Vascular smooth muscle cell growth kinetics in vivo in aged rats.

Age is a risk factor in the development of atherosclerosis. In this study we investigated the hypothesis that proliferation of vascular smooth muscle cells (SMCs), an integral part of atherosclerotic plaque formation, changes with age. SMC growth kinetics of old rats (21-24 months) were compared to those of young adult rats (3-4 months). Rat aortas were denuded of their endothelium and the animals were killed after [3H]thymidine and Evans blue injections at 0-28 days after denudation. Incorporation of [3H]thymidine into SMC peaked in the young animals by day 2, whereas the older animals responded to endothelial removal with greater incorporation at day 2 and a more sustained rate of incorporation peaking at day 4. The [3H]thymidine incorporation curves decreased sharply from their peaks at 2 and 4 days, respectively, and paralleled each other after day 7. [3H]Thymidine uptake reflected the subsequent SMC intimal growth as measured morphometrically, with old animals showing greater numbers of intimal SMC than did the younger animals. The difference in response of SMC to injury with age suggests that aging produces a change in the vascular SMC that enhances proliferation. This change in response implies that the more pronounced atherosclerotic plaque growth seen with aging may be a result of an age-related increase in response to injury rather than merely the accumulation of time-related intimal change.     

Vascular smooth muscle cells and calcification in atherosclerosis

Vascular calcification is a prominent feature of atherosclerosis but the mechanisms underlying vascular calcification are still obscure. Since bone-associated proteins such as osteonectin, osteocalcin, and matrix Gla protein have been detected in calcified vascular tissues, calcification has been considered to be an organized, regulated process similar to mineralization in bone tissue. Vascular smooth muscle cells (VSMCs) are currently considered to be responsible for the formation of vascular calcifications. Apoptosis of VSMCs appears to be a key factor in this process, while other factors including cell-cell interactions (macrophages and VSMCs), lipids, and plasma inorganic phosphate levels modulate the calcification process. The focus of this review is on the role of VSMCs in the development of calcifications in atherosclerotic plaques.     

Vascular smooth muscle cell senescence in atherosclerosis

Markers of cell senescence have been identified in both the blood and vessel wall of patients with atherosclerosis. In particular, vascular smooth muscle cells (VSMCs) derived from human plaques show numerous features of senescence both in culture and in vivo. This review summarises the evidence for VSMC senescence in atherosclerosis, and outlines the mechanisms and triggers leading to their senescence.     

Vascular smooth muscle phenotype and growth behaviour can be influenced by macrophages in vitro

The effect of macrophages on rabbit vascular smooth muscle phenotype and proliferative ability was examined using ultrastructural morphometry. The volume fraction of myofilaments (Vv myo) in smooth muscle cells (SMC) from 9-week-old rabbit aorta in vivo was 39.5 +/- 1.2%. After seeding the enzymatically isolated SMC at 4 X 10(5) cells/ml in primary culture, the Vv myo was 38.9 +/- 1.2% on day 3 dropping to 29.9 +/- 2.0% by day 5. On day 6 the Vv myo was 29.2 +/- 1.8%, and the cells began to proliferate. Confluency was reached after less than 24 h proliferation and the Vv myo rose abruptly on day 7 to 36.9 +/- 1.9%. When the SMC were co-cultured with macrophages, the Vv myo fell to 31.2 +/- 0.9% on day 3 and to 25.9 +/- 0.5% on day 5 at which time cells commenced proliferation. Confluency occurred on day 6 but the SMC Vv myo did not rise throughout the rest of the culture period (27.4 +/- 1.8% and 26.9 +/- 1.3% on days 7 and 9, respectively) and the cells, unlike the controls, continued to proliferate, becoming multi-layered. Early phenotypic modulation in sparsely seeded SMC (8 X 10(4) cells/ml) co-cultured with macrophages was also found using fluorescent labelled antibodies to smooth muscle myosin. Measurement of proliferation by cell counts (and tritiated thymidine autoradiography) showed that macrophages stimulated SMC in primary culture to proliferate at a significantly greater rate than control cells grown alone in 5% whole blood serum (WBS). Proliferation of subcultured SMC co-cultured with macrophages was also stimulated.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pasteur effect in vascular and intestinal smooth muscle

The increase in lactate production on changing from aerobic to anaerobic conditions, i.e. the Pasteur effect, has been reported to be small in vascular muscle and especially in aorta. It has been suggested that this may be an artefact caused by damage to the intimal endothelium. We have compared the Pasteur effect in different kinds of pig arteries, but also in rabbit colon. The aerobic lactate production in 60 min was 11-15 mumol/g in the aorta and the carotid artery, but 3 mumol/g in the mesenteric and renal arteries and 4 mumol/g in the rabbit colon. The increase in lactate production under anaerobic conditions was 12-20 mumol/g/60 min in the carotid artery, aorta and rabbit colon and 10 mumol/g/60 min in the mesenteric and renal arteries. When calculated in per cent, the Pasteur effect was greater in the mesenteric artery than in the aorta, but the actual rise in lactate production in mumol/g was higher in the aorta and carotid artery. The high aerobic lactate production of smooth muscle in vitro may be related to its low ability to oxidize glucose; some other substrates may be preferentially oxidized when present in vitro or in vivo.     

Vascular smooth muscle growth in genetic hypertension: evidence for multiple abnormalities in growth regulatory pathways.

OBJECTIVE: To gain insight into the mechanisms which contribute to the development of vascular hypertrophy in the spontaneously hypertensive rat (SHR). DESIGN: These experiments were performed under conditions which most closely mimic the growth of smooth muscle in blood vessels, i.e. once cell-cell contact has been achieved. METHODS: A comparison of the growth characteristics (growth rates and cell density at quiescence) of vascular smooth muscle cells (VSMC) from SHR and normotensive Wistar-Kyoto (WKY) rats. RESULTS: In the presence of foetal calf serum (1, 2.5, 5 and 10%), early passaged VSMC from SHR exhibited higher growth rates and reached higher densities at quiescence than VSMC from WKY rats. Accelerated growth rates could not be attributed to differences in cell-cell interactions. Also, growth rates and cell density at quiescence appear to be regulated by distinct mechanisms. Transforming growth factor-beta 1 (TGF-beta 1) caused an inhibition of serum-stimulated proliferation of confluent VSMC from WKY rats. In contrast, TGF-beta 1 had little, if any, inhibitory action upon the growth of VSMC from SHR. Scatchard analysis of 125I-TGF-beta 1 binding to VSMC from both strains yielded a single class of high affinity binding sites. CONCLUSIONS: VSMC from SHR exhibit enhanced proliferation, attain a higher cell density at quiescence and are less susceptible to growth inhibition by TGF-beta 1 than VSMC from WKY rats. All these characteristics of SHR VSMC may contribute to the development of vascular hypertrophy in this strain.     

Myenteric denervation of rat jejunum alters calcium responsiveness of intestinal smooth muscle

Long-term myenteric and extrinsic denervation of a segment of rat jejunum results in increased stress generation by the longitudinal muscle layer of the denervated segment 15 days after denervation. This study examined whether alterations in the properties of either cell membrane calcium channels and/or sarcoplasmic reticular Ca²⁺-adenosine triphosphatase (ATPase) contribute to the increased stress development. The effects of the calcium channel blocker nifedipine and the sarcoplasmic reticular Ca²⁺-ATPase inhibitor cyclopiazonic acid on the contractile activity of denervated and control smooth muscle were determined. The ability of nifedipine to inhibit KCl-induced contractions was significantly increased in denervated tissues; however, there was no difference in the potency of nifedipine when tissues were stimulated with carbachol. Calcium concentration-response curves obtained in the presence of either KCl or carbachol were determined in tissues previously depleted of calcium. Long-term denervated tissues showed an increased sensitivity to calcium and a decreased maximum contractile response after stimulation with carbachol. Cyclopiazonic acid inhibited repletion of intracellular calcium stores of control muscle but had no effect in denervated tissue. Long-term denervation of a segment of rat small intestine results in profound alterations in calcium metabolism at the cell membrane and, to a lesser extent, at the sarcoplasmic reticulum of smooth muscle cells of the longitudinal muscle layer.     

The physiology of intestinal smooth muscle

Conclusion Intestinal smooth muscle is an extremely complex structure not only because of the many interactions between the different types of cells, but also because of the many peculiarities of smooth-muscle cells themselves Only an intensive study on the cellular level with biophysical and biochemical technics will provide us with an understandable picture of smooth-muscle cells and the tissues which they compose.     

Stretch-dependent modulation of contractility and growth in smooth muscle of rat portal vein

Increased intraluminal pressure of the rat portal vein in vivo causes hypertrophy and altered contractility in 1 to 7 days. We have used organ cultures to investigate mechanisms involved in this adaptation to mechanical load. Strips of rat portal vein were cultured for 3 days, either undistended or loaded by a weight. Length-force relations were shifted toward longer length in stretched cultured veins compared with freshly dissected veins, whereas the length-force relations of unstretched cultured veins were shifted in the opposite direction. This occurred after culture either with or without 10% FCS to promote growth. The wet weight of loaded veins increased by 56% in the presence of FCS, whereas that of undistended control veins increased by 24%. No weight increase was seen in serum-free culture. The dry/wet weight ratio decreased during culture with FCS but was not affected by stretch. Electron microscopy revealed increased cell cross-sectional area in stretched relative to unstretched veins, and protein contents were greater, as were [(3)H]thymidine and [(3)H]leucine incorporation rates. Growth responses were associated with the activation of stretch-sensitive extracellular signal-regulated kinases 1 and 2 and were inhibited by herbimycin A and PD 98059, inhibitors of extracellular signal-regulated kinases 1 and 2. The results demonstrate that by culture of whole vascular tissue, smooth muscle cells are maintained in the contractile phenotype and respond to stretch with a physiological adaptation involving hypertrophy/hyperplasia and remodeling of the contractile system, similar to that in vivo. Mechanical stimulation and growth factors are both required for functionally significant growth.     

Leukocyte adherence in venules of rat skeletal muscle following thermal injury

An intravital microscopic study of rat skeletal muscle microvasculature was undertaken to explore the relationship of adherent leukocytes to the venous microcirculation following thermal injury. WBC adherence was increased in venules in the acute period following thermal injury. This increase was not dependent upon changes in wall shear stress. The presence of sticking leukocytes was associated with a disruption of the normal relationship between venular radius and flow. In contrast to previous studies, there was no evidence for erythrocyte sludging or diameter changes in venules following thermal injury.     

血管平滑肌细胞表型转化与血管钙化

血管钙化是一种由细胞所介导、主动的生物矿化过程,可增加心血管疾病的患病率和死亡率,并严重危害人类的健康和生活。越来越多的研究证实血管平滑肌细胞(Vascular smooth muscle cell,VSMC)及表型转化（phenotypic switching）在血管钙化的发生发展中具有重要作用。本文将阐述VSMC的表型转化,向骨/软骨化表型转化不同时期的标志蛋白分子,并探讨其表型转化的调控因素,进而深入认识血管钙化的发病过程。     

Vascular smooth muscle transmembrane potentials during hypotensive stress

Although transmembrane potential (Em) appears to be an important regulator of vascular smooth muscle (VSM) contractile force in vivo, little is known of the electrophysiological changes in VSM during low flow states. In rats, VSM in small mesenteric veins depolarizes in the compensatory stage of hemorrhagic hypotension while the onset of peripheral vascular decompensation is associated with VSM hyperpolarization. Pretreatment with 30 mg/kg i.v. of methylprednisolone prevents the Em changes in mesenteric veins of hemorrhaged rats and reduces the depolarization of isolated vessels in response to norepinephrine, suggesting that some therapeutic agents may act by modifying the electrophysiological responses of VSM during the stress. Finally, adrenergic depolarization under resting conditions may contribute to a reduced ability of spontaneously hypertensive rats (SHR) to tolerate blood loss by reducing their compensatory venoconstrictor reserve relative to normotensive WKY controls. Although these observations suggest that vascular responses to hypotensive stress are related to the effects of the sympathetic nervous system upon VSM Em, further studies are required fully to elucidate the relative role of Em-dependent and Em-independent mechanisms in controlling active VSM tone during low flow states.     

Vascular smooth muscle cell proliferation is effectively suppressed by the non-specific growth factor inhibitor suramin

The purpose of this study was to investigate the effects of the non-specific growth factor inhibitor suramin on smooth muscle cell proliferation in vitro and in vivo. Cultured vascular smooth muscle cells (VSMC) were stimulated by platelet-derived growth factor (PDGF) and cellular DNA synthesis assessed by [3H]-thymidine uptake. Suramin dose-dependently inhibited DNA synthesis in VSMC, and 100 microM of suramin completely suppressed the PDGF-AB-induced cellular DNA synthesis. Rabbit carotid arteries were injured by the balloon catheter, and then suramin locally delivered using a porous balloon catheter over ten minutes. Three weeks after the vascular injury, the extent of intimal thickening was compared between the suramin-treated and control rabbits. The neointimal formation triggered by balloon-mediated vascular injury was suppressed significantly and dose-dependently by locally infused suramin, and the intima to media area ratios of the control and 1 mM suramin-treated animals were 48.8+/-14.9 and 12.2+/-6.0%, respectively (p < 0.01. n = 6 for each group). These results suggest that one time local administration of suramin was sufficient to suppress neointimal formation after balloon-mediated vascular injury, and that pharmacological intervention targeting the growth factor's signaling pathways could be a promising approach to prevent smooth muscle cell proliferation in various proliferative vascular diseases.     

Vascular smooth muscle cell phenotypes in primary pulmonary hypertension.

Primary pulmonary hypertension (PPH) is associated with specific structural alterations, including cellular intimal thickening, intimal fibrosis, and plexiform lesions. To determine the phenotypes of smooth muscle cells (SMCs) in such lesions, the authors conducted an immunohistochemical analysis of lung tissues from two patients with PPH, using two antimuscle actin antibodies, HHF35 and CGA7, and two anti-SMC myosin heavy chain markers, anti-SM1 and anti-SM2 antibodies and related antibodies. Cells that stained positive (+) with HHF35, CGA7, anti-SM1, and anti-SM2 were considered to be SMCs of a mature state. Conversely, those that stained positive with HHF35 and anti-SM1, but weakly positive (+/-) or negative (-) with CGA7 and anti-SM2, were considered to be SMCs exhibiting an immature state. Cellular intimal thickening was composed of SMCs of an immature phenotype (HHF35+, CGA7+/-, SM1+, SM2+/-), accompanied by the expression of fibronectin and the presence of macrophages; intimal fibrosis contained mature SMCs (HHF35+, CGA7+, SM1+, SM2+); and plexiform lesion consisted of proliferative endothelial cells (von Willebrand factor-positive cells, proliferating cell nuclear antigen-positive cells) and underlying immature SMCs (HHF35+, CGA7-, SM1+, SM2-) associated with fibronectin expression and macrophage infiltration. These findings suggest that smooth muscle cells with specific phenotypes may contribute to the development of specific vascular lesions in primary pulmonary hypertension.