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An empirical method for converting nucleolar counts to neuronal numbers.

Our goal is to estimate neuronal numbers from counts of nucleolar profiles. The primary difficulty is that a simple way to convert the counts to the numbers, especially when multiple nucleoli are present in a nucleus, is not available. In this paper, we propose a solution. The formula is N = n X [N(c.f.)/n(c.f.)] where N = the true number of neurons, n = the number of nucleolar profiles in these neurons, N(c.f.) = the number of neurons used to estimate the correction factor and n(c.f.) = the number of nucleolar profiles found in the neurons that make up N(c.f.). The constraints are that the neurons identified for N(c.f.) be representative of the entire population, N; that the nucleolar profiles be counted by the same criteria when n(c.f.) is determined as when n is determined, and that when the correction factor [N(c.f.)/n(c.f.)] is calculated, the nucleolar profiles in each neuron be counted only once. The advantages are simplicity, and generality; the latter resulting from the empirical nature of the correction factor which can calibrate for multiple nucleoli, split nucleoli, invisible fragments, nucleolar size changes, section thickness differences and any other factors that cause n to deviate from N.

Animals↗

Functional and phenotypic characterization of human B lymphocyte subsets isolated by unit gravity sedimentation.

We report the use of unit gravity sedimentation with a CelSep apparatus to generate two volumetrically similar but functionally and phenotypically distinct subsets of human peripheral blood B cells. One subset, comprised of small B lymphocytes, underwent a significant size change in response to anti-mu, proliferated synergistically to low concentrations of anti-mu plus B cell growth factor (BCGF) or phorbol myristate acetate plus BCGF, and could be induced to produce immunoglobulin in response to pokeweed-mitogen-derived T-lymphocyte-replacing factors. These cells were primarily sIg+, B1+, B2+, and were virtually free of monocytes (less than 0.01%). Unlike these resting B lymphocytes, the large cells proliferated directly to BCGF, without displaying synergy with anti-mu. These cells displayed very little B2 (less than 7%), did not increase in volume in the presence of anti-mu, and made more immunoglobulin in response to TRF than the small resting B lymphocytes. However, neither population synthesized immunoglobulin spontaneously. This technique, which is highly reproducible, not equipment intensive, and produces high cell recovery (greater than 90%), allows for a precise analysis of the steps involved in the maturation of a resting B lymphocyte to an immunoglobulin-secreting cell.

B-Lymphocytes↗

Cholinergic amacrine cells in the rat retina.

Staining of rat retinal wholemounts with a monoclonal antibody against choline-acetyl-transferase (ChAT) reveals two matching populations of amacrine cells in pigmented and albino rat retinae. One population has cell bodies in the inner nuclear layer (INL). Their dendrites are confined to a narrow stratum in the outer half of the inner plexiform layer (IPL). The other, displaced, population has cell bodies in the ganglion cell layer (GCL) with dendrites stratifying in the middle of the IPL. The density changes with eccentricity, ranging from 1,700 cells/mm2 centrally to 600 cells/mm2 in the periphery. Presumptive cholinergic cells were filled with the fluorescent dye Lucifer yellow. Both subpopulations have the same "starburstlike" morphology as described for rabbit cholinergic amacrine cells (Famiglietti, '83; Tauchi and Masland, '84; Masland et al., '84b). Their dendritic tree sizes change with eccentricity and range from 160 to 300 microns in diameter. Counterstaining of Lucifer yellow-filled cells by ChAT immunohistochemistry did not yield an unequivocal double staining. Nevertheless, indirect evidence of same soma size, same number and form of primary dendrites, same level of stratification, and the good fit into the cholinergic mosaic makes it very likely that the "starburstlike" amacrine cells in the rat use acetylcholine as their transmitter. A comparison with the rabbit cholinergic system strengthens this assumption and reveals a striking similarity between both species.

Animals↗

Spontaneous length variation in microsatellite DNA from human T-cell clones.

Recently, much interest has been focused on instability of microsatellite DNA sequences such as di- and tri-nucleotide repeats in human cancers. Certain tumors show an increased frequency of mutation leading to repeat length variation at microsatellite loci, and it is thought that such instability may be a marker for the transformed phenotype. However, the spontaneous frequency by which repetitive DNA such as CA-repeats undergoes size changes in normal human somatic cells is not known. Therefore, it is not possible to decide if there is an increase in the frequency of microsatellite mutation in specific tumors or if the change observed simply reflects the frequency of microsatellite mutation in the cell population from which the tumor originates. To investigate this we have established panels of T-lymphocyte clones from 28 healthy males and determined the spontaneous length variations at three CA-repeat markers that are often used to investigate satellite instability: D2S123, D9S180, and D10S197. We found 3 T-cell clones with altered microsatellite size in a total of 178. This corresponds to a background frequency of 3 somatic microsatellite mutations in 1,028 alleles studied, i.e., 2.9 x 10(-3). This frequency is comparable to that found in many tumors of the breast, brain, ovary, and skin but is considerably lower than the frequency of microsatellite mutation in tumors related to hereditary non-polyposis colorectal cancer.

Cell Transformation, Neoplastic↗

Endotoxin-induced size change in bone marrow progenitors of granulocytes and macrophages.

Injection of 5 mug endotoxin to adult C57BL mice caused a marked increase in the sedimentation velocity of granulocytic and macrophage progenitor (colony-forming) cells in the bone marrow. This change was maximal two days after injection and was not accompanied by corresponding changes in total marrow nucleated cell populations. The endotoxin-induced shift was not dependent on the presence of the thymus but did not occur in mice challenged after preinjection with endotoxin. No changes in buoyant density, cell cycle status, pattern of differentiation and responsiveness of granulocytic and macrophage progenitor cells were observed after the injection of endotoxin. The increased sedimentation velocity of progenitor cells appears to indicate an increase in cell volume but the mechanisms involved have not been identified.

Animals↗

Separation and size distribution of red blood cells of diverse size, shape, and origin by flow/hyperlayer field-flow fractionation.

Red blood cells (RBCs) from human, equine, canine, feline, and bovine whole blood samples have been separated and characterized by high-speed flow/hyperlayer field-flow fractionation (Fl/HyFFF). The elution-based separation of RBCs by this method is based mainly on the size and shape of the cell particles. The typical separation time for RBCs is less than 3 min. Size distributions can be derived from the fractograms of cell samples using a calibration plot based on retention data for uniform polystyrene beads. The method is shown to be effective both to separate and to characterize cell populations, including those with cells of abnormal shape and size. In order to investigate differences in cell morphology, shape and size changes induced by 500,000 Da Dextran were monitored. The changes in the Fl/HyFFF elution profiles indicate that the RBCs decrease in size but become partially aggregated in the presence of the dextran. These changes were found to depend on polymer concentration and specific blood samples. Some of the results from Fl/HyFFF were compared with those from the Coulter counter and from microscopy.

Animals↗

A Sau3A polymorphism in the 5' end of the IT15 gene that nonrandomly segregates with the Huntington disease trinucleotide expansion.

Genomic clones encompassing the Huntington disease (HD) mutation were used to isolate a probe that detects size changes in the restriction fragments that contain the HD trinucleotide repeat (TNR). This probe also detects a frequent Sau3A polymorphism (allele sizes 1.8-kb and 2.7kb), which maps approximately 950bp from the TNR. Examination of a number of HD families established that the frequency of the Sau3A alleles did not differ significantly between control and HD populations; however, the HD expansion was always present on a chromosome that contained the 1.8-kb Sau3A allele. This association between a specific allele and the HD TNR expansion was significant and could provide a clue to the chromosomal elements that produce the trinucleotide expansion on the Huntington disease chromosome.

Alleles↗

Size and contrast have only small effects on the responses to faces of neurons in the cortex of the superior temporal sulcus of the monkey.

There is a population of neurons in the cortex in the middle and anterior part of the superior temporal sulcus (STS) of the monkey with responses which are selective for faces. To investigate whether the responses of these neurons show some of the perceptual properties of face recognition such as tolerance to changes in the size and contrast of the face, the effects of alteration of the size and contrast of an effective face stimulus on the responses of these neurons were analysed quantitatively in macaque monkeys. First, it was shown that the majority of these neurons had responses which were relatively invariant with respect to the size of the stimulus. The median size change tolerated with a response of greater than half the maximal response was 12 times. Second, it was found that for a few of these neurons, the size of the face did affect the neuronal response. For most of these neurons, it was found that when the size of the image and its distance were altered, the neuronal response was related to the retinal angle subtended by the image. But for four neurons the absolute size of the image determined the magnitude of the neuronal response, independently of the distance of the image. Thus these four neurons showed size constancy. It is suggested that these neurons would be useful as part of a face recognition system, because only objects in a certain absolute size range should normally be classified as faces. Third, the responses of the neurons were relatively invariant with respect to the contrast of the face. The mean contrast at which the neurons still responded with more than half the maximal response was 0.26. Fourth, the responses of the neurons were relatively invariant with respect to the sign of the contrast of the face, that is the neurons responded to negative as well as to positive images of faces. Fifth, the neurons typically responded to a face when the information in it had been reduced from 3D to a 2D representation in gray on a monitor, with a response which was on average 0.5 that to a real face. These results show that the responses of these neurons have some of the invariant properties with respect to size and contrast alteration shown by face perception, and show that their processing is at a level which would be useful in face recognition.(ABSTRACT TRUNCATED AT 400 WORDS)

Action Potentials↗

Spontaneous closure of atrial septal defects.

Atrial septal defects (ASDs) are found more frequently in the pediatric population than in adults, and improved diagnostic techniques with echocardiography (2DE) and Doppler facilitate diagnosis so that repair is possible at an optimal time. The purpose of our investigation was to study the size of ASDs at diagnosis, how size changes during follow-up, and to explore the relationship between size at diagnosis and need for surgery. We reviewed the medical records of all patients in Iceland with the diagnosis of ASD born between 1984 and 1993. ASD was confirmed by 2DE in all patients and defects smaller than 4 mm were excluded. ASD size was measured by 2DE from subxyphoid long and short axis views. There were 91 patients-29 males and 62 females. Four patients died from causes other than the heart defect and had not been operated upon. Seven patients with ASD primum and sinus venosus defects were excluded from analysis. There were 29 patients with a 4 mm defect, 17 patients with 5 or 6 mm defects, 8 patients with 7 or 8 mm defects, and 26 patients had defects >8 mm. In the 4 mm group, in 26 patients (89%) the ASD closed spontaneously or decreased in size, and 1 patient had been operated upon. In the 5 or 6 mm group, 15 of 19 ASDs (79%) had closed spontaneously, and 2 patients (9.5%) had been operated upon. In the 7 or 8 mm group, 1 of 6 ASDs (16.6%) had closed spontaneously and 3 had been closed surgically. In the >8 mm group, 1 of 24 ASDs had closed spontaneously and 20 (91%) had been closed surgically. We conclude that defects smaller than 6 mm in diameter are very likely to close spontaneously although follow-up is necessary. Defects larger than 8 mm have a high probability requiring operative closure.

Blood Flow Velocity↗

Cochlear nucleus cell size changes in the dalmatian: model of congenital deafness.

We assessed cellular changes in one population of neurons of the cochlear nucleus associated with a form of genetic deafness in deaf dalmatians. Spheric cells from deaf dalmatians and age-matched control (hearing) dogs were analyzed morphometrically. The somatic silhouette of these cells was reduced by 22.1% to 38.1%. The effect on cell size was greater with increased duration of deafness. Because the deaf dalmatian exhibits progressive degeneration of the auditory periphery, shrinkage of spheric cells may reflect the initial influence of attenuated activity of auditory nerve fibers, and sensorineural degeneration with longer periods of deafness.

Age Factors↗

Longitudinal radiographic assessment of dense bone islands of the jaws.

OBJECTIVES: This study assessed dense bone island (DBI) inception and growth in jaws. STUDY DESIGN: A population of 2991 patients (age range 5 to 35 years) was studied with at least two panoramic radiographs taken 1 to 10 years apart for each patient. Lesions were digitized and measured with computer measuring software. The size of the earliest DBI was compared with subsequent DBI measurements after the latter measurement was normalized with reference to the size of the nearest tooth. RESULTS: Sixty-nine patients with DBIs were identified. A total of 3.1% of the patients with DBI were found in the 5- to 10-year age range, with the first patient in the DBI group found at 9.4 years. A significantly higher proportion (p = 0.002) of the unaffected patients (26%) was found in the same 5-to 10-year age range. The proportions of patients in the DBI and unaffected groups in older age ranges were similar. Assessment of DBI size changes showed that 43% of the 53 DBIs detected between 9.4 and 19 years enlarged (mean change, 212%), and 17% diminished (mean change, 49%). In comparison, 29.4% of 17 DBIs detected between 20 and 35 years enlarged (mean change, 153%), and 5.9% diminished (mean change, 60%). CONCLUSION: Our results indicate that DBIs are labile lesions, develop during early adolescence, and retain a potential for enlargement, or to a lesser extent shrinkage, into adulthood.

Adolescent↗

Temperature-induced fusion of small unilamellar vesicles formed from saturated long-chain lecithins and diheptanoylphosphatidylcholine.

Small unilamellar vesicles which form when gel-state long-chain phosphatidylcholines are mixed with micellar short-chain lecithins undergo an increase in size as the long-chain species melts to its liquid-crystalline form. Analysis of the vesicle population with quasi-elastic light scattering shows that the particle size increases from 90-A radius to greater than 5000-A radius. Resonance energy transfer experiments show total mixing of lipid probes with unlabeled vesicles only when the Tm of the long-chain phosphatidylcholine is exceeded. This implies that the large size change represents a fusion process. Aqueous compartments are also mixed during this transition. 31P NMR analysis of the vesicle mixtures above the phase transition shows a great degree of heterogeneity with large unilamellar particles coexisting with oligo- and multilamellar structures. Upon cooling the vesicles below the Tm, the original size distribution (e.g., small unilamellar vesicles) is obtained, as monitored by both quasi-elastic light scattering and 31P NMR spectroscopy. This temperature-induced fusion of unilamellar vesicles is concentration dependent and can be abolished at lower total phospholipid concentrations. It occurs over a wide range of long-chain to short-chain ratios and occurs with 1-palmitoyl-2-stearoylphosphatidylcholine and dimyristoylphosphatidylcholine as well. Characterization of this fusion event is used to understand the anomalous kinetics of water-soluble phospholipases toward these unusual vesicles.

Kinetics↗

Character displacement in some Cnemidophorus lizards revisited: a phylogenetic analysis.

Ecological studies have demonstrated the role of competition in structuring communities; however, the importance of competition as a vehicle for evolution by natural selection and speciation remains unresolved. Study systems of insular faunas have provided several well known cases where ecological character displacement, coevolution of competitors leading to increased morphological separation, is thought to have occurred (e.g., anoline lizards and geospizine finches). Whiptail lizards (genus Cnemidophorus) from the islands of the Sea of Cortez and the surrounding mainland demonstrate a biogeographic pattern of morphological variation suggestive of character displacement. Two species of Cnemidophorus occur on the Baja peninsula, one relatively large (Cnemidophorus tigris) and one smaller (Cnemidophorus hyperythrus). Oceanic islands in the Sea of Cortez contain only single species, five of six having sizes intermediate to both species found on the Baja peninsula. On mainland Mexico C. hyperythrus is absent, whereas C. tigris is the smaller species in whiptail guilds. Here we construct a phylogeny using nucleotide sequences of the cytochrome b gene to infer the evolutionary history of body size change and historical patterns of colonization in the Cnemidophorus system. The phylogenetic analysis indicates that (i) oceanic islands have been founded at least five times from mainland sources by relatives of either C. tigris or C. hyperythrus, (ii) there have been two separate instances of character relaxation on oceanic islands for C. tigris, and (iii) there has been colonization of the oceanic island Cerralvo with retention of ancestral size for Cnemidophorus ceralbensis, a relative of C. hyperythrus. Finally, the phylogenetic analysis reveals potential cryptic species within mainland populations of C. tigris.

Animals↗

Patterns of instability of expanded CAG repeats at the ERDA1 locus in general populations.

A highly polymorphic CAG repeat locus, ERDA1, was recently described on human chromosome 17q21.3, with alleles as large as 50-90 repeats and without any disease association in the general population. We have studied allelic distribution at this locus in five human populations and have characterized the mutational patterns by direct observation of 731 meioses. The data show that large alleles (>/=40 CAG repeats) are generally most common in Asian populations, less common in populations of European ancestry, and least common among Africans. We have observed a high intergenerational instability (46. 3%+/-5.1%) of the large alleles. Although the mutation rate is not dependent on parental sex, paternal transmissions have predominantly resulted in contractions, whereas maternal transmissions have yielded expansions. Within this class of large alleles, the mutation rate increases concomitantly with increasing allele size, but the magnitude of repeat size change does not depend on the size of the progenitor allele. Sequencing of specific alleles reveals that the intermediate-sized alleles (30-40 repeats) have CAT/CAC interruptions within the CAG-repeat array. These results indicate that expansion and instability of trinucleotide repeats are not exclusively disease-associated phenomena. The implications of the existence of massively expanded alleles in the general populations are not yet understood.

Alleles↗

Size changes of protamine 1 mRNA provide a molecular marker to monitor spermatogenesis in wild-type and mutant mice.

We utilized a cDNA encoding the cysteine-rich, tyrosine-containing mouse protamine, mouse protamine 1 (MP1), to detect the presence of several classes of differentiating germ cells in testicular extracts from wild-type and male sterile mutant mice. This assay is based on the changes in the poly (A) length of MP1-mRNA during spermatogenesis. Testicular extracts of sexually mature CD-1 mice contain a heterogeneous population of protamine-1 mRNA ranging in length from 450 to 580 nucleotides. When the protamine-1 probe was hybridized to testicular RNA preparations from 16- to 20-day-old animals, no MP1-mRNA was detected. Twenty-four-day-old mice contain only the 580-nucleotide form of MP1-mRNA. This size class of protamine mRNA is also present in purified populations of round spermatids, whereas elongating spermatids and residual bodies contain mRNAs ranging from 450 to 580 nucleotides in length, which are identical in size to those present in the testes of sexually mature animals. When the protamine cDNA probe was used to examine the progression of spermiogenesis in three male sterile mouse mutants, blind sterile (bs), quaking (qk) and testicular feminization (Tfm), the results demonstrated that each mutant is pathologically distinct. Analysis of the bs mutant revealed a diminution in the amount of both size classes of MP1-mRNA, in agreement with the cytological reports of reduced numbers of haploid spermatogenic cells in these animals. The presence of both size classes of protamine mRNA in the qk mutant indicates that germ-cell differentiation has proceeded at least to the step-12 spermatid in these animals.(ABSTRACT TRUNCATED AT 250 WORDS)

Androgen-Insensitivity Syndrome↗

Genome size variation in North American minnows (Cyprinidae). II. Variation among 20 species.

Genome sizes (nuclear DNA contents) from 200 individuals representing 20 species of North American cyprinid fishes (minnows) were examined spectrophotometrically. The distributions of DNA values of individuals within populations of the 20 species were essentially continuous and normal; the distribution of DNA values among species was continuous and overlapping. These observations suggest that changes in DNA quantity in cyprinids are small in amount, involve both gains and losses of DNA, and are cumulative and independent in effect. Significant heterogeneity in mean genome size occurs both between individuals within populations of species and among species. The former averages maximally around 6% of the cyprinid genome and is nearly the same as the amount of DNA theoretically needed for the entire cyprinid structural gene component. The majority of the DNA content variation among the 20 species is distributed above the level of individuals within populations. Comparisons of average genome size difference or distance between individuals drawn from different levels of taxonomic organization indicate that considerably greater divergence in genome size has occurred in the extremely speciose cyprinid genus Notropis as compared with other North American cyprinid genera. This may suggest that genome size change is concentrated in speciation episodes. Finally, no associations were found between interspecific variation in genome size and five life-history characters. This suggests that much of the variation in genome size within and among the 20 species may be phenotypically inconsequential.

Animals↗

Age-related changes in glomerular volume and hydroxyproline content in rat and human.

Total 4-hydroxyproline content and volume were measured in the same sample of microdissected glomeruli obtained fro rat and human outer or inner cortex. Glomerular volume was determined by computer-assisted image analysis, and 4-hydroxyproline was measured by a highly sensitive gas-liquid chromatographic method. Results were expressed as weight of basement membrane material by comparison with the amount of 4-hydroxyproline in purified basement membrane/mesangial matrix preparations. Microanalyses were possible in samples containing as few as eight human glomeruli. Rat glomerular size increased sevenfold between 5 wk and 2 yr of age, with volume being consistently 36 to 45% greater in inner than in outer cortex glomeruli. Basement membrane material content per glomerulus markedly increased with age (12-fold); however, when expressed per unit volume, this change was greatly reduced (2-fold). Expressed per volume, inner and outer cortex glomerular content of basement membrane material was always similar, regardless of age. Therefore, a greater glomerular size, in itself, does not accelerate the rate of basement membrane material deposition. Glomerular size distributions (measured by skewness and kurtosis) did not change, indicating that, although glomerular volume increases with age, aging does not appear to cause the emergence of distinct glomerular populations within an age group. Basement membrane material accumulation is probably a generalized change. Human glomeruli increased sevenfold in size from infancy to adulthood and then declined during senescence. Contrary to that in the rat, glomerular basement membrane material content appeared to closely follow size changes, thus, varying little from infancy to senescence if expressed per unit of glomerular volume.

Adolescent↗