Mutation frequency declines during spermatogenesis in young mice but increases in old mice

Five percent of live-born human offspring will have a genetic disorder. Of these, 20% are because of germ-line de novo mutations. Several genetic diseases, such as neurofibromatosis and Duchenne muscular dystrophy, are associated with a high percentage of de novo germ-line mutations. Until recently, a direct analysis of spontaneous mutation frequencies in mammalian germ cells has been prevented by technical limitations. We have measured spontaneous mutation frequencies in a lacI transgene by using enriched populations of specific spermatogenic cell types. Similar to previously published results, we observed a lower mutation frequency for seminiferous tubule cell preparations, which contain all stages of spermatogenesis, relative to somatic tissues. We made the unexpected observation of a decline in mutation frequency during spermatogenesis, such that the mutation frequencies of type B spermatogonia and all subsequent stages of spermatogenesis are lower than the frequency for primitive type A spermatogonia. In addition, spermatogenic cells from old mice have significantly increased mutation frequencies compared with spermatogenic cells from young or middle-aged mice. Finally, the mutation frequency was observed to increase during spermiogenesis in postreplicative cell types when spermatogenic cells were obtained from old mice.     

Can tetraplex recombination models explain observations in induced mitotic gene conversion?

Induced mitotic gene conversion studies on the CYC1 gene of yeast have shown that the actual base pair changes, the types of changes (base substitution, deletion or addition) and the distances between mutations all affect gene conversion yields. In crosses between mutations less than four bases apart, gene conversion rates are as low as back mutation rates. The same mutants crossed to alleles more than five bases away may recombine 50-fold more. In crosses between mutations five or more base pairs apart, recombination rates varying by up to ten-fold are observed when comparing mutations at the same codon sites. The actual mutations in crosses affect recombination rates at these distances. The data rules out models in which mutants are repaired independently. Models with large gaps at the initiation site are ruled out if the mutants are within the gap. Recombination models are favoured in which both mutations can interact at a distance to affect the probability of recombination; such interactions may reflect the geometry of the recombinational junctions. The specific interactions proposed are that the actual mutations, and residual mismatches arising on excision resynthesis, affect both the further migration of the recombinational junction, and the probability that excision-repair will detect and correct residual mismatches. Junction models in which interactions are expected include those composed of base tetraplexes. The data is interpreted in terms of these models. Meiotic recombination data is consistent with these models.     

Molecular typing shows a high level of HLA class I incompatibility in serologically well matched donor/patient pairs: implications for unrelated bone marrow donor selection

In comparison with HLA-matched sibling bone marrow transplants, unrelated donor transplants are associated with increased graft-versus-host disease and graft failure. This is likely in part due to HLA incompatibilities not identified by current matching strategies. High resolution DNA-based typing methods for HLA class II loci have improved donor selection and treatment outcome in unrelated donor bone marrow transplantation. By using DNA-based typing methods for HLA-A and -B on a cohort of 100 potential bone marrow donor/patient pairs, we find that serological typing for HLA class I is limited in its ability to identify incompatibilities in unrelated pairs. Furthermore, the incompatibilities identified are associated with the presence at high frequency of alloreactive cytotoxic T-lymphocyte precursors. DNA typing also indicates that HLA-C mismatches are common in HLA-A and -B serologically matched pairs. Such mismatches appear to be significantly less immunogenic with respect to cytotoxic T-lymphocyte recognition, but are expected to influence natural killer cell activity. Thus, improved resolution of HLA class I shows many previously undisclosed mismatches that appear to be immunologically functional. Use of high resolution typing methods in routine matching is expected to improve unrelated donor selection and transplant outcome.     

Testosterone has potent, selective effects on the morphology of pelvic autonomic neurons which control the bladder, lower bowel and internal reproductive organs of the male rat

Although gonadal steroids are important determinants of the development and activity of various neuronal circuits in the brain and spinal cord, their function has been relatively poorly studied in the peripheral nervous system. In the present work, the effects of pre- and postpubertal castration were examined on the morphology of autonomic neurons that supply pelvic visceral organs in male rats. These neurons were identified by peripheral injection of fluorescent retrograde tracers and, in the major pelvic ganglion, were further classified as sympathetic or parasympathetic by means of immunostaining for tyrosine hydroxylase. Sizes of ganglion cell somata were indicated by areas of nucleated profiles in cryosections. The results show that, irrespective of whether castration was carried out at two or seven weeks-of-age, noradrenergic pelvic neurons that supply the vas deferens, prostate gland, urinary bladder or colon achieved only approximately 60% of the size of those in unoperated controls. In contrast, cholinergic pelvic neurons were unaffected by castration unless they supplied reproductive targets. Pre- and paravertebral sympathetic neurons that supplied the pelvic viscera were only slightly smaller following castration or were unchanged, depending on their target. All effects of castration were prevented by testosterone replacement following surgery. Androgen receptor-immunoreactivity was particularly prevalent in the nuclei of some pelvic ganglion neurons. The studies suggest that circulating androgens are essential for the maturation and maintenance of the structure of select groups of autonomic neurons that supply the viscera. The presence of androgen receptor immunoreactivity in many of these neurons indicates that direct neuronal effects of androgens are possible. However this does not exclude other less direct mechanisms of steroid action on neurons, such as by an effect on target organs, neurotrophic factor release or peripheral vascular supply. These studies point to the androgenic steroids as potentially important determinants of autonomic reflex function.     

Impairment and disability: their relation during stroke rehabilitation

Migration of non-volatile and volatile compounds from 'microwave and roasting bags' (MRB), made of Nylon 6,6 (and some Nylon 6), into chicken meat, skin, and juices during roasting (200 degrees C/2 h) in a conventional oven was determined. For measurement of migration of non-volatile compounds, cooked chicken was freeze-dried, extracted with methanol after addition of 2-azacyclononane (internal standard) and the extract cleaned-up using liquid-solid adsorption chromatography (silica gel). High performance liquid chromatography (HPLC) in the reverse phase mode using a linear gradient of methanol in water was used to quantify seven Nylon 6 and Nylon 6,6 cyclic monomers and oligomers of molecular mass up to 678 daltons. Migration into chicken was 7.48 micrograms/g (8.26 mg/bag; 3.94 micrograms/cm2), 16% of the total non-volatile compounds contained in the MRB material. Individual migrants were also quantified. Migration of one volatile compound, 2-cyclopentyl cyclopentanone, into the roast chicken parts was measured. Extraction with diethyl ether, using a modified Likens-Nickerson system of concurrent steam distillation-solvent extraction with an internal standard (cyclohexanone) was performed for 10 h. Gas chromatography/mass spectrometry (GC/MS) in the selected ion mode (SIM) was used for quantification. An average of 14.0 (+/- 4.36) micrograms/bag (or micrograms/chicken) migrated, being 0.08% of the total 2-cyclopentyl cyclopentanone present in MRB. Loss of volatile compounds to the atmosphere is believed to have occurred since there was another, more volatile compound (cyclopentanone), present in MRB, at levels higher than 2-cyclopentyl cyclopentanone, but this was not detected in roast chicken. In general, the transference of MRB components into roast chicken can be considered not to present a hazard.     

Choline derived from the phosphatidylcholine specific phospholipase D is not directly available for the CDP choline pathway in phorbol ester-treated C3H10T1/2 Cl 8 fibroblasts

To clarify the mechanism of capsaicin-induced primary neuronal cell death, newborn and adult rats were given a subcutaneous injection of capsaicin (50 mg/kg). Neonatal capsaicin injection induced neuronal apoptosis in the trigeminal ganglion. Apoptotic neurons had peripheral stacks of long parallel endoplasmic reticulum that are characteristic to primary neurons of the B-type, and exhibited nucleoplasmic condensation, nuclear shrinkage and cytoplasmic fragmentation. Light microscopically, apoptotic neurons exhibited a sign of DNA fragmentation as revealed by a nick end labelling method. The proportion of apoptotic cells was quite low during the first 12 h after capsaicin injection (<1%), rapidly increase to 10.44% by 24 h, and decreased to 0.29% by 48 h. Normal and vehicle control levels of apoptosis were <1%. Nerve growth factor (NGF, 0.5 mg/kg) simultaneously administered with capsaicin reduced the incidence of apoptosis by about 35% at 24 h post-injection. Neonatal transection of the infraorbital nerve induced neuronal apoptosis similar to that produced by the neonatal capsaicin in the maxillary division of the trigeminal ganglion. Unlike capsaicin, however, the neurotomy-induced apoptosis was seen in neurons of both the A- and B-types. Neither the capsaicin injection nor the neurotomy induced apoptosis in adult rats, though mitochondrial swelling similar to that seen at 0.5 h after neonatal capsaicin was observed after capsaicin injection in adults. The results indicate that the capsaicin-induced and nerve injury-induced primary neuronal damages in newborn rats share a common final pathway, apoptosis.     

Macular hole secondary to retrobulbar needle perforation

An unusual case of cerebral toxoplasmosis leading to a fatal vehicular crash is presented. Massive intracerebral hemorrhage, a rare complication of cerebral toxoplasmosis in persons with acquired immunodeficiency syndrome (AIDS), caused sudden onset of increased intracranial pressure that led to loss of control of the vehicle and an ensuing fatal traffic crash.