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Methodological problems in placental morphometry: apologia for the use of stereology based on sound sampling practice.

Several problem areas in morphometry of human and animal placentae are reviewed. Attention is given to methods of tissue processing (handling, mode of fixation, embedding, shrinkage) and sampling (of organs, tissue blocks, sections, micrographs). Principal sources of bias and sampling variability are identified and the crucial importance of randomized sampling is emphasized. Methods for obtaining structural quantities from sections are compared. The case is made for estimating absolute values (volumes, surface areas, lengths, numbers, thicknesses) using stereological principles rather than relying on planar data (profile areas, perimeter lengths, numbers, apparent thicknesses). Absolute values may be obtained simply and efficiently without resort to expensive measuring devices. Finally, morphological descriptors suitable for correlating with functional data or for comparing normal and diseased organs are surveyed.

Delivery, Obstetric↗

PubMed1991

A useful programme in BASIC for axonal morphometry with introduction of new cytoskeletal parameters.

Interest in the structure of axons and quantification of their components has been growing over the last years. However, the existing literature contains few reports of available computer programmes to facilitate such studies. This paper presents a fully comprehensive BASIC programme for the morphometric analysis of electron micrographs of cross-sectional nerve fibres. From drawings of fibre and axonal contours and dots of the microtubules and neurofilaments, the programme calculates the following parameters: area, diameter and form factor of the fibres and axons, number and density of microtubules and neurofilaments, proportion between microtubules and neurofilaments (R-proportion), myelin thickness and the diameter of the axon relative to its sheath (g-ratio). The programme also introduces three new parameters to analyse the degree of uniformity of microtubule and neurofilament distribution: distances between microtubules and between neurofilaments, equilateral index and cytoskeletal intermingling index. The programme is written in Microsoft BASIC Interpreter for Apple Macintosh (Microsoft Corporation) but can be used on other computers. Although the programme has been tested on adult rat optic nerve fibres, it can be used for different projects concerning axonal morphometry.

Animals↗

PubMed1989

Intrinsic and extrinsic factors collaborate to generate driving forces for neural tube formation in the chick: a study using morphometry and computerized three-dimensional reconstruction.

The formation of the neural tube, the rudiment of the entire central nervous system, is one of the earliest morphogenetic movements. The origin of the driving forces for this process remains uncertain, but recent studies suggest the involvement of both intrinsic and extrinsic factors. In the present study, we have used morphometry, analysis of stereopair photographs of whole embryos, and computerized three-dimensional reconstruction to investigate the factors which constitute the bulk of the driving forces for neural tube formation in the developing midbrain of Hamburger and Hamilton stages 5-9 chick embryos. Results support the notion that neural tube formation is driven by a coordinated interplay of intrinsic and extrinsic forces. Initial bending of the neural plate along the midline of the embryo and uplifting of the neural folds is accomplished primarily through the combined action of intrinsic forces (resulting from apical constriction of neuroepithelial cells) and extrinsic forces (mostly a passive consequence of head-fold formation). However, once in the uplifted position, curling over of neural folds and closure of the neural tube is driven largely by apical constriction-mediated (intrinsic) forces that are generated by cells in the midlateral walls of the forming neural tube.

Animals↗

PubMed1995

DATAPLOT: a graphical display package for bacterial morphometry and fluorimetry data.

A microbiological image processing system has been developed, which is used to determine both morphometry and fluorimetry data on (mainly faecal) bacteria. A special program for display of these data has been developed for this system. Besides probability density estimates of any one or two parameters and scatter plots of any two or three parameters in the distribution, it can generate scatter plots using the actual bacterial shapes, rather than arbitrary symbols. A particularly powerful feature of the program is the possibility to overlay a contour plot of the probability density distribution of the bacteria in parameter space, with a scatter plot of the actual bacterial shapes involved. In this paper, the program's organization, options, and the algorithms used are discussed.

Algorithms↗

PubMed1988

Vertebral morphometry in diagnosis of spinal fractures.

Vertebral morphometry was performed on lateral thoracic and lumbar radiographs of 153 women with a preliminary diagnosis of spinal osteoporosis. Measurements included anterior and posterior vertebral height, width, area, wedge angle, percent reduction of anterior to posterior height (PRH) and percent difference in anterior height between adjoining vertebrae (PDAH). A vertebral fracture was identified if any of the measurements which control for interindividual variation in vertebral size (wedge angle, PRH, PDAH) were below the normal range. Among individuals with mild osteoporosis (0-2 fractures) PDAH identified 86% of the fractures and 95% of the individuals with fractures. Other measurements identified less than 71% of the fractures and less than 80% of the individuals with fractures. Although the results reflect a relative rather than a true sensitivity it appears that PDAH is the better diagnostic measurement for fractures in the earlier stages of spinal osteoporosis.

Aged↗

PubMed1984

Ready evaluation of lung alveolar toxic damage with histological sections morphometry.

In this experiment we aimed at developping a method of ready evaluation of lung alveolar toxic damage using morphometric measurements. Swiss-Webster mice were treated with different doses of diquat (i.p.) and the left lung was fixed at 0, 3, and 5 days after treatment. The technique of morphometry of histological sections embedded in glycol methacrylate (GMA) was used to quantify possible modifications of alveolar cells. Quantitative evaluation includes: alveolar macrophages, polymorphonuclear cells, type II and intermediate pneumocytes. The results indicate a lung dose-dependent response, observed in both inflammatory and epithelial regeneration components of the alveolar primary reaction. This method could be used to readily evaluate the pulmonary toxic potential of any substance.

Animals↗

PubMed1994

In vivo morphometry of planum temporale asymmetry in first-episode schizophrenia.

Magnetic resonance imaging is a promising tool for in vivo analysis of the neuropathology underlying schizophrenia. One of the most consistent features emerging from the majority of published studies is the lateralization of pathological findings, and this has led to hypotheses of impaired hemispheric specialization in schizophrenia. In previous work, we have validated morphometry of supratemporal language-related cortex using high-resolution magnetic resonance imaging and analyzed the relation of the so defined planum temporale to functional parameters of hemispheric specialization. In the present study, we examined planum temporale structural asymmetry in first-episode schizophrenics. Asymmetry coefficients obtained in these patients did not differ significantly from those in equally right-handed controls and were not correlated to standard psychopathological measures. These data are contrasted with other studies reporting lateralized brain pathology in schizophrenia with special emphasis on methodological considerations in neuroimaging procedures.

Adult↗

PubMed1987

Changes in the distribution of extracellular matrix vesicles during healing of rat tibial bone (computerized morphometry and electron microscopy).

A study of the distribution of extracellular matrix vesicles during the first 3 weeks of healing of adult rat tibial bone was performed by transmission electron microscopy in combination with computerized morphometry. Bone injury comprised removal of the marrow followed by regeneration of the tissue via a phase of primary mineralization. A total number of 39,498 vesicles were traced on electron micrographs and sorted according to their diameters, distance from the calcified front and types. The different vesicular types were defined as follows: (a) vesicles with electron lucent contents, i.e. "empty", (b) vesicles with amorphous electron opaque contents, i.e. "amorphic", (c) vesicles containing crystalline depositions, i.e. "crystal", and (d) vesicles containing crystalline structures with ruptured membranes i.e. "rupture". The vesicles were studied on the days 3, 6, 14 and 21 after bone injury. Most of the vesicles were concentrated between diameters of 0.07 and 0.17 micron. Most of the vesicles were found in a distance less than 3 microns from the calcified front. The sequence of changes of distances from the calcified front and of the vesicular diameters were recorded as follows: "rupture", "crystal", "amorphic" and "empty", the "rupture" type being the closet to the front and of the largest diameter in each day. The results of the present study confirm the accepted hypothesis on calcification via extracellular matrix vesicles. It is thought that the cell secretes "empty" vesicles that accumulate Ca and Pi forming amorphous calcium phosphate that is then converted to hydroxyapatite. This is followed by rupture of the vesicular membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Differences in cortical bone in overloaded and underloaded femurs from ovariectomized rats: comparison of bone morphometry with torsional testing.

Osteopenia and mechanical incompetence are the defining features of osteoporosis, yet the effects of changes in bone structure on mechanical properties are not completely understood. The primary objective of this study was to determine if in a rodent model, changes in cortical bone structure, as measured by static morphometry, would correlate with functional properties, as measured by torsional testing. For this, cortical bone structure and stiffness were determined in overloaded and underloaded limbs in rats rendered ovarian hormone-deficient by ovariectomy (OVX). Rats were OVX and six weeks later a hind limb was immobilized by casting for an additional six weeks. In all cases the femur from the underloaded, casted limb was compared with that from the contralateral, overloaded limb. The success of this experimental protocol was confirmed by differences in femoral cancellous bone volume by microradiography and single photon absorptiometry, with the overloaded limb having a greater bone volume and bone mineral density than the contralateral, immobilized controls. Morphometric differences in the femoral diaphyseal cortical bone included greater endocortical perimeter and endocortical osteoclast surface, less cortical area, less minimum cortical width, and less minimum cortical width at fracture planes when the bones were tested for stiffness in the underloaded, compared with the contralateral, overloaded limbs. Using a torsional test, the ultimate torque to failure and stiffness were less in the underloaded femurs compared with the contralateral, overloaded femurs. These results emphasize the importance of mechanical loading on bones from a gonad-deficient animal.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Standardization of specimen preparation, staining, and sampling methods improves automated sperm-head morphometry analysis.

OBJECTIVES: To evaluate the accuracy and performance of the CellForm-Human automated sperm morphometry instrument (Motion Analysis Corp., Santa Rosa, CA) using different specimen preparation, staining, and analysis techniques. SETTING: Clinical and research andrology and in vitro fertilization laboratories. PATIENTS: Individuals who were undergoing semen evaluation and infertility work-up. RESULTS: The percentage of normal sperm detected by CellForm-Human was not different for washed specimens compared with nonwashed controls. Washing and resuspension to a standard concentration in medium significantly increased spermatozoan density and homogeneity on the slide. Comparison of digitization errors and morphometric measures of sperm stained by the Papanicolaou (PAP) method, the hematoxylin portion of the PAP method, and a new method developed by us, GZIN (pronounced Gee-ZIN), showed that GZIN produced larger measures for length, width, area, and perimeter, but not for width/length, and also produced less variability for most measures than PAP or hematoxylin. The number of digitization errors was significantly less for GZIN than for PAP. Visual inspection revealed that GZIN produced a more consistent stain over the entire sperm head than hematoxylin. The number of sperm analyzed affected the percentage of normal sperm detected. The % normal stabilized only after at least 200 sperm had been analyzed for each man. CONCLUSIONS: Technical variability arising from semen preparation and slide staining methods can be reduced when specimens are washed and resuspended to a standard concentration before smearing. At least 200 sperm should be analyzed to obtain a stable estimate of the percentage of normal sperm. More sperm should be analyzed if the % normal is < 50%. Morphometric measurements are more accurate and precise when sperm are stained with GZIN than when stained with PAP or hematoxylin.

Cytological Techniques↗

PubMed1991

Histology and morphometry of Strongylus vulgaris-mediated equine mesenteric arteritis.

Histological and morphometric evaluation of equine cranial mesenteric arteries was performed on 239 and 89 arteries, respectively. Histological examination revealed that thrombosis and the severity of inflammation varied on a seasonal basis and were directly associated with larval presence. Intimal and adventitial fibrosis were generally of greater severity than medial fibrosis. Fibrosis of the vasa vasorum was less frequent than fibrosis of the artery itself. Morphometry revealed a significant increase in intimal, adventitial and, to a lesser extent, medial area in affected as compared with normal arteries. This change was due to the accumulation of collagen and was considered to result in decreased arterial elasticity. The luminal area varied widely among affected arteries.

Animals↗

PubMed1992

Quantitative morphometry of the adult human bladder.

The primary objective of the present retrospective study was to characterize the effects of aging and BPH on bladder morphometry. Eighty-six bladder specimens were obtained from the autopsy archives of the Milwaukee County Medical Complex. The bladder specimens were divided into 4 groups based upon age and gender: Group I: males between the ages of 35-45 years; Group II: males between the ages of 65-75 years; Group III: females between the ages of 35-45 years; and Group IV: females between the ages of 65-75 years. The age groups were selected in order to identify a group of males with and without BPH. The area density of smooth muscle:connective tissue was determined in bladder specimens using color assisted computer image analysis. Masson-trichrome and double immunoenzymatic staining techniques were used to discriminate the smooth muscle and connective tissue elements of the bladder. The area density of smooth muscle:connective tissue in the Masson-trichrome stained sections was significantly greater in Group I vs. Group II (2.90 +/- 0.22 vs. 2.33 +/- 0.16) and in Group III vs. Group IV (2.85 +/- 0.13 vs. 2.03 +/- 0.20). Aging was associated with a decrease in the area density of smooth muscle:connective tissue ratio in both males and females. The area density of smooth muscle:connective tissue was not significantly different in younger males and females (Group I vs. Group III) and older males and females (Group II vs. Group IV). The present morphometric study suggests that aging and not BPH, is associated with a relative increase in detrusor fibrosis. Infravesical obstruction in BPH may effect bladder function via mechanisms unrelated to the histologic composition of the bladder.

Adult↗

PubMed1990

Nuclear morphometry as a prognostic indicator for genitourinary rhabdomyosarcoma: a preliminary investigation.

Rhabdomyosarcoma of the urogenital tract is a malignant mesenchymal tumor seen primarily in childhood. Multimodal therapy, encompassing surgery, radiotherapy and chemotherapy, has dramatically improved the survival of patients with this disease. However, the quest for markers of tumor aggression is important to decrease the morbidity of treatment given to patients with good prognosis tumors, while at the same time intensifying treatment of tumors with poor prognosis. Using archival tumor specimens from 13 patients with genitourinary rhabdomyosarcoma, a multivariate analysis of multiple nuclear shape descriptors was done with the Hopkins Morphometry System. Three nuclear shape descriptors clearly separated patients with no evidence of disease recurrence or progression from those with recurrent disease, progressive disease or death of disease. These nuclear shape descriptors were standard error of the chain code standard deviation analysis (p = 0.010), range of the feret ellipticity distribution (p = 0.016) and standard error of the chain code range analysis (p = 0.037). With multivariate analysis these shape descriptors taken together separated patients with good and poor prognoses to a level of significance of p = 0.007. Thus, nuclear morphometric analysis may prove to be useful as an individual prognostic indicator in childhood genitourinary rhabdomyosarcoma and warrants further analysis in a much larger, blinded, controlled study.

Cell Nucleus↗

PubMed1985

Ultrastructural morphometry of cultivated smooth muscle cells from normotensive and hypertensive rats.

Cultivated aortic smooth muscle cells (SMC) derived from normotensive and hypertensive rats were analyzed by transmission electron microscopy and ultrastructural morphometry. SMC cultures obtained from hypertensive rats showed an increase in cell size and a higher percentage of large, often polynuclear cells. The stereological data of mitochondria, rough endoplasmic reticulum (RER) and secondary lysosomes were determined by measuring area according to the "Delesse principle". The organelle contents of small and large cells of each group were only slightly different. Significant changes were found between SMC from normotensive and hypertensive rats: The volume density Vv of mitochondria per unit cytoplasm volume was increased up to 48% in small and 50% in large SMC from hypertensive rats compared to those of control animals. The numerical density Nv of secondary lysosomes per unit cytoplasm volume was increased up to 68% in small and 267% in large SMC from hypertensive rats. The content of rough endoplasmic reticulum varied tremendously between the individual cells. All these stereological data represent relative parameters of cell organelles relative to the unit cytoplasm volume. Therefore the differences between the absolute values would be even higher since SMC from hypertensive rats are shown to have a higher mean cell size and volume. We conclude that the effect of experimental hypertension, resulting in an activation of SMC, persists even when the cells are transferred to in vitro conditions.

Animals↗

PubMed1992

Morphometrical investigation of medulloblastoma nuclei by S.A.M. (Shape Analytical Morphometry) software system.

In order to characterize medulloblastomas and to get over the difficulties sometimes encountered in differential diagnosis, a double morphometric procedure has been applied to its nuclei. The first consisted of size measurements (maximum diameter, area and perimeter), the latter is represented by S.A.M. (Shape Analytical Morphometry) software-system specifically implemented to describe shape of biological structure by analytical parameters. Analytical and dimensional parameters submitted to Hotelling's multivariate discriminant analysis gave the best results when used together in convenient discriminant subsets, thereby allowing a good distinction between medulloblastoma in comparison with neuroblastoma, Ewing's tumor, lymphoblastic and lymphocytic lymphoma. These results underline the usefulness of morphometric characterization also for practical diagnostic purposes.

Cell Nucleus↗

PubMed1992

Biological image analysis for microscopical diagnosis: the work-station S.A.M. (Shape Analytical Morphometry).

The S.A.M. (Shape Analytical Morphometry) software system and its related work-station give a generalized and easy-to-handle tool to face a classic and intriguing problem in biomedical morphological diagnosis. What is the shape of an object in a microscopic image? How can we understand the relationship between size and shape? According to Holloway: "Measurements such as length, width, height, whether in chords or arcs only describe space, ... and further run into the abyss of allometric correction ... if additional information (shape?) to size is expected, some method of allometric correction must be used." The S.A.M. software system assumes a logic architecture able to separate and to parametrize independently shape characteristics in terms of allometry and local pertubation by analytical procedures (polynomials, parabolic fitting, Fourier analysis) in addition to the classic evaluations of size and density carried out by image analyzers for microscopical diagnosis.

Diagnosis, Computer-Assisted↗

PubMed1992

Morphometry and prognosis in cancer of the pancreatic head.

The histological slides of 39 cases of cancer of the pancreatic head were analysed using an interactive image analyser system. Some 14 cases were classified as periampullary, 25 as ductal pancreatic cancer. All cases had undergone radical tumor resection according to Whipple's procedure. Morphometric data, tumor size and metastatic nodal involvement were correlated to prognosis. Univariate statistical analysis showed that the classical differentiation between ductal and periampullary cancer was a weaker prognosticator than morphometric variables. In fact, multivariant statistical analysis showed that the morphometric variable irregularity was the best prognosticator (p = 0.0001). No other variable added significant prognostic information. Irregularity is a newly developed variable describing the nuclear shape corrected for roundness. We conclude that morphometry can be of essential prognostic information for the clinician in cancer of the pancreatic head.

Carcinoma, Intraductal, Noninfiltrating↗

PubMed1992

Cytonuclear morphometry in the assessment of dysplasia in colorectal adenomatous polyps. A pilot study.

In order to objectify the degree of dysplasia, results of nuclear and cellular morphometry were compared with assessed grades of dysplasia in 32 colorectal adenomatous polyps. Of these adenomas 8 showed mild, 17 moderate and 7 severe dysplasia (according to blind duplicate assessments by two pathologists). Using a microscope video-overlay interactive digitizing measurement system, 100 nuclei were measured in each specimen according to a strict measurement protocol. In a stepwise discriminant analysis the best discriminating features appeared to be mean stratification index, N/C ratio, mean contour ratio and mean form AR. Overall with these features 65.6% correct jackknifed classification of the 32 cases could be achieved. On the other hand, a clear three-group distinction could not be obtained, even with the best set of discriminating features. Similarly to gastric dysplasia, the morphometric features might be used to design a two-grade (low, high) rather than a three-grade system to assess the degree of dysplasia. These findings make clear that objectifying the degree of dysplasia in adenomatous polyps is possible by means of interactive morphometric analysis.

Adenoma↗