DE102006015550B4 - Method and device for coupling an electric field into a physiological and electrically conductive medium - Google Patents

Abstract

contraption for coupling an electric field into a physiological one and electrically conductive Medium, characterized in that a first with alternating voltage (5) acted upon coil (4) made of an electrically conductive material is arranged on a ferromagnetic transformer core (3) and that a closed conductor loop (2) made of an electrically conductive Material as a second coil through the transformer core (3) passing to transformer-like Coupling of an electric field in the conductor loop (2) is provided, in which arranged the physiological medium is.

Description

The The invention relates to a method for coupling an electrical Field in a physiological and electrically conductive medium. Such methods are used, for example, for efficient production or for targeted influencing of cell cultures, microorganisms, Yeasts or mushrooms used.

The The invention further relates to a device which is for coupling of an electric field in a physiological and electrical conductive medium is trained.

such Devices are in addition to the generation of electric fields in ionic conductive Cell culture media or microorganism suspensions also for the treatment of extremities or other parts of the body used by humans or animals.

Farther Such devices play a role in tissue engineering. The method Tissue engineering is based on living cells of an organism outside of a body to cultivate, optionally with extracellular biological components or synthetic species, and the bioartificial regenerates or to rebuild constructs into an organism.

All in all Such devices thus find their use in bioprocess engineering in a broader sense as well as in reconstructive surgery and the Orthopedics.

in the State of the art are various methods and devices for the coupling of electromagnetic fields into physiological Media known.

Under physiological media should be understood in a broader sense media used or used in connection with living matter such as nutrient media or culture media. Furthermore, as physiological media as well Be considered living beings or parts thereof.

in the State of the art is known to be mechanically induced electrical For example, fields functional requirements for the bone structure convey what is as a starting point for research as well as for therapy is to be considered by electromagnetic fields. The coupling of electromagnetic Fields for stimulating bone healing or for treatment of Pseudoarthroses are well-known applications, and there are several Devices for implementing these methods.

Also it is known that fields generally have an influence on metabolic and transport operations of Have cells and a stimulation and inhibition is thereby possible.

The Interaction of electromagnetic fields with vital systems However, according to the state of science, this is done only by the electrical system Field. One knows the inductive coupling (IC), the capacitive coupling (CC) and the direct current (DC) and distinguishes the devices in principle according to the mode of action the coupling of the fields in connection with the type of used Field itself.

The most common method is inductive coupling. After DE 195 25 654 A1 For example, a method for mechanically influencing biological objects by means of magnetic particles in an inhomogeneous magnetic field is known. The device used for this purpose consists of a cylindrical coil with a non-closed ferrite core, between the ends of the ferrite core, the sample with the biological system and the magnetizable particles contained therein, the beads, between a flat pole and a pointed pole are arranged.

This Device and the associated method is particularly liable the disadvantage of creating an inhomogeneous magnetic field, resulting in the suitability of the device for reproducible scientific investigations greatly reduced.

The underlying method of stimulation of cells will also referred to as indirect coupling method, since the desired electrical Field through a time varying magnetic Field in the medium is caused.

After US 2004/0248291 A1 is a method for the cultivation of cells or cell culture media and cell cultures and a device known. The stimulation of cell growth occurs in vitro by means of a magnetic field. The device contains a cell culture container for the storage of the cell culture solution with the cells to be cultured and the spherical cell culture carriers to which the cells are adherent. Furthermore, the device includes a magnetic field generator for generating the magnetic field for the culture solution. The coil which generates the magnetic field is in this case arranged partially around the cell culture vessel or, alternatively, introduced into the cell culture medium in rod form. Further embodiments are to arrange a plurality of coils circular segment shape around the culture vessel around.

Mostly become in the inductive coupling so-called Helmholzanordnungen used by coils.

These consist of two current-carrying coils with parallel concentric Orientation. It is along and near the central axis of the Coils generated a homogeneous magnetic field that model well leaves. However, interaction with vital systems occurs indirectly the induced electric field, as already stated.

After DE 41 07 028 A1 A method is known for enhancing the production of extracellular enzymes in aerobic submucosal growth of fungal strains by electrostimulation, in which the enhancement of the production of extracellular enzymes is achieved by means of pulsed flows of Helmholtz coils.

adversely In the principle of inductive coupling is the indirect nature of Interaction, which ultimately leads to a hardly writable system. The Coupled energies are hard to determine and therefore the Dosage in an individual case a discretionary issue. The required power for generating a magnetic field in a Helmholzanordnung is also very high. At the same time resulting small effective electrical field strengths Thus, the efficiency of the treatment is low.

Farther disadvantageous is the size or the construction volume of the required devices.

As well is to be regarded as disadvantageous in the method of inductive coupling, that the fields are very difficult to model, which is especially true at basic research is a disadvantage of central importance. The reproducibility under otherwise comparable conditions is from the exact execution depending on the coil construction and thus hardly or only given under special conditions.

The Coupling of an electric field takes place through time-variable Magnetic fields. In electrically conductive tissue or nutrient medium thereby also generate eddy currents, which react on the magnetic field. The distribution of the electric fields is thus of the construction and the arrangement of coils as well as anatomy and conductivity dependent on the exposed tissue and therefore difficult to generalize. To estimate the induced electrical field strengths In addition to the magnetic flux density, the temporal change must also be considered be known of the magnetic field. Generally, the steeper the Flanks of the pulse are, even the induced field strength all the more is higher. Furthermore, it is difficult to determine the actual electric field strength in the Near the To determine and control cells. It is also problematic the duration of the induced electric fields. It exists thus significant differences in the shape, distribution and strength of the Fields, which ultimately includes, for example, in medical technology Use case endogenous electric fields only insufficiently Pulsed magnetic fields can be reproduced.

A Another method is the direct coupling of the current into the physiological medium.

After DE 102 52 773 A1 For example, a device and method for cell stimulation is known in which cell stimulation is performed by direct electrical stimulation without damage to the cells, cell-contactless, but with electrodes in the medium.

All Generally, electric fields in this method by the direct application of currents via electrodes cathodic with an increase and anodic with a decrease are coupled to the local pH and in addition gradients of oxygen concentration being constructed. An implantation of electrodes in organic Material also has an additional Risk of infection besides the disadvantages of electrochemical Decomposition or electrochemical degradation of components of the Cell culture medium or the cell culture itself.

These Effects arise in part from the fact that electrodes charge in the medium primarily by Faradaysche processes, since the for a charge required by a physiological response far exceeds that of achieved by loading and unloading the double-layer capacity can be.

Consequently The stimulation comes through direct coupling in the rarest make for use. For one, the probability of a direct damage by electroporation big and on the other hand leads the direct contact of the electrodes to redox processes in the physiological Environment that as an impairment to be considered.

Furthermore, the electrode material used for use in biological tissue must be made inert, and it should also be biocompatible. Can be used platinum and iridium. For the production of bioelectrodes platinum is therefore very often used, since it is very resistant to corrosion and is also well-suited due to low threshold potentials for electrical stimulation. An additional platinization of the surface, ie application of a rough noble metal layer, the Oberflä more than 1,000 times larger. As a result, the current density decreases and the electrochemical processes run longer in the reversible range. However, the use of platinum has the disadvantage that this metal is very expensive.

at the capacitive coupling over External capacitor plates, the electric field from the outside into a introduced between the capacitor plates lying system. Consequently In the case of capacitive coupling, the fields are covered by dielectrics, often combined several, coupled, their susceptibility often not is known. This is a modeling of the process and the Parameters difficult.

The Distribution of strength of the electric field and the treatable tissue are strong of the frequency and the passive electrical properties of the Tissue dependent.

By the capacitive charge processes become in contrast to direct coupling, no chemical changes in the medium, it can but only low charge densities are generated. For larger charge densities it comes to dielectric breakdown-like phenomena and for inserting Faraday reactions. The capacities of Capacitor plates are generally very small, so only that way transmit high frequencies can be which are above the physiologically justified frequency band.

The The object of the invention is a method and a device as possible lossless coupling of a - preferably homogeneous - electrical To provide field in physiological media.

The The object of the invention is achieved by a device for coupling of an electric field in a physiological and electrical conductive medium solved, which is characterized in particular by a first with AC voltage acted upon coil on a "transformer core", such as a ferromagnetic toroid, is arranged and that a closed Conductor loop as a second coil through the transformer core and the toroid going through to transformer-like Coupling of an electric field provided in the conductor loop is, in which the physiological medium is arranged. toroid is called each torus and in the topological sense a torus homeomorphs Body.

The Concept of the present invention is that an electrical Field through a transformer-like Provided or generated in the physiological medium coupling becomes. The generated electric field is in a preferred design the conductor loop formed by the medium of homogeneous nature and is used for positive or negative interaction with the vital components.

To A preferred embodiment of the invention is the conductor loop designed as an electrically non-conductive hose with an electrically conductive medium. The transformer-like Principle implies that the conductor loop at least once and dependent on the translation ratio too repeatedly penetrates the toroid to the electric field in the Medium of the conductor loop according to the transformer principle to bring.

Farther it is advantageous to use part of the conductor loop as bioreactor train. Also, it is advantageous to have the conductor loop in length and Cross-section to perform so that it is high impedance, with an electrical resistance greater than 1 kΩ. in the Bioreactors are components for tempering the medium as well options for monitoring and control of the relevant parameters of the physiological system intended. It is possible the electrical stimulation beneficial with other stimulating Components, such as a control of hydrostatic pressure or a device for deforming the cells with the purpose of mechanical Combine stimulation. Thus, especially in structures of the Tissue engineering one optimized influence on the generation of the constructs achieved become.

The spatial Separability of stimulation source and culture space will be beneficial used to the coupling of the electric field in a structure to realize that allows the vital component at the same time microscopy with stimulation (in situ microscopy).

According to one embodiment of the invention, the conductor loop has a length of 90 centimeters and is formed in the region of penetration of the toroid as a silicone tube with a length of 40 centimeters and a diameter of 10 millimeters and a model resistance R _M of 4.95 kΩ.

To an alternative embodiment of the device is the toroid designed such that limbs or extremities or Body Parts of humans or animals to form a conductor loop through the toroid can be passed are. With this training of the toroid are, for example, orthopedic treatments with the device according to the invention possible.

The object of the invention is further by a method for the treatment of an electric conductive cell culture medium dissolved with a device, which is characterized in that the cells or cell assemblies are introduced into an electrically conductive cell culture medium and that the cell culture medium positioned in the conductor loop and with an electric field E of 0.5 V / m at a generator frequency f of 75 Hz is applied.

In alternative embodiments of this method for treatment electrically conductive Cell culture media with a device according to the invention becomes an electrical Field of corresponding higher or higher lesser strength generated as 0.5 V / m, and the generator frequency is higher or lower than 75 Hz depending chosen by cell type. The Optima for field strength E and frequency f are according to the specifics of each physiological systems or cell types or microorganism species determine.

In Further embodiments, the generator, the AC voltage generated, which is applied to the first coil on the toroid, one superimposed from several frequencies Generate waveform that leads to temporally uneven field strengths in the Medium leads.

To an exemplary embodiment of the invention are called cells Saos-2 ATCC number HTB-85 Cells on glass slides used with approx. 12 mm diameter. After a further advantageous Embodiment of the method are used as cells microorganisms. As a cell culture medium for the cells or cell aggregates becomes McCoy's, for example 5a medium used.

in the Tissue engineering is an exemplary embodiment of this kind that allows specific cell carriers (scaffolds) that are (bio) chemical can be functionalized and with multipotent progenitor cells of a tissue to be replaced are colonized to influence purposefully. After an embodiment The invention is used for the purpose of forming a tissue engineering construct for the therapy of Bone defects a porous one carrier from a bioresorbable plastic, such. B. a knit polyhydroxybutyric (PHB) or polylactic acid combined with components of the extracellular matrix, such as collagen type I or chondroitin sulfate (CS). After colonization with autologous mesenchymal precursor cells from bone marrow and / or fatty tissue, the culture is perfused Bioreactors. In the culture medium, for. Dulbecc's modified Eagle Medium (DMEM) with autologous Serum becomes an electric field according to the invention essential Principle of transformer-like Incoupling produces that differentiation into osteoblastic cells favors.

• • Die erfindungsgemäße Vorrichtung ist einfacher im Aufbau und benötigt weniger Platz und Bauvolumen.
• • Die Einkopplung des elektrischen Feldes kann räumlich getrennt von anderen apparativen Komponenten oder dem Bioreaktor erfolgen.
• • Weiterhin besteht kein unmittelbarer Zusammenhang zwischen der Größe des zur Einkopplung genutzten Transformators und dem Volumen, in dem das homogene elektrische Feld vorliegt.
• • Von Vorteil ist ebenso, dass ein höherer Wirkungsgrad und damit eine geringere erforderliche Primärleistung für die Erzeugung des elektrischen Feldes anwendbar ist. Von besonderem Interesse ist dabei, dass durch die geringere Verlustleistung – im Vergleich zu Helmholzspulen – für Vorrichtungen nach dem erfindungsgemäßen Prinzip keine Wasserkühlung der Anordnung notwendig ist, da sich das physiologische Medium während der Behandlung mit dem elektrischen Feld nicht zu stark aufheizt.

The device according to the invention has the following advantages in particular over the Helmholtz coils predominantly used in the prior art and a coupling according to the inductive coupling principle:

• The device according to the invention is simpler in construction and requires less space and volume.
• • The coupling of the electric field can be spatially separated from other apparatus components or the bioreactor.
• • Furthermore, there is no direct correlation between the size of the transformer used for coupling and the volume in which the homogeneous electric field is present.
• • Another advantage is that a higher efficiency and thus a lower required primary power for the generation of the electric field is applicable. Of particular interest is that due to the lower power loss - compared to Helmholtz coils - for devices according to the principle of the invention no water cooling of the arrangement is necessary because the physiological medium does not heat up too much during treatment with the electric field.

Further Advantages over known methods are that in comparison to the methods No damaging electrodes required according to the direct coupling principle are and thus also the negative effects of the occurrence of electrochemical Reactions and reaction products are excluded. Farther are no sensitive dielectrically coated in comparison to the capacitive coupling Electrodes required in the medium. Besides, the transmission is a wide frequency band, especially of deep, physiological relevant frequencies compared to the capacitive coupling method, possible.

The electric field strength according to the principle of the invention is adjustable in a wide range, and it is in particular also high field strengths compared to the methods of capacitive coupling and inductive Coupling feasible.

Ultimately leads the consistent application of the transformer-like coupling to a much weaker Magnetic field as a side effect. It only creates the induced Current surrounding magnetic field, and there is no magnetic field in the nutrient medium, from which an electric field is induced. It only happens very much weak eddy currents on, and thus there is only a slight warming in the magnetic field Field.

Another advantage is that the electric Compared to inductive coupling, the field is easily calculable, which is important in terms of planning of trials or therapeutic treatment.

The electric field is over size spatial Areas homogeneous, which is a very special advantage is.

methodical The decisive advantage is that a largely pure Stimulation by an electric field, unlike mostly magnetic fields associated alternative principles of action is possible.

One Another advantage of the inventive device design is that the principle can be used for closed systems is because the conductor loop itself is closed and thus the requirement the microbiological sterility in applications in bioprocess engineering or medicine easy way becomes feasible.

Further Details, features and advantages of the invention will become apparent the following description of embodiments with reference on the associated Drawings. Show it:

1 : Device for coupling an electric field,

2 : Bioreactor with meandering channel and connected tube, both filled with physiological medium, to form a conductor loop,

3 : Bioreactor with meandering channel with circular channel widening for receiving cell culture carriers,

4 : Bioreactor with meandering channel with wells for receiving cell culture carriers.

In 1 a device according to the invention for coupling an electric field into a physiological and electrically conductive medium is shown.

Apparative core of the device is the toroid 3 which consists of a ferromagnetic material. The sink 4 is wound on this toroid and can with a primary alternating voltage 5 be charged. The thereby generated magnetic field in the toroid 3 leads to an induced voltage in the conductor loop 2 that the toroid 3 penetrates and is self-contained. The bioreactor 1 forms part of the conductor loop 2 and is equipped in the usual way with means for tempering and the like.

The sink 4 is in the embodiment with ten turns and the conductor loop 2 executed as a single turn.

Further embodiments are that the coil 4 one to one hundred turns.

Flows an alternating current 5 through the coil 4 , so in the closed ring with a conductive medium, a voltage is induced. Accordingly, the alternating electric field is created in the nutrient medium in the conductor loop 2 and it flows in this one stream. The resulting electric field is homogeneous and therefore well describable. The current in the nutrient medium within the conductor loop 2 surrounds itself with a magnetic field, which is however negligibly weak. Decisive effect is the emergence of the alternating electric field in the nutrient medium.

The bioreactor 1 has according to the embodiment according to 2 meandering channels with circular channel broadening 8th for the cell culture carriers 9 on and consists of a base plate 11 and a cover plate, not shown. The meandering channels 6 with the channel widenings 8th for the cell culture carriers 9 are in the base plate 11 incorporated.

The cover plate is made of a translucent material, allowing the observation of the processes in the bioreactor 1 is possible by means of a microscope, not shown.

The bioreactor 1 Also contains components, not shown, for temperature control, pH measurement and adjustment as well as options for sterile loading and unloading with sample volume and consists in the usual way of sterilizable material.

Furthermore, to ensure the homogeneity of the physiological medium and to stimulate the bioreactor 1 running processes means for moving the physiological medium in the form of a hose or diaphragm pump, for example, provided.

3 shows the base plate 11 a bioreactor 1 with meandering channel 6 , with circular channel extensions 8th for receiving cell culture carriers 9 Is provided.

The channel extensions 8th Although they are detrimental to the homogeneity of the field, but serve the safe spacing of the form-fitting in the channel broadening 8th held cell culture carrier 9 , The cell cultures are again on the cell culture carriers 9 placed. This will ensure that the cell culture carriers 9 also with moving physiology schem medium can not move to each other and a shearing of the adherent cell cultures from the cell culture carriers 9 by a relative movement of adjacent cell culture carriers 9 is excluded. Furthermore, by fixing the cell culture carrier 9 in the channel widenings 8th Microscopic examination of cell cultures is reproducible even over longer periods of time.

4 shows a bioreactor 1 with meandering channel 6 with depressions 10 for receiving cell culture carriers 9 , This alternative embodiment of the invention has a uniform as possible cross-section and thus leads to an improvement in the homogeneity of the electric field within the conductor loop 2 in the bioreactor 1 , The fixation of the cell culture carrier 9 is through depressions 10 in the base plate 11 of the bio-rector 1 within the meandering channel 6 achieved, which analogous to the design of the base plate 11 to 3 a positive fixation of the same is achieved. In contrast to the aforementioned embodiment but can the channel cross-section of the meandering channel 6 are chosen equal or approximately equal over the entire length of the channel, creating a more homogeneous electric field in the bioreactor 1 is reachable.

The bioreactors according to 3 and 4 have connecting pieces 7 for a hose on which the conductor loop 2 closes.

1

bioreactor

2

conductor loop

3

Transformer core, Torus, Toroid

4

Kitchen sink

5

AC signal

6

meandering channel

7

spigot

8th

channel extensions

9

Cell culture carriers

10

wells in the bottom of the bioreactor plate

11

baseplate

Claims (17)

Device for coupling an electric field into a physiological and electrically conductive medium, characterized in that a first one with alternating voltage ( 5 ) applied coil ( 4 ) of an electrically conductive material on a ferromagnetic transformer core ( 3 ) and that a closed conductor loop ( 2 ) of an electrically conductive material as a second coil through the transformer core ( 3 ) passing through the transformer-like coupling of an electric field in the conductor loop ( 2 ) is provided, in which the physiological medium is arranged. Device according to claim 1, characterized in that the transformer core ( 3 ) is designed as a toroid. Device according to claim 1 or 2, characterized in that the conductor loop ( 2 ) is designed as an electrically non-conductive hose with an electrically conductive medium. Device according to one of claims 1 to 3, characterized in that the conductor loop ( 2 ) several times the transformer core ( 3 ) penetrates. Device according to one of claims 1 to 4, characterized in that a part of the conductor loop ( 2 ) as bioreactor ( 1 ) is trained. Device according to one of claims 1 to 5, characterized in that the bioreactor ( 1 ) Has means for microscopic observation of the stimulated cells in situ. Device according to one of claims 1 to 6, characterized in that the bioreactor ( 1 ) from an electrically non-conductive base plate ( 11 ) and a cover plate is constructed. Device according to one of claims 1 to 7, characterized in that the bioreactor ( 1 ) Comprises means for acting on the physiological medium with intermittent hydrodynamic pressure. Device according to one of claims 1 to 8, characterized in that the bioreactor ( 1 ) Has means for deforming the cells for the purpose of mechanical stimulation. Device according to one of claims 1 to 9, characterized in that the conductor loop ( 2 ) in the bioreactor ( 1 ) as a meandering channel ( 6 ) is trained. Device according to one of claims 1 to 10, characterized in that the with AC voltage ( 5 ) applied coil ( 4 ) has one to one hundred turns. A method for treating an electrically conductive cell culture medium with a device according to one of claims 1 to 11, characterized in that a) cells or cell clusters are introduced or positioned in an electrically conductive cell culture medium and that b) the cell culture medium with the rows or the cell clusters in the conductor loop ( 2 ) and that the rows or cell assemblies c) by means of a transformer-like coupling d) with an electric field E of 0.5 V / m at e) a generator frequency f of 75 Hz are applied. Method according to claim 12, characterized in that that the alternating voltage consists of several frequencies with one superimposed Waveform with irregular waveform with weak field strengths is formed. Method according to claim 12 or 13, characterized that microorganisms are used as cells. Method according to one of claims 12 to 14, characterized that as cells multipotent progenitor cells on biocompatible Scaffolding used become. Method according to claim 15, characterized in that that mesenchymal progenitor cells differentiate into osteoblasts become. Method according to claim 15 or 16, characterized that as biocompatible scaffolds Polyhydroxybutyric acid embroidery (PHB) or polylactic acid combined with components of the extracellular matrix, especially collagen type I or chondroitin sulfate (CS) can be used.