Searching for the Optimal Donor for Allogenic Adipose-Derived Stem Cells: A Comprehensive Review.

Adipose-derived stem cells comprise several clinically beneficial qualities that have been explored in basic research and have motivated several clinical studies with promising results. After being approved in the European Union, UK, Switzerland, Israel, and Japan, allogeneic adipose-derived stem cells (darvadstrocel) have been recently granted a regenerative medicine advanced therapy (RMAT) designation by US FDA for complex perianal fistulas in adults with Crohn's disease. This huge scientific step is likely to impact the future spread of the indications of allogeneic adipose-derived stem cell applications. The current knowledge on adipose stem cell harvest describes quantitative and qualitative differences that could be influenced by different donor conditions and donor sites. In this comprehensive review, we summarize the current knowledge on the topic and propose donor profiles that could provide the optimal initial quality of this living drug, as a starting point for further applications and studies in different pathological conditions.

Stem cell therapy applied for digestive anastomosis: Current state and future perspectives.

BACKGROUND: Digestive tract resections are usually followed by an anastomosis. Anastomotic leakage, normally due to failed healing, is the most feared complication in digestive surgery because it is associated with high morbidity and mortality. Despite technical and technological advances and focused research, its rates have remained almost unchanged the last decades. In the last two decades, stem cells (SCs) have been shown to enhance healing in animal and human studies; hence, SCs have emerged since 2008 as an alternative to improve anastomoses outcomes. AIM: To summarise the published knowledge of SC utilisation as a preventative tool for hollow digestive viscera anastomotic or suture leaks. METHODS: PubMed, Science Direct, Scopus and Cochrane searches were performed using the key words "anastomosis", "colorectal/colonic anastomoses", "anastomotic leak", "stem cells", "progenitor cells", "cellular therapy" and "cell therapy" in order to identify relevant articles published in English and Spanish during the years of 2000 to 2021. Studies employing SCs, performing digestive anastomoses in hollow viscera or digestive perforation sutures and monitoring healing were finally included. Reference lists from the selected articles were reviewed to identify additional pertinent articles.Given the great variability in the study designs, anastomotic models, interventions (SCs, doses and vehicles) and outcome measures, performing a reliable meta-analysis was considered impossible, so we present the studies, their results and limitations. RESULTS: Eighteen preclinical studies and three review papers were identified; no clinical studies have been published and there are no registered clinical trials. Experimental studies, mainly in rat and porcine models and occasionally in very adverse conditions such as ischaemia or colitis, have been demonstrated SCs as safe and have shown some encouraging morphological, functional and even clinical results. Mesenchymal SCs are mostly employed, and delivery routes are mainly local injections and cell sheets followed by biosutures (sutures coated by SCs) or purely topical. As potential weaknesses, animal models need to be improved to make them more comparable and equivalent to clinical practice, and the SC isolation processes need to be standardised. There is notable heterogeneity in the studies, making them difficult to compare. Further investigations are needed to establish the indications, the administration system, potential adjuvants, the final efficacy and to confirm safety and exclude definitively oncological concerns. CONCLUSION: The future role of SC therapy to induce healing processes in digestive anastomoses/sutures still needs to be determined and seems to be currently far from clinical use.

Vía totalmente extraperitoneal extendida (eTEP) para la corrección de hernias ventrales: resultados preliminares / Extended Totally Extraperitoneal (eTEP) Approach for Ventral Hernia Repair: Initial Results

INTRODUCCIÓN: Desde la primera eventroplastia laparoscópica se han desarrollado múltiples procedimientos en cirugía miniinvasiva de la pared abdominal. En 2017 se publica el acceso endoscópico totalmente extraperitoneal extendido (eTEP) para la reparación abdominal. Se presentan los resultados de la implementación de la técnica en 2 centros por un único cirujano. MÉTODOS: Estudio prospectivo descriptivo de la implementación de la vía eTEP con o sin liberación de transverso (TAR) para defectos de la pared abdominal. La técnica quirúrgica se inicia con el acceso al espacio entre el músculo recto y la vaina posterior, uniendo dicho espacio con el preperitoneo de la línea media y el espacio retrorrectal contralateral. En la cavidad creada se realiza la disección y reducción del saco herniario. Se puede realizar adicionalmente una liberación posterior de componentes tipo TAR. Se realiza el cierre del peritoneo y del defecto fascial y se extiende una prótesis en el espacio definitivo. RESULTADOS: Se intervinieron 40 casos, 20 casos con defectos supraumbilicales, 10 casos infraumbilicales y 10 en la línea semilunar. Dieciséis casos asociaron TAR. El tiempo quirúrgico medio fue de 126 minutos. El dolor mediano por la escala EVA al alta fue de 3. La estancia mediana fue de un día y el seguimiento medio de 10 meses. Un caso mostró recidiva y 2 pacientes requirieron reintervención. CONCLUSIONES: La reparación de hernias primarias e incisionales abdominales mediante eTEP es segura y reproducible. Los resultados preliminares muestran buen control del dolor postoperatorio y baja estancia

INTRODUCTION: Since the first laparoscopic incisional hernia repair, several minimally invasive procedures have been developed in abdominal wall repair. In 2017, the extended totally extraperitoneal (eTEP) approach for abdominal wall repair was published. We present the results from eTEP implementation at two medical centers by one surgeon. METHODS: Prospective descriptive study of the implementation of the eTEP approach, with transversus abdominis release (TAR) when needed. The surgical technique was initiated by accessing the space between the rectus abdominis muscle and posterior rectus sheath, connecting this space with the fatty preperitoneal space at the midline and the contralateral retrorectal space. Identification and dissection of the hernia sac is performed in the created cavity. Additionally, posterior component release in a TAR fashion could be done. Finally, closure of posterior plane and linea alba is completed and mesh prosthesis is deployed along the whole dissected space. RESULTS: Forty patients underwent an eTEP procedure with 20 supraumbilical defects, 10 infraumbilical and 10 lateral hernias. Sixteen cases required a TAR technique. Mean operative time was 126 minutes. Median pain reported the first postoperative day was 3 on the visual analogue scale. Median length of stay was 1 day and mean follow-up was 10 months. Only one patient developed recurrence, and two patients underwent reoperation. CONCLUSIONS: Implementation of eTEP in abdominal wall repair is safe. Preliminary outcomes of the eTEP approach in ventral hernia repair show good pain control with less hospital stay

Extended Totally Extraperitoneal (eTEP) Approach for Ventral Hernia Repair: Initial Results. / Vía totalmente extraperitoneal extendida (eTEP) para la corrección de hernias ventrales: resultados preliminares.

INTRODUCTION: Since the first laparoscopic incisional hernia repair, several minimally invasive procedures have been developed in abdominal wall repair. In 2017, the extended totally extraperitoneal (eTEP) approach for abdominal wall repair was published. We present the results from eTEP implementation at two medical centers by one surgeon. METHODS: Prospective descriptive study of the implementation of the eTEP approach, with transversus abdominis release (TAR) when needed. The surgical technique was initiated by accessing the space between the rectus abdominis muscle and posterior rectus sheath, connecting this space with the fatty preperitoneal space at the midline and the contralateral retrorectal space. Identification and dissection of the hernia sac is performed in the created cavity. Additionally, posterior component release in a TAR fashion could be done. Finally, closure of posterior plane and linea alba is completed and mesh prosthesis is deployed along the whole dissected space. RESULTS: Forty patients underwent an eTEP procedure with 20 supraumbilical defects, 10 infraumbilical and 10 lateral hernias. Sixteen cases required a TAR technique. Mean operative time was 126minutes. Median pain reported the first postoperative day was 3 on the visual analogue scale. Median length of stay was 1 day and mean follow-up was 10 months. Only one patient developed recurrence, and two patients underwent reoperation. CONCLUSIONS: Implementation of eTEP in abdominal wall repair is safe. Preliminary outcomes of the eTEP approach in ventral hernia repair show good pain control with less hospital stay.

Clinical Implications of NRAS Overexpression in Resectable Pancreatic Adenocarcinoma Patients.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal forms of cancer, and its incidence is rising worldwide. Although survival can be improved by surgical resection, when detected at an early stage, this type of cancer is usually asymptomatic, and disease becomes only apparent after metastasis. Adjuvant treatment does not improve survival, thus after surgery there is a lack of predictive and prognosis biomarkers to predict treatment response and survival. The mitogen-activated protein-kinase and phosphoinositide 3-kinase signalling pathways play a crucial role in cancer development and progression. Especially, activated RAS proteins promote cell proliferation through constitutive stimulation of the downstream effectors RAF-MEK-ERK and PI3K-AKT. Mutational status of NRAS is required in several types of cancer like colorectal or cutaneous melanoma. However, mutations in this gene are very scarce in PDAC patients, and NRAS determination is not usually performed in clinical practice for this kind of tumor. In this study, we analyse the association between NRAS protein expression and progression-free survival and overall survival of an homogenous cohort of pancreatic ductal adenocarcinoma patients from a single-centre. Interestingly, we found that patients with high expression not only showed longer progression-free survival than those patients with low expression (22 versus 9 months, respectively) (P = 0.013), but also longer overall survival (43 versus 19 months, respectively) (P = 0.020). These results confirm NRAS expression could be used to differentiate patients according to their prognosis. Proportional hazard model revealed NRAS expression together with grade of differentiation as pathological variables to predict patient's outcome.

A Step-By-Step Surgical Protocol for the Treatment of Perianal Fistula with Adipose-Derived Mesenchymal Stem Cells.

Anal fistula is a challenging condition both for surgeons and patients. Recurrent fistula, Crohn's disease, or autoimmune disorders add further complexity to this situation. Numerous clinical trials have now demonstrated that cell-based therapy appears to be a good complement to fistulous surgery. As in any new treatment, especially that involving living cells, appropriate application is paramount to achieve optimal outcomes. As stem cell-based treatments are gaining a strong foothold in fistula management worldwide, we herein aim to share our mesenchymal stem cell surgical protocol. With the goal of optimizing results of this emerging therapy, we have improved and refined our protocol over the past 17 years of working with stem cells in clinical trials. The protocol consists of nine reproducible steps for mesenchymal stem cell application inside the fistulous tract, and has proven to be safe and effective in several studies, including international phase III clinical trials.

Rat model of anal sphincter injury and two approaches for stem cell administration.

AIM: To establish a rat model of anal sphincter injury and test different systems to provide stem cells to injured area. METHODS: Adipose-derived stem cells (ASCs) were isolated from BDIX rats and were transfected with green fluorescent protein (GFP) for cell tracking. Biosutures (sutures covered with ASCs) were prepared with 1.5 x 106 GFP-ASCs, and solutions of 106 GFP-ASCs in normal saline were prepared for injection. Anorectal normal anatomy was studied on Wistar and BDIX female rats. Then, we designed an anal sphincter injury model consisting of a 1-cm extra-mucosal miotomy beginning at the anal verge in the anterior middle line. The sphincter lesion was confirmed with conventional histology (hematoxylin and eosin) and immunofluorescence with 4', 6-diamidino-2-phenylindole (commonly known as DAPI), GFP and α-actin. Functional effect was assessed with basal anal manometry, prior to and after injury. After sphincter damage, 36 BDIX rats were randomized to three groups for: (1) Cell injection without repair; (2) biosuture repair; and (3) conventional suture repair and cell injection. Functional and safety studies were conducted on all the animals. Rats were sacrificed after 1, 4 or 7 d. Then, histological and immunofluorescence studies were performed on the surgical area. RESULTS: With the described protocol, biosutures had been covered with at least 820000-860000 ASCs, with 100% viability. Our studies demonstrated that some ASCs remained adhered after suture passage through the muscle. Morphological assessment showed that the rat anal anatomy is comparable with human anatomy; two sphincters are present, but the external sphincter is poorly developed. Anal sphincter pressure data showed spontaneous, consistent, rhythmic anal contractions, taking the form of "plateaus" with multiple twitches (peaks) in each pressure wave. These basal contractions were very heterogeneous; their frequency was 0.91-4.17 per min (mean 1.6980, SD 0.57698), their mean duration was 26.67 s and mean number of peaks was 12.53. Our morphological assessment revealed that with the aforementioned surgical procedure, both sphincters were completely sectioned. In manometry, the described activity disappeared and was replaced by a gentle oscillation of basal line, without a recognizable pattern. Surprisingly, these findings appeared irrespective of injury repair or not. ASCs survived in this potentially septic area for 7 d, at least. We were able to identify them in 84% of animals, mainly in the muscular section area or in the tissue between the muscular endings. ASCs formed a kind of "conglomerate" in rats treated with injections, while in the biosuture group, they wrapped the suture. ASCs were also able to migrate to the damaged zone. No relevant adverse events or mortality could be related to the stem cells in our study. We also did not find unexpected tissue growths. CONCLUSION: The proposed procedure produces a consistent sphincter lesion. Biosutures and injections are suitable for cell delivery. ASCs survive and are completely safe in this clinical setting.

UNR/CDSE1 expression as prognosis biomarker in resectable pancreatic ductal adenocarcinoma patients: A proof-of-concept.

Pancreatic ductal adenocarcinoma is an aggressive form of pancreatic cancer and the fourth leading cause of cancer-related death. When possible, curative approaches are based on surgical resection, though not every patient is a candidate for surgery. There are clinical guidelines for the management of these patients that offer different treatment options depending on the clinical and pathologic characteristics. However, the survival rates seen in this kind of patients are still low. The CDSE1 gene is located upstream of NRAS and encodes an RNA-binding protein termed UNR. The aim of this study was to analyze UNR expression and its correlation with outcome in patients with resectable pancreatic ductal adenocarcinoma (PDAC). For this, samples from resectable PDAC patients who underwent duodenopancreatectomy were used to evaluate UNR protein expression by immunohistochemistry using a tissue microarray. Here, we observed that low UNR expression was significantly associated with shorter progression-free survival after surgery (P = 0.010). Moreover, this prognostic marker remained significant after Cox proportional hazards model (P = 0.036). We further studied the role of CDSE1 expression in patient's prognosis using data from public repositories (GEO and TGCA), confirming our results. Interestingly, CDSE1 expression correlated with that of genes characteristic of an immunogenic molecular subtype of pancreatic cancer. Based on these findings, UNR may be considered a potential prognostic biomarker for resectable PDAC and may serve to guide subsequent adjuvant treatment decisions.

Treatment of Crohn's-Related Rectovaginal Fistula With Allogeneic Expanded-Adipose Derived Stem Cells: A Phase I-IIa Clinical Trial.

: The aim of this clinical trial was to determine the safety and feasibility of expanded allogeneic adipose-derived stem cells to treat Crohn's-related rectovaginal fistula (CRRVF). We designed a phase I-II clinical trial (https://ClinicalTrials.gov, NCT00999115) to treat 10 patients with CRRVF. Patients receiving biological therapy during follow-up were excluded. Curettage was performed, and a vaginal or rectal flap was added if the surgeon considered it necessary. The therapeutic protocol included intralesional injection of 20 million stem cells in the vaginal walls (submucosal area) and fistula tract. Healing was evaluated 12 weeks later. If the fistula had not healed, a second dose of 40 million stem cells was administered. Patient follow-up was 52 weeks from last cell injection. Healing was defined as re-epithelialization of both vaginal and rectal sides and absence of vaginal drainage. Cytokines and immunological blood tests were monitored. Serious adverse events or rejection issues were not observed. Five patients were excluded because biologic drugs were required to treat a Crohn's disease flare-up during follow-up. Cytokine profiles and immunotoxicity assays showed no statistically significant alterations. Sixty percent of the nonexcluded patients achieved a complete healing. Expanded allogeneic adipose-derived stem-cell injection is a safe and feasible therapy for treating CRRVF, and the healing success rate seems promising (60%). The results of this trial encourage further exploration into this therapy. SIGNIFICANCE: This may be the first publication in which allogeneic stem cells to treat rectovaginal fistula in Crohn´s disease seem to be a feasible and safe treatment. Additional studies are necessary to confirm the efficacy profile of the allogeneic stem cells strategy in a controlled design.

Adipose-derived stem cells and platelet-rich plasma for preventive treatment of bisphosphonate-related osteonecrosis of the jaw in a murine model.

OBJECTIVES: The main challenge in treating bisphosphonate-related osteonecrosis of the jaw (BRONJ) is the absence of an effective established treatment. We aimed to compare different potentially preventive treatments for BRONJ after dental extractions in zoledronic acid (ZA)-treated animals. We studied the local application of different combinations of adipose-derived stem cells (ASCs) with or without previous stimulation with bone morphogenetic protein 2 (BMP-2) and platelet-rich plasma (PRP) in rats. MATERIAL AND METHODS: Fifty-six male Wistar rats were treated with ZA for 9 weeks. Dental extractions were performed in the eighth week, and the animals were divided into 4 groups. In group 1 (n = 14), alveolar coverage with mucoperiosteal flap was performed. In group 2 (n = 14), PRP was applied over the sockets and covered with the flap. In group 3 (n = 15), allogeneic ASCs with PRP were applied and covered with the flap. In group 4 (n = 13), animals were treated with ASCs cultured with BMP-2, PRP, and flap coverage. Histologic, fluorescence, and radiologic studies of the maxillae were performed. RESULTS: ASC-treated animals showed lower frequency of osteonecrosis (14% vs 50%, p = 0.007) and greater bone turnover (p = 0.024) and osteoclast count (p = 0.045) than those not receiving the ASC treatment. CONCLUSIONS: In this high-risk model, ASC-based treatments seem to prevent BRONJ more effectively than mucosal flap with or without PRP. The combination of ASCs and PRP appears to be synergistic, and the addition of BMP-2 could further improve the results.

Coloduodenal fistula in a patient with Crohn’s disease / Fístula coloduodenal en un paciente con enfermedad de Crohn

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Sutures enriched with adipose-derived stem cells decrease the local acute inflammation after tracheal anastomosis in a murine model.

OBJECTIVES: Local inflammatory reaction and tension are the main causes of postoperative complications after tracheal surgery. Adipose-derived stem cells (ASCs) are known to have immunomodulatory activity. The exact mechanism of this activity is not known, although it is possible that they modulate the function of different cells involved in the immune response. Little is known of their impact on acute inflammation, especially in the problematic tracheal area. We aimed to study the effect of ASCs applied locally in an animal model of tracheal resection and anastomosis. METHODS: ASCs from the subcutaneous fat of BDIX rats were infected for expression of the enhanced green fluorescent protein (eGFP) and were cultured with Polyglactin 910 sutures to obtain biosutures (ASC-coated sutures). After tracheal resection, 90 BDIX rats (syngeneic, autologous model) underwent anastomosis with biosutures (1.5 10(6) cells/biosuture [Group 1] or 0.5 10(6) cells/biosuture [Group 2]) or conventional sutures (Group 3). The animals were killed after 1, 4, 10, 30 or 60 days and histological and immunofluorescence studies were performed on the anastomotic areas. Inflammatory cell densities were graded semiquantitatively by the pathologist in a blinded fashion. RESULTS: In the early period (1 and 4 days), the biosuture groups presented an atypical pattern of acute inflammation, characterized by the almost complete absence of neutrophils, and the presence of abundant lymphocytes and plasma cells, compared with the control group (P < 0.05). Moreover, abundant macrophages/monocytes were immunolocated around blood vessels near the biosutures and between biosuture threads 1 day after anastomosis, whereas the presence of macrophages/monocytes in animals treated with conventional sutures was discrete (P < 0.05). No differences were observed in the later period. No side effects in the biosuture groups were found. CONCLUSIONS: Biosutures are a comfortable way of stem cell delivery to the surgical field without modification of the operative protocol. ASCs suppress the local acute inflammatory reaction (increased macrophage migration and decreased neutrophil infiltration) in the tracheal anastomosis and cause an early switch from acute to chronic inflammation.

Stem cell therapy for digestive tract diseases: current state and future perspectives.

Diseases of the digestive tract are complex and encompass a broad spectrum of different pathogeneses (inflammatory, ischemic, neoplastic, and functional deficit). The digestive tract is not a sterile environment, and its organs are composed of tissues with different embryologic origin and different morphologic and functional complexity. As a consequence, the management of these diseases is often challenging. Stem cell (SC) therapy has yielded some promising results in preclinical studies, and, recently, some approaches have been tested clinically. Indeed, during the last 5 years, the number of clinical trials with SCs for treatment of digestive tract diseases has increased 10-fold. The most advanced programs involve liver failure, Crohn's disease, and fistulous disease and are now in phase III of development. If progress continues and the preliminary results are confirmed, SC therapy will become a clinical reality in the near future. In this review we examine the basic concepts of the SC therapy, analyze the potential benefits of SCs in diseases of the digestive tract, and summarize current experience in the field and the future perspectives.