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PubMed1990

The TEACL method of DNA-DNA hybridization: technical considerations.

This paper emanated from a conference concerning the value, accuracy, and technical considerations of DNA-DNA hybridization for evolutionary studies. Our laboratory has been performing the so-called TEACL (tetraethylammonium chloride) method, and we have amassed sufficient data to indicate that this method is very powerful if performed properly with correct analyses. Here we address five technical considerations: (1) We present empirical data that size correction for tracer length is legitimate and accurate. (2) We show that the error of delta Tm measurement does not significantly increase with increasing distance up to at least 10 degrees C. (3) The error distribution for delta Tm does not deviate from the expected normal distribution indicating parametric statistics are probably legitimate for analyses. (4) Using a known phylogeny we examined the resolving power of the technique by showing that at least five taxa can be correctly placed in phylogenies with a maximum delta Tm of 2.5 degrees C. (5) To date, all our data sets based on DNA-DNA hybridization are very robust with respect to analytical procedures in that every algorithm used on the data sets has yielded identical trees with nearly identical branch lengths. Nevertheless, we point out that theoretical analyses of distance data (as generated by DNA-DNA hybridization) are lacking, especially with regard to tests of the molecular clock hypothesis.

Algorithms↗

PubMed1975

An automated, direct method for measuring adipocyte cell size.

Current methods for direct determination of fat cell size are optical and non-automated. They are thus tedious, but have the advantage of providing estimates of the variance of a cell size distribution, as well as a measure of the mean cell size. An indirect method based on counting the number of osmium-fixed fat cells derived from a known wet weight of adipose tissue is also available. This method is automated and thus rapid, but does not provide information about the variance of the cell size distribution. In the present paper we describe a direct, automated method of fat cell sizing that provides estimates for both mean cell size and the variance of the cell size distribution. The correlation between our method and in indirect method based on counting osmium-fixed cells was 0.96. Transformation of the volume distribution to diameters indicated that cell diameters appeared to be normally distributed, confirming the observations of others.

Adipose Tissue↗

PubMed1991

A nonparametric approach to analyzing large human populations.

There are many statistical techniques that require the assumption that the population being studied is normally distributed--regression analysis, multivariate analysis, time series analysis, and so on. Unfortunately, as the development of survey sampling has long acknowledged, large human populations are usually stratified into several different subpopulations. Since the boundaries between the strata are somewhat blurred, they are not independent, so the overall distribution of the population tends to be multimodal rather than normal. In this paper, a technique is developed to find these multimodal techniques using nonparametric density estimation. Its effectiveness is demonstrated by means of an example.

Analysis of Variance↗

PubMed1995

The twitch interpolation technique for study of fatigue of human quadriceps muscle.

The aim of the study was to examine if the twitch interpolation technique could be used to objectively measure fatigue in the quadriceps muscle in subjects performing submaximally. The 'true' maximum isometric quadriceps torque was determined in 21 healthy subject using the twitch interpolation technique. Then an endurance test was performed in which the subjects made repeated isometric contractions at 50% of the 'true' maximum torque for 4 s, separated by 6 s rest periods. During the test, the force response to single electrical stimulation (twitch amplitude) was measured at 50% and 25% of the estimated maximum torque. In 10 subjects, the test was repeated 2-4 weeks later. Twitch amplitudes at 50% of maximum torque declined exponentially with time in 20 of 21 subjects. The distribution of the exponential rate constant was skewed with a mean of 4.6 h-1 and range of 0.3-21.5 h-1. After logarithmical transformation, the distribution of the exponential rate constant fitted closely to a normal distribution, and the inter-individual variation was SD = 1.15 compared to an intra-individual variation of 0.29. The coefficient of correlation for repeated determination was 0.91 (P < 0.001, n = 10). In conclusion, the twitch technique can be used for objectively measuring fatigue of the quadriceps muscle.

Adult↗

PubMed1995

Architectural changes of the cortico-spinal system in the dystrophin defective mdx mouse.

The mutant mdx mice which lack the protein dystrophin are an animal model of Duchenne muscular dystrophy. We studied the organization of the cortico-spinal (CS) system in mdx mice using the horseradish peroxidase retrograde tracing technique. Tracer injections were placed in the cervical spinal cord of mutant and control mice. The tangential and radial distribution of CS labeled neurons were similar in mdx and normal mice. Conversely, the absolute number and the cell packing density of labeled CS neurons were considerably lower in mdx than in controls. In mdx, the average size of CS cells was smaller while the perikaryal sizes displayed a normal distribution. In addition, CS neurons of mdx appeared round-shaped compared to the pyramidal cells labeled in control animals. The structural modifications described here should prompt a reconsideration of the involvement of central nervous system in the dystrophin deficient mdx mice.

Animals↗

PubMed1987

Identification of rat osteoclasts in bone smears with quantification of acid phosphatase activity in vitamin D deficiency.

Vitamin D deficiency may depress bone formation but its effect on bone resorption is not well defined. As an index of bone resorption, the activity of acid phosphatase, a lysosomal enzyme found in osteoclasts, was quantitated in situ from a bone tissue smear preparation. Activity of the enzyme, measured by integrative microdensitometry increased linearly from 25 to 60 min. The distribution of activity in osteoclasts quantitated appeared to follow a normal distribution with a median value of 0.19 integrated optical density units. Animals treated with vitamin D or 1,25 dihydroxycholecalciferol had significantly increased activity of acid phosphatase in osteoclasts compared to animals which were vitamin D deficient. The increase of acid phosphatase activity averaged 63%. Vitamin D or its metabolites may have a permissive effect on the action of parathyroid hormone or act directly to increase bone resorption.

Acid Phosphatase↗

PubMed2005

Synthesis and characterization of porous beta-tricalcium phosphate blocks.

Porous beta-tricalcium phosphate (beta-TCP) blocks with four different macropore sizes (pore larger than 50 microm were synthesized using "calcium phosphate emulsions", and characterized by optical, geometrical, gravimetric, and radiological methods. The reproducibility of the synthesis method was excellent. Moreover, the macropore size could be easily controlled without modifying the microporosity (pore smaller than 50 microm) or the total porosity (microporosity+macroporosity). Based on the initial composition of the blocks and their final apparent density, the microporosity, macroporosity, and the total block porosity were calculated to be close to 21%, 54%, and 75%, respectively. These values were confirmed by microcomputed tomography (microCT). The mean macropore diameters were close to 150, 260, 510 and 1220 microm, as measured optically. Consistently lower values (25% lower) were obtained by microCT, but the linear correlation between microCT and optical method was high (r(2)>0.97). The macropore size distribution calculated from microCT scans appears to be narrow and normally distributed. The very good correlation between the results of the various methods and the possibility to determine the pore size distribution suggest that microCT is an ideal tool to non-destructively characterize macroporous calcium phosphate bone substitutes.

Biocompatible Materials↗

PubMed1997

Saccadic reaction times: a statistical analysis of multimodal distributions.

The distributions of saccadic reaction times (SRT) often deviate from unimodal normal distributions. An excess-mass procedure was used to detect peaks in 963 data sets containing 90,927 reaction times from 170 subjects. About 55% showed one, 30% two, 12% three and 3% four peaks. According to their clustering along the reaction time scale the modes could be classified into express (90-120 msec), fast regular (135-170 msec) and slow regular (200-220 msec) modes. Among the unimodal distributions 29% had peaks in the range of the express mode and 46% had peaks in the range of the fast regular mode. Therefore, 87% of the data sets support the notion of saccadic reaction time distributions being the superposition of three modes. All experimental distributions were fitted by as many gamma distributions as determined by the excess-mass test. The significance of the multimodality for saccade generation processes is discussed.

Fixation, Ocular↗

PubMed1997

Measurement of patient positioning errors in three-dimensional conformal radiotherapy of the prostate.

PURPOSE/OBJECTIVE: To determine the spatial distribution of setup errors for patients treated with six-field, three-dimensional (3D) conformal radiation therapy for prostate cancer. METHODS AND MATERIALS: Port films for 50 patients were analyzed retrospectively. The port films were digitized and compared, using image registration software, to simulator films (representing the ideal treatment position). Patient positioning uncertainty for a given setup was determined using port films from three projections, two obliques, and one lateral. A total of 1239 port films and 300 simulator films were analyzed for the study. Patient position was analyzed for out-of-plane rotations and time trends over the course of treatment. RESULTS: The distribution of systematic setup errors for the 50 patients, defined as the mean patient displacement for the treatment course, had a mean and standard deviation (SD) of (-0.1 +/- 1.9) mm, (0.4 +/- 1.4) mm, and (-0.3 +/- 1.3) mm in the mediolateral (ML), superior-inferior (SI) and anterior-posterior (AP) directions, and (-0.1 +/- 0.2) for rotational errors. The distribution of random setup errors about the mean approximated a normal distribution and the standard deviations for the population of patients in the ML, SI, and AP directions, were 2.0 mm, 1.7 mm, and 1.9 mm, respectively. The distribution of out-of-plane rotations had 1 SD of 0.9 degrees and 0.6 degrees about the SI and AP axes. Ten of the 50 patients demonstrated a statistically significant time trend in their setup position resulting in shifts ranging from 2 to 7 mm. CONCLUSIONS: The setup verification protocol appears to minimize systematic setup errors to a level that approaches the sensitivity of the image registration technique. The random day to day fluctuations, represented by the average values of the standard deviations, are minor in comparison to the currently used margins, which further emphasizes the effectiveness of this protocol in conjunction with the use of the immobilization device.

Analysis of Variance↗

PubMed1998

Assessment of neuropsychological changes in patients with arteriovenous malformation (AVM) after radiosurgery.

PURPOSE: The purpose of this study was to investigate neuropsychological effects of radiosurgery in patients with cerebral arteriovenous malformation (AVM), with special focus on attention and memory. This report describes the study setup and presents the first results during a follow-up of up to 1 year. MATERIALS AND METHODS: Seventy-nine patients were studied before, acutely after radiosurgery, and during the regular follow-up (subacute phase: Weeks 6-12, chronic phase: Months 6-12). Radiosurgery was performed using a modified linear accelerator (minimum doses to the target volume: 15-22 Gy, median 20 Gy). Estimated whole brain dose was 0.5 to 2 Gy. Neuropsychological testing included assessment of general intelligence (Wechsler Adult Intelligence Scale), attention (modified Trail-Making Test A, Digit Symbol Test, D2 Test, Wiener Determination Machine) and memory (Rey Auditory Verbal Learning Test, Benton Visual Retention Test). During follow-up, alternate test versions were used. Neuropsychological deficits were defined as a test score of at least one standard deviation (SD) below the mean of the normal distribution. RESULTS: The pretherapeutic evaluation revealed marked deviations from the normal population; 24% had deficits in intelligence (range 23-31% in different subtests), attention (35%, 23-59%) and memory (48%, 31-61%). The overall percentage of aberrant results was reduced by 12% (memory) to 14% (attention) in the chronic phase up to 12 months after therapy. The improvement in test scores was significant (p < 0.05) in 3 of 4 subtests of attention functions. CONCLUSIONS: The acute tolerance of radiosurgery seems to be very good in these patients, showing no relevant increase in number of patients with neuropsychological deficits. Although the long-term follow-up needs to be further increased, our data indicate a tendency to slight improvement in the overall neuropsychological performance of AVM patients in the chronic phase after radiosurgery.

Adult↗

PubMed2000

Predicting long-term response to selection.

Lande's equation for predicting the response of trait means to a shift in optimal trait values is tested using a stochastic simulation model. The simulated population is finite, and each individual has a finite number of loci. Therefore, selection may cause allele frequencies and distributions to change over time. Since the equation assumes constant genetic parameters, the degree to which such allelic changes affect predictions can be examined. Predictions are based only on information available at generation zero of directional selection. The quality of the predictions depends on the nature of allelic distributions in the original population. If allelic effects are approximately normally distributed, as assumed in Lande's Gaussian approximation to the continuum-of-alleles model, the predictions are very accurate, despite small changes in the G matrix. If allelic effects have a leptokurtic distribution, as is likely in Turelli's 'house-of-cards' approximation, the equation underestimates the rate of response and correlated response, and overestimates the time required for the trait means to reach their equilibrium values. Models with biallelic loci have limits as to the amount of trait divergence possible, since only two allelic values are available at each of a finite set of loci. If the new optimal trait values lie within these limits, predictions are good, if not, singularity in the G matrix results in suboptimal equilibria, despite the presence of genetic variance for each individual trait.

Gene Frequency↗

PubMed2006

Proton permeation into single vesicles occurs via a sequential two-step mechanism and is heterogeneous.

This article describes the first single-vesicle study of proton permeability across the lipid membrane of small (approximately 100 nm) uni- and multilamellar vesicles, which were composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). To follow proton permeation into the internal volume of each vesicle, we encapsulated carboxyfluorescein, a pH-sensitive dye whose fluorescence was quenched in the presence of excess protons. A microfluidic platform was used for easy exchange of high- and low-pH solutions, and fluorescence quenching of single vesicles was detected with single-molecule total internal reflection fluorescence (TIRF) microscopy. Upon solution exchange and acidification of the extravesicular solution (from pH 9 to 3.5), we observed for each vesicle a biphasic decay in fluorescence. Through single-vesicle analysis, we found that rate constants for the first decay followed a Poisson distribution, whereas rate constants for the second decay followed a normal distribution. We propose that proton permeation into each vesicle first arose from formation of transient pores and then transitioned into the second decay phase, which occurred by the solubility-diffusion mechanism. Furthermore, for the bulk population of vesicles, the decay rate constant and vesicle intensity (dependent on size) correlated to give an average permeability coefficient; however, for individual vesicles, we found little correlation, which suggested that proton permeability among single vesicles was heterogeneous in our experiments.

Cell Membrane Permeability↗

PubMed2007

The relationship between the pleiotropic phenotypic effects of a mutation fixed by selection.

A pleiotropic model of mutation is presented that allows for correlations between the effects of a new mutation and for the distribution of mutational effects to vary from being leptokurtic to normally distributed. Using this model I quantify how selection transforms the correlation between the effects of a new (random) mutation into the correlation between the effects of a mutation that is fixed by selection and contributes to an adaptation. Results suggest that under most conditions the correlation between the effects of a fixed mutation is less than the correlation between the effects of a new mutation. I also generalize previous results that quantified the expected size of a fixed mutation's effect on a character given an observed effect of that mutation on another character. In agreement with previous results, work here suggests that as the observed effect becomes large and beneficial the expected effect on another character approaches the expected effect of a new (random) mutation given the observed effect. Lastly, these theoretical results are related to recent empirical work that found beneficial mutations had a positive correlation in their pleiotropic effects.

Models, Genetic↗

PubMed2002

Elevated monocyte chemotactic protein-1 in amniotic fluid is a risk factor for pregnancy loss.

OBJECTIVE: Pregnancy loss after mid-trimester amniocentesis occurs in 0.5-1% of cases and is frequently attributed to the procedure. Accumulating evidence implicates a pre-existing, but clinically silent, intra-amniotic inflammation in the etiology of adverse pregnancy outcome after mid-trimester amniocentesis. Monocyte chemotactic protein-1 (MCP-1) is a potent chemokine produced by a wide variety of cells during the course of an inflammatory response. This study was designed to assess if the amniotic fluid concentration of this chemokine identifies patients at risk for spontaneous abortion and/or fetal death. METHOD: A retrospective case-control study of women who had a mid-trimester amniocentesis was designed. Cases (n = 10) consisted of patients who had a spontaneous pregnancy loss after the procedure, while the control group (n = 84) consisted of patients who had a normal pregnancy outcome after mid-trimester amniocentesis. MCP-1 was measured by a specific enzyme immunoassay (sensitivity, 18.3 pg/ml). The Kolmogorov-Smirnov test was utilized to assess normal distribution of the data. Logarithmic transformation was applied to achieve normality. Statistical analysis was performed using Student's t test. A receiver operating characteristic (ROC) curve analysis was used to select a cut-off to dichotomize amniotic fluid concentrations of MCP-1. RESULTS: MCP-1 was detectable in all amniotic fluid samples. Patients who had a mid-trimester amniocentesis and a subsequent pregnancy loss had a higher mean amniotic fluid log MCP-1 concentration than those with a normal pregnancy outcome (pregnancy loss, mean 2.95 +/- 0.19 pg/ml vs. normal outcome, mean 2.78 +/- 0.19 pg/ml; p = 0.01). A cut-off of > 765 pg/ml was selected by ROC curve analysis (area under the curve, 0.74; p = 0.01). An amniotic fluid concentration of MCP-1 above this level was strongly associated with pregnancy loss (odds ratio, 7.35; 95% confidence interval, 1.7-31.1), a sensitivity of 70%, and a specificity of 76%. CONCLUSION: A subset of women who had a pregnancy loss after a mid-trimester amniocentesis had higher concentrations of the chemokine MCP-1 than those who had a normal pregnancy outcome. Subclinical intra-amniotic inflammation is a risk factor for pregnancy loss after mid-trimester amniocentesis. This observation may have medicolegal and clinical implications. An elevated MCP-1 concentration in amniotic fluid of patients with a pregnancy loss after a mid-trimester amniocentesis indicates that a pathological condition was present at the time of the procedure.

Abortion, Spontaneous↗

PubMed1986

Evaluation of serum CA125 values in healthy individuals and pregnant women.

CA125 is an antigenic determinant associated with human epithelial ovarian carcinoma. This study was undertaken to evaluate the distribution of serum CA125 levels and the effect of smoking on these levels among healthy individuals and clarify the relation of maternal serum CA125 level and pregnancy. Among 552 healthy individuals, the distribution of serum CA125 values was demonstrated to resemble logarithmic normal distribution. Analysis of variance about age and sex revealed apparent elevation of values for women under 49 years of age in comparison with those for women over 50 years of age and men. Values for these two groups were 143 units/ml for the former and 32 units/ml for the latter, with a 99.7% confidence limit. These values exclude 99.3% of the former and 99.7% of the latter. Serum CA125 values were not affected by smoking. The measurement of serum CA125 levels in 71 pregnant women disclosed a significant elevation during the first trimester in comparison with nonpregnant women under 49 years of age. These results indicate that CA125 values must be deliberatively evaluated in young women, especially during first trimester of pregnancy.

Adult↗

PubMed2003

A test of linkage for complex discrete and continuous traits in nuclear families.

This article presents a score test for genetic linkage in nuclear families which applies to any trait having a distribution belonging to the exponential family, which includes binary and normal distributions, and distributions which are skewed or have nonnormal kurtosis. The specific distribution need not be specified and the method applies to sibships of arbitrary size. Tests of complex genetic effects are given, including unspecified mode of inheritance or additive, dominant, overdominant, and recessive modes of inheritance, covariates, multiple-locus models, including gene-gene interactions, and gene-environment interactions. The relation of our method to the Haseman-Elston methods is studied theoretically and by simulation.

Computer Simulation↗

PubMed2006

Physiological range of peak cardiac power output in healthy adults.

AIMS: The purpose of this study was to indicate the normal range for peak cardiac power output (CPO(peak)) in healthy adults and to explore age- and sex-related variations of this parameter. METHODS AND RESULTS: Using the non-invasive exponential CO(2) rebreathing technique [J.G. Defares, J Appl Physiol13 (1958) 159], cardiac output was measured at an exercise intensity determined to coincide with > or =95% of peak oxygen consumption in 102 healthy adults (mean +/- SD, age 43 +/- 13 years, body mass 74 +/- 13 kg). Peak cardiac power was then computed from measurements of peak cardiac output (Q(Tpeak)) and peak mean arterial pressure (MAP(peak)) using the equation described by Cooke et al. [Heart79 (1998) 289]. Peak oxygen consumption in the study population was 2.42 (+/-0.74) l min(-1) and subjects achieved 101 +/- 7% of this value during the measurement of . was 17.3 (+/-4) l min(-1), and CPO(peak) was computed as 4.5 (+/-1.2) W. CPO(peak) ranged from 3.11 to 7.94 W in men and 2.53 to 5.57 W in women. Additionally, ageing appears to be associated with a significant loss of peak cardiac power in men that is not apparent in women. CONCLUSION: Although the sample size remains moderate, the CPO values attained were normally distributed and these values provide a useful indication of the normal range for CPO(peak) in healthy adults.

Adult↗

PubMed2001

Biological sensor for sucrose availability: relative sensitivities of various reporter genes.

A set of three sucrose-regulated transcriptional fusions was constructed. Fusions p61RYTIR, p61RYlac, and p61RYice contain the scrR sucrose repressor gene and the promoterless gfp, lacZ, and inaZ reporter genes, respectively, fused to the scrY promoter from Salmonella enterica serovar Typhimurium. Cells of Erwinia herbicola containing these fusions are induced only in media amended with sucrose, fructose, or sorbose. While a large variation in sucrose-dependent reporter gene activity was observed in cells harboring all gene fusions, fusions to the inaZ reporter gene yielded a much wider range of activity and were responsive to lower levels of sucrose than either lacZ or gfp. The lacZ reporter gene was found to be more efficient than gfp, requiring approximately 300-fold fewer cells for a detectable response over all concentrations of sucrose. Similarly, inaZ was found to be more efficient than lacZ, requiring 30-fold fewer cells at 1.45 microM sucrose and 6,100-fold fewer cells at 29 mM sucrose for a quantifiable response. The fluorescence of individual cells containing p61RYTIR was quantified following epifluorescence microscopy in order to relate the fluorescence exhibited by populations of cells in batch cultures with that of individual cells in such cultures. While the mean fluorescence intensity of a population of individual cells increased with increasing concentrations of sucrose, a wide range of fluorescence intensity was seen among individual cells. For most cultures the distribution of fluorescence intensity among individual cells was log-normally distributed, but cells grown in intermediate concentrations of sucrose exhibited two distinct populations of cells, one having relatively low fluorescence and another with much higher fluorescence. When cells were inoculated onto bean leaves, whole-cell ice nucleation and gfp-based biological sensors for sucrose each indicated that the average concentration of sucrose on moist leaf surfaces was about 20 microM. Importantly, the variation in green fluorescent protein fluorescence of biosensor cells on leaves suggested that large spatial variations in sugar availability occur on leaves.

Bacterial Proteins↗