Stereology and morphometry of lung tissue.

This chapter deals with the stereological quantification of structural characteristics of the lung. The aim of design-based stereological methods is the unbiased and efficient estimation of structural features without making any assumptions on the underlying nature of the biological sample. The methods are based on rigorous sampling of location and orientation, the application of appropriate test systems, and the controlling of the precision of the estimates. Here, we describe the workflow from the fixation of the lung over the processing of the tissue samples to gaining estimates on the structural properties of the lung. Specifically, this chapter deals with methods for estimating the reference volume, sampling location, and sampling orientation, estimating volumes and surface areas of alveolar compartments, estimating total alveolar number, performing stereology at light and electron microscopic level, and dealing with technical problems such as tissue shrinkage. The procedures are illustrated using a worked example from the authors' own laboratory.

[1]  H. Bachofen,et al.  Electron microscopy of rapidly frozen lungs: evaluation on the basis of standard criteria. , 1982, Journal of applied physiology: respiratory, environmental and exercise physiology.

[2]  M. Ochs,et al.  A brief update on lung stereology , 2006, Journal of microscopy.

[3]  H. Fehrenbach.  Animal models of pulmonary emphysema: a stereologist's perspective , 2006, European Respiratory Review.

[4]  H J G Gundersen,et al.  The smooth fractionator , 2002, Journal of microscopy.

[5]  C. Hsia.  Quantitative morphology of compensatory lung growth , 2006, European Respiratory Review.

[6]  H. Gundersen,et al.  The isector: a simple and direct method for generating isotropic, uniform random sections from small specimens , 1992 .

[7]  M. Ochs,et al.  Studying lung ultrastructure , 1998 .

[8]  H. Gundersen,et al.  Notes on the estimation of the numerical density of arbitrary profiles: the edge effect , 1977 .

[9]  H. Gundersen,et al.  Stereological estimation of the volume‐weighted mean volume of arbitrary particles observed on random sections * , 1985, Journal of microscopy.

[10]  E. Weibel,et al.  Is Length an Appropriate Estimator to Characterize Pulmonary Alveolar Capillaries? A Critical Evaluation in the Human Lung , 2010, Anatomical record.

[11]  E. Weibel,et al.  Architecture of the Human Lung , 1962, Science.

[12]  Adrian Baddeley,et al.  Stereology for Statisticians , 2004 .

[13]  E. Weibel,et al.  American Thoracic Society Documents An Official Research Policy Statement of the American Thoracic Society/European Respiratory Society: Standards for Quantitative Assessment of Lung Structure , 2010 .

[14]  Matthias Ochs,et al.  The number of alveoli in the human lung. , 2004, American journal of respiratory and critical care medicine.

[15]  V. Howard,et al.  Unbiased Stereology: Three-Dimensional Measurement in Microscopy , 1998 .

[16]  J R Nyengaard,et al.  Tissue shrinkage and unbiased stereological estimation of particle number and size * , 2001, Journal of microscopy.

[17]  T. Mattfeldt,et al.  Estimation of surface area and length with the orientator , 1990, Journal of microscopy.

[18]  D M Hyde,et al.  Total number and mean size of alveoli in mammalian lung estimated using fractionator sampling and unbiased estimates of the Euler characteristic of alveolar openings. , 2004, The anatomical record. Part A, Discoveries in molecular, cellular, and evolutionary biology.

[19]  P. Woodruff,et al.  Quantitative morphology using bronchial biopsies , 2006, European Respiratory Review.

[20]  H. Gundersen,et al.  Sampling for stereology in lungs , 2006, European Respiratory Review.

[21]  M. Ochs,et al.  Visualization and quantitative analysis of nanoparticles in the respiratory tract by transmission electron microscopy , 2007, Particle and Fibre Toxicology.

[22]  M. Ochs,et al.  Microscopy-based quantitative analysis of lung structure: application in diagnosis. , 2011, Expert opinion on medical diagnostics.

[23]  E. Weibel.  Why measure lung structure? , 2001, American journal of respiratory and critical care medicine.

[24]  Peter R. Mouton,et al.  Unbiased Stereology: A Concise Guide , 2011 .

[25]  K. Pinkerton,et al.  Asthma: a comparison of animal models using stereological methods , 2006, European Respiratory Review.

[26]  T. Mayhew,et al.  Taking tissue samples from the placenta: an illustration of principles and strategies. , 2008, Placenta.

[27]  E R Weibel,et al.  A simple tool for stereological assessment of digital images: the STEPanizer , 2011, Journal of microscopy.

[28]  M. Ochs.  Stereological analysis of acute lung injury , 2006, European Respiratory Review.

[29]  E. Weibel.  Morphometry of the Human Lung , 1965, Springer Berlin Heidelberg.

[30]  H. J. G. Gundersen,et al.  The new stereological tools: Disector, fractionator, nucleator and point sampled intercepts and their use in pathological research and diagnosis , 1988, APMIS : acta pathologica, microbiologica, et immunologica Scandinavica.

[31]  Matthias Ochs,et al.  How much is there really? Why stereology is essential in lung morphometry. , 2007, Journal of applied physiology.

[32]  D. C. Sterio.  The unbiased estimation of number and sizes of arbitrary particles using the disector , 1984, Journal of microscopy.

[33]  M. Ochs,et al.  Stereological estimation of the volume weighted mean volumes of alveoli and acinar pathways in the rat lung to characterise alterations after ischaemia/reperfusion , 1999, Journal of anatomy.

[34]  E. Weibel,et al.  Variation of lung volume after fixation when measured by immersion or Cavalieri method. , 2003, American journal of physiology. Lung cellular and molecular physiology.

[35]  W. Scherle,et al.  A simple method for volumetry of organs in quantitative stereology. , 1970, Mikroskopie.

[36]  L M Cruz-Orive,et al.  Estimation of surface area from vertical sections , 1986, Journal of microscopy.