Chondrocytes Cell Profile Report

Introduction

Cartilage is a type of connective tissue in the body that contains specialised cells called chondrocytes. As seen in figure 4.4, these chondrocytes are situated inside cavities called lacunae and are embedded in the cartilaginous extracellular matrix (ECM) [1] in which they help to produce and maintain. Cartilage has many functions which may vary from embryo to adulthood. In embryos, its main functions are to provide structural support and to prepare for the formation of the axial skeleton during a process called endochondral ossification [2]. Endochondral ossification is a vital process which occurs during the development of many bones in the body. In this way, hyaline cartilage, which is in an abundance in embryos and neonatal babies is replaced by bone through a series of steps. In human embryos, this process occurs at around ten to eleven weeks [3], while mice embryos usually commence endochondral ossification at around 15.5 days (Figure 1 [4]). Figure 1 explains the progression of the chondrocytes throughout the ossification, starting from the mesenchymal cells which then differentiate into chondrocytes and chondroblasts (details of which will be discussed later in the report). The differentiated cells proliferate and chondrocytes near the middle of the bone (Primary ossification centre [4]) secrete matrix. Cells in the primary ossification centre are labelled as hypertrophic once they have matured and are larger than those in the zone of proliferation. The various zones can be seen in figure 2[5] and figure 3.4. Hypertrophic chondrocytes then go on to die through apoptosis, leaving behind a cavity for osteoblasts and blood vessels to enter [3]. The end result is the formation of the bone with hyaline cartilage remaining at the ends of the long bones. Endochondral ossification is a tightly regulated process involving various interactions between transcription factors such as Sox9, growth factors, and the extracellular matrix [6]. Some of the signalling pathways affected are the Indian Hedgehog signalling pathway (Ihh), bone morphogenic proteins (BMPs) and Fibroblast growth factors (FGFs) [7][8]. These pathways are also essential for the development of chondrocytes, or chondrogenesis. As with all cell types, chondrocytes journey through a sequence of events leading from fertilization to blastocyst stage and then to gastrulation within the first 14-16 days. Totipotent cells of the zygote are able to differentiate into the three germ layers, and in this case the mesoderm. From there, development continues through to the paraxial mesoderm, onto Somitogenesis, and finally differentiates into the sclerotome, from which chondrocytes are derived. Again, these are regulated through many factors like Sonic hedgehog signalling (Shh) and BMP. Chondrocytes begin as mesenchymal progenitor cells and are differentiated by BMP-induced signalling of Sox9 and TGF-beta [8]. At this stage, they are considered chondroblasts. Chondroblasts can be considered chondrocytes once they embed themselves into the ECM and are surrounded by the lacunae.

On the other hand, cartilage in adults may have several functions depending on the type of cartilage and its location in the body. Some of these functions include acting as cushioning for joints and bones and working to communicate with the surrounding tissues and other cells in the body [9]. Cartilage in adults is generally less plentiful than in earlier stages of life due to the previously mentioned process of endochondral ossification. The three types of cartilage are hyaline, elastic, and fibrocartilage [1]. These differ in the constituents of their matrices, including the type and amount of collagen present [9]. Hyaline cartilage is the most common type, and is found in structures like synovial joints, the nose and the trachea (which can be seen in figure 3.1.). Elastic cartilage, as its name suggests, contains elastic fibres and are the most flexible type. The external ear is an example of an area which contains elastic cartilage. The last type is fibrocartilage, which differs from the other two as it contains type 1 collagen in conjunction with collagen type two [1]. It is found predominantly in the vertebral column in the intervertebral discs and can withstand more pulling or compressive forces than the other two. While the three types of cartilage have some differences, they mainly share similar compositions of collagen, peptidoglycans (aggrecan), chondrocytes and the ECM. The ECM has an important role in the transportation and communication of substances. As cartilage is avascular [9] and does not contain nerve endings, chondrocytes must gain nutrients through diffusion through the ECM, hence the reason why cartilage is slow repairing. Consequently, these properties explain why Osteoarthritis (OA) is such a slow-healing disease and why sufferers are unaware of their condition until the cartilage has completely worn away and the bones rub together in a painful manner. The topic of Osteoarthritis will be further analysed in the discussion section.

Results

Figure 3.1 shows an overall image of a mouse embryo section taken with no scanning objective. The slide was placed over the light on the microscope. Since it is a sagittal section, there are many structures that can be identified including the brain, heart, and liver. The use of Haemoxylin/Eosin stain enables a better viewing of the tissue types in the section. The dark blue structures stained with Alcian blue specify the cartilage structures. As can be seen in the figure, cartilage is present in many different areas of the embryo and is a prominent feature that will mostly become bone (pre-ossified). Some structures, like the trachea for example, will maintain some cartilage and not proceed with ossification.

Figure 3.2 is a snapshot of the upper region of the vertebral column, where the blue areas indicate the presence of chondrocytes. The image was taken at 4X objective and enables for a wider view of the vertebrae. From this image, the gradation of colour of resting chondrocytes and pre-hypertrophic/hypertrophic can be seen from an overall point of view.

Figure 3.3 shows the same vertebrae at a higher objective (10X). In this image, the zones mentioned in figure 2 are more visible and individual chondrocytes can be seen. The red dots also show the beginnings of red blood cell accumulation, which is another indication that the mouse embryo is still in the early stages of development as blood vessels have not formed/intruded.

Figure 3.4 is an image of the mouse embryo at 40X objective. This image clearly shows the individual chondrocytes surrounded by the lacunae. Near the region of (a), it can be seen that