This is the first double-blinded, randomized, placebo-controlled trial investigating the effects of intralesional MSC injection in patients with tendinopathy. Although no meaningful safety issues were noted during the trial and subsequent follow-up, MSC injection was not superior to the control treatment regarding improvement of pain and shoulder function in patients with partial thickness tears of the supraspinatus tendon. Changes in lesion size, as assessed by MRI, also did not differ between groups. There were also no differences in the occurrence of adverse events among groups.
Randomized, controlled clinical trials of stem cell transplantation for tendon defects
When the potential clinical applicability of stem cell treatment was introduced in 2004, many researchers and clinicians were skeptical about illusive promises, such as stem cell treatment for spinal cord injury or stroke, made by flamboyant pioneers of stem cell research. Because there seemed a long way with many difficult, if not impossible, challenges inducing transplanted stem cells to differentiate into functioning neuronal tissues in the damaged central nervous system. However, the positive results of anal fistula trial4 lead to an idea that, contrary to highly specialized central nervous tissues, connective tissues may be effectively treated with stem cells. The authors undertook several animal experiments13 and a subsequent phase I clinical trial14 to acquire positive results. The current phase II clinical trial was carried out with a strong expectation of revealing positive outcomes, extrapolating previous positive results.
In most previous clinical trials that have applied MSCs in tendon disorders, MSCs were implanted as an adjuvant treatment during surgical repair of the tendon. However, recent trials and systematic reviews have disputed the long-term clinical efficacy of repairing nontraumatic rotator cuff tears surgically.15,16 Thus, along with efforts to improve surgical outcomes in patients with tendinopathies using MSC, similar work should be performed for nonsurgical methods. Studies that have described the nonoperative use of MSCs in patients with tendinopathy8,14,17 are all single-arm design which is improper for studies assessing pain as an outcome measure.18 The current study was the first double-blinded, randomized, controlled clinical trial to investigate the effects of MSCs in tendinopathy.
Potential pitfalls in study design
Although pioneering work often fails, the negative results of the current study are disappointing considering the promising reports of previous single-arm studies. Successful works are based on trial and error, and we hope that the current study can serve as a foothold to the progress of regenerative medicine in tendinopathy by concisely reviewing the possible causes of the negative results. There were pitfalls in three domains of the study design in the current clinical trial: setting of the inclusion criteria, the control intervention, and estimating the sample size.
The inclusion criteria could not guarantee that the participants all had chronic intractable conditions. Although the mean duration of the symptoms and age in each group did not differ statistically, statistical insignificance cannot assure homogeneity of the participants when the sample size is small. All participants in the group injected with MSCs had a duration of symptoms longer than a year, whereas four patients in the active control group and one patient in the control group had symptoms for less than a year. The uneven distribution of participants with potential for spontaneous recovery may have balanced out the effects of MSCs. Similarly, the different age distributions in each group may have contributed to the lack of differences in the results. All but one patient was 60 years old or older in the MSC injection group, whereas only two patient in the control group and no patients in the active control group were of that age. Additionally, there were no minimum criteria for pain severity. A considerable portion of participants had only mild pain (VAS ≤ 4) at baseline, which left only a small margin for improvement and acted as a ceiling. (Supplementary Data)
In the context of inclusion criteria, we did not exclude volunteers with calcifications when it did not seem to cause any symptoms and many participants had one or multiple small calcifications in the treated tendon. Even a dormant calcification might interfere with the action of the injected MSCs by altering the microenvironment the MSC interacts with. Conversely, the MSC injection might have provoked the existing calcification to progress to a different phase on its natural course which would have ultimately affected the clinical course of the participants. Secondary analysis is warranted, focusing on the outcomes influenced by calcification.
The control groups did not have a purely sham intervention. In the active control group, fibrin glue was injected, and in both control groups, the tendon was pierced. Although fibrin glue has many advantages as a scaffold, it is naturally bioactive and can stimulate cell adhesion and growth.10 The escalated pain level at 3 days postinjection in both intervention and active control groups (Supplementary Data) is indirect proof of the bioactivity of fibrin glue. Penetrating the tendon is an unnatural stimulus to the tendon, clearly distinct from the causative stimulus of the tear in the supraspinatus tendon. Tendon fenestration itself can have therapeutic effects in chronic tendinopathy.19 Thus, the therapeutic effects of the control intervention could have diluted the effects of MSCs.
Target sample size is another limitation of the current study. The target sample size was estimated based on a clinical trial that compared platelet-rich plasma and sham injection in epicondylitis of the elbow.9 This study was the most appropriate reference in the existing literature at the time of conception of the current study because it was the only comparison study that concerned non-surgical application of regenerative medicine in chronic tendinopathy.20 We are unsure whether sample size estimation based on any previous study would be valid when there is no precedent on nonsurgical stem cell treatment for tendinopathy owing to the fundamental difference between stem cell therapy and other regenerative treatments.
Why not in tendinopathy?
Despite all the potential pitfalls discussed above, the most plausible explanation for the current results is that MSCs were not more effective than the control treatments, at least not enough to overcome the potential pitfalls discussed above. Thus, we should speculate as to why no positive results were obtained for patients with tendinopathy, despite positive findings in other disease conditions.
After report of the feasibility of expanding stem cells ex vivo and treating leukemia,21 stem cells have been applied in various clinical entities, including hematologic disorders, anal fistula in Crohn’s disease,4 and bone fractures,5 showed promising results in clinical trials. Blood cells are not attached to the extracellular matrix (ECM) and can function individually to contribute to the system. Thus, in hematologic disorders, stem cells avoid the problem of anoikis22 and only need to proliferate and differentiate into functional blood cells. In Crohn’s fistula, the goal of the transplanted MSCs is to produce areolar tissue that simply occupies space and therefore occludes the fistula without the necessity to functionally interact with the surrounding environment. However, in tendinopathy, the stem cells must interact with the ECM and differentiate into tenocytes, which produce and organize the ECM.
In regard of interaction between cells and the ECM, the bone is more similar to the tendon because both tissues function to transmit force via the ECM. However, the bone has a higher metabolic turnover rate and is more vascularized compared with the tendon, which makes the environment more amicable to introduced stem cells. Additionally, the bone transmits compressive force, which can be conveyed without tissue connection. Thus, in cases of fracture, stem cells can receive adequate physical stimulus once the fracture gap is filled with an appropriate scaffold. In contrast, the tendon transmits tensile force, and in tendinopathy, the fibrous ECM that undertakes the tendon’s function is disrupted. A linear fibrous environment23 and tensile loading24 are critical in the tenogenic differentiation of MSCs. The lack of both components in tendinopathy is a major impediment to establishing efficient methods for stem cell therapy in patients with tendinopathy. There should be substantial improvement in fundamental issues of stem cell therapy, as discussed below, to overcome these tendon-specific challenges.
Underlying issues of intralesional injection of stem cells in tendon tears
There are many factors that could be optimized to improve the outcomes of the current intervention. The material, i.e., stem cells, can be either augmented in potency or manipulated for tendon regeneration.25 The MSCs could be replaced with stem cells of higher hierarchy, such as embryonic stem cells, umbilical cord stem cells, or induced pluripotent stem cells with consideration that higher potency is accompanied by teratogenicity and tumorigenicity issues to be addressed. Alternatively, the stem cells can be harnessed towards tenogenic differentiation26 at the cost of somewhat decreased mitotic potential.
The microenvironment of the tendon that interferes the native regenerative process will also impede the effect of the administered MSCs. Preparing the lesion site congenial to the effects of MSC might be necessary. For instance, peppering technique to prompt some intrinsic healing processes19 or injecting the MSC with supplementary biological substances27 might augment the effect of MSC. Or with multiple MSC injections, separate injections may have priming or boosting effects. There can be limitless administrating methods by combining various routes, scaffolds, volumes, numbers, intervals and adjuvants of injections. Further investigations are required to establish the optimum method.
The scaffold would be of immense significance in the action of MSC. The primary usage of a scaffold is to retain the MSC within the lesion. There were a couple of cases where we could visualize the injectate leaking through a small fissure in the tendon which was not visible before the injection. More cases with such leaks could have occurred that were not captured in sonographic surveillance. It takes a few minutes for fibrinogen to interact with thrombin to form a clot and adhere to the surrounding tissue. Ideally, the scaffold should solidify shortly after administration and at the same time, be malleable enough to conform to the contour of the defect. Additionally, it should be permeable to cytokines and transmit tensile force that stimulate the mechanoreceptors of the MSC to steer it toward tenogenic differentiation.
Whether to mobilize or immobilize the shoulder after the injection could influence the effect of the intervention. In the current trial, participants were educated gentle range of motion exercise and instructed to perform the exercise within tolerable range three times a day. This mobilization would be disadvantageous for holding the injected MSC in site. On the other hand, adequate tensile loading is known to induce tenogenic differentiation of MSC and to refine the extracellular structure to resemble the tendon. Immobilization has detrimental effect on the tendon28 and clinical evidence also support functional rehabilitation rather than strict immobilization in tendon injury.29 Obliging non-painful mobilization after the intervention seems reasonable. However, the intervention is administrating exogenous cells and traditional concepts of mobilizing the tendon might not apply analogously.
Minor safety issues
All participants had pain at the injection site after the intervention, which should be related to the intervention. Transiently elevated pain can be regarded as an unavoidable cost of the intervention. A single-arm study that adopted a similar MSC injection without scaffolds in the rotator cuff tendon reported no such pain.8 This may be because the participants underwent arthroscopy, and pain after the intervention was considered natural. The possibility that fibrin glue may trigger a pain-generating process when injected in the shoulder is worth knowing for future investigators. All participants’ pain was manageable with conventional conservative treatment and did not recur during the 2-year follow-up period. Thus, we consider intratendinous injection of MSC to be safe and believe that pain should not be a constraint to future studies of the application of MSCs in tendons.
Our view point on current clinical implications and the future
The current results reporting lesser but nonsignificant clinical effects of MSC injection compared with the control treatment are worth noting. Literature suggesting the potential competence of MSCs in tissue regeneration is rapidly accumulating. Patients with chronic diseases are open to and easily influenced by such information, without proper interpretation. This may result in an inappropriate medical demand for stem cell therapy, and ill-considered applications of MSCs have already spread in clinical practice.30 Our current findings suggest that in nonsurgical treatment of tendinopathy, the use of MSCs should be confined to research purposes or rigorously selected clinical situations.
The vast discrepancy between preclinical studies that report promising results and insufficient clinical trials cannot be explained solely by the uniqueness of the human body. Clinical trials applying stem cells on tendon lesions has started as early as 2010, according the clinicaltrials.gov, and presumably there would have been more unregistered trials even before. However, the status of most previous trials are unknown, recruiting overdue, terminated or completed without subsequent reports. Not being able to share the details of an unsuccessful clinical trial causes insufficiency in growth of knowledge built up based on trial and error. As discussed above, there are many facets in stem cell therapy for tendinopathy, and literally infinite number of cases in treatment options remains to be investigated. Distribution of all clinical trials, although unsuccessful, followed by a vigorous networked discussion about the results is warranted to develop the optimum method of stem cell treatment for tendinopathy.