Venoms can deleteriously affect any physiological system reachable by the bloodstream, including directly interfering with the coagulation cascade. Such coagulopathic toxins may be anticoagulants or procoagulants. Snake venoms are unique in their use of procoagulant toxins for predatory purposes. The boomslang (Dispholidus typus) and the twig snakes (Thelotornis species) are iconic African snakes belonging to the family Colubridae. Both species produce strikingly similar lethal procoagulant pathologies. Despite these similarities, antivenom is only produced for treating bites by D. typus, and the mechanisms of action of both venoms have been understudied. In this study, we investigated the venom of D. typus and T. mossambicanus utilising a range of proteomic and bioactivity approaches, including determining the procoagulant properties of both venoms in relation to the human coagulation pathways. In doing so, we developed a novel procoagulant assay, utilising a Stago STA-R Max analyser, to accurately detect real time clotting in plasma at varying concentrations of venom. This approach was used to assess the clotting capabilities of the two venoms both with and without calcium and phospholipid co-factors. We found that T. mossambicanus produced a significantly stronger coagulation response compared to D. typus. Functional enzyme assays showed that T. mossambicanus also exhibited a higher metalloprotease and phospholipase activity but had a much lower serine protease activity relative to D. typus venom. The neutralising capability of the available boomslang antivenom was also investigated on both species, with it being 11.3 times more effective upon D. typus venom than T. mossambicanus. In addition to being a faster clotting venom, T. mossambicanus was revealed to be a much more complex venom composition than D. typus. This is consistent with patterns seen for other snakes with venom complexity linked to dietary complexity. Consistent with the external morphological differences in head shape between the two species, CT and MRI analyses revealed significant internal structural differences in skull architecture and venom gland anatomy. This study increases our understanding of not only the biodiscovery potential of these medically important species but also increases our knowledge of the pathological relationship between venom and the human coagulation cascade.
Intellectual impairment is a strongly disabling feature of Down's syndrome, a genetic disorder of high prevalence (1 in 700-1000 live births) caused by trisomy of chromosome 21. Accumulating evidence shows that widespread neurogenesis impairment is a major determinant of abnormal brain development and, hence, of intellectual disability in Down's syndrome. This defect is worsened by dendritic hypotrophy and connectivity alterations. Most of the pharmacotherapies designed to improve cognitive performance in Down's syndrome have been attempted in Down's syndrome mouse models during adult life stages. Yet, as neurogenesis is mainly a prenatal event, treatments aimed at correcting neurogenesis failure in Down's syndrome should be administered during pregnancy. Correction of neurogenesis during the very first stages of brain formation may, in turn, rescue improper brain wiring. The aim of our study was to establish whether it is possible to rescue the neurodevelopmental alterations that characterize the trisomic brain with a prenatal pharmacotherapy with fluoxetine, a drug that is able to restore post-natal hippocampal neurogenesis in the Ts65Dn mouse model of Down's syndrome. Pregnant Ts65Dn females were treated with fluoxetine from embryonic Day 10 until delivery. On post-natal Day 2 the pups received an injection of 5-bromo-2-deoxyuridine and were sacrificed after either 2 h or after 43 days (at the age of 45 days). Untreated 2-day-old Ts65Dn mice exhibited a severe neurogenesis reduction and hypocellularity throughout the forebrain (subventricular zone, subgranular zone, neocortex, striatum, thalamus and hypothalamus), midbrain (mesencephalon) and hindbrain (cerebellum and pons). In embryonically treated 2-day-old Ts65Dn mice, precursor proliferation and cellularity were fully restored throughout all brain regions. The recovery of proliferation potency and cellularity was still present in treated Ts65Dn 45-day-old mice. Moreover, embryonic treatment restored dendritic development, cortical and hippocampal synapse development and brain volume. Importantly, these effects were accompanied by recovery of behavioural performance. The cognitive deficits caused by Down's syndrome have long been considered irreversible. The current study provides novel evidence that a pharmacotherapy with fluoxetine during embryonic development is able to fully rescue the abnormal brain development and behavioural deficits that are typical of Down's syndrome. If the positive effects of fluoxetine on the brain of a mouse model are replicated in foetuses with Down's syndrome, fluoxetine, a drug usable in humans, may represent a breakthrough for the therapy of intellectual disability in Down's syndrome.
deficiency and decreased adiposity are associated with induction of thermogenic gene expression in subcutaneous white adipose and brown adipose tissue. Am J Physiol Endocrinol Metab 308: E159-E171, 2015. First published November 25, 2014; doi:10.1152/ajpendo.00056.2014.-The Rarrelated orphan receptor-␣ (Ror␣) is a nuclear receptor that regulates adiposity and is a potential regulator of energy homeostasis. We have demonstrated that the Ror␣-deficient staggerer (sg/sg) mice display a lean and obesity-resistant phenotype. Adaptive Ucp1-dependent thermogenesis in beige/brite and brown adipose tissue serves as a mechanism to increase energy expenditure and resist obesity. DEXA and MRI analysis demonstrated significantly decreased total fat mass and fat/lean mass tissue ratio in male chow-fed sg/sg mice relative to wt mice. In addition, we observed increased Ucp1 expression in brown adipose and subcutaneous white adipose tissue but not in visceral adipose tissue from Ror␣-deficient mice. Moreover, this was associated with significant increases in the expression of the mRNAs encoding the thermogenic genes (i.e., markers of brown and beige adipose) Ppar␣, Err␣, Dio2, Acot11/Bfit, Cpt1, and Cidea in the subcutaneous adipose in the sg/sg relative to WT mice. These changes in thermogenic gene expression involved the significantly increased expression of the (cell-fate controlling) histone-lysine N-methyltransferase 1 (Ehmt1), which stabilizes the Prdm16 transcriptional complex. Moreover, primary brown adipocytes from sg/sg mice displayed a higher metabolic rate, and further analysis was consistent with increased uncoupling. Finally, core body temperature analysis and infrared thermography demonstrated that the sg/sg mice maintained greater thermal control and cold tolerance relative to the WT littermates. We suggest that enhanced Ucp1 and thermogenic gene expression/activity may be an important contributor to the lean, obesityresistant phenotype in Ror␣-deficient mice.retinoic acid receptor-related orphan receptor-␣; obesity; adiposity; adipose tissue; uncoupling protein-1 OBESITY IS A METABOLIC DISEASE that affects more than 10% of the global population. Energy imbalance as a result of excess dietary energy intake or genetic susceptibility results in increased energy storage and adiposity. Current strategies to ameliorate this disease include increasing energy expenditure through physical exercise and reducing energy consumption. However, the success rate is limited by compliance and the genetic disposition to maintain energy balance.Adaptive thermogenesis is an adrenergic/sympathetic-dependent mechanism utilized by mammals to increase energy expenditure in response to excessive dietary intake and/or cold stimulus. Thermogenesis occurs in both brown adipose tissue and beige fat [white adipose tissue cells that acquire a brown adipose phenotype (37, 41)]. This process involves increased (fatty acid-dependent) expression and activity of the thermogenic mitochondrial uncoupling protein (Ucp1). The function of Ucp1 is to me...
BackgroundAdvances in 3D shape capture technology have made powerful shape analyses, such as geometric morphometrics, more feasible. While the highly accurate micro-computed tomography (µCT) scanners have been the “gold standard,” recent improvements in 3D surface scanners may make this technology a faster, portable, and cost-effective alternative. Several studies have already compared the two devices but all use relatively large specimens such as human crania. Here we perform shape analyses on Australia’s smallest rodent to test whether a 3D scanner produces similar results to a µCT scanner.MethodsWe captured 19 delicate mouse (Pseudomys delicatulus) crania with a µCT scanner and a 3D scanner for geometric morphometrics. We ran multiple Procrustes ANOVAs to test how variation due to scan device compared to other sources such as biologically relevant variation and operator error. We quantified operator error as levels of variation and repeatability. Further, we tested if the two devices performed differently at classifying individuals based on sexual dimorphism. Finally, we inspected scatterplots of principal component analysis (PCA) scores for non-random patterns.ResultsIn all Procrustes ANOVAs, regardless of factors included, differences between individuals contributed the most to total variation. The PCA plots reflect this in how the individuals are dispersed. Including only the symmetric component of shape increased the biological signal relative to variation due to device and due to error. 3D scans showed a higher level of operator error as evidenced by a greater spread of their replicates on the PCA, a higher level of multivariate variation, and a lower repeatability score. However, the 3D scan and µCT scan datasets performed identically in classifying individuals based on intra-specific patterns of sexual dimorphism.DiscussionCompared to µCT scans, we find that even low resolution 3D scans of very small specimens are sufficiently accurate to classify intra-specific differences. We also make three recommendations for best use of low resolution data. First, we recommend that extreme caution should be taken when analyzing the asymmetric component of shape variation. Second, using 3D scans generates more random error due to increased landmarking difficulty, therefore users should be conservative in landmark choice and avoid multiple operators. Third, using 3D scans introduces a source of systematic error relative to µCT scans, therefore we recommend not combining them when possible, especially in studies expecting little biological variation. Our findings support increased use of low resolution 3D scans for most morphological studies; they are likely also applicable to low resolution scans of large specimens made in a medical CT scanner. As most vertebrates are relatively small, we anticipate our results will bolster more researchers in designing affordable large scale studies on small specimens with 3D surface scanners.
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