Decellularized Bovine Pericardial Patch Loaded With Mesenchymal Stromal Cells Enhance the Mechanical Strength and Biological Healing of Large-to-Massive Rotator Cuff Tear in a Rat Model.

PURPOSE: The purpose of this study was to determine whether the addition of decellularized bovine pericardial patch loaded with mesenchymal stromal cells enhanced bone-to-tendon healing and improved the biomechanical strength of large-to-massive rotator cuff tears in a small animal model. METHODS: Adipose-derived mesenchymal stromal cells (MSCs) from rat inguinal fat were isolated, cultured, and loaded onto decellularized bovine pericardium patches. To simulate large-to-massive tears, rats were managed with free cage activity for 6 weeks after tear creation. A total of 18 rats were randomly allocated to repair-only (control), repair with pericardial patch augmentation (patch), or repair with MSC loaded pericardial patch augmentation (patch-MSC). Each group had 6 rats (one shoulder of each rat was used for histological evaluation and another for biomechanical evaluation). MSCs seeded on the pericardial patches were traced on four shoulders from 2 other rats at 4 weeks after surgery. Histological evaluation for bone-to-tendon healing and biomechanical testing was carried out at 8 weeks after repair. RESULTS: MSCs tagged with a green fluorescent protein were observed in the repair site 4 weeks after the repair. One shoulder each in the control and patch groups showed complete discontinuity between the bone and tendon. One shoulder in the control group showed attenuation with only a tenuous connection. Fibrocartilage and tidemark formation at the bone-to-tendon interface (P = .002) and collagen fiber density (P = .040) and orientation (P = .003) were better in the patch-MSC group than in the control or patch group. Load-to-failure in the patch-MSC and patch groups was higher than that in the control group (P = .001 and .009, respectively). CONCLUSION: Decellularized bovine pericardial patches loaded with adipose-derived and cultured mesenchymal stromal cells enhanced healing in terms of both histology and mechanical strength at 8 weeks following rotator cuff repair in a rat model. CLINICAL RELEVANCE: Large-to-massive rotator tears need a strategy to prevent retear and enhance healing. The addition of decellularized bovine pericardial patch loaded with MSCs can enhance bone-to-tendon healing and improve biomechanical healing of large-to-massive rotator cuff tears following repair.

Extracellular Vesicles Delivered by Injectable Collagen Promote Bone-Tendon Interface Healing and Prevent Fatty Degeneration of Rotator Cuff Muscle.

PURPOSE: This study aimed (1) to confirm the maintenance of the extracellular vesicles (EVs) delivered via injectable collagen at the application site, and (2) to evaluate the effect of EVs derived from the human umbilical cord-derived mesenchymal stem cells and loaded in an injectable collagen gel after rotator cuff repair (RCR). METHODS: Rabbits (n = 20) were assigned to normal (N), repair-only (R), and those administered with injectable collagen after repair (RC), and EV-laden injectable collagen after repair (RCE) groups. The EVs isolated by ultra-centrifugation from the human umbilical cord-derived mesenchymal stem cells spent medium were mixed with collagen and administered accordingly. After 12 weeks, the rabbits were sacrificed to evaluate the healing of the bone-to-tendon junction and the fatty degeneration of muscle. Histomorphometric scoring for bone-tendon interface, fatty infiltration (%), and biomechanical tests were performed. Separately, groups of 3 rabbits were assigned to 3 different time points to evaluate maintenance of green fluorescence-labeled EVs with injectable collagen via tracking on the bursal side of the rotator cuff (3 groups: 3 days, 2, and 4 weeks). RESULTS: The EVs delivered by injectable collagen remained until 4 weeks at the bursal side of the cuff tissue. The RCE group showed a significantly greater histomorphometric total score (P < .001, and P = .013, respectively) and significantly lower fatty degeneration than the RC and R groups (P = .001, and P = .013, respectively). The biomechanical tests revealed significant growing trends in load-to-failure and stiffness (P = .002, and P = .013, respectively), in the R, RC, RCE, and N groups. CONCLUSIONS: EVs mounted in injectable collagen remained at the repair site for at least 4 weeks after application. Furthermore, they effectively promote bone-to-tendon healing via collagen maturation in bone-tendon interface and preventing fatty degeneration of rotator muscle after RCR as compared with collagen-only or repair-only groups. CLINICAL RELEVANCE: The combination of collagen with EVs significantly promotes rotator cuff healing demonstrating potential clinical application during partial rotator cuff tear or after RCR.

Allogeneic epithelial cell sheet transplantation for preventing esophageal stricture after circumferential ESD in a porcine model: preliminary results.

OBJECTIVES: Circumferential endoscopic submucosal dissection (ESD) for large lesions induces severe stricture, requiring subsequent treatment. We aimed to evaluate the efficacy of allogeneic epithelial cell sheet transplantation in preventing esophageal stricture after circumferential ESD in a porcine model. MATERIALS AND METHODS: A total of 15 conventional pigs underwent a 4 cm long circumferential ESD in the mid-esophagus. Out of these animals, 11 were immediately subjected to allogeneic oral mucosal cell sheet transplantation at the resection site, whereas four pigs underwent circumferential ESD only. We performed upper endoscopy 1 and 2 weeks after ESD and assessed the degree of esophageal stricture and histologic characteristics. RESULTS: Dysphagia scores and weight change ratios recorded 1 and 2 weeks after ESD did not differ between the two groups. The stricture rate 2 weeks after ESD was 100% in the control group and 90.9% in the cell sheet group (p = 1.000). The median mucosal constriction rates of the control and cell sheet groups were 73.5% (range 63.0-80.0%) and 53.8% (37.5-73.3%, p = .018), respectively. With regard to microscopic measurements, the length of re-epithelialization was greater in the cell sheet group than in the control group (2,495 µm vs. 369 µm, p = .008). Median fibrosis thickness and degree of muscle damage were not significantly different between groups. CONCLUSIONS: Although allogeneic epithelial cell sheet transplantation showed greater re-epithelialization and less mucosal constriction of post-ESD ulcers, it was not sufficiently effective in preventing post-ESD stricture.

The effect of ionized collagen for preventing postoperative adhesion.

BACKGROUND: Collagen exhibits ideal multifactorial action for preventing tissue adhesions. This study examined the efficacy of ionized collagen in preventing tissue adhesion after surgical procedures. MATERIALS AND METHOD: Ionized collagen was prepared using the esterification technique of natural collagen. Three forms of collagen materials (membrane, film, and gel) were compared with three commercialized materials (oxidized regenerated cellulose membrane [OC membrane], hyaluronic acid and carboxymethylcellulose film, and gel [HC film and HC gel]) in a rat cecum abrasion model. Antiadhesive activity and histologic findings were assessed. RESULT: The incidence of adhesion was reduced significantly in all test groups compared to the sham-operated control group (100% in control, 14.3% in collagen membrane, 63.6% in collagen film, 25.0% in collagen gel, 55.6% in OC membrane, 75% in HC film, and 83.3% in HC gel). All collagen materials of the three forms exhibited a significant reduction in adhesion grade compared with the sham operation, whereas no significant difference was found among these three different forms. The collagen membrane showed significantly less adhesion grade, less inflammation and more regenerative features compared to widely used conventional materials. CONCLUSIONS: This preclinical investigation indicated that ionized collagen materials readily formed clinically suitable shapes for easy handling without the need for any complex processing and effectively reduced postoperative tissue adhesion profiles compared to conventional antiadhesive agents.

Collagen esterification enhances the function and survival of pancreatic ß cells in 2D and 3D culture systems.

Collagen, one of the most important components of the extracellular matrix (ECM), may play a role in the survival of pancreatic islet cells. In addition, chemical modifications that change the collagen charge profile to a net positive charge by esterification have been shown to increase the adhesion and proliferation of various cell types. The purpose of this study was to characterize and compare the effects of native collagen (NC) and esterified collagen (EC) on ß cell function and survival. After isolation by the collagenase digestion technique, rat islets were cultured with NC and EC in 2 dimensional (2D) and 3 dimensional (3D) environments for a long-term duration in vitro. The cells were assessed for islet adhesion, morphology, viability, glucose-induced insulin secretion, and mRNA expression of glucose metabolism-related genes, and visualized by scanning electron microscopy (SEM). Islet cells attached tightly in the NC group, but islet cell viability was similar in both the NC and EC groups. Glucose-stimulated insulin secretion was higher in the EC group than in the NC group in both 2D and 3D culture. Furthermore, the mRNA expression levels of glucokinase in the EC group were higher than those in the NC group and were associated with glucose metabolism and insulin secretion. Finally, SEM observation confirmed that islets had more intact component cells on EC sponges than on NC sponges. These results indicate that modification of collagen may offer opportunities to improve function and viability of islet cells.

Stem cell sheet interpositioned between the tendon and bone would be better for healing than stem cell sheet overlaid above the tendon-to-bone junction in rotator cuff repair of rats.

BACKGROUND: Although stem cells might enhance natural enthesis healing in surgical rotator cuff repair, not much attention has been given to the delivery and location of delivering stem cells. The purpose of this study to know where to locate those stem cells during repair. METHODS: Animal model of chronic rotator cuff tear was created in 24 rats. Adipose-derived stem cells were engineered as a sheet and transplanted 1) between a torn tendon and humerus (interposition group) or 2) over a repaired tendon-to-bone junction (overlay group) at the time of surgical repair. Tracking of stem cells with overexpression of green fluorescent protein (GFP) were carried out at the time of sacrifice in additional 4 shoulders in each group. Histological and Biomechanical evaluation was performed to compare the differences in tendon-to-bone healing. RESULTS: Histology showed increased fibrocartilage, a clear boundary at the mineralized fibrocartilage, abundant collagen type III, and higher total scores, especially in the interposition group. GFP-overexpression was observed at the transplanted site at 2 weeks after repair. Although two groups where stem cell sheets applied showed higher load to failure than the repair-only group, the load to failure was not different between the interposition and overlay group. CONCLUSION: In the chronic rotator cuff repair model, stem cell sheets enhanced regeneration of the tendon-to-bone junction. This regeneration was effective when the stem cell sheet was interpositioned at the tendon-to-bone interface. LEVEL OF EVIDENCE: Basic Science Study; In Vivo Animal Model; Histology and Biomechanics.

Enhanced Differentiation Capacity and Transplantation Efficacy of Insulin-Producing Cell Clusters from Human iPSCs Using Permeable Nanofibrous Microwell-Arrayed Membrane for Diabetes Treatment.

Although pancreatic islet transplantation is a potentially curative treatment for insulin-dependent diabetes, a shortage of donor sources, low differentiation capacity, and transplantation efficacy are major hurdles to overcome before becoming a standard therapy. Stem cell-derived insulin-producing cells (IPCs) are a potential approach to overcoming these limitations. To improve the differentiation capacity of the IPCs, cell cluster formation is crucial to mimic the 3D structure of the islet. This study developed a biodegradable polycaprolactone (PCL) electrospun nanofibrous (NF) microwell-arrayed membrane permeable to soluble factors. Based on the numerical analysis and experimental diffusion test, the NF microwell could provide sufficient nutrients, unlike an impermeable PDMS (polydimethylsiloxane) microwell. The IPC clusters in the NF microwells showed higher gene expression of insulin and PDX1 and insulin secretion than the PDMS microwells. The IPC clusters in the NF microwell-arrayed membrane could be directly transplanted. Transplanted IPC clusters in the microwells survived well and expressed PDX1 and insulin. Additionally, human c-peptide was identified in the blood plasma at two months after transplantation of the membranes. The NF microwell-arrayed membrane can be a new platform promoting IPC differentiation capacity and realizing an in situ transplantation technique for diabetic patients.

Dysfunctional pancreatic cells differentiated from induced pluripotent stem cells with mitochondrial DNA mutations.

Diabetes mellitus (DM) is a serious disease in which blood sugar levels rise abnormally because of failed insulin production or decreased insulin sensitivity. Although many studies are being conducted for the treatment or early diagnosis of DM, it is not fully understood how mitochondrial genome (mtDNA) abnormalities appear in patients with DM. Here, we induced iPSCs from fibroblasts, PBMCs, or pancreatic cells of three patients with type 2 DM (T2D) and three patients with non-diabetes counterpart. The mtDNA mutations were detected randomly without any tendency among tissues or patients. In T2D patients, 62% (21/34) of iPSC clones harbored multiple mtDNA mutations, of which 37% were homoplasmy at the 100% mutation level compared to only 8% in non-diabetes. We next selected iPSC clones that were a wild type or carried mutations and differentiated into pancreatic cells. Oxygen consumption rates were significantly lower in cells carrying mutant mtDNA. Additionally, the mutant cells exhibited decreased production of insulin and reduced secretion of insulin in response to glucose. Overall, the results suggest that screening mtDNA mutations in iPSCs from patients with T2D is an essential step before pancreatic cell differentiation for disease modeling or autologous cell therapy. [BMB Reports 2022; 55(9): 453-458].

The Outcomes and Quality of Pancreatic Islet Cells Isolated from Surgical Specimens for Research on Diabetes Mellitus.

Isolating a large quantity of high-quality human islets is a prerequisite for diabetes research. Human islets are typically isolated from the pancreases of brain-dead donors, making research difficult due to low availability. Pancreas tissue discarded after surgical resection may be a good alternative source of islet cells. To test this hypothesis, we isolated islets from discarded surgical specimens and evaluated the islet yield and quality as well as islet cell preparations. Eighty-two segmental pancreases were processed using the Ricordi automated method, and islet yield and quality were investigated. The mean age of patients was 54.6, and the cohort included 32 diabetes patients. After purification, partially resected pancreases yielded an average of 59,593 ± 56,651 islet equivalents (IEQs) and 2546 IEQ/g of digested pancreas, with 71.5 ± 21% purity. Multivariate analysis revealed that diabetes (p = 0.0046) and the lobe used (p = 0.0156) significantly altered islet yield. Islets transplanted into diabetic mice displayed good viability and in vitro glucose responses, DNA/RNA quality, mitochondrial function, and glucose control, even though these results were dependent on islet quality. Isolated cells also maintained high viability and function even after cryopreservation. Our findings indicate that pancreatic tissue discarded after surgery can be a valuable source of islets for diabetes research.

3D-Printed Coaxial Hydrogel Patches with Mussel-Inspired Elements for Prolonged Release of Gemcitabine.

With the aim of fabricating drug-loaded implantable patches, a 3D printing technique was employed to produce novel coaxial hydrogel patches. The core-section of these patches contained a dopamine-modified methacrylated alginate hydrogel loaded with a chemotherapeutic drug (Gemcitabine), while their shell section was solely comprised of a methacrylated alginate hydrogel. Subsequently, these patches were further modified with CaCO3 cross linker and a polylactic acid (PLA) coating to facilitate prolonged release of the drug. Consequently, the results showed that addition of CaCO3 to the formula enhanced the mechanical properties of the patches and significantly reduced their swelling ratio as compared to that for patches without CaCO3. Furthermore, addition of PLA coating to CaCO3-containing patches has further reduced their swelling ratio, which then significantly slowed down the release of Gemcitabine, to a point where 4-layered patches could release the drug over a period of 7 days in vitro. Remarkably, it was shown that 3-layered and 4-layered Gemcitabine loaded patches were successful in inhibiting pancreatic cancer cell growth for a period of 14 days when tested in vitro. Lastly, in vivo experiments showed that gemcitabine-loaded 4-layered patches were capable of reducing the tumor growth rate and caused no severe toxicity when tested in mice. Altogether, 3D printed hydrogel patches might be used as biocompatible implants for local delivery of drugs to diseased site, to either shrink the tumor or to prevent the tumor recurrence after resection.

Improvement of the therapeutic capacity of insulin-producing cells trans-differentiated from human liver cells using engineered cell sheet.

BACKGROUND: Although pancreatic islet transplantation therapy is ideal for diabetes patients, several hurdles have prevented it from becoming a standard treatment, including donor shortage and low engraftment efficacy. In this study, we prepared insulin-producing cells trans-differentiated from adult human liver cells as a new islet source. Also, cell sheet formation could improve differentiation efficiency and graft survival. METHODS: Liver cells were expanded in vitro and trans-differentiated to IPCs using adenovirus vectors carrying human genes for PDX1, NEUROD1, and MAFA. IPCs were seeded on temperature-responsive culture dishes to form cell sheets. Differentiation efficiency was confirmed by ß cell-specific gene expression, insulin production, and immunohistochemistry. IPC suspension was injected by portal vein (PV), and IPC sheet was transplanted on the liver surface of the diabetic nude mouse. The therapeutic effect of IPC sheet was evaluated by comparing blood glucose control, weight gain, histological evaluation, and hepatotoxicity with IPC injection group. Also, cell biodistribution was assessed by in vivo/ex vivo fluorescence image tagging. RESULTS: Insulin gene expression and protein production were significantly increased on IPC sheets compared with those in IPCs cultured on conventional culture dishes. Transplanted IPC sheets displayed significantly higher engraftment efficiency and fewer transplanted cells in other organs than injected IPCs, and also lower liver toxicity, improved blood glucose levels, and weight gain. Immunohistochemical analyses of liver tissue revealed positive staining for PDX1 and insulin at 1, 2, and 4 weeks after IPC transplantation. CONCLUSIONS: In conclusion, cell sheet formation enhanced the differentiation function and maturation of IPCs in vitro. Additionally, parameters for clinical application such as distribution, therapeutic efficacy, and toxicity were favorable. The cell sheet technique may be used with IPCs derived from various cell sources in clinical applications.

Acetazolamide-eluting biodegradable tubular stent prevents pancreaticojejunal anastomotic leakage.

Postoperative pancreatic fistula at the early stage can lead to auto-digestion, which may delay the recovery of the pancreaticojejunal (PJ) anastomosis. The efficacy and safety of an acetazolamide-eluting biodegradable tubular stent (AZ-BTS) for the prevention of self-digestion and intra-abdominal inflammatory diseases caused by pancreatic juice leakage after PJ anastomosis in a porcine model were investigated. The AZ-BTS was successfully fabricated using a multiple dip-coating process. Then, the drug amount and release profile were analyzed. The therapeutic effects of AZ were examined in vitro using two kinds of pancreatic cancer cell lines, AsPC-1 and PANC-1. The efficacy of AZ-BTS was assessed in a porcine PJ leakage model, with animals were each assigned to a leakage group, a BTS group and an AZ-BTS group. The overall mortality rates in these three groups were 44.4%, 16.6%, and 0%, respectively. Mean α-amylase concentrations were significantly higher in the leakage and BTS groups than in the AZ-BTS group on day 2-5 (p < 0.05 each all). The luminal diameters and areas of the pancreatic duct were significantly larger in the leakage group than in the BTS and AZ-BTS groups (p < 0.05 each all). These findings indicate that AZ-BTS can significantly suppress intra-abdominal inflammatory diseases caused by pancreatic juice leakage and also prevent late stricture formation at the PJ anastomotic site in a porcine model.

Evaluation of Multi-Layered Pancreatic Islets and Adipose-Derived Stem Cell Sheets Transplanted on Various Sites for Diabetes Treatment.

Islet cell transplantation is considered an ideal treatment for insulin-deficient diabetes, but implantation sites are limited and show low graft survival. Cell sheet technology and adipose-derived stem cells (ADSCs) can be useful tools for improving islet cell transplantation outcomes since both can increase implantation efficacy and graft survival. Herein, the optimal transplantation site in diabetic mice was investigated using islets and stem cell sheets. We constructed multi-layered cell sheets using rat/human islets and human ADSCs. Cell sheets were fabricated using temperature-responsive culture dishes. Islet/ADSC sheet (AI sheet) group showed higher viability and glucose-stimulated insulin secretion than islet-only group. Compared to islet transplantation alone, subcutaneous AI sheet transplantation showed better blood glucose control and CD31+ vascular traits. Because of the adhesive properties of cell sheets, AI sheets were easily applied on liver and peritoneal surfaces. Liver or peritoneal surface grafts showed better glucose control, weight gain, and intraperitoneal glucose tolerance test (IPGTT) profiles than subcutaneous site grafts using both rat and human islets. Stem cell sheets increased the therapeutic efficacy of islets in vivo because mesenchymal stem cells enhance islet function and induce neovascularization around transplanted islets. The liver and peritoneal surface can be used more effectively than the subcutaneous site in future clinical applications.

Spheroid Fabrication Using Concave Microwells Enhances the Differentiation Efficacy and Function of Insulin-Producing Cells via Cytoskeletal Changes.

Pancreatic islet transplantation is the fundamental treatment for insulin-dependent diabetes; however, donor shortage is a major hurdle in its use as a standard treatment. Accordingly, differentiated insulin-producing cells (DIPCs) are being developed as a new islet source. Differentiation efficiency could be enhanced if the spheroid structure of the natural islets could be recapitulated. Here, we fabricated DIPC spheroids using concave microwells, which enabled large-scale production of spheroids of the desired size. We prepared DIPCs from human liver cells by trans-differentiation using transcription factor gene transduction. Islet-related gene expression and insulin secretion levels were higher in spheroids compared to those in single-cell DIPCs, whereas actin-myosin interactions significantly decreased. We verified actin-myosin-dependent insulin expression in single-cell DIPCs by using actin-myosin interaction inhibitors. Upon transplanting cells into the kidney capsule of diabetic mouse, blood glucose levels decreased to 200 mg/dL in spheroid-transplanted mice but not in single cell-transplanted mice. Spheroid-transplanted mice showed high engraftment efficiency in in vivo fluorescence imaging. These results demonstrated that spheroids fabricated using concave microwells enhanced the engraftment and functions of DIPCs via actin-myosin-mediated cytoskeletal changes. Our strategy potentially extends the clinical application of DIPCs for improved differentiation, glycemic control, and transplantation efficiency of islets.

Coaxial mussel-inspired biofibers: making of a robust and efficacious depot for cancer drug delivery.

Biopolymer-based hydrogels have emerged as promising platforms for drug delivery systems (DDSs) due to their inherent biocompatibility, tunable physical properties and controllable degradability. Yet, drug release in majority of these systems is solely contingent on diffusion of drug molecules through the hydrogel, which often leads to burst release of drugs from these systems. Herein, inspired by the chemistry of mussel adhesive proteins, a new generation of coaxial hydrogel fibers was developed that could simultaneously exert both affinity and diffusion control over the release of chemotherapeutic drugs. Specifically, dopamine-modified alginate hydrogel along with chemotherapeutic drugs (doxorubicin or gemcitabine) was used as the main core component to confer affinity-controlled release, while a methacrylated-alginate hydrogel was used as the shell composition to provide the controlled diffusion barrier. It was shown that our coaxial mussel-inspired biofibers yielded biocompatible hydrogel fibers (as indicated by comprehensive in vitro and in vivo experiments) with favourable properties including controlled swelling, and enhanced mechanical properties, when compared against single fibers made from unmodified alginate. Notably, it was observed that these coaxial fibers were capable of releasing the two drugs in a slower manner, when compared to single fibers made from pure alginate, which was partly attributed to stronger interactions of drugs with dopamine-modified alginate (the core element of coaxial fibers) as observed from zeta-potential measurements. It was further shown that these drug-loaded coaxial fibers had optimal anticancer activity both in vitro and in vivo using various pancreatic cancer cell lines. Most remarkably, drug loaded coaxial fibers, particularly doxorubicin-containing fibers, had higher anticancer effect in vivo compared to systemic injection of equivalent dosage of the drugs. Altogether, these biocompatible and robust hydrogel fibers may be further used as neoadjuvant or adjuvant therapies for controlled delivery of chemotherapeutic drugs locally to the tumor sites.

Engineered Cell Sheets for the Effective Delivery of Adipose-Derived Stem Cells for Tendon-to-Bone Healing.

BACKGROUND: Efforts are being made to treat rotator cuff tears (RCTs) that exhibit poor healing and high retear rates. Tendon-to-bone healing using mesenchymal stem cells is being explored, but research is needed to establish effective delivery options. PURPOSE: To evaluate the effects of an adipose-derived stem cell (ADSC) sheet on mesenchymal stem cell delivery for tendon-to-bone healing of a chronic RCT in rats and to demonstrate that ADSC sheets enhance tendon-to-bone healing. STUDY DESIGN: Controlled laboratory study. METHODS: Mesenchymal stem cells were obtained from rat adipose tissue, and a cell sheet was prepared using a temperature-responsive dish. To evaluate the efficacy of stem cells produced in a sheet for the lesion, the experiment was conducted with 3 groups: repair group, cell sheet transplantation after repair group, and cell sheet-only group. Histological, biomechanical, and micro-computed tomography (micro-CT) results were compared among the groups. RESULTS: Hematoxylin and eosin staining for histomorphological analysis revealed that the cell sheet transplantation after repair group (5.75 ± 0.95) showed statistically significant higher scores than the repair (2.75 ± 0.50) and cell sheet-only (3.25 ± 0.50) groups (P < .001). On safranin O staining, the cell sheet transplantation after repair group (0.51 ± 0.04 mm2) had a larger fibrocartilage area than the repair (0.31 ± 0.06 mm2) and cell sheet-only (0.32 ± 0.03 mm2) groups (P = .001). On micro-CT, bone volume/total volume values were significantly higher in the cell sheet transplantation after repair group (23.98% ± 1.75%) than in the other groups (P < .039); there was no significant difference in the other values. On the biomechanical test, the cell sheet transplantation after repair group (4 weeks after repair) showed significantly higher results than the other groups (P < .005). CONCLUSION: Our study shows that engineered stem cells are a clinically feasible stem cell delivery tool for rotator cuff repair. CLINICAL RELEVANCE: This laboratory study provides evidence that ADSCs are effective in repairing RCTs, which are common sports injuries.

A Novel Biodegradable Tubular Stent Prevents Pancreaticojejunal Anastomotic Stricture.

Stricture of pancreatic-enteric anastomoses is a major late complication of a pancreaticoduodenectomy for the treatment of a periampullary tumor and can lead to exocrine and endocrine insufficiency such as malnutrition and diabetes mellitus. We investigated the safety and efficacy of a biodegradable tubular stent (BTS) for preventing a pancreaticojejunostomy (PJ) anastomotic stricture in both a rat and porcine model. The BTS was manufactured using a terpolymer comprising poly p-dioxanone, trimethylene carbonate, and glycolide. A cohort of 42 rats was randomized into 7 groups of 6 animals each after BTS placement into the duodenum for the biodegradation assay. A total of 12 pigs were randomized equally into a control and BTS placement group. The effectiveness of the BTS was assessed by comparing radiologic images with histologic results. Surgical procedures and/or BTS placements were technically successful in all animals. The median mass losses of the removed BTS samples from the rat duodenum were 2.1, 6.8, 11.2, 19.4, 26.1, and 56.8% at 1, 2, 3, 4, 6, and 8 weeks, respectively. The BTS had completely degraded at 12 weeks in the rats. In the porcine PJ model, the mean luminal diameter and area of the pancreatic duct in the control group was significantly larger than in the BTS group (all p < 0.05). BTS placement thus appears to be safe and effective procedure for the prevention of PJ anastomotic stricture. These devices have the potential to be used as a temporary stent placement to treat pancreatic-enteric anastomoses, but further investigations are required for optimization in human.

Long-term reversal of diabetes by subcutaneous transplantation of pancreatic islet cells and adipose-derived stem cell sheet using surface-immobilized heparin and engineered collagen scaffold.

OBJECTIVE: Esterified collagen (EC) can be functionalized with heparin to enhance islet graft stability. Growth factors secreted by human adipose-derived stem cells (hADSCs) can bind efficiently to EC-heparin (EC-Hep), which enhances revascularization and cell protection. We investigated the therapeutic potential of a combined heparin-esterified collagen-hADSC (HCA)-islet sheet to enhance islet engraftment. RESEARCH DESIGN AND METHODS: This study was designed to assess the efficiency of using EC-Hep as a scaffold for subcutaneous islet transplantation in diabetic athymic mice. After the hADSC-cocultured islets were seeded in the EC-Hep scaffold, islet function was measured by glucose-stimulated insulin secretion test and growth factors in the culture supernatants were detected by protein array. Islet transplantation was performed in mice, and graft function and survival were monitored by measuring the blood glucose levels. ß-Cell mass and vascular densities were assessed by immunohistochemistry. RESULTS: The EC-Hep composite allowed sustained release of growth factors. Secretion of growth factors and islet functionality in the HCA-islet sheet were significantly increased compared with the control groups of islets alone or combined with native collagen. In vivo, stable long-term glucose control by the graft was achieved after subcutaneous transplantation of HCA-islet sheet due to enhanced capillary network formation around the sheet. CONCLUSIONS: The findings indicate the potential of the HCA-islet sheet to enhance islet revascularization and engraftment in a hADSC dose-dependent manner, following clinical islet transplantation for the treatment of diabetes mellitus.

Effect of immobilized cell-binding peptides on chitosan membranes for osteoblastic differentiation of mesenchymal stem cells.

Two cell-binding domains from FGF-2 (fibroblast growth factor-2) were shown to increase cell attachment and osteoblastic differentiation. Two synthetic peptides derived from FGF-2, namely residues 36-41 (F36; PDGRVD) and 77-83 (F77; KEDGRLL), were prepared and their N-termini further modified for ease of surface immobilization. Chitosan membranes were used in the present study as mechanical supportive biomaterials for peptide immobilization. Peptides could be stably immobilized on to the surface of chitosan membranes. The adhesion of mesenchymal stem cells to the peptide (F36 and F77)-immobilized chitosan membrane was increased in a dose-dependent manner and completely inhibited by soluble RGD (Arg-Gly-Asp) and anti-integrin antibody, indicating the existence of an interaction between F36/F77 and integrin. Peptide-immobilized chitosan supported human bone-marrow-derived mesenchymal-stem-cell differentiation into osteoblastic cells, as demonstrated by alkaline phosphate expression and mineralization. Taken together, the identified peptide-immobilized chitosan membranes were able to support cell adhesion and osteoblastic differentiation; thus these peptides might be useful as bioactive agents for osteoblastic differentiation and surface-modification tools in bone regenerative therapy.

Pancreatic Tissue-Derived Extracellular Matrix Bioink for Printing 3D Cell-Laden Pancreatic Tissue Constructs.

The transplantation of pancreatic islets is a promising treatment for patients who suffer from type 1 diabetes accompanied by hypoglycemia and secondary complications. However, islet transplantation still has several limitations such as the low viability of transplanted islets due to poor islet engraftment and hostile environments. In addition, the insulin-producing cells differentiated from human pluripotent stem cells have limited ability to secrete sufficient hormones that can regulate the blood glucose level; therefore, improving the maturation by culturing cells with proper microenvironmental cues is strongly required. In this article, we elucidate protocols for preparing a pancreatic tissue-derived decellularized extracellular matrix (pdECM) bioink to provide a beneficial microenvironment that can increase glucose sensitivity of pancreatic islets, followed by describing the processes for generating 3D pancreatic tissue constructs using a microextrusion-based bioprinting technique.