Favorably tipping the balance between cytopathic and regulatory T cells to create transplantation tolerance

Therapeutic application of broadly reactive anti-T cell antibodies can lead not only to potent immunosuppression but also to profound and long-lived T cell depletion. We reasoned that a strategy that almost exclusively targets activated cytopathic donor reactive T cells and spares immunoregulatory networks might prove to be an exceptionally potent and highly selective means of producing long-term engraftment and tolerance. Herein we show that the combined administration of rapamycin and agonist IL-2- and antagonist IL-15-related cytolytic fusion proteins provides for long-term engraftment/tolerance in exceptionally stringent allotransplant models by (1) limiting the early expansion of activated T cells, (2) preserving and even exaggerating their subsequent apoptotic clearance, and (3) further amplifying the depletion of these activated T cells by antibody-dependent mechanisms, while (4) preserving CD4+CD25+ T cell-dependent immunoregulatory networks.     

Kinetic fluorescence RT-PCR is highly sensitive for detection of germ-cell-tumor-specific transcripts in peripheral blood

The aim of our study was to determine whether conventional staging in patients with testicular germ-cell-tumors (GCT) could be supplemented by quantification of beta-human choriogonadotropin mRNA levels in peripheral blood using kinetic fluorescence RT-PCR. Blood samples from 41 patients with GCT of different clinical stages (CS) were pre-therapeutically examined by kinetic fluorescence RT-PCR with the LightCycler for beta-human chorionic gonadotropin (beta-HCG) mRNA expression levels. The controls comprised of samples taken from patients 3 months after treatment, from patients with inflammatory testicular diseases or non-germ-cell-tumors and from healthy males (n=66). Six positive results [cut-off level: normalized beta-HCG mRNA (Nbeta-HCG) >400 relative gene expression (RGE)] were found in controls (specificity 90.9%, 95% CI: 76.9-97.3%). The overall ratio of positive PCR results in the group of GCT patients was 82.92% (34/41) (CS I 18/23, CS IIa-b 6/7, CS >IIb 10/11) (sensitivity 82.9%, 95% CI: 65.1-91.2%). The average Nbeta-HCG level in patients with clinical stage I tumors was 63772.0+/-125720.5 (mean +/- standard deviation) relative gene expression (RGE), 35076.0+/-52253.5 RGE in those with CS IIa-b tumors and 87298.3+/-120895.3 RGE in those with CS >IIb tumors. Kinetic fluorescence RT-PCR for tumor-specific gene products is, in contrast to qualitative RT-PCR, a promising approach to improve conventional staging in clinical low-stage testicular germ-cell-tumors. With high specificity, its sensitivity is higher than that of the corresponding serum tumor marker (82.92% vs 48.72%).     

Effect of methoxime combined with anticholinergic, anticonvulsant or anti-HCN drugs in tabun-poisoned mice

The effect of methoxime combined with a) atropine, b) benactyzine, c) atropine and natrium thiosulphate, d) atropine and diazepam on antidotal treatment effectiveness was studied in tabun-poisoned mice. In addition, the influence of pretreatment consisiting of pyridostigmine, benactyzine and trihexyphenidyle (PANPAL) administered 2 hours before tabun intoxication on the treatment effectivity of methoxime combined with e) atropine or f) benactyzine was tested. The most efficacious therapeutic mixture in non-pretreated mice was methoxime, atropine and diazepam. Natrium thiosulphate did not significantly increase neither decrease the antidotal treatment efficacy in comparison with methoxime and atropine alone. Pretreatment with PANPAL significantly decreased tabun toxicity (nearly 4 times in methoxime and benactyzine combination and more than 4 times in atropine and methoxime mixture). The present study demonstrates that the tabun toxicity in mice is more effectively reduced when PANPAL prophylactically is administered than in case of treatment with methoxime and cholinergic drug alone. We established that anticholinergic drug option in the therapeutic mixture of methoxime and anticholinergic drug did not cause the difference in the antidotal treatment effectivities.     

Tumor necrosis factor-mediated inhibition of interleukin-18 in the brain: a clinical and experimental study in head-injured patients and in a murine model of closed head injury.

Tumor necrosis factor (TNF) and interleukin-(IL)-18 are important mediators of neuroinflammation after closed head injury (CHI). Both mediators have been previously found to be significantly elevated in the intracranial compartment after brain injury, both in patients as well as in experimental model systems. However, the interrelation and regulation of these crucial cytokines within the injured brain has not yet been investigated. The present study was designed to assess a potential regulation of intracranial IL-18 levels by TNF based on a clinical study in head-injured patients and an experimental model in mice. In the first part, we investigated the interrelationship between the daily TNF and IL-18 cerebrospinal fluid levels in 10 patients with severe CHI for up to 14 days after trauma. In the second part of the study, the potential TNF-dependent regulation of intracerebral IL-18 levels was further characterized in an experimental set-up in mice: (1) in a standardized model of CHI in TNF/lymphotoxin-alpha gene-deficient mice and wild-type (WT) littermates, and (2) by intracerebro-ventricular injection of mouse recombinant TNF in WT C57BL/6 mice. The results demonstrate an inverse correlation of intrathecal TNF and IL-18 levels in head-injured patients and a TNF-dependent inhibition of IL-18 after intracerebral injection in mice. These findings imply a potential new anti-inflammatory mechanism of TNF by attenuation of IL-18, thus confirming the proposed "dual" function of this cytokine in the pathophysiology of traumatic brain injury.     

Translocation (6;9)(p23;q34) in acute nonlymphocytic leukemia: Three new cases

Several recent reports have described cases of acute nonlymphocytic leukemia with a unique chromosome translocation, t(6;9)(p23;q34). We have studied three additional patients who have acute nonlymphocytic leukemia and t(6;9)(p23;q34). Our findings provide additional support for the suggestion that this translocation is yet another distinct cytogenetic abnormality associated with myeloproliferative disorders.     

Infrared spectra and crystallinity of isotactic poly(methyl methacrylate) and of its deuterated analogues

Infrared spectra of partially crystalline and amorphous samples of isotactic poly(methyl methacrylate) (i-PMMA), poly[methyl (α,α,α-²H₃)methacrylate] (i-PMMA-α-CD₃), poly-[(²H₃)methyl methacrylate] (i-PMMA-OCD₃), and poly[(²H₃)methyl (α,α,α,2,2-²H₅)methacrylate] (i-PMMA-D₈) were measured. Infrared spectra of the pure crystalline forms of these substances were obtained by digital separation. It was found that in acetonitrile, CDCl₃, and toluene solutions, i-PMMA has the same conformational structure as in the amorphous state; conformational structures of crystalline i-PMMA are not present in the solutions in measurable amounts.     

Phenotypic dynamics of tumor progression in human malignant melanoma

The phenotypic changes in human melanoma cells during the course of tumor progression were studied with monoclonal antibodies (MAbs) against the melanoma-associated antigens (MAA) M.2.2.4, H.2.8.10, K.1.2, A.1.43, and A.10.33, and HLA-(A,B,C and D). Cryostat sections of 172 primary melanomas of the skin, 157 melanoma metastases and 56 nevi were investigated with an indirect immunoperoxidase method. Phenotypic heterogeneity was observed within lesions at all stages, and also within different tumors of the same patients. Despite this heterogeneity, principles of antigen expression were found. From the reaction pattern of MAbs, the following classifications of antigens were derived: "constitutive" markers of nevomelanocytic cells (M.2.2.4 and H.2.8.10) were found expressed over a wide range of local and systemic tumors. One MAA, K.1.2 (Suter et al., 1985), that declines with progression of melanoma, was classified as an "early" antigen, whereas MAA that appear in primary melanoma in proportion to invasiveness, and which are expressed in metastases of lymph nodes and visceral organs (A.1.43, and A.10.33), were classified as "late" markers of tumor progression. HLA-antigens were classified as "intermediate" markers, HLA-A,B,C, as an "early-intermediate", and HLA-DR as a "late-intermediate" marker. The occurrence of class II HLA, A.1.43-, and A.10.33-positive tumor cells in primary melanoma indicates a high metastatic potential of tumors, independent of tumor thickness. The data show that local and systemic progression of melanoma is associated with qualitative changes in tumor cells which can be recognized by MAbs.