Macrophage and myofibroblast involvement in ischemic acute renal failure is attenuated by endothelin receptor antagonists

BACKGROUND: Endothelin (ET) may be a mediator of injury following ischemia-induced acute renal failure (ARF). ET receptor (ETR) antagonists have been reported to increase survival rates and lower serum creatinines when administered postrenal ischemia-reperfusion injury in the rat. Renal cellular and extracellular matrix responses to this therapy have not been addressed. METHODS: We investigated the use of ETR antagonists, PD 156707 (ETA) and SB 209670 (ETA and ETB) in the treatment of sublethal postischemic ARF. The right kidney of female Sprague-Dawley rats weighing approximately 200 g was removed. After five days, the left renal pedicle was occluded for 45 minutes. Twenty-four hours after renal ischemia, one of two ETR antagonists, PD 156707 (N = 7) or SB 209670 (N = 8), was administered. Experimental animals were compared with an ischemic group receiving only saline (N = 9). Three nephrectomized groups that did not undergo ischemia but that received infusions of saline (N = 6), PD 156707 (N = 6), and SB 209670 (N = 6), respectively, were also studied. Animals were sacrificed one week postischemia. Quantitation of monocytes and macrophages (Mo/Mphi), alpha-smooth muscle actin-positive myofibroblasts, and collagens type III and IV was performed by immunohistochemical staining. Cell kinetics were examined by staining for apoptosis with terminal deoxyuridine triphosphate (dUTP) nick end labeling and for proliferation with proliferating cell nuclear antigen. RESULTS: All ischemic groups of rats initially developed raised serum creatinine levels; however, no significant difference was observed between the groups (Kruskal-Wallis). Creatinines returned to preischemic values in all groups by the time of sacrifice. No significant difference in kidney weights or body weights was found between groups. Histologically, infiltration of Mo/Mphi was significantly reduced in groups treated with ETR antagonists (P < 0.001). The presence of myofibroblasts was also significantly reduced in the antagonist-treated groups (P < 0. 001). This was also paralleled by reduced quantities of collagen IV in the treated rat groups (P < 0.001). The interstitial area was also significantly greater in the saline group (P < 0.001). The amount of collagen III did not significantly differ between rat groups. Apoptosis was reduced (P < 0.001) by treatment with ETR antagonists, whereas proliferation was enhanced (P < 0.005). All non-ischemic groups showed no variation in any parameter studied at this time point. CONCLUSIONS: Treatment of ischemic ARF in the rat with ETR antagonists PD 156707 and SB 209670 attenuated cellular infiltration and matrix accumulation. An advantage of one antagonist over the other could not be determined in this study. The marked discrepancy between function and pathology (former unchanged, latter markedly improved) may be due to the time frame of this experiment, and longer outcome measures need to be assessed.     

Using sequence logos and information analysis of Lrp DNA binding sites to investigate discrepanciesbetween natural selection and SELEX

In vitro experiments that characterize DNA-protein interactions by artificial selection, such as SELEX,are often performed with the assumption that the experimental conditions are equivalent to natural ones. To test whether SELEX gives natural results, we compared sequence logos composed from naturally occurring leucine-responsive regulatory protein (Lrp) binding sites with those composed from SELEX-generated binding sites. The sequence logos were significantly different, indicating that the binding conditions are disparate. A likely explanation is that the SELEX experiment selected for a dimeric or trimeric Lrp complex bound to DNA. In contrast, natural sites appear to be bound by a monomer. This discrepancy suggests that in vitro selections do not necessarily give binding site sets comparable with the natural binding sites.     

Stuttering therapy in the elementary school setting: guidelines for clinician-teacher collaboration

This article describes guidelines for developing an effective collaborative/consultative approach to the treatment of stuttering in an elementary school setting. Four primary areas of shared clinician/teacher responsibility are discussed, which include: education, observation, facilitation of fluent speech, and generalization of speech fluency. Although the speech-language pathologist should initiate the teacher's involvement in a child's therapy, both the clinician and teacher should work toward developing a mutually influential partnership, through which the specific needs of individual children who stutter can be addressed in a comprehensive manner.     

Information content of individual genetic sequences

Related genetic sequences having a common function can be described by Shannon's information measure and depicted graphically by a sequence logo. Though useful for many purposes, sequence logos only show the average sequence conservation, and inferring the conservation for individual sequences is difficult. This limitation is overcome by the individual information ( R i) technique described here. The method begins by generating a weight matrix from the frequencies of each nucleotide or amino acid at each position of the aligned sequences. This matrix is then applied to the sequences themselves to determine the sequence conservation of each individual sequence. The matrix is unique because the average of these assignments is the total sequence conservation, ad there is only one way to construct such a matrix. For binding sites on polynucleotides, the weight matrix has a natural cut off that distinguishes functional sequences from other sequences. R i values are on an absolute scale measured in bits of information so the conservation of different biological functions can be compared with one another. The matrix can be used to rank-order the sequences, to search for new sequences, to compare sequences to other quantitative data such as binding energy or distance between binding sites, to distinguish mutations from polymorphisms, to design sequences of a given strength, and to detect errors in databases. The R i method has been used to identify previously undescribed but experimentally verified DNA binding sites. The individual information distribution was determined for E. coli ribosome binding sites, bacterial Fis binding sites, and human donor and acceptor splice junctions, among others. The distributions demonstrate clearly that the consensus sequence is highly unusual, and hence is a poor method to describe naturally occurring binding sites.     

Occupational exposures and amyotrophic lateral sclerosis. A population-based case-control study

This population-based case-control study was conducted in three countries in western Washington State to evaluate associations between workplace exposures and the risk of amyotrophic lateral sclerosis (ALS). Cases (n = 174) were all newly diagnosed with ALS by neurologists during 1990-1994, and controls (n = 348), who were matched according to age (+/-5 years) and sex, were identified via random-digit dialing or Medicare enrollment files. Four industrial hygienists blindly assessed detailed lifetime job histories for exposures to metals, solvents, and agricultural chemicals. Case-control comparisons were made for jobs held between 15 years of age and 10 years prior to the cases' dates of diagnosis. After adjustment for age and education, ever exposure to agricultural chemicals was associated with ALS (odds ratio (OR) = 2.0, 95% confidence interval (CI) 1.1-3.5); this association was observed separately in men (OR = 2.4, 95% CI 1.2-4.8) but not in women (OR = 0.9, 95% CI 0.2-3.8). Among men, the odds ratio for low exposure to agricultural chemicals (below the median level for exposed controls) relative to no exposure was 1.5 (95% CI 0.4-5.3), and for high exposure, it was 2.8 (95% CI 1.3-6.1) (p for trend = 0.03). Similar analyses based on the panel's assessment of exposures to metals and solvents showed no associations. These findings suggest an association between ALS and agricultural chemicals in men.     

Specific detection of the genus Serpulina, S. hyodysenteriae and S. pilosicoliin porcine intestines by fluorescent rRNA in situ hybridization

PURPOSE: Our purpose was to evaluate the utility of spectral imaging for multicolor, multichromosome enumeration in human interphase cell nuclei. METHODS: Chromosome-specific probes labeled with different fluorochromes or nonfluorescent haptens were obtained commercially or prepared in-house. Metaphase spreads, interphase lymphocytes, or blastomeres cells were hybridized with either 7 or 11 distinctly different probes. Following 46 hr of hybridization, slides were washed and detected using either a filter-based quantitative image processing system (QUIPS) developed in-house or a commercial spectral imaging system. RESULTS: The filter-based fluorescence microscope system is preferred for simultaneous detection of up to seven chromosome targets because of its high sensitivity and speed. However, this approach may not be applicable to interphase cells when 11 or more targets need to be discriminated. Interferometer-based spectral imaging with a spectral resolution of approximately 10 nm allows labeling of chromosome-specific DNA probes with fluorochromes having greatly overlapping emission spectra. This leads to increases in the number of fluorochromes or fluorochrome combinations available to score unambiguously chromosomes in interphase nuclei. CONCLUSIONS: Spectral imaging provides a significant improvement over conventional filter-based microscope systems for enumeration of multiple chromosomes in interphase nuclei, although further technical development is necessary in its application to embryonic blastomeres. When applied to preconception/preimplantation genetic diagnosis, presently available probes for spectral imaging are expected to detect abnormalities responsible for 70-80% of spontaneous abortions caused by chromosomal trisomies.