Histomorphometry represents a microscopic planimetry or stereology. It is used to study homeostasis of the skeleton, mainly in metabolic disorders of bone. The technique is occasionally used to evaluate the skeletal status and treatment effect of rickets and osteomalacia associated with tumors.
For histomorphometry, undecalcified bone sections stained with techniques that enable the visualization of calcified and uncalcified osteoid such as von Kossa’s or Goldner’s (modified trichrome) stains are used. Another frequently used technique is tetracycline-pulse labeling for epifluorescence.
Two short cycles (3 days) of tetracycline approximately 2 weeks apart allow two separate mineralization fronts to be visualized as epifluorescence lines outlining the bone trabeculae of cancellous medullary and endosteal cortical bone. This is due to the ability of tetracyclines to incorporate at the border of osteoid, which is actively mineralized.
The width between the two lines of fluorescence (pulse labels) reflects the rate of mineralization.
A number of parameters can be assessed by histomorphometry of bone.
Total bone volumeThe content of mineralized and non-mineralized bone calculated as the percentage of the total area of tissue examined.
Cancellous bone volume
Mineralized and nonmineralized cancellous bone calculated as the percentage of the total area of tissue examined or as the percentage of the area of the medullary cavity.
Trabecular osteoid volume
Nonmineralized cancellous bone calculated as a percentage of total cancellous bone.
Trabecular osteoid surface: The percent of cancellous bone surface covered by osteoid.
Mineralizing surface: The proportion of trabecular osteoid surface that exhibits tetracycline labeling.
Mineral apposition value: The distance (in microns) between two tetracycline-pulse labels per day.
Bone formation value: The mineralized bone produced in 1 year.
Mineralization lag time: The time (delay) of mineralization of the matrix after its deposition by osteoblasts.
Trabecullar resorptive surface: The percent of bone surface that shows current or prior osteoclastic activity.
Cortical resorptive surface: The percent of the cortical surface that shows current or prior osteoclastic activity.
Periosteal resorptive surface: The percent of periosteal surface with osteoclastic activity.
Trabecular osteoclast count: The number of osteoclasts per area (millimeters squared) of medullary cavity or the number of osteoclasts per length (in centimeters) of the trabecular bone.
Osteoclastic index: The number of osteoclasts per length of the trabecular surface with evidence of present or prior resorptive activity.
Cortical porosity: The percentage of the cortex that contains pores without osteocytic cells.
Accurate measurements of bone resorption usually require specimens taken from two consecutive biopsies.
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