8 0 nylon suture

Ligation surgeries were performed on left and right side of C57BL/6 mice as described above and previously^{43 (link)}. Immediately after severing the epigastric artery, a one millimeter FTY720-loaded or unloaded PLAGA thin film was placed immediately adjacent to the collateral vessel entering the inguinal fat pad. The thin film was placed underneath the fascial layer so as to minimize displacement away from the inguinal fat pad and region of interest (Figure 4A,B). The left side of the mouse had the FTY720 loaded PLAGA film while the right side had an unloaded PLAGA film. Sham surgeries were performed on separate mice on both the left and right side of C57BL/6 mice. Again, the left side had the drug loaded thin film and the right side had the unloaded thin polymer film. Incisions were closed with 8-0 nylon suture (Ethicon, Somerville, NJ). The process described above was also repeated with sham surgeries on the left and right side in a separate set of experiments. These procedures were repeated in three mice (n = 3).

Eight-10 week old female C57B6l/6J mice were obtained from Jackson Laboratories (Bar Harbor, ME). Animals were housed at up to 5 mice per cage with ad lib food and water and a 12-hour light-dark cycle. Briefly, the distal FDL was isolated in the right hind-paw. The tendon was lacerated and repaired with 8-0 nylon suture (Ethicon Inc., Summerville, NJ) using a modified Kessler technique [20 (link), 21 (link)]. The skin incisions were closed using 5-0 nylon suture (Ethicon Inc., Summerville, NJ). To protect the distal repair, the FDL was released at the proximal myotendinous junction. Mice were randomly assigned to vehicle or Mmp9 inhibitor treatment (Supplemental Table 1). The Mmp9 activity inhibitor Ro 32-3555 (R&D Systems, Minneapolis, MN) was diluted in normal saline and administered via i.p. injection (10mg/kg) [22 (link)]. Mice received a dose every 24 hours starting immediately following surgery and continuing through day 8 post-surgery. The vehicle treated group received daily i.p. injections of saline.

Animal care and treatment were conducted in conformity with approved protocols and animal welfare regulations of Augusta University Institutional IACUC Committees. All animal procedures conformed to the NIH guidelines. C57BL/6 mice (2-3-month-old) were purchased from Jackson Laboratory (Bar Harbor, ME) and maintained under controlled environmental conditions. Mice were anesthetized with an intraperitoneal injection of 100 mg/kg ketamine and 10 mg/kg xylazine. The mice were orally intubated with a 24 gauge tube and ventilated with room air using a Harvard Rodent Ventilator (Model 55-7058, Holliston, MA). The thorax was opened by a lateral thoracotomy, and the heart was exposed by a pericardial incision. An 8-0 nylon suture (Ethicon, Somerville, NJ) was placed under the left coronary artery and then threaded through a small plastic PE10 tubing to form a snare for reversible left coronary artery occlusion. The left coronary artery was occluded for 45 min, followed by reperfusion by removing the tubing. The chest was closed and the mice allowed to recover. A sham-operated group consisted of mice that underwent the thoracotomy procedure but were not subjected to ischemia and reperfusion. Animals were sacrificed at the indicated times after reperfusion.

Partial ligation of the L5 spinal nerve was performed as previously described in 7 week aged rats.^{13 (link)} Animals were anesthetized with 1.75-3% isoflurane in oxygen. A 2.5 cm incision was made lateral to the lumbar spine and the right L6-transverse process was removed to expose the L5 spinal nerve. One third of the L5 spinal nerve was ligated with 8-0 nylon suture (Ethicon, Cincinnati, OH, USA) under a dissecting microscope. To avoid paralysis, the L4 spinal nerve was left untouched. The skin was closed with 5-0 suture and treated with a topical antibiotic (neomycin, polymyxin and bacitracin, Johnson & Johnson, New Brunswick, NJ, USA). After surgery and emergence from anesthesia, animals were returned to paired housing. Animals were weighed regularly for 1 week following surgery and a 30-40 g weight gain was considered evidence of normal surgical recovery.

To evaluate the effect of C-MSC-Exo on cardiac repair after induction of
myocardial infarction, mice were subjected to acute MI as previously
described(38 (link), 39 (link)). Briefly, C57BL/6 mice were anesthetized with an
intraperitoneal injection of 100 mg/kg ketamine and 10 mg/kg xylazine. The mice
were orally intubated with a 24 gauge tube and ventilated with room air using a
Harvard Rodent Ventilator (Model 55–7058, Holliston, MA). The thorax was
opened by a lateral thoracotomy and the heart was exposed by a pericardial
incision. An 8–0 nylon suture (Ethicon, Somerville, NJ) was placed under
the left anterior inferior artery (LAD) for permanent ligation. Immediately
after coronary occlusion, mice were injected intramuscularly with 30 μl
PBS or C-MSC-Exo (50 μg, 30 μl) in the infarct border zone. The
chest was closed by layers, and the mice were allowed to recover. Animals were
sacrificed one month after LAD ligation for tissue harvesting and histological
assay. Animals were treated according to approved protocols and animal welfare
regulations of the Institutional Animal Care and Use Committee of the Medical
College of Georgia.

To assess the efficacy of clodronate liposome-mediated depletion of macrophages within the inguinal fat pad, we performed direct injection of clodronate loaded liposomes and control liposomes (PBS-loaded liposomes) into the inguinal fat pad. A one centimeter incision was made above the epigastric artery and blunt dissection was used to locate the collateral vessel entering the side of the inguinal fat pad. Using a 27G syringe (BD Soloshot™ 301793, Franklin Lakes, NJ), 100 microliters of clodronate-loaded liposomes and control liposomes (ClodronateLiposomes.org, Haarlem, The Netherlands) were injected directly into the inguinal fat pad at the insertion site of the collateral vessel. Successful injection was confirmed by visual observation of a bubble formed within the inguinal fat pad. Clodronate liposomes were injected into the right inguinal fat pad of the mouse, and control liposomes were injected into the left inguinal fat pad. After injection the incision was closed with 8-0 nylon suture (Ethicon).

Male C57/BL6, aged 10 to 11 weeks, were anaesthetized and intubated as described above. A left lateral thoracotomy was then performed. Pericardium was gently pulled off and an 8-0 Nylon suture (Ethicon, Inc. Johnson & Johnson Co., USA) was used to ligate the left anterior descending coronary artery against a PE10 tubing, which was removed after 30 min occlusion. Successful performance of coronary artery occlusion was verified by visual inspection (by noting the development of a pale colour in the distal myocardium upon ligation). The chest was then closed using 7–0 sutures around adjacent ribs, and the skin was closed with 6–0 suture. Buprenorphine was administered subcutaneously for a minimum of 1 day at BID dosing. For the animal group treated with patch, a second thoracotomy was performed one week post the incidence of I/R and the prepared collagen patch was sutured (at two points) onto the surface of ischaemic myocardium. Sham-operated controls consisted of age-matched mice that underwent identical surgical procedures (two thoracotomies) with the exception of LAD ligation (Extended Data Fig. 4).