S. Hassenstein

Smooth muscle cell proliferation and restenosis after stand alone coronary excimer laser angioplasty

It has been shown that coronary excimer laser angioplasty can remove atherosclerotic intracoronary tissue. Stand alone coronary excimer laser angioplasty was successfully performed in a 53 year old white man with 90% stenosis of the left anterior descending coronary artery and exertional angina (Canadian Cardiovascular Society class III). The lesion was reduced to a 30% residual stenosis with use of a 1.2 mm and subsequently a 1.8 mm diameter laser catheter. Early follow-up angiography 24 h later revealed persistent patency and unchanged lesion diameter of the target vessel. The patient was free of symptoms during the 2 month follow-up period, but died suddenly while playing in a tennis tournament 63 days after the procedure. Postmortem histologic examination revealed 80% restenosis at the lesion site without plaque disruption or thrombosis. Specific staining of the histologic specimen for smooth muscle cells using alpha-actin revealed significant smooth muscle cell proliferation at the site of coronary excimer laser angioplasty. However, most of the vessel narrowing appeared to be due to underlying fibrotic plaque as a result of insufficient tissue ablation. This was probably related to the size of the currently available catheters, which are too small to create a large channel.

Inhibition of cellular proliferation after experimental balloon angioplasty by low-molecular-weight heparin.

Background The proliferative response induced by balloon angioplasty is known to be an important factor in the development of restenosis after successful coronary angioplasty. Methods and Results To study the effects of low-molecular-weight heparin (LMWH) on cellular proliferation after experimental balloon angioplasty, LMWH (3.9 kd, 400 anti-Xa units/kg/day) was given to 20 male New Zealand White rabbits. After an intimal fibromuscular plaque was induced by electrical stimulation in the right carotid artery, LMWH was applied during the 7 days after balloon dilatation. As the control group, 20 other rabbits underwent balloon angioplasty without application of LMWH. The vessels were excised 3, 7, 14, and 28 days after balloon treatment. During the final 18 hours before the rabbits were killed, bromodeoxyuridine was applied. Intimal wall thickness increased from 13±5 cell layers (preangioplasty control group) to 20±6 cell layers in the LMWH-treated group at 28 days (p < 0.05). In contrast, histological examination of control animals 28 days after angioplasty revealed a significant increase to 35±15 cell layers (p < 0.01). Immunohistological quantification showed a significant increase (p < 0.001) of cells undergoing DNA synthesis at 3 (10.2±4.2%) and 7 (7.7±4.8%) days after balloon dilatation in control animals. In contrast, at 3 and 7 days after balloon treatment, the percentage of cells undergoing DNA synthesis in LMWH-treated rabbits was lower (3 days, 2.7±1.8%; 7 days, 1.9±0.3%) than the corresponding untreated controls but showed a significant increase (p < 0.01) compared with the preangioplasty controls. The differences between the two groups were statistically significant, however (3 days, p < 0.01; 7 days, p < 0.05). As early as 14 days after angioplasty, the extent of cellular proliferation was normalized and was comparable to the preintervention levels in both groups. Conclusions Our data indicate that the proliferative response after balloon angioplasty can be reduced in vivo by early treatment with LMWH and thus encourage further clinical investigations.

Morphological changes and smooth muscle cell proliferation after experimental excimer laser treatment.

BackgroundLittle is known about the mechanism(s) in the development of restenosis after excimer laser angioplasty. Thus, the rationale of this study was to determine the time course of intimal and medial smooth muscle cell (SMC) proliferation and histomorphological changes after experimental excimer laser treatment. Methods and ResultsLaser ablation was performed in the right carotid artery of 34 New Zealand White rabbits after development of a fibromuscular plaque by repeated weak electrical stimulations. The vessels were excised 3, 7, 14, 21, 28, and 42 days after excimer laser treatment. Staining of a-actin was used to identify SMCs. In five rabbits (15%), a stenosis of more than 50% of luminal area was due to intimal proliferation of SMCs, and in four other rabbits, a total occlusion was due to organized thrombi. After the initial ablation of the preformed plaque (13±6 intimal SMC layers) a continuous increase of intimal wall thickness was found from 7 + 6 SMC layers at 7 days to 28±5 intimal SMC layers at 28 days after excimer laser ablation (p < 0.01). After 42 days, no additional increase of intimal thickening occurred. After bromodeoxyuridine labeling, the extent of cell proliferation (percent of cells undergoing DNA synthesis) in the intima and media was determined using a monoclonal antibody against bromodeoxyuridine. Immunohistological quantification of SMC proliferation in the intima revealed a significant increase of cells undergoing DNA synthesis at 3 (p <0.05) and 14 (p <0.01) days after laser treatment. Medial proliferation of SMCs was delayed and had a significant increase 7 days (p < 0.05) after intervention. Twenty-one days after laser treatment, SMC proliferation in the intima as well as in the media was normalized. ConclusionsThe proliferative response of SMCs after experimental excimer laser treatment will occur as a dynamic process with a maximum of SMCs undergoing DNA synthesis during 14 days after laser ablation, resulting in an increase of intimal thickening within 4 weeks after laser treatment. The extent of intimal hyperplasia due to SMC proliferation after excimer laser treatment is comparable with the effect of transluminal balloon angioplasty in this experimental model.

Vascular injury and time course of smooth muscle cell proliferation after experimental holmium laser angioplasty.

BackgroundIn vitro experiments have shown that holmium laser energy can effectively ablate even calcified plaque in human arterial vessels. Because high-energy densities from holmium lasers can easily be transmitted through quartz fibers, this solid-state laser has been suggested as an alternative intraluminal treatment of atherosclerotic plaque. Methods and ResultsTo develop an intimal plaque, 35 New Zealand White rabbits underwent electrical stimulation of their right carotid artery for 28 days. Subsequently, in 25 rabbits, holmium laser angioplasty (wavelength, 2.12 pm; pulse duration, 150 jasec; energy density, 350 mJ/mm2) was performed. To study the morphological results, the vessels were excised after 7, 14, 28, and 42 days. Cross sections were analyzed in regard to laser-specific injury. Staining of α-actin was used to identify smooth muscle cells (SMCs). After bromodeoxyuridine labeling, the extent of proliferation (number of cells undergoing DNA synthesis) was determined by using a monoclonal antibody. Holmium laser ablation resulted in an initial decrease of the numbers of intimal cell layers in the early group (7 days after treatment: 5±1 cell layers with 76±39 μm; control: 13±3 cell layers with 144±44 μm). Quantification of SMCs undergoing DNA synthesis in the intima (control: 51±19 cells/mm2) showed a significant increase of labeled cells after 7 (216±74 cells/mm2, p=0.003) and 14 days (281±139 cells/mm2, p=0.011). Integrity of the internal elastic lamina was disrupted in all animals after intervention. Seven and 14 days after treatment, a considerable reduction of medial cell nuclei was found in 10 of 12 animals. SMC proliferation in the medial layer was increased within the first 2 weeks after laser ablation (168± 113 cells/mm2; control: 8±4 cells/mm2; p=0.023). Six weeks after holmium laser angioplasty, SMC proliferation had returned to control levels in the intima and remained increased in the medial layer. This proliferative response resulted in a significant increase of intimal thickening within 6 weeks after laser ablation (30±6 cell layers, 375±97 μm resp.; p=0.001 each). ConclusionsHolmium laser treatment leads to considerable vessel wall injury and results in SMC proliferation in the intimal and medial layer with a maximum of proliferative activity within the first 2 weeks. Subsequently, this results in considerable intimal and medial hyperplasia within 6 weeks after treatment.

Detection of macrophages following experimental balloon angioplasty

Ziel der vorliegenden Studie war es, die intimalen Makrophagen im zeitlichen Verlauf nach experimenteller Ballonangioplastie quantitativ zu erfassen. Nach Erzeugung einer Plaque in der Arteria carotis durch Elektrostimulation wurde bei 32 Kaninchen eine Ballonangioplastie durchgefuhrt. Die Gefase wurden 3, 7, 14, 21, 28 bzw. 42 Tage nach Intervention histologisch aufgearbeitet. Unter Verwendung eines monoklonalen Antikorpers (RAM 11) erfolgte die prozentuale Bestimmung der Makrophagen in der Neointima. Ausgehend von 2,4± 2,6 % in der Kontrollgruppe fand sich ein signifikanter Anstieg der Makrophagen vom 7. (7,9± 2,1 %) bis zum 21. (6,9± 3,1 %) Tag nach Intervention. Am 28. und 42. Tag nach Dilatation war die Anzahl der intimalen Makrophagen wieder mit der Kontrollgruppe vergleichbar.

In vitro Alexandrite laser angioplasty: A study on fibre conduction and tissue effects

Pulsed ultra-violet excimer laser radiation is capable of tissue ablation with only minimal thermal injury of adjacent tissue structures. Since difficult fibre optic coupling of energy was observed, alternative Q-switched laser sources capable of ablation of atherosclerotic plaque are under current investigation. To evaluate tissue effects of Alexandrite laser radiation, 160 arterial segments with macroscopic evidence of atherosclerotic disease were treated. The laser light was transmitted via silica based quartz fibres with different diameters. Using the Q-switched Alexandrite laser at the fundamental wavelength (748 nm) with a pulse duration of 300 ns the energy density threshold for tissue ablation was found to be in the range of 63 to 126 J cm−2 using a 300μm fibre. On macroscopic examination only limited thermal and acoustic injury was found in crater adjacent tissue structures. Crater edges were even and did not reveal signs of crater charring or debris in the crater lumen. However, the histological cross-sections revealed thermal injury extending from 100 up to 200μm lateral into adjacent tissue. The crater margins revealed fissuring as a result of shock wave injury. Thermal damage was most evident if irradiation of atherosclerotic tissue was performed in blood.

Smooth excimer laser coronary angioplasty (SELCA) and conventional excimer laser angioplasty: Comparison of vascular injury and smooth muscle cell proliferation

Although the excimer laser, which utilizes ‘non-thermal ablation effects’, has achieved encouraging results in early clinical trials, the long-term results have failed to show any advantage over conventional percutaneous transluminal coronary angioplasty (PTCA).A new system, Smooth Excimer Laser Coronary Angioplasty (SELCA), has been developed to reduce the tissue damage in the vessel wall caused by shock waves and vapour bubbles.SELCA (wavelength 308 nm, pulse duration 115 ns, repetition rate 150 Hz and energy density 50 mJ mm-2) lowers the amount of shock wave formation and pressure peak amplitude in the surrounding tissue by about eight times when compared to the conventional 308 nm excimer laser (ELCA). In this preclinical evaluation, this new system was compared to ELCA. Fifty New Zealand White rabbits were stimulated by repeated weak DC impulses for a period of 28 days in order to form an atherosclerotic plaque in the right carotid artery. The vessels were excised 3, 7,14 and 28 days after laser irradiation for immunohistochemical analysis.SELCA and ELCA laser treatment lead to a decrease in maximal intimal wall thickness 3 days after intervention (control: 177±4 µm; SELCA: 131±22µm; ELCA: 120 ±33µm). In the period between 3 and 28 days, a moderate increase in intimal wall thickness was observed after SELCA treatment compared to a significant increase after ELCA (28 days after intervention: SELCA: 157±22µm; ELCA: 274 ±28µm). Bromodeoxyuridine (BrdU) was applied 18 and 12 h before excision of the vessels in order to determine the percent of cells undergoing DNA synthesis. The percent of BrdU labelled SMC in the intima (control: 13 ± 2 cells mm-2) increased in both groups after 3 days (SELCA: 248 ± 107 cells mm-2; ELCA: 162 ± 41 cells mm-2) and 7 days (SELCA: 162± 55 cells mm-2; ELCA: 279 ± 119 cells mm-2).The present results demonstrate that vascular wall injury and increase in intimal wall thickness following SELCA are reduced in comparison to the results achieved with the conventional technique. Further trials are necessary to assess whether these improvements will lead to more favourable long-term results after excimer laser angioplasty.

Immunohistochemical detection of proliferation of smooth muscle cells after experimental angioplasty: comparison of BrdU incorporation and PCNA expression

Restenosierung durch Proliferation glatter Muskelzellen limitiert in bis zu 30% der Falle den klinischen Nutzen der Ballonangioplastie [7]. Die auslosenden Faktoren und die Regelmechanismen dieser Wachstumsvorgange sind Gegenstand zahlreicher experimenteller Studien. Somit werden exakte Analysen der Kinetik der Zellproliferation zunehmend wichtiger.

Effect of low molecular weight heparin on intimal smooth muscle cell proliferation after experimental balloon angioplastyEffekt von niedermolekularem Heparin auf die Proliferation glatter Muskelzellen nach experimenteller Angioplastie

Die Entwicklung von Restenosen nach primar erfolgreicher perkutaner transluminaler Koronarangioplastie (PICA) stellt auch weiterhin das ungeloste Problem dieser invasiven kardiologischen Therapie dar [7].

Effekt von niedermolekularem Heparin auf die Proliferation glatter Muskelzellen nach experimenteller Angioplastie

Die Entwicklung von Restenosen nach primar erfolgreicher perkutaner transluminaler Koronarangioplastie (PICA) stellt auch weiterhin das ungeloste Problem dieser invasiven kardiologischen Therapie dar [7].