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Iskalni niz: "avtor" (Damijan Miklavčič) .

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Cell size dynamics and viability of cells exposed to hypotonic treatment and electroporation for electrofusion optimization
Marko Ušaj, Katja Trontelj, Rosana Hudej, Maša Kandušer, Damijan Miklavčič, 2009, izvirni znanstveni članek

Povzetek: Background. Various electrofusion parameters have to be adjusted to obtain theoptimal electrofusion efficiency. Based on published data, good electrofusion conditions can be achieved with the hypotonic treatment. However, the duration of the hypotonic treatment before electroporation and buffer hypoosmolarity have to be adjusted in order to cause cell swelling, to avoid regulatory volume decrease and to preserve cell viability. The aims of our study were to determine cell size dynamics and viability of four different cell lines in hypotonic buffer and to study the influence of the electroporation on the selected cell line in hypotonic buffer. Materials and methods. Cell size dynamics of different cell lines exposed to hypotonic buffer and electroporation were analyzed by time-resolvedcell size measurements. The viability of hypotonically treated oržand electroporated cells was determined 24 h after the experiment by a modified crystal violet (CV) viability assay. Results. In our experimental conditions the hypotonic treatment at 100 mOsm was efficient for CHO, V79 and B16-F1 cell lines. The optimal duration of the treatment was between two and five minutes. On the other hand the same hypotonic treatment did not cause cell swelling of NS1 cells. Cell swelling was also observed after electroporation of B16-F1 in isotonic buffer and it was amplified when hypotonic buffer was used. In addition, the regulatory volume decrease was successfully inhibited with electroporation. Conclusions. Cell size dynamicsin hypotonic conditions should be studied for each cell line since they differ in their sensitivity to the hypotonic treatment. The inhibition of cell regulatory volume decrease by electroporation may be beneficial in achieving higher electrofusion efficiency. (Abstract truncated at 2000 characters)
Ključne besede: hypotonic treatment, cell swelling, regulatory volume decrease, cell size measurements, viability, electrofusion, electroporation
Objavljeno v DiRROS: 08.03.2024; Ogledov: 69; Prenosov: 27
.pdf Celotno besedilo (263,01 KB)

3.
Optimization of electrode position and electric pulse amplitude in electrochemotherapy
Anže Županič, Selma Čorović, Damijan Miklavčič, 2008, izvirni znanstveni članek

Povzetek: Background. In addition to the chemotherapeutic drug being present within the tumor during electric pulse delivery, successful electrochemotherapy requires the entire tumor volume to be subjected to a sufficiently high electric field,while the electric field in the surrounding healthy tissue is as low as possible to prevent damage. Both can be achieved with appropriate positioning of the electrodes and appropriate amplitude of electric pulses. Methods. We used 3D finite element numerical models and a genetic optimization algorithm to determine the optimum electrode configuration and optimum amplitude of electric pulses for treatment of three subcutaneous tumor models of different shapes and sizes and a realistic brain tumor model acquired from medical images. Results. In all four tumor cases, parallel needle electrode arrays were a better choice than hexagonal needle electrode arrays, since their utilization required less electric current and caused less healthy tissue damage. In addition, regardless of tumor geometry or needle electrode configuration, the optimum depth of electrode insertion was in all cases deeper than the deepest part of the tumor. Conclusions. Our optimization algorithm was able to determine the best electrode configuration in all four presented models and with further improvement it could be a useful tool in clinical electrochemotherapy treatment planning.
Objavljeno v DiRROS: 08.03.2024; Ogledov: 55; Prenosov: 16
.pdf Celotno besedilo (248,14 KB)

4.
Numerical modeling in electroporation-based biomedical applications
Nataša Pavšelj, Damijan Miklavčič, 2008, izvirni znanstveni članek

Povzetek: Background. Numerous experiments have to be performed before a biomedical application is put to practical use in clinical environment. As a complementary work to in vitro, in vivo and medical experiments, we can use analytical and numerical models to represent, as realistically as possible, real biological phenomena of, in our case, electroporation. In this way we canevaluate different electrical parameters in advance, such as pulse amplitude, duration, number of pulses, or different electrode geometries. Suchnumerical models can contribute significantly to the understanding of an experiment and treatment planning as well as to the design of new electroporation devices and electrodes. Methods. We used commercially available modeling software, based on finite element method. We constructed a model of a subcutaneous tumor during electrochemotherapy (EMAS) and a model ofskin during gene electrotransfer (COMSOL Multiphysics). Tissue-electrode geometries, pulse parameters and currentvoltage measurements from in vivo experiments were used to develop and validate the models. Results. To describeadequately our in vivo observations, a tissue conductivity increase during electroporation was included in our numerical models. The output currents of the models were compared to the currents and the voltages measuredduring in vivo experiments and a good agreement was obtained. Also, when comparing the voltages needed for a successful electropermeabilization assuggested by the models, to voltages applied in experiments and achieving a successful electrochemotherapy or in vivo gene electrotransfer, good agreementcan be observed. Conclusions. Modeling of electric current and electric field distribution during cell and tissue electroporation proves to be helpful in describing different aspects of the process and allowing us to design electrodes and electroporation protocols as a part of treatment planning.
Ključne besede: electroporation, gene electrotransfer, electrochemotherapy, subcutaneous tumor, finite-element method
Objavljeno v DiRROS: 07.03.2024; Ogledov: 63; Prenosov: 22
.pdf Celotno besedilo (549,62 KB)

5.
Electrochemotherapy of tumours
Gregor Serša, Maja Čemažar, Damijan Miklavčič, Zvonimir Rudolf, 2006, pregledni znanstveni članek

Povzetek: Electroehemotherapy consists of chemotherapy followed by local application of electrie pulses to the tumour to increase drug delivery into cells. Drug uptake can be inereased by electroporation for only those drugs whose transport through the plasma membrane is impeded. Among many drugs that have been tested so far, only bleomycin and cisplatin found their way from preclinical testing to clinical trials. In vitro studies demonstrated several fold inerease of their cytotoxicity after electroporation of cells. In vivo, electroporation of tumours after local or systemic administration of either ofthe drugs, i.e. electrochemotherapy, proved to be an effective antitumour treatment. In preclinical studies on several tumour models, electrochemotherapy either with bleomycin or cisplatin was elaborated and parameters for effective local tumour control were determined. In veterinary medicine, electrochemotherapy also proved to be effective in the treatment of primary tumours in cats, dogs and horses. In human clinical studies, electrochemotherapy was performed on the patients with progressive disease andaccessible tumour nodules of different malignancies. All clinical studies demonstrated that electrochemotherapy is an effective treatment for local tumour control in cancer patients.
Objavljeno v DiRROS: 15.02.2024; Ogledov: 113; Prenosov: 31
.pdf Celotno besedilo (225,92 KB)

6.
A web-application that extends functionality of medical device for tumor treatment by means of electrochemotherapy
Ivan Pavlović, Peter Kramar, Selma Čorović, David Cukjati, Damijan Miklavčič, 2004, izvirni znanstveni članek

Povzetek: Electrochemotherapy (ECT) is a novel method for efficient tumor treatment in clinical environment. It combines local drug delivery and application of shorthigh voltage pulses, which permeabilize the plasma membrane by electroporation. Drug can enter only the cells with permeabilzed membrane. Recently, medical device CliniporatorTM for controlled electroporation was developed. Here, we present a web-application that extends the functionality of this medical device. The aim of the application is to collect, store and toallow the analysis of every ECT application using this medical device. The application helps transferring data collected by devčce during the electroporation process to the central database, and enables filling of medical records through the web forms. The application is based on technologies ASP, HTML, Flash, JavaScript, XML and others. The application main advantages are easy and rapid data access, scalability and independence of client computer operating system as well as easy application debugging and upgrading.
Ključne besede: neoplasms- drug therapy, drug delivery systems, elektroporation instrumentation
Objavljeno v DiRROS: 07.02.2024; Ogledov: 126; Prenosov: 36
.pdf Celotno besedilo (133,39 KB)

7.
Tumor blood flow modifying effects of electrochemotherapy : a potential vascular targeted mechanism
Gregor Serša, Maja Čemažar, Damijan Miklavčič, 2003, izvirni znanstveni članek

Povzetek: Background. The aim of this study was to determine the tumor blood flow modifying, and potential vascular targeted effect of electrochemotherapy with bleomycin or cisplatin. Materials and methods. Electrochemotherapy was performed by application of short intense electric pulses to the tumors after systemic administration of bleomycin or cisplatin. Evaluated were antitumor effectiveness of electrochemotherapy by tumor measurement, tumor blood flow modifying effect by Patent blue staining technique, and sensitivity of endothelial and tumor cells to the drugs and electrochemotherapy by clonogenicity assay. Results. Electrochemotherapy was effective in treatment of SA-1 tumors in A/J mice resulting in substantial tumor growth delay and also tumor cures. Tumor blood flow reduction following electrochemotherapy correlated well with its antitumor effectiveness. Virtually complete shut downof the tumor blood flow was observed already at 24 h after electrochemotherapy with bleomycin whereas only 50% reduction was observed after electrochemotherapy with cisplatin. Sensitivity of human endothelial HMEC-1 cells to electrochemotherapy suggests a vascular targeted effect for electrochemotherapy in vivo with bleomycin as well as with cisplatin. Conclusion. These results show that in addition to direct electroporation of tumor cells, other vascular targeted mechanisms are involved in electrochemotherapy with bleomycin or cisplatin, potentially mediated by tumorblood flow reduction, and enhanced tumor cell death as a result of endothelial damage by electrochemotherapy.
Objavljeno v DiRROS: 06.02.2024; Ogledov: 102; Prenosov: 21
.pdf Celotno besedilo (205,34 KB)

8.
Cell electropermeabilization to small molecules in vitro : control by pulse parameters
Alenka Maček Lebar, Damijan Miklavčič, 2001, izvirni znanstveni članek

Povzetek: A systematic study concerning the role of the different electric field parameters (pulse number, duration and amplitude) on electropermeabilization of DC3F cells to small molecules (propidium iodide) and on cell viability is presented. Cell permeabilization and viability dependence on the pulse amplitude was determined by twenty different sets of electrical parameters. The number of pulses varied between 1 and 64 and pulse duration between 20 micros and 1 ms. The most important parameter was the pulse amplitude because at triggered the electropermeabilization process and the process of cell death. Either in the case of electropermeabilization as well as in the case ofcell viability experiments, the parameter Uso (the pulse amplitude leading to permeabilization or to the death of 50% of cell population) was not changedif the set of electrical parameters consisted of more than 16 pulses. This was independent of the pulse duration. The efficiency of permeabilizationwas enhanced by using of longer pulses. Such a systematic study of the influence of different electric field parameters on electropermeabilization and cell viability may serve as a base for optimization of the electropermeabilization conditions for different applications.
Objavljeno v DiRROS: 25.01.2024; Ogledov: 121; Prenosov: 32
.pdf Celotno besedilo (508,35 KB)

9.
First experience with a novel luminescence-based optical sensor for measurement of oxygenation in tumors
Tomaž Jarm, Hotimir Lešničar, Gregor Serša, Damijan Miklavčič, 2001, izvirni znanstveni članek

Povzetek: Background. The purpose of this preliminary study was to evaluate a novel luminescence-based fiber-optic sensor (OxyLite system) for the measurement of partial pressure of oxygen (pO2) in tumors and for the detection of changes inpO2 as a function of time. The new method was used simultaneously with the laser Doppler flowmetry method for the measurement of relative tissue perfusion. Materials and methods. Blood perfusion and pO2 were measured continuously via fiber-optic sensors inserted into SA-1 tumors in anesthetizedAžJ mice. The changes in blood flow and oxygenation of tumors wereinduced by transient changes of the parameters of anesthesia and by injection of a vasoactive drug hydralazine. Results. Both optical methods usedin the study successfully detected the induced changes in blood flow and pO2. The measurements of pO2 were well correlated with measurements of microcirculatory blood perfusion. In the majority of pO2 measurements, we observed an unexpected behavior of the signal during the stabilization processimmediately after the insertion of the probe into tumor. This behaviourof the pO2 signal was most probably caused by local tissue damage induced by the insertion of the probe. Conclusion. The novel luminescence-based optical oximetry can reliably detect local pO2 changes in tumors as a function of time but some aspects of prolonged pO2 measurement by this method require further investigation.
Ključne besede: sarcoma experimental, blood supply, laser-doppler flowmetry, oxygen, luminiscence
Objavljeno v DiRROS: 25.01.2024; Ogledov: 118; Prenosov: 34
.pdf Celotno besedilo (231,19 KB)

10.
Electroporator for in vitro cell permeabilization
Marko Puc, Karel Flisar, Stanislav Reberšek, Damijan Miklavčič, 2001, izvirni znanstveni članek

Povzetek: The use of high voltage electric pulse technology, electroporation, in cell biology, biotechnology and medicine has attracted an enormous interest. Electroporation is a transient phenomenon that increases the permeability of cell plasma membrane. In the state of high permeability, the plasma membrane allows small and large molecules to be introduced into the cytoplasm, althoughthe cell plasma membrane represents a considerable barrier for them inits normal state. The effectiveness of electroporation depends on many parameters that can be divided into the parameters of the electric field and the parameters that define the state of cells and their surrounding i.e, temperature, osmotic pressure, etc. In this article, we present a prototype electroporator GT-1 for in vitro electropermeabilization that we have developed. Our electroporator offers a vast flexibility of parameters and can generate high and low voltage pulses, of which the latter ones are used for electrophoretic transfer of charged molecules through permeabilized cell plasma membrane.
Ključne besede: electroporation instrumentation methods, cell membrane permeabilization
Objavljeno v DiRROS: 25.01.2024; Ogledov: 140; Prenosov: 36
.pdf Celotno besedilo (239,19 KB)

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