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Journal of Culture Collections
National Bank for Industrial Microorganisms and Cell Cultures
ISSN: 1310-8360
Vol. 4, Num. 1, 2005, pp. 3-16
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Journal of
Culture Collections, Volume 4, No. 1, 2004-2005, pp. 3-16
MCCOY AND
MCCOY-PLOVDIV CELL LINES IN EXPERIMENTAL AND DIAGNOSTIC PRACTICE PAST,
PRESENT AND PERSPECTIVES
Marian Draganov1*, Marianna Murdjeva2
and Teodora Michailova-Topalska1
1The University of Plovdiv, Developmental Biology Department, 24 Tzar Assen Str.,
4000 Plovdiv, Bulgaria. e-mail: mmd@pu.acad.bg;
2Medical University, Faculty of Medicine, Department of Microbiology and Immunology,
15A V. Aprilov Str., 4000 Plovdiv, Bulgaria
Code Number: cc05001
Summary
The McCoy
cell line has almost 50 years history. The cells are widely applied in the
diagnostics and culture of various microorganisms with medical importance. The
cell line is included in laboratory and diagnostic tests which are the basis
for study of interactions between various pathogens and host cells leading to
cytotoxic damage or cell death. With its importance in experimental and
diagnostic laboratories, McCoy cell line is among the most popular cell
cultures HeLa, HEp-2, Vero, CaCo-2, 3T3, MDCK. An alternative for application
of McCoy is the serum-free strain McCoy-Plovdiv. It is cultured in completely
defined, serum- and protein-free medium. It keeps the properties of the
parental line but also offers new opportunities.
Historical data
McCoy cell line is created in the Tissue Culture
Laboratory, Department of Anatomy, the University
of ,
Galveston,
Texas.
In 1957 Pomerat et al. [116] published own investigations on radiation influence
upon cells in tissue culture conditions. McCoy cells were among the available
cell cultures and they were announced for the first time in research
literature: Synovial fluid - McC. A strain developed in this laboratory in
October 1955 from cells in synovial fluid from the knee joint of a patient with
a diagnosis of degenerative arthritis.
By chromosomal
examination of eight cell strains Hsu et al. [63] found that even in more
recently established cell lines such as McCoy, which chromosomes were analyzed
approximately half a year after the primary cultures had been made, nearly all
the cells showed heteroploid constitution.
In 1960 Defendi et al. [22] published data concerning
identification of cell lines in culture on the basis of morphological, immunological
and karyological criteria. As a result of these studies McCoy cell line, placed
at disposal from two different laboratories (University of Texas, Houston and
the Wistar Institute, Philadelphia), showed distinctions giving grounds to
categorize the cell line as McCoy A (human cells) and McCoy B (mouse cells
possessing a marker chromosome, characteristic for L mouse fibroblasts). There
are no data in literature whether any contamination of the original human cell
line McCoy has emerged or when and under which circumstances an eventual
contamination has lead to the peculiar karyotype and the presence of mouse
antigens.
In American Type Culture Collection (ATCC) the cells
have been left by the Center for Disease Control, Cell Culture Department, Atlanta, Georgia
in March 1984 [2], where they have been registered with No CRL-1696. They were
described as obtained from a mouse (Mus
musculus), from unknown tissue, as adherent cells with fibroblast-like
morphology. McCoy B subline is mainly distributed among the laboratories. This
was confirmed also for McCoy, given us by National Bank of Industrial
Microorganisms and Cell Cultures (NBIMCC) in Bulgaria
[29].
Based on information in the ATCC Catalogue [1],
Nogueira et al. [102] showed McCoy cell line as a hybrid lineage with markers
from human cells and mouse cells, confirming this with the fact that these
cells express human CD4 receptors [101].
Culture medium
McCoy cell line can be cultured in various media
Minimum Essential Medium (MEM), Dulbeccos Modified Eagles Medium (DMEM),
Roswell Park Memorial Institute (RPMI) 1640, medium 199 and others
(Table 1). The most commonly used medium is MEM, but it varies in the
amount of serum and other supplements.
McCoy cells and microbial culture
McCoy cells
are applied for culture of various microorganisms, which are dependent in their
development on the eukaryotic host cell. During their interaction with
pathogens the cells are subjected to various changes leading often to lethality
of infected cells.
Chlamydia
trachomatis.At the
beginning of 60s of 20th century, the popularity of McCoy cells has
increased considerably after the report of Gordon et al. [51] that cobalt-60
irradiated cultures of synovial fibroblasts strongly rise
their susceptibility to infection with chlamydial strains. This has been
confirmed in another publication of Gordon et al. [52] and allowed the
introduction of McCoy cells as a method for diagnosis of genital and ocular
infections caused by chlamydia [59].Numerous
investigations with various agents followed, aiming the increase of cell
susceptibility to infection with chlamydial strains.Pretreatment of McCoy cells with diethylaminoethyldextran [20],
cycloheximide [143, 15], cytohalasine B [137], 5-iodo-2-deoxyuridine [75],
cycloheximideand centrifugation [151],polybrene [123], mitomicin C [152] fascilitates more
successful infection of cell cultures. Inoculation and isolation of
chlamydia in hen embryos has been replaced by cell cultures - commonly McCoy
[142]. The attention was driven towards searching new cell lines which may be
used for chlamydial culture with diagnostic purpose or study the life cycle of
bacteria in vitro. Croy et al. [18]
examined the susceptibility to infection with trachoma TW-3 (type C) and UW-5
(type E) ofo eleven cell lines -
HeLa 229, HeLa M, HEp-2, FT, BHK-21, Vero, MK-2, MPK, L-WO5A2, McCoy and L-929;
Rota [120] compared five cell lines - BHK-21, CHO, HeLa S3, McCoy, OWMK and two
diploid strains, ST/BTL and WI-38, for their ability to be infected with
trachoma strains B serotype; the monkey cell line BGM was studied for isolation
of C. trachomatis by Krech et
al. [74]. At present McCoy, HeLa 229 and BGMK are the most commonly used cells
for maintenance of C. trachomatis
growth [12].
C. trachomatis
growth in McCoy cell culture, non-treated with cycloheximide, is deeply
influenced by the lack of glucose and minimal changes in aminoacids in the
environment and blood plasma.This is supported
by the production of abnormal forms with lower infectivity [55]. McCoy cells
show great capacity in terms of aminoacid concentration, which is important for
the development of microorganisms [56].
Table
1.Media for culture of McCoy cells.
Medium
|
Supplements
|
References
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Gey salt solution
|
45 % human ascitic fluid, 5 % chick embryonic
extract
|
116, 63
|
Medium 199
|
1 % horse serum
|
22
|
MEM
|
10 % FCS
|
1, 2
|
MEM
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5 % FCS, 2 mM L-glutamine, 100 μg/ml streptomycin
|
25
|
MEM
|
5 % FCS, 10 mmol/l L-glutamine, 200 U/ml
penicillin, 200 μg/ml streptomycin
|
125
|
EMEM
|
10 % FCS, 10 mg/l gentamicin, 2 mM glutamin,
1 % non-essential aminoacids
|
126
|
Eagles MEM with Hanks salts
|
10 % FCS, 10 μg/ml gentamicin, 50 μg/ml vancomycin, 2 mM glutamin
|
10
|
EMEM
|
10 % FBS, 1 % L-glutamine w/v, 10 % (v/v)
non-essential aminoacids and 1 % (w/v) of antibiotic/antibiotic mixture,
containing penicillin, streptomycin and amphotericin B.
|
42
|
GMEM (Glasgow modification of Eagles medium)
|
10 % new born calf serum, 25 mM glucose
|
91
|
RPMI 1640
|
10 % FCS, 30 mmol/l glucose, 10 μg/ml gentamicin, 2 mmol/l L-glutamin
|
82
|
RPMI 1640
|
10 % FCS
|
133
|
DMEM
|
4 % FCS
|
138
|
DMEM
|
10 % FCS
|
147
|
DMEM-H
|
584 mg/l L-glutamine, 4500 mg/l glucose, 10
mM HEPES, 10 % FCS
|
127
|
Liverpool-Waymouth medium
|
10 % FCS
|
70
|
SF-3
|
|
119
|
|
|
|
The cell
culture serves as a standard for comparative detection of C. trachomatis from genital specimens by Polimerase Chain Reaction
(PCR), Ligase Chain Reaction (LCR) and cell culture [132, 99, 66, 80]. With the encroach of DNA amplification techniques in
this field the cell cultures are applied less frequently, usually in
specialized reference laboratories [12]. McCoy cells possess advantages which
can not be duplicated by noncultural techniques, for example the culture may:
i. preserve microorganisms and even allow them to
multiply, ii. serve for examination of the
interactions microorganisms-host cell which helps studying the bacterial
biology and pathological effects as a result of bacterial metabolism, iii. allow testing for susceptibility to various antimicrobials.
The
information elucidating complex chlamydia-host interactions has been obtained
mainly form investigations on cell cultures as an in vitro model of
infectious process. Various cell lines (HeLa, HEp2, HeLa 229, CHO, VERO, BGMK) are applied now for culture of cells with chlamydia
and studying the chlamydial pathogenesis. McCoy remains the leading cell line
in these investigations.
The
interaction between bacteria and host cells takes place in the following
sequence: adhesion of microorganisms to the eukaryotic cell and entering it,
intracellular development with bacterial amplification and releasing of new
chlamydial bodies out of the cell.
Studying the
kinetics of attachment and ingestion of C.
trachomatis serotype L1 by monolayers of McCoy cells Söderlund et al.
[135] proved that the accumulation of bacteria in cells needs 3 hours when the
incubation is at 37 oC and cannot take place at 4 oC.
They also established that chitobiose and chitotriose reduced association of C. trachomatis with McCoy cells. The
precise mechanism through which the elementary bodies (EBs) attach to the cell
and penetrate into it is not completely understood. Thermolabile proteins
mediate adhesion of many C. trachomatis
serotypes to a common receptor on McCoy and HeLa cells [148]. Hodinka et al.
[61] performed ultrastructural studies upon endocytosis of C. trachomatis on McCoy cells. Following attachment to a non-well
defined receptor on host cell surface, the bacterium internalizes in the cell.
The internalization of C. trachomatis
serotype L2 in McCoy cells may be realized through phagocytosis and pinocytosis
[117]. In cytoplasm the EBs are membrane confined and their aggregation and
fusion take place by means of cytosol anexins which participate selectively in
the endosomal aggregation and escape the fusion with lysosomes during
chlamydial infection [84]. In intracellular distribution and localization of
chlamydia-containing vesicles actin and clathrin [83, 85], anexins together
with the level of free intracellular calcium ions [84], and the host cell
cytoskeleton [127] participate. It has been established that the way of
accumulation and development of serotype L in HeLa cells is different from the
one of serotype E in McCoy cells. These differences concern the cell types,
respectively epithelial and fibroblast [127]. Entering the cell, bacteria begin
multiplication. Van Ooij et al. [144] found that C. trachomatis serotype LGV L2 fusion does not take place at 32 oC
in HeLa, McCoy and CHO-K1 cell lines and requires synthesis of bacterial
proteins.
As a result
of infection with C. trachomatis,
McCoy cells produce interferon and nitric oxide in the absence of exogenous
cytokines [25]. During chlamydial infection in eukaryotic cells, proapoptotic
stimuli are induced, leading to apoptosis in non-infected adjacent cells [126].
The treatment with antioxidants reduces the degree of apoptosis. HeLa 229,
HEp-2 and McCoy have been used by Shaw et al. [133] for characterization of
proteases secreted by chlamydia.
Chlamydia
psittaci. For in vitro isolation, culture, typing and
studying the bacterium-host cell interaction and intracellular life of
C. psittaci various fibroblast (McCoy, L-929, BHK-21)
or epithelial (HeLa and BGM) cell cultures have been used [37]. The McCoy cell
line is applied for characterization of C. psittaci isolates from horses
[150] and pigeons [122]. Cultures facilitate studies on C. psittaci in
ruminal and abomasal contents [3]. The ovine abortion isolate of C. psittaci,
S26/3, may be cultured in McCoy cells which serve for investigation of
biochemical properties of Major Outer Membrane Protein (MOMP) [153] and
clarifying the antigenic organization of N-terminal part of the membrane
proteins 90, 91A and 90B of C. abortus [149]. McCoy cells are the standard
for development of new non-cultural methods for detection of chlamydia in bull
semen fluid [28], introduction of PCR for diagnosis of enzootic abortion in
ewes (EAE), [17] and/or development of new vaccines [48].
Chlamydia
pneumonie. C. pneumonie can be hardly cultured in vitro. Cles et al. [14] have
established that this respiratory pathogen is better cultured in non-treated HL
cells than in HeLa 229, McCoy, BHK-21. Roblin et al.
[118] proved that HEp-2 was the most sensitive cell line (among HeLa 229,
McCoy, HL, HEp-2, HTED) for infection with TW-183 strain and two clinical
isolates. Although less frequently McCoy cells are applied for maintenance of
some strains as C. pneumonie TW 183
strain [50].
Chlamydia pecorum.McCoy
cell line is used for culture, identification and testing of C. pecorum [113, 114].
McCoy cells are applied for in vitro investigation of C.
trachomatis [8, 9] and C. pecorum [113] persistence.
Helicobacter pylori.H. pylori participates
in the pathogenesis of chronic superficial gastritis and ulcer [36]. Korych et
al. [72], using in vitro cell models as VERO and McCoy cell lines, found that H. pylori strainscaused cytotoxic effect on cells with changes in cell cytoplasm
and morphology. The authors suggested that morphological changes in cell
cultures support the idea for the pathogenic activity of bacterium on gastric
mucosa. Similar association between cytotoxic strains and activity of gastritis
was established by Hua-Xiang Xia et al. [64]. They also proved that McCoy cells
are more sensitive than HeLa cells in detection of H. pylori cytotoxicity in
vitro.
Gardnerella vaginalis.McCoy
cells are applied as an in vitro cell
model for testing the adhesion-receptor mechanism of interaction between G. vaginalis (a causative agent of
bacterial vaginosis) and host cells as well as for various factors that inhibit
pathogen adherence [129, 131]. The pathogen adhesion to vaginal epithelial
cells, McCoy cells and red blood cells, was studied by electron microscopy
[130].
Clostridium difficille.C. difficile causes
a severe disease of the colon pseudomembranous colitis. This bacterium
produces two toxins - A and B, which are the main virulent factors [140]. They
do not affect membrane permeability of intestinal cells and McCoy cells but
inhibit protein synthesis of the latter [96]. In vitro toxins A and B cause cytotoxic effect which is manifested
by change in morphology (shrinkage and roundness) of McCoy cells [5]. During
cell intoxication, reorganization of cytoskeleton microfilaments occurs. The
effects are mainly due to cytotoxin B 1000 times more toxic than toxin A.
Toxin A damages phosphorilation of intracellular proteins in contrast to
cytotoxin L (released by C. sordelli,
a pathogen capable of producing gas gangrene in humans). The latter acts
through phosphorilation of pp80c on McCoy cells [128].
Various
cell cultures have been used for detection of cytopathic effect of C. difficile. McCoy cells as suspension
or monolayer may replace HeLa [86]. Comparative studies on several cell lines: african green monkey kidney (AGMK), MRC-5, primary rhesus
monkey kidney (RMK) and Vero proved that only Vero could be used as equivalent
to McCoy in detection of C. difficile
toxin from stool filtrates [87].
The
detection of cytotoxic effect on monolayers of McCoy culture cells is widely
applied in the diagnostics of C. difficile
infections [13]. This method is evaluated as a standard [23] and helps
elucidation of etiology and pathogenesis of C. difficile -associated
diarrhea [141].
McCoy cells are a comparative standard for various
immunological and molecular methods (TCD Toxin A
Enzyme Immunoassay (EIA), Toxins A/B Enzyme-linked Immuno-sorbent Assay
(ELISA), PCR) for detection of C. difficile
toxins [71, 88, 89, 90, 115].
Bacillus cereus and other Bacillus spp.According to
Jackson
[65] McCoy cell line may be a rapid test for screening and detection of
enterotoxin-producing B. cereus as a
cause of toxin mediated foodborne disease. Studying the cultural supernatants
of 30 bacteria, the author established the cytotoxic effect of progressive
damaging the McCoy cell monolayer.
This cell culture system is an excellent opportunity
for testing the cytotoxicity of Bacillus
isolates but also allows studying the mechanism of action of the cytotoxic
components. Combining Methylthiazoletetrazolium (MTT) cytotoxic test, Confocal
scanning laser microscopy and Scaning electron microscopy, Lindsay et al. [79]
showed that cytotoxic effects of Bacillus
spp. isolates caused for three hoursirreversible
morphological changes leading to cell membrane damage, linkage of cell content
and necrosis.
Trichomonas vaginalis.Clinical
isolates and strains of T. vaginalis
are successfully maintained in serum-free culture media [26] or as co-culture
with eukaryotic McCoy cells [47, 15]. The results obtained are similar with
those in conventional conditions of culture. The application of cell cultures
in this field gives the chance to study the contact interaction with its
intimate mechanism between this parasite and the eukaryotic cells. Thus the
role of some soluble products of T.
vaginalis was established. The example is the cell detaching factor
(CDF), which causes monolayer damage of McCoy, HEp2, CHO cell cultures and
human skin fibroblasts for 6 hours only [45]. The parasite develops in physical
contact with McCoy cells and the cell death is a result of production of
contact-dependent cytotoxicity [46]. In a co-culture of T. vaginalis (a highly vurulent strain) with McCoy Roussel et al.
[121] proved that the cytopathic effect is specifically inhibited by
monosaccharides, N-acetylglucosamine and manose. The researchers suggested that
the cytopathic effect is mediated by a manose/N-acetyl-glucosamine-binding
lectin. As it is known, there are molecules in serum, which may interact and
bind directly to T. vaginalis [112]. Meysick et al. [93] examined
the growth kinetics of T. vaginalis
in McCoy cell culture in serum-free conditions. They obtained lower peak
of vaginalis
concentration and prolonged doubling time compared to the serum-containing
system and the conventional culture of the parasite. It has been determined
that serum proteins could interact with enzymes and directly with T. vaginalis.
Viruses. According
to Consales et al., [16] rabies infected McCoy cells may provide a useful assay
system based on induction of cytopathic effect, high virus production and
sensitivity to interferon. The authors proved cytopathic changes 24 to 72 hours
after infection. The viral titre grew with the number of passages reaching
maximum after the third one. This sensitivity was confirmed by Nogueira [100]
in isolation of Rabies virus from central nervous system of a patient with
rabies. A comparative study proved that McCoy cells are with higher sensitivity
and specificity than N2A cells (a mouse neuroblastoma), which have been
accepted as a reference culture [101]. This implies McCoy cells as an effective
model for Rabies virusisolation.
There are
data that HIV-1 is success-fully replicated in McCoy cells [102] and the cell
line is a suitable model for its isolation. It can be used for studying the
dynamics of viral infection together with pharmacological testing of drugs as
well as analysis of the immune response in vaccine therapies.
In
contrast to Rabies virus, Measles virus does not induce cytopathic effect in
McCoy cells but leads to the development of persistent infection which is
maintained by an antiviral factor [125].
McCoy
cell culture and antibacterial agents.
A
number of active substances with various origin are examined in the straggle
against pathogens plant extracts [145, 24], human defensin and porcine
protegrin [156], microbial producers [62, 67], recombinant mouse
interferon-gamma [25], semisynthetic [24], or synthetic ones [76, 155]. The
ideal agent would be the one with no or minimal cytotoxicity and high
antibacterial activity. This implies the idea that in searching such drugs
combined and parallel investigations for toxicity on eukaryotic cells on one
hand and antimicrobial effect on the other should be performed. The minimal
inhibitory concentration (MIC) of chloramphenicol for C. trachomatis in McCoy cell culture varies widely in terms of
antibiotic preparation, duration of treatment and method of infection [60]. The
considerable variations in accumulation of macrolides depend on the ability of
the tested McCoy, HeLa 229F and HeLa 229W cells to accumulate the drug [73]. In
vitro cultures allow to obtain the minimal inhibitory concentrations for
normal (MICN) and abnormal inclusions (MICA) of C. trachomais when treated with various antimicrobials [62].
De la Maza et al. [21] found approximately 50 % inhibition of cell growth
in McCoy cells, using 10 U/ml mouse recombinant interferon-gamma whereas
at 1 U/ml over 95 % inhibition of chlamydial inclusions had been
observed.
In vitro
studies concerning inhibitory activity of various antibiotics against C. trachomatis in McCoy cells give
satisfactory prelimenary information about the activity of the tested drugs
which may be a basis for further clinical investigations. Lefevre et al. [78]
showed that the lowest MIC values had been established for clarythromycin and
sparfloxacin which were with the highest activity in clinical trials. Moulding in vivo the in vitro conditions of the cell culture test system McCoy, Lampe et
al. [76] proved that chlorhexidine gluconate gel killed C. trachomatis serovar D and F at concentrations applicable in
genital tract of women.
Electron
microscopic studies on McCoy cells established a realtionship between the
applied concentrations of doxycyclin, erythromycin and ofloxacin and the
changes in cell cycle of C. trachomatis [19].
Studying the antichlamydial activity of doxycyclin, erythromycin, ofloxacin and
trovafloxacin, Jones et al. [67] established that all 19 strains of C. trachomatis were sensitive to
trovafloxacin. In vitro activity of a
new fluoroquinalone - ABT-492, was examinedon McCoy [54].
The
sensitivity of chlamydia to β-lactam antibiotics is due to the presence of
penicillin-binding proteins (PBPs). Using C.
trachomatis 434 serotype L2, cultured in McCoy cell monolayer, the binding
of seven β-lactams
to chlamydial PBPs and their antichlamydial activity were examined in vitro [138].
Mast
acids and monoglycerides inactivate C. trachomatis
in vitro. The highly specific
antichlamydial effect of monocaprin is combined with cytotoxic changes in McCoy
cell monolayer at high concentration whereas at 50 μg/ml and
lower, lysis has not been observed [11].
Sokyleszczyrska
et al. [136] studied the effect of antibacterial and antitoxic serum against C. difficile by neutralization test on
McCoy cell line. Martirosian et al. [88] moulded the influence of dioctahedral
upon ten toxigenic strains of C.
difficile and eight enterotoxigenic strains of Bacteroides fragilis using McCoy and HT 29/C1 cell lines.
There are microorganisms with ability to survive and multiply in eukaryotic
cells. That is why it is important to know for the antimicrobial agent to have
good penetration, accumulation and intracellular activity. Pascual et al. [105] examined in a series of
experiments lomefloxacin and temafloxacin penetration in human neutrophils and
peripheral macrophages, accumulation of fluconazole in human polymorphonuclear
leucocytes [106], accumulation and intracellular activity of trovafloxacin, a
new ketolide, HMR 3647, obtained by erythromycin A and linezolid in human
phagocytes [107, 108, 109]. This allows elucidating the intracellular activity
of antimicrobial agents and enriches the information about their intracellular
pharmacology in macrophages and non-phagocytic cells as McCoy cells which are
used in the studies as comparative cell culture. It has been proved that
ofloxacin is carried by liposomes and its accumulation in McCoy cells is 2.6
fold higher than the one of free drug [43].
Microbicides are strategy with great potential for prevention of
sexually transmitted diseases (STD). An ideal topical microbicide should not
only kill STD-causing pathogens and be potentially spermicidal, but also should
not disrupt the normal flora of the vagina or rectum and cause cytotoxicity to
the vaginal or rectal epithelium [6]. Two cecropin peptides D2A21 and D4E1 and
gel formulations containing 0.1 to 2 % D2A21 act as effective topical
microbicides against two urogenital strains of C. trachomais serovars (UW-3/Cx) and F (UW-6/Cx). This study was
performed on McCoy cell test system combining preinoculation MCC-tests and
postinoculation MIC-test [6].
Long treatment with given antimicrobials may evoke resistance of many
microorganisms. In vitro serial
passage of C. trachomatis and C. pneumoniae with McCoy cell monolayer
established that increasing the number of passages developed resistance to the
examined fluoro-quinolones only in C. trachomatis
[97]. Analysing quinolone resistance determinating regions of two
quinolone-resistant C. trachomatis
mutants, the authors proved the presence of a point mutation in the DNA-girase
coding gene. They also assumed other, unknown mechanisms for the high level of
resistance [97].
McCoy cell line - cytotoxicity and
cell compatibility in vitro assessment
Cell
cultures as method for in vitro study
of the interaction between various substances and cells have already gone
through a rapid development in the years. Numerous cell test models and methods
for detection of cell response after the treatment have been created. McCoy
cells are the suitable and widely applied in
vitro cell test system for these investigations. McCoy cell culture has
been used for investigation of the inhibitory effects of chloramphenicol [104]
and cephalotin [103].
Using
Neutral Red (NR) and MTT tests, Varanda et al. [145] studied the influence of a
new isocoumarin (Paepalantine), isolated by them, with
antimicrobial activity on McCoy cell line. Devienne et al. [24], applying the
same culture test system, examined in
vitro the cytotoxic effect of natural and semisynthetic isocoumarins of Paepalanthus bromelioides and the
structural parameters influencing the cytotoxicity of isocoumarins, similar to
paepalantine.
Vento
et al. [146] studied the effect of dexamethazone on cell division and
macromolecular synthesis in McCoy cell system. Fighetti at al. [41] established
that various concentrations of cadmium caused cell damage, induced reduction of
metaphase number and shortened the metaphasic chromosomes.
Microorganisms
produce and secrete in the environment various substances, some of which cause
cytotoxic or necrotic effect on eukaryotic cells. A rapid and easy way for
detecting such virulent factors is the application of in vitro sensitive cell test systems from animal and human cell
lines. Balaji et al. [4] reported a cytopathic effect in monolayers of McCoy,
HEp-2, HeLa confluent cultures after treatment with Burkholderia pseudomallei supernatants.
Cytopathic
effect was detected during treatment of McCoy cells with virulent factors,
produced by C. difficile [86], B. cereus [65]; hemolysin,
proteases and cytotoxin of Aeromonas
hydrophilia and A. sobria
[7, 81], phenol acids from Scrophularia
frutescens [49].
Examining
48 chemicals on MEIC programme (The Multicenter Evaluation of In Vitro Cytotoxicity), Shrivastava et
al. [134] used cell cultures of primary hepatocytes and McCoy and MDBK cell
lines. They found significant correlation between in vitro and in vivo
values.
Newly
obtained silicone polymers, designed for contact eye lenses, have been
investigated on McCoy cells for cellular compatibility [95].
The
interaction of cells with endoplasmic reticulum and the intercellular
interactions are of great interest concerning the processes of cell
differentiation and signal transduction. Latz et al. [77] studied the adhesion
of McCoy and rabbit lens epithelial (LE) cells on modified acrylic polymers
(EF35) and found different adhesive mechanism in substituted polymers, leading
to an increased and sustained activation of integrin mediated kinases and
changes in the cytoskeletons of McCoy and rabbit LE cells.
Other applications. Apoptosis may be induced by various physiological and pathological stimuli.
Low-temperature shock causes
DNA fragmentation in McCoy cells as well as morphological and biochemical
changes, characteristic of apoptosis [111] as a result of activation of
mechanisms with participation of Ca 2+ and protein kinase C
[110]. Investigations on apoptotic processes, induced by a new group of immune
response modifiers, were carried out on McCoy cells and other cell cultures
[92].
The
culture medium, presence of serum and supplements influence the development of
every cell culture. They affect cell growth, cell yield and production of
specific cell products.
Cell culture
media contain glutamine as a permanent component which is the main energy
source and biosynthetic precursor of cell growth. It is believed that
adaptation of McCoy cells to a medium where glutamine is replaced by glutamate
or 2-octa-ketoglutarate, would promote cell yield [57]. The effect of lactat
and ammonium was studied on McCoy and other eight cell lines. It was confirmed
that the cell yield is influenced by the accumulation of ammonium in the medium
as a result of glutamine metabolism and its chemical degradation [58]. In biotechnological
aspect the production of biological components can be limited by ammonium
accumulation. McCoy and MDCK cell lines have been used as model cultures by
McDermott et al. [91] for clarifying the metabolic changes concerning cell
adaptation to glutamine-free medium and the role of glutamate-transport system.
Hanotte et
al. [53] studied the develop-ment of McCoy cells and other animal cell cultures
on microbeads Cytodex 3.
In
several publications the metabolism of polyamines in McCoy cell culture was
elucidated [38, 39, 40].
The McCoy-Plovdiv cell strain a
serum free and protein free culture
The
serum-free cell culture McCoy-Plovdiv was derived from the McCoy cell line
[31]. The cells are cultured solely in HD medium which is chemically defined,
serum-free and does not contain additional proteins. The processes of freezing
and thawing are completed in the same medium [34]. The morphological and
karyological analysis confirmed the origin of McCoy-Plovdiv cell line [29]. The
cell kinetics [33], the proliferative activity [44], the dynamics of cell
monolayer and the postconfluent culture conditions [30] were studied. It was
experimentally proved that McCoy-Plovdiv cells showed equivalent sensitivity
and specificity as McCoy cells in detection of C. trachomatis from genital specimens of patients [98]. Cytotoxic
studies established that McCoy-Plovdiv cells are more susceptible than McCoy to
in vitro testing of chemicals [27].
The McCoy-Plovdiv cell test system for development of cytotoxic tests with vital
dyes is in progress [94].
There are convincing data about advantages of
McCoy-Plovdiv cell system in co-culture with T. vaginalis [32].
Hopeful results have been obtained for McCoy-Plovdiv cells as an appropriate
substrate for antinuclear antibody detection [124] and a protocol for
application of the serum-free cells in the screening of antinuclear antibodies
has been proposed [35].
Additionally,
McCoy-Plovdiv cells might be an adequate cell line for serum-free culture of
nanobacteria, a recently characterized group of extremely small bacteria,
capable of precipitating calcium salts and implicated in the pathogenesis of
human renal and gall stones and calcific atherosclerosis [68; 69]. Now
nanobacteria are cultured by using cell culture media under mammalian cell
culture conditions or 3T6 fibroblast monolayers. It has been established that
the stone formation by nanobacteria is low in the presence of serum in the
culture but extensive and rapid in serum-free conditions [69]. Thus
McCoy-Plovdiv cell line may provide a good opportunity to study nanobacteria
and their interactions with the cells.
Conclusion
McCoy cell line occupies an important place among the
most popular cell cultures - HeLa, HEp-2, Vero, CaCo-2, 3T3,
MDCK, with its application in experimental and diagnostic laboratories. As
recent reports show the cells are actually important for culturing viruses,
chlamydia, vaccine studies, development of models for C. trachomatis or cytotoxic activity [4, 48, 101, 102, 139, 154]. A new direction of McCoy application is the creation
of McCoy-Plovdiv serum-free cell line.
Acknowledgments.
This work was supported by Grant No 12/2002 from the Scientific and
Developmental Work, University
of .
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