The antithrombin binding regions of heparin mediate fetal growth and reduced placental damage in the RUPP model of preeclampsia†.

Preeclampsia is a serious hypertensive disorder of pregnancy, which is only cured with delivery of the placenta, thereby commonly necessitating preterm birth of the fetus. Low-molecular-weight heparin (LMWH) has demonstrated potential to reduce the incidence of preeclampsia in high-risk pregnant women, although the underlying mechanism by which LMWH protects against preeclampsia is unknown. Given the complex structure and biologic actions of heparin, we tested the hypothesis that heparin can mediate preeclampsia prevention via nonanticoagulant pathways. We compared the effects of a nonanticoagulant, glycol-split LMWH (gsHep)-rendered nonanticoagulant through disruption of the antithrombin binding regions-with the LMWH dalteparin in the rat reduced uterine perfusion pressure (RUPP) surgical model of preeclampsia. Although RUPP animals exhibit significantly elevated blood pressure and reduced plasma levels of placental growth factor (PGF) compared to sham, neither dalteparin nor gsHep treatment significantly impacted these parameters. However, the observed positive correlation between PGF levels and number of viable fetuses in RUPP-induced animals suggests that reduced PGF levels were predominately due to placental loss. Daily subcutaneous injections of low-dose dalteparin but not gsHep significantly restored fetal growth that was impaired by RUPP surgery. Placentas from RUPP animals exhibited an abnormal labyrinth structure, characterized by expanded sinusoidal blood spaces, relative to sham-operated animals. Morphometric analysis demonstrated that dalteparin but not gsHep treatment normalized development of the labyrinth in RUPP-exposed conceptuses. These data suggest that the antithrombin-binding regions of LMWH are required to confer its protective effects on fetal growth and placental development.

Role of dalteparin sodium on the growth of cancer cells and tumor-associated angiogenesis in A549 human lung cancer cell line and grafted mouse model.

PURPOSE: To investigate the effects of dalteparin sodium on the expression of vascular endothelial growth factor (VEGF), VEGF receptor (VEGFR), and hypoxia-inducible factor 1α (HIF-1α) in A549 human lung cancer (LC) cell line and a human A549-grafted nude mouse model. MATERIALS AND METHODS: A549 human lung adenocarcinoma cell line was divided into control group, treated using normal saline (NS); and dalteparin sodium groups, receiving 5, 15, 50, and 150 IU/ml of dalteparin sodium, respectively. Human A549-grafted nude mouse was induced through subcutaneous (SC) injection of A549 (5 × 106/0.2 ml) into the right armpit, and randomly assigned into control group (n = 6) receiving intraperitoneal (i.p.) injection of NS, cisplatin (DDP) group (n = 6, 3 mg/kg DDP alone, i.p., for 3 days), low molecular weight heparin (LMWH) group (n = 6) receiving SC injection of 1500 IU/kg dalteparin sodium for 35 days, and DDP plus LMWH group (n = 6, 3 mg/kg DDP, i.p., for 3 days, followed by SC injection of 1500 IU/kg dalteparin sodium for 35 days). RESULTS: Significant difference was noted in the messenger RNA expression of VEGF, VEGFR, and HIF-1α after treating with heparin with a concentration of 15, 50, or 150 IU/ml in the A549 cell line at 24 and 48 h, respectively. In the human A549-grafted nude mouse model, a significant reduction was noted in the expression of VEGF, VEGFR, and HIF-1α in the tumor mass harvested from the mice receiving administration of dalteparin sodium plus DDP. CONCLUSION: Dalteparin sodium had the inhibitory effects on the growth of human LC A549 cells in vitro and A549 LC xenograft model, which could be enhanced when administrated together with DDP.

FGF-2-containing dalteparin/protamine nanoparticles (FGF-2&D/P NPs) ameliorate UV-induced skin photoaging in hairless mice.

UVB exposure penetrates deeply into the dermis to alter skin barrier function, which is a primary factor in skin photoaging. We previously reported that dalteparin and protamine nanoparticles (D/P NPs) are effective carriers of FGF-2. This study aimed to examine the ability of FGF-2-containing D/P NPs (FGF-2&D/P NPs) to ameliorate UVB-induced skin photoaging in hairless mice. Dorsal skin of HR-1 hairless mice were exposed to UVB irradiation 5 days/week for 8 weeks (UV (+): final total, 2700 mJ/cm2). Mice were divided into four groups: Non-UVB (UV (-)) + saline, UV (+) + saline, UV (+) + FGF-2&D/P NPs, UV (+) + FGF-2, and UV (+) + D/P NPs, and following UVB irradiation, FGF-2&D/P NPs, FGF-2, and D/P NPs were applied to the groups of mice just after each UVB irradiation. Each group was subjected to evaluation of skin changes (elasticity), and histological examination using hematoxylin & eosin and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining. UVB irradiation of mice significantly induced a decline in elasticity and acanthosis, which was alleviated by application of FGF-2&D/P NPs. Furthermore, TUNEL-staining showed the proportions of apoptotic dermal fibroblast cells (DFCs) and epidermal keratinocyte cells (EKCs) in the UV (+) + FGF-2&D/P NPs group were significantly lower than those in the UV (+) + saline, UV (+) + FGF-2, and UV (+) + D/P NPs groups. Thus, FGF-2&D/P NPs may be effective in preventing skin photoaging accelerated by UVB irradiation such as declining elasticity, acanthosis, and apoptosis of DFCs and EKCs.

Effects of glycol-split low molecular weight heparin on placental, endothelial, and anti-inflammatory pathways relevant to preeclampsia.

Low molecular weight heparin (LMWH) is being investigated as a potential preventative therapy against preeclampsia. There is evidence suggesting that LMWH may prevent preeclampsia through anticoagulation-independent mechanisms. In this study, we compared the in vitro placental, endothelial, and anti-inflammatory effects of an LMWH (dalteparin) with a nonanticoagulant, glycol-split heparin derivative (gsHep). In contrast with dalteparin, gsHep did not interact with antithrombin III, possess significant anti-Factor Xa activity, or significantly prolong in vitro plasma clotting time. However, dalteparin and gsHep were otherwise mechanistically similar, both interacting with soluble fms-like tyrosine kinase-1 (sFlt1) and promoting release of the pro-angiogenic protein placental growth factor, but not the antiangiogenic sFlt1, from healthy placental villous explants. Placental explant media pretreated with dalteparin or gsHep significantly stimulated endothelial cell tube formation compared to untreated explants. Lastly, dalteparin and gsHep both significantly suppressed inflammation by inhibiting complement activation and leukocyte adhesion to endothelial cells that were activated using serum from preeclamptic women. Our data suggest that nonanticoagulant heparin derivatives may be utilized as a tool to distinguish the anticoagulation-independent mechanisms of LMWH, and provide insight into the role of anticoagulation in the prevention of preeclampsia.

Stability of Dalteparin 1,000 Unit/mL in 0.9% Sodium Chloride for Injection in Polypropylene Syringes.

The stability of dalteparin 1,000 units/mL in 0.9% sodium chloride for injection stored in polypropylene syringes under refrigeration was examined. Dalteparin 1,000-units/mL syringes were prepared by adding 9 mL of 0.9% sodium chloride for injection to 1 mL of dalteparin sodium 10,000 unit/mL from commercial single-use syringes. Compounded solutions in 0.5-mL aliquots were transferred to 1-mL polypropylene syringes and sealed with a Luer lock tip cap and stored at refrigerated temperatures (2°C to 8°C) with ambient fluorescent light exposure. Syringes from three batches of dalteparin 1,000 units/mL were potency tested in duplicate by a stability-indicating high-performance liquid chromatography assay using a 0.5-mL sample at specified intervals. Visual and pH testing were performed on each batch. Samples were visually inspected for container integrity, color, and clarity. Samples for pH testing were prepared using a 1:1 dilution of dalteparin 1,000 units/mL in sterile water for injection and underwent duplicate analysis at each time point. High-performance liquid chromatography analyses showed a remaining percent of the initial dalteparin content at day 30 of 94.88% ± 2.11%. Samples remained colorless and clear with no signs of container compromise and no visual particulate matter at each time point. Throughout the 30-day study period, pH values remained within 0.3-pH units from the initial value of 5.84. Dalteparin 1,000 unit/mL in 0.9% sodium chloride for injection, packaged in 1-mL polypropylene syringes was stable for at least 30 days while stored at refrigerated conditions with ambient fluorescent light exposure.

Analysis of contributing factors influencing thromboembolic events after total knee arthroplasty.

BACKGROUND: Venous thromboembolic events (VTE) are a known and well-described complication following total knee arthroplasty (TKA). We sought to validate the American College of Chest Physicians thromboprophylaxis recommendations after elective TKA, paying special attention to our dose adjustments for weight, and their impact on VTE in our population. METHODS: We retrospectively investigated risk factors in patients undergoing TKA, focusing mainly on symptomatic VTE occurrence rates from deep vein thrombosis (DVT) or pulmonary embolism (PE). The anticoagulation protocol consisted of starting low molecular-weight heparin (LMWH) therapy, with dalteparin administered 12 h after surgery in patients who received general anesthesia or 24 h later in patients who received single-dose regional anesthesia. RESULTS: Data from 346 patients (mean age 66.8 [range 24-91] yr) who underwent primary or revision TKA depicted an overall symptomatic VTE rate of 15%. The proximal DVT rate was 1.7%, and the nonfatal PE rate was 0.9%. The mean time to VTE diagnosis was 5.6 days. The first dalteparin dose was administered 19.5 (range 10-48) h after surgery in patients without VTE and 22.6 (range 11.5-52) h after surgery in patients with VTE (p = 0.003). With a first dose of dalteparin administered 12 h postoperatively, patients presented significantly lower DVT and PE rates than if it was administered 24 h postoperatively (8.5% v. 16.3%, p = 0.048). CONCLUSION: Delayed administration of LMWH has deleteriously impacted the VTE rate after TKA at our institution. Prompt initiation of LMWH (≤ 12 h after surgery) is appropriate, without increasing the risk of major bleeding.

CONTEXTE: Les événements thromboemboliques veineux (ETV) sont une complication connue et bien décrite de la chirurgie pour prothèse totale du genou (PTG). Nous avons voulu valider les recommandations de l'American College of Chest Physicians en matière de thromboprophylaxie après la PTG non urgente en portant une attention particulière à l'ajustement des doses selon le poids et à leur impact sur les ETV dans notre population. MÉTHODES: Nous avons analysé de manière rétrospective les facteurs de risque chez des patients soumis à une PTG en nous attardant principalement aux taux d'ETV symptomatiques sous forme de thrombose veineuse profonde (TVP) ou d'embolie pulmonaire (EP). Le protocole d'anticoagulothérapie prévoyait l'administration d'une héparine de bas poids moléculaire (HBPM), la daltéparine, 12 h après la chirurgie chez les patients ayant reçu une anesthésie générale, ou 24 h après chez les patients ayant reçu une anesthésie locorégionale à dose unique. RÉSULTATS: Les données provenant de 346 patients (âgés en moyenne de 66,8 and [éventail 24-91 and]) ayant subi une PTG primaire ou une révision de PTG ont révélé un taux d'ETV symptomatique global de 15 %. Le taux de TVP proximal a été de 1,7 % et le taux d'EP non fatale a été de 0,9 %. Le temps moyen avant le diagnostic d'ETV a été 5,6 jours. La première dose de daltéparine avait été administrée 19,5 h (éventail 10-48 h) après la chirurgie chez les patients n'ayant pas présenté d'ETV et 22,6 h (éventail 11,5-52 h) après la chirurgie chez les patients ayant manifesté un ETV (p = 0,003). Avec une première dose de daltéparine administrée 12 h après l'intervention, les patients ont présenté des taux de TVP et d'EP significativement moindres que si elle leur avait été administrée 24 h après l'intervention (8,5 % c. 16,3 %, p = 0,048). CONCLUSION: L'administration retardée de l'HBPM a produit des effets défavorables pour ce qui est des taux d'ETV après la PTG dans notre établissement. L'instauration rapide de l'HBPM (≤ 12 h après la chirurgie) est appropriée et n'accroît pas le risque d'hémorragie majeure.

Dalteparin or vitamin K antagonists to prevent recurrent venous thromboembolism in cancer patients: a patient-level economic analysis for France and Austria.

BACKGROUND: International guidelines recommend extended duration secondary prophylaxis in cancer patients who develop primary venous thromboembolism (VTE). Agent selection is guided in part by one large randomized trial (i.e., CLOT; Lee et al., N Engl J Med 349:146-53, 2003) which demonstrated that dalteparin reduced the relative risk of recurrence by 52% compared with oral vitamin K antagonists (VKA; HR = 0.48, 95% CI, 0.30 to 0.77). In a subgroup analysis from that same trial, patients with renal impairment also derived benefit with dalteparin (VTE rates = 3% vs. 17%; p = 0.011). To measure the economic value of secondary VTE prophylaxis with dalteparin, a patient-level pharmacoeconomic analysis was conducted from the Austrian and French healthcare system perspectives. METHODS: Chapter 1 Healthcare resource use collected during the CLOT trial was extracted and converted into direct cost estimates. Incremental cost differences between the dalteparin and VKA groups were then combined with health state utilities to measure the cost per quality-adjusted life year (QALY) gained. RESULTS: The dalteparin group had significantly higher costs than the VKA group in both countries (Austria: dalteparin = €2687 vs. VKA = €2012; France: dalteparin = €2053 vs. VKA = €1352: p < 0.001). However, when the incremental costs were combined with the utility gain, dalteparin had a cost of €6600 and €4900 per QALY gained in Austria and France, respectively. The analyses in patients with renal impairment suggested an even better economic profile, with the cost per QALY gained being less than €4000 in both countries. CONCLUSIONS: Secondary prophylaxis with dalteparin is a cost-effective alternative to VKA for the prevention of recurrent VTE in patients with cancer.

A retrospective analysis of outcomes of dalteparin use in pediatric patients: a single institution experience.

BACKGROUND: Dalteparin is a commonly used low molecular weight heparin (LMWH) with extensive safety data in adults. With distinct advantages of once daily dosing and relative safety in renal impairment, it has been used off-label in pediatric practice; however, age-based dosing guidelines, safety and efficacy data in children are evolving. OBJECTIVES: To report our institutional experience with the use of dalteparin in the treatment and prophylaxis of venous thromboembolism (VTE) in pediatric patients. PATIENTS/METHODS: Retrospective chart review of all children (0-18years) that received dalteparin from December 1, 2000 through December 31, 2011. Doses per unit body weight per day (units/kg/day) were calculated for age-based group comparisons. RESULTS: Of 166 patients identified, 116 (70%) received prophylactic doses while 50 (30%) received therapeutic doses of dalteparin. Infants (<1year) required significantly higher weight-based dosing to achieve therapeutic anti-Xa levels compared to children (1-10years) or adolescents (>10-18years) (mean dose units/kg/day; 396.6 versus 236.7 and 178.8 respectively, p<0.0001). Overall response rate, including complete and partial thrombus resolution, was 83%. Bleeding complications were minor and the rates were similar in therapeutic and prophylaxis patients. No significant differences in dosing or bleeding events were noted based on obesity or malignancy. CONCLUSIONS: In our experience, dalteparin is effective for prophylaxis and therapy of VTE in pediatric patients. Dosing should be customized in an age-based manner with close monitoring of anti-Xa activity in order to achieve optimal levels, prevent bleeding complications, and to allow full benefit of prevention or therapy of thrombotic complications.

The role of hepcidin in chronic mild stress-induced depression.

Depression is one of the most prevalent challenges of mental conditions. Yet its exact etiology has not been clear. Chronic stress increases the production of cytokines, which can lead to depression. Hepcidin, an iron modulator, is involved in the inflammation process as well as iron homeostasis. This study was designed to investigate the role of hepcidin, on stress-induced depression. 60 male wistar rats were entered the experiment. We used a chronic unpredictable mild stress (for 28 days) as a rat model of depression. In stressed group, three subgroups were treated with three different doses of dalteparin (a hepcidin inhibitor): 70IU/kg, 100IU/kg and 140IU/kg daily, for 4 weeks. The animals in the stressed group had more depressive-like behavior than the control group. Moreover, chronic mild stress produced an increased serum interleukin-6 levels. These effects were accompanied by an obvious increase in hepcidin mRNA level and iron content in the hippocampus. These changes were blocked by the injection of dalteparin. In conclusion, inhibition of hepcidin may reduce many pathological changes seen in stress-induced depressive disorders.

Platelet-rich plasma-containing fragmin-protamine micro-nanoparticles promote epithelialization and angiogenesis in split-thickness skin graft donor sites.

BACKGROUND: Platelet-rich plasma (PRP) contains multiple growth factors, and fragmin-protamine micro-nanoparticles (F-P M-NPs) significantly enhance and stabilize growth factors. The purpose of this study was to evaluate the effects of PRP-containing F-P M-NPs (PRP&F-P M-NPs) on wound repair in split-thickness skin graft (STSG-) donor sites (DS). MATERIALS AND METHODS: A total of 56 inbred male rats were anesthetized and split-thickness skin graft donor site (STSG-DS) were created with a Padgett dermatome. PRP&F-P M-NPs, F-P M-NPs, PRP, and saline (control) were then intradermally injected evenly into the STSG-DSs. On 3, 4, 5, 7, and 10 d after creation of STSG-DS, skin sample sections were stained with hematoxylin and eosin to evaluate reepithelialization and angiogenesis. RESULTS: Treatment of STSG-DS with PRP&F-P M-NPs effectively promoted epithelialization and new vessel formation compared with those treated with PRP, F-P M-NPs, and control (saline). CONCLUSIONS: The intradermal injection of PRP&F-P M-NPs promotes epithelialization and angiogenesis in STSG-DS wounds.

Identification of pro- and anti-proliferative oligosaccharides of heparins.

Heparins, unfractionated heparin (UFH) and low molecular weight heparins (LMWHs), are heterogeneous mixtures of anticoagulant and non-anticoagulant oligosaccharides. In addition to their well-known anticoagulant effect, heparins have shown to mediate a wide range of non-anticoagulant effects, including the modulation of cellular growth. However, contradictory results have been reported with regard to their effects on cellular proliferation, with some studies suggesting anti-proliferative while others indicating pro-proliferative effects. This study investigated the proliferation of human colonic epithelial cancer cells in the presence of UFH and LMWHs (enoxaparin and dalteparin). In our experimental setting, all heparins caused a dose-dependent reduction in cellular growth, which correlated well with the induction of cell cycle arrest in the G1 phase and which was not associated with significant changes in cell viability. The effects on cellular proliferation of 14 different oligosaccharides of enoxaparin obtained through ion-exchange chromatography were also assessed. Surprisingly, only two oligosaccharides showed distinctive anti-proliferative effects while the majority of oligosaccharides actually stimulated proliferation. Interestingly, the smallest oligosaccharide devoid of any anticoagulant activity showed the strongest anti-proliferative effect. Notably, heparins are currently standardised only according to their anticoagulant activity but not based on other non-anticoagulant properties. Our results indicate that slight differences in the composition of heparins' non-anticoagulant oligosaccharides, due to different origins of material and preparation methods, have the potential to cause diverse effects and highlight the need for additional characterisation of non-anticoagulant activities.

Pharmacodynamics of multi-dose low molecular weight heparin in healthy horses.

OBJECTIVES: To compare the pharmacodynamics of once daily and twice daily administration of low molecular weight heparin (LMWH) administration in horses. STUDY DESIGN: Randomized cross over study. ANIMALS: Adult mixed breed healthy mares (n = 6). METHODS: LMWH (dalteparin) was administered (50 U/kg subcutaneously) either every 12 or 24 hours for 3 consecutive days. Anti-factor Xa activity was measured before and at select time points after LMWH administration. Packed cell volume (PCV), platelet count, partial thromboplastin time (PTT), and anti-thrombin (AT) activity were monitored throughout the study. RESULTS: No changes in PCV, platelet count, or AT activity were detected with either frequency of daily LMWH administration. Values for PTT increased throughout the study but never exceeded the normal reference interval. Anti-factor Xa activity was maintained within or above the suggested thromboprophylactic range (0.1-0.2 U/mL) when LMWH was administered twice daily, but fell below this range ≈ 16 hours after administration when given once daily. For both once and twice daily LMWH administration, the area under the curve was significantly greater after the last dose of LMWH when compared to the first dose. CONCLUSIONS: Administration of LMWH once or twice daily for 3 consecutive days appears to be safe in healthy adult horses. Anti-factor Xa activity was maintained within or above the suggested thromboprophylactic range for 24 hours with twice daily LMWH administration but not with once daily administration.

Effective expansion of human adipose-derived stromal cells and bone marrow-derived mesenchymal stem cells cultured on a fragmin/protamine nanoparticles-coated substratum with human platelet-rich plasma.

Fragmin/protamine nanoparticles (F/P NPs) can be stably coated onto plastic surfaces and used as a substratum for the absorption and controlled release of growth factors (GFs) secreted from human platelet-rich plasma (PRP). In this study, we investigated the capability of F/P NP-coated plates to act as a substratum for the proliferation of human adipose-derived stromal cells (ASCs) and bone marrow-derived mesenchymal stem cells (BMSCs) with GFs in PRP. Both cell types adhered well to the F/P NP-coated plates and grew optimally, with a doubling time of 30 and 32 h in low-concentration PRP (0.5%) medium supplemented with 5 ng/ml fibroblast growth factor-2 (FGF-2) on the F/P NP-coated plates. These cells maintained their multilineage potential for differentiation into adipocytes or osteoblasts. Furthermore, ASCs and BMSCs grew well in medium without PRP and FGF-2 on F/P NP-coated plates pretreated with PRP and FGF-2 in a concentration-dependent manner. Thus, F/P NP-coated plates are a useful substratum for the adherence and proliferation of ASCs and BMSCs in low-concentration PRP medium supplemented with FGF-2. No xenogeneic serum is required.

Attenuation of limb loss in an experimentally induced hindlimb ischemic model by fibroblast growth factor-2/fragmin/protamine microparticles as a delivery system.

Fibroblast growth factor-2 (FGF-2) is a well-characterized protein that is used in the treatment of healing-impaired wounds. We previously reported that fragmin/protamine microparticles (F/P MPs) are useful as biodegradable carriers for the controlled release of cytokines. We examined the ability of FGF-2-containing (FGF-2/) F/P MPs to prevent limb loss in an experimentally induced ischemic hindlimb model using adult Balb/c-nu/nu male mice. One day after inducing ischemia, intramuscular injections of 100 µL of FGF-2/F/P MPs turbid suspension (10 µg/mL FGF-2 and 6 mg/mL F/P MPs) were administered into eight sites of the ischemic hindlimb. A 100-µL suspension of each of the following-10 µg/mL FGF-2, 6 mg/mL F/P MPs, and phosphate-buffered saline (PBS; the control)-was similarly injected into the hindlimb. From 5 days onward after the injections, recovery from ischemia was observed in the FGF-2/F/P MP-treated group, but only partial recovery occurred in the FGF-2-treated group. The F/P MP-treated and PBS-treated groups (i.e., control) exhibited no recovery from the ischemia. The histological evaluations of the hindlimbs also confirmed that the capillary (i.e., mature vessels) density was significantly higher in the FGF-2/F/P MP-treated group than in the other groups. The mice injected with FGF-2/F/P MPs also recovered hindlimb blood flow, as reflected by oxygen saturation and surface temperature evaluation. Our present approach using FGF-2/F/P MPs could be considered a valuable option for the therapeutic treatment of peripheral ischemic diseases.

PRP&F/P MPs improved survival of dorsal paired pedicle skin flaps in rats.

BACKGROUND: Skin flap necrosis is a problem encountered postoperatively. The purpose of this study was to evaluate the effects of platelet-rich plasma containing fragmin/protamine microparticles (PRP&F/P MPs) on viability in a rat dorsal paired pedicle skin (DPPS) flap. MATERIALS AND METHODS: Two symmetrical adjoining rectangular flaps (8 × 2 cm each) were drawn on the rat dorsum. Two days after PRP&F/P MPs-, PRP-, F/P MPs-, and saline (control)-injections (n = 8 each), flaps were elevated as a random pattern flap without the lateral thoracic, posterior intercostal, and deep circumflex iliac vessels. The flaps were immediately sutured back and the flap survival area was measured 7 d after flap elevation. RESULTS: The flap survival rate in PRP&F/P MPs-injected groups (73.1% ± 4.2%) was significantly higher than those in PRP (64.9% ± 4.0%), F/P MPs (59.4 ± 4.5%), and control (61.2% ± 4.2%) groups. Histologic observation of the flaps showed survived thick granulation tissue and neovascularization in PRP&F/P MPs-injected groups. CONCLUSIONS: When PRP&F/P MPs are administered 2 d before the flap elevation, the improved flap survivals are observed. The pre-injection of PRP&F/P MPs may thus represent a promising treatment to prevent skin flap necrosis in reconstructive surgery.

Efficacy of oral colon-specific delivery capsule of low-molecular-weight heparin on ulcerative colitis.

Low-molecular-weight heparin has the potential for the treatment of ulcerative colitis, and targeted drug delivery to the colon is important for topical treatment of this disease, so low-molecular-weight heparin oral colon-specific delivery capsule was prepared, and the in vitro and in vivo drug release behavior was investigated. The macroscopical and histological scoring systems, wet colon mass index and myeloperoxidase activity were assessed to evaluate the efficacy of the capsule after administered orally to experimental colitis mice. Serum levels, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and a link factor of blood coagulation and inflammation factor Xa (FXa) were assayed by enzyme-linked immunosorbent assay. The expression of Musashi-1 (as an intestinal stem cell marker) in the colons was assessed by immunohistochemical analysis. The in vitro and in vivo drug release studies clearly indicated that the specific coated capsules were capable of protecting low-molecular-weight heparin from releasing in stomach and small intestine, while specifically delivering at colon. The oral colon-specific delivery capsule of low-molecular-weight heparin could attenuate macroscopic and histological features of colitis. The results showed that low-molecular-weight heparin oral colon-specific delivery capsule significantly decreased the serum levels of TNF-α, IL-6 as well as FXa, while increased the expression of Musashi-1 in colon compared with acetic acid-induced ulcerative colitis model group. The results showed that low-molecular-weight heparin oral colon-specific delivery capsule had the potential for treatment of inflammatory bowel disease.

Dalteparin, a low-molecular-weight heparin, promotes angiogenesis mediated by heparin-binding VEGF-A in vivo.

Tumors are angiogenesis dependent and vascular endothelial growth factor-A (VEGF-A), a heparin-binding protein, is a key angiogenic factor. As chemotherapy and co-treatment with anticoagulant low-molecular-weight heparin (LMWH) are common in cancer patients, we investigated whether angiogenesis in vivo mediated by VEGF-A is modulated by metronomic-type treatment with: (i) the LMWH dalteparin; (ii) low-dosage cytostatic epirubicin; or (iii) a combination of these two drugs. Using the quantitative rat mesentery angiogenesis assay, in which angiogenesis was induced by intraperitoneal injection of very low doses of VEGF, dalteparin sodium (Fragmin(®) ) and epirubicin (Farmorubicin(®) ) were administered separately or in combination by continuous subcutaneous infusion at a constant rate for 14 consecutive days. Dalteparin was administered at 27, 80, or 240 IU/kg/day, i.e., doses that reflect the clinical usage of this drug, while epirubicin was given at the well-tolerated dosage of 0.4 mg/kg/day. While dalteparin significantly stimulated angiogenesis in an inversely dose-dependent manner, epirubicin did not significantly affect angiogenesis. However, concurrent treatment with dalteparin and epirubicin significantly inhibited angiogenesis. The effect of dalteparin is the first demonstration of a proangiogenic effect of any LMWH in vivo. The fact that co-treatment with dalteparin and epirubicin significantly inhibited angiogenesis suggests a complex drug effect.

Weight-adjusted dalteparin for prevention of vascular thromboembolism in advanced pancreatic cancer patients decreases serum tissue factor and serum-mediated induction of cancer cell invasion.

The aim of the present study was to assess the role of tissue factor and serum-induced cell invasion in patients with advanced pancreatic cancer (APC). A cohort of 39 patients with APC, without thrombosis, receiving chemotherapy, were entered in a randomized controlled trial (ISRCTN = 76464767) of thromboprevention with weight-adjusted dalteparin (WAD). A total of 19 patients received WAD, the remaining 20 acting as a control group. Serum from baseline and week 8 was analysed for circulating-tissue factor antigen using ELISA. Circulating-tissue factor antigen rose from 324 pg/ml, [interquartile range (IQR) 282-347 pg/ml] to 356 pg/ml, (IQR 319-431 pg/ml) in controls (C), and decreased in the dalteparin-treated group (D) from 336 pg/ml (IQR 281-346 pg/ml) to 303 pg/ml (IQR 274-339 pg/ml). The difference in median percentage change between D and C was statistically significant [-4.0 (D) vs. 4.7 (C); P = 0.005, n = 39]. Serum-induced cellular invasion of MIA-Paca-2 cells in response to patient serum was studied using a Boyden chamber assay in 30 patients (14 WAD and 16 C). The median percentage change between C and D was significant [+54.9 (C) vs. -21.9 (D) P = 0.025, n = 30]. There was a weak correlation between BB-tissue factor reduction and cellular invasion reduction (Spearman) [0.384 (P = 0.037, n = 30)]. APC patients treated with WAD have lower tissue factor antigen levels and attenuated induction of cellular invasion in their blood. These assays may provide useful markers to guide appropriate dalteparin (and other low-molecular weight heparin) dosing schedules to optimize anticancer effects of dalteparin in APC.

Low molecular weight heparin suppresses lymphatic endothelial cell proliferation induced by vascular endothelial growth factor C in vitro.

BACKGROUND: Pancreatic cancer is one of the most aggressive human malignancies. Lymphangiogenesis plays an important role in lymph node metastasis of many solid tumors. It is well known that low molecular weight heparins (LMWHs) can inhibit cell growth, cell invasion and angiogenesis, which are key processes in tumor progression. METHODS: We measured the expression of vascular endothelial growth factor C (VEGF-C) in pancreatic cancer cells (PANC-1) using reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting. We used an in vitro assay to evaluate the anti-lymphangiogenic effect of an LMWH, Fragmin, on human lymphatic endothelial cell (HLEC) proliferation. RESULTS: Fragmin at a low concentration can effectively inhibits HLEC proliferation induced by VEGF-C. VEGF-C secreted by PANC-1 cells stimulated HLEC proliferation. Low concentration LMWH suppressed HLEC proliferation induced by VEGF-C but did not affect proliferation or VEGF-C expression of PANC-1 cells, whereas high concentrations of LMWH inhibited PANC-1 cell proliferation. CONCLUSIONS: These results suggest that VEGF-C released by cancer cells plays an important role in promoting HLEC proliferation. The LMWH Fragmin has anti-lymphangiogenic effects and may inhibit lymphatic metastasis in pancreatic cancer.

Short- and long-term efficacy of aspirin and clopidogrel for thromboprophylaxis for mechanical heart valves: an in vivo study in swine.

OBJECTIVE: In the interest of exploring alternatives to warfarin, we tested the hypothesis that clopidogrel combined with aspirin is effective for thromboprophylaxis of mechanical valves using a swine model. METHODS: Adult swine underwent heterotopic implantation of a modified bileaflet mechanical valved conduit bypassing the ligated, native descending thoracic aorta. Animals were randomized to no anticoagulation (n = 7), 175 U/kg dalteparin administered subcutaneously twice daily (n = 9), 325 mg of aspirin (n = 6), 75 mg of clopidogrel (n = 6), or 325 mg of aspirin and 75 mg of clopidogrel daily (n = 6) and survived for 30 days. Additionally, 11 animals were randomized to no anticoagulation (n = 5) or 325 mg of oral aspirin and 75 mg of clopidogrel daily (n = 6) and survived for 150 days. RESULTS: At 30 days, we observed 216 +/- 270 mg of thrombus for the no anticoagulation group, 53 +/- 91 mg for the dalteparin group, 33 +/- 23 mg for the aspirin group, 25 +/- 10 mg for the clopidogrel group, and 17 +/- 9 mg for the combined aspirin and clopidogrel group, respectively (P < .01 for clopidogrel and aspirin vs no anticoagulation). At 150 days, we observed 223 +/- 200 mg of thrombus for the no anticoagulation group and 4 +/- 4 mg for the aspirin and clopidogrel group (P = .02). Mean platelet deposition on the valve was 4.1 x 10(9) +/- 3.6 x 10(9) for the no anticoagulation and 6.81 x 10(7) +/- 1.4 x 10(8) for the combined aspirin and clopidogrel groups, respectively (P = .03). No major hemorrhagic events were observed. CONCLUSIONS: Effective short- and long-term thromboprophylaxis of mechanical valves can be achieved by using dual-antiplatelet therapy in this porcine model. Prospective human trials should be conducted with combination aspirin and clopidogrel as an alternative to warfarin in patients with bileaflet mechanical aortic valves.