Role of Oxidative Stress in Peyronie’s Disease: Biochemical Evidence and Experiences of Treatment with Antioxidants

Background: Peyronie’s disease (PD) is a chronic inflammatory condition affecting adult males, involving the tunica albuginea of the corpora cavernosa of the penis. PD is frequently associated with penile pain, erectile dysfunction, and a secondary anxious–depressive state. The etiology of PD has not yet been completely elucidated, but local injury is generally recognized to be a triggering factor. It has also been widely proven that oxidative stress is an essential, decisive component in all inflammatory processes, whether acute or chronic. Current conservative medical treatment comprises oral substances, penile injections, and physical therapy. Aim: This article intends to show how antioxidant therapy is able to interfere with the pathogenetic mechanisms of the disease. Method: This article consists of a synthetic narrative review of the current scientific literature on antioxidant therapy for this disease. Results: The good results of the antioxidant treatment described above also prove that the doses used were adequate and the concentrations of the substances employed did not exceed the threshold at which they might have interacted negatively with the mechanisms of the redox regulation of tissue. Conclusions: We believe new, randomized, controlled studies are needed to confirm the efficacy of treatment with antioxidants. However, we consider the experiences of antioxidant treatment which can already be found in the literature useful for the clinical practice of urologists in the treatment of this chronic inflammatory disease.

The disease affects the tunica albuginea of the corpora cavernosa of the penis, and in its natural evolution can also involve the tissue of the corpora cavernosa and the intercavernous septum. The disease's hallmark is an excessive production of collagen, which causes localized fibrosis and results in penile deformation. Most of the time, the clinical consequence is a penile bend that can be more or less pronounced, and is sometimes associated with penile shortening, penile torsion of various degrees, divots, hourglass deformity, etc. The disease is frequently associated with penile pain, erectile dysfunction, and a secondary anxious-depressive state. It has been found that PD patients are more likely to have psychiatric disorders (anxiety disorder, depression, substance abuse, alcohol abuse, self-injurious behavior, etc.) than men without PD [14,15]. It is therefore possible that psychological distress can negatively affect the correct daily intake of drugs and facilitate Transforming growth factor beta-1 TGF-beta-1 Chemotactic action on neutrophils, monocytes, lymphocytes, and fibroblasts. It induces the production of collagen by fibroblasts. It stimulates the proliferation of fibroblasts and the transformation of fibroblasts into myofibroblasts. It induces collagen synthesis and deposition. It induces the production of ROS. It increases the synthesis of tissue inhibitors of matrix metalloproteinase (TIMP-1). It inhibits the production of matrix metalloproteinases (MMPs) with collagenase action (MMP-1, MMP-8, and MMP-13). It induces the production of MMP-2 and MMP-9 (with elastase activity). It induces the activation of NF-kB. It induces osteogenesis in PD plaque. It inhibits production of plasminogen activator inhibitor-1 (PAI-1).

The First Studies on the Use of Antioxidants in Patients with PD
The use of antioxidants in the treatment of PD dates back over 70 years: specifically, vitamin E was the first antioxidant to be successfully used [89]. It was not chosen for its antioxidant properties, however, but merely because it had already been used with good results in other fibrotic disorders, such as Dupuytren's contracture and primary fibrositis. At the time, it was postulated that fibrotic diseases, including PD, were due to a metabolic disorder caused by vitamin E deficiency [89].
Many years later, in recent years of the second millennium, propolis began to be used-likewise, not for its known antioxidant properties, but simply because a patient with PD, who had been treated with propolis (hospital in Havana, Cuba) by gastroenter-

The First Studies on the Use of Antioxidants in Patients with PD
The use of antioxidants in the treatment of PD dates back over 70 years: specifically, vitamin E was the first antioxidant to be successfully used [89]. It was not chosen for its antioxidant properties, however, but merely because it had already been used with good results in other fibrotic disorders, such as Dupuytren's contracture and primary fibrositis.
At the time, it was postulated that fibrotic diseases, including PD, were due to a metabolic disorder caused by vitamin E deficiency [89].
Many years later, in recent years of the second millennium, propolis began to be used-likewise, not for its known antioxidant properties, but simply because a patient with PD, who had been treated with propolis (hospital in Havana, Cuba) by gastroenterologists because he was affected by Giardiasis, had noticed a significant improvement in his penile curvature. The urologists of the same hospital where the patient was treated, having observed the clinical evidence, began to treat PD patients with propolis (in oil form: propoleum). All the studies published by these Cuban urologists, which include both the exclusive use of propolis and treatment in association with laser therapy, showed significant improvement in terms of both plaque volume reduction and improvement in penile curvature [90][91][92][93].
Subsequently, in the early 2000s, carnitine was used to treat PD, specifically for its antioxidant properties, both alone (orally) and together with penile injections of a calcium channel blocker (Verapamil) [94,95]. Both studies achieved good results.
A number of studies then followed in which pentoxifylline was successfully used to treat PD [29,[96][97][98].
In other studies, a number of antioxidants (vitamin B3, propionyl-L-carnitine, L-arginine) were used in association with other substances (verapamil, tadalafil) or penile traction [46,99,100]. All of these studies achieved excellent results. Furthermore, these studies introduced the concept of "combined" or "multimodal" antioxidant treatment to the literature.

Multimodal Treatment with Antioxidants
Multimodal or combined treatment is a therapeutic practice which has already been used in other fields of medicine, such as oncology (polychemotherapy) and the treatment of infections (antibiotic combination therapy), for instance, in the treatment of tuberculosis or even simply in the treatment of drug-resistant bacteria, e.g., for amoxicillin-resistant germs, through the association of amoxicillin with clavulanic acid. The aim is to obtain a better therapeutic result than is achievable by administering a single substance.
In our case, this is particularly advantageous, since many antioxidants also have antiinflammatory and antifibrotic properties that differ slightly (see Table 2A,B), and therefore combining them makes it possible to reduce the dose of each antioxidant (and minimize the possibility of adverse effects from an overdose) and to tackle PD and its various biochemical mechanisms, interfering in different ways with the many "chemical messengers" involved.
For instance, bilberry is capable of inhibiting the production of bFGF, unlike other antioxidant substances. Boswellia, superoxide dismutase and diclofenac cannot reduce the synthesis of PAI-1 as other substances do. Pentoxifylline, silymarin, boswellia, coenzyme Q10, carnitine, and Ginkgo biloba, unlike other antioxidant substances, are able to contrast the vasoconstriction caused by high concentrations of nitroxide radicals (NO•-), and thanks to their properties, they can cause vasodilation at the disease site [101,102], thus preventing hypoxia and tissue damage and contrasting the outcomes of PD.
Thus, it appears that multimodal antioxidant therapy can be used as a therapeutic strategy to maximize the end effects of treatment.

Brief Narrative Review of Peyronie's Disease Treatment with Antioxidants
Although a number of guidelines contain no recommendation for the use of antioxidants in the treatment of PD, they have been used in several therapeutic experiences, either alone or in combination with other substances [168,169].
We searched the PubMed database for articles on the topic "antioxidant treatment in Peyronie's disease" and found 21 articles on this topic. We considered the following articles eligible: randomized and/or controlled clinical trials; case reports containing more than one case report. After screening the 21 articles, we excluded 4 of them: 2 articles because they reported a single clinical case, and 2 clinical studies because they did not have a control group.
We shall, of course, cite only a selection of the scientific literature on the topic, including randomized studies and a number of controlled studies, as well as three case report studies in which the complete regression of plaque was achieved [29,30,98,[170][171][172][173][174][175][176]. Other minor studies cited in the References section of this article are not described here for reasons of space, despite the fact that most of them are controlled trials and despite their positive results [90][91][92][93][94][95][96][97][177][178][179] (Figure 2).
We shall, of course, cite only a selection of the scientific literature on the topic, including randomized studies and a number of controlled studies, as well as three case report studies in which the complete regression of plaque was achieved [29,30,98,[170][171][172][173][174][175][176]. Other minor studies cited in the References section of this article are not described here for reasons of space, despite the fact that most of them are controlled trials and despite their positive results [90][91][92][93][94][95][96][97][177][178][179] (Figure 2). A randomized, double-blind, placebo-controlled study by Riedl (2005) used Superoxide dismutase to treat PD patients. Superoxide dismutase was used as topical gel once a day for eight weeks, reaching the following statistically significant results: pain reduction in 52.6% of treated cases, compared to 20% in the control group; plaque volume reduction in 47% of treated cases, compared to the control group where, on the contrary, plaque volume grew in 8% of cases; reduction in the degree of penile curvature (between 5 and 30 degrees) in 23% of treated cases, compared to the control group in which, instead, curvature increased in 10% of cases [170].
In a randomized controlled study by Favilla et al. (2014), the authors treated PD patients for 12 weeks with oral antioxidants (vitamin E, para-aminobenzoic acid, propolis, blueberry anthocyanins, soy isoflavones, muira puama, damiana, persea americana) associated with weekly verapamil injections. The control group only received weekly verapamil injections [171]. After treatment, there were no statistically significant differences for some endpoints (e.g., reduction in plaque size = −29% (group receiving oral antioxidants + verapamil injections) and −38% (group receiving only verapamil injections); improvement in penile curvature = −11.9 degrees (group receiving oral antioxidants + verapamil injections) and −10.8 degrees (group receiving only verapamil injections)).
Regarding the other endpoints, in the group receiving oral antioxidants + verapamil injections, there were statistically significantly better results compared to the control group (only verapamil injections) in terms of: penile pain reduction, orgasmic function, intercourse satisfaction, and overall satisfaction. In conclusion, the study showed that combined therapy made it possible to obtain better results [171]. Full-text articles assessed for eligibility n = 17 Studies included in qualitative synthesis n = 9 Full-text articles excluded for reasons of space, although they are randomized and/or controlled trials = 8 Records excluded with reasons of : case report of a single case = 2, study without control group = 2 A randomized, double-blind, placebo-controlled study by Riedl (2005) used Superoxide dismutase to treat PD patients. Superoxide dismutase was used as topical gel once a day for eight weeks, reaching the following statistically significant results: pain reduction in 52.6% of treated cases, compared to 20% in the control group; plaque volume reduction in 47% of treated cases, compared to the control group where, on the contrary, plaque volume grew in 8% of cases; reduction in the degree of penile curvature (between 5 and 30 degrees) in 23% of treated cases, compared to the control group in which, instead, curvature increased in 10% of cases [170].
In a randomized controlled study by Favilla et al. (2014), the authors treated PD patients for 12 weeks with oral antioxidants (vitamin E, para-aminobenzoic acid, propolis, blueberry anthocyanins, soy isoflavones, muira puama, damiana, persea americana) associated with weekly verapamil injections. The control group only received weekly verapamil injections [171]. After treatment, there were no statistically significant differences for some endpoints (e.g., reduction in plaque size = −29% (group receiving oral antioxidants + verapamil injections) and −38% (group receiving only verapamil injections); improvement in penile curvature = −11.9 degrees (group receiving oral antioxidants + verapamil injections) and −10.8 degrees (group receiving only verapamil injections)).
Regarding the other endpoints, in the group receiving oral antioxidants + verapamil injections, there were statistically significantly better results compared to the control group (only verapamil injections) in terms of: penile pain reduction, orgasmic function, intercourse satisfaction, and overall satisfaction. In conclusion, the study showed that combined therapy made it possible to obtain better results [171].
In a randomized controlled study by Alizadeh et al. (2014), the authors treated patients with PD for six months, dividing them into three treatment groups:
In their retrospective control group study, Gallo et al. (2019) treated PD patients for 6 months, dividing them into three groups: first group: oral therapy with arginine and pentoxifylline; second group: oral therapy with arginine and pentoxifylline + intralesional injections with verapamil (every other week, 12 total injections); third group: oral therapy with arginine and pentoxifylline + intralesional injections with verapamil (every other week, 12 total injections) + penile traction therapy with a penile extender, applied daily for about 2-8 h [172]. The therapeutic results for the various endpoints, compared with the baseline data, were as follows: Penile pain: first group = resolution of pain in 100% of cases; second group = resolution of pain in 100% of cases; third group = resolution of pain in 100% of cases [172].
The authors conclude that oral therapy alone can simply block disease progression, association between oral therapy and verapamil injections enables only slight improvement, and the combination of oral therapy, verapamil injections, and penile traction therapy is the only conservative approach leading to optimal results.
In our 2016 retrospective control group study (Paulis et al., 2016), we treated 206 patients with PD using the following combined therapy: group A: 112 patients, pentoxifylline (perilesional injections) twice a month for 6 months + oral pentoxifylline + oral propolis + oral blueberry + oral vitamin E + diclofenac gel for 6 months); group B: 94 patients undertaking the same therapy as group A but with no pentoxifylline injections; group C: 101 patients with PD and no treatment [30].
After treatment, a better response was observed in group A, where the combined therapy was bolstered by the association with pentoxifylline injections: a reduction in plaque volume in 100% of cases compared to the 79.7% obtained in group B; a mean reduction in plaque volume in 46.9% of cases compared to 24.8% of group B; an improvement in curvature in 96.8% of cases compared to 56.4% of group B; a mean reduction in the angle of the curve = −10.1 degrees compared to −4.8 degrees in group B; recovery of normal penile rigidity in patients with ED was obtained in 56% of cases compared to 23.5% of group B [30].
In 2013, we published a controlled study  which, in contrast with other articles in the literature, proved that oral vitamin E, associated with other substances (oral propolis + oral blueberry + verapamil injections) + topical diclofenac (multimodal therapy) for 6 months, was statistically effective in curing PD. In particular, vitamin E was shown to improve the therapeutic result: increasing the reduction in the volume of plaque after treatment (from 35.8% to 50.2%); increasing the percentage of patients who achieved an improvement in their penile curvature after treatment (from 48.4% to 96.6%); and increasing the degree of reduction in the curve (from 6.7 degrees to 12.2 degrees) [173].
Finally, we cite three case report articles of ours in which, overall, thanks to multimodal antioxidant treatment, we obtained the complete regression of plaque in eight patients suffering from PD [174][175][176].
In all patients, we used most of the antioxidants mentioned above (vitamin E, vitamin C, propolis, bilberry, silymarin, Ginkgo biloba, coenzyme Q10, boswellia, superoxide dismutase, diclofenac and pentoxifylline). In two cases, the combined oral and topical therapy was not associated with pentoxifylline injection therapy because the patients did not give their consent.
Treatment duration varied from 28 months to 53 months, most likely depending on plaque volume, since the patient who was cured in 28 months initially had plaque which was measured by ultrasound to be 122 mm 3 , while the patient whose treatment took 53 months had an initial plaque volume measuring 733 mm 3 and received no penile injections because of his refusal to undergo injection therapy.
The time period required for treatment was necessarily long due to the nature of the disease, i.e., its being a chronic (not an acute) inflammation, which therefore requires adequate treatment time. All patients, in any case, were informed before starting treatment that treatment would have to be long, due to the intrinsic characteristics of PD.
In all cases, at the end of treatment, patients experienced the complete regression of both their baseline penile pain and the penile curvature caused by the disease.

Discussion
Oxidative stress evidently plays a leading role in Peyronie's disease, a disease where there is an evident redox imbalance caused by the production of reactive species, following trauma to the penis in a genetically predisposed male.
It is also evident that oxidative stress plays a very large role not only in the onset, but especially in the progression of PD, resulting in local tissue damage and the formation of fibrotic plaque and its possible calcification [20,21,23].
From the literature, it is therefore clear that the most important physiopathogenic events are the following: penile injury (including minor trauma), formation of blood collection in the corpus cavernosum, recruitment of inflammatory cells, release of ROS and cytokines, activation of transcription factor NF-kB, the production of iNOS and the subsequent overproduction of radical NO, and the overproduction and local deposition of collagen (plaque formation) ( Figure 1). All the antioxidants we use in the treatment of PD, as indicated in Table 2A,B, can interfere with the disease's basic pathophysiological mechanisms, and this explains the good therapeutic outcomes in the literature.
The good results of the antioxidant treatment described above also prove that the doses used were adequate and the concentrations of the substances employed did not exceed the threshold beyond which they might have instead interacted negatively with the mechanisms of redox regulation of the tissues (see "antioxidant paradox") [180,181]. It is very likely that larger doses would have rendered the treatment ineffective or-worse still-favored disease progression.
Several antioxidant substances, some of which we use to treat PD, have been successfully used in other PD-related diseases. Silymarin, when given in a dose of 140 mg thrice daily for 3 months as an adjuvant for glycemic control, lipid profile, and insulin resistance, proved to have a beneficial efficacy [182]. Additionally, Daflon 500 mg (micronized a purified flavonoid fraction of Rutaceae aurantiae, consisting of 90% diosmin and 10% hesperidin) given twice daily for 45 days is helpful in reducing glucose level and the risk of cardiovascular disease [183].
In our clinical practice, in particular in the treatment of PD, we never doubted that our antioxidant doses were adequate and not excessive, especially since in our first experiences, we used at most three or four substances (vitamin E, propolis, blueberry, diclofenac) [173,177]. Over the following years, we gradually introduced other antioxidant substances in our combined PD therapy, with the aim of obtaining even more positive results; we always used low doses, even for these additional substances. Since in the past few years the results of our treatments have proved ever more effective and with no appreciable side effects, we are convinced we are not causing any damage to the redox system. On the contrary, the excellent results achieved in our most recent studies confirm that the most effective treatment for PD is a multimodal treatment, in which the various substances combine, with their different individual properties, in contrasting the multiple physiopathogenic mechanisms at play, with the aim of achieving better therapeutic results than those which could be obtained using a single or a very small number of substances [184].
We believe that many current articles on the conservative treatment of PD are lacking, and unable to accurately evaluate the volume of plaque before and after the treatment. The plaque volume must be evaluated only with a very sensitive ultrasound system and a three-dimensional evaluation with the formula of the ellipsoid (volume = 0.524 × length × width × thickness). The change in plaque volume is an important endpoint that should always be evaluated, together with the other parameters, for a correct evaluation of the therapeutic result. We propose that all future studies concerning the conservative treatment of PD include this very important parameter.

Conclusions
Oxidative stress represents the fundamental chemical environment for the onset and progression of Peyronie's disease, and-more broadly-for chronic inflammatory states, degenerative diseases, and malignant neoplasms. We think the good clinical results obtained using antioxidant therapy confirm the important role played by oxidative stress in PD. We also believe that the experiences of antioxidant treatments already present in the literature represent an important aid to the clinical practice of urologists for the treatment of this chronic inflammatory disease which, incidentally, is not rare, as its prevalence, especially in the Western world, is similar to that of diabetes mellitus (=10.5% worldwide) [185].
Treating PD with antioxidants directly interferes with the most important mechanisms of inflammation, thus treating the disease directly and not just its symptoms. We therefore hold that PD is a disease which can be cured, in contrast to the affirmation of authors who consider it an incurable condition [186]. We advocate this position in view of the fact that in the field of andrology, the prevailing background belief is that the gold standard of PD is surgery-basically, "treating the curve" and not "treating the disease"-whereas we and other authors support the latter [187].
We recommend that only patients with active (unstabilized) PD be treated with medical therapy. In the case of large penile plaques, we recommend adding periodic perilesional injections with pentoxifylline to the oral antioxidant substances.
In any case, we believe further studies are needed to both improve the understanding of the pathophysiology of Peyronie's disease and to implement new randomized controlled trials to confirm the efficacy of treatment with antioxidants. We also propose multi-center studies involving the combined use of antioxidants in order to demonstrate more clearly that PD can be cured.