IFN-gamma promotes Fas ligand- and perforin-mediated liver cell destruction by cytotoxic CD8 T cells

To study liver cell damage by CTL, CD8 T cells from P14 TCR transgenic (tg) mice specific for the gp33 epitope of lymphocytic choriomeningitis virus with either deficiency in IFN-gamma (P14.IFN-gamma(null)), functional Fas ligand (P14.gld), or perforin (P14.PKO) were transferred into H8 tg mice ubiquitously expressing gp33 Ag. Treatment of H8 recipient mice with agonistic anti-CD40 Abs induced vigorous expansion of the transferred P14 T cells and led to liver cell destruction determined by increase of glutamate dehydrogenase serum levels and induction of caspase-3 in hepatocytes. Liver injury was mediated by the Fas/Fas ligand (FasL) pathway and by perforin, because P14.gld and P14.PKO T cells failed to induce increased glutamate dehydrogenase levels despite strong in vivo proliferation. In addition, H8 tg mice lacking Fas were resistant to the pathogenic effect of P14 T cells. Besides FasL and perforin, IFN-gamma was also required for liver cell damage, because P14.IFN-gamma(null) T cells adoptively transferred into H8 mice failed to induce disease. Moreover, Fas expression on hepatocytes from H8 recipient mice was increased after transfer of wild-type compared with P14.IFN-gamma(null) T cells, and wild-type P14 T cells expressed higher levels of FasL than P14 T cells lacking IFN-gamma. Thus, our data suggest that IFN-gamma released by activated CD8 T cells upon Ag contact facilitates liver cell destruction.     

IFN-gamma production by Th1 cells generated from naive CD4+ T cells exposed to norepinephrine

During activation in vivo, naive CD4(+) T cells are exposed to various endogenous ligands, such as cytokines and the neurotransmitter norepinephrine (NE). To determine whether NE affects naive T cell differentiation, we used naive CD4(+) T cells sort-purified from either BALB/c or DO11.10 TCR-transgenic mouse spleens and activated these cells with either anti-CD3/anti-CD28 mAbs or APC and OVA(323-329) peptide, respectively, under Th1-promoting conditions. RT-PCR and functional assays using selective adrenergic receptor (AR) subtype antagonists showed that naive CD4(+) T cells expressed only the beta 2AR subtype to bind NE and that stimulation of this receptor generated Th1 cells that produced 2- to 4-fold more IFN-gamma. This increase was due to more IFN-gamma produced per cell upon restimulation instead of more IFN-gamma-secreting cells, as determined by IFN-gamma-specific immunofluorescence and enzyme-linked immunospot. In contrast, Th1 cell differentiation was unaffected when naive T cells were exposed to NE and activated either in the presence of a neutralizing anti-IL-12 mAb or by APC from IL-12-deficient mice. Moreover, the addition of IL-12 to the IL-12-deficient APC cultures restored the ability of NE to increase Th1 differentiation. Taken together, these results indicate that a possible link may exist between the signaling pathways used by NE and IL-12 to increase naive CD4(+) T cell differentiation to a Th1 cell.     

Comparative results of thyroid ultrasound and laboratory studies in patients exposed to radiation

The paper presents the data of thyroid ultrasound and laboratory studies in 233 persons exposed to radiation: 63 X-ray physicians, 36 Chernobyl accident liquidators, 31 patients with lymphogranulomatosis, receiving radiotherapy, and 103 individuals (a control group) who were physicians of various specialties whose professional activity is unassociated with radiation factors. Thyroid changes were found in 67.7% of the examinees after gamma teletherapy and in 31% in the control group; among X-rat physicians and Chernobyl accident liquidators, the occurrence of thyroid changes was equal--44.4%. Direct radiation exposure of the thyroid in a dose of above 30 Gy led to the development of postradiation fibrosis (22.5%) and postradiation thyroiditis (32.3%) with thyroid dysfunction that is most commonly presented by hypothyroidism (22.45%). Autoimmune thyroiditis (11.1%) more predominantly with subclinical hypothyroidism, and diffuse thyroid hyperplasia (19.4%) resulted from mixed internal and external radiation. Thyroid changes revealed in X-ray physicians mainly appeared as nodular masses (22.2%) without functional impairments but to be followed up. It is recommended that the annual preventive examination protocol be supplemented by thyroid ultrasound studies in X-ray physicians.     

Microvascular remodeling and accelerated hyperemia blood flow restoration in arterially occluded skeletal muscle exposed to ultrasonic microbubble destruction

We showed previously that microbubble destruction with pulsed 1-MHz ultrasound creates a bioeffect that stimulates arteriogenesis and a chronic increase in hyperemia blood flow in normal rat muscle. Here we tested whether ultrasonic microbubble destruction can be used to create a microvascular remodeling response that restores hyperemia blood flow to rat skeletal muscle affected by arterial occlusion. Pulsed ultrasound (1 MHz) was applied to gracilis muscles in which the lateral feed artery was occluded but the medial feed artery was left intact. Control muscles were similarly occluded but did not receive ultrasound, microbubbles, or both. Hyperemia blood flow and number of smooth muscle (SM) alpha-actin-positive vessels, >30-mum arterioles, and capillaries per fiber were determined 7, 14, and 28 days after treatment. In ultrasound-microbubble-treated muscles, lateral region hyperemia blood flow was increased at all time points and restored to normal at day 28. The number of SM alpha-actin vessels per fiber was increased over control in this region at days 7 and 14 but decreased by day 28, when larger-diameter arterioles became more prevalent in the medial region. The number of capillaries per fiber was increased over control only at day 7 in the lateral region and only at days 7 and 14 in the medial region, indicating that the angiogenesis response was transient and likely did not contribute significantly to flow restoration at day 28. We conclude that ultrasonic microbubble destruction can be tailored to stimulate an arteriogenesis response that restores hyperemia blood flow to skeletal muscle in a rat model of arterial occlusion.     

The thyroid and adrenal glands in rats chronically exposed to an intense electric field

Two month-old male Long-Evans rats were exposed for 8 hours a day during 4 weeks to an ambient electric field (50 kV/m). The growth rate of electric field-exposed rats was identical to the controls' one. No significant effect of exposure was observed on the levels of various hormones: plasma and adrenal corticosterone; plasma TSH, ACTH, T3 and T4; adrenal epinephrine and norepinephrine. Histological studies of thyro?d and adrenals showed a normal appearance. In adrenals, the levels of various components and the activities of enzymes which have a role on stero?dogenesis were not modified. It is concluded that, in the present experiment conditions, the daily exposure of the rat to a severe electric field does not affect the normal state of thyro?d and adrenals.     

The cell biology of T-dependent B cell activation

The requirement that CD4+ helper T cells recognize antigen in association with class II Major Histocompatibility Complex (MHC) encoded molecules constrains T cells to activation through intercellular interaction. The cell biology of the interactions between CD4+ T cells and antigen-presenting cells includes multipoint intermolecular interactions that probably involve aggregation of both polymorphic and monomorphic T cell surface molecules. Such aggregations have been shown in vitro to markedly enhance and, in some cases, induce T cell activation. The production of T-derived lymphokines that have been implicated in B cell activation is dependent on the T cell receptor for antigen and its associated CD3 signalling complex. T-dependent help for B cell activation is therefore similarly MHC-restricted and involves T-B intercellular interaction. Recent reports that describe antigen-independent B cell activation through coculture with T cells activated by anti-T-cell receptor or anti-CD3 antibodies suggest that cellular interaction with T cells, independent of antigen presentation or lymphokine secretion, induces or triggers B cells to become responsive to T-derived lymphokines, and that this may be an integral component of the physiological, antigen- and MHC-restricted T-dependent B cell activation that leads to antibody production.     

Universal vaccine carrier. Liposomes that provide T-dependent help to weak antigens

The design of a thymus-dependent synthetic vaccine that will provide a universal T cell epitope for B cell epitopes is described in this study. Simultaneous incorporation into liposomes of both a peptide recognized by Th lymphocytes and a lipophilic hapten and the IgG antibody responses to this hapten were assessed in outbred mice. DNP-aminocaproyl phosphatidylethanolamine (DNP-CapPE) is a well characterized T-independent hapten Ag. HA2 peptide derived from the hemagglutinin protein of influenza virus contains amino acid sequences recognized by Th and T cytotoxic lymphocytes. In addition, HA2 contains a sequence of hydrophobic amino acids near the carboxyl terminus, allowing its incorporation into liposomes. Results of immunization show that (i), when DNP-CapPE is carried by liposomes without the HA2 peptide, an IgM antibody response is induced, (ii) liposomes carrying both HA2 and DNP-CapPE elicit an IgG antibody response to DNP in a dose-dependent fashion for both HA2 and DNP, (iii) the liposomes must be processed intracellularly in order to elicit a response, (iv) the system leads to a memory response for DNP, and (v) all of the IgG subclasses are elicited. These data suggest that liposomes containing the HA2 peptide exhibit a T-dependent carrier effect for a T-independent Ag. The significance of these findings is discussed in conjunction with the characteristics of the liposome model used.     

Corticosteroid and thyroid responses of larval and juvenile turbot exposed to the water-soluble fraction of crude oil

Turbot larvae (24–590 degree C days; 2–32 days post-hatch) and juveniles (1345 degree C days; 98 days post-hatch), were exposed for 6 h to 25, 33 and 50% water-soluble fraction (WSF) of crude oil in either static or flow-through test systems. Larvae showed generalized primary endocrine responses, identified by elevated whole body cortisol content from as early as 2 days post-hatch. In older larvae and juveniles, the response was related to the WSF concentration. This dose-response relationship was not apparent in younger and yolk-sac larvae. Whole body thyroxine content of turbot larvae exposed to the WSF of crude oil was increased, but triiodothyronine content remained stable. Aromatic hydrocarbon concentrations [benzene, toluene, ethylbenzene and xylene (BTEX) and naphthalenes] remained constant during flow-through tests, but 65% of the initial level of BTEX hydrocarbons and 40% of the naphthalenes were lost during static exposures. Larval mortalities increased with exposure to an increasing concentration of crude oil WSF. Larval activity was significantly reduced even at the lowest WSF concentration.     

Engineering of macrophages to produce IFN-gamma in response to hypoxia

Activation of murine macrophages (Mphi) requires the collaboration of signals derived from the immune system and the environment. In this study, we engineered a murine Mphi cell line to become activated in response to an environmental signal, hypoxia, as the sole stimulus. Hypoxia is a condition of low oxygen tension, occurring in several pathological tissues, which acts in synergy with IFN-gamma to induce full Mphi activation. We transfected the ANA-1 murine Mphi cell line with a construct containing the IFN-gamma gene controlled by a synthetic promoter inducible by hypoxia (HRE3x-Tk), and we characterized the cellular and molecular biology of the engineered Mphi under normoxia or hypoxia. Engineered Mphi in normoxia expressed basal levels of IFN-gamma mRNA and protein that were strongly augmented by shifting the cells to hypoxia. Furthermore, they responded to the synthesized IFN-gamma with induction of IFN-responsive factor-1 and 2'-5'-oligoadenylate synthase expression. Under normoxic conditions, the engineered Mphi had a significant constitutive level of Ia Ags and Fc receptors. Hypoxia induced further augmentation of Ia and Fc expression. Finally, hypoxia induced inducible NO synthase expression, and subsequent reoxygenation led to the production of NO. In conclusion, the engineered Mphi, which produce IFN-gamma in an inducible manner, express new biochemical and functional properties in response to low oxygen environment as the sole stimulus, thereby circumventing the need for costimulation by other immune system-derived signals.     

Immunoregulation of T-dependent responses by a cloned dendritic cell

We have described a cloned dendritic cell, Clone Den-1, which is a potent accessory cell for some T-dependent immune responses. Clone Den-1 activates T cells in both autologous and allogeneic mixed lymphocyte reactions, but cannot present soluble antigen to T cells that co-recognize nominal antigen plus I-Ab gene products. In addition, Clone Den-1 or factors produced by it can reconstitute plaque-forming cell responses by accessory cell-depleted B plus T cells to the T-dependent antigen sheep red blood cells. The relationship of this clone to heterogeneous populations of dendritic cells and other accessory cells is discussed.