Introduction and Function of Osteoblast
Osteoblasts (OB) are mainly differentiated from the mesenchymal progenitor cells in the inner and outer periosteum and bone marrow and can specifically secrete a variety of biologically active substances, regulating and affecting the formation and reconstruction of bone. At different stages of maturation, osteoblasts manifest in four different forms in the body, namely pre-osteoblasts, osteoblasts, bone cells, and formation cells. Pre-osteoblasts, which are precursors of osteoblasts, differentiate from stromal stem cells and develop along the osteoblast lineage, on the outside of osteoblasts that cover the surface of the bone formation. Mature osteoblasts are monolayers of cells located on the surface of the bone that carries important functions in the synthesis of bone matrix. Osteoblasts are mature and ultimate differentiated cells in the osteogenesis of osteoblasts. Bone cells are embedded in mineralized bone tissue, and superficial bone cells still retain part of the osteoblast structure. A squamous cell is a layer of flat or rectangular cells arranged on the surface of most of the adult bone. Active osteoblasts are fusiform, conical or cuboid, and the cytoplasm is basophilic. The nucleus is located at the end of the cell, the nucleolus is obvious, and the surface has short protrusions connected to adjacent cells. Electron microscopy has a typical protein synthesis structure in the cytoplasm - abundant rough endoplasmic reticulum and ribosome, Golgi is more developed. In biochemistry and histochemistry, osteoblasts are rich in alkaline phosphatase (ALP) and have glycogen present. Osteoblasts can be activated by different kinds of hormones (such as parathyroid hormone and prostaglandin E2) and growth factors (such as insulin-like growth factor, transforming growth factor-β, bone morphogenetic protein) to increase the level of cytosolic cAMP, and stimulates the synthesis of DNA and collagen. Osteoblasts are the main functional cells of bone formation and are responsible for the synthesis, secretion, and mineralization of the bone matrix. Human and animal bone tissue is continuously reconstructed. The bone reconstruction process includes the decomposition and absorption of bone and the formation of new bone. Osteoclasts are responsible for bone breakdown and absorption, and osteoblasts are responsible for new bone formation. Osteoclasts are attached to the old bone area, secreting acidic substances to dissolve minerals, secreting protease to digest the bone matrix, and forming a bone resorption lacuna; thereafter, the osteoblasts migrate to the absorbed site, secreting the bone matrix, and the bone matrix is mineralized and form new bones.
Expression Maker of Osteoblast
Now, the different markers of osteoblasts expressed at different stages are introduced in detail, which plays an important role in the development and function of osteoblasts. Alkaline phosphatase (ALP): early osteogenic markers, ALP is mainly distributed on the cell membrane, promote cell maturation, calcification, quantitative detection of ALP can reflect the differentiation level of osteoblasts. The higher expression of ALP activity is an early indicator of osteogenic differentiation and maturation. When ALP activity is enhanced, bone formation is enhanced, and bone mineralization is promoted. Therefore, ALP activity is a good indicator of the degree of osteoblast differentiation and functional status. Kaplow is used for staining ALP, and brownish is positive for ALP and green is the nucleus. Positive staining is mainly distributed at the edge of the new trabecular bone. Runx2: Runx2 is expressed in the early stage of wound healing. The early expression is high expressed in the nucleus. By using HE stains and found that Runx2 mainly distributed around the new trabecular bone and margin. Early promote differentiation, late inhibition of differentiation. Runx2 is expressed in OB, chondrocytes, myoblasts, and fibroblasts differentiated from MSCs and are an important transcription factor that determines the differentiation of MSCs into OB and OB development. Runx2 can be used with osteoblast-specific cis-acting elements. Binding promotes the transcription of OCN, osteopontin, bone protein and Coll, indicating that Runx2 not only regulates the differentiation of OB but also participates in the regulation of OB function. OCN: Osteoblast secreted protein, osteocalcin is the main component of bone non-collagen protein and is a specific secreted protein of bone tissue. At present, serum osteocalcin is a marker for the function of osteoblasts. After osteocalcin synthesis, osteocalcin is mostly deposited in the bone matrix, and a small part is released into the blood. The total amount of osteocalcin synthesized by the calcium domain in the blood circulation is highly correlated, and it responds to transient changes in bone metabolism. ConI: In the osteogenesis stage, type I collagen is the main collagen, which is secreted by osteoblasts. Type I collagen is the most important fiber collagen component in the bone matrix. The expression of extracellular matrix and alkaline phosphatase gene, as well as the expression of osteocalcin and osteopontin genes during matrix mineralization, are the main manifestations of osteoblast differentiation.
Research Status of Osteoblast
The main sources of osteoblasts are bone, periosteum, bone marrow and extra-bone tissue. Domestic and foreign literature reports that animals (rats, mice, chickens, rabbits) and human embryonic skulls or skulls of newborn animals are common sources of osteoblasts. Robey (1985) used collagenase to treat cancellous bone to remove connective tissue and bone marrow hematopoietic tissue and then cultured the bone to obtain more pure osteoblasts. Riccio et al. (1991) cultured fibroblast-like cells obtained from human embryonic skulls by adding β-glycerophosphate for 3 weeks, showing cell matrix calcification, indicating that the obtained cells are osteoblasts with strong differentiation ability. Malekzadeh et al. (1998) cultured cells obtained by digesting fetal skull with ethylenediaminetetraacetic acid and collagenase in vitro and found that they can proliferate more than 20-fold on artificial materials and have high ALP activity. Wang Haibin et al (1985) took human cancellous bone to establish an in vitro culture model of osteoblasts and obtained a large number of purified osteoblasts. The differentiation process of osteoblasts is affected and regulated by genetic factors, hormone levels, and cell regulatory factors. The regulation of osteoblast proliferation is mainly regulated by regulation of the cell cycle, that is, the replication of DNA and cell division by the action of mitotic mitogens. The effects of hormone levels and cell regulatory factors on the proliferation and differentiation of osteoblasts are described below. Nuclear Binding Factor-α 1 (CBF-α 1): CBF-α 1 is specifically expressed by osteoblasts and is a factor determining osteoblast differentiation, and its regulated osteoblast differentiation pathway is irreplaceable (Tou et al. 2001). cbf-α 1 is a key gene for bone formation and determines the development and differentiation of osteoblasts, which plays an important role in maintaining normal bone growth and development (David et al., 2000). The results have shown that in addition to regulating osteoblast differentiation, CBF-α 1 also regulates the function of differentiated osteoblasts and the expression of other growth factors, thereby controlling the physiological processes of postnatal bone formation and development (Tanara et al., 2001). CBF-α 1 not only plays a specific role in the regulation of osteoblast differentiation, but also plays an indispensable role in cartilage formation and endochondral ossification. The CBF-α 1 transcription factor and its regulation of osteoblast differentiation have been widely recognized and accepted, but the molecular regulation of cbf-α 1 gene expression and its induction of osteoblast differentiation needs further exploration. Ogawa et al. (1993) first cloned CBF-α 1/p56 cDNA from mouse fibroblasts and found that they were expressed in T lymphocyte cell line, NIH3T cell, thymus and testis tissue. According to Duey et al. (1998), higher CBF-α 1 mRNA expression was found in the skull, midshaft bone and limb bone mesenchymal cell aggregates during embryonic development at 12d after mating in mice. It is a dual-energy precursor cell that can differentiate into osteoblasts and chondrocytes. However, overexpression of CBF-α 1 also stimulates osteoclastic bone resorption. Xiao et al analyzed the cbf-α 1 gene structure and the expression of CBF-α 1 isoforms in mice, rats and humans, and found that type II CBFα 1 was expressed in osteoblasts of all species, type III CBF-α 1 is expressed in mouse and rat but not human osteoblasts. Therefore, type II CBF-α 1 may play an even more important role in the differentiation of osteoblasts. Komori et al. (1997) used a knockout mouse model to find that CBF-α 1 knockout heterozygous mice were significantly blocked at birth and showed clinical signs like human clavicular skull dysplasia syndrome. The homozygous CBF-α 1 knockout mice developed no osteoblasts and bone tissue. Lengner et al. (2002) found that overexpression of CBF-α 1 affected the maturation of adult rat osteoblasts, increased bone formation and bone resorption, enhanced bone metabolism, and reduced bone mineralization. Clark et al. (2004) placed a type I collagen sponge containing CBF-α 1 plasmid in the bone injury site and found that the recovery of the plasmid insert group was faster than that of the non-implanted group, and there was significant chondrogenesis and wound healing in the bone. Tissue engineering is an interdisciplinary subject in bioengineering, materials science, cell and molecular biology, chemical engineering, and surgical medicine. It uses tissue engineering methods to induce osteoblasts, bone growth factors that specifically induce cells, and has excellent biological activity. HA/TCP porous scaffold materials with biodegradability, mechanical strength, osteoconductivity, and osteoinductive ability, etc., after constructing tissue engineering materials, they will have excellent ectopic and ortho-inducing ability to promote bone defects, repair and reconstruction, its development prospects are very attractive. In the future, the main research directions for osteoblasts in tissue engineering are: Large-scale production of engineered tissues requires suitable, healthy and expandable seed cells; Application of cell growth factors to tissue engineering technology, to study its regulation mechanism and release regulation on cell regulatory and sustained-release carriers; Bioreactor and osteoblast/scaffold structure cryopreservation of bionic cells that can be cultured and scaled. If there is a breakthrough in the exploration and research in these fields, it will show a whole new aspect to animals, especially to human health.