ISSN 0006-2979, Biochemistry (Moscow), 2025, Vol. 90, No. 10, pp. 1366-1375 © Pleiades Publishing, Ltd., 2025.
Russian Text © The Author(s), 2025, published in Biokhimiya, 2025, Vol. 90, No. 10, pp. 1463-1473.
1366
Transmembrane Transport of Water and Urea
in Rat Corneal Endothelial Cells
Lyubov E. Katkova
1,2,a#
, Galina S. Baturina
1,2#
, Igor A. Iskakov
3
,
and Evgeny I. Solenov
1,4,b
*
1
Institute of Cytology and Genetics, Siberian Branch of the Russian Academy of Sciences,
630090 Novosibirsk, Russia
2
Novosibirsk National Research State University (NSU), 630090 Novosibirsk, Russia
3
Federal State Autonomous Institution “National Medical Research Center
“Interbranch Scientific and Technical Complex “Eye Microsurgery” named after Academician S. N. Fedorov”
of the Ministry of Health of the Russian Federation, Novosibirsk Branch,
630096 Novosibirsk, Russia
4
Novosibirsk State Technical University (NSTU), 630087 Novosibirsk, Russia
a
e-mail: ile@academposter.ru
b
e-mail: eugsol@bionet.nsc.ru
Received July 3, 2025
Revised September 12, 2025
Accepted September 25, 2025
Abstract— This study investigated permeability of the apical and basolateral membranes of rat corneal endo-
thelial cells to water and urea. We demonstrated that the apparent water permeability of the basolateral mem-
brane of endothelial cells (4.43E−05 ± 7.57E−07cm/s) is more than three times higher than that of the apical
membrane (1.21E−05 ± 1.03E−07cm/s). Permeability of the basolateral membrane to urea (1.23E−04 ± 1.56E−06
cm/s) was statistically significantly higher than that of the apical membrane (9.52E−05 ± 1.02E−06 cm/s) by
approximately 30%. We examined contribution of the phloretin-inhibited urea transport across the apical
and basolateral membranes in these cells. Phloretin at concentration of 0.1 mM significantly reduced urea
permeability by more than 20% through both the apical and basolateral membranes. The results suggest that
the compositions of transporters involved in water transport in the apical and basolateral membranes differ
significantly. It is hypothesized that high apparent water permeability of the basolateral membrane of endo-
thelial cells is due to contribution of the concomitant water transport with ions involved in active transport
processes. Presence of the phloretin-sensitive urea transporters in the plasma membrane of endothelial cells,
likely involved in its transcellular transport, has been demonstrated. The results indicate potential significance
of urea for corneal function.
DOI: 10.1134/S0006297925601881
Keywords: corneal endothelium, transmembrane transport, water permeability, urea permeability, phloretin
* To whom correspondence should be addressed.
# These authors contributed equally to this study.
INTRODUCTION
Corneal transparency in many animals and hu-
mans is achieved through mechanisms that maintain
osmotic balance in the stroma, which contains hydro-
philic proteoglycans [1, 2]. Disruption of the balance
between water and osmotically active substances can
cause stromal swelling, leading to uneven spacing
between collagen fibers, increased light scattering,
and reduced corneal transparency. Corneal edema is
counteracted by the multilayered epithelium, which
exhibits low water permeability and low rate of ion
transport, and the endothelium, which is character-
ized by higher permeability and active ion transport
processes in the cells. Corneal endothelium is a mono-
layer of tightly connected cells that separates stroma
from anterior chamber of the eye. Endothelium can
maintain stromal thickness and hydration levels by
transporting water and ions. This function is typically
WATER AND UREA FLUX IN CORNEAL ENDOTHELIUM 1367
BIOCHEMISTRY (Moscow) Vol. 90 No. 10 2025
described in terms of the “pump-leak” model, accord-
ing to which the flow of water into the stroma, de-
termined by osmotic and hydrostatic pressure gradi-
ents, is balanced by active transport (pump) of ions
and associated water into the anterior chamber of
the eye [3].
Permeability of endothelium to water is largely
determined by the presence of water channels in the
apical and basolateral cell membranes, represent-
ed by aquaporin 1 (AQP1) [4]. Immunohistochemical
studies indicate presence of AQP3 in the plasma mem-
brane, but functional contribution of this channel in
the corneal endothelial cells remains unexplored [5].
AQP3 water channels are permeable to water, glycer-
ol, urea, and hydrogen peroxide [6, 7].
Transport of water and solutes across a mono-
layer with tight junctions between the cells, as in the
case of the corneal endothelium, depends on permea-
bilities of both basolateral and apical cell membranes,
which could differ significantly. For example, studies
of anion transport across the corneal endothelium
have shown that permeability of the basolateral mem-
brane to HCO
3
−
is significantly higher than that of
the apical membrane [8]. However, transport of water
and electrically neutral molecules in the corneal en-
dothelial cells remains poorly understood.
Urea appears to be an important and well-reg-
ulated agent necessary for eye function. Disruption
of its metabolism, particularly decrease in the urea
concentration in tear fluid in dry eye syndrome [9],
indicates its possible role in regulating osmotic bal-
ance of the cornea. However, current research on the
role of urea transport in the mechanism of stromal
osmoregulation is virtually absent in the literature.
Expression of the SLC14a1 gene, which encodes the
urea transporter UT-A (UT, urea transporters), has
been demonstrated in the corneal endothelial cells
[10], which is present in significant amounts in the
basolateral membrane of endothelial cells, indicating
its possible importance for urea transport and mainte-
nance of stromal osmotic balance [11]. Studies of mice
with knockout of the SLC14a1 gene have revealed ab-
normalities in the corneal endothelium leading to its
edema [12]. Phloretin (2′,4′,6′-trihydroxy-3-p-hydroxy-
phenylpropiophenone) is a plant-derived flavonoid
widely used in research as a pharmacological agent
that inhibits various transport pathways, including
urea transporters [13, 14]. In particular, UT-B and UT-A
exhibit high sensitivity to phloretin, even at low con-
centrations [15, 16]. Several studies have also shown
modulating effect of phloretin on certain aquaglycero-
porins [17-19].
In this study, we investigated permeabilities of
the apical and basolateral surfaces of rat corneal en-
dothelial cells to water and urea. Given wide variety
of the molecular forms of potential urea transporters,
the study of urea transport was limited to examin-
ing possible contribution of their phloretin-sensitive
forms.
MATERIALS AND METHODS
Animals. Experiments were conducted on isolat-
ed corneal endothelial cells from Wistar rats. Animals
aged 3 months were obtained from the vivarium of
conventional animals at ICG SB RAS (Novosibirsk).
Preparation of corneal endothelium. To obtain
a preparation of cells with an accessible basolateral
surface, the endothelium was applied to a coverslip
covered with a poly-D-lysine solution (0.1% (w/v),
Sigma- Aldrich, USA) using the imprint method [20].
The coverslip with endothelial cells was then placed
in a flow chamber of a microscope. To obtain prepa-
ration of cells with an accessible apical surface, a
fragment of a Descemet’s membrane with cells ori-
ented with the apical surface toward the perfusing
medium was placed on a coverslip and placed in the
flow chamber of the microscope, where the fragment
was mechanically held using a nylon mesh.
Measurement of cell volume. In the microscope
chamber with stopped flow, cells were loaded with a
fluorescent dye calcein (Calcein AM, 10
−5
M, 20 min,
37°C). Changes in cell volume were determined using
a method based on the quenching effect of calcein
fluorescence by cytoplasmic proteins [21-23].
Design of the flow chamber has been described
previously [22]. The experimental setup was based on
a fluorescence microscope Observer-Z1 (objective Fluar
25/0.8 M27, Zeiss, Germany). The chamber volume was
~50 µL, and solution flow rate was 15 mL/min, ensur-
ing rapid (t
1
/
2
< 70 ms) medium exchange. Temperature
was maintained at 36.8 ± 0.2°C. To determine the rates
of changes in cell volume reflecting permeabilities to
water and urea, cells were equilibrated in a PBS me-
dium (280 mOsm/kg H
2
O), the medium was changed
to a “PBS + mannitol” (560 mOsm/kg H
2
O) (mannitol,
Sigma-Aldrich), and the cells were equilibrated in this
hypertonic medium. The medium was then changed
to an isotonic “PBS + urea” (560 mOsm/kg H
2
O), re-
turned to PBS medium (280 mOsm/kg H
2
O), and in-
crease in fluorescence was recorded. Emitted light was
detected using a photodetector based on a photomul-
tiplier tube (PMT) equipped with a field diaphragm
to measure fluorescence intensity in the region of in-
terest. Signal recording was performed using a digital
oscilloscope AKTAKOM ASK-3102 with an interval of
10 ms throughout the experiment at low excitation
light intensity, which prevented photobleaching of the
fluorophore during the experiment. The signal was
recorded with 8-bit resolution and saved to a com-
puter.
KATKOVA et al.1368
BIOCHEMISTRY (Moscow) Vol. 90 No. 10 2025
Calculation of membrane permeability coeffi-
cients. Water permeability was calculated based on
the flow equation (1):
dV/dt = −S ∙ V
w
∙ Pf ∙ gradΦ, (1)
where Pf is osmotic permeability coefficient (cm/s), S
is surface area, Vw is molar volume of water, and Φ
is osmotic pressure.
Fluorescence profile F(t) in the initial section was
approximated by a linear function F(t) = A + Kr ∙ t.
The value of Kr was taken as an approximate estimate
of dV/dt, initial rate of change in cell volume. Given
that, as we have previously shown, V/V
0
= F/F
0
[23],
where V
0
is the initial cell volume and F
0
is the ini-
tial fluorescence signal level, Pf (osmotic permeability
coefficient) corresponds to:
dV/dt = V
0
∙ Kr = Pf ∙ S ∙ V
w
∙ ∆C, (2)
Pf = V
0
∙ Kr/(S ∙ V
w
∙ ∆C), (3)
where Kr is linear regression coefficient of the ini-
tial section of the fluorescence profile (F/F
0
), and ∆C
is difference in osmotic concentrations (Osm/kg H
2
O)
across the plasma membrane of the cell.
Permeability coefficient for urea was determined
by analyzing kinetics of the cell volume increase in a
hypertonic medium during the isotonic entry of urea.
The increase in cell volume in such system can be
approximately described by the equation (4):
V/V
0
≈ 1 + C
u
/Π
out
≈ 1 + [(S ∙ P
u
∙ ∆C)/(V
0
∙ Π
out
)] ∙ t; (4)
from which it follows:
∆C= C
out
− C
u
; (5)
(V/V
0
− 1)/t => (F/F
0
)/t = Kr
u
; (6)
P
u
≈ Kr
u
∙ (V
0
∙ Π
out
)/(S ∙ C
out
). (7)
where Kr
u
is regression coefficient of the cell volume
in the presence of urea, S is surface area, C
u
is urea
concentration in the cell, C
out
is urea concentration in
the medium, Π
out
is osmotic concentration of the me-
dium, V
0
is initial cell volume, and P
u
is permeability
coefficient for urea.
Experimentally, the area of the plasma mem-
brane surface of the cell open for exchange (S) was
determined by the area in an image of the endotheli-
um stained with calcein, since thickness of these cells
(size along the Z-axis) (3-5µm) is significantly smaller
than the dimensions along the XY axes. The cells were
considered flat, and no correction for the surface cur-
vature was made [24].
Permeability coefficients were calculated using the
average value of S = 2.5E−06 ± 1.3E−08cm
2
(M±stan-
dard error of the mean (SEM), n = 100) (Fig. 1).
Statistical analysis. Statistical analysis of flu-
orescence profiles was performed using Origin 5.0.
Statistical calculations were performed using the
Statistica 6.0 software package for Windows. The ob-
tained data were analyzed using Student’s t-test. Re-
sults are presented as M ± SEM. Results were consid-
ered significant at p < 0.05. Normality of distribution
was tested using the Shapiro–Wilk test in the Statis-
tica 6.0 program.
RESULTS
Averaged osmotic permeability coefficients and
urea permeability in the intact preparations of cor-
neal endothelial cells were as follows: for the apical
membrane (M ± SEM) – Pf = 1.2E−05 ± 1.03E−07 cm/s,
n = 30; P
u
= 9.5E−05 ± 1.02E−06 cm/s, n = 30; for the
basolateral membrane – Pf = 4.4E−05 ± 7.57E−07 cm/s,
n = 36; P
u
= 1.2E−04 ± 1.56E−06 cm/s, n = 36.
In this study, to avoid the influence of errors
arising from determining cell surface area (S) in the
calculations of permeability coefficients (Pf, P
u
), the
linear regression coefficient of the initial section of
the fluorescence profile (Kr) was used as an experi-
mental estimate of the initial rate of change in cell
volume (dV/dt) to assess the rate of transmembrane
fluxes of water and urea.
As we have previously shown, rapid osmotic
entry of water into the cell, on the one hand, ini-
tially causes the effect of increased water permeabil-
ity, which is not reproduced upon repeated shocks,
and on the other hand, activates the mechanism of
regulatory volume decrease [23, 25]. To avoid such
effects, which significantly distort the results of deter-
mining the value of water permeability, in this study,
Fig. 1. Rat corneal endothelium preparation stained with
calcein. Scale bar: 50 µm.
WATER AND UREA FLUX IN CORNEAL ENDOTHELIUM 1369
BIOCHEMISTRY (Moscow) Vol. 90 No. 10 2025
Fig. 2. Dynamics of cell volume decrease during water outflow through the basolateral (1) and apical (2) surfaces into a
hypertonic medium. a) Fluorescence decrease profile; b) initial sections of the profile. The arrow indicates the moment of
switching from isotonic PBS solution (280 mOsm/kg H
2
O) to hypertonic solution (560 mOsm/kg H
2
O).
Fig. 3. Dynamics of cell volume increase during urea entry through the basolateral (1) and apical (2) surfaces under iso-
tonic urea gradient conditions. a) Fluorescence increase profile; b) initial sections of the profile. The arrow indicates the
moment of isotonic switching from hypertonic PBS solution (PBS + mannitol, 560mOsm/kg H
2
O) to hypertonic urea solution
(PBS + urea, 560 mOsm/kg H
2
O).
we used hypertonic exposure, which allows obtaining
more reproducible results. According to the obtained
results, the intensity of osmotic water outflow from
cells with increase in osmotic pressure in the external
environment through the apical and basolateral mem-
branes differed significantly. With the same osmotic
gradient, the flow through the basolateral surface was
more than 3-fold higher than through the apical sur-
face (Table 1, Fig. 2).
During the isotonic entry of urea and associated
water into the corneal endothelial cells, the rate of
volume increase was higher when water entered
through the basolateral cell surface compared to the
same flow through the apical surface.
Intensity of urea transport through the basolat-
eral surface in these experiments exceeded transport
through the apical surface less significantly than for
water, being only 30% higher (Table 1, Fig. 3).
Phloretin present in the medium at concentration
of 0.1mM significantly reduced the rate of urea trans-
port by more than 20% through both the apical and
basolateral membranes (Table 2, Fig. 4).
From the obtained results, one can see that
with the same osmotic gradients, the resulting wa-
ter flows through the basolateral membrane are sev-
eral times greater compared to the flows through
the apical surface of the cell. Such asymmetry of
flows apparently indicates significant difference in
the composition of transporters involved in water
transport in the apical and basolateral membranes.
The more intense total water flow through the ba-
solateral membrane is obviously explained by the
KATKOVA et al.1370
BIOCHEMISTRY (Moscow) Vol. 90 No. 10 2025
Fig. 4. Effect of phloretin on the dynamics of cell volume increase during urea entry through the basolateral (c, d) and
apical (a, b) surfaces under isotonic urea gradient conditions. 1 – Control; 2 – effect of phloretin (0.1mM). a,c)Fluorescence
increase profile; b, d) initial sections of the profile.
Table 1. Linear regression coefficients of the initial sections of the profiles of cell volume dynamics during
osmotic water outflow (Kw) or urea entry under its isotonic gradient conditions (Ku)
Apical membrane (n = 23) Basolateral membrane (n=29) p
K
w
−0.15 ± 5.1E−04 −0.56 ± 6.7E−03 p = 8.06E−03
K
u
0.12 ± 6.6E−04 0.15 ± 1.2E−03 p = 1.18E−02
Table 2. Effect of phloretin on the dynamics of urea entry
Apical membrane (n=23) p Basolateral membrane (n=29) p
K
u
0.12 ± 6.6E−04
6.72E−05
0.15 ± 1.2E−03
4.86E−03
K
u
(0.1 mM
phloretin)
0.08 ± 4.7E−04 0.12 ± 1.3E−03
significant water flows associated with the ions in-
volved in the active transport processes in this part
of the cell. At the same time, urea flows through
the apical and basolateral membranes differ by less
than 30% and have a similar degree of inhibition by
phloretin (Table 1).
WATER AND UREA FLUX IN CORNEAL ENDOTHELIUM 1371
BIOCHEMISTRY (Moscow) Vol. 90 No. 10 2025
These results allow us to assume existence of the
phloretin-sensitive urea transporters in the endotheli-
al cells that carry out its transcellular transport; mo-
lecular organization of these transporters and signifi-
cance of the urea flow through the endothelium are
currently unknown and require further investigation.
DISCUSSION
In this study, we determined the differences in
the integral values of osmotic water and urea fluxes
across the apical and basolateral membrane of corne-
al endothelial cells based on the kinetics of changes in
cell volume during rapid changes in osmotic pressure
in the perfusing solution. To the best of our knowl-
edge, this is the first study of urea transport across
the apical and basolateral membranes of corneal en-
dothelial cells.
According to the obtained results, under the same
osmotic gradient, the water flux through the basolat-
eral surface of corneal endothelial cells exceeded the
flux through the apical surface by more than three-
fold. In many cell types, such as epithelial cells, the
area of the basolateral membrane significantly ex-
ceeds the area of the apical membrane, and due to
this, the apical surface acts as a rate-limiting barrier
for transcellular transport. However, the corneal en-
dothelium consists of a monolayer of flat hexagonal
cells, and in this cell type, there is likely no significant
excess of basolateral membrane area over the api-
cal membrane. According to our data, similar to the
water transport, intensity of urea transport through
the basolateral surface of corneal endothelial cells ex-
ceeded the flux through the apical surface. The ob-
tained results suggest that the pathways for water,
urea, and presumably ion transport across the apical
and basolateral plasma membranes of corneal endo-
thelial cells have different conductivities and different
compositions of transporters.
Water transport across the corneal endothelium
largely depends on the presence of water channels, as
well as HCO
3
−
and Cl
−
transporters [26-28]. The struc-
ture of water transport pathways in the mammalian
eye, according to various authors, may include six
[4, 29, 30] or nine [31] types of aquaporins (AQP).
AQP1, AQP4, and AQP7 are expressed in the corne-
al endothelium [31-33]. The recent data obtained by
immunohistochemical methods indicate presence of
the aquaglyceroporin AQP3 in the corneal endothelial
cells, which, in addition to water, can conduct some
neutral molecules such as urea and glycerol [5]. AQP1
is widely present on both apical and basolateral mem-
branes of endothelial cells [34], although its function-
al contribution remains largely unexplored. The mod-
els with knockout of the AQP1 gene showed almost
no phenotypic differences from the wild-type mice,
except that the cornea thickness in the knockout ani-
mals was reduced [32]. In the later works, the authors
conclude that the function of endothelial AQP1 and
AQP4 is minimal under physiological conditions and
is, likely, manifested during edema or other patholog-
ical conditions [33, 35].
In addition to the transport through water
channels, water can be transported across the plas-
ma membrane in a bound state, such as during ion
transport or with flows of electroneutral molecules,
for example, urea. Thus, high permeability of the
basolateral membrane to water could be associated
with high metabolic activity of the corneal endothelial
cells, which requires intense ion flows generated in
the reactions during both ATP synthesis and its hy-
drolysis [36, 37]. The literature data on composition
and expression of basolateral membrane transporters
in the corneal endothelial cells support the assump-
tion of their role in the transport of bound water
and increased apparent water permeability of this
part of the cell plasma membrane. High expression
and activity of the cotransporters NBC-1 (Na
+
/HCO
3
−
)
and NKCC1 (Na
+
/K
+
/2Cl
−
), as well as of the cAMP-ac-
tivated chloride ion transporter that is not a CFTR
channel, have been demonstrated on the basolateral
membrane [38, 39].
High expression of the SLC4A11 protein, which
has been shown to play a role in the transport of H
+
,
NH
3
, and water, has been demonstrated in the baso-
lateral membrane of corneal endothelial cells [40-42].
Mutations in the SLC4A11 cause congenital hereditary
endothelial dystrophy and, in some cases, Fuchs endo-
thelial corneal dystrophy [43-47]. However, due to the
wide variety of transporters present in the basolateral
membrane, determining their functional contribution
will require further investigations.
As with water permeability, urea flux through the
basolateral membrane of the corneal endothelial cells
is higher compared to the apical membrane under
the same gradients. The difference between the urea
fluxes is less significant, which could indicate differ-
ences in the number of molecular structures involved
in urea transport. It should be noted that the plas-
ma membrane of these cells has high permeability to
urea. As can be seen from our results, permeability of
the corneal endothelial cells to urea is comparable to
permeability of the collecting duct epithelium of the
kidney [48]. It could be assumed that such high per-
meability indicates a similar set of transporters and
that intensive urea exchange in the cornea, as in the
kidney, could be associated with its role in maintain-
ing the level of osmotic pressure, in this case in the
stroma. Since the degree of inhibition of urea flux-
es through the apical and basolateral membranes by
phloretin is similar, it is most logical to assume that
KATKOVA et al.1372
BIOCHEMISTRY (Moscow) Vol. 90 No. 10 2025
molecular structure of these transporters could be the
same. It is possible that aquaporins are involved in
urea transport. For example, it has been shown that
deletion of the aquaporin genes AQP3 and/or AQP4
disrupts urea transport in the kidney [49].
Phloretin is considered as a potential inhibitor
of some aquaporins, particularly aquaglyceroporins
[50]. Studies have been conducted on the inhibition
by phloretin (0.1mM) of water and urea permeability
of the plasma membrane of Xenopus oocytes injected
with RNA encoding aquaporins 3 or 9. It has been
shown that phloretin inhibits 86% of water transport
and about 20% of urea transport in the case of AQP9.
In the case of AQP3, suppression of both water and
urea transport by about 10% has been demonstrated
[17, 18]. Functional study of the effect of phloretin on
the model of oocytes expressing AQP4 did not reveal
its effect on water permeability [19]. Thus, phloretin
is ineffective as a blocker of aquaporins expressed in
the corneal endothelium.
Urea transporters (UT) facilitate water diffusion
across the cell membranes, as shown in the exper-
iments with expression of UT-B, UT-A2, and UT-A3
transporters in the Lithobates oocytes [51]. The issue
regarding the pathways of urea transport in the cor-
neal endothelial cells is complicated. Currently, there
is little information about the molecular transporters
capable of urea transport expressed in the corneal en-
dothelium on the apical and basolateral membranes.
The most significant indication of the involvement of
urea transporters in the regulation of stromal osmotic
balance is expression of the SLC4a1 gene, which en-
codes the urea transporter UT-A in the corneal en-
dothelial cells. Expression of UT-A in mice results in
the synthesis of several proteins – UT-A, UT-A1, and
UT-A3 – due to alternative splicing, but knockout
of the gene results in complete suppression of the
phloretin-sensitive urea transport. In the UT-A1/3
−/−
mice, low cell permeability to urea, which was not
inhibited by phloretin, was observed, allowing the
authors to suggest that this flux was not due to any
known urea transporter [52].
The results of this study indicate a significantly
higher permeability of the basolateral membrane com-
pared to the apical membrane for water, and suggest
that high permeability to urea indicates its intensive
transport in the rat corneal endothelial cells, which is
mediated through the transporters with similar per-
meability in the apical and basolateral membranes
and sensitive to inhibition by phloretin. The results
obtained in this study suggest that investigations of
the urea transporters in endothelial cells could im-
prove diagnosis of the functional state of endotheli-
um and help to identify molecular mechanisms that
are promising as targets for therapeutic interventions
in the disruption of stromal osmotic balance.
Abbreviations
AQP aquaporin
UT urea transporters
Contributions
L. E. Katkova and G. S. Baturina – equal contribution,
obtaining experimental data, discussion of research
results, writing the article text; I. A. Iskakov – discus-
sion of research results; E. I. Solenov – concept, ob-
taining experimental data, writing the article text.
Funding
This work was carried out within the framework of
the state assignment of ICG SB RAS “Mechanisms of
Genetic Control of Development, Physiological Process-
es, and Animal Behavior” FWNR-2022-0019.
Ethics approval and consent to participate
All procedures performed in studies involving an-
imals were in accordance with the following doc-
uments: “Rules of Good Laboratory Practice in the
Russian Federation,” approved by Order no. 199n of
the Ministry of Health of the Russian Federation dat-
ed 01.04.2016; interstate standards GOST 33215-2014
“Guidelines for the Care and Use of Laboratory Ani-
mals” (Rules for equipping premises and organizing
procedures, effective date 01.07.2015). The conditions
for keeping animals and the experimental procedures
performed were approved by the Bioethics Commis-
sion of ICGSBRAS (protocol no.115 dated 20.12.2021).
Conflict of interest
The authors of this work declare that they have no
conflicts of interest.
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