Impact of Zinc Fingers and Homeoboxes 3 on the Regulation of Mesenchymal Stem Cell Osteogenic Differentiation

Abstract

We propose zinc fingers and homeoboxes 3 (ZHX3) as new osteogenic markers for mesenchymal stem cells (MSC). ZHX3 mRNA expression was upregulated within 1-6 h after incubation of MSCs in the osteogenic induction medium, and reached maximum levels after 24 h of incubation. Two to 4 days later, ZHX3 mRNA levels had decreased sharply. Maximal mRNA levels were 3- to 5-fold higher than those in the undifferentiated state. In contrast, Runt-related transcription factor2 (RUNX2) mRNA expression was downregulated at 2-4 h after incubation, and levels were only enhanced 1.4-fold after 12 and 24 h of incubation. Further, Osterix mRNA levels increased only 1.6-fold after 4 and 24 h of incubation. Thus, ZHX3 expression may be a better marker of MSC osteogenic differentiation than RUNX2 or Osterix expression at the initial stage of differentiation. Knockdown of ZHX3 using 2 distinct small interfering RNA (siRNA) oligonucleotides had little effect on cell morphology or on MSC proliferation, regardless of the differentiation state of the cells. However, ZHX3 siRNAs suppressed Osterix, but not RUNX2 mRNA expression, within 1 h of osteogenic differentiation, and this suppression was sustained for at least 24 h. The 2 ZHX3 siRNAs also suppressed alkaline phosphatase induction and matrix mineralization (assessed using alizarin red staining), and, further, suppressed the calcium content of the cultures at a later stage of differentiation (days 6-21). The effects of ZHX3 siRNAs on the osteogenic differentiation were comparable to those of RUNX2 and Osterix siRNAs. These findings suggest that ZHX3 is involved in the switch from the undifferentiated state of MSC to an osteogenic program, and that ZHX3 may be useful as an early osteogenic differentiation marker.

The Effect of Quercetin on the Osteogenesic Differentiation and Angiogenic Factor Expression of Bone Marrow-Derived Mesenchymal Stem Cells

“…These results indicated that quercetin could not only promote the proliferation of rBMSCs but also enhance osteogenic differentiation at various stages, especially at the middle and late stages; additionally, the concentration of 2 μM was found to be the optimum concentration. However, previous studies have shown that the cellular morphology is one of aspects, which can influence the osteogenic differentiation of MSCs [ 61 – 63 ]; the other studies also showed that some transcription factors (TFs) and Chinese medicine, such as zinc fingers and homeoboxes 3 (ZHX3), Runx2 and Tinospora cordifolia, could promote the osteogenic differentiation of MSCs by regulating the gene expression and secretion of osteogenic markers without significant effect on the cellular morphology [ 64 – 66 ]. Present study also showed that quercetin could stimulate the osteogenic differentiation of rBMSCs by promoting the gene expression and secretion of osteogenic markers without obvious effect on the cellular morphology.…”

ZHX3 promotes the progression of urothelial carcinoma of the bladder via repressing of RGS2 and is a novel substrate of TRIM21

“…By screening the UCB dataset from TCGA, we identified that zinc finger and homeobox 3 ( ZHX3 ) was either gained or amplified in 59.6% (243/408) of patients with UCB. The ZHX3 gene, which is mapped to chromosome 20q12, has been found to be associated with many kinds of biological processes, such as cell differentiation and metabolism 8,9 . It has been suggested that ZHX3 is a ubiquitous transcriptional repressor expressed in human cells and tissues 10 .…”

<p>Attenuated ZHX3 expression serves as a potential biomarker that predicts poor clinical outcomes in breast cancer patients</p>

“…From their identification, ZHX family members have been identified as transcriptional repressors through interacting with the α-subunit of the nuclear factor Y (NF-YA) and forming homodimers and heterodimers with each other 9–13. Increasing evidence has revealed that ZHX factors are major transcriptional mediators involved in the events including development and differentiation of hematopoietic cells, maintenance of neural progenitors and osteogenic differentiation of mesenchymal stem cells 8,14,15. Dysregulation of ZHX factors has been found to be correlated with initiation and progression of diverse diseases such as neurological, hematological and glomerular diseases 8,16,17.…”