Poly (2-hydroxyethyl methacrylate) (HEMA) has been used as a clinical material, in the form of a soft hydrogel, for various surgical procedures, including endovascular surgery of liver. It is a clear liquid compound and, as a soft, flexible, water-absorbing material, has been used to make soft contact lenses from small, concave, spinning molds. Primary rat hepatocyte spheroids were created on a poly-HEMA-coated surface with the intention of inducing hepatic tissue formation and improving liver functions. We investigated spheroid formation of primary adult rat hepatocyte cells and characterized hepatic-specific functions under the special influence of fetal calf serum (FCS) and nonparencymal cells (NPC) up to six days in different culture systems (e.g., hepatocytes + FCS, hepatocytes – FCS, NPC + FCS, NPC – FCS, co-culture + FCS, co-culture – FCS) in both the spheroid model and sandwich model. Immunohistologically, we detected gap junctions, Ito cell/Kupffer cells, sinusoidal endothelial cells and an extracellular matrix in the spheroid model. FCS has no positive effect in the sandwich model, but has a negative effect in the spheroid model on albumin production, and no influence in urea production in either model. We found more cell viability in smaller diameter spheroids than larger ones by using the apoptosis test. Furthermore, there is no positive influence of the serum or NPC on spheroid formation, suggesting that it may only depend on the physical condition of the culture system. Since the sandwich culture has been considered a “gold standard” in vitro culture model, the hepatocyte spheroids generated on the poly-HEMA-coated surface were compared with those in the sandwich model. Major liver-specific functions, such as albumin secretion and urea synthesis, were evaluated in both the spheroid and sandwich model. The synthesis performance in the spheroid compared to the sandwich culture increases approximately by a factor of 1.5. Disintegration of plasma membranes in both models was measured by lactate dehydrogenase (LDH) release in both models. Additionally, diazepam was used as a substrate in drug metabolism studies to characterize the differences in the biotransformation potential with metabolite profiles in both models. It showed that the diazepam metabolism activities in the spheroid model is about 10-fold lower than the sandwich model. The poly-HEMA-based hepatocyte spheroid is a promising new platform towards hepatic tissue engineering leading to in vitro hepatic tissue formation.

Till date, no bioartificial liver (BAL) procedure has obtained FDA approval or widespread clinical acceptance, mainly because of multifactorial limitations such as the use of microscale or undefined biomaterials, indirect and lower oxygenation levels in liver cells, short-term undesirable functions, and a lack of 3D interaction of growth factor/cytokine signaling in liver cells. To overcome preclinical limitations, primary rat liver cells were cultured on a naturally self-assembling peptide nanoscaffold (SAPN) in a clinically relevant bioreactor for up to 35 days, under 3D interaction with suitable growth factors and cytokine signaling agents, alone or combination (e.g., Group I: EPO, Group II: Activin A, Group III: IL-6, Group IV: BMP-4, Group V: BMP4 + EPO, Group VI: EPO + IL-6, Group VII: BMP4 + IL-6, Group VIII: Activin A + EPO, Group IX: IL-6 + Activin A, Group X: Activin A + BMP4, Group XI: EPO + Activin A + BMP-4 + IL-6 + HGF, and Group XII: Control). Major liver specific functions such as albumin secretion, urea metabolism, ammonia detoxification, phase contrast microscopy, immunofluorescence of liver specific markers (Albumin and CYP3A1), mitochondrial status, glutamic oxaloacetic transaminase (GOT) activity, glutamic pyruvic transaminase (GPT) activity, and cell membrane stability by the lactate dehydrogenase (LDH) test were also examined and compared with the control over time. In addition, we examined the drug biotransformation potential of a diazepam drug in a two-compartment model (cell matrix phase and supernatant), which is clinically important. This present study demonstrates an optimized 3D signaling/scaffolding in a preclinical BAL model, as well as preclinical drug screening for better drug development.

Application of a nephrotoxicity- and atherogenicity-free, MMF-based i.s. induction/maintenance protocol in elderly recipient of kidneys from elderly cadaver donors leads to improved long-term outcomes which are comparable with data from young recipients who have received allografts from young cadaver donors.

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The metabolic risk for patients undergoing abdominal cancer resection increases in the perioperative period and malnutrition may be observed. In order to prevent further weight loss, the guidelines recommend for high-risk patients the placement of a needle catheter jejunostomy (NCJ) for supplementing enteral feeding in the early and late postoperative period. Our aim was to evaluate the safety of NCJ placement and its potential benefits regarding the nutritional status in the postoperative course. We retrospectively analyzed patients undergoing surgery for upper gastrointestinal cancer, such as esophageal, gastric, and pancreato-biliary cancer, and NCJ placement during the operation. The nutritional parameters body mass index (BMI), perioperative weight loss, phase angle measured by bioelectrical impedance analysis (BIA) and the clinical outcome were assessed perioperatively and during follow-up visits 1 to 3 months and 4 to 6 months after surgery. In 102 patients a NCJ was placed between January 2006 and December 2016. Follow-up visits 1 to 3 months and 4 to 6 months after surgery were performed in 90 patients and 88 patients, respectively. No severe complications were seen after the NCJ placement. The supplementing enteral nutrition via NCJ did not improve the nutritional status of the patients postoperatively. There was a significant postoperative decline of weight and phase angle, especially in the first to third month after surgery, which could be stabilized until 4–6 months after surgery. Placement of NCJ is safe. In patients with upper gastrointestinal and pancreato-biliary cancer, supplementing enteral nutrition during the postoperative course and continued after discharge may attenuate unavoidable weight loss and a reduction of body cell mass within the first six months.

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Much effort has been directed towards the optimization of the capture of in vivo hepatocytes from their microenvironment. Some methods of capture include an ex vivo cellular model in a bioreactor based liver module, a micropatterned module, a microfluidic 3D chip, coated plates, and other innovative approaches for the functional maintenance of primary hepatocytes. However, none of the above methods meet US Food and Drug Administration (FDA) guidelines, which recommend and encourage that the duration of a toxicity assay of a drug should be a minimum of 14 days, to a maximum of 90 days for a general toxicity assay. Existing innovative reports have used undefined extracellular matrices like matrigel, rigid collagen, or serum supplementations, which are often problematic, unacceptable in preclinical and clinical applications, and can even interfere with experimental outcomes. We have overcome these challenges by using integrated nanostructured self-assembling peptides and a special combination of growth factors and cytokines to establish a proof of concept to mimic the in vivo hepatocyte microenvironment pattern in vitro for predicting the in vivo drug hepatotoxicity in a scalable bioartificial liver module. Hepatocyte functionality (albumin, urea) was measured at days 10, 30, 60, and 90 and we observed stable albumin secretion and urea function throughout the culture period. In parallel, drug metabolizing enzyme biomarkers such as ethoxyresorufin-O-deethylase, the methylthiazol tetrazolium test, and the lactate dehydrogenase test were carried out at days 10, 30, 60, and 90. We noticed excellent mitochondrial status and membrane stability at 90 days of culture. Since alpha glutathione S-transferase (GST) is highly sensitive and a specific marker of hepatocyte injury, we observed significantly low alpha GST levels on all measured days (10, 30, 60, and 90). Finally, we performed the image analysis of mitochondria-cultured hepatocytes at day 90 in different biophysical parameters using confocal microscopy. We applied an automatic algorithm-based method for 3D visualization to show the classic representation of the mitochondrial distribution in double hepatocytes. An automated morphological measurement was conducted on the mitochondrial distribution in the cultured hepatocytes. Our proof of concept of a scalable bioartificial liver modular device meets FDA guidelines and may function as an alternative model of animal experimentation for pharmacological and toxicological studies involving drug metabolism, enzyme induction, transplantation, viral hepatitis, hepatocyte regeneration, and can also be used in other existing bioreactor modules for long-term culture for up to 90 days or more.