Table 1.

MiRNAs identified in chondrogenic differentiation of stem cells, target mRNA, effect of miRNA modulation on chondrogenesis and potential use of identified miRNAs to enhance chondrogenic differentiation

MiRNA	Expression during chondrogenesis	mRNA targets of miRNA and mRNA function in chondrogenesis	Reported effect of miRNA modulation on chondrogenesis	Potential use of miRNA in inducing chondrogenesis
miR-29a	miR-29a was reported to be down-regulated during chondrogenic differentiation of human MSCs (hMSCs) [58, 59].	miR-29a was demonstrated to directly target the 3’UTR of FOXO3A. Transcription factor FOXO3A was observed to be up-regulated during chondrogenesis, and modulation of FOXO3A expression was found to regulate SOX9, AGCAN and COL2A1 expression, with FOXO3A binding sites also identified within the genomic sequences of these genes [59]. miR-29a is likely to be down-regulated during chondrogenesis enabling derepression of FOX3A expression.	The overexpression of miR-29a in hMSCs, using pre-miR-29a, resulted in inhibition of chondrocyte-specific markers and a suppressive effect on chondrogenic differentiation. [59].	Decrease endogenous miR-29a levels with a miR-29a inhibitor.
miR-140-3p	miR-140-3p was reported to be up-regulated during chondrogenic differentiation of hMSCs [60].	mRNA targets of miR-140-3p remain unknown [60].	–	Increase miR-140-3p levels with a miR-140-3p mimic.
miR-140-5p	miR-140-5p was reported to be up-regulated during chondrogenesis of hMSCs [60, 61] and equine cord blood mesenchymal stromal cells [62].	MiR-140-5p was demonstrated to directly target the 3’UTR of RALA. RALA encodes Ras-related protein Ral-A (RALA), a small GTPase which functions to bind and hydrolyse guanosine triphosphate. RALA has been shown to interact with the exocyst complex in mediating cytoskeletal and secretory pathways [63]. RALA has been shown to be involved in TGF-β signalling through internalisation of membrane receptor activin type II [64] and also may be involved with the trafficking and secretion of glycosaminoglycans [65].	Inhibition of endogenous miR-140-5p in differentiating hMSCs, using anti-miR-140-5p, resulted in impaired chondrogenesis with an observed down-regulation of SOX9 and aggrecan. Knockdown of RALA resulted in the up-regulation of SOX9 [60].	Increase miR-140-5p levels with a miR-140-5p mimic.
miR-145	miR-145 was reported to be down-regulated during chondrogenic differentiation of murine MSCs [66].	miR-145 was demonstrated to directly target the 3’UTR of Sox9 [66]. SOX9 is required for aggrecan [67], Col2a1 [68], Col9a1 [69] and Col11a2 expression [70] and has been shown to bind to chondrocyte-specific enhancer elements in all of these genes. miR-145 is likely to be down-regulated during chondrogenesis enabling derepression of Sox9 expression.	Overexpression of miR-145 in C3H10T1/2 cells, using pre-miR-146, resulted in the down-regulation of chondrogenic differentiation, evidenced by down-regulation of Sox9 protein and Col2a1, Agcan, Col9a2, Col11a1 and COMP mRNA. Endogenous inhibition of miR-145, using anti-miR-145, resulted in enhancement of chondrogenic differentiation, evidenced by the up-regulation of Sox9 protein and Col2a1, Agcan, Col9a2, Col11a1 and COMP mRNA [66].	Decrease endogenous miR-145 levels with a miR-145 inhibitor.
miR-146a	miR-146a was reported to be down-regulated in chondrogenic epiphyseal cell populations isolated from the epiphyses of human foetal femora [71].	miR-146a was suggested to target positive mediators of chondrogenic signalling, SMAD2 and SMAD3 [71]. miR-146a is likely to be down-regulated during chondrogenesis enabling derepression of SMAD2 and SMAD3 expression.	Overexpression of miR-146a in cells derived from the epiphyses of human foetal femora, using miR-146a mimic, resulted in SOX9 down-regulation, indicating the negative effect of miR-146a on chondrogenesis [71].	Decrease endogenous miR-146a levels with a miR-146a inhibitor.
miR-146b	miR-146b was reported to be down-regulated during chondrogenic differentiation of human SSCs [72•].	miR-146b was suggested to target early chondrogenic transcription factor SOX5 [72•]. Early transcription factor SOX5 is co-expressed with SOX6 and SOX9 to enhance Col2a1 expression [73] and to enable SOX9 binding to the AGCAN enhancer [67]. miR-146b is likely to be down-regulated during chondrogenesis, enabling derepression of SOX5 expression.	Overexpression of miR-146b in human SSCs, using miR-146b mimic, resulted in down-regulation of SOX5 [72•].	Decrease endogenous miR-146b levels with a miR-146b inhibitor.
miR-193b	miR-193b was reported to be up-regulated during chondrogenic differentiation of human adipose-derived stem cells (hADSCs) [74, 75] and ATDC5 cells [74].	miR-193b was demonstrated to directly target the 3’UTRs of Tgfb2 and Tgfbr3 [74].	Overexpression of miR-193b in ATDC cells, using miR-193b mimic, resulted in the down-regulation of chondrogenic differentiation, evidenced by the down-regulation of early chondrogenic markers col2a1, sox9 and comp as well as Tgfb2 and Tgfbr3. Inhibition of endogenous miR-193b, using anti-miR-193b, resulted in the enhancement of chondrogenic differentiation, evidenced by the up-regulation of the early chondrogenic markers and Tgfb2 and Tgfbr3 [74].	Decrease endogenous miR-193b levels with a miR-193b inhibitor.
miR-194	miR-194 was reported to be down-regulated during chondrogenic differentiation of hADSCs [76].	miR-194 was demonstrated to directly target the 3’UTR of SOX5 [76]. Early transcription factor SOX5 is co-expressed with SOX6 and SOX9 to enhance Col2a1 expression [73] and to enable SOX9 binding to the AGCAN enhancer [67]. miR-194 is likely to be down-regulated during chondrogenic differentiation enabling derepression of SOX5 expression.	Overexpression of miR-194 in hADSCs, using pre-miR-194, resulted in the down-regulation of chondrogenic differentiation, evidenced by the down-regulation of the chondrogenic markers COL2A1, COL9A2, COL11A1, AGC1 and COMP. Inhibition of endogenous miR-194, using anti-miR-194, resulted in enhanced chondrogenesis evidenced by up-regulation of chondrogenic markers [76].	Decrease endogenous miR-194 levels with a miR-194 inhibitor.
miR-221	miR-221 was reported to be up-regulated during JNK inhibitor-induced chondrogenic differentiation inhibition in chick limb bud mesenchymal cells [77].	miR-221 was demonstrated to directly target MDM2 [77].	Silencing of miR-211 in hMSCs resulted in the up-regulation of chondrogenic markers such as COL2A1 and SOX9 [78]. In an in vivo cartilage defect model, miR-221 silenced and alginate encapsulated hMSCs, generated cartilaginous tissue with enhanced cartilage repair [79••].	Decrease endogenous miR-221 levels with a miR-221 inhibitor.
miR-495	miR-495 was reported to be down-regulated during chondrogenic differentiation of hMSCs [80•].	miR-495 was demonstrated to directly target the 3’UTR of SOX9 [80•]. SOX9 is required for aggrecan [67], Col2a1 [68], Col9a1 [69] and Col11a2 expression [70] and binds to chondrocyte-specific enhancer elements in all of these genes. miR-495 is likely to be down-regulated during chondrogenesis enabling derepression of SOX9 expression.	Overexpression of miR-495 in hMSCs during chondrogenic differentiation, using miR-495 mimic, resulted in the down-regulation chondrogenic differentiation, evidenced by down-regulation of SOX9, COL2A1 and AGCAN mRNA. Inhibition of endogenous miR-495, using anti-miR-495, resulted in the enhancement of chondrogenic differentiation, evidenced by up-regulation of SOX9, COL2A1 and AGCAN mRNA [80•].	Decrease endogenous miR-495 levels with a miR-495 inhibitor.