Cell population kinetics of collagen scaffolds in ex vivo oral wound repair.

Biodegradable collagen scaffolds are used clinically for oral soft tissue augmentation to support wound healing. This study sought to provide a novel ex vivo model for analyzing healing kinetics and gene expression of primary human gingival fibroblasts (hGF) within collagen scaffolds. Sponge type and gel type scaffolds with and without platelet-derived growth factor-BB (PDGF) were assessed in an hGF containing matrix. Morphology was evaluated with scanning electron microscopy, and hGF metabolic activity using MTT. We quantitated the population kinetics within the scaffolds based on cell density and distance from the scaffold border of DiI-labled hGFs over a two-week observation period. Gene expression was evaluated with gene array and qPCR. The sponge type scaffolds showed a porous morphology. Absolute cell number and distance was higher in sponge type scaffolds when compared to gel type scaffolds, in particular during the first week of observation. PDGF incorporated scaffolds increased cell numbers, distance, and formazan formation in the MTT assay. Gene expression dynamics revealed the induction of key genes associated with the generation of oral tissue. DKK1, CYR61, CTGF, TGFBR1 levels were increased and integrin ITGA2 levels were decreased in the sponge type scaffolds compared to the gel type scaffold. The results suggest that this novel model of oral wound healing provides insights into population kinetics and gene expression dynamics of biodegradable scaffolds.

Soft tissue volume augmentation by the use of collagen-based matrices in the dog mandible -- a histological analysis.

OBJECTIVES: The aim was to test, whether or not soft tissue volume augmentation with a specifically designed collagen matrix (CM), leads to ridge width gain in chronic ridge defects similar to those obtained by an autogenous subepithelial connective tissue graft (SCTG). MATERIAL AND METHODS: In six dogs, soft tissue volume augmentation was performed by randomly allocating three treatment modalities to chronic ridge defects [CM, SCTG and sham-operated control (Control)]. Dogs were sacrificed at 28 (n = 3) and 84 days (n = 3). Descriptive histology and histomorphometric measurements were performed on non-decalcified sections. RESULTS: SCTG and CM demonstrated favourable tissue integration, and subsequent re-modelling over 84 days. The overall mean amount of newly formed soft tissue (NMT) plus bone (NB) amounted to 3.8 ± 1.2 mm (Control), 6.4 ± 0.9 mm (CM) and 7.2 ± 1.2 mm (SCTG) at 28 days. At 84 days, the mean NMT plus NB reached 2.4 ± 0.9 mm (Control), 5.6 ± 1.5 mm (CM) and 6.0 ± 2.1 mm (SCTG). Statistically significant differences were observed between CM/SCTG and Control at both time-points (p < 0.05). CONCLUSION: Within the limits of this animal model, the CM performed similar to the SCTG, based on histomorphometric outcomes combining NB and NMT.

Evaluation of the tissue reaction to a new bilayered collagen matrix in vivo and its translation to the clinic.

This study evaluates a new collagen matrix that is designed with a bilayered structure in order to promote guided tissue regeneration and integration within the host tissue. This material induced a mild tissue reaction when assessed in a murine model and was well integrated within the host tissue, persisting in the implantation bed throughout the in vivo study. A more porous layer was rapidly infiltrated by host mesenchymal cells, while a layer designed to be a barrier allowed cell attachment and host tissue integration, but at the same time remained impermeable to invading cells for the first 30 days of the study. The tissue reaction was favorable, and unlike a typical foreign body response, did not include the presence of multinucleated giant cells, lymphocytes, or granulation tissue. In the context of translation, we show preliminary results from the clinical use of this biomaterial applied to soft tissue regeneration in the treatment of gingival tissue recession and exposed roots of human teeth. Such a condition would greatly benefit from guided tissue regeneration strategies. Our findings demonstrate that this material successfully promoted the ingrowth of gingival tissue and reversed gingival tissue recession. Of particular importance is the fact that the histological evidence from these human studies corroborates our findings in the murine model, with the barrier layer preventing unspecific tissue ingrowth, as the scaffold becomes infiltrated by mesenchymal cells from adjacent tissue into the porous layer. Also in the clinical situation no multinucleated giant cells, no granulation tissue and no evidence of a marked inflammatory response were observed. In conclusion, this bilayered matrix elicits a favorable tissue reaction, demonstrates potential as a barrier for preferential tissue ingrowth, and achieves a desirable therapeutic result when applied in humans for soft tissue regeneration.

A bioreactor test system to mimic the biological and mechanical environment of oral soft tissues and to evaluate substitutes for connective tissue grafts.

Gingival cells of the oral connective tissue are exposed to complex mechanical forces during mastication, speech, tooth movement and orthodontic treatments. Especially during wound healing following surgical procedures, internal and external forces may occur, creating pressure upon the newly formed tissue. This clinical situation has to be considered when developing biomaterials to augment soft tissue in the oral cavity. In order to pre-evaluate a collagen sponge intended to serve as a substitute for autogenous connective tissue grafts (CTGs), a dynamic bioreactor system was developed. Pressure and shear forces can be applied in this bioreactor in addition to a constant medium perfusion to cell-material constructs. Three-dimensional volume changes and stiffness of the matrices were analyzed. In addition, cell responses such as cell vitality and extracellular matrix (ECM) production were investigated. The number of metabolic active cells constantly increased under fully dynamic culture conditions. The sponges remained elastic even after mechanical forces were applied for 14 days. Analysis of collagen type I and fibronectin revealed a statistically significant accumulation of these ECM molecules (P < 0.05-0.001) when compared to static cultures. An increased expression of tenascin-c, indicating tissue remodeling processes, was observed under dynamic conditions only. The results indicate that the tested in vitro cell culture system was able to mimic both the biological and mechanical environments of the clinical situation in a healing wound.

Soft tissue volume augmentation by the use of collagen-based matrices: a volumetric analysis.

OBJECTIVES: The aim was to test whether or not soft tissue augmentation with a newly developed collagen matrix (CM) leads to volume gain in chronic ridge defects similar to those obtained by an autogenous subepithelial connective tissue graft (SCTG). MATERIAL AND METHODS: In six dogs, soft tissue volume augmentation was performed by randomly allocating three treatment modalities to chronic ridge defects (CM, SCTG, sham-operated control). Impressions were taken before augmentation (baseline), at 28, and 84 days. The obtained casts were optically scanned and the images were digitally analysed. A defined region of interest was measured in all sites and the volume differences between the time points were calculated. RESULTS: The mean volume differences per area between baseline and 28 days amounted to a gain of 1.6 mm (CM; SD+/-0.9), 1.5 mm (SCTG; +/-0.1), and a loss of 0.003 mm (control; +/-0.3). At 84 days, the mean volume differences per area to baseline measured a gain of 1.4 mm (CM; +/-1.1), 1.4 mm (SCTG; +/-0.4), and a loss of 0.3 mm (control; +/-0.3). The differences between CM and SCTG were statistically significant compared with control at 28 and 84 days (p<0.001). CONCLUSION: Within the limits of this animal study, the CM may serve as a replacement for autogenous connective tissue.

Neuronal nitric oxide synthase inhibitor has a neuroprotective effect in a rat model of brain injury.

Purpose: The aim of the present study was to assess the effects of neuronal nitric oxide synthase (NOS I) inhibitors and a combination of NOS I and NOS II inhibitors on lesion volume after experimental brain injury. Methods: Cold lesion of the brain was induced by application of a precooled (-78 degrees C) copper cylinder to the intact dura of the rat for 6 s. Brains were removed 24 h after the injury and lesion volume determined using the triphenyltetrazolium-chloride method. Results: The specific NOS I inhibitor 3-bromo-7-nitroindazole (Br-7-NI) reduced lesion volume significantly by 21 % compared with the vehicle control. In contrast, 7-nitroindazole had no effect on lesion volume. When aminoguanidine, a specific NOS II inhibitor, was adminis-tered after Br-7-NI, lesion volume was significantly reduced but not significantly more than with either compound alone. Conclusion: Brain injury after cold lesion is partly mediated by NOS I activity and can be attenuated successfully with Br-7-NI, while coin-hibition of NOS II does not improve the outcome significantly.