xt7sf7667h0r https://exploreuk.uky.edu/dips/xt7sf7667h0r/data/mets.xml University of Kentucky. Office of the Vice President for Research and Graduate Studies Kentucky University of Kentucky. Office of the Vice President for Research and Graduate Studies 1993 volumes. Call Number: LD2762.9 .K38c. Accession Number: 2018ua049 journals LD2762.9 .K38c English Office of the Vice President for Research and Graduate Studies Contact the Special Collections Research Center for information regarding rights and use of this collection University of Kentucky Odyssey Magazine Research -- Kentucky -- Periodicals Education -- Research -- Periodicals Research -- Kentucky Odyssey: The Magazine of University of Kentucky Research,Summer/Fall 1993 text images Odyssey: The Magazine of University of Kentucky Research,Summer/Fall 1993 1993 1993 2026 true xt7sf7667h0r section xt7sf7667h0r

. THE NLXGAZINE OF UNIVERSITY OF KENTUCKY RESEARCH 7 . SUMMERi/FVAVLLWIQQS .

A Ncw Dinwnsion in the Operating R( )()1112
EH) Modvls as Visual Aids
in Rcmnslrucliw Surgery

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Innqu'xx IILII [III’ 11mm |\II\ Im~ mmlv
ln\\AIholic drinkers, So the suggesv
tion is that metabolism is primed to eliminate
alcohol and other calories."

\Vhat happens to other food sources when
alcohol is added to the diet? Another aspect of

Joltnstone‘s project will examine the relatiotr

ship between normal or moderate drinkingI and
the intake of protein. carliohydrates. fat and
dietary cholesterol,

Hypoﬂwsized Relationship for Non-Alcoholirs behuom Alcohol Comumption and Body lr’Voz'ghl

Body Weight Index

I Males
[:l Females

Abstainer Low

Moderate Heavier

Alcohol Consumption

drinkers [and to waste
than (1'0 modm‘ate and

non—alrohalir (172' ItlfﬂiiS‘.

Plimz'natr alcohol and
other calories. ”

“There has been a specific suggestion that
male drinkers tend to he heayy meat eaters. A
hamburger and steak-type diet is Very common
among heayy drinkers. \Ve also know that
alcohol is yery similar to carbohydrates in many
ways. so carlxrhydrates o\'erall tend to decline
in the diet with alcohol."

It helps to associate alcohol. like food. \\ ith
appetite. says
loltnstone. rather
than with addio
tion. “\\"e find clinr
. _ call\. wlten we
all types 0/ (alarm, mare look at alcoholic
' ‘ populations. that
as people try to
abstain. they start
to cra\'e sugar par—
80 [hr suggestion as that “““1/‘11‘13‘1‘7‘1"‘1“

s . sources of carbt»
hydrates. And. in
fact. we think car—
bohydrates may
modify their crayv
ing. Appetite regtt—
lation. which
seems to he unbalanced in some way for
alcoholics. may be in part a function of the
relationship het\\ een alcohol and sugars and
other carht rhydrates. So there may he a suggese
tion as far as the de\'el( rpment ofalcoholism that
there is some role t>fcarlxrhydrate food sources
in that process.”

Thus. one implication for johnstone‘s find
ings might he the deyelopment of nutritional
therapies for alcoholics. “If we understand
alct )h< )1 consumption and other appetite hehay—
iors. we may understand how the craving for
alcohol deyelops." he says.

johnstone anticipates a continued high inter—
est iii the subject ofalcohol and diet, especially
in light of the recent controyersies about the
role of alcohol in breast cancer and heart
disease. "If this research has sortie kind of
significance for understanding how alcoholics
deyelop an uncontrolled appetite for alcohol.
that's fine. But i think the implications for the
general population of normal drinkers is also
very important. particularly when it comes to
understanding obesity and cltronic diseases".

“80W people my heavy

tmtaboltwn is primed to

BeerInamsesRisktnyoﬂiLung
wrdBrmstCanoerinOldemnen

B eer may he a factor for increased risk ofhoth lung and breast cancer
among pt>stmenopausal women. according to two recent epide-
miological studies.

in the studies supported by the National Cancer Institute. nearly
42.000 women in lowa filled out a questionnaire that included
questions on nutrition. alcohol consumption and other he: lth informa-
tion. After analyzing the data. the scientists discovered that postmeno-
pausal women who drank one or more beers a week were at double
the risk for lung cancer. compared to women who drank less than an
average of one beer a week. The findings, howeyer. only held true for
postmenopausal women who smoked. the researchers note.

"We don‘t really know why alcohol. and specifically beer. increases
the risk for lung cancer for female smokers." says Thomas Sellers. one
of the epidemiologists who helped with the studies. which were
conducted at the [Tniyersity of Minnesota. “I ioweyer. we do have some
theories." says Sellers. He hypothesizes that beer depletes the stores of
vitamin A in the liver Vitamin A is an antioxidant. a chemical that
neutralizes the activities of free radicals. substances which are potential
carcinogens. Beer may also exacerbate the deleterious effects of
smoking.

In a second study. the Minnesota epidemiologists found that women
who drank one beer a day had a 50 percent greater risk for hr east
cancer than women who drank less than one beer a day. Among
women who were also taking estrogen supplements. this risk rose to
80 percent.

“Beer may stimulate pr‘olactin. a hormone that causes the mammary
gland cells to divide more rapidly." explains Sellers. Cells that diyide
more rapidly are considered more likely to become malignant.

This research was published in the September 1992 Annals of
Izpidemz'o/ogt:

—‘[l((1)'[‘tIIkP’n/Iﬂl‘g,ﬁ‘mll NIH Healthline, illart‘h/A/nil I 993

§ ODYSSEY

Inn/41' WW

transgenic manipulation:

a process which involves altering an
animal ’5 genetic system by the
injection ofsome other mammalian
genetic material

In September of 1991. University of Kentucky
researcher Brett Spear obseived some veiy
unusual behavior by one of his transgenic lab
mice. Instead of scarnpering around in its cage.
the mouse was pulling itself along. with some
difficulty. by its front legs.

"There wasn‘t a complete loss of muscle
control in this mouse—he would pull his back
legs away if I touched them—but there was a
noticeable paralysis." says Spear. a molecular
biologist who has been at UK since 1989. "We
were curious why this had happened. We
wanted to know if this was a result of our
genetic manipulations with these animals."

Spear and others at [TR were intrigued by this
discovery because it has important implications
for researchers trying to discover the causes of
degenerative diseases of the aging neryous
system. such as Alzheimer‘s disease. Parkinson‘s
t“ disease and amyotrophic lateral sclerosis
(ALS). What all three of these diseases
have in common is that
they are inevitably

progressive: the
cells just shrivel
and disap—
pear. and

ODYSSEY 6

Building a Better Mouse
to Trap Genetic Secrets

the patient eventually dies from the disease.

"This animal looked to have symptoms simi—
lar to those of people who have motor—neuron
degenerative disease." says Hi neurologist
Edward Kasarskis. who examined the mouse
soon after Spear noticed its disability. "Although
we dont know yet in what ways this discovery
may be important. it is clearly a significant
development. Ifthe disease is found to parallel
ALS. for example. Spear's work may become
veiy important as it relates to our understanding
of this disease." The progressively paralyzing
disease ALS became known as Lou Gehrig‘s
disease for the New York Yankees star who
died of the illness in 1941. Faulty genes are
blamed for about five to 10 percent of ALS
cases. and the disease affects an estimated
§().00() Americans at any given time. according
to the ALS Association.

Spear and his associates were extremely
interested in the possibilities of this finding. As
so often happens in research. however. they
aren't certain just where they went right.

“We certainly weren‘t tiying to make an
animal that would develop this hind—limb pa—
ralysis." says Spear. “This was totally unex-
pected. But we are trying to take advantage of
it to understand something about mouse devel—
opment as well as human genetic diseases."
Only one group of mice being used in Spears
transgenic research has shown hind—limb pa-
ralysis. To date, of the 587’ pups that have been
born in this group. 104 have had the defect.

Along with researchers in the Department of
Anatomy and Neurology. Spear has looked at
cells within the spinal cord of these animals
using electron microscopy techniques. What

they‘ve found is that the motor neurons degen—
erate in the same way that those of people
deteriorate that have similar degeneratiye dis
eases.

The original line of research. funded for fiye
years by the \itti<)ri;tl Institutes of Health.
in\'( )lyed genetic manipulation of mice in order
to understand how genes are switched on in the
li\'er. Spears research team. which includes
graduate students 'l‘ennore Ramesh and Ke Li.
and technicians Amy Ileis and Mike Green. is
now working intensely in both areas.

The Tm nsgen 11- rlIouse: Bar/{gamma

Recent adyances in biotechnology haye made
it possible to use mice to study a yariety of
diseases. in 1981) and 1981. the introduction of
cloned DNA into the mouse through microin—
jection of fertilized eggs was achieved. This
method attracted widespread attention when it
was found that the procedure noticeably
changed the mouses hereditary makeatp.

(iene expression was first demonstrated in
1981 by the dramatic growth oftransgenic mice

()rIe pipette ([e/I) holds the mouse embryo 1' n plaee wilh a slight
vaeaum as [he ol/zerpipelte (rig/II), (In ullraf/ine needle, is used to
iujeel a small amount (if/”Warez! I).\H irzlo the ("1111's [)ronza'lei.

harboring the human growth hormone gene,
These "supermice."
nearly twice as large

“Our major goal,

as normal mice. capr

tured headlines

of (011736, "I, l00k_ around the world.
lng‘ai the hlnd_leg‘ SithL‘ [11:11 littlc.

. "transgenics"—the
ljaﬂllySZS oft/1656 creation ofan animal
, '. . , whichcontains genes
171366, 25 t0 identlﬁ‘ from another organ—

. ism has helped to
the pamwlargme revolutionize the
we’re disrupting in
the mouse... ”

study of genetics and
illness.

"Transgenic mice
haye been made that
mimic certain human disorders." says Spear.
“Mice have been generated that are genetically
programmed to haye cancer. arthritis. obesity.
diabetes. and other diseases."

Thousands ofyarieties ofthese "guinea mice“
are now proyiding models for testing possible
treatments. In a I SiVea'sci “Yale/Reportarticle

which appeared in Noyember of 1991. Charlene
Crabb reports that scientists haye protected
mice from cardit)yascular disease by giying
them genes for high—density lipoproteins. or
HDL. the "good" cholesterol that has long been
suspected of reducing the buildup of fatty tissue
in the arteries.

A HarVard Medical School team has patented
transgenics that reliably develop cancers. and
researchers in California have created mice
carrying the gene for beta-amyloid protein. the
principal component of the plaques riddling the
brains ofpeople withAlzheimer‘s. Says Richard
Palmiter of the l'niyersity of \‘é’ashington. a
pioneer in the field. "The transgenic mouse is a
wonderful experimental tool to study almost
any question of genetics."

To try to understand disease processes. the
mouse has been a frequent choice for research-
ers because it bears a striking biological similar:

cmz/mrrer/ rm [rage N

- ODYSSEY

ODYSSEY 8

'I he transgenic mouse research team i nol tales
(from left [0 rig/1.!)Biell Spear, principal investiga—
lor; Ke Li, from China; technician Amy Ellis, from
Danville, Kentucky; and 'I‘ennore Ra mesh, _ /rom
India.

ity to humans. “There are very few animal
models we can use to study neurtrdegeneratiye
disorders." Spear says. “For mammalian sysi
tems. the mouse is probably the best choice to
work with. Its nervous system is highly analo-
gous to the human neryous system, it‘s a
relatively small animal. and mice breed faster
than any other mammal Understanding how
biological systems function in mice is directly
applicable to our understanding of gene func—
tion in humans."

But why use mice at all? Why not utilize the
technology of computer modeling instead?

“Computer modeling can go only so far," says
[7K Veterinarian Ward Crowe. "It's only as
sophisticated as the human mind that programs
it." Nothing. at this time. Crowe says. can
parallel the highly complex biological system of
the mouse. “Perhaps one day researchers will
have an alternatiye so that animal models won‘t
be necessary," Crowe adds. “Until then. it is
essential that laboratory animals be used for the
betterment of both humans and animals."

How Transgenic Mire Are illade

Spear explains that the creation ol'a transgenic
mouse starts with DNA. the basic molecular
material that determines heredity in animals.
The researchers use "recombinant DNA." which
is prepared in the laboratory by breaking tip and
splicing DNA from seyeral species. “We do this
in a petri dish. in bacteria. which allows us to
manipulate the DIVA in yarious ways." Spear
says.

The next step inyolyes setting up mouse
matings. The day before the researchers are
ready to do a particular experiment. they pair off
a number of normal mice. placing a male and a
female mouse together in a cage. "'l‘welye
hours or so later. we identify which females
have mated by the presence of a 'copulation
plug.” Spear explains. This mucousilike plug.
formed in the presence of semen. lets the
researchers know which females haye mated.

After a period of gestation. those female mice
are then painlessly euthanized and the embryos
remoyed. At this time the fertili/ed embryos are
placed into a petri dish.

The next step in\'olyes injecting the prepared
DNA into these embryos. This is where senior
research analyst Mike Green comes in. (ireen
performs the delicate task of microiniecting the
embryos in UKs Transgenic Mouse Facility. a
ﬁOO-square—foot lab in the Combs Cancer Build—
ing.

“Ifwe can identify this gene, ”Spear
says, “it would be relatively easy to
identijy the parallel gene in humans
involved in the nervous system.
Hopefully, we will eventually come to
understand how this gene functions
and why . . . ”

Green sits at what appears to be a standard
micrr )SCOPC. except for the two extended handles
on either side of it. “These are micrtHnanipulav
tors." Green explains "They control two lines
that come into the microscope. Each line holds
and controls a pipette—a long. thin glass cylin-
der." The pipette on the left is a needle with a
finely polished end so that with a slight vacuum
pressure it holds the embryo in place, The other
pipette is an ultrarfine needle. through which a
small amount of prepared DNA is injected. “This
takes extensive training and a really steady pair
of hands." Spear says.

The embryos. now containing the genetic
information Spear wishes to implant. are ready
to be surgically placed into other female mice.
To maximize the possibility that the embryo cells
will grow and divide properly. Spear and his staff
set up a second round of matings. This time.
though. females that are hormonally ready to
nrate are paired up with vasectomized males.

“\‘i'e do this." Spear explains. "so that the
females will actively mate with none of their
eggs becoming fertilized. But they are physi~
ologically ready to accept fertilized embryos."
The genetically engineered embryos are then
surgically implanted by Green into the oviducts
of these female mice, 'l‘wenty—one days later. a
litter of pups is born. llairless and pink at birth.
each resembles a small jelly bean. And for two
weeks all of them appear to be completely
normal baby mice.

\V'hen these mice are two weeks old. research—
ers take a small tail biopsy to check whether or
not the injected genetic material exists inside
each mouse. Any part of the body could be
biopsied. since ideally each cell contains the
transgenic information. but a tiny piece ofthe tail
is taken because. Spear says. “they have very
little feeling there."

Because the injected DNA may be lost before
becoming part of a chromosome. only 10 to 20
percent of the mice biopsied will contain the
genetic material injected. However. the good
news for the researchers is that once these 10 to
20 percent have been identified. each of these
can start a "founder line." passing this DNA on

conlrmred on page [H

\
5

Smith“ research analysl Mike Green in UK ’s Transgenic Mousp Facility,

“midis/red in I 98 9

9 ODYSSEY

ODYSSEY 1f)

Once this gene is identified,

then studies can be done tofuither
understand human genetic disarders
that aﬂect the nervous system.

to their offspring and eyentually generating
hundreds ofoffspring who contain the genetic
triaterial.

Although a great deal of work will be neces
sary to identify the gene responsible for the
neurssihle solution
to try to re\erse the man's seemingly hopeless
e1mditil1 )gy ol' sterei>lithographyi

"’l‘he images \\ e had gotten ol‘the mans lace
lri nn (i.»\'l' scans and \lagnetie Resonanee Image
ing \yere dil'l'it'ult to interpret." reealls lleniy
\ast'one/_ a [K plastit' surgeon “hose spe~
eialty is eranii rlaeial surgery and mierosurgeiy
"So I ealled l)r. Knapp and asked him to make
.1 computer model ol' the mandihle. a threer
Llllllt‘ll\l(ll1;ll model ol'the missing pieee l eould
e\amine heliore the surgery.”

Knapp. dlt‘et‘toi‘11ll'K‘s(lentet'lin'lilomedk
eal lingineering. \\as eonlident sueh a tnodel
eould he eonstrueted through the eomputer—
i/.ed steret )li1111 >graphy \\ ork heing Ll( me at l 'K‘s
(enter it 11' R1 iht )ties and \lanulaeturing Systems
((TRXlSl. Stere<>lithography is a system that
enahles engineers to dralt an ohieet on a
1‘1 imputer sereen. then \\;1teh as a maehine
ereates a lullAsi/ed. threerdimensk)nal plastic
model ol‘ the object.

\'ase< )ne/ gaye Knapp some (I;\'l' seans ol'the
patients head. and Knapp‘s' group was off and
running. The lirst step \\'as to ei‘eate the three
dimensional model of the head. "\Vith this
pr1 >eess." Knapp explains. "we get a 5—D image,
look at the undamaged side ol the laee#tlie
uninjured jaw in this ease—and llip the image
met." The llipped—m er image is What the jaw

mull/11ml of] [urge l.’

A New Dimension
in the Operating Room

(Jun/(ts Knapp, Ilil‘m'lm‘ (1/ l 'l\”.\‘ (killer/hr Bionzvdiml 1ingimwing. \ils behind
s17 W11! (my/1'1" and Immlm mode/s built (1/ l 'K and um] (Is s‘lngim/ (lids. SIM/1
mode/s Ill/mu [he Viagra); {11 .w'. ins/171d tl/‘I'nmgim', (/11) Ill/rd (linu’nsimz.

ll ODYSSEY

ODYSSEY 12

“This equipment was obviously built for
industrial applications, butDr. Knapp and
Dr. Vasconez saw that it could he used for
medical purposes. We did the plastiejaw in
about seven hours. ”

looked like before it was damaged.

Next. the computer image was sent to CRMS,
where it was used to guide a laser system.
Though the next step#the “growing" of the
jaw~might seem like science fiction, Knapp
characterizes it as a "pretty str