Biomedical subjects

J R Nyengaard

Publications and source records attributed to J R Nyengaard.

At least 19 recordsLinked to original sources

PubMed2006

Cartilage thickness in the hip joint measured by MRI and stereology--a methodological study.

OBJECTIVE: The purpose of this study was to develop a precise and efficient method for estimating the thickness of the articular cartilage in the hip joint and hence three different stereologic methods were tested based on Magnetic Resonance Imaging. DESIGN: Twenty two females and four males with hip dysplasia underwent MRI. The thickness of the femoral and acetabular cartilage was estimated. RESULTS: The results for all three methods showed that the observed total variance on cartilage thickness is small. The mean thickness of the acetabular cartilage measured by the three different methods ranged between 1.15 mm and 1.46 mm. The mean thickness for the femoral cartilage measured by the three different methods ranged between 1.18 mm and 1.78 mm. The measurements took 15-20 min per hip to carry out. CONCLUSION: Methods 1 and 3 are as precise but we favour method 3 because the measurements are done on images obtained through the center of the femoral head which means that the cartilage surface is intersected perpendicular and partial volume effect avoided. We suggest that this method can be advantageous for assessing the progression of osteoarthritis in dysplastic hips after periacetabular osteotomy.

Adult↗

PubMed2006

Using biased image analysis for improving unbiased stereological number estimation - a pilot simulation study of the smooth fractionator.

The smooth fractionator was introduced in 2002. The combination of a smoothing protocol with a computer-aided stereology tool provides better precision and a lighter workload. This study uses simulation to compare fractionator sampling based on the smooth design, the commonly used systematic uniformly random sampling design and the ordinary simple random sampling design. The smooth protocol is performed using biased information from crude (but fully automatic) image analysis of the fields of view. The different design paradigms are compared using simulation in three different cell distributions with reference to sample size, noise and counting frame position. Regardless of clustering, sample size or noise, the fractionator based on a smooth design is more efficient than the fractionator based on a systematic uniform random design, which is more efficient than a fractionator based on simple random design. The fractionator based on a smooth design is up to four times more efficient than a simple random design.

Journal Article↗

PubMed2006

A new fractionator principle with varying sampling fractions: exemplified by estimation of synapse number using electron microscopy.

The quantification of ultrastructure has been permanently improved by the application of new stereological principles. Both precision and efficiency have been enhanced. Here we report for the first time a fractionator method that can be applied at the electron microscopy level. This new design incorporates a varying sampling fraction paradigm. The method allows for systematic random sampling from blocks of variable slab thickness, thereby eliminating the need for exhaustive serial sectioning through an entire containing space. This novel approach acknowledges the inaccuracy inherent in estimating the total object number using section sampling fractions based on the average thickness of sections of variable thicknesses. As an alternative, this approach estimates the correct particle section sampling probability based on an estimator of the Horvitz-Thompson type, resulting in a theoretically more satisfying and accurate estimate of the expected number of particles for the defined containing space.

Animals↗

PubMed2006

Direct and efficient stereological estimation of total cell quantities using electron microscopy.

Stereological estimation of total subcellular quantities in bioscience is presented in this report. Special emphasis is placed on the use of electron microscopy, which under certain circumstances may be combined with light microscopy. Three strategies based on the Cavalieri principle, the disector and local stereological probes through arbitrarily fixed points for estimation of total quantities inside cells are presented. The quantities comprise (total) number, length, surface area, volume or 3D spatial distribution for organelles as well as total amount of gold particles, various compounds or certain cytochemical markers.

Animals↗

PubMed2005

Prospective bone density changes after periacetabular osteotomy: a methodological study.

We used computed tomography (CT) and 3D design-based sampling principles (stereology) to estimate changes in acetabular bone density after periacetabular osteotomy. We included six consecutive patients with hip dysplasia in the study. Baseline density was measured within the first 7 days following periacetabular osteotomy and compared with bone density 2 years later. Double measurements were performed on three patients, and the coefficient of error of the mean was estimated to 0.05. Bone density in zone 1 increased significantly in the anteromedial quadrant as well as in the posteromedial quadrant of the acetabulum. In the anterolateral and the posterolateral quadrant, bone density was unchanged. In zone 2 and 3, bone density was unchanged 2 years postoperatively. We suggest that the observed increase in bone density medially represents a remodelling response to an altered load distribution after osteotomy. The method used is a precise tool to estimate changes in acetabular bone density.

Acetabulum↗

PubMed2005

Design-based stereological estimation of the total number of cardiac myocytes in histological sections.

BACKGROUND: Counting the total number of cardiac myocytes has not previously been possible in ordinary histological sections using light microscopy (LM) due to difficulties in defining the myocyte borders properly. AIM: To describe a method by which the total number of cardiac myocytes is estimated in LM sections using design-based stereology. MATERIALS AND METHODS: From formalin-fixed left rat ventricles (LV) isotropic uniformly random sections were cut. The total number of myocyte nuclei per LV was estimated using the optical disector. Two-microm-thick serial paraffin sections were stained with antibodies against cadherin and type IV collagen to visualise the intercalated discs and the myocyte membranes, respectively. Using the physical disector in "local vertical windows" of the serial sections, the average number of nuclei per myocyte was estimated. RESULTS: The total number of myocyte nuclei in LV was 34.1 x 10(6) (0.08) (mean (coefficient of variation)), and the mean number of nuclei per myocyte 1.85 (0.03). Combining these estimates the total number of myocytes in LV was calculated to be 18.5 x 10(6) (0.09). CONCLUSIONS: This new method is applicable to a range of experiments focusing on myocyte proliferation and death.

Animals↗

PubMed2004

Postnatal growth of cardiomyocytes in the left ventricle of the rat.

We studied the development of myocytes and interstitium using perfusion-fixed left ventricles obtained from normal female Wistar rats at 5 days (n = 5), 25 days (n = 5), and 125 days (n = 5) of age. Using design-based stereological methods and light microscopy, we estimated the following parameters: volume of left ventricle made up by myocytes, myocyte nuclei, and interstitium; total numbers of myocyte and non-myocyte nuclei; mean volumes of myocyte nuclei; the total volume, surface area, and length of fibers; and the mean star volumes of fibers. Some derived parameters were also calculated, namely, the mean myocardium volume per nucleus and the mean fiber cross-sectional area. We found that postnatal myocyte growth after day 5 in the young rat is largely hypertrophic, while interstitial growth is hyperplastic. The increase in left ventricular mass was 10-fold over the ages studied, whereas total length, surface area, and volume of fibers increased approximately 3-, 8-, and 11-fold over the period. Relative rates of growth implied that fiber growth was dominated by an increase in length compared to other dimensions. The total number of myocyte nuclei ( approximately 30 x 10(6)) did not change between 5 and 25 days of age, but then almost doubled in 125-day-old rats. The number of non-myocyte nuclei increased 9-fold over the period studied in an exponential manner. The mean myocyte nucleus volume tripled between the ages of 5 and 25 days and then remained the same. The volume-weighted mean nucleus volume was highly variable and showed no significant trend with age. Our results provide support for the claim made by some researchers that myocyte proliferation had ceased by day 5 after birth, but do not provide evidence for binucleation of myocytes between 5 and 25 days after birth. Reported numbers of myocyte nuclei express a net growth and do not rule out both myocyte death and creation throughout the early postnatal period. We clearly detect an increase in the number of myocyte nuclei from 25 to 125 days but are not able to state whether this increase reflects myocyte proliferation or myocyte binucleation. To determine this would require special double-staining techniques to delineate intercalated discs and myocyte membranes. Our results agree very well with the limited data available from other studies that have used unbiased stereology. It is important to use unbiased designs that consider anisotropy as well as heterogeneity of the heart in sampling schemes, and to determine absolute quantities to avoid problems associated with interpreting changes in density (the "reference trap").

Age Factors↗

PubMed2004

Volume estimation from projections.

We describe a new estimator of the volume of axially convex objects from total vertical projections with known position of the vertical axis. The estimator combines the Cavalieri method with the known formula for area in terms of the support function of a convex body. We examine the accuracy of the proposed estimator for ellipsoidal objects having exactly known support function and volume. In addition, we illustrate practical problems of accuracy by implementing the method for some biological products.

Biology↗

PubMed2004

Shape modelling of spatial particles from planar central sections -- a case study.

In this paper we develop statistical tools for shape modelling of spatial particles from central sections through the particles. The particles are assumed to be star-shaped with respect to a reference point inside the particles and are modelled as stochastic deformations of spheres centred at the reference points. The resulting particles are rotation invariant with respect to the reference point. As an illustration, the model is applied to study shape differences between neurons in the Granular and CA1 layer in the human hippocampus.

Animals↗

PubMed2003

Gentamicin used as an adjunct to GTR.

OBJECTIVES: To evaluate in a discriminating "capsule" model whether local application of gentamicin may have an added effect on bone formation produced by Bio-Oss and guide tissue regeneration (GTR). MATERIAL AND METHODS: Thirty male 3-month-old Wistar rats were used. After elevation of muscle-periosteal flaps, a rigid hemispherical Teflon capsule, loosely packed with 0.025 g of Bio-Oss impregnated with 2 mg/ml gentamicin sulfate (Garamycin), was placed with its open part facing the lateral bone surface of the mandibular ramus (test) in one side of the jaw. A capsule filled only with Bio-Oss (control) was placed on the contralateral side of the jaw. After healing periods of 1, 2 and 4 months, groups of 10 animals were sacrificed and the specimens were processed for histological examination. The volumes of (1) the space created by the capsule, (2) newly formed bone, (3) Bio-Oss particles, (4) loose connective tissue, and (5) acellular space in the capsule were estimated by a point-counting technique in three to four histological sections of each specimen, taken by uniformly random sampling. RESULTS: The histological evaluation showed limited but increasing bone fill in the capsules from 1 to 4 months in both the test and control sides. After 4 months, the newly formed bone occupied 11.9% (CV: 0.39) of the space created by the capsules at the test sides versus 13.2% (CV: 0.41) at the control sides. There was no statistical significant difference between test and control specimens at any observation time (p>0.05). CONCLUSION: It is concluded that local application of gentamicin has no added effect on bone formation when combined with Bio-Oss and GTR.

Animals↗

PubMed2002

Stereological estimation of the total number of ECL cells and related parameters using the smooth, vertical fractionator in the rat oxyntic mucosa.

During the last 10 years many attempts have been made to estimate the number of enterochromaffin-like (ECL) cells in various animal studies. This is the first presentation of an unbiased stereological estimator of the total number of histamine-positive ECL cells per rat and linked to estimators of related parameters: total volume of the oxyntic mucosa, total oxyntic mucosal surface area, total oxyntic serosal surface area, surface amplification factor, average thickness of the oxyntic mucosa, total and mean volume of the ECL cells, total number of oxyntic glands and pits, mean number of ECL cells per gland, and mean number of ECL cells and glands per oxyntic serosal surface area. This study is the first application of the smooth fractionator and includes a description of all sources of sampling variance in the smooth fractionator design with newly developed predictors.

Animals↗

PubMed2001

Total regional and global number of synapses in the human brain neocortex.

An estimator of the total number of synapses in neocortex of human autopsy brains based on unbiased stereological principles is described. Each randomly chosen cerebral hemisphere was stratified into the four major neocortical regions. Uniform sampling with a varying sampling fraction in each region of neocortex was performed. The total volume of each neocortical region was estimated using point counting according to Cavalieri's principle. The ethanolic phosphotungstic acid staining technique was modified for synapses in human autopsy brains. The numerical density of synapses in each neocortical region studied was estimated using the disector at the electron microscopical level. The total number of neocortical synapses in each region was estimated as the product of the total volume of neocortex and the numerical density of synapses. The influence of the postmortem fixation delay on the number of synapses was investigated in five large mammals (one dog, one cow, and three pigs), the brains of which were kept under conditions similar to those under which human corpses are normally kept. The apparent decrease of 3.9% in the numerical density of synapses in the large mammals following a 2-day fixation delay was not significant. The average total number of synapses in the neocortex of five young male brains was 164 x 10(12) (CV = 0.17). An analysis of the precision of the estimate of the total number of synapses in neocortex indicates that blocks represent both the major source of variation and the largest workload. Using eight blocks per brain the imprecision of the estimate is, however, only 66% of the total variance.

Adult↗

PubMed2001

Direct evidence of the importance of stromal urokinase plasminogen activator (uPA) in the growth of an experimental human breast cancer using a combined uPA gene-disrupted and immunodeficient xenograft model.

Several studies have indicated an interaction between tumor cells and infiltrating stromal cells regarding the urokinase plasminogen activation (uPA) system. By developing combined uPA gene-disrupted and immunodeficient mice, we have studied the role of stromal uPA for the growth of the MDA-MB-435 BAG human tumor xenograft. Subcutaneous tumor growth and lung metastasis were compared between wild-type immunodeficient mice and mice with the combined deficiencies. Tumor growth was evaluated by volume measurements and plasma beta-galactosidase activity and metastasis was evaluated by counting lung surface metastases. Although no differences appeared in primary tumor take between the two groups of mice, a significant difference was observed in primary tumor growth, with tumors in uPA-/- mice growing significantly more slowly. In addition, a nonsignificant trend toward fewer lung metastases in uPA-/- mice was observed. The present data points to a critical role of stromal-derived uPA in the primary tumor growth of MDA-MB-435 BAG xenografts, whereas only a trend toward fewer lung metastases in uPA gene-disrupted mice was found.

Animals↗

PubMed2001

Stereological estimates of number and length of capillaries in subdivisions of the human hippocampal region.

The hippocampal formation is a neuroanatomically well-defined region of the brain involved in memory processes. In view of the functional importance of the region and its involvement in a number of brain pathologies, including Alzheimer's disease and temporal lobe epilepsy, a quantitative description of its vascular supply represents an important first step in evaluating the involvement of vascular changes in these phenomena. Unbiased estimates of the length and connectivity of the vascular supply of brain regions have not been described previously. The total number, total length, and distribution of the diameters of capillaries were estimated in the five major subdivisions of the hippocampal formation (fascia dentata, hilus, CA3-2, CA1, and subiculum) in 5 normal males, 52-84 years of age. These estimates were used to derive several other structural parameters. Both the primary and the derived parameters were used to make inter- and intra-individual comparisons. For each of the five major subdivisions from each individual, the volume was estimated using the Cavalieri principle. The total capillary length was estimated on 3-microm-thick plastic isotropic uniform random sections. Using a topological definition of a capillary unit and the optical disector, total capillary number was estimated in 40-microm-thick plastic sections. Length-and number-weighted three-dimensional diameter distributions were obtained from the thin and thick plastic sections, respectively. In each subdivision the total length of capillaries was correlated with previously obtained data on the number of neurons in the same subdivisions of the same individuals. Intersubdivisional differences were observed, in that the hilus of the dentate gyrus had fewer capillaries per unit volume than the other four subdivisions. Interindividual comparisons indicate that the interindividual variances are of a magnitude suitable for sensitive group comparisons. The design-based stereological methods that were used in the analyses can provide a basis for a new unbiased approach to the estimation of vascular parameters in well-defined regions of the brain.

Aged↗

PubMed2001

Birth weight has no influence on glomerular number and volume.

It has been proposed that low birth weight (LBW) results in a reduction in glomerular number that may, in turn, predispose an individual to develop hypertension in adulthood. Glomerular number is reduced in animal models of intra-uterine malnutrition using a variety of techniques. However, the relevance of such extreme models to man is uncertain. The purpose of this study was to evaluate whether animals with naturally occurring LBW, which have not received any manipulation in utero, have a reduction in glomerular number, altered glomerular volume and abnormal urine albumin excretion. Litters from female rats delivering at term on the same day were weighed and sexed at birth. From each litter 2 males with the lowest birth weight (LBW n = 18) and 2 males with a birth weight closest to the litter mean [normal birth weight (NBW) n = 18] were selected and cross-fostered onto periparturient lactating dams. LBW rats weighted 6.7 +/- 0.6 g compared with 7.2 +/- 0.6 g for NBW rats (P = 0.03). After weaning all rats were weighed weekly and underwent metabolic studies at 4, 8, 12 and 16 weeks. Following perfusion fixation, glomerular number and mean glomerular volume were estimated using standard stereological techniques. There was no significant difference between LBW and NBW rats with respect to glomerular number (24,499 +/- 2,078 vs. 24,825 +/- 1,818), mean glomerular volume and urine albumin excretion, and no rats had a glomerular number outside the normal range. This study suggests that naturally occurring LBW has little influence on renal development, glomerular number and volume.

Animals↗

PubMed2001

Morphology of renal afferent arterioles and glomeruli, heart weight, and blood pressure in primates.

In a Caribbean outbred population of African green monkeys (Cercopithecus aethiops), 5 to 10% of feral adults have elevated blood pressure (BP). We have investigated whether the increased pressure is associated with abnormal renal afferent arteriole structure or glomerular number. In seven young adult (aged 7 to 13 years) male monkeys with consistently high BP (mean BP, 111 mm Hg; ketamine anesthesia) and seven controls (mean BP, 81 mm Hg), the morphology of the renal vasculature has been analyzed in three cortical zones. In each animal, the left kidney vasculature was fixed while relaxed and at known intravascular pressure, and afferent arteriolar diameter and media cross-sectional area were estimated. The right kidney was perfusion-fixed and prepared for unbiased stereologic estimation of glomerular number and size. No difference was found in afferent arteriole lumen diameter or media cross-sectional area, or in glomerular number or size, between the high BP group and controls. There was no difference in heart weight between the two groups, but there was a negative correlation between left ventricle heart weight and afferent arteriole diameter (controls: r = -0.81, P = .025; all animals: r = -0.70, P = .005, slope about 3.5% reduction in lumen diameter for 10% increase in heart weight). The results suggest that cardiac mass and renal afferent arteriole structure may be controlled by a common mechanism unrelated to BP measured in anesthesia. However, the lack of conscious measurements prevents conclusions as to whether this mechanism involves ambulatory BP.

Animals↗

PubMed2001

Tissue shrinkage and unbiased stereological estimation of particle number and size.

This paper is a review of the stereological problems related to the unbiased estimation of particle number and size when tissue deformation is present. The deformation may occur during the histological processing of the tissue. It is especially noted that the widely used optical disector may be biased by dimensional changes in the z-axis, i.e. the direction perpendicular to the section plane. This is often the case when frozen sections or vibratome sections are used for the stereological measurements. The present paper introduces new estimators to be used in optical fractionator and optical disector designs; the first is, as usual, the simplest and most robust. Finally, it is stated that when tissue deformation only occurs in the z-direction, unbiased estimation of particle size with several estimators is possible.

Brain↗

PubMed2001

Deproteinized bovine bone used as an adjunct to guided bone augmentation: an experimental study in the rat.

BACKGROUND: Promising results have been reported following treatment of periodontal and peri-implant bone defects with deproteinized bovine bone grafts, but their influence on bone formation has not been clarified. PURPOSE: The goal of this study was to examine whether implantation of deproteinized bovine bone (Bio-Oss, Geistlich AG, Wolhusen, Switzerland) influences bone formation when used as an adjunct to guided bone augmentation (GBA). MATERIALS AND METHODS: A rigid, hemispherical, Teflon capsule was loosely packed with a standardized quantity of Bio-Oss and placed with its open part facing the lateral surface of the mandibular ramus (test) in 30 rats. At the contralateral side of the jaw, an empty capsule was placed (control). Groups of 10 animals were sacrificed after 1, 2, and 4 months. The volumes of the space created by the capsule and of the (1) newly formed bone, (2) remaining Bio-Oss particles, (3) soft connective tissue, and (4) acellular space in the capsule were estimated by a point-counting technique in three or four histologic sections, taken by uniformly random sampling. RESULTS: Bone formation at 1 month was limited in both tests and controls. After 2 months, the mean volume of the newly formed bone occupied 9.0% of the space created by the capsule in the test specimens compared with 23.8% in the control specimens (p < .01). After 4 months, the respective figures were 11.6% (tests) versus 38.7% (controls) (p < .01). CONCLUSION: It can be concluded that Bio-Oss, used as an adjunct to GBA, interferes with bone formation.

Animals↗