Traumatic brain injury and bone healing: radiographic and biomechanical analyses of bone formation and stability in a combined murine trauma model.

INTRODUCTION: The combination of traumatic brain injury (TBI) and long-bone fractures has previously been reported to lead to exuberant callus formation. The aim of this experimental study was to radiographically and biomechanically study the effect of TBI on bone healing in a mouse model. MATERIALS AND METHODS: 138 female C57/Black6N mice were assigned to four groups (fracture (Fx) / TBI / combined trauma (Fx/TBI) / controls). Femoral osteotomy and TBI served as variables: osteotomies were stabilized with external fixators, TBI was induced with controlled cortical impact injury. During an observation period of four weeks, in vivo micro-CT scans of femora were performed on a weekly basis. Biomechanical testing of femora was performed ex vivo. RESULTS: The combined-trauma group showed increased bone volume, higher mineral density, and a higher rate of gap bridging compared to the fracture group. The combined-trauma group showed increased torsional strength at four weeks. DISCUSSION: TBI results in an increased formation of callus and mineral density compared to normal bone healing in mice. This fact combined with a tendency towards accelerated gap bridging leads to increased torsional strength. The present study underscores the empirical clinical evidence that TBI stimulates bone healing. Identification of underlying pathways could lead to new strategies for bone-stimulating approaches in fracture care.

Versatile attosecond beamline in a two-foci configuration for simultaneous time-resolved measurements.

We present our attoline which is a versatile attosecond beamline at the Ultrafast Laser Physics Group at ETH Zurich for attosecond spectroscopy in a variety of targets. High-harmonic generation (HHG) in noble gases with an infrared (IR) driving field is employed to generate pulses in the extreme ultraviolet (XUV) spectral regime for XUV-IR cross-correlation measurements. The IR pulse driving the HHG and the pulse involved in the measurements are used in a non-collinear set-up that gives independent access to the different beams. Single attosecond pulses are generated with the polarization gating technique and temporally characterized with attosecond streaking. This attoline contains two target chambers that can be operated simultaneously. A toroidal mirror relay-images the focus from the first chamber into the second one. In the first interaction region a dedicated double-target allows for a simple change between photoelectron/photoion measurements with a time-of-flight spectrometer and transient absorption experiments. Any end station can occupy the second interaction chamber. A surface analysis chamber containing a hemispherical electron analyzer was employed to demonstrate successful operation. Simultaneous RABBITT measurements in two argon jets were recorded for this purpose.

[Management of complications following resection and defect reconstruction of tumors near the hip joint]. / Komplikationsmanagement nach Resektion und Defektrekonstruktion hüftgelenknaher Tumoren.

BACKGROUND: Tumorous destruction of the periacetabular region and the proximal femur are a consequence of either primary malignant bone tumor manifestation or metastatic disease, which is observed much more frequently and occurs typically in these skeletal segments. Pathological fractures of the proximal femur and periacetabular regions of the pelvis have a high incidence and ultimately lead to severe pain and immobilization. TREATMENT METHODS: Advanced resection techniques and different types of defect reconstruction, allowing for oncologically sufficient resection of extensive tumors have contributed to a marked increase in the limb salvage rate. However, these procedures are associated with an increasing rate of several, sometimes severe intraoperative and postoperative complications. COMPLICATIONS: Compared to elective total hip arthroplasty, the rate of postoperative deep infections, dislocations, the incidence of pathological and periprosthetic fractures and the prevalence of deep vein thrombosis are increased with high rates of postoperative mortality and local tumor recurrence, being the most serious complications. Pelvic involvement and subsequent periacetabular resection have the highest complication rate when compared to proximal femur resection with endoprosthetic treatment. CONCLUSION: In order to minimize the risk of these intraoperative and postoperative complications wide resection and advanced reconstruction as well as complicated palliative stabilization due to malignant bone tumor growth around the hip joint should be performed in musculoskeletal tumor centers with profound expertise in osteosynthetic and endoprosthetic reconstruction of the pelvis and the proximal femur. Only in specialized centers an effective, multidisciplinary emergency management of these complications and, more importantly, reliable prevention of complications can be ensured.

Smoking is associated with impaired long-term glucose metabolism in patients with type 1 diabetes mellitus.

BACKGROUND AND AIMS: Smoking is known to negatively influence glucose metabolism both in healthy subjects and in patients with diabetes. The aim of this study was to compare glycemic control in patients with type 1 diabetes mellitus who were smokers with those who did not smoke during a prospective long-term follow-up. METHODS AND RESULTS: In a single center, 763 patients with type 1 diabetes mellitus were included, 160 (21.0%) of them were smokers. Patients were treated with intensive insulin therapy according to existing guidelines. Glucose control was monitored quarterly, diabetes related complications and cardiovascular risk factors were assessed at least once a year. Glucose control in smokers was significantly worse than in non-smokers at baseline and during follow-up (mean HbA1c during 5047 patient-years of follow-up 7.9 ± 1.3% in smokers and 7.3 ± 1.1% in non-smokers, p < 0.001) despite a higher insulin dosage in smokers (0.71 ± 0.30 U/kg vs. 0.65 ± 0.31 U/kg in non-smokers, p = 0.046). HDL cholesterol was lower in smokers at baseline (1.53 ± 0.45 vs. 1.68 ± 0.51 in non-smokers, p = 0.048). Diabetes related complications tended to occur with a higher frequency in smokers, with a significant difference in macroalbuminuria (9.8% vs. 4.8% in non-smokers, p = 0.047). CONCLUSION: Smoking is associated with worse glucose control in patients with type 1 diabetes mellitus despite the same treatment strategies as in non-smokers. Hyperglycemia, therefore, may contribute to an earlier incidence of diabetes related complications in these patients, in addition to direct toxic effects of smoking.

Vitamin E prevents extensive lipid peroxidation in patients with hypertension.

Oxidative modification of low-density lipoprotein (LDL) increases atherogenic potential to induce the accumulation of lipids and cells in the vascular wall. Previous studies reveal that hypertensive patients have a higher susceptibility to LDL oxidation. As animal models indicate that vitamin E protects LDL from oxidation, here we study the influence of vitamin E on the resistance of LDL to oxidation (lag time) in 47 subjects (31 normotensive, 16 hypertensive) before and after oral administration of vitamin E (400 IE) daily for two months. LDL was isolated and oxidised by incubation with copper ions. The time course of oxidation was measured by continuous photometric monitoring of diene formation at 234 nm. At the beginning of this study, normotensive subjects showed a lag time of 108 +/- 26 minutes and hypertensive patients a lag time of 85 +/- 24 minutes (P<0.05). Vitamin E caused a significant increase in the lag time in both groups: normotensive subjects 128 +/- 33, hypertensives patients 114 +/- 27 minutes (P<0.01). At completion of the study, lag times in both groups were similar (P=not significant). The data presented here suggests that vitamin E protects against the increased risk of vascular disease in patients with hypertension by reducing the susceptibility to oxidative modification of LDL. Vitamin E may therefore act as an inhibitor of atherogenesis.

LDL-oxidation and -subgroups in normo- and hypertensive patients with and without a family history of hypertension.

OBJECTIVES: Oxidative modification of low-density-lipoprotein (LDL) increases its atherogenic potential to induce the accumulation of lipids and cells in the vascular wall. Patients have different lipoprotein profiles according to their LDL-subgroup pattern. The subgroup of LDL, which is most susceptible to oxidation, is most likely the dense LDL3 subfraction. In order to study an assumed association between hypertension, LDL subgroup distribution and the susceptibility of LDL to oxidation, 14 normotensive patients without family histories of hypertension (NT), 13 normotensive patients with family histories of hypertension (NT-FH), 10 hypertensive patients without family histories of (HT) and 11 hypertensive patients with family histories of hypertension (HT-FH) were evaluated. PATIENTS AND METHODS: LDL was oxidatively modified by incubation with copper ions (1.6 microM/L). The course of LDL-oxidation was measured in vitro by continuous photometric monitoring and the quantitative distribution of 3 LDL-subgroups by capillary isotachophoresis (ITP). RESULTS: The lag-phases of NT-FH and hypertensive patients were shorter than those of the control group (NT: 116 +/- 36 minutes; NT-FH 92 +/- 32 minutes, p < 0.05; HT: 95 +/- 41 minutes; HT-FH: 76 +/- 33 minutes, p < 0.05). Compared to NT a significant difference in the relative preponderance of LDL3 subgroup was observed for HT-FH (23.5 +/- 4.6% versus NT: 19.3 +/- 6.6%), additionally, statistical analysis showed a similar trend amongst the other patient groups (NT-FH: 20.4 +/- 7.4%, HT: 21.4 +/- 4.6%). CONCLUSIONS: The increased occurrence of the LDL3 subgroup might contribute to a higher susceptibility to LDL oxidation and therefore create an increased risk of vascular disease in the genotypic and phenotypic hypertensive patient population.

[Homocysteine: a cardiovascular risk factor?]. / Homozystein: Ein kardiovaskulärer Risikofaktor?

Elevated plasma homocysteine levels may lead to an increased risk for atherosclerosis. Besides genetic factors a deficiency of folate, vitamin B6 and/or vitamin B12 may lead to an increase in the plasma concentration of this sulfur containing amino acid. Homocysteine may enhance by several direct and/or indirect mechanisms the pathogenesis of atherosclerosis. In this review selected aspects of homocysteine in relation to clinical practice will be discussed.

Is there a role for the ob gene product leptin in essential hypertension?

In this study we wanted to evaluate the relationship between the ob gene product leptin and blood pressure, as well as plasma renin activity and plasma aldosterone levels. We studied 139 subjects with a mean+/-SD age of 50 +/-14 years and a body mass index of 26.5+/-5.3 kg/m2; 110 subjects had essential hypertension and 29 were healthy nonhypertensive controls. Blood pressure was measured in resting conditions in the morning and blood was drawn for the determination of the plasma renin activity, aldosterone, and leptin levels. The mean blood pressure of the population was 155/97 mm Hg. The relationship between these parameters was studied by univariate regression analysis according to gender and, whenever indicated, adjusted for age and body mass. The mean+/-SEM plasma leptin level in the whole population was 9.5+/-0.6 ng/mL (range, 1.1-43.3). Subjects with stage I hypertension had significantly higher plasma leptin levels than normotensive subjects. Systolic blood pressure correlated with the plasma leptin levels and the leptin levels adjusted for body weight in women (r = 0.422, P < .01) and nonhypertensive men (r = 0.644, P = .03) only. Plasma renin activity (r = 0.329, P = .03) and aldosterone levels (r = 0.342, P = .026) correlated with the leptin concentration. A significant relationship between the peripheral expression of the ob gene product leptin and systolic blood pressure was found in women and nonhypertensive men. In view of the multiple functions of leptin a causal relationship is postulated and potential mechanisms may involve modulatory effects of leptin on neuropeptide Y, angiotensinogen gene expression, the modulation of the autonomous nervous system, or effects on the pituitary adrenal axis. Direct relationships between both plasma renin activity and aldosterone levels and leptin support the potential importance of the relationship between leptin and blood pressure. Our observation may be of future importance for the understanding of the link between the increase in blood pressure and increasing body weight.

Sperm allocation in the simultaneously hermaphroditic land snail Arianta arbustorum.

To test the idea that individuals of the simultaneously hermaphroditic land snail Arianta arbustorum can control the number of spermatozoa in their spermatophores, we investigated whether they differentially release sperm to virgin or nonvirgin partners with respect to the potential risk of sperm competition in a given mating. The number of sperm transferred ranged from 802 620 to 3 968 800 (X= 2 185 100; N=91), but was related neither to the mating history of the partner nor to copulation duration. This indicates that individuals of A. arbustorum are not able to adjust sperm expenditure to the mating history of the partner. Furthermore, the number of sperm transferred was correlated neither with the size of the donor nor with the size of the recipient. It has been proposed that the sexual conflict between the two genders in simultaneous hermaphrodites could be resolved by gamete trading. Theory predicts that sperm trading should occur in hermaphrodites in which the female role controls fertilization, for example in gastropods with a gametolytic gland and/or sperm storage such as A. arbustorum. To see whether sperm trading occurs, we also examined whether individuals of A. arbustorum adjust the number of sperm they release to the number they receive from their mating partner. There was a high degree of reciprocity in spermatophore transfer: in 45 of the 46 mating pairs investigated both partners delivered a spermatophore that contained spermatozoa. The numbers of sperm transferred by the two mating partners were not correlated, however. This indicates that sperm trading does not occur in this simultaneously hermaphroditic land snail. Copyright 1998 The Association for the Study of Animal Behaviour.

Can low density lipoprotein influence microvascular caliber?

The intracellular free calcium concentration ([Ca2+]i) plays an important role in the regulation of vascular tone. In vascular smooth muscle cells (VSMCs) LDL causes changes in vascular tone by increasing [Ca2+]i. Pericytes are regarded as the microvascular counterpart of VSMCs and implicated in the regulation of microvascular cell biology under normal and pathological conditions (e.g., diabetes mellitus, arterial hypertension, arteriosclerosis). For this reason pericytes and VSMCs were compared in their ability to increase [Ca2+]i after stimulation with LDL. Single VSMCs and pericytes were loaded with 2 microM of the Ca2+-sensitive dye Indo-1/AM. Fluorescence was recorded at 405 nm (Ca2+-bound) and 485 nm (Ca2+-free). Cells in suspension were loaded with 2 microM of the calcium ionophore FURA-2 AM (excitation wavelengths: 340 and 380 nm, emission 505 nm). Basal [Ca2+]i levels were significantly higher in single pericytes (165 +/- 38 nmol/L, n = 41) than in VSMCs (150 +/- 39 nmol/L, n = 40, P = 0.0038). In cell suspensions the following values were obtained: Pericytes (113 +/- 27 nmol/L, n = 36) and VSMCs (109 +/- 26 nmol/L, n = 28), which are statistically not significant. For all concentrations of LDL used (except at 1 microg/ml n-LDL), the increase above basal values was significant and both cell types showed a clear dose-dependent reaction pattern. This study shows for the first time that pericytes and VSMCs increase their [Ca2+]i in a similar way after LDL stimulation. In analogy to aortic smooth muscle cells, our results indicate that LDL mediated [Ca2+]i changes in pericytes in the microvascular bed may cause vasoconstriction leading to impairment of blood flow in the microvasculature.

Single pericytes and pericytes in suspension are stimulated in a similar way by low-density lipoprotein.

BACKGROUND: Pericytes are regarded as the microvascular counterpart of smooth muscle cells and implicated in the regulation of blood pressure at the microvascular level. Ca2+ plays an important role in biochemical processes involved in blood pressure regulation and can be activated by low-density lipoprotein (LDL) cholesterol. OBJECTIVE: To determine whether stimulation either of single cells or of cells in suspension by LDL would produce any difference in the increase in cytosolic free calcium levels ([Ca2+]i). DESIGN AND METHODS: Single pericytes were loaded with 2 micromol/l of the Ca2+-sensitive dye Indo-1/AM. The Indo-1 fluorescence was recorded at 405 nm (Ca2+-bound) and 485 nm (Ca2+-free) after stimulation with LDL. Pericytes in suspension were loaded with 2 micromol/l of the Ca2+-sensitive dye FURA-2/AM. The FURA-2 fluorescence kinetics were recorded at 340-380 nm. Ratios of fluorescence at the two wavelengths were transformed to [Ca2+]i. RESULTS: Basal [Ca2+] levels appeared to be higher in single cells (148+/-13 nmol/l, n = 20) than they were in cells in suspension (128+/-8 nmol/l, n = 25; P= 0.0078). After stimulation with LDL the increase in [Ca2+]i in both systems was about 220% above baseline. A clear dose dependency was seen for both systems. CONCLUSIONS: Single pericytes and pericytes in suspension increase their [Ca2+]i after stimulation with LDL dose-dependently. Even though single-cell measurements revealed some technical limitations, their responses were comparable to those obtained in a cell suspension. In analogy to aortic smooth muscle cells, our results indicate that LDL might also play a blood-pressure-regulatory role in the microvasculature.

Inhibitory action of silibinin on low density lipoprotein oxidation.

Low density lipoprotein (LDL) oxidation and smooth muscle cell growth represent key events in atherogenesis. Any mean to reduce these two phenomena may decrease the risk of coronary artery disease and atherosclerosis in general. The effects of silibinin (CAS 22888-70-6) on LDL oxidation and proliferation of vascular smooth muscle cells were evaluated in vitro. Silibinin (50-200 mumol/l) prolonged the lag times of both LDL autooxidation and oxidation by copper by > 50%, as assessed by recordings of diene formation. However, silibinin (up to 500 mumol/l) did not interfere with LDL-stimulated radiolabeled thymidine incorporation. These findings indicate that silibinin, apart from its hepatoprotective effects, has inhibitory properties on LDL oxidation in vitro. Therefore silibinin might represent a novel tool in the prevention and therapy of atherosclerosis.

Ethanol suppresses smooth muscle cell proliferation in the postprandial state: a new antiatherosclerotic mechanism of ethanol?

Epidemiologic studies suggest that moderate amounts of ethanol may reduce cardiovascular risk. The mechanisms of the alcohol-associated risk reduction are not known exactly. Vascular smooth muscle cell proliferation represents an important phenomenon in the pathogenesis of atherosclerosis. Recently, it was suggested that metabolic changes during the postprandial phase may be important in the pathogenesis of atherosclerosis. Therefore, we evaluated the effect of postprandial plasma with and without ethanol on the proliferation of rat vascular smooth muscle cells. Identical meals containing 1 g fat/kg body wt were given with and without ethanol (38 +/- 0.5 g) to eight healthy young men. Blood was drawn hourly during an 8-h postprandial period; the plasma was separated and added to the cell cultures (0.3%, by vol). The proliferative response (DNA synthesis) of these cells was assessed by measuring the incorporation of [methyl-3H]thymidine. The maximal blood ethanol concentration of 11.5 +/- 0.6 mmol/L (mean +/- SEM) was attained within the first hour. The ingestion of the meal with ethanol led to a 20% reduction in the capacity of postprandial plasma to induce thymidine incorporation into smooth muscle cells compared with the meal without ethanol (P < 0.05). These results suggest that ethanol may reduce cardiovascular risk by modulating vascular muscle cell growth during the postprandial period. Considering the amount of time humans spend in the postprandial state during their lifetimes, these findings may be of great importance in the pathogenesis of atherosclerosis.

Low-density lipoprotein-induced formation of nitric oxide by cultured vascular smooth muscle cells of the rat.

There is evidence that low-density lipoprotein (LDL) plays a crucial role in atherogenesis. On the cellular level, LDL has been shown to activate a number of mechanisms involved in atherogenesis and vasoconstriction. Local immoderate vasoconstriction is physiologically antagonized by nitric oxide, which is released from the endothelium. To find out whether LDL also influences the synthesis of nitric oxide in vascular smooth muscle cells, both the conversion of arginine to citrulline and the production of nitrite were determined as a measure of nitric oxide formation. After incubation of rat vascular smooth muscle cells with native LDL (25 micrograms mL-1) for 24 h, the production of both L-[14C]-citrulline [39600 (3600) cpm mg-1 cell protein] and nitric oxide [2.95 (0.56) mumol L-1] were about twice and 1.5-fold the amount of the corresponding values in untreated cells (mean +/- SD, P < 0.05, n = 4). Oxidized LDL was less effective than the native form. The presence of the arginine analogue NG-methyl-L-arginine reduced citrulline production dose-dependently but augmented DNA synthesis, both induced by LDL. In addition, the lipoprotein caused a 1.6-fold increase in cyclic GMP production following a 24-h incubation [control = 10.9 (3.8) pmol mg-1 cell protein, P = 0.016]. The results suggest that native LDL might partly impair its atherogenic potential on the vasculature by stimulating the production by smooth muscle cells of both nitric oxide and cyclic GMP.

In vitro antioxidant activity of calcium antagonists against LDL oxidation compared with alpha-tocopherol.

The dose-dependent (1, 5, 10, 50 microM) antioxidative activity of calcium antagonists (verapamil, diltiazem, nifedipine, amlodipine, isradipine or lacidipine) and alpha-tocopherol against copper-induced LDL (0.25 mg/ml) oxidation was compared by measuring the diene formation and the content of TBARS. For diltiazem no antioxidant effect could be found, whereas the other calcium antagonists and alpha-tocopherol have demonstrated antioxidant activity at least at concentrations of 10 and 50 microM: alpha-tocopherol > lacidipine > nifedipine > isradipine, verapamil, amlodipine. Additionally, alpha-tocopherol and lacidipine were able to attenuate LDL-oxidation significantly at 1 and 5 microM. These results indicate in vitro antioxidative activity of calcium antagonists especially from the dihydropyridine-type with greatest activity for the strongly lipophilic lacidipine. This might be one possible antiatherogenic mechanism of calcium antagonists, since oxidative modification enhances the atherogenic potential of LDL.

[The metabolic syndrome: common etiology for distinct cardiovascular risk factors?]. / Metabolisches Syndrom: gemeinsame Ursache für unterschiedliche kardiovaskuläre Risikofaktoren?

Hypertension, dyslipidemias, glucose intolerance and obesity are among the most important cardiovascular risk factors. There is growing evidence for the concept of a relationship between blood pressure regulation and metabolic changes. The combination of hypertension and the metabolic changes mentioned above has been named metabolic syndrome in the literature. The central role of insulin resistance and consecutive hyperinsulinemia in the metabolic syndrome has been shown in epidemiological, clinical, genetic and animal studies. The metabolic syndrome can be demonstrated in about one half of the hypertensive population. This pathophysiological concept has to be taken into consideration in the therapy and prevention of the different cardiovascular risk factors.

Low density lipoprotein subfractions and [Ca2+]i in vascular smooth muscle cells.

Several studies have established that plasma low density lipoprotein (LDL) consists of various discrete subfractions. Using a variety of techniques (analytical ultracentrifugation, equilibrium density gradient ultracentrifugation, and gradient gel electrophoresis), LDL has been fractionated into a maximum of seven subclasses that differ in particle size, density, and physiochemical composition. Recently, a predominance of smaller denser LDL particles has been associated with an increased risk of coronary artery disease. However, other lipoprotein changes, such as elevated triglycerides and lower HDL cholesterol levels, have been shown in patients with a predominance of the smaller denser LDL subfractions. Thus, it is unclear whether the enhanced atherogenic potential is induced by the LDL subfraction pattern per se or by concomitant lipoprotein changes. Because intracellular free Ca2+ is an important second messenger involved in atherogenesis and regulation of vascular tone, we studied the influence of three LDL subfractions (very light [LDL1], 1.030 to 1.033 g/mL; light [LDL2], 1.033 to 1.040 g/mL; and dense [LDL3], 1.040 to 1.045 g/mL) on [Ca2+]i in vascular smooth muscle cells (VSMCs) cultured from rat aorta. LDL subfractions were isolated by density gradient ultracentrifugation from human EDTA-plasma (n = 15). [Ca2+]i was measured by fura 2 fluorescence. Basal [Ca2+]i was 77 +/- 6 nmol/L. Stimulation of VSMCs with dense LDL3 caused a significantly (P < .05) more pronounced increase (+71 +/- 13 nmol/L) compared with LDL1 (+38 +/- 8 nmol/L) and LDL2 (+36 +/- 9 nmol/L). To further investigate the mechanisms leading to the stimulation of [Ca2+]i by LDL subfractions, we incubated VSMCs with the Ca2+ antagonists nifedipine, diltiazem, and verapamil in concentrations up to 10 mumol/L.(ABSTRACT TRUNCATED AT 250 WORDS)

Inositol Trisphosphate Metabolism in Subcellular Fractions of Barley (Hordeum vulgare L.) Mesophyll Cells.

Phosphatases in cytosolic fractions, vacuoles, and vacuolar membranes from barley (Hordeum vulgare L.) leaves were found to dephosphorylate inositol 1,4,5-trisphosphate (IP3). 1,4-inositol bisphosphate (1,4-IP2) is the main product of IP3 dephosphorylation by the cytosolic fraction. The activity was strictly Mg2+ dependent. In contrast, IP3 dephosphorylation activity of both the soluble vacuolar and the tonoplast fractions was inhibited up to 50% by Mg2+. When vacuolar membranes were incubated with IP3, 1,4-IP2 was produced only under neutral and slightly alkaline conditions. Under acidic conditions, however, dephosphorylation yielded putative 4,5-inositol bisphosphate. Li+ (20 mM) and Ca2+ (100 [mu]M) strongly inhibited activity in the soluble vacuolar fraction but had only a slight effect on the activities of the cytosolic and tonoplast fractions.