Androgen receptor YAC transgenic mice carrying CAG 45 alleles show trinucleotide repeat instability

X-linked spinal and bulbar muscular atrophy (SBMA) is caused by a CAG repeat expansion in the first exon of the androgen receptor (AR) gene. Disease-associated alleles (37-66 CAGs) change in length when transmitted from parents to offspring, with a significantly greater tendency to shift size when inherited paternally. As transgenic mice carrying human AR cDNAs with 45 and 66 CAG repeats do not display repeat instability, we attempted to model trinucleotide repeat instability by generating transgenic mice with yeast artificial chromosomes (YACs) carrying AR CAG repeat expansions in their genomic context. Studies of independent lines of AR YAC transgenic mice with CAG 45 alleles reveal intergenerational instability at an overall rate of approximately 10%. We also find that the 45 CAG repeat tracts are significantly more unstable with maternal transmission and as the transmitting mother ages. Of all the CAG/CTG repeat transgenic mice produced to date the AR YAC CAG 45 mice are unstable with the smallest trinucleotide repeat mutations, suggesting that the length threshold for repeat instability in the mouse may be lowered by including the appropriate flanking human DNA sequences. By sequence-tagged site content analysis and long range mapping we determined that one unstable transgenic line has integrated an approximately 70 kb segment of the AR locus due to fragmentation of the AR YAC. Identification of the cis - acting elements that permit CAG tract instability and the trans -acting factors that modulate repeat instability in the AR YAC CAG 45 mice may provide insights into the molecular basis of trinucleotide repeat instability in humans.      

Development ofEimeria dispersa Tyzzer, 1929, from bobwhite quail (Colinis virginianus), in bovine kidney cell cultures

Summary The development ofEimeria dispersa Tyzzer, a parasite of bobwhite quail, in Madin-Darby bovine kidney cell cultures was investigated. Excysted sporozoites were inoculated into Leighton tubes containing cell monolayers on glass coverglasses and maintained in minimum essential medium supplemented with heat-inactivated fetal calf serum. Sporozoites became intracellular within 2 h. Sporozoite-shaped schizonts, schizonts with developing merozoites, and mature first-generation schizonts were seen 24 h postinoculation. Intracellular first-generation merozoites, second-generation trophozoites, and early second-generation schizonts containing two nuclei were first observed 72 h postinoculation. Second-generation schizonts containing developing merozoites as well as mature second-generation schizonts were first seen 96h postinoculation. Gametogony was not observed.DM developing merozoite - HN host nucleus - IM intracellular merozoite - M merozoite - N nucleus - R refractile body - RB residual body - V parasitophorous vacuole     

Glomerular permeability: ultrastructural quantitative studies relating proteinuria to pathologic features in murine lupus nephritis.

The pathogenesis of proteinuria associated with immune complex disease is incompletely understood. A quantitative electron-microscopic study was undertaken to determine the relative contribution of lesions in capillary loops and mesangial basement membrane areas and their possible correlations to urinary protein excretion data. Pathologic features including the loss of foot processes (and slit diaphragms), the formation of junctional complexes in visceral epithelium, and the distribution of immune complexes in basement membrane were assessed in glomeruli of mice with lupus nephritis. Swiss albino mice served as control animals. In control animals the distribution of split pores per unit length of basement membrane was approximately 60% higher in capillary loop compared to mesangial basement membrane areas. In mice with lupus nephritis, the reduction in the number of slit pores per unit length of basement membrane to 30% or less of normal, the formation of epithelial junctions, and the relative distribution of immune complexes were not statistically different in capillary versus mesangial basement membrane areas. Animals with murine lupus showed poorly selective proteinuria, but the correlation between features studied and extent of protein excretion was poor. The results of these studies 1) establish the relative distribution of slit pores in mesangial and peripheral loop basement membrane, 2) demonstrate that glomerular changes associated with immune complex deposition are comparable in capillary and mesangial basement membrane areas, and 3) are consistent with a focal and nonuniform alteration in glomerular permeability properties associated with immune complex disease.     

The effect of dietary supplementation with n-3 polyunsaturated fatty acids on the synthesis of interleukin-1 and tumor necrosis factor by mononuclear cells

We examined whether the synthesis of interleukin-1 or tumor necrosis factor, two cytokines with potent inflammatory activities, is influenced by dietary supplementation with n-3 fatty acids. Nine healthy volunteers added 18 g of fish-oil concentrate per day to their normal Western diet for six weeks. We used a radioimmunoassay to measure interleukin-1 (IL-1 beta and IL-1 alpha) and tumor necrosis factor produced in vitro by stimulated peripheral-blood mononuclear cells. With endotoxin as a stimulus, the synthesis of IL-1 beta was suppressed from 7.4 +/- 0.9 ng per milliliter at base line to 4.2 +/- 0.5 ng per milliliter after six weeks of supplementation (43 percent decrease; P = 0.048). Ten weeks after the end of n-3 supplementation, we observed a further decrease to 2.9 +/- 0.5 ng per milliliter (61 percent decrease; P = 0.005). The production of IL-1 alpha and tumor necrosis factor responded in a similar manner. Twenty weeks after the end of supplementation, the production of IL-1 beta, IL-1 alpha, and tumor necrosis factor had returned to the presupplement level. The decreased production of interleukin-1 and tumor necrosis factor was accompanied by a decreased ratio of arachidonic acid to eicosapentaenoic acid in the membrane phospholipids of mononuclear cells. We conclude that the synthesis of IL-1 beta, IL-1 alpha, and tumor necrosis factor can be suppressed by dietary supplementation with long-chain n-3 fatty acids. The reported antiinflammatory effect of these n-3 fatty acids may be mediated in part by their inhibitory effect on the production of interleukin-1 and tumor necrosis factor.     

Coccidioidin and merthiolate in previously sensitized animals.

The effect of merthiolate, which is used as a preservative in skin test materials, on skin test reactions was determined in guinea pigs. In four groups of animals, merthiolate in basal medium produced skin tests at 24 and 48 h characterized by erythema and/or induration in an intermediate region, i.e., 5 plus or minus 2.2 mm. One of the four groups of animals was a nonsensitized control group. The other three groups were subcutaneously sensitized with (i) merthiolate and saline, (ii) killed Coccidioides immitis arthrospores, and (iii) merthiolate with killed C. immitis arthrospores. Coccidioidin only and merthiolate in coccidioidin produced positive delayed results in groups 3 and 4, which were sensitized with arthrospores. A synergistic effect of merthiolate and coccidioidin was observed in animals of group 4 sensitized by merthiolate with killed C. immitis arthrospores. This effect was observed at 24 h when positive reactions of coccidioidin with merthiolate were significantly greater than skin tests with plain coccidioidin.     

Cyclosporine and alpha-interferon do not attenuate morphine withdrawal in rats but do impair thermoregulation

Immunomodulating drugs as diverse as alpha-interferon and cyclosporine have been reported to attenuate physical signs of morphine withdrawal in rats. On the basis of these results, the immune system has been claimed to be involved in opiate addiction. To assess whether this is the case, the effects of alpha-interferon and cyclosporine were studied on objective signs of morphine withdrawal in morphine-dependent rats. Rats made dependent upon morphine by implantation of a 75-mg morphine pellet were challenged three days later by naloxone (1 mg/kg). Pretreatment with alpha-interferon (150 U/g) or cyclosporine (15 mg/kg) did not attenuate the reduction in body weight or the behavioral suppression induced by naloxone in morphine-dependent rats trained to press a lever for food reinforcement on a fixed-ratio 10 schedule. Alpha-interferon pretreatment blocked the capacity of naloxone to decrease body temperature in these rats and actually induced an hyperthermic response. In contrast, cyclosporine tended to enhance the drop in body temperature induced by naloxone. This last effect was more striking when the rats were placed in a cold room at 3.5 degrees C. Cyclosporine by itself induced a drop in body temperature in normal rats exposed to 3.5 degrees C. These results indicate that alpha-interferon and cyclosporine impair thermoregulation but do not directly interfere with morphine withdrawal signs.     

Correlation of proliferation of lung epithelium with intramuscular sensitization and complement-fixing antibody to respiratory syncytial virus in the golden hamster.

Intramuscular sensitization of hamsters with several forms of respiratory syncytial virus (RSv) caused proliferation of lung epithelium. In contrast, intranasal injection of live virus rarely resulted in this phenomenon. A correlation existed between proliferation of lung epithelium and presence of complement-fixing antibody, but not between lung disease and delayed skin reactions. Complement-fixing antibody to RSv was found to be independent of the influence of the thymus.     

Spleen cell factor inhibits lymphoproliferation, abnormal Ia expression and overt autoimmunity in MRL-lpr mice

Autoimmune MRL-lpr have an abnormal pattern of lymphokine production. In our attempt to repair this defect, MRL-lpr mice were prophylactically treated daily with a lectin stimulated rat spleen cell product rich in interleukin-2. Therapy inhibited the lymphoid hyperplasia of the unique lymphocytes regulated by the lpr gene, suppressed the enhanced supranormal expression of Ia on peritoneal macrophages and protected this strain from autoimmune renal injury. Purified recombinant interleukin-2 alone did not prevent autoimmune disease expression. Thus, a spleen cell product other than interleukin-2 can ameliorate the aggressive course of lymphoproliferation and autoimmunity in MRL-lpr mice.