4 0 silk suture

Under isoflurane anesthesia, each rat (n = 37) was subjected to a laparotomy that exposed the left uterine horn. A 20 mm × 10 mm full-thickness antemesometrial uterus wall segment was excised from the uterus horn leaving the blood vessel filled uterus mesometrium intact. The segment was replaced by an enzyme treated recellularized scaffold (n = 19; Group 1; Figure 1(C)) or with an enzyme treated acellular scaffold (n = 18; Group 2). Each graft was sutured using 6-0 non-absorbable polypropylene (Ethicon, Raritan, USA) by continuous sutures along the mesodermal side, while interrupted sutures were used for the proximal and distal anastomosis sites (Figure 1(D)). The abdominal muscle layer was then closed with 4-0 silk sutures (Ethicon) and the skin with titanium clips (Reflex7, Gaithersburg, USA). All animals received analgesics and antibiotics post-surgically (buprenorphine 0.05 mg/kg; carprofen 5 mg/kg; sulfamethoxazole 100 mg/kg; trimethoprim 20 mg/kg). Transplanted scaffolds in Group 1 and 2 were evaluated after euthanasia 14 days (n = 7/group), 30 days (n = 6/group), and 120 days (Group 1, n = 6; Group 2, n = 5) after transplantation. The spleen from every animal at every time point was also explanted and weighed.

Abcc6−/− and WT mice were unilaterally nephrectomised under isoflurane inhalation anaesthesia (2.2–2.6% isoflurane in 100% O₂). Hind-paw withdrawal, blink reflex and respiratory rate were monitored in these mice. A 1.5-cm incision was made through the skin and abdominal muscle caudal to the rib cage. The renal artery and vein were ligated with 4-0 silk sutures (Ethicon) and the left kidney was removed. Skin and muscle layers were closed separately with 4-0 silk sutures. In addition, a 1-cm incision was made in the back, at the base of the neck to implant DOCA pellets subcutaneously (Innovative Research of America, Sarasota, Florida, USA), to provide a dose of 1 mg/kg/day. Mice were then administered water containing 0.9% NaCl and 0.2% KCl. Upon recovery, they were monitored each day for 19 days to measure SBP, DBP and HR using tail-cuff plethysmography⁴⁸ .

Prior to the study, the mice were adapted to the maintenance environment for 1 week. Thirty mice were then divided into the sham group, the model group, and the LIFU treatment group, with 10 mice in each group. Then, we used the CCI surgical method [24 (link)] to establish CNP models. Specifically, the sciatic nerve in the CCI group and the treatment group was ligated for 90 days. The mice were anesthetized with isoflurane (Sigma-Aldrich, St. Louis, Missouri, USA) and laid on a heating pad. In the prone position, the surgical site was prepared by shaving the posterolateral side of the right hind limb, and then, three applications of 75% ethanol were applied to the site. An incision (1 cm) was then made proximal to the right hind limb. The nerve was then uncovered using a splitting approach on the bicep femoris, and three ligatures (gut ligatures 6.0, Jinhuang, Shanghai, China) were tied at 1 mm long intervals. The deep and superficial muscles were reapproximated by applying an interrupted stitching technique using 4.0 chromic gut (Stoelting Co, Wood Dale, IL). The skin was closed using 4.0 silk sutures (Ethicon, Somerville, NJ). For the sham group, the right sciatic nerve was exposed using the same methods, but the nerve was not ligated. After surgery, the mice were returned to their original cages to continue feeding.