2025 年 7 巻 3 号 p. 112-119
Synthetic surgical meshes are widely used for hernia repair; they effectively reduce recurrence rates. Mesh implantation in the abdominal wall is typically employed to study mesh toxicity; however, this site has limitations when evaluating their physical properties. This study proposed the buccal region as a novel implantation site and compared its histopathological outcomes with those of the conventional abdominal method. Three types of synthetic meshes: polypropylene (PP), polytetrafluoroethylene (PTFE), and polyvinylidene fluoride (PVDF), were implanted in the buccal and abdominal areas of the subjects. Tissue samples were collected and analyzed at 1–2 weeks post implantation. Compared to the abdominal implantation group, the buccal implantation group exhibited muscle layer adhesion and tissue infiltration on both mesh surfaces, although granulation and fibrosis were generally reduced, especially at 2 weeks. Among the meshes, PP demonstrated the highest pathology scores, whereas PTFE exhibited the lowest scores. Buccal implantation allows continuous mesh contact with the tissues involved in mastication and facial movements; this promotes mesh expansion, contraction, and enhanced tissue infiltration. This approach mitigates the disadvantages of conventional abdominal implantation such as mesh detachment and adjacent organ invasion. Therefore, buccal implantation provides a stable platform for pathological evaluation and enables the assessment of various physical properties after long-term implantation, making it a promising new model for assessing mesh biocompatibility.
Synthetic mesh evaluation compared the conventional abdominal wall implantation with a new buccal region method.
Three mesh types (polypropylene [PP], polytetrafluoroethylene [PTFE], and polyvinylidene fluoride [PVDF]) were assessed for bioreactivity, with PTFE showing the lowest invasiveness.
The buccal implantation method improved the reproducibility of histopathological analysis and minimized animal invasiveness.
This new transplantation site is valuable for directly assessing the biocompatibility of mesh materials and offers a more stable and less harmful alternative to traditional methods.
Synthetic medical meshes are widely used in surgical procedures to repair and reinforce weakened or damaged tissues. Advances in materials and manufacturing techniques have improved their safety and performance. These meshes have a wide range of applications, including the prevention of internal organ displacement caused by damage to the walls of the thoracic cavity, abdominal cavity, or diaphragm, such as in inguinal hernia [1, 2], pelvic organ prolapse (POP) [3], and hiatal hernia [4]. The primary challenge is to ensure durability and resistance to luminal pressure while minimizing the invasiveness to the surrounding tissues in direct contact with the mesh. Over the past 50 years, the development of synthetic meshes, mainly in Western countries, has progressed to include materials such as polypropylene (PP) [5], polyester (PE), polytetrafluoroethylene (PTFE) [6], polyvinylidene fluoride (PVDF) [7,8,9], and polylactic acid (PLA) [10]; meshes are also classified based on functionality, such as absorbable [11] and non-absorbable meshes. The required properties include sufficient tensile strength to prevent internal organ displacement caused by abdominal or organ pressure and to effectively cover and protect hernia sites, moderate elasticity and flexibility similar to those of thin skeletal muscles such as the abdominal muscles, and long-term durability. Generally, synthetic filaments outperform naturally derived silk braids because of their higher tensile strength, lower immunogenicity, and greater ease of processing into monofilaments. However, developing synthetic meshes with reduced biological invasiveness remains necessary, as their stability increases their invasiveness into the surrounding tissues, leading to various complications, such as chronic pain, infection, and inflammation, and persistent complaints [12].
Synthetic meshes, particularly tension-free vaginal meshes (TVM), are increasingly used to repair POP caused by childbirth and aging. Studies have reported their effectiveness, including lower recurrence rates than conventional methods such as vaginal hysterectomy with vaginal wall plasty [13]. Sacrovaginal fixation, in which a synthetic mesh is placed between the bladder and vagina or between the vagina and rectum with one side suspended over the sacral area to prevent organ prolapse, has become the mainstay of this technique. The utilization of laparoscopic and robot-assisted surgery offers advantages, including reduced infection rates, less bleeding, and shorter hospital stays [14]. In recent years, demand in Japan has increased for meshes composed of materials such as PP, PE, PTFE, and PVDF due to their stability, flexibility, and low bioinvasiveness. PTFE meshes have been applied in hiatal hernia repair [15], while PVDF meshes have been used for abdominal wall and inguinal hernia repair [16, 17]. However, the use of PTFE meshes for POP repair requires careful monitoring of national and international trends [18, 19], as well as further preclinical and clinical studies.
Pathological analysis in the nonclinical evaluation of implanted synthetic meshes enables gross and histological assessment of changes in both the organism and graft, as well as their interaction [20]. Such an evaluation provides detailed insights into local immune responses and infection following implantation, influenced by differences in the graft site and synthetic mesh properties, the process of complex formation between the filaments comprising the mesh and the tissue of biological origin, and the degenerative state of the filament surface itself. However, as non-clinical in vivo evaluations of synthetic meshes predominantly focus on abdominal wall hernia repair and biological reactivity, such as inflammatory and foreign body responses, many studies have employed implantation methods [21] within and outside of the abdominal muscles. Although this approach considers the effects on not only the muscle layer but also the subcutaneous tissue and intra-abdominal environment, the adhesion and adherence to the muscle layer are always unilateral and transient and are insufficient for evaluating the properties of the entire synthetic mesh. For POP repair, continuous evaluation of mesh fiber deterioration is necessary, considering their contact with highly motile tissues, such as the colon and vagina, as well as the stretching response of the mesh fibers themselves. To address this, this study utilized the subcutaneous buccal region of the face as a novel implantation site, where the entire mesh surface maintains close contact with the biological tissue, while a continuous stretching load is reproduced. In this model, the graft was positioned between the muscle layers of the plastyma muscle and temporalis muscles, both of which are involved in mastication. We speculated that the localized nature of the mesh and moderate mobility could be reproduced. The biocompatibility of the two materials widely used in TVM cases, PP and PTFE, was compared to that of an unapproved PVDF mesh, mainly through histopathological analysis.
This experiment was conducted in accordance with the “Laboratory Animal Welfare Regulations” of the Biotechnical Center of SLC (SLC Corporation, Shizuoka, Japan) and followed the ARRIVE guidelines for reporting animal experiments [22, 23].
AnimalsAll procedures were performed by the Contract Research Department of the Biotechnical Center of the SLC Corporation (Shizuoka, Japan). Fifteen female Sprague-Dawley rats (12 weeks old) were purchased from SLC Corporation and acclimated for 1 week before the experiments commenced at 13 weeks. Animals were kept in a room in a barrier facility maintained at 23 ± 2.0°C, with 50 ± 10% humidity and a 12-hr light/dark cycle (lights on from 7:00 to 19:00). They were kept in TPX floor flat-bottom cages (W21.8 × D37.3 × H18.1 cm) containing small soft fir wood chips (Soft tip, SLC Japan, Inc.). The animals were allowed free access to food (Labo MR Stock; Nippon Nosan Kogyo, Kanagawa, Japan) and chlorinated well water to meet the 2023 Ministry of Health, Labour and Welfare standards. They were housed in groups of up to three per cage. Post-surgery, Diet Gel® 76A (EP Trading, Tokyo, Japan) was added as needed to assist with feeding difficulties, and the animals continued to feed ad libitum.
Preparation of synthetic meshThree types of sterile synthetic meshes were prepared: PP (Johnson & Johnson K.K. Ethicon, Tokyo, Japan), PTFE, and PVDF (Kono Seisakusho, Chiba, Japan). All meshes were shaped convexly and cut to rectangular dimensions (approximately 5 mm wide × 10 mm long, adjusted to 2.5 mm wide at the tip) to fit the implantation site—the supraorbital to auricular region above the zygomatic bone under the facial skin. The meshes for abdominal wall implantation were cut into 10 × 10 mm pieces. Care was taken to avoid protrusions from the cut edges and minimize physical irritation to the insertion site (Fig. 1B, 1C).
Experimental protocol. After implanting various synthetic meshes (B, scale bar represents 5 mm) into the buccal region and abdominal cavity wall (day 0), the surgical sites were harvested on days 7 and 14, fixed in formalin, and pathological tissue specimens were prepared. Postoperative behavioral observations were conducted daily, and videos of feeding behavior were recorded every 7 days to monitor postoperative recovery. Three types of synthetic mesh were implanted and sutured into the buccal region (C) or abdominal cavity wall (D). The platysma muscle was incised to expose the temporalis ligament, onto which a pre-fragmented synthetic mesh was wrapped and implanted. Meanwhile, in the intra-abdominal implantation group, the skin at the abdominal midline and costal margin was incised in a T-shape in the supine position. Two types of mesh per animal were sutured to the left and right intra-abdominal walls, with the four corners of the rectangle secured using suture thread. PTFE: polytetrafluoroethylene; PVDF: polyvinylidene fluoride; PP: polypropylene.
Eighteen animals were divided into two experimental groups: (1) buccal subcutaneous implantation (12 animals) and (2) intraperitoneal implantation (6 animals). Pathological evaluations were performed on two animals per mesh type. In the buccal subcutaneous implantation group, one of each of the three different sterile synthetic meshes was implanted in the left buccal region. Following disinfection of the left buccal area with 0.5% Hexac® alcohol (Yoshida Pharmaceutical, Tokyo, Japan) under isoflurane general inhalation anesthesia (1.5–2.0%), a skin incision was made approximately 2 cm above the buccal area over the temporalis muscle. The broad cheek muscle was incised to expose the temporalis ligament, around which a small piece of synthetic mesh was wrapped for implantation (Fig. 2C). In the intraperitoneal implantation group, the animals were placed in a supine position under similar anesthesia and disinfection conditions. A T-shaped incision was made along the abdominal midline and the skin at the rib margin, followed by cuts through the linea alba, transverse abdominis muscle, and rectus abdominis muscle. Three different types of mesh (PP, PTFE, and PVDF) were then placed on the inner abdominal wall on both sides of the incision, and sutures (Brade silk 4–0, Nitcho Industry Co., Ltd., Tokyo, Japan) were used to secure the four corners of the rectangle (Fig. 2D). After implantation, the muscle and skin layers were closed using the same sutures. During the surgery, the animals were kept warm using a heater. Buprenorphine (0.05 mg/ml/kg B.W.; Otsuka Pharmaceutical, Tokyo, Japan) and enrofloxacin (5 mg/ml/kg B.W.; Kyoritsu Pharmaceutical, Tokyo, Japan) were administered subcutaneously for postoperative pain relief and infection prevention, respectively. General symptoms were observed daily postoperatively, body weight was measured (GX-3000; A & D Corporation, Tokyo, Japan), and feeding behavior was recorded weekly via video (GZ-F200 video camera; JVC Kenwood, Kanagawa, Japan). Chewing frequency (cycles per min) was measured and compared (Fig. 1A). Four weeks postoperatively, the animals were euthanized under isoflurane anesthesia and released via abdominal aortotomy. For the buccal subcutaneous implantation group, the entire skin and muscle layers from the orbit to the posterior auricle and from the parietal region to the mandibular neck were sampled. In the intraperitoneal implantation group, the entire skin and muscle layers from the orbit to the umbilical region were sampled, ensuring that the entire implantation material was sampled. The specimens were then immersed in 10% neutral buffered formalin (Fujifilm Wako Pure Chemical Industries, Tokyo, Japan) and fixed for histopathological analysis.
Rat body weight changes with buccal implants and abdominal implants groups, chewing frequency in buccal implants rats. Body weight measurements and behavioral observations were performed every 7 days starting from the day of implantation surgery (day 0). Feeding behavior was recorded and chewing frequency was measured (B). (A, **; P<0.01 vs. abdominal groups: B, *; P<0.05 vs. 7 days).
The experimental process was conducted at Sept. Sapie Co., Ltd., Pathology and Analysis Center (Tokyo, Japan). The formalin-fixed specimens were sectioned into three segments (approximately 5 mm wide × 2.0 cm) from the orbit to the ear canal; the rostral and caudal sections were thinly sectioned. The specimens were dehydrated, embedded in paraffin, thin-sectioned (6 µm), and stained with hematoxylin and eosin (Muto Chemical, Tokyo, Japan). Histopathological evaluation was performed by two blinded experts, one from the Japanese Society of Toxicological Pathology and the other from the Japanese Society of Veterinary Pathology, who independently assessed pathological changes, including granuloma formation, fibrosis, granulocyte and macrophage infiltration, and foreign body giant cell formation, by comparison with the area without the mesh, using a 5-point scale (0: no change, 1: slight, 2: mild, 3: moderate, 4: severe). A typical example of pathological evaluation has been added as a supplementary figure. All inflammatory parameters except fibrosis were grouped as “extent of granuloma” and compared across the three types of meshes.
Statistical analysisAll data are presented as mean ± standard error. Statistical analyses were performed using Statcel 4 (OMS Publishing, 2017) or R® (ver. 4.0.2, Statistical Analysis Research Institute, Inc.). Changes in body weight and chewing frequency were evaluated using paired t-tests, followed by one-way analysis of variance with duplicate measures. Comparison of scores in the histopathological analysis was performed using Tukey’s multiple comparison test for post-hoc analysis. Statistical significance was set at P<0.05.
The mean change in body weight every 7 days showed a significant increase in the buccally implanted group (n=12) compared with the abdominally implanted groups (A, **; P<0.01 vs. abdominal groups). Masticatory behavior analysis revealed an increase in chewing frequency per min between postoperative days 7 and 14 (Fig. 2B; P<0.05).
Pathological analysisWe compared the granulation tissue formation and fibrosis around the implantation site between the buccal and abdominal wall groups during the first and second weeks after synthetic mesh implantation. At 1 week, cellular infiltration (granulocytes, lymphocytes, macrophages, and foreign body giant cells) was enhanced in both groups within the outlined areas and was evaluated pathologically (Fig. 3, upper section). Foreign body giant cells appeared sporadically (Fig. 3, upper section; arrowheads). Connective tissue formation progressed around the masticatory muscles in the buccal group (Fig. 3, upper section A, C, E), whereas the abdominal group showed adhesion to the intra-abdominal adipose tissue (Fig. 3, upper section G, I, K). In the buccal PTFE group, infiltration was milder (Fig. 3, upper section, A and B), whereas the abdominal PP group showed prominent granuloma formation (Fig. 3, upper section, L; arrow).
Synthetic mesh surrounding tissue images after implantation in the buccal region (left) and abdominal wall (right). The upper section shows the buccal implant group (A–F) and the abdominal wall implant group (G–L) at 1 week and 2 weeks (lower section) post-implantation, with low-magnification images (×20, A, C, E, G, I, K) and high-magnification images (×400, B, D, F, H, J, L). Foreign body giant cells (arrowheads) and granulomas (arrows) are shown in the fibrotic periphery. The mesh implantation area is outlined with dashed lines (A, C, E, J, I, K), and each fiber section is indicated with an asterisk (B, D, F, H, J, L). PTFE: polytetrafluoroethylene; PVDF: polyvinylidene fluoride; PP: polypropylene.
By 2 weeks, cellular infiltration increased in both groups, particularly in the abdominal wall group (Fig. 3, lower section). The extent of granuloma scores (Fig. 4, upper section) ranged from 5 to 10 in all mesh groups at 1 week, with higher scores in the buccal group, particularly in the PVDF group (P<0.05). At 2 weeks, the scores remained at approximately 10, with upward trends in all abdominal wall groups and a significant increase in the PTFE buccal group (P<0.05). Fibrosis scores (Fig. 4, lower section) showed no differences at 1 week (1.5–2.5), but increased in the abdominal wall group by 2 weeks (3.0–4.0), especially in the PP and PVDF groups (P<0.01)
Pathological evaluation of synthetic mesh implantation in the buccal region and abdominal wall. Scores for extent of granuloma and fibrosis 1 week and 2 weeks after implantation are shown (*; P<0.05, **; P<0.01, n.s.; no significant). PTFE: polytetrafluoroethylene; PVDF: polyvinylidene fluoride; PP: polypropylene.
The in vivo properties of synthetic meshes require high biocompatibility with the surgical site and minimal invasiveness to the adjacent organs. Consequently, their physical properties must differ from those of tissue-substituting synthetic fiber fabrics used for hernia repair or thoracic and abdominal wall reinforcement [24] as well as from anti-adhesion materials used to prevent internal organ adhesion following cancer resection or transplantation [25], both of which are widely employed in current surgical practice. Additionally, improvements in mesh size and shape are necessary to make the material suitable for use in laparoscopic and robot-assisted surgeries. The two materials selected for this study (PP and PTFE) are widely used in POP cases and are associated with low postoperative complications such as organ adhesions and infections [26]. Internally absorbable meshes are also being developed to prevent delayed mesh-related infections [27, 28] and surgical site pain [29]. Findings from this buccal implantation model, together with insights into bone formation and repair in maxillofacial plastic surgery [30], suggest potential applications for facial muscle augmentation and the modulation of subtle changes in facial expression.
Body weight changes and ingestive behaviors were assessed over time to evaluate the invasiveness of the synthetic mesh implantation procedure. The results demonstrated a significant weight gain compared to the abdominal implant groups, regardless of the synthetic mesh material and shape (Fig. 2A). Additionally, masticatory behavior analysis revealed increased chewing frequency over time in the buccally implanted group, indicating that the maintenance of normal ingestive behavior contributed to the observed weight gain (Fig. 2A). Although mesh implantation between the broad neck and temporalis muscles under the facial skin may interfere with masticatory behavior during feeding or cause postoperative pain, the significant weight gain and normal ingestive behavior observed from postoperative days 7 to 14 suggest that the effect of the mesh implantation was minimal.
Histopathological evaluation revealed increased connective tissue formation within 14 days of implantation in the buccal implantation group (Fig. 4). In contrast, intraperitoneal implantation resulted in limited connective tissue formation between the abdominal wall surface and biological tissues, with a unilateral adhesive surface, high incidence of detachment from the target surface, and adhesion due to intraperitoneal adipose tissue. A stable synthetic mesh material can be assessed by suturing it to the biological side, making it a suitable model for abdominal wall hernia repair procedures, such as the intraperitoneal onlay mesh technique [31]. However, it is necessary to develop a method to reproduce techniques such as POP repair, given the wide variation in the degree of adhesion and contact between the entire synthetic mesh and living tissue. The buccal implantation method developed in this study (Fig. 1) involved inserting the mesh into a confined space between two muscle layers, thereby preventing displacement and reproducing the biological response to both sides of the mesh from the tissue side with high probability. Simultaneously, the deterioration caused by the physical stimulation of the mesh fibers owing to muscle movement can be considered a contributing factor. Furthermore, the increased opportunity for contact between both sides of the synthetic mesh and the surrounding biological tissue promotes fixation through active connective tissue formation. This approach may provide a novel assessment system for reducing postoperative pain, inflammation, and overall invasiveness, while enabling highly reproducible histopathological evaluations.
Exposure of epitopes resulting from fibrinogen adsorption and denaturation on filament surfaces has been reported to trigger inflammatory responses initiated by the transplantation of low-immunogenicity biomaterials [32]. This process is believed to promote the accumulation of the same cells through interactions with the phagocytic surface antigen Mac-1, leading to a hyper-inflammatory effect. The foreign body response via the innate immune system to the synthetic mesh involves protein adsorption onto the filament surface, acute and chronic inflammatory responses, atypical giant cell formation, and fibrosis, ultimately leading to fibrous capsule formation [33]. In recent years, non-clinical studies have increasingly focused on combining different filament materials [34] and designing synthetic meshes that optimize adhesion to biological tissues and minimize inflammatory responses [21, 35, 36]. Efforts have been made to induce tissue regeneration by applying stem cells and adhesive factors to synthetic meshes used in regenerative medicine [37]. In summary, synthetic meshes and suture materials are recognized as foreign substances, provoking inflammation that can result in granulation and fibrosis, and excessive reactions may cause contracture and adhesion. Therefore, preventive measures and development of better biocompatible materials remain essential.
A synthetic mesh transplantation model was created by certified laboratory animal technicians (Senior Laboratory Animal Technicians, Japanese Society for Laboratory Animal Resources) to ensure consistent reproducibility. However, insufficient data exist on the effects of technicians’ variability and inflammation due to surgical procedures for medical device development applications, requiring additional investigation using a sham operation group. Furthermore, the biocompatibility of the synthetic mesh was evaluated up to two weeks post-transplantation. However, because foreign body reactions, fibrosis, and mesh degradation/integration are chronic processes, long-term observation is essential for evaluating safety and efficacy. Although this novel buccal transplantation model has the potential to mimic the histological characteristics of human POP repair, direct comparative data are lacking, representing a critical future challenge.
The buccal implantation of synthetic meshes provides a stable and reproducible model for evaluating biocompatibility. Placement between the masticatory muscles allows tissue infiltration on both sides of the mesh with reduced fibrosis and minimal impact on feeding behavior, unlike abdominal implantation, which induces higher fibrosis and complications. Among the evaluated meshes, PTFE elicited the lowest inflammatory response. This novel approach mitigates mesh displacement issues and replicates physiological conditions more closely, making it a valuable model for long-term pathological and physical assessments.
This study was funded by Sept. Sapie Co., Ltd., Tokyo, Japan.
M. Kobayashi and M. Ishii served as staff members and CEO in Sept. Sapie Co., Ltd. All other authors declare no conflicts of interest.
We thank Prof. Hakamata (Nippon Veterinary and Life Science University) for valuable guidance in this study and Mrs. Kamata (Nippon Veterinary and Life Science University) for her assistance with animal care and management.
