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CARDIOVASCULAR JOURNAL OF AFRICA • Volume 31, No 2, March/April 2020
86
AFRICA
activity in liver tissue homogenates was significantly increased
in the veh control, Mel, NMel and NRF groups compared to
the nicotine-treated group. CAT activity was also increased in
the veh control group compared to the water control (Table 4).
Lipid peroxidation
TBARS levels in serum of the nicotine-treated group were
significantly increased when compared to the RF, veh control,
water control, RUF, Mel and NMel treatment groups. TBARS
levels were also significantly increased in the NRF- and NRUF-
treated groups compared to the RF treatment group (Table 5).
Supplementary
in vitro
investigations
Based on the effects of RF on nicotine-induced vascular
changes in the
in vivo
investigations, RF was selected for
performing additional
in vitro
investigations. According to
separate dose–response experiments for the NO production and
necrosis investigations (data not shown), nicotine was used at a
concentration of 100
μ
M and RF at a concentration of 0.015
mg/ml. Nicotine at a concentration of 100
μ
M over a treatment
period of 24 hours resulted in significant reduction in NO
production, as indicated by DAF-2/DA fluorescence (Fig. 4),
and an increase in necrosis, as indicated by PI fluorescence (Fig.
5), when compared to controls.
AECs were pre-treated for one hour with 0.015 mg/ml RF,
followed by the addition of 100
μ
M nicotine for a further 24
hours. Pre-treatment with 0.015 mg/ml RF was associated
with a modest but significant increase in NO production in
nicotine-injured cells compared to cells treated with nicotine
Table 4. Effects of melatonin and rooibos (fermented and unfermented)
treatment on SOD activity (U/mg protein) and CAT activity
(
μ
mole/min/
μ
g) in liver tissue homogenates of all treatment groups
Treatment group SOD activity (U/mg protein) CAT activity (
μ
mole/min/
μ
g)
Veh control
155.3
±
6.7
*
868.3
±
138.4
*#
Water control
135.5
±
5.0
427.2
±
51.4
Nicotine
121.4
±
14.7
295.8
±
76.7
Mel
133.4
±
10.4
597.7
±
98.2
*
RF
192
±
21.1
*#$
597.5
±
111.7
RUF
182.7
±
20.4
#
515.6
±
111.9
NMel
180.8
±
9.0
*
565.8
±
87
*
NRF
160.7
±
6.4
*
727.3
±
158.6
*
NRUF
164.7
±
16.6
191.9
±
27.6
Values are mean
±
SEM of 9–10 rats per group.
*
Significantly different compared to nicotine treatment group (
p
<
0.05);
#
significantly different compared to water control group (
p
<
0.05);
$
significantly different compared to Mel treatment group (
p
<
0.05).
Table 5. Effects of melatonin and rooibos treatment on lipid
peroxidation in serum of all treatment groups
Treatment groups
TBARS (
μ
mol MDA equivalents/l)
Veh control
2.907
±
0.2
Water control
2.997
±
0.3
Nicotine
4.615
±
0.3
#$&@!*
Mel
2.829
±
0.9
RF
2.472
±
0.3
RUF
3.350
±
0.3
NMel
3.411
±
0.3
NRF
3.772
±
0.3
#
NRUF
3.707
±
0.1
#
Values are mean
±
SEM of 10 rats per group.
#
Significantly different compared to RF (
p
<
0.05);
$
significantly different
compared to veh control (
p
<
0.05);
&
significantly different compared to water
control (
p
<
0.05);
@
significantly different compared to RUF (
p
<
0.05);
!
signifi-
cantly different compared to Mel (
p
<
0.05);
*
significantly different compared to
NMel (
p
<
0.05).
Phe (
μ
M)
0.0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1.0 1.1
% Cumulative contraction
150
125
100
75
50
25
0
NRF
Nicotine
NRUF
NMel
*
#
$
ACh (M)
1.0
×
10
-8
3.2
×
10
-8
1.0
×
10
-7
3.2
×
10
-7
1.0
×
10
-6
3.2
×
10
-6
1.0
×
10
-5
3.2
×
10
-5
% Cumulative relaxation
0
20
40
60
80
100
120
*
NRF
Nicotine
NRUF
NMel
Fig. 3.
(A) Contractile responses of aortic rings harvested from nicotine, NMel, NRF and NRUF-treated rats following cumulative
Phe administration (*
p
<
0.05 NMel vs NRF and NRUF;
#
p
<
0.05 NMel vs nicotine;
$
p
<
0.05 NRF, NRUF vs nicotine). (B)
Relaxation response of aortic rings harvested from nicotine, NMel, NRF and NRUF-treated rats following cumulative ACh
administration (*
p
<
0.05 NMel and NRF vs nicotine and NRUF).
A
B