What Is the Connection Between Osteoblasts and Osteocytes?

Osteoblasts (osteoblast, OB) are mainly differentiated from 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 process of bone formation and reconstruction.

Chinese name: osteoblast
Foreign name: osteoblast

Origin: Pluripotent bone marrow stromal cells
cell cycle: 20 24
Function: The main functional cells of bone formation

Osteoblasts and their functions

At different maturity stages, osteoblasts show 4 different morphologies in vivo, namely preosteoblast, osteoblast, osteocyte and bonelining cell. Pre-osteoblasts are precursors of osteoblasts, differentiated by stromal stem cells, developed along the osteoblast lineage, and are located outside the osteoblasts that cover the bone-forming surface. Mature osteoblasts are monolayer cells located on the surface of bones, and they play an important role in synthesizing bone matrix.

Bone cells are mature and ultimate differentiated cells in the osteoblastic lineage. Bone cells are embedded in mineralized bone tissue, and superficial bone cells still retain part of the osteoblast structure. Team cells are a layer of flat or rectangular cells arranged on the surface of most bones of an adult. Active osteoblasts are spindle-shaped, cone-shaped, or cubic, with basophilic cytoplasm. The nucleus is located at one end of the cell, the nucleoli is obvious, and short protrusions on the surface connect with adjacent cells. Under the electron microscope, the cytoplasm has a typical protein synthesis structure-rich rough endoplasmic reticulum and ribosome, and the Golgi apparatus is more developed.

In biochemistry and histochemistry, osteoblasts are rich in alkalinephosphatase (ALP) and have glycogen. 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-, bone morphogenetic protein), and increase the level of cytoplasmic cAMP Stimulate DNA and collagen synthesis (Aronow et al., 1990).

Osteoblasts are the main functional cells of bone formation and are responsible for the synthesis, secretion and mineralization of bone matrix. Human and animal bone tissues are constantly being reconstructed. The process of bone reconstruction includes the breakdown and absorption of bones and the formation of new bones. Osteoclasts are responsible for bone breakdown and absorption, while osteoclasts are responsible for new bone formation. Osteoclasts are attached to the old bone area, secreting acidic substances to dissolve minerals, secreting proteases to digest the bone matrix, and forming bone resorption pits; thereafter, osteoblasts migrate to the site of absorption, secrete bone matrix, and mineralize Form new bone. The balance between osteogenesis and osteogenesis is the key to maintaining normal bone mass.

During bone formation, osteoblasts undergo four stages of osteoblast proliferation, extracellular matrix maturation, extracellular matrix mineralization, and osteoblast apoptosis (Tong Anli et al., 1999). The phenotype development of osteoblasts in the in vitro culture system is similar to the development and differentiation in vivo, and they have to undergo three stages: cell proliferation, extracellular matrix maturation and matrix mineralization. Type I corogen and some non-collagen proteins are markers of osteoblast differentiation, which can reflect the phenotypic characteristics of osteoblast differentiation.

Osteoblasts in vitro culture

The main sources of osteoblasts are bone, periosteum, bone marrow and extra bone tissue. Human embryo skulls or skulls of newborn animals are common sources of osteoblasts. Robey (1985) treated cancellous bone mass with collagenase to remove connective tissue and bone marrow hematopoietic tissue, and then cultured the processed bone mass to obtain more pure osteoblasts. The fibroblast-like cells obtained from human embryo skull were cultured for 3 weeks after being induced to differentiate by adding -glycerol phosphate. Cell calcification was seen, indicating that the cells obtained were osteoblasts with strong differentiation ability. Cells obtained by digesting fetal skulls with ethylenediaminetetraacetic acid and collagenase were cultured in vitro, and it was found that they can continuously proliferate more than 20 times on artificial materials and have high ALP activity. Human cancellous bone was used to establish an osteoblast culture model in vitro and a large number of purified osteoblasts were obtained. Adult female rat cancellous bone was selected to obtain a larger number of osteoblasts in a short period of time. Cells in the periosteum have long been shown to form bone and cartilage under appropriate conditions. Vacanti et al. (1993) isolated osteoblasts from the periosteal periosteum of newborn calves and planted them on porous polyglycolic acid scaffolds. After 7-10 days, osteoblasts proliferated.

High-purity osteoblasts were obtained from the periosteal membrane on the inner side of the upper end of the tibia of New Zealand rabbits using collagenase short-time pre-digestion. Bone marrow is divided into two major systems: hematopoietic and matrix. Its osteogenic ability is derived from the matrix. Zhang Yingang et al. (2000) cultured fetal rabbit long tube bone marrow cells in vitro and proved that they have the characteristics of osteocyte lineage and can be used as seed cells for repairing bone defects. Pericutaneous cells, vascular endothelial cells, fibroblasts, and myoblasts have all been reported to be transformed into osteoblasts.

Regulatory factors of osteoblast proliferation and differentiation

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 through the regulation of the cell cycle, that is, the regulation of cell replication and DNA division under the action of mitogen. The effects of hormone levels and cell regulatory factors on the proliferation and differentiation of osteoblasts will be described below.

Osteoblast nuclear binding factor

Nuclear binding factor-1 (CBF-1) is specifically expressed by osteoblasts and is a factor determining osteoblast differentiation, and the osteoblast differentiation pathway regulated by it is irreplaceable (Tou et al., 2001). CBF-1 is a key gene for bone formation and determines the occurrence and differentiation of osteoblasts. It plays an important role in maintaining normal bone growth and development (David et al., 2000).

The results of the study have demonstrated that in addition to regulating osteoblast differentiation, CBF-1 also regulates the functions of differentiated osteoblasts and the expression of other growth factors, thereby controlling the physiological processes of bone formation and development after birth (Tamara et al., 2001). CBF-1 not only plays a specific regulatory role in osteoblast differentiation, but also plays an indispensable role in the process of cartilage formation and endochondral ossification. CBF-1 transcription factor and its regulation of osteoblast differentiation are widely recognized and accepted, but the molecular mechanism of CBF-1 gene expression regulation and its induction of osteoblast differentiation needs to be further explored.

Ogawa et al. (1993) first cloned CBF-1 / p56 cDNA from mouse fibroblasts and found that it was expressed in T lymphocyte strains, NIH3T cells, thymus, and testis tissues. Duey et al. (1998) found that during the embryonic development of mice at 12.5 days after mating, higher CBF-1 mRNA expression was found in the skull, central axis, and limb bone mesenchymal cell aggregation regions, and these cells were able to differentiate into Dual-energy precursor cells of osteoblasts and chondrocytes. However, overexpression of CBF-1 can also stimulate osteoclastic bone resorption. Xiao et al. (1999) analyzed the CBF-1 gene structure of mice, rats, and humans and the expression of three CBF-1 subtypes, and found that type II CBF1 is expressed in osteoblasts of all species and type III CBF- 1 is expressed in mouse and rat but not human osteoblasts. Therefore, type II CBF-1 may play a more important role in the differentiation of osteoblasts. Komori et al. (1997) used a knockout mouse model and found that CBF-1 knockout heterozygous mice were significantly obstructed at birth and showed clinical symptoms similar to human clavicle skull dysplasia syndrome. However, homozygous mice with CBF-1 gene knockout were born without osteoblasts and bone tissue. Lengner et al. (2002) found in the study of transgenic mice that the overexpression of CBF-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-1 plasmid in the bone injury site, and found that the recovery of the plasmid insertion group was faster than that of the non-plasmid group, and there was obvious cartilage formation at the edge of the bone injury wound Viable osteoblasts were observed, while the control group had fewer active osteoblasts and no new bone formation, suggesting that the local application of CBF-1 can promote the recovery of bone injury.

D Osteoblast vitamin D receptor

In recent years, with the continuous research and application of vitamin D in bone metabolism diseases, it has been found that vitamin D can regulate the microenvironment of bone and affect the function of bone cells. Vitamin D is a steroid hormone with multiple physiological functions. 1.25 (OH) 2D3 is the most active metabolite of vitamin D, mainly acting on mature cells. Studies have shown that the reason why 1.25 (OH) 2D3 can regulate the function of osteoblasts is mainly through the receptor-mediated gene pathway, VDR (Norman et al., 2002). Guo Lijuan et al. (2005) showed that 1.25 (OH) 2D3 can stimulate the expression of MT1-MMP in human osteoblasts. MT1-MMP can directly or indirectly break down the unmineralized bone matrix. Therefore, 1.25 (OH) 2D3 can stimulate human The expression of MT1-MMP in osteocytes degrades bone matrix such as unmineralized type I collagen and initiates bone resorption. Vitamin D receptor (VDR) is a nuclear macromolecule that mediates the biological effects of 1,25 (OH) 2D3. VDR is a gene transcription regulatory protein, which belongs to the steroid hormone receptor superfamily and is located in the nucleus of the cell. It is one of the more widely studied genes related to bone metabolism. VDR is essentially a ligand-dependent nuclear transcription factor, which plays an important role in maintaining the body's calcium and phosphorus metabolism and regulating cell proliferation and differentiation. It has become a hot topic in the field of bone and endocrinology in recent years (Gu Fengying et al., 2008). VDR exists not only in osteoblasts, but also in osteoclasts. The role of VDR in bone tissue is two-way. VDR on osteoblasts can regulate VDR on osteoclasts, which can inhibit its proliferation and promote its differentiation, thus playing a two-way regulatory role on bone synthesis and catabolism ( Gu Fengying et al. (2008). VDR on osteoblasts can promote the synthesis of osteopontin (OPN) and osteocalcin (OC), and participate in bone formation and mineralization. OPN is a matrix protein secreted by osteoblasts, which is very important for cell adhesion and migration. 1.25 (OH) 2D3 combines with VDR, induces osteoclasts to move to the surface of bone matrix, combines with OPN, removes aging bone tissue, and synthesizes new bone tissue. OC is mainly synthesized and secreted by osteoblasts. It is the most abundant non-collagen protein in bone tissue. Most of OC is deposited in the extracellular bone matrix. A small amount of newly synthesized OC is released into the blood circulation. The content of OC in serum is synthesized with osteoblasts. The total amount is positively correlated, which can specifically reflect the activity of osteoblasts, and is a specific indicator of the state of bone regeneration and bone formation in the body. It can regulate the formation of mineral crystals and promote the mineralization of bone matrix.

Osteoblast calcitonin

Calcitonin is a peptide hormone secreted by thyroid C cells. It is an important hormone to maintain calcium and phosphorus metabolism in the body. Calcitonin participates in bone metabolism by inhibiting the activity and number of osteoclasts and promoting the formation of osteoblasts (Zhang Yongli et al. , 2005): In recent years, it has been widely used to treat diseases characterized by acute or chronic bone loss, such as deformable osteitis, senile osteoporosis, hypercalcemia, and malignant osteolysis. Calcitonin is a powerful osteoclast inhibitor that directly, rapidly and widely inhibits bone resorption. The results show that calcitonin not only affects osteoclasts, but also has a direct effect on osteoblasts (Xiao Yonghua et al., 2002). As early as 1991, Farrry et al. Found that calcitonin can directly affect human osteoblasts and stimulate the proliferation and differentiation of osteoblasts. It is inferred that calcitonin may be the main factor that promotes the proliferation, differentiation and mineralization of osteoblasts. one. Kobayashi et al. (1994) showed that calcitonin can directly affect

Bone cells (MC3T3E1), stimulate the expression of insulin-like growth factor (IGF-I), fos tumor gene (c-fos), type collagen and osteocalcin mRNA in mouse osteoblasts, stimulate mouse osteoblast proliferation and Differentiation. Drissi et al. (1997) showed that human osteosarcoma cells and human osteoblasts can express calcitonin and exogenous calcitonin-related peptide (CGRP). Calcitonin has a stimulating effect on the proliferation, differentiation and mineralization of rat osteoblasts cultured in vitro, and can also prevent the apoptosis of osteoblasts (Zhu Jianmin et al., 2001). The results of CGRP research show that exogenous calcitonin can increase the number and size of osteoblast colonies. Lian Kai et al. (2002) cultured osteoblasts derived from cranium of SD rats with CGRP culture solution. The results confirmed that exogenous CGRP can promote osteoblast proliferation in a dose-dependent manner. Imai et al. (2002) used a genetic recombination method to cultivate transgenic rats in which osteoblasts can secrete a large amount of CGRP. It was found that osteoblast activity was greatly enhanced, bone synthesis significantly exceeded bone resorption, and bone volume was significantly increased compared with the wild type control And further shows that osteoblasts can enhance the activity of themselves and surrounding cells by means of autocrine CGRP. CGRP not only acts as a neurotransmitter. However, how CGRP participates in the normal metabolism of osteoblasts, and the specific mechanism of its role needs further research.

Osteoblast transforming growth factor

Transforming growth factor- (TGF-) is a growth factor with more content in osteoblasts. Osteoblasts themselves can synthesize TGF-, and there are specific receptors for TGF- on the cell membrane of osteoblasts. TGF- can act on osteoblasts and regulate their proliferation and differentiation (Lu Weizhong et al., 2000). The transforming growth factor- (TGF-) family includes TGF-s, bone morphogenetic proteins 2-7 (BMPs 2-7), actin, and inhibitors. TGF- is an important local factor of bone metabolism, which can stimulate the proliferation and differentiation of a variety of bone tissue cells. TGF- is an inactive macromolecular complex in the early stage of synthesis, and there is a large amount of inactive TGF- in the bone matrix. , when the pH is reduced or plasmin and cathepsin are activated, inactive TGF- can be activated to regulate the formation of new bone in the bone resorption zone. TGF- stimulates DNA synthesis and cell proliferation in non-transformed osteoblasts. TGF- isoforms TGF1, TGF-2, and TGF-3 may play different functions in regulating osteoblastic factor secretion, extracellular interstitial production, and cell maturation (Fagenholz et al., 2001). TGF- is also a powerful osteoblast chemokine, which can move and aggregate osteoblasts from a low TGF- concentration to a high TGF- concentration area, and increase the number of migrated cells by 4 times (Lind , 1998). In vivo tests have shown that TGF- has a strong osteogenic and chondrogenic effect. Noda et al. (1989) first injected exogenous TGF- directly into the skulls of mice and rabbits and found that TGF- has a significant promotion effect on bone formation and a significant increase in osteogenesis. Marcellic et al. (1996) found that TGF- injected under the rat femur periosteum induced differentiation of periosteal mesenchymal cells into osteoblasts and chondrocytes, and stimulated the proliferation of these cells and the extracellular matrix characteristic of bone and cartilage. Protein synthesis. Richards et al. (1999) found that the distraction performed on the tibia of test rabbits can make osteoblasts in the stretched area proliferate actively and enhance osteogenesis. The culture of osteoblast-like cells found that TGF- has a direct and potent chemotactic effect on osteoblast-like cells. Therefore, the increase of TGF- during distraction osteogenesis has a positive effect on mesenchymal cells and osteoblasts. Migration may be an important moderator.

Osteoblast bone morphogenetic protein

Bone morphogenetic protein (BMP) is a kind of bone growth factor that is closely related to osteoinduction. It is the main induced repair factor in bone repair. BMP is a recognized high-efficiency osteoinductive factor, which is widely present in human and animal bone tissues, but is mainly concentrated in the cortical bone of the backbone. It is produced by human and animal osteoblasts and neoplastic osteoblasts, which diffuse into the cancellous bone and bone marrow as bone reconstruction progresses. BMP has the ability to induce osteoblast differentiation and to induce osteogenesis in vitro (Chen et al., 1997). Bone morphogenetic proteins are members of the transforming growth factor- (TGF-) superfamily and are a family of substances. At least 15 BMPs have been discovered so far. Urist (1965) first used decalcified bone matrix to induce ectopic osteogenesis in muscle. The results of this test suggest that the bone matrix may contain an active protein that has the ability to direct the differentiation of undifferentiated mesenchymal cells. For osteoblasts and the ability to form bone tissue, it was later named BMP. With the development of molecular biology and genetic engineering, BMP-13 has been discovered by 1996, and corresponding cDNA clones have been obtained. Among BMP213, the research on osteogenesis of BMP-2 has been reported the most (Wzney et al., 1998: Cleste et al., 1990: Qaynak et al., 1990: Oaynak et al., 1992). The results of in vitro and in vivo experiments have demonstrated that BMP-2 has the ability to promote osteoblast differentiation and induce osteogenesis in vitro (Chen et al., 1997). BMP-2 is one of the most important extracellular signaling molecules that promote bone formation and induce osteoblast differentiation. It exerts its osteogenic effect by activating Smads signaling and regulating osteogenic gene transcription. Studies have found (Liu et al., 2007) that BMP-2 can up-regulate the expression of 66 genes, including 13 related transcription factors such as Smad6, Smad7, and Msx2. Gazzerro et al. (1999) found that BMP-2 can reduce the expression of collagenase 11 mRNA in cells and thus reduce the expression of collagenase in osteoblasts, which is beneficial to matrix synthesis. Hay et al. (1999) added recombinant human BMP-2 (50 g / L) to human newly generated osteoclast cells (HNC) and continued to culture for 3 to 7 days, which could affect immature HNC cells and enhance their ALP. active. Matrix mineralization increased significantly at week 2, and osteocalcin mRNA and protein levels and calcium content in the matrix increased at week 3, suggesting that osteoblasts were fully differentiated. Ming Liu et al. (2006) found that local injection of BMP-2 can significantly increase the local cortical thickness, bone density, and bone mechanical strength of the upper femur in elderly rats. This study also shows that BMP-2 plays an important role in osteogenesis. The research of osteoblasts is still a hot topic, such as the research in bone tissue engineering, the research on medical prevention and treatment of osteoporosis, etc. However, there are still many problems. Many studies are still in the preliminary exploration stage, and systematic and in-depth research is needed. I believe that osteoblasts will have very good application prospects.