[Rose-scented geranium essential oil from Algeria (Pelargonium graveolens L'Hérit.): Assessment of antioxidant, anti-inflammatory and anticancer properties against different metastatic cancer cell lines]. / Essence aromatique du Géranium Odorant (Pelargonium graveolens L'Hérit.) d'Algérie : exploration des propriétés antioxydante, anti-inflammatoire et anticancéreuse (anti-angiogénique et cytotoxique), in vitro et in ovo, vis-à-vis de différentes lignées cellulaires cancéreuses métastasiques.

INTRODUCTION: The study of natural products is one of the strategies implemented for the discovery of new compounds that can be used in cancer therapy. Aromatic herbs and medicinal plants found in Algeria and their anti-angiogenesis and cytotoxic potentials against cancer have not been much explored. OBJECTIVES: Our work aimed to evaluate the antioxidant, anti-inflammatory and anticancer properties of the essential oil (EO) extracted from rose-scented geranium (Pelargonium graveolens) and its major (citronellol) and characteristic (linalool) constituents. RESULTS: The chemical composition of EO was determined with chromatographic analysis and revealed the presence of citronellol as the major compound (25.84%). A strong chelating power of terpene alcohols (IC50=1.58±0.23mg/mL for citronellol) was found, with a significant difference (P<0.05) compared with the standard antioxidants used (L-ascorbic acid and butylated hydroxyanisole). The EO is distinguished by an interesting anti-inflammatory effect with the lowest IC50 (4.63±0.3mg/mL), and it constitutes a good stabilizer of the erythrocyte membrane. Citronellol also exhibited the best anti-inflammatory effect (IC50=0.74±0.09mg/mL). We also assessed the anticancer effect of EO on two main pathways involved in cancer development, angiogenesis and cell proliferation, using in ovo bioassays with a chorio-allantoic membrane (CAM) of chicken eggs and in vitro assays of its cytotoxicity on different metastatic breast cancer (MDA-MB-231), gastric (AGS) and melanoma (MV3) cell lines. In the CAM model, the density of micro-vessels is 75±10 in the group supplemented with EO compared to 140±9 for the control group (b-FGF). In addition, the EO significantly reduced the number of newly formed vessels. The cytotoxicity was evaluated using the cell proliferation inhibition method and cell viability was measured using the MTT test. Results revealed that the treatment of cancer lines with different concentrations of EO reduces the rate of cell viability in a dose-dependent manner. EO showed the greatest cytotoxicity on the AGS line with an inhibition rate of 92.87±0.13% at the highest dose (4µL/mL), followed by the MV3 line (88.76±0.96%). CONCLUSION AND PROSPECTS: Data demonstrated that rose-scented geranium EO has an antitumor potential on metastatic cancer cell lines. It is distinguished by its antiproliferative, anti-angiogenic, and anti-inflammatory activities. Medicinal plants might contain new molecules, with new structures, which could become lead candidate among future anticancer drugs.

Avoiding COVID-19 complications with diabetic patients could be achieved by multi-dose Bacillus Calmette-Guérin vaccine: a case study of beta cells regeneration.

Diabetes mellitus (DM) is one of the major risk factors for COVID-19 complications as it is one of the chronic immune-compromising conditions especially if patients have uncontrolled diabetes, poor HbA1c and/or irregular blood glucose levels. Diabetic patients' mortality rates with COVID-19 are higher than those of cardiovascular or cancer patients. Recently, Bacillus Calmette-Guérin (BCG) vaccine has shown successful results in reversing diabetes in both rats and clinical trials based on different mechanisms from aerobic glycolysis to beta cells regeneration. BCG is a multi-face vaccine that has been used extensively in protection from tuberculosis (TB) and leprosy and has been repositioned for treatment of bladder cancer, diabetes and multiple sclerosis. Recently, COVID-19 epidemiological studies confirmed that universal BCG vaccination reduced morbidity and mortality in certain geographical areas. Countries without universal policies of BCG vaccination (Italy, Nederland, USA) have been more severely affected compared to countries with universal and long-standing BCG policies that have shown low numbers of reported COVID-19 cases. Some countries have started clinical trials that included a single dose BCG vaccine as prophylaxis from COVID-19 or an attempt to minimize its side effects. This proposed research aims to use BCG vaccine as a double-edged weapon countering both COVID-19 and diabetes, not only as protection but also as therapeutic vaccination. The work includes a case study of regenerated pancreatic beta cells based on improved C-peptide and PCPRI laboratory findings after BCG vaccination for a 9 year old patient. The patient was re-vaccinated based on a negative tuberculin test and no scar at the site of injection of the 1st BCG vaccination at birth. The authors suggest and invite the scientific community to take into consideration the concept of direct BCG re-vaccination (after 4 weeks) because of the reported gene expressions and exaggerated innate immunity consequently. As the diabetic MODY-5 patient (mutation of HNF1B, Val2Leu) was on low dose Riomet® while eliminating insulin gradually, a simple analytical method for metformin assay was recommended to ensure its concentration before use as it is not approved yet by the Egyptian QC labs.

[Background and current use of adjuvants for regional anesthesia : From research to evidence-based patient treatment]. / Hintergrund und aktueller Einsatz von Adjuvanzien für die Regionalanästhesie : Aus der Forschung in die evidenzbasierte Patientenversorgung.

The discovery of the local anaesthetic effect by blocking sodium ion channels was a milestone in anaesthesia but was soon limited by sometimes life-threatening toxic effects of the local anaesthetics. By developing novel local anaesthetics and also by adding so-called adjuvants, attempts have been made to limit these life-threatening events. This article focuses on the historic background and the current state of the use of these adjuvants for regional anaesthesia. Adding epinephrine, clonidine or dexmedetomidine, but only as a single dose, results in a faster onset, longer duration of action and increased intensity of neuronal blockade of regional anaesthesia. The benefits of adding sodium bicarbonate, on the other hand, are relatively minor and, therefore, clinically negligible. Although increasing evidence in the literature suggests an improvement and prolongation of the analgesic effect after axonal administration of opioids, which can also be given continuously, systemic effects are not fully ruled out due to the increased incidence of central side effects. The partial local anaesthetic effects of opioids cannot always be distinguished from opioid receptor-specific effects. Mechanistic studies postulate a functional coupling of opioid receptors in injured rather than in intact peripheral nerves. Recent studies have identified glucocorticoid and mineralocorticoid receptors predominantly on peripheral nociceptive nerve fibers. This is consistent with numerous clinical reports of a marked prolongation of the local anaesthetic effect. In addition to the known genomic effects of steroids that occur via a change in gene expression of pain-sustaining protein structures, faster non-genomic effects are also discussed, which occur via a change in intracellular signaling pathways. In summary, new insights into mechanisms and novel results from clinical trials will help the anaesthesiologist in the decision to use adjuvants for regional anaesthesia which, however, requires to weigh the individual patient's benefits against the risks.

Pain control in inflammation governed by selectins.

Opioid-containing immune cells migrate preferentially to inflamed sites, where they release beta-endorphin which activates peripheral opioid receptors to inhibit pain. Immunocyte recruitment is a multistep, sequential engagement of various adhesion molecules located on immune cells and vascular endothelium. Selectins mediate the initial phase of immunoctye extravasation into inflamed sites. Here we show that anti-selectin treatment abolishes peripheral opioid analgesia elicited either endogenously (by stress) or by corticotropin-releasing factor. This results from a blockade of the infiltration of immunocytes containing beta-endorphin and the consequent decrease of the beta-endorphin content in the inflamed tissue. These findings indicate that the immune system uses mechanisms of cell migration not only to fight pathogens but also to control pain in injured tissue. Thus, pain is exacerbated by measures inhibiting the immigration of opioid-producing cells or, conversely, analgesia might be conveyed by adhesive interactions that recruit those cells to injured tissue.

Lycopene nanoparticles promotes osteoblastogenesis and inhibits adipogenesis of rat bone marrow mesenchymal stem cells.

OBJECTIVE: Lycopene is a carotenoid and antioxidant with potent singlet oxygen quenching ability that reduces oxidative stress and promotes bone health. However, the cellular mechanisms by which lycopene influences bone metabolism are not known. MATERIALS AND METHODS: The present study investigated the effects of lycopene nanoparticles on the differentiation of rat bone marrow-derived mesenchymal stem cells into osteoblasts or adipocytes. RESULTS: In osteogenic medium, lycopene supplementation dose-dependently enhanced osteoblast differentiation, as evidenced by the transcription of Alpl, Runx2, Col1a1, Sp7, and Bglap, higher alkaline phosphatase activity, osteocalcin secretion and extracellular matrix mineralisation seen with Alizarin red S staining, and increased haem oxygenase levels. By contrast, lycopene in adipogenic medium inhibited adipocyte differentiation evidenced by decreases in the transcription of Tnfsf11, Tnfrsf11b, Pparg, Lpl, and Fabp4 and reduced fat accumulation observed by Oil Red O staining. CONCLUSIONS: Lycopene nanoparticles may promote bone health and are considered as a potential candidate for the prevention and/or treatment of bone loss conditions.

Antinociception by neutrophil-derived opioid peptides in noninflamed tissue--role of hypertonicity and the perineurium.

Inflammatory pain can be controlled by intraplantar opioid injection or by secretion of endogenous opioid peptides from leukocytes in inflamed rat paws. Antinociception requires binding of opioid peptides to opioid receptors on peripheral sensory nerve terminals. In the absence of inflammation, hydrophilic opioid peptides do not penetrate the perineurial barrier and, thus, do not elicit antinociception. This study was designed to examine the conditions under which endogenous, neutrophil-derived hydrophilic opioid peptides (i.e. Met-Enkephalin and beta-endorphin) can raise nociceptive thresholds in noninflamed tissue in rats. Intraplantar injection of the chemokine CXCL2/3 (macrophage inflammatory protein-2) induced selective neutrophil recruitment without overt signs of inflammation or changes in mechanical nociceptive thresholds (paw pressure threshold). Following intraplantar injection of hypertonic saline, the perineurial barrier was permeable for hours and intraplantar injection of opioid peptides increased mechanical nociceptive thresholds. While formyl-Met-Leu-Phe (fMLP) triggered opioid peptide release from neutrophils in vitro, nociceptive thresholds were unchanged in vivo. In vitro, hypertonicity interfered with fMLP-induced p38 mitogen activated kinase (MAPK) phosphorylation and opioid peptide release from neutrophils. These inhibitory effects were fully reversible by washout. In vivo, return to normotonicity occurred within 30min while the perineurium remained permeable for hours. Under these conditions, fMLP triggered MAPK phosphorylation and induced opioid peptide-mediated increases in nociceptive thresholds in the noninflamed paw. Taken together, antinociception mediated by endogenous opioids in noninflamed tissue has two important requirements: (i) opening of the perineurial barrier for opioid peptide access and (ii) opioid peptide release from neutrophils involving p38 MAPK.

AlphaIIbbeta3 priming and clustering by orally active and intravenous integrin antagonists.

BACKGROUND: Drugs that block platelet-platelet and platelet-fibrin interactions via the alpha(IIb)beta(3) (glycoprotein IIb/IIIa) receptor are used daily in patients undergoing percutaneous coronary interventions. Along with expected increases in spontaneous bleeding, clinical trials have revealed a surprising increase in thrombosis when these drugs are used without other anticoagulants. A better understanding of their mechanisms can minimize these risks. OBJECTIVES: This study tested the hypothesis that interventions designed to block fibrinogen binding inevitably leave the alpha(IIb)beta(3) receptor in an activated state. It compared the effects on platelet function and alpha(IIb)beta(3) conformation of the orally active compounds orbofiban and roxifiban, the i.v. agents eptifibatide and tirofiban, and echistatin, an arginine-glycine-aspartate (RGD) disintegrin. METHODS: The integrin antagonist concentrations required to saturate platelets and to block platelet-platelet and platelet-fibrin interactions were determined by flow cytometry, aggregometry, and clot-based adhesion assays, respectively. Analytical ultracentrifugation measured each antagonist's effects on the solution structure of alpha(IIb)beta(3). Fluorescence anisotropy provided equilibrium and kinetic data for integrin:antagonist interactions. RESULTS: Both orally active drugs bound more tightly and inhibited platelet aggregation and adhesion to fibrin more effectively than echistatin. Analytical ultracentrifugation yielded this order for perturbing alpha(IIb)beta(3) conformation (priming) and promoting oligomerization (clustering): echistatin > eptifibatide > orbofiban > tirofiban > roxifiban. Roxifiban was also most effective at disrupting the rapidly forming/slowly dissociating alpha(IIb)beta(3):echistatin complex. CONCLUSIONS: Our results suggest that the same molecular mechanisms that enable glycoprotein IIb/IIIa inhibitors to bind tightly to the alpha(IIb)beta(3) receptor and block fibrinogen binding contribute to their ability to perturb the resting integrin's conformation, thus limiting the safety and efficacy of both oral and i.v. integrin antagonists.

Spontaneous recanalization in deep venous thrombosis: a prospective duplex ultrasound study.

AIM: The aim of this study was to evaluate the dynamics of the recanalization process (spontaneous fibrinolysis) in completely occlusive deep venous thrombosis (DVT) using duplex ultrasound examination and to investigate the influence of different factors on the evolution of thrombus regression. METHODS: This longitudinal prospective study was done with 74 consecutive patients with completely occlusive acute multilevel DVT, confirmed by echo duplex scan after 1, 3, 6, and 12 months. At each re-evaluation, the degree and the type of recanalization were determined. Efficacy of tinzaparin (175 IU/kg, s.c., q.d. for 7-14 days) and continued with warfarin (12 months at INR 2-3) as well as patients' compliance with compressive elastic hosiery wearing were carefully followed. Relationship between the degree and pattern of recanalization and patients' age, gender, as well as thrombosis etiology and location were determined. RESULTS: Sixty-four patients completed the study. The mean recanalization rate was 39.7% at 1, 64.8% at 3, 82% at 6, and 90.3% at 12 months. Marginal recanalization was more frequently observed, but recanalization pattern was changing during follow-up. CONCLUSIONS: In the case of efficient anticoagulant and compressive therapy, the spontaneous recanalization process of DVT is important from the very first month of evolution, but an almost complete re-permeabilization is observed only after 12 months of treatment. The unilocular, marginal pattern of thrombus lysis is often observed and has better evolution than the multilocular cavernous one. The dynamics of recanalization are characterized by distal-to-proximal extension and in the first 6 months are significantly influenced by patient's gender and thrombosis etiology.

Interleukin-1 beta contributes to the upregulation of kappa opioid receptor mrna in dorsal root ganglia in response to peripheral inflammation.

During local painful inflammation, axonal transport of opioid receptors from dorsal root ganglia toward the periphery is increased, associated with a higher receptor density and enhanced efficacy of opioid analgesics at the injured site. To examine whether this increase is related to transcription, mRNA of the kappa opioid receptor in lumbar dorsal root ganglia was quantified by real time light cycler polymerase chain reaction. In dorsal root ganglia of naive rats, kappa opioid receptor mRNA expression was three-fold higher than previously shown for delta opioid receptor and two times lower than mu opioid receptor mRNA, respectively. After induction of unilateral paw inflammation by Freund's complete adjuvant, kappa opioid receptor mRNA was significantly upregulated with a peak at 12 h in ipsilateral dorsal root ganglia. This effect could be mimicked by intraplantar injection of the proinflammatory cytokine interleukin-1 beta. Kappa opioid receptor mRNA upregulation lasted longer in interleukin-1 beta-treated rats compared with Freund's complete adjuvant-treated rats. Furthermore, a significant increase in kappa opioid receptor positive neurons was detected by immunohistochemistry 24 h after local injection of Freund's complete adjuvant or interleukin-1 beta. In Freund's complete adjuvant-induced inflammation, kappa opioid receptor upregulation was blocked by treatment with interleukin-1 receptor antagonist without changing the leukocyte infiltration in the paw. In conclusion, kappa opioid receptor mRNA and protein in dorsal root ganglia are upregulated in response to peripheral inflammation. This effect can be mimicked by a single local injection of interleukin-1 beta, and Freund's complete adjuvant-induced upregulation in kappa opioid receptor mRNA and protein can be prevented by treatment with interleukin-1 receptor antagonist. These data suggest that the peripheral production of the proinflammatory cytokine interleukin-1 beta is a specific inducer of kappa opioid receptor expression in the dorsal root ganglia.

Inhibitory effect of C-reactive protein on the release of tissue factor pathway inhibitor from human endothelial cells: reversal by low molecular weight heparin.

AIM: The effects of C-reactive protein (CRP) and low molecular weight heparin (LMWH) on the release of tissue factor pathway inhibitor (TFPI) from human umbilical vein endothelial cells (HUVECs) were examined. METHODS: Confluent HUVECs were resuspended and plated in 48-well plates coated with fibronectin polymer. Cells were allowed to attach for 3 h; they were then washed, and fresh medium with or without CRP at different concentrations (0 to 20 ng/mL) was added. At 4 h, TFPI released in the medium was measured using a commercial TFPI ELISA kit for total TFPI antigen. In parallel assays, wells containing HUVECs and CRP were treated with tinzaparin at 1 mg/mL. RESULTS: Data showed that CRP significantly inhibited TFPI release from HUVECs in a concentration-dependent manner. In contrast, LMWH increased endothelial TFPI release. The amount of endothelial TFPI released was dependent on the heparin molecular weight distribution, with minimal effect at 3000 Da and maximum at 8000 to 12,000 Da. LMWH effectively reversed the inhibitory effects of CRP on TFPI release from HUVECs. CONCLUSIONS: These findings support the hypothesis that CRP may play a direct role in promoting a hypercoagulable state by decreasing the release of the natural anticoagulant TFPI, which can be counteracted by LMWH.

Pro-angiogenesis action of thyroid hormone and analogs in a three-dimensional in vitro microvascular endothelial sprouting model.

AIM: Our laboratory has recently demonstrated the pro-angiogenesis effects of thyroid hormone in the chick chorioallantoic membrane model. METHODS: Generation of new blood vessels from existing vessels was promoted two- to three-fold by either L-thyroxine (T4) or 3,5,3'-triiodo-L-thyronine (T3) at total hormone concentrations of T7-T9 M. RESULTS: T4-agarose, a formulation of thyroid hormone that does not cross the cell membrane, produced a potent pro-angiogenesis effect comparable to that obtained with T3 or T4. In the present investigation, T3, T4, T4-agarose, and basic fibroblast growth factor, each added to vascular endothelial growth factor, produced comparable pro-angiogenesis effects in the in vitro three-dimensional human microvascular endothelial sprouting model. The pro-angiogenesis effect of the thyroid hormone analogs was blocked by PD 98059, an inhibitor of the mitogen-activated protein kinase (MAPK; ERK1/2) signal transduction cascade. A specific avb3 integrin antagonist (XT199) also inhibited the pro-angiogenesis effect of either thyroid hormone analogs or T4-agarose. Tetrac, a thyroid hormone analog that blocks cell surface-initiated actions of T4 and T3, inhibited the pro-angiogenesis response of thyroid hormone. CONCLUSIONS: T4, T3, and T4-agarose are pro-angiogenic in the three-dimensional human microvascular endothelial sprouting model, an action that is initiated at the plasma membrane, involves avb3 integrin receptors, and is MAPK-dependent.

Selective alpha(v)beta(3)-receptor blockade reduces macrophage infiltration and restenosis after balloon angioplasty in the atherosclerotic rabbit.

BACKGROUND: alpha(v)beta(3)-Integrin receptors are upregulated in atherosclerotic arteries and play a key role in smooth muscle cell and possibly inflammatory cell migration. We hypothesized that after balloon angioplasty (BA) of atherosclerotic arteries, selective inhibition of the alpha(v)beta(3)-receptor by XT199, a small-molecule, non-peptide-selective alpha(v)beta(3)-receptor antagonist, would reduce restenosis. METHODS AND RESULTS: After induction of focal atherosclerosis, rabbits underwent femoral BA and received XT199 (2.5 mg/kg IV bolus plus 2.5 mg. kg(-1). d(-1) IV; n=19) or vehicle (n=20) for 14 days. At 28 days after BA, the XT199 group had a larger lumen (0.75+/-0.26 versus 0.57+/-0.20 mm(2), P=0.03) and a smaller neointimal area (0.49+/-0.18 versus 0.68+/-0.25 mm(2), P=0.01) than the vehicle group. Angiographic analysis confirmed a 30% to 40% reduction in restenosis. Arteries harvested at 28 days after BA did not show a reduction in intima plus media smooth muscle cell content but did show a 50% reduction in macrophage cell density in the XT199 group (716+/-452 versus 1458+/-989 cells/mm(2), P<0.006). Neovessel density at 28 days was also reduced (23+/-42 versus 58+/-46 vessel cross sections/mm(2), P<0.02). Early after BA (ie, 3 to 7 days), there was a decrease in intracellular adhesion molecule-1 and vascular cell adhesion molecule-1 expression, indicative of a reduction in vascular cell activation. CONCLUSIONS: Selective alpha(v)beta(3)-receptor blockade for 14 days after BA in the focally atherosclerotic rabbit significantly reduced restenosis and limited macrophage infiltration and neovascularization in the vessel wall.

Comparative antiplatelet efficacy of a novel, nonpeptide GPIIb/IIIa antagonist (XV454) and abciximab (c7E3) in flow models of thrombosis.

Glycoprotein (GP) IIb/IIIa is pivotal in homotypic platelet aggregation and may also be involved in the heterotypic adhesion of leukocytes and tumor cells to platelets. This study was primarily undertaken to compare the antiplatelet efficacy of a novel, nonpeptide GPIIb/IIIa antagonist, XV454, to that of abciximab in 2 flow models of platelet thrombus formation: (1) direct shear-induced platelet aggregation imposed by a cone-and-plate rheometer and (2) platelet adhesion onto von Willebrand factor (vWF)/collagen I followed by aggregation in a perfusion system. XV454 inhibited platelet aggregation in a concentration-dependent manner in both experimental models. Maximal inhibition of aggregation was achieved by XV454 at approximately 70% receptor occupancy, which is lower than the >/=85% previously reported for abciximab. At similar levels of receptor blockade (approximately 45%), XV454 appeared to be relatively more effective than abciximab in suppressing platelet aggregation. Neither XV454 nor abciximab inhibited platelet adhesion to collagen. Pretreatment of surface-adherent platelets with either XV454 or abciximab inhibited the attachment of monocytic THP-1 cells under flow. In contrast, the rapidly reversible GPIIb/IIIa inhibitor orbofiban failed to suppress these heterotypic interactions. These findings demonstrate that XV454 is a potent GPIIb/IIIa antagonist with a long receptor-bound lifetime like abciximab and may be beneficial for the treatment/prevention of thrombotic complications.

Elevation of plasma von Willebrand factor and tumor necrosis factor-a in obese subjects and their reduction by the low molecular weight heparin tinzaparin.

AIM: Increased plasma-soluble von Willebrand factor (vWF) level, a marker of vascular endothelial cell dysfunction, is a predictor of atherosclerotic cardiovascular disease. We compared associations between vWF level and markers of inflammation as well as the effects of LMWH in obese as compared to healthy human subjects. METHODS: Plasma samples were obtained from healthy volunteers (n=32) and obese subjects (n=12) before and after administration of a single subcutaneous dose of tinzaparin, given at 75 IU/kg once a day, a deep vein thrombosis prophylaxis dose. Plasma samples were analyzed for vWF and tumor necrosis factor-alfa (TNF-alfa) using specific and sensitive ELISA. RESULTS: Obese subjects showed relatively higher plasma levels of TNF-alfa compared with normal-weight subjects. Regression analysis showed that plasma vWF levels to be directly associated with the presence of higher plasma levels of TNF-alfa in these obese subjects. Tinzaparin significantly reduced elevated plasma levels of both vWF and TNF-a levels (P<0.01). CONCLUSIONS: Plasma values of vWF and TNF-alfa are higher in obese than in normal-weight individuals. Treatment with tinzaparin lowers plasma levels of TNF-alfa in both obese and normal-weight subjects. The levels of vWF were higher in obese subjects than in normal-weight ones, which might be due to the higher levels of circulating TNF-alfa. Tinzaparin reduced vWF levels in these obese subjects.

Pharmacodynamic effects of low molecular weight heparin in obese subjects following subcutaneous administration of 75 IU/kg on plasma tissue factor pathway inhibitor and nitric oxide.

AIM: Pharmacodynamic effects of the low molecular weight heparin tinzaparin on plasma levels of tissue factor pathway inhibitor (TFPI) and nitric oxide (NO) were compared in obese subjects, as well as in normal healthy controls. METHODS: Obese (n = 13) patients received a single 75 IU/kg SC injection (the deep vein thrombosis prophylaxis dose) of tinzaparin. Blood samples were obtained pre- and postadministration of drug and at different intervals over 24 h and assayed for total TFPI and NO stable metabolites (nitrates and nitrites) plasma levels, using a specific immunoassay and calorimetric methods. RESULTS: Mean maximum plasma TFPI levels approached 150-230 ng/ml at the 0.8 h and up to 5 h posttinzaparin dose compared to basal TFPI levels of 35-90 ng/mL. Plasma TFPI levels were still about 2-fold above basal levels at 12 h and fell to basal levels at 16 h after tinzaparin dose. Basal plasma levels of NO, but not TFPI, were significantly lower (P < 0.01) in obese patients compared to controls. Similar TFPI (3-fold above basal at peak) and NO pharmacodynamic profiles for tinzaparin at 75 anti-Xa IU/kg were demonstrated in obese and in normal healthy subjects. Plasma NO (nitrate + nitrite) showed a lag time of about 5-6 h posttinzaparin followed by a steady increase with a peak at 12-15 h and slow decline with a significant residual level at 24 h in obese and healthy subjects. CONCLUSIONS: Data suggest a normal responsiveness of vascular endothelial cells and other cellular compartments to tinzaparin with regard to the pharmacodynamic profiles of plasma TFPI and NO in obese subjects.

Antithrombotics in thrombosis and cancer.

Many cancer patients have a hypercoagulable state, with recurrent thrombosis due to the impact of cancer cells and chemotherapy or radiotherapy on the coagulation cascade. Studies have demonstrated that unfractionated heparin (UFH) or its low molecular weight fractions interfere with various processes involved in tumour growth and metastasis. These include fibrin formation; binding of heparin to angiogenic growth factors, such as basic fibroblast growth factor (FGF2) and vascular endothelial growth factor (VEGF); modulation of tissue factor; and perhaps other more important modulatory mechanisms, such as enhanced tissue factor pathway inhibitor (TFPI) release and inhibition of various matrix-degrading enzymes. Clinical trials have suggested a clinically relevant effect of low molecular weight heparin (LMWH), as compared to UFH, on the survival of cancer patients with deep vein thrombosis. Similarly, the impact of warfarin on the survival of cancer patients with thromboembolic disorders was demonstrated. Studies from our laboratory demonstrated a significant role for LMWH, warfarin, anti-VIIa, and LMWH-releasable TFPI on the regulation of angiogenesis, tumour growth, and tumour metastasis. Thus, modulation of tissue factor/VIIa non-coagulant activities by LMWH, warfarin, anti-VIIa, or TFPI might be a useful therapeutic method for the inhibition of angiogenesis associated with human tumour growth and metastasis. Additionally, antiplatelet drugs could have an impact on tumour metastasis, and the combination of antiplatelets and anticoagulants at adjusted doses might provide greater benefits to cancer patients.

Differential antiplatelet efficacy for various GPIIb/IIIa antagonists: role of plasma calcium levels.

OBJECTIVES: The present study was undertaken to determine the effects of free ionized calcium influenced by either the anticoagulant used (citrate vs. heparin) or directly varying the calcium levels after treatment of blood with citrate on the antiplatelet efficacy of two classes of GPIIb/IIIa antagonists. METHODS: The platelet effects of changes in plasma [Ca(++)] with the different GPIIb/IIIa antagonists were determined using light transmittance aggregometry, direct binding kinetics, and (125)I-fibrinogen binding to activated human platelets. RESULTS: A significantly higher IC50s was shown with heparin (free ionized calcium=1.1 mM) as compared to that with citrate (free ionized calcium=0.12 mM) with class II GPIIb/IIIa antagonists (P<0.01) such as Orbofiban, and Integrilin. In contrast, class I GPIIb/IIIa antagonists such as Roxifiban and Abciximab showed no significant changes in their IC50s in either citrate or heparin. Similar data were shown with other non-calcium chelating anticoagulant such as PPACK as compared to that with heparin. Additionally, similar data were shown with regard to the [Ca(++)] sensitivity for GPIIb/IIIa antagonists from Class II but not Class I in the changes in IC50 values required for the inhibition of (125)I-fibrinogen binding to activated human gel filtered platelets. Additionally, examples from Class I GPIIb/IIIa antagonists such as (3)H-active form of Roxifiban showed no significant changes in its platelet binding affinity in response to change in [Ca(++)]. In contrast, GPIIb/IIIa antagonists from class II such as (3)H-active form of Orbofiban demonstrated significant changes (P<0.01) in its platelet binding kinetics and antiplatelet efficacy in response to changes in Ca(++) concentrations. CONCLUSIONS: These data suggest the impact of the method of blood collection or changes in plasma calcium levels on the antiplatelet efficacy for class II but not class I GPIIb/IIIa antagonists depending on their platelet binding kinetics.

Selective alpha v beta 3 integrin blockade potently limits neointimal hyperplasia and lumen stenosis following deep coronary arterial stent injury: evidence for the functional importance of integrin alpha v beta 3 and osteopontin expression during neointima formation.

UNLABELLED: Lumen loss from vascular restenosis remains a leading cause of chronic revascularization failure. OBJECTIVE: We hypothesized that cell-matrix adhesion, migration, and differentiation events that underlie restenosis are mediated by alpha v beta 3 integrin-ligand interactions. METHODS: Using immunohistochemistry and in situ hybridization, we examined the spatial and temporal vessel wall expression of alpha v beta 3 and osteopontin following deep coronary arterial injury. Cell migration and adhesion assays were performed to demonstrate the affinity and specificity of XJ 735 for various vessel wall integrins. The effects of XJ 735 (a selective cyclic Arg-Gly-Asp (RGD) peptidomimetic alpha v beta 3 antagonist) on neointimal hyperplasia and lumen stenosis were tested in a porcine coronary injury model. Normolipemic swine underwent oversized stent injury followed by XJ 735 administration (9 animals, 28 lesions; 1 mg/kg bolus + 7 days 4 mg/kg/d infusion + 21 days 2 mg/kg i.v. bolus 12 hourly) or placebo (10 animals, 30 arterial lesions). RESULTS: Maximal alpha v beta 3 immunoreactivity was observed between 7-14 days following injury in the neointima, media, and adventitia. Maximal osteopontin mRNA signal in the neointima, media, and adventitia was observed at 14, 7 and 28 days respectively. IC50 for XJ 735 alpha v beta 3-mediated inhibition of human and porcine endothelial cell adhesion, and vascular smooth muscle cell migration, ranged from 0.6 to 4.4 microM. In contrast, IC50 for porcine or human alpha IIb/beta 3, alpha 4 beta 1, alpha v beta 5, and alpha 5 beta 1 inhibition exceeded 100 microM. Steady state XJ 735 plasma levels exceeded 5 microM. Despite slightly higher injury scores in XJ 735 treated animals, significant reductions in mean neointima area (43% reduction; p = 0.0009), and mean percent lumen stenosis (approximately 2.9 fold reduction; p = 0.04) were observed in XJ 735 treated animals. XJ 735 treatment did not significantly alter the relative size of the arterial injury and reference sites (geometric remodeling). Comparison of neontima area vs. injury score regression lines revealed significant reductions in slope (p = 0.0001) and intercept (p = 0.0001) for XJ 735. CONCLUSIONS: Selective alpha v beta 3 blockade is an effective anti-restenosis strategy that potently limits neointimal growth and lumen stenosis following deep arterial injury. The co-ordinate spatial and temporal upregulation of alpha v beta 3 expression following vessel wall injury, and the high affinity and specificity of XJ 735 for alpha v beta 3, confirms the importance of this integrin in adhesive and migratory cell-matrix events underlying coronary restenosis.

Asperger's syndrome: diagnosis, comorbidity and therapy.

Asperger's syndrome (AS), a behavioral disorder that is related to autism, is associated with abnormal social functioning and repetitive behaviors but not with a decrease in intelligence or linguistic functionality. This article reviews the clinical diagnosis of AS and discusses the comorbid disorders that may be present with AS, as well as the efficacy, safety, and tolerability of pharmacotherapies given to AS patients, as reported in preclinical and clinical studies. AS may be present with several comorbid disorders including: attention deficit hyperactivity disorder, anxiety, schizophrenia, bipolar disorder, depression, and Tourette's syndrome. The difficulty in distinguishing AS from autism results in treating the comorbid disorder symptoms, rather than treating the symptoms of AS. Accordingly, there is a great need to further understand the psychobiology of AS and its association with other disorders, which should expand the pharmacological and non-pharmacological therapeutic options and improve the quality of life for AS patients.

Ultrastructural changes associated with myocardial apoptosis, in failing rat hearts induced by volume overload.

BACKGROUND: Myocardial apoptosis has been discussed to play a pivotal role in the development and progression of congestive heart failure (CHF). However, recently there is doubt on the evidence of myocardial apoptosis in heart failure as information on ultrastructural changes by electron microscopy is still scarce. This project therefore aimed to detect direct morphological evidence of myocardial apoptosis in an experimental heart failure model. METHOD: Following IRB approval, an aortocaval fistula (ACF) was induced in male Wistar rats using a 16G needle. 28±2days following ACF rats were examined by hemodynamic measurements, Western blot, immunofluorescence confocal and electron microscopic analysis. RESULTS: Within 28±2days of ACF heart (3.8±0.1 vs. 6.6±0.3mg/g) and lung (3.7±0.2 vs. 6.9±0.5mg/g) weight indices significantly increased in the ACF group accompanied by a restriction in systolic (LVEF: 72±2 vs. 39±3%) and diastolic (dP/dtmin.: -10,435±942 vs. -5982±745mmHg/s) function (p<0.01). Activated caspase-3 was significantly increased in failing hearts concomitant with mitochondrial leakage of cytochrome c into the cytosol. Finally, electron microscopy of the left ventricle (LV) of ACF rats revealed pronounced ultrastructural changes in >70% of examined cardiomyocytes, such as nuclear chromatin condensation, myofibril loss and disarray, contour irregularities and amorphous dense bodies, mitochondriosis and damaged cell-cell-contacts between cardiomyocytes. CONCLUSIONS: Volume overload induced heart failure is associated with activation of the mitochondrial apoptotic pathway. In addition, electron microscopy of the LV revealed direct ultrastructural evidence of extended myocardial apoptosis in ACF rats.