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September 1, 2012 | Veterinary Record | 219
ResearchResearch
PAPER
Acute phase protein concentration and metabolic
status affect the outcome of treatment in cows with
clinical and subclinical endometritis
M. Heidarpour, M. Mohri, A. H. Fallah-Rad, F. Dehghan Shahreza, M. Mohammadi
Veterinary Record
(2012) 171, 219
cite as doi:
10.1136/vr.100947
M. Heidarpour,
M. Mohri,
A. H. Fallah-Rad,
F. Dehghan Shahreza,
M. Mohammadi,
Ferdowsi University of
Mashhad, Iran
E-mail for
correspondence:
heidarpour@um.ac.ir
This is a summary
of a paper that is
published in full at
veterinaryrecord.
bvapublications.com
Published Online First
August 4, 2012
Context
Metabolic-related risk factors for uterine infections
include subclinical ketosis, hypocalcaemia, hypomag-
nesaemia and hepatocellular damage. It has also been
reported that the severity of uterine bacterial contamina-
tion, as determined by the bacterial growth density, was
correlated with the peripheral circulating concentrations
of acute phase proteins. Although the relationships of
metabolic parameters and acute phase proteins with the
occurrence of endometritis have been investigated, no
study has compared these parameters between clinical
and subclinical endometritis, or evaluated the effects of
these parameters on the outcome of treatment.
The aim of this study was to investigate the role of
acute phase protein concentration and metabolic status in
the establishment and persistence of clinical and subclini-
cal endometritis in dairy cows.
Main conclusion
Improved liver function and a decrease in the acute phase
protein concentration might favour the resolution of endo-
metritis after treatment.
Approach
The study involved 30 healthy cows, 30 cows with sub-
clinical endometritis and 30 cows with clinical endometri-
tis. Cows were in their second parity, kept in an open shed,
and received a well-balanced total mixed ration. They were
first examined between 21 and 33 days in milk. The cows
with subclinical endometritis received one of two treat-
ments: 500 mg of cloprostenol intramuscularly (Estroplan;
Parnell) on days 1 and 14, or 500 mg of intrauterine ben-
zathine cephapirin in 19.6 g ointment base (Metricure;
Intervet) and then 500 mg of cloprostenol intramuscularly,
seven days after the benzathine cephapirin injection.
Treatment protocols for cows with clinical endometri-
tis were: 500 mg of cloprostenol intramuscularly on days
1 and 14 or 1 mg/kg of sodium-ceftiofur intramuscularly
(Excenell; Pfizer Animal Health) for three consecutive days
and then 500 mg of cloprostenol intramuscularly seven
days after the last ceftiofur injection. After treatment and
taking the second samples (on day 7 after treatment) by
cytobrush from cows with subclinical endometritis, two
groups of animals were selected: cows with successful
treatment (defined as the presence of less than 18 per cent
polymorphonuclear [PMN] cells in endometrial samples)
and animals with unsuccessful treatment (more than 18
per cent PMN cells in endometrial samples). The same
criteria were applied for cows with clinical endometritis
and two groups were selected: successful treatment (no
abnormal discharge and less than 18 per cent PMN cells
in endometrial samples) and unsuccessful treatment (with
abnormal discharge and/or PMN more than 18 per cent in
endometrial samples).
Blood was taken at diagnosis (before treatment)
fromthe jugular vein of all cows with clinical and sub-
clinical endometritis and on day 7 after treatment. Blood
samples were also taken from the control group (n=30,
healthy cows with no clinical or subclinical endometri-
tis) on the day of the first examination. The activities of
alkaline phosphatase (ALP), aspartate aminotransferase
(AST), gamma glutamyl transferase, and the concentra-
tions of bilirubin, albumin, calcium, magnesium, phos-
phorous and β-hydroxybutyrate (BHB) in serum samples
were measured using commercial kits and an autoanalyser
(3000 Targa; Biotecnica). Serum total sialic acid (TSA)
concentration was determined using a method described
by Sydow in 1985. Serum haptoglobin concentration was
determined with a bovine haptoglobin ELISA kit (Bio-X
Diagnostics) according to the manufacturer’s instruction.
Analyses of variance with multiple comparison test (Tukey
Honestly) were used to compare means among the trial
groups. Student’s t test compared means before and after
treatment in cows with clinical and subclinical endometri-
tis. Statistical significance was P<0.05.
Results
Cows in the clinical and subclinical endometritis
groups had significantly higher BHB, haptoglobin and
TSA concentrations compared with the healthy con-
trol group of animals. A significantly lower serum
calcium concentration and a significantly higher serum
AST activity in the clinical endometritis group were
observed when compared with subclinical endometritis
and healthy groups. The comparison of parameters before
treatment indicated that cows suffering from clinical and
subclinical endometritis with lower concentrations of
hepatic markers (AST and ALP in clinical and subclini-
cal endometritis groups, respectively) and haptoglobin
showed a better response to further treatment, and endo-
metritis was not detected at the second examination.
Moreover, decreased concentrations of BHB, haptoglobin
and TSA and decreased concentrations of haptoglobin,
TSA and AST were observed after successful treatment for
endometritis in cows with clinical and subclinical endo-
metritis cows, respectively.
Interpretation
Dairy cows undergo substantial metabolic and physiologi-
cal adaptations during the transition from pregnancy to
lactation. Metabolic disorders play the most important role
in the pathology of the transition period. The results sug-
gest that higher acute phase protein concentration, lower
energy status (as indicated by increased BHB concentra-
tion), impaired liver function and decreased serum calcium
could be risk factors for the establishment and persistence
of endometritis.
Significance of findings
Endometritis is a significant cause of decreased reproduc-
tive performance in dairy cows and delayed breeding has a
marked economic effect. The above results should there-
fore be considered in the prevention and management of
this infection.
group.bmj.com on September 2, 2012 - Published by veterinaryrecord.bmj.comDownloaded from
doi: 10.1136/vr.100947
2012 2012 171: 219 originally published online August 4,Veterinary Record
M. Heidarpour, M. Mohri, A. H Fallah-Rad, et al.
subclinical endometritis
treatment in cows with clinical and
metabolic status affect the outcome of
Acute-phase protein concentration and
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September 1, 2012 | Veterinary Record
Paper
Paper
M. Heidarpour, M. Mohri, A. H Fallah-Rad, F. Dehghan Shahreza, M. Mohammadi
Acute-phase protein concentration and
metabolic status affect the outcome of
treatment in cows with clinical and subclinical
endometritis
The aim of this study was to investigate the role of acute-phase protein concentration
and metabolic status in the establishment and resistance of clinical endometritis (CE) and
subclinical endometritis (SE) in dairy cows. We also characterised the treatment-related
changes in the concentration of acute-phase proteins and metabolic variables in dairy
cows affected by CE and SE. Cows of the SE and CE groups presented a significantly higher
β-hydroxybutyrate (BHB), haptoglobin and total sialic acid (TSA) concentrations compared
with a healthy group of animals. A significantly lower serum calcium concentration, and
a significantly higher serum aspartate aminotransferase activity in the CE group, were
observed when compared with SE and healthy groups. The comparison of parameters before
treatment indicated that cows suffering from CE or SE with lower concentrations of hepatic
and inflammatory markers showed a better response to further treatment, and endometritis
was not detected in the second examination. Moreover, decreased concentrations of BHB,
acute-phase proteins and hepatic markers were observed after successful treatment for
endometritis in CE and SE cows. The results obtained in this study suggest that improved liver
function and a decrease in the acute-phase protein concentration might favour the resolution
of endometritis after treatment.
Introduction
Reproductive performance of dairy cows after the voluntary wait-
ing period is greatly related to the health status of the uterus after
calving. Bacterial contamination of the uterine lumen in the first
two weeks after parturition occurs in 80–90 per cent of dairy cattle
(Sheldon and others 2009). For several weeks after parturition, there
is a cycle of bacterial contamination, clearance and recontamination.
In many animals, this bacterial contamination is gradually resolved
by uterine involution, passage of lochia out of the uterus, and through
the mobilisation of immune defences. However, failure to resolve the
contamination can compromise uterine function, and the persistence
of pathogenic bacteria for at least three weeks postpartum, causes
endometritis in 10–20 per cent of postpartum dairy cattle (Sheldon
and others 2009). Endometritis is defined as inflammation of the
endometrium without systemic signs, and is associated with delayed
uterine involution (Sheldon and others 2006). Endometritis has been
subdivided into clinical and subclinical categories (Sheldon and others
2006). Clinical endometritis is defined as the presence of a purulent
uterine discharge detectable in the vagina 21 days or more, postpar-
tum, or mucopurulent discharge detectable in the vagina after 26 days
postpartum (Leblanc and others 2002). Subclinical endometritis (SE)
is defined as the presence of >18 per cent polymorphonuclear (PMN)
cells in uterine cytology samples collected 21–33 days postpartum, or
>10 per cent PMNs in samples collected at days 34–47 (Kasimanickam
and others 2004). Cows with SE do not have uterine discharge; how-
ever, the severity of the disease is still considered sufficient to impair
reproductive performance (Sheldon and others 2006).
Dairy cows undergo substantial metabolic and physiological adap-
tations during the transition from pregnancy to lactation. Essentially,
all dairy cows experience a period of insulin resistance, reduced food
intake, negative energy balance (NEB), hypocalcaemia and reduced
immune function soon before, or in the weeks after calving (Leblanc
2010). Coordinated shifts in nutrient partitioning must occur in
order to meet the increased demand for energy and other nutrients
necessary for foetal growth and lactation (Sordillo and Aitken 2009).
Metabolic disorders play the most important role in the pathology of
the transition period. The energy deficiency often causes the atony or
hypotony of the uterus, and this is the main cause for its late involu-
tion and cleaning (Kaczmarowski and others 2006). Metabolic-related
risk factors for uterine infections include subclinical ketosis (Hammon
and others 2006), hypocalcaemia (Mateus and De Costa 2002),
Veterinary Record (2012) doi: 10.1136/vr.100947
M. Heidarpour, DVM, DVSc,
M. Mohri, DVM, DVSc,
A. H Fallah-Rad, DVM, PhD
F. Dehghan Shahreza, DVM,
M. Mohammadi, DVM,
Department of Clinical Sciences, School
of Veterinary Medicine, Ferdowsi
University of Mashhad, Mashhad, Iran
M. Mohri
Center of Excellence in Ruminant
Abortion and Neonatal Mortality, School
of Veterinary Medicine, Ferdowsi
University of Mashhad,
Mashhad, Iran
E-mail for correspondence:
heidarpour@um.ac.ir
Provenance: not commissioned;
externally peer reviewed
Accepted June 20, 2012
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Paper
Veterinary Record | September 1, 2012
Paper
hypomagnesemia (Burke and others 2010) and hepatocellular dam-
age (ie, increased glutamate dehydrogenase and aspartate aminotrans-
ferase (AST)) (Burke and others 2010). It has also been reported that
the severity of uterine bacterial contamination, as determined by the
bacterial growth density, was correlated with the peripheral circulating
concentrations of acute-phase proteins (Sheldon and others 2001).
Although the relationships of metabolic parameters and acute-
phase proteins with the occurrence of endometritis have been investi-
gated, no study has compared these parameters between clinical and
SE, nor have the effects of these parameters on the outcome of treat-
ment been evaluated. Therefore, the objectives of this study were (1)
to evaluate the metabolic state and acute phase proteins concentra-
tions of cows with clinical and SE and to compare them with those
of healthy animals; and (2) to compare these parameters in cows that
recovered from clinical and SE after treatment with those suffering
from persistent endometritis.
Materials and methods
Cows
The study was conducted from April to September 2010 in a large
commercial dairy herd (Mashhad, northeast Iran). Cows (n=90)
were in their second parity, kept in an open shed and received a
well balanced total mixed ration (TMR) ration, and were milked
three times a day. Average daily milk production was 31–38 kg/cow.
Voluntary waiting period was 45 days, and average annual culling
rate was 26 per cent. All data were recorded in a computerised sys-
tem. First examination took place between 21 and 33 days in milk
(DIM). If the cows had any apparent sickness, they were excluded
from the experiment, and if not, the first cytological sample was
taken. Second examination and sampling were made seven days
after treatment. The cows having peripartum diseases, such as milk
fever, ketosis, displaced abomasums and retained fetal membranes
(RFM) were excluded from the experiment. Cows with a history of
systemic or intrauterine antibiotic therapy within seven days prior to
enrolment, reproductive hormone administration in the current lac-
tation prior to enrolment, or abnormal genitalia including adhesions,
laceration and pyometra, were also excluded from the study.
Clinical endometritis
The presence of clinical endometritis (CE) was determined by finding
pus in the lumen of the vagina by withdrawing the contents of the
vagina by hand. First, the perineum and vulva were cleansed with a
paper towel, and a clean, lubricated, gloved hand was inserted through
the vulva. The vagina was assessed for any signs of damage or injury
by palpation of the lateral, dorsal and ventral walls. Animals with
palpable vaginal injury were excluded from the study. The mucus con-
tent of the vagina was withdrawn manually for examination. The
vaginal mucus was characterised using an endometritis scoring system
(Williams and others 2005). Character score was assigned as follows:
unaffected animals, (0) clear or translucent mucus; (1) mucus contain-
ing flecks of white or off-white pus; (2) mucopurulent exudates con-
taining ≤50 per cent white or off-white mucopurulent material; and
(3) exudate containing >50 per cent purulent material usually white
or yellow, but occasionally sanguinous. CE was defined by the pres-
ence of >50 per cent purulent uterine discharge (score 3) detectable
in the vagina 21 days or more, postpartum, or mucuopurulent dis-
charge (score 2 or 3) detectable in the vagina after 26 days postpartum
(Sheldon and others 2006).
Subclinical endometritis
In animals without signs of CE, subclinical disease was diagnosed
by measuring the proportion of PMNs present in endometrial cytol-
ogy samples. Endometrial cytology samples were collected using the
modified cytobrush technique as described by Kasimanickam and
others (2004). Smears were prepared by rolling the brush on a micro-
scope slide, dried and fixed immediately after collection. Samples
were stained (Giemsa at a dilution of 5 per cent in buffer solution) and
evaluated by 400× magnification of a bright light microscope (Zeiss,
Germany) in the lab.
In total, 300 cells were counted under the microscope to deter-
mine the proportion of PMNs. The endometrial cytology slides were
assessed twice by a clinician. Cows with >18 per cent PMNs were
diagnosed as SE cows.
Treatments
At the conclusion of the first examination, the SE cows received
one of two treatments: 500 mg of cloprostenol intramuscular (im)
(Estroplan, Parnell, Australia) on days 1 and 14, or 500 mg of benza-
thine cephapirin in 19.6 g ointment base intrauterine (iu) (Metricure,
Intervet, Canada) and then 500 mg of cloprostenol im seven days after
benzathine cephapirin injection.
Treatment protocols for CE cows were as below: 500 mg of
cloprostenol im (Estroplan, Parnell, Australia) on days 1 and 14, or
Na-ceftiofur intramuscular (1 mg/kg, Excenell, Pfizer, Animal health
SA, USA, Kalamazoo, Michigan) for three consecutive days, and then
500 mg of cloprostenol, intramuscular, seven days after the last ceft-
iofur injection.
After treatment, and taking the second samples (on day seven after
treatment) by cytobrush from SE cows, two groups of animals were
selected: cows with successful treatment (ST group, PMN less than 18
per cent in endometrial samples), and animals with unsuccessful treat-
ment (UST group, PMN more than 18 per cent in endometrial sam-
ples). The same procedure was applied for CE cows, and two groups
were selected: ST (ST group, no abnormal discharge and PMN less
than 18 per cent in endometrial samples) and UST (UST group, with
abnormal discharge and/or PMN more than 18 per cent in endome-
trial samples).
Sampling
Blood was taken from the jugular vein of all cows with CE and SE at
the time of diagnosis (before treatment) and on day seven after treat-
ment. All tubes were immediately placed on ice and were transferred
to the laboratory. Serum was separated after spinning at 1800 g for
10 minutes and stored at −20°C until analysis. Blood samples were
also taken from the control group (n=30, healthy cows with no clinical
or SE based on clinical evaluation and endometrial cytology described
above) on the day of first examination.
Biochemical analysis
The activities of alkaline phosphatase (ALP), AST, gamma glutamyl
transferase (GGT), and the concentrations of bilirubin, albumin, cal-
cium, magnesium, phosphorous and β-hydroxybutyrate (BHB) in
serum samples were measured with commercial kits (Pars Azmoon,
Iran) for ALP, AST, GGT, bilirubin, albumin, calcium, magnesium
and phosphorous (Randox, Antrim, UK for BHB) using an autoana-
lyser (Biotecnica, Targa 3000, Rome, Italy). The intra- and inter-assay
coefficients of variation were, respectively, 3.25 per cent and 3.75
per cent for ALP, 2.48 per cent and 1.49 per cent for GGT, 2.51 per
cent and 2.48 per cent for AST, 2.27 per cent and 2.54 per cent for
bilirubin, 0.91 per cent and 1.12 per cent for albumin, 1.45 per cent
and 1.53 per cent for calcium, 0.92 per cent and 1.45 per cent for
magnesium, 1.61 per cent and 2.22 per cent for phosphorous, and
5.25 per cent and 3.78 per cent for BHB. Control serum (Randox
control sera, Antrim, UK) was used for controlling measurement
accuracy.
Serum total sialic acid (TSA) concentration was determined by a
method previously described by Sydow (1985). Briefly, a mixture of
0.2 ml serum and 1.5 ml 5 per cent perchloric acid (HClO4) was incu-
bated for 5 minutes at 100°C, cooled, and then centrifuged at 2500×g
for 4 minutes. Next, 0.2 ml of Ehrlich’s reagent was added to 1.0 ml
of the clear supernatant, and heated for 15 minutes at 100°C. After
the mixture was cooled, 1.0 ml distilled water was added to it. The
optical density was measured at 525 nm. The amount of TSA was
determined by use of a standard curve developed from a standard sam-
ple of n-acetyl neuraminic acid.
Serum haptoglobin concentration was determined by Bovine
Haptoglobin ELISA kit (Bio-X Diagnostics, Jemelle, Belgium) accord-
ing to the manufacturer’s instruction.
Statistical procedures
Values are reported in the text and tables as means±sd. Analyses of
variance with multiple comparison test (Tukey Honestly) were used
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September 1, 2012 | Veterinary Record
to compare means among the trial groups (healthy control, CE and SE
cows). We also used Student’s t test to compare means before and after
treatment in cows with CE and SE. All analyses were performed with
the statistics package SPSS (release 16, SPSS Inc, Chicago, Illinois,
USA). Statistical significance was taken to be indicated by P<0.05.
Results
The overall incidence of CE and SE in the cows was 11.83 per cent
(71/600) and 13.5 per cent (81/600), respectively. The incidence of
treatment success within CE and SE groups was 48.3 per cent and
58 per cent, respectively. Of the examined cows, 30 fit the selection
criteria for CE (15 cows in ST group and 15 cows in UST group), 30
were chosen for SE (15 cows in ST group and 15 cows in UST group),
and 30 were selected as healthy cows.
Metabolic variables and acute-phase proteins in cows
with CE and SE
Table 1 shows mean±sd of the values of metabolic variables and acute-
phase proteins in healthy, CE and SE cows. Cows of the SE group
presented a significantly higher BHB (P<0.01), haptoglobin (P<0.01)
and TSA (P<0.05) concentrations compared with the healthy group,
while the CE group presented a significantly higher (P<0.01) BHB,
haptoglobin and TSA concentrations compared with the healthy and
SE groups. The CE cows had a lower (P<0.01) sera calcium and greater
(P<0.05) sera AST compared with other groups.
Metabolic variables and acute-phase proteins’
concentration before and after treatment in cows
with CE
The comparison of measured parameters at the time of diagnosis
(before treatment) indicated that cows suffering from CE in the ST
group had lower AST (P<0.01, Table 2) and haptoglobin concentra-
tions (P<0.05, Table 2) compared with the UST group.
After ST for CE, mean serum BHB, haptoglobin and TSA con-
centrations dropped significantly (P<0.05, Table 2). If treatment was
not successful, no significant difference was seen between parameters
measured before and after treatment (Table 2).
Metabolic variables and acute-phase proteins before
and after treatment in cows with SE
The comparison of measured parameters at the time of diagnosis
(before treatment) indicated that cows suffering from SE in the ST
group had lower ALP (P<0.05, Table 3) and haptoglobin concentra-
tions (P<0.01, Table 3) compared with the UST group.
Mean serum haptoglobin (P<0.001) and TSA (P<0.05) concentra-
tions and AST (P<0.05) activity decreased after ST for SE (Table 3).
However, this was not the case in cows with UST, and no significant
difference was seen between parameters measured before and after
treatment (Table 3).
Discussion
The results of the current study suggest the existence of an association
between endometritis and increased acute-phase protein and BHB
concentration. The results also indicate that CE was associated with
impaired liver function, coupled with low serum calcium concentra-
tion. The working hypothesis, that acute-phase protein concentration
and metabolic status affect the outcome of treatment in dairy cows
with endometritis, was supported by the results of this study.
A part of the innate immune response to bacterial infections is the
elaboration of pro-inflammatory cytokines, which are potent stimu-
lants of acute-phase proteins’ synthesis (Williams and others 2005).
Haptoglobin is a major acute-phase protein in numerous species of
animals. In ruminants, its circulating level is negligible in healthy ani-
mals, but increases following immune stimulation (Murata and others
2004). Sialic acid is localised at the end chain of many acute-phase
proteins. Therefore, the detection of sialic acid may be a valuable indi-
cator for diagnosis and prognosis of inflammatory diseases (Motoi
and others 1984). Several previous reports link increased acute-phase
proteins concentration with the presence of uterine disease, and hap-
toglobin has been suggested as an early predictor of clinical metritis
and endometritis (Drillich and others 2007, Huzzey and others 2009,
Burke and others 2010). Acute-phase proteins have very high sensitiv-
ity in detecting many conditions that alter the health of the animal,
and in providing evidence that an animal has subclinical inflamma-
tion or infection. As summarised by [b10]Kent (1992), acute-phase
proteins quickly and precisely demonstrate the presence of infectious
TABLE 1: Mean±sd of metabolic variables and acute-phase
proteins in cows with CE and SE at the time of diagnosis
(21–33 days postpartum)
Healthy (n=30) SE (n=30) CE (n=30)
ALP (IU/l) 29.27±9.93 29.74±10.28 32.18±9.08
AST (IU/l) 66.33±21.64a70.6±22.3a85.17±17.72b
GGT (IU/l) 27.77±8.93 25.2±11.21 30.6±10.66
Bilirubin (mg/dl) 0.48±0.14 0.45±0.19 0.55±0.22
Albumin (mg/dl) 3.52±0. 33 3.61±0. 37 3.53±0.26
Calcium (mg/dl) 11.16±1.74a10.78±1.67a9. 21±0.81b
Phosphorous (mg/dl) 6.63±0.68 6.51±0.75 6.39±1.04
Magnesium (mg/dl) 3.16±0.44 3.36±0.62 3.27±0.60
BHB (mg/dl) 0.39±0.08a0.71±0.24b1.11±0.63c
Haptoglobin (μg/ml) 11.08±14.25a54.53±23.21b84.68±66.96c
TSA (mg/dl) 57.80±38.35a99.55±57. 4 7 b178.84±73.48c
In each row, means with different superscripts a, b, c, show significant
differences (P<0.05).
ALP, alkaline phosphatase; AST, aspartate aminotransferase; BHB,
β-hydroxybutyrate; CE, clinical endometritis; GGT, gamma glutamyl transferase;
SE, subclinical endometritis; TSA, total sialic acid.
TABLE 2: Comparison of the level of metabolic variables and
acute-phase proteins in CE cows, at the time of diagnosis (before
treatment, 21–33 days postpartum), and after treatment
Parameters Before treatment After treatment
ALP (IU/l)
S T 30.61±9.15 37.69±14.41
UST 33.75±9.0 5 35.15±10.70
AST (IU/l)
S T 75.00±10.75A70.53±23.21A
UST 95.33±17.71B90.86±21.05B
GGT (IU/l)
S T 30.40±11.63 34.73±13.53
UST 30.80±9.99 28.06±15.58
Bilirubin (mg/dl)
S T 0.52±0.21 0.62±0. 21
UST 0.58±0.23 0.69±0.27
Albumin (mg/dl)
S T 3.53±0.30 3.48±0.27
UST 3.54±0.23 3.64±0. 32
Calcium (mg/dl)
S T 9.13±0.62 8.98±0.83
UST 9.28±0.98 9.14±0.72
Phosphorous (mg/dl)
S T 5.98±0.77 6.56±0. 74
UST 6.79±1.12 6.62±0.92
Magnesium (mg/dl)
S T 3.26±0.47 3.41±0.45
UST 3.26±0.71 2.88±0.47
BHB (mg/dl)
S T 1. 26±0.73a0.80±0.29b
UST 0.96±0.50 0.87±0.15
Haptoglobin (μg/ml)
S T 55.18±64.84Aa 14.82±11.83Ab
UST 114.23±56.69B72.60±42.82B
TSA (mg/dl)
S T 160.35±82.63a8 7.75 ±4 2.08Ab
UST 190.98±74.96 177.14±67.16B
ALP, alkaline phosphatase; AST, aspartate aminotransferase; BHB,
β-hydroxybutyrate; CE, clinical endometritis; GGT, gamma glutamyl transferase;
ST, cows with successful treatment (n=15); TSA, total sialic acid; UST, cows with
unsuccessful treatment (n=15).
In each row, means with different lowercase superscripts a, b, show significant
differences (P<0.05).
In each column, means with different uppercase superscripts A, B, show
significant differences (P<0.05).
No significant difference was observed within UST group before and after
treatment.
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and inflammatory conditions. The authors believe that this is the first
study that has measured TSA concentration in cows with endometri-
tis. In the present study, serum haptoglobin and TSA were significant-
ly higher in CE and SE groups compared with the healthy group. It
has been reported that the severity of uterine bacterial contamination,
as determined by the bacterial growth density, was correlated with the
peripheral circulating concentrations of acute-phase proteins (Sheldon
and others 2001). Therefore, higher acute-phase proteins in CE cows
compared with SE cows may be related to more severe uterine bacte-
rial contamination and subsequently more inflammatory response.
A correlation has been described between acute-phase protein con-
centration and the severity of disease, with serum acute-phase protein
concentrations being higher in severe cases, or in animals with clinical
signs of disease compared with those without clinical signs (Ceron
and others 2005). Acute-phase proteins are also useful for prognostic
purposes. In the present study, SE and CE cows with low haptoglobin
concentration showed a better response to further treatment, and
endometritis was not detected in the second examination. In addition,
after ST, mean serum haptoglobin and TSA concentrations dropped
significantly. The postpartum uterine involution is too important for
uterine cleaning from bacteria that gained access to the uterus (Arthur
and others 1996). Uterine involution is associated with a decrease in
the concentrations of acute-phase proteins (Sheldon and others 2001).
Thus, the efficacy of treatment in cows with low haptoglobin concen-
trations might be a consequence of better uterine involution in cows
with a mild inflammatory response.
The peripartum period is of critical importance to subsequent
health and fertility. Most cows enter a state of NEB associated with
many metabolic changes. Non-esterified fatty acids and BHB concen-
trations elevate, indicative of lipid mobilisation and fatty acid oxida-
tion. NEB reduces the ability of the uterus to recover after calving,
and may result in persistent inflammatory-mediated damage (Wathes
and others 2007). In the current study, serum BHB concentration,
as indicator of energy status, was significantly higher in CE and SE
groups compared with the healthy group, and after ST in CE cows, its
concentration decreased. The results of a study performed by Wathes
and others (2009) showed an ongoing inflammatory response in the
uteri of cows with severe NEB (evidenced by decreased plasma IGF-I
together with increased non-esterified fatty acids (NEFA) and BHB
concentrations). NEB may, therefore, prevent cows from mounting
an effective immune response to the microbial challenge experienced
after calving (Wathes and others 2009). In addition, subclinical ketosis
(serum BHB>12.5–15 mg/dl) is associated with increased risk of SE
(Hammon and others 2006).
Elevated serum activity of AST in CE cows indicated hepatic tis-
sue damage. Serum AST activity was also higher in the CE group
compared with SE cows. This result showed that the challenge to
liver function among cows diagnosed with CE was greater. Similarly,
Burke and others (2010) reported higher serum AST concentration in
cows with high PMN in cytological samples. In early postpartum,
the liver coordinates the extensive biochemical and morphological
modifications required via upregulation of genes involved in fatty
acid oxidation and gluconeogenesis, and downregulation of triacylg-
lycerol synthesis (Loor and others 2005). However, postpartum dis-
eases may have a detrimental effect on liver function. A previous
report implied that liver function was compromised because of sys-
temic toxins from postpartum disease (Sweeney and others 1988).
Moreover, postpartum accumulation of excessive fat impairs liver
function, compromising glucose production and increasing inflam-
matory responses (Wathes and others 2007). More efficacy of treat-
ment in endometric cows with mild hepatic tissue damage (lower
concentration of AST in CE cows with ST, and lower concentration
of ALP in SE cows with ST), indicate the role of the liver in coor-
dinating the extensive modifications which the cow must make in
order to recover from endometritis.
Myometrial contraction plays an important role in uterine involu-
tion (Arthur and others 1996). This contraction of the smooth mus-
cular layer of the uterus is mediated by several factors, including oes-
trogens, prostaglandins, oxytocin and minerals, like calcium (Arthur
and others 1996). Decreased blood calcium concentration was related
to poor uterine involution (Jonsson 1999). In the current study, serum
calcium concentration was significantly lower in CE groups com-
pared with the SE and healthy groups. These findings show a rela-
tionship between blood calcium concentrations and the establishment
and severity of endometritis. Similarly, Mateus and De Costa (2002)
reported that plasma calcium concentrations were significantly lower
in cows with severe endometritis than in cows with mild endometri-
tis, and in normal puerperium cows.
Conclusion
The results of the present study suggest that increased acute-phase
protein concentration, lower energy status (as indicated by increased
BHB concentration), impaired liver function, and decreased serum
calcium could be risk factors for the establishment and persistence
of endometritis. Moreover, improved liver function and a decrease in
the acute-phase protein concentration might favour the resolution of
endometritis after treatment.
Acknowledgements
This study was supported by a research fund from the Ferdowsi
University of Mashhad (project No 16748/2).
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doi: 10.1136/vr.100947
2012 2012 171: 219 originally published online August 4,Veterinary Record
M. Heidarpour, M. Mohri, A. H Fallah-Rad, et al.
subclinical endometritis
treatment in cows with clinical and
metabolic status affect the outcome of
Acute-phase protein concentration and
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