Programm–2017

Recellularization of rat livers: morphology and function

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The Journal of Tissue Engineering and Regenerative Medicine accepted our paper Evolution of graft morphology and function after recellularization of decellularized rat liversfor publication.

Decellularization of livers is a well-established procedure. Data on different reseeding techniques or the functional evolution and re-organization processes of repopulated grafts remains limited.
We established a proprietary, customized bioreactor to repopulate decellularized rat livers (n=21) with primary rat hepatocytes (150 x 106 cells) via the hepatic artery and to subsequently evaluate graft morphology and function during seven days of ex vivo perfusion. Grafts were analyzed at 1h, 6h, 12h, 24h, 3d, 5d and 7d after recellularization (all n=3) by immunohistologic evaluation, hepatocyte-related enzyme (AST, ALT, LDH) and albumin measurement in the perfusate.
To the best of our knowledge, this is the first available protocol for repopulation of rat livers via the hepatic artery. Within the first 24 hours after repopulation, the hepatocytes seemed to migrate out of the vascular network and form clusters in the parenchymal space around the vessels. Graft function increased for the first 24 hours after repopulation with a significantly higher function compared to standard 2D culture after 24 hours. Thereafter, graft function constantly decreased with significantly lower values after six and seven days of perfusion, although histologically viable hepatocytes were found even after this period. Our data suggests that due to a constant loss of function, repopulated grafts should potentially be implanted as soon as cell engraftment and graft re-organization are completed.

Authors are Antje Butter, Khalid Aliyev, Karl-Herbert Hillebrandt, Nathanael Raschzok, Martin Kluge, Nicolai Seiffert, Peter Tang, Hendrik Napierala, Muhammad Imtiaz Ashraf, Anja Reutzel-Selke, Andreas Andreou, Johann Pratschke, Igor Maximilian Sauer, and Benjamin Struecker.

Neujahrsessen 2017

Einladung Neujahrsessen 2017

Visit of Jean-Philippe Courtois and Renate Radon

Jean-Philippe Courtois (Executive Vice President and President, Microsoft Global Sales, Marketing and Operations), Renate Radon (Senior Director Public Sector, Microsoft Deutschland) and other high-ranking representatives of Microsoft visited us in order to see the HoloLens in use for preoperative planning and during surgery. We had a very fruitful discussion on Mixed Reality and Virtual Reality in medicine and are looking forward to a strong collaboration!

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Vascular anatomy of the juvenile Göttingen minipig

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Lab Animal accepted our „Computed tomography-based survey of the vascular anatomy of the juvenile Göttingen minipig“ for publication.
Over the past 50 years, image-guided procedures have been established for a wide range of applications. The development and clinical translation of new treatment regimens necessitate the availability of suitable animal models. The juvenile Göttingen minipig presents a favourable profile as a model for human infants. However, no information can be found regarding the vascular system of juvenile minipigs in the literature. Such information is imperative for planning the accessibility of target structures by catheterization. We present here a complete mapping of the arterial system of the juvenile minipig based on contrast-enhanced computed tomography. Four female animals weighing 6.13 ± 0.72 kg were used for the analyses. Imaging was performed under anaesthesia, and the measurement of the vascular structures was performed independently by four investigators. Our dataset forms a basis for future interventional studies in juvenile minipigs, and enables planning and refinement of future experiments according to the 3R (replacement, reduction and refinement) principles of animal research.

Authors are J. Siefert, K.H. Hillebrandt, M. Kluge, D. Geisel, P. Podrabsky, T. Denecke, M. Nösser, J. Gassner, A. Reutzel-Selke, B. Strücker, M.H. Morgul, S. Guel-Klein, J.K. Unger, A. Reske, J. Pratschke, I.M. Sauer, and N. Raschzok.
Update: Now available via Lab Anim. 2016 Dec 8. pii: 0023677216680238. [Epub ahead of print]

Monitoring of hepatocyte transplantation by MRI

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A new book on Hepatocyte Transplantation Methods and Protocols, part of the series: Methods in Molecular Biology, Vol. 1506 P. Stock, B. Christ (Eds.), Springer, will be available end of November, 2016. We contributed a chapter on Preclinical swine models for monitoring of hepatocyte transplantation by MRI (authors: Nathanael Raschzok, Ulf Teichgräber, Johann Pratschke, and Igor M. Sauer) and are proud to provide the cover image.
This volume features up-to-date protocols for the isolation, preservation, and validation of various cell sources comprising large and small animal models, examining the impact of cell transplantation on acute and chronic liver diseases. Hepatocyte Transplantation: Methods and Protocols guides readers through laboratory protocols for the generation of humanized livers for the assessment of biological actions in vivo and techniques to monitor cell engraftment after cell transplantation in vivo are described and procedures for computational analyses of hepatocyte transplantation. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Comprehensive and practical, Hepatocyte Transplantation: Methods and Protocols is an essential resource for researchers and clinicians to assess the biological as well as the therapeutic potential of hepatocyte transplantation.

Staatsexamen

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More than 100 days of loops of learning and 3 days conquering more than 300 questions: 5 faces.
Congratulations!
We are all looking forward to having you back in the lab!

HCC: miRNA profiles of the tumor-surrounding tissue

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The presence of hepatocellular carcinoma (HCC) is a significant complication of cirrhosis because it changes the prognosis and the treatment of the patients. By now, contrast-enhanced CT and MR scans are the most reliable tools for the diagnosis of HCC; however, in some cases, a biopsy of the tumor is necessary for the final diagnosis. The aim of the study was to develop a diagnostic tool using the microRNA (miRNA) profiles of the tissue surrounding the HCC tumor combined with clinical parameters in statistical models. At a transplantation setting, 32 patients with HCC and cirrhosis (B) were compared to 22 patients suffering from cirrhosis only (A). The diagnosis and exclusion of HCC was confirmed following the histopathological examination of the explanted liver. The HCC patients were significantly older than the patients with cirrhosis only (B: 60.6 and A: 49.9, p<0.001) and showed higher levels of ALT (A: 0.76μkat/l, B: 1.02μkat/, p=0.006) and AFP (A: 5.8ng/ml, B: 70.3ng/ml, p<0.001), whereas the bilirubin levels were higher in the cirrhosis only group (p=0.002). Using age (cut-off 50.23years) and AFP (cut-off 4.2ng/ml) thresholds, the levels of expression of miR-1285-3p and miR-943 differentiated between the patients with HCC and cirrhosis from those with cirrhosis only with an accuracy of 96.3%. This is the first report about the use of stepwise penalized logistic regression and decision tree analyses of miRNA expressions in the tumor-surrounding tissue combined with clinical parameters for the diagnosis of HCC.

Authors are Mehmet Haluk Morgul, Sergej Klunk, Zografia Anastasiadou, Ulrich Gauger, Corinna Dietel, Anja Reutzel-Selke, Philipp Felgendref, Hans-Michael Hau, Hans-Michael Tautenhahn, Rosa Bianca Schmuck, Nathanael Raschzok, Igor Maximillian Sauer, and Michael Bartels.
Exp Mol Pathol. 2016 Aug 20;101(2):165-171. doi: 10.1016/j.yexmp.2016.07.014. [Epub ahead of print]

Bile: miRNA Pattern post OLT

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BIOMARKERS accepted our latest paper on "Bile: miRNA Pattern and Protein Based Biomarkers May Predict Acute Cellular Rejection after Liver Transplantation" for publication. Authors are Rosa Bianca Schmuck, Anja Reutzel-Selke, Nathanael Raschzok, Mehmet Haluk Morgul, Benjamin Struecker, Steffen Lippert, Cynthia de Carvalho Fischer, Moritz Schmelzle, Sabine Boas-Knoop, Marcus Bahra, Andreas Pascher, Johann Pratschke, and Igor M. Sauer.

Bile rather than blood depicts the local inflammation in the liver and may improve prediction and diagnosis of acute cellular rejection (ACR) after liver transplantation (OLT). Secretome and miRNAs were analyzed during the first two weeks and on clinical suspicion of ACR in the bile of 45 OLT recipients. Levels of CD44, CXCL9, miR-122, miR-133a, miR-148a and miR-194 were significantly higher in bile of patients who developed ACR within the first 6 months after OLT and during ACR. Analysis of secretome and miRNA in bile could further our understanding of the local inflammatory process during rejection.

Biomarkers. 2016 Aug 5:1-9. [Epub ahead of print]

Hepatocyte isolation after laparoscopic liver resection

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Tissue Engineering, Part C: Methods accepted our paper entitled "Hepatocyte isolation after laparoscopic liver resection" for publication. Authors are Horner R*, Kluge M*, Gassner J, Nösser M, Major RD, Reutzel-Selke A, Leder AK, Struecker B, Morgul MH, Pratschke J, Sauer IM, Raschzok N (*contributed equally).

Liver tissue obtained from partial hepatectomy is a common source for isolation of primary human hepatocytes. Until now, liver resections were most commonly performed by conventional open surgery. Although the laparoscopic approach is currently emerging in liver surgery, data on the outcome of hepatocyte isolation from laparoscopically resected liver tissue is not available.
A total of 22 hepatocyte isolations were performed using the two-step collagenase perfusion technique from October 2015 until March 2016. Liver tissue was obtained from n=15 open liver resections (OLR) and n=7 laparoscopic liver resections (LLR). Isolation parameters (cell yield, viability, percoll survival) were assessed and hepatocyte function (plating efficiency, urea, albumin, and aspartate aminotransferase) was measured over a culture period of 6 days (OLR: n=13; LLR: n=3).
Total cell yield (OLR: 36.81 ± 6.77 x106 cells/g vs. LLR 16.84 ± 10.66 x106 cells/g, p=0.0318) as well as viable yield (OLR 31.70 ± 6.05 x106 cells/g vs. LLL 14.70 ± 9.89 x106 cells/g, p=0.0260) were significantly higher in the OLR group. Subgroup analysis revealed that the worse outcome of isolation of laparoscopically resected liver tissue was associated with right-lateral laparoscopic liver resections, while hepatocyte isolation from left-lateral laparoscopic liver resections was as effective as from open surgery. Hepatocyte function did not differ between hepatocytes from openly resected versus left-lateral laparoscopically resected liver tissue.
We here present the first data on hepatocyte isolation from laparoscopic liver surgery. While the overall outcome is worse compared to open surgery, our data suggest that liver tissue from laparoscopic resection of the left lobe is an excellent source for primary human hepatocytes.

Five years BIH Charité Clinician Scientist Program

BIH Charité Clinician Scientist Symposium

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Dr. rer. medic. Dipl.-Ing. Annekatrin Leder

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Today, Anne Leder successfully defended her thesis "summa cum laude"!

Her work entitled "Entwicklung und Evaluierung eines mikroskaligen, Oligonukleotid-gekoppelten Eisenoxidpartikels zur Stimulation kultivierter humaner Hepatozyten" deals with particle-based delivery systems for therapeutic manipulation and tracking of transplanted cells by magnetic resonance imaging (MRI) based on multifunctional, silica based micron-sized iron oxide-containing particles (sMPIO) that combine fluorescence imaging, MRI tracking, and on-the-spot targeting of specific microRNAs on a particle surface for therapeutic manipulation by RNA interference.

CONGRATULATIONS !

SPARK Berlin supports Fikatas Knot

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The Berlin Institute of Health (BIH) and Stiftung Charité teamed up with Stanford University School of Medicine to initiate SPARK Berlin.

We are very pleased to announce that Dr. Panagiotis Fikatas' project “Device for ready-prepared surgical knots” was selected for both, funding and mentorship. 

SPARK was created to overcome the hurdles associated with translating academic discoveries into therapeutics and diagnostics that address unmet medical needs.
The SPARK mission is to help academics overcome the obstacles involved in moving their early discoveries from bench to bedside, to educate faculty, postdoctoral fellows and graduate students on the translational research process and path to clinical application, so that development of promising discoveries becomes second nature to our institution, and to develop more cost-effective and innovative approaches to drug development .

Congratulations!

Ethicon Zukunfts-Award 2016 | Dr. Fikatas

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During the 133. Meeting of the Deutsche Gesellschaft für Chirurgie Dr. Panagiotis Fikatas was awarded with the Ethicon Zukunfts-Award 2016 (2nd place) – Congratulations!

International Medical Tournament 2016

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Karl Hillebrandt and his team won this year's International Medical Tournament (English League) in Novosibirsk, Russia!

Congratulations!


The International Medical Tournament is a team competition for future doctors, which provides unique professional experience such as solving complex diagnostic cases from real clinical practice without any risks, presentation and discussion of solutions among students and experts.
The main goal of the competition is to help medical students gather and develop lacking skills for future medical practice, critical thinking in diagnostic process, abilities of teamwork, presentation and rhetorical skills. 

NeoHybrid liver graft – proof of concept

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Cells Tissues Organs accepted our latest paper on "Allogeneic liver transplantation and subsequent syngeneic hepatocyte transplantation in a rat model – proof of concept for in vivo tissue engineering" for publication.
Authors are Susanne Rohn, Jan Schroeder, Henriette Riedel, Dietrich Polenz, Katarina Stanko, Anja Reutzel-Selke, Peter Tang, Lydia Brusendorf, Nathanael Raschzok, Peter Neuhaus, Johann Pratschke, Birgit Sawitzki, Igor M. Sauer, and Martina T. Mogl.

Aim of the project was the evaluation of a new concept for in vivo tissue engineering using autologous primary human hepatocytes and hepatic progenitor cells isolated from diseased livers explanted during orthotopic liver transplantation (LTx). Cells will be isolated and infused into the spleen for repopulation of the allogeneic liver graft. The latter is serving as biological matrix for the engraftment of autologous cells. Once these cells have engrafted, it is assumed that autologous cells will repopulate the allogeneic liver, since they should have a selective advantage due to their autologous origin. It is postulated that this will lead to a neo-hybrid liver graft, reducing immunogenicity and inducing immunoregulation thus minimizing the need for extensive immunosuppression and eventually inducing operational tolerance.
We therefore developed a new rat model for combined liver and liver cell transplantation under stable immunosuppression. Immunohistochemistry demonstrated the engraftment of transplanted cells, as confirmed by fluorescence in-situ hybridization, showing repopulation of the liver graft with 15.6 % male cells (± 1.8 SEM) at day 90. The quantitative PCR revealed 14.15 % (mean ± 5.09 SEM) male DNA at day 90. Engraftment of transplanted autologous cells after combined liver and cell transplantation was achieved for up to 90 days under immunosuppression. Immunohistochemistry indicated cell proliferation, and the fluorescence in-situ hybridization results were partly confirmed by quantitative RT-PCR. This new protocol in rats appears feasible to address long-term function and eventually induction of operational tolerance in the future.

LTx – microRNA signatures in peripheral blood ?

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BIOMARKERS accepted our latest paper on „microRNA signatures in peripheral blood fail to detect acute cellular rejection after liver transplantation“ for publication. Authors are N. Raschzok, A. Reutzel-Selke, R. Schmuck, L. Tannus, M. Morgul, C. Dietel, A. Leder, B. Struecker, S. Lippert, H. Sallmon, M. Schmelzle, M. Bartels, S. Jonas, J. Pratschke, and I.M. Sauer.

We investigated whether microRNA signatures in whole blood samples are associated with acute cellular rejection (ACR) after liver transplantation. Blood samples were collected using Paxgene technology and analyzed by microarrays and qRT-PCR.
microRNA signatures failed to distinguish between 19 patients with ACR and 16 controls. Let-7b-5p and let-7c were up-regulated in a subgroup of patients with ACR during the 6th and 7th postoperative day but failed in an independent validation of 20 patients.
microRNA signatures in whole blood processed by Paxgene technology are not suited for detection of ACR after liver transplantation.

Single Pass Albumin Dialysis – Dose finding study

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Artificial Organs accepted our paper „Single Pass Albumin Dialysis (SPAD) – A dose finding study to define optimal albumin concentration and dialysate flowfor publication.
Authors are R.B. Schmuck, G.-H. Nawrot, P. Fikatas, A. Reutzel-Selke, J. Pratschke, and I.M. Sauer.

Aim of these studies was to define the optimal conditions for SPAD in a standardized experimental set-up. Albumin concentration was adjusted to either 1%, 2%, 3%, or 4%, while the flow rate of the dialysate was kept constant at a speed of 700 ml/h. The flow rate of the dialysate was altered between 350, 500, 700, and 1000 ml/h, whereas the albumin concentration was continuously kept at 3%.
This study revealed that the detoxification of albumin bound substances could be improved by increasing the concentration of albumin in the dialysate with an optimum at 3%. A further increase of the albumin concentration to 4% did not lead to a significant increase in detoxification. Furthermore, we observed a gradual increase of the detoxification efficiency for albumin bound substances, from 350 ml/h to 700 ml/h (for bilirubin) or 1000 ml/h (for bile acids) of dialysate flow. Water-soluble toxins (ammonia, creatinine, urea, uric acid) were removed almost completely, regardless of albumin concentration or flow rate.

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Although we do hate Facebook, feel kind of robbed and dislike the music –
We have to admit: 7,201,027 clicks (as of January 25th, 2016) are impressive…

Feature in Berliner Zeitung on Tissue Engineering

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Recellularized liver created in our lab featured in Berliner Zeitung.

ESOT | YPT – Interview with Karl Hillebrandt

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Being the winner of the Rising Stars Video Session organised by the YPT Committee at the ESOT2015 Brussels Congress Karl Hillebrandt gave an interview for the ESOT | YPT webpage.
His abstract "Optimized decellularization of rat livers byarterial and portal venous perfusion underoscillating pressure conditions" and the accompanying video were the most voted at the Rising Stars Session, where the audience voted live for the best video abstract. You can read Karl's abstract in the special ESOT2015 issue of Transplant International.