**6. Ex Vivo Model**

Four-hour preincubation of cultured cells with verapamil led to complete prevention of the serum atherogenic effect [65]. Thus, verapamil possesses not only an atherosclerotic effect in culture causing the regression of atherosclerotic manifestations at the cellular level but also elicits an anti-atherogenic, i.e. preventive effect, eliminating atherogenic potential of the serum.

**ANTI-ATHEROSCLEROTIC ANTI-ATHEROGENIC**

Atherosclerotic plaque Uninvolved intima

Standard (nonatherogenic) serum Atherogenic patients' serum

but also anti-atherogenic, i.e. preventive effect at the arterial cell level.

Cholesterol fall Prevention of cholesterol accumulation

The effect of several calcium antagonists on primary cholesterol accumulation in cultured cells induced by the patients' serum was tested. Verapamil and nifedipine completely inhib‐ ited the accumulation of intracellular cholesterol induced by the serum while other calcium antagonists: diltiazem, nicardipine, isradipine, darodipine rather substantially reduced cho‐ lesterol accumulation [65]. As it is known, the examined calcium antagonists manifested an‐ ti-atherogenic action in vivo inhibiting the development of experimental atherosclerosis in animals [66,67]. Thus, our in vitro data obtained on cellular model correspond to the in vivo observations. One can conclude that calcium antagonists elicit not only anti-atherosclerotic

Nitrates and beta-blockers have been tested to reveal their action on atherosclerotic cellular indices. Nitrates had no effect on proliferative activity of atherosclerotic cells and practically did not affect the cholesterol level [55]. On the other hand, all the examined beta-blockers, propranolol, alprenolol, metoprolol, pindolol, and timolol, more or less increased athero‐ sclerotic manifestations, i.e. all of these drugs exhibited atherogenic activity in culture [55,60]. If beta-blockers manifest a similar action in vivo, one may assume that these drugs are atherogenic and realize the atherogenic action at the arterial cell level. Apparently, ni‐

The influence of cardiovascular drugs on atherosclerosis-related effects of each other was studied. The study was focused on metoprolol, nifedipine and nitroglycerin, the drugs widely used in clinic [55]. Metoprolol caused an elevation of intracellular cholesterol, nifedi‐ pine reduced the cholesterol level while nitroglycerin was without effect on this index. The use of nifedipine on the background of metoprolol did not modify the anti-atherosclerotic action of the calcium antagonist. In this combination atherogenic action of metoprolol was not revealed. The application of metoprolol in combination with nitroglycerin led to the elimination of an atherogenic effect of the beta-blocker. Nifedipine used together with meto‐ prolol and nitroglycerin was just as effective as in the absence of these drugs. Thus, nifedi‐ pine produces its anti-atherosclerotic effects both by itself and in combination with widely used nitrates and beta-blockers. These data suggest one important conclusion. Atherogenic

Regression Prevention

194 Current Trends in Atherogenesis

**Table 2.** Anti-atherosclerotic and anti-atherogenic drug effects in culture

trates are neutral, indifferent in this respect.

All the above conclusions and hypotheses are based on the data obtained in in vitro experi‐ ments. Obviously, the question arises, whether anti-atherosclerotic effects of calcium antago‐ nists and atherogenic effects of beta-blockers can be manifested in vivo and what is the optimal anti-atherosclerotic therapy based on calcium antagonists and other drugs.

To optimize anti-atherosclerotic and anti-atherogenic drug therapy, ex vivo model was de‐ veloped. In case of ex vivo model not drug but blood serum taken from patients after oral drug administration is added to cultured cells.

Calcium antagonists, verapamil and nifedipine, and beta-blockers, propranolol and pindo‐ lol, were examined using ex vivo model. Within 2-4 hrs after nifedipine or verapamil admin‐ istration, the patient's serum had anti-atherosclerotic properties, i.e. it was able to cause a fall in the intracellular cholesterol and inhibited atherosclerotic cell proliferation [55,56]. On the contrary, the serum of patients who received propranolol or pindolol was pro-athero‐ genic. Its pro-atherogenic properties manifested themselves at the arterial cell level in the rise of intracellular cholesterol and stimulation of cell proliferation [55,56]. This finding al‐ lows to assuming that not only in vitro, but in vivo as well, calcium antagonists and betablockers are anti-atherosclerotic and atherogenic drugs, respectively.

The effect of nifedipine on serum properties during a prolonged course was assessed. A pa‐ tient was on nifedipine for 7 days. He received 20 mg doses three times a day with an 8-hr interval. Twenty-eight days after regular nifedipine therapy the initial atherogenicity of the patient's serum was substantially lower than at the beginning. Directly after a dose of nifedi‐ pine the atherogenicity was practically completely eliminated [65]. On the contrary, as a re‐ sult of a prolonged therapy with a beta-blocker, propranolol, patient's serum acquired stable atherogenic properties. At the beginning of the course the serum of this patient was nona‐ therogenic, however, 28 days of regular propranolol therapy led to the emergence of athero‐ genicity revealed even before the drug administration [65]. Thus, a single dose of betablockers brings about temporary atherogenicity of the serum. Prolonged therapy with betablockers leads to the emergence of stable atherogenic properties of patients' blood serum.

with another dose of krill meat. Blood serum samples were added to a culture of subendo‐ thelial cells isolated from uninvolved human aortic intima, and intracellular cholesterol ac‐ cumulation was assessed in each case. Anti-atherogenic activity of krill meat was evaluated by the ability to reduce serum atherogenicity which manifested in cholesterol accumulation in cultured cells (Figure 2). The dose-effect dependence was revealed by comparing the effi‐ cacy of two doses. The efficacy of each dose was evaluated by the analysis of at least 6 sera obtained from different patients. In can be seen from that the krill meat elicits an anti-athe‐ rogenic effect at a dose of 10-20 g, half maximum effect was reached at a dose of 30 g and the

Use of Natural Products for Direct Anti-Atherosclerotic Therapy

http://dx.doi.org/10.5772/52967

197

We believe that this approach will be useful in the development and optimization of anti-

maximum effect - at a dose of 50 g.

involved intima of the human aorta.

be a good alternative.

atherosclerotic and anti-atherogenic diet therapies.

**Figure 2.** Antiatherogenic effect of krill meat on blood serum of atherosclerotic patients

**8. Natural Products for Anti-Atherosclerotic Therapy**

Blood serum atherogenicity was determined using primary culture of cells derived from un‐

Anti-atherogenic effects of dietary products promote the development of anti-atherosclerotic therapy on the basis of natural products. Atherosclerosis develops over many years, so the anti-atherosclerotic therapy should be long-term or even lifelong therapy. For such a longterm therapy conventional medicine might not work. Drugs based on natural products can
