Development of animal models with chronic kidney disease-mineral and bone disorder based on clinical characteristics and pathogenesis

Abstract

Chronic kidney disease–mineral and bone disorder (CKD-MBD) is a systemic complication of chronic kidney disease (CKD), resulting in high morbidity and mortality. However, effective treatment strategies are lacking. The pathogenesis of CKD-MBD is unclear but involves feedback mechanisms between calcium, phosphorus, parathyroid hormone, vitamin D and other factors, in addition to FGF23, Klotho, Wnt inhibitors, and activin A. Construction of a perfect animal model of CKD-MBD with clinical characteristics is important for in-depth study of disease development, pathological changes, targeted drug screening, and management of patients. Currently, the modeling methods of CKD-MBD include surgery, feeding and radiation. Additionally, the method of CKD-MBD modeling by surgical combined feeding is worth promoting because of short time, simplicity, and low mortality. Therefore, this review based on the pathogenesis and clinical features of CKD-MBD, combined with the current status of animal models, outlines the advantages and disadvantages of modeling methods, and provides a reference for further CKD-MBD research.

Full text

Development of animal models
with chronic kidney disease-
mineral and bone disorder based
on clinical characteristics
and pathogenesis
Biyu Tan
1
†
, Weili Tang
2
†
, Yan Zeng
1
, Jian Liu
1
, Xiaomei Du
1
,
Hongwei Su
3
, Xianlun Pang
2
*, Lishang Liao
4
*
and Qiongdan Hu
1
*
1
Department of Nephrology, The Affiliated Traditional Chinese Medicine Hospital, Southwest Medical
University, Sichuan, China,
2
Department of Orthopedics, The Affiliated Traditional Chinese Medicine
Hospital of Southwest Medical University, Sichuan, China,
3
Department of Urology, The Affiliated
Traditional Chinese Medicine Hospital, Southwest Medical University, Sichuan, China,
4
Department of
Neurosurgery, The Affiliated Traditional Chinese Medicine Hospital, Southwest Medical University,
Sichuan, China
Chronic kidney disease–mineral and bone disorder (CKD-MBD) is a systemic
complication of chronic kidney disease (CKD), resulting in high morbidity and
mortality. However, effective treatment strategies are lacking. The pathogenesis
of CKD-MBD is unclear but involves feedback mechanisms between calcium,
phosphorus, parathyroid hormone, vitamin D and other factors, in addition to
FGF23, Klotho, Wnt inhibitors, and activin A. Construction of a perfect animal
model of CKD-MBD with clinical characteristics is important for in-depth study
of disease development, pathological changes, targeted drug screening, and
management of patients. Currently, the modeling methods of CKD-MBD include
surgery, feeding and radiation. Additionally, the method of CKD-MBD modeling
by surgical combined feeding is worth promoting because of short time,
simplicity, and low mortality. Therefore, this review based on the pathogenesis
and clinical features of CKD-MBD, combined with the current status of animal
models, outlines the advantages and disadvantages of modeling methods, and
provides a reference for further CKD-MBD research.
KEYWORDS
chronic kidney disease-mineral and bone disorder, renal osteodystrophy, animal
models, clinical characteristics, pathogenesis
Frontiers in Endocrinology frontiersin.org01
OPEN ACCESS
EDITED BY
Zhongjian Xie,
Central South University, China
REVIEWED BY
Zhenglin Zhu,
Chongqing Medical University, China
Saswat Kumar Mohanty,
Brown University, United States
*CORRESPONDENCE
Xianlun Pang
tsgxkfw@swmu.edu.cn
Lishang Liao
13982751916@163.com
Qiongdan Hu
huqiongdan@swmu.edu.cn
†
These authors have contributed equally to
this work
RECEIVED 21 December 2024
ACCEPTED 10 March 2025
PUBLISHED 25 March 2025
CITATION
Tan B, Tang W, Zeng Y, Liu J, Du X, Su H,
Pang X, Liao L and Hu Q (2025) Development
of animal models with chronic kidney
disease-mineral and bone disorder based on
clinical characteristics and pathogenesis.
Front. Endocrinol. 16:1549562.
doi: 10.3389/fendo.2025.1549562
COPYRIGHT
©2025Tan,Tang,Zeng,Liu,Du,Su,Pang,
Liao and Hu. This is an open-access article
distributed under the terms of the Creative
Commons Attribution License (CC BY). The
use, distribution or reproduction in other
forums is permitted, provided the original
author(s) and the copyright owner(s) are
credited and that the original publication in
this journal is cited, in accordance with
accepted academic practice. No use,
distribution or reproduction is permitted
which does not comply with these terms.
TYPE Review
PUBLISHED 25 March 2025
DOI 10.3389/fendo.2025.1549562

1 Introduction
Chronic kidney disease–mineral and bone disorder (CKD-
MBD) significantly increases the incidence and mortality of
fractures and cardiovascular diseases in patients, with high
hospitalization rates and low quality of life (1–3), as well as
incurring high medical costs and heavy social burdens. CKD-
MBD accompanied by abnormal laboratory indicators, bone
lesions, and calcification of blood vessels or other soft tissues has
a prevalence of 33.3%–81% in developing countries (4,5). The
complicated pathogenesis of CKD-MBD involves fibroblast growth
factor 23 (FGF23), a-Klotho (Klotho), Wnt inhibitors, activin A,
and other factors. In addition to the general clinical manifestations
of CKD disorders of calcium and phosphorus metabolism,
secondary hyperparathyroidism (SHPT), persistent high levels of
parathyroid hormone (PTH), abnormal vitamin D (VD)
metabolism, bone abnormalities (manifested as bone turnover,
mineralization, bone mass, linear bone growth, or bone strength
abnormalities), and vascular or other soft tissue calcification are
caused by CKD-MBD (6–9). Currently, the treatment of CKD-
MBD is still complex, including dietary and lifestyle changes,
adjustment of dialysis schedules, and the use of phosphate
binders, VD, and calcimimetic agents (10). Management of
patients with CKD-MBD faces higher demands because of
considerations of therapeutic goals, adverse drug reactions, and
health economics (11–14). Establishing a stable animal model of
CKD-MBD based on its pathogenesis and clinical characteristics is
crucial for the study of the disease, and provides an effective
experimental tool for screening and testing of clinically effective
drugs. Therefore, this review summarizes the current status of
animal models of CKD-MBD and provides an overview of the
pathogenesis, evaluation methods, modeling, and common
problems of CKD-MBD.
2 Pathogenesis of CKD-MBD
The onset and progression of CKD-MBD involves feedback
mechanisms between phosphate, calcium, PTH, VD, and other key
factors (15,16). FGF23, Klotho, Wnt inhibitors, activin A and
circulating inflammatory biomarkers play different roles in the
pathogenesis of CKD-MBD (Figure 1). FGF23 is derived from
osteoblasts and plays an important role in VD and phosphate
metabolism (17–19). By targeting proximal renal tubular
epithelial cells, FGF23 decreases the surface expression of the
sodium/phosphate cotransporter proteins NaPi-2a and NaPi-2c,
thereby reducing renal phosphate reabsorption (20,21). At the
same time, FGF23 reduces intestinal phosphate absorption by
down-regulating 1,25-hydroxylase activity and increasing 24-
hydroxylase activity, thereby decreasing 1,25-dihydroxy-vitamin
D (1,25-(OH)
2
D) synthesis (22–24). According to the early stages
of CKD, the compensatory elevation of FGF23 levels can counteract
hyperphosphatemia. Nevertheless, the prolonged FGF23 overdose
reduces phosphate reabsorption by impairing the ability of the
FIGURE 1
The pathogenesis of CKD-MBD. ActA, Activin A; ActRIIA, activin II type A receptor; Klotho, a-Klotho; FGFR1, FGF23 receptor 1; FGF23, fibroblast
growth factor 23; uPA, urokinase plasminogen activator; uPAR, soluble urokinase receptor; suPAR, soluble urokinase plasminogen activator receptor;
DKK1, Dickkopf-1; LRP 5/6, lipoprotein receptor-related protein 5/6; Fzd, Frizzled; Ca, calcium; VDR, vitamin D receptor; P, phosphorus; 1,25-
(OH)
2
D, 1,25-dihydroxy-vitamin D.
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parathyroid gland to respond to calcium and vitamin D receptor
(VDR) signaling pathway, thus exacerbating SHPT (25). Klotho is a
calcium–phosphorus regulatory protein that has the ability to
increase urinary phosphorus and prevent urinary calcium loss. It
is tissue specific for FGF23, converting FGF23 receptor 1 (FGFR1)
to a specific receptor for FGF23. In Klotho-deficient mice, vascular
calcification, hyperphosphatemia due to abnormal calcium/
phosphate metabolism, and shortened lifespan characterize the
development (26,27). The FGF23–Klotho axis can be disrupted
in early CKD, which is characterized by decreased Klotho
expression and increased FGF23 levels in serum. In the absence
of Klotho, FGFR1 is underexpressed in the parathyroid glands and
serum FGF23 levels are elevated, leading to a series of mineral
metabolism disorders, SHPT, vascular calcification, and cardiac
hypertrophy. Exogenous Klotho may ameliorate or prevent the
progression of CKD-MBD (28). The Wnt signaling pathway (Wnt/
beta-catenin) promotes bone formation and can affect bone
remodeling by modulating the biological function of osteoblasts
and osteoclasts. Wnt inhibitors (a combination of wingless and int)
play a role in the pathogenesis of CKD-MBD. Wnt inhibitors,
including Dickkopf-1 (Dkk1) and sclerostin, are secreted at
increased levels in response to renal injury (29,30).
Overexpression of Dkk1 leads to decreased levels of beta-catenin,
which reduces the number of osteoblasts, inhibits bone formation,
and induces osteoclast differentiation and promotes bone
resorption, leading to severe bone metabolic disorders (31).
Activin A originates in renal-injured peritubular myofibroblasts
and acts through the activin II type A receptor (ActRIIA) (32). In a
mouse model, activation and inhibition of ActRIIA using the ligand
trap RAP-011 (a fusion of the soluble extracellular structural
domain of ActRIIA to a mouse IgG-Fc fragment) were separately
evaluated for their roles in the pathogenesis of CKD-MBD (33).
Activation of ActRIIA decreased Klotho expression and induced
osteodystrophy and fibrosis, whereas inhibition of ActRIIA
signaling was observed to reverse and ameliorate these changes
(33,34). Soluble urokinase receptor (uPAR) and soluble urokinase
plasminogen activator receptor (suPAR), whose important cellular
source is immature myeloid cells in the bone marrow (35), refer to
circulating inflammatory biomarkers that play a pivotal role in the
pathogenesis of renal diseases (36–39). As a cell membrane receptor
distributed on the cell membranes of a wide range of
immunoreactive cells and vascular endothelial cells, uPAR is
involved in extracellular matrix degradation, inflammatory
responses and tissue fibrosis by regulating the fibrinogen
activation system (40). As the soluble form of uPAR shed in body
fluids, suPAR, which is present in the peripheral blood circulation,
is associated with inflammation and immune activation (41,42). By
virtue of impeding the formation of podocyte peduncles through
activation of b3 integrin on glomerular podocyte membranes,
suPARisabletoimpairglomerularfiltration and even cause
pathological outcomes such as severe renal failure (36). However,
there are currently no animal models that perfectly fit the clinical
characteristics of CKD-MBD, due to the complex pathogenesis.
3 Modeling of CKD-MBD
Animal models were constructed by mimicking the
development of human CKD-MBD. Most CKD-MBD animal
models were formed by extending the modeling time of CKD
animal models. However, the construction of animal models is
required to be completed in a short period to facilitate experimental
studies. Therefore, methods such as surgery, special diet, and
radiation are sometimes adopted to accelerate the progression of
CKD-MBD.
3.1 Surgical intervention
The surgical intervention method means that the kidney of the
subject is removed or destroyed in some way, and varying degrees of
kidney damage were caused, leading to CKD-MBD. 5/6
nephrectomy (Nx), unilateral ureteral obstruction (UUO), and
electrocautery are used as common surgical modeling methods.
3.1.1 5/6 Nx
5/6 Nx is a widely used method. After removing 5/6 of rat
kidneys, the residual renal units have the function of systemic blood
filtration, thus leading to glomerular hyperfiltration, which further
destroys glomerulosclerosis and the residual renal units, and results
in the interstitial fibrotic lesions of chronic renal failure
characterized mainly by glomerular hypertrophy and sclerosis
(43,44). Researchers have been preparing models of kidney
diseases by 2/3 or 3/4 nephrectomy since 1889 when the first
kidney-related animal models were created. However, no
significant signs of proteinuria, hypertension, or myocardial
hypertrophy were observed. Hence, Chauntin et al. (45) proposed
the 5/6 Nx modeling method in 1932. This procedure was
performed by first removing 2/3 of the kidney on one side of
Wistar rats and then the entire kidney on the opposite side 1 week
later. The rats were also characterized by significant proteinuria,
nitrogen retention, hypertension, and cardiac hypertrophy after
successful modeling. Jablonski et al. (46) reported that 150-day-old
female Wistar rats were selected for a long-term renal
osteodystrophy (ROD) model by the 5/6 Nx method in 1993
(Figure 2a). Blood samples were analyzed intermittently after
surgery, and the rats were killed when 340 days old. Samples of
the skull, residual kidney tissue, and bilateral femur, PTH, VD,
alkaline phosphatase (ALP), calcium, and phosphorus were found
to have varying degrees of change in the operated rats, and SHPT
was observed, with a significant decrease in ALP indicating long-
term obstruction of bone formation. The majority of the tested
animals had reduced transverse cross-sectional area of the diaphysis
and markedly increased bone resorption, with fibrous osteitis and
long bone chondromalacia. In 2018, after constructing a CKD-
MBD model by 5/6 Nx based on 8-week-old SD rats, the researchers
found the elevated serum creatinine (Scr), phosphorus and intact
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parathyroid hormone (iPTH) levels and the decreased blood
calcium levels compared with sham-operated rats (47,48). At the
same time, the model rats showed severe renal tubular injury and
inflammatory interstitial cell infiltration. They also had extensive
glomerulosclerosis, which was accompanied by a large number of
dilated renal tubules and interstitial fibrosis. In addition, these
CKD-MBD model rats displayed the significantly reduced bone
mineral density. In 2021, founded on building a new CKD-MBD rat
model using 5/6 Nx, Linna Liu et al. (49) revealed that serum urea
nitrogen (BUN)and Cr levels were significantly elevated. At the 16th
week of the experiment, the model rats showed significant lesions in
the mesorectum of the aorta. Furthermore, the expression of bone
morphogenetic protein 7 (BMP-7) was significantly down-regulated
in the vertebrae of the CKD-MBD model rats, and the values of
FIGURE 2
Simple models of CKD-MBD. (a) CKD-MBD model of 5/6 Nx; (b) CKD-MBD model of UUO; (c) CKD-MBD model of electrocautery; (d) CKD-MBD
model of electrocautery combined with left nephrectomy; (e) CKD-MBD model of adenine alone diet; (f)CKD-MBD model of high-phosphorus diet;
(g) CKD-MBD model of Cy/+ rat fed with casein diet; (h) CKD-MBD model of whole-body radiation in a puppy; (i) CKD-MBD model of localized
radiation in rats. DBA/2, Dilute brown non-Agouti; Cy/+, a genetic model of polycystic kidney disease; SD, Sprague-Dawley; Nx, Nephrectomy; HPD,
high-phosphate diet; SCD, standard chow diet; CKD-MBD, chronic kidney disease-mineral and bone disorder.
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BMD, BV/TV, Tb, and Tb.Th were significantly decreased, and
Tb.Sp was significantly increased. This model shows development
of mixed bone impairments in uremic animals and can be used to
study the early effects of CKD-MBD and the effects of different
treatment options on bone. The subtotal nephrectomy animal
model, a classic and mature model of CKD, has been mainly
applied to the study of the pathological mechanisms of chronic
renal failure. The model is equivalent to CKD stage 5 and GFR value
is less than 15 ml/min/1.73m
2
. Researchers have used this model to
evaluate and analyze the changes in bone tissue due to kidney injury
as they have gained a better understanding of CKD. Typical bone
abnormalities and vascular calcification are difficult to induce by 5/
6-NX (50), owing to the high renal compensatory capacity of
experimental rats, whose serum PTH, calcium, phosphorus, and
ALP are not markedly altered. Therefore, more time is needed for
developing the model of CKD-MBD. Infection and excessive blood
loss are risks involved in this method of modeling, with greater
surgical difficulty of control and high mortality (51,52).
3.1.2 UUO
CKD is a state of progressive renal fibrosis (53). The UUO model
is well established and has been used to explore renal
tubulointerstitial injury and progressive fibrosis (54). The
hyperplasia of the renal tubular and interstitial cells and the
aggregation and infiltration of macrophages and monocytes in the
renal parenchyma induced by the UUO model ultimately lead to
CKD-MBD due to tubulointerstitial fibrosis and tubular atrophy
caused by the activation of the RAS system (55–57). For instance, 6-
week-old male Sprague–Dawley (SD) rats (160–200 g) were
anesthetized with isoflurane. The left nephron and ureter were
exposed with a side abdominal incision, and the left ureter was
ligated in two places with 3-0 sutures. After surgery, the rats were fed
a high-phosphorus and low-calcium diet (1.2% Pi and 0.6% calcium)
for 8 weeks (Figure 2b)(58). Histopathological analysis using micro-
computed tomography (CT) and immunohistochemistry showed
increased bone resorption in UUO model rats. However, the levels
of Scr, phosphorus, intact PTH, and FGF23 were unremarkable in
UUO model rats. The inconsistency in these results might stem from
the compensatory renal excretory function of the contralateral
kidney, rendering it difficult to ascertain the metabolic state of the
bones solely based on serum biochemical markers. No further studies
on vascular calcification in this model were performed at that time.
The UUO model was dramatically disrupted by subtle biochemical
changes and should be used with caution in studies of CKD-MBD.
The UUO model is equivalent to CKD stage 4 and GFR in 15-29 ml/
min/1.73m
2
. The UUO model is an ideal renal injury model to study
the rapid progression of renal fibrosis with little effect on total
glomerular filtration rate.
3.1.3 Electrocautery combined with
single nephrectomy
Electrocautery refers to the establishment of a model of CKD-
MBD using needle-type electrocautery to damage the kidney cortex,
causing inflammatory reactions and cortical fibrosis, resulting in
elevated Scr and BUN. The renal failure model established by
electrocoagulation of the bilateral renal cortex was first proposed
by Gagnon et al. (59). Five-week-old C57/BL6 male mice were
anesthetized with ether, whose kidneys were exposed through a 2-
cm-long waist incision. The perirenal adipose tissue and capsule
were then peeled off and the renal epidermis was subjected to
electrocoagulation and electrocautery to a depth of ~1 mm, leaving
the renal cortex 2 mm from the perirenal hilum unabraded. The
same manipulations were performed on the left renal cortex 10 days
later (Figure 2c). Scr and BUN levels were significantly increased 4
weeks after bilateral renal cortices were electrocoagulated. The
model is equivalent to CKD stage 4 and GFR in 15-29 ml/min/
1.73m
2
. The previous method of bilateral renal cortical
electrocoagulation was modified by Lund et al. (60). The whole
cortex of the right kidney was cauterized, excluding a 2-mm zone
around the hilum, followed by total left nephrectomy 2 weeks later
(Figure 2d). Elevated levels of BUN, serum phosphorus and PTH,
hyperparathyroidism, conspicuous signs of femoral osteodystrophy
(including reduced number and area of osteoblasts, lower rates of
bone formation and bone mineralization deposition, decreased
bone density, severe fibrosis around the trabeculae, and enlarged
bone marrow cavities filled by fibrotic cells) were visualized after
successful modeling. Renal electrocautery combined with single
nephrectomy is a practical method for constructing CKD-MBD
models. The model is equivalent to CKD stage 5 and GFR value is
less than 15 ml/min/1.73m
2
. Restrictions of the electrocautery
method are not just that complications can be caused at the
beginning of the procedure, but also that the mortality of the
subject is increased by complications of surgery, anesthesia, and
late CKD (61).
3.2 Feeding intervention
Feeding modeling is a method by which animals are treated
with various nephrotoxic drugs, foods, or special diets, causing renal
unit injury, CKD, and then CKD-MBD. The causative agents
include adenine, high-phosphate diet, casein diet, and doxorubicin.
3.2.1 Adenine diet
By generating 2,8-dihydroxyadenine in vivo through the action
of xanthine oxidase that is deposited in the glomerular and
interstitial parts of the kidney, adenine helps form a foreign body
granulomatous inflammation and block the lumen of the renal
tubules to cause the corresponding cystic dilatation of the lumen of
the renal tubules. As the disease progresses, a large number of lost
renal units lead to CKD-MBD (62). Acute renal failure occurred in
patients with Lesch–Nyhan syndrome treated with adenine in 1974
(63). An animal model of renal injury induced by adenine diet was
first reported in 1986 (64). Male Wistar rats (~110 g) were fed
particles containing 0.75% adenine (adenine dose: 270–320 mg/kg/
day). Eight-week-old male SD rats (~200 g) were fed an adenine-
containing diet (0.75% adenine) for 4 weeks (65), and the rats had
raised levels of Scr, PTH, and phosphorus, reduced serum 1,25(OH)
2
D
3
, increased osteoid on the trabecular surface, active osteoblasts,
and reduced cancellous bone mineral density (Figure 2e). In
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addition, 12-week-old male Wistar-Jcl rats were fed an adenine diet
and exhibited a high turnover type of ROD (66). Male Wistar rats
were fed a diet containing 0.25% adenine and low in vitamin K (67)
to generate a CKD-MBD model. Chronic renal failure in rats can be
induced by adenine diet, and also hyperparathyroidism and
disorders of bone and calcium–phosphorus metabolism can be
caused by this diet. Thus, a stable, highly reproducible, and highly
transformative animal model of CKD-MBD was successfully
established. More severe bone disease and vascular calcification
can be manifested by the adenine-induced CKD-MBD animal
model without invasive surgery (68,69), and thus the difficulty of
modeling and mortality are significantly reduced. However, some
critical issues in elucidating the pathophysiological mechanisms of
CKD-MBD exist in this model. Theoretically, bone metabolism is
affected by systemic toxicity or organ-specific damage caused by
adenine (62). The model is equivalent to CKD stage 5 and GFR
value is less than 15 ml/min/1.73m
2
. Chronic renal failure is caused
first, with renal bone disease then being caused by chronic renal
failure as the first possible pathway. Bone metabolism directly
affected by adenine is the second possible pathway, but the direct
mechanisms by which it affects bone metabolism have not been
reported. Weight loss, malnutrition, and systemic inflammation can
be induced by an adenine diet.
3.2.2 High-phosphorus diet
High-phosphorus diet increases the risk of decreased renal
function (70) and has detrimental effects on bone health (71,72).
By increasing blood phosphorus levels, inhibiting calcium-sensitive
receptor and vitamin D activation, and stimulating the
overproduction of PTH and FGF23, high-phosphorus diets cause
calcium and phosphorus metabolism disorders, enhanced bone
resorption and mineralization disorders. Meanwhile, calcium
phosphate deposition triggers ectopic calcification of the
vasculature and soft tissues, ultimately leading to CKD-MBD
(73). A novel model of CKD-MBD using only a high-phosphorus
diet has been created in recent years (61). DBA/2 mice were fed with
a high-phosphorus diet (20.2 g phosphorus, 9.4 g calcium,0.7 g
magnesium, and 500 IU/kg vitamin D3) for 4 or 7 days, followed by
standard chow diet (7.0 g phosphorus, 10.0 g calcium, 2.2 g
magnesium, and 1000 IU/kg vitamin D3), and followed until day
84 (Figure 2f). The experimental mice were found to develop
phosphate nephropathy, as demonstrated by tubular atrophy,
interstitial fibrosis, reduced glomerular filtration rate, elevated
serum urea, as well as SHPT, arterial calcification, and reduced
tibial bone volume and mineralization. The model is equivalent to
CKD stage 5 and GFR value is less than 15 ml/min/1.73m
2
. The
high mortality in animals due to surgical modeling was reduced
because the model excluded the effects of surgical intervention, and
because the low turnover bone disease was described for the first
time. In addition, progression of CKD-MBD was better simulated
by this model, rendering it a new, noninvasive, easy-to-perform,
and reproducible model. However, limitations of the model include
prolonged breeding and close monitoring of animals. The mortality
of the mice fed high-phosphorus diet for >10 days increases rapidly.
Lastly, researchers found that the model differed in susceptibility to
vascular calcification by comparing CKD patients to this CKD
mouse model (61).
3.2.3 Casein diet
Casein diet by regulating phosphorus and calcium intake result
in hyperphosphatemia and increased PTH secretion, accompanied
by renal fibrosis and mineral metabolism disorders, which
ultimately leads to CKD-MBD (74,75). The heterozygous Cy/+
rat is a genetic model of polycystic kidney disease that can be
developed into CKD-MBD on a special diet (75). Male Cy/+ rats are
consistent in the progressive development of nephropathy and have
several features of advanced CKD-MBD (76). Cy/+ rats were fed
with a high-casein diet (18% casein-based protein, 0.7% phosphate,
0.7% calcium, and 5% fat) and were sampled at 10, 34, and 38 weeks
of age, which showed persistent azotemia beginning at 10 weeks of
age, hyperphosphatemia, and hyperparathyroidism at 34 weeks of
age, vascular calcification at 38 weeks of age, and uremia at ~40
weeks of age (Figure 2g). Dietary protein type affecting the
progression of CKD-MBD and renal dysfunction were confirmed
by this model, and casein was introduced as a novel dietary
modeling method, alongside further exploration of the
mechanisms involved; namely that a casein-based diet increases
the concentration of FGF23, resulting in hyperphosphatemia to
complete the modeling. The model is equivalent to CKD stage 5 and
GFR value is less than 15 ml/min/1.73m
2
. This model, as the first
CKD-MBD model that occurs spontaneously under a normal
phosphorus diet without surgical or pharmacological
involvement, can simulate the development of human CKD,
study the early changes of CKD-MBD, and evaluate the effects of
different dietary regimens on the course of CKD-MBD.
3.3 Radiation
Radiation is an animal modeling method to induce CKD-MBD,
similar to bone disease due to chronic renal failure. The systemic
radiation contributes to anemia and immunosuppression by
destroying the hematopoietic function of the bone marrow (77).
At the same time, systemic radiation induces oxidative stress and
inflammation (78), which exacerbates mineral metabolism
disorders, ultimately leading to CKD-MBD. The dose and timing
of radiation are important because too small a dose makes it difficult
to produce visible kidney damage, while too large a dose can cause
gastrointestinal damage (79,80). Two-day-old puppies were
exposed to sublethal doses of 60Co gamma radiation in 1981
(Figure 2h)(81), inducing varying degrees of renal failure, with
hyperparathyroidism, altered osteochondrosis, increased bone
remodeling, and reduced bone mineral density. The model is
equivalent to CKD stage 5 and GFR value is less than 15 ml/min/
1.73m
2
. Therefore, the radiation method can be applied to the study
of CKD-MBD in humans. The local radiation induces renal failure
by directly damaging renal tissues, which in turn leads to CKD-
MBD. The direct damage to renal tissues can cause renal
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hypoplasia, which in turn triggers inflammatory reactions and
fibrosis, as well as disorders of mineral metabolism, ultimately
leading to the pathologic features of CKD-MBD (82).
The method of inducing impaired bone metabolism by
establishing renal injury through localized radiation to the
kidneys based on whole-body radiation was proposed by Ming-
Yu Wu et al. (82). The bilateral kidneys of 3-month-old male SD
rats (280–310 g) were exposed through longitudinal incisions on
both sides of the spine. The rats were fixed in the lateral recumbent
position on a radiation rat mold, and the tissues other than the
kidneys were shielded by lead plates. The exposed kidneys were
irradiated by gamma rays at 15 Gy (Figure 2i). Indicators of bone
mass, three-point bending load on the femur, and compressive load
on the lumbar spine were significantly reduced in the subjects after
3 months of local kidney irradiation. Bone morphology tests
showed diluted bone trabeculae and accelerated bone conversion.
In addition, renal-radiation-injury-induced bone metabolism
disorders are similar in clinical manifestations to renal bone
disease due to chronic renal failure, with frequent manifestations
of osteoporosis, fibrous osteitis, resting bone disease, and
osteochondrosis, which predispose to fractures. The model is
equivalent to CKD stage 5 and GFR value is less than 15 ml/min/
1.73m
2
. In conclusion, definite bone changes can be caused by
systemic radiation and the modeling process is similar to the
progression of renal failure. However, clinical features more
similar to CKD-MBD are demonstrated by localized radiation of
the kidneys. Shortcomings of the radiation method are the long
modeling times due to slowly developing radiation damage to the
kidneys. Different nephrotoxicity thresholds are available in
different radiological entities (83), so the difficulty of modeling is
heightened by the radiation dose and length of time required
for radiation.
3.4 Improved models
The improved versions of the models refer to the derivation or
combination of the above models to obtain a model with a shorter
modeling time, greater efficiency, and greater suitability for
research purposes.
3.4.1 Partial nephrectomy combined with high-
phosphorus diet
5/6 NX brings severe damage to renal function, such as
increasing the burden on the residual kidneys and decreasing the
glomerular filtration rate to a high degree. Founded on this, a high-
phosphorus diet further exacerbates the abnormalities of bone
metabolism and vascular calcification, thus leading to the
development of CKD-MBD (84). Modeling with a high-
phosphorus diet based on a 5/6 Nx model can achieve better
modeling results for CKD-MBD in a short period. Male Wistar
rats (200–225 g) received a high-phosphorus diet (0.8% calcium and
0.93% phosphorus) for 3 weeks before starting two-stage 5/6 Nx
(85). Two branches of the left renal artery were ligated under
anesthesia, followed by removal of the right kidney 2 weeks later
(Figure 3a). Execution and sampling were performed at 6 and 12
weeks, followed by testing that revealed elevated PTH,
hyperphosphatemia, hypocalcemia, and SHPT, as well as
increased mineral deposition rates, bone formation rates,
osteoblast perimeter, and erosion perimeter. The model can be
used to assess the effects of dietary phosphorus and the severity of
renal failure on morphological changes in bone histology and
various biochemical markers. Similarly, Scr, FGF23, and
phosphorus were significantly elevated, and total serum ALP
activity was increased with severe SHPT in 5/6 Nx mice fed a
high-phosphorus diet (2.0% calcium, 1.25% phosphorus, 20%
lactose, and 600 IU VD/kg) for 8 weeks postoperatively (86). Low
total and cortical bone density of the spine and proximal tibial
epiphysis, as well as significant signs of impaired bone
mineralization were detected. The CKD-MBD rat model was
developed by Linna Liu et al. (49) using 5/6 Nx combined with a
high-phosphorus diet (0.5 g sodium dihydrogen phosphate, 0.5 g
sodium dihydrogen phosphate). Alternatively, male SDT and SD
rats were used by Kentaro Watanabe et al. and were divided into
experimental and control groups (48). The experimental group had
2/3 of their left kidney removed at 8 weeks of age, followed by CKD
being established by total nephrectomy of the right kidney 1 week
later. Then CKD-MBD animal model was established by feeding a
high-phosphorus diet (1.0% calcium and 1.2% phosphate) at 10
weeks of age. SDT-Nx rats that had undergone 5/6 Nx compared
with SD nephrectomy rats by 20 weeks of age showed more
dramatic changes in CKD-MBD parameters, including vascular
calcification, serum PTH, FGF23, serum calcium and phosphorus
levels, and urinary excretion of calcium and phosphorus. Partial
nephrectomy combined with a high-phosphorus diet is regarded as
an optimal mouse model of chronic renal failure, with
characteristics such as malnutrition, hypertension, and disturbed
calcium and phosphorus metabolism, thus making an ideal model
for studying CKD-MBD. The model is equivalent to CKD stage 5
and GFR value is less than 15 ml/min/1.73m
2
. SDT-Nx rats can also
be used to examine the pathophysiology of CKD-MBD.
3.4.2 Unilateral nephrectomy combined with
adenine diet
Left nephrectomy aggravates the burden on the remaining
kidneys, resulting in reduced renal function. Moreover, because
adenine is nephrotoxic, an adenine diet causes interstitial fibrosis
and tubular damage, which further impairs residual renal function and
triggers abnormal mineral metabolism, ultimately leading to CKD-
MBD (51). The experiment was conducted using 200–220 g Male SD
rats (87) housed at 22 ± 3°C with 50± 0% humidity on a 12-h light/
dark cycle, and were fed standard rat chow of specificpathogen-free
grade. The rats were subjected to left-sided nephrectomy on day 7 and
given 2% adenine (150 mg/kg/day) on days 8–21 (Figure 3b). Renal
insufficiency, tubular interstitial injury, disturbance of calcium and
phosphorus metabolism, and bone abnormalities were found 3 weeks
after the induction of renal injury. The model is equivalent to CKD
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stage5andGFRvalueislessthan15ml/min/1.73m
2
.Significant
vascular calcification was present in this modified CKD-MBD rat
model due to the use of adenine, and renal injury and bone
abnormalities were more easily studied.
3.4.3 Nephrectomy combined with Adriamycin
The compensatory hyperfiltration state of the residual kidneys after
left nephrectomy results in intraglomerular high pressure, proteinuria
and oxidative stress, which further harms renal structure. Additionally,
FIGURE 3
Improved models of CKD-MBD. (a) CKD-MBD model of partial nephrectomy combined with high-phosphorus diet; (b) CKD-MBD model of
unilateral nephrectomy combined with adenine diet; (c) CKD-MBD model of nephrectomy combined with doxorubicin; (d) CKD-MBD model of
LDLR
-
/
-
mice with electrocautery combined with nephrectomy and special dietary intervention; (e) CKD-MBD model of fed with high-phosphorus
and adenine diet. SD, Sprague-Dawley; SDT, spontaneously diabetic Torii; LDLR=low density lipoprotein receptor; Nx, Nephrectomy; HPD, high-
phosphate diet; SCD, standard chow diet; SPF, specific pathogen-free; CKD-MBD, chronic kidney disease-mineral and bone disorder.
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adriamycin causes glomerular shrinkage, glomerulosclerosis, tubular
atrophy and tubulointerstitial fibrosis (88), and left nephrectomy
combined with adriamycin leads to deterioration of renal function
and the development of bone pathology, ultimately leading to CKD-
MBD. Nephrotoxic drugs were used by Liu et al. (89)basedon1/2
nephrectomy; i.e., the surgical modeling method was combined with the
drug–food modeling method. Thus, a new, short-term model of CKD-
MBD was formed. Male SD rats of 120–150 g were selected and
anesthetized under pentobarbital (60 mg/kg) for left nephrectomy
with intravenous doxorubicin (dissolved in 0.9% saline, 5 mg/kg),
whereby an ROD model was created, and experimental sampling was
performed at various times after surgery (Figure 3c). Marked increases in
BUN, Scr, Uric Acid(UA), and Urea-Creatinine Ratio (UCR), and
decreases in serum albumin were indicated in subjects with late ROD
(1 week < duration of disease ≤1 month). Overt renal injury was also
found. Low transforming bone lesions were revealed by bone
morphology that were characterized by marked decreases in bone
formation rate, osteoclasts, osteoblasts, and trabecular volume
thickness, as well as a significant increase in osteoid volume. This
experimental model is proposed as a highly reproducible model of
kidney injury, whose time to induction is short and time to injury is
predictable and consistent. Therefore, this model can also be used to test
interventions that exacerbate or prevent kidney injury. The model is
equivalent to CKD stage 4 and GFR in 15-29 ml/min/1.73m
2
.
Additionally, the process of human CKD development can be better
simulated on account of the high similarity between the type of
structural and functional impairment of the model and that of human
chronic proteinuria nephropathy.
3.4.4 Electrocautery combined with
nephrectomy and special dietary intervention in
LDLR
-
/
-
mice
By directly injuring the renal tissues, both left nephrectomy and
electrocautery trigger inflammatory reactions and fibrosis, causing
hyperphosphatemia, increased PTH secretion and abnormal bone
metabolism, ultimately leading to CKD-MBD (50). Therefore, better
modeling results will be achieved by combining the two procedures.
LDLR
-
/
-
mice were used by Davies in the preparation of this model
(90), with standard diet-fed mice being given a high-fat diet for 2
weeks at 10 weeks of age. The experimental manipulations were
carried out following the procedure previously described by Gagnon
(59) and Lund (60) at 12 weeks of age. The rate of mineral deposition
in the cancellous bone of the distal femur was significantly reduced,
osteoblasts were reduced, bone formation was slowed, and low
turnover of osteodystrophy was observed in mice fed a high-fat
diet. The right kidney of 12-week-old LDLR
-
/
-
mice was subjected to
electrocautery through a 2-cm lateral incision pair (91), with mild
and moderate kidney injury being produced according to the degree
of cautery. The left kidney was removed through a similar incision in
mice at 14 weeks, followed feeding a high-fat diet until 22 or 28 weeks
(Figure 3d). The final experiment showed suppressed bone formation
rate, decreased cortical bone density, decreased bone area, increased
osteoclast secretion, and vascular calcification. In addition, BUN,
calcium, phosphate, and PTH were elevated at week28. The degree of
renal injury was lowered by reducing the area of electrocautery in the
right kidney. Early CKD (stages 2 and 3) in the model group was
judged by utilizing inulin clearance, and CKD-MBD appeared early.
The model is equivalent toCKD stage 5 and GFR value is less than 15
ml/min/1.73m
2
.
3.4.5 Combined high-phosphorus and
adenine diet
Adenine-fed renal pathology stems from the formation of 2,8-
dihydroxyadenine, an adenine metabolite that crystallizes in renal
tubules, which leads to an inflammatory response, oxidative stress,
tubular atrophy and renal parenchymal fibrosis. A high-phosphorus
diet increases the phosphorus load in the body and damaged kidneys
can’t excrete phosphorus efficiently, leading to elevated blood
phosphorus and stimulating increased secretion of PTH, which
triggers disorders of mineral metabolism and ultimately leads to
CKD-MBD. Eight-week-old male C57BL/6J mice were used by
Takashi et al. in 2017 (73). A novel CKD mouse model with
adenine and high-phosphate diet for assessing the progression of
hyperphosphate and associated mineral bone disease, and the longer
the high-phosphorus diet was fed, the greater the volume of
calcification was found. Twenty-week-old male C57/BL6J mice were
recently used to create a CKD-MBD model (92)inwhichCKDwas
induced by feeding a 0.2% adenine and 0.8% phosphorus diet for 6
weeks, followed by induction of CKD-MBD by feeding a 0.2% adenine
and 1.8% phosphorus diet for 6 weeks (Figure 3e). Elevated creatinine
and phosphorus, decreased calcium, SHPT, thin and irregular femoral
cortex, visibly reduced cortical bone mineral density and cortical bone
thickness, and reduction in bone volume and trabecular number were
detected after successful modeling. The model is equivalent to CKD
stage 5 and GFR value is less than 15 ml/min/1.73m
2
.The
complications associated with medial arterial calcification and ROD
in patients with CKD are mimicked by this modeling approach, and
developed severe vascular calcification without surgery.
4 Quality evaluation of animal models
of CKD-MBD
No clear assessment criteria are seen in animal models with CKD-
MBD, and serum biochemical tests, renal histopathological tests, bone
tissue-related index tests, and vascular calcification are commonly used
for the evaluation and diagnosis of CKD-MBD (93,94).
4.1 Serum biochemical tests
CKD-MBD is treated as a disease secondary to CKD. CKD-
MBD is regarded as a secondary disease of chronic kidney disease
(CKD), and its animal model needs to be evaluated by serum
biochemical tests to reflect the key indicators of renal function and
mineral metabolism disorders. The commonly used assays include
serum Scr, BUN, calcium, phosphorus, 1,25-(OH)
2
D, PTH (95,96)
and ALP (73,97,98). Additionally, in order to improve the validity
and reproducibility of the model, it is recommended to incorporate
statistical analyses to validate the sensitivity and specificity of these
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assays, and to ensure the consistency of the experimental conditions
(e.g., feed formulations, surgical procedures) in order to minimize
bias due to individual differences.
4.2 Renal histopathology
Renal histopathology serves as an important tool for evaluating
animal models of CKD-MBD. Pathological changes in the kidneys,
including glomeruli, tubules, and interstitium, are usually visualized
under light microscopy after hematoxylin–eosin staining,
peroxynitrite Schiff staining, and Masson staining. To enhance
the clinical relevance of the model, quantitative analyses (e.g.,
glomerular sclerosis rate, percentage of fibrotic area) should be
combined to quantify the pathological changes and compared with
the pathological characteristics of human CKD-MBD patients to
validate the representativeness of the model.
4.3 Bone tissue-related indicators
Bone tissue-related index tests have been used as an important
indicator for the diagnosis of CKD-MBD. The lumbar spine, femur,
and tibia are often tested in experiments, with the femur being the
most commonly tested (75,87).The presence of abnormalities in
bone transformation, mineralization, bone volume, linear bone
growth, or bone strength is clarified by detecting bone mineral
density (99), hematoxylin–eosin staining of bone sections (87,100),
Masson staining, tartrate-resistant acid phosphatase (TRAP staining),
Goldner’sstaining(101), immunohistochemical staining, and micro-
CT, and the type of bone transformation can be predicted by the
results of the assay (102–104). By combination with the dynamic
bone metabolism markers detection, the progression pattern of bone
lesions can be observed through long-term follow-up, thus enhancing
the validity, time-dependence and clinical relevance of the model.
4.4 Vascular calcification
Damage to the cardiovascular system in CKD-MBD is
considered an important factor in mortality (1). Experimental
models are often assessed by vascular calcium content
measurement, Von Kossa staining of aortic segments, and
percentage of aortic calcified plaque area (50,105,106). To
improve the reproducibility and clinical relevance of the model, it
can be combined with vascular endothelial function test as well as
inflammatory factor test to comprehensively reflect the pathological
mechanism of vascular calcification and to be compared with the
vascular lesion characteristics of human CKD-MBD patients.
4.5 Other assessment methods
The establishment of CKD-MBD animal models should also
consider the validity, reproducibility and clinical relevance. Firstly,
in terms of the model validity, it can be statistically verified that the
key features of the model (e.g., serum biochemical indexes,
pathological changes and vascular calcification) are at the
expected level and ensure that the experimental results are
significant and biologically meaningful. Secondly, in terms of the
model reproducibility, a standardized operation procedure,
encompassing experimental design, animal selection, surgical
operation and detection methods, can be established to verify the
stability and consistency of the model through multiple batches of
experiments. Finally, the clinical relevance of the model can be
compared with the clinical data and pathological features of human
CKD-MBD patients to verify whether the animal model can
accurately simulate the pathophysiological process of
human disease.
A well-established model of CKD-MBD should resemble the
alterations in bone, renal function, and electrolytes of clinical
patients, in addition to the above-mentioned indicators being
examined. Also, the model should be altered by chronic renal
failure and not by other diseases or drugs. The model is required
to be broadly representative, highly stable, and reproducible, while
simple to operate, take a short time to achieve, and be capable of
representing renal bone disease from various causes.
5 Discussion and conclusion
The clinical presentation of patients with CKD-MBD varies with
the main metabolic abnormalities and characteristic bone disease of the
patient. CKD-MBD is primarily characterized by ROD, leading to
weakness, fractures, bone and muscle pain, and ischemic necrosis.
Hyperconversion osteodystrophy, hypoconversion osteodystrophy,
mixed ROD, and b2-microglobulin amyloidosis osteodystrophy are
specifically included in ROD. The treatment of CKD-MBD has so far
focused on phosphate retention, abnormal VD metabolism, and PTH
disruption, but the strategies have largely proved to be unsuccessful.
Recently, a single modeling method has been found to have a long
modeling time and other obvious shortcomings as the animal models of
CKD-MBD have been improved. For example, the mortality rate of 5/6
Nx is high, the experimental subjects of high-phosphorus diet modeling
are limited and have different sensitivities, and the radiation dose and
duration of radiation modeling are difficult to control. The modified
modeling methods were found to be noticeably more advantageous
than single modeling methods concerning the modeling time.
Unilateral nephrectomy combined with adenine diet is recommended
considering the cost of modeling and the difficulty of the operation.
Previous surgical modeling methods have resulted in high mortality
rates due to postoperative infections and other factors. However,
penicillin given to SD rats after unilateral nephrectomy reduced the
risk of infection and mortality. Adenine diet given a few days after
nephrectomy was effective in preventing the rats from developing rapid
malnutrition and death. Unilateral nephrectomy combined with
adenine diet for modeling is simple to perform, low in cost and
mortality, and deserves to be further promoted or improved upon.
However, no widely accepted method of model preparation
exists at present. Further practical studies on the pathogenesis of
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CKD-MBD will provide new ideas for animal modeling. For
instance, proliferation and differentiation of osteoblasts and
osteoclasts can be promoted by thyroid hormone (TH); the bone
turnover rate is high and the bone remodeling time is short in the
hyperthyroid state. Also, previous experiments have shown that the
bone turnover rate of rats can be increased by enhancing TH.
Therefore, can we supplement TH by thyroxine liquid gavage or
other ways based on an adenine diet to cause a high-conversion
CKD-MBD animal model? Bone metabolism can be regulated by
PTH through different signaling pathways; for example, the Gq/
PLC/PKC signaling pathway can be upregulated by increasing PTH,
thereby inhibiting bone formation. Therefore, can we upregulate the
Gq/PLC/PKC signaling pathway by administering PTH
intramuscularly based on the CKD rat model to inhibit bone
formation, thus creating a low conversion CKD-MBD animal
model with more similar clinical manifestations and more severe
vascular calcification? Excessive PTH stimulates increased bone
fibroplasia and osteoid formation, and tends to slow down bone
mineralization if accompanied by low blood calcium and
phosphorus. Then, can we make a mixed CKD-MBD model
based on the model of SHPT by decreasing blood calcium and
phosphorus in experimental animals through subcutaneous
injection of calcitonin, which subsequently leads to insufficient
bone mineralization? Finally, osteoarthropathy can be caused by
continuous aggregation of b(2)-microglobulin; therefore, can we
develop b(2)-microglobulin amyloid osteoarthropathy in
experimental animals by local injection of b2-microglobulin at
the joints? If the above methods are feasible, they will be more
convenient than the existing methods of CKD-MBD modeling,
more consistent with clinical characteristics and pathogenesis, and
more targeted to create different bone transition types required for
experiments. These modeling methods still need to be further
studied. Researchers should start from the pathogenesis of CKD-
MBD and experimental purposes, find some modeling methods to
establish more consistent clinical features and pathogenesis of
human CKD-MBD, to provide more new strategies for clinical
treatment and disease prevention.
The various methods of modeling CKD-MBD, including
surgical, drug and food, radiation, and modified modeling
methods, have been established from the pathogenesis and
clinical features of CKD-MBD (Table 1). Every modeling
approach has its own advantages; for example, models with slow
disease progression are advantageous because they are more likely
to translate to chronic kidney disease in humans, while surgical,
radiation, and high-dose adenine models may more often simulate
kidney disease after acute kidney injury. However, many animal
models of CKD-MBD have the disadvantages of long modeling
time, difficult handling, and high mortality (Table 2). Therefore,
subject selection should be considered by the laboratory
workers, including the species, age, weight, and whether the
animals have underlying diseases. The operation method,
technical requirements, anesthesia dose, time and content of the
extraction, and mortality should be controlled by the experimenter
for each modeling method.
TABLE 1 Summary of CKD-MBD modeling methods and disease progression-related parameters.
Models Modeling
time
parameter variation Disease
severity
References
5/6 Nx 13 weeks ↑Scr,↑P,↓Ca,↑iPTH, Renal tubular injury, inflammatory interstitial cell infiltration, increased
bone resorption, fibrous osteitis and long bone chondromalacia
CKD stage 5 (46,50–52)
UUO 8 weeks ↑BUN, slightly elevated iPTH, no change in Ca and P, tubulointerstitial fibrosis and tubular
atrophy, increased bone resorption
CKD stage 4 (55–58)
Electrocautery 12 weeks ↑P,↑iPTH, no change in Ca, significant depressions in osteoblast number, perimeters, bone
formation rates, and mineral apposition rates
CKD stage 4 (59–61)
Adenine diet 4 weeks ↑Scr,↑P,↑iPTH, ↓1,25(OH)
2
D
3
, increased osteoid on the trabecular surface, active osteoblasts,
and reduced cancellous bone mineral density
CKD stage 5 (65–69)
High-
phosphorus diet
12 weeks ↑Scr,↑P,↓Ca,↑iPTH, renal tubular atrophy, interstitial fibrosis, vascular calcification, and
decreased tibial bone volume and mineralization
CKD stage 5 (61)
casein diet 40 weeks ↑Scr,↑BUN,↑P,↓Ca,↑iPTH, SHPT, vascular calcification CKD stage 5 (75,76)
Whole
body radiation
Within
2 years
↑P,↓Ca, SHPT, Osteochondrosis changes, increased bone remodeling, decreased bone density CKD stage 5 (81)
Local radiation 12 weeks Accelerated bone turnover, osteoporosis, fibrous osteitis, resting bone disease
and osteochondrosis
CKD stage 5 (82)
5/6 Nx+HPD 12 weeks ↑Scr,↑P,↓Ca,↑iPTH, increased rates of mineral deposition, bone formation, osteoblast
circumference and erosion circumference
CKD stage 5 (48,49,85,86)
left
nephrectomy
+adenine
3 weeks ↑Scr,↑P,↓Ca,↑iPTH, tubular interstitial injury, bone abnormalities CKD stage 5 (87)
(Continued)
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TABLE 1 Continued
Models Modeling
time
parameter variation Disease
severity
References
Left
nephrectomy
+Adriamycin
3 weeks ↑Scr,↑BUN, Renal inflammatory cell infiltration, renal tubular collapse, and low transforming
bone lesions
CKD stage 4 (90)
Electrocautery
+left nephrectomy
28 weeks ↑Scr,↑P,↓Ca,↑iPTH, Vascular calcification, decreased cortical bone density, decreased bone
area and increased osteoclasts
CKD stage 5 (91)
Adenine
+phosphorus diet
12 weeks ↑Scr,↑P,↓Ca,↑iPTH, thin femoral cortex, reduced cortical bone mineral density and cortical
bone thickness, and reduction in bone volume and trabecular number
CKD stage 5 (92)
5/6 Nx: 5/6 nephrectomy; UUO: unilateral ureteral obstruction; HPD: high-phosphate diet;↑: increased; ↓: decreased; Scr: serum creatinine; P: phosphate; Ca: calcium; iPTH: intact parathyroid
hormone; BUN: urea nitrogen; SHPT: secondary hyperparathyroidism; CKD: chronic kidney disease.
TABLE 2 Common animal modeling methods for CKD-MBD and their advantages and disadvantages.
Models Species Advantages shortcomings Application type References
5/6 Nx Rat stable, widely used, classic
and mature
long modeling time, operation
difficulty, high mortality, insufficient
bone abnormalities and
vascular calcification
Chronic renal failure, uremic
mixed bone disease, and
rapid progression of
renal fibrosis
(46,50–52)
UUO Rat mature Unknown bone metabolic state Rapid progression of
renal fibrosis
(55–58)
Electrocautery Mouse reproducible, little bleeding long modeling time, early
Complications, difficult
Uremic mixed bone disease (59–61)
Adenine diet Rat stable, Simple, high repeatability,
low mortality, Significant bone
disease and vascular calcification
unknown mechanism, weight loss,
malnutrition, systemic
inflammatory response
Chronic renal failure, highly
transformed bone disease
(65–69)
High-phosphorus diet Mouse simple, high repeatability, low
mortality, similar clinical feature
long modeling time, limited
experimental subject,
different susceptibility
Lowly transformed
bone disease
(61)
casein diet Rat simple, no surgery, no drug,
simulate disease progression,
evaluate diet plan
long modeling time, high Cost Chronic renal failure, highly
transformed bone disease
(75,76)
Whole body radiation puppy definite osteomalacic changes,
Similar renal failure progression
high mortality, limited exposure to
dogs and radiation sources
Chronic renal failure, uremic
mixed bone disease
(81)
Local radiation Rat similar clinical feature long modeling time, the dose and
duration of radiation are difficult
to control
Chronic renal failure, uremic
mixed bone disease
(82)
5/6-Nx+HPD Rat similar clinical features, drastic
parameter change,
pathophysiology examination
SD +Nx: insufficient vascular
calcification; SDT +Nx: unknown
mechanism, basic disease
Chronic renal failure, highly
transformed bone disease
(48,49,85,86)
left
nephrectomy+adenine
Rat significant bone damage and
skeletal abnormalities
high risk of surgery Uremic mixed bone disease (87)
Left
nephrectomy
+Adriamycin
Rat short molding time, drastic
parameter change, high
repeatability, highly similarity,
predictable damage
unknown mechanism, Adriamycin’s
batch difference, individual
response differences
Lowly transformed
bone disease
(90)
electrocautery
+left nephrectomy
Mouse clear mechanism, obvious
bone lesions
narrow application range, the
complexity of modeling,
high mortality
Lowly transformed
bone disease
(91)
Adenine
+phosphorus diet
Mouse simple, no surgery, no drug,
severe vascular calcification
narrow application range, unknown
mechanism, weight loss, malnutrition,
systemic inflammatory response
Uremic mixed bone disease (92)
5/6 Nx: 5/6 nephrectomy; UUO: unilateral ureteral obstruction; HPD: high-phosphate diet.
Tan et al. 10.3389/fendo.2025.1549562
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This review summarizes the current methods for modeling
CKD-MBD, their advantages and disadvantages, and scope of
application according to the pathogenesis and clinical
characteristics of CKD-MBD, combined with the serum
biochemical indexes, vascular calcification, and pathological
changes of kidney and bone, which provides a more convenient
reference for researchers to select, establish, and customize animal
models for CKD-MBD research.
Author contributions
BT: Writing –original draft, Writing –review & editing. WT:
Writing –original draft, Writing –review & editing. YZ: Writing –
original draft. JL: Writing –original draft. XD: Writing –original
draft. HS: Writing –original draft. XP: Writing –original draft,
Writing –review & editing. LL: Writing –original draft, Writing –
review & editing. QH: Writing –original draft, Writing –review
& editing.
Funding
The author(s) declare that financial support was received for the
research and/or publication of this article. This work was supported
by the National Natural Science Foundation of China (82205002).
Sichuan Science and Technology Program (2022YFS0621). Science
and Technology Department of Sichuan Province
(2023NSFSC0655). Science and Technology Cooperation Projects
of the First People’s Hospital of Suining and the Southwest Medical
University (2022SNXNYD03). Southwest Medical University
Technology Program (2023ZD008). Southwest Medical University
Technology Program (2024ZXYZX02). Southwest Medical
University Technology Program (2023ZYQJ04), and Science and
Technology Research Special Project of Sichuan Administration of
Traditional Chinese Medicine (2024MS524).
Conflict of interest
The authors declare that the research was conducted in the
absence of any commercial or financial relationships that could be
construed as a potential conflict of interest.
Generative AI statement
The author(s) declare that no Generative AI was used in the
creation of this manuscript.
Publisher’s note
All claims expressed in this article are solely those of the authors
and do not necessarily represent those of their affiliated
organizations, or those of the publisher, the editors and the
reviewers. Any product that may be evaluated in this article, or
claim that may be made by its manufacturer, is not guaranteed or
endorsed by the publisher.
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