Fig 1.
Flow chart summarizing the experimental protocol.
Fig 2.
Template adapted from the DIN 50125 with standardized dimensions for the esophagus, ureter and skin samples.
Fig 3.
Osmotic stress protocol with different polyethylengylcol (PEG) concentrations for 3A) esophagus 3B) ureter 3C) skin samples.
Fig 4.
Boxplot from esophagus, ureter and skin samples in the native and acellular condition 4A) cross section 4B) elastic modulus 4C) ultimate tensile stress 4D) maximum strain.
Table 1.
Mean and standard deviation with the p value for the comparison native versus acellular condition.
Table 2.
P value for the comparison between the different tissues.
Fig 5.
Hematoxylin-eosin stained-samples of 5A) native esophagus 5B) acellular esophagus 5C) native ureter 5D) acellular ureter 5E) native skin 5F) acellular skin.
In spite of the removal of the cellular structures the respective tissue layers remained intact structurally. A marked thinning was observed in the tunica media of the esophagi and some thinning in the ureters. In the acellular skin samples the epidermis was completely removed. Black rectangles indicate the regions where samples for electron microscopy were obtained (also refer to Figs 9 to 13). Scale bar: 300 μm (5A,B), 100 μm (5C,D), 300 μm (5E,F).
Fig 6.
Anti-type I collagen staining of 6A) native esophagus 6B) acellular esophagus 6C) native ureter 6D) acellular ureter 6E) native skin 6F) acellular skin.
Collagens were observed throughout the native and acellular scaffolds but appeared to be more condensed in the latter, especially in the tunica muscularis (6B,D) and in the subepidermal regions (6F). Scale bar 100 μm.
Fig 7.
Anti-elastin staining of 7A) native esophagus 7B) acellular esophagus 7C) native ureter 7D) acellular ureter 7E) native skin 7F) acellular skin.
Elastic fibers were observed in the submucosal regions, the tunica muscularis, to some extent in the adventitia and in blood vessels (7A-D). The skin samples showed low quantities of elastic fibers, mostly in the vessels and dermal perivascular regions (Fig 7E,F). Scale bar 100 μm.
Fig 8.
Anti-fibronectin staining of 8A) native esophagus 8B) acellular esophagus 8C) native ureter 8D) acellular ureter 8E) native skin 8F) acellular skin.
Fibronectins were found throughout the ureters in the native (8A,C) and the acellular condition (8B,D), especially in the muscosal and muscular layers. A marked fibronectin washout was observed in the acellular (8F) compared to the native skin (8E) samples, especially in the dermal regions. Scale bar 100 μm.
Fig 9.
Scanning electron microscopy of 9A) native esophagus 9B) acellular esophagus 9C) native ureter 9D) acellular ureter 9E) native skin 9F) acellular skin.
Type I collagens appeared remain largely intact in the 1000x magnification without noticeable major direction in the tunica muscularis of the esophagi and the ureters but in the dermal skin layer. Scale bar 10 μm.
Fig 10.
Scanning electron microscopy of 10A) native esophagus 10B) acellular esophagus 10C) native ureter 10D) acellular ureter 10E) native skin 10F) acellular skin.
In the higher 10,000x magnification type I collagen clotting was observed (interrupted circles) accompanied by a loss of cross-linking collagens. Scale bar 1 μm.
Fig 11.
Scanning electron microscopy of the dermal areas of 11A) native skin 11B) acellular skin.
Intact fibrils were observed at the 50,000x-magnification with the collagen-characteristic D-period at a 67-nm distance (white arrows), resembling the findings from transmission electron microscopy. Scale bar 150 nm.
Fig 12.
An emptied hair follicle root sheath is depicted in the subepidermal region of porcine skin.
The surrounding tissue appears to be loosened and may be indicative of notching effects in skin samples. Scale bar 5 μm (1000x magnification).
Fig 13.
Transmission electron microscopy of 13A-D esophagi (tunica muscularis), 13E-H ureters (tunica muscularis), 13I-L skin (dermis) with focus on type I collagens.
Left column (13A,C,E,G,I,K): native condition, right column (13B,D,F,H,J,L): acellular condition. Intact collagen fibers and fibrils were observed throughout the native and acellular scaffolds with the collagen-characteristic D-period at a 67-nm distance. Scale bar 1250 nm (13A,B,E,F,I,J; magnification 9700x) and 200 nm (13C,D,G,H,K,L; magnification 66,000x).