An improved method of preparing nuclei for absorption cytophotometry

An improved method of isolating nuclei from tissue for Feulgen-DNA measurements has been developed. The optimal nuclear isolation medium (NIM) was found to be a solution of 2% polyethylene glycol (PEG) and 0.6% NP40 in phosphate-buffered saline. The disaggregation procedure consists of gentle mechanical disruption of the tissues, followed by a 10 min incubation in the NIM at room temperature. The mixture is then syringed four times through a 27-gauge needle, and the nuclei are placed onto slides with a cytocentrifuge. Nuclei prepared in NIM without PEG had obvious DNA leakage and tended to form clumps. Addition of PEG to the NIM led to separation of nuclei without any DNA leakage, thus greatly increasing the accuracy of the DNA cytophotometry results. G0/G1 nuclei at the appropriate ploidy levels were found for non-transformed and transformed tissues prepared with this technique. In addition, S-phase liver nuclei prepared in this manner showed the expected incorporation of (3H) thymidine after a 1/2 hr pulse in vivo.     

Comparative Toxicities of Salts on Microbial Processes in Soil

Soil salinization is a growing threat to global agriculture and carbon sequestration, but to date it remains unclear how microbial processes will respond. We studied the acute response to salt exposure of a range of anabolic and catabolic microbial processes, including bacterial (leucine incorporation) and fungal (acetate incorporation into ergosterol) growth rates, respiration, and gross N mineralization and nitrification rates. To distinguish effects of specific ions from those of overall ionic strength, we compared the addition of four salts frequently associated with soil salinization (NaCl, KCl, Na₂SO₄, and K₂SO₄) to a nonsaline soil. To compare the tolerance of different microbial processes to salt and to interrelate the toxicity of different salts, concentration-response relationships were established. Growth-based measurements revealed that fungi were more resistant to salt exposure than bacteria. Effects by salt on C and N mineralization were indistinguishable, and in contrast to previous studies, nitrification was not found to be more sensitive to salt exposure than other microbial processes. The ion-specific toxicity of certain salts could be observed only for respiration, which was less inhibited by salts containing SO₄²⁻ than Cl⁻ salts, in contrast to the microbial growth assessments. This suggested that the inhibition of microbial growth was explained solely by total ionic strength, while ion-specific toxicity also should be considered for effects on microbial decomposition. This difference resulted in an apparent reduction of microbial growth efficiency in response to exposure to SO₄²⁻ salts but not to Cl⁻ salts; no evidence was found to distinguish K⁺ and Na⁺ salts.     

Steroidogenic acute regulatory protein/aldosterone synthase mediates angiotensin II-induced cardiac fibrosis and hypertrophy

Molecular Biology Reports - Aldosterone produced in adrenal glands by angiotensin II (Ang II) is known to elicit myocardial fibrosis and hypertrophy. This study was designed to test the hypothesis...     

Insights into the evolution of mammalian telomerase: Platypus TERT shares similarities with genes of birds and other reptiles and localizes on sex chromosomes

ABSTRACT: BACKGROUND: The TERT gene encodes the catalytic subunit of the telomerase complex and is responsible for maintaining telomere length. Vertebrate telomerase has been studied in placental mammals, fish, and the chicken, but less attention has been paid to other vertebrates. The platypus occupies an important evolutionary position, providing unique insight into the evolution of mammalian genes. We report the cloning of a platypus TERT (pTERT) ortholog, and provide a comparison with genes of other vertebrates. RESULTS: The pTERT encodes a protein with the high homology to marsupial TERT and avian TERT. Like the TERT of sauropsids and marsupials, as well as that of sharks and echinoderms, pTERT contains extended variable linkers in the N-terminal region suggesting that they were present already in basal vertebrates and lost independently in placental mammals and ray-finned fish. Several alternatively spliced pTERT variants structurally similar to avian TERT variants were identified. Telomerase activity is expressed in all platypus tissues similarly to cold-blooded animals and murine rodents. pTERT was localized on pseudoautosomal regions of sex chromosomes X3/Y2, expanding the homology between human chromosome 5 and platypus sex chromosomes. The synteny analysis suggests that TERT co-localized with sex-linked genes in the last common mammalian ancestor. Interestingly, female platypuses express higher levels of telomerase in heart and liver tissues than do males. CONCLUSIONS: pTERT shares many features with TERT of the reptilian outgroup, suggesting that pTERT represents the ancestral mammalian TERT. Features specific to TERT of eutherian mammals have, therefore, evolved more recently after the divergence of monotremes.