Analysis of the Optimum Tapering Angle in Microanastomosis Using Computational Fluid Dynamics

Background: In free flap transfer, size discrepancy between the vascular pedicle and recipient vessel can create a problem for microsurgeons and sometimes induces postoperative thrombus formation. When there is a major difference between the diameters of the vascular pedicle and the recipient vessel, the larger vessel is often tapered to perform the anastomosis properly. However, the decision on the tapering angle used depends mostly on the operator’s experience. In this study, computational fluid dynamics (CFD) was used to investigate the optimum tapering angle. Methods: Using ANSYS ICEM 16.0 (ANSYS Japan, Tokyo, Japan), simulated vessels of diameters 1.5 mm and 3.0 mm were designed and then used to produce four anastomosis models with the 3.0-mm vessel tapered at angles of 15º, 30º, 60º, and 90º (no tapering). Venous perfusion with a mean value of 13.0 mL/min was simulated, and this was passed through the four anastomosis models in both the forward direction (F), from the smaller to the larger vessel, and the retrograde direction (R), from the larger to the smaller vessel. The velocity, wall shear stress (WSS), and oscillatory shear index (OSI) were measured in these eight patterns and then analyzed using OpenFOAM version 5. Results: The decrease in velocity was limiting. The WSS was greater in the R direction than the F direction at every tapering angle. The OSI also tended to be almost the same in the F direction, and lower at smaller tapering angles in the R direction. And, it was greater in the F direction than in the R direction at every tapering angle. The OSI values for 15º and 30º were almost identical in the R direction. Conclusion: The risk of thrombus formation is thought to be lower when tapering is used for anastomosis if the direction of flow is from the larger to the smaller vessel, rather than vice versa. These results also suggest that the optimum tapering angle is approximately 30º in both directions

Protein Crosslinking by Transglutaminase Controls Cuticle Morphogenesis in Drosophila

Transglutaminase (TG) plays important and diverse roles in mammals, such as blood coagulation and formation of the skin barrier, by catalyzing protein crosslinking. In invertebrates, TG is known to be involved in immobilization of invading pathogens at sites of injury. Here we demonstrate that Drosophila TG is an important enzyme for cuticle morphogenesis. Although TG activity was undetectable before the second instar larval stage, it dramatically increased in the third instar larval stage. RNA interference (RNAi) of the TG gene caused a pupal semi-lethal phenotype and abnormal morphology. Furthermore, TG-RNAi flies showed a significantly shorter life span than their counterparts, and approximately 90% of flies died within 30 days after eclosion. Stage-specific TG-RNAi before the third instar larval stage resulted in cuticle abnormality, but the TG-RNAi after the late pupal stage did not, indicating that TG plays a key role at or before the early pupal stage. Immediately following eclosion, acid-extractable protein from wild-type wings was nearly all converted to non-extractable protein due to wing maturation, whereas several proteins remained acid-extractable in the mature wings of TG-RNAi flies. We identified four proteins—two cuticular chitin-binding proteins, larval serum protein 2, and a putative C-type lectin—as TG substrates. RNAi of their corresponding genes caused a lethal phenotype or cuticle abnormality. Our results indicate that TG-dependent protein crosslinking in Drosophila plays a key role in cuticle morphogenesis and sclerotization

A soluble form of human nectin-2 impairs exocrine secretion of pancreas and formation of zymogen granules in transgenic mice

Transgenic mouse lines expressing a soluble form of human nectin-2 (hNectin-2Ig Tg) exhibited distinctive elevation of amylase and lipase levels in the sera. In this study, we aimed to clarify the histopathology and to propose the transgenic mouse lines as new animal model for characteristic pancreatic exocrine defects. The significant increase of amylase and lipase levels in sera of the transgenic lines approximately peaked at 8 weeks old and thereafter, plateaued or gradually decreased. The histopathology in transgenic acinar cells was characterized by intracytoplasmic accumulation of abnormal proteins with decrease of normal zymogen granules. The hNectin-2Ig expression was observed in the cytoplasm of pancreatic acinar cells, which was consistent with zymogen granules. However, signals of hNectin-2Ig were very weak in the transgenic acinar cells with the abnormal cytoplasmic accumulaion. The PCNA-positive cells increased in the transgenic pancreas, which suggested the affected acinar cells were regenerated. Acinar cells of hNectin-2Ig Tg had markedly small number of zymogen granules with remarkable dilation of the endoplasmic reticulum (ER) lumen containing abundant abnormal proteins. In conclusion, hNectin-2Ig Tg is proposed as a new animal model for characteristic pancreatic exocrine defects, which are due to the ER stress induced by expression of mutated cell adhesion molecule that is a soluble form of human nectin-2

Comparison of Antera 3D® and TcPO2 for Evaluation of Blood Flow in Skin

Background: There is a need for quick skin blood flow tests that can be performed in the wound healing field. Antera 3D® is a compact scanner using multispectral imaging. It can perform quick assessment of skin conditions. The purpose of the present study was to investigate the ability of the Antera 3D® to assess skin blood flow in comparison with transcutaneous partial pressure of oxygen (TcPO2) measurements. Methods: This study was conducted on 13 patients with a history of lower extremity ulcers. Measurements of hemoglobin average level (hereafter, Hb score) measured by Antera 3D® and TcPO2 measured by a transcutaneous blood gas monitor were obtained at the same sites on the dorsal foot and lower leg. Differences in Hb score and TcPO2 were analyzed by t-test for each measurement site and for the presence of peripheral arterial disease (PAD). The correlation between TcPO2 and Hb score was analyzed by Pearson’s correlation coefficient. Results: Twenty-four limbs were tested. Hb score was higher (P < 0.001) and TcPO2 was lower (P = 0.056) in the dorsal foot compared to the lower leg. In the dorsal foot, Hb score was higher (P = 0.023) and TcPO2 was lower (P = 0.046) in patients with PAD compared to those without PAD. A significant negative correlation (r = –0.68; 95% confidence interval –0.85 to –0.38, P < 0.001) between TcPO2 and Hb score was observed in the dorsal foot. Conclusion: The negative correlation between TcPO2 and Hb score may reflect compensatory peripheral vasodilation due to occlusion or stenosis of central arteries. This study showed that Hb score measured by Antera 3D® may be related to skin blood flow

The life span of the <i>Da-GAL4>UAS-TG IR</i> flies.

<p>(A) The life span of the RNAi flies was compared with those of the control flies, <i>Da-GAL4>+</i>, <i>Da-GAL4>UAS-lacZ IR</i> and <i>+>UAS-TG IR</i>. Sixty adult flies were collected and maintained at 25°C. The number of surviving flies was recorded daily. The means ± S. D. of four independent experiments were plotted. <i>Da</i>, <i>Da-GAL4</i>. (B) Phenotypes of <i>Tub-GAL80^ts; Da-GAL4>UAS-TG IR</i>. <i>Tub-GAL80^ts; Da-GAL4</i> flies were crossed with the <i>UAS-TG IR</i> flies in 20 vials and maintained at 18°C. The suppression of <i>TG</i> by RNAi was triggered by increasing the temperature to 29°C. The ratios flies with abnormal wings (square) and abnormal abdominal cuticles (circle) to total adult flies are indicated (upper panel). The number of adult flies born from each vial is indicated (lower panel).</p

Identification of TG substrates associated with cuticle formation.

<p>The wings of wild-type and <i>Da-GAL4>UAS-TG IR</i> flies were collected at indicated times after eclosion. Wing proteins were extracted and subjected to SDS-PAGE. TG antigen was detected by Western blotting (upper panels). Loaded proteins were stained with Coomassie Brilliant Blue R-250 (lower panels). <i>Da, Da-GAL4.</i></p

Acid-extractable wing proteins from <i>TG</i>-RNAi flies identified by mass spectrometry.

<p>RFABP, retinoid and fatty acid binding protein; NP, no phenotypic difference observed.</p

Binding of Cpr97Eb and Clect27 proteins to chitin.

<p>Cpr97Eb and Clect27 proteins were mixed with chitin, and unbound (UB) and bound (B) fractions were subjected to SDS-PAGE. The bound fraction was eluted by 2% SDS. Z2A was used as negative control for chitin binding. Proteins were detected by Coomassie Brilliant Blue R-250 staining (Clect27 and Z2A) or by anti-6×His tag antibody (Cpr97Eb).</p

Phenotypes of TG substrate RNAi flies.

<p>Phenotypes of the <i>MS1096-GAL4>UAS-Cpr97Eb IR</i> (A), <i>MS1096-GAL4>UAS-Clect27 IR</i> (B), <i>Da-GAL4>UAS-LSP2 IR</i> (D) and <i>Da-GAL4>UAS-Cpr76Bd IR</i> (E) flies. The control flies, <i>MS-GAL4</i>>+ (C) and <i>Da-GAL4</i>>+ (F), are also indicated. Each fly was laid at 25°C. <i>MS, MS1096-GAL4; Da, Da-GAL4</i>.</p