Zeng Dingyi, Yubo, Zheng Xiaowei, Qi Guoxian, Wang
(Department of Cardiology, The First Affiliated Hospital of China
Medical University, Shenyang 110001)
(Examination Department of The First Affiliated Hospital of China
Song Ji'ai, Shi Yuxiu
(Fundamental Medical Institute, The First Affiliated Hospital of
China Medical University)
Xuezhikang, with HMG-CoA reductase inhibitor as the main ingredient,
contains many kinds of necessary amino acids and unsaturated fatty
acids. This study aims at investigating the effects of Xuezhikang
on the proliferation and migration of vessel smooth muscle cells
healthy and pure New Zealand white rabbits were chosen with body
weight 2.57 ± 0.17 kg. They were divided into three groups
in random. The first group was the control group taking normal feed.
The second group acted as hypercholesterol group fed with normal
feed plus 1.5% cholesterol. And the third group was the treatment
(Xuezhikang) group taking normal feed plus 1.5% cholesterol and
0.8 g Xuezhikang per kilogram weight per day. Every rabbit was raised
in a single cage with daily feed of 150 g and the treatment lasted
for 12 weeks.
the morning, blood samples were taken from ear mid-artery on fast
animals before, 4 weeks, 8 weeks and 12 weeks after the study to
determine the concentrations of serum TC, TG and HDL-C. Whereas
LDL-C concentration was computed with the help of Friedewald Formula.
At the end of 12 weeks, all rabbits were killed by bloodletting
and aortic arch was taken as samples maintained 2.5% glutaraldehyde
ready for the preparation of transmission electron microscopic samples.
In Vitro Study
Rabbit chest artery SMC was taken and cultivated by line-plot sticking
method. Under 5% CO2 concentration, 15% calf serum RPMI 1640 culture
solution was used and set still for 3-week cultivation. Then 0.1%
EDTA and 0.25% pancreatic enzymes digesting solution were used to
digest new SMC generations. And only well developed 3 ~ 6 generations
of SMC were used in the study. After digestion, SMCs were made into
2´105/ml suspended cell solution. Then, they were added to
24-hole plate for 24-hour incubation. As soon as 24 hours passed,
fresh 0.5% calf serum and RPMI 1640 culture solution with various
concentrations of Xuezhikang were replaced for another 24-hour cultivation.
Six hours before the reaction, each hole was added with 0.5 mg/ml
3HTdR. Cell scraper was utilized to strip cells and micro-porous
membrane used to collect those cells. After that, PBS was used to
wash membrane film repeatedly and the washed out was put in several
bottles. Then, these bottles were placed in over under temperature
of 70oC for certain period. Line 3H-TdR mixing rate of Beckman 6500
Liquid Flashing Instrument was used to determine the concentration.
The cell suspended solution incubated by RPMI 1640 for 24 hours
was then incubated for another 72 hours with fresh 15% calf serum
and various concentrations of Xuezhikang. Finally, haemocytes counting
plate was utilized to count VSMC numbers.
Result of Serum Lipid
There was no obvious difference of the data
among each group before the treatment. However, serum TC, TG and
LDL-C concentrations of hypercholesterol and Xuezhikang groups rose
significantly after the treatment (P < 0.05). But the above concentrations
in Xuezhikang group were remarkably lower than that of hypercholesterol
(P < 0.05). Whereas no significant differences of HDL-C level
existed between hypercholesterol group, Xuezhikang group and the
control group (P > 0.05). Nevertheless, a clear downturn of HDL-C
level occurred in hypercholesterol group.
Effects of Xuezhikang Concentration on the Proliferation of VSMC
Under In-Vitro Conditions (See the following Table)
The Effects of Xuezhikang Concentration on the Proliferation
of VSMC Under In-Vitro Conditions
With the increase of Xuezhikang concentration, VSMC counts gradually
reduced and 3H-TdR mixing rate also gradually went down (P <0.05).
Electron Microscope Observation
The internal plastic plate (IPP) in the control group appeared intact,
plane and even in thickness. Mid-membrane SMC contracted and inter-paralleled
with IPP. Whereas underlying fissure of endothelium of hypercholesterol
group expanded significantly. IPP appeared uneven in thickness and
broke or even seemed like worm-bite form. Mid-membrane SMC was seen
rose and intruded into underlying fissures through broken IPP. Synthesized
surface form dominated SMC and endoplasmic reticulum increased and
dilated. Plastiosomes increased and swelled. Meanwhile, ridges reduced
or vanished or even bubbled. And lipid droplets came into being
in endochylema with difference in both number and size. Partial
cytoplasm lowered its density or concentrated. Myofilament dense
body became vague or even disappeared. But the control group showed
very little of the above changes or closed to normal.
study demonstrated that Xuezhikang not only had effective performance
to regulate lipid level, but also significantly inhibits the proliferation
and migration of VSMC. This may be one of the important mechanism
of Xuezhikang to prevent the formation of atherosclerosis and the
re-occurrence or development of vessel restenosis after antioplasty
or endangium resection.
reductase inhibitor is the speed retarding enzyme for cholesterol
synthesis and mevalonic acid is the product of HMG-CoA reductase
catalytic reaction. And mevalonic acid metabolism is the necessity
for DNA synthesis and cell proliferation.
may directly inhibits the proliferation, migration and gene expression
of SMC by suppressing mevalonic acid metabolism. Or it indirectly
inhibits the proliferation and migration of SMC by adjusting lipid
metabolism, improving functions of vessel endothelial cells, reducing
the concentrations of plasma ET-1 and TXB2, increasing plasma
PGI2 and NO and decreasing the release of platelet source growing
JH, Fuster V, Badimon L. Syndromes of accelerated atherosclerosis:
role of vascular injury and smooth muscle cell proliferation.
J Am Coll Cardiol, 1990, 15:1667-1687.
A. Relationship between mevalonate pathway and arterial myocyte
proliferation: in vitro studies with inhibition of HMG-CoA
reductase. Atherosclerosis, 1993, 101:117-125.