MiR-214 is essential in stimulating and maintaining various physiological processes, specifically cellular proliferation, followed by tissue formation, which supports growth and development. Numerous studies have revealed the significance of miR-214 in regulating its downstream targets, subsequently triggering bone development (Fig. 2). It plays an imperative role in bone remodelling and chondrocyte differentiation (Iaquinta et al., 2021).

MiR-214 interacts with various intracellular downstream targets, including Col2a1, Mgp and Sox9 in ATDC5 chondrogenic cell lines, PTEN in pre-osteoclasts, ATF4 in human periodontal ligament stem cells, and aids in the development of tissues. It can either positively (indicated in green) or negatively (indicated in red) regulate cellular differentiation.

In recent years, miR-214 has been found to be involved in regulating intricate bone formation and resorption mechanisms. Yao et al. (2017) identified that miR-214 targets ATF4 to suppress the differentiation of human periodontal ligament stem cells into osteoblastic cells, thereby inhibiting bone formation via the miR-214/ATF4 axis. Likewise, in human MSCs (hMSCs), miR-214 overexpression inhibited osteogenesis by targeting β-catenin (Li et al., 2017). A study demonstrated that the downregulation of miR-214 triggered by physical exercise generated-strain on osteoblasts could stimulate osteogenesis in vivo and in vitro, followed by an upregulation of bone markers including ALP, ATF4 and Osx. However, miR-214 overexpression inhibited the expression of ALP, ATF4, and β-catenin, thus reducing osteogenic differentiation (Yuan et al., 2019). Another study by Li et al. (2016) identified that exosomal miR-214 derived from osteoclast induces osteoclast differentiation and suppresses osteoblast differentiation via targeting osteoblast-specific markers including ALP, COL, OPN, OC, and bone sialoprotein (BSP) in the osteoblastic cells, thereby reducing bone formation via increasing resorption. Their results suggested that miR-214 interacts with PTEN and triggers osteoclastogenesis via the Akt/PI3K signalling. Further, osteoclast-specific markers such as tartrate-resistant-acid phosphatase 5a (TRAcP5), semaphorin-4D (Sema4D), and cathepsin K (CTSK) were detected in serum EVs produced by osteoclasts, which could be transmitted to osteoblastic cells to suppress osteogenesis (Li et al., 2016).

Reports have also suggested the participation of miR-214 in the regulation of cartilage formation. Roberto et al. (2018) demonstrated an elevated expression of miR-214 in undifferentiated ATDC5 cells (chondrogenic cell line). Increased miR-214 was observed to target Col2a1 (collagen type II alpha 1), Mgp (Matrix gla protein), and Sox9 (SRY-box transcription factor 9). These genes regulated the differentiation of chondrogenic cells into mature chondrocytes, which was, however, restricted by the inhibition of their activities in the presence of miR-214 (Roberto et al., 2018).

The role of miR-214 in bone physiology is well recognized, as discussed above, but recent studies have shown its participation in regulating different pathological states such as osteoporosis, osteoarthritis, and cancers, including osteosarcoma and Ewing’s sarcoma (Fig. 3).

MiR-214 controls several life-threatening diseases that mostly affect bone tissue homeostasis and prominent molecular mechanisms involved in bone growth and development. It significantly inhibits HMGA1 activity in Ewing’s sarcoma cells to prevent tumor growth and TRAF3 in mesenchymal stem cells from inducing their differentiation into osteosarcoma cells. Additionally, miR-214 obstructs the functions of IKKβ in chondrocytes to impede cartilage degradation. miR-214 also restricts the expression of Osterix in osteoblasts to suppress their differentiation, which culminates in osteoporosis, followed by leakage of Ca²⁺ ions.

Osteoporosis is the most prominent and well-known skeletal anomaly that causes fragility in bones leading to enhanced chances of fracture occurrence (Ensrud and Crandall, 2017). miR-214 has been linked to the reduced differentiation of MSCs into osteoblasts and a rapid decline in the mineralization of the bone matrix, thereby triggering osteoporosis (Ge et al., 2017). Mohamad et al. (2019) showed the regulatory association between miR-214 and Osx in promoting primary osteoporosis, characterized by reduced bone mass and which mostly affected the trabecular and cortical bones. They identified that the expression profiles of miR-214 and Osx are inversely correlated. Further, they reported a reduction in serum calcium levels and an increase in ALP levels in osteoporotic patients (Sözen et al., 2017; Mohamad et al., 2019). Apart from that, a study found YAP1 (Yes-associated protein 1), a Hippo pathway component, to be a potential target of miR-214 in osteoporotic patients. In particular, YAP1 favours the differentiation of hMSCs into osteocytes. However, miR-214 binds to YAP1 and downregulates its expression, which hinders the differentiation of hMSCs, confirming that miR-214 inhibits YAP1, thereby inducing osteoporosis (Zhong et al., 2021).

Osteoarthritis is the most typical form of bone disability that affects and structurally modifies the whole joint. The structural modification comprises the formation of osteophytes, inflammation in the synovial portions, and degradation of cartilaginous tissues, thereby severely affecting the quality of life (Charlier et al., 2019). A study highlighted the intricate function of miR-214-3p in osteoarthritis. In chondrocytes, the Inhibitor of nuclear factor kappa-B kinase subunit β (IKKβ), an important member of the nuclear factor kappa-B (NF-kB) family, serves as an initiator of the NF-kB pathway. Activation of the NF-kB signalling pathway disrupts processes such as extracellular matrix (ECM) metabolism, followed by the degradation of chondrocytes, thus leading to osteoarthritis. However, miR-214-3p directly targeted and inhibited the activity of IKKβ to exert a chondroprotective effect that aids in preventing the disruption of ECM and chondrocyte apoptosis (Choi et al., 2019; Cao et al., 2021).

Osteosarcoma refers to bone cancer that mostly occurs in children, including adolescents, and primarily originates from a single cell in the marrow of long bones, giving rise to a large heterogeneous tumor mass (Brown et al., 2017). Recent research has identified the participation of miR-214 in regulating osteosarcoma (Rehei et al., 2018). miR-214 was found to act as an oncogene and directly regulated the expression of TNF receptor-associated factor 3 (TRAF3) in osteosarcoma cell lines and patient tissue samples. TRAF3, a member of the TRAF family of adapter proteins, is found in almost all bone cells and tissues. When miR-214 was expressed, it reduced the expression levels of TRAF3, thereby leading to osteosarcoma (Rehei et al., 2018). Similarly, Cheng et al. (2020) reported that miR-214-3p directly inhibits Fibronectin type III domain-containing protein 5 (FNDC5)/irisin in osteosarcoma cells to promote their migration and invasion. This study indicated that FNDC5 could act as a prospective inhibitor to obstruct the viability and procession of the osteosarcoma cells that are reversed by miR-214-3p (Cheng et al., 2020).

Ewing's sarcoma (EWS) refers to a malignant bone tumor, commonly occurring in the ribs, tibia, pelvis, and femur regions and majorly affects children and young adults. Histologically, EWS belongs to a diverse group of tiny round cell sarcomas, and each cell is morphologically similar to the other (Grünewald et al., 2018). A recent study suggested the regulatory role of miR-214-3p in EWS (de Feo et al., 2022). EWS has two prominent hallmarks, Cluster of differentiation 99 (CD99) and Ewing’s sarcoma-friend leukaemia integration 1 (EWS-FLI1). While EWS-FLI1 is an anomalous transcription factor, CD99 is a cell-surface protein molecule (Sen et al., 2018; Manara et al., 2016). It was determined that higher expression levels of CD99 and EWS-FLI1 are exclusively responsible for causing EWS malignancy. Typically, CD99 interacts and binds to EWS-FLI1 to promote malignancy during EWS. In addition, these hallmarks have been described to repress the activity of miR-214-3p, a tumor suppressor in EWS. miR-214-3p has been shown to target HMGA1 (High-mobility group AT-hook 1), which is a small histone protein molecule that engages in tumor growth and metastasis. miR-214-3p overexpression suppressed HMGA1 expression to restrain EWS, but this was opposed by inhibition of miR-214-3p activity in the presence of EWS-FLI1 and CD99, which led to severe EWS malignancy (de Feo et al., 2022).

Together, the findings discussed above illustrate the diverse functions of miR-214 in regulating a wide range of physiological processes in the skeletal system of our body and development at a molecular level.

LncRNAs are a class of ncRNAs that are greater than 200 nucleotides in length and perform several functions, including chromatin remodelling, epigenetic modifications on DNA, serving as protein scaffolds, and acting as miRNA sponges (Paraskevopoulou and Hatzigeorgiou, 2016). Over the past years, several lncRNAs have been identified to promote or suppress osteoblast differentiation. While the lncRNAs MALAT1 (metastasis-associated lung adenocarcinoma transcript 1) and H19 (imprinted maternally expressed transcript) promoted osteoblast differentiation (Xiao et al., 2017; Liang et al., 2016), lncRNAs HOTAIR (HOX transcript antisense RNA) and DANCR (differentiation antagonizing non-protein-coding RNA) inhibited osteoblast differentiation (Wei et al., 2017; Zhu and Xu, 2013). Several lncRNAs that target miR-214 have been reported to date. For instance, lncRNA KCNQ1OT1 sponged miR-214 and upregulated BMP2 expression, thereby stimulating the osteogenic differentiation of BMSCs (Wang et al., 2019a). Similarly, lncRNA XIST (X-inactive specific transcript) stimulated the osteogenic differentiation of periodontal ligament stem cells (PDLSCs) by sponging miR-214-3p (Feng et al., 2020). Stromal cell-derived factor-1 (SDF1) regulated osteoblast differentiation by stimulating BMP2 expression (Hosogane et al., 2010). A recent study demonstrated that lncRNA H19 targets miR-214-5p upon SDF1 exposure and increases BMP2 expression, hence promoting the osteoblast differentiation of BMSCs (He et al., 2021).

Li et al. (2020) demonstrated that lncRNA LOC100506178 sponged miR-214-5p and promoted BMP2 expression, subsequently favoring the differentiation of BMSCs towards osteogenic lineage. Furthermore, they identified that lncRNA LOC100506178 overexpression in BMSCs and transplantation of the same in nude mice reverses the inhibitory effect of miR-214-5p and promoted ectopic bone formation in vivo (Li et al., 2020). While miR-214 was often reported to negatively regulate osteoblast differentiation, Yang et al. (2021) identified that miR-214 could promote osteoblast differentiation by targeting thioredoxin-interacting protein (TXNIP). TXNIP is responsible for bone loss during osteoporosis, hence using a rat osteoporosis model, they found that lncRNA MEG3, which sponges miR-214, was upregulated in the osteoporosis condition. Further, siRNA-mediated silencing of lncRNA MEG3 or miR-214 overexpression led to increased expression of OPG, thereby enhancing osteoblast differentiation. Thus, lncRNA MEG3/miR-214/TXNIP is important in understanding and treating osteoporosis (Yang et al., 2021).

Long-intervening ncRNAs (lincRNAs) are a unique group of lncRNAs without protein-coding ability. LincRNAs have been recently identified and are known to play a role in several diseases, including osteosarcoma (Deniz and Erman, 2017). A recent study revealed that LINC00657 induced osteogenic differentiation of BMSCs via targeting miR-214-3p, and upregulating BMP2 expression (Li et al., 2022). Since lincRNAs are only gaining interest recently, we speculate the involvement of many novel lincRNAs that would be important regulators of osteoblast differentiation.

CircRNAs are single-stranded, ncRNA molecules with persistent loop configuration and are known to regulate gene expression. They are highly stable molecules with high tissue specificity, making them ideal for therapeutics. Although circRNAs have been discovered for a long time, their role in human physiology and disease has only recently been investigated (Verduci et al., 2021). CircRNAs can sponge miRNAs and regulate osteoblast differentiation. For instance, circ_0006873, circ_FAT1, and circ_0066523 promoted osteoblast differentiation by sponging their respective target miRNAs (Ye et al., 2021; Xin et al., 2021; Lv et al., 2022).

Another study in a mice osteoporosis model demonstrated that circ_ITCH1 sponges miR-214 and upregulates the expression of YAP1. Higher expression of YAP1 corresponds to enhanced osteoblast differentiation, making circ_ITCH1 an important biomarker and a therapeutic target for the treatment of osteoporosis (Zhong et al., 2021). Runx3 is a positive mediator of osteoblast differentiation and a direct target of miR-214 (Wang et al., 2017). Circ_33287 was observed to sponge miR-214-3p and protected Runx3 from miR-214-3p mediated inhibition, thereby promoting osteogenic differentiation in maxillary sinus membrane stem cells. Further, overexpression of circ_33287 and miR-214-3p inhibition was identified to stimulate ectopic bone formation in vivo. Hence, circ_33287/miR-214-3p/Runx3 axis presents a novel molecular approach for bone regeneration in the posterior maxilla (Peng et al., 2019). Studies have demonstrated that circRNAs could regulate the proliferative and invasive capacity of osteosarcoma cells (Ji et al., 2020). Mao et al. (2021) identified the competing endogenous RNA (ceRNA) role of circ_XPR1 in osteosarcoma cells. They reported that circ_XPR1 sponges miR-214-5p and promotes the proliferative capacity of osteosarcoma cells via regulating the expression of DEAD-Box Helicase 5 (DDX5), a target of miR-214-5p. Their findings suggested that circ_XPR1/miR-214-5p/DDX5 axis may serve as an important therapeutic target for osteosarcoma (Mao et al., 2021). Table 1 summarizes the involvement of ncRNA/mir-214/mRNA axes in bone-related diseases.

The above-discussed reports highlighted the emerging role of ncRNAs targeting miR-214 in treating various bone-related disorders. However, many of these ncRNAs are still in the preclinical stage due to the challenges in developing RNA-based therapeutics. Challenges such as degradation by RNases and the inability to cross cell membranes are a few concerns that have to be addressed for the efficient development of ncRNA-based therapeutics.