Clinical Hemorheology and Microcirculation - Volume 67, issue 3-4

Authors: Zou, Jie | Wang, Weiwei | Neffe, Axel T. | Xu, Xun | Li, Zhengdong | Deng, Zijun | Sun, Xianlei | Ma, Nan | Lendlein, Andreas

Article Type: Research Article

Abstract: Polymeric matrices mimicking multiple functions of the ECM are expected to enable a material induced regeneration of tissues. Here, we investigated the adipogenic differentiation of human adipose derived mesenchymal stem cells (hADSCs) in a 3D architectured gelatin based hydrogel (ArcGel) prepared from gelatin and L-lysine diisocyanate ethyl ester (LDI) in an one-step process, in which the formation of an open porous morphology and the chemical network formation were integrated. The ArcGel was designed to support adipose tissue regeneration with its 3D porous structure, high cell biocompatibility, and mechanical properties compatible with human subcutaneous adipose tissue. The ArcGel could support initial …cell adhesion and survival of hADSCs. Under static culture condition, the cells could migrate into the inner part of the scaffold with a depth of 840±120 μm after 4 days, and distributed in the whole scaffold (2 mm in thickness) within 14 days. The cells proliferated in the scaffold and the fold increase of cell number after 7 days of culture was 2.55±0.08. The apoptotic rate of hADSCs in the scaffold was similar to that of cells maintained on tissue culture plates. When cultured in adipogenic induction medium, the hADSCs in the scaffold differentiated into adipocytes with a high efficiency (93±1%). Conclusively, this gelatin based 3D scaffold presented high cell compatibility for hADSC cultivation and differentiation, which could serve as a potential implant material in clinical applications for adipose tissue reparation and regeneration. Show more

Keywords: Mesenchymal stem cells, gelatin based scaffold, adipose tissue regeneration, adipogenic differentiation

DOI: 10.3233/CH-179210

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 297-307, 2017

Authors: Krüger-Genge, A. | Hiebl, B. | Franke, R.P. | Lendlein, A. | Jung, F.

Article Type: Research Article

Abstract: In drug eluting stents the cytostatic drugs Sirolimus or Tacrolimus are used to inhibit blood vessel restenosis by limiting the proliferation of smooth muscle cells. However, the cytostatic activity of both drugs was shown to be not cell specific and could also affect the stent endothelialisation, respectively. Currently, only limited in vitro data are available about the impact of Sirolimus and Tacrolimus on endothelial cell proliferation over a broad concentration range. To answer this question the following study was performed. Commercially obtained HUVEC were expanded with DMEM cell culture medium (GIBCO, Germany) supplemented with 5 vol% fetal calf …serum on non-coated regular polystyrene-based 24-multiwell plates. For drug testings 2×104 cells/cm2 were seeded and grown for 24 h until 30–40% of the multiwell surfaces were covered and then exposed to Sirolimus (1.0×10–11 – 1.0×10–5  mol/l) or Tacrolimus (2.0×10–8 – 6.2×10–5  mol/l), both dissolved in DMSO. 12, 24 and 48 h after adding the drugs cell numbers per area were quantified by counting the cells in six wells with four fields of view per well, representing 0.6 mm2 , using a confocal laser microscope. After 48 h of cell growth in the drug-free cell culture medium, the HUVEC number increased from 2.0×104 to 3.55×104 cells/cm2 (mean cell doubling time: 53.6 h, n = 6). At lower concentrations (≤2.0×10–6  mol/l) Tacrolimus reduced the number of adherent HUVEC significantly less than Sirolimus (p  < 0.05). However, at higher concentrations (≥2.07×10–5  mol/l) the effect of Tacrolimus on the number of adherent endothelial cells was significantly greater than that of Sirolimus (p  < 0.05). At the highest concentration applied (6.22×10–5  mol/l), Tacrolimus induced detachment of all HUVECs within 12 h after drug application. The number of adherent HUVEC decreased only slightly (about 9%) after Sirolimus application at the highest concentration (1.09×10–5  mol/l). These data show that in a non-flow model the cytostatic drug Tacrolimus reduced the number of adherent endothelial cells less than Sirolimus, as long as the drug concentration did not surpass 10–6  mol/l. At the limits of solubility, Sirolimus (1×10–5  mol/l) reduced the number of adherent endothelial cells less than Tacrolimus (6×10–5  mol/l), which induced detachment of endothelial cells. Show more

Keywords: Endothelial cells, Sirolimus, Tacrolimus, DMSO

DOI: 10.3233/CH-179211

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 309-318, 2017

Authors: Broer, P. Niclas | Aung, Thiha | Heidekrueger, Paul I. | Prantl, L. | Narayan, Deepak

Article Type: Research Article

Abstract: OBJECTIVE: The interleukin-1 receptor antagonist (IL-1RA) contributes to tumor survival and progression in multiple cancer entities. IL-1RA polymorphisms influence IL-1RA expression patterns and function. A known polymorphism was correlated with clinical outcomes in melanoma patients with particularly aggressive disease. METHODS: DNA of 343 controls and 97 melanoma patients with poor prognostic indicators (time from diagnosis to death, nodal status, metastasis) was analyzed for a variable number of tandem repeat polymorphisms (VNTR) of the IL-1RA gene. Five alleles containing two (allele 2), three (allele 4), four (allele 1), five (allele 3) or six (allele 5) 86-bp repeats were targeted …via PCR amplification. RESULTS: Genotype 1/2 is less common in the melanoma patient group vs. the control (28.8% vs. 39.6%; p  = 0.06). Significant was the stage of the melanoma in order to predict the survivability (p  = 0.008). The 1/1 and 1/2 genotype appeared to have lower hazards ratios than the 2/2 genotype (p  > 0.05). CONCLUSIONS: Compared to the general population, the distribution of alleles coding for IL-1RA is different in melanoma patients. This alteration and the potential impact on tumor protein function and systemic inflammatory response may warrant further investigation. Show more

Keywords: Melanoma, interleukin-1 receptor antagonist, IL-1RA, polymorphism, inflammation

DOI: 10.3233/CH-179212

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 319-326, 2017

Authors: Lutter, Anne-Helen | Scholka, Jenny | Richter, Heiko | Anderer, Ursula

Article Type: Research Article

Abstract: BACKGROUND: Tetrazolium-based assays are optimized to assess proliferation/toxicity of monolayer or suspension cells in microtiter plates. With regard to tissue engineering and regenerative medicine the need for in vivo like 3D microtissues has an increasing relevance. Applying tetrazolium-based assays to 3D culture systems is technically more challenging. The composed microenvironment may influence the assay standards, e.g. equal distribution of tetrazolium. OBJECTIVE: Evaluation of membrane-impermeable tetrazolium salt-based assays with regard to spheroid culture (3D) of human chondrocytes. METHODS: Chondrocytes were isolated from human articular cartilage. XTT, WST-1, and WST-8 were applied to monolayer cells (2D, varying …cell numbers) and spheroids (3D, different sizes) in 96well plates. Formazan formation was measured spectrophotometrically after different incubation periods. Evaluation was done using phase contrast microsopy (toxicity), analyzing the correlation of cell number and absorbance signals (Gompertz function), and document signal over background ratio. RESULTS: In monolayer culture the assays showed a correlation between seeded cell numbers and absorption data. Spheroid sizes are directly related to the starting cell number. A correlation between size and absorbance was only detectable starting from 10,000 cells/aggregate. Phase contrast microscopy of monolayer cells revealed strong toxicity effects of the WST-1 (4 h) and XTT (8 h) assay and no signs of toxicity using WST-8. CONCLUSION: The WST-8 assay is non-toxic and revealed the highest sensitivity in comparison to the XTT or WST-1 assay. There is evidence, that only cells of the outer rim of spheroids are able to convert membrane-impermeable tetrazolium salts to formazans. Show more

Keywords: XTT, WST-1, WST-8, human chondrocytes, spheroid, 3D culture, 2D culture

DOI: 10.3233/CH-179213

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 327-342, 2017

Authors: Anadol, Remzi | Gori, Tommaso

Article Type: Research Article

Abstract: Coronary scaffolds have been recently developed to address the long-term limitations of metallic drug eluting stents. Concerns have however been expressed on the safety of these devices, with evidence of both early and late scaffold thrombosis. While early thrombosis has been associated with incomplete scaffold expansion, leading to flow disturbances, blood recirculation, and platelet activation, the pathophysiology of late events remains less understood. Recent cases series have shown that malapposition and scaffold dismantling might play a role in this phenomenon, an observation that further confirms the importance of an accurate implantation. Further, the role of dual antiplatelet therapy, and whether …prolonging it may reduce event rates, remains to be elucidated. As well, the role of inflammatory phenomena has been proposed but never demonstrated. This brief review summarizes the current evidence on these phenomena. Show more

Keywords: Bioresorbable scaffolds, stent thrombosis

DOI: 10.3233/CH-179214

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 343-346, 2017

Authors: Mrowietz, C. | Franke, R.P. | Pindur, G. | Wolf, U. | Jung, F.

Article Type: Research Article

Abstract: The Laser Doppler technique (Laser-Doppler-Fluxmetry, LDF), a noninvasive method to estimate skin blood flow (LDF), is frequently used in research and clinical routine [1 ]. Here, the measurements were carried out with a new Laser Doppler system, the DOP-system, which allows to measure frequency spectra in four different frequency windows according to the velocities in venules (low velocity), capillaries (low to medium velocities), and in arteries (with high and very high velocities). However, the diagnostic reliability or the effectiveness of the LDF has not yet been evaluated sufficiently, which is indispensable, where medical diagnostics and therapy controls are concerned. For …a valid interpretation of LDF values of individual patients, the knowledge of the reference range and the variability of the measured parameters is required. In four successive studies the reference range (62 apparently healthy subjects), the circadian variability (8 subjects), the variability from day-to-day (6 subjects) and over one year with monthly measurements (6 subjects) were evaluated. With the knowledge of the reference range, microcirculatory disorders can now be diagnosed using the DOP method. Following a standard measurement procedure there was no dependence of the measured data on the day or season of measurement. Show more

Keywords: Laser Doppler Flux, reference range, variability, skin perfusion

DOI: 10.3233/CH-179215

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 347-353, 2017

Authors: Ranieri, M. | Wohlgemuth, W. | Müller-Wille, R. | Prantl, L. | Kehrer, A. | Geis, S. | Klein, S. | Lamby, P. | Schiltz, D. | Uller, W. | Aung, T. | Dolderer, J.H.

Article Type: Research Article

Abstract: This article presents our experience in managing peripheral vascular malformations of upper and lower extremities over a 4-year period in a series of 46 patients of the Department of Plastic Surgery treated in the Interdisciplinary Center of Vascular Anomalies (ICVA) at the University of Regensburg. The patients presented vascular malformations of upper and lower extremity and were selected from our prospective vascular anomalies file archive from 2012 to 2016. During this period in the ICVA at University of Regensburg were performed more than 1400 radiological interventional treatments in patients with vascular malformations. The purpose of this retrospective study was …to review combined embolotherapy, sclerotherapy (embolo/sclerotherapy), and surgical procedures (surgical excision and soft tissue reconstruction) to manage vascular malformations. Treatments were principally induced to reduce pain, daily physical limitations, social discomfort and recover tegument continuity after ulceration. The 46 patients were first examined with noninvasive radiological procedures. After diagnosis was posed, embolo/sclerotherapy, surgical procedures and clinically as well as radiological follow-ups were coordinated and established by the multidisciplinary team. All vascular malformations were categorized according to the classification approved at the April 2014 General Assembly of International Society for the Study of Vascular Anomalies (ISSVA) in Melbourne, Australia. Arteriovenous malformations (AVMs) were further classified following the Cho-Do and Schobinger classification. Embolo/sclerotherapy shows to be the most appropriate procedure in vascular malformations treatment. Nevertheless was found that in case of complications or lack of improvement as well as to improve functional or aesthetical results, a following partial or complete surgical excision and immediate soft tissue reconstruction seems to be the gold-standard treatment. In addition, the precise clinical and radiological diagnosis as well as an intensive postoperative patient care have a significant positive influence on the clinical outcome and patient satisfaction while decreasing morbidity and recurrence during early and late follow-up. Vascular malformations require a multidisciplinary approach and individual treatment after complex excision and indispensable reconstruction. Show more

Keywords: Vascular malformations, vascular malformations surgical therapy, vascular malformations soft tissue reconstruction, interdisciplinary treatment vascular malformations

DOI: 10.3233/CH-179216

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 355-372, 2017

Authors: Haimerl, M. | Brünn, K | Poelsterl, S. | Beyer, L.P. | Wiesinger, I. | Stroszczynski, C. | Jung, E.-M. | Wiggermann, P.

Article Type: Research Article

Abstract: PURPOSE: To compare the diagnostic performance of real-time maximum liver capacity (LiMAx) with dynamic contrast-enhanced ultrasound (CEUS)-based liver microcirculation. MATERIALS AND METHODS: 23 patients underwent liver function capacity (LiMAx) test and consecutive or previous CEUS examinations. A bolus injection of 1.4 ml sulfur hexafluoride microbubbles was administered for CEUS measurements (1–6 MHz) and quantitative perfusion analysis (TIC) was performed with an integrated perfusion software using stored cine-loops. Two perfusion-parameters, time to peak (TtoP) and area under the curve (Area), were evaluated in liver parenchyma and portal vein using TIC analysis. To compare quantification parameters, patients were classified in patients representing …a healthy population (LiMAx value >315 μg/kg/h) and those representing patients with liver disease (LiMAx value <315 μg/kg/h). RESULTS: Comparing perfusion parameters derived from portal vein measurements, TtoP and Area were higher in patients with normal liver function TtoP: 25.0±8.4 s, Area: 1483±920 a.u. compared to patients with impaired liver function TtoP: 22.4±14.0 s; Area 1351±1212 a.u. This difference however was not statistically significant (p  = 0.52, p  = 0.48). In parameters derived from measurements in liver parenchyma TtoP was higher (38.5±11.3 s) and Area was lower (999±632 a.u.) in patients with normal liver function compared to patients with impaired liver function (TtoP; 30.6±11.0 s, p  = 0.156; Area: 1202±719 a.u.) (p  = 0.16, p  = 0.56). In a simple linear regression model, none of the perfusion parameters measured in portal vein (TtoP portal, Area portal) and liver parenchyma (TtoP liver, Area liver) correlated significantly with respective LiMAx values (p  = 0.194–0.950). CONCLUSION: Within the framework of this study, CEUS-based perfusion parameters were not able to assess severity of liver disease, assessed with LiMAx- test. Show more

Keywords: Liver capacity, LiMAx, CEUS, microcirculation, liver function

DOI: 10.3233/CH-179217

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 373-382, 2017

Authors: Pregler, B. | Beyer, L.P. | Wiesinger, I. | Haimerl, M. | Stroszczynski, C. | Jung, E.M. | Wiggermann, P.

Article Type: Research Article

Abstract: PURPOSE: To determine the influence of pre-interventionally existing portal vein thrombosis on the ablation success of percutaneous tumor ablation of HCC. MATERIALS AND METHODS: 15 patients with HCC and pre-existing portal vein thrombosis underwent thermal tumor ablation. We retrospectively analyzed the pre- and post-interventionally performed CT and MRI scans in terms of technical success as well as the complication rate. The portal vein thrombosis was classified into segmental, lobar and central thrombus. RESULTS: In 13/15 cases (87%) complete ablation with no evidence of residual tumor tissue was seen 6 weeks after the procedure in contrast-enhanced MRI …scans and contrast-enhanced ultrasound (CEUS). No major and 2 minor complications were observed after the ablation procedure. CONCLUSION: Reduced perfusion due to pre-interventionally existing portal vein thrombosis has no significant impact on the ablation success or the complication rate. Show more

Keywords: HCC, portal vein thrombosis, ablation, RFA, MWA

DOI: 10.3233/CH-179218

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 383-388, 2017

Authors: Zengel, Pamela | Reichel, Christoph Andreas | Vincek, Teresa | Clevert, Dirk André

Article Type: Research Article

Abstract: PURPOSE: Chronic recurrent parotitis (CRP) is a non-obstructive disease with episodes characterized by painful swelling of the parotid gland. It presents in both a juvenile and an adult form, with no clear information on its actual origin. Diagnosis is based on patient medical history and ultrasound examination but is frequently not correctly identified. Acoustic Radiation Force Impulse Imaging (ARFI) is a novel ultrasound elastography technology that has recently been implemented in the diagnostic work-up of patients with malignancies. This study aimed to answer whether ARFI can reasonably be employed in the initial examination and follow-up during therapy in patients with …CRP. MATERIAL AND METHODS: Mechanical tissue properties of the salivary glands were analyzed by ARFI in 37 parotid glands of patients with CRP. RESULTS: Having integrated ARFI into our diagnostic protocol for CRP, affected parotid glands were found to exhibit lower tissue elasticity compared to both healthy contralateral glands in the same individuals as well as those of healthy individuals. Most importantly, this method enabled us to quantitatively assess the patient benefit of therapy regarding the recovery of the glands’ diseased parenchyma. CONCLUSIONS: ARFI provides a quick, easy, and reliable diagnostic tool for the assessment of disease severity and progression in patients with CRP that can be seamlessly implemented into preexisting ultrasound protocols. Show more

Keywords: Chronic recurrent parotitis, ultrasound, elastography, ARFI imaging, tissue elasticity

DOI: 10.3233/CH-179219

Citation: Clinical Hemorheology and Microcirculation, vol. 67, no. 3-4, pp. 389-397, 2017