Treatment of non-hypertrophic pseudoarthrosis of long bones with a Tissue Engineered Product loaded with autologous bone marrow-derived Mesenchymal Stromal Cells: Results from a phase IIa, prospective, randomized, parallel, pilot clinical trial comparing to iliac crest autograft.

BACKGROUND: Atrophic pseudoarthrosis is a serious complication with an incidence of 5-10 % of bone fractures located in the diaphysis of long bones. Standard treatments involve aggressive surgical procedures and re-interventions requiring the use of autografts from the iliac crest as a source of bone-forming biological activity (Standard of Care, SoC). In this context, regenerative ex vivo expanded osteogenic cell-based medicines could be of interest. Particularly, Mesenchymal Stromal Cells (MSC) offer new prospects to promote bone tissue repair in pseudoarthrosis by providing biological activity in an osteoconductive and osteoinductive environment. METHODS: We conducted a phase IIa, prospective, randomised, parallel, two-arms, open-label with blinded assessor pilot clinical trial to compare SoC vs. a tissue-engineered product (TEP), composed of autologous bone marrow (BM)-derived MSCs loaded onto allogeneic decellularised, lyophilised spongy bone cubes, in a cohort of 20 patients with non-hypertrophic pseudoarthrosis of long bones. Patients were followed up for 12 months. Radiological bone healing was evaluated by standard X-ray and computed tomography (CT) scanning. Quality of life was measured using the EUROQOL-5D questionnaire. RESULTS: Ten patients were randomized to TEP and 10 to SoC with iliac crest autograft. Manufacturing of TEP was feasible and reproducibly achieved. TEP implantation in the bone defect was successful in all cases and none of the 36 adverse events (AE) reported were related to the treatment. Efficacy analyses were performed in the Full Analysis Set (FAS) population, which included 17 patients after 3 patients withdrew from the study. The degree of consolidation, estimated by measuring Hounsfield units (HU) on CT, showed no significant differences between the two treatment groups at 12 months post treatment (main efficacy variable) (p = 0.4835) or at 6 months. CONCLUSIONS: Although only a small number of patients were included in our study, it is notable that no significant differences were observed between the experimental treatment and SoC, thus suggesting TEP as an alternative where autograft is not available or contraindicated.

PyMCGPU-IR Monte Carlo code test for occupational dosimetry.

PyMCGPU-IR is an innovative occupational dose monitoring tool for interventional radiology procedures. It reads the radiation data from the Radiation Dose Structured Report of the procedure and combines this information with the position of the monitored worker recorded using a 3D camera system. This information is used as an input file for the fast Monte Carlo radiation transport code MCGPU-IR in order to assess the organ doses, Hp(10) and Hp(0.07), as well as the effective dose. In this study, Hp(10) measurements of the first operator during an endovascular aortic aneurysm repair procedure and a coronary angiography using a ceiling suspended shield are compared to PyMCGPU-IR calculations. Differences in the two reported examples are found to be within 15%, which is considered as being very satisfactory. The study highlights the promising advantages of PyMCGPU-IR, although there are still several improvements that need to be implemented before its final clinical use.

Preclinical Development of a Therapy for Chronic Traumatic Spinal Cord Injury in Rats Using Human Wharton's Jelly Mesenchymal Stromal Cells: Proof of Concept and Regulatory Compliance.

(1) Background: the use of Mesenchymal Stromal Cells (MSC) in emerging therapies for spinal cord injury (SCI) hold the potential to improve functional recovery. However, the development of cell-based medicines is challenging and preclinical studies addressing quality, safety and efficacy must be conducted prior to clinical testing; (2) Methods: herein we present (i) the characterization of the quality attributes of MSC from the Wharton's jelly (WJ) of the umbilical cord, (ii) safety of intrathecal infusion in a 3-month subchronic toxicity assessment study, and (iii) efficacy in a rat SCI model by controlled impaction (100 kdynes) after single (day 7 post-injury) and repeated dose of 1 × 106 MSC,WJ (days 7 and 14 post-injury) with 70-day monitoring by electrophysiological testing, motor function assessment and histology evaluation; (3) Results: no toxicity associated to MSC,WJ infusion was observed. Regarding efficacy, recovery of locomotion was promoted at early time points. Persistence of MSC,WJ was detected early after administration (day 2 post-injection) but not at days 14 and 63 post-injection. (4) Conclusions: the safety profile and signs of efficacy substantiate the suitability of the presented data for inclusion in the Investigational Medicinal Product Dossier for further consideration by the competent Regulatory Authority to proceed with clinical trials.

SARS-CoV-2/COVID-19 pandemic: first wave, impact, response and lessons learnt in a fully integrated Regional Blood and Tissue Bank. A narrative report.

BACKGROUND: The COVID-19 pandemic is placing blood and tissue establishments under unprecedented stress, putting its capacity to provide the adequate care needed at risk. Here we reflect on how our integrated organisational model has faced the first impact of the pandemic and describe what challenges, opportunities and lessons have emerged. MATERIALS AND METHODS: The organisational model of the Catalan Blood and Tissue Bank (Banc de Sang i Teixits, BST) is described. The new scenario was managed by following international recommendations and considering the pandemic in a context of volatility, uncertainty, complexity, and ambiguity (VUCA), allowing rapid measures to be taken. These aimed to: ensure donor safety, promote proper responses to patients' needs, ensure the health and well-being of personnel, and prepare for future scenarios. RESULTS: The BST has adapted its activities to the changes in demand. No shortage of any product or service occurred. Donor acceptance, safety and wellbeing were maintained except for tissue donation, which almost completely stopped. To support the health system, several activities have been promoted: large-scale convalescent plasma (CP) production, clinical trials with CP and mesenchymal stromal cells, massive COVID-19 diagnoses, and participation in co-operative research and publications. Haemovigilance is running smoothly and no adverse effects have been detected among donors or patients. DISCUSSION: Several elements have proven to be critical when addressing the pandemic scenario: a) the early creation of a crisis committee in combination with technical recommendations and the recognition of a VUCA scenario; b) identification of the strategies described; c) the integrated donor-to-patient organisational model; d) active Research and Development (R&D); and e) the flexibility of the staff. It is essential to underline the importance of the need for centralised management, effective contingency strategies, and early collaboration with peers.

Randomized clinical trial: expanded autologous bone marrow mesenchymal cells combined with allogeneic bone tissue, compared with autologous iliac crest graft in lumbar fusion surgery.

BACKGROUND CONTEXT: Although autogenous iliac crest bone graft (AICBG) is considered the gold-standard graft material for spinal fusion, new bone substitutes are being developed to avoid associated complications and disadvantages. By combining autologous bone marrow mesenchymal stromal cells (MSCs) expanded ex vivo and allogenic cancellous bone graft, we obtain a tissue-engineered product that is osteoconductive and potentially more osteogenic and osteoinductive than AICBG, owing to the higher concentration of MSCs. PURPOSE: This study aimed to evaluate the feasibility and safety of implanting a tissue-engineered product consisting of expanded bone marrow MSCs loaded onto allograft bone (MSC+allograft) for spinal fusion in degenerative spine disease, as well as to assess its clinical and radiological efficacy. STUDY DESIGN/SETTING: A prospective, multicenter, open-label, blinded-reader, randomized, parallel, single-dose phase I-II clinical trial. PATIENT SAMPLE: A total of 73 adult patients from 5 hospitals, with Meyerding grade I-II L4-L5 degenerative spondylolisthesis and/or with L4-L5 degenerative disc disease who underwent spinal fusion through transforaminal lumbar interbody fusion (TLIF). OUTCOME MEASURES: Spinal fusion was assessed by plain X-ray at 3, 6, and 12 months and by computed tomography (CT) at 6 and 12 months post-treatment. An independent radiologist performed blinded assessments of all images. Clinical outcomes were measured as change from baseline value: visual analog scale for lumbar and sciatic pain at 12 days, 3, 6, and 12 months posttreatment, and Oswestry Disability Index and Short Form-36 at 3, 6, and 12 months posttreatment. METHODS: Patients who underwent L4-L5 TLIF were randomized for posterior graft type only, and received either MSC+allograft (the tissue-engineered product, group A) or AICBG (standard graft material, group B). Standard graft material was used for anterior fusion in all patients. Feasibility was measured primarily as the percentage of randomized patients who underwent surgery in each treatment group. Safety was assessed by analyzing treatment-emergent adverse events (AEs) for the full experimental phase and appraising their relationship to the experimental treatment. Outcome measures, both radiological and clinical, were compared between the groups. RESULTS: Seventy-three patients were randomized in this study, 36 from the MSC+allograft group and 37 from the AICBG group, and 65 were surgically treated (31 group A, 34 group B). Demographic and comorbidity data showed no difference between groups. Most patients were diagnosed with grade I or II degenerative spondylolisthesis. MSC+allograft was successfully implanted in 86.1% of randomized group A patients. Most patients suffered treatment-emergent AEs during the study (88.2% in group A and 97.1% in group B), none related to the experimental treatment. X-ray-based rates of posterior spinal fusion were significantly higher for the experimental group at 6 months (p=.012) and 12 months (p=.0003). CT-based posterior fusion rates were significantly higher for MSC+allograft at 6 months (92.3% vs 45.7%; p=.0001) and higher, but not significantly, at 12 months (76.5% vs 65.7%; p=.073). CT-based complete response (defined as the presence of both posterior intertransverse fusion and anterior interbody fusion) was significantly higher at 6 months for MSC+allograft than for AICBG (70.6% vs 40%; p=.0038), and remained so at 12 months (70.6% vs 51.4%; p=.023). Clinical results including patient-reported outcomes improved postsurgery, although there were no differences between groups. CONCLUSIONS: Compared with the current gold standard, our experimental treatment achieved a higher rate of posterior spinal fusion and radiographic complete response to treatment at 6 and 12 months after surgery. The treatment clearly improved patient quality of life and decreased pain and disability at rates similar to those for the control arm. The safety profile of the tissue-engineered product was also similar to that for the standard material, and no AEs were linked to the product. Procedural AEs did not increase as a result of BM aspiration. The use of expanded bone marrow MSCs combined with cancellous allograft is a feasible and effective technique for spinal fusion, with no product-related AEs found in our study.

Evaluation of a cell-based osteogenic formulation compliant with good manufacturing practice for use in tissue engineering.

Proper bony tissue regeneration requires mechanical stabilization, an osteogenic biological activity and appropriate scaffolds. The latter two elements can be combined in a hydrogel format for effective delivery, so it can readily adapt to the architecture of the defect. We evaluated a Good Manufacturing Practice-compliant formulation composed of bone marrow-derived mesenchymal stromal cells in combination with bone particles (Ø = 0.25 to 1 µm) and fibrin, which can be readily translated into the clinical setting for the treatment of bone defects, as an alternative to bone tissue autografts. Remarkably, cells survived with unaltered phenotype (CD73+, CD90+, CD105+, CD31-, CD45-) and retained their osteogenic capacity up to 48 h after being combined with hydrogel and bone particles, thus demonstrating the stability of their identity and potency. Moreover, in a subchronic toxicity in vivo study, no toxicity was observed upon subcutaneous administration in athymic mice and signs of osteogenesis and vascularization were detected 2 months after administration. The preclinical data gathered in the present work, in compliance with current quality and regulatory requirements, demonstrated the feasibility of formulating an osteogenic cell-based tissue engineering product with a defined profile including identity, purity and potency (in vitro and in vivo), and the stability of these attributes, which complements the preclinical package required prior to move towards its use of prior to its clinical use.

First-in-human PeriCord cardiac bioimplant: Scalability and GMP manufacturing of an allogeneic engineered tissue graft.

BACKGROUND: Small cardiac tissue engineering constructs show promise for limiting post-infarct sequelae in animal models. This study sought to scale-up a 2-cm2 preclinical construct into a human-size advanced therapy medicinal product (ATMP; PeriCord), and to test it in a first-in-human implantation. METHODS: The PeriCord is a clinical-size (12-16 cm2) decellularised pericardial matrix colonised with human viable Wharton's jelly-derived mesenchymal stromal cells (WJ-MSCs). WJ-MSCs expanded following good manufacturing practices (GMP) met safety and quality standards regarding the number of cumulative population doublings, genomic stability, and sterility. Human decellularised pericardial scaffolds were tested for DNA content, matrix stiffness, pore size, and absence of microbiological growth. FINDINGS: PeriCord implantation was surgically performed on a large non-revascularisable scar in the inferior wall of a 63-year-old male patient. Coronary artery bypass grafting was concomitantly performed in the non-infarcted area. At implantation, the 16-cm2 pericardial scaffold contained 12·5 × 106 viable WJ-MSCs (85·4% cell viability; <0·51 endotoxin units (EU)/mL). Intraoperative PeriCord delivery was expeditious, and secured with surgical glue. The post-operative course showed non-adverse reaction to the PeriCord, without requiring host immunosuppression. The three-month clinical follow-up was uneventful, and three-month cardiac magnetic resonance imaging showed ~9% reduction in scar mass in the treated area. INTERPRETATION: This preliminary report describes the development of a scalable clinical-size allogeneic PeriCord cardiac bioimplant, and its first-in-human implantation. FUNDING: La Marató de TV3 Foundation, Government of Catalonia, Catalan Society of Cardiology, "La Caixa" Banking Foundation, Spanish Ministry of Science, Innovation and Universities, Institute of Health Carlos III, and the European Regional Development Fund.

Enhanced fully cellulose based forward and reverse blood typing assay.

This study presents an enhanced paper-based analytical device (PAD) for forward and reverse group blood typing. The proposed PAD uses a novel methodology, which provides highly reliable results on a fully cellulose based device. The PAD was printed on different cellulose substrates. These substrates were made of different cellulose fibers (sisal and eucalyptus), different grammages, refining steps, and wet additive content. Best parameters were chosen to achieve high reliability on both forward and reverse blood typing. The substrates were patterned with five hydrophilic channels and two hydrophobic areas. For reverse blood typing, the hemoagglutination reaction took place on the hydrophobic surface of the paper before being transferred to the paper web, where together with the forward blood typing tests were all washed with saline solution to read the results by elution. This device allows direct read-out of results; the stains show were agglutination happens. Different blood types were in full agreement between the reverse and forward method and in agreement with traditional methods. The time and simplicity of this methodology confirmed its utility.

A novel, fully digital approach to quantifying volume gain after soft tissue graft surgery. A pilot study.

OBJECTIVE: Measuring soft tissue thickness after mucogingival surgery has traditionally been performed by means of a calibrated transgingival probe. The main aim of this study was to apply a non-invasive technique based on digital images formatted as Standard Tessellation Language (STL) files to quantify soft tissue volume after connective tissue grafting. CLINICAL INNOVATION REPORT: Ten patients who presented Cairo Class I gingival recession were selected for connective tissue grafting using the tunnel technique. In all patients, the initial position of the gingiva and quantity of keratinized tissue were recorded, and gingival recession was scanned with an intra-oral scanner. Six months after surgery, the same intra-oral parameters were recorded and compared with the initial registers using digital volumetric analysis software. RESULTS: Complete root coverage was obtained in most patients (90%), mean coverage being of 2.70 mm with a mean increase in volume of 115.49 mm3 in the treated areas. No pattern was identified that indicated a statistically significant relation between gingival recession and coverage volume in mm3 . CONCLUSIONS: Digital processing of pre- and post-treatment images makes it possible to measure the volume of tissue gained after tissue graft surgery simply and non-invasively. The technique is an objective and reproducible method for measuring soft tissue thickness.

Diagnostic Accuracy of CYFRA21-1 in the Differential Diagnosis of Pleural Effusions.

BACKGROUND/AIM: Approximately 20% of pleural effusions are associated with cancer; about 50% require invasive procedures to perform diagnosis. Determination of the concentration of soluble cytokeratin 19-fragments (CYFRA21-1) may help identify patients with malignant effusions. However, pathologies other than cancer can increase its concentration. The identification of these possible false positives with routine tests CRP, ADA, % polymorphonuclear cells (PN) may improve diagnostic accuracy. This study aimed to determine the diagnostic accuracy of CYFRA21-1 in the detection of malignant pleural effusions and the possible false positives. MATERIALS AND METHODS: Analysis of CYFRA21-1, adenosine deaminase (ADA), C-reactive protein (CRP), and the percentage of polymorphonuclear leukocytes (PN%) in the fluid from 643 consecutive undiagnosed pleural effusions was performed. RESULTS: CYFRA21-1 showed 38.7% sensitivity and 97.3% specificity at 175 ng/ml cut-off. Effusions not suspicious of a false-positive showed 39.0% sensitivity and 98.2% specificity, while effusions suspicious of false positive showed lower sensitivity (36.4%) and specificity (95.0%). CONCLUSION: The diagnostic accuracy of CYFRA21-1 in pleural effusions can be improved by classification according to the possibility of false positives.

Dissection of the inferior mesenteric vein versus of the inferior mesenteric artery for the genitourinary function after laparoscopic approach of rectal cancer surgery: a randomized controlled trial.

BACKGROUND: Total Mesorectal Excision (TME) is the standard surgical technique for the treatment of rectal cancer. However, rates of sexual dysfunction ofup to 50% have been described after TME, and rates of urinary dysfunction of up to 30%. Although other factors are involved, the main cause of postoperative genitourinary dysfunction is intraoperative injury to the pelvic autonomic nerves. The risk is particularly high in the inferior mesenteric artery (IMA). The aim of this study is to compare pre- and post-TME sexual dysfunction, depending on the surgical approach usedin the inferior mesenteric vessels: either directly on the IMA, or from the inferior mesenteric vein (IMV) to the IMA. METHODS: Prospective, randomized,controlled study of patients with rectal adenocarcinoma with neoadjuvant chemoradiotherapy, who will be randomly assigned to one of two groups depending on the surgical approach to the inferior mesenteric vessels. The main variable is pre- and postoperative sexual dysfunction; secondary variables are visualization and preservation of the pelvic autonomic nerves, pre- and postoperative urinary dysfunction, and pre- and postoperative quality of life. The sample will comprise 90 patients, 45 per group. DISCUSSION: The aim is to demonstrate that the dissection route from the IMV towards the IMA favors the preservation of the pelvic autonomic nerves and thus reducesrates of sexual dysfunction post-surgery. TRIAL REGISTRATION: Ethical and Clinical Research Committee, Parc Taulí University Hospital: ID 017/315. ClinicalTrials.gov TAU-RECTALNERV-PRESERV-2018 (TRN: NCT03520088 ) (Date of registration 04/03/2018).

Compliance with Good Manufacturing Practice in the Assessment of Immunomodulation Potential of Clinical Grade Multipotent Mesenchymal Stromal Cells Derived from Wharton's Jelly.

BACKGROUND: The selection of assays suitable for testing the potency of clinical grade multipotent mesenchymal stromal cell (MSC)-based products and its interpretation is a challenge for both developers and regulators. Here, we present a bioprocess design for the production of Wharton's jelly (WJ)-derived MSCs and a validated immunopotency assay approved by the competent regulatory authority for batch release together with the study of failure modes in the bioprocess with potential impact on critical quality attributes (CQA) of the final product. Methods: The lymphocyte proliferation assay was used for determining the immunopotency of WJ-MSCs and validated under good manufacturing practices (GMP). Moreover, failure mode effects analysis (FMEA) was used to identify and quantify the potential impact of different unexpected situations on the CQA. Results: A production process based on a two-tiered cell banking strategy resulted in batches with sufficient numbers of cells for clinical use in compliance with approved specifications including MSC identity (expressing CD73, CD90, CD105, but not CD31, CD45, or HLA-DR). Remarkably, all batches showed high capacity to inhibit the proliferation of activated lymphocytes. Moreover, implementation of risk management tools led to an in-depth understanding of the manufacturing process as well as the identification of weak points to be reinforced. Conclusions: The bioprocess design showed here together with detailed risk management and the use of a robust method for immunomodulation potency testing allowed for the robust production of clinical-grade WJ-MSCs under pharmaceutical standards.

Influence of Transmucosal Area Morphology on Peri-Implant Bone Loss in Tissue-Level Implants.

PURPOSE: To analyze peri-implant bone loss around two types of tissue-level implants (convergent and divergent transmucosal morphology) placed in the same region of either the maxilla or mandible in a single surgical session. MATERIALS AND METHODS: This prospective study included 21 patients who each received two implants, one with a convergent transmucosal collar (n = 21) the other with a divergent collar (n = 21). Implants were placed in a single surgical session, by the same clinician, in the same clinical setting at the Prosthodontic and Occlusion Unit, University of Valencia (Spain). The implants (n = 42) were restored with splinted crowns screwed directly onto the prosthetic platforms. Bone loss analysis consisted of measurements taken from periapical radiographs 24 months after loading. Statistical analysis evaluated the homogeneity of the groups using Pearson's chisquare test; bone loss was compared with the Wilcoxon test. Statistical significance was set at 5% (α = .05), with a confidence interval of 95% and a power of 57%. RESULTS: Implants with convergent transmucosal morphology presented less mean peri-implant bone loss (0.29 ± 0.34 mm) than those with divergent transmucosal morphology (0.60 ± 0.63 mm), with a statistically significant difference between the two types of implants (P = .031). CONCLUSION: Less peri-implant bone loss occurs around supracrestal implants with convergent transmucosal morphology than divergent transmucosal morphology.

Influence of Implant-Prosthetic Connection on Peri-Implant Bone Loss: A Prospective Clinical Trial with 2-Year Follow-up.

PURPOSE: To analyze peri-implant bone loss around six different types of implant-prosthetic connections over a 2-year follow-up period. MATERIALS AND METHODS: A total of 120 implants were divided into six groups (n = 20), three with crestally and three with supracrestally placed implants, with different internal connections, placed in posterior sectors (molar and premolar), and bearing one-piece CAD/CAM restorations screwed directly to each implant's prosthetic platform. Bone height was measured from parallelized periapical radiographs taken at the moment of loading and 2 years later. RESULTS: The patient sample included 61 subjects, 36 men and 25 women. Significant differences were found in bone loss between groups, the implant with internal hex and convergent transmucosal collar showing the least bone loss (P < .001) in comparison with the other designs. Supracrestally placed implants underwent less bone loss than crestally placed implants (P = .025). CONCLUSION: Peri-implant bone loss is influenced by the level of implant placement in relation to the bone crest (crestal or supracrestal) and by the morphology of the prosthetic platform.

Levels of IL-17F and IL-33 correlate with HLA-DR activation in clinical-grade human bone marrow-derived multipotent mesenchymal stromal cell expansion cultures.

BACKGROUND AIMS: Multipotent mesenchymal stromal cell (MSC)-based medicines are extensively investigated for use in regenerative medicine and immunotherapy applications. The International Society for Cell and Gene Therapy (ISCT) proposed a panel of cell surface molecules for MSC identification that includes human leukocyte antigen (HLA)-DR as a negative marker. However, its expression is largely unpredictable despite production under tightly controlled conditions and compliance with current Good Manufacturing Practices. Herein, we report the frequency of HLA-DR expression in 81 batches of clinical grade bone marrow (BM)-derived MSCs and investigated its impact on cell attributes and culture environment. METHODS: The levels of 15 cytokines (interleukin [IL]-1ß, IL-4, IL-6, IL-10, IL-17A, IL-17F, IL-21, IL-22, IL-23, IL-25, IL-31, IL-33, interferon-γ, soluble CD40 ligand and tumor necrosis factor-α) were determined in sera supplements and supernatants of BM-MSC cultures. Identity, multipotentiality and immunopotency assays were performed on high (>20% of cells) and low (≤20% of cells) HLA-DR+ cultures. RESULTS: A correlation was found between HLA-DR expression and levels of IL-17F and IL-33. Expression of HLA-DR did neither affect MSC identity, in vitro tri-lineage differentiation potential (into osteogenic, chondrogenic and adipogenic lineages), nor their ability to inhibit the proliferation of stimulated lymphocytes. DISCUSSION: Out of 81 batches of BM-MSCs for autologous use analyzed, only three batches would have passed the ISCT criteria (<2%), whereas 60.5% of batches were compliant with low HLA-DR values (≤20%). Although a cause-effect relationship cannot be drawn, we have provided a better understanding of signaling events and cellular responses in expansion culture conditions relating with HLA-DR expression.

The Place of Transanal Endoscopic Surgery in the Treatment of Rectourethral Fistula.

OBJECTIVE: To assess the role of transanal endoscopic operation (TEO) or transanal endoscopic microsurgery (TEM) in rectourethral fistulas (RUF). RUF may appear after radical prostatectomy. Their treatment represents a challenge; many therapies have been proposed, from conservative to aggressive surgical approaches. Transanal endoscopic surgery (TEO or TEM) is a minimally invasive technique to access the site of the RUF to perform repair. MATERIALS AND METHODS: This is an observational study with prospective data collection, conducted between September 2006 and December 2015. All patients were diagnosed with RUF following management of prostate cancer. Conservative treatment was administered in the form of urinary and fecal diversion with cystotomy and terminal colostomy, to achieve total urinary and fecal exclusion. If the fistula persisted, it was treated by TEO or TEM, with or without biological mesh interposition. If this failed, gracilis muscle was applied as salvage therapy. RESULTS: Ten patients were diagnosed with RUF. In 1 patient (1 of 10), the fistula healed with bladder catheterization alone. In another patient (1 of 9), it resolved after total urinary and fecal exclusion. Eight patients underwent repair by TEO or TEM, 4 with biological mesh interposition; all 4 presented recurrence. In the other 4 patients treated via TEO or TEM, 2 had early recurrence, whereas the others had healed at follow-up visits after 4-6 months (2 of 8)-a success rate of 25%. The 6 patients who recurred were treated with gracilis muscle interposition via a transperineal approach. CONCLUSION: The low rate of positive results obtained by TEO or TEM argues against its use as technique of choice in RUF, and against the use of biological meshes.

Clinical translation of a mesenchymal stromal cell-based therapy developed in a large animal model and two case studies of the treatment of atrophic pseudoarthrosis.

Pseudoarthrosis is a relatively frequent complication of fractures, in which the lack of mechanical stability and biological stimuli results in the failure of bone union, most frequently in humerus and tibia. Treatment of recalcitrant pseudoarthrosis relies on the achievement of satisfactory mechanical stability combined with adequate local biology. Herein we present two cases of atrophic pseudoarthrosis that received a tissue-engineering product (TEP) composed of autologous bone marrow-derived mesenchymal stromal cells (BM-MSC) combined with deantigenized trabecular bone particles from a tissue bank. The feasibility of the treatment and osteogenic potential of the cell-based medicine was first demonstrated in an ovine model of critical size segmental tibial defect. Clinical-grade autologous BM-MSC were produced following a good manufacturing practice-compliant bioprocess. Results were successful in one case, with pseudoarthrosis resolution, and inconclusive in the other one. The first patient presented atrophic pseudoarthrosis of the humeral diaphysis and was treated with osteosynthesis and TEP resulting in satisfactory consolidation at month 6. The second case presented a recalcitrant pseudoarthrosis of the proximal tibia and the Masquelet technique was followed before filling the defect with the TEP. This patient presented a neuropathic pain syndrome unrelated to the treatment that forced the amputation of the extremity 3 months later. In this case, the histological analysis of the tissue formed at the defect site provided evidence of neovascularization but no overt bone remodelling activity. It is concluded that the use of expanded autologous BM-MSC to treat pseudoarthrosis was demonstrated to be feasible and safe, provided that no clinical complications were reported, and early signs of effectiveness were observed. Copyright © 2016 John Wiley & Sons, Ltd.

Assessment of weaning indexes based on diaphragm activity in mechanically ventilated subjects after cardiovascular surgery. A pilot study. / Avaliação dos índices de desmame com base na atividade do diafragma em pacientes submetidos à ventilação mecânica após cirurgia cardiovascular. Um estudo piloto.

OBJECTIVE: The aim of this pilot study was to evaluate the feasibility of surface electromyographic signal derived indexes for the prediction of weaning outcomes among mechanically ventilated subjects after cardiac surgery. METHODS: A sample of 10 postsurgical adult subjects who received cardiovascular surgery that did not meet the criteria for early extubation were included. Surface electromyographic signals from diaphragm and ventilatory variables were recorded during the weaning process, with the moment determined by the medical staff according to their expertise. Several indexes of respiratory muscle expenditure from surface electromyography using linear and non-linear processing techniques were evaluated. Two groups were compared: successfully and unsuccessfully weaned patients. RESULTS: The obtained indexes allow estimation of the diaphragm activity of each subject, showing a correlation between high expenditure and weaning test failure. CONCLUSION: Surface electromyography is becoming a promising procedure for assessing the state of mechanically ventilated patients, even in complex situations such as those that involve a patient after cardiovascular surgery.

OBJETIVO: Avaliar a viabilidade do uso de índices derivados do sinal de eletromiografia de superfície para predizer desfechos do processo de desmame em pacientes mecanicamente ventilados após cirurgia cardíaca. MÉTODOS: Foram incluídos dez pacientes em pós-operatório de cirurgia cardiovascular que não cumpriram os critérios para extubação precoce. Os sinais da eletromiografia de superfície foram registrados, assim como as variáveis ventilatórias durante o processo de desmame, sendo o momento do procedimento determinado pela equipe médica, segundo sua experiência. Avaliaram-se diversos índices da atividade dos músculos respiratórios obtidos a partir da eletromiografia de superfície com uso de técnicas de processamento lineares e não lineares. Compararam-se dois grupos: pacientes com e sem sucesso no desmame. RESULTADOS: Os índices obtidos permitiram estimar a atividade diafragmática de cada paciente, demonstrando uma correlação entre atividade elevada e falha do teste de desmame. CONCLUSÃO: A eletromiografia de superfície está se tornando um procedimento promissor para avaliar as condições de pacientes ventilados mecanicamente, mesmo em condições complexas, como as que envolvem aqueles após cirurgia cardiovascular.

Assessment of biodistribution using mesenchymal stromal cells: Algorithm for study design and challenges in detection methodologies.

BACKGROUND AIMS: Biodistribution of candidate cell-based therapeutics is a critical safety concern that must be addressed in the preclinical development program. We aimed to design a decision tree based on a series of studies included in actual dossiers approved by competent regulatory authorities, noting that the design, execution and interpretation of pharmacokinetics studies using this type of therapy is not straightforward and presents a challenge for both developers and regulators. METHODS: Eight studies were evaluated for the definition of a decision tree, in which mesenchymal stromal cells (MSCs) were administered to mouse, rat and sheep models using diverse routes (local or systemic), cell labeling (chemical or genetic) and detection methodologies (polymerase chain reaction [PCR], immunohistochemistry [IHC], fluorescence bioimaging, and magnetic resonance imaging [MRI]). Moreover, labeling and detection methodologies were compared in terms of cost, throughput, speed, sensitivity and specificity. RESULTS: A decision tree was defined based on the model chosen: (i) small immunodeficient animals receiving heterologous MSC products for assessing biodistribution and other safety aspects and (ii) large animals receiving homologous labeled products; this contributed to gathering data not only on biodistribution but also on pharmacodynamics. PCR emerged as the most convenient technique despite the loss of spatial information on cell distribution that can be further assessed by IHC. DISCUSSION: This work contributes to the standardization in the design of biodistribution studies by improving methods for accurate assessment of safety. The evaluation of different animal models and screening of target organs through a combination of techniques is a cost-effective and timely strategy.