1. Ossification is a bone formation process. It is a process where other connective tissue is replaced by
bone connective tissue. There are two types of ossification, namely intramembranous ossification and
endochondral ossification.
Intramembranous ossification
Intramembranous ossification is a process where the mesenchyme will be replaced with bone tissue.
This ossification does not use the cartilage model. It forms flat bones of skull, facial bones, mandible
and clavicle (collarbone). There are four main stages in intramembranous ossification.
The first stage is the formation of ossification centre. This formation starts with mesenchymal cells
which are centrally located migrates and cluster together to form ossification centres. More than one
ossification centres will form. Then, the mesenchymal cells in ossification centres will differentiate
into osteoprogenitor cells and later these osteoprogenitor cells differentiate into osteoblasts.
Osteoblast then will secrete osteoid. Osteoblasts will continuously produce more osteoid until they are
surrounded by osteoid. Once they are surrounded by osteoid, they will separate from each other but
still remain attached by thin cytoplasmic process.
Next stage is calcification. In this stage, calcium phosphates are deposited in the osteoid to calcify
bone matrix. This calcification of bone matrix traps osteoblasts in spaces. Osteoblast are now called as
osteocytes and housed in lacuna.
Formation of trabeculae is the third stage of intramembranous ossification. More osteoprogenitor
cells rest on surface of calcified bone matrix and differentiate osteoblasts. Osteoblasts add more
osteoid and causes bone matrix to enlarge. The bone matrix from adjacent ossification centres fuse to
form trabeculae. In the spaces between trabeculae, the blood vessels within mesenchyme start to
grow. Meanwhile, the mesenchyme remaining in between trabeculae differentiate into red bone
marrow.
The last stage of intramembranous process is the formation of periousteum. The mesenchyme at the
periphery of the bone condenses to form periosteum. Initially, bone consists of trabeculae only. Then,
spaces between trabeculae at outer part of bone are filled with bone matrix to form compact bone.
Meanwhile, the remaining trabeculae in centres will form spongy bone.
Endochondral ossification
Endochondral ossification occurs within cartilage. Cartilage model is needed for bone formation. The
endochondral ossification results formation of long bones, vertebrae and pelvis.
In long bones formation, during early embryonic development, limb buds which contain mesenchyme
appear. The mesenchyme will form bones of the limb. There are six stages occurred in endochondral
ossification.
1. Formation of hyaline cartilage model.
Mesenchymal cells aggregate and proliferate at the site of future bone. Mesenchymal cells
differentiate into chondroblast, where chondroblasts secrete cartilage matrix (ECM) to form a hyaline
cartilahe model. Now, chondroblasts that are surrounded by ECM are called chondrocytes.
Perichondrium also develops around the cartilage model.

2. 2. Growth of hyaline cartilage model.
Basically, hyaline cartilage model grows in length and width, involving two mechanisms of growth
namely interstitial growth and appositional growth.
For growth in length, we called it as interstitial growth. Here, chondrocytes in the cartilage model do
several works. One is undergoing mitosis and the other one is secreting cartilage matrix. New
cartilage matrix is formed inside the existing cartilage model where we called as ‘growth from
within’. This will thus cause the cartilage to increase in length.
The second mechanism is appositional growth, where it will cause the cartilage to increase in width.
New chondrboblasts are formed by the inner chondrogenic layer of perichondrium. These new
chondroblasts will deposit new cartilage matrix on the outer surface of the existing cartilage model,
causing increasing in width of cartilage.
3. Formation of periosteal bone collar.
Cells in the perichondrium of the diaphysisof the model form osteoblasts. Thus, perichondrium will
become periosteum.Osteoblasts form a layer of bone around the diaphysis which is called as
periosteal bone collar.
4. Calcification of cartilage in centre of diaphysis.
Chondrocytes in the centre of the cartilage model mature & enlarges (hypertrophy).Simultaneously,
surrounding cartilage matrix begins to calcify. Periosteal bone collar & calcified cartilage does not
allow diffusion of nutrients.Chondrocytes that does not receive nutrients will eventually die.Calcified
cartilage breaks down because no chondrocytes to maintain cartilage.Walls of lacuna breakdown thus,
adjacent lacunae become confluent, formingcentral cavity with remnants of calcified cartilage.
5. Formation of primary ossification centre.
In this stage, periosteal bud (blood vessels) enter central cavity of the cartilage model while bringing
osteoprogenitorcelss and hemopoietic cells. Osteoprogenitor cells will form periosteum while
hemopoietic cells form bone marrow. Here, the osteoprogenitor cells rest on surface of remnants of
calcified cartilage and differentiate into osteoblasts. The osteoblasts then lay down osteoid on surface
of remnants of calcified cartilage while simultaneously removing the remnants of calcified cartilage.
This site is called as the primary ossification centre which it is the first site where cartilage is replaced
by bone tissue. After that, osteoid becomes calcified while osteoblasts continue laying down more
osteoid on the calcified bone matrix. The calcified matrix then enlarges to form trabeculae. This
process continues. The primary ossification centre grows towards epiphyses. Eventually, all cartilage
of diaphysis is replaced by bone (trabeculae).Then, osteoclasts resorb trabeculae in the diaphysis, thus
forming medullary cavity.
6. Formation of secondary ossification centre.

3. Secondary ossification centre, which occurs in epiphyses, is the centre of the bone formation that
appears later than the primary ossification centre. The bone formation of secondary ossification centre
is similar to the primary ossification centre except perioestal bone collar is not formed. Here,
chondrocytes in cartilage hypertrophy; the cartilage matrix calcifies and chondrocytes die. The
calcified cartilage matrix will be broken down.Also, the blood vessels together with osteoprogenitor
cells that differentiate into osteoblast will enter epiphyses. Similar to the primary ossification centre,
these osteoblasts lay down osteoid on the remants of calcified cartilage before removes the calcified
cartilage to form trabeculae.
In simple words, the only cartilage that remains is articular cartilage, which located at the articular
surface at ends of bone and epiphyseal cartilage, which locates between the epiphyses and diaphysis.