Asher Meshorer

Suppression of experimental allergic encephalomyelitis in rhesus monkeys by a synthetic basic copolymer

Abstract Experiments with rhesus monkeys demonstrate that a random basic copolymer of amino acids, denoted Cop 1, suppresses experimental allergic encephalomyelitis (EAE) when administered to the animals after clinical symptoms have already appeared. Fifteen daily injection of this material caused reversal of disease symptoms in the two treated monkeys, one of which suffered relapse 38 days later. When no treatment was given, the control monkeys deteriorated rapidly and died within a short time after onset of symptoms. Histological tests indicate EAE lesions in the brains of all monkeys except the monkey saved by Cop 1.

Oral treatment of mice with copolymer 1 (glatiramer acetate) results in the accumulation of specific Th2 cells in the central nervous system.

Mucosal administration of copolymer 1 (Cop 1, Copaxone(R), glatiramer acetate) suppresses experimental autoimmune encephalomyelitis (EAE), and is currently tested for its efficacy in the treatment of multiple sclerosis (MS). Here we demonstrate that oral treatment with Cop 1 induces, in mice, specific Th2 cells in the central nervous system (CNS), as manifested by their isolation from brains of actively sensitized Cop 1-fed mice, as well as, by the localization of orally induced Cop 1 specific suppressor cells in the brain, after their passive transfer to the periphery. Feeding with Cop 1 results in the accumulation in the CNS of cells that secrete Th2 cytokines in response to either Cop 1 or the autoantigen myelin basic protein (MBP), even in Th1 shifting environment, which consequently would lead to therapeutic effect in the MS diseased organ.

The safety of ultrasound in fetal monitoring

Abstract Since ultrasonic (Doppler) fetal monitoring systems are technically feasible, it is most important to establish whether repeated prolonged irradiations are safe to both the mother and the fetus. The safety of ultrasonic exposures was investigated in this study with particular regard to structural changes, congenital malformation, and chromosomal aberration. Intensity levels of 164, 272, 490, 1,050 mW. per square centimeter have been used at a frequency of 2.28 megahertz (MHz), for 5 or 60 minutes, either singly or for 5 consecutive exposures. At an intensity of 1,050 mW. per square centimeter, tissue damage was recorded proportional to the duration of exposure. Macroscopic and microscopic examinations including chromosomal analysis showed these tissue effects to be identical with those found in overheating. It has been found that an intensity level of 490 mW. per square centimeter did not cause critical tissue temperature rise. There were no pathologic findings on the macroscopic and microscopic examinations in the mothers, fetuses, and neonates, and no congenital malformations were recorded. Moreover, there were no pathologic findings in neonates of the third generation produced by brother-sister cross-mating from the fetuses which had been irradiated. All of the exposed mothers were able to conceive again, producing healthy offspring. Chromosome squash preparations were done from irradiated fetuses, revealing an equal incidence of chromosome aberrations as compared with that of the control group.

T-cells specific for soluble recombinant oligodendrocyte-specific protein induce severe clinical experimental autoimmune encephalomyelitis in H-2b and H-2s mice

To investigate the immunogenicity and encephalitogenicity of oligodendrocyte-specific protein (OSP), recombinant soluble mouse OSP (smOSP) was produced from a synthetic gene engineered to lack the sequences coding for the hydrophobic transmembrane domains of the native molecule. SmOSP was immunogenic and encephalitogenic for SJL/J, C3H.SW and C57BL/6J mice, but not PL/J or BALB/c mice. SmOSP-specific T-cells from SJL/J, C3H.SW and C57BL/6J mice induced severe chronic clinical experimental autoimmune encephalomyelitis upon transfer. These findings indicate that autoimmune T-cell responses to OSP should be investigated in the context of multiple sclerosis.

Chemoimmunotherapy Reduces the Progression of Multiple Myeloma in a Mouse Model

Multiple myeloma (MM) is a B-cell malignancy characterized by clonal proliferation of malignant plasma cells in the bone marrow. Recently, we showed a correlation between increased ratios of functional regulatory T cells (Treg) and disease progression in a unique mouse model that mimics the human disease. Cyclophosphamide (CYC) is a cytotoxic alkylating agent widely used in chemotherapeutic regimens. Low-dose CYC was previously reported to selectively reduce Treg levels and to contribute to immunostimulation. Our objectives were (a) to determine whether treatment using a low-dose CYC could reduce MM progression and (b) to further characterize the modes of action underlying these effects. We found that both low- and high-dose CYC given to sick mice with hind limb paralysis resulted in the disappearance of the paralysis, the replacement of plasma tumor cells in the bone marrow by normal cell populations, and a significant prolongation of survival. However, only low-dose CYC treatment decreased the incidence of MM. Low-dose CYC rendered Tregs susceptible to apoptosis because of the downregulation of Bcl-xL and CTLA-4 in these cells, and a decreased production of interleukin 2 by effector CD4 cells. Moreover, using this treatment, we noted the recovery of IFN-γ–producing natural killer T cells and maturation of dendritic cells. Treatment of tumor-bearing mice with repeated administrations of low-dose CYC at longer time intervals (coinciding with the blocked renewal of Tregs) resulted in reduced tumor load, and the prevention or delay of disease recurrence, thereby breaking immune tolerance against MM tumor cells. Cancer Prev Res; 3(10); 1265–76. ©2010 AACR.

Amelioration of brain pathology and behavioral dysfunction in mice with lupus following treatment with a tolerogenic peptide

OBJECTIVE Central nervous system (CNS) involvement in systemic lupus erythematosus (SLE) is manifested by neurologic deficits and psychiatric disorders. The aim of this study was to examine SLE-associated CNS pathology in lupus-prone (NZBxNZW)F1 (NZB/NZW) mice, and to evaluate the ameliorating effects of treatment with a tolerogenic peptide, hCDR1 (human first complementarity-determining region), on these manifestations. METHODS Histopathologic analyses of brains from lupus-prone NZB/NZW mice treated with vehicle, hCDR1, or a control scrambled peptide were performed. The messenger RNA expression of SLE-associated cytokines and apoptosis-related molecules from the hippocampi was determined. Anxiety-like behavior was assessed by open-field tests and dark/light transfer tests, and memory deficit was assessed using a novel object recognition test. RESULTS Infiltration was evident in the hippocampi of the lupus-afflicted mice, and the presence of CD3+ T cells as well as IgG and complement C3 complex deposition was observed. Furthermore, elevated levels of gliosis and loss of neuronal nuclei immunoreactivity were also observed in the hippocampi of the mice with lupus. Treatment with hCDR1 ameliorated the histopathologic changes. Treatment with hCDR1 down-regulated the high expression of interleukin-1beta (IL-1beta), IL-6, IL-10, interferon-gamma, transforming growth factor beta, and the proapoptotic molecule caspase 8 in the hippocampi of the mice with lupus, and up-regulated expression of the antiapoptotic bcl-xL gene. Diseased mice exhibited increased anxiety-like behavior and memory deficit. Treatment with hCDR1 improved these parameters, as assessed by behavior tests. CONCLUSION Treatment with hCDR1 ameliorated CNS pathology and improved the tested cognitive and mood-related behavior of the mice with lupus. Thus, hCDR1 is a novel candidate for the treatment of CNS lupus.

Exogenous Dp71 is a dominant negative competitor of dystrophin in skeletal muscle.

Dystrophin, the protein which is absent or non-functional in Duchenne muscular dystrophy, consists of four main domains: an N-terminal actin binding domain, a rod shaped domain of spectrin-like repeats, a cysteine-rich domain and a unique C-terminal domain. In muscle, dystrophin forms a linkage between the cytoskeletal actin and a group of membrane proteins (dystrophin associated proteins). The N-terminal domain binds to the cytoskeletal actin and the association with the dystrophin associated proteins is mediated mainly by the cysteine-rich and C-terminal domains of dystrophin. The dystrophin gene also encodes two isoforms of non-muscle dystrophins and a number of smaller products consisting of the two C-terminal domains with different extensions into the spectrin-like repeat domain. Dp71, which consist of the C-terminal and the cysteine-rich domains of dystrophin, is the major product of the gene in all non-muscle tissues tested so far, but it is absent in differentiated skeletal muscle. In an attempt to understand the functions of Dp71, we produced transgenic mice over-expressing this protein in several tissues. The highest levels of exogeneous Dp71 were detected in skeletal muscle, in association with the sarcolemma. This resulted in muscle damage similar to that found in mice which lack dystrophin. The data indicates that Dp71 competes with dystrophin for the binding to the dystrophin associated proteins. Since Dp71 lacks the actin binding domain, it cannot form the essential linkage between the dystrophin associated proteins complex and the cytoskeleton.

A daidzein-daunomycin conjugate improves the therapeutic response in an animal model of ovarian carcinoma.

The use of daunomycin against neoplasms is limited due to its severe cardiotoxicity. The cytotoxicity of daunomycin can be minimized by linking it to an affinity tag. Since ovarian cancer cells are sensitive to isoflavone action, we synthesized a daidzein daunomycin conjugate. In MLS human ovarian cancer cells, the conjugate was shown to have a larger cytotoxic effect than daunomycin per se at a low concentration. The conjugate was then tested in vivo in mice carrying MLS xenografts. Tumour growth in the groups of conjugate and daunomycin was inhibited by >50% as compared to vehicle treated mice. In contrast to daunomycin treated mice, no weight reduction or death was seen in mice treated with the conjugate. In vivo imaging of the fluorescence signal generated by daunomycin indicated uptake of both conjugate and daunomycin by the tumour. Tumour fluorescence was, however, higher in the conjugate treated mice than in the daunomycin treated mice, thus suggesting specific delivery of the drug to the tumour. Histological examination of myocardial tissue indicated that only the daunomycin, but not conjugate treated mice showed cardiac damage. These results indicate that targeting of daunomycin via carboxymethyldaidzein retains daunomycins cytotoxic effects while averting its toxicity in an ovarian xenograft.

Histological findings in rabbits which died with symptoms of mucoid enteritis

Histological and aetiological similarities between rabbit mucoid enteritis and human ulcerative colitis are briefly discussed. Escherichia coli seems to be associated with the rabbit disease, and treatment aimed at this organism has been followed by a period of 4 years free from mucoid enteritis.

4-Nitrobenzylidene malononitrile reduces apoptosis-mediated liver injury in mice.

BACKGROUND Apoptosis plays a role in experimental and clinically related liver damage. Inhibitors of tyrosine kinases were shown to modulate apoptosis induced by different agents in various cell types. AIMS Investigation of the effect of 4-nitrobenzylidene malononitrile (belonging to the tyrphostins family which are selective inhibitors of protein tyrosine kinases) on apoptosis-mediated acute liver injury. METHODS Two murine experimental models exhibiting apoptosis-mediated liver injury were used: (1) mice treated with tumor necrosis factor-alpha and D-galactosamine; and (2) mice treated with anti-Fas antibody. Liver injury was assessed by serum levels of transaminases and by microscopic analysis. Apoptosis was assessed by labeling of apoptotic cells in the liver by the TUNEL assay and by determination of caspase-3 activity. RESULTS Pretreatment of mice with 4-nitrobenzylidene malononitrile reduced tumor necrosis factor-alpha/D-galactosamine-induced hepatotoxicity. TUNEL positive cells in sections from livers treated with vehicle (control), 4-nitrobenzylidene malononitrile, tumor necrosis factor-/d-galactosamine and tumor necrosis factor-alpha/D-galactosamine and 4-nitrobenzylidene malononitrile, were >0.2, >0.2, 49+/-2.3 and 4+/-0.2 per mm(2), respectively. 4-Nitrobenzylidene malononitrile also reduced hepatotoxicity induced by anti-Fas antibody. Caspase-3 activation induced by either tumor necrosis factor-alpha/D-glactosamine or by anti-Fas treatment, was reduced by pretreatment with N-nitrobenzylidene malononitrile. CONCLUSIONS The findings may provide a base for development of a new therapeutic modality to reduce apoptosis-mediated liver damage.