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Bone & Joint Research
Vol. 12, Issue 8 | Pages 486 - 493
4 Aug 2023
Yamanaka Y Tajima T Tsujimura Y Naito T Mano Y Tsukamoto M Zenke Y Sakai A

Aims. Dupuytren’s contracture is characterized by increased fibrosis of the palmar aponeurosis, with eventual replacement of the surrounding fatty tissue with palmar fascial fibromatosis. We hypothesized that adipocytokines produced by adipose tissue in contact with the palmar aponeurosis might promote fibrosis of the palmar aponeurosis. Methods. We compared the expression of the adipocytokines adiponectin and leptin in the adipose tissue surrounding the palmar aponeurosis of male patients with Dupuytren’s contracture, and of male patients with carpal tunnel syndrome (CTS) as the control group. We also examined the effects of adiponectin on fibrosis-related genes and proteins expressed by fibroblasts in the palmar aponeurosis of patients with Dupuytren’s contracture. Results. Adiponectin expression in the adipose tissue surrounding the palmar aponeurosis was significantly lower in patients with Dupuytren’s contracture than in those with CTS. The expression of fibrosis-related genes and proteins, such as types 1 and 3 collagen and α-smooth muscle actin, was suppressed in a concentration-dependent manner by adding AdipoRon, an adiponectin receptor agonist. The expression of fibrosis-related genes and proteins was also suppressed by AdipoRon in the in vitro model of Dupuytren’s contracture created by adding TGF-β to normal fibroblasts collected from patients with CTS. Conclusion. Fibrosis of the palmar aponeurosis in Dupuytren’s contracture in males may be associated with adiponectin expression in the adipose tissue surrounding the palmar aponeurosis. Although fibroblasts within the palmar aponeurosis are often the focus of attention when elucidating the pathogenesis of Dupuytren’s contracture, adiponectin expression in adipose tissues warrants closer attention in future research. Cite this article: Bone Joint Res 2023;12(8):486–493


Bone & Joint Research
Vol. 12, Issue 8 | Pages 455 - 466
1 Aug 2023
Zhou H Chen C Hu H Jiang B Yin Y Zhang K Shen M Wu S Wang Z

Aims. Rotator cuff muscle atrophy and fatty infiltration affect the clinical outcomes of rotator cuff tear patients. However, there is no effective treatment for fatty infiltration at this time. High-intensity interval training (HIIT) helps to activate beige adipose tissue. The goal of this study was to test the role of HIIT in improving muscle quality in a rotator cuff tear model via the β3 adrenergic receptor (β3AR). Methods. Three-month-old C57BL/6 J mice underwent a unilateral rotator cuff injury procedure. Mice were forced to run on a treadmill with the HIIT programme during the first to sixth weeks or seventh to 12th weeks after tendon tear surgery. To study the role of β3AR, SR59230A, a selective β3AR antagonist, was administered to mice ten minutes before each exercise through intraperitoneal injection. Supraspinatus muscle, interscapular brown fat, and inguinal subcutaneous white fat were harvested at the end of the 12th week after tendon tear and analyzed biomechanically, histologically, and biochemically. Results. Histological analysis of supraspinatus muscle showed that HIIT improved muscle atrophy, fatty infiltration, and contractile force compared to the no exercise group. In the HIIT groups, supraspinatus muscle, interscapular brown fat, and inguinal subcutaneous white fat showed increased expression of tyrosine hydroxylase and uncoupling protein 1, and upregulated the β3AR thermogenesis pathway. However, the effect of HIIT was not present in mice injected with SR59230A, suggesting that HIIT affected muscles via β3AR. Conclusion. HIIT improved supraspinatus muscle quality and function after rotator cuff tears by activating systemic sympathetic nerve fibre near adipocytes and β3AR. Cite this article: Bone Joint Res 2023;12(8):455–466


Bone & Joint Research
Vol. 10, Issue 10 | Pages 650 - 658
1 Oct 2021
Sanghani-Kerai A Black C Cheng SO Collins L Schneider N Blunn G Watson F Fitzpatrick N

Aims. This study investigates the effects of intra-articular injection of adipose-derived mesenchymal stem cells (AdMSCs) and platelet-rich plasma (PRP) on lameness, pain, and quality of life in osteoarthritic canine patients. Methods. With informed owner consent, adipose tissue collected from adult dogs diagnosed with degenerative joint disease was enzymatically digested and cultured to passage 1. A small portion of cells (n = 4) surplus to clinical need were characterized using flow cytometry and tri-lineage differentiation. The impact and degree of osteoarthritis (OA) was assessed using the Liverpool Osteoarthritis in Dogs (LOAD) score, Modified Canine Osteoarthritis Staging Tool (mCOAST), kinetic gait analysis, and diagnostic imaging. Overall, 28 joints (25 dogs) were injected with autologous AdMSCs and PRP. The patients were followed up at two, four, eight, 12, and 24 weeks. Data were analyzed using two related-samples Wilcoxon signed-rank or Mann-Whitney U tests with statistical significance set at p < 0.05. Results. AdMSCs demonstrated stem cell-like characteristics. LOAD scores were significantly lower at week 4 compared with preinjection (p = 0.021). The mCOAST improved significantly after three months (p = 0.001) and six months (p = 0.001). Asymmmetry indices decreased from four weeks post-injection and remained significantly lower at six months (p = 0.025). Conclusion. These improvements in quality of life, reduction in pain on examination, and improved symmetry in dogs injected with AdMSCs and PRP support the effectiveness of this combined treatment for symptom modification in canine OA for six months. Cite this article: Bone Joint Res 2021;10(10):650–658


Bone & Joint Research
Vol. 12, Issue 9 | Pages 580 - 589
20 Sep 2023
Dai X Liu B Hou Q Dai Q Wang D Xie B Sun Y Wang B

Aims. The aim of this study was to investigate the global and local impact of fat on bone in obesity by using the diet-induced obese (DIO) mouse model. Methods. In this study, we generated a diet-induced mouse model of obesity to conduct lipidomic and 3D imaging assessments of bone marrow fat, and evaluated the correlated bone adaptation indices and bone mechanical properties. Results. Our results indicated that bone mass was reduced and bone mechanical properties were impaired in DIO mice. Lipidomic sequencing and bioinformatic analysis identified 373 differential lipids, 176 of which were upregulated and 197 downregulated. Functional enrichment analysis revealed a significant downregulation of the pathways: fat digestion and absorption (ko04975) and lipolysis regulation in adipocytes (ko04923) in DIO mice, leading to local fat accumulation. The use of 3D imaging confirmed the increase in fat accumulation within the bone marrow cavity of obese mice. Conclusion. Our study sheds light on the intricate interplay between fat and bone, and provides a non-toxic and non-invasive method for measuring marrow adipose tissue. Cite this article: Bone Joint Res 2023;12(9):580–589


Bone & Joint Research
Vol. 11, Issue 2 | Pages 112 - 120
16 Feb 2022
Vittrup SØ Hanberg P Knudsen MB Tøstesen SK Kipp JO Hansen J Jørgensen NP Stilling M Bue M

Aims. Prompt and sufficient broad-spectrum empirical antibiotic treatment is key to preventing infection following open tibial fractures. Succeeding co-administration, we dynamically assessed the time for which vancomycin and meropenem concentrations were above relevant epidemiological cut-off (ECOFF) minimal inhibitory concentrations (T > MIC) in tibial compartments for the bacteria most frequently encountered in open fractures. Low and high MIC targets were applied: 1 and 4 µg/ml for vancomycin, and 0.125 and 2 µg/ml for meropenem. Methods. Eight pigs received a single dose of 1,000 mg vancomycin and 1,000 mg meropenem simultaneously over 100 minutes and 10 minutes, respectively. Microdialysis catheters were placed for sampling over eight hours in tibial cancellous bone, cortical bone, and adjacent subcutaneous adipose tissue. Venous blood samples were collected as references. Results. Across the targeted ECOFF values, vancomycin displayed longer T > MIC in all the investigated compartments in comparison to meropenem. For both drugs, cortical bone exhibited the shortest T > MIC. For the low MIC targets and across compartments, mean T > MIC ranged between 208 and 449 minutes (46% to 100%) for vancomycin and between 189 and 406 minutes (42% to 90%) for meropenem. For the high MIC targets, mean T > MIC ranged between 30 and 446 minutes (7% to 99%) for vancomycin and between 45 and 181 minutes (10% to 40%) for meropenem. Conclusion. The differences in the T > MIC between the low and high targets illustrate how the interpretation of these results is highly susceptible to the defined MIC target. To encompass any trauma, contamination, or individual tissue differences, a more aggressive dosing approach may be considered to achieve longer T > MIC in all the exposed tissues, and thereby lower the risk of acquiring an infection after open tibial fractures. Cite this article: Bone Joint Res 2022;11(2):112–120


Bone & Joint Research
Vol. 9, Issue 12 | Pages 857 - 869
1 Dec 2020
Slullitel PA Coutu D Buttaro MA Beaule PE Grammatopoulos G

As our understanding of hip function and disease improves, it is evident that the acetabular fossa has received little attention, despite it comprising over half of the acetabulum’s surface area and showing the first signs of degeneration. The fossa’s function is expected to be more than augmenting static stability with the ligamentum teres and being a templating landmark in arthroplasty. Indeed, the fossa, which is almost mature at 16 weeks of intrauterine development, plays a key role in hip development, enabling its nutrition through vascularization and synovial fluid, as well as the influx of chondrogenic stem/progenitor cells that build articular cartilage. The pulvinar, a fibrofatty tissue in the fossa, has the same developmental origin as the synovium and articular cartilage and is a biologically active area. Its unique anatomy allows for homogeneous distribution of the axial loads into the joint. It is composed of intra-articular adipose tissue (IAAT), which has adipocytes, fibroblasts, leucocytes, and abundant mast cells, which participate in the inflammatory cascade after an insult to the joint. Hence, the fossa and pulvinar should be considered in decision-making and surgical outcomes in hip preservation surgery, not only for their size, shape, and extent, but also for their biological capacity as a source of cytokines, immune cells, and chondrogenic stem cells. Cite this article: Bone Joint Res 2020;9(12):857–869


Bone & Joint Research
Vol. 8, Issue 7 | Pages 313 - 322
1 Jul 2019
Hanberg P Lund A Søballe K Bue M

Objectives. Meropenem may be an important drug in the treatment of open tibial fractures and chronic osteomyelitis. Therefore, the objective of this study was to describe meropenem pharmacokinetics in plasma, subcutaneous adipose tissue (SCT), and cancellous bone using microdialysis in a porcine model. Methods. Six female pigs were assigned to receive 1000 mg of meropenem intravenously over five minutes. Measurements of meropenem were obtained from plasma, SCT, and cancellous bone for eight hours thereafter. Microdialysis was applied for sampling in solid tissues. The meropenem concentrations were determined using ultra-high-performance liquid chromatography. Results. The penetration of meropenem into cancellous bone, expressed as the ratio of plasma to cancellous bone area under the concentration-curve from zero to the last measured value, was incomplete and delayed. The time with concentration above the minimal inhibitory concentration (T. >MIC. ), for an MIC of 0.5 μg/ml, was shorter for cancellous bone in comparison with both plasma and SCT. For MICs above 0.5 μg/ml, T. >MIC. in cancellous bone was only shorter than SCT. Considering an MIC of 4 μg/ml, no animals achieved the target of 40% T. >MIC. in plasma and cancellous bone, while less than 20% achieved it in SCT. Conclusion. The main finding of this study was short T. >MIC. in cancellous bone after intravenous administration of 1000 mg meropenem. Consequently, in order to achieve sufficient tissue concentration in the cases of open tibial fractures and chronic osteomyelitis, supplemental application of meropenem may be necessary. Cite this article: P. Hanberg, A. Lund, K. Søballe, M. Bue. Single-dose pharmacokinetics of meropenem in porcine cancellous bone determined by microdialysis: An animal study. Bone Joint Res 2019;8:342–348. DOI: 10.1302/2046-3758.87.BJR-2018-0308.R1


Bone & Joint Research
Vol. 8, Issue 2 | Pages 73 - 80
1 Feb 2019
Zhang J Hao X Yin M Xu T Guo F

Long non-coding RNAs (lncRNAs) are transcripts longer than 200 nucleotides with limited coding potential, which have emerged as novel regulators in many biological and pathological processes, including growth, development, and oncogenesis. Accumulating evidence suggests that lncRNAs have a special role in the osteogenic differentiation of various types of cell, including stem cells from different sources such as embryo, bone marrow, adipose tissue and periodontal ligaments, and induced pluripotent stem cells. Involved in complex mechanisms, lncRNAs regulate osteogenic markers and key regulators and pathways in osteogenic differentiation. In this review, we provide insights into the functions and molecular mechanisms of lncRNAs in osteogenesis and highlight their emerging roles and clinical value in regenerative medicine and osteogenesis-related diseases. Cite this article: J. Zhang, X. Hao, M. Yin, T. Xu, F. Guo. Long non-coding RNA in osteogenesis: A new world to be explored. Bone Joint Res 2019;8:73–80. DOI: 10.1302/2046-3758.82.BJR-2018-0074.R1


Bone & Joint Research
Vol. 13, Issue 7 | Pages 321 - 331
3 Jul 2024
Naito T Yamanaka Y Tokuda K Sato N Tajima T Tsukamoto M Suzuki H Kawasaki M Nakamura E Sakai A

Aims

The antidiabetic agent metformin inhibits fibrosis in various organs. This study aims to elucidate the effects of hyperglycaemia and metformin on knee joint capsule fibrosis in mice.

Methods

Eight-week-old wild-type (WT) and type 2 diabetic (db/db) mice were divided into four groups without or with metformin treatment (WT met(-/+), Db met(-/+)). Mice received daily intraperitoneal administration of metformin and were killed at 12 and 14 weeks of age. Fibrosis morphology and its related genes and proteins were evaluated. Fibroblasts were extracted from the capsules of 14-week-old mice, and the expression of fibrosis-related genes in response to glucose and metformin was evaluated in vitro.


Bone & Joint Research
Vol. 12, Issue 12 | Pages 722 - 733
6 Dec 2023
Fu T Chen W Wang Y Chang C Lin T Wong C

Aims

Several artificial bone grafts have been developed but fail to achieve anticipated osteogenesis due to their insufficient neovascularization capacity and periosteum support. This study aimed to develop a vascularized bone-periosteum construct (VBPC) to provide better angiogenesis and osteogenesis for bone regeneration.

Methods

A total of 24 male New Zealand white rabbits were divided into four groups according to the experimental materials. Allogenic adipose-derived mesenchymal stem cells (AMSCs) were cultured and seeded evenly in the collagen/chitosan sheet to form cell sheet as periosteum. Simultaneously, allogenic AMSCs were seeded onto alginate beads and were cultured to differentiate to endothelial-like cells to form vascularized bone construct (VBC). The cell sheet was wrapped onto VBC to create a vascularized bone-periosteum construct (VBPC). Four different experimental materials – acellular construct, VBC, non-vascularized bone-periosteum construct, and VBPC – were then implanted in bilateral L4-L5 intertransverse space. At 12 weeks post-surgery, the bone-forming capacities were determined by CT, biomechanical testing, histology, and immunohistochemistry staining analyses.


Bone & Joint Research
Vol. 11, Issue 12 | Pages 854 - 861
1 Dec 2022
Park TJ Park SY Cho W Oh H Lee HJ Abd El-Aty AM Bayram C Jeong JH Jung TW

Aims

Myokine developmental endothelial locus-1 (DEL-1) has been documented to alleviate inflammation and endoplasmic reticulum (ER) stress in various cell types. However, the effects of DEL-1 on inflammation, ER stress, and apoptosis in tenocytes remain unclear.

Methods

Human primary tenocytes were cultured in palmitate (400 μM) and palmitate plus DEL-1 (0 to 2 μg/ml) conditions for 24 hours. The expression levels of ER stress markers and cleaved caspase 3, as well as phosphorylated 5' adenosine monophosphate-activated protein kinase (AMPK) and autophagy markers, were assessed by Western blotting. Autophagosome formation was measured by staining with monodansylcadaverine, and apoptosis was determined by cell viability assay and caspase 3 activity assay.


Bone & Joint Research
Vol. 11, Issue 10 | Pages 715 - 722
10 Oct 2022
Matsuyama Y Nakamura T Yoshida K Hagi T Iino T Asanuma K Sudo A

Aims

Acridine orange (AO) demonstrates several biological activities. When exposed to low doses of X-ray radiation, AO increases the production of reactive radicals (radiodynamic therapy (AO-RDT)). We elucidated the efficacy of AO-RDT in breast and prostate cancer cell lines, which are likely to develop bone metastases.

Methods

We used the mouse osteosarcoma cell line LM8, the human breast cancer cell line MDA-MB-231, and the human prostate cancer cell line PC-3. Cultured cells were exposed to AO and radiation at various concentrations followed by various doses of irradiation. The cell viability was then measured. In vivo, each cell was inoculated subcutaneously into the backs of mice. In the AO-RDT group, AO (1.0 μg) was locally administered subcutaneously around the tumour followed by 5 Gy of irradiation. In the radiation group, 5 Gy of irradiation alone was administered after macroscopic tumour formation. The mice were killed on the 14th day after treatment. The change in tumour volume by AO-RDT was primarily evaluated.


Aims

This study examined whether systemic administration of melatonin would have different effects on osseointegration in ovariectomized (OVX) rats, depending on whether this was administered during the day or night.

Methods

In this study, a titanium rod was implanted in the medullary cavity of one femoral metaphysis in OVX rats, and then the rats were randomly divided into four groups: Sham group (Sham, n = 10), OVX rat group (OVX, n = 10), melatonin day treatment group (OVX + MD, n = 10), and melatonin night treatment group (OVX + MN, n = 10). The OVX + MD and OVX + MN rats were treated with 30 mg/kg/day melatonin at 9 am and 9 pm, respectively, for 12 weeks. At the end of the research, the rats were killed to obtain bilateral femora and blood samples for evaluation.


Bone & Joint Research
Vol. 13, Issue 4 | Pages 137 - 148
1 Apr 2024
Lu Y Ho T Huang C Yeh S Chen S Tsao Y

Aims

Pigment epithelium-derived factor (PEDF) is known to induce several types of tissue regeneration by activating tissue-specific stem cells. Here, we investigated the therapeutic potential of PEDF 29-mer peptide in the damaged articular cartilage (AC) in rat osteoarthritis (OA).

Methods

Mesenchymal stem/stromal cells (MSCs) were isolated from rat bone marrow (BM) and used to evaluate the impact of 29-mer on chondrogenic differentiation of BM-MSCs in culture. Knee OA was induced in rats by a single intra-articular injection of monosodium iodoacetate (MIA) in the right knees (set to day 0). The 29-mer dissolved in 5% hyaluronic acid (HA) was intra-articularly injected into right knees at day 8 and 12 after MIA injection. Subsequently, the therapeutic effect of the 29-mer/HA on OA was evaluated by the Osteoarthritis Research Society International (OARSI) histopathological scoring system and changes in hind paw weight distribution, respectively. The regeneration of chondrocytes in damaged AC was detected by dual-immunostaining of 5-bromo-2'-deoxyuridine (BrdU) and chondrogenic markers.


Bone & Joint Research
Vol. 12, Issue 9 | Pages 601 - 614
21 Sep 2023
Gu P Pu B Liu T Yue D Xin Q Li H Yang B Ke D Zheng X Zeng Z Zhang Z

Aims

Mendelian randomization (MR) is considered to overcome the bias of observational studies, but there is no current meta-analysis of MR studies on rheumatoid arthritis (RA). The purpose of this study was to summarize the relationship between potential pathogenic factors and RA risk based on existing MR studies.

Methods

PubMed, Web of Science, and Embase were searched for MR studies on influencing factors in relation to RA up to October 2022. Meta-analyses of MR studies assessing correlations between various potential pathogenic factors and RA were conducted. Random-effect and fixed-effect models were used to synthesize the odds ratios of various pathogenic factors and RA. The quality of the study was assessed using the Strengthening the Reporting of Observational Studies in Epidemiology using Mendelian Randomization (STROBE-MR) guidelines.


Bone & Joint Research
Vol. 13, Issue 4 | Pages 169 - 183
15 Apr 2024
Gil-Melgosa L Llombart-Blanco R Extramiana L Lacave I Abizanda G Miranda E Agirre X Prósper F Pineda-Lucena A Pons-Villanueva J Pérez-Ruiz A

Aims

Rotator cuff (RC) injuries are characterized by tendon rupture, muscle atrophy, retraction, and fatty infiltration, which increase injury severity and jeopardize adequate tendon repair. Epigenetic drugs, such as histone deacetylase inhibitors (HDACis), possess the capacity to redefine the molecular signature of cells, and they may have the potential to inhibit the transformation of the fibro-adipogenic progenitors (FAPs) within the skeletal muscle into adipocyte-like cells, concurrently enhancing the myogenic potential of the satellite cells.

Methods

HDACis were added to FAPs and satellite cell cultures isolated from mice. The HDACi vorinostat was additionally administered into a RC injury animal model. Histological analysis was carried out on the isolated supra- and infraspinatus muscles to assess vorinostat anti-muscle degeneration potential.


Bone & Joint Research
Vol. 11, Issue 9 | Pages 652 - 668
7 Sep 2022
Lv G Wang B Li L Li Y Li X He H Kuang L

Aims

Exosomes (exo) are involved in the progression of osteoarthritis (OA). This study aimed to investigate the function of dysfunctional chondrocyte-derived exo (DC-exo) on OA in rats and rat macrophages.

Methods

Rat-derived chondrocytes were isolated, and DCs induced with interleukin (IL)-1β were used for exo isolation. Rats with OA (n = 36) or macrophages were treated with DC-exo or phosphate-buffered saline (PBS). Macrophage polarization and autophagy, and degradation and chondrocyte activity of cartilage tissues, were examined. RNA sequencing was used to detect genes differentially expressed in DC-exo, followed by RNA pull-down and ribonucleoprotein immunoprecipitation (RIP). Long non-coding RNA osteoarthritis non-coding transcript (OANCT) and phosphoinositide-3-kinase regulatory subunit 5 (PIK3R5) were depleted in DC-exo-treated macrophages and OA rats, in order to observe macrophage polarization and cartilage degradation. The PI3K/AKT/mammalian target of rapamycin (mTOR) pathway activity in cells and tissues was measured using western blot.


Bone & Joint Research
Vol. 11, Issue 7 | Pages 453 - 464
20 Jul 2022
Wang H Shi Y He F Ye T Yu S Miao H Liu Q Zhang M

Aims

Abnormal lipid metabolism is involved in the development of osteoarthritis (OA). Growth differentiation factor 11 (GDF11) is crucial in inhibiting the differentiation of bone marrow mesenchymal stem cells into adipocytes. However, whether GDF11 participates in the abnormal adipogenesis of chondrocytes in OA cartilage is still unclear.

Methods

Six-week-old female mice were subjected to unilateral anterior crossbite (UAC) to induce OA in the temporomandibular joint (TMJ). Histochemical staining, immunohistochemical staining (IHC), and quantitative real-time polymerase chain reaction (qRT-PCR) were performed. Primary condylar chondrocytes of rats were stimulated with fluid flow shear stress (FFSS) and collected for oil red staining, immunofluorescence staining, qRT-PCR, and immunoprecipitation analysis.


Bone & Joint Research
Vol. 9, Issue 10 | Pages 719 - 728
1 Oct 2020
Wang J Zhou L Zhang Y Huang L Shi Q

Aims

The purpose of our study was to determine whether mesenchymal stem cells (MSCs) are an effective and safe therapeutic agent for the treatment of knee osteoarthritis (OA), owing to their cartilage regeneration potential.

Methods

We searched PubMed, Embase, and the Cochrane Library, with keywords including “knee osteoarthritis” and “mesenchymal stem cells”, up to June 2019. We selected randomized controlled trials (RCTs) that explored the use of MSCs to treat knee OA. The visual analogue scale (VAS), Western Ontario and McMaster University Osteoarthritis Index (WOMAC), adverse events, and the whole-organ MRI score (WORMS) were used as the primary evaluation tools in the studies. Our meta-analysis included a subgroup analysis of cell dose and cell source.


Bone & Joint Research
Vol. 10, Issue 10 | Pages 668 - 676
1 Oct 2021
Liu L Li Z Chen S Cui H Li X Dai G Zhong F Hao W Zhang K Liu H

Aims

Acquired heterotopic ossification (HO) is a debilitating disease characterized by abnormal extraskeletal bone formation within soft-tissues after injury. The exact pathogenesis of HO remains unknown. It was reported that BRD4 may contribute to osteoblastic differentiation. The current study aims to determine the role of BRD4 in the pathogenesis of HO and whether it could be a potential target for HO therapy.

Methods

Achilles tendon puncture (ATP) mouse model was performed on ten-week-old male C57BL/6J mice. One week after ATP procedure, the mice were given different treatments (e.g. JQ1, shMancr). Achilles tendon samples were collected five weeks after treatment for RNA-seq and real-time quantitative polymerase chain reaction (RT-qPCR) analysis; the legs were removed for micro-CT imaging and subsequent histology. Human bone marrow mesenchymal stem cells (hBMSCs) were isolated and purified bone marrow collected during surgeries by using density gradient centrifugation. After a series of interventions such as knockdown or overexpressing BRD4, Alizarin red staining, RT-qPCR, and Western Blot (Runx2, alkaline phosphatase (ALP), Osx) were performed on hBMSCs.