Adv Physiol Educ 29: 75–82, 2005;
doi:10.1152/advan.00035.2004.
Staying Current
CFTR in cystic fibrosis and cholera: from membrane transport to clinical practice
Barbara E. Goodman and William H. Percy
Division of Basic Biomedical Sciences, University of South Dakota School of Medicine, Vermillion, South Dakota
Submitted 24 August 2004; accepted in final form 9 February 2005
salty sweat; enterocyte; airway epithelia; mucus
BACKGROUND
This information has been used in a 90-min lecture-style
presentation using PowerPoint slides for first year medical
students and has been team taught by the membrane transport
and gastrointestinal physiologists. The presentation occurs either immediately following the membrane transport portion of
the physiology course or at the end of the gastrointestinal
physiology portion of the course, which follows the general
physiology portion (membrane transport and nerve, muscle,
and autonomic physiology). The presentation begins with a
discussion of the involvement of cystic fibrosis (CF) transmembrane conductance regulators (CFTR) transport in the
intestine and how cholera affects intestinal transport and is then
extended to CFTR transport in various organ systems in CF.
Handouts are given to the students with the major points about
the effects of transport changes in each disease state. Basic
information explaining how various treatments work is presented as a logical extension of the membrane transport processes. Student learning objectives for cholera are 1) describe
the transport implications of cholera for both secretory and
absorptive epithelia in the gastrointestinal tract. Differentiate
between which transporters are stimulated and which transporters are inhibited; 2) describe how CFTR is involved in cholera;
and 3) explain how cholera is treated and why. Student learning objectives for CF are 1) describe how CFTR is involved in
the formation of salty sweat in CF patients; 2) explain how
CFTR is involved in increased thickness of mucus in the upper
Address for reprint requests and other correspondence: B. E. Goodman, Division of Basic Biomedical Sciences, Univ. of South Dakota School of Medicine,
414 E. Clark St., Vermillion, SD 57069 (E-mail: [email protected]).
airways of the lungs in CF patients; 3) identify the clinical
problems that commonly lead to death in CF patients; 4)
explain why CF patients might be reproductively sterile; 5)
understand what causes meconium ileus in newborns and
CF-related pancreatic damage in adults; and 6) explain how
cholera and CF might be related.
HISTORY OF CHOLERA
According to the World Health Organization, both Hippocrates (466–377 BCE) and Galen (129–216 CE) described
an illness that appeared to be cholera (37). In addition, a
cholera-like malady has been known throughout the Ganges
River area since antiquity. However, modern understanding of
cholera began in the early 19th century during which researchers into the first pandemic (worldwide epidemic which started
in 1817) made progress toward understanding the causes and
treatment of the disease. In the 1850s, John Snow made the
initial link between transmission of cholera and the consumption of fecally contaminated water by deducing that the spread
of cholera on Broad Street in London was due to raw sewage
in the drinking water (21). When the handle was removed from
the offending water pump, the epidemic ended. Subsequently
in 1883, Robert Koch was the first researcher to identify the
causative organism as Vibrio cholerae 01 (14). Koch described
cholera in the Egyptian city of El Tor where he found ulcerations of the intestinal mucosa of affected individuals and
postulated that an increased leak of plasma into the intestinal
lumen caused their diarrhea (1). Cholera is now defined as any
infection of the gastrointestinal tract by the bacterium V.
cholerae. The organism responsible for the seventh pandemic,
now in progress on earth, is known as V. cholerae 01, biotype
El Tor (37).
PREVALENCE OF CHOLERA
According to data from the World Health Organization, the
current seventh pandemic began in 1961 when epidemic cholera first appeared in Celebes, Indonesia (37). The disease
spread to other countries in eastern Asia including Bangladesh,
India, and the USSR, Iran, and Iraq by 1965–1966. In 1970,
cholera reached West Africa, where it had not been seen for
more than 100 years. In 1991, cholera appeared in Latin
America for the first time in more than a century. Until 1992,
only this one serogroup (V. cholerae serogroup 01) had been
implicated in causing epidemic cholera. Other serogroups had
caused sporadic cases of diarrhea, but not epidemic cholera.
Later in 1992, additional outbreaks of cholera began in India
and Bangladesh caused by a previously unrecognized serogroup of V. cholerae, designated 0139. It is unclear whether
this new serogroup from Southeast Asia will spread to other
regions of the world. It is estimated that 120,000 people die
from cholera worldwide each year (37).
1043-4046/05 $8.00 Copyright © 2005 The American Physiological Society
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Goodman, Barbara E., and William H. Percy. CFTR in cystic
fibrosis and cholera: from membrane transport to clinical practice. Adv
Physiol Educ 29: 75–82, 2005; doi:10.1152/advan.00035.2004.—We
have used a brief analysis of transport via cystic fibrosis (CF)
transmembrane conductance regulators (CFTRs) in various organ
systems to highlight the importance of basic membrane transport
processes across epithelial cells for first-year medical students in
physiology. Because CFTRs are involved in transport both physiologically and pathologically in various systems, we have used this
clinical correlation to analyze how a defective gene leading to defective transport proteins can be directly involved in the symptoms of
cholera and CF. This article is a “Staying Current” approach to
transport via CFTRs including numerous helpful references with
further information for a teaching faculty member. The article follows
our normal presentation which begins with a discussion of the involvement of CFTR transport in the intestine and how cholera affects
intestinal transport, extends to CFTR transport in various organ
systems in CF, and concludes with the logic behind many of the
treatments that improve CF. Student learning objectives are included
to assist in assessment of student understanding of the basic concepts.
Staying Current
76
CFTR IN TEACHING MEMBRANE TRANSPORT
CHOLERA: THE DISEASE
Fig. 1. Schematic of the secretory cells in the small intestine (intestinal crypt
cells), the chemicals that regulate their secretory processes, and some of the
transport proteins involved. The second messengers Ca2⫹ and cAMP are the
links between the membrane receptors on the basolateral side and the effects
on transport on the apical side of these cells. CFTR, cystic fibrosis (CF)
transmembrane conductance regulators. (Modeled after figures in Ref. 15.)
the intestinal lumen through CFTRs located on the luminal
surface of intestinal crypt cells is continuously activated by
intracellular cAMP causing osmotic diarrhea drawing water
into the lumen and leading to the characteristic large volumes
of watery diarrhea.
How is cholera prevented? The major prevention against
epidemic cholera is a safe water supply, good sanitation and
disposal of excreta, and safe food (37). Thus effective control
measures include proper case management, surveillance, reporting, sufficient emergency supplies, and access to health
facilities, whereas ineffective measures include massive efforts
to give antibiotics to entire communities, travel restrictions,
and mass vaccinations. Two new oral vaccines have been
developed that may provide protection for up to 50% of
individuals ⬎5 yr of age for as long as 3 yr after immunization
(38). However, these vaccines only protect against V. cholerae
01, are not effective for sustained protection in children ⬍2
years of age, and may lead to travelers becoming overconfident
and not following simple safety rules.
How is cholera treated?
Attempts to stop the spread of cholera have been variably
successful; however, a major success story has been treatment
efforts that have drastically decreased mortality during the
current pandemic. While cholera used to have a mortality
rate ⬎20%, with the development of oral rehydration therapy
(ORT), the fatality rate for cholera has dropped to about 1%
(14). “ORT is, in fact, the quintessential example of a primary
care intervention: it is a simple technology; it can be made
available in the home, encouraging the involvement of the
mother and other family members to provide effective care;
it is inexpensive; and it effectively treats a common problem.” (7).
In about 1,000 BCE, an Indian physician named Sushruta
recommended that his patients with diarrhea drink large
amounts of tepid water with dissolved rock salt and molasses
(7). Effective treatment of cholera involves rapid replacement
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What causes cholera? Vibrios are one of the most common
organisms in surface waters of the world (both marine and
freshwater) (37). Vibrios are acquired by eating contaminated
food or drinking contaminated water and are transmitted from
person-to-person by the fecal-oral route. V. cholerae is endemic to areas including India, Asia, Africa, the Mediterranean, and more recently, South and Central America, and the
United States. In the Asian countries like India, cholera outbreaks are more prevalent in the late summer and during the
monsoon season (June through August). A type of Vibrio is
associated with shellfish, especially raw oysters. No major
outbreaks of cholera have occurred in the United States since
1911; however, sporadic cases have occurred between 1973
and 1991 and have been mostly associated with the consumption of raw shellfish or shellfish either improperly cooked or
recontaminated after proper cooking. The illness is caused by
ingestion of viable bacteria, which attach to the wall of the
gastrointestinal tract and there produce cholera toxin (37).
Recently, investigators found that the infectivity of cholera
may be enhanced with the organisms more likely to cause
epidemics after various genes that increase survivability are
turned on while passing through the human gut (24).
What are the symptoms (and prognosis) of cholera? Cholera
is an acute illness characterized by sudden onset of watery
diarrhea with a rice water appearance (flakes of mucus and
epithelial cells) and a fishy odor (37). The onset of the illness
has an incubation period varying from 6 h to 5 days. The major
complication of infection by cholera is massive loss of body
fluid due to the diarrhea and subsequent dehydration. It is
possible for a healthy person infected with cholera to become
hypotensive within 1 h of the onset of symptoms and to die
within 2–3 h if untreated. More commonly, the disease
progresses from the first liquid stool to hypovolemic shock
within 4–12 h, with death following in 18 h to several days.
Adults with cholera sometimes excrete more than 1 liter of
fluid every hour for several days (if they are adequately
rehydrated) (37).
How does cholera toxin cause loss of body fluid? Cholera
toxin has binding and enzymatically active subunits that activate the adenylate cyclase system of cells in the intestinal
mucosa leading to increased levels of intracellular cAMP (11).
The effect is dependent on a specific receptor, a monosialoganglioside (GM1), present on the luminal surface of epithelial
cells. The A1 subunit of the toxin, once it enters the cell,
enzymatically transfers ADP-ribose from NAD to the ␣Ssubunit of a stimulatory G protein (GS). When ADP-ribose is
bound, the intrinsic GTPase activity of the ␣S-subunit is
inhibited and the GTP-coupled form is permanently activated.
Once activated, the ␣S-subunit of the GS attaches to the
catalytic subunit of adenylate cyclase and leads to its continuous activation producing abnormally high levels of cAMP.
What specific transport processes are affected by cholera
toxin in the small intestine? Intestinal crypt cells, the primary
secretory cells found in the small intestinal mucosa, respond to
numerous secretagogues including acetylcholine, prostaglandins, and vasoactive intestinal peptide and the second messengers Ca2⫹ and cAMP to lead to chloride secretion through
CFTRs located on the apical (luminal) side of the cells (see
Fig. 1). In the presence of cholera toxin, chloride transport into
Staying Current
CFTR IN TEACHING MEMBRANE TRANSPORT
HISTORY OF CF
The disease now known as CF has long been identified by
health workers as indicated by the old adage from Northern
European folklore “Woe to that child which when kissed on the
forehead tastes salty. He is bewitched and soon must die” (35).
The first major contribution to determining the cause of CF
Fig. 2. Schematic of the absorptive cells in the small intestine (enterocytes)
and the transport proteins involved in net solute transport from the apical side
to the basolateral side of the epithelium. A: types of pathways involved in the
uptake of sodium by the cell. Electrogenic transport pathways (1 and 2) lead
to a net movement of charge across the membrane, and electroneutral transport
pathways (3 and 4) do not contribute to a charge difference. B: Na⫹-substrate
cotransporters are available for the absorption of nutrients from the intestinal
lumen. (Modeled after figures in Ref. 15.)
occurred in 1938 when Dorothy Andersen (after performing
autopsies on infants and children with the disease and reviewing their case histories) provided a comprehensive description
of their symptoms and the changes produced by the disease in
various organs. Andersen noted that there was almost always
destruction of the pancreas accompanied by infection of and
damage to airways in the lungs. Andersen named the disease
“cystic fibrosis of the pancreas.” Subsequently in 1946, researchers deduced that CF was inherited and results from an
autosomal recessive mutation. In 1948, there was a devastating
heat wave in New York City and hospitals saw a disproportionate number of children with CF who had become dehydrated from losing excessive salt in their sweat (26). This
observation by P. A. di Sant’Agnese at Columbia Hospital led
to his presentation of his findings to the American Pediatrics
Society in 1953 and the development of the cornerstone diagnosis for CF, the sweat chloride test.
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of lost ions and fluid either intravenously or orally with a
solution containing sodium chloride, sodium HCO⫺
3 , potassium chloride, and D-glucose. In severe cases, antibiotics may
be used to reduce the volume and duration of diarrhea and the
period of Vibrio excretion leading to infection of others.
Tetracycline is the usual antibiotic of choice, but organismal
resistance to tetracycline is increasing. Oral rehydration therapy involves oral rehydration solution (ORS), the recipe for
which is approved by the World Health Organization (37).
ORS is effective in preventing dehydration, although the fecal
purging rate and duration of diarrhea are unaffected. In the
United States, ORS is only available by prescription because it
has higher sodium levels than other rehydration solutions used
in children and has the potential for severe salt-loading in
children if used for other kinds of secretory diarrheas besides
cholera. The approved recipe for ORS is (in mM) 111 Dglucose, 90 sodium, 20 potassium, 80 chloride, and 30 citrate
(37). Recently, there has been controversy about whether
reduced osmolarity ORS that still has approximately equal
amounts of sodium and D-glucose but lower total osmolarity
might be better for children with cholera in addition to other
diarrheal diseases (10).
Scientific research set the stage for the use of oral rehydration therapy instead of more costly intravenous and sterile
rehydration of cholera patients. Enterocytes (intestinal epithelial cells that participate in the absorption of the breakdown
products of ingested nutrients) have several different types of
transport proteins that facilitate the absorption of sodium and
other organic molecules (see Fig. 2). Sodium is known to enter
epithelial cells from the lumen via epithelial sodium channels
(ENaCs–facilitated diffusion), sodium-dependent substrate cotransporters (secondary active transporters), electroneutral sodium-chloride cotransporters (secondary active transporters),
and coupled sodium/proton exchangers and chloride/HCO⫺
3
exchangers (secondary active transporters). Intracellular cAMP
is known to inhibit both the electroneutral sodium-chloride
cotransporters and the sodium/proton exchangers leading to
decreased cellular uptake of sodium and its transport all the
way across enterocytes. Thus in addition to enhancing chloride
secretion into the intestinal lumen via activating cAMP in the
secretory crypt cells, cholera toxin also inhibits absorption via
electroneutral sodium-chloride cotransport and sodium/proton
exchange out of the intestinal lumen across the absorptive
enterocytes, both of which lead to more osmotically driven
water in the lumen and the resultant watery diarrhea. However,
the sodium-organic substrate (L-amino acids, D-sugars, oligopeptides, etc.) cotransporters are unaffected by cAMP. Thus
the approved recipe for ORS is specifically designed to activate
sodium and substrate absorption out of the intestinal lumen and
to lead to absorption of water molecules osmotically following
these solutes. Therefore, simply drinking ORS can be a viable
and vital way to quickly rehydrate an individual suffering from
infection with cholera.
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CFTR IN TEACHING MEMBRANE TRANSPORT
CF: THE DISEASE
Fig. 3. Schematic hydropathy plot of CFTR in a cell membrane. There are two
membrane-spanning domains (MSD1 and MSD2), two nucleotide-binding
domains (NBD1 and NBD2), and one regulatory (R) domain. Note the sites for
ATP binding and hydrolysis on NBD1 and NBD2 and the phosphorylation
sites on the R domain. In the absence of phosphorylation of the R domain, the
channel is closed and chloride transport ceases. Sequential phosphorylation of
the R domain and subsequent binding of ATP to sites on the nucleotide binding
fold increases the open probability of the channel. (Modeled after Figure 1 in
Ref. 30.)
How are CFTRs in normal, healthy people regulated? Figure 3 shows a schematic of the CFTR and its regulatory sites.
Various studies of CFTR protein function have shown that in
the absence of phosphorylation of the R domain, the channel is
closed and chloride ion transport ceases (12). cAMP stimulates
protein kinase A to phosphorylate one to four serine residues
on the R domain and depending on how many sites are
phosphorylated, the CFTR may be activated to varying degrees. Sequential phosphorylation of at least three sites increases the likelihood of high affinity binding of ATP to the
nucleotide binding fold and enhances the open probability of
the channel. When the nucleotide binding folds on CFTR bind
ATP, a conformational change occurs. ATP hydrolysis has
been shown to be necessary for the conformational change in
the pore and may be located in or near the membrane-spanning
portion of the pore. These changes may lead to as many as
three different gating states of the pore including closed, open
only briefly, and open for long periods of time. In addition, the
open probability and voltage-dependent fast gate of CFTR may
be dependent on tyrosine phosphorylation.
CFTRs are cAMP and protein kinase A regulated chloride
channels, which also regulate numerous other ion channels.
Recent studies (20) have shown that at least 13 different
transporters interact with CFTR by being inhibited or activated
and that a common motif for protein interaction called the PDZ
binding domain is likely involved. Evidence indicates that
CFTRs play important roles in the transcellular secretion of
⫺
HCO⫺
3 by serving as the conductive pathway for HCO3 exit
⫺
across the apical membranes in HCO3 -secreting cells (17).
At the cellular level, how do mutations leading to CF cause
the transport problems in the various organs/systems that are
recognized as symptoms of CF? Briefly, in the lung upper
airways, there is decreased Cl⫺ (and HCO⫺
3 ) secretion into the
lumen, increased Na⫹ absorption through ENaCs out of the
lumen, possibly due to loss of an inhibitory influence of
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According to data from the Cystic Fibrosis Foundation,
more than 10 million white Americans are unknowing, asymptomatic carriers of a defective gene for CF and one of every
3,200 live Caucasian births in the United States has CF (6).
One thousand new cases are diagnosed annually with 30,000
children and adults in the United States having CF. “People
with CF have a variety of symptoms including: very saltytasting skin; persistent coughing, at times with phlegm; wheezing or shortness of breath; an excessive appetite but poor
weight gain; and greasy, bulky stools. Symptoms vary from
person to person due, in part, to the more than 1,300 mutations
of the CF gene.” (6)
What are the major common clinical manifestations of CF?
One of the major problems with CF for many individuals is
blockage of the outflow of digestive enzymes from the exocrine pancreas into the small intestine and the resultant pancreatitis that can lead to the cystic changes in the pancreas
previously noted by Andersen (2). A second major difficulty in
CF (and probably the most devastating for the long-term health
of the individual) is the accumulation of heavy, dehydrated
mucus in airways and the resultant changes in the capability of
the lungs to fight infections. A third major change that leads to
the ability to use the sweat chloride test as the standard
diagnostic test for the disease is the abnormally salty sweat of
CF patients. A fourth clinical manifestation is sterility in both
males and females that is particularly prevalent and may be
irreversible in males. A fifth manifestation that may be recognized less frequently than some of the others is abnormal
secretion in the small intestine. This aspect has been previously
described in detail in the sections on cholera. These common
pathophysiological problems lead to the several major symptoms of CF described previously.
How do the various CF mutations affect the function of
CFTR proteins? The various mutations (⬎1,300) that have
been shown to cause CF have been categorized into four
classes (36). Class I mutations cause defective protein production with a total loss of functional CFTRs. One defect produces
a truncated CFTR due to premature stop mutants. This defect
may be corrected by certain antibiotics that bypass the shortened protein to make full-length functional CFTR. These
antibiotics have restored 25–35% CFTR protein function when
only 10% correction appears to be enough for noticeable
improvement. Class II mutations cause defective protein processing leading to CFTR that is not in its correct location in the
cell or that is different from CFTRs in normal individuals (less
glycoproteins and gangliosides on the cell surface in CF cells).
The most common mutation found in 70% of CF patients (the
⌬F508 deletion) is one of the Class II mutations. Class III
mutations cause defective regulation of channel opening of
CFTR by changes in the nucleotide binding fold or regulatory
(R) domain of CFTR (see Fig. 3) (30). Class IV mutations
cause defective ion conduction through CFTRs. These mutations are in membrane-spanning domains of the protein and
thus affect the pore that normally allows ion fluxes. Class I and
II mutations generally lead to the more serious phenotype of
the disease and have accompanying pancreatic insufficiency.
Class III and IV mutations generally lead to a less serious
phenotype with normal pancreatic function.
Staying Current
CFTR IN TEACHING MEMBRANE TRANSPORT
Fig. 4. This model of NaCl transport across ductal cells in sweat glands of
normal and CF individuals demonstrates the differences in salt reabsorption
there that leads to the elevated sweat NaCl in CF patients. Na⫹ ions utilize
ENaCs to enter the cells from the luminal side and Na⫹-K⫹-ATPases to leave
the cells on the blood side. Cl⫺ ions predominantly utilize CFTRs to enter the
cells from the luminal side and other Cl⫺ transporters to leave the cells on the
blood side. In CF, the lack of functional CFTRs on the luminal side of the cell
compromises the movement of cationic Na⫹ ions and its accompanying
anionic Cl⫺ ions leading to less reabsorption of salt out of the lumen. Salt
concentrations in CF sweat can be up to 3–5 times normal. (Modeled after
Figure 1 in Ref. 27.)
destruction and pathogenesis. Constipation and distal intestinal
obstruction syndrome also occur in older CF patients (CF
patients have also been shown to have loss of CFTR function
in their colon) with rectal prolapse being common (usually
before pancreatic enzyme therapy). All of these situations are
based on lack of Cl⫺ secretion. These intestinal CFTRs are the
same ones involved in excessive chloride secretion under the
effects of cholera toxin. Recently, investigators (4) have shown
that while prostaglandin-stimulated electrolyte secretion in the
intestine is decreased in CF patients, prostaglandin inhibition
of neutral sodium absorption is unaffected. Thus this antiabsorptive response may contribute to the low level of intestinal
disease in CF patients. In the reproductive tract, absence of fine
ducts (i.e., vas deferens) developmentally renders 95% of CF
males infertile. The speculation is that the faulty development
is due to blockage of the ductules in utero so that most CF
males are aspermic or hypospermic. CF women show a significantly lower fertility rate due to physical mucus plugs in the
Fallopian tubes which block sperm from fertilizing ova. These
transport problems are based on the lack of Cl⫺ secretion.
Figure 5 summarizes the important transport implications of
CFTR in various organs. Thus in multiple organs, ion and
water imbalance contributes to the pathophysiology by failing
to dilute the native mucus sufficiently to maintain its normal
fluidity. This dehydrated mucus becomes very viscous and
relatively immovable.
Therefore, what are the basic goals of clinical studies to help
treat patients with CF? The major problems that need to be
addressed to assist CF patients to live longer are to 1) fight the
chronic lung infections that inevitably lead to lung damage and
eventually death, 2) thin the dangerously thick CF mucus
particularly in the upper airways of the lungs, 3) reduce the
inflammatory response particularly in the lungs which leads to
tissue destruction, and ultimately to be able to 4) correct the
basic cellular defect (i.e., gene therapy or selective drugs).
Thus various efforts are occurring to use or develop therapies
for treating CF that 1) correct the abnormal gene via genetic
manipulation, 2) correct the abnormal protein via protein
rescue/activation, 3) alter ion transport and abnormal mucus
secretion via selective drugs, 4) decrease infection and inflammation leading to tissue destruction via anti-inflammatory and
anti-infective agents, or 5) overcome organ destruction leading
to respiratory failure via lung transplantation. In summary, the
various stages of the lung pathogenesis seen in CF are developed from the abnormal gene that translates the abnormal
protein that exhibits abnormal transport of ions, which leads to
changes in the luminal fluid, causing infection followed by
inflammation and finally destruction of lung tissue. The only
recourse that one might have may be lung transplantation.
Cells in the upper airways of the lungs of CF patients secrete
a mucus layer that is dehydrated, thicker, stickier, and more
difficult to clear than in healthy individuals. This layer provides a hospitable environment for pathogens (particularly
Pseudomonas aeruginosa and Staphylococcus aureus), which
leads to the recurrent infections and progressive loss of respiratory function due to bronchiectasis (destruction of muscular
and elastic components of airway walls due to chronic inflammation). Now that digestive problems in CF patients are
relatively easily managed if treated before induction of damage
to the pancreas, lung impairment in CF patients causes ⬎90%
of the disability and death due to the disease. How can ion
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functional CFTRs and/or ENaCs stimulating the activity of
CFTR up to sixfold (18), and normal, but not CF, CFTR may
activate water permeability through aquaporins there (28). In
sweat glands, sweat is normally produced at the base of the
glands and then passes through a narrow duct in which reabsorption of salt occurs [in CF, abnormal Cl⫺ absorption out of
the duct via defective CFTRs leads to excessive Na⫹ and Cl⫺
(3–5 times normal concentration) in sweat] (Fig. 4 and Ref.
27). In the liver of CF patients, plugging of small bile ducts
based on lack of Cl⫺ secretion impedes digestion and disrupts
liver function; however, liver involvement may be asymptomatic and may be slowly or not at all progressive. In the
pancreas, occlusion of ducts (and lack of HCO⫺
3 ) prevents
pancreatic enzymes from reaching the lumen of the intestine
and may lead to premature activation of digestive enzymes
inside the pancreas causing pancreatitis that may also cause
diabetes mellitus. Lack of digestive enzymes in the intestinal
lumen can cause symptoms including frequent, loose, oily, and
malodorous stools caused by steatorrhea (fat in stool due to
lack of active pancreatic lipases). Approximately 80–85% of
CF patients have pancreatic insufficiency, which leads to
decreased availability of both digestive enzymes and HCO⫺
3 in
the intestinal lumen and clinical maldigestion and malabsorption based on the lack of Cl⫺ secretion. Fortunately, oral
pancreatic enzyme supplementation after meals appears to
override the digestive problems. In the intestine, obstruction of
the gut by a “thick, dehydrated, rubbery, tarry, tenacious,
mucoid plug” (26) leads to a condition known as meconium
ileus that necessitates surgery in 10% of newborns with CF. In
addition, knockout mice with no CFTRs (model developed in
1992) will die from intestinal obstruction by 5 wk of age, and
their intestines exhibit an inflammatory state leading to an
innate immune response, which may cause additional tissue
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CFTR IN TEACHING MEMBRANE TRANSPORT
transport and fluid balance across the upper airway epithelium
be improved in CF patients? Better dilution of the thick, sticky
mucus lining the upper airways will help lower the chances of
various bacteria colonizing the upper airways and leading to
destruction of lung tissue. This enhanced dilution can be
facilitated by simply manipulating concentrations of salts and
water across airway epithelia. In other words, net solute movement is accompanied by net water movement by osmosis (see
Fig. 6). There are several possible ways to modify salt (and
water) movement across the airway epithelial cells. One way
would be to use aerosolized amiloride to inhibit ENaCs and
thereby decrease sodium reabsorption out of the lumen of the
upper airways (13). Less sodium leaving means less water
leaving and leads to a larger airway liquid volume. Another
way would be to increase chloride secretion either through the
CFTR (using various pharmacological agents to rescue the
Fig. 6. This schematic shows the location of various transport proteins in the
epithelia of the upper airways of the lungs and how they may be regulated to
improve solute and water balance in airway fluid in CF patients.
function of mutant CFTR) (13); through other chloride channels (ATP, UTP, and UDP may work via various purinergic
receptors to open other Ca2⫹-activated chloride channels) (19);
or by facilitating K⫹ secretion (activation of basolateral Ca2⫹activated K⫹ channels by benzimidazolone may promote a
sustained Cl⫺ secretory response by providing the accompanying cations needed for net transport) (9). All of these methods lead to more ion secretion into the lumen and more airway
liquid volume. Another way might be to increase HCO⫺
3
secretion because functional CFTRs may facilitate Cl⫺-coupled HCO⫺
3 transport, causing abnormally acid luminal fluid in
CF patients, which causes precipitation of mucins, plugs various ducts, and facilitates bacterial infections (3).
Although the manipulations described above would likely
lead to changes in water levels in airway liquid and may assist
in diluting the airway liquid so that it is less thick and
tenacious, are there other more indirect methods that can
change the characteristics of the airway liquid? The traditional
treatment for CF patients has been postural drainage and chest
percussion (chest physical therapy) where the patients lie with
their heads tilted downward and someone pounds gently and
rapidly on their back and chest to physically loosen and clear
the secretions in the lung airways. More recently, the purulent
airway secretions that have become viscoelastic due to polymerized DNA from the degenerating leukocytes in the lung
airways have been treated with recombinant human deoxyribonuclease, thereby reducing the viscosity of the sputum (8).
Recurrent infections in the lungs lead to a chronic burden of
neutrophils and their resultant neutrophil elastase in upper
airways. Thus secretory leukoprotease inhibitor (produced by
cells of the mucosal surface) can be used to protect against the
assault of neutrophil elastase on airway epithelial cells (23).
Alternatively, excess neutrophil elastase can be diminished by
high-dose ibuprofen (which decreases neutrophil aggregation
and adherence) taken by patients with mild lung disease for a
long time (34). Neutralizing the destructive neutrophil elastase
by any of these methods may slow the progression of the lung
epithelial damage in CF patients. Similarly it has been shown
that CF sputum samples contain filamentous actin; thus human
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Fig. 5. This diagram summarizes the effects of altered CFTR function in CF in several important organ systems.
Staying Current
CFTR IN TEACHING MEMBRANE TRANSPORT
CFTRs in the cell membrane (as in the ⌬F508 mutation) there
is less infection (25). Thus diminished numbers of functional
CFTRs in CF heterozygotes may have decreased their susceptibility to typhoid fever and preserved the gene at high numbers
in the population.
CFTR AND CHOLERA REVISITED
Secretory diarrhea is the leading cause of infant death and
has devastating effects on adults in developing countries. It is
now known that CFTR is the transporter required for chloride
(and accompanying water) secretion both in the intestines and
in the upper airways of the lungs. Mutations in CFTR lead to
various expressions of CF which is still a lethal genetic disease.
Ma et al. (22) have utilized high throughput screening techniques to identify high-affinity blockers of cholera toxininduced intestinal fluid secretion via CFTRs. In fact, recently a
thiazolidinone-type CFTR blocker has been shown to reduce
cholera toxin-induced ion and fluid secretion in mouse intestine
(31). Thus CFTR inhibitors are being developed as drugs to
block intestinal fluid secretion in cholera and other secretory
diarrheas. In addition, these compounds might be useful to
establish the CF phenotype in cells and animal models to
investigate more specifically the mechanisms of lung tissue
destruction in patients with CF.
Thus a brief investigation into how CFTRs are involved in
transport aberrations in both infections by V. cholerae and the
genetic disease CF can help teach students about direct relevance of membrane transport physiological principles to human disease. In addition, the transport principles represented in
these two examples are amazingly logical and relevant to basic
concepts of diffusion and osmosis already understood by the
students.
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plasma gelsolin (protein that severs actin filaments) has also
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The ultimate goal for CF patients would be a cure by
correcting or replacing the genetic or protein defect (16, 33).
What about gene therapy for CF patients? Human gene therapy
studies for CF began in 1993 with investigations into their
safety and with the first aerosolized CF gene therapy protocol
used in 1995. Recent evidence shows that only 6–10% of cells
with functional CFTRs will correct the Cl⫺ transport defect but
100% of cells need functional CFTRs to correct the Na⫹
transport defect. Insertion of normal CFTRs into airway cells
would be an ideal way to cure the disease; however, finding a
good vector for insertion of the gene, which will lead to
long-term improvement, has been difficult. Viral vectors (adenovirus, modified adenoviruses, retrovirus) that can be genetically modified to be replication deficient with CFTR cDNA
plus a promoter are highly efficient but immunogenic and
therefore only effective in the short term. Nonviral vectors
(cationic liposome-DNA complexes or molecular conjugates)
have greater safety and ease of preparation but low transduction efficiency. Molecular conjugates can be constructed so
they are targeted to airway epithelial cells via specific receptormediated endocytosis. Since most cells in epithelial tissue are
replaced every few months, gene therapy would still have to be
administered multiple times a year.
Why is the disease CF so prevalent in humans? There have
been several theories about the high incidence of mutations
leading to CF in Northern Europeans and their descendants.
One theory ties together the relationships between CF and
cholera. This theory states that heterozygotes in the population
have ⬃50% functional CFTRs compared with healthy individuals and have a selective advantage in resisting death by
cholera because of lower enhanced chloride secretory diarrhea
during infection with the disease. Thus these individuals had
heterozygote advantage and were more likely to pass on the
gene for CF to their offspring. This theory, while reasonable
and interesting, does not totally explain the inheritance of CF,
because cholera epidemics did not strike Northern Europe until
the 19th century. However, diarrheal diseases caused by other
organisms, such as Escherichia coli, have affected European
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genetic advantage in ileal or colonic epithelia of the null CF
mouse for acute responses to secretory diarrhea (5). Another
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were searching for evidence about other adult lung diseases in
CF heterozygotes and found that only a handful had asthma
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asthma. Thus there may be some as yet unknown link between
defective chloride transport and protection against asthma. A
third theory is related to increased resistance to infectious
diseases that helps to maintain the mutant CFTR alleles at high
levels. It has been shown that Salmonella typhi uses CFTR to
enter and infect cells and that when there are fewer functional
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