Bone IMS - Cassani

Remember

Correct periosteum, nor periostium

Same with endosteum, not endostium

  • Bone tissue is a type of connective tissue, together with cartilage they are specialized connective tissue

    • Is a structural component of the bones

    • Has a mineralized ECM

    • Is continously remodelled

  • Functions of bone tissue

    • Provide a scaffold for our body

    • protects some organs

    • We have insertion of ligaments and tendons in the bone allowing for locomotion

    • stores minerals, mostly calcium and phosphate

    • Hematopoiesis happens in the red bone marrow

    • Site of energy storage - yellow bone marrow, rich and abundant in lipids

  • Common features among bone and cartilage

    • cells embeded in ECM

    • cells trapped within lacunae

    • have a bilayered connective tissue covering - perisotium supplying it

  • Differences among bone and cartilage

    • in cartilage we have perychordium, while in bone we have ostechordium

    • cells more abundant compared to ECM

    • the bone tissue is vascularized

    • higher metabolic rate and better regenerative properties than cartilage

  • Composition of ECM

    • Inorganic:

      • Crystals of hydroxyapatite - calcium and phosphates, their presence is responsible for rigidity and stiffness, also storage of minerals

    • Organic (35%):

      • mainly composed by collagen fibers:

        • mostly type I

        • but also type V present

      • proteoglycans

        • assembled in

      • multiadhesive proteins

        • stabilise the structure of ECM

        • Most different is:

          • osteonoctin

          • osteopontin

      • bone specific protein

        • osteocalcin - binds ca2+ ions, inhibits calcification of blood vessles

      • growth factoes and cytokines

  • Without the organic components the bone becomes very fragile

  • Without the inorganic components the bone becomes noodly

  • Bone tissue cells

    • osteogenic cells (osteoprogenitors) - give rise to osteoblasts

      • line the periostium

      • located at the surface of the bone

      • derived from mesenchymal stem cells in bone marrow

      • the expression of TF Runx2 is crucial for osteoblast differentiation

        • after knock-out of the gene the osteoblasts are not present in the skeleton

      • several proteins and enzymes are also required for osteoblast maturation

        • collagen

        • alkaline phoshpatase

        • BMP

        • Wnt

    • osteoblasts

      • synthetise and secrete all the components of ECM

      • located on the surface of the bone

      • after secreting ECM most of osteoblasts die, small portion differentiates

      • look like large polygonal or cuboidal cells

      • abundant RER and GA and secretory vesicles

      • Secretory vescicles are especially important during calcification

    • bone-lining cells

      • located near osteoblasts

      • secretory activity can be reaquired when needed

      • lose the secreting ECM ability

      • can sense the osteocy

    • Osteocytes

      • mature cells of the bone

      • Osteocytes can regulate the pre-osteoblasts activity

        • they use mechanosensors

        • secrete a number of factors to regulate the activity

      • derived from osteoblasts

      • trapped in lacunae

      • They have a globular or flattened body

      • have dendrytes, which extend on the ECM and allow for contact in between the osteocytes and other cells

      • have reduced ability to synthetise proteins

    • osteoclasts

      • cells responsible for resorbtion of the bone

      • canaliculi - cavities in which the prolongement of the cells are located

        • between them are gap junctions,which allows for the exchange of info and molecules

      • destroy the tissue, during a process called bone remodelling

      • are derived from the hematopoietic mononuclea cells (GMP)

      • made from the fusion of different cells

      • not derived from mesenchymal origin, they derive from cells of hematopoietic origin

      • when concentration of CA2+ is low in blood, calcitonin is produced stimulating the differentiation of osteoclasts and degradation of the bone, releasing calcium into the blood

      • osteolasts can be found below

      • podosomes are a structure that allows osteoclasts to adhere to the bone surface

      • Osteoclastogenesis

        • M-CSF acts on receptors in progenitors, which leads to proliferation and differentiation to monocytes and then osteocytes

        • RANKL - produced mainly by osteoblasts, stroma cells and immune cells, able to act with the receptor determining the differentiation onto osteoclast

  • Organic components are called osteoid

  • Mineralisation need to elaborate

  • We can distinguish 3 states of the osteocytes

    • quiescent

      • very little cytoplasm around nucleus

      • GA and RER low in abundance

    • formative

      • characterised by the central position at the basal part of the nucleus in the cells

      • cells usually aquire the ability to secrete

    • resorptive

      • increased GA and RER occupying most of the volume of the cells

      • cells able to secrete things into the ECM

  • There are 3 specialize membrane domains

    • Ruffled border- formed at the level of the cells, by microvilli like processes, where bone resorption happens

    • sealing zone/clear zone

      • corresponds to the area of the membrane that firmly attaches to the tissue,

      • devoid of organells

    • basolateral region

      • exocytosis happens of digested materials into the blood

  • Bone resorption

    • starts the deminoralisation of the bone occurs

      • at the ruffle border pH decreases

      • this dissolved the crystals

        • release of calcium and phosphate

  • Resorption lacunae also called Howship lacunae

  • TRAP Staining

    • TRAP staining stands for tartrate-resistant acid phosphatase staining.

    • It is a histological staining technique used to identify and visualize osteoclasts, which are cells responsible for bone resorption.

    • Osteoclasts contain high levels of tartrate-resistant acid phosphatase (TRAP) enzyme, which is the target of this staining method.

    • TRAP staining is commonly used in bone research and pathology to study bone remodeling, osteoporosis, and other bone-related diseases.

    • The staining procedure involves the use of a TRAP-specific substrate that reacts with the TRAP enzyme present in osteoclasts.

    • The substrate is usually a diazonium salt, which forms a colored precipitate upon reaction with TRAP.

    • The stained osteoclasts appear as dark purple or red cells under a microscope, allowing for their identification and quantification.

    • TRAP staining can be performed on tissue sections or cultured cells, depending on the experimental setup.

    • The staining results can be further analyzed using image analysis software to measure osteoclast number, size, and activity.

    • TRAP staining is often combined with other staining techniques, such as hematoxylin and eosin staining, to provide additional information about the bone tissue and surrounding cells.

    • Overall, TRAP staining is a valuable tool in bone research, providing insights into osteoclast biology and the pathogenesis of bone diseases.

  • If the osteoclast activity is increased the pathologies may occur

    • osteoporosis

    • Paget disease of the bone

      • first phase of significatn reduction of bone density

      • followed by dramatic deposition of bone mass

      • results in bone deformities

    • rheumatoid arthritis

      • autoimmune disease

  • If the osteoclast activity is decreased the pathologies may occur:

    • osteopetrosis

      • opposite to osteoporosis

      • huge increase in bone mass

      • May be hereditary:

        • Autosomal recessive osteopetrosis (ARO)

          • reduced osteoclast activity

          • no cure and therapy

          • main defects:

            • bone defects

              • increase in the mass of the bone

            • hematological defects

            • neurological defects

          • there are types of ARO

            • osteoclast-poor - osteoclasts are very few

              • due to genetic defects in the RANK and RANKL genes

            • osteoclast-rich - osteoclasts are present

  • HSCT in RANKL-dependent ARO

  • Types of bones

    • long bones

      • middle portion shaft - diaphysis

      • two ends - epiphysis

    • short bones

    • flat bones

    • irregular bones

  • Bone classification

    • sponge bone (cancellous bone)

      • made by traveculae

      • spaces occupied by blood vessels and bone marrow

    • compact bone

      • the outer layer of the bone

      • higher bone mass

  • Long bone structure:

    • diaphysis

    • eppiphysis x2

      • sometimes on the surface cartilage may occur

    • inside the diaphysis is a cavity,through which goes

      • bone marrow

      • blood vessels

    • covered on the external surface by periosteum

    • the cavities are covered by connective tissue called endosteum

  • Blood supply of the long bone

    • arteries enter the bone at the level of diaphysis through foramina

    • they divide forming a network of blood vessels

  • Bone membranes

    • periosteum

      • dense regular connective tissue

      • outer layer - more fibrous

      • inner layer - whith osteogenic layer

      • rich in blood vessels and nerves, also lymphatic

        • this is as far as lymphatic system goes

      • bound to the bone through Sharpey’s fibers

    • endosteum

      • thinner thatn periosteum

      • possess osteogeneic cells

  • Bone marrow cavity

    • red bone marrow

      • hematopoietic place

    • yellow bone marrow

      • rich in fat droplets

  • Microscopic mineralised bone

    • Woven bone (immature)

      • turns into lamellar bone

      • contains high amount of cells per volume copared to the mature bone

      • randomly distributed cells

      • less strong bone than the lamellar bone

    • Lamellar bone (mature)

      • composed by structural units called osteon

        • a system of circular lamellae jak słoje drzewa

        • inside is the Haversian cannal

          • inside tha cannal are blood vessels and nerves

        • the system is also called Haversian system

      • Volkmann cannal

        • connect 2 Haversian cannals

  • Lamellae system

    • osteon

    • outer circuferential lamellae

      • located in the most peripheral part of the bone

      • just below periosteum

    • internal circumferential lamellae

      • line the medullary cavity

    • interstitial lamellae

      • located in between osteons

      • derived from remodelling of the previous osteons

  • How the development of the new osteon occurs

    • in the beginning a resorption cavity is formed

      • a long circular cavity, like a tunnel

    • blood vessels and surrounding tissue occupies the cannal

    • osteoblasts are formed and multiply forming a bone

    • we can differentiate a new bone cone and a resorption cone

  • Spongy bone

    • made by irregular lamellae

    • don’t have Haversian cannal and so the moelcules are supplied through the cavities

  • Ossification

    • intramembous ossification (direct)

      • starts from mesenchyme

      • mesenchymal cells proliferate and form a small cluster of cells

      • cells start to differentiate into the progenitors and osteoblast forming a ossification center

      • They secrete osteoid which traps osteoblasts

      • the trabecular matrix and peristeum form

    • endochondral ossification (indirect)

      • takes place starting from a template of cartilage tissue

      • at first a bone collar is fomed in the perichondrium region

      • it’s creation is parallel to initial calification of ECM in cartilage

      • the calcification makes the cells to become hypertrophic

      • that causes them to die

      • at the same time blood vessels make osteoblast progenitors arrive at first ossification center

      • then a secondary ossification center

  • Long bone can grow

    • longitudinal growth

      • only during puberty

    • appositional growth

      • throughout the entire life

  • Vascular invasion at the level of epiphyseal plate

  • Bone repair

    • release of vasodilatation of blood vessels

    • formation of hematoma

    • reqruitment of progenitors from the blood

    • formation of a soft callus

      • made by cartilage and soft connective tissue

    • removal of the soft callus

    • formation of a travecular bone (sponge bone)

    • remodelling of the bone fragment

    • deposition of compact bone at the peripheries

  • Optimum Ca2+ levels in blood - 9-11mg/100ml of blood

  • Bone deposition affected by

    • Ca and P

    • Vit D

    • Vit C

    • Vit A

    • Vit B12

    • Growth horone

    • calcitonin

    • parathormone

    • sex hormones