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Biomedical subjects

L L Mays-Hoopes

Publications and source records attributed to L L Mays-Hoopes.

7 recordsLinked to original sources

Preparation of spermatogonia, spermatocytes, and round spermatids for analysis of gene expression using fluorescence-activated cell sorting.

A new method is described for the purification of spermatogenic cell populations from mouse testis. Through use of this method, it is possible to purify leptotene, zygotene, and pachytene primary spermatocytes as well as round spermatids from adult mouse testis. In addition, spermatogonial populations can be purified from mice at 9 days postpartum. The leptotene and zygotene primary spermatocytes that can be prepared by this method are impossible to separate successfully by the unit gravity method. The cells were used to prepare RNA for reverse transcriptase-polymerase chain reactions.

Animals↗

DNA methylation in aging and cancer.

DNA methylation processes appear to be disturbed in senescing mice and also in some tumor cells. Although it is possible that age-related demethylation of DNA could predispose cells to carcinogenic transformation, present evidence does not favor this hypothesis.

Aging↗

DNA methylation in aging of mice.

Methylation of cytidine residues of DNA appears to be involved in the control of gene expression; therefore, hypomethylation of the DNA can be considered to be an active rather than a passive process. Previous studies of mammalian DNA methylation during aging have produced an assortment of results. In this study, we have examined the change in DNA base composition, including the change in 5-methyldeoxycytidine (m5dC) contents with age of mice. Livers pooled from 6 mice from each of six age groups between 6 and 31 months have been subjected to a sensitive analytical technique (HPLC). The DNA composition of different age groups is very consistent in most aspects. The ratio of (dA + dT)/(dG + dC + m5dC) as well as the sum of dC and m5dC remain constant throughout the animal's lifespan. However, a consistent gradual decline in m5dC content is noted as the age increases to 24 months. Thus, the 6-month-old animal pool exhibits the largest amount of m5dC (1.67 +/- 0.2%), which is reduced consistently as the animal's age reaches 24 months. This decrease in m5dC is accompanied by an increase in dC. No further decrease in m5dC occurs after 24 months; in fact, the data could indicate an increase after that age. No dTs are apparently produced by deamination of m5dC.

Aging↗

Function and fidelity of aging tRNA: in vivo acylation, analog discrimination, synthetase binding, and in vitro translation.

The liver transfer RNAs for valine and lysine were completely acylated in vivo, as judged by periodate oxidation, at 4 and 24 months of age in male Sprague-Dawley rats. In vitro acylation capacity for whole tRNA populations from rat livers is decreased, but this is evidently not deleterious in vivo. Several halogenated phenylalanines were synthesized and their effects upon acylation capacity for phenylalanine were examined. Synthetases bound to young (3 month) and old (24 month) tRNAs discriminated differently between p-chlorophenylalanine and authentic phenylalanine; synthetase with young tRNA was less able to discriminate than with old tRNA. Purified tRNAphe from old rats did not form ultraviolet-induced crosslinks to purified phenylalanyl tRNA synthetase as well as young tRNAphe. In vitro translation of encephalomyocarditis virus, hemoglobin, and ovalbumin mRNAs was effective, using tRNAs of young or old Sprague-Dawley or Fischer 344 rat livers, although, when the old tRNA was supplied, the product synthesized per unit tRNA was reduced. All of the protein products were synthesized with all tRNAs, as shown by sodium dodecyl sulfate polyacrylamide gel electrophoresis. We conclude that tRNA is capable of normal functions in livers of aging rats, is probably modification deficient, and is unlikely to produce protein errors.

Acylation↗

Methylation and rearrangement of mouse intracisternal a particle genes in development, aging, and myeloma.

Sequences of DNA that hybridize on Southern blots with cloned intracisternal A-particle (IAP) sequences have been examined in genomic DNAs of neonatal mice, livers of adult mice (3, 6, 12, 18, 24, and 26 months old), and the solid myeloma tumor MOPC-315. The isoschizomers HpaII (CCGG or mCCGG) and MspI (CCGG or CmCGG) were used to assess methylation. All the DNAs produced a major 0.5-kilobase MspI fragment that hybridizes with IAP probe. Only the myeloma DNA, and to a much lesser degree DNA from senescent mouse liver, produced this fragment in HpaII digest; the other DNAs all had IAP sequences resistant to HpaII digestion. These sequences thus become fully methylated to CmCGG early and remain so in adult life, except in the myeloma cells that are expressing the IAP genes. An increase in MspI-sensitive sites in IAP gene-containing DNA was observed in aging mice. The probe used to assess methylation, a 0.8-kilobase fragment produced by BamHI-HindIII double digestion, is common to several cloned IAP genes and is part of a region of DNA which is conserved in genomes of all mouse tissues. The probe hybridized to 1.5- and 1.4-kilobase doublet bands produced by BamHI, HindIII, and EcoRI triple digestions of neonatal DNA. These two bands were found in neonatal livers of Swiss Webster, BALB/c, and C57BL/6J mouse strains, showed less in adult liver, and were barely detectable in senescent livers from C57BL/6J mice.

Aging↗