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African Journal of Biomedical Research
Ibadan Biomedical Communications Group
ISSN: 1119-5096
Vol. 5, Num. 3, 2002, pp. 131-135
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African Journal of Biomedical Research, Vol. 5, No. 3, Sept, 2002, pp.
131-135
HAEMATOLOGICAL CHANGES ACCOMPANYING
PROLONGED OCULAR
CHLORAMPHENICOL ADMINISTRATION IN LABORATORY RABBITS.
SABA A.B1*., AWE,
O.E2. AKINLOYE, A.K1., OLADELE
G.M1
1 Faculty
of
Veterinary Medicine, University of Ibadan Ibadan.; 2 Department of
Pharmacology /Therapeutics, College of Health Sciences, Ladoke Akintola University,
Ogbomosho
* Author for
correspondence
Accepted in final form:
March 2002
Code Number: md02027
The
toxic effect of ocular chloramphenicol on haematological parameters was
studied in laboratory rabbits; Oryctolagus cuniculus while the haemotoxic
effect
of oral chloramphenicol
provided the basis for comparison. 20 adult male rabbits were randomly but equally
divided into two main groups based on the route of administration of the drug
(i.e ocular or oral). In each group of ten rabbits equal number of rabbits were
randomly divided into test (n=5) and control (n=5) subgroups. Oral chloramphenicol
was administered at a dosage of 500mg twice daily for 21 days. Drops of ocular
chloramphenicol were administered on the conjunctiva of the animals thrice daily
over the same period of time. The control animals were administered with
0.9% physiological saline orally and distilled water administered ocularly.
Ocular
chloramphenicol produced no significant changes in the haematological parameters
evaluated on the 11th and 22nd days. Conversely oral
chloramphenicol was observed to significantly (P<0.05) reduced the mean
total erythrocyte count, PCV, mean corpuscular haemoglobin, and neutrophils
progressively by the
11th and 22nd days. Ocular chloramphenicol was confirmed
to have no dose-dependent haemotoxic effect however the possibility of idiosyncratic
aplastic anaemia is highlighted
in this study.
Key words: Ocular chloramphenicol,
Haemotoxicity, Rabbits.
INTRODUCTION
The clinical use of systemic chloramphenicol has been plagued
by the established
cases of haemotoxic effect of the drug. It has been reported that of the entire
drug that may be responsible for pancytopaenia, chloramphenicol is the most common
cause (Wallerstain: Kasper; Brown and Morrison, 1969) Investigations showed that
chloramphenicol induced toxicity is traceable to
bone marrow depression (Robbana Barnat, 1997., Holt, 1998). The clinical pictures
include reticulocytopaenia (within 5 to 7 days of initiation of
therapy). This is followed by decrease in hemoglobin, an increase in plasma
iron, cytoplasm vacoulation of early erythroid forms and granulocytes forms. The
dose-related bone marrow depression by chloramphenicol has been reported to progress
to fatal aplasia (Daum, 1979). As a result of this toxic effect chloramphenicol
is used with a lot of caution. It is only employed in treatment of certain diseases
where the risk benefit of the drug out weighs the risk of the potential toxicities
especially in case of typhoid fever.
The aforementioned effects of systemic chloramphenicol
is widely accepted, however the degree of safety of the ocular use of this
same drug is enmeshed in
controversy. While some workers believe that ocular chloramphenicol could be
as toxic as systemic chloramphenicol, others believe the contrary. Lazarov and
Amichai (1996) reported adverse skin reactions due to eye drops of chloramphenicol.
Laporte et al (1998) also confirmed an association between ocular chloramphenicol
and aplastic anaemia. At present, topical ocular chloramphenicol is widely used
in the U.K for the treatment of conjunctivitis, whereas it is very rarely prescribed
for this indication in the U.S. Reyner and Buckley (1996) reported that the possible
haemotoxic effect of chloramphenicol led to the prohibition of the use this drug
in the U.S.
We present in this study
an attempt to investigate the effect of ophthalmic chloramphenicol on haematological
parameters using animal
model- (laboratory rabbits). The effect of oral chloramphenicol on the haematological
parameters is intended to provide a basis for comparison between ocular and systemic
chloramphenicol.
MATERIALS AND METHODS
Experimental
Animals: 20 adult male rabbits kept on commercially prepared rabbits
growers mash (Guinea feeds, Nigeria) ad lib and allowed to fresh water
without restriction. The twenty rabbits were randomly but equally divided into
two groups based on the route of administration of the drug; ocular or oral (i.e
Groups I and II respectively). In each group (I or II) 10 rabbits were randomly,
but equally selected into test and control subgroups. In other words, ocularly
administered rabbits consist of the test I animals (n=5), control I animals (n=5)
and orally administered rabbits consist of the test II animals (n=5) and control
II animals (n=5). The animals were allowed to stabilize for four weeks during
which they were dewormed with levamisole
hydrochloride (LevajectR, Pantex. Holland).
Administration of Drug: In
group I, the conjunctiva of the five rabbits in the test subgroup were on each
occasion exposed and generally flooded with ocular chloramphenicol (0.5% chloramphenicol
eye drop U.S.P Mubai, India) three times daily for a period of 21 days while
drops of distilled water were similarly instilled on the eye of other five
rabbits in the control
subgroup thrice daily for the same period of time.
In group II, oral dose of
500mg of chloramphenicol
(chloramphenicol palmitate oral suspension, vardhman, Bombay, India) was administered
twice daily to each rabbit in the test subgroup for a period of 21 days while
the rabbits in the control subgroup were administered orally with 0.9% physiological
saline for the same period of time.
Collection of Blood Samples: Blood
samples were collected from all rabbits from both groups (I and II) on day
zero, 11th day and 22nd day; a day after the last administration
of the drug. This was achieved by
first anaesthetizing the animals with ether. Capillary tubes were introduced
into the orbital sinus systematically in a way that allowed blood to collect
in
the lithium-heparinised tubes.
Haematology: Haemoglobin
concentration (HBC) was determined by cyanomethaemoglobin method, packed cell
volume (PCV) by capillary
tube method, total erythrocyte
count (TEC) by the
haemocytometer method, total leucocyte count (TLC) and its differentials by
Giemsa stains slides methods (Jain, 1986). The mean corpuscular volume (MCV)
and mean corpuscular haemoglobin concentration (MCHC) and mean corpuscular
haemoglobin (MCH) were calculated from the data obtained.
Statistical Analysis: Results
are expressed as the mean ± standard error of mean. Differences between means
of values of test and control animals were statistically determined by the
students t-test. Differences are considered significant at p<0.05 probability
level (Bailey, 1992).
RESULTS
The effect of ocular chloramphenicol on erythrocyte
indices: The mean value of PCV and mean corpuscular hemoglobin
of rabbits in the test group increased through the period of administration
of ocular chloramphenicol. However the differences are not significant
(Table 1) when compared with the control values. Conversely mean value
of MCV and HBC of the same rabbits decreased insignificantly (P >0.05)
(Table 1) through the same period of time. By the 11th day
of ocular administration of the drug, the mean total circulating erythrocyte
count
had decreased while MCHC increased. These various changes are not statistically
significant (P>0.05) when compared with the control values at
any of the days.
The effect of ocular chloramphenicol
on total leukocyte
count and its differentials: There was slight increase in mean of basophils,
eosinophils and lymphocytes covering the period of drug administration in the
test group. The differences are not significant (P>0.05) when compared with
the control
values (Table 2). By the 11th day of administration of ocular chloramphenicol,
neutrophils was found to have decreased while the monocytes and total leucocytes
count increased but the reverse was the case by the 22nd days. However,
these changes are not statistically significant (P>0.05) when compared with
the control values (Table 2).
The Effect of oral chloramphenicol
on erythrocyte
indices: The
mean value of circulating erythrocytes, PCV and mean corpuscular haemoglobin
decline consistently through the period of oral administration of
chloramphenicol. The difference are statistically significant in TEC, PCV and
MCH (P<0.05) when compared with the values obtained for control group (Table
1). MCHC and MCV were found to have decreased by the 11th day and
by
the 22nd day: these values have increased. The changes were not statistically
significant (P>0.05) over the values obtained for the control
group (Table1). HBC increased by the 11th day and was observed to
have decreased by the 22nd day but these changes are not statistically
significant (P>0.05) Table (1).
The effect of oral chloramphenicol
on leucocyte count
and its differential: The total leucocyte count, neutrophils, eosinophils,
monocytes and basophils decreased consistently through the period of oral administration
of
the drug. The changes in the mean values when compared with control values are
only significant (P<0.05) for neutrophils. (Table 2).
Table
1. The mean (± S.E.M) of total erythrocyte counts and other haematological
indices of rabbits before, during and after administration with ocular or oral
chloramphenicol.
Parameter
|
Day
|
Ocular chloramphenicol (n=5)
|
Oral Chloramphenicol (n=5)
|
Test group I
|
Control group I
|
Test group II
|
Control group II
|
RBC 106/ul)
PCV (%)
MCV (fl)
MCH(Pg)
MCHC (g/dl)
HBC(g/dl)
|
0
11
22
0
11
22
0
11
22
0
11
22
0
11
22
0
11
22
|
5.76 ±0.11
4.90±±0.12
5.27±0.10
25.25±1.00
25.53±0.29
26.00±0.18
54.22±3.41
52.28±0.93
49.60±0.79
7.93±0.36
8.03±0.29
8.05±0.14
31.27±0.45
31.44±0.27
31.06±0.49
20.82±0.39
20.77±0.56
20.69±0.47
|
5.40±0.12
5.13±0.16
521±0.12
25.75±0.36
26.50±0.43
24.50±0.74
46.82±1.12
52.18±1.18
47.03±0.90
8.10±0.29
7.83±0.32
7.45±0.23
30.90±0.44
31.27±0.79
30.47±0.54
21.00±0.33
20.98±0.60
20.70±0.49
|
4.92±0.11
*4.50±0.16
*4.45±0.05
26.11±0.08
*25.19±0.2
*24.70±0.1
53.24±0.56
51.72±0.64
53.17±0.85
7.57±0.14
7.45±0.12
*7.41±0.14
31.27±0.67
31.06±0.64
31.24±0.70
20.72±0.33
21.11±0.29
20.60±0.36
|
5.01±0.09
4.98±0.11
5.04±0.03
26.23±0.09
26.21±0.15
26.45±0.14
51.73±0.89
49.66±0.92
50.95±0.77
7.63±0.11
7.68±0.11
7.90±0.16
31.12±0.64
31.05±0.72
31.15±0.74
20.99±0.35
20.50±0.29
21.00±0.30
|
n=Number of animals in the group; *=Implies significant
differences (P<0.001) exists between means values of test and control
animals; **= Implies significant
differences (P<0.05) exists between mean values of test and control animals.
Table
2: The mean
(± S.EM) of total white blood cell count and the different cells of rabbits before,
during and after
administration with ocular or oral chloramphenicol
Parameter
|
DAY
|
Ocular chloramphenicol
|
Oral Chloramphenicol
|
Test group I
n=5
|
Control group I
n=5
|
Test group II
n=5
|
Control groupII
n=5
|
Total WBC count 103/ul)
Neutrophils (%)
Lymphocytes (fl)
Monocytes (Pg)
Eosinophils (%)
Basophils (%)
|
0
11
22
0
11
22
0
11
22
0
11
22
0
11
22
0
11
22
|
6.64±0.61
6.82±0.80
6.74±0.50
45.04±1.7
43.07±0.5
45.34±1.1
40.36±0.6
41.04±0.7
42.03±0.7
9.34±0.16
9.72±0.28
9.65±0.33
1.51±0.16
1.53±0.20
1.62±0.10
5.34±0.32
5.65±0.22
5.87±0.56
|
6.73±0.53
6.92±0.51
7.13±0.45
42.75±1.1
44.26± .8
43.88±0.8
41.12±0.6
39.75±0.9
42.91±0.7
9.57 ± 0.2
9.26 ± 0.5
9.10 ± 0.6
1.27± 0.24
1.93± 0.28
1.50± 0.21
5.07± 0.21
5.33± 0.28
5.25± 0.63
|
7.88± 0.32
6.24± 0.57
5.70± 0.40
44.10±0.8
**40.22±0.5
**40.13±0.5
41.42±0.36
43.25±0.76
41.76±0.54
9.16 ± 0.23
8.79 ± 0.34
8.46 ± 0.38
1.82±0.19
1.67±0.21
1.52±0.11
5.27±0.14
5.25±0.33
5.01±0.28
|
6.99±0.31
7.16±0.33
5.04±0.03
42.21±0.8
42.73±0.6
42.50±0.7
41.93±0.3
41.62±0.8
42.54±0.7
8.95±0.19
9.23±0.26
9.01±0.32
1.53±0.23
1.55±0.23
1.64±0.18
5.14±0.24
5.36±0.18
5.13±0.27
|
n = Number of rabbits
in the group.
** = Implies significant
difference (P< 0.05 )
exists between means of test and control animal
The mean circulating lymphocytes
increased by the 11th day of drug administration and was found to
have decreased by the 22nd day. Statistical comparism of these
values with the control values shows that the differences are not significant
(P>0.05). (Table 2).
DISCUSSION
There was no specific pattern
of effect of ocular chloramphenicol on the erythrocyte indices, leucocyte count
and its differential cells. Much of the changes observed were not consistent
or statistically significant thus creating an impression that these changes
are merely due to chances. It will be right to affirm based on this study that
ocular chloraphenicol may not have haemotoxic effects. This assertion is similar
to the opinion of Field et al (1999) that ophthalmic chloramphenicol
is demonstrably effective, safe, cost-effective treatment for most superficial
eye infection. Walker (1998) had earlier reported that ocularly administered
chloramphenicol failed to accumulate to detectable levels in the blood to warrant
the drug to
be a risk factor for inducing dose-related bone marrow toxicity.
On the other hand, systemic
chloramphenicol had long been confirmed to exhibit haemotoxic effects. This
was clearly observed in this study; by 11th day of administering Chloramphenical
orally; the circulating RBC, WBC, neutrophilis, PCV and mean corpuscular haemoglobin
levels reduced significantly and these reductions were progressively so observed
by the 22nd day. The effect of systemic Chloramphenicol on the blood has been
attributed to inhibition of mitochondrial protein synthesis in the myeloid
tissue (Wakabayashi, 1999) or due to membrane stabilising effect of cells by
Chloraphenicol (Wuc, 1996).Holt, (1998) further observed that these toxic effects
are exerted at the differentiation stage of the committed marrow progenitor
cells rather than at the replicative sage of the stem cells. Haemotoxicty of
systemic Chloramphenicol has also been reported to be presented in two forms,
(I) dose-dependent bone marrow depression and (ii) fatal idiosyncratic aplastic
anaemia (Holt et al. 1997). Even though ocular chloramphenicol may yield
low detectable levels in the blood (Walker, 1998) which may not be significant
to precipitate dose-dependent bone marrow depression, the incidence of aplastic
anemia is a simple possibility. This is so when considering the fact that aplastic
anaemia is a factor of hypersensivity or idiosyncrasy rather than dose-dependence.
In this same line of argument, McGhee (1996) proved that the risk of chloramphenicol-induced
idiosyncratic aplastic anaemia exists with topical ophthalmic therapy with
the minimum risk of death (equalling that of systemic penicillin therapy being
1 in 50,000 to 90,000. This figure he stressed is comparable to the risk of
fatal anaphylaxis resulting from any
route (i.e 1 in 100,000). Laporte et al (1998) also gave credence to this
fact when he confirmed a probable association between ocular chloramphenicol
and fatal idiosyncratic aplastic anaemia. Field et al (1999) hinted that
even in UK where ocular chloramphenicol is freely prescribed, the use of the
drug is being reviewed starting from 1995.
In conclusion, it is very
pertinent to establish that in the clinical use of ocular chloramphenicol the
benefit of its proved safety, tolerance, cost and efficacy must be weighed
viz a viz the remote risk of serious adverse effect of drug induced aplastic
anaemia. This risk-benefit assessment is therefore the duty of every prescribing
physician.
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