In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration
Abstract Background Low back pain (LBP) is predominantly caused by degeneration of the intervertebral disc (IVD) and central nucleus pulposus (NP) region. Conservative treatments fail to restore disc function, motivating the exploration of nucleic acid therapies, such as the use of microRNAs (miRNAs...
Saved in:
| Main Authors: | , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Wiley
2024-12-01
|
| Series: | JOR Spine |
| Subjects: | |
| Online Access: | https://doi.org/10.1002/jsp2.1366 |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| _version_ | 1846107527098400768 |
|---|---|
| author | Tara Ní Néill Marcos N. Barcellona Niamh Wilson Fergal J. O'Brien James E. Dixon Caroline M. Curtin Conor T. Buckley |
| author_facet | Tara Ní Néill Marcos N. Barcellona Niamh Wilson Fergal J. O'Brien James E. Dixon Caroline M. Curtin Conor T. Buckley |
| author_sort | Tara Ní Néill |
| collection | DOAJ |
| description | Abstract Background Low back pain (LBP) is predominantly caused by degeneration of the intervertebral disc (IVD) and central nucleus pulposus (NP) region. Conservative treatments fail to restore disc function, motivating the exploration of nucleic acid therapies, such as the use of microRNAs (miRNAs). miRNAs have the potential to modulate expression of discogenic factors, while silencing the catabolic cascade associated with degeneration. To deliver these miRNAs, nonviral cell penetrating peptides (CPPs) are gaining favor given their low immunogenicity and strong targeting ability. Single miRNA therapies have been investigated for IVD repair, however dual miRNA delivery strategies have not been commonly examined and may augment regeneration. Materials and methods Transfection of four pro‐discogenic miRNAs (miRNA mimics:140‐5p; 149‐5p and inhibitors: 141‐3p; 221‐3p) and dual delivery of six miRNA pairings was performed using two CPPs, RALA and GET peptide (FLR), in primary rat NP monolayer culture, and in an ex vivo organ culture model of rat caudal discs. Protein expression of discogenic (aggrecan, collagen type II, and SOX9) and catabolic markers (ADAMTS5 and MMP13) were assessed. Results Monolayer investigations signified enhanced discogenic marker expression following dual miRNA delivery, signifying a synergistic effect when compared to single miRNA transfection. Utilization of an appropriate model was emphasized in our ex vivo organ culture experiment, revealing the establishment of a regenerative microenvironment characterized by reduced catabolic enzyme activity and enhanced matrix deposition, particularly following concurrent delivery of FLR‐miRNA‐149‐5p mimic and miRNA‐221‐3p inhibitor. Bioinformatics analysis of miRNA‐149‐5p mimic and miRNA‐221‐3p inhibitor identified distinct targets, pathways, and interactions, suggesting a mode of action for this amplified response. Conclusion Our findings suggest the potential of FLR‐miRNA‐149‐5p + miRNA‐221‐3p inhibitor to create an anti‐catabolic niche within the disc to foster regeneration in moderate cases of disc degeneration, which could be utilized in further studies with the overarching aim of developing treatments for LBP. |
| format | Article |
| id | doaj-art-d9e4e24821bf4c0fbb4262e3a0d23d35 |
| institution | Kabale University |
| issn | 2572-1143 |
| language | English |
| publishDate | 2024-12-01 |
| publisher | Wiley |
| record_format | Article |
| series | JOR Spine |
| spelling | doaj-art-d9e4e24821bf4c0fbb4262e3a0d23d352024-12-26T11:06:35ZengWileyJOR Spine2572-11432024-12-0174n/an/a10.1002/jsp2.1366In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regenerationTara Ní Néill0Marcos N. Barcellona1Niamh Wilson2Fergal J. O'Brien3James E. Dixon4Caroline M. Curtin5Conor T. Buckley6Trinity Centre for Biomedical Engineering Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of Dublin Dublin IrelandTrinity Centre for Biomedical Engineering Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of Dublin Dublin IrelandTrinity Centre for Biomedical Engineering Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of Dublin Dublin IrelandTrinity Centre for Biomedical Engineering Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of Dublin Dublin IrelandRegenerative Medicine and Cellular Therapies The University of Nottingham Biodiscovery Institute (BDI), School of Pharmacy, University of Nottingham Nottingham UKTrinity Centre for Biomedical Engineering Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of Dublin Dublin IrelandTrinity Centre for Biomedical Engineering Trinity Biomedical Sciences Institute, Trinity College Dublin, The University of Dublin Dublin IrelandAbstract Background Low back pain (LBP) is predominantly caused by degeneration of the intervertebral disc (IVD) and central nucleus pulposus (NP) region. Conservative treatments fail to restore disc function, motivating the exploration of nucleic acid therapies, such as the use of microRNAs (miRNAs). miRNAs have the potential to modulate expression of discogenic factors, while silencing the catabolic cascade associated with degeneration. To deliver these miRNAs, nonviral cell penetrating peptides (CPPs) are gaining favor given their low immunogenicity and strong targeting ability. Single miRNA therapies have been investigated for IVD repair, however dual miRNA delivery strategies have not been commonly examined and may augment regeneration. Materials and methods Transfection of four pro‐discogenic miRNAs (miRNA mimics:140‐5p; 149‐5p and inhibitors: 141‐3p; 221‐3p) and dual delivery of six miRNA pairings was performed using two CPPs, RALA and GET peptide (FLR), in primary rat NP monolayer culture, and in an ex vivo organ culture model of rat caudal discs. Protein expression of discogenic (aggrecan, collagen type II, and SOX9) and catabolic markers (ADAMTS5 and MMP13) were assessed. Results Monolayer investigations signified enhanced discogenic marker expression following dual miRNA delivery, signifying a synergistic effect when compared to single miRNA transfection. Utilization of an appropriate model was emphasized in our ex vivo organ culture experiment, revealing the establishment of a regenerative microenvironment characterized by reduced catabolic enzyme activity and enhanced matrix deposition, particularly following concurrent delivery of FLR‐miRNA‐149‐5p mimic and miRNA‐221‐3p inhibitor. Bioinformatics analysis of miRNA‐149‐5p mimic and miRNA‐221‐3p inhibitor identified distinct targets, pathways, and interactions, suggesting a mode of action for this amplified response. Conclusion Our findings suggest the potential of FLR‐miRNA‐149‐5p + miRNA‐221‐3p inhibitor to create an anti‐catabolic niche within the disc to foster regeneration in moderate cases of disc degeneration, which could be utilized in further studies with the overarching aim of developing treatments for LBP.https://doi.org/10.1002/jsp2.1366anti‐catabolismcell penetrating peptidescell therapiesmicroRNAregeneration |
| spellingShingle | Tara Ní Néill Marcos N. Barcellona Niamh Wilson Fergal J. O'Brien James E. Dixon Caroline M. Curtin Conor T. Buckley In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration JOR Spine anti‐catabolism cell penetrating peptides cell therapies microRNA regeneration |
| title | In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration |
| title_full | In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration |
| title_fullStr | In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration |
| title_full_unstemmed | In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration |
| title_short | In vitro and ex vivo screening of microRNA combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration |
| title_sort | in vitro and ex vivo screening of microrna combinations with enhanced cell penetrating peptides to stimulate intervertebral disc regeneration |
| topic | anti‐catabolism cell penetrating peptides cell therapies microRNA regeneration |
| url | https://doi.org/10.1002/jsp2.1366 |
| work_keys_str_mv | AT taranineill invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration AT marcosnbarcellona invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration AT niamhwilson invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration AT fergaljobrien invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration AT jamesedixon invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration AT carolinemcurtin invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration AT conortbuckley invitroandexvivoscreeningofmicrornacombinationswithenhancedcellpenetratingpeptidestostimulateintervertebraldiscregeneration |