Fig 1.
Characterization of the prosthetic materials.
Scanning electron microscopy (SEM) images (20x magnification) of: (A) the laminar expanded polytetrafluoroethylene (ePTFE). A further magnified view (2000x) is shown in the box; (B) The bare reticular polypropylene alone (PP, Surgipro); (C) DynaMesh (PP interwoven with polyvinylidene fluoride, PVDF) and (D) TiMESH (PP coated with titanium). Note that for DynaMesh (C) the PVDF side (designed to be viscerally deployed) is shown.
Fig 2.
Illustration depicting the design of the culture chambers (A) and the experimental setup (B).
Fig 3.
In vitro mesothelialization of meshes.
Scanning electron microscopy (1000x magnification) of the mesh fragments seeded with omental mesothelial cells. The micrographs are representative of every study group: Preclude (laminar e-PTFE); Surgipro (bare reticular PP alone), DynaMesh (PP interwoven with PVDF) and TiMESH (PP coated with titanium) at 1, 2 and 3 days after seeding.
Fig 4.
Surgical technique, laparoscopy and adhesion formation.
Macroscopic appearance of the meshes after implantation and laparoscopic analysis in the different experimental groups: Preclude (P, laminar e-PTFE); Surgipro (S, bare reticular PP alone) and the two hybrid reticular meshes: DynaMesh (D, PP interwoven with PVDF) and TiMESH (T, PP coated with titanium). Percentage of the mesh areas covered by adhesions for each experimental group at 7 and 14 days post-implantation are shown. Reticular meshes (Surgipro, DynaMesh and TiMESH) showed no differences between each other at any study time point, but differences were significantly higher for all of them compared to the laminar Preclude (*p<0.05, **p<0.01, ***p<0.001).
Fig 5.
Morphological studies of the implanted meshes.
Light microscopy (Masson´s trichrome staining, 100x magnification) and scanning electron microscopy micrographs of the surface of the materials (500x magnification) in adhesion-free and adhesion-covered areas for each group at 14 days post-implantation. The Preclude group showed no adhesion-covered areas in any animal. f: mesh filaments, P: Preclude.
Fig 6.
Immunolabeling for collagens type I and III and Picrosirius red staining in implants from the different study groups (100x magnification). f: mesh filaments, P: Preclude.
Fig 7.
Foreign-body reaction of the different meshes.
RAM-11 monoclonal antibody immunohistochemical staining showing the presence of labeled macrophages (→) in proximity to the different mesh materials (200x magnification): (A) Preclude, (B) Surgipro, (C) DynaMesh and (D) TiMESH. f: mesh filaments, P: Preclude. (E) Positive cell counts per field recorded after 14 days of implantation. The results are shown as the means ± standard error of the mean for the different study groups: P (Preclude), S (Surgipro), D (DynaMesh) and T (TiMESH). *p<0.05.
Fig 8.
Collagen 1 and 3 mRNA expression levels determined by RT-PCR in the implant areas.
Relative mRNA levels of collagen 1, collagen 3 and the Col1/Col 3 ratio in implant samples from the different experimental groups. Gene expression was normalized to the expression recorded for the reference gene GAPDH. Agarose gel electrophoresis of RT-PCR products is shown. Each pair of lanes is representative of each experimental group: P, Preclude; S, Surgipro; D, DynaMesh; T, TiMESH. Mw: Molecular weight markers. N: Negative. Collagen 1 mRNA expression was statistically greater in the hybrid meshes DynaMesh (*, p<0.05) and TiMESH (**, p<0.01) compared to Surgipro and Preclude. No differences were found in collagen 3 mRNA expression levels or the Col1/Col3 ratio.
Fig 9.
Morphological studies and foreign-body reaction in adhesion tissue.
Representative light microscopy micrographs (A and E, Masson´s trichrome staining, 100x magnification; C, hematoxylin-eosin staining, 100x magnification) and immunohistochemical labeling of rabbit macrophages (→) using the RAM-11 monoclonal antibody (B, D and F, 320x magnification) in adhesion tissue formed to the different meshes (A and B, Surgipro; C and D, DynaMesh; E and F, TiMESH).
Fig 10.
Collagen expression in adhesion tissue.
Immunohistochemical labeling for collagen type I (A, D and G, 200x magnification), collagen type III (B, E and H, 200x magnification), and Picrosirius red staining (C, F and I, 100x magnification) in adhesion tissues from animals implanted with different meshes (A, B and C, Surgipro; D, E and F, DynaMesh; G, H and I, TiMESH).
Fig 11.
Collagen 1 and 3 mRNA expression levels determined by RT-PCR in adhesion tissue.
Relative mRNA levels of collagen 1, collagen 3 and Col1/Col 3 ratios in adhesion tissues from the different experimental groups that developed adhesions (Surgipro, DynaMesh and TiMESH). Gene expression was normalized to the expression recorded for the reference gene GAPDH. Agarose gel electrophoresis of RT-PCR products is shown: S, Surgipro; D, DynaMesh; T, TiMESH. Mw: Molecular weight markers. N: Negative. No significant differences were found for collagen 1 or collagen 3 mRNA expression or for the Col 1/Col 3 ratios.
Fig 12.
Morphological studies in non-injured omentum.
Light microscopy micrographs (A, B and D, Masson´s trichrome staining, 100x magnification; C, hematoxylin-eosin, 100x magnification) of samples of omentum (non-injured and not involved in adhesions) from animals implanted with different meshes: (A) Preclude, (B) Surgipro, (C) DynaMesh and (D) TiMESH.
Fig 13.
Collagen 1 and 3 mRNA expression levels determined by RT-PCR in non-injured omentum.
Relative mRNA levels of collagen 1, collagen 3 and Col1/Col 3 ratios in samples of omentum (non-injured and not involved in adhesions) from animals implanted with different meshes. Gene expression was normalized to the expression recorded for the reference gene GAPDH. Agarose gel electrophoresis of RT-PCR products is shown: P, Preclude; S, Surgipro; D, DynaMesh; T, TiMESH. Mw: Molecular weight markers. N: Negative. All reticular meshes showed statistically significant higher collagen 1 mRNA expression levels compared to the laminar Preclude (**, p<0.01 vs. Surgipro and *, p<0.05 vs. DynaMesh, TiMESH). Col 1/ Col 3 ratios were greatest in TiMESH vs. Preclude or Surgipro (*, p<0.05).