Clinical Hemorheology and Microcirculation - Volume 73, issue 1

Authors: Schaible, J. | Stroszczynski, C. | Beyer, L.P. | Jung, E.M.

Article Type: Research Article

Abstract: OBJECTIVE: The use of a standardized dynamic contrast enhanced ultrasound (CEUS) protocol to determine perfusion of hepatocellular carcinomas (HCC) using quantitative analysis. METHODS: Retrospective analysis of 27 patients with hepatocellular carcinoma examined by a standardized protocol (bolus injection of 2.4 ml sulphur hexafluoride microbubbles, arterial phase to portal venous phase using continuous CINE-loops over 1 minute, short CINE-loops after 2, 3, 4, 5 and 6 minutes and additional single images for B-mode, CCDS). The examination was performed by an experienced sonographer with a high resolution, multi-frequency transducer (1–6 MHz). The reading of the reference imaging (contrast enhanced CT and MRI) …and histopathology, if necessary, was performed independently. Retrospective analysis of the CEUS DICOM loops was done using time intensity curve analysis (TIC) with evaluation of the time to peak (TTP) and the area under the curve (AUC). RESULTS: All tumor lesions were characterized by CEUS, based on typical contrast patterns for HCC lesions with arterial enhancement and wash out in the late phase, corresponding to MRI with liver specific contrast agent or contrast enhanced CT. Mean TTP (SD) in the tumor centre (C) was 19.93 (11.31), in the periphery (P) 22.94 (9.44) and in the normal liver tissue (LT) 28.19 (11.34) with significant differences between all zones C/P (p  = 0.013), C/LT (p  = 0.005) and P/LT (p  = 0.022). AUC mean (SD) in the tumor centre (C) was 660.03 (292.64), in the periphery (P) 586.04 (237.01) and in the normal liver tissue (LT) 484.20 (236.99), also with significant differences between all zones C/P (p  = 0.001), C/LT (p  < 0.001) and P/LT (p  < 0.001). CONCLUSION: TIC-analysis is an easy-to-use tool for the dynamic evaluation of microvascularization in hepatocellular carcinoma and allows a fast and cost-efficient quantitative analysis. Show more

Keywords: CEUS, Contrast enhanced ultrasound, hepatocellular carcinoma, TIC-analysis, quantitative analysis

DOI: 10.3233/CH-199221

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 95-104, 2019

Authors: Chaloupka, M. | Bischoff, R. | Pfitzinger, P. | Lellig, E. | Ledderose, S. | Buchner, A. | Schlenker, B. | Stief, C. | Clevert, D.-A. | Apfelbeck, M.

Article Type: Research Article

Abstract: INTRODUCTION: Multiparametric-Magnetic Resonance Imaging (mpMRI)-Ultrasound fusion guided biopsy (Fbx) has emerged as the new standard of risk stratification for prostate cancer (PCa) with superior detection rates of clinically significant PCa than randomized biopsy. In the present study, we evaluated patients with suspicion of clinically significant PCa on mpMRI, but histopathologically proven Gleason 6 PCa in Fbx. MATERIAL AND METHODS: Between 2015 and 2019, 849 patients underwent Fbx and concurrent systematic 12-core biopsy at our department. 234 patients were diagnosed with Gleason 6 PCa in either mpMRI-targeted and/or concurrent systematic biopsy. Patients were analyzed regarding PSA, mpMRI findings according …to PI-RADS classification, histopathological results of Fbx and systematic 12-core biopsy. 99/234 patients were also analyzed in regards of histopathology of the whole-mount specimen of subsequent radical prostatectomy (RP). RESULTS: In 131/234 patients (56%), Gleason 6 PCa was detected in the mpMRI target. In 103/234 patients (44%), Gleason 6 PCa was detected in the concurrent systematic 12-core biopsy with negative mpMRI-targeted biopsy. Men with evidence of Gleason 6 in the mpMRI target had significantly higher amounts of overall positive biopsies (median 4 vs. 2, p  < 0.001) and higher maximum tumor infiltration per biopsy core (30% vs. 20%, p  < 0.001) compared to men with negative mpMRI-targeted biopsy. Detection of Gleason 6 in mpMRI Target lesions correlated significantly with the PI-RADS score (p  < 0.001). Patients with positive mpMRI-target had significantly higher tumor infiltration in whole-mount specimen after prostatectomy (20% vs. 15%, p  = 0.0026) compared to men without detection of Gleason 6 in mpMRI-targeted biopsy but in additional systematic biopsy. CONCLUSION: Detection of Gleason 6 PCa in mpMRI-targeted biopsy indicates higher tumor burden compared to detection of Gleason 6 PCa in concurrent systematic biopsy and negative mpMRI-targeted biopsy. Show more

Keywords: Low-risk prostate cancer, multiparametric MRI, fusion biopsy, active surveillance, PI-RADS classification

DOI: 10.3233/CH-199223

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 105-111, 2019

Authors: Schiltz, Daniel | Taeger, Christian D. | Biermann, Niklas | Ranieri, Marco | Klein, Silvan | Prantl, Lukas | Geis, Sebastian

Article Type: Research Article

Abstract: BACKGROUND: Reconstruction of soft tissue defects with free flaps is a common procedure in plastic and reconstructive surgery. Most postoperative complications occur within the first 48–72 hours after surgery. After postoperative complications, short perfusion restoration times may improve flap survival rates by up to 30–50%. Ratiometric fluorescence imaging is an additional or alternative method of postoperative flap monitoring. OBJECTIVE: To test the efficacy and utility of transepidermal oxygen flux imaging to evaluate postoperative skin oxygenation of free and local flaps in the first 48 hours after surgery. METHODS: The study included 32 patients (aged between 18 …and 80 years; mean age 52.9) with a tissue defect covered with a free flap transplant at the Department of Plastic and Reconstructive Surgery of the University Medical Center Regensburg. Postoperative oxygen flux was measured with the ‘VisiSens system’ placed on the vascular pedicle as well as on the peripheral and central part of the flap. RESULTS: Values of oxygen flux were higher in case of flap congestion (0.069±0.012) or flap necrosis (0.155±0.083) than in cases without any complications (0.061±0.006). Flux values of different areas of the same flap showed only minimal differences (central part: 0.065±0.008, peripheral part: 0.070±0.009, vascular pedicle: 0.056±0.004); the level of significance was p  = 0.904. CONCLUSION: Imaging transepidermal oxygen flux by ratiometric luminescence seems to be a reliable alternative, indirect method of postoperative flap monitoring with regard to microcirculatory function and flap viability. Show more

Keywords: Postoperative free flap monitoring, microcirculatory function, tissue oxygenation, tissue perfusion, flux imaging, flap monitoring, transcutaneous oxygen measurement

DOI: 10.3233/CH-199225

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 113-123, 2019

Authors: Brebant, V. | Heine, N. | Lamby, P. | Heidekrueger, PI | Forte, A.J. | Prantl, L. | Aung, T.

Article Type: Research Article

Abstract: BACKGROUND: A physiological and minimal invasive form of surgery with minimal risk to treat lymphedemas is the so-called supermicrosurgical lymphovenous anastomosis (LVA) where a lymph vessel is connected with a venule. METHODS: 30 patients (between 2018 and 2019) with secondary upper extremity lymphedema refractory to conservative therapy (manual lymph drainage and compression therapy were operated using the “simplified lymphovenous anastomosis” method). For the assessment of lymphatic supermicrosurgery, an operating microscope in which a near-infrared illumination system is integrated (Leica M530 OHX with glow technology ULT530, Leica Microsystems) and the IC-FlowTM Imaging System(Diagnostic Green)/Visionsense System (Medtronic) together with …a ZEISS S8 microscope was used. Augmented reality intraoperative indocyanin green (ICG) lymphography-navigated modified “simplified lymphovenous anastomosis” were performed on the Leica microscope. All patients were informed about Off-label-use of ICG lymphography. RESULTS: 57 LVAs were performed with modified “simplified lymphovenous anastomosis” lymphography-guidance on 30 upper extremities. All patients showed good patency after lymphovenous anastomosis. CONCLUSIONS: Supermicrosurgery in the case of LVA is minimally invasive, highly effective, and shows a very low complication rate. The surgeon/equipment-related factors restrict the pratice of LVA, and its effectiveness limited by technical constraints. Show more

Keywords: Lymphedema, lymph node transplantation, lymph vessel transplantation, LVA, augmented reality, indocyanine green (ICG)

DOI: 10.3233/CH-199220

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 125-133, 2019

Authors: Apfelbeck, M. | Chaloupka, M. | Schlenker, B. | Stief, C.G. | Clevert, D.-A.

Article Type: Research Article

Abstract: INTRODUCTION: Focal therapy (FT) of the prostate for low risk prostate cancer (PCa) is an alternative to traditional definite treatment options like external beam radiotherapy or radical prostatectomy. However, follow up after FT is still challenging and is subject to current studies. Significance of imaging after FT such as multiparametric MRI (mpMRI) is currently not well established. In this study, we aimed to evaluate the efficacy of alternative imaging during the follow up of low risk PCa treated with focal HIFU therapy using CEUS and image fusion. MATERIALS AND METHODS: Retrospective single arm study in patients with uni- …or bilateral, low or intermediate risk prostate cancer treated with HIFU at our institution between October 2016 and January 2018. CEUS in combination with image fusion using an axial T2-weighted MRI sequence was performed during follow up 3, 6, 9 and 12 months after the therapy. RESULTS: 4 consecutive patients with Gleason score (GS) 6 and 4 patients with GS 7a prostate cancer were included in the study. Hemiablation was performed in 7 patients with unilateral tumor. One patient underwent whole gland treatment due to histological proven bilateral PCa. Mean patient age at time of therapy was 70.3 (54–83) years and mean Prostate-specific antigen (PSA) level prior treatment was 7.8 ng/ml (2.1–14.4), after 3 months mean PSA level was 3.9 ng/ml (0.1–7.2), after 6 months 3.5 ng/ml (0.2–6.0), after 9 months 3.1 ng/ml (0.2–6.8) and 3.3 ng/ml (0.2–6.1) after 12 months. CEUS showed no signs of microvascularisation after 3, 6, 9 and 12 months in the ablated zone. 3 months posttreatment the necrotic tissue was still visible in the B-mode scan, although with no signs of vascularization performing CEUS. After 6 months the ablated side of the prostate was almost completely atrophic. And after 9 months the necrotic tissue was completely resolved. Between 9 and 12 months no changes in microvascularisation and perfusion could be shown. CONCLUSIONS: MpMRI/CEUS image fusion is a cost-effective and feasible technique to monitor the perfusion of the ablation zone after focal therapy of the prostate. Show more

Keywords: Prostate cancer, high intensity focused ultrasound therapy, contrast enhanced ultrasound, image fusion, follow up

DOI: 10.3233/CH-199222

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 135-143, 2019

Authors: Goetz, J. | Kaisermayer, E. | Haase, H. | Jünger, M. | Riebe, H.

Article Type: Research Article

Abstract: BACKGROUND: Although the efficacy of compression therapy through knee-length compression stockings with an interface pressure of 18–21 mmHg for leg oedemas has been confirmed by many studies, 91% of patients are still treated with a higher interface pressure. METHOD: In this prospective open randomised monocentric study 19 patients (age ≥65, movement restrictions and symptomatic leg oedema) received knee-length compression stockings with an interface pressure of 18–21 mmHg (stocking type 1 ) and 23–32 mmHg (stocking type 2 ). On two consecutive days each of the two compression stocking types were worn for at least 8 hours. After this test period, both …stocking types were assessed subjectively and skin changes were recorded. RESULT: A significantly (p -value <0.001) subjective improvement of the symptoms and complaints based on leg oedema was reported with the knee-length compression stockings. The Stocking type 1 recorded a significantly (p  = 0,045) better wearing comfort. In addition, the Stocking type 2 was significantly too large amongst women in the front foot area (p  = 0.044). The most common side effects were constrictions on the proximal lower leg (stocking type 1 = 73,7% (14/19); stocking type 2 = 78,9% (15/19)). Subjects with arthritis (p  = 0.006), hallux valgus (p  = 0.034) and/or digitus flexus (p  = 0.021) found the socking type 1 significantly more comfortable. CONCLUSION: In order to achieve optimal patient compliance it is recommended to prescribe knee-length compression stockings with an interface pressure of 18–21 mmHg if the following criteria are met: age ≥65 years, female sex, arthritis, digitus flexus (claw toe) or hallux valgus. Show more

Keywords: Chronic leg oedema, compression stocking therapy, chronic venous insufficiency, quality of life, wearing comfort

DOI: 10.3233/CH-199207

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 145-156, 2019

Authors: Asam, Claudia | Buerger, Korbinian | Felthaus, Oliver | Brébant, Vanessa | Rachel, Reinhard | Prantl, Lukas | Witzgall, Ralph | Haerteis, Silke | Aung, Thiha

Article Type: Research Article

Abstract: BACKGROUND: Doxorubicin is a cytostatic drug from the group of anthracycline antibiotics that is widely used as a chemotherapeutic agent. Side effects of the active substance include cardiotoxicity and nephrotoxicity. Doxorubicin-treated renal epithelial cells and (sarcoma) tumors are examined by correlative light and electron microscopy (CLEM) to investigate the subcellular localization of doxorubicin. METHODS: The kidney epithelial cell line MDCK II (Madin-Darby Canine Kidney) grown on culture dishes were treated with doxorubicin. Subsequently, the cells are analyzed by means of fluorescence and transmission electron microscopy (TEM). In vivo , alveolar rhabdomyosarcoma (RH 30) tumor cells are transferred to …the chorioallantoic membrane (CAM) of the chicken embryo. Doxorubicin is injected into a vein of the chicken embryo. After 24 hours, the tumor is removed and examined using CLEM. RESULTS: The kidney epithelial cells and the doxorubicin-injected tumors show a clear staining of the cell nucleus, which correlates with electron-dense regions (heterochromatin). High-resolution TEM shows that doxorubicin treatment leads to an enormous stress situation with an increased formation of membrane blebbings. CONCLUSIONS: CLEM is a promising new method to visualize the pattern of fluorescing drugs (e.g. doxorubicin) in renal epithelial cells and tumors, and to localize the drug in its subcellular context combined with high resolution. Show more

Keywords: Correlative light and electron microscopy (CLEM), chorioallantoic membrane (CAM), doxorubicin, chemotherapy, tumor, sarcoma, Epon

DOI: 10.3233/CH-199212

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 157-167, 2019

Authors: Haileka, V. | George, S. | Steinbrecht, S. | Jung, F. | Reinehr, R. | Küpper, J.-H.

Article Type: Research Article

Abstract: BACKGROUND: In most clinical studies older people are underrepresented compared to the demographic reality. However, risk for some severe diseases like cancer typically increase with age. Most insight into cancer treatment comes from mixed-age patient cohorts, leading to a lack of detailed understanding of cancer drug effects in the elderly population. There is growing evidence that cancer drug effects can be influenced by dehydration conditions often found in older people. Colon cancer remains the second leading cause of death by cancer in Europe. Inter- and intra-heterogeneity of tumors contribute to why some individuals do not respond to specific cancer therapies …or may often suffer a relapse. OBJECTIVE: Our study applies an in vitro drug test system for simulating treatment with cytostatics of colorectal cancer in elderly patients with dehydration condition. METHODS: Two well-known colon cancer cell lines, Caco-2 and RKO, harboring defined cancer-related mutations, were step-wisely adapted from routine culture medium to a severe hyperosmotic condition (397 mOmol/kg) by adding sodium chloride to the medium. We investigated the effects of these cell culture conditions, which should mimic cellular dehydration in elderly people, on the growth characteristics of the cells. Therefore, cell proliferation was investigated by measuring population doubling times. Furthermore, we investigated how the metabolic activity of the cells was influenced by treatment with different concentrations of cyclophosphamide (CPA) under normal and hyperosmotic conditions. RESULTS: We found that Caco-2 and RKO cell lines have an identical cell doubling time of 23 hours in normosmotic medium. However, hyperosmotic medium lifted the doubling time of Caco-2 cells to 31 hours while that of RKO cells did not change. Despite reduced cell proliferation rates, hyperosmotic medium sensitized Caco-2 cells to treatment with 10 mM CPA for 48 hours as measured by metabolic activity assays on ATP levels. CONCLUSIONS: The two investigated colon cancer cells lines reacted differently to hyperosmotic conditions. Only the growth of Caco-2 cells was reduced by increased osmolality. Despite this reduced growth their sensitivity to an alkylating cytostatic agent was even slightly increased. We are now in line to examine these effects in more detail and with more tumor cell lines. Show more

Keywords: Colon cancer, cyclophosphamide, dehydration, hyperosmotic, osmolality

DOI: 10.3233/CH-199210

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 169-176, 2019

Authors: Neuber, Christin | Schulze, Sabine | Förster, Yvonne | Hofheinz, Frank | Wodke, Johanna | Möller, Stephanie | Schnabelrauch, Matthias | Hintze, Vera | Scharnweber, Dieter | Rammelt, Stefan | Pietzsch, Jens

Article Type: Research Article

Abstract: Biomaterials coated with artificial extracellular matrices (aECM) are intended to support the healing of critical size bone defects. This pilot study investigated (i) the feasibility of dual-tracer PET/CT imaging for functional characterization of biomaterial-assisted bone healing in a rat femoral defect model and (ii) the bone healing ability of polycaprolactone-co -lactide (PCL) scaffolds, coated with various aECM consisting of collagen type I (Col) and glycosaminoglycans (GAGs) such as chondroitin sulfate (CS) or polysulfated hyaluronan (sHA3). [18 F]FDG and [18 F]fluoride PET 4 and 8 weeks after implantation of aECM-coated PCL scaffolds, which provide an in vivo measure of cellular …activation and bone mineralization, respectively, combined with CT imaging (in vivo/ex vivo ) and histological/immunohistochemical investigations (ex vivo ) showed that coating with CS in particular is beneficial for bone healing. The possible involvement of COX-2 and TGase 2, key enzymes of inflammation and ECM remodeling, in these processes offers starting points for targeted adjuvant therapy in the course of various bone healing phases. Our investigations show the feasibility of the selected dual-tracer approach for PET/CT imaging. In principle, this approach can be extended by further PET tracers for the functional characterization of physiological processes such as hypoxia/reperfusion or selected molecular players. Show more

Keywords: Artificial extracellular matrices, bone healing, [18F]fluorodeoxyglucose, [18F]fluoride, glycosaminoglycans, inflammation, metabolism, molecular imaging, revascularization

DOI: 10.3233/CH-199208

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 177-194, 2019

Authors: Nies, A. | Proft, L. | Nehring, M.E. | Gruber, C. | Sievers, H. | Hünigen, H. | Rodrigues, A.G. | Gemeinhardt, O. | Mrowietz, C. | Jung, F. | Hiebl, B.

Article Type: Research Article

Abstract: BACKGROUND: In cardiovascular research small pigs breeds like Göttingen® minipigs (GM) are established animal models, but systematic data about the micromorphology of the GM vasculature at different ages are scarce. OBJECTIVE: The study was aimed at gaining knowledge about the micromorphology of the femoral artery (FA) from German Landrace pigs (DL) and GM during the period of growth over a body weight range of 10–40 kg. METHODS: FA samples from DL aged two or three months were compared to GM ones, aged 18 or 40 months using transmitted light microscopy. RESULTS: All FA samples …showed typical characteristics of muscular arteries. Growth was associated with increased vessel wall thickness. In the GM this resulted in a slight decrease of the luminal diameter (LD), while in the DL pigs, an increase of the LD and smooth muscle cell content (10%) with decreased elastic fiber content (10%) has been detected. In contrast, within the 22 months lasting growth period of the GM, the tunica media content of smooth muscle cells and elastic fibers remained stable. CONCLUSIONS: FA maturation strongly depends on the pig breed and age. It can be different from what is described in humans. Show more

Keywords: Maturation, micromorphology, muscular artery type, porcine femoral artery, tunica media

DOI: 10.3233/CH-199219

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 195-201, 2019

Authors: Monteleone, Adrian | Schary, Weronika | Fath, Andreas | Wenzel, Folker

Article Type: Research Article

Abstract: INTRODUCTION: Since the beginning of industrial production in 1950, plastic production has continued to grow strongly worldwide and is now at 322 million tonnes in the year 2015. From these very high production volumes ever larger quantities are found in the environment. There the plastics degradate to microplasticity and spread ubiquitously in the world. The present work deals with the possible uptake of microplastic particles in human organisms. For the detection of these plastic particles, an extraction method was developed and validated. MATERIALS AND METHODS: Biological materials consist of human blood (healthy volunteers, n  = 4) and different tissues …of pigs and cattles. Various lysis solutions were tested for degradation efficiency of biological material and for effects on the plastics. The mass loss, surfaces and structure variations as well as the physicochemical spectrum of the material were observed after treatment by atomic force (AFM) and electron microscopy (EM) and Fourier transform infrared spectrometry (FTIR). RESULTS: The different plastic types as polyamide (PA), polycarbonate (PC), polyethylene (PE), polypropylene (PP), polystyrene (PS) and polyvinyl chloride (PVC) could be clearly differentiated and identified by FTIR. Regarding the surface control, especially PVC showed detectable alterations: After extraction an irregular surface structure caused by protuberances or bubbles could be observed. However, instead of these alterations an equivalent count of plastic particles was found in correlation to the applied plastic amount (recovery rate overall was 99,12±0,67%). CONCLUSION: The applied method can be used for plastic extractions from human or animal tissues without remarkable effects on the plastics. Show more

Keywords: Microplastic, microparticel, extraction method, human tissue

DOI: 10.3233/CH-199209

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 203-217, 2019

Authors: Sun, Xianlei | Tung, Wingtai | Wang, Weiwei | Xu, Xun | Zou, Jie | Gould, Oliver E. C. | Kratz, Karl | Ma, Nan | Lendlein, Andreas

Article Type: Research Article

Abstract: Electrospinning has attracted significant attention as a method to produce cell culture substrates whose fibrous structure mimics the native extracellular matrix (ECM). In this study, the influence of E-modulus of fibrous substrates on the lineage commitment of human adipose-derived stem cells (hADSCs) was studied using fiber meshes prepared via the electrospinning of a polyetheresterurethane (PEEU) consisting of poly(ρ -dioxanone) (PPDO) and poly(ɛ -caprolactone) (PCL) segments. The PPDO: PCL weight ratio was varied from 40:60 to 70:30 to adjust the physiochemical properties of the PEEU fibers. The cells attached on stiffer PEEU70 (PPDO:PCL,= 70:30) fiber meshes displayed an elongated morphology compared to …those cultured on softer fibers. The nuclear aspect ratio (width vs. length of a nucleus) of hADSCs cultured on softer PEEU40 (PPDO:PCL = 40:60) fibers was lower than on stiffer fibers. The osteogenic differentiation of hADSCs was enhanced by culturing on stiffer fibers. Compared to PEEU40, a 73% increase of osteocalcin expression and a 34% enhancement of alkaline phosphatase (ALP) activity was observed in cells on PEEU70. These results demonstrated that the differentiation commitment of stem cells could be regulated via tailoring the mechanical properties of electrospun fibers. Show more

Keywords: Electrospun fibers, polyetheresterurethane, E-modulus, mesenchymal stem cells, osteogenic differentiation

DOI: 10.3233/CH-199206

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 219-228, 2019

Authors: Tung, Wing Tai | Wang, Weiwei | Liu, Yue | Gould, Oliver E. C. | Kratz, Karl | Ma, Nan | Lendlein, Andreas

Article Type: Research Article

Abstract: The mechanical properties of electrospun fiber meshes typically are measured by tensile testing at the macro-scale without precisely addressing the spatial scale of living cells and their submicron architecture. Atomic force microscopy (AFM) enables the examination of the nano- and micro-mechanical properties of the fibers with potential to correlate the structural mechanical properties across length scales with composition and functional behavior. In this study, a polyesteretherurethane (PEEU) polymer containing poly(p -dioxanone) (PPDO) and poly(ɛ -caprolactone) (PCL) segments was electrospun into fiber meshes or suspended single fibers. We employed AFM three point bending testing and AFM force mapping to measure the …elastic modulus and stiffness of individual micro/nanofibers and the fiber mesh. The local stiffness of the fiber mesh including the randomized, intersecting structure was also examined for each individual fiber. Force mapping results with a set point of 50 nN demonstrated the dependence of the elasticity of a single fiber on the fiber mesh architecture. The non-homogeneous stiffness along the same fiber was attributed to the intersecting structure of the supporting mesh morphology. The same fiber measured at a point with and without axial fiber support showed a remarkable difference in stiffness, ranging from 0.2 to 10 nN/nm respectively. For the region, where supporting fibers densely intersected, the stiffness was found to be considerably higher. In the region where the degrees of freedom of the fibers was not restricted, allowing greater displacement, the stiffness were observed to be lower. This study elucidates the relationship between architecture and the mechanical properties of a micro/nanofiber mesh. By providing a greater understanding of the role of spatial arrangement and organization on the surface mechanical properties of such materials, we hope to provide insight into the design of microenvironments capable of regulating cell functionality. Show more

Keywords: Biomaterials, AFM, electrospinning, elastic modulus, force mapping

DOI: 10.3233/CH-199201

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 229-236, 2019

Authors: Zou, Jie | Wang, Weiwei | Nie, Yan | Xu, Xun | Ma, Nan | Lendlein, Andreas

Article Type: Research Article

Abstract: Laminin-5 (Ln-5), an important extracellular matrix (ECM) protein, plays a critical role in regulating the growth and differentiation of mesodermal tissues, including bone. Ln-5 can be secreted by the mesenchymal stem cells (MSCs), and Ln-5 promotes MSCs osteogenic differentiation. It has been demonstrated that a substrate’s surface topography could regulate MSC secretion and differentiation. A better understanding of the mechanism of how Ln-5 and surface roughness regulate MSC osteogenic differentiation would guide the design of surface topography and coatings of orthopedic implants and cell culture substrates. However, few studies have investigated the relationship between surface roughness and the secretion of …Ln-5 in MSC osteogenic differentiation. Whether substrate surface topography regulates MSC differentiation via regulating Ln-5 secretion and how surface topography contributes to the secretion of Ln-5 are still not known. In this study, the influence of microscale roughness at different levels (R0, R1 and R2) on the secretion of Ln-5 of human bone marrow MSCs (hBMSCs) and subsequent osteogenic differentiation were examined. hBMSCs spreading, distribution and morphology were greatly affected by different roughness levels. A significantly higher level of Ln-5 secretion was detected on R2, which correlated to the local cell density regulated by the rough surface. Ln-5 binding integrins (α 2 and α 3) were strongly activated on R2. In addition, the results from hBMSCs on R0 inserts with different cell densities further confirmed that local cell density regulated Ln-5 secretion and cell surface integrin activation. In addition, the mineralization level of MSCs on R2 was remarkably higher than that on R0 and R1. These results suggest that hBMSC osteogenic differentiation level on R2 roughness was enhanced via increased Ln-5 secretion that was attributed to rough surface regulated local cell density. Thus, the microroughness could serve as effective topographical stimulus in cell culture devices and bone implant materials. Show more

Keywords: Mesenchymal stem cells, roughness, cell density, laminin-5, osteogenesis

DOI: 10.3233/CH-199205

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 237-247, 2019

Authors: Schulz, Christian | Kammerer, Sarah | Küpper, Jan-Heiner

Article Type: Research Article

Abstract: BACKGROUND: Human hepatocyte in vitro cell culture systems are important models for drug development and toxicology studies in the context of liver xenobiotic metabolism. Often, such culture systems are used to elucidate the biotransformation of xenobiotics or drugs and further investigate drug and drug metabolite effects on biological systems in terms of potential therapeutic benefit or toxicity. Human hepatocytes currently used for such in vitro studies are mostly primary cells or cell lines derived from liver cancers. Both approaches have limitations such as low proliferation capacity and progressive dedifferentiation found in primary cells or lack of liver functions …in cell lines, which makes it difficult to reliably predict biotransformation of xenobiotics in patients. In order to overcome these limitations, HepaFH3 cells and Upcyte® hepatocytes representing primary-like hepatocytes of the first and second generation are increasingly used. Based on primary human hepatocyte cells transduced for stable expression of Upcyte® proliferation genes, they are mitotically active and exhibit liver functions over an extended period, making them comparable to primary human hepatocytes. These hepatocyte models show active liver metabolism such as urea and glycogen formation as well as biotransformation of xenobiotics. The latter is based on the expression, activity and inducibility of cytochrome P450 enzymes (CYP) as essential phase I reaction components. However, for further characterisation in terms of performance and existing limitations, additional studies are needed to elucidate the mechanisms involved in phase I reactions. One prerequisite is sufficient activity of microsomal NADPH-cytochrome P450 reductase (POR) functionally connected as electron donor to those CYP enzymes. OBJECTIVE: For Upcyte® hepatocytes and HepaFH3 cells, it is so far unknown to what extent POR is expressed, active, and may exert CYP-modulating effects. Here we studied POR expression and corresponding enzyme activity in human hepatoblastoma cell line HepG2 and compared this with HepaFH3 and Upcyte® hepatocytes representing proliferating primary-like hepatocytes. METHODS: POR expression of those hepatocyte models was determined at mRNA and protein level using qRT-PCR, Western Blot and immunofluorescence staining. Kinetic studies on POR activity in isolated microsomes were performed by a colorimetric method. RESULTS: The investigated hepatocyte models showed remarkable differences at the level of POR expression. Compared to primary-like hepatocytes, POR expression of HepG2 cells was 4-fold higher at mRNA and 2-fold higher at protein level. However, this higher expression did not correlate with corresponding enzyme activity levels in isolated microsomes, which were comparable between all cell systems tested. A tendency of higher POR activity in HepG2 cells compared to HepaFH3 (p  = 0.0829) might be present. Compared to primary human hepatocyte microsomes, POR activity was considerably lower in all hepatocyte models. CONCLUSION: In summary, our study revealed that POR expression and activity were clearly detectable in all in vitro hepatocyte models with the highest POR expression in cancer cell line HepG2. However, POR activity was lower in tested hepatocyte models when compared to human primary hepatocyte microsomes. Whether this was caused by e.g. polymorphisms or metabolic differences of investigated hepatocyte models will be target for future studies. Show more

Keywords: Biotransformation, CYP, cytochrome P450 monooxygenase, HepaFH3, HepG2, hepatocytes, NADPH-cytochrome P450 reductase, POR, Upcyte® hepatocytes, xenobiotics

DOI: 10.3233/CH-199226

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 249-260, 2019

Authors: Lamby, P. | Krüger-Genge, A. | Franke, R.P. | Mrowietz, C. | Falter, J. | Graf, S. | Schellenberg, E.L. | Jung, F. | Prantl, L.

Article Type: Research Article

Abstract: Repeated injections of iodinated contrast media (CM) can lead to a deterioration of the renal blood flow, can redistribute blood from the renal cortex to other parts of the kidney and can cause small decreases of the blood flow in cortical capillaries, a significant reduction in blood flow in peritubular capillaries and a significant reduction in blood flow in the vasa recta. Therefore, a study in pigs was designed, to show whether the repeated injection of CM boli, alone, can cause a reduction of oxygenation in the cortico-medullar renal tissue – the region with the highest oxygen demand in the …kidney - of pigs. While the mean pO2 -value had only decreased by 0.3 mmHg from 29.9±4.3 mmHg to 29.6±4.3 mmHg (p  = 0.8799) after the tenth Iodixanol bolus, it decreased by 5.9 mmHg from 34.0±4.3 mmHg to 28.1±4.3 mmHg after the tenth Iopromide bolus (p  = 0.044). This revealed a remarkable difference in the influence of these CM on the oxygen partial pressure in the kidney. Repeated applications of CM had a significant influence on the renal oxygen partial pressure. In line with earlier studies showing a redistribution of blood from the cortex to other renal areas, this study revealed that Iodixanol – in contrast to Iopromide - induced no changes in the pO2 in the cortico-medullar region which confirms that Iodixanol did not hinder the flow of blood through the renal micro-vessels. These results are in favor of a hypothesis from Brezis that a microcirculatory disorder might be the basis for the development of CI-AKI. Show more

Keywords: Contrast agent, Iodixanol, Iopromide, tissue oxygen partial pressure, CI-AKI, contrast-induced acute kidney injury

DOI: 10.3233/CH-199009

Citation: Clinical Hemorheology and Microcirculation, vol. 73, no. 1, pp. 261-270, 2019