Assessment of mesh shrinkage using fibroblast-populated collagen matrices: a proof of concept for in vitro hernia mesh testing.

PURPOSE: This study uses free-floating contractile fibroblast-populated collagen matrices (FPCMs) to test the shrinkage of different hernia mesh products. We hope to present this model as a proof of concept for the development of in vitro hernia mesh testing-a novel technology with interesting potential. METHODS: FPCMs were formed by seeding Human Dermal Fibroblasts into collagen gels. FPCMs were seeded with three different cell densities and cast at a volume of 500 µl into 24-well plates. Five different mesh products were embedded within the collagen constructs. Gels were left to float freely within culture media and contract over 5 days. Photographs were taken daily and the area of the collagen gel and mesh were measured. Media samples were taken at days 2 and 4 for the purposes of measuring MMP-9 release. After 5 days, dehydrated FPCMs were also examined under light and fluorescence microscopy to assess cell morphology. RESULTS: Two mesh products-the mosquito net and large pore lightweight mesh were found to shrink notably more than others. This pattern persisted across all three cell densities. There were no appreciable differences observed in MMP-9 release between products. CONCLUSIONS: This study has successfully demonstrated that commercial mesh products can be successfully integrated into free-floating contractile FPCMs. Not only this, but FPCMs are capable of applying a contractile force upon those mesh products-eliciting different levels of contraction between mesh products. Such findings demonstrate this technique as a useful proof of concept for future development of in vitro hernia mesh testing.

The experimental methodology and comparators used for in vivo hernia mesh testing: a 10-year scoping review.

PURPOSE: Before being marketed, hernia mesh must undergo in vivo testing, which often includes biomechanical and histological assessment. Currently, there are no universal standards for this testing and methods vary greatly within the literature. A scoping review of relevant studies was undertaken to analyse the methodologies used for in vivo mesh testing. METHODS: Medline and Embase databases were searched for relevant studies. 513 articles were identified and 231 duplicates excluded. 126 papers were included after abstract and full text review. The data extraction was undertaken using standardised forms. RESULTS: Mesh is most commonly tested in rats (53%). 78% of studies involve the formation of a defect; in 52% of which the fascia is not opposed. The most common hernia models use mesh to bridge an acute defect (50%). Tensile strength testing is the commonest form of mechanical testing (63%). Testing strip widths and test speeds vary greatly (4-30 mm and 1.625-240 mm/min, respectively). There is little consensus on which units to use for tensile strength testing. Collagen is assessed for its abundance (54 studies) more than its alignment (18 studies). Alignment is not measured quantitatively. At least 21 histological scoring systems are used for in vivo mesh testing. CONCLUSIONS: The current practice of in vivo mesh testing lacks standardisation. There is significant inconsistency in every category of testing, both in methodology and comparators. We would call upon hernia organisations and materials testing institutions to discuss the need for a standardised approach to this field.

In vitro characterisation of low-cost synthetic meshes intended for hernia repair in the UK.

PURPOSE: Low-cost meshes (LCM) were repurposed for the repair of hernias in the developing world. In vivo studies have shown LCM to have comparable results to commercial meshes (CM) at a fraction of the cost. However, little has been done to characterise the mechanical and biocompatible properties of LCM, preventing its clinical use in the UK. The objectives of the research are to assess mechanical and ultrastructural properties of two UK-sourced low-cost meshes (LCM) and the characterisation of the LCMs in vitro biocompatibility. METHODS: Mechanical properties of the two LCM were measured through uniaxial tensile test and ultrastructure was evaluated with Scanning Electron Microscopy. LIVE/DEAD® Viability/Cytotoxicity Assay kit and alamarBlue were used to assess cellular viability and proliferation, respectively. Images were acquired with a fluorescence microscope and analysed using ImageJ (NIH, USA). RESULTS: LCM1 and LCM2 were both multifilament meshes, with the first having smaller pores than the latter. LCM1 exhibited significantly higher tensile strength (p < 0.05) than LCM2 but significantly lower extensibility (p < 0.0001), while Young's Modulus of the two samples was not significantly different. No significant difference was found in the cellular viability and morphology cultured in LCM1 and LCM2 conditioned media. Metabolic assay and fluorescence imaging showed cellular attachment and proliferation on both LCMs over 14 days. CONCLUSION: The characterisation of the two UK-sourced LCMs showed in vitro biocompatibility and mechanical and ultrastructural properties comparable to the equivalent CM. This in vitro data represents a step forward for the feasibility of adopting LCM for surgical repair of hernias in the UK.

Application of Demirjian's and Cameriere's Method in Dental Age Estimation of 8-16 Year Old Adolescents from Hunan Han Nationality.

ABSTRACT: Objective To apply Demirjian's and Cameriere's method for dental age estimation of adolescents from Hunan Han nationality, and compare the accuracy of the two methods. Methods A total of 480 orthopantomograms of?8-16 year?old adolescents from Hunan Han nationality?with no special diseases and good nutritional status were collected?by Xiangya Stomatological Hospital of Central South University from January, 2016 to July, 2017, among them 236 males and 244 females. The dental age of each adolescent was determined by Demirjian's method and Cameriere's method, respectively, and the paired t-test of the estimated dental age and the chronological age determined by the two methods was conducted by SPSS 20.0 software to compare the difference between estimated dental age and chronological age. Results Mean chronological age of males and females was 11.91 and 11.88 years, respectively. The estimated dental age determined by Demirjian's method showed an underestimate of chronological age by an average of 0.11 years （males） and 0.15 years （females）, while the estimated dental age determined by Cameriere's method showed an underestimate of chronological age by an average of 0.83 years （males） and 0.72 years （females）. Conclusion Demirjian's method is more accurate than Cameriere's method in dental age estimation of adolescents from Hunan Han nationality, therefore more suitable for dental age estimation of adolescents in this region.

Development of a conjunctival tissue substitute on the basis of plastic compressed collagen.

Ocular surface disorders, such as pterygium, cicatricial pemphigoid and external disruptions, can cause severe inflammation, scarring, fornix shortening as well as ankyloblepharon. Current treatments do not resolve these conditions sufficiently. The aim of this study was to evaluate clinical applicability and suitability of plastic compressed collagen to serve as a substrate for the expansion of human conjunctival epithelial cells in order to develop an epithelialized conjunctival substitute for fornix reconstruction. Human conjunctival epithelial cells were expanded on plastic compressed collagen gels. Epithelial cell characteristics were evaluated by haematoxylin and eosin staining, electron microscopy and cytokeratin expression. The expression of putative epithelial progenitor cell markers p63α, ABCG2 and CK15 was assessed by immunostaining. The proliferative capacity and clonal growth of the cells was evaluated before (P0) and after expansion (P1) on the plastic compressed collagen gels by colony forming efficiency assay. The potential clinical applicability of this gel substitutes was evaluated by assessment of their biomechanical properties as well as their surgical handling. Human conjunctival epithelial cells cultured on plastic and plastic compressed collagen gels formed a confluent cell layer and expressed CK19. The cells showed expression of the putative epithelial progenitor cell markers p63α, ABCG2 and CK15 and sustained colony forming ability. The compressed collagen gels showed a high ultimate tensile strength and elasticity and the surgical handling of gels was comparable to amniotic membrane. An epithelialized conjunctival tissue construct on the basis of compressed collagen might therefore be a promising alternative bioartificial tissue substitute for conjunctival reconstruction. Copyright © 2015 John Wiley & Sons, Ltd.

3D culture model of fibroblast-mediated collagen creep to identify abnormal cell behaviour.

Native collagen gels are important biomimetic cell support scaffolds, and a plastic compression process can now be used to rapidly remove fluid to any required collagen density, producing strong 3D tissue-like models. This study aimed to measure the mechanical creep properties of such scaffolds and to quantify any enhanced creep occurring in the presence of cells (cell-mediated creep). The test rig developed applies constant creep tension during culture and measures real-time extension due to cell action. This was used to model extracellular matrix creep, implicated in the transversalis fascia (TF) in inguinal hernia. Experiments showed that at an applied tension equivalent to 15% break strength, cell-mediated creep over 24-h culture periods was identified at creep rates of 0.46 and 0.38%/h for normal TF and human dermal fibroblasts, respectively. However, hernia TF fibroblasts produced negligible cell-mediated creep levels under the same conditions. Raising the cell culture temperature from 4 to 37 °C was used to demonstrate live cell dependence of this creep. This represents the first in vitro demonstration of TF cell-mediated collagen creep and to our knowledge the first demonstration of a functional, hernia-related cell abnormality.