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. 2021 Aug 13;18(6):1009–1020. doi: 10.1007/s13770-021-00378-5

Polydeoxyribonucleotide and Polynucleotide Improve Tendon Healing and Decrease Fatty Degeneration in a Rat Cuff Repair Model

Jung-Taek Hwang 1,✉, Sang-Soo Lee 1,#, Sang Hak Han 2,✉, Binod Sherchan 1, Jiss Joseph Panakkal 1
PMCID: PMC8599544  PMID: 34387852

Abstract

Background:

After surgical repair of chronic rotator cuff tears, healing of the repaired tendons often fails and is accompanied by high-level fatty degeneration. Our purpose was to explore the effects of polydeoxyribonucleotide (PDRN) and polynucleotide (PN) on tendon healing and the reversal of fatty degeneration in a chronic rotator cuff tear model using a rat infraspinatus.

Methods:

Sixty rats were randomly assigned to the following three groups (20 rats per group: 12 for histological evaluation and 8 for mechanical testing): saline + repair (SR), PDRN + repair (PR), and PN + repair (PNR). The right shoulder was used for experimental intervention, and the left served as a control. Four weeks after detaching the infraspinatus, the torn tendon was repaired. Saline, PDRN, and PN were applied to the repair sites. Histological evaluation was performed 3 and 6 weeks after repair and biomechanical analysis was performed at 6 weeks.

Results:

Three weeks after repair, the PR and PNR groups had more CD168-stained cells than the SR group. The PR group showed a larger cross-sectional area (CSA) of muscle fibers than the SR and PNR groups. Six weeks after repair, the PR and PNR groups showed more adipose cells, less CD68-stained cells, and more parallel tendon collagen fibers than the SR group. The PR group had more CD 68-stained cells than the PNR group. The PR group showed a larger CSA than the SR group. The mean load-to-failure values of the PR and PNR groups were higher than that of the SR group, although these differences were not significant.

CONCLUSION:

PDRN and PN may improve tendon healing and decrease fatty degeneration after cuff repair.

Supplementary Information

The online version contains supplementary material available at 10.1007/s13770-021-00378-5.

Keywords: Polydeoxyribonucleotide, Polynucleotide, Tendon healing, Fatty degeneration, Chronic rotator cuff tear, Rat model

Introduction

Rotator cuff tears are common causes of shoulder pain and dysfunction; their prevalence is approximately 50% in adults over the age of 70 years [1]. Although the techniques and instruments used for rotator cuff repair have markedly improved, recent studies found that about 50% of repaired cuffs do not adequately heal, regardless of the repair technique employed [2, 3].

After rotator cuff tears, there is often muscle fibers atrophy, fibrosis, and fat accumulation within and around the muscle fibers. This set of pathophysiological changes is often termed "fatty degeneration" [4–6], and is frequently aggravated in chronic rotator cuff tears in elderly patients. Preoperative fatty degeneration of rotator cuff muscles is an important prognostic factor to predict repair failure and poor functional outcomes [7, 8].

Several studies have sought to biologically enhance tendon-to-bone healing and reduce fatty degeneration, in efforts to lower the failure rate of rotator cuff repair [3, 6, 9, 10]. One study found that tendon-to-bone healing was enhanced by local administration of autologous platelet-rich plasma (PRP). Healing was assessed both histologically and biologically after repair of chronic rotator cuff tears in a rabbit model [9]. Another study suggested that local administration of adipose-derived stem cells (ADSCs) might improve muscle function and tendon healing and decrease fatty infiltration after repair of chronic rotator cuff tears in a rabbit model [3]. Conversely, some studies have reported that after arthroscopic rotator cuff repair, PRP treatment did not improve tendon healing or functional recovery [11–13].

Similarly, polydeoxyribonucleotide (PDRN) is a tissue regeneration activator that is composed of a mixture of nucleotides and activates adenosine A2A receptors, stimulating vascular endothelial growth factor (VEGF) expression and the activity of fibroblasts [14, 15]. This compound is extracted from the sperm of trout bred for consumption. The drug is obtained by a classified extraction process to purify the substance from proteins and ensures a very high percentage of polydeoxyribonucleotides. The chemical structure of PDRN consists of low-molecular weight DNA, and it is a linear polymer of deoxyribonucleotides with a chain length ranging between 50 and 2000 bp. It is likely that PDRN is cleaved by active cell membrane enzymes, providing a source of purine and pyrimidine deoxyribonucleosides and deoxyribonucleotides that can increase the proliferation and activity of cells in different tissues [16]. Previous studies have shown that PDRN improves wound healing especially in diabetic rat and mouse models [9, 14, 15]. Recently, PDRN has been used to aid in the regeneration of tendons or ligaments. There are several preclinical studies regarding rotator cuff tear and PDRN [17–19]. In these previous studies, the torn rotator cuffs were not repaired and PDRN was used combined with umbilical cord blood-derived mesenchymal stem cell (UCB-MSC) or microcurrent. In addition, polynucleotides (PNs) are composed of polynucleotides (20 mg/ml), polymeric molecules that are able to bond to a large amount of water to reorganize their structure by orienting and coordinating water molecules to form a 3-D gel. They undergo enzymatic cleavage and progressively release both water molecules and smaller oligonucleotides that retain moisturizing and viscoelastic properties, thereby maintaining the effect over a long time period [20, 21]. These properties of PN suggests that it has a similar effect as PDRN with a later onset and a longer duration.

The aim of this study was to determine the effects of PDRN and PN on tendon healing and the reversal of fatty degeneration in a chronic rotator cuff tear model using the rat infraspinatus muscle. We hypothesized that local administration of PDRN and PN in the subacromial space would enhance tendon-to-bone healing and mitigate fatty degeneration.

Materials and methods

This animal study was performed in accordance with the ethical standards in the 1964 Declaration of Helsinki and conducted in accordance with guidelines and approval of the Institutional Animal Care and Use Committees (IACUC) of the authors’ institution.

Allocation of experimental rats

For biomechanical testing, a priori power analysis revealed that a minimum of 8 rats per group was necessary to detect a significant difference in peak load to failure (mean difference: 10.0 N, standard deviation: 5.5 N) with a power of 0.8 and an alpha of 0.05, assuming a 25% drop-out rate [1, 3, 22]. For histological testing, we assigned 6 rats per group at 3 weeks and 6 weeks, based on previous studies [1, 2, 6]. Therefore, 60 male Sprague–Dawley rats (300 ± 5 g) were randomly allocated to three groups (20 rats per group: 12 for histological and 8 for mechanical testing): saline + repair (SR), PDRN + repair (PR), and PN + repair (PNR). All experimental procedures were performed in the right shoulders; the left shoulders underwent sham operations as a control (Fig. 1).

Fig. 1.

Fig. 1

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Flow diagram. PDRN: polydeoxyribonucleotide, PN: polynucleotide, Rt: right

Chronic rotator cuff tear model

Under intraperitoneal pentobarbital anesthesia (50 mg/kg) and sterile draping, a 2-cm skin incision was made along the spine of the scapula to the posterolateral angle of the acromion, and then the infraspinatus muscle and tendon were exposed after incising and retracting the deltoid muscle. In all rats, a chronic infraspinatus tear was created in the right shoulder by completely detaching the infraspinatus tendon at its insertion site into the greater tuberosity, wrapping the torn tendon with a 6-mm long silicone Penrose hose (6-mm outer diameter, Sewoon Medical Co., Ltd., Cheonan, Korea), and suturing them with No. 5–0 Prolene (Ethicon, Johnson & Johnson, New Brunswick, NJ, USA) to inhibit adhesion to the surrounding soft tissue, and it was left alone for 4 weeks [23, 24]. The deltoid muscle was sutured with a No. 5–0 Monocryl absorbable suture (Ethicon, Johnson & Johnson), and the skin was sutured with No. 5–0 Prolene. The left shoulder underwent a sham operation (skin incision and closure only).

Infraspinatus repair and injection of PDRN or PN

Four weeks after infraspinatus detachment, the torn tendon was repaired. A modified Mason-Allen stitch with a No. 5–0 Prolene (Ethicon) was used for repair in a transosseous manner by passing the suture through a bone tunnel created in the greater tuberosity using a 22-gauge needle. Immediately after the deltoid muscle was repaired using a No. 5–0 Monocryl absorbable suture (Ethicon), we injected 0.5 mL saline, PDRN, or PN into the subacromial space of the appropriate groups in a ballooning manner between the two sides of the sutured deltoid. Therefore, there was no loss of the injection material and the injection material reached the repair site. The skin was sutured using No. 5–0 Prolene (Ethicon).

Histological testing

Histological evaluation was performed 3 and 6 weeks after repair, and biomechanical analysis was performed at 6 weeks. Rats (6 per group for each period) were intraperitoneally anesthetized with pentobarbital and euthanized with carbon dioxide; then, the proximal humerus including the greater tuberosity with the entire attached infraspinatus tendon of both shoulders of each rat was harvested. The specimens were fixed in neutral buffered 10% formalin (pH 7.4) and decalcified for 24 h (Formical-2000, Decal Chemical Corporation, Tallman, NY, USA). Each specimen was embedded in paraffin, and two blocks were created. The specimen was horizontally cut at a point approximately 3 mm medial to the musculotendinous junction. The proximal portion was made as a paraffin block. The distal portion was cut longitudinally at a midpoint of the repair site (the infraspinatus tendon and the greater tuberosity). Two distal portions were prepared in one paraffin block. Then, 5 µm-thick sections were cut in the coronal plane from the tendon-to-bone junction; these were stained with hematoxylin and eosin (H&E) and Masson’s trichrome. We assessed collagen fiber continuity and parallel orientation in an average field of 100 × from the Masson’s trichrome-stained sections, and vascularity and cellularity in an average field of 200 × from the sections of H&E-stained sections. We graded each of these parameters semiquantitatively using 4 stages (present with < 25% proportion (grade 0), 25%–50% proportion (grade 1), 50%–75% proportion (grade 2), and > 75% proportion (grade 3)) [22]. We also cut 5-µm-thick sections in the sagittal plane to a point approximately 3 mm medial to the musculotendinous junction of the infraspinatus and stained the sections with H&E. We used theses sections to assess the extent of fatty degeneration of the infraspinatus muscle and calculated the cross-sectional area of muscle fibers. With an average field of 200 × from these sections, the histological findings were graded on a four-scale system (grades 0, 1, 2, and 3), where a grade 0 = no fat deposits and 3 = fat droplets found in most fibers [10]. The CSA of muscle fiber was calculated using Aperio ImageScope v12.1 (Leica Biosystems, Nussloch, Germany) [25, 26] (Fig. 2A). One hundred fibers in infraspinatus were selected in the average field of 400 × and the average cross-sectional area of the one hundred fibers was defined as the CSA [26].

Fig. 2.

Fig. 2

Fig. 2

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The measurement of CSA and the numbering of macrophage. A The measuring of CSA on H&E stain (× 400) of musculotendinous region using Aperio ImageScope v12.1 (Leica Biosystems, Nussloch, Germany). B The numbering of macrophages on CD68 stain (× 400) using an antigen counter (UTHSCSA ImageTool 3.0, The University of Texas Health Science Center at San Antonio, TX, USA). CSA: cross-sectional area of muscle fiber

In addition, CD68 and CD168 staining was performed by immunohistochemistry (IHC) in sagittal sections. CD68 staining was adopted to detect macrophages that suggested degeneration and CD168 staining was adopted to detect macrophages that suggested regeneration [6, 27, 28]. Sections of formalin-fixed, paraffin wax-embedded infraspinatus muscle tissue were stained in the Bond-Max automatic immunostaining device (Leica Biosystem, Newcastle, UK) using a bond polymer intensity detection kit (Leica Biosystem) for formalin-fixed, paraffin-embedded tissue sections. Antibodies against cluster of differentiation 68 (CD68; 47,850, Leica Biosystem, Newcastle, UK, RTU), and 168 (CD168; ab124729, Abcam, Cambridge, UK, RTU) were used. These sections were counterstained with Harris hematoxylin. From CD68 and CD168 immunohistochemical staining, the findings were verified each in an average field of 400 × as the number of stained macrophages using an antigen counter (UTHSCSA ImageTool 3.0, The University of Texas Health Science Center at San Antonio, TX, USA) (Fig. 2B). All of these analyses were performed by one pathologist who was blinded to the experimental conditions. In all grades and measurements, the pathologist selected an average field and performed the work in the field. The average of three measured values of the histological grading and measuring was used for quantitative analysis.

Mechanical testing

Rats (8 per group) were intraperitoneally anesthetized and euthanized with carbon dioxide, and the proximal humerus including the greater tuberosity with the entire attached infraspinatus tendon of both shoulders of each rat was harvested. The harvested tendon with the bone was wrapped in saline soaked gauze and placed in an icebox. Ten minutes before testing, each sample was placed at room temperature. Biomechanical testing was performed with the aid of an Instron 3343 device (Instron, Norwood, MA, USA) (Fig. 3A). The proximal humerus of the specimen was hung on a metal device (rat muscle fixation device), which bored a hole, having been previously fixed onto a pneumatic grip (ISG Inc., Sungnam, Korea). The muscular end of each sample was wrapped in a thin layer of dry gauze and fixed between the two hard rubber layers of a second pneumatic grip (Fig. 3B). Each tendon was initially preloaded to 0.1 N, followed by 10 cycles of preconditioning (cycling between 0.1 and 0.5 N at a strain rate of 0.4%/s). After a 300-s hold to attain equilibrium, each 600-s stress-relaxation experiment began with a ramp to 5% strain at 5%/s, followed by a return to gauge length and a 60-s hold. Finally, each specimen was quasi-statically tested to failure at a rate of 0.3%/s. The key data recorded included the mode of tearing and the peak load to failure [2].

Fig. 3.

Fig. 3

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The material testing machine. A Instron 3343 equipped with a pneumatic grip and a metal device for humeral head (rat muscle fixation device). B Tensile load is being applied to the infraspinatus of rat

Statistical analysis

One-way analysis of variance, followed by Bonferroni post hoc testing or Kruskal–Wallis analysis, followed by Mann–Whitney post hoc testing was used to analyze the values between the groups according to normality. The Mann–Whitney U test or independent t test was performed to compare the values between the operated side and the control side according to normality. Interclass correlation coefficients (ICCs) were used to assess intraobserver reliability for histological and immunohistochemical evaluation. The statistical analysis was performed using IBM SPSS Statistics 22 (IBM Corp., Armonk, NY, USA). A P < 0.05 was considered statistically significant.

Results

One specimen in the SR group, two in the PR group, and two in the PNR group showed a wound infection at the time of reattachment or harvest. Three rats from the SR, PR, and PNR groups died during the operation. One specimen form the SR group was excluded because an acromial fracture had occurred during the operation. Therefore, these 15 rats were excluded from the following assessment, and 45 rats were included in further analysis (3-week histological evaluation: n = 4, 4, 4; 6-week histological evaluation: n = 4, 4, 4; 6-week mechanical evaluation: n = 7, 7, 7 for SR, PR, PNR groups), as shown in Table 1.

Table 1.

Complications and number of rats included in each analysis

Complication SR group PR group PNR group
Wound infection 1 2 2
No recovery from anesthesia 3 3 3
Acromial fracture 1 0 0
3-week histological evaluation 4 4 4
6-week histological evaluation 4 4 4
6-week mechanical evaluation 7 7 7
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SR Saline + Repair, PR PDRN + Repair, PNR PN + Repair

Histological testing

Overall histological grades are reported in Fig. 4. In addition, IHC evaluations are shown Table 2. At 3 weeks, the tendon fibers at the tendon-to-bone region were poorly organized, and fiber continuity with bone had not yet been observed in the SR group. For IHC testing of the musculotendinous region, the largest mean number of CD68-stained cells were showed in the SR group and the smallest number in the PNR group are shown among the 3 groups, while the largest mean number of CD168-stained cells in the PR group and the smallest number in the SR group are shown. Six weeks after repair, tendon-to-bone integration was markedly improved with more collagen fibers and parallel orientation in all groups. The overall results of histological and IHC evaluation at 6 weeks were similar to those at 3 weeks with a greater improvement (Supplemental figure 1A–F).

Fig. 4.

Fig. 4

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Histologic Gradings. A–E At 3 weeks. F–J 6 weeks. 1: * H&E stain at musculotendinous region, a four scale system (grades 0, 1, 2, 3), where a grade 0 = no fat deposits and 3 = fat droplets found in most fibers. 2–5: § H&E and Masson’s Trichrome stains at tendon-to-bone junction, < 25% of proportion: Grade 0 (G0: absent or minimal), 25–50%: Grade 1 (G1: mild degree), 50–75%: Grade 2 (G2: moderate degree), > 75%: Grade 3 (G3: severe (marked) degree). SR: Saline + Repair, PR: PDRN + Repair, PNR: PN + Repair. Sham: left shoulder of SR groups at 3 weeks and 6 weeks

Table 2.

Immunohistochemistry (S100 and CD 68) and cross-sectional area at musculotendinous region

Groups (n) CD68 (n) CD168 (n) CSA(µm2)
3 weeks after repair
 Sham (4) 0.0 ± 0.0 0.0 ± 0.0 3825 ± 231
 SR (4) 18.3 ± 10.6 3.0 ± 0.8 2288 ± 226
 PR (4) 11.8 ± 7.3 7.3 ± 2.6 3110 ± 180
 PNR (4) 9.5 ± 6.0 6.3 ± 1.0 2566 ± 318
6 weeks after repair
 Sham (4) 0.0 ± 0.0 0.0 ± 0.0 3933 ± 218
 SR (4) 16.3 ± 5.4 4.0 ± 0.8 2603 ± 354
 PR (4) 9.3 ± 1.0 5.3 ± 1.5 3389 ± 192
 PNR (4) 7.0 ± 0.8 4.8 ± 2.1 2866 ± 361
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SR Saline + Repair, PR PDRN + Repair, PNR PN + Repair, CSA cross-sectional area of the muscle fiber

Statistical analysis revealed that there was no difference between the ratios of adipose cells of the sham and PR group on H&E staining of the musculotendinous region at 3 and 6 weeks, suggesting a greater improvement of fatty degeneration than in the SR and PR groups. Three weeks after repair, the PR and PNR groups had more CD168-stained cells than the SR group. The PR group showed a larger cross-sectional area (CSA) of muscle fibers than the SR and PNR groups. Six weeks after repair, the PR and PNR groups showed less adipose cells, less CD68-stained cells, and more parallel tendon collagen fibers than the SR group. PR group had more CD 68-stained cells than PNR group. PR groups showed a larger CSA than the SR group. Evidence of vascularization was noted by the presence of blood vessels, and cells were present in the irregularly arranged fibrovascular interface tissues. There is no statistically significant difference in vascularity and cellularity among the SR, PR, and PNR groups according to Kruskal–Wallis testing (Fig. 5).

Fig. 5.

Fig. 5

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Comparison of outcome in histologic results between the groups. SR: Saline + Repair, PR: PDRN + Repair, PNR: PN + Repair. Sham: left shoulder of SR groups at 3 weeks and 6 weeks, MT: musculotendinous region, TB: tendon-to-bone junction, V: vascularity, C: cellularity, Co: continuity of collagen fiber, Pa: parallel orientation of collagen fiber, CSA: cross-sectional area of muscle fiber. One-way analysis of variance or Kruskal–Wallis analysis was used to analyze the values among the SR, PR, and PNR groups. p < .05

The intraobserver reliability of histological grading for the the fat ratio on H&E staining was excellent (ICC of 0.94). The intraoberver reliability of histological grading for tendon healing on H&E staining and Masson’s Trichrome staining was excellent (ICC of 0.92). The intraobserver reliability of numbering for adipose cells for H&E staining and macrophages for the CD68 and CD168 staining was excellent (ICC of 0.91, 0.95 and 0.93). The intraobserver reliability of measuring for CSA of muscle fibers for H&E staining was also excellent (ICC of 0.93).

Mechanical testing

The mechanical data are shown in Table 3. The failure modes were four insertional and three midsubstance tears in the SR group, three insertional and four midsubstance tears in the PR group, and two insertional and five midsubstance tears in the PNR group. All control failures were midsubstance tears. Insertional tearing is suggestive of relatively poor tendon-to-bone healing while midsubstance tears indicate good healing. Midsubstance tearing was more common in the PR and PNR groups (57.1% and 71.4%) than in the SR group (42.9%). The load-to-failure values were higher in the PR and PNR groups (17.5 ± 7.9 N and 17.2 ± 8.7 N) than in the SR group (11.3 ± 5.8 N), although the differences did not reach statistical significance (one-way ANOVA; P = 0.245, post-hoc test: P = 0.418 and P = 0.476, respectively) (Tables 3 and 4). The load-to-failure values of the control (left side) specimens were greater than those of the test specimens, but the differences were not significant in either the PR or PNR group (p = 0.157 and p = 0.104, respectively), suggesting that tendon healing was good.

Table 3.

Outcome of mechanical testing

Outcome SR group (n = 7) PR group (n = 7) PNR group (n = 7)
Load to failure-operated side (N) 11.3 ± 5.8 17.5 ± 7.9 17.2 ± 8.7
Load to failure-control side (N) 21.5 ± 4.1 23.2 ± 6.0 24.6 ± 6.9
P value .002 .157 .104
Tear pattern Insertional: Midsubstance – operated side (n) 4: 3 3: 4 2: 5
Tear pattern Insertional: Midsubstance – control side (n) 0: 7 0: 7 0: 7
P value .023 .060 .141
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SR Saline + Repair, PR PDRN + Repair, PNR PN + Repair, P < .05

Table 4.

Comparison of outcome in mechanical testing between the treated right sides

P value Load to failure (N) Tear pattern
SR group Vs PR group .418 .606
SR group Vs PNR group .476 .298
PR group Vs PNR group 1.000 .591
One-way ANOVA or Kruskal–Wallis testing .245 .574
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SR Saline + Repair, PR PDRN + Repair, PNR PN + Repair, P < .05

Discussion

In this study, there was no difference between the ratios of adipose cells of the sham and PNR group on H&E staining of the musculotendinous region at 3 and 6 weeks, suggesting a greater improvement of fatty degeneration than in the SR and PR groups. Three weeks after repair, the PR and PNR groups had more CD168-stained cells than the SR group. The PR group showed a larger cross-sectional area (CSA) of muscle fibers than the SR and PNR group. Six weeks after repair, the PR and PNR groups showed more adipose cells, less CD68-stained cells, and more parallel tendon collagen fibers than the SR group. PR group had more CD 68-stained cells than PNR group. PR groups showed a larger CSA than the SR group. The overall results of histological and immunohistochemical evaluation at 6 weeks were similar to those at 3 weeks with a greater improvement. On biomechanical evaluation, the mean load-to-failure of the PR (17.5 ± 7.9 N) and the PNR (17.2 ± 8.7 N) groups were higher than that of the SR group (11.3 ± 5.8 N), although these differences were not significant (one-way ANOVA; p = 0.245, post-hoc test: P = 0.418 and P = 0.476, respectively). Considering these findings, local administration of PDRN and PN into the subacromial space in a ballooning manner may improve tendon healing and reduce fatty degeneration after rotator cuff repair. In addition, lower vascularity and cellularity suggest a reduced inflammation or foreign body reaction. But these parameters showed no significant difference among the three groups.

Despite many developments in surgical techniques and instrumentation, the failure rate after repair of chronic rotator cuff tears remains rather high [3, 29]. In addition, fatty degeneration in chronic rotator cuff tears is considered irreversible, even when cuff repair is successful. Earlier reports have shown that fatty degeneration persisted at long-term follow-up [30].

Various biological methods have been used to enhance tendon healing and reduce fatty degeneration, such as local administration of PRP, stem cells, and growth factors [3, 19, 31, 32]. Chung et al. [22] showed that tendon-to-bone healing was enhanced after local administration of autologous PRP, as assessed both histologically and biomechanically in a rabbit model of chronic rotator cuff tears. Vieira et al. [32] demonstrated that ADSCs enhanced tendon healing, as evidenced histologically, in a rabbit model of Achilles tendon rupture. Kaux et al. [31] found that a local injection of VEGF-111 improved the early healing phase of surgically sectioned Achilles tendons in rats. On the other hand, several clinical studies have shown that PRP does not aid in rotator cuff healing [11–13].

PDRN is a tissue regeneration activator that is composed of a mixture of nucleotides and activates adenosine receptors of fibroblasts to stimulate VEGF expression and fibroblast activity, in turn enhancing collagen fiber synthesis [9, 14, 15]. With the ligand-receptor type of mechanism of action, the effects of PDRN might last longer than those of PRP, stem cells, or growth factors. Moreover, the effects of PN can last even longer than those of PDRN because this biomaterial provides a suitable microenvironment in for injured regions through extracellular matrix formation. Additionally, PN is capable of scaffolding the created 3-D formation as a water-soluble nucleic acid with a nature of gel-type nature, and PN is physiologically present in the extracellular matrix, which constitutes fundamental nutrition for cells and tissues. Therefore, PDRN and PN may aid in tendon healing and reverse fatty degeneration, more effectively than PRP, stem cells, or growth factors.

Several studies have shown that PDRN and PN stimulate wound healing and tissue regeneration [9, 14, 15, 34, 35]. Altavilla et al. [14]. used an immunostaining method to study the effect of PDRN on wound healing in a diabetic mouse model. Chung et al. [9]. found that PDRN improved the survival of random pattern skin flaps in rats. Gennero et al. [36]. showed that PDRN inhibited cartilage degradation in an experimental culture. Guizzardi et al. [35]. reported that PN induced rapid bone regeneration in an experimental study on rats and that the combination of PN and deproteinated porcine cortical bone showed faster bone regeneration than PN alone or deproteinated porcine cortical bone alone. In addition, Kwon et al. [17–19]. performed three animal studies using rabbit cuff tear model. They showed PDRN combined with UCB-MSC or microcurrent was effective.

Our study has several limitations. First, we did not measure blood biological markers [growth factors, e. g., VEGF, fibroblast growth factor (FGF), or insulin-like growth factor (IGF)]. As PDRN and PN both stimulate growth factor production [9, 15, 35], such measurements would have been useful, and are required in future studies. Second, in humans, chronic rotator cuff tears develop slowly over a long period of time, and are influenced by various factors [16]. We repaired torn rat infraspinatus tendons only 4 weeks after surgical detachment. However, the time course of tendon injury and healing in rats may be 2- to threefold shorter than that in humans [37, 38]. Although the exact difference between rats and humans remains unknown, the week interval we employed was based on the data of previous studies [39]. The infraspinatus of rats was adopted as the specimen in this study because of its biomechanical similarity to that of humans [40, 41]. Third, H&E staining was used to detect adipose tissue in this study. Although oil red O can make fat more visible in fresh frozen tissue, its use is limited. Fourth, PDRN and PN were administered as single injections. Multiple injections might further enhance tendon healing and reverse fatty degeneration. The effects of multiple injections should be tested in a future study.

In conclusion, histological and biomechanical tests showed that PDRN and PN administered into the subacromial space improved tendon-to-bone healing and reduced fatty degeneration in a rat model of chronic infraspinatus tears. Thus, PDRN and PN may enhance rotator cuff healing in humans, which remains poor even after successful cuff repair. However, our results should be interpreted with caution in terms of clinical applications, as animal studies have intrinsic limitations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (ZIP 7112 kb) (6.9MB, zip)

Acknowledgements

We thank Jun Sub Jung, a research agent for valuable advices. Jung-Taek Hwang have received the National Research Foundation of Korea (NRF) Grant. This research was supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT (NRF-2016R1C1B2007014).

Compliance with ethical standards

Conflict of interest

The authors have no financial conflicts of interest.

Ethical statement

This animal study was conducted in accordance with guidelines and approval of the Institutional Animal Care and Use Committees (IACUC) of Hallym University (Hallym-2014–22). The present study had been performed in accordance with the ethical standards in the 1964 Declaration of Helsinki. The present study had been carried out in accordance with relevant regulations of the US Health Insurance Portability and Accountability Act (HIPAA). Details that might disclose the identity of the subjects under study should be omitted. This study was presented as a podium in 2016 AAOS and as an e-poster in 2019 ISAKOS.

Trial registration

Rat muscle fixation device (KR Design Registration 30–0854878).

Footnotes

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Jung-Taek Hwang and Sang-Soo Lee are co-first authors.

Change history

7/8/2023

A Correction to this paper has been published: 10.1007/s13770-023-00558-5

Contributor Information

Jung-Taek Hwang, Email: drakehjt@hanmail.net.

Sang Hak Han, Email: hue-2@hanmail.net.

References

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