Study on tumor microvasculature damage induced by alternate cooling and heating.

Tumor vasculature damage induced by various thermal treatments has been studied in vivo via laser confocal microscopy. Murine mammary carcinoma 4T1 was implanted in the nude mice dorsal skin fold window chamber. The implanted tumor was treated by alternate cooling and heating. Results showed that the treatment was much more effective as compared with that of cooling or heating alone, especially in damaging the tumor vasculature. In general, tumor vascular response to thermal stimuli was heterogeneous. All the treatments of hyperthermia at 42 degrees C (for 1 h), alternate cooling at 1 degrees C and heating at 42 degrees C (for 1/2 h each) and that of -10 degrees C/42 degrees C (for 1/2 h each) enhanced liposome extravasation. Pre-cooling tumor at 1 degrees C preserved most of the vascular integrity but partially inhibited the effect of post-hyperthermia at 42 degrees C. On the other hand, cooling at -10 degrees C for 1/2 h before heating at 42 degrees C caused severe vessel damage. Histo-pathological analyses further confirmed the effect as rare tumor vessel recurrence and large necrotic tumor tissue areas shown on the 7th day after the treatment.

Clinical features, topographic patterns on DWI and etiology of thalamic infarcts.

Thalamic infarcts may lead to diverse neurological disturbances, which easily results in misdiagnosis. Diffusion-weighed magnetic resonance imaging (DWI) is sensitive for the early diagnosis of the infarct and identification of the territory involved. The aim of this study was to analyze the clinical features, topographic appearance on DWI and etiology of thalamic infarcts. We reviewed clinical data, vascular risk factors, topographic patterns and etiology of thalamic infarcts. The patients were divided into 2 groups according to DWI patterns: isolated thalamic infarcts (ISO-TH) and combined thalamic infarcts (COM-TH). The former were further subdivided into 2 subgroups: inferolateral isolated thalamic infarcts (INF-TH) and non-inferolateral isolated thalamic infarcts (NON-INF) according to the vascular territories. The Patients were also divided according to etiology based on TOAST classification. The association of clinical features, DWI patterns and etiology was analyzed. Twenty nine patients were included, among which, 23 (79.3%) were ISO-TH and 6 (20.7%) were COM-TH. The most common territory involved in the ISO-TH was inferolateral territory [n=17 (73.9%)], followed by tuberothalamic artery territory [n=3 (13.0%)], and posterior choroidal artery territory [n=2 (8.7%)]. In COM-TH, the most common territory also was the inferolateral territory (n=3), followed by posterior choroidal artery territory (n=1). In 2 patients, the lesions involved more than one vascular thalamic territory. Significant association between small-vessel occlusion (SVO) and ISO-TH (INF-TH+NON-IFN) infarcts were found. Our study suggested that SVO was more prevalent in ISO-TH, and COM-TH needed more etiological examination. DWI might provide meaningful clues about etiology of thalamic infarcts.

Cold stimulus fingertip lacticemy test--an effective method to monitor acute therapeutic intervention on primary Raynaud's phenomenon and systemic sclerosis.

OBJECTIVES: The recently developed cold stimulus fingertip lacticemy test (CS-FTL) provides biochemical assessment of peripheral perfusion in patients with Raynaud's phenomenon (RP). We evaluated how the CS-FTL test can assess the acute effect of nifedipine in microvascular dynamics on primary RP and RP secondary to SSc. METHODS: A double-blinded controlled trial with crossover design was performed in 20 primary RP and 20 SSc patients. Patients received one single sublingual placebo or 10 mg nifedipine capsule, with crossover after a 15-day washout period. FTL was determined in resting conditions (pre-CS-FTL) and 10 min after CS (post-CS-FTL), before and 1 h after drug administration. Percent variation in post- vs pre-CS-FTL was expressed as deltaCS-FTL. RESULTS: Before intervention, CS induced FTL decrease in primary RP (deltaCS-FTL = -21.3 +/- 13.0%) and FTL increase in SSc patients (deltaCS-FTL = +24.5 +/- 21.2%). Placebo had no effect on pre-CS-FTL, post-CS-FTL and deltaCS-FTL in primary RP and SSc. Nifedipine induced a significant decrease in pre-CS-FTL (1.94 +/- 0.45 vs 1.57 +/- 0.41 mg/dl; P = 0.005) and post-CS-FTL (1.53 +/- 0.35 vs 1.32 +/- 0.37 mg/dl; P = 0.004) in primary RP and a significant decrease in post-CS-FTL (3.18 +/- 1.43 vs 2.56 +/- 1.30 mg/dl; P = 0.028) and deltaCS-FTL (+15.9 +/- 24.7% vs -12.9 +/- 16.6%; P = 0.001) in SSc. CONCLUSIONS: The CS-FTL test was able to demonstrate and quantify a dual effect of nifedipine on the biochemical dimension of peripheral perfusion in primary RP and in SSc patients in which there was a significant improvement in tissue perfusion in resting conditions and after exposure to a CS. The CS-FTL test will enrich the armamentarium for investigation and clinical evaluation of conditions associated with RP.

Intravenous immunoglobulins reverse acute vaso-occlusive crises in sickle cell mice through rapid inhibition of neutrophil adhesion.

Previous studies using intravital microscopy in a sickle cell disease (SCD) mouse model suggest that adherent white blood cells (WBCs) play a key role in vaso-occlusion by capturing circulating red blood cells (RBCs) in venules. Commercial intravenous immunoglobulin (IVIG) given before the inflammatory stimuli increased microcirculatory blood flow and survival. To mimic the clinical situation in which SCD patients seek medical attention after the onset of symptoms, we developed an in vivo model in which the therapeutic intervention (eg, IVIG) was administered after in the inflammatory challenge. In this setting, IVIG rapidly (<10 minutes) reduced adherent leukocyte numbers and dramatically inhibited interactions between RBCs and WBCs, resulting in improved microcirculatory blood flow and survival of sickle cell "Berkeley" mice. Longer survival correlated positively with blood flow (P=.001) and negatively with the number of adherent leukocytes (P=.001) and RBC-WBC interactions (P=.002). Using multichannel digital fluorescence videomicroscopy, we found that IVIG affected specifically the recruitment of neutrophils. Moreover, further analyses of leukocyte behavior revealed that IVIG significantly increased rolling velocities, indicating that it alters adhesion pathways involved in slow rolling. These data suggest that the potential therapeutic benefits of IVIG in SCD crises should be evaluated in a clinical trial.

Angiogenesis in cancer: molecular mechanisms, clinical impact.

BACKGROUND: Angiogenesis, the formation of new blood vessels from the endothelium of the existing vasculature, is fundamental in tumor growth, progression, and metastasis. Inhibiting tumor angiogenesis is a promising strategy for treatment of cancer and has been successfully transferred from preclinical to clinical application in recent years. Whereas conventional therapeutic approaches, e.g. chemotherapy and radiation, are focussing on tumor cells, antiangiogenic therapy is directed against the tumor supplying blood vessels. MATERIALS AND METHODS: This review will summarize important molecular mechanisms of tumor angiogenesis and advances in the design of antiangiogenic drugs. Furthermore, clinical implications of antiangiogenic therapy in surgical oncology will be discussed. RESULTS: First antiangiogenic drugs have been approved for treatment of advanced solid tumors in several countries. Leading antiangiogenic drugs are designed to inhibit vascular endothelial growth factor-mediated tumor angiogenesis. Combining antiangiogenic agents with conventional chemotherapy or radiation is currently investigated clinically with great emphasis to realize a multimodal tumor therapy, targeting both the tumor cell and tumor vascular compartment. CONCLUSION: Antiangiogenic tumor therapy represents a promising strategy for treatment of cancer and will most likely exhibit its clinical potential in combination with established standard tumor therapies in the future.

Fabry disease mimicking multiple sclerosis.

Fabry disease is an X-linked recessive lysosomal storage disorder resulting from the deficiency of alpha-galactosidase. This disease causes endothelial vasculopathy and affects multiple organ systems. Hemizygous male patients represent the classical renal, cardiac and neurological symptoms of disease. Heterozygous female carriers are frequently asymptomatic, but cerebrovascular events in females are as frequent as in males. Even if rarely seen, neurological damage is an important cause of morbidity. Severe neurological signs that are due to multifocal small vessel occlusions may be present without major thrombosis. In this report, we present a 33-year-old female patient with recurrent neurological deficits secondary to multifocal small vessel involvements. The case had previously been misdiagnosed as multiple sclerosis. Cerebral MRI revealed hyperintense lesions located in bilateral thalamus, supratentorial areas, and left cerebellum. Laboratory and radiological investigations were performed for differential diagnosis, but the etiology could not be identified. During follow-up period, skin lesions and proteinuria were detected. The dermatological, neurological, laboratory, and radiological findings were all suggestive of Fabry disease and the diagnosis was confirmed by subsequent enzyme assays. Fabry disease should be considered in young patients with unexplained stroke-like episodes, especially in those who have infarction in the vertebrobasilar arterial system, angiokeratomas, and proteinuria.

Vitamin C supplementation could reverse diabetes-induced endothelial cell dysfunction in mesenteric microcirculation in STZ-rats.

This paper was aimed to study whether vitamin C supplementation reverses the diabetes-induced endothelial cell dysfunction occurred in streptozotocin (STZ)-rats or not. The animals were divided into four groups: control and diabetes rats (DM, using iv. injection of 50 mg/kg.bw STZ), and two DM rats treated with vitamin C (1 g/l) starting on day 2 (DM + VitC(day2)) and week 6th after STZ-injection (DM + VitC(6wks)). The mesenteric microcirculation was observed using fluorescence videomicroscopy. Based on the recorded videoimages, microvascular responses to acetylcholine (Ach; 10-5 M) and number densities of leukocyte adhesion in venules were evaluated using the Global Lab II image software. In DM group, blood glucose and glycosylated hemoglobin were significantly increased, while the body weight and plasma vitamin C levels were decreased significantly compared to their controls. Ach-induced vasodilation was decreased, while the number of leukocyte adhesion was increased significantly compared to their controls (p<0.01). These abnormalities induced by DM were prevented by supplementation of vitamin C in DM + VitC(day2) group. Six-weeks delayed treatment of vitamin C (DM + VitC(6wks)) demonstrated increase in the Ach-induced vasodilation with significant decrease in the leukocyte adhesion. It was indicated that vitamin C supplementation could reverse diabetes-induced endothelial cell dysfunction in mesenteric microcirculation.

Effect of Jianweiyuyang granule on gastric ulcer recurrence and expression of VEGF mRNA in the healing process of gastric ulcer in rats.

AIM: To investigate the effect of Jianweiyuyang (JWYY) granule on gastric ulcer recurrence and its mechanism in the treatment of gastric ulcer in rats. METHODS: Gastric ulcer in rats was induced according to Okeba's method with minor modification and the recurrence model was induced by IL-1beta. The expression of vascular endothelial growth factor mRNA (VEGF mRNA) was examined by reverse transcription polymerase chain reaction in gastric ulcer and microvessel density (MVD) adjacent to the ulcer margin was examined by immunohistochemistry. RESULTS: MVD was higher in the JWYY treatment group (14.0+/-2.62) compared with the normal, model and ranitidine treatment groups (2.2+/-0.84, 8.8+/-0.97, 10.4+/-0.97) in rats (P<0.01). The expression level of VEGF mRNA in gastric tissues during the healing process of JWYY treatment group rats significantly increased compared with other groups (normal group: 0.190+/-0.019, model group: 0.642+/-0.034, ranitidine group: 0.790+/-0.037, P<0.01). CONCLUSION: JWYY granules can stimulate angiogenesis and enhance the expression of VEGF mRNA in gastric ulcer rats. This might be the mechanism for JWYY accelerating the ulcer healing, and preventing the recurrence of gastric ulcer.

Preventing and treating actions of compound lian zhu capsule on micrangium lesions in diabetic rats.

Tissue culture, biochemical techniques and radioimmunoassay were used to study the effects of Compound Lian Zhu Capsule on micrangium lesions in diabetic rats. The results indicated that blood sugar, glycosylated hemoglobin (GHb), urinary protein and malondialdehyde (MDA) contents, aldose reductase (AR) activity and 3H-TdR incorporation rate in the vascular smooth muscle cell (VSMC) were significantly higher, and plasma NO content in the diabetes mellitus (DM) group were significantly lower than those in the normal control group (both P < 0.05). The above indexes in the Chinese medicine (TCM) treatment group were improved significantly as compared with the DM group, with no significantly differences, except urine volume and urinary protein, as compared with the normal control group. It is suggested that Compound Lian Zhu Capsules cansignificantly alleviate the complicated lesions of the micrangium in diabetic rats.

Hyperbaric oxygen reduces blood-brain barrier damage and edema after transient focal cerebral ischemia.

BACKGROUND AND PURPOSE: Hyperbaric oxygen (HBO) has been shown to protect the brain parenchyma against transient focal cerebral ischemia, but its effects on the ischemic microcirculation are largely unknown. We examined the potential of HBO to reduce postischemic blood-brain barrier (BBB) damage and edema. METHODS: Wistar rats and C57/BL6 mice underwent occlusion of the middle cerebral artery (MCAO) for 2 hours. Forty minutes after filament introduction, animals breathed either 100% O2 at 3.0 atmospheres absolute (ata; HBO group) or at 1.0 ata (control) for 1 hour in an HBO chamber. In rats, MRI was performed 15 minutes after MCAO and after 15 minutes and 3, 6, 24, and 72 hours of reperfusion. In mice, BBB permeability for sodium fluorescein was measured after 24-hour reperfusion. RESULTS: Increased BBB permeability on postcontrast T1-weighted (T1w) images had a biphasic pattern. HBO reduced volumes and intensity of enhancement. Mean abnormal enhancing volumes were 71+/-10 mm3 (control) versus 47+/-10 mm3 (HBO) at 15 minutes; 111+/-21 mm3 versus 69+/-17 mm3 3 hours; 147+/-44 mm3 versus 83+/-21 mm3 6 hours; 150+/-37 mm3 versus 89+/-14 mm3 24 hours; and 322+/-52 mm3 versus 215+/-21 mm3 72 hours (all P<0.05). Interhemispheric quotients of mean gray values on T1w were at 1.73+/-0.11 versus 1.57+/-0.07 15 minutes; 1.74+/-0.07 versus 1.60+/-0.06 at 3 hours; 1.77+/-0.07 versus 1.62+/-0.06 at 6 hours; 1.79+/-0.10 versus 1.60+/-0.05 at 24 hours; and 1.81+/-0.10 versus 1.62+/-0.07 at 72 hours (all P<0.05). HBO-treated mice had significantly lower postischemic BBB permeability than mice treated with either normobaric hyperoxia or room air. Vasogenic edema assessed on T2w images and histologic sections was significantly lower in HBO-treated rats. CONCLUSIONS: Intraischemic HBO therapy reduces early and delayed postischemic BBB damage and edema after focal ischemia in rats and mice.

Tumor angiogenesis and dynamic CT in colorectal carcinoma: radiologic-pathologic correlation.

AIM: To investigate the correlation between microvessel density and spiral CT perfusion imaging in colorectal carcinoma. METHODS: Thirty-seven patients, with histologically proven colorectal carcinoma, underwent water enema spiral CT scan. The largest axial surface of the primary tumor was searched on unenhanced spiral CT images. At this level, the enhanced dynamic scan series was acquired. Time-density curves (TDC) were created from the region of interest drawn over the tumor, target artery by Toshiba Xpress/SX spiral CT with perfusion functional software. Then the perfusion was calculated. Microvessel density (MVD) was evaluated using immunohistochemical staining of surgical specimens with anti-CD34, and then MVD was correlated with perfusion. RESULTS: MVD of colorectal carcinomas was 33.11-173.44, mean 87.28, and perfusion was 15.60-64.80 mL/min/100 g, mean 39.74 mL/min/100 g. MVD and perfusion were not associated with invasive depth, metastasis and disease stage, and they all decreased with increasing Dukes' stage, but no significant correlation was found between them (r = 0.18, P = 0.29). CONCLUSION: There is no significant correlation between MVD and perfusion. Neovascularizaton and perfusion are highly presented in early colorectal carcinoma. CT perfusion imaging may be more suited for assessing tumorigenesis in colorectal carcinoma than histological MVD technique.

Folic acid supplementation can reduce the endothelial damage in rat brain microvasculature due to hyperhomocysteinemia.

To evaluate the effects of dietary folic acid supplementation on the cerebral vascular damage induced by hyperhomocysteinemia, rats were fed a diet containing 3.0 g/kg homocystine for 2 wk and then either 3.0 g/kg homocystine or 3.0 g/kg homocystine plus 0.008 g/kg folic acid for 8 wk. Control rats consumed the AIN-93 Maintenance diet throughout the experiment. The cerebral expression of glucose transporter-1 was measured by Western blot analysis and cerebrovascular structural alterations were evaluated by electron microscopy. The homocystine diet significantly increased the plasma levels of homocysteine and TBARS and decreased the cerebral expression of glucose transporter-1 (GLUT-1) with a concomitant increase in the percentage of damaged cerebral vessels. The inclusion of dietary folic acid for 8 wk caused plasma homocysteine levels to be the same as in control rats and it significantly upregulated the cerebral expression of GLUT-1 that was significantly reduced by hyperhomocysteinemia. Folic acid supplementation also significantly decreased the incidence of damaged vessels due to hyperhomocysteinemia. These results and the electron microscopy findings suggested that folic acid supplementation might reduce the detrimental effects on the endothelium caused by experimentally induced hyperhomocysteinemia.

[Taohong Siwu decoction II inhibits the angiogenesis and the expressions of VEGF and KDR/FLK-1 in C57BL/6J mice bearing B16 melanoma].

OBJECTIVE: To investigate the effects of Taohong Siwu decoction II on B16 melanoma in mice and the underlying mechanism. METHOD: C57BL/6J mice bearing B16 melanoma were used in this study. The experimental groups were treated respectively with Taohong Siwu decoction II at doses of 2.5, 5 and 10 g x kg(-1) and cyclophosphamide at 0.05 g x kg(-1), Taohong Siwu decoction II at 10 g x kg(-1) plus cyclophosphamide at 0.025 mg x kg(-1). The tumor volume and weight, the expression of VEGF and KDR/FLK-1 and the amount of MVD were measured. RESULT: The tumor volume and weight, the expression of VEGF and KDR/FLK-1 and the amount of MVD were reduced significantly after treatment with Taohong Siwu decoction II at the doses of 5 and 10 g x kg(-1) and Taohong Siwu decoction II combined with cyclophosphamide as compared with normal saline-treated group (P < 0.01). CONCLUSION: Taohong Siwu decoction II can inhibit the growth of B16 melanoma in mice and attenuate the expressions of VEGF and KDR/FLK-1, suggesting that Taohong Siwu decoction II produces the antitumor effect via a possible antiagiogenisis mechanism.

Combined photodynamic and photothermal induced injury enhances damage to in vivo model blood vessels.

BACKGROUND AND OBJECTIVES: The degree of port wine stain (PWS) blanching following pulsed dye laser (PDL) therapy remains variable and unpredictable. Because of the limitations of current PDL therapy, alternative treatment approaches should be explored. The objective was to evaluate a novel methodology for selective vascular damage, combined photodynamic (PDT) and photothermal (PDL) treatment, using the in vivo chick chorioallantoic membrane (CAM) model. STUDY DESIGN/MATERIALS AND METHODS: Thirty microliters of benzoporphyrin derivative monoacid ring A (BPD) solution was administered intraperitoneally into chick embryos at day 12 of development. Study groups were: (1) control (no BPD, no light); (2) BPD alone; (3) continuous wave irradiation (CW) alone (576 nm, 60 mW/cm2, 125 seconds); (4) CW + PDL; (5) BPD+PDL; (6) PDT (BPD+CW); (7) PDL alone (585 nm, 4 J/cm(2)); and (8) PDT+PDL (BPD + CW followed immediately by PDL). Vessels were videotaped prior to, and at 1 hour post-intervention and then assessed for damage based on the following scale: 0, no damage; 1, coagulation; 1.5, vasoconstriction; 2.0, coagulation+vasoconstriction; 2.5, angiostasis; 3.0, hemorrhage. Damage scores were weighted by vessel "order." RESULTS: PDT + PDL resulted in significantly (P < 0.01) more severe vascular damage than was observed in any other study group: 127% more than PDT, 47% more than PDL alone. CONCLUSIONS: PDT + PDL is a novel and promising approach for selective vascular damage and may offer a more effective method for treatment of PWS and other vascular skin lesions.

[Histological investigation of the micromorphological effects of the application of a laser needle--results of an animal experiment]. / Histologische Untersuchungen zu mikromorphologischen Einflüssen von Lasernadelstrahlung--Ergebnisse einer tierexperimentellen Untersuchung.

In an experimental animal study (Sus scrofa domesticus) we investigated the effects of the new technique of laser needle stimulation (wavelength: 685 nm; energy density: 4.6 kJ/cm2 per point; application duration: 20 min). The results revealed changes in microcirculatory parameters of the skin resulting in an increase in blood flow. However, the quality and intensity of the laser light did not induce micromorphological alterations in the skin.

SU5416 selectively impairs angiogenesis to induce prostate cancer-specific apoptosis.

We have previously demonstrated the differential expression in tumor-associated blood vessels of two vascular endothelial growth factor receptors (VEGFRs), VEGFR1 and VEGFR2, during initiation and progression of prostate cancer in the genetically engineered transgenic adenocarcinoma of the mouse prostate (TRAMP) mouse model. In our "progression switch" model, expression of VEGFR1 is associated with early and more differentiated disease, whereas expression of VEGFR2 is associated with advanced and more poorly differentiated disease. To test the hypothesis that stage-specific inhibition of vascular endothelial growth factor signaling could be used as therapy for autochthonous prostate cancer, we initiated a preclinical trial with SU5416, a potent antiangiogenic small molecule inhibitor of VEGFR associated tyrosine kinase activity. In our early intervention trial, administration of SU5416 to TRAMP mice did not appear to influence angiogenesis or tumor progression between 10 and 16 weeks of age, a time corresponding to high levels of VEGFR1 expression. In our late intervention trial, however, we observed a significant decrease in tumor-associated mean vessel density, increased apoptotic index, and pronounced regions of cell death when SU5416 was administered to TRAMP mice between 16 and 22 weeks of age, a time corresponding to high levels of VEGFR2 expression. These results clearly demonstrate that therapy directed specifically against the VEGFR signaling axis can dramatically impair angiogenesis and induce apoptosis of autochthonous spontaneous and progressive prostate cancer.

Augmentation of endothelial cell monolayer permeability by hyperthermia but not tumor necrosis factor: evidence for disruption of vascular integrity via VE-cadherin down-regulation.

Following isolated limb perfusion (ILP) with hyperthermia (H T), TNF and melphalan, there is immediate tumor softening secondary to augmentation of capillary leak in the tumor neovasculature. TNF can induce vascular permeability but is always used with HT during ILP and the contribution of the latter on permeability is not known. This study characterizes the effects of HT and TNF on vascular permeability in vitro. Permeability across confluent human umbilical vein endothelial cells exposed to HT (40 degrees C) with or without 0.1-1000 ng/ml TNF was assessed by quantitating flux of albumin bound Evan's Blue dye from the upper to lower chamber. Immunofluorescent staining for VE-cadherin and F-actin was performed after human umbilical vein endothelial cells (hUVECs) were exposed to these conditions. HT induced a significant and reversible increase in permeability compared to untreated hUVECs (p<0.001) whereas barrier function was not altered by TNF. Untreated hUVECs had uniform cell surface staining for VE-cadherin, the primary endothelial intercellular adhesion molecule, with colocalization of F-actin cytoskeletal elements. HT resulted in a marked decrease in VE-cadherin staining and contraction of F-actin at sites of endothelial cell-cell separation. These data demonstrate that under conditions relevant to those used in ILP, HT but not TNF contributes to a rapid and reversible change in endothelial cell permeability in association with a down regulation of VE-cadherin. These data support the use of HT in isolation perfusion and demonstrate a novel mechanism for alterations in microvascular permeability by HT.

Mechanisms of endotoxin tolerance in human intestinal microvascular endothelial cells.

Lipopolysaccharide (endotoxin) tolerance is well described in monocytes and macrophages, but is less well characterized in endothelial cells. Because intestinal microvascular endothelial cells exhibit a strong immune response to LPS challenge and play a critical regulatory role in gut inflammation, we sought to characterize the activation response of these cells to repeated LPS exposure. Primary cultures of human intestinal microvascular endothelial cells (HIMEC) were stimulated with LPS over 6-60 h and activation was assessed using U937 leukocyte adhesion, expression of E-selectin, ICAM-1, VCAM-1, IL-6, IL-8, manganese superoxide dismutase, HLA-DR, and CD86. Effect of repeat LPS stimulation on HIMEC NF-kappaB and mitogen-activated protein kinase (MAPK) activation, generation of superoxide anion, and Toll-like receptor 4 expression was characterized. LPS pretreatment of HIMEC for 24-48 h significantly decreased leukocyte adhesion after subsequent LPS stimulation. LPS pretreatment inhibited expression of E-selectin, VCAM-1, IL-6, and CD86, while ICAM-1, IL-8, and HLA-DR were not altered. Manganese superoxide dismutase expression increased with repeated LPS stimulation, with a reduction in intracellular superoxide. NF-kappaB activation was transiently inhibited by LPS pretreatment for 6 h, but not at later time points. In contrast, p44/42 MAPK, p38 MAPK, and c-Jun N-terminal kinase activation demonstrated inhibition by LPS pretreatment 24 or 48 h prior. Toll-like receptor 4 expression on HIMEC was not altered by LPS. HIMEC exhibit endotoxin tolerance after repeat LPS exposure in vitro, characterized by diminished activation and intracellular superoxide anion concentration, and reduced leukocyte adhesion. HIMEC possess specific mechanisms of immunoregulatory hyporesponsiveness to repeated LPS exposure.