Induction of Skin Allograft Chimerism by Pharmacological Mobilization of Endogenous Bone Marrow-Derived Stem Cells.

Skin substitutes including allografts remain a standard therapeutic approach to promote healing of both acute and chronic large wounds. However, none have resulted in the regrowth of lost and damaged tissues and scarless wound healing. Here, we demonstrate skin allograft chimerism and repair through the mobilization of endogenous bone marrow-derived stem and immune cells in rats and swine. We show that pharmacological mobilization of bone marrow stem cells and immune cells into the circulation promotes host repopulation of skin allografts and restoration of the skin's normal architecture without scarring and minimal contracture. When skin allografts from DA rats are transplanted into GFP transgenic Lewis recipients with a combination of AMD3100 and low-dose FK506 (AF) therapy, host-derived GFP-positive cells repopulate and/or regenerate cellular components of skin grafts including epidermis and hair follicles and the grafts become donor-host chimeric skin. Using AF combination therapy, burn wounds with skin allografts were healed by newly regenerated chimeric skin with epidermal appendages and pigmentation and without contracture in swine.

A phase I, first-in-human study to evaluate the safety and tolerability, pharmacokinetics, and pharmacodynamics of MRG-001 in healthy subjects.

Preclinical studies demonstrate that pharmacological mobilization and recruitment of endogenous bone marrow stem cells and immunoregulatory cells by a fixed-dose drug combination (MRG-001) improves wound healing, promotes tissue regeneration, and prevents allograft rejection. In this phase I, first-in-human study, three cohorts receive subcutaneous MRG-001 or placebo, every other day for 5 days. The primary outcome is safety and tolerability of MRG-001. Fourteen subjects received MRG-001 and seven received a placebo. MRG-001 is safe over the selected dose range. There are no clinically significant laboratory changes. The intermediate dose group demonstrates the most significant white blood cell, stem cell, and immunoregulatory cell mobilization. PBMC RNA sequencing and gene set enrichment analysis reveal 31 down-regulated pathways in the intermediate MRG-001 dose group compared with no changes in the placebo group. MRG-001 is safe across all dose ranges. MRG-001 may be a clinically useful therapy for immunoregulation and tissue regeneration (ClinicalTrials.gov: NCT04646603).

Major Improvement in Wound Healing Through Pharmacologic Mobilization of Stem Cells in Severely Diabetic Rats.

Current therapeutic strategies for diabetic foot ulcer (DFU) have focused on developing topical healing agents, but few agents have controlled prospective data to support their effectiveness in promoting wound healing. We tested a stem cell mobilizing therapy for DFU using a combination of AMD3100 and low-dose FK506 (tacrolimus) (AF) in streptozocin-induced type 1 diabetic (T1DM) rats and type 2 diabetic Goto-Kakizaki (GK) rats that had developed peripheral artery disease and neuropathy. Here, we show that the time for healing back wounds in T1DM rats was reduced from 27 to 19 days, and the foot wound healing time was reduced from 25 to 20 days by treatment with AF (subcutaneously, every other day). Similarly, in GK rats treated with AF, the healing time on back wounds was reduced from 26 to 21 days. Further, this shortened healing time was accompanied by reduced scar and by regeneration of hair follicles. We found that AF therapy mobilized and recruited bone marrow-derived CD133+ and CD34+ endothelial progenitor cells and Ym1/2+ M2 macrophages into the wound sites, associated with enhanced capillary and hair follicle neogenesis. Moreover, AF therapy improved microcirculation in diabetic and neuropathic feet in GK rats. This study provides a novel systemic therapy for healing DFU.

Pharmacological Mobilization and Recruitment of Stem Cells in Rats Stops Abdominal Adhesions After Laparotomy.

Adhesions are a very common complication in the abdominal surgery. Animal studies and human trials have evaluated strategies designed to reduce and prevent postsurgical adhesions but few have an evidence base that justifies routine use. A strategy to prevent adhesions effectively remains an urgent need. We studied a reproducible model of intra-peritoneal adhesion formation in rats using laparotomy with several peritoneal sutures to produce the adhesions. Here we show that entraining endogenous stem cells into injury sites using the combined effect of AMD3100 and low-dose FK-506 (AF) can reduce the adhesion score significantly and abolish peritoneal adhesions in 45% of animals in a rat model of severe postsurgical intra-abdominal adhesions, compared with saline controls. Searching for mechanisms, we found AF treatment dramatically increased SDF-1 expressing cells, HGF expressing Ym1+ M2 macrophages and CD133+ stem cells in the injury sites of peritoneal surface at day 5 post-operation. Our results demonstrate that medically induced recruitment of autologous stem cells using AF significantly reduced postsurgical intra-abdominal adhesions. These findings suggest a novel effective therapeutic approach to preventing adhesions in patients.

Choledochoscope with stent placement for treatment of benign duodenal strictures: A case report.

BACKGROUND: Endoscopically or fluoroscopically guided dilation is a safe and effective alternative to surgery for patients with benign strictures of the gastric outlet. CASE SUMMARY: We describe two cases where a novel approach with a Spyglass® choledochoscope in assessing the extent of benign duodenal strictures and aiding in placement of duodenal stents for treatment of the strictures. Choledochoscope-guided wire and stent placement was successful in all cases, leading to symptom resolution related to benign duodenal obstruction. No major adverse events were observed. CONCLUSION: Choledochoscope-guided assessment and endoscopic therapy is a viable approach in relieving duodenal obstruction, if the conventional combined fluoroscopic and endoscopic methods fail.

Endoscopic vacuum assisted wound closure (EVAC) device to treat esophageal and gastric leaks: assessing time to proficiency and cost.

BACKGROUND: Endoluminal vacuum therapy (EVAC) is an emerging procedure used to treat anastomotic leaks and/or perforations that would otherwise require surgery. The aim of this study was to determine time to proficiency in EVAC and the cost effectiveness of the procedure. METHODS: We retrospectively reviewed a prospectively maintained IRB approved database for all patients undergoing EVAC after esophageal and gastric complications between October 2013 and December 2017. Proficiency was determined by obtaining predicted estimates and analyzing the point at which average procedure time plateaued based on case volume. Total cost was calculated based on supplies and location where the procedure was performed. RESULTS: There were 50 patients (17 males, 33 female), with a mean age of 52.1 years. EVAC was placed in 23 (46%) patients with esophageal injuries and 28 (56%) with gastric injuries. Two advanced endoscopists performed all EVAC procedures in this study (1 surgeon, 1 gastroenterologist). The average procedure time for all patients was 43.5 min and the average wheel in/wheel out time for all patients was 75.6 min. Analysis of the trend based on average procedure times for EVAC revealed that proficiency was obtained after 10 cases. Total cost of the procedure is significantly lower in the GI lab compared to the operating room ($4528 vs. $11889). The majority of EVAC were performed in the GI lab (62%) compared to the operating room (38%). CONCLUSION: Successful outcomes in managing anastomotic leaks or intestinal perforations non-operatively has led to an increased interest in EVAC. For advanced endoscopists, time to proficiency is approximately 10 cases. Performing the procedure in the GI lab has a 2.5 reduction in total cost compared to the operating room.

Stem cells in burn wound healing: A systematic review of the literature.

INTRODUCTION: Severe burns are often associated with high morbidity and unsatisfactory functional and esthetic outcomes. Over the last two decades, stem cells have generated great hopes for the treatment of numerous conditions including burns. The aim of this systematic review is to evaluate the role of stem cell therapy as a means to promote burn wound healing. METHODS: Comprehensive searches in major databases were carried out in March 2017 for articles on stem cell therapy in burn wound healing. In total 2103 articles were identified and screened on the basis of pre-determined inclusion and exclusion criteria. RESULTS: Fifteen experimental and two clinical studies were included in the review. The majority of studies reported significant improvement in macroscopic burn wound appearance as well as a trend toward improved microscopic appearance, after stem cell therapy. Other parameters evaluated, such as re-vascularization, collagen formation, level of pro- and anti-inflammatory mediators, apoptosis and cellular infiltrates, yielded heterogeneous results across studies. CONCLUSION: Stem cell therapy appears to exert a positive effect in burn wound healing. There is, therefore, justification for continued efforts to evaluate the use of stem cells as an adjunct to first-line therapies in burns.

Primary and Rescue Endoluminal Vacuum Therapy in the Management of Esophageal Perforations and Leaks.

BACKGROUND: To investigate the efficacy of primary and rescue endoluminal vacuum (EVAC) therapy in the treatment of esophageal perforations and leaks. METHODS: We conducted a retrospective review of a prospectively gathered, Institutional Review Board (IRB) approved database of EVAC therapy patients at our center from July 2013 to September 2016. RESULTS: In all, 13 patients were treated for esophageal perforations or leaks. Etiologies included iatrogenic injury (n = 8), anastomotic leak (n = 2), Boerhaave syndrome (n = 1), and bronchoesophageal fistula (n = 2). In total, 10 patients underwent primary treatment and three were treated with rescue therapy. Mean Perforation Severity Scores (PSSs) in the primary and rescue treatment groups were 7 and 10, respectively. Average defect size was 2.4 (range: 0.5-6) cm. The rescue group had a shorter mean time to defect closure (25 vs. 33 days). In all, 12 of 13 defects healed. One death occurred following the implementation of comfort care. One therapy-specific complication occurred. Hospital length of stay (LOS) was longer in the rescue group (72 vs. 53 days); however, the intensive care unit (ICU) duration was similar between groups. Totally, 10 patients (83%) resumed an oral diet after successful defect closure. CONCLUSION: Utilized as either a primary or rescue therapy, EVAC therapy appears to be beneficial in the management of esophageal perforations or leaks.

Endolumenal Vacuum Therapy and Fistulojejunostomy in the Management of Sleeve Gastrectomy Staple Line Leaks.

Laparoscopic sleeve gastrectomy (LSG) is the most common bariatric surgery performed for morbid obesity. Leaks of the vertical staple line can occur in up to 7% of cases and are difficult to manage. Endolumenal vacuum (EVAC) therapy and fistulojejunostomy (FJ) have separate documented uses to heal these complicated leaks. We aim to show the benefit of using EVAC with FJ in the treatment of LSG staple line leaks. The patient presented with an LSG chronic leak. EVAC therapy was initiated but failed to close the fistula after 101 days. EVAC therapy was abandoned, and FJ was performed to resolve the leak. Postoperatively, no leak was encountered requiring any additional procedures. Based on our findings, we conclude that EVAC therapy facilitates in resolving leaks that restore gastrointestinal continuity and maintain source control. It promotes healing and causes reperfusion of ischemic tissue and fistula cavity debridement.

Pharmacological Mobilization of Endogenous Bone Marrow Stem Cells Promotes Liver Regeneration after Extensive Liver Resection in Rats.

Rapid regeneration of the remnant liver is critical for preventing liver failure and promoting recovery after extensive liver resection. Numerous studies have demonstrated the involvement of bone marrow-derived stem cells in liver regeneration and the potential benefits of bone marrow stem cell therapy. To avoid the preparation of stem cells, we proposed in this study to mobilize endogenous bone marrow stem cells pharmacologically with a combination of AMD3100 (A), an antagonist of CXCR4 and low-dose FK506 (F). Here we show that AF combination therapy significantly increased lineage negative (Lin-) CD34+ and Lin-CD133+ stem cells in peripheral blood and enhanced recruitment of CD133+ cells into the remnant liver in a rat model of 85% partial hepatectomy. Recruiting CD133+ stem cells in the remnant liver was associated with increased proliferation of hepatic oval cells and paralleled the increased SDF-1, CXCR4 and HGF expression. Importantly, AF combination therapy increased the number of Ki67 positive hepatocytes and BrdU incorporation in the remnant liver and improved serum levels of albumin. Our results demonstrate that pharmacological mobilization of endogenous bone marrow stem cells with AF combination therapy can enhance endogenous stem cell mobilization to promote liver regeneration and improve liver function after extensive hepatectomy.

Fibrosis in small syngeneic rat liver grafts because of damaged bone marrow stem cells from chronic alcohol consumption.

A patient with liver failure due to chronic and acute alcohol abuse under consideration for an urgent liver transplant shortly after stopping alcohol may have residual abnormalities that threaten transplant success, particularly for a small graft. To address this, we studied a model in which reduced-size (50%) Lewis rat livers are transplanted into green fluorescence protein transgenic Lewis recipients after they are fed alcohol or a control diet for 5 weeks. Here we show that normal small Lewis grafts transplanted to alcohol-fed Lewis hosts developed fibrosis, whereas no fibrosis was observed in control-fed recipients. Host-derived CD133 + 8-hydroxy-2'-deoxyguanosine (8-OHdG) cells were significantly increased in livers recovered from both alcohol-fed and control recipients, but only alcohol-fed recipients demonstrated co-staining (a marker of oxidative DNA damage). α smooth muscle actin (α-SMA) staining, a marker for myofibroblasts, also co-localized with CD133 + cells only in the livers of alcohol-fed recipients. Immunostaining and polymerase chain reaction analysis confirmed that chronic alcohol consumption decreased the proportion of bone marrow stem cells (BMSCs) expressing CD133, c-Kit, and chemokine (C-X-C motif) receptor 4 markers and caused oxidative mitochondria DNA (mtDNA) damage. Culture of CD133 + cells from normal rats with medium containing 3% ethanol for 48 hours resulted in elevated mitochondrial 8-OHdG and mtDNA deletion, and ethanol exposure diminished CD133 expression but dramatically increased α-SMA expression. In conclusion, oxidative mtDNA damage and deletions occur in BMSCs of chronic alcohol-fed recipients, and these damaged cells mobilize to the small liver grafts and become myofibroblasts where they play a key role in the subsequent development of fibrosis. Liver Transplantation 23 1564-1576 2017 AASLD.

A study of the clinical utility of a 20-minute secretin-stimulated endoscopic pancreas function test and performance according to clinical variables.

BACKGROUND AND AIMS: Direct pancreas juice testing of bicarbonate, lipase, or trypsin after stimulation by secretin or cholecystokinin is used to determine exocrine function, a surrogate for diagnosing chronic pancreatitis (CP). Endoscopic pancreas function tests (ePFTs), where a peak bicarbonate concentration (PBC) ≥80 mEq/L in pancreas juice is considered normal, are now used more frequently. In this ePFT, aspirates start 35 minutes after secretin administration because pancreas output peaks 30 minutes after secretagogue administration. The performance of ePFT in a cohort of patients with a presumptive diagnosis of CP referred to a pancreas clinic for consideration of an intervention including total pancreatectomy and islet autotransplantation was studied, compared with EUS, ERCP, histology, and consensus diagnosis. The effect of sedation, narcotic use, aspirate volume, body mass index, age, and proton pump inhibitors (PPIs) on test performance is reported. METHODS: After a test dose, synthetic human secretin was administered intravenously, and 30 minutes later sedation was achieved with midazolam and fentanyl or propofol. A gastroscope was advanced to the major papilla where 4 continuous aspiration samples were performed at 5-minute intervals in sealed bottles. PBC ≥80 mEq/L was normal. RESULTS: Eighty-one patients had ePFTs from August 2010 through October 2015. Twenty-seven patients (33%) were diagnosed with CP. Eighteen of the 27 patients with CP and 1 of the 54 patients without CP had an abnormal ePFT, producing a sensitivity of 66% (95% CI, 46.0-83.5), specificity 98% (95% CI, 90.1-99.9), positive predictive value 94.7% (95% CI, 74-99.9), and negative predictive value 85.5% (95% CI, 74.2-93.1). ERCP and PBC concordance was generally poor, but none of the patients without CP had major EUS changes, and only 3 patients with a PBC <80 mEq/L had a normal EUS. The PBC was affected by narcotics and PPI use. CONCLUSION: A 20-minute ePFT after secretin administration had a marginal sensitivity for diagnosis of CP. The diagnosis of CP should not rely on a single study and certainly not a PFT. The duodenal aspirate volume did not correlate with the PBC, which contrasts with current secretin-enhanced MRCP knowledge; therefore, further studies on this subject are warranted. Neither type of sedation, BMI, nor age affected test performance. Narcotics and PPIs may affect the PBC, so borderline results should be interpreted with caution in these groups.

Management of gastric leaks after sleeve gastrectomy with endoluminal vacuum (E-Vac) therapy.

BACKGROUND: Sleeve gastrectomy has become a popular weight loss procedure, but it is associated with staple line leak resulting in high morbidity and mortality. Current management options range from endoscopic techniques (predominantly stent placement) to surgical intervention. OBJECTIVE: The purpose of this study was to recognize endoluminal vacuum (E-Vac) therapy as a viable option for use in anastomotic leaks of sleeve gastrectomies. SETTING: This study took place at Baylor University Medical Center at Dallas, Texas. METHODS: Retrospective and prospectively gathered registries for use of E-Vac therapy were queried to identify 35 patients. Using upper gastrointestinal series (UGI) and esophagogastroduodenoscopy, 9 of these patients were identified with a staple line leak from laparoscopic sleeve gastrectomy (LSG). E-Vac therapy was used to resolve the leak. RESULTS: Nine patients were treated with E-Vac therapy. Eight of 9 patients were admitted from outside hospitals with a mean of 61 days (5-233) after LSG. During treatment, an average of 10.3 procedures per patient was done to place and exchange the Endo-SPONGE. All 9 patients had resolution of leaks confirmed by upper gastrointestinal series, after undergoing E-Vac therapy for an average of 50 days. Six of 9 patients had laparoscopic procedures before their admission. During admission, 5 of the 9 patients had self-expanding metal stents placed with failure of leak resolution. Discharge disposition included 2 patients sent to rehabilitation facilities, 1 death not attributable to E-Vac, and 6 patients went home. CONCLUSION: E-Vac therapy is a viable option for patients with staple line leak after LSG.

Effect of the Duration of Chronic Pancreatitis on Pancreas Islet Yield and Metabolic Outcome Following Islet Autotransplantation.

BACKGROUND: Total pancreatectomy (TP) with islet autotransplantation (IAT) is a highly selected treatment for severe pain associated with chronic pancreatitis (CP) after exhausting medical and endoscopic therapies. The effect of duration of CP on TP-IAT has not been clarified. METHODS: Retrospective review of a consecutive cohort undergoing TP-IAT was performed. Patients were classified according to islet dose of <2500 IEQ/kg, 2500 to 5000 IEQ/kg, and >5000 IEQ/kg. Islet yield and metabolic outcomes were compared to disease duration of CP. RESULTS: A total of 76 CP patients underwent TP-IAT. Longer disease duration was associated with lower islet yield transplanted (Spearman's correlation = -0.24; p = 0.04) for total cohort. Highest absolute value of the coefficient was found in patients with hereditary CP when study subjects were classified by the etiology of CP (correlation = -0.72; p = 0.02). Higher islet yields were significantly associated with better metabolic outcomes (7.6 ± 1.1 vs 6.6 ± 1.1% of HbA1c post-TPIAT in patients with <2500 and >5000 IEQ/kg transplanted, respectively; p = 0.04). CONCLUSIONS: The duration of CP could affect islet yield and metabolic outcomes. The time since the diagnosis of CP should be considered when selecting patients for islet autotransplantation.

Surgical management of early and late ureteral complications after renal transplantation: techniques and outcomes.

BACKGROUND: In this study, we present our experience with ureteral complications requiring revision surgery after renal transplantation and compare our results to a matched control population. METHODS: We performed a retrospective analysis of our database between 1997 and 2012. We divided the cases into early (<60 d) and late repairs. Kaplan-Meier and Cox proportional hazards models were used to compare graft survival between the intervention cohort and controls generated from the Scientific Registry of Transplant Recipients data set. RESULTS: Of 2671 kidney transplantations, 51 patients were identified as to having undergone 53 ureteral revision procedures; 43.4% of cases were performed within 60 d of the transplant and were all associated with urinary leaks, and 49% demonstrated ureteral stenosis. Reflux allograft pyelonephritis and ureterolithiasis were each the indication for intervention in 3.8%; 15.1% of the lesions were located at the anastomotic site, 37.7% in the distal segment, 7.5% in the middle segment, 5.7% proximal ureter, and 15.1% had a long segmental stenosis. In 18.9%, the location was not specified. Techniques used included ureterocystostomy (30.2%), ureteroureterostomy (34%), ureteropyelostomy (30.1%), pyeloileostomy (1.9%), and ureteroileostomy (3.8%). No difference in overall graft survival (HR 1.24 95% CI 0.33-4.64, p = 0.7) was detected when compared to the matched control group. CONCLUSION: Using a variety of techniques designed to re-establish effective urinary flow, we have been able to salvage a high percentage of these allografts. When performed by an experienced team, a ureteric complication does not significantly impact graft survival or function as compared to a matched control group.

Cationic poly(2-aminoethylmethacrylate) and poly(N-(2-aminoethylmethacrylamide) modified cellulose nanocrystals: synthesis, characterization, and cytotoxicity.

Cellulose nanocrystals (CNCs) continue to gain increasing attention in the materials community as sustainable nanoparticles with unique chemical and mechanical properties. Their nanoscale dimensions, biocompatibility, biodegradability, large surface area, and low toxicity make them promising materials for biomedical applications. Here, we disclose a facile synthesis of poly(2-aminoethylmethacrylate) (poly(AEM)) and poly(N-(2-aminoethylmethacrylamide) (poly(AEMA)) CNC brushes via the surface-initiated single-electron-transfer living radical polymerization technique. The resulting modified CNCs were characterized for their chemical and morphological features using a combination of analytical, spectroscopic, and microscopic techniques. Zeta potential measurements indicated a positive surface charge, and further proof of the cationic nature was confirmed by gold deposition as evidenced by electron microscopy. The cytotoxicity of these cationic modified CNCs was evaluated utilizing a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in two different cell lines, J774A1 (mouse monocyte cells) and MCF-7 (human breast adenocarcinoma cells). The results indicated that none of the cationic modified CNCs decreased cell viability at low concentrations, which could be suitable for biomedical applications.

Arteriosclerosis in kidneys from healthy live donors: comparison of wedge and needle core perioperative biopsies.

CONTEXT: Although risks associated with live kidney donation are low, there are few pathologic studies of kidneys from live donors, and possible risk factors for development of hypertension or renal insufficiency remain unknown. There are many studies of histopathologic changes in deceased donor kidneys and how these changes affect subsequent graft function; most are based on wedge rather than needle core biopsies. OBJECTIVE: To examine the frequency and severity of arterial fibrointimal thickening and other pathologic lesions in kidneys from healthy live donors and compare wedge and needle core biopsies as methods for evaluating these changes. DESIGN: For 36 of 332 live donor renal transplantations performed from January 2004 through November 2006, a wedge biopsy of the transplanted kidney was done prior to and/or after implantation, and a needle core biopsy was done postimplantation or during the ensuing 7 days. For these 36 allografts, we compared pathologic features of the wedge and core perioperative biopsies. RESULTS: Findings on core and wedge biopsies were similar, except for arterial fibrointimal thickening. Moderate thickening (Banff cv2) was present on 13 core biopsies, and mild thickening (cv1) was present on another 10; by contrast, no wedge biopsies showed cv2 lesions, and only 8 showed cv1. Arterial thickening on core but not wedge biopsies correlated significantly with increasing patient age. CONCLUSIONS: The findings indicate that needle core biopsies are superior to wedge biopsies for evaluating vascular changes in donor kidneys, and they suggest a need for studies correlating such changes with long-term outcomes of live donors, particularly older donors.