Biomedical subjects

Thomas Knoll

Publications and source records attributed to Thomas Knoll.

34 records · Page 2Linked to original sources

PubMed2004

In vitro and ex vivo gene delivery into proximal tubular cells by means of laser energy--a potential approach for curing cystinuria?

Cystinuria is the cause of 1-2% of stones observed in adults and about 10% of those occurring in children. Recurrent stone formation and multiple operations cause considerable morbidity. We investigated the transfection efficiency of naked plasmid DNA in porcine kidney cells by applying holmium laser (Ho:YAG) energy in vitro as well as ex vivo in a porcine kidney papilla model. In the in vitro experiments, naked plasmid DNA was added to LLC-PK1 cells suspended in a medium and Ho:YAG laser applied with varying pulses. The transfection efficiency was measured by the expression of EGFP reporter gene in the cells by FACScan analysis and fluorescence microscopy. Ex vivo, papilla from porcine kidney was excised and naked plasmid DNA was added to the tissue in the medium. The laser was then applied and the cryosectioned tissue observed under fluorescence microscope. The efficiency of transfection in vitro significantly improved with the increase in impulses (P<0.01). Transfection at 50 impulses averaged 0.7+/-0.3%, at 200 impulses 28.3+/-7.7%, and at 500 impulses 36.1+/-3.1%. The cell mortality rate increased with higher pulse rate up to 70%. Ex vivo trials showed transfection in extended regions of the tissue and also in the peripheral layers of the papilla. Our study indicates that the transfection of benign kidney cells by Ho:YAG is a promising new gene transfer strategy. The ex vivo trials showed that peripheral renal tissue layers are susceptible to transfection by Ho:YAG applied from the papillary surface.

Cystinuria↗

PubMed2004

Micelle delivery of doxorubicin increases cytotoxicity to prostate carcinoma cells.

The use of doxorubicin as a chemotherapeutic agent is hindered by its toxic side effects on the normal cells of the body. The objective of this study was to determine if micelle-delivered doxorubicin could increase the effectiveness of doxorubicin against prostate carcinoma cells. Rat prostate carcinoma cells (MatLu) were cultured under standard conditions. Phosphate-buffered saline (PBS), doxorubicin and/or micelle solution (Pluronic 10500 solution) was added to the cell suspensions and incubated for 3 h. After incubation, cells were washed twice. Analysis consisted of: 1) immediate cell count and 2) proliferation assay at 24 and 144 h. After 24 h, samples with micelle-incorporated doxorubicin had 75% (10% pluronic with 10 microg/ml doxorubicin) and 80% (1% pluronic with 10 microg/ml doxorubicin) cell proliferation results compared with the control group. After 144-h incubation, these same two groups demonstrated cell proliferation results of only 30 and 43% of the control group. The in vitro cytotoxicity of doxorubicin against prostate carcinoma cells was dramatically increased by incorporating the molecule with polymeric micelles.

Analysis of Variance↗

PubMed2004

Expression of pro-angiogenic growth factors VEGF, EGF and bFGF and their topographical relation to neovascularisation in prostate cancer.

The aim of the study was to quantify the expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF) in prostate cancer and adjacent non-tumorous tissue in a standardized experimental set-up and to evaluate the paracrine effects of three endothelial stimuli on neovascularisation. Immunohistochemical staining of prostate cancer (PCa) specimens for VEGF, bFGF, EGF and the endothelial marker CD31 was performed (n=56). Sections were analyzed for growth factor-positive cancer/epithelial cells as well as staining intensity in (I) malignant and (II) non-tumorous tissue. Within PCa the topographic relationship (TR) of maximum microvessel density (MWD) and maximum expression of each growth factor was assessed. The number of VEGF- and EGF-positive cells in PCa was significantly enhanced compared with non-tumorous tissue (p<0.0001), whereas there was no difference in staining intensity. In contrast, the staining intensity of bFGF sections revealed a stronger expression in non-tumorous tissue compared with PCa (p<0.0001). In benign glands, VEGF, EGF and bFGF expression is chiefly restricted to basal cells. VEGF and EGF displayed a close TR in 65 and 57% of cases, respectively, whereas bFGF revealed a close TR in only 43% of PCa specimens. The results outline the relationship of the investigated growth factors and angiogenesis in PCa, which is strongest for VEGF and EGF. The relevance of VEGF and EGF is underlined by the increased number of positive cancer cells. Although previously reported to be a pro-angiogenic growth hormone, bFGF appears to play an assimilably minor role in the angiogenesis of PCa.

Aged↗

PubMed2004

High-intensity focused ultrasound for ex vivo kidney tissue ablation: influence of generator power and pulse duration.

BACKGROUND AND PURPOSE: The therapeutic application of noninvasive tissue ablation by high-intensity focused ultrasound (HIFU) requires precise physical definition of the focal size and determination of control parameters. The objective of this study was to measure the extent of ex-vivo porcine kidney tissue ablation at variable generator parameters and to identify parameters to control lesion size. MATERIALS AND METHODS: The ultrasound waves generated by a cylindrical piezoceramic element (1.04 MHz) were focused at a depth of 100 mm using a parabolic reflector (diameter 100 mm). A needle hydrophone was used to measure the field distribution of the sound pressure. The morphology and extent of tissue necrosis were examined at generator powers of up to 400 W (P(el)) and single pulse durations of as long as 8 seconds. RESULTS: The two-dimensional field distribution resulted in an approximately ellipsoidal focus of 32 x 4 mm (-6 dB). A sharp demarcation between coagulation necrosis and intact tissue was observed. Lesion size was controlled by both the variation of generator power and the pulse duration. At a constant pulse duration of 2 seconds, a generator power of 100 W remained below the threshold doses for inducing a reproducible lesion. An increase in power to as high as 400 W induced lesions with average dimensions of as much as 11.2 x 3 mm. At constant total energy (generator power x pulse duration), lesion size increased at higher generator power. CONCLUSIONS: This ultrasound generator can induce defined and reproducible necrosis in ex-vivo kidney tissue. Lesion size can be controlled by adjusting the generator power and pulse duration. Generator power, in particular, turned out to be a suitable control parameter for obtaining a lesion of a defined size.

Animals↗

PubMed2003

Influence of acute selective endothelin-receptor-A blockade on renal hemodynamics in a rat model of chronic allograft rejection.

We have recently demonstrated up-regulation of renal endothelin (ET) synthesis in a rat model of chronic renal allograft rejection. Treatment with a selective ET-A receptor antagonist improved survival and reduced functional and morphological kidney damage. However, the underlying mechanisms have not yet been elucidated, as ET exhibits both hemodynamic and inflammatory properties. Therefore, in the present study we investigated acute hemodynamic effects of the selective ET-A receptor antagonist LU 302146 (LU) on chronic renal allograft rejection in rats. Experiments were performed in the Fisher-to-Lewis model of chronic renal allograft rejection. Lewis-to-Lewis isografts served as controls. After 2, 12, and 24 weeks, hemodynamic measurements were performed on anesthetized animals. Measurement of mean arterial pressure (MAP) was performed via a catheter in the femoral artery. Renal blood flow (RBF) was measured by an ultrasonic flow probe placed around the renal transplant artery. Medulla blood flow (MBF) and cortex blood flow (CBF) were determined with laser Doppler probes. Hemodynamic response upon intravenous bolus injection of LU (50 mg/kg) was investigated. The application of LU was followed by a decline in MAP that reached statistical significance only in isografts (ISOs) after 12 weeks and allografts (ALLOs) after 24 weeks. RBF slightly decreased in all groups; however, without reaching statistical significance. MBF showed a small increase in ALLO12 and ALLO24 whereas CBF slightly decreased in all groups. Acute ET-A receptor blockade does not induce important hemodynamic effects in kidneys undergoing chronic rejection. The lack of response to ET-A receptor blockade suggests that the beneficial effect of ET receptor antagonists in this model is likely to be due to improvement of renal morphology.

Animals↗

PubMed2003

The role of the carbachol test and concomitant diseases in patients with nonobstructive urinary retention undergoing sacral neuromodulation.

The aim of this study was to evaluate the percutaneous nerve evaluation (PNE) test success in patients with nonobstructive urinary retention. A total of 24 PNE tests were performed in patients with nonobstructive urinary retention and in 18 patients, a carbachol test was performed during urodynamics. The diagnosis relating to the acontractile detrusor was also assessed and compared to the outcome of the PNE test. The PNE test was successful in eight of 24 patients (33.3%) with the the highest success rate being observed in patients after hysterectomy (80%). It was successful in five of 12 patients with negative carbachol tests and in three of six patients with positive carbachol tests. We conclude that sacral neuromodulation is an effective treatment option in patients with nonobstructive urinary retention. PNE tests should be performed in all patients with therapy resistant nonobstructive urinary retention, because predictive factors do not exist.

Adult↗

PubMed2003

Measurement of renal anatomy for prediction of lower-pole caliceal stone clearance: reproducibility of different parameters.

BACKGROUND AND PURPOSE: The architecture of the lower renal pole seems to have a considerable influence on caliceal stone clearance after different therapeutic modalities. The published data are partially inconsistent, and publications on reproducibility are completely lacking. The aim of this study was to evaluate the intraobserver and interobserver reproducibility of different measures of lower-calix anatomy. MATERIALS AND METHODS: We studied the intraobserver and interobserver reproducibility of parameters describing the lower-pole anatomy that are significant for treatment outcome. Forty renal units without urologic disease were analyzed by five independent urologists. Infundibular length (IL), infundibular width (IW), and lower infundibulopelvic angles (LIP) were measured by the Elbahnasy (LIP I), Keeley (LIP II), and Gupta (LIP III and LIP IV) methods. Statistical analysis of each parameter and investigator was performed. RESULTS: All LIP angles showed low interobserver correspondence: correlation coefficients (CC) did not exceed 0.44 (P < 0.05). Even the relatively clearly defined parameters IW and IL achieved CCs of only 0.63 and 0.49, respectively. The intraobserver correlation achieved better results: 0.73 (LIP I), 0.84 (LIP II), 0.73 (LIP III), 0.65 (LIP IV), 0.88 (IL), and 0.82 (IW). With the Elbahnasy method, almost all renal units were classified as favorable for stone persistence after shockwave lithotripsy. With the Keeley and Gupta methods, more than 50% of the kidneys were defined as having low clearance probability. CONCLUSIONS: Assessment of the chosen parameters is difficult and shows high interobserver variation. Inexperience in measuring the specific angles and low imaging quality can limit correct evaluation. The large number of kidneys with anatomy inappropriate for clearance of lower-pole stones may explain the poor outcome of shockwave treatment for stones in a lower calix. Prospective studies will determine the clinical value of anatomic assessments.

Humans↗

PubMed2003

Oxidative stress in chronic renal allograft nephropathy in rats: effects of long-term treatment with carvedilol, BM 91.0228, or alpha-tocopherol.

Oxidative stress is markedly increased after kidney transplantation and may participate in the development and/or progression of chronic renal allograft nephropathy. In the present study we sought to assess the nephroprotective potential of antioxidative treatment in renal allograft recipients. Experiments were performed in the Fisher-Lewis rat model of chronic renal allograft nephropathy, with isografted Lewis rats serving as controls. Allografted rats were orally treated with carvedilol, an antihypertensive drug with antioxidative properties (25 mg/kg/d), its purely antioxidative derivative BM 91.0228 (5 mg/kg/d), alpha-tocopherol (100 mg/kg/d), a combination of propranolol/doxazosine (10/5 mg/kg/d), or vehicle for 24 weeks. At the end of the study, oxidative status and influence of antioxidative treatment were assessed in transplanted animals. Chronic allograft nephropathy was characterized by a marked increase of markers for oxidative stress (increased plasma and kidney levels of malondialdehyde, reduced glutathione, and tocopherol levels in renal allografts). Treatment with carvedilol, BM 91.0228, and tocopherol significantly improved antioxidative status of allograft kidney recipients. In addition, carvedilol reduced elevated blood pressure in allografted rats. However none of the drugs had a beneficial influence on functional and morphologic renal changes. Our data thus demonstrate that long-term treatment with the antioxidants carvedilol, BM 91.0228, or alpha-tocopherol does not prevent development of chronic transplant nephropathy, despite an improvement of antioxidative status.

Animals↗

PubMed2002

Flexible ureterorenoscopy for the treatment of lower pole calyx stones: influence of different lithotripsy probes and stone extraction tools on scope deflection and irrigation flow.

The 7.5 and 9.0 F flexible ureterorenoscopes are used widely in endourology. These scopes, along with new available tools and probes, enable intrarenal ureteroscopic access to renal stones in the lower pole. Following individual clinical experiences the infundibulopelvic angle has to be respected for the selection of different scopes, tools and probes. The aim of this study was to determine the influence of different scopes, tools and probes on the angle of the active and passive flexion of the flexible ureterorenoscope. The angles of the active and passive flexion and maximum irrigation flow of different flexible ureterorenoscope (9.0 F Wolf, 7.5 F Olympus, 7.5 F Storz) were measured repeatedly in vitro using nine different inserted tools and/or probes. In addition, the maximum allowing for easy passage of the nine different tools/probes through the channel of the scope were measured. The Olympus scope without working instrument present, allows for the greatest deflection. The flexible Lithoclast, the non-nitinol baskets, and the 365 microns laser probes significantly inhibit the deflection of all scopes. The greatest deflection with inserted working tool was possible using the Wolf and Storz scopes. Both nitinol tools as well as the 200 microns laser probe had only minimal influence on the deflection. The 200 microns laser probe reduces the irrigation flow from 50 to 28 ml/min. In contrast, the baskets reduce the irrigation flow from 50 to 2 ml/min. There are significant differences of the active and passive flexion depending on the ureterorenoscopes itself and the different tools. In order to perform a successful flexible ureterorenoscopic stone treatment in the lower pole, the inhibition of the flexion of the different scopes together with the individual infundibulopelvic angle must be respected when selecting different tools and probes.

Alloys↗

PubMed2002

Evaluation of superficial papillary ablation by endoscopic lasers in an ex vivo kidney model.

BACKGROUND AND PURPOSE: Endoscopic examinations of stone-forming kidneys show a coincidence of plaques and microliths on the surface of and within papillary epithelial tissue. These calcifications are thought to be precursors of calcium oxalate urolithiasis. We hypothesized that minimally invasive endoscopic laser ablation of microliths and necrotic cell layers enables epithelial regeneration and prevents recurrent urolithiasis. The aim of this study was to determine the most suitable laser type and dose intensity for selective superficial cell ablation. MATERIALS AND METHODS: Conventional Nd:YAG (1-40 W) or Ho:YAG (0.5-3 J/single impulse) lasers were used endoscopically on an ex vivo blood-perfused porcine kidney model. Defined doses were applied to the papillary surface in the contact and noncontact modes for 10 to 30 seconds. Papillae were excised after treatment and histopathologically analyzed in continuous sections. Lesions were microscopically assessed with the aid of a Leica Quantimed computer program. RESULTS: Depending on the time and dose, vaporization by the Nd:YAG laser caused large tissue defects and coagulation necrosis at energy levels over 5 W (contact and noncontact mode). Lower energy levels with tissue contact produced only superficial cell defects (<20 cell layers) but more extensive coagulation necrosis, whereas no histologic effects were observed at the same energy level without contact. In contrast, independent of delivered energy but dependent on time, Ho:YAG laser application caused pure tissue loss without relevant coagulation necrosis. The generation of small lesions (6-10 cell layers) without tissue contact was possible at energy levels under 2 J. CONCLUSIONS: Selective superficial papillary cell ablation is possible. Low-energy Nd:YAG treatment in the contact mode and Ho:YAG treatment in the noncontact mode led to superficial vaporization with no (Ho:YAG) or minimum (Nd:YAG) coagulation defects.

Animals↗

PubMed2002

The low normothermia concept--maintaining a core body temperature between 36 and 37 degrees C in acute stroke unit patients.

Elevated body temperature increases mortality and worsens outcome in acute stroke patients. In animal models of stroke, even slight hypothermia was shown to be neuroprotective. Pharmacological treatment alone (paracetamol, metamizol) usually fails to lower core body temperature below 37 degrees C. The purpose of this study was to establish the feasibility and safety of continuous body surface cooling towards low normothermic temperatures in noncomatose, nonventilated stroke unit patients. Eighteen acute stroke patients (15 ischemic infarcts, 3 hemorrhages) with baseline body core temperatures >37.0 degrees C (taken in the urinary bladder) were laid on a water-perfused cooling mattress and received pethidine and dihydroergotoxine in order to avoid shivering and peripheral vasoconstriction. The target range for core body temperature was between 36 and 37 degrees C for 24 hours. None of the patients was treated with antipyretic drugs during the cooling period. Median baseline National Institutes of Health Stroke Scale score (NIHSSS) was 15.5 (8-24). Three patients had core temperatures >38 degrees C. A temperature in the target range could be reached within 3.3 hours (median) and maintained in all but two patients. Major procedure-related adverse events were vomiting (n = 2), drop in mean arterial blood pressure >20% (n = 2), pneumonia (n = 2), and a rise in central venous pressure >20 cm H2O (n = 3) totaling 9 events in 8 of 18 patients (44%). No patient died within the first week; mortality after three months was 12%. Continuous body core temperature reduction of 1-2 degrees C may safely be attained by a cooling mattress in nonventilated stroke unit patients. Critically high temperature values can be avoided. The neuroprotective potential of this method has to be assessed in a controlled trial.

Acute Disease↗

PubMedDate not supplied

Intrinsic presence of poly (ADP-ribose) is significantly increased in malignant prostate compared to benign prostate cell lines.

Poly(ADP-ribosyl)ation plays important roles in cellular DNA repair mechanisms as well as in cellular proliferation and genomic stability. Four carcinoma cell lines, LNCaP, PC-3, C4-2 and MatLu, and one prostate epithelial cell line, PNT1A, were tested using the H10 antibody to poly(ADP-ribose) by Western blotting and FACS analysis for basal and activated (10 mM H2O2) PARP activity. In both analyses, higher levels of basal polymer were present in the LNCaP cell line than in the prostate epithelial cell line or the MatLu cell an line. LNCaP cells demonstrated greater than 90% positive analyzed cells in FACS analysis both before and after treatment with H2O2. PNT1A epithelial cells demonstrated evidence of polymer presence only after treatment with H2O2. Microsatellite instability due to increased oxidative damage in tumor cells has been proposed as a means for accumulation of genetic changes. As oxidative stress can stimulate poly(ADP-ribose) reactions, an increased presence of the polymer may indicate a high level of oxidative damage occurring in the tumor cells, contributing to higher levels of genetic instability towards progression.

Adenocarcinoma↗

PubMedDate not supplied

Expression of different vascular endothelial markers in prostate cancer and BPH tissue: an immunohistochemical and clinical evaluation.

BACKGROUND: The microvessel density (MVD) is often used as a quantitative parameter for angiogenesis. The aim of this study was to evaluate the three most commonly used endothelial markers of microvessels, CD31, CD34 and vWF, in benign and malignant prostatic tissue. MATERIALS AND METHODS: Fifteen benign hyperplastic prostate and 54 prostate cancer specimens were immunohistochemically stained for CD31/CD34/vWF. The MVD obtained by each marker was quantified by a vessel count in standardized grids within the area of maximum angiogenesis. The data on MVD was interrelated and compared to tumor staging, grading and clinical follow-up. RESULTS: A significant correlation of the CD31/CD34/vWF-MVD data was observed in BPH tissue, but not in PCa. In PCa, the most sensitive marker for newly derived blood vessels was CD34. While CD34-MVD demonstrated a significant association with tumorgrading and PSA follow-up, CD31- or vWF-MVD did not. CONCLUSION: CD34 is a suitable marker for the immunohistochemical visualization of microvessels in benign and malignant prostate tissue.

Aged↗

PubMedDate not supplied

Acoustic energy: a new transfection method for cancer of the prostate, cancer of the bladder and benign kidney cells.

BACKGROUND: Clinical use of gene therapy is limited by the poor efficacy and accuracy of intracellular DNA delivery. Known concepts of DNA transfection have not yet become clinical routine in the treatment of human disorders. We therefore focused on new transfection methods using different forms of acoustic energy as potentially safe and topographically applicable methods for gene delivery in the field of urology. MATERIALS AND METHODS: Three different cell lines (prostatic and urothelial cancer, benign kidney) were transfected by different forms of acoustic energy. The effect of several parameters of electromagnetic shock wave treatment (number and frequency of impulses, energy flow density and plasmid concentration) as well as focused ultrasound on the transfection rate was assessed in a standardized experimental setup. The transfection rate was measured through reporter genes (pEGFP) by FACScan. Transfection by lipofectamine and electroporation served as positive controls. RESULTS: All cell lines were transfectable by acoustic energy. Maximum transfection rate was achieved using focused ultrasound (49.5o%; 200 W, 500 ms, 200 microg/ml DNA). 31.3% of kidney cells were transfected by electromagnetic shock waves (1500 impulses, 200 microg/ml DNA, 0.5 mJ/mm2 energy density, 2 Hz). Plasmid strand breaks were identified as a limiting factor of the transfection rate. CONCLUSION: Transfection by acoustic energy, especially focused ultrasound, can be achieved at a high level in different cell lines. The possible topical application to urological organs and the low level of side-effects make acoustic energy a promising new gene therapy treatment option in urology.

Acoustics↗

PubMedDate not supplied

Identification of metastasis-associated genes in prostate cancer by genetic profiling of human prostate cancer cell lines.

OBJECTIVES: Prostate cancer (PCa) is a heterogeneous tumour entity with known interindividual differences in biological behaviour regarding tumour aggressiveness and metastatic potentiaL To date, the prediction of the metastatic status of patients with PCa has not been possible. To identify the molecular causes behind these differences, the gene expression profiles of two cell lines (LNCaP and LNCaP C4-2) with different metastatic potentials were examined using DNA microarray technology. MATERIALS AND METHODS: LNCaP and LNCaP C4-2 cells were cultured under standard conditions. RNA was isolated using Trizol extraction. After processing the total RNA according to the manufacturer's instructions, we performed Affymetrix GeneChip analysis with HG-U133A chips. Data analysis was performed using NetAffx, dChip, GenMAPP and OMIM software. RESULTS: After statistical evaluation of the raw data, we obtained a set of 158 differently expressed probe sets in the LNCaP and LNCaP C4-2 cells. The search for genes associated with proliferation, cell metabolism, growth factors, metastatic potential and tumour progression in this list revealed a number of 42 differently expressed probe sets. The comparison of this list of probe sets with the literature resulted in a list of 14 differently expressed genes which could well contribute to the metastatic potential and progression of PCa. Of these 14 genes only 6 (Cip1, IGF-1, NK4, CXCL 12, ILGF2R, RHOE) have already been associated with PCa, whereas the other 8 genes (FSTL-1, SOCS-2, Midkine, Thrombospondin 1, Secretory leukocyte protease inhibitor, Desmoglein 2, MLT 1, PTPRF) had not been previously related to PCa. CONCLUSION: DNA microarray technology offers the possibility of screening a large number of genes with regard to alterations in the expression level or mutations. In this study, we identified 14 genes that are most probably associated with the higher metastatic potential of LNCaP C4-2 cells as compared to LNCaP cells. Eight of these 14 genes are potential new molecular markers for assessing the metastatic potential of PCa, or may serve as therapeutical targets.

Apoptosis↗

PubMedDate not supplied

Prostate cancer therapy: standard management, new options and experimental approaches.

The management of prostate cancer is one of the core tasks for urologists today. This review focuses on therapeutic options for the curative and palliative treatment of prostate cancer. Within the area of urological competence, radical prostatectomy remains the standard procedure for the curative treatment of localised prostate cancer. Recently, interest in minimally-invasive procedures such as brachytherapy, focused ultrasound and cryotherapy has increased considerably. Established palliative treatment strategies include hormonal treatment schemes as the gold standard. Chemotherapeutic regimes are used for the hormone refractory disease. In addition to both the standard and new urological therapeutic options, promising experimental systemic strategies for the generalised disease are presented in this review.

Brachytherapy↗