30 hr without TCGF 93% of the cells were in the G1 phase of the cell cycle in comparison to 38% in control cultures containing TCGF. Direct evidence that B6.1 cells completed one cell cycle after removal of TCGF before they accumulated in the G1 phase was obtained with a technique combining the use of BUdR, a thymidine analog, and the DNA binding dye Hoechst 33342. When TCGF was added again to cultures of B6.1 cells that were arrested in the G1 phase, the majority of cells entered the S phase in a synchronous fashion after a lag phase of 10 to 12 hr. The duration of this lag phase was independent of the concentration of TCGF used to restimulate quiescent B6.1 cells. However, the number of cells entering S phase 10 hr after re-addition of TCGF was concentration dependent. Additional experiments indicated that the cytolytic activity was not dependent on TCGF, since no decrease in the activity of B6.1 cells was noted after these cells had been deprived of TCGF for up to 30 hr.

The requirement for the signals in induction of cytolytic T lymphocytes (CTL) has been investigated. C57BL/6 X CBA/T6 F1 spleen cells stimulated with the lectin leukoagglutinin (L-A) failed to show CTL activity in a PHA-facilitated assay, although L-A-activated splenic T cells were able to respond to T cell growth factor (TCGF). Concanavalin A (Con A) on the other hand was able to induce cytolytic activity from CTL-P, as well as to render splenic T cells responsive to TCGF. Furthermore, L-A-activated splenic T cells could generate cytolytic activity upon subsequent culture in secondary mixed leukocyte culture supernatant (2 degrees MLC SN). In contrast, EL-4-derived SN (EL-4 SN) was unable to induce cytolytic activity from L-A-activated spleen cells. In addition, proliferation of L-A-activated spleen cells cultured in EL-4 SN was similar to those cultured in 2 degrees MLC SN. Nonactivated spleen cells were totally unresponsive to both SN in proliferation and generation of CTL. Analysis of T cell clones for the production of a factor necessary for induction of cytolytic activity revealed that both cytolytic and noncytolytic T cell clones were able to produce a factor(s) for the generation of cytolytic activity from L-A-activated T cells. On the other hand, SN from certain antigen-stimulated T cell clones produced factors capable of inducing cytocytic activity by L-A-activated T cells only in the presence of EL-4 SN. Neither EL-4 SN nor cloned T cell SN alone had such a capacity. The nature of the necessary lymphokines in the SN from the clone cells or from the EL-4 is unknown. In the case of the EL-4 SN, it is not known whether the presence of TCGF plays a role or whether that role is perhaps more differentiative than proliferative. This study provides evidence that the induction of CTL from CTL-P can be dissociated into activation, which is required to render T cells responsive to second signals, and differentiation, which is mediated by two different factors.

The present study investigates the regulatory effects of glycosaminoglycans such as heparin and heparan sulfate on T cell proliferation induced by thymic stromal cell monolayer or its derived T cell growth factor (TCGF). A thymic stromal cell clone (MRL104.8a) supported the growth of Ag-specific, IL-2-dependent Th cell clone (9-16) in the absence of Ag and IL-2 by producing a unique TCGF designated as thymic stroma-derived T cell growth factor (TSTGF). The addition of heparin to cultures in which the growth of 9-16 Th cells was otherwise stimulated by the MRL104.8a monolayer or a semipurified sample of the TSTGF resulted in heparin dose-dependent inhibition of 9-16 Th proliferation. The dose of heparin required for inducing 50% reduction of TSTGF-induced proliferation of Th at a given cell number was found to be proportional to the magnitude of the TSTGF added to cultures, suggesting that heparin exerted its inhibitory effect by binding to the TSTGF rather than by acting on Th cells. A similar growth-inhibiting effect of heparin was observed in IL-7-dependent proliferation of pre-B cell line or Th, but not in IL-2-dependent T cell proliferation or IL-3-dependent myeloid cell proliferation. A strong affinity of TSTGF and IL-7 for heparin was confirmed by the fact that both TSTGF and IL-7 adhered to columns of heparin-agarose and were eluted by salt. When various glycosaminoglycans were tested for the heparin-like Th growth-regulatory capacity, heparan sulfate exhibited Th growth-inhibiting ability comparable to that observed for heparin. These results indicate that the activity of thymic and/or bone marrow stroma-derived lymphocyte growth factor (TSTGF/IL-7) but not of Th-producing TCGF (IL-2) is negatively regulated by heparin or heparan sulfate, which would represent major glycosaminoglycans in the extra-cellular matrix of stromal cells.

Supernatants of human PBL cultures stimulated with PHA contain a lymphokine named T-cell growth factor (TCGF). To remove the contaminant PHA from these supernatants, we have designed a simple and effective method based on the binding properties of this lectin to chicken red blood cells. This procedure completely removes the functional activities of a PHA solution, both in lectin-induced cellular cytotoxicity against erythroid targets and in DNA synthesis of fresh PBL. The TCGF activity remaining in the supernatants after absorption with CRBC is dependent on the cellular concentration employed in the preparation of the supernatants. The absorption procedure does not remove significant TCGF activity in the supernatants prepared at high cellular concentrations (5 X 10(6) cells/ml), whereas in those prepared at low cellular concentrations (1 X 10(6) cells/ml) a partial loss of TCGF activity is detected.

The stimulation of DNA synthesis in lymphocyte populations was previously shown to depend strongly on the intracellular glutathione (GSH) level. Since T cell growth is known to depend on interleukin 2 (IL-2), the experiments in this report were designed to determine whether intracellular GSH depletion may inhibit IL-2 production or the IL-2 dependent DNA synthesis. Our experiments revealed that IL-2 production and DNA synthesis of mitogenically stimulated splenic T cells have indeed different requirements for GSH. The addition of relatively high concentrations of GSH (5 mM) to cultures of concanavalin A (Con A)-stimulated splenic T cells was found to augment strongly the DNA synthesis but inhibited the production of IL-2. Moderate intracellular GSH levels, however, are apparently not inhibitory for IL-2 production, since intracellular GSH depletion by cysteine starvation or by graded concentrations of DL-buthionine sulfoximine (BSO) had virtually no effect on IL-2-specific mRNA expression and the production of T cell growth factor (TCGF). The DNA synthesis activity, in contrast, was strongly suppressed after GSH depletion with either method. As in cultures of splenic T cells, GSH depletion had no substantial effect on the induction of IL-2 mRNA and TCGF production in several mitogenically stimulated T cell clones. Taken together, our experiments suggest that complex immune response may operate best at intermediate GSH levels that are not too high to inhibit IL-2 production but sufficient to support DNA synthesis.

T cells obtained from pleural effusion of patients with tuberculous pleurisy were stimulated in vitro with PPD. The culture supernatant was shown to be substituted for T cells in PWM-induced IgG production by human B cells. As many as 0.5 to 3 x 10(9) lymphocytes were obtained from 1 patient, and about 85% of them were E-rosette positive, making it possible to purify human helper T cell factor(s) (ThF). Helper activity was sensitive to heating at 70 degrees C for 5 min and treatment with trypsin. ThF did not have conventional Ig determinants and was recovered in the 50 to 67% ammonium sulfate fraction. The purification was done by successive DEAE-Sephadex and CM-Sephadex ion-exchange chromatography, Sephadex G-100 gel filtration, and isoelectric focusing. It was shown that ThF activity was eluted by gel filtration in the fraction with a m.w. of about 20,000. Isoelectric focusing of this fraction revealed that ThF activity distributed in the pl range of 6.5 to 8.0, whereas TCGF activity was focused in a single peak at pI of 6.5, indicating the existence of a helper factor without TCGF activity.

Lymphocytes from peripheral blood, lymph node, spleen and tumour of 7 patients with various carcinomas (2 lung, 3 colon, 1 gastric and 1 parotid tumour) were cultured for 15 days in conditioned media containing T-cell growth factor (TCGF; Interleukin 2) after which their cytotoxic activity against autologous tumour (and in some instances, autologous normal) cells and allogeneic tumour targets was evaluated in a short-term 51Cr-release assay. Significant cytotoxicity against autologous tumour targets was detected in at least one effector preparation from all of the patients, under conditions where, in some cases, other autologous cells (normal lung, PHA-transformed lymphocytes) were resistant. This cytotoxicity also generally extended to allogeneic tumour targets, but lysis of K562, a cell line sensitive to natural killing, occurred in only 3 of 19 effector cell preparations. The data are consistent with a polyclonal expansion of cytotoxic T-cells of tumour-bearing patients which includes the amplification of a population recognitive of antigens expressed on autologous neoplastic cells.

Clones of human cytotoxic T cells (Tc) specific for Epstein Barr virus (EBV) were isolated from peripheral blood lymphocyte (PBL) cultures stimulated repeatedly with autologous EBV-transformed lymphoblastoid cell line (LCL) cells in vitro. The method employed to clone EBV-specific Tc was a limiting dilution technique utilizing T cell growth factor (TCGF). The EBV specificity of Tc clones was determined by showing that they were significantly cytotoxic for autologous LCL cells but not for either autologous PBL or (natural killer-sensitive) K-562 cells. Eight EBV-specific Tc clones derived from a single donor exhibited distinct cytotoxic patterns against allogeneic LCL targets. Two clones were cytotoxic to LCL targets sharing both HLA-A26 and B15 antigens with effectors, and killing by two other clones was strongly restricted to autologous LCL cells. The four remaining clones showed cytotoxicities against various allogeneic LCL targets irrespective of HLA antigen expression. Eight EBV-specific Tc clones derived from a second donor also exhibited a wide spectrum of cytotoxicity to allogeneic LcL targets. We conclude that EBV-specific Tc, induced in vitro, consist of a number of clones with respect to restrictions imposed by the major histocompatibility complex. The determinants regulating these restrictions may include not only private HLA antigenic determinants that are defined by the HLA serotyping, but also undefined HLA antigenic determinants.

Immunodeficient ageing (C57BL/10 x DBA/2)F1 mice were treated by a single injection of synthetic thymic hormones and 4 days later their thymus and spleen cells were assayed in vitro for T cell activities. A few nanograms of THF-gamma 2 were found to raise the frequency of mitogen-responsive T cells in thymus and spleen cell populations as well as the frequency of cytokine-producing splenic T cells, up to the levels observed in young mice. Moreover, injection of THF-gamma 2 was found to restore T cell growth factor (TCGF) production by mitogen-stimulated spleen cells. Also, the helper activity of spleen cells was enhanced by this treatment and increased with increasing the THF-gamma 2 dose over a wide range. Similarly, the effects of thymopentin and thymosin-alpha 1 on T helper cell activity increased with increasing the injected dose, but the efficiencies of THF-gamma 2 and thymopentin were, respectively, 400-fold and eight-fold greater than that of thymosin-alpha 1.

Three-week-old chicks were inoculated orally with CAV and killed at various times postinoculation (PI). The spleens were removed, the cells were stimulated with concanavalin A, and lymphocyte transformation responses were determined. Supernatants from these cultures were also assayed for T-cell growth factor (TCGF) and interferon. Adherent macrophages from spleen or bone marrow were assayed for interleukin-1 production, Fc receptor expression, phagocytosis, and bactericidal activity. All CAV-inoculated chickens developed CAV antibodies, but no anemia was seen. Controls remained CAV-antibody-negative throughout the experiment. CAV-inoculated chickens showed significant differences from controls in their lymphocyte transformation responses and in production of TCGF and interferon. Differences were greatest at 14, 21, and 28 days PI. Significant differences were also observed in interleukin-1 production by spleen macrophages, as well as in Fc receptor expression, phagocytosis, and bactericidal activity of bone-marrow macrophages.

Acquired immune deficiency syndrome (AIDS) is characterized by severe depletion of OKT4+ T lymphocytes and leukemia is associated with abnormal proliferation of maturation-arrested lymphocytes. Human T-lymphotropic virus type III (HTLV-III) or lymphoadenopathy virus (LAV) and type I (HTLV-I) are etiologically linked to AIDS and adult T-cell leukemia/lymphoma, respectively. T-cell growth factor (TCGF, also known as interleukin 2) is required for the growth of activated T-cells, which play an important role in immune regulation. gamma-Interferon (IFN-gamma) is also implicated in immune modulation. It was possible that T-cell depletion in acquired immune deficiency syndrome could be due to an impairment of TCGF synthesis and that adult T-cell leukemia could be due to unregulated production of TCGF. The results reported here show that the transcription of the TCGF gene was not impaired in cultured HTLV-III-infected cells. Paradoxically, the TCGF gene in HTLV-I-infected cells was transcriptionally inactive. The reverse was the case for the gamma-interferon gene--it was actively transcribed in HTLV-I-infected cells but not in the HTLV-III-infected and virus-producing H9 and H4 cell line. No evidence was obtained suggesting abnormal regulation of the TCGF or of the IFN-gamma gene consequent to HTLV-III infection. It thus appears that in both HTLV-III and HTLV-I infection, growth control and immune regulatory mechanisms may bypass a modulatory role of TCGF or of IFN-gamma.

A cloned T cell hybridoma (123) was shown to produce T cell growth factor (TCGF). Supernatants of cultures of hybridoma-123 that had been stimulated with concanavalin A caused T cell blasts or a cloned T cell line to proliferate, allowed a mitogenic response to concanavalin A by thymocytes and by lymph-node cells depleted of accessory cells by treatment with anti-Ia serum and complement, and permitted the generation of both polyclonal and antigen-stimulated cytotoxic T lymphocytes in cultures of thymus cells. These observations suggest that the hybridoma is producing a factor with an identical spectrum of activities to that associated with TCGF derived from mitogen-stimulated spleen cells and indicate that the T cell is the source of TCGF. Together with other evidence that the same hybridoma produces activity affecting B lymphocytes, myeloid progenitor cells, and pluripotential stem cells, these experiments confirm the role of the activated T cell as a regulator of hemopoietic and lymphoid systems. T cell hybridomas should prove invaluable for biochemical and genetic analysis of these factors.

Interleukin-2 (IL-2 or T cell growth factor, TCGF) is a lymphokine absolutely required for clonal expansion of activated T-lymphocytes with specific cytotoxic, helper, and suppressor functions. The capability of production of and responsiveness to IL-2 in 15 normal children, 28 newly diagnosed, and 22 hyposensitized (greater than 1 yr) children with asthma was studied. The results demonstrated (1) The lymphocytes from newly diagnosed patients produced a much greater amount of IL-2 when they were stimulated with specific allergen (house dust) but not with phytohemagglutinin (PHA) than did those from both hyposensitized patients and normal subjects. (2) The responsiveness to IL-2 of activated lymphocytes from hyposensitized patients was much more vigorous than those from both new patients and normal subjects, whether the responder lymphocytes were pretreated with PHA or house dust. Thus, hyposensitization is able to modulate the immunobiologic functions of T cells with respect to production of and responsiveness to IL-2, and such alterations may contribute to the immunologic changes occurring in patients receiving long-term hyposensitization.