Bone Lab

Learning Objectives

  • Observe and describe the processes of bone remodeling and bone growth.
  • Name the different histological regions of bone.
  • Explain the process of bone turnover.
  • Describe the calcification processes of cartilage and bone.
  • Identify some key pathological examples relevant to bone histology.

Keywords

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Pre-Lab Reading

Introduction

The skeletal system, which is made up of bone and cartilage, serves three primary functions:

  • Mechanical support for sites of muscle attachment
  • Protection of vital organs
  • Ion reserve of Ca and PO for metabolic functions

Bone Cells and Matrix

Bone is a tissue in which the extracellular matrix has been hardened to accommodate a supporting function. The fundamental components of bone, like all connective tissues, are cells and matrix. There are three key cells of bone tissue. They each have unique functions and are derived from two different cell lines.

  • Osteoblasts synthesize the bone matrix and are responsible for its mineralization. They are derived from osteoprogenitor cells, a mesenchymal stem cell line.
  • Osteocytes are inactive osteoblasts that have become trapped within the bone they have formed.
  • Osteoclasts break down bone matrix through phagocytosis. Predictably, they are derived from the monocyte (macrophage) cell line. Think of osteoclasts as the "bone version" of the macrophage. Their activity occurs along their ruffled border, and the space between the osteoclast and the bone is known as Howship's lacuna.

The balance between osteoblast and osteoclast activity governs bone turnover and ensures that bone is neither overproduced nor overdegraded. These cells build up and break down bone matrix, which is composed of:

  • Osteoid, which is the unmineralized matrix composed of type I collagen and glycosaminoglycans (GAGs).
  • Calcium hydroxyapatite, a calcium salt crystal that gives bone its strength and rigidity.

Bone is divided into two types that are different structurally and functionally. Most bones of the body consist of both types of bone tissue:

  • Compact bone, or cortical bone, mainly serves a mechanical function. This is the area of bone to which ligaments and tendons attach. It is thick and dense.
  • Trabecular bone, also known as cancellous bone or spongy bone, mainly serves a metabolic function. This type of bone is located between layers of compact bone and is thin and porous. Located within the trabeculae is the bone marrow.

Macroscopic Bone Structure

Long bones are composed of both cortical and cancellous bone tissue. They consist of several areas:

  • The epiphyses are at the ends of the long bone and are the parts of the bone that participate in joint surfaces.
  • The diaphysis is the shaft of the bone and has walls of cortical bone and an underlying network of trabecular bone.
  • The epiphyseal growth plate lies at the interface between the shaft and the epiphysis and is the region in which cartilage proliferates to cause the elongation of the bone.
  • The metaphysis is the area in which the shaft of the bone joins the epiphyseal growth plate.

Different areas of the bone are covered by different tissue:

  • The epiphyses are lined by a layer of articular cartilage, a specialized form of hyaline cartilage, which serves as protection against friction in the joints.
  • The outside of the diaphysis is lined by periosteum, a fibrous external layer onto which muscles, ligaments, and tendons attach.
  • The inside of the diaphysis, at the border between the cortical and cancellous bone and lining the trabeculae, is lined by endosteum.

Microscopic Bone Structure

Compact bone is organized as parallel columns, known as Haversian systems, which run lengthwise down the axis of long bones. These columns are composed of lamellae, concentric rings of bone, surrounding a central channel, or Haversian canal, that contains the nerves, blood vessels, and lymphatic system of the bone. The parallel Haversian canals are connected to one another by the perpendicular Volkmann's canals.

The lamellae of the Haversian systems are created by osteoblasts. As these cells secrete matrix, they become trapped in spaces called lacunae and become known as osteocytes. Osteocytes communicate with the Haversian canal through cytoplasmic extensions that run through canaliculi, small interconnecting canals.

The layers of a long bone, beginning at the external surface, are therefore:

  • Periosteal surface of compact bone
  • Outer circumferential lamellae
  • Compact bone (Haversian systems)
  • Inner circumferential lamellae
  • Endosteal surface of compact bone
  • Trabecular bone

Bone Development

Bone development begins with the replacement of collagenous mesenchymal tissue by bone. This results in the formation of woven bone, a primitive form of bone with randomly organized collagen fibers that is further remodeled into mature lamellar bone, which possesses regular parallel rings of collagen. Lamellar bone is then constantly remodeled by osteoclasts and osteoblasts. There are two different methods by which bone is produced from mesenchymal tissue:

  • Endochondral ossification is the process by which cartilage is progressively replaced by bone at the epiphyseal growth plates. This occurs in long bones, the vertebrae, and the pelvis.
  • Intramembranous ossification is the process by which mesenchymal tissue is directly replaced by bone without an intermediate cartilage step. It occurs most notably in the bones of the skull.

Bone Remodeling

In adults, after growth has ceased, bone is formed by the osteoblasts only where it was previously resorbed by the osteoclasts. This follows a specific sequence of events, and takes about three months in total to complete:

  • Activation - In the process of activation, osteoblasts induce osteoclasts to break down bone matrix. This occurs via the Receptor Activator for NFkB-Ligand (RANK-L) signaling pathway, in which RANK-L on the surface of osteoblasts binds to RANK on osteoclasts to turn them on. This process lasts for approximately 3 days.
  • Resorption - In resorption, the ruffled border of the osteoclast forms a sealing zone which isolates the area of bone erosion. Organic acids and lysosomal enzymes dissolve the mineral component and break down the organic matrix, respectively. This process occurs at approximately 14 days.
  • Reversal - Over time, osteoblasts begin to replace osteoclasts at the site of bone turnover.
  • Formation - Osteoblasts begin to lay down new lamellar bone on top of old bone. In doing so, cement lines are created to mark the borders between old and new bone matrix. This can take up to 70 days to complete.

Pre-Lab Quiz

  1. Distinguish the following: Haversian canal, Volkmann's canal, canaliculi.
  2. Answer:
  3. Name some bones that are formed through endochondral ossification and some that are formed through intramembranous ossification. What is the key difference between these two types of bone formation?
  4. Answer:
  5. Name and describe the key steps in the sequence of bone remodeling.
  6. Answer:

Slides

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  1. Macroscopic Organization of Bone
  2. Long Bone
  3. Trabecular Bone
  4. Compact Bone
  5. Haversian System
  6. Osteoblasts
  7. Osteoblast EM
  8. Osteoclasts
  9. Osteoclast EM
  10. Bone Remodeling
  11. Intramembraneous Ossification
  12. Endochondrial Ossification
  13. Chondrocyte Growth Sequence

Virtual Microscope Slides

  1. Microscopic Organization of Bone
  2. Observe this slide at low magnification. The calcified bone is purple. The non-calcified elements are light blue. Find areas of compact bone and trabecular bone.
  3. Haversian Systems
  4. Pick a Haversian canal and observe it at high magnification.
  5. Endochondrial Ossification
  6. This slide shows endochondral ossification, the process by which cartilage is calcified to form bone. Begin by distinguishing bone from the surrounding muscle tissue and from the cartilage epiphyses. Indicate the regions in which cartilage is being concerted into bone. Identify the specific areas where chondrocytes are resting, proliferating, maturing, undergoing hypertrophy, and calcifying.

Pathology

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  1. Osteosarcoma
  2. Paget's Disease

Quiz

  1. Name the precursor to this cell.
  2. Answer: This is an osteocyte, which is an osteoblast that has become encased in bone. The embryonic precursor to both of these is the osteoprogenitor cell, a mesenchymal stem cell.
  3. Identify the labeled cells.
  4. Answer: A = osteoclast; note multiple nuclei. B = osteoblast
  5. What will these cells secrete?
  6. Answer: Lysosomal hydrolases, collagenase, acid pH
  7. What is the pale, blue-staining region?
  8. Answer: Epiphyseal Plate
  9. What general process causes the circular formations?
  10. Answer: Bone remodeling
  11. What type of bone formation is taking place?
  12. Answer: Endochondral Ossification
  13. What is the function of this cell and what is its embryological precursor?
  14. Answer: This is an osteoclast, which resorbs bone. It's embryological precursor is a monocyte. In contrast, the precursors of osteoblasts are mesencyhmal stem cells.
  15. This slide shows a resorption canal, where new bone is being laid down. Identify A, B, C, D and E
  16. Answer: A = osteoblasts, B = osteocytes, C = osteoid, D = cement line E = bone
  17. Parathyroid hormone increases the activity of osteoclasts and causes bone to be broken down. However, with an infusion of PTH, there is an initial increase in the creation of new bone matrix. Explain this observation in terms of what you know about osteoclast activation.
  18. Answer: Osteoclast activation requires a previous osteoblast activation. For this reason, the brief surge in osteoblast activity associated with PTH causes a transient increase in bone production before a large increase in bone resorption takes place.