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The aim of this study was to investigate the potential role of transvaginal mesh bacterial colonization in the development of mesh-related complication (MRCs).An observational and exploratory study was performed including 49 patients indicated for mesh removal and 20 women of whom vaginal tissue was retrieved during prolapse surgery as a reference cohort.Homogenized samples were used for quantitative microbiological culture. Inflammation and fibrosis were semi-quantitatively histologically scored, Gram staining and Fluorescence in situ Hybridization (FISH) was used to detect bacteria and bacterial biofilms.Of the 49 patients, 44 (90%) samples were culture positive, with a higher diversity of species, more Gram-negative bacteria and polymicrobial cultures in the MRC cohort compared to the reference cohort, with mostly staphylococci, streptococci, Actinomyces spp., Cutibacterium acnes and Escherichia coli. Patients with clinical signs of infection or exposure had the highest bacterial counts. Histology demonstrated moderate to severe inflammation in the majority of samples. Gram staining showed bacteria in 57% of culture positive samples and in selected samples FISH illustrated a polymicrobial biofilm.In this study we observed distinct differences in bacterial numbers and species between patients suffering from MRCs compared to a reference cohort. Bacteria were observed at the mesh-tissue interface in a biofilm. These results strongly support the potential role of bacterial mesh colonization in the development of MRCs.

Objective: This study investigates whether noninvasive focal depth (FD) measurements correlate with vaginal wall epithelial thickness (ET). If FD accurately reflects ET of the vaginal wall, this would allow noninvasive longitudinal assessment of (newly developed) treatment modalities aiming to increase ET, without the need for invasive biopsies. Methods: Fourteen women, median age 62 years (inter quartile ranges: 57-65), undergoing vaginal prolapse surgery because of anterior and/or posterior compartment pelvic organ prolapse were included. We used the CytoCam, a handheld video microscope based on incident dark field imaging, and performed FD measurements of the vaginal wall before surgery. Histology was performed on tissue that was removed during the surgical procedure, and ET was measured in stained sections. We compared ET with FD interindividually, and determined the expected linear correlation and agreement between the two measurements. Results: Seventeen ET measurements (mean 125 mu m +/- 38.7, range 48-181 mu m) were compared with 17 FD measurements (mean 128 mu m +/- 34.3, range 68-182 mu m). The lineair correlation between the two measurements was strong (r = 0.902, P < 0.01). Bland-Altman analysis demonstrated a mean difference of 13.5 mu m when comparing ET to FD. Conclusions: The results demonstrate good agreement between ET and FD measurements. We consider the mean difference demonstrated with Bland-Altman analysis acceptable for these measurements. This suggests that FD accurately reflects ET, which further supports the use of FD to measure ET of the vaginal wall. For a complete assessment of the vaginal wall, FD measurements are preferably combined with the assessment of vaginal angioarchitecture.

Objective In sheep of reproductive age, we aimed to document decrease in epithelial thickness, glycogen amount, and other vaginal changes after castration and the effect of Er:YAG laser as used clinically. Methods On day 0, 16 sheep underwent ovariectomy. They were randomized to sham or three vaginal Er:YAG laser applications at monthly intervals. Primary outcome was vaginal epithelial thickness (d60, d71, d73, d77, and d160). Secondary outcomes included indicators of atrophy (vaginal health index = VHI), pH, cytology, morphology at the above time points, microcirculation focal depth (FD; d70 and d160), and at sacrifice (d160) vaginal dimensions and active and passive biomechanical testing. Results Menopausal changes between 60 and 160 days after ovariectomy included a progressive decrease in epithelial thickness, in VHI, FD, glycogen, elastin content and vasculature, and an increase in pH and collagen content. In lasered animals, the first day a few white macroscopic foci were visible and an increase in pH was measured. Both disappeared within 3 days. Seven days after laser the epithelial thickness increased. At sacrifice (d160), there were no differences between sham and laser group in vaginal dimensions, morphometry, mitotic and apoptotic activity, active contractility, vaginal compliance, except for a lower blood vessel density in the lamina propria of the midvagina in the laser group. Conclusions In reproductive sheep, ovariectomy induces vaginal atrophy evidenced in different outcome measurements. Vaginal Er:YAG laser induced visual impact, a short-term increase in epithelial thickness yet no long-term changes compared to sham therapy in menopausal controls.

Pelvic organ prolapse (POP) is a multifactorial condition characterized by the descent of the pelvic organs due to the loss of supportive tissue strength. This is presumably caused by the decreased fibroblast function and the subsequent change in the quality of the extracellular matrix. The correction of POP using an implant intends to provide mechanical support to the pelvic organs and to stimulate a moderate host response. Synthetic polypropylene (PP) implants were commonly used for the correction of prolapse. Although they were successful in providing support, these implants have been associated with clinical complications in the long term due to substantial foreign body response and inappropriate tissue integration. The complications can be avoided or minimized by engineering a biocompatible and fully absorbable implant with optimized mechanical and structural characteristics that favor more appropriate cellular interactions with the implant. Therefore, in this study, we evaluated implants comprised of poly-4-hydroxybutyrate (P4HB), a fully absorbable material with high mechanical strength, as an alternative to PP. The P4HB implants were knitted in four unique designs with different pore shapes ranging from a more rectangular geometry- as it is in PP implant- to a rounded geometry, to determine the effect of the implant structure on the textural and mechanical properties and subsequent cell-matrix interaction. The cellular response was investigated by seeding primary vaginal fibroblasts isolated from patients with POP. P4HB favored cellular functions more than PP, as indicated by greater cell attachment and proliferation (P < 0.01), and significantly more collagen deposition (P4HB vs PP, 11.19 μg vs 6.67 μg) at 28 days culture (P < 0.05). All P4HB implants had higher strength and lower stiffness than the PP scaffold. The material and the design of the implant also influenced the behavior of vaginal fibroblasts. The aspect ratio of the vaginal POP fibroblasts cultured on the PP implant (1.61 ± 0.75) was significantly (P < 0.005) smaller than those cultured on P4HB implants (average 2.31 ± 0.09). The P4HB structure with rounded pores showed the lowest stiffness and highest fibroblast attachment and proliferation (P < 0.01). Overall, P4HB induces more matrix deposition compared to PP and knit design can further optimize cell behavior.

OBJECTIVE The aim of the study was to evaluate if vaginal focal depth measurement could be a noninvasive method to quantify vaginal wall thickness. METHODS Postmenopausal women undergoing topical estrogen therapy because of vaginal atrophy (VA) were recruited. VA was diagnosed based on the presence of symptoms and vaginal pH at least 5.5. The control group consisted of women above 40 years without VA. Focal depth measurements were performed before and after treatment using the Cytocam-Incident Dark Field device assessing the distance between the subepithelial microcirculation and the epithelial surface. Measurements were performed before and after treatment in the intervention group and at two different time points in the control group. Vaginal pH was measured. Symptoms were evaluated using the most bothersome symptom approach. RESULTS Eight women with VA and nine controls were included. Pretreatment median focal depth was not significantly different between both groups. Pretreatment focal depth more than doubled after a median of 7 weeks of topical estrogen treatment (80 μm [interquartile range 80-120 μm] vs 220 μm [148-248 μm], P = 0.02), whereas the measurements in the control group did not change. Pretreatment vaginal pH differed between both groups (5.5 vs 5.1, respectively, P

The extracellular matrix of fascia-like tissues is a resilient network of collagenous fibers that withstand the forces of daily life. When overstretched, the matrix may tear, with serious consequences like pelvic organ prolapse (POP). Synthetic implants can provide mechanical support and evoke a host response that induces new matrix production, thus reinforcing the fascia. However, there is considerable risk of scar formation and tissue contraction which result in severe complications. Matrix producing fibroblasts are both mechanosensitive and contractile; their behavior depends on the implant's surface texture and mechanical straining. Here we investigate the effect of both in a newly-designed experimental setting. Electrospun scaffolds of Nylon and PLGA/PCL and a non-porous PLGA/PCL film were clamped like a drumhead and seeded with fibroblasts of POP patients. Upon confluency, scaffolds were cyclically strained for 24 or 72 h at 10% and 0.2 Hz, mimicking gentle breathing. Non-loading condition was control. Strained fibroblasts loosened their actin-fibers, thereby preventing myofibroblastic differentiation. Mechanical loading upregulated genes involved in matrix synthesis (collagen I, III, V and elastin), matrix remodeling (α-SMA, TGF-β1, MMP-2) and inflammation (COX-2, TNF-α IL8, IL1-β). Collagen genes were expressed earlier under mechanical loading and the ratio of I/III collagen increased. Matrix synthesis and remodeling were stronger on the electrospun scaffolds, while inflammation was more prominent on the non-porous film. Our findings indicate that mechanical straining enhances the regenerative potential of fibroblasts for the regeneration of fascia-type tissues and limit the risk of scar tissue formation. These effects are stronger on an electrospun texture. Statement of significance: Pelvic organ prolapsed is a dysfunctional disease in female pelvic floor that can reduce the quality of life women. Currently, trans-vaginal knitted meshes are used to anatomically correct the dysfunctional tissues. However, the meshes can create sever adverse complications in some patients (e.g. chronic pain) in longer-term. As an alternative, we developed nanofibrous matrices by electrospinning based on different materials. We designed an in-vitro culture system and subjected cell-seeded matrices to cyclic mechanical loading. Results revealed that gentle straining of POP-cells on electrospun matrices, advances their regenerative potential at morphological and gene expression levels. Our findings, provide a proof-of-concept for using electrospun matrices as an alternative implant for pelvic floor repair, given that the parameters are designed efficiently and safely.