1 Introduction THE PROPOSITIONthat electricity may act as a healing agent in medicine went for the past two centuries through several cycles of enthusiastic acceptance and total rejection. One of the reasons for a sceptical attitude is the fact that basic mechanisms of interactions of electrical fields with biological systems are still poorly understood. In addition, claims that electricity could cure almost any disease made the medical profession even more suspicious. To aggravate the issue some recent epidemiological studies tried to show that electromagnetic fields might even be harmful by producing cancer and leukaemia (WERTHEIMER and LEEPER, 1979; SAVITZ et al., 1990). Thus we are confronted with the puzzling question: How does electricity affect the human body--is it a panacea, placebo or poison? For a general and conclusive answer we shall still have to wait some time. However, as this question is of great medical and economic relevance, research in bioelectricity is gaining priority in biomedical and engineering sciences. One of the fundamental problems in this area is the effect of electrical currents on cell proliferation. Does electricity accelerate or retard the cell cycle; does it affect it at all? We made an attempt to provide evidence that electrical currents might work in both directions; that they can accelerate cell growth and division when the rate is too low and inhibit when it becomes abnormally high. ElecReceived l Oth April 1991

9 IFMBE: 1992

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tricity thus would be acting as a modifying agent by normalising abnormal cell profileration. We discuss this problem on two paradigms: healing of chronic wounds and tumour growth retardation due to applied electrical direct current. Finally we propose a unifying hypothesis which could offer one of the possible explanations why externally applied electrical currents might normalise cell proliferation.

2 W o u n d healing The healing of injuries inflicted on skin and soft tissue is one of the rare regenerative capabilities of the human body. Wounds may appear on the body both without external injury or surgical intervention and due to surgery (e.g. amputations), all of which may not heal. Such chronic wounds may last for months or years and therefore represent medically, socially and economically an important problem. Two major groups of patients are susceptible to chronic wounds:

(a) spinal cord injured patients who develop pressure sores (deeubitus ulcers) due to their immobility and increased pressure over extended periods of time on bone tuberocities (trochanter, sacrum, heels) (b) patients with peripheral vascular diseases who develop ischaemic ulcers on the lower extremities due to

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various pathological conditions, the most common being diabetes mellitus; in a large proportion of such patients their condition impairs to such an extent that amputation is required with the usual complication of the stump wound refusing to heal.

I001 90 80 70

g~ One of the modalities to initiate wound healing is the application of electrical currents. Various currents and stimulation sites are currently being used for this purpose (KAADA, 1983; CARLEY and WAINAPEL, 1985; JIVEGARD et al., 1987; VODOVNIK et al., 1988; 1991; KLOTrI and FEEDAR,1988; IERAN,1990). As supporting evidence for our hypothesis regarding the normalising effects of electrical currents and fields on cell proliferation we examine only their simplest form: direct current (DC) applied across the wound. There exist several investigations which report improved wound healing following the application of DC electrotherapy (WoLcOTT et al., 1969; GAULT and GATENS, 1976; CARLEY and WAINAPEL, 1985; VODOVNIK et al., 1991). However, they usually lack a careful experimental design and comparison with a control group, which is probably one of the major reasons that electrotherapy of chronic wounds is met with scepticism by the medical community. In the next paragraph we give a brief account of our recent results in treating pressure sores by DC electrotherapy. Spinal cord injured patients admitted to the Rehabilitation Institute in Ljubljana for treatment of pressure sores were assigned into a control group and an experimental group. Groups of patients were matched by age, diagnosis, initial wound size and initial wound depth to obtain statistically nonsignificant differences of these parameters among both groups (the study was designed and carried out as a double-blind). Patients in the control group received conventional treatment while patients in the experimental group were subjected, in addition to equally careful conventional treatment, to two hours of electrotherapy daily until the closure of the wound. The current (0.6mA) was applied across the wounds by means of selfadhesive skin electrodes (Pals Plus*). In our former studies we found that the healing process in individuals can be well fitted with an exponential curve (STEFANOVSKAe t al., 1987; KARBA et al., 1991). If So is the initial wound surface area and S the surface area at time t, the reduction of wound size may be expressed as S

=

S 0 exp ( - t / z )

where z is the time constant of the process. To quantify the healing process we introduced the relative healing rate 0 [per cent per day], the reciprocal value of the time constant, 0 = 1/z

x

100 per cent = 1/t In (So~S) x 100 per cent

The values of the sore size together with other data from patients participating in the study were collected weekly. Methods used for assessment of electrotherapy effectiveness are described in detail elsewhere (VoDOVNIK and STEFANOVSI(A, 1989). The results of a study including 49 patients in a control group (conventional treatment) and 18 patients in a group treated by DC electrotherapy are summarised in Fig. 1 A uniform presentation of the results is a very difficult task due to different initial wound size and grade; the location of the wound and various other factors influencing the *Pals Plus is a trademark of Axelgaard Manufacturing Co. Ltd.

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60

50 0-.~.0

3O 0"-

2O 10 0

30

60

90

120

150

180

t im e , days

Fig. 1

The healing process of decubitus ulcers in patients subjected to conventional treatment (control group, n = 49, Q ) and in patients with adjuvant DC eIectrotherapy o f 0"6mA, two hours daily (n = 18, 0 ) . Data points are mean values of the groups with standard error of the mean vertical bars. The wound healing exponential curves with relative healing rate ( 0 : 2 . 2 + 3.2 per cent per day in control; 3"7+_3"5 per cent per day in electrotherapy group; mean + SD) were determined for both groups from individually obtained relative healing rates of each patient

healing process caused rather large variations in the presented results. For each wound individually, relative healing rate 0 was determined, resulting in 2.2+3.2 (mean • SD), n = 4 9 in control group and 3-7_+ 3.5, n = 18 in the group subjected to electrotherapy. Test of variances (F-test) implied equal variances of both samples (P[F48, t7 ~< 0.90); therefore means were statistically evaluated employing single-sided Student' t-tests, indicating that the relative healing rate of the group subjected to electrotherapy in addition to conventional treatment was higher in comparison with the control group with 0-90 < P [ T ~< t] < 0-95. 3 Tumour growth retardation Although qualitative differences between normal and tumour cells have been identified a specific cancer therapy which would act selectively on tumour cells has not been found. Under the term cancer we include various diseases to which infinite ability of dividing, invasion of surrounding tissues and capability of travelling to distant sites (metastases) are common and mutual. Conventional approaches in containment of cancer, e.g. surgery, chemotherapy, immunotherapy and their combinations, do not satisfy the needs. Therefore new treatment modalities are sought. One of the potential local therapeutical agents is electrotherapy by means of low-intensity direct current (DC). The idea of employing DC in cancer therapy, although not new, has not received the expected attention (HUMPHREYand SEAL,1959; DAVID et aL, 1985; MARINO et al., 1986; NORDENSTROM, 1989; SER~A and MIKLAV~I~, 1990; MIKLAV(~I~ et al., 1990). In the next paragraph we present one of our recent results on an in vivo tumour model. Solid subcutaneous tumours were initiated by subcutaneous injection of 5 x l05 fibrosarcoma Sa-1 viable tumour cells dorsolaterally in A/J syngeneic mice. After one week of growth tumours reached the initial volume Vo of approximately 4 0 m m a, mean tumour diameter 4.2 mm. On day D O animals were randomly divided into two groups; control (Vo = 41.2 + 5.8mm3; n - - 1 7 ) and electrotherapy (Vo = 39-0 -t- 4-6mm3; n = 29). Single-shot electrotherapy (ET) was performed on D o with 0.6mA of

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i~176 25o~-

200 E ~ 150 o 0

E

50

0 9

I

I

5

I0

days after treatment

Fig. 2

Tumour growth curves for control (n = 17, 0 ) and group subjected to single-shot electrotherapy (n = 29, A) of 1 h duration on day 0 with 0.6mA. Vertical bars: standard error of the mean

1 h duration. Electrical current was delivered via metal electrodes (Pt/Ir alloy, 90/10 per cent; diameter 0.Tmm, 18mm length) which were introduced through small skin incisions and placed subcutaneously cranially and caudally, each 8-10ram away from the tumour edges. During the ET the animals were firmly fixed. No anaesthesia was employed. Tumour growth and the effectiveness of electrotherapy was assessed by determination of tumour volume (V = rcabc/6) on each subsequent day by measurement of three mutally orthogonal diameters (a, b and c) by means of a vernier calliper gauge. The results were statistically evaluated by the Mann-Whitney rank-sum test (two-tailed on DOand single-tailed on Di-D7). Tumour growth curves (Fig. 2) and statistical evaluation (p--0.2797 on Do and p--0-0001 on D1-DT) showed faster tumour growth retardation caused by single-shot electrotherapy of 1 h duration. 4 Factors affecting cell proliferation The two sets of experiments described in the previous paragraphs, although hardly comparable, add supporting evidence to the observation that reasonably weak electrical currents of the same intensity may either enhance or inhibit cell proliferation in the case of chronic wounds and tumour growth, respectively. In the tissue, according to its needs, the cells constantly grow and divide, this process being highly regulated (PARDEE, 1989; LASKEYet al., 1989; MURRAYand KIRSCHNER, 1989; MCINTOSH and KOONCE, 1989; HARTWELLand WEINERT, 1989; O'FARRELLet al., 1989). Most normal cells are differentiated, having specific morphology and function. During the process of differentiation, normal cells mature and tend to lose the ability of proliferation. In the tissue where the cells have to be replaced, new cells proliferate from undifferentiated precursor cells. The process of cell proliferation can therefore be dissected into cell growth and division, where the cells have to proceed MBEC

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through the cell cycle and differentiation into tissue specific cells. In pathological conditions cell proliferation can be suppressed, as in the condition of chronic ulcers, or enhanced where proliferation is continuous and independent of the requirements for new cells, as in the case of neoplastic growth. Both conditions have their rationale in hampered regulation of the cell cycle, which is a sophisticated process, still not well understood, but under intensive investigation (MURRAYand KIRSCHNER, 1991). One of the well known cancer cell properties is their lack of contact inhibition. Normal cells in tissue divide until neighbouring cells are touched, while malignant cells grow irrespective of cell contact and invade the neighbouring tissue. Cell division is regulated on the cell nucleus, on cytoplasmatic and on the cell membrane level. In addition, environmental factors such as nutrient and oxygen supply are needed for succesful conductance of a cell through the cell cycle. Growth factors which are synthesised and secreted by various cells can both stimulate and inhibit cell proliferation. These factors activate synthesis of other factors which drive the cells over the restriction point into cell division, or may help the cells to progress through the cell cycle (MIYAZAKI and HORIO, 1989; SORRENTINO, 1989; BASERGA, 1990). Some of these factors are platelet-derived growth factor (PDGF), fibroblast growth factors (FGF-ct, FGF-fl), epidermal growth factor (EGF), transforming growth factors (TGF-~, TGF-fl), interluekin-1 (IL-1), interleukin-2 (IL-2), nerve growth factor (NGF), hematopoetic growth factors (IL-3, GM-CSF, M-CSF, G-CSF) and insulin-like growth factors (IGF-I, IGF-II). Cells compete for these factors, which bind to specific receptors on cell membrane. The signals are then transduced into the cell by different mechanisms. Some of the transducers are tirosin kinase, protein kinase C and cyclic AMP (cAMP). The cAMP receptor proteins are important in molecular control of growth and differentiation in normal development, malignant transformation, and suppression of malignancy. The restoration of the normal functional balance of cAMP receptor isoforms can lead to the restoration of intracellular regulatory molecules that induce differentiation and could be of importance and of great value in cancer therapy (CHO-CHUNG, 1990). Recent investigations have produced evidence that the maturation promoting factor (MPF) is a major regulator of mitosis. It has also been found that MPF consists of two proteins: the cdc 2 (cell division cycle or cell division control) protein; and cycline--a protein which cyclically appears in interphase, activates the cdc 2 and is abruptly degraded after the cell has entered mitosis (MURRAYand KmSCHNER,1991). The synthesis of these proteins is strongly dependent again on the presence of nutrients, oxygen, hormones and a proper ionic environment. All these factors influence DNA replication either by direct binding to DNA as promoters or through perturbation of intracellular ionic composition. It has been shown that elevated intracellular Ca 2 + and H § levels are necessary for the transcription of genes for DNA replication (EBERHARDand HOLZ, 1988). It has also been suggested that one of the most important ions regarding cell mitotic activity seems to be Na § At low intracellular concentrations of Na + the cell proliferation is blocked and the cell resides in the G O state. When the concentration of Na § increases, the mitotic switch shifts to the Gl metabolic mode. From several experimental data CONE concluded that the 'absolute intracellular concentrations of Na § is the key factor involved in control of mitogenesis activation in somatic cells' (CONE, 1969; 1970; 1971; CONE and CONE, 1976). The importance of the plasma membrane as the factor 1992

C E23

affecting cell growth and division has been pointed out where all extracellular signals are perceived and transduced into the cell (NICOLSON, 1976a; b). External and internal factors which influence cell cycle may also have an effect on the transmembrane potential, which according to some experimental data is reduced in the cells which underwent neoplastic transformation and are extremely high in nondividing cells. 5 Transmembrane potential and its biological implications From the many factors affecting cell proliferation we will concentrate on the transmembrane potential (TMP). It is quite possible that the factors mentioned are interdependent and only an orchestrated activity of all of them triggers the cell cycle. However, the TMP presents an experimentally convenient integrated parameter amenable to measurement, and it might reflect the status of the ionic environment inside and outside the plasma membrane as well as the properties of the membrane. The transmembrane potential of non-excitable cells has long been shadowed by the role of TMP in excitable cells. WILLIAMS(1970) has collected and reviewed the data existing on TMPs of various nonexcitable cells. His general conclusion was that the TMP in nonexcitable cells is consistently negative (inner to outer) but of lower value than in excitable cells and that TMP is of the same origin in both cases. In equilibrium (resting TMP), such as in excitable cells, the approximation of the Goldman equation RT PK[K+]o + PNa[Na+]o TMP = - - In p~[K+] ~+ pN,[Na+] ~ describes the TMP of non-excitable cells sufficiently well. Similarly, such as in the case of excitable cells, the membrane ion permeability to K + is much higher than its permeability to Na +, resulting in values of ratio p = PNa/PK in the range 0.006-0.36, which is somewhat higher than in excitable cells where p = 0.01. The profile of the electric potential across the membrane and in its vicinity, due to membrane surface charge density, should not be overlooked because of its possible important biological implications of electric potential slope and inner (~kl) and outer (fro) membrane potential (Fig. 3). The results of measurements and the profile of the electric potential has been reported by several authors (HEINRICHet al., 1982; IGLI(3 et al., 1987; McLAUGHLIN, 1989). Without going beyond the scope of the paper we would like to point out that the ionic current I across the

IiJjiiiiEiiiiiii!iiiiiiiiiiiiiiiiiiiil

TMP -

~ii~iiiiiiiiii!iiiilil i~iiiiii~iiiii~iiiiii~iiiii~i ~iii;iiiii~iiiiiiii~i

#i

iii~i~iii iiiiiiii~iiii iiiiiii~iiiiiiiiii~ii iiiiii~iiiiiiiiiiiiiiiiiiiiiiiiiiiiiiiiii~ Co outer solution

ci membrane

inner s o l u t i o n

Fig. 3 Profile of the electric potential across the cell membrane and in its vicinity (schematically). The meaning of the symbols is given in the text

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membrane functionally depends on inner ~ki and outer ~ko potential, bulk ionic concentrations in the cell ci and outer millieu co, as well as on the TMP which is measured experimentally by means of microelectrodes. Furthermore, the interdependence of both membrane potentials (~'i, 0o), TMP and concentrations (q, Co) should be taken into account. The complex relationships, which are reflected in the merely sketched functional dependence of ionic current across the plasma membrane, give us the enormous possible relationships of TMP such as with conformational changes of membrane proteins, surface membrane charge and ionic distributions, membrane permeability, ion channels' conductivity and several others. It has been shown experimentally that neoplastic transformation of the cell was accompanied by changes in cell surface charge density and TMP resulting in lower TMP after the cell had undergone the neoplastic transformation (PRtCE et al., 1987). TMP and ionic potassium and sodium concentrations were related to DNA synthesis (ORR et al., 1972; McDONALD et al., 1972; ZS-NAGY et 'al., 1981). Simple monovalent ion regulation of genetic translation systems was reported (Douzou and MAUREL,1977). TMP and ionic intra and extracelullar concentrations were closely related to mitogenesis and oncogenesis (CONE, 1971; CAMERON et al., 1980; LEEFERT, 1980; BORGENS, 1982) and to lateral diffusion of cell surface antigens (EDIDIN and WEI, 1977). A close relationship between TMP alteration and the cell cycle has been found (SACHSet al., 1974; STAMBROOK et al., 1974; BOONSTRAet al., 1981). Low TMP was measured during late G1 phase, followed by abrupt elevation of TMP in the transition of the cell from the G1 to the S phase. The TMP then remained elevated throughout the cell cycle, again decreasing in the next G~ phase. A very strict relationship between contact inhibition and TMP elevation, and lack of contact inhibition in transformed cells being accompanied by low TMP, with no TMP elevation, has been reported as well (BINGGELIand WEINSTEIN,1985; CONE and TONGIER,1973). Many of the well known cell cycle regulatory mechanisms have already been connected to monovalent ion concentrations and concomitantly to TMP, and other relationships have been proposed (CONE, 1971; DONALDSON, 1975; NIEMTZOW 1985; DE LOOF, 1986), giving us a reasonably solid base to believe that the TMP could provide us with a good insight into the very complex life of the cell and its 'cycle'. 6 Normalisation of transmembrane potential When a cell is in its normal equilibrium state external electric fields with intensities within physiological limits do not affect general functions of the cell. FINDL (1987) proposed for such a situation the term 'electrical homeostasis'. The cell in its electro-homeostatic state compensates external fields by adjusting its internal miUieu in such a way that external fields produce no permanent effect on cellular processes. However, redistribution of integral membrane components, changes in cell shape, cell migration and orientation as well as other changes on the membrane level due to exposure of the cell to externally applied electrical fields have been reported (JAEEE, 1977; JAFEE and NUCCITELLI, 1977; POD and ROBINSON, 1977; POO, 1981; ROBINSON, 1985). Possible targets of the externally applied electrical currents/fields are summarised in Fig. 4. From the results which we gave in previous paragraphs regarding healing of chronic wounds and tumour growth retardation we can expect an ultimate effect of electrical Cellular Engineering Special Feature

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currents on cell mitotic activity, enhanced in the former and retarded cell proliferation in the latter case. In an attempt to explain the two diametrically opposite effects of electrical currents we assume that in wounds and tumours the cell has shifted away from its electro-

cell membrane

externally applied electrical field

fluidity receptors cell microenvironment

conductivity and permeability ion currents transmembrane potential

ion concentration surface contact growth factors hormones nutrients

Fig. 4

facing side depolarised by A Ve (in our two cases we have estimated AVe being in the 5 - 1 0 m V range). In a normal cell we assume--for reasonably small changes of AVe--a linear relationship between changes in T M P and transmembrane current. On the analogy of

intracelIular~ metabolite and concei~ t i o /

ONA synthesis

1

mitosis on or off

Schematic flowchart illustrating possible targets and the influences of an externally applied electrical field E o on a cell through effects on the microenvironment and via the plasma membrane. Transmembrane potential ( T M P ) is controlled and determined by the external environment and membrane structure and properties

homeostasis into a 'stressed' state (FINDL, 1987). Thus it seems that external currents are able to shift the cell from its stressed state back to or towards the electrohomeostatic state. To explain how such shifts might be achieved we propose the following hypothesis. Let us assume a simplified situation where a spherical isolated (nonconductive membrane) cell of radius r having a resting T M P of magnitude V, is exposed to a homogenous field E o (Fig. 5). It has been shown that the T M P in a homogeneous field (JAFFE and NUCCITELLI,1977; ROmNSON, 1985) changes according to the following approximate equation: Vm = I1, - l ' 5 r E cos |

= 1,1,+ AVe cos O

where O is the latitude of the sphere. The transmembrane potential thus varies sinusoidally on the cell surface so that the anode-facing side is hyperpolarised and the cathode-

externally applied electrical field Eo +'rr/2

Vr+Av e0

rl,,j ~

--

--

neurons we also suppose that the I / V characteristic has a negative slope which means that an increased depolarisation would increase the transmembrane current, whereas hyperpolarisation would decrease the current. The normal situation in Fig. 6 is depicted with a resting potential V, (state A) and the corresponding current I,. If the cell is exposed to an external electrical field the anodal side of the cell will become hyperpolarised and the transmembrane current will decrease by Alh. Without affecting the essence of our hypothesis we assume the cell as twodimensional. If G is the conductance per angular unit we obtain in the normal resting state a decrease in current across the membrane AIh n/2

AI h = -GAVe

fd hi2

cos O dO

-

On the cathodal side, however, the cell will become equally depolarised and an increase of current AIa will be observed.

f nt2 AIa = GAVe

d-n~2

cos 19 dO

The total change in ionic current integrated over the whole cell membrane due to the application of the external field will therefore be zero because

-- ,rrV r +/W e

IA/hl = [Alal

The external field did not change the overall ionic transmembrane current and the cell remained in its electrohomeostatic state. Around this state we assumed a linear relationship.

- ,Tr/2 Vr Fig. 5

MBEC

x/2

AI = + GAVe

Spherical cell with membrane potential V,. When an external electrical field E o is applied, the anode-facing side becoms hyperpolarised and the cathode-facing side becomes depolarised equally by A Ve

Cellular Engineering Special Feature

J-n~2

cos O dO

with G being constant. It is quite unlikely that such linearity would hold for large changes in A V e or for changes in

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II, from its normal resting value. Increased hyperpolarisation will decrease the current to a small, constant level whereas large depolarisations will produce a maximum, limited transmembrane current. We believe therefore that

184 A i ____,r

7 Conclusions We are quite aware of the flaws in the proposed hypothesis. There is no experimental proof as yet that cells in chronic wounds have an increased TMP, and no direct experimental data exist about the nonlinear characteristics between ionic current and TMP. But we believe that the indirect evidence for these assumptions is strong enough to justify the hypothesis. To our knowledge this is the first attempt to propose a unified approach for two diametrically opposite applications of electrical currents. The model offers a plausible though hypothetical explanation of the apparent contradiction that external fields can either enhance or reduce cell proliferation. It also offers a rationale for further basic research on the effects of external electric fields on cell proliferation as well as encouragement for continued clinical application of electric currents.

Acknowledgments--The authors wish to express their appreV r - A Vc

Fig. 6

Vr

Vr - t N w

In a normal cell (I1,; state A) when exposed to an electrical field E o the hyperpolarisation (+AVe) on the anodal side and depolarisation ( - A V e ) on the cathodal side of the cell produce zero net ion flux (I = AI h - Ale). Thus there is little or no influence of E o on the cell. When a cancerous cell (V~ -- AVe; state B) is exposed to Eo, the ion flux on the hyperpolarised side (AI~) is larger than on the deport the net ion flux (1) drives the cell larised one (AIa), towards the homeostatic state (11, ; state A). When a nondividing cell (V, + AVw ; state C) is exposed to Eo, the ion flux on the depolarised side (AI'n) is larger than on the hyperpolarised side (Arh) ; the net ion flux (I) driving the cell again towards the homeostatic state (V~ ; state A). The direction of ion flux (I) is defined as a movement of cations into the cell .

the nonlinear I / V curve depicted in Fig. 6 is quite realistic and physiologically acceptable. Point A in Fig. 6 may thus represent the state of electrical homeostasis while points B and C represent the stressed situations where the resting potentials V~- AV~ and V~+ AVw are too low (cancer) or too high (chronic wounds), respectively. Let us suppose we have a stressed cell with a resting potential increased by AVw from the normal value V~. This cell could represent a hyperpolarised cell of a chronic wound. If the cell is exposed to an external field, hyperpolarisation by AVe on the anodal side and depolarisation by AVe on the cathodal side will be observed again as in the normal cell, the previous case, but the suggested nonlinear I / V characteristic causes different ion fluxes on the anodal (AI;,) and cathodal sides (AE,), the latter being higher.

ciation to their colleagues from the Laboratory of Biocybernetics, Faculty of Electrical & Computer Engineering, Ljubljana, especially to R. Karba, M. Sc., and P. Kro~elj, Dipl. Eng., to colleagues from the Tumour Biology Laboratory of the Institute of Oncology, Ljubljana, and to A. Igli~, M.Sc., from the Institute of Biophysics, School of Medicine, Ljubljana, who all contributed to the work presented. This work was supported in part by the Ministry of Science & Technology of the Republic of Slovenia, by the National Institute for Disability & Rehabilitation Research, Washington DC, USA, and by the Commission of the European Communities, Directorate-General for Science, Research & Development, International Scientific Cooperation, Brussels, Belgium. This paper was presented at the IFMBE Satellite Symposium on Cellular Engineering in Medicine, 1st European Conference on Biomedical Engineering, Nice, France, 21st February 1991.

References BASERGA, R. (1990) The cell cycle: myths and realities. Cancer Res., 50, 6769-6771.

BIN~GEL[, R. and WEINST~IN, R. C. (1985) Deficits in elevating membrane potential of rat fibrosarcoma cells after cell contact. Ibid., 45, 235-241. BOONSTRA, J., MUMMERY, C. L., LEON, G. J., TERTOOLEN, L. G. J., VAN DER SAAG, P. T. and DE LAAT, S. W. (1981) Cation transport and growth regulation in neuroblastoma cells. Modula-

tions of K + transport and electrical membrane properties during the cell cycle. J. Cell Physiol., 107, 75-83. BORGENS, R. B. (1982) What is the role of naturally produced electric current in vertebrate regeneration and healing. Int. Rev. Cytol., 76, 245-298. CAMERON,I. L., SMITH,N. K. R., POOL, T. B. and SPARKS,R. L. (1980) Intracellular concentration of sodium and other elements as related to mitogenesis and oncogenesis in vivo. I AE,[ > I AI~I Cancer Res., 40, 1493-1500. CARLEY, P. J. and WAINAPEL, S. F. (1985) Electrotherapy for The net ionic flux will increase, thus driving the resting acceleration of wound healing: low intensity direct current. potential closer to the homeostatic V~. Arch. Phys. Med. Rehabil., 66, 443-446. In an analogous way it may be postulated that the C~O-CHUNG, Y. S. (1990) Rote of cyclic AMP receptor proteins in resting potential of a cancerous cell might be V~- AVe. growth, differentiation, and suppression of malignancy: new Again, exposure to an external field will change the transapproaches to therapy. Cancer Res., 50, 7093-7100. membrane potentials equally (by AVe) on the cathodal and CONE, C. D. Jr (1969) Electroosmotic interactions accompanying anodal sides. However, the cathodal side of the cell will mitosis initiation in sarcoma cells in vitro. Trans. N Y Acad. now experience a smaller increase in ionic flux compared Sci., 31, 404-427. with the anodal side. This will result in CONE,C. D. Jr. (1970) Oncogenesis. Oncol., 24, 438-470. CONE, C. D. Jr (1971) Unified theory on the basic mechanism of IAI;I < IAI~I normal mitotic control and oncogenesis. J. Theor. Biol., 30, 151-181. thus reducing the overall transmembrane current and CONS, C. D. and TONGIER, M. Jr (1973) Contact inhibition of increasing the TMP, which consequently represents a shift division: involvement of the electrical transmembrane potentowards the homeostatic state. tial. J. Cell Physiol., 82, 373-386. CE26 MBEC Cellular Engineering Special Feature July 1992

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WmLIAMS, J. A. (1970) Origin of transmembrane potentials in non-excitable cells. J. Theor: Biol., 28, 287-296. WOLCOTT, L. E., WHEELER, P. C., HARDWICKE,H. M. and ROWLEY, B. A. (1969) Accelerated healing of skin ulcers by electrotherapy: preliminary clinical results. South Med. J., 62, 795-801. ZS-NAGY, [., LUSTYIK, G., Zs-NAGY, V., ZAR'ANDAI, I . and BERTONI-FREDDARI,C. (1981) lntracellular Na+: K + ratios in human cancer cells as revealed by energy dispersive X-ray microanalysis. J. Cell Biol., 90, 769-777.

Damijan Miklav~i~ was born in 1963 in Ljubljana, Slovenia (then part of Yugoslavia). He received a B.Sc. in 1987 and an M.Sc. in Electrical Engineering in 1991 from the University of Ljubljana, Faculty of Electrical & Computer Engineering. He is currently working towards his Ph.D degree and holds a position of teaching and research assistant at the Faculty of Electrical & Computer Engineering. His prime interests are in the field of biomedical engineering.

Authors" biographies Lojze Vodovnik, Dipl.Eng., D.Sc. completed his studies at the Faculty of Electrical Engineering, University of Ljubljana, Slovenia, (then part of Yugoslavia), where he is Professor of Biocybernetics & Neurocybernetics. He was research associate and visiting professor from 1964 to 1969 at Case Western Reserve University, Cleveland and World Rehabilitation Fund Fellow in 1981 at Rancho Los Amigos Hospital, Los Angeles, USA. He is a Fellow of the IEEE, senior member of the Bioengineering Society, member, Board of Directors, European Society for Engineering & Medicine, and a member of the Slovenian Academy of Science & Arts.

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Gregor Ser~a was born in Ljubljana, Slovenia (then part of Yugoslavia), in 1956. He received a Masters degree in 1984 and Doctorate degree in Biomedical Sciences in 1988, both from the Medical Faculty, University of Ljubljana. From 1986 to 1987 he was visiting researcher at MD Andreson Hospital, Department of Experimental Radiotherapy, Texas, USA. He is currently working in the department of Tumour Biology & Biotherapy at the Institute of Oncology, Ljubljana, Slovenia. His interests are tumour biology, experimental radiotherapy, biological response modifiers and electrotherapy in cancer treatment.

Cellular Engineering Special Feature

July 1992