Triglyceride-rich lipoproteins prevent septic death in rats.

Triglyceride-rich lipoproteins bind and inactive bacterial endotoxin in vitro and prevent death when given before a lethal dose of endotoxin in animals. However, lipoproteins have not yet been demonstrated to improve survival in polymicrobial gram-negative sepsis. We therefore tested the ability of triglyceride-rich lipoproteins to prevent death after cecal ligation and puncture (CLP) in rats. Animals were given bolus infusions of either chylomicrons (1 g triglyceride/kg per 4 h) or an equal volume of saline for 28 h after CLP. Chylomicron infusions significantly improved survival (measured at 96 h) compared with saline controls (80 vs 27%, P < or = 0.03). Chylomicron infusions also reduced serum levels of endotoxin, measured 90 min (26 +/- 3 vs 136 +/- 51 pg/ml, mean +/- SEM, P < or = 0.03) and 6 h (121 +/- 54 vs 1,026 +/- 459 pg/ml, P < or = 0.05) after CLP. The reduction in serum endotoxin correlated with a reduction in serum tumor necrosis factor, measured 6 h after CLP (0 +/- 0 vs 58 +/- 24 pg/ml, P < or = 0.03), suggesting that chylomicrons improve survival in this model by limiting macrophage exposure to endotoxin and thereby reducing secretion of inflammatory cytokines. Infusions of a synthetic triglyceride-rich lipid emulsion (Intralipid; KabiVitrum, Inc., Alameda, CA) (1 g triglyceride/kg) also significantly improved survival compared with saline controls (71 vs 27%, P < or = 0.03). These data demonstrate that triglyceride-rich lipoproteins can protect animals from lethal polymicrobial gram-negative sepsis.

Human very low density lipoproteins and chylomicrons can protect against endotoxin-induced death in mice.

Endotoxemia stimulates many physiologic responses including disturbances in lipid metabolism. We hypothesized that this lipemia may be part of a defensive mechanism by which the body combats the toxic effects of circulating endotoxin. We tested the effects of mixtures of endotoxin, lipoproteins, and lipoprotein-free plasma and determined the ability of varying concentrations of human very low density lipoproteins (VLDL) and chylomicrons, as well as low density lipoproteins (LDL) and high density lipoproteins (HDL), and of the synthetic lipid emulsion SOYACAL to prevent endotoxin-induced death in mice. This study demonstrates that the triglyceride-rich VLDL and chylomicrons, as well as cholesterol-rich LDL and HDL, and cholesterol-free SOYACAL can protect against endotoxin-induced death. Protection required small amounts of lipoprotein-free plasma, and depended on the incubation time and the concentration of lipoprotein lipid. Despite stringent techniques to prevent exogenous endotoxin contamination eight of ten duplicate VLDL preparations contained endotoxin (5,755 +/- 3,514 ng endotoxin/mg triglyceride, mean +/- SEM) making the isolation of endotoxin-free VLDL difficult. In contrast, simultaneous preparations of LDL and HDL were relatively free of endotoxin contamination (3 +/- 3 and 320 +/- 319 ng/mg total cholesterol, respectively), suggesting that the contamination of VLDL occurs in vivo and not during the isolation procedure. These observations suggest a possible role for increased triglyceride-rich lipoproteins in the host's defense against endotoxemia and infection.

The effect of oxidized lipids in the diet on serum lipoprotein peroxides in control and diabetic rats.

The levels of oxidized serum lipoproteins are increased in humans and animals with diabetes. We have examined the contribution of dietary oxidized lipids on the levels of oxidized lipoproteins. In both control and streptozocin induced diabetic rats, the oxidized lipid content of mesenteric lymph chylomicrons (CM) increased when increasing quantities of oxidized lipids were administered intragastrically. However, at all levels of administered oxidized lipids, the quantity of oxidized lipids in CM was greater in the diabetic animals. These results indicate that oxidized lipids are absorbed and packaged into CM and suggest that there is increased absorption of oxidized lipids in diabetic animals. In nondiabetic rats fed a fat-free diet, the levels of oxidized lipids in their serum lipoproteins were very low. When oxidized lipids were added to the diet, the quantity of peroxides in serum lipoproteins increased about fivefold. In diabetic animals fed a fat-free diet, there were also very low levels of oxidized lipids in their serum lipoproteins, and there was no difference between control and diabetic rats. However, when diabetic animals were fed a diet containing oxidized lipids, the quantity of oxidized lipids in their serum lipoproteins increased 16-fold and were significantly greater than in controls. Thus, in both control and diabetic rats the quantity of oxidized lipids in the diet largely determines the levels of oxidized lipids in circulating lipoproteins. However, in diabetic animals the effect of diet is more pronounced. Together with the CM studies, these results demonstrate that dietary oxidized lipids make a major contribution to the levels of oxidized lipids in circulating lipoproteins and indicate that increased absorption of oxidized lipids in diabetic animals may play a role in the elevation of oxidized lipoproteins observed in this disorder.

Endotoxin and cytokines increase hepatic messenger RNA levels and serum concentrations of apolipoprotein J (clusterin) in Syrian hamsters.

Infection and inflammation induce alterations in hepatic synthesis and plasma concentrations of the acute phase proteins. Our results show that apolipoprotein (apo) J is a positive acute phase protein. Endotoxin (LPS), tumor necrosis factor (TNF), and interleukin (IL)-1 increased hepatic mRNA and serum protein levels of apo J in Syrian hamsters. Hepatic apo J mRNA levels increased 10- to 15-fold with doses of LPS from 0.1 to 100 micrograms/100 g body weight within 4 h and were elevated for > or = 24 h. Serum apo J concentrations were significantly increased by 16 h and further elevated to 3.3 times that of control, 24 h after LPS administration. Serum apo J was associated with high density lipoprotein and increased fivefold in this fraction, after LPS administration. Hepatic apo J mRNA levels increased 3.5- and 4.6-fold, with TNF and IL-1, respectively, and 8.2-fold with a combination of TNF and IL-1. Serum apo J concentrations were increased 2.3-fold by TNF, 79% by IL-1, and 2.9-fold with a combination of TNF and IL-1. These results demonstrate that apo J is a positive acute phase protein.

Rabbit aorta and human atherosclerotic lesions hydrolyze the sphingomyelin of retained low-density lipoprotein. Proposed role for arterial-wall sphingomyelinase in subendothelial retention and aggregation of atherogenic lipoproteins.

Aggregation and retention of LDL in the arterial wall are key events in atherogenesis, but the mechanisms in vivo are not known. Previous work from our laboratories has shown that exposure of LDL to bacterial sphingomyelinase (SMase) in vitro leads to the formation of LDL aggregates that can be retained by extracellular matrix and that are able to stimulate macrophage foam cell formation. We now provide evidence that retained LDL is hydrolyzed by an arterial-wall SMase activity. First, we demonstrated that SMase-induced aggregation is caused by an increase in particle ceramide content, even in the presence of excess sphingomyelin (SM). This finding is compatible with previous data showing that lesional LDL is enriched in SM, though its ceramide content has not previously been reported. To address this critical compositional issue, the ceramide content of lesional LDL was assayed and, remarkably, found to be 10-50-fold enriched compared with plasma LDL ceramide. Furthermore, the ceramide was found exclusively in lesional LDL that was aggregated; unaggregated lesional LDL, which accounted for 20-25% of the lesional material, remained ceramide poor. When [3H]SM-LDL was incubated with strips of rabbit aorta ex vivo, a portion of the LDL was retained, and the [3H]SM of this portion, but not that of unretained LDL, was hydrolyzed to [3H]ceramide by a nonlysosomal arterial hydrolase. In summary, LDL retained in atherosclerotic lesions is acted upon by an arterial-wall SMase, which may participate in LDL aggregation and possibly other SMase-mediated processes during atherogenesis.

Chylomicrons alter the fate of endotoxin, decreasing tumor necrosis factor release and preventing death.

The hypertriglyceridemia of infection was traditionally thought to represent the mobilization of substrate to fuel the body's response to the infectious challenge. However, we have previously shown that triglyceride-rich lipoproteins can protect against endotoxin-induced lethality. The current studies examine the mechanism by which this protection occurs. Rats infused with a lethal dose of endotoxin preincubated with chylomicrons had a reduced mortality compared with rats infused with endotoxin alone (15 vs. 76%, P < 0.001). Preincubation with chylomicrons increased the rate of clearance of endotoxin from plasma and doubled the amount of endotoxin cleared by the liver (30 +/- 1 vs. 14 +/- 2% of the total infused radiolabel, P < 0.001). In addition, autoradiographic studies showed that chylomicrons directed more of the endotoxin to hepatocytes and away from hepatic macrophages. Rats infused with endotoxin plus chylomicrons also showed reduced peak serum levels of tumor necrosis factor as compared with controls (14.2 +/- 3.3 vs. 44.9 +/- 9.5 ng/ml, mean +/- SEM, P = 0.014). In separate experiments, chylomicrons (1,000 mg triglyceride/kg) or saline were infused 10 min before the infusion of endotoxin. Chylomicron pretreatment resulted in a reduced mortality compared with rats infused with endotoxin alone (22 vs. 78%, P < 0.005). Therefore, chylomicrons can protect against endotoxin-induced lethality with and without preincubation with endotoxin. The mechanism by which chylomicrons protect against endotoxin appears to involve the shunting of endotoxin to hepatocytes and away from macrophages, thereby decreasing macrophage activation and the secretion of cytokines.

Chylomicron and chylomicron remnant metabolism in STZ-induced diabetic rats.

The small intestine is an important source of plasma lipoproteins in various diabetic animal models. This increase in intestinally derived lipids originate from diet and/or primary lipid synthesis, and these lipids are transported to the plasma as chylomicrons (CM). The understanding of the metabolism of these triglyceride-rich particles has assumed considerable importance. When [14C]cholesterol and [3H]triglyceride-labeled normal CM were injected into rats, we found no difference in either initial plasma clearance or in the hepatic uptake between control and diabetic rats. However, the clearance rate and hepatic uptake were dependent on the triglyceride concentration administered. Both the initial clearance and hepatic uptake in control and diabetic rats slowed to a similar extent with increasing triglyceride dose demonstrating the influence of the size of the endogenous triglyceride pool on the metabolic rate of CM. No difference was found in the clearance of CM remnants between control and diabetic rats when examined both in vivo and in liver perfusion experiments. Furthermore, with affinity chromatography, we found that the increase in serum triglycerides levels in diabetic rats was due to triglyceride-rich very-low-density lipoproteins and/or CM and not to the accumulation of remnants, which supports the observation that remnant clearance is not impaired. Despite the absence of alterations in bulk CM metabolism, we observed an increase in CM-CM remnant binding to the endothelium in hearts of diabetic rats.(ABSTRACT TRUNCATED AT 250 WORDS)

Structure of plaque at carotid bifurcation: high-resolution MRI with histological correlation.

BACKGROUND AND PURPOSE: The composition of carotid atherosclerosis was visualized by using 3D MRI at high resolution with 200-micrometer (3) voxels. Magnetic resonance signal characteristics were correlated with plaque components, including collagenous cap, necrotic core, and calcification, to define resolution and other requirements for future clinical carotid MRI. METHODS: Twenty-one en bloc carotid endarterectomy specimens were imaged ex vivo by 3D gradient-echo MRI by using a 1.5-T clinical scanner with repetition time, echo time, and flip angle of 40 ms, 18 ms, and 20 degrees, respectively. Plaques were placed in Gd-saline and imaged in a solenoid radiofrequency coil. For quantitative tissue-specific signal analysis, techniques were developed to match tissue sections analyzed by MRI and histology. RESULTS: Three-dimensional imaging resolved complex morphological features not visualized by density- or T(2)-weighted 2D spin-echo imaging. The collagenous cap, necrotic core, and areas of focal calcification showed differing signal characteristics: mean contrast-to-noise ratio for cap versus underlying core was 20. The signal distributions for media and necrotic core overlapped but were resolvable in most specimens. The signal from thrombus was variable. CONCLUSIONS: En bloc specimens provide a useful model for studying plaque MRI. By use of isotropic submillimeter resolution, the collagenous cap and underlying necrotic core typically can be distinguished, and calcification can be identified. Thrombus displays a wide variation in signal intensity. The techniques presented could facilitate future clinicohistological correlation studies for atherosclerotic plaque MRI.

Preoperative balloon-tipped catheter placement for proximal control in visceral artery aneurysm surgery.

A 13 cm false aneurysm of the common hepatic artery developed after repeated episodes of pancreatitis in a 47-year-old man with diabetes who was undergoing chronic hemodialysis. A balloon-tipped catheter was positioned in the common hepatic artery before operation. This maneuver allowed proximal control of the aneurysm and suture closure of the hepatic artery defect with minimal dissection and blood loss in this high-risk patient.

Hypercholesterolemia and early restenosis after carotid endarterectomy.

Cellular proliferation in response to endothelial injury has been examined extensively in experimental animals. Under certain conditions (e.g., hypercholesterolemia and hypertension), this response can be exaggerated and develop into lesions that resemble early atherosclerosis. The injury caused by endarterectomy in human beings and the repair of the arterial wall that ensues may be analogous to the animal models. Presumably, those patients with an exaggerated proliferative response manifest myointimal hyperplasia and recurrent stenosis. To determine potential causes of recurrent stenosis after carotid endarterectomy, we studied 31 patients with early restenosis (group I), 35 patients with later restenosis (group II), and compared them with a control group of 100 consecutive patients who underwent uncomplicated carotid endarterectomy (group III). The known risk factors for atherosclerosis were analyzed. There was no significant difference in the male-to-female ratio, number of cigarettes smoked, or incidence of diabetes mellitus. However, the serum cholesterol level for group I was 282 +/- 57 mg/dl (p less than 0.001 versus controls) while the serum cholesterol level in group II was not significantly elevated over that of the control group. Both groups I and II had a higher incidence of hypertension (p less than 0.005 for both versus controls). There were no differences in the severity of hypertension. The data suggest that hypercholesterolemia has a strong association with early restenosis after carotid endarterectomy but not with late recurrent disease and that hypertension, even when treated, may be associated with both early and late recurrent stenosis.

Comparative effects of hypertension and nicotine on injury-induced myointimal thickening.

Although hypertension has been shown to increase myointimal thickening after arterial injury, a similar effect of nicotine has not been demonstrated. To compare the influence of nicotine with the known effect of hypertension, a standard balloon catheter injury was created in the aortas of four groups of experimental animals. A preliminary group (P) (n = 11) consisted of adult Wistar Kyoto rats used to determine the degree of denudation and the time course of arterial healing; group I (n = 10) consisted of adult Wistar Kyoto rats; group II (n = 13) consisted of spontaneously hypertensive rats; and group III (n = 16) consisted of Wistar Kyoto rats fed nicotine. Endothelial denudation was achieved after an identical injury with an aortic balloon catheter; completeness of denudation was confirmed in group P with Evans blue stain. Injured vessels were fixed by perfusion at 2- and 9-week intervals. Intima to media (I/M) ratios were measured at 82 points per animal by use of light microscopy. Computer-assisted statistical analysis was performed with the t test and Wilcoxon nonpaired rank sum test. Serum nicotine levels were maintained in group III at 73.3 +/- 36.7 ng/ml, which is comparable to levels in human smokers. There were no significant differences in I/M ratios between any of the groups at 2 weeks. At 9 weeks both groups II and III had significantly higher I/M ratios than did control subjects (p = 0.039). The nicotine-treated group had greater myointimal thickening than did the hypertensive animals (p = 0.02). Since there was no hypertension or tachycardia in nicotine-treated rats, we postulate that our data reflects the desquamating effect of nicotine.

Sepsis increases endocytosis of endotoxin into hepatocytes.

BACKGROUND: Chylomicrons bind endotoxins and accelerate their clearance from plasma to the liver. This results in reduced mortality from septic shock in a rodent model. We hypothesized that the clearance of the LPS-chylomicron (LPS-CM) complex by hepatocytes is due to receptor-mediated endocytosis and that sepsis up-regulates this process. METHODS: Three groups of Sprague-Dawley rats; (1) control; (2) pretreated with 10 micrograms/kg LPS 24 hours before treatment; and (3) pretreated with 17-alpha-ethinyl estradiol (EE, 5 mg/kg subcutaneously for 3 days), were infused with labeled I125-LPS alone or with I125-LPS bound to chylomicron. Livers were removed 2.5, 15, and 30 minutes after LPS injection, and hepatic endosomes were isolated from the liver homogenates by serial ultracentrifugation in sucrose gradients. RESULTS: The injection of I125-LPS-CM complexes resulted in higher levels of endosomal I125-LPS in all groups, as compared with I125-LPS alone. In addition, the endosomal uptake of I125-LPS was markedly increased by both LPS and EE pretreatments. CONCLUSIONS: These data strongly suggest a primary role for receptor-mediated endocytosis in the increased clearance of LPS when bound to chylomicron. In addition, exposure to LPS appears to increase the accumulation of LPS in endosomes by a mechanism similar to that of EE, which is known to up-regulate receptor-mediated lipoprotein uptake. This endogenous pathway for the catabolism of endotoxins may provide a teleological explanation for the hypertriglyceridemia observed during sepsis.

"Angiography" by magnetic resonance imaging: detailed vascular anatomy without ionizing radiation or contrast media.

Using software to transform magnetic resonance imaging (MRI) tomographic data into three-dimensional projections, we have produced "angiograms" of the abdominal aorta in 18 patients with occlusive (15) and aneurysmal (3) disease. This information may be displayed to demonstrate flow, cross-sectional diameter, or aortic surface anatomy. To test the accuracy of the MRI studies, we compared them at selected points with the transected aorta and with routine aortograms. Relative cross-sectional diameter estimated by MRI was within 15% of measured aortic diameter in 14 of the 18 vessels and enabled correct detection of thrombus in 16. We conclude that MRI may accurately image the diseased abdominal aortic wall. In addition, an angiographic display format may aid in the interpretation of these studies. Although MRI is not likely to replace aortography, it could be used adjunctively to define aortic wall thickness and the presence of thrombus.

Triglyceride-rich lipoproteins improve survival when given after endotoxin in rats.

BACKGROUND: Triglyceride-rich lipoproteins have been shown to bind bacterial endotoxin and inhibit its activity in vitro and to protect animals from death when administered before a lethal injection of endotoxin. We now demonstrate that triglyceride-rich lipoproteins can neutralize the toxic effects of endotoxin already in circulation. METHODS: Rats were infused with a lethal dose of endotoxin, followed at various time intervals by an infusion of either mesenteric lymph containing nascent chylomicrons (1 gm chylomicron triglyceride/kg) or an equal volume of normal saline solution. Survival was measured at 48 hours. The experiment was then repeated, substituting the synthetic triglyceride-rich lipid emulsion (1 gm/kg) for chylomicrons. We also measured the clearance and tissue distribution of radioiodinated endotoxin in rats treated subsequently with chylomicrons or saline solution. RESULTS: Chylomicron infusions significantly improved survival when given up to 30 minutes after a lethal dose of endotoxin (p < 0.05). Chylomicrons accelerated endotoxin clearance from the blood and increased endotoxin uptake by the liver. The synthetic triglyceride-rich lipid emulsion significantly improved survival when given up to 15 minutes after a lethal dose of endotoxin (p < 0.05). CONCLUSIONS: Triglyceride-rich lipoproteins and synthetic triglyceride-rich lipid emulsions significantly improve survival of rats when given after a lethal dose of endotoxin. Lipoprotein treatment accelerates endotoxin clearance to the liver, which may account for the observed protection. These data suggest a possible therapeutic role for triglyceride-rich lipoproteins or synthetic lipid emulsions in the treatment of the endotoxemia of gram-negative sepsis.

Saphenous vein bypass to pedal arteries. An aggressive strategy for foot salvage.

Patients with forefoot ischemia and severe tibial artery disease present a major challenge to revascularization and foot preservation. Encouraged by the success of saphenous vein bypass to the more proximal arteries of the lower leg, we extended this technique to the pedal arteries. Between February 1986 and September 1988, we performed 26 bypasses in 24 men (mean age, 66 years) with critical foot ischemia. Sixty-three percent of the patients had diabetes mellitus, 50% had hypertension, and 71% were actively smoking. Angiography invariably revealed multiple tibial artery occlusions with reconstitution of the pedal arteries. The foot salvage rate was 83% (mean survival, 14 months), primary patency was 83% (mean survival, 9 months), and survival was 86% (mean, 12 months). There were 11 wound complications (42%); two resulted in disruptions of the distal anastomosis and eventual graft failure. Bypass to the pedal arteries yields a high rate of foot salvage and is comparable with more proximal bypass procedures; however, wound complications are common and require special technical considerations.

Detection of clinically silent infarcts after carotid endarterectomy by use of diffusion-weighted imaging.

BACKGROUND AND PURPOSE: Intraprocedural transcranial Doppler sonography has identified multiple microembolic events during and immediately after carotid endarterectomy (CEA) or angioplasty, yet the rate of clinically evident stroke is small. To determine the significance of the transcranial Doppler sonography findings, we examined patients by use of diffusion-weighted imaging and fluid-attenuated inversion recovery MR imaging before and immediately after CEA for evidence of clinically silent ischemic events. METHODS: Twenty-five patients with atherosclerotic disease of the carotid arteries underwent diffusion-weighted imaging and fluid-attenuated inversion recovery MR imaging performed, on average, 3 days before and 12 hours after CEA. Diffusion-weighted images were acquired in three orthogonal directions at b = 900. Pre- and postoperative neurologic examinations were performed by the same physician. RESULTS: After endarterectomy, 4.0% of the patients (one of 25 patients) showed a single, cortical focus of restricted diffusion and new fluid-attenuated inversion recovery hyperintensity, measuring <1 cm in diameter, ipsilateral to the CEA. The postoperative neurologic examination showed no change in status from the preoperative baseline state. This patient had an intraoperative course complicated by the development of a large luminal thrombus, necessitating thrombectomy. CONCLUSION: The use of diffusion-weighted imaging may serve to improve conspicuity of clinically silent infarcts after CEA. An important next step is to determine the risk factors that predispose to detectable parenchymal ischemic events.

Assessment of carotid artery stenosis by MR angiography: comparison with x-ray angiography and color-coded Doppler ultrasound.

PURPOSE: To compare magnetic resonance angiography (MRA) with duplex Doppler ultrasound (US) and x-ray angiography (XRA) in the evaluation of the carotid bifurcation. METHODS: The carotid arteries of 61 patients were studied using MRA, US, or XRA; 31 of the patients underwent all three examinations. MRA included both 2D and 3D time-of-flight sequences. Internal and external carotid artery origins were graded normal, mild, moderate, severe, or critical stenosis, or complete occlusion by each of the three studies. RESULTS: Spearman rank correlations of both internal and external carotid artery grades were 0.85 (MRA and XRA), 0.69 (MRA and US), and 0.73 (XRA and US). For internal carotid artery origins only, the correlations were 0.94 (MRA and XRA), 0.85 (MRA and US), and 0.82 (XRA and US). Of discrepancies in internal carotid artery interpretation greater than one grade, seven resulted from US error, three from MRA error, and one from XRA error. A 2-cm partially thrombosed aneurysm detected by US and MRA was missed by XRA. Of 16 possible ulcers on XRA, 11 were noted by MRA, none by US. CONCLUSIONS: MRA and XRA are similar in assessment of carotid bifurcation stenosis. MRA, like US, permits direct visualization of plaque. This preliminary study suggests that MRA may be used to clarify equivocal findings of US, or replace XRA in presurgical planning.

Ischemia of the upper extremity: significance of proximal arterial disease.

Thirty-four patients had upper extremity ischemia due to proximal arterial disease, including subclavian compression at the thoracic outlet, innominate or subclavian atherosclerosis, and other causes. Nineteen lesions caused distal embolization. Although successful arterial reconstructions were performed in all but one patient, there were five major amputations. Four were associated with a delay in diagnosis, which allowed the formation of repeated distal emboli. Since overlooked proximal brachiocephalic arterial lesions can produce devastating consequences, we believe these vessels should be examined in all cases of hand and arm ischemia.