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P. Coutinho, C. Qiu, S. Frank, C. M. Wang, T. Brown, C. R. Green, D. L

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Presentation on theme: "P. Coutinho, C. Qiu, S. Frank, C. M. Wang, T. Brown, C. R. Green, D. L"— Presentation transcript:

1 Limiting burn extension by transient inhibition of Connexin43 expression at the site of injury 
P. Coutinho, C. Qiu, S. Frank, C.M. Wang, T. Brown, C.R. Green, D.L. Becker  British Journal of Plastic Surgery  Volume 58, Issue 5, Pages (July 2005) DOI: /j.bjps Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions

2 Figure 1 Antisense reduces Cx43 protein expression. While Cx43 puncta (green) are absent in the blister region (b) of control and treated wounds at 1d pw, they are present at the wound edge (we) (a, b). Levels of Cx43 in this region are higher in controls wounds (a), than in treated wounds (b). At 2d pw, when levels of Cx43 have increased in the wound epidermis, levels of Cx43 are still slightly higher in control wounds (c), when compared to treated wounds (d). At 4d pw, no difference in Cx43 expression is detected between control (e) and treated wounds (f), suggesting that the antisense is no longer effective at this time. The eschar (e) on the wound surface is indicated (d, e and f). Scale bar=50μm. British Journal of Plastic Surgery  , DOI: ( /j.bjps ) Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions

3 Figure 2 Macroscopic appearance of the progress wound healing. Within 6h a distinct difference can be seen macroscopically, treatment with Cx43 antisense (AS) significantly reduces the size and area covered by the blister and amount of fluid it contains compared to sense (S) controls. At 4 days the tissue in centre of the control wounds is clearly dying and turning into eschar whereas this is not so apparent following antisense treatment where the tissue appears to be healthier. Once the eschar forms little difference can be seen through the scabs, but these fall off the treated wounds sooner leaving a smaller and flatter area of scar, which has a better appearance at both 12 and 22d. Scale bar 1mm. British Journal of Plastic Surgery  , DOI: ( /j.bjps ) Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions

4 Figure 3 Antisense treatment affects wound architecture. Within 1d pw, a noticeable difference is detected between control and treated wounds. The blister (b) is significantly smaller in treated wounds at 1d pw, compared to controls. From 1d pw onwards, arrows point to the leading edge of the migrating wound epidermis. By 4d pw, the leading edges in treated wounds have migrated significantly, while less migration has occurred in the control wounds, which appears to have more extensive damage. Wound closure is almost complete by 7d in treated wounds, while a significant degree of re-epithelialisation is still required in controls. At 12d, wound closure is complete in treated wounds in contrast to controls. In addition, the eschar has detached and the wound region is marked by a small area of granulation tissue. In controls, a large granulation tissue area is apparent and the eschar (e) is still firmly attached to the surface of the skin. Images selected for this figure are typical for the time points examined. Scale bars=200μm. British Journal of Plastic Surgery  , DOI: ( /j.bjps ) Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions

5 Figure 4 Reduced neutrophil infiltration in Cx43 antisense treated wounds. Neutrophil numbers counted in a standardised deep dermal wound region of sense and antisense treated wounds, from 6h to 2d pw. Numbers of neutrophils were reduced in the treated wounds compared to controls at early injury time points (6h, 1d pw). However, at the last time point (2d pw), neutrophil numbers were increased in treated wounds, compared to controls. The rise in neutrophil numbers at this time may compensate for the reduced numbers at early time points. The data represent the mean±SEM values of five animals. The statistical significance of the data (*) was analysed using the Wilcoxon signed rank correlation test, with a criteria level of p<0.05. British Journal of Plastic Surgery  , DOI: ( /j.bjps ) Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions

6 Figure 5 Cx43 antisense treatment accelerates wound re-epithelialisation. A schematic diagram illustrates the method of measurement for re-epithelialisation in burn wounds (A). The red dotted line indicates the length of wound bed, and the blue dotted line from the leading edge of the epidermis to the end of thickened epidermis, indicates the re-epithelialised region. A histogram shows the comparison of re-epithelialisation rates in Cx43 sense and antisense treated wounds from 1 to 12d post-wounding (B). The histogram shows that antisense treated wounds have accelerated rates of re-epithelialisation compared to controls. Values shown in the histogram are mean±SEM values for five animals. The statistical significance of the data (*p<0.05) is shown. British Journal of Plastic Surgery  , DOI: ( /j.bjps ) Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions

7 Figure 6 Cx43 antisense treatment increases epidermal proliferation. A schematic diagram shows a typical wound at 4–7d pw (A) and 12d pw (B) in a control wound. While wound closure is far from complete at 4–7d, re-epithelialisation is almost complete by 12d. The eschar is shown firmly attached to the wound surface at 4–7d, but almost detached at 12d. The dotted lines mark the injured region and the locations of the black boxes in the proliferation zone and peripheral wound region indicate the areas in which counts were made. Histograms show the distribution of Ki67 positive cells in these areas. In the proliferation zone, increased numbers of Ki67 positive cells were detected in antisense treated wounds from 4 to 12d pw (C). However, in the peripheral wound region, an increased level of proliferation was found in treated wounds at early time points and was followed by a reduction in levels compared to controls at later times. Values shown in the histogram are mean±SEM for five animals. The statistical significance of the data (*) is shown for a criteria level of p<0.05. British Journal of Plastic Surgery  , DOI: ( /j.bjps ) Copyright © 2005 The British Association of Plastic Surgeons Terms and Conditions


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