Traditional Animal Models Research Articles

Modelling of feather pecking behavior in beak-trimmed and non-beak-trimmed crossbred laying hens: Variance component and trait-based approach

Because of a ban on the use of beak trimming in some European countries, feather pecking is becoming a substantial problem in the layer industry, both from animal welfare and economic points of view. The feather condition score (FCS) as a measure of feather damage has been shown to be closely related to feather pecking behavior in laying hens housed in groups. To obtain a better understanding of genetic and other biological mechanisms underlying feather pecking behavior, data on FCS of a population of 2,724 female offspring from crossing 50 male W1 and 907 female WB purebred lines were used. The offspring of 25 sires were beak-trimmed, and the offspring of another 25 sires were non-beak-trimmed. Titers of plasma natural antibody (NAb) isotypes IgM and IgG binding keyhole limpet hemocyanin at 24 wk of age were measured. Feather condition was scored at 53 wk of age. In the first part of the present study, we estimated genetic parameters for FCS with 2 variance components models: a traditional linear animal model and a model combining direct and associative genetic effects. In the second part of the present study, a trait-based analysis for FCS was conducted to investigate whether NAb isotype titers can explain variation in FCS among individuals, by fitting a linear mixed model. Though the estimated associative genetic variance was substantial, associative effects for FCS were not statistically significant in both populations (P = 0.09 in beak-trimmed birds, and P = 0.08 in non-beak-trimmed birds). This suggests an insufficient number of records on FCS. Individual's NAb isotypes titers did not show direct effect for FCS of itself, but individual's IgG titers showed a suggestive effect on the FCS of cage mates (associative effect) in beak-trimmed laying hens, which need further confirmation.

Development and characterization of a Yucatan miniature biomedical pig permanent middle cerebral artery occlusion stroke model

BackgroundEfforts to develop stroke treatments have met with limited success despite an intense need to produce novel treatments. The failed translation of many of these therapies in clinical trials has lead to a close examination of the therapeutic development process. One of the major factors believed to be limiting effective screening of these treatments is the absence of an animal model more predictive of human responses to treatments. The pig may potentially fill this gap with a gyrencephalic brain that is larger in size with a more similar gray-white matter composition to humans than traditional stroke animal models. In this study we develop and characterize a novel pig middle cerebral artery occlusion (MCAO) ischemic stroke model.MethodsEleven male pigs underwent MCAO surgery with the first 4 landrace pigs utilized to optimize stroke procedure and 7 additional Yucatan stroked pigs studied over a 90 day period. MRI analysis was done at 24 hrs and 90 days and included T2w, T2w FLAIR, T1w FLAIR and DWI sequences and associated ADC maps. Pigs were sacrificed at 90 days and underwent gross and microscopic histological evaluation. Significance in quantitative changes was determined by two-way analysis of variance and post-hoc Tukey’s Pair-Wise comparisons.ResultsMRI analysis of animals that underwent MCAO surgery at 24 hrs had hyperintense regions in T2w and DWI images with corresponding ADC maps having hypointense regions indicating cytotoxic edema consistent with an ischemic stroke. At 90 days, region of interest analysis of T1 FLAIR and ADC maps had an average lesion size of 59.17 cc, a loss of 8% brain matter. Histological examination of pig brains showed atrophy and loss of tissue, consistent with MRI, as well as glial scar formation and macrophage infiltration.ConclusionsThe MCAO procedure led to significant and consistent strokes with high survivability. These results suggest that the pig model is potentially a robust system for the study of stroke pathophysiology and potential diagnostics and therapeutics.

Potential of Birds to Serve as a Pathology-Free Model of Type 2 Diabetes, Part 1: Is the Apparent Absence of theRAGEGene a Factor in the Resistance of Avian Organisms to Chronic Hyperglycemia?

Diabetes mellitus is a global pandemic that accounts for ever-increasing rates of morbidity and mortality and consumes a growing share of national health care budgets. In spite of concerted efforts, a solution to this problem has not yet been found. One reason for this situation is lack of good animal models. Such models have been used successfully in many areas of biomedical research, but they have proven less than satisfactory in studies on diabetic complications. In this article, we propose to supplement traditional animal models of diabetes that use longitudinal, prospective studies of sick animals (mammals) with retrospective/comparative investigations of healthy animals (birds). Avians are promising models for such studies because they live healthy lives with chronic hyperglycemia that would be fatal to humans. We outline the advantages of the new perspective and show how, by implementing this approach, we observed that birds appear to be missing an important gene linked to diabetic complications. The protein encoded by this gene is a receptor for advanced glycation end products (RAGEs). Although the absence of RAGEs from birds has yet to be confirmed at the protein level, other differences between humans and birds may also be important in accounting for the ability of birds to live with chronic hyperglycemia. Two such additional such characteristics are currently being explored, and it is probable that more will emerge in time. We believe that the proposed perspective may improve the understanding of diabetes mellitus and may help in developing new means for controlling and preventing diabetic complications.

Genomic evaluation of both purebred and crossbred performances

BackgroundFor a two-breed crossbreeding system, Wei and van der Werf presented a model for genetic evaluation using information from both purebred and crossbred animals. The model provides breeding values for both purebred and crossbred performances. Genomic evaluation incorporates marker genotypes into a genetic evaluation system. Among popular methods are the so-called single-step methods, in which marker genotypes are incorporated into a traditional animal model by using a combined relationship matrix that extends the marker-based relationship matrix to non-genotyped animals. However, a single-step method for genomic evaluation of both purebred and crossbred performances has not been developed yet.ResultsAn extension of the Wei and van der Werf model that incorporates genomic information is presented. The extension consists of four steps: (1) the Wei van der Werf model is reformulated using two partial relationship matrices for the two breeds; (2) marker-based partial relationship matrices are constructed; (3) marker-based partial relationship matrices are adjusted to be compatible to pedigree-based partial relationship matrices and (4) combined partial relationship matrices are constructed using information from both pedigree and marker genotypes. The extension of the Wei van der Werf model can be implemented using software that allows inverse covariance matrices in sparse format as input.ConclusionsA method for genomic evaluation of both purebred and crossbred performances was developed for a two-breed crossbreeding system. The method allows information from crossbred animals to be incorporated in a coherent manner for such crossbreeding systems.

Intravital two-photon imaging to understand bacterial infections of the mammalian host.

Intravital two-photon microscopy (2PM) is an advanced fluorescence based imaging technique that allows for a cinematic study of physiological events occurring within tissues of the live animal. Based on this real-time imaging platform, the pathophysiology of bacterial infections can be studied in the most relevant of model systems-the live host. Whereas traditional animal models of host-pathogen interaction studies rely on end stage analysis of dissected tissues, noninvasive intravital imaging allows for real-time monitoring of infection during shorter or extended time frames. Here we describe the use of advanced surgical techniques for initiation of spatially and temporally well-controlled kidney infections in rats, and how the bacterial whereabouts can be studied while at the same time monitoring the host's altered tissue homeostasis based on real-time deep tissue imaging on the 2PM platform. Whereas this chapter focuses on pyelonephritis induced by uropathogenic Escherichia coli (UPEC) in rats, the major concepts can easily be translated to numerous infections in a variety of organs.

The ups and downs of modelling mood disorders in rodents.

The wide spectrum of disruptions that characterizes major depressive disorder (MDD) and bipolar disorder (BD) highlights the difficulties researchers are posed with as they try to mimic these disorders in the laboratory. Nonetheless, numerous attempts have been made to create rodent models of mood disorders or at least models of the symptoms of MDD and BD. Present antidepressants are all descendants of the serendipitous findings in the 1950s that the monoamine oxidase inhibitor iproniazid and the tricyclic antidepressant imipramine were effective antidepressants. Thus, the need for improved animal models to provide insights into the neuropathology underlying the disease is critical. Such information is in turn crucial for identifying new antidepressants and mood stabilisers. Currently, there is a shift away from traditional animal models to more focused research dealing with an endophenotype-style approach, genetic models, and incorporation of new findings from human neuroimaging and genetic studies. Such approaches are opening up more tractable avenues for understanding the neurobiological and genetic bases of these disorders. Further, such models promise to yield better translational animal models and hence more fruitful therapeutic targets. This overview focuses on such animal models and tests and how they can be used to assess MDD and BD in rodents.

P5-HT1A receptors in mood and anxiety: recent insights into autoreceptor versus heteroreceptor function

Serotonin (5-HT) neurotransmission is intimately linked to anxiety and depression and a diverse body of evidence supports the involvement of the main inhibitory serotonergic receptor, the serotonin-1A (5-HT(1A)) subtype, in both disorders. In this review, we examine the function of 5-HT(1A) receptor subpopulations and re-interpret our understanding of their role in mental illness in light of new data, separating both spatial (autoreceptor versus heteroreceptor) and the temporal (developmental versus adult) roles of the endogenous 5-HT(1A) receptors, emphasizing their distinct actions in mediating anxiety and depression-like behaviors. It is difficult to unambiguously distinguish the effects of different populations of the 5-HT(1A) receptors with traditional genetic animal models and pharmacological approaches. However, with the advent of novel genetic systems and subpopulation-selective pharmacological agents, direct evidence for the distinct roles of these populations in governing emotion-related behavior is emerging. There is strong and growing evidence for a functional dissociation between auto- and heteroreceptor populations in mediating anxiety and depressive-like behaviors, respectively. Furthermore, while it is well established that 5-HT(1A) receptors act developmentally to establish normal anxiety-like behaviors, the developmental role of 5-HT(1A) heteroreceptors is less clear, and the specific mechanisms underlying the developmental role of each subpopulation are likely to be key elements determining mood control in adult subjects.

Microdissection of Miniature Pig Ear

ObjectiveTo investigate the suitability of miniature pigs as an animal model for otological research. MethodsMicrodissection of the temporal bone was performed on 10 miniature pigs and recorded on photographs. ResultsThe morphology and measurement of the external, middle and inner ear and the lateral recess of the miniature pigs were obtained by microdissection. ConclusionCompared to traditional animal models, the miniature pig may be a better model for biomedical research because of its many similarities in physiological functions with humans. Similarities of the temporal bone structures, including the external, middle and inner ear and the lateral recess, between the miniature pig and human make the animal a potentially useful model for otological research.

Ionizing radiation induced biological effects in three-dimensional cell cultures

In vitro three-dimensional cell culture (TDCC) is a new cell culture model that owns its unique perspectives by mimicking features of the in vivo environment and is believed to have the potential to bridge the gap between traditional cell culture and animal models in biological and medicinal studies. Despite the generally harmful and potentially lethal effects, ionizing radiation also demonstrates health benefits in radiotherapy for cancers. This review mainly introduced the three-dimensional cell culture model and different biological effects induced by ionizing radiation, as well as their potential mechanism(s) as compared to traditional monolayer cultures.

The temporal bone microdissection of miniature pigs as a useful large animal model for otologic research

Conclusion: Compared with traditional animal models, the miniature pig may be a better model for biomedical research because its morphology has many similarities with that of humans. Objective: To investigate the suitability of the miniature pig as an animal model for otological research as regards morphology. Methods: Microdissection of the temporal bone of 10 miniature pigs was performed and recorded on photographs. Results: The morphology and measurements of the external, middle, and inner ear, and the lateral recess of the miniature pigs were completed by microdissection. The temporal bone structures, including the external, middle, inner ear, and the lateral recess, were similar in the miniature pig and humans.

Assessment of Juvenile Pigs to Serve as Human Pediatric Surrogates for Preclinical Formulation Pharmacokinetic Testing

Pediatric drug development is hampered by biological, clinical, and formulation challenges associated with age-based populations. A primary cause for this lack of development is the inability to accurately predict ontogenic changes that affect pharmacokinetics (PK) in children using traditional preclinical animal models. In response to this issue, our laboratory has conducted a proof-of-concept study to investigate the potential utility of juvenile pigs to serve as surrogates for children during preclinical PK testing of selected rifampin dosage forms. Pigs were surgically modified with jugular vein catheters that were externalized in the dorsal scapular region and connected to an automated blood sampling system (PigTurn-Culex-L). Commercially available rifampin capsules were administered to both 20 and 40kg pigs to determine relevant PK parameters. Orally disintegrating tablet formulations of rifampin were also developed and administered to 20kg pigs. Plasma samples were prepared from whole blood by centrifugation and analyzed for rifampin content by liquid chromatography-tandem mass spectrometry. Porcine PK parameters were determined from the resultant plasma-concentration time profiles and contrasted with published rifampin PK data in human adults and children. Results indicated significant similarities in dose-normalized absorption and elimination parameters between pigs and humans. Moreover, ontogenic changes observed in porcine PK parameters were consistent with ontogenic changes reported for human PK. These results demonstrate the potential utility of the juvenile porcine model for predicting human pediatric PK for rifampin. Furthermore, utilization of juvenile pigs during formulation testing may provide an alternative approach to expedite reformulation efforts during pediatric drug development.

Hopes and Challenges in using miRNAs as Translational Biomarkers for Drug-Induced Liver Injury

There is a need for better biomarkers of drug-induced liver injury (DILI) to guide risk assessment and patient management. Over the past 3 years, both animal and clinical studies have provided proof-of-concept data showing that a subset of miRNAs appear to offer unique advantages over the conventional DILI biomarkers, such as enhanced sensitivity and specificity, reduced inter-individual variations, the potential to differentiate lethal and nonlethal liver injury, and the ability to reflect the patterns and even the etiology of liver injury. Notably, many studies have demonstrated that level of miR-122, a liver-enriched miRNA accounting for approximately 70% of total hepatic miRNAs, was increased many fold in the blood when DILI occurred. However, currently available data are predominantly based on animal models and not human samples. Due to the lack of a standard quantification method for miRNAs and confirmatory studies using a comprehensive list of drugs and patients, the true value of all reported miRNA biomarkers remains to be carefully assessed. An outstanding challenge is to examine if miRNAs are also useful for idiosyncratic DILI, which constitutes the major part of clinical DILI cases but generally cannot be recapitulated in traditional animal models or in clinical trials (the latter due to its relative rarity).

The Openness is There.

The Openness is There.

Zebrafish embryos as an alternative model for screening of drug-induced organ toxicity

Zebrafish have initially emerged as model in developmental biology. The ease of maintenance, high number of offspring, transparency of the eggs and its suitability in mutagenicity screens to unravel developmental pathways contributed to its popularity. Today, it can be considered as one of the best characterised vertebrate models and more than 1,000 labs worldwide use the zebrafish (Strahle et al. 2012). At around 1990 zebrafish were discovered also as a toxicology model (Gorge and Nagel 1990), and the popularity is still increasing (Busch et al. 2011). A major reason for the attractiveness of this small teleost is associated with the suitability of its embryonic life stages. Early life stages are considered to sentinel no or less pain or discomfort when exposed to chemicals. Hence, according to European regulation (EU 2010), they are accepted as alternatives to animal experiments (Embry et al. 2010; Halder et al. 2010). Furthermore, the zebrafish embryo model has the principal capacity of high-throughput analysis and can be used in, for example, screenings for low toxicity of drug candidates. Zebrafish embryos provide an enormous versatility of applications in both environmental and human hazard assessment ranging from acute systemic toxicity (Ali et al. 2011; Lammer et al. 2009; Padilla et al. 2012), chronic toxicity (Volz et al. 2011), teratogenicity (Gustafson et al. 2012; Selderslaghs et al. 2009), neuractivity/neurotoxicity (Kokel et al. 2010; Selderslaghs et al. 2010), and endocrine disruption (Brion et al. 2012; Thienpont et al. 2011) to specific organ toxicities (Berghmans et al. 2008; Parng et al. 2002). As outlined by the paper of Driessen et al. in this issue of Archives in toxicology, hepatotoxicity is one of the major concerns of organ toxicity in drug development. Identification of hepatoxicity as early as possible in the drug development pipeline would reduce the costs associated with development of new medicines. Traditional animal models are time and cost intensive and subject to ethical concerns. Genuine cell-based in vitro methods do not provide the complexity of animal models and may provide only limited predictivity. Therefore, the zebrafish embryo has been proposed also as a better model to identify organ toxicity. As shown by Driessen et al., zebrafish embryos can indeed be used to identify hepatotoxicity. However, it is important to translate the observations appropriately, since histopathological effects or individual gene responses may differ among vertebrates. Identification of key events and conserved mechanisms can be supported by toxicogenomic analysis, and hence, it is not surprising that the number of studies in zebrafish embryos supported or accompanied by toxicogenomic analysis is increasing (e.g. Buttner et al. 2012; Hermsen et al. 2012; Pelayo et al. 2012; Schiller et al. 2012; Yang et al. 2007). Transcriptional profiling of whole embryos is even able to reveal organ-specific profiles—as shown in the Driessen paper in this issue and by various other studies (e.g. Kluver et al. 2011; Yang et al. 2007). With the advent of next generation sequencing (NGS), new perspectives for transcriptome profiling by RNA-sequencing (RNA-seq) are provided also for the zebrafish embryo model (Aanes et al. 2011; Vesterlund et al. 2011). With RNA-seq, novel transcripts from annotated or non-annotated regions not available in predefined probe sets, alternative splicing forms and rare transcripts can be detected. This could be important for the comparative analysis and identification of conserved key toxicity pathways in vertebrates. At present, S. Scholz (&) Department of Bioanalytical Ecotoxicology, UFZ Helmholtz Centre for Environmental Research, Permoserstr. 15, 04318 Leipzig, Germany e-mail: Stefan.Scholz@ufz.de

Noninvasive bioluminescence imaging of dengue virus infection in the brain of A129 mice

Dengue virus (DENV) infection is one of the most important public health threats globally; however, no vaccines or effective antivirals are currently available. The bioluminescence imaging technique has emerged as a powerful tool for studies on viral pathogenesis in vitro and in vivo. In this study, using a recombinant DENV that stably expressed Renilla luciferase (Rluc-DENV), we used bioluminescence for imaging of DENV infection in the brain of A129 mice that lacked type I interferon receptors. Upon intracranial inoculation with Rluc-DENV, A129 mice developed typical neurological symptoms and rapidly succumbed to viral infection. Real-time bioluminescence intensity analysis revealed the replication kinetics of Rluc-DENV in the brain of A129 mice. Linear regression analyses showed a good correlation between photon flux and viral titers (R(2) = 0.9923). Finally, the bioluminescence model was validated using a known mouse monoclonal antibody, 2A10G6, and the therapeutic effects of this neutralizing antibody were readily monitored by live imaging in the same animal. The noninvasive bioluminescence imaging of DENV infection as described here shows distinct advantages over traditional animal models and provides a powerful tool for potential antiviral or vaccine assays against DENV infection in vivo.

Time-lapse ultrashort pulse microscopy of infection in three-dimensional versus two-dimensional culture environments reveals enhanced extra-chromosomal virus replication compartment formation

The mechanisms that enable viruses to harness cellular machinery for their own survival are primarily studied in cell lines cultured in two-dimensional (2-D) environments. However, there are increasing reports of biological differences between cells cultured in 2-D versus three-dimensional (3-D) environments. Here we report differences in host-virus interactions based on differences in culture environment. Using ultrashort pulse microscopy (UPM), a form of two-photon microscopy that utilizes sub-10-fs pulses to efficiently excite fluorophores, we have shown that de novo development of extra-chromosomal virus replication compartments (VRCs) upon murine cytomegalovirus (mCMV) infection is markedly enhanced when host cells are cultured in 3-D collagen gels versus 2-D monolayers. In addition, time-lapse imaging revealed that mCMV-induced VRCs have the capacity to grow by coalescence. This work supports the future potential of 3-D culture as a useful bridge between traditional monolayer cultures and animal models to study host-virus interactions in a more physiologically relevant environment for the development of effective anti-viral therapeutics. These advances will require broader adoption of modalities, such as UPM, to image deep within scattering tissues.

Contributions of animal models to the study of mood disorders

Mood disorders are a leading cause of morbidity and mortality, yet their underlying pathophysiology remains unclear. Animal models serve as a powerful tool for investigating the neurobiological mechanisms underlying psychiatric disorders; however, no animal model developed to date can fully mimic the "corresponding" human psychiatric disorder. In this scenario, the development of different animal models contributes to our understanding of the neurobiology of these disorders and provides the possibility of preclinical pharmacologic screening. The present review seeks to provide a comprehensive overview of traditional and recent animal models, recapitulating different features and the possible pathologic mechanisms of mood disorders emulated by these models.

The role of the anterior, mediodorsal, and parafascicular thalamus in instrumental conditioning

The traditional animal model of instrumental behavior has focused almost exclusively on structures within the cortico-striatal network and ignored the contributions of various thalamic nuclei despite large and specific connections with each of these structures. One possible reason for this is that the thalamus has been conventionally viewed as a mediator of general processes, such as attention, arousal and movement, that are not easily separated from more cognitive aspects of instrumental behavior. Recent research has, however, begun to separate these roles. Here we review the role of three thalamic nuclei in instrumental conditioning: the anterior thalamic nuclei (ANT), the mediodorsal (MD), and parafascicular thalamic nuclei (PF). Early research suggested that ANT might regulate aspects of instrumental behavior but, on review, we suggest that the types of tasks used in these studies were more likely to recruit Pavlovian processes. Indeed lesions of ANT have been found to have no effect on performance in instrumental free-operant tasks. By contrast the mediodorsal thalamus (MD) has been found to play a specific and important role in the acquisition of goal-directed action. We propose this role is related to its connections with prelimbic cortex (PL) and present new data that directly implicates this circuit in the acquisition of goal-directed actions. Finally we review evidence suggesting the PF, although not critical for the acquisition or performance of instrumental actions, plays a specific role in regulating action flexibility.

Evolution of Tumor Model: From Animal Model of Tumor to Tumor Model in Animal

Patient derived xenograft (PDX) is defined as a growth of patients’ tumor in the xenograft setting. The evolution of cancer model in animal has a century old history. The most single reason that exerted the pressure on the traditional animal model of cancer to evolve to PDX is that the traditional models have not delivered as expected and traditional models have not predicted clinical success. In spite of well above 50 drugs developed and approved for oncology over the last several decades, there remains a nirking paucity of clinical success as a reminder that this war on cancer riding on the animal model is far from won. In a backbreaking attempt to analyze the failure, the limitation of the “model” system appeared to be the most rational cause of this shortcoming. It was more of a failure to test a drug rather than a failure to make a drug that stunted our collective growth and success in cancer research. PDX is the product of this age-old failure and its fitness is currently tested in virtually all organ-type solid tumors. This review will present and appraise PDX model in the context of its evolution, its future promise, its limitations and more specifically, the current content of PDX in different solid tumors including breast, lung, colorectal, prostrate, GBM, pancreatic, hepatocellular carcinoma and melanoma.

Comparative Antithrombotic and Bleeding Effects of Two U.S. Generic Enoxaparins

Abstract 1172 Background:Low molecular weight heparins (LMWHs) are complex biologic drugs that exhibit heterogeneity in terms of saccharide chain length and in the composition (sulfate, acetyl), content, and location of functional groups. Such heterogeneity impacts the biologic activity of LMWHs as there is a certain threshold chain length required for thrombin inhibitory activity and a particular sequence is required for interaction with antithrombin. In July 2010 the US Food and Drug Administration published requirements necessary to demonstrate the ‘sameness' of generic LMWHs with the originator LMWH. We undertook this study to compare the primary anticoagulant effect of thrombosis prevention and the primary adverse effect of bleeding of two FDA approved generic enoxaparins. Methods:Four batches of commercially available Sandoz US generic enoxaparin (Princeton, NJ) and two batches of Watson US generic enoxaparin (Parsippany, NJ) were compared. All products were obtained from hospital pharmacies as pre-filled syringes containing 40 mg of the drugs. The molecular weight profile of each batch was determined by HPLC in relation to well-defined heparin fractions and by utilizing the US Pharmacopeia method. In vitro activity was determined by supplementing each LMWH batch to normal human plasma over a range of concentrations (0–10 mg/ml) and analyzing these samples using aPTT, anti-FXa and anti-FIIa assays. Hemorrhagic activity was measured using a rat tail transection model five minutes after administration of a 2 mg/kg intravenous dose (n=8 rats/batch). Upon completion of the bleeding model, antithrombotic activity was assessed using a jugular vein clamping model (∼90 minutes post-dosing). Blood samples collected from treated rats were used to estimate circulating blood levels of LMWH using anti-FXa and anti-FIIa assays. Results:The two groups of generic enoxaparins exhibited a similar molecular weight profile with mean molecular weights of 4,270 ± 20 Da for the Sandoz products and 4,420 ± 80 Da for the Watson products. In vitro activities were similar between batches of the same product, but the individual products differed considerably (p=0.01). In the aPTT and anti-FIIa assays, the Watson LMWH produced significantly more activity than the Sandoz LMWH at concentrations ≥5 μg/ml. At 5 μg/ml, a clotting time of 74.0 ± 16.6 sec was observed with Watson batches compared to clotting times ranging from 52.9 to 54.9 sec for Sandoz LMWHs. While a similar pattern was observed with the anti-FXa assay, the differences between products were not statistically significant. In the bleeding model, all LMWHs prolonged the bleeding time compared to vehicle control. One batch of the Watson LMWH, however, produced a significantly longer bleeding time compared to all other samples tested (33.8 ± 5.1 min vs. a range of 11.9 ± 2.4 to 18.3 ± 4.3 minutes for the other samples; p=0.003). The same batch produced significantly more antithrombotic activity (6.3 ± 0.7 clampings) compared to the other samples (range of 4.0 ± 0.6 to 4.6 ± 0.5 clampings; p<0.001). One batch of Sandoz LMWH produced a significantly smaller prolongation of bleeding time compared to other samples. Circulating drug levels determined by anti-FXa and anti-FIIa activities were comparable in all treatment groups but did not appear to correlate with hemorrhagic or antithrombotic activities. Conclusion:The expectation was that all batches of all generic enoxaparins would produce the same in vitro, in vivo, and ex vivo outcomes. The findings of this study suggest that incorporation of traditional animal models in the development of generic enoxaparins may be of value as multiple biological effects of LMWHs, not fully addressed with the anti-FXa and anti-FIIa activities, contribute to the overall antithrombotic activity of these LMWHs. This study demonstrates that all generic enoxaparins available today may not necessarily be the same. These findings underscore the importance of in vivo studies in animal models to demonstrate the bioequivalence of the generic products. Disclosures:Jeske:Sanofi-Aventis, Paris, France: Research Funding. Walenga:Sanofi-Aventis, Paris, France: Research Funding. Escalante:Sanofi-Aventis, Paris, France: Research Funding. Hoppensteadt:Sanofi-Aventis, Paris, France: Research Funding. Cunanan:Sanofi-Aventis, Paris, France: Research Funding. Kahn:Sanofi-Aventis, Paris, France: Research Funding. Paulus:Sanofi-Aventis, Paris, France: Research Funding. Fareed:Sanofi-Aventis, Paris, France: Research Funding. Bakhos:Sanofi-Aventis, Paris, France: Research Funding.