Method for determining at least one type and/or condition of cells and system

Abstract

The invention relates to a method for discriminating cells of a cellular structure, notably of a cellular tissue, comprising the steps consisting in determining (12) a frequency spectrum of the impedance of the cellular structure; defining (22) at least one model of the impedance of the cellular structure including a constant phase element (30); determining (44) the impedance of the constant phase element (30) which optimizes the correlation of each model of the impedance of the cellular structure with the spectrum; and deducing (66), from the impedance of the constant phase element (30) or from the impedances of the constant phase elements (30), an item of information on the cells of the cellular structure. The invention also relates to a system for implementing the method for discriminating cells of a cellular structure.

Claims

1. A method for discriminating cells of a cellular structure, the method comprising: operating a first device applied to the cellular structure within an animal to make in vivo measurements of impedance of the cellular structure contacted by the first device, wherein the first device that is applied to the cellular structure within the animal comprises a plurality of electrodes to contact the cellular structure and an internal electronic control to measure a discrete impedance spectrum of the cellular structure, wherein the internal electronic control measures the discrete impedance spectrum at least in part by applying to the cellular structure, via a first portion of the plurality of electrodes, an alternating current at multiple frequencies and calculating an impedance of the cellular structure at each of the multiple frequencies, the multiple frequencies not being continuous; and discriminating, with a second device, the cells of the cellular structure within the animal, wherein the discriminating comprises: computing, from the discrete impedance spectrum, at least one parameter that is representative of the cellular structure contacted by the plurality of electrodes; comparing the at least one parameter that is representative of the cellular structure to a plurality of reference values; and determining based at least on a result of the comparing, one or more items of information regarding the cellular structure.

2. The method of claim 1, wherein operating the first device to make in vivo measurements of the impedance of the cellular structure comprises operating a surgical device within the animal to make the in vivo measurements of the impedance of the cellular structure, wherein a portion of the surgical device that is disposed within the animal includes the internal electronic control.

3. The method of claim 2, wherein the surgical device is a guidewire or a catheter.

4. The method of claim 1, wherein determining the one or more items of information regarding the cellular structure comprises determining, based at least on the result of the comparing, a cellular composition of the cellular structure.

5. The method of claim 4, wherein determining the cellular composition of the cellular structure comprises determining, based at least on the result of the comparing, one or more types of cells present in the cellular structure.

6. The method of claim 4, wherein determining the cellular composition of the cellular structure comprises determining, based at least on the result of the comparing, one or more conditions of cells present in the cellular structure.

7. The method of claim 1, wherein computing the at least one parameter from the discrete impedance spectrum comprises computing an effective capacitance of the cellular structure from the discrete impedance spectrum.

8. The method of claim 1, wherein computing the at least one parameter from the discrete impedance spectrum comprises computing, from the discrete impedance spectrum, an effective capacitance of the cellular structure and at least one other parameter of the cellular structure.

9. The method of claim 1, wherein computing the at least one parameter from the discrete impedance spectrum comprises identifying, from the discrete impedance spectrum, an equivalent circuit model for the cellular structure, the equivalent circuit model including at least one constant phase element.

10. The method of claim 1, further comprising receiving the discrete impedance spectrum from the first device at the second device while the first device is disposed at least partially within the animal.

11. The method of claim 1, wherein: the plurality of reference values are stored in a data structure together with items of information; comparing the at least one parameter to the plurality of reference values comprises identifying at least one reference value, of the plurality of reference values, that is a match to the at least one parameter; and determining the one or more items of information regarding the cellular structure based at least on a result of the comparing comprises retrieving from the data structure one or more items of information associated with the at least one reference value identified as a match.

12. The method of claim 11, where the plurality of reference values are values determined from testing performed on one or more tissues of known compositions, in known media, and with known test conditions.

13. A system for discriminating cells of a cellular structure, the system comprising: a first surgical device for which at least a part is arranged to be disposed within an animal to make in vivo measurements of impedance of the cellular structure within the animal that is contacted by the part, the part of the first surgical device arranged to be disposed within the animal comprising a plurality of electrodes to contact the cellular structure and an internal electronic control to measure a discrete impedance spectrum of the cellular structure, wherein the internal electronic control measures the discrete impedance spectrum at least in part by applying to the cellular structure, via a first portion of the plurality of electrodes, an alternating current at multiple frequencies and calculating an impedance of the cellular structure at each of the multiple frequencies, the multiple frequencies not being continuous; and a second device, arranged to be disposed outside of the animal, to receive the discrete impedance spectrum from the surgical device and to discriminate the cells of the cellular structure within the animal, wherein the discriminating comprises: computing, from the discrete impedance spectrum, at least one parameter that is representative of the cellular structure contacted by the plurality of electrodes; comparing the at least one parameter that is representative of the cellular structure to a plurality of reference values; and determining, based at least on a result of the comparing, one or more items of information regarding the cellular structure.

14. The system of claim 13, wherein the second device is arranged to receive the discrete impedance spectrum from the surgical device at least partially via a wired connection.

15. The system of claim 13, wherein the surgical device is a guidewire, a catheter, or a probe.

16. The system of claim 13, wherein determining the one or more items of information regarding the cellular structure comprises determining, based at least on the result of the comparing, a cellular composition of the cellular structure and/or a number of cellular layers of the cellular structure.

17. At least one non-transitory computer-readable storage medium having encoded thereon executable instructions that, when executed by at least one processor, cause the at least one processor to carry out a method for discriminating cells of a tissue within an animal, the method comprising: receiving, from a first surgical device disposed at least partially within the animal to make in vivo measurements of impedance of the tissue within the animal contacted by the first surgical device, at a second device, a discrete impedance spectrum of the tissue, the discrete impedance spectrum indicating impedance of the tissue at multiple frequencies, the multiple frequencies not being continuous; and discriminating, with the second device, the cells of the tissue within the animal, wherein the discriminating comprises: computing, from the discrete impedance spectrum, at least one parameter that is representative of the tissue contacted by the first surgical device; comparing the at least one parameter that is representative of the tissue to a plurality of reference values for a plurality of known tissue compositions; and determining, based at least on a result of the comparing, a composition of the tissue.

18. The at least one non-transitory computer-readable storage medium of claim 17, wherein receiving the discrete impedance spectrum from the surgical device comprises receiving the discrete impedance spectrum from a guidewire, a catheter, or a probe.

19. The at least one non-transitory computer-readable storage medium of claim 17, wherein receiving the discrete impedance spectrum from the surgical device comprises receiving the discrete impedance spectrum from a surgical device for which at least a part is arranged to be disposed within the animal to make in vivo measurements of impedance of a cellular structure within the animal that is contacted by the part, the part of the surgical device arranged to be disposed within the animal comprising a plurality of electrodes to contact the cellular structure and an internal electronic control to measure a discrete impedance spectrum of the cellular structure, wherein the internal electronic control measures the discrete impedance spectrum at least in part by applying to the cellular structure, via a first portion of the plurality of electrodes, an alternating current at multiple frequencies and calculating an impedance of the cellular structure at each of the multiple frequencies, the multiple frequencies not being continuous or pseudo-continuous.

20. The method of claim 1, wherein the multiple frequencies are not pseudo-continuous.

21. The system of claim 13, wherein the multiple frequencies are not pseudo-continuous.

22. The at least one non-transitory computer-readable storage medium of claim 17, wherein the multiple frequencies are not pseudo-continuous.

Description

(1) Other features and advantages of various embodiments will become apparent on reading the following detailed description, said description referring to the attached drawings in which:

(2) FIG. 1 is a flow diagram of an exemplary method for discriminating cells in a tissue;

(3) FIG. 2 is a flow diagram of an exemplary method for determining, from a frequency spectrum, the impedance of a tissue;

(4) FIG. 3 represents an exemplary frequency spectrum of the modulus of the impedance of a cell tissue;

(5) FIGS. 4 to 7 illustrate exemplary models of the impedance of the tissue, that may be implemented in the method of FIG. 1, including a constant phase element;

(6) FIG. 8 shows an example, in diagram form, of effective capacitances of cellular structures determined by means of the method of FIG. 1;

(7) FIG. 9 illustrates an exemplary system for implementing the method of FIG. 1;

(8) FIGS. 10A and 10B show amplitude and phase spectra for experimental data;

(9) FIGS. 11A and 11B, and 12A to 12F show various parameters distributions;

(10) FIGS. 13 to 15 show distributions of values representative of effective capacitance for different cell types;

(11) FIGS. 16 and 17 show examples of systems made in accordance with the present invention.

(12) In some embodiments, a method allows for a discrimination of cells of a cellular structure, notably of a cell tissue. Discrimination should be understood here to include the possibility, given by this method, of distinguishing cells, for example of determining the type of cells of the tissue. More generally, the discrimination made possible by the method consists in determining at least one item of information relating to cells in a tested tissue. Examples of information items that may be determined by virtue of this method are given later.

(13) The cell discrimination method 10, as illustrated schematically in FIG. 1, comprises a first step 12 of determining a frequency spectrum of the impedance of a cellular structure to be tested. Hereinbelow, the structure of a tissue of cells will be taken as example of cellular structure.

(14) Spectrum should be understood here to include a set of pairs of values of the impedance of the tissue, the latter being able to be complex, and of the corresponding frequency. This spectrum may thus be discrete and comprise only a finite number of pairs. These pairs may notably be separated by several Hz, even by several tens of Hz, even by several hundreds of Hz. However, preferably, the spectrum determined in this step is continuous, pseudo-continuous or discretized, over a frequency band. Pseudo-continuous should be understood to mean that the spectrum is determined for successive frequencies separated by 100 Hz or less, preferably by 10 Hz or less, preferably even by 1 Hz or less. The frequency band over which the impedance of the tissue is determined extends, for example, from 10 kHz, preferably 100 kHz. In effect, at low frequencies, the membrane of the cells of the tissue acts as an electrical insulator, so that the impedance is very high and, above all, varies little. Moreover, the frequency band over which the impedance of the tissue is determined extends, for example, up to 100 MHz, preferably 1 MHz. In effect, at high frequencies, the wall of the cells that makes up the tissues become transparent from an electrical point of view. The measured impedance is therefore no longer representative of the cell wall. This spectrum may be a frequency spectrum of the real part and/or of the imaginary part and/or of the modulus and/or of the phase of the complex impedance of the cellular structure.

(15) This first step 12 of determination of a frequency spectrum of the impedance of the tissue may notably be performed as described hereinbelow.

(16) First of all, during a step 14, two, preferably three, even more preferably four electrodes are placed in contact with the tissue to be tested, the electrodes being linked to an alternating current generator. The measurement with four electrodes is preferred because it makes it possible to implement two electrodes to pass the current into the tissue to be tested and to measure the potential difference between the other two electrodes. This makes it possible to improve the accuracy of the measurement. Then, during a step 16, an alternating current is applied between the electrodes implanted in the tissue. Then, by varying the frequency of the current applied during a step 18, the corresponding voltage is measured, at the terminals of the electrodes for different frequencies. Finally, during a step 20, the ratio between the voltage measured and the current applied is calculated, for each of the frequencies for which the measurement has been performed. This ratio gives the impedance of the tissue tested, as a function of the measurement frequency. The calculated ratios make it possible to define a frequency spectrum of the impedance of the tissue.

(17) When the spectrum is continuous or pseudo-continuous, it may be represented as illustrated in FIG. 3, in the form of a curve giving, in this particular case, the modulus of the impedance of the tissue as a function of the frequency, the latter being plotted according to a logarithm scale. It should be noted here that a logarithmic scale is used on the x axis.

(18) In a step 22 of the discrimination method 10, different models of the impedance of the tissue, that is to say different electrical circuits that may model the tissue, are then chosen. Here, models are chosen that include a constant phase element, and not a capacitance. In effect, it has been found that a constant phase element models more realistically the behaviour of the tissue than a capacitance.

(19) A constant phase element (or CPE) has an impedance Z.sub.CPE of the form:

(20) $\begin{matrix} Z_{CPE} = \frac{1}{{(j)}^{} Q_{0}} & [1] \end{matrix}$

(21) in which: j is the square root of 1 (j.sup.2=1); is the specific pulsing of the current (=2f, in which f is the frequency of the current); Q.sub.0 is a real parameter of the constant phase element, also referred to as pseudo-capacitance; and is another real parameter of the constant phase element, lying between 0 and 1, such that the phase .sub.CPE of the constant phase element is equal to /2.

(22) Hereinafter in the description, a constant phase element whose impedance is given by the equation [1] above is chosen by way of example.

(23) The models of the impedance of the tissue may notably be chosen from those described hereinbelow, with respect to FIGS. 4 to 7. Obviously, the simpler the model, the simpler the calculations. However, a complex model may better correlate to the spectrum of the impedance obtained by the measurement and therefore give more accurate results.

(24) According to a first model 24 illustrated in FIG. 4, the impedance of the cell tissue is modelled by a first resistance 26 mounted in series with a parallel connection 28 of a constant phase element 30 and of a second resistance 32.

(25) In this case, the total resistance Z.sub.tot of the cell tissue is of the form:

(26) $\begin{matrix} Z_{tot} = R_{1} + \frac{R_{2}}{1 + {(j)}^{} Q_{0} R_{2}}, & [3] \end{matrix}$
in which: Z.sub.tot is the total impedance of the first model 24 representing the cell tissue; R1 and R2 are the resistance values of the first 26 and second 32 resistances.

(27) Such a model describes particularly well a tissue covering measurement electrodes, like a set of individual parallel mountings, each individual mounting being made up of an individual resistance in series with a parallel mounting of an individual resistance and of an individual capacitance. Such a mounting may make it possible to model a distribution of the time constant over all of the surface of the measurement electrodes, according to different circuits in parallel whose parameters may be different, each of these circuits in parallel representing a cell of the tissue. Thus, the fact that the cells of the tissue may exhibit different electrical properties, notably a different resistance and/or capacitance, may be modelled.

(28) A second model 34, illustrated in FIG. 5, complements the model 24 of FIG. 4, by the series mounting of a second constant phase element 36. The impedance Z.sub.CPE,2 of this second constant phase element 36 may also be chosen to be of the form:

(29) 0 $\begin{matrix} Z_{CPE, 2} = \frac{1}{{(j)}^{} Q_{1}}, & [4] \end{matrix}$
in which: is a real parameter lying between 0 and 1, such that the constant phase of this second constant phase element is equal to /2; and Q.sub.1 is a pseudo-capacitante (real number) of the constant phase element, also referred to as Q.sub.d1 for double layer pseudo-capacitance.

(30) The total impedance Z.sub.tot of the tissue according to this second model 34 is therefore given by the following equation:

(31) $\begin{matrix} Z_{tot} = \frac{1}{{(j)}^{} Q_{1}} + R_{1} + \frac{R_{2}}{1 + {(j Q_{0})}^{} R_{2}} . & [5] \end{matrix}$

(32) A variant 34 of the second model 34 is shown in FIG. 5A, and differs from the model of FIG. 5 by the addition of a capacitance C in parallel with the circuit of FIG. 5, for a better fit of the impedance curve at high frequencies.

(33) A third model 38, illustrated in FIG. 6, corresponds to the model of FIG. 4, mounted in parallel with a third resistance 40, of resistance R.sub.3. In this case, the total impedance Z.sub.tot of the tissue is given by the equation:

(34) $\begin{matrix} \frac{1}{Z_{tot}} = \frac{1}{R_{3}} + \frac{1}{R_{1} + \frac{R_{2}}{1 + {(j Q_{0})}^{} R_{2}}} . & [6] \end{matrix}$

(35) Finally, a fourth exemplary model 42 is illustrated in FIG. 7. This model 42 comprises, as illustrated, a first resistance 26, mounted in parallel with a series mounting of a constant phase element 30 and of a second resistance 32.

(36) The total impedance Z.sub.tot of the tissue is given, for this model 42, by the equation:

(37) $\begin{matrix} \frac{1}{Z_{tot}} = \frac{1}{R_{1}} + \frac{R_{2}}{1 + {(j Q_{0})}^{} R_{2}} & [7] \end{matrix}$

(38) The discrimination method then continues with a step 44, during which, for each model chosen in step 22, the impedance of the constant phase element 30 and all other components of the model are determined so that the impedance of the model matches to some extent the spectrum determined in step 12.

(39) This step of improving the matching of the model of the impedance of the tissue with the spectrum determined in the step 12 may be implemented by any optimization method known by those skilled in the art. By way of example, the least squares method may be implemented, which allows for a practical and relatively simple implementation of this step 44.

(40) An intermediate step 46 of the discrimination method 10 may then be provided. This step 46 consists in determining the model which seems to improve the matching between the model and the measured impedance. This model may for example be that which minimizes the standard deviation with the measured spectrum. Hereinafter in the description, the case in which the model 24 is retained as that correlating best to the measured spectrum of the impedance of the tissue is assumed.

(41) During a step 48, an effective capacitance of the cell tissue is deduced from the parameters of the impedance of the constant phase element and from the corresponding model.

(42) Theoretically, this effective capacitance is representative of a set of individual capacitances of elements of the cell structure. The effective capacitance is representative of distributed local capacitances of elements of the cell structure. These elements of the cell structure may notably be all or some of the nuclei of the cells of the cellular structure and also other parts of the cells such as the golgi apparatus, vesicles, mitochondrion, lysosome and other elements which may play a role in membrane interaction. The effective capacitance may also be influenced by the geometry of cells and the space between cells. The effective capacitance is a model which allows for a representation of the electrical membrane behaviour of a part or of all of a cellular structure. This model makes it possible to relevantly discriminate the cells. It differs from the membrane capacitance at least for the reason that it does not take for value the resultant capacitance of the electrical measurement but rather is given by a distributed model that is equivalent to a distribution of local capacitances.

(43) More practically, this effective capacitance is determined by identifying the impedance of the chosen cellular structure with a model comprising individual parallel mountings, each individual mounting comprising at least one individual resistance and one individual capacitance. Each mounting may notably comprise, preferably consist of, a first individual resistance in series with a parallel mounting of an individual capacitance with a second individual resistance. These individual mountings aim to model the behaviour of each cell of the cellular structure.

(44) In the case of the model 24 (or 34 or 34), the determination of the effective capacitance may notably be performed as follows. The impedance of the model 24 with a constant phase element is compared with the impedance of an equivalent or identical model, in which the constant phase element is replaced by an effective capacitance. The calculation, strictly speaking, of the effective capacitance may then be performed by comparing the real part and/or the imaginary part and/or the phase and/or the modulus of the impedance of the model chosen for the cellular structure with a constant phase element with the identical model in which the constant phase element is replaced by an effective capacitance.

(45) In the case of the model 24 (or 34 or 34), for example, by introducing a time constant

(46) $_{0} = C_{eff} \frac{R_{1} R_{2}}{R_{1} + R_{2}}$
into the equation of the admittance of the model 24, directly deduced from the equation [3], the equation [8] below is obtained:

(47) $\begin{matrix} Y_{tot} = \frac{1}{R_{1}} [1 - \frac{R_{2}}{R_{1} + R_{2}} {(1 + \frac{R_{1} R_{2}}{R_{1} + R_{2}} {Q_{0} (j)}^{})}^{- 1}] = \frac{1}{R_{1}} [1 - \frac{R_{2}}{R_{1} + R_{2}} {(1 + {(j_{0})}^{})}^{- 1}], & [8] \end{matrix}$

(48) from which a formula for the effective capacitance may be deduced, in the form:

(49) $\begin{matrix} C_{eff} = Q_{0}^{1 /} {(\frac{1}{R_{1}} + \frac{1}{R_{2}})}^{(- 1) /} & [9] \end{matrix}$

(50) In the case where another model of impedance of the cellular structure with a constant phase element is chosen, it is possible to determine a corresponding equation of the effective capacitance. To do this, it is sufficient to calculate the impedances R.sub.1, R.sub.2, Z.sub.CPE and Z.sub.CPE,2, if appropriate, of the model 24 or 34 or 34, as a function of the parameters of the chosen model, for the model 24 or 34 or 34 to be electrically equivalent to the model of the impedance of the chosen cellular structure. The effective capacitance may then be calculated by replacing R.sub.1, R.sub.2, Z.sub.0 and with the corresponding values, expressed as a function of the parameters of the chosen model.

(51) The cell discrimination method 10 then continues with a step 66 of deduction of an item of information on the cells of the tissue, from the effective capacitance determined previously.

(52) This deduction may be made by comparing the value of the effective capacitance determined in the step 48 with pre-established effective capacitance values. The pre-established effective capacitance values may have been obtained during tests performed on tissues of known compositions (e.g., types of cells and/or conditions of cells), in known media, and with known test conditions. The pre-established values may be grouped together in a database (or other form of data structure or data storage) of effective capacitance values, grouping together the effective capacitances measured for different types of cells and/or different conditions of different cells and/or in different test conditions. The effective capacitance value may be compared to a database of effective capacitances of cell type and condition susceptible to be found in the present measurement.

(53) For the comparison, the effective capacitance Ceff may be used together with other parameters. The comparison may not be an exact match and includes the determination whether the effective capacitance value falls or not within a pre-determined range.

(54) It is thus possible to discriminate the cells of the tissue, that is to say to determine at least one of the following items of information: the type of cells in the tissue; the composition of the tissue, notably if the latter is composed of different types of cells or of cells in different conditions; the number of layers of cells present in the tissue; and/or the condition of the cells, notably if the cells are in a healthy condition, in an inflamed condition, in a degenerated condition, notably if there are one or more cancerous cells, in an infected condition or if they are differentiated.

(55) As an example, FIG. 8 represents, in diagram form, the effective capacitances 68, 70, 72, 74 determined in the context of a test conducted according to the method described previously.

(56) In the context of a test, cells were cultivated until the confluence of the cells was obtained. In the case of the exemplary test which was conducted, two days of culture were required in an incubator at 37 C. and 5% CO.sub.2, to obtain, by confluence, the tissues to be tested. The determination of the spectrum of the impedance of the different tissues to be tested was performed using an impedance spectroscopy system. The spectrum was determined between 1 kHz and 10 MHz, by applying an alternating voltage estimated to be fairly low so as not to electrically excite the cells being studied, but sufficient to have correct measurements. In the example of the test conducted, an amplitude of 20 mV of the alternating voltage was retained.

(57) The effective capacitance 68 is that of the test medium, static, alone. This test medium is a cell culture medium. The effective capacitance 70 is that of bovine aortic endothelial cells (BAEC). The effective capacitance 72 is that of bovine aortic smooth muscle cells (BAOSMC). Finally, the effective capacitance 74 is that of blood platelets (or thrombocytes). As this diagram shows, the effective capacitances of the different types of cells exhibit values clearly different from one another, which makes it possible to effectively distinguish between the different types of cells with accuracy, without risk of confusion.

(58) Thus, one advantage of the discrimination method of some embodiments described above is that it allows for the discrimination of cells in a cellular structure, notably in a confluent and single-layer cellular structure, covering the electrodes, from a simple measurement of a frequency spectrum of an impedance of the structure to be tested. The results obtained are accurate. There is no need to proceed with a normalization of the measured impedance, nor to proceed with a reference measurement in the absence of any sample to be tested. The method may thus be implemented in vivo, that is to say without the need for prior sampling of cells or of a cellular structure to be tested.

(59) It should be noted, in the case where an effective capacitance is determined, a single value for effective capacitance, at one time, may be sufficient to discriminate the cells of the tissue. This is in contrast with other techniques performing an analysis of a capacitance that require multiple determinations of capacitance over time. In these embodiments, the parameters of the chosen model of the impedance of the cellular structure to be tested may also be compared to pre-established values to specify the result of the comparison of the effective capacitance. For example, when the cells are inflamed, the junction between the cells is looser. The resistance at low frequencythat is to say the resistance 32 of the model 24 for exampleis then lower, compared to healthy cells. A comparison of the value of this resistance with a value pre-established for healthy, non-inflamed cells may then make it possible to determine the inflamed condition of these cells.

(60) It should also be noted that other parameters of a model of impedance, aside from effective capacitance, may be considered to discriminate the cells. In some embodiments, these other parameters may also make it possible to determine additional items of information on the cellular structure tested. Thus, for example, R.sub.2 or the sum R.sub.1+R.sub.2 of the resistances 26, 32 of the model 24 may be considered to determine the thickness of the cellular structure. To do this, in some embodiments the values R.sub.2, and possibly R.sub.1, are determined, notably concomitantly with the determination of the impedance of the constant phase element, so as to optimize the correlation of the model 24 with the measured impedance spectrum. The value R.sub.2 or the sum R.sub.1+R.sub.2 may then be compared to corresponding values, predetermined in known conditions, for example in vitro. These predetermined values may notably be stored in database (or another data structure) form.

(61) As conditioned previously, the method may easily be implemented in the context of devices that may be implanted in the human body or applied to the human body.

(62) By way of example, FIG. 9 illustrates an example 100 of a system for implementing a method of some embodiments as described previously.

(63) The system 100 essentially comprises means 102 for measuring the impedance of a cellular structure 104, here a single-layer tissue of confluent cells, dipped in a medium 105, for example blood, and an electronic control unit 106, linked to the measurement means 102, to implement the method and discriminate the cells of the cellular structure 104 as a function of the measured impedance.

(64) The measurement means 102 here comprise an electrical generator 108 of alternating current, linked to two electrodes 110, 112 in contact with the cellular structure 104. The measurement means 102 also comprise a device 114 for determining the intensity passing through the cellular structure 104, linked to said cellular structure 104 by two electrodes 116, 118 in contact with the cellular structure 104. The electronic control unit 106 is linked to the electrical generator 108 and to the intensity measurement device 114, in order to be able to determine the impedance of the cellular structure 104, for example from the measurement of the voltage and of the intensity at the terminals of the electrodes 110, 112, 116, 118.

(65) The electrodes 110, 112, 116, 118 consist of an electrically conductive material, such as gold for example.

(66) Here, advantageously, the measurement means 102 further comprise a medical device 120 that may be implanted in the human body, here a stent 120, or that may be applied to the human body. In this case, the electrodes 110, 112, 116, 118, the alternating voltage generator and the intensity measurement device may be fixed onto this medical device. The medical device is for example as described in the application FR3026631 A1 MEDICAL DEVICE PROVIDED WITH SENSORS HAVING VARIABLE IMPEDANCE filed on 2014 Oct. 3, the entire contents of which, and in particular the discussion of implantable medical devices including measurement devices, are incorporated herein by reference.

(67) In this case, the alternating electrical generator 108 may include an armature, such as the body of the medical device or an antenna electrically insulated from the body of the medical device, adapted to emit an electrical current under the effect of an electromagnetic field emitted by an interrogation unit external to the stent 120. The electrodes may then form a sensor with variable impedance, the impedance of which varies as a function of the cellular structure which covers them. Finally, the electronic control unit may receive an item of information relating to the impedance between the electrodes, notably by emission of a magnetic field by an antenna fixed onto the body of the implantable medical device 120.

(68) The stent 120 may thus make it possible to check the correct progress of the healing of the endothelium, after the stent 120 has been fitted. In effect, such a stent 120, in cooperation with the electronic control unit, makes it possible to determine, by implementing the method of FIG. 1, whether the cellular structure which is formed on the surface of the endothelium essentially comprises healthy endothelial cells, inflamed endothelial cells, smooth muscle cells and/or platelets.

(69) The invention is not limited to the examples described hereinabove and numerous variants are possible, while within the scope of the definition given by the attached claims.

(70) Thus, for example, it is possible to choose a single model of the impedance of the tissue in the step 22. In this case, it is not necessary to carry out the optimization for a number of models. The method is therefore simpler and faster to implement in this case. It is notably possible to proceed in this way when a model is considered as more relevant.

(71) Moreover, in the examples described, the discrimination of the cells is based essentially on the calculated effective capacitance and on its comparison with pre-established values. As a variant, however, it is possible to proceed with the discrimination of the cells from parameters of the chosen model of the impedance of the cellular structure. However, it seems that the comparison of just the value of the effective capacitance is both simple and allows for a reliable discrimination of the cells.

(72) FIG. 16 shows an example of a system 300 made in accordance with the present invention. This system comprises a measurement module 301 with may be part of an implanted device, for example a stent, or of a device for in vitro cultivation of cells.

(73) The measurement module comprises at least two electrodes and may be as described above with reference to FIG. 9.

(74) The system 300 also comprises an internal processing unit 302 that is configured for example to generate an impedance spectrum from data from the measurement module.

(75) The system 300 may comprise an emitter 303 to wirelessly transmit data (the data from the measurement module 301 and/or the impedance spectrum determined by the internal processing unit 302) to a receiver 304, which may be external to the body in case the measurements take place in vivo. The transmission may take place under any wireless protocol such as RFID, NFC, Bluetooth, Wifi, either radio or Infrared, inter alia. In some embodiments, the transmission may include transmission via one or more wired and/or wireless local and/or wide-area networks, including the Internet.

(76) The system 300 may comprise an external processing unit 305 to compute the impedance spectrum (in the case of receiving from the emitter 303 the data from the measurement module 301) and/or the various parameters and effective capacitance C.sub.eff based on the received data and display means 306 such as a LCD screen to display information relating to the type and/or condition of cells determined based upon comparison of a value representative of C.sub.eff with reference data. To determine the various parameters and effective capacitance, the external processing unit 305 may be configured with information regarding one or more equivalent circuit models for an impedance, and determine the parameters of at least one of the model(s), such as in the manner discussed above. The external processing unit 305 may also be configured to select one of the models, following determination of the parameters of the model(s), as a model from which to determine the effective capacitance, as discussed above. The external processing unit may make the selection based on a degree of fit between the equivalent circuit model and the impedance spectrum. The system may provide, based on the at least one type and/or condition of cells thus identified, information representative of an evolution of a healing process, for example, information regarding a current status of an area in which (e.g., tissue to which) a procedure was performed (including positioning of an implant such as a stent) and/or provide information regarding a change over time in the status of the area that may be reflective of a response to the procedure in the area, such as a healing or scarring response.

(77) The external processing unit may be a special-purpose device that includes specialized hardware such as an ASIC, EEPROM, or other component specially configured to perform the operations of the external processing unit described above. In other embodiments, the external processing unit may be a general-purpose device such as a laptop or desktop personal computer, a server, a smart/mobile phone, a personal digital assistant, a tablet computer, or other computing device including mobile computing devices. In the case that the external processing unit is implemented with a general-purpose device, the general-purpose device may include one or more processors and a non-transitory computer-readable storage medium (e.g., an instruction register, an on-chip cache, a memory, a hard drive, a removable medium such as an optical medium) having encoded thereon instructions for execution by the processor(s), where the instructions cause the processor to carry out the operations described above as performed by the external processing unit. The internal processing unit may, in some embodiments, be any appropriate IC chip or other hardware component with processing capabilities. The external and internal processing units may be located proximate to one another (e.g., within a same room, or within 5 feet) or may be located remote (e.g., in different parts of a building or complex of buildings) or geographically remote (e.g., miles apart) from one another, such as in the case that the external processing unit is implemented in a server and data is transmitted via one or more networks or the Internet.

(78) In a variant, as shown in FIG. 17, part of the processing is carried out in a distant server 310 to which data is transmitted via the internet for example.

EXAMPLES

(79) FIGS. 10A and 10B show a collection of amplitude and phase of an impedance spectra measured for cellular structures comprising respectively three cell types, i.e. platelets, smooth muscle cells and endothelial cells.

Comparative Examples

(80) First, an equivalent circuit model without CPE is used, consisting of a double layer capacitance C.sub.d1 in series with a solution resistance in series with a R.sub.0C.sub.mix (R.sub.0 resistance in parallel with C.sub.mix capacitance).

(81) Then, the C.sub.mix parameter describing the impact of the cells layers on the complex impedance is computed.

(82) The result of the distribution of C.sub.mix for two cell types is shown in FIG. 11A. It is possible to distinguish between the two cell types. However, if adding a third cell type the three cell types cannot be distinguished any longer, as shown in FIG. 11B.

(83) If one uses a more sophisticated approach and implement CPE elements into the equivalent circuit model, and uses for example the model 34 shown in FIG. 5, there are six parameters describing the system, i.e. R.sub.0, R.sub.inf, Q.sub.0, , Q.sub.dl and .

(84) These parameters can be computed so that the impedance of the equivalent circuit model best fit the experimental impedance spectra curves of FIGS. 10A and 10B.

(85) Then, one can display for each parameter the distribution of this parameter for the three cell types, as shown in FIGS. 12A to 12F.

(86) One can see that for each parameter the three cell types cannot be distinguished clearly, and no linear combination of these parameters can provide the cell discrimination that is looked for.

Examples According to the Invention

(87) FIG. 13 shows the distribution of a value representative of the effective capacitance C.sub.eff for the three cell types, determined based on the formula [9] above.

(88) One can see that it is possible to clearly distinguish between all three cell types. The precision is over 90%. The differentiation between cells is significantly improved compared to FIGS. 12A-12F.

(89) If the equivalent circuit is the one 34 of FIG. 5A, one obtains the C.sub.eff distribution of FIG. 14.

(90) If one considers R.sub.0-R.sub.inf is large in respect to R.sub.inf, the equation [9] can be simplified as C.sub.eff=Q.sub.0.sup.1/R.sub.1.sup.(1-)/

(91) The resulting distribution of C.sub.eff is shown in FIG. 15. One can see that the three cell types can still be distinguished with a precision of about 85%.

(92) The distributions shown in FIGS. 13 to 15 may serve as reference data for cell type determination.

(93) For example, an impedance spectrum may be measured in similar conditions as the impedance spectra of FIGS. 10A and 10B, and based on this spectrum the values of parameters R.sub.0, R.sub.inf, Q.sub.0, , Q.sub.dl and are determined. This determination may be based on least square fitting of the impedance curves of amplitude and phase with the equivalent circuit model 34 of FIG. 5.

(94) Then, once the parameter values R.sub.0, R.sub.inf, Q.sub.0 and are known, the effective capacitance C.sub.eff can be computed and the value compared with the distribution of FIG. 13 to determine to what cell type it corresponds. For example, a low value of C.sub.eff in nF/cm.sup.2 will indicate that the cells are of first type; a value between about 50 and about 100 that the cells are of type 3, and a value of over about 100 that the cells are of type 2.

Method for determining at least one type and/or condition of cells and system

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Abstract

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