On the relationship between viscoelasticity and water diffusion in soft biological tissues.

Magnetic resonance elastography (MRE) and diffusion-weighted imaging (DWI) are complementary imaging techniques that detect disease based on viscoelasticity and water mobility, respectively. However, the relationship between viscoelasticity and water diffusion is still poorly understood, hindering the clinical translation of combined DWI-MRE markers. We used DWI-MRE to study 129 biomaterial samples including native and cross-linked collagen, glycosaminoglycans (GAGs) with different sulfation levels, and decellularized specimens of pancreas and liver, all with different proportions of solid tissue, or solid fractions. We developed a theoretical framework of the relationship between mechanical loss and tissue-water mobility based on two parameters, solid and fluid viscosity. These parameters revealed distinct DWI-MRE property clusters characterizing weak, moderate, and strong water-network interactions. Sparse networks interacting weakly with water, such as collagen or diluted decellularized tissue, resulted in marginal changes in water diffusion over increasing solid viscosity. In contrast, dense networks with larger solid fractions exhibited both free and hindered water diffusion depending on the polarity of the solid components. For example, polar and highly sulfated GAGs as well as native soft tissues hindered water diffusion despite relatively low solid viscosity. Our results suggest that two fundamental properties of tissue networks, solid fraction and network polarity, critically influence solid and fluid viscosity in biological tissues. Since clinical DWI and MRE are sensitive to these viscosity parameters, the framework we present here can be used to detect tissue remodeling and architectural changes in the setting of diagnostic imaging. STATEMENT OF SIGNIFICANCE: The viscoelastic properties of biological tissues provide a wealth of information on the vital state of cells and host matrix. Combined measurement of viscoelasticity and water diffusion by medical imaging is sensitive to tissue microarchitecture. However, the relationship between viscoelasticity and water diffusion is still poorly understood, hindering full exploitation of these properties as a combined clinical biomarker. Therefore, we analyzed the parameter space accessible by diffusion-weighted imaging (DWI) and magnetic resonance elastography (MRE) and developed a theoretical framework for the relationship between water mobility and mechanical parameters in biomaterials. Our theory of solid material properties related to particle motion can be translated to clinical radiology using clinically established MRE and DWI.

[Position paper of the Berlin-Brandenburg Surgical Society-On the future of surgical further training]. / Positionspapier der Berlin-Brandenburgischen Chirurgischen Gesellschaft ­ zur Zukunft der chirurgischen Weiterbildung.

BACKGROUND: Surgical further training faces the challenging task of reconciling technological advancements and patient safety, particularly in the context of the planned hospital reform. Additionally, the generation shift and evolving expectations of Generations Y and Z in the workplace present further challenges. In response to these demands, the Berlin-Brandenburg Surgical Society (Berlin-Brandenburgische Chirurgische Gesellschaft, BCG) initiated a structured discussion and developed a position paper during the Neuhardenberg talks (Neuhardenberger Gespräche). METHODOLOGY: Within the framework of the Neuhardenberg talks, four sessions with keynote presentations and discussions took place. Based on the main discussion points, theses and positions were subsequently formulated and digitally voted on. RESULTS: The results reveal a clear consensus favoring flexible working hours models, earlier specialization options and the integration of external rotations in surgical further training. Regarding talent acquisition and early recruitment of residents, there was a clear consensus supporting the promotion of employee engagement and structured early recruitment of students. There was unanimous agreement on the introduction of training associations as an effective means to ensure high-quality surgical further training. DISCUSSION: One of the central points in the discussions was that high-quality surgical further training will only be achievable within training associations, especially given the impending hospital reform. The BCG plans to develop a modular further training association to make surgical further training in Berlin/Brandenburg fit for the future.

From morbidity reduction to cost-effectiveness: Enhanced recovery after surgery (ERAS) society recommendations in minimal invasive liver surgery.

PURPOSE: Minimal-invasive liver surgery (MILS) reduces surgical trauma and is associated with fewer postoperative complications. To amplify these benefits, perioperative multimodal concepts like Enhanced Recovery after Surgery (ERAS), can play a crucial role. We aimed to evaluate the cost-effectiveness for MILS in an ERAS program, considering the necessary additional workforce and associated expenses. METHODS: A prospective observational study comparing surgical approach in patients within an ERAS program compared to standard care from 2018-2022 at the Charité - Universitätsmedizin Berlin. Cost data were provided by the medical controlling office. ERAS items were applied according to the ERAS society recommendations. RESULTS: 537 patients underwent liver surgery (46% laparoscopic, 26% robotic assisted, 28% open surgery) and 487 were managed by the ERAS protocol. Implementation of ERAS reduced overall postoperative complications in the MILS group (18% vs. 32%, p = 0.048). Complications greater than Clavien-Dindo grade II incurred the highest costs (€ 31,093) compared to minor (€ 17,510) and no complications (€13,893; p < 0.001). In the event of major complications, profit margins were reduced by a median of € 6,640. CONCLUSIONS: Embracing the ERAS society recommendations in liver surgery leads to a significant reduction of complications. This outcome justifies the higher cost associated with a well-structured ERAS protocol, as it effectively offsets the expenses of complications.

Optimizing environmental enrichment for Sprague Dawley rats: Exemplary insights into the liver proteome.

BACKGROUND: Considering the expected increase in the elderly population and the growing emphasis on aging-related biomedical research, the demand for aged laboratory animals has surged, challenging established husbandry practices. Our objective was to establish a cost-effective method for environmental enrichment, utilizing the liver as a representative organ to assess potential metabolic changes in response to differing enrichment levels. METHODS: We conducted a six-month study involving 24 male Sprague Dawley rats, randomly assigned to four environmental enrichment groups. Two groups were housed in standard cages, while the others were placed in modified rabbit cages. Half of the groups received weekly playtime in an activity focused rat housing unit. We evaluated hormone levels, playtime behavior, and subjective handling experience. Additionally, liver tissue proteomic analysis was performed. RESULTS: Initial corticosterone levels and those after 3 and 6 months showed no significant differences. Yet, testosterone levels were lower in the control group by the end of the study (p = 0.007). We observed 1871 distinct proteins in liver tissue, with 77% being common across groups. In gene ontology analysis, no specific pathways were overexpressed. In semiquantitative analysis, we observed differences in proteins associated in lipid metabolism such as Apolipoprotein A-I and Acyl-CoA 6-desaturase, which were lower in the control group (p = 0.024 and p = 0.009). Rats in the intervention groups with weekly playtime displayed the least amount of reported distress during inspection or upon room entry and were less prone to accepting treats. Removing animals from their enclosure was most effortless for those in the large cage group. Over time, there was a decrease in conflicts among rats that interacted only twice weekly during playpen time. DISCUSSION: In summary, refining husbandry practices for aging rats is both simple and budget-friendly, with no apparent adverse effects on stress levels, animal development, or relevant metabolic changes in the liver.

Extended Right Hepatectomy following Clearance of the Left Liver Lobe and Portal Vein Embolization for Curatively Intended Treatment of Extensive Bilobar Colorectal Liver Metastases: A Single-Center Case Series.

BACKGROUND: Two-staged hepatectomy (TSH) including portal vein embolization (PVE) may offer surgical treatment for extensive bilobar colorectal liver metastases (CRLM). This study aimed to investigate the feasibility and outcomes of extended right hepatectomy (ERH) within TSH including PVE for patients with extended CRLM. METHODS: We retrospectively collected data of patients who underwent TSH for extended CRLM between 2015 and 2021 at our institution. Clearance of the left liver lobe (clear-up, CU) associated with PVE was followed by ERH. RESULTS: Minimally invasive (n = 12, 46%, MIH) or open hepatectomy (n = 14, 54%, OH) was performed. Postoperative major morbidity and 90-day mortality were 54% and 0%. Three-year overall survival was 95%. Baseline characteristics, postoperative and long-term outcomes were comparable between MIH and OH. However, hospital stay was significantly shorter after MIH (8 vs. 15 days, p = 0.008). Additionally, the need for intraoperative transfusions tended to be lower in the MIH group (17% vs. 50%, p = 0.110). CONCLUSIONS: ERH following CU and PVE for extended CRLM is feasible and safe in laparoscopic and open approaches. MIH for ERH may result in shorter postoperative hospital stays. Further high-volume, multicenter studies are required to evaluate the potential superiority of MIH.

Proteomic analysis of decellularized mice liver and kidney extracellular matrices.

BACKGROUND: The extracellular matrix (ECM) is a three-dimensional network of proteins that encases and supports cells within a tissue and promotes physiological and pathological cellular differentiation and functionality. Understanding the complex composition of the ECM is essential to decrypt physiological processes as well as pathogenesis. In this context, the method of decellularization is a useful technique to eliminate cellular components from tissues while preserving the majority of the structural and functional integrity of the ECM. RESULTS: In this study, we employed a bottom-up proteomic approach to elucidate the intricate network of proteins in the decellularized extracellular matrices of murine liver and kidney tissues. This approach involved the use of a novel, perfusion-based decellularization protocol to generate acellular whole organ scaffolds. Proteomic analysis of decellularized mice liver and kidney ECM scaffolds revealed tissue-specific differences in matrisome composition, while we found a predominantly stable composition of the core matrisome, consisting of collagens, glycoproteins, and proteoglycans. Liver matrisome analysis revealed unique proteins such as collagen type VI alpha-6, fibrillin-2 or biglycan. In the kidney, specific ECM-regulators such as cathepsin z were detected. CONCLUSION: The identification of distinct proteomic signatures provides insights into how different matrisome compositions might influence the biological properties of distinct tissues. This experimental workflow will help to further elucidate the proteomic landscape of decellularized extracellular matrix scaffolds of mice in order to decipher complex cell-matrix interactions and their contribution to a tissue-specific microenvironment.

miRNA as potential biomarkers after liver transplantation: A systematic review.

BACKGROUND: Liver transplantation is a life-saving therapy for end-stage liver disease patients, but acute cellular rejection (ACR) and graft complications remain significant postoperative challenges. Early and accurate diagnosis is crucial for timely intervention and improved patient outcomes, but their diagnosis rely currently on invasive biopsy sampling, thus prompting the search for non-invasive Biomarkers. MicroRNA (miRNA) have emerged as promising biomarkers in various pathological conditions, and their potential utility in diagnosing acute cellular rejection after liver transplantation has gained significant interest. METHODS: This systematic review of PubMed, Web of Science, and the ClinicalTrials.gov registry analyzes studies exploring miRNA as biomarkers for ACR and graft dysfunction in liver transplantation (PROSPERO ID CRD42023465278). The Cochrane Collaboration tool for assessing risk of bias was employed. Population data, identified miRNA and their dynamic regulation, as well as event prediction were compared. Data extraction and quality assessment were performed independently by two reviewers. RESULTS: Thirteen studies were included in this systematic review. Various investigated miRNAs were upregulated in association with acute cellular rejection, like miR-122, miR-155, miR-181, miR-483-3p, and miR-885-5p, demonstrating great biomarker potential. Additionally, several studies conducted target gene analysis, revealing insights into cellular mechanisms linked to ACR. Moreover, various miRNA were also capable of predicting different organ complications following transplantation, expanding their versatility. Remaining challenges include the standardization of miRNA profiling, the need for functional validation, and the necessity for long-term studies. CONCLUSION: The results highlight the potential of miRNA as specific, non-invasive biomarkers for ACR and graft dysfunction following liver transplantation. However, further research is needed to validate these findings and establish standardized diagnostic panels to incorporate them into clinical practice and explore miRNA-based therapies in the future.

Impact of complexity in minimally invasive liver surgery on enhanced recovery measures: prospective study.

BACKGROUND: Adherence to enhanced recovery after surgery (ERAS) protocols is crucial for successful liver surgery. The aim of this study was to assess the impact of minimally invasive liver surgery complexity on adherence after implementing an ERAS protocol. METHODS: Between July 2018 and August 2021, a prospective observational study involving minimally invasive liver surgery patients was conducted. Perioperative treatment followed ERAS guidelines and was recorded in the ERAS interactive audit system. Kruskal-Wallis and ANOVA tests were used for analysis, and pairwise comparisons utilized Wilcoxon rank sum and Welch's t-tests, adjusted using Bonferroni correction. RESULTS: A total of 243 patients were enrolled and categorized into four groups based on the Iwate criteria: low (n = 17), intermediate (n = 81), advanced (n = 74) and expert difficulty (n = 71). Complexity correlated with increased overall and major morbidity rate, as well as longer length of stay (all P < 0.001; standardized mean difference = 0.036, 0.451, 0.543 respectively). Adherence to ERAS measures decreased with higher complexity (P < 0.001). Overall adherence was 65.4%. Medical staff-centred adherence was 79.9%, while patient-centred adherence was 38.9% (P < 0.001). Complexity significantly affected patient-centred adherence (P < 0.001; standardized mean difference (SMD) = 0.420), but not medical staff-centred adherence (P = 0.098; SMD = 0.315). Postoperative phase adherence showed major differences among complexity groups (P < 0.001, SMD = 0.376), with mobilization measures adhered to less in higher complexity cases. CONCLUSION: The complexity of minimally invasive liver surgery procedures impacts ERAS protocol adherence for each patient. This can be addressed using complexity-adjusted cut-offs and 'gradual adherence' based on the relative proportion of cut-off values achieved.

Minimal Invasive Versus Open Surgery for Colorectal Liver Metastases: A Multicenter German StuDoQ|Liver Registry-Based Cohort Analysis in Germany.

Objective: To compare the outcome of minimally invasive liver surgery (MILS) to open liver surgery (OLS) for resection of colorectal liver metastases (CRLM) on a nationwide level. Background: Colorectal cancer is the third most common malignancy worldwide. Up to 50% of all patients with colorectal cancer develop CRLM. MILS represents an attractive alternative to OLS for treatment of CRLM. Methods: Retrospective cohort study using the prospectively recorded German Quality management registry for liver surgery. Propensity-score matching was performed to account for variance in the extent of resection and patient demographics. Results: In total, 1037 patients underwent liver resection for CRLM from 2019 to 2021. MILS was performed in 31%. Operative time was significantly longer in MILS (234 vs 222 minutes, P = 0.02) compared with OLS. After MILS, median length of hospital stay (LOS) was significantly shorter (7 vs 10 days; P < 0.001). Despite 76% of major resections being OLS, postoperative complications and 90-day morbidity and mortality did not differ. The Pringle maneuver was more frequently used in MILS (48% vs 40%, P = 0.048). After propensity-score matching for age, body mass index, Eastern Cooperative Oncology Group, and extent of resection, LOS remained shorter in the MILS cohort (6 vs 10 days, P < 0.001) and operative time did not differ significantly (P = 0.2). Conclusion: MILS is not the standard for resection of CRLM in Germany. Drawbacks, such as a longer operative time remain. However, if technically possible, MILS is a reasonable alternative to OLS for resection of CRLM, with comparable postoperative complications, reduced LOS, and equal oncological radicality.

Rise of tissue- and species-specific 3D bioprinting based on decellularized extracellular matrix-derived bioinks and bioresins.

Thanks to its natural complexity and functionality, decellularized extracellular matrix (dECM) serves as an excellent foundation for creating highly cell-compatible bioinks and bioresins. This enables the bioprinted cells to thrive in an environment that closely mimics their native ECM composition and offers customizable biomechanical properties. To formulate dECM bioinks and bioresins, one must first pulverize and/or solubilize the dECM into non-crosslinked fragments, which can then be chemically modified as needed. In bioprinting, the solubilized dECM-derived material is typically deposited and/or crosslinked in a layer-by-layer fashion to build 3D hydrogel structures. Since the introduction of the first liver-derived dECM-based bioinks, a wide variety of decellularized tissue have been employed in bioprinting, including kidney, heart, cartilage, and adipose tissue among others. This review aims to summarize the critical steps involved in tissue-derived dECM bioprinting, starting from the decellularization of the ECM to the standardized formulation of bioinks and bioresins, ultimately leading to the reproducible bioprinting of tissue constructs. Notably, this discussion also covers photocrosslinkable dECM bioresins, which are particularly attractive due to their ability to provide precise spatiotemporal control over the gelation in bioprinting. Both in extrusion printing and vat photopolymerization, there is a need for more standardized protocols to fully harness the unique properties of dECM-derived materials. In addition to mammalian tissues, the most recent bioprinting approaches involve the use of microbial extracellular polymeric substances in bioprinting of bacteria. This presents similar challenges as those encountered in mammalian cell printing and represents a fascinating frontier in bioprinting technology.

Development and systematic evaluation of decellularization protocols in different application models for diaphragmatic tissue engineering.

BACKGROUND: Tissue engineered bioscaffolds based on decellularized composites have gained increasing interest for treatment of various diaphragmatic impairments, including muscular atrophies and diaphragmatic hernias. Detergent-enzymatic treatment (DET) constitutes a standard strategy for diaphragmatic decellularization. However, there is scarce data on comparing DET protocols with different substances in distinct application models in their ability to maximize cellular removal while minimizing extracellular matrix (ECM) damage. METHODS: We decellularized diaphragms of male Sprague Dawley rats with 1 % or 0.1 % sodium dodecyl sulfate (SDS) and 4 % sodium deoxycholate (SDC) by orbital shaking (OS) or retrograde perfusion (RP) through the vena cava. We evaluated decellularized diaphragmatic samples by (1) quantitative analysis including DNA quantification and biomechanical testing, (2) qualitative and semiquantitative analysis by proteomics, as well as (3) qualitative assessment with macroscopic and microscopic evaluation by histological staining, immunohistochemistry and scanning electron microscopy. RESULTS: All protocols produced decellularized matrices with micro- and ultramorphologically intact architecture and adequate biomechanical performance with gradual differences. The proteomic profile of decellularized matrices contained a broad range of primal core and ECM-associated proteins similar to native muscle. While no outstanding preference for one singular protocol was determinable, SDS-treated samples showed slightly beneficial properties in comparison to SDC-processed counterparts. Both application modalities proved suitable for DET. CONCLUSION: DET with SDS or SDC via orbital shaking or retrograde perfusion constitute suitable methods to produce adequately decellularized matrices with characteristically preserved proteomic composition. Exposing compositional and functional specifics of variously treated grafts may enable establishing an ideal processing strategy to sustain valuable tissue characteristics and optimize consecutive recellularization. This aims to design an optimal bioscaffold for future transplantation in quantitative and qualitative diaphragmatic defects.

Effect of Preoperative Chemotherapy on the Isolation Outcome of Primary Human Hepatocytes.

Primary human hepatocytes isolated from surgically resected liver tissue are an essential resource for pharmaceutical and toxicological studies. Patients undergoing partial liver resections have often received preoperative chemotherapy. The aim of our study was to investigate whether preoperative chemotherapy has effects on the outcome of cell isolation or the metabolic function of cultured hepatocytes. Liver specimens from 48 patients were used for hepatocyte isolation. Out of these, 21 patients had prior chemotherapy, with fluoropyrimidine-based regimen in 14 patients. Viability and cell yield as parameter for the outcome of isolation, as well as transaminase levels, urea or albumin secretion to the culture medium were not different between hepatocytes from pretreated and untreated donor. Furthermore, the transcription levels of cytochrome P450 (CYP) 1A2, CYP 2B6, and CYP 3A4 of cultured hepatocytes were not affected by prior chemotherapy of the tissue donors. In conclusion, hepatocytes from tissue donors that underwent fluoropyrimidine-based chemotherapy regimens before isolation seem to perform as well as hepatocytes without preoperative chemotherapy exposure. Our results suggest that hepatocytes from patients who received combination chemotherapy before liver resection are an uncompromised resource for pharmacological and toxicological studies. Impact statement Isolated primary human hepatocytes are an essential resource for pharmacological and toxicological studies. Our results present further evidence that isolated hepatocytes from patients who received combination chemotherapy before liver resection are an uncompromised resource for pharmacological and toxicological studies-especially when fluoropyrimidine-based regimens are used.

The influence of the COVID-19 pandemic on surgical therapy and care: a cross-sectional study.

BACKGROUND: Due to the COVID-19 pandemic, an extensive reorganisation of healthcare resources was necessary-with a particular impact on surgical care across all disciplines. However, the direct and indirect consequences of this redistribution of resources on surgical therapy and care are largely unknown. METHODS: We analysed our prospectively collected standardised digital quality management document for all surgical cases in 2020 and compared them to the years 2018 and 2019. Periods with high COVID-19 burdens were compared with the reference periods in 2018 and 2019. RESULTS: From 2018 to 2020, 10,723 patients underwent surgical treatment at our centres. We observed a decrease in treated patients and a change in the overall patient health status. Patient age and length of hospital stay increased during the COVID-19 pandemic (p = 0.004 and p = 0.002). Furthermore, the distribution of indications for surgical treatment changed in favour of oncological cases and less elective cases such as hernia repairs (p < 0.001). Postoperative thromboembolic and pulmonary complications increased slightly during the COVID-19 pandemic. There were slight differences for postoperative overall complications according to Clavien-Dindo, with a significant increase of postoperative mortality (p = 0.01). CONCLUSION: During the COVID-19 pandemic we did not see an increase in the occurrence, or the severity of postoperative complications. Despite a slightly higher rate of mortality and specific complications being more prevalent, the biggest change was in indication for surgery, resulting in a higher proportion of older and sicker patients with corresponding comorbidities. Further research is warranted to analyse how this changed demographic will influence long-term patient care.

Robot-assisted pancreatic surgery-optimized operating procedures: set-up, port placement, surgical steps.

Even in most complex surgical settings, recent advances in minimal-invasive technologies have made the application of robotic-assisted devices more viable. Due to ever increasing experience and expertise, many large international centers now offer robotic-assisted pancreatic surgery as a preferred alternative. In general however, pancreatic operations are still associated with high morbidity and mortality, while robotic-assisted techniques still require significant learning curves. As a prospective post-marketing trial, we have established optimized operating procedures at our clinic. This manuscript intends to publicize our standardized methodology, including pre-operative preparation, surgical set-up as well as the surgeons' step-by-step actions when using pancreatic-assisted robotic surgery. This manuscript is based on our institutional experience as a high-volume pancreas operating center. We introduce novel concepts that should standardize, facilitate and economize the surgical steps in all types of robotic-assisted pancreatic surgery. The "One Fits All" principle enables single port placement irrespective of the pancreatic procedure, while the "Reversed 6-to-6 Approach" offers an optimized manual for pancreatic surgeons using the robotic console. Novel and standardized surgical concepts could guide new centers to establish a robust, efficient and safe robotic-assisted pancreatic surgery program.

Robotic-assisted pancreatic surgery in the elderly patient: experiences from a high-volume centre.

BACKGROUND: Robotic-assisted pancreatic surgery (RPS) has fundamentally developed over the past few years. For subgroups, e.g. elderly patients, applicability and safety of RPS still needs to be defined. Given prognosticated demographic developments, we aim to assess the role of RPS based on preoperative, operative and postoperative parameters. METHODS: We included 129 patients undergoing RPS at our institution between 2017 and 2020. Eleven patients required conversion to open surgery and were excluded from further analysis. We divided patients into two groups; ≥ 70 years old (Group 1; n = 32) and < 70 years old (Group 2; n = 86) at time of resection. RESULTS: Most preoperative characteristics were similar in both groups. However, number of patients with previous abdominal surgery was significantly higher in patients ≥ 70 years old (78% vs 37%, p < 0.0001). Operative characteristics did not significantly differ between both groups. Although patients ≥ 70 years old stayed significantly longer at ICU (1.8 vs 0.9 days; p = 0.037), length of hospital stay and postoperative morbidity were equivalent between the groups. CONCLUSION: RPS is safe and feasible in elderly patients and shows non-inferiority when compared with younger patients. However, prospectively collected data is needed to define the role of RPS in elderly patients accurately. Trial registration Clinical Trial Register: Deutschen Register Klinischer Studien (DRKS; German Clinical Trials Register). Clinical Registration Number: DRKS00017229 (retrospectively registered, Date of Registration: 2019/07/19, Date of First Enrollment: 2017/10/18).

Intrahepatic De Novo Tumors in Liver Recipients are Highly Associated With Recurrent Viral Hepatitis.

BACKGROUND/AIMS: Long-term survival of liver transplant recipients is endangered by tumorigenesis at different sites. Little is known about primary de novo tumors developing in the graft. METHODS: We analyzed the follow-up data of 2731 liver recipients that were transplanted between 1988 and 2019 at our institution (Charité - Universitätsmedizin Berlin, Department of Surgery). All cases with new intrahepatic tumors during follow-up were identified. RESULTS: A total of nine patients were diagnosed at a median of 16 years (range, 2-24 years) after surgery. Eight patients presented with hepatocellular carcinoma (HCC), and one patient presented with epithelioid hemangioendothelioma (EHE). All eight HCC patients had a recurrence of the initial disease that had caused liver failure before transplantation. This was associated with viral reinfection with either HCV or HBV in seven cases. Of the nine patients, three underwent surgical resection and only one patient was alive at data abstraction. CONCLUSION: Intrahepatic de novo neoplasms in the liver graft need to be considered in the long-term follow-up of liver recipients and were strongly associated with recurrent viral hepatitis in our study. Although prognosis of this rare complication is generally poor, patients may benefit from surgical resection of localized disease.

In vitro recellularization of decellularized bovine carotid arteries using human endothelial colony forming cells.

BACKGROUND: Many patients suffering from peripheral arterial disease (PAD) are dependent on bypass surgery. However, in some patients no suitable replacements (i.e. autologous or prosthetic bypass grafts) are available. Advances have been made to develop autologous tissue engineered vascular grafts (TEVG) using endothelial colony forming cells (ECFC) obtained by peripheral blood draw in large animal trials. Clinical translation of this technique, however, still requires additional data for usability of isolated ECFC from high cardiovascular risk patients. Bovine carotid arteries (BCA) were decellularized using a combined SDS (sodium dodecyl sulfate) -free mechanical-osmotic-enzymatic-detergent approach to show the feasibility of xenogenous vessel decellularization. Decellularized BCA chips were seeded with human ECFC, isolated from a high cardiovascular risk patient group, suffering from diabetes, hypertension and/or chronic renal failure. ECFC were cultured alone or in coculture with rat or human mesenchymal stromal cells (rMSC/hMSC). Decellularized BCA chips were evaluated for biochemical, histological and mechanical properties. Successful isolation of ECFC and recellularization capabilities were analyzed by histology. RESULTS: Decellularized BCA showed retained extracellular matrix (ECM) composition and mechanical properties upon cell removal. Isolation of ECFC from the intended target group was successfully performed (80% isolation efficiency). Isolated cells showed a typical ECFC-phenotype. Upon recellularization, co-seeding of patient-isolated ECFC with rMSC/hMSC and further incubation was successful for 14 (n = 9) and 23 (n = 5) days. Reendothelialization (rMSC) and partial reendothelialization (hMSC) was achieved. Seeded cells were CD31 and vWF positive, however, human cells were detectable for up to 14 days in xenogenic cell-culture only. Seeding of ECFC without rMSC was not successful. CONCLUSION: Using our refined decellularization process we generated easily obtainable TEVG with retained ECM- and mechanical quality, serving as a platform to develop small-diameter (< 6 mm) TEVG. ECFC isolation from the cardiovascular risk target group is possible and sufficient. Survival of diabetic ECFC appears to be highly dependent on perivascular support by rMSC/hMSC under static conditions. ECFC survival was limited to 14 days post seeding.

Engineering an endothelialized, endocrine Neo-Pancreas: Evaluation of islet functionality in an ex vivo model.

Islet-based recellularization of decellularized, repurposed rat livers may form a transplantable Neo-Pancreas. The aim of this study is the establishment of the necessary protocols, the evaluation of the organ structure and the analysis of the islet functionality ex vivo. After perfusion-based decellularization of rat livers, matrices were repopulated with endothelial cells and mesenchymal stromal cells, incubated for 8 days in a perfusion chamber, and finally repopulated on day 9 with intact rodent islets. Integrity and quality of re-endothelialization was assessed by histology and FITC-dextran perfusion assay. Functionality of the islets of Langerhans was determined on day 10 and day 12 via glucose stimulated insulin secretion. Blood gas analysis variables confirmed the stability of the perfusion cultivation. Histological staining showed that cells formed a monolayer inside the intact vascular structure. These findings were confirmed by electron microscopy. Islets infused via the bile duct could histologically be found in the parenchymal space. Adequate insulin secretion after glucose stimulation after 1-day and 3-day cultivation verified islet viability and functionality after the repopulation process. We provide the first proof-of-concept for the functionality of islets of Langerhans engrafted in a decellularized rat liver. Furthermore, a re-endothelialization step was implemented to provide implantability. This technique can serve as a bioengineered platform to generate implantable and functional endocrine Neo-Pancreases.

The human liver matrisome - Proteomic analysis of native and fibrotic human liver extracellular matrices for organ engineering approaches.

The production of biomaterials that endow significant morphogenic and microenvironmental cues for the constitution of cell integration and regeneration remains a key challenge in the successful implementation of functional organ replacements. Despite the vast development in the production of biological and architecturally native matrices, the complex compositions and pivotal figures by which the human matrisome mediates many of its essential functions are yet to be defined. Here we present a thorough analysis of the native human liver proteomic landscape using decellularization and defatting protocols to create extracellular matrix scaffolds of natural origin that can further be used in both bottom-up and top-down approaches in tissue engineering based organ replacements. Furthermore, by analyzing human liver extracellular matrices in different stages of fibrosis and cirrhosis, we have identified distinct attributes of these tissues that could potentially be exploited therapeutically and thus require further investigation. The general experimental pipeline presented in this study is applicable to any type of tissue and can be widely used for different approaches in regenerative medicine and in the construction of novel biomaterials for organ engineering approaches.

Magnetic resonance elastography quantification of the solid-to-fluid transition of liver tissue due to decellularization.

Maintenance of tissue extracellular matrix (ECM) and its biomechanical properties for tissue engineering is one of the substantial challenges in the field of decellularization and recellularization. Preservation of the organ-specific biomatrix is crucial for successful recellularization to support cell survival, proliferation, and functionality. However, understanding ECM properties with and without its inhabiting cells as well as the transition between the two states lacks appropriate test methods capable of quantifying bulk viscoelastic parameters in soft tissues. We used compact magnetic resonance elastography (MRE) with 400, 500, and 600 Hz driving frequency to investigate rat liver specimens for quantification of viscoelastic property changes resulting from decellularization. Tissue structures in native and decellularized livers were characterized by collagen and elastin quantification, histological analysis, and scanning electron microscopy. Decellularization did not affect the integrity of microanatomy and structural composition of liver ECM but was found to be associated with increases in the relative amounts of collagen by 83-fold (37.4 ± 17.5 vs. 0.5 ± 0.01 µg/mg, p = 0.0002) and elastin by approx. 3-fold (404.1 ± 139.6 vs. 151.0 ± 132.3 µg/mg, p = 0.0046). Decellularization reduced storage modulus by approx. 9-fold (from 4.9 ± 0.8 kPa to 0.5 ± 0.5 kPa, p < 0.0001) and loss modulus by approx. 7-fold (3.6 kPa to 0.5 kPa, p < 0.0001), indicating a marked loss of global tissue rigidity as well as a property shift from solid towards more fluid tissue behavior (p = 0.0097). Our results suggest that the rigidity of liver tissue is largely determined by cellular components, which are replaced by fluid-filled spaces when cells are removed. This leads to an overall increase in tissue fluidity and a viscous drag within the relatively sparse remaining ECM. Compact MRE is an excellent tool for quantifying the mechanical properties of decellularized biological tissue and a promising candidate for useful applications in tissue engineering.