Lymphokine activity production in graft-versus-host reactions across minor histocompatibility antigen barriers.

Activated T cells responding to murine minor histocompatibility antigens (HA) were characterized according to the patterns of lymphokine activity production. Although B10.D2/nSN and BALB/c are mutually non-reactive in mixed lymphocyte reaction (MLR), graft-versus-host reaction (GVHR) can be induced by the injection of a large amount of B10.D2/nSN lymphoid cells into irradiated BALB/c recipient mice. Spleen cells from such GVHR mice spontaneously produced interleukin 3 (IL-3)-dependent cell-stimulating activity in cultures, but did not produce interleukin 2 (IL-2). Normal B10.D2/nSN spleen cells also produced IL-3-like activity, but not IL-2 in MLR supernatants, in response to irradiated BALB/c splenocytes. In addition, B-cell stimulatory factor-1 (BSF-1)/interleukin 4 (IL-4) and colony-stimulating factor (CSF) activity were detected in MLR supernatants. The properties of the produced lymphokine activities were similar to those produced in syngeneic transplant mice and syngeneic MLR, but a difference in the time course of lymphokine production existed between GVHR and syngeneic transplant mice. These results indicate that T cells may be activated in vivo in allogeneic transplantation when the donor and the recipient are matched for major HA, and are non-reactive in MLR. Also, the character of lymphokine-producing T cells activated by minor HA may not be qualitatively different from those responding to irradiated syngeneic cells.     

Immunohistochemical study of synphilin-1 in brains of patients with dementia with Lewy bodies - synphilin-1 is non-specifically implicated in the formation of different neuronal cytoskeletal inclusions

The caudal part of the fastigial nucleus, or the fastigial oculomotor region (FOR), plays an important role in executing accurate saccades. Inactivation of a monkey FOR leads to dysmetric saccades. Currently available data suggest that the dysmetria could be described as a parametric, uniform change in saccadic gain or, alternatively, as a constant error in the specification of the saccadic goal. To discriminate between these two possibilities, we examined the effect of FOR inactivation in the monkey. After a unilateral injection of muscimol into the FOR, ipsiversive saccades overshot a target. Gains were similar for movements of different sizes. The overshoot increased proportionately with the target distance and had a very small constant component. The present study indicates that the hypermetria of ipsiversive saccades after inactivation of the monkey FOR is primarily due to a uniform gain increase for all sizes of saccades.     

TCR repertoire in early fetal mouse thymus

We investigated the rearrangement and expression of TCR genesin mouse fetal thymus organ culture, a system that avoids subsequententry of hematopoietic precursor cells. The first observablerearranged TCR gene was homogeneous V2-J2, detectable as earlyas fetal day 11 (d11) in the thymic primordla. The productiveTCR was homogeneous V5-J1, first detectable in d13 thymocytes,followed by adult-type TCR (V4 and V7). Sequence analysis ofTCR revealed five types of V-J junctional sequences. In thevery early stage, a homogeneous V-J junction is generated viaa short homology sequence in the coding region (Type I), whilea short homology sequence in the P-nucleotlde rather than thecoding region is used in the following stage (Type II). In thelater embryonic stages, diverse V-J junctions are generatedby well-known mechanisms, such as P-nucleotide (Type III), N-regioninsertion (Type IV) or trimming of the coding ends (Type V).These findings suggest that the generation of homogeneous TCR (V2 and V5) in the early fetal stages is due to the intrinsicrearrangement mechanisms and is in stage specific manner.     

Transforming growth factor beta is protective in host resistance against Listeria monocytogenes infection in mice.

The role of transforming growth factor beta (TGF-beta) in host resistance against Listeria monocytogenes infection was studied with mice. The constitutive expression of TGF-beta 1 mRNA was observed in the spleens and livers of mice before and after infection. Injecting the mice with anti-TGF-beta 1 peptide serum resulted in diminished antilisterial resistance, whereas the administration of human platelet-derived TGF-beta 1 enhanced the resistance. Moreover, mice were protected against lethal infection when treated with TGF-beta 1. These results suggest the TGF-beta 1 might be involved in antilisterial resistance. On the other hand, injecting the mice with TGF-beta 1 resulted in a decrease in the titers of endogenous gamma interferon, tumor necrosis factor alpha, and interleukin-6, which are crucial in antilisterial resistance, in sera and in extracts of spleen and liver. Thus, a complicated mechanism might be involved in the role of TGF-beta 1 in host resistance against L. monocytogenes infection.     

Differing Patterns of P-Selectin Expression in Lung Injury

Using two models of acute lung inflammatory injury in rats (intrapulmonary deposition of immunoglobulin G immune complexes and systemic activation of complement after infusion of purified cobra venom factor), we have analyzed the requirements and patterns for upregulation of lung vascular P-selectin. In the immune complex model, upregulation of P-selectin was defined by Northern and Western blot analysis of lung homogenates, by immunostaining of lung tissue, and by vascular fixation of ¹²⁵I-labeled anti-P-selectin. P-selectin protein was detected by 1 hour (long before detection of mRNA) and expression was sustained for the next 7 hours, in striking contrast to the pattern of P-selectin expression in the cobra venom factor model, in which upregulation was very transient (within the 1st hour). In the immune complex model, injury and neutrophil accumulation were P-selectin dependent. Upregulation of P-selectin was dependent on an intact complement system, and the presence of blood neutrophils was susceptible to the antioxidant dimethyl sulfoxide and required C5a but not tumor necrosis factor α. In contrast, in the cobra venom factor model, upregulation of P-selectin, which is C5a dependent, was also dimethyl sulfoxide sensitive but neutrophil independent. Different mechanisms that may explain why upregulation of lung vascular P-selectin is either transient or sustained are discussed.     

Bradykinin Stimulates Type II Alveolar Cells to Release Neutrophil and Monocyte Chemotactic Activity and Inflammatory Cytokines

In the present study, we evaluated the potential of bradykinin (BK) to induce the release of neutrophil and monocyte chemotactic activity (NCA and MCA) and cytokines from an alveolar type II epithelial cell line, A549 cells. BK stimulated A549 cells to release NCA and MCA in a dose- and time-dependent manner (P < 0.001). Checkerboard analysis revealed that both NCA and MCA involved chemotactic and chemokinetic activity. Molecular sieve column chromatography showed three molecular weight masses (near 19 kd, 8 kd, and 400 d) for NCA and several molecular weight peaks (near 66 kd, 25 kd, 19 kd, 16 kd, and 400 d) for MCA. The release of NCA and MCA was inhibited by cycloheximide and lipoxygenase inhibitors (P < 0.01). The NCA and MCA were inhibited by leukotriene B4 (LTB4) receptor antagonist (P < 0.01), and the concentration of LTB4 was high enough for NCA and MCA. Antibodies to interleukin (IL)-8 and granulocyte colony-stimulating factor (G-CSF) attenuated NCA (P < 0.01), and antibodies to monocyte chemotactic protein-1 (MCP-1), G-CSF, and transforming growth factor (TGF)-β attenuated MCA (P < 0.01). The levels of IL-8, G-CSF, MCP-1, and TGF-β increased time dependently (P < 0.01). BK also stimulated the release of ILeukin-6 from A549 cells (P < 0.001). The receptors responsible for the release of NCA, MCA, and individual chemokines involved both BKB1 and BKB2 receptors. These data suggest that BK may stimulate alveolar type II pneumocytes to release inflammatory cytokines, which then may modulate the lung inflammation.     

An immunodominant haptenic epitope of carbamazepine detected in serum from patients given long-term treatment with carbamazepine without allergic reaction

An anticarbamazepine antibody was detected in the serum of a patient with severe carbamazepine-induced serum sickness. We found that the patient's T cells and IgG antibody recognized an epitope which appeared in subjects showing an allergic reaction, as well as that in subjects who showed no allergic reaction, after long-term carbamazepine therapy. These results show that an anti-carbamazepine immune response does not occur in the majority of subjects who undergo long-term carbamazepine therapy without developing allergic symptoms, although the immunodominant haptenic epitope of carbamazepine is present in their sera.     

Comparative studies with various substrains of Mycobacterium bovis BCG on the production of an antigenic protein, MPB70.

A protein, isolated and purified from the unheated culture filtrate of Mycobacterium bovis BCG (substrain Tokyo 172) and designated MPB70, elicited a delayed skin reaction in guinea pigs sensitized with viable cells of BCG but not in those sensitized with heat-killed cells. The skin reaction reached the maximum 4 to 8 weeks after the inoculation of the BCG and then decreased gradually, resulting in conversion to negative after 20 weeks, whereas the skin reaction to purified protein derivative (PPD) continued to be positive. Guinea pigs immunized with viable cells of various substrains of BCG were skin tested with MPB70 and PPD. Guinea pigs immunized with the BCG substrain Tokyo 172 and the substrain Moreau (Brazil) showed strong delayed skin reactions to both MPB70 and PPD. On the other hand, guinea pigs immunized with the Pasteur substrain 1173P2, the Glaxo substrain 1077, the Copenhagen substrain 1331, the Tice substrain, or the Beijing substrain 64-42 showed negative skin reactions to MPB70, whereas they were strongly positive to PPD. In a two-dimensional acrylamide gel electrophoretic analysis of proteins from the culture filtrates of the BCG substrains, the culture filtrates of the Tokyo and Moreau substrains showed the spot of MPB70 on the gel slabs, whereas those of the other BCG substrains did not.     

The role of the solitary and paramedian reticular nuclei in mediating cardiovascular reflex responses from carotid baro- and chemoreceptors

1. With dye-filled micro-electrodes single neurones in the medulla of anaesthetized paralysed cats were identified which: (a) fired rhythmically in synchrony with or were modulated by the cardiac cycle, and which ceased firing with occlusion of the ipsilateral common carotid artery (carotid sinus baroreceptor neurones); (b) were excited by stimulation of carotid body chemoreceptors by close intra-arterial injection of lobeline into the thyroid artery (carotid body chemoreceptor neurones).2. Twelve carotid baroreceptor neurones were identified, in thirty-three cats, nine of which were localized in the intermediate area of the nucleus of the solitary tract (NTS) within 1 mm ahead of or behind the obex; three units were located either in the parahypoglossal area or the dorsal portion of the paramedian reticular nucleus (PRN).3. Of the twenty-one carotid chemoreceptor neurones which were identified, thirteen were localized in the NTS, three in the parahypoglossal area and four in the dorsal PRN.4. Bilateral lesions of the paramedian reticular area of medulla destroying the PRN, abolished or reversed the depressor response to electrical stimulation of myelinated fibres of the carotid sinus nerve (CSN), attenuated the depressor response to carotid sinus stretch and augmented the pressor response to chemoreceptor stimulation by lobeline. Such lesions did not significantly alter the reflex heart rate responses.5. Small lesions of the NTS within an area 1 mm rostral to the obex abolished all reflex blood pressure and heart rate responses to electrical stimulation of the CSN or natural stimulation of carotid baro- or chemoreceptors.6. Baroreceptors and chemoreceptors of the CSN project both to the intermediate zone of the NTS and to more medial areas of the medulla, particularly the dorsal PRN and parahypoglossal area.7. The PRN serves to mediate the reflex depressor, but not cardio-vagal, response from myelinated baroreceptors and buffers the pressor responses from chemoreceptors; it may serve as an important area integrating cardiovascular activity descending from forebrain, brain stem and cerebellum with baroreceptor reflexes.8. Cardiovascular reflex responses arising from non-myelinated baroreceptors and all chemoreceptors are mediated by neurones in the intermediate area of the NTS.     

Interaction of Arc with CaM kinase II and stimulation of neurite extension by Arc in neuroblastoma cells expressing CaM kinase II

We investigated the relationship between Arc (activity-regulated cytoskeleton-associated protein) and Ca(2+)/calmodulin-dependent protein kinase II (CaM kinase II). Arc and CaM kinase II were concentrated in the postsynaptic density. These proteins were accumulated after electroconvulsive treatment. Arc increased about 2.5-fold within 30 min and was maintained at this level for 8h after the stimulation. CaM kinase II also increased within 30 min and remained at this level for at least 24h. The interaction of Arc with CaM kinase II was demonstrated using GST-Arc fusion protein, and confirmed in neuroblastoma cells by immunoprecipitation. We examined the function of Arc by introducing Arc cDNA into neuroblastoma cells expressing CaM kinase II. The cells expressing both Arc and CaM kinase II had longer neurites than those expressing CaM kinase II alone. Arc itself did not promote neurite outgrowth. The growth of neurites by Arc was completely blocked by treatment with KN62, an inhibitor of CaM kinases. These results indicated that Arc potentiated the action of CaM kinase II for neurite extension.