Vulva-vaginal atrophy (VVA) is one of the common consequences of estrogen deficiency, especially after menopause. Several studies have evaluated the effects of hyaluronic acid (HA) on the physical and sexual symptoms associated with VVA and have achieved promising results. However, most of these studies have focused on the subjective assessment of the symptomatic response to topical formulations. Nevertheless, HA is an endogenous molecule, and it is logical that it works best if injected into the superficial epithelium. Desirial® is the first cross-linked hyaluronic acid administered via vaginal mucosal injection. The purpose of this study was to investigate the impact of multiple intravaginal intramucosal injections of specific cross-linked hyaluronic acid (DESIRIAL®, Laboratoires VIVACY) on several core clinical and patient-reported results.
Cohort two-center pilot study. The selected results included changes in vaginal mucosal thickness, collagen formation biomarkers, vaginal flora, vaginal pH, vaginal health index, symptoms of vulvovaginal atrophy and sexual function 8 weeks after Desirial® injection. The patient’s overall impression of improvement (PGI-I) scale was also used to assess patient satisfaction.
A total of 20 participants were recruited from 19/06/2017 to 05/07/2018. At the end of the study, there was no difference in the median total vaginal mucosa thickness or procollagen I, III, or Ki67 fluorescence. However, COL1A1 and COL3A1 gene expression increased statistically significantly (p = 0.0002 and p = 0.0010, respectively). The reported dyspareunia, vaginal dryness, genital itching, and vaginal abrasions were also significantly reduced, and all female sexual function index dimensions were significantly improved. Based on PGI-I, 19 patients (95%) reported varying degrees of improvement, of which 4 (20%) felt slightly better; 7 (35%) was better, and 8 (40%) was better.
Multi-point intravaginal injection of Desirial® (a cross-linked HA) was significantly associated with the expression of CoL1A1 and CoL3A1, indicating that collagen formation was stimulated. In addition, VVA symptoms were significantly reduced, and patient satisfaction and sexual function scores were significantly improved. However, the total thickness of the vaginal mucosa did not change significantly.
Vulva-vaginal atrophy (VVA) is one of the common consequences of estrogen deficiency, especially after menopause [1,2,3,4]. Several clinical syndromes are associated with VVA, including dryness, irritation, itching, dyspareunia, and recurrent urinary tract infections, which can have a significant negative impact on women’s quality of life [5]. However, the onset of these symptoms may be subtle and gradual, and begin to become apparent after other menopausal symptoms subsided. According to reports, up to 55%, 41%, and 15% of postmenopausal women suffer from vaginal dryness, dyspareunia, and repeated urinary tract infections, respectively [6,7,8,9]. Nevertheless, some people believe that the actual prevalence of these problems is higher, but most women do not seek medical help due to symptoms [6].
The main content of VVA management is symptomatic treatment, including lifestyle changes, non-hormonal (such as vaginal lubricants or moisturizers and laser treatment) and hormone treatment programs. Vaginal lubricants are mainly used to relieve vaginal dryness during sexual intercourse, so they cannot provide an effective solution to the chronicity and complexity of VVA symptoms. On the contrary, it is reported that vaginal moisturizer is a kind of “bioadhesive” product that can promote water retention, and regular use can improve vaginal irritation and dyspareunia [10]. Nevertheless, this has nothing to do with the improvement of the overall vaginal epithelial maturity index [11]. In recent years, there have been multiple claims to use radiofrequency and laser to treat vaginal menopausal symptoms [12,13,14,15]. Nevertheless, the FDA has issued warnings to patients, emphasizing that the use of such procedures may lead to serious adverse events, and has not yet determined the safety and effectiveness of energy-based devices in the treatment of these diseases [16]. Evidence from a meta-analysis of several randomized studies supports the effectiveness of topical and systemic hormone therapy in alleviating VVA-related symptoms [17,18,19]. However, a limited number of studies have evaluated the sustained effects of such treatments after 6 months of treatment. In addition, their contraindications and personal choice are limiting factors for the widespread and long-term use of these treatment options. Therefore, there is still a need for a safe and effective solution to manage VVA-related symptoms.
Hyaluronic acid (HA) is a key molecule of extracellular matrix, which exists in various tissues including vaginal mucosa. It is a polysaccharide from the glycosaminoglycan family, which plays an important role in maintaining water balance and regulating inflammation, immune response, scar formation and angiogenesis [20, 21]. Synthetic HA preparations are provided in the form of topical gels and have the status of “medical devices”. Several studies have evaluated the impact of HA on the physical and sexual symptoms associated with VVA and have achieved promising results [22,23,24,25]. However, most of these studies have focused on the subjective assessment of the symptomatic response to topical formulations. Nevertheless, HA is an endogenous molecule, and it is logical that it works best if injected into the superficial epithelium. Desirial® is the first cross-linked hyaluronic acid administered via vaginal mucosal injection.
The purpose of this prospective dual-center pilot study is to explore the impact of multi-point intravaginal intramucosal injections of specific cross-linked hyaluronic acid (DESIRIAL®, Laboratoires VIVACY) on the core results of several clinical and patient reports, and to evaluate the feasibility of the evaluation evaluation Sex these results. The comprehensive results selected for this study included changes in vaginal mucosal thickness, biomarkers of tissue regeneration, vaginal flora, vaginal pH and vaginal health index 8 weeks after Desirial® injection. We measured the results reported by several patients, including changes in sexual function and the reporting rate of VVA-related symptoms at the same point in time. At the end of the study, the patient’s overall impression of improvement (PGI-I) scale was used to assess patient satisfaction.
The study population consisted of postmenopausal women (2 to 10 years old after menopause) who were referred to a menopause clinic with symptoms of vaginal discomfort and/or dyspareunia secondary to vaginal dryness. Women must be ≥ 18 years old and <70 years old and have a BMI <35. Participants came from one of 2 participating units (Centre Hospitalier Régional Universitaire, Nîmes (CHRU), France and Karis Medical Center (KMC), Perpignan, France). Women are considered eligible if they are part of a health insurance plan or benefit from a health insurance plan, and they know that they can participate in the 8-week planned follow-up period. Women participating in other studies at the time were not eligible to be recruited. ≥ Stage 2 apical pelvic organ prolapse, stress urinary incontinence, vaginismus, vulvovaginal or urinary tract infection, hemorrhagic or neoplastic genital lesions, hormone-dependent tumors, genital bleeding of unknown etiology, recurrent porphyria, Uncontrolled epilepsy, cardiac conduction disorders, recurrent angina pectoris, rheumatic fever, previous vulvovaginal or urogynecological surgery, hemostatic disorders, and the tendency to form hypertrophic scars were considered as exclusion criteria. Women taking antihypertensive, steroidal and non-steroidal anti-inflammatory drugs, anticoagulants, major antidepressants or aspirin, and known local anesthetics linked to HA, mannitol, betadine, lidocaine, amide or Women who are allergic to any of the excipients in this medicine are considered to be ineligible for this study.
At baseline, women were asked to complete the Female Sexual Function Index (FSFI) [26] and use the 0-10 visual analog scale (VAS) to collect information related to VA symptoms (dyspareunia, vaginal dryness, vaginal abrasions, and genital itching) information. The pre-intervention evaluation included checking the vaginal pH, using the Bachmann Vaginal Health Index (VHI) [27] for clinical evaluation of the vagina, Pap smear to assess vaginal flora, and vaginal mucosal biopsy. Measure the vaginal pH near the planned injection site and in the vaginal fornix. For vaginal flora, Nugent score [28, 29] provides a tool to quantify the vaginal ecosystem, where 0-3, 4-6 and 7-10 points represent normal flora, intermediate flora and vaginosis, respectively. All assessments of the vaginal flora are performed in the Bacteriology Department of the CHRU in Nimes. Use standardized procedures for vaginal mucosal biopsy. Perform a 6-8 mm punch biopsy from the area of the planned injection site. According to the thickness of the basal layer, the middle layer and the superficial layer, the mucosal biopsy was evaluated histologically. Biopsy is also used to measure COL1A1 and COL3A1 mRNA, using RT-PCR and procollagen I and III immunotissue fluorescence as a surrogate for collagen expression, and the fluorescence of the proliferation marker Ki67 as a surrogate for mucosal mitotic activity. Genetic testing is carried out by the BioAlternatives laboratory, 1bis rue des Plantes, 86160 GENCAY, France (agreement is available upon request).
Once the baseline samples and measurements are completed, the cross-linked HA (Desirial®) is injected by one of the 2 trained experts according to the standard protocol. Desirial® [NaHa (sodium hyaluronate) cross-linked IPN-Like 19 mg/g + mannitol (antioxidant)] is an injectable HA gel of non-animal origin, for single use and packaged in pre-packed Syringe (2 × 1 ml). It is a Class III medical device (CE 0499), used for intramucosal injection in women, used for biostimulation and rehydration of the mucosal surface of the genital area (Laboratoires Vivacy, 252 rue Douglas Engelbart-Archamps Technopole, 74160 Archamps, France). Approximately 10 injections, each 70-100 µl (0.5-1 ml in total), are performed on 3-4 horizontal lines in the triangular area of the posterior vaginal wall, the base of which is at the level of the posterior vaginal wall, and the apex at 2 cm above ( figure 1).
The end-of-study evaluation is scheduled for 8 weeks after enrollment. The evaluation parameters for women are the same as those at baseline. In addition, patients are also required to complete the Overall Improving Impression (PGI-I) Satisfaction Scale [30].
In view of the lack of prior data and the pilot nature of the research, it is impossible to conduct a formal prior sample size calculation. Therefore, a convenient sample size of a total of 20 patients was selected based on the capabilities of the two participating units and was sufficient to obtain a reasonable estimate of the proposed outcome criteria. Statistical analysis was performed using SAS software (9.4; SAS Inc., Cary NC), and the significance level was set at 5%. The Wilcoxon signed rank test was used for continuous variables and the McNemar test was used for categorical variables to test the changes at 8 weeks.
The research was approved by Comité d’ethique du CHU Carémeau de Nimes (ID-RCB: 2016-A00124-47, protocol code: LOCAL/2016/PM-001). All study participants signed a valid written consent form. For 2 study visits and 2 biopsies, patients can receive compensation of up to 200 Euros.
A total of 20 participants were recruited from 19/06/2017 to 05/07/2018 (8 patients from CHRU and 12 patients from KMC). There is no agreement that violates a priori inclusion/exclusion criteria. All injection procedures were safe and sound and were completed within 20 minutes. The demographic and baseline characteristics of the study participants are shown in Table 1. At baseline, 12 out of 20 women (60%) used the treatment for their symptoms (6 hormonal and 6 non-hormonal), while at week 8 only 2 patients (10%) were still treated like this ( p = 0.002).
The results of clinical and patient report results are shown in Table 2 and Table 3. One patient refused the W8 vaginal biopsy; the other patient refused the W8 vaginal biopsy. Therefore, 19/20 participants can obtain complete histological and genetic analysis data. Compared with D0, there was no difference in the median total thickness of the vaginal mucosa at week 8. However, the median basal layer thickness increased from 70.28 to 83.25 microns, but this increase was not statistically significant (p = 0.8596). There was no statistical difference in the fluorescence of procollagen I, III or Ki67 before and after treatment. Nevertheless, COL1A1 and COL3A1 gene expression increased statistically significantly (p = 0.0002 and p = 0.0010, respectively). There was no statistically significant change, but it helped to improve the trend of vaginal flora after Desirial® injection (n = 11, p = 0.1250). Similarly, near the injection site (n = 17) and the vaginal fornix (n = 19), the vaginal pH value also tended to decrease, but this difference was not statistically significant (p = p = 0.0574 and 0.0955) (Table 2 ).
All study participants have access to patient-reported results. According to PGI-I, one participant (5%) reported no change after injection, while the remaining 19 patients (95%) reported varying degrees of improvement, of which 4 (20%) felt slightly better; 7 (35 %) is better, 8 (40%) is better. The reported dyspareunia, vaginal dryness, genital itching, vaginal abrasions, and FSFI total scores as well as their desire, lubrication, satisfaction, and pain dimensions were also significantly reduced (Table 3).
The hypothesis supporting this study is that multiple Desirial® injections on the posterior wall of the vagina will thicken the vaginal mucosa, lower vaginal pH, improve vaginal flora, induce collagen formation and improve VA symptoms. We were able to demonstrate that all patients reported significant improvements, including dyspareunia, vaginal dryness, vaginal abrasions, and genital itching. VHI and FSFI have also been significantly improved, and the number of women who need alternative therapies to control their symptoms has also been significantly reduced. Relatedly, it is feasible to collect information about all the results determined at the beginning and be able to provide interventions for all study participants. In addition, 75% of study participants reported that their symptoms improved or were much better at the end of the study.
However, despite the slight increase in the average thickness of the basal layer, we could not prove a significant effect on the total thickness of the vaginal mucosa. Although our study was unable to evaluate the effectiveness of Desirial® in improving vaginal mucosal thickness, we believe that the results are relevant because the expression of CoL1A1 and CoL3A1 markers was statistically significantly increased in W8 compared to D0. Means collagen stimulation. However, there are some issues to consider before considering its use in future research. First, is the 8-week follow-up period too short to prove an improvement in total mucosal thickness? If the follow-up time is longer, the changes identified in the base layer may have been implemented in other layers. Secondly, does the histological thickness of the mucosal layer reflect tissue regeneration? The histological evaluation of vaginal mucosal thickness does not necessarily consider the basal layer, which includes the regenerated tissue in contact with the underlying connective tissue.
We understand that the small number of participants and the lack of a priori formal sample size are the limitations of our research; nonetheless, both are standard features of the pilot study. It is for this reason that we avoid extending our findings to claims of clinical validity or invalidity. However, one of the main advantages of our work is that it allows us to generate data for several results, which will help us calculate the formal sample size for future deterministic research. In addition, the pilot allows us to test our recruitment strategy, churn rate, feasibility of sample collection and result analysis, which will provide information for any further related work. Finally, the series of results we evaluated, including objective clinical results, biomarkers, and patient-reported results evaluated using validated measures, are the main strengths of our research.
Desirial® is the first cross-linked hyaluronic acid administered via vaginal mucosal injection. In order to deliver the product through this route, the product must have sufficient fluidity so that it can be easily injected into the specialized dense connective tissue while maintaining its hygroscopicity. This is achieved by optimizing the size of the gel molecules and the level of gel cross-linking to ensure high gel concentration while maintaining low viscosity and elasticity.
A number of studies have evaluated the beneficial effects of HA, most of which are non-inferiority RCTs, comparing HA with other forms of treatment (mainly hormones) [22,23,24,25]. The HA in these studies was administered locally. HA is an endogenous molecule characterized by its extremely important ability to fix and transport water. With age, the amount of endogenous hyaluronic acid in the vaginal mucosa decreases sharply, and its thickness and vascularization also decrease, thereby reducing plasma exudation and lubrication. In this study, we have demonstrated that Desirial® injection is associated with a significant improvement in all VVA-related symptoms. These findings are consistent with a previous study conducted by Berni et al. As part of Desirial® regulatory approval (undisclosed-supplementary information) (Additional File 1). Although only speculative, it is reasonable that this improvement is secondary to the possibility of restoring the transfer of plasma to the vaginal epithelial surface.
Cross-linked HA gel has also been shown to increase the synthesis of type I collagen and elastin, thereby increasing the thickness of surrounding tissues [31, 32]. In our study, we did not prove that the fluorescence of procollagen I and III is significantly different after treatment. Nevertheless, COL1A1 and COL3A1 gene expression increased statistically significantly. Therefore, Desirial® may have a stimulating effect on the formation of collagen in the vagina, but larger studies with longer follow-up are needed to confirm or refute this possibility.
This study provides baseline data and potential effect sizes for several results, which will help future sample size calculations. In addition, the study proved the feasibility of collecting different results. However, it also highlights several issues that need to be carefully considered when planning future research in this area. Although Desirial® seems to significantly improve VVA symptoms and sexual function, its mechanism of action is unclear. As can be seen from the significant expression of CoL1A1 and CoL3A1, there seems to be preliminary evidence that it stimulates the formation of collagen. Nevertheless, procollagen 1, procollagen 3 and Ki67 did not achieve similar effects. Therefore, additional histological and biological markers must be explored in future research.
Multi-point intravaginal injection of Desirial® (a cross-linked HA) was significantly associated with the expression of CoL1A1 and CoL3A1, indicating that it stimulates collagen formation, significantly reduces VVA symptoms, and uses alternative treatments. In addition, based on the PGI-I and FSFI scores, patient satisfaction and sexual function improved significantly. However, the total thickness of the vaginal mucosa did not change significantly.
The data set used and/or analyzed during the current study can be obtained from the corresponding author upon reasonable request.
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