biofheattarnsfer

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REA T TRANSFER N BIOMEDICAL ENGINEERING ANDBIOTECHNOLOGY

Keywords: bioheat transfer, thermal biological sytstems. hermal evolution, hyperthermia, hypothermia,cryosurgery.cryobiologyABSTRACT

Heat transferplays an important role inbiological systems.The goal of this article is tointroduce researchers n the general area ofengineering heat transfer to the area of bioheattransfer.To this end will review selected opicsfrom eachof the different areas hat comprise hefield of biological heat transfer. Prior to adiscussionof the heat ransfer n organisms hatexist on earth now, I will introduce the area ofthermal evolution. Than I will describeselectedtopics of research in each of the followingtemperature anges:a) temperatures igher thanthe physiological temperatureof warm-bloodedorganisms, rom about40 c to about 120c, b)temperaturesn the physiological angeof warm-bloodedorganisms, rom about 36 c to about40c and c) temperatures lower than thephysiological temperature of warm-bloodedorganisms, rom about 36 c to - 273 c. In the

. Proceedingsf the~ ASMFJJSME oint Thennal EngineeringConferenceMarch 15-19, 1999,SanDiego, California

Boris RUBINSKYDepartment f MechanicalEngineeringUniversity of Califomia at BerkeleyBerkeleyCA 94720. [email protected]

first range of temperature he review focusesonthe use of high temperaturesn medicine o treatundesirable issue and on the study of thermalaccidents. n the second range of temperaturesthe review focuseson three scales:a) microscale,in which I win discuss molecular motors, b)mesoscale n which I will discuss the heattransfer between blood and tissue and c) themacroscale n which I will discuss the area ofwhole body heat ransfer. The third temperaturerange can be subdivided into two; temperaturesabove freezing and below freezing.Temperaturesn this rangeare mportant becausethey can be used o preservebiological tissueandcells and because hey can be also used todestroy undesirable issue. Here I will discussthe thennal mechanismsof cell damage n thistemperature range and also cold and freezetolerant animals.

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INTRODUCTIONLife has evolved in relation to thetemperatureand the thermal environmentof ourplanet. It is no coincidence hat the optical nerveis affected by radiation only in the narrowwavelength range of between 0.38J1m and0.76Jun. This range, known as visible light,corresponds o the peak of solar radiation. It isalso no coincidence that all warm bloodedanimals have a deep body temperature ofbetween36 c and 40 c. This temperatures 15c above the average temperate zonetemperatureof 22 c. The cooler environmentservesas a heat sink to warm-blooded rganismsand allows them to function effectively at aconstant temperature, while continuouslyproducing heat. From thermal considerations,tis obvious that warm blooded animals, whichhave an aerobic metabolism,continuously bumoxygen and produce heat. could not maintaintheir current temperature f the temperatureofthe environment would have been 40 c, forinstance. n a different thermal environmentwewould have evolved differently, and as thethermal environment of our planet changesterrestrial organisms change. In studying thebroad area of bioheat transfer I find that theresearch on life processes n relation to thecurrent thermal environmentof our planetcan bedivided into three emperatureanges:

Temperatureshigher than the physiologicaltemperature of warm-blooded organisms,from about40 c to about 120c.Temperatures n the physiological range ofwarm-bloodedorganisms, rom about 36 cto about 40 c.Temperatures ower than the physiologicaltemperature of warm-blooded organisms,from about 36 c to - 273 c.Distinctly different bioheat ransferproblemsareencountered n each of these three temperatureranges. The goal of this article is to introduceresearchers n the general area of engineeringheat transfer to the area of bioheat transfer.Therefore n this paper will describe he topicsonly briefly and the goal will be to introduceasmany topics as this spacepermits. I will discussresearch opics and problems typical to each ofthe three temperatureangesof relevance obioheat transfer isted above. However, before Iproceedwith a discussionon the bioheat ransferproblems in organisms hat live in the current

thermal environment I want to address briefly theexciting topic of thermal evolutionTHERMAL EVOLUI10NOrigin of liteOne of the most exciting questions inthe history of science s the origin of life. Haslife originated n a cold or in a hot environment?Has ife originated n the watecsof hot springs orgeothermal entsor in cool poolSor cold oceans.Proponents f the hot origin of life point to thefact that many of the organisms hat Populate heearliestbranchesof the tree of life live today inextreme thermal environmental conditions(Forterre, 1996; Vargas et aL, 1998; Vogel,1999). These organisms, known ashyperthermophiles,hrive between80 c and 90c. Support or the proponents f the cold originof life has comes rom a recent study by Galtieret at. (1999). GaItier and his colleagues haveapproachedhe questionabout the thermal originof life from a molecularperspective.They havetracked the evolution of two temperaturesensitive ibosomeRNA (rRNA) molecules rombacteria o mammals. (The RNA is composedofnucleotidebases hat form temperaturesensitivebonds. Some of the bonds withstand hightemperatures better than others. The basesguanine G) and cytosine C) form a strong bondwhile adenine A) and uracil (U) form a weakerbond. Studieshave shown hat the rRNA of heatloving organismshas more G and C than A andU, presumablebecause he G-c bond is morestableat higher temperatureshan the weaker A-U bond (Vogel, 1999).)Using a Markov model,GaItier et at. (1999) have evaluated the G-cnucleotide content of rRNA in the organismswhoseevolution they have tracked. Their resultsshow that "The inferred G+C content of thecommon ancestor o extant life forms appearsincompatiblewith survival at high temperature".The ancestral organisms apparently lived atmoderate temperatures. With respect tohyperthermophilicspecies hey favor the notionthat "extant hyperthermophilic species evolvedfrom mesophilic organisms via adaptation tohigh temperatures" Altho~gh much excitingwork is being done n studying the thenna1originof life, the answersare not yet conclusive. Boththe molecular and the cellular research nto theorigin of life are open areas of considerableinterest to the heat transfer community.Scientists from the heat transfer andthermodynamic analysis community couldcontribute from their perspectiveand training tothe search or the thermalorigin of life.


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Origin of endothermyAnother interestingquestion n thennalevolution relates to the origin of endothermicorganisms.Severalbooksand reviewshave beenwritten on this topic (Ruben, 1996; Fastovsky,1996; Randolph, 1994). Depending on themethod of thennoregulation, n organismcan beclassified as either endothermicor ectothennic.Endothermic organismsgenerateheat from thebody while ectothennic ones obtain heat fromtheir environment Maintaining a constantbodytemperature is essential to the survival ofendothermic animals. Enzymes n endothermicorganisms, for example, denature duringhyperthermia, when the body temperature isesabove 40 C. The first organ to be affected bytemperaturesabove 40 c is the brain. Seriousdisruptions leading to deathensue f the brain isexposed to high temperatures,or a prolongedperiod. On the other hand, hypothermia lowerthan normal body temperature) an causeequallydisastrous esults n slowing metabolicactivitiesand impairing brain function. In contrast toectothenns,which control their body temperaturemostly by behavioral adaptation, endothermsmaintain it by physiologicalmeans, hrough heiraerobic metabolism. Endothermic animalsrequire more energy expenditure thanectothennic ones since they have to maintaintheir constant temperature elevated over theaverage environmental temperature. Largeamounts of energy are spent by endothennsonthermoregulation and this seems to suggestinefficiency of maintaining such a highmetabolic rate. Moreover as more time is spenton obtaining food, there is also higher risk ofexposure to danger such as predators andweather hazards. What are the origins ofendothenny? Speculationshave been made onthe origin of endothenny as the selection of ahigh stable temperature for important lifeprocesses such as enzymatic catalysis,independenceof daily timing and resistance ofreezing. However, hermoregulation lone maynot justify its evolution. Metabolic incrementcannot have solely come from attempt o attainthermoregulationalone since n the initial stagesof the evolution of endothenny, he mechanismwould not have been effective in attainingthennoregulation. In addition endothermymayhave first evolved during the Mesozoic which isone of the most thermally stableperiods n earthhistory. Thermodynamic studies were madewhich compared oxygen consumption ofectothenns and endotherms during differentactivities. These studies suggest that the

increased aerobic metabolism of endothermsover the anaerobicmetabolismof ectotherms sthe main factor in the evolution of theendotherms. ven hough he cost of maintaininga high aerobicmetabolic ate s significant. thereare also a numberof advantageshat offset thecost. Sustainedstaminacan assist herbivores toescape arnivoresand help predators n capturingtheir pray. The key step in evolution may havebeen the elevation of metabOlic rates withaerobic respiration and thermoregulation mayhave been a consequence f the elevation ofmetabolic rate with increased aerobicmetabolismcapacity. The study of the evolutionof endothermy s another exciting topic of heatand mass transfer that should be studied byresearchersn heat ransferand hermodynamics.TEMPERATURES IN THEPHYSIOLOGICAL RANGE OF WARMBLOODED ORGANISMSUnderstanding he thermal behavior ofwarm-bloodedorganisms endothenns)can leadto a better understandingof life processes ngeneral and has applications in health care,engineering for human comfort and medicine.The study of heat transfer in warm-bloodedorganisms akes place at three different lengthscales:molecular microscale),whole organsandtissues (mesoscale) and whole body(macrosca1e). t the molecular scales there isextensive research in the areas ofthermodynamics f proteins assembly,molecularconversion (photosynthesis,ATP (adenine hreephosphate) production and molecular motors),ionic pumps (active transport of ions) andcellular metabolism (efficient combustion atroom temperature). At the mesoscale here isinterest in understandingheat transfer betweenblood vessels and the surrounding tissue,measuring blood flow with thermal clearance,understanding he thermal effect of malignanttissue, respiratory heat transfer, and even heattransfer in teeth. The thermal macroscaleresearchbas been the realm of the traditionalbio-thermal engineer. Extensive work was donein designing thermal comfort systems (beating,air conditioning and thermal comfort), studyingthermoregulation, heat transfer in extremeenvironments either n space,deep n the ocean,or during fires), developingan understanding orthe heat ransfer n newborn,elderly and sick. Inthe following sections I will introduce a fewexamples of the thermal work in each of thesethree scales.


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Thermal processes at the microscaleBiological systems produce energy.store energy and convert energy to work. This is

what any traditional inanimate thermal energyconversion system does. through variousprocesses of combustion. energy storage andmechanical motors and pumps for converting theenergy to work. However. there is a majordifference between biological systems andinanimate systems. Biological energyconversions systems are true nanodevices. whichfunction at constant temperatures of about 37 cto 40 oCt with superior efficiency. In contrastinanimate energy conversion systems aremacroscale device in which the combustionprocess takes place hundreds of degrees abovethe body temperature with lower efficiencies.Nature has designed compact motors known asproteins (kinesin's motor domain is 350 aminoacids) that convert chemical energy at roomtemperature from adenine triphosphate (ATP)hydrolysis to work with an efficiency as high as50% in the case of the myosin molecule. (Vale.1993). Protein motors produce musclecontraction, cause organelle motion and cellmotion. Attempts are currently made tounderstand how these motors work and to designthermodynamic models for these motors. Theseproteins work like stepper motors and a typicaldesign can be found in Vale (1993). Recently.Kitamura et al. (1999) have used a scanningprobe with a fluorescent label to detect thatmyosin moves along an actin filament withsingle mechanical steps of approximately 5.3nanometers. Protein motors have also otherfunctions. They activate ionic pumps across cellmembranes. Woiling in reverse these proteinpumps can use trans-membrane ion gradients tomanufacture ATP, the fuel of the biologicalsystem. Probably some of the most intriguingstudies in this field are by the group of Oster.(Wang et al., 1998; Elston et al.. 1998; Elston etal.. 1997; Doering et al.. 1995. For instance inElson et al. (1998), they have developed anequivalent thermodynamic and mechanicaldescription of a proton pump and engine. Theydescribe how a flow of protons. driven by thedifference in Gibbs free energy across themitochondrial membrane can produce threeATP's per twelve protons passing through themotor. 1be motor protein known as ATPase isshown to be composed of two counter-rotatingassemblies, one a stator and the second a rotor.1be motor must generate sufficient torque toproduce three A TP's per revolution, an amountof work equi valent to 20 lea per ATP molecule.

(ka is the Boltzman constant and T is theabsolute eMperature;at room temperature thevalue is 0.6 kcal mol -1 ). Elson et at. (1998)show that the motor is reversible and can actboth as a pump and as a means o convert ionicflow into stored useful energy, the A TPmolecule.While the goal of modem engineeringis to design more efficient energy conversionsystems and to build smaller and smallermicromechanical ystems, t is difficult to ignorethe fact that suchsystemsalreadyexist in nature.Thermal processes t the mesoscaleProbably the most extensive body ofwork produced by the bioengineering heattransfer community is in the mesoscale. Muchresearchhas been dedicated to modeling heattransfer n biological systems. Tissue s differentfrom other materialsbecause f the presence f avascular system and its associatedblood flow,metabolic heat transfer and heterogeneousthermal properties.The effect of blood flow ismost difficult to account or and researchers aveproposed different models. More detailedreviews of bioheat ransfer models can be foundelsewhere Chato, 1981; Shitzer and Eberhard,1985; Baish et aI., 1986; Chamy, 1992; Eto andRubinsky, 1996). The most widely used hermalmodel is commonly known as the bioheatequation (pennes, 1948). This formulationincludes a source/sink erm, which accounts orthe heat transfer from blood perfusion. Thegeneral formulation s,aT. .P,C'T=V(k,VT)+q_,+liJpbCb(T. -T,,)at,where p is density, c, heat capacity, T,temperature,, time, k, thermal conductivity, m,volumetric blood flow and the subscripts. , standfor tissue,b for blood, a for arterial blood and v,for venousblood. The bioheat equation assumesthat all the thennal effects of the blood can beabsorbed n the last term of the equation, asource erm. Here t is assumed hat blood entersany volume at tissue at the arterial temperatureand leaves at the venous temperature whiletransferringall the blood energy nstantaneously.Although it is widely used, the equation hasshortcomings.One difficulty is the determinationof the blood perfusion rate, and the fact that theblood flow is given a scalar erm. when in fact itis a vectorial quantity. There are other models.The Wulff (1974) model describes he tissue as aporous media and attributes the blood a Darcy'velocity. Chen and Holmes (1980) expandon the


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Pennes equation and add two new termsincluding the Darcy velocity in a porousmaterialand a term that describes the directionalenhancedheat transfer due to the blood flow.Weinbaumand Jiji, (1985) developeda seriesofnew bioheat equations hat originally dispensedwith the volumetric blood flow term andincluded only the directional flow of blood andrecently developed a new significance to thevolumetric blood flow term.

The need to analyze biological heattransfer at the blood vessel evel has generatedseries of mathematical models describing thedirect heat transfer interaction between bloodand issue.Chen and Holmes 1980) were amongthe rust to perform a simple energybalance or asingle blood vessel embedded n tissue. Thesolution is a simple exponential hermal decay,typical of heat exchangers.n analyzing varioustypical blood vesselparameters hey recognizedthat only blood vessels arger than about 80 J1111are thermally significant, while smaller bloodvesselsare not. Chato (1980) performed at thesame time a similar analysis treating the tissueand the blood as heat exchangers whileWeinbaumet at. (1984) approachedhe problemas an entry length problem. In tissue, bloodvesselswith diameters arger than the thermallysignificant blood vessels occur as vein-arterypairs. Mitchell and Myers (1968) were the firstto analyze the local heat counter-currentheattransfer, between he arterial and venous bloodflow in the pair. This counter current heattransfer is one of the most important aspect ofheat transfer in biological systems at themesoscale.

The area of bioheat transport haswitnessed in the last two decades, ncreasedactivity. Much of the work is related todevelopingmathematicalmodels or the effect ofblood flow on heat transfer n tissue.This workwas triggered by the interest in hyperthermiatreatment of malignant tissues, a topic I willdiscuss ater. While many advanceswere madein modeling heat transfer in tissue, this area isnot sufficiently understood.Thermal processes t the macroscaleCurrently most of the industrialapplications of bioheat transfer are in themacroscale ange. These relate primarily to thedesign of heating, ventilation and air-conditioning systems. The design requires agood understanding of the whole body heat

transfer and what is considered o be thennalcomfort. A ~ood summary of the field can befound in the ASHRAE Fundamentals (1997).The worlc in this field includes: evaluation ofwhat constituteshuman hermal comfort, studieson the effect of clothing on heat transfer to thehuman body (Xu and Werner, 1997),mathematicalmodeling of heat transfer in thewhole humanbody (Yokoyamaet al, 1997).Themathematical models of the' human bodydescribe segmentsof the body as a lumpedthermal mass and were developed first byStolwijk and Hardy (1966). Currently theresearch ocuseson expanding hese models ntosmaller thermal compartmentsand developingmodels for thermoregulation. Improvedexperimental thermal mannequins are beingdesigned o measure he heat transfer betweenthe body and the environment (Holmer andNilsson 1995). Studies are also carried out insuch areas as office and vehiclemicroenvironment thermal control andthermoregulation. There are importantapplications of this field in medicine. Heattransferand temperatures re mportant in sportsmedicine (Hargreavesand Febbraio, 1998) andin anesthesiology Saito et aI. 1998). Anotherimportantapplication relates o the change n thethermal behavior of humans with aging. Thethermal behavior of humanschangeswith age nboth the newborn (Benedict and Talbot, 1915)and the elderly (Shock. 1961; Florez-Duquet, etaI. 1998). t is important for the bioheat transfercommunity to worlc in the area of medicalresearch.Temperatures nd heat transfer playasignificant role in diseaseand dead, particular inthe extreme age groups, (O'Neal et ai, 1984;Kenney and Havenith, 1993), and many of theengineeringaspectsof the medical heat transferareahave not been esolved.TEMPERA WRES LOWER THAN THEPHYSIOLOGICAL TEMPERATURE OFWARM BLOODED ANIMALS.Low temperature heat transfer isaffectedby the fact that water s the largest issuecomponent Water freezes at 0 c and thebehavior of the biological system changesentirely when the water is removed from thesystemas inert ice. Therefore ow temperaturesbiological heat transfer can be divided in twotemperature ranges: temperatures above thefreezing temperature or water and temperaturesbelow the freezing temperature or water. Here Iwill discuss he behavior of biological systems n


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these two temperature anges in two separatesections.Biological systems in the temperature rangebetween normal mammalian physiologicaland 0 GC.Studying life in the temperature angeof between he normal physiological emperatureof warm blooded animals and the change ofphase temperature of water has practicalapplications in developing methods forpreserving mammalian cells and tissues byrefrigeration. Cell and tissue refrigeration isimportant in biomedicine and biotechnology.will begin with a discussionof phenomena hatoccur when the temperature of biologicalsystems are reduced from their normaltemperature nd than will discussorganism hathave evolved the ability to live under conditionsof low temperatures.Understandinghow theseorganismssurvive may lead to the developmentof techniques or preservation f cells and organsby refrigerationChanges n biological systemsduring cooling.Cells are entities in which anintracellular solution is separated from theextracellular milieu by a membrane. Theintracellular composition is entirely differentfrom the extracellular composition. Cell andsubcellular membranesmake life possible byisolating the interior of the cell and subcellularcomponents from their exterior. Thereby,membranes reserveand control the cell identityin the face of uncontrollable changes n theenvironment. The cell membrane acts as abarrier. Since the interaction and the masstransfer between he interior and the exterior ofthe cell is important, he cell membrane lso actsas a gate hat regulates he mass ransfer.This isdone through a variety of channels,pumps andother methods of controlled mass ransfer. Thecell membranealso servesas an anchor to thecytoskeleton hat provides he structural ntegrityof the cell. Cell membranes are made ofphospholipidsarranged n a bilayer configurationin which the hydrophobicheadsof the moleculesface outward of the bilayer and the hydrophilictails face inward. This configuration producesamembrane hat is impermeableo water.The cellmembrane s than spanby transportproteins hatform channelsand pumps.There is a very largevariety of different molecular species ofmembrane hospholipids.

In a nonnally functioning cell, lipids(fats) are in' a liquid phase. However, lipidsundergophase ransition, upon cooling. This isthe familiar phenomenon f fat solidifying in acooking pan after it cools. The cooled lipidbilayer can undergo several different types ofphase transfonnation. Some of the newstructures that fonD are known as invertedmicelles,hexagonal I structuresor cubic phasesSiegel, (1994). A detailed review of the processof lipid phase ransition can be found in Quinn(1985). Studying he phase ransition in the lipidbilayer is an interesting area of research withinthe realm of the heat ransfer engineer. Some ofthe most common methods for studying thesephase transformations utilize the opticalproperties of membranes, particular in theinfrared range Tablin, F et aI., 1996).During cooling, important changesoccur in the cell membranestructure and in thecell membrane unction (Drobnis, E Z et al.1993). The membrane lipids undergo phasetransition as the cell cools. 1be new lipidconfiguration fails to anchor tightly themembraneproteins and new openings form inthe cell membrane.The membrane ooses theability to isolate the cell interior from theexterior. Uncontrolled mass transfer begins totake place between he cell interior and exterior,which leads o cell damage McGrath, 11. 1997).The membrane roteins and the binding between

the cytoskeleton and the membrane are alsoaffectedby cooling. 1be mass ransfer activity ofthe membrane proteins involves chemicalreactions related to the A TP molecule. Areduction in temperature reduces thethermodynamic efficiency of these membraneproteinsand urther reduces he ability of the cellmembrane to regulate the intracellularcomposition. The bonds between the cellmembrane and the cytoskeleton are chemicalbonds that are temperature dependent. As thetemperature s reduced hese bonds weaken andthe cytoskeleton fails. Obviously all thesetemperature elated phenomena ausedamage oa mammaliancell that is cooled to temperatureslower than he normal physiological temperature.How can a refrigerated mammalian cell beprotected rom these modesof damageand howcan one preserve cells in a refrigerated state?Nature has found solutions. Some of them wejust begin to understandand some are not yetunderstood. Probably the animal model mostrelevant to the study of refrigeration ofmammalian issue s the hibernator.


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HibernatorsMammals normally maintain heir bodytemperature constant during the cold wintermonths by increasing their metabolism tocompensate or the increase n heat oss and byadaptationchanges. Adaptationchanges ncludeadditional layers of insulation on the bodysurface or habitat changes. When a warmblooded animal temperatures reducedbelow thephysiological temperature it will eventuallyloose its ability to control the metabolismanddie. However some mammals, he hibernators,have evolved mechanisms hat allow them tosurvive at low temperature. here are a numberof differences between obliged homeotherms(regular mammals) and hibernators.When anobliged homeotherm s artificially cooled below20C to 25 c all measurable igns of brain andnervous activity cease. The hibernators,however, an ower theircore temperatureo afew degreesabove he ambient and enter a stateof torpor. The hibernators' brain continues tofunction at these reduced temperatures.Mostmammals retain a relative constantheart rate astheir temperature s lowered, until they reach atemperature f irreversibledamage.n contrast nhibernators here is a gradual decreasen heartrate with temperature. The heart of thehibernators can maintain rhythmic beating attemperaturesnear 0 C. As the environmentaltemperature drops, an obliged homeothermicmammal will attempt to retain a constant highphysiological core temperature.Unlike obligedhomeotherms, the hibernator thermoregulatorymechanism allows the animal to retain athermoregulated emperature, ut at a level thatis significantly reducedand close to that of theenvironment. Therefore the heat loses in ahibernator are much smaller than in a mammalunder the same hermal conditions. Because heheat osses are small the hibernatorcan survivethe long winter months without metabolic heatgenerationand without the consequent eed forfood. Obviously hibernatorscan survive at lowtemperatures. But how can the hibernatorssurvive the fatal change hat cells experienceatlow temperatures? he mechanism s related, atleast in part, to molecular changes.Obviouslymaintaining a fluid membrane at lowertemperatures s essential. It appears hat inhibernators the cell membrane lipid phasetransition temperature is reduced duringhibernation.This is accomplished y changes nthe ipid compositionof the cell membrane itherprior to entering hibernation or throughout the

period.The secondmechanismmay relate to theprotein mas~ transfer carriers across the cellmembrane. Apparently in hibernators at lowtemperatures,while the metabolism s reducedthe activity of the ionic pumpsand the ability ofthe mitochondria o produceA TP is maintained.The fundamental mechanisms by whichhibernators achieve survival at reducedtemperature re not yet fully understood. 'Thereis an extremely small community of scientistsworking n this exciting bio-thermal areaand theengineering heat transfer community cancontribute o this research.A good review of thisarea can be found in several books (Wang,1982);Carey,C., et at. (Ed.), 1993).Antifreeze proteins and trehaloseIn response to low environmentaltemperatures,many fish and plants produceproteins with special properties. These proteinsare known as antifreeze proteins. There areseveral reviews on these proteins and theirproperties (Ananthanarayanan,1989). Here Iwill addressonly their low temperature elatedproperties. n a later section I will discuss theirfreezing related properties.Studies have shownthat antifreeze proteins protect the integrity ofthe oolema (membrane) n oocytes (unfertilizedeggs) at low temperatures (Rubinsky, et aI.1990), they reduce ionic leakag~ rom the cellmembraneat low temperature Rubinsky et aI.,1992) and inhibit ion transport across the cellmembrane (Negulescu et aI., 1992). It isconceivable hat these proteins have evolved toinhibit the mass ransfer across ow temperatureimpaired cell membranes (Rubinsky et aI.,1991).Studieswith liposomesand platelets haveshown that while the antifreeze proteins inhibitmass ransfer across he cell membrane they donot modify the lipid phase ransition of the cellmembrane Hays et aI., 1997; Hays, et al.1996). Because antifreeze proteins block iontransport hey were successfully used in organpreservation during hypothermia and inpreservation f such difficult cells to preserve,asplatelets. n addition to antifreezeproteins t hasbeen also shown that the sugar trehalose hasbeneficial roperties or preservationf cells na hypothermic state (Ring and Danks 1998).Trehalose s being produced by organisms innature Singer, et aI. 1998), and itwas shown tostabilize he cell membraneagainstcold damage(Beattieet aI., 1997).


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BIOWGICAL SYTEMS ATTEMPERATURES LOWER THAN THEFREEZING TEMPERATURE.Freezing of biological systems isanother reaof biologicalheat ransfer o whichscientists rom the heat ransfercommunity havemade major contribution. Freezing has aparadoxicaleffect on biological systems.On onehand freezing can be used o preservebiologicalmaterials and on the other hand freezing candestroy biological material. The preservationofbiological materials n a frozenstate s known ascryopreservation and the controlled destructionof biological materials s known as cryosurgeryand is used for treatment of cancer and otherundesirable tissues. To successfully eitherpreserve or destroy biological materials byfreezing n is important o developa fundamentalunderstandingof the processeshat occur duringfreezing. There are severalextensive eviewsandbooks on the fundamentals and the use offreezing in cryosurgery and cryopreservation(pegg and Karrow, 1987; McGrath and Diller,1988; Diller, 1992; Onik et at. 1994; Diller1997; Rubinsky, 1998). Here I will reviewbriefly some of the fundamentalaspectsof thefield.

Studying he processof freezingof cellsin a cellular suspension and in tissue hasidentified several universal mechanisms ofdamage. t was observed hat when a biologicalmedium begins to freeze, reezing usually startsin the solution that surrounds he cell. Thevolume of the extracellular solution is muchgreater than that of any cell and therefore theprobability of extracellularsolution nucleation smuch greater han hat of intracellularnucleation.When the extracellular solution begins o freezethe two phasemixture of ice and solution s in astate of thermodynamicequilibrium. In contrast,the unfrozen solution inside the cells isthermodynamicallysupercooled. herefore, hereis a difference in Gibbs free energy between hesolution inside the cell and that outside.The cellmembrane has a greater permeability to waterthan o other solutes.Therefore, o equilibrate hedifference between the intracellular andextracellular Gibbs free energy water will leavethe cell. As the cell dehydrates, theconcentrationof solutes nside he cell increases.The increased ntracellular concentration auseschemical damage o intracellularproteins.This isthe first mode of damageduring freezingof cellsin solution or in tissue.The chemicaldamage s afunction of concentrationand emperature.When

the dehydrating ells are rapidly cooled to lowertemperatureShe chemical damage is reduced.The higher the cooling rate (rate of temperaturechange) uring freezing he shorter he period oftime cells are exposed o the lethal combinationof high intracellular concentration and highsubzero emperaturesor shorter periods of time.Therefore, an increase in cooling rate willincrease ell survival. However, water transportacross the cell membrane s time dependentConsequently,when the cooling rates are highenough the intracellular solution will besufficiently supercooled o nucleate and freezerapidly intracellularly. This is the second modeof damageduring freezing. The two modes ofdamage described above occur in cells insolution and in tissue. A typical cell survivalcurve that depicts cell survival as a function ofcooling rate hasan nverseU shape.Cell survivalis low first at low cooling rates, ncreases owardan optimum with an ncrease n cooling rates andthandrops as highercooling rates that lead to theformation of intracellular ice are reached.Eachcell has its own survival curve. These curvesdependon the cell size, membranepermeabilityand intracellular nucleation sites. Thece areadditional modesof freezing damage. RecentlyIshiguro and Rubinsky (1994), (1998) haveshown that cells are also damaged by themechanical nteraction betweenextracellular iceand the cell. In addition, in tissue, thedehydrationof the cells results in a defonnationof the extracelular matrix, destruction ofvasculatureand damage o the functionality ofthe organ. Equationsdescribing the process offreezing n cells and tissuescan be found amongother referencesalso in (Rubinsky and Pegg,1988; Rubinsky, 1989; Bischoff and Rubinsky,1993). Experimentally the most important toolfor studying freezing is the cryomicroscope. Areview of the history of cryomicroscopy can befound n McGrath and Diller (1988).Mechanisms of freeze avoidance and freezetolerance n nature.

In nature, many cold-blooded animalsand plants are exposed to temperaturesbelowfreezing. How do theSe'organisms survive.Apparently, there are two mechanisms, reezeavoidanceand freeze olerance. Freezing can beavoided by reducing the change of phasetempecature f the biological solution. One ofthe mechanisms o reduce the change of phasetemperature s colligative i.e. by increasing theconcentrationof solutes nside the tissue. Thereare animals that during winter generate high


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concentrations of intracellular solutes, such asglycerol (Storey, 1997). Certain insects canproduce in winter high, molar concentrations fglycerol (Storey, 1997; Duman, 1991). Whileavoiding freezing, the high concentrationsofglycerol also stops the life processes n theseorganisms, making them extremely susceptibleto destruction. This mechanism of freezeavoidance s effective in insectsbecause f theirlarge numbers. Other organisms,such as deepocean fish, survive in a thermodynamicallysupercooled state. They can survive in athermodynamically supercooled state becausethere are no nucleationsites,deep n the ocean. Tnucleated these fish would instantaneouslyfreeze. Surface fish and plants, which have tosurvive in the presence f nucleating ce crystalsin the sea water have developed a differentmechanism of freeze avoidance.They use theantifreeze proteins, discussedearlier, to depressthe freezing temperature f their body fluids in anon-colligative way (DeVries, 1971).Antifreezeproteins have the unusual ability to depress hefreezing emperature y up to three ordersofmagnitude more than expected rom coUigativeprinciples. It is not yet understoodhow this isaccomplished. There are theories which claimthat the antifreezeproteins depress he freezingtemperature by binding to certain facets of icecrystals (Raymond and DeVries, 1977). Inaddition to their ability to depress non-colligatively the freezing temperature ntifreezeproteins also have the ability to inhibitrecrystalization once ice forms (Knight et at.1984). Understanding how antifreeze proteinsfunction may have many engineeringapplications. In particular, it could lead to thedesign of low temperature efrigeration cyclesworking with water. Recrystalization nhibitionhas already found applications n the frozen foodindustry where it is desired to inhibit therecrysta1ization f frozen oods.

Certain animals are freeze resistantThe best known freeze resistant animal is thewood frog (RQJUlSylvatica) (Storey. 1997).Studies have shown that as the frog begins tofreeze it produces arge (half mol) quantities ofglucose.The glucose s produced n the iver andis distributed by the blood circulation to theremainder of the body. The frog can survivefreezing to temperaturesn which up to 60% ofthe body fluids are frozen. It cannot, however.survive freezing to lower temperatures.Thefreezing is extracellular and the cells in fact donot freeze, they only dehydrate.The purposeof

the glucose. s to decrease he temperature atwhich 60 % of the body fluids are frozen (Storeyet ai, 1992) In addition, the frogs produce icenucleating proteins that are distributed in theblood vessels nd ensure hat the freezing startsextracellularly Storey, 1997).TEMPERATURES HIGHER THAN mEPHYSIOLOGICAL TEMPERATURE OFWARM-BLOOD ORGANISMS.Research opics in bioheat transfer inthe temperature range higher than thetemperature of warm blooded animals dealprimarily with three topics: high temperatureorganisms,use of high temperature n medicineand accidents t high temperature. The researchon high temperature organisms,(hyperthermophils) has become inportant inrelation to the technique of PCR (polymerasechain reaction) {Huber and Stetter, 1998). Thetechnique f PeR useshigh temperatureso meltthe DNA and o producenew generationof DNAusing enzymes from the high temperatureorganisms usually hyperthermophilic bacteria).While the high temperature ycling of the PCR snot strictly a bioheat transfer problem, devicesfor PeR that do precise thermal cycling are ofinterest Also finding methods to preciselydetermine he temperature at which the DNAstrandmelts s an interesting problem of precisedetectionof phase ransformation n very smallsampleswith high temperature esolution.

A good review of high temperaturebioheatproblemscan be found in Diller (1992).In medicine elevated temperatures are use todestroy undesirable issues n a controlled way.During accidents at elevated temperatures, hetemperaturehas a similar effect on biologicaltissues ike in medical applications. except thathere the application of heat is not controlled.Therefore n medicine t is important to preciselycontrol the application of heat while in the studyof accidents t is important to understand therelation between emperatureand the magnitudeof damage. There are several methods forgeneratinghigh temperatures hyperthermia) intissue in medicine. They include: laser tissueinteraction Galletti. 1997; Cubedduet aI. 1997;Van HiUegersberg, 1997), focused ultrasound(Uchida, et aI. 1998; Albu Barkman, et aI.1998), electrical inductance heating, contactheatingwith hot objects and electrical resistanceheating. There are two areas of engineeringinterest in the study of high temperaturebiological heat transfer. The first area is of a


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biological nature and relates o understanding fthe mechanisms of damage by elevatedtemperatures.The biological studies follow thepioneering work of Henriqueand Moritz (1947).They attempt to develop correlations betweentemperature, ime of exposureand damage n anArhenius type plot. The secondarea relatesdirectly to heat transfer and deals with thepropagationof heat nto tissue and mathematicalmodels of the heat transfer process (Diller,1992). The strong interest in hyperthermictreatmentof malignant umors has generatedherenewed interest in developing models forbioheat transfer discussed n one of the earliersections.CONCLUSIONThis article is, by necessity, very briefintroduction into the many opics of heat ransferin biomedical engineering and biotechnology.There is no doubt that this limited spacecannotdo ustice neither to all the topics n the field norto many researchershat have contributed o thework in the filed. My hope s that this paperwillserve as an introduction into the area ofbiological heat transfer o our colleagues n thegeneral ield of heat ransfer.

Albu Barkman, C., Kirkhom, T., Almquist, L-0., Holmer, N-G., 1998, "Measurements f thethennal focus of an experimental focusedultrasound thermotherapy ystem." InternationalJournal of Hyperthermia,Vol. 14, pp. 383-393.Ananthanarayanan, V.S. 1989, "Antifreezeproteins: structural diversity and mechanismofaction" Life CMm. Reprts.Vol. 7, pp. 1-32.ASHRAE Handbook of Fundamentals 1997)ASHRAE Press.Baish, J.W., Ayyaswamy, P.S., Foster, K.R.,(1986), "Heat transport mechanismsn vasculartissues:a model comparson." of Biomech.Bog.ViI. 108, pp. 324-31.Beattie, G M., Crowe, J H., Lopez, A D., Cirulli,V., Ricordi, C., Hayek, A., 1997, "Trehalose:Acryoprotectant lhat enhances recovery andpreserves funclion of human pancreatic sletsDiabetes,Vol. 46, pp.

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