Canine olfactory ensheathing cells from the olfactory mucosa can be engineered to produce active chondroitinase ABC

A multitude of factors must be overcome following spinal cord injury (SCI) in order to achieve clinical improvement in patients. It is thought that by combining promising therapies these diverse factors could be combatted with the aim of producing an overall improvement in function. Chondroitin sulphate proteoglycans (CSPGs) present in the glial scar that forms following SCI present a significant block to axon regeneration. Digestion of CSPGs by chondroitinase ABC (ChABC) leads to axon regeneration, neuronal plasticity and functional improvement in preclinical models of SCI. However, the enzyme activity decays at body temperature within 24–72 h, limiting the translational potential of ChABC as a therapy. Olfactory ensheathing cells (OECs) have shown huge promise as a cell transplant therapy in SCI. Their beneficial effects have been demonstrated in multiple small animal SCI models as well as in naturally occurring SCI in canine patients. In the present study, we have genetically modified canine OECs from the mucosa to constitutively produce enzymatically active ChABC. We have developed a lentiviral vector that can deliver a mammalian modified version of the ChABC gene to mammalian cells, including OECs. Enzyme production was quantified using the Morgan-Elson assay that detects the breakdown products of CSPG digestion in cell supernatants. We confirmed our findings by immunolabelling cell supernatant samples using Western blotting. OECs normal cell function was unaffected by genetic modification as demonstrated by normal microscopic morphology and the presence of the low affinity neurotrophin receptor (p75NGF) following viral transduction. We have developed the means to allow production of active ChABC in combination with a promising cell transplant therapy for SCI repair

Connexin 36 Expression Regulates Neuronal Differentiation from Neural Progenitor Cells

Background: Gap junction communication has been shown in glial and neuronal cells and it is thought they mediate interand intra-cellular communication. Connexin 36 (Cx36) is expressed extensively in the developing brain, with levels peaking at P14 after which its levels fall and its expression becomes entirely neuronal. These and other data have led to the hypothesis that Cx36 may direct neuronal coupling and neurogenesis during development. Methodology/Principal Findings: To investigate Cx36 function we used a neurosphere model of neuronal cell development and developed lentiviral Cx36 knockdown and overexpression strategies. Cx36 knockdown was confirmed by western blotting, immunocytochemistry and functionally by fluorescence recovery after photobleaching (FRAP). We found that knockdown of Cx36 in neurosphere neuronal precursors significantly reduced neuronal coupling and the number of differentiated neurons. Correspondingly, the lentiviral mediated overexpression of Cx36 significantly increased the number of neurons derived from the transduced neurospheres. The number of oligodendrocytes was also significantly increased following transduction with Cx36 indicating they may support neuronal differentiation. Conclusions/Significance: Our data suggests that astrocytic and neuronal differentiation during development are governed by mechanisms that include the differential expression of Cx36

The different molecular forms of urine neutrophil gelatinase-associated lipocalin present in dogs with urinary diseases

Neutrophil gelatinase-associated lipocalin (NGAL) is a useful biomarker for the early prediction of renal diseases. NGAL may exist as monomer, dimer and/or NGAL/MMP-9 complex forms in humans. In this study, the existence of various forms of NGAL in urine (uNGAL) was determined and whether these forms are related to the different urinary diseases found in dogs is further discussed

Assessment of Heavy Metals in Water, Sediment, and Fishes of a Large Tropical Hydroelectric Dam in Sarawak, Malaysia

Bakun Hydroelectric Dam in Sarawak is one of the world highest concrete rock filled dams.This paper reports the heavy metals concentrations in water, sediment, and fishes of Bakun Dam.Water and sediment samples were collected from11 stations and 6 fish species were caught. The samples were digested with open acid digestion and the metals contents were analysed using an atomic absorption spectrophotometer and mercury analyser. The method was validated based on certified reference materials. A higher concentration of Fe andMn was detected in downstreamwater with significant longitudinal variation. Cu, Zn, andHg were present in trace amount. All elements analysed were consistently found in sediment with no risk of contamination. For fish, Hemibagrus planiceps was characterised by higher affinity for Hg accumulation. The concentrations detected in all fish species were within the permissible guideline of 0.5mg/kg. The health risk assessment suggested that Barbonymus schwanenfeldii, Puntioplites waandersii, Cyclocheilichthys apogon, and Hemibagrus planiceps were characterised by hazard index > 1 implying possible adverse effects.The amount of fish recommended for adults and children was in the range of 500–775 g/week and 33–135 g/week, respectively

Abnormal scaffold attachment factor 1 expression and localisation in spinocerebellar ataxias and huntington’s chorea

SAFB1 is a DNA and RNA binding protein that is highly expressed in the cerebellum and hippocampus and is involved in the processing of coding and non‐coding RNAs, splicing and dendritic function. We analyzed SAFB1 expression in the post‐mortem brain tissue of spinocerebellar ataxia (SCA), Huntington’s disease (HD), Multiple sclerosis (MS), Parkinson’s disease patients and controls. In SCA cases, the expression of SAFB1 in the nucleus was increased and there was abnormal and extensive expression in the cytoplasm where it co‐localized with the markers of Purkinje cell injury. Significantly, no SAFB1 expression was found in the cerebellar neurons of the dentate nucleus in control or MS patients; however, in SCA patients, SAFB1 expression was increased significantly in both the nucleus and cytoplasm of dentate neurons. In HD, we found that SAFB1 expression was increased in the nucleus and cytoplasm of striatal neurons; however, there was no SAFB1 staining in the striatal neurons of controls. In PD substantia nigra, we did not see any changes in neuronal SAFB1 expression. iCLIP analysis found that SAFB1 crosslink sites within ATXN1 RNA were adjacent to the start and within the glutamine repeat sequence. Further investigation found increased binding of SAFB1 to pathogenic ATXN1‐85Q mRNA. These novel data strongly suggest SAFB1 contributes to the etiology of SCA and Huntington’s chorea and that it may be a pathological marker of polyglutamine repeat expansion diseases

Separate elements of episodic memory subserved by distinct hippocampal-prefrontal connections

Episodic memory formation depends on information about a stimulus being integrated within a precise spatial and temporal context, a process dependent on the hippocampus and prefrontal cortex. Investigations of putative functional interactions between these regions are complicated by multiple direct and indirect hippocampal–prefrontal connections. Here application of a pharmacogenetic deactivation technique enabled us to investigate the mnemonic contributions of two direct hippocampal–medial prefrontal cortex (mPFC) pathways, one arising in the dorsal CA1 (dCA1) and the other in the intermediate CA1 (iCA1). While deactivation of either pathway impaired episodic memory, the resulting pattern of mnemonic deficits was different: deactivation of the dCA1→mPFC pathway selectively disrupted temporal order judgments while iCA1→mPFC pathway deactivation disrupted spatial memory. These findings reveal a previously unsuspected division of function among CA1 neurons that project directly to the mPFC. Such subnetworks may enable the distinctiveness of contextual information to be maintained in an episodic memory circuit

The effectiveness of dietary approaches to stop hypertension (DASH) counselling on estimated 10-year cardiovascular risk among patients with newly diagnosed grade 1 hypertension : a randomised clinical trial

The Dietary Approaches to Stop Hypertension (DASH) has been shown to lower blood pressure in the West. However, the real-life impact of DASH on reducing cardiovascular (CV) risk in routine clinical setting has not been studied. Methods A parallel-group, open-labelled, physician-blinded, randomised controlled trial was conducted in January–June 2013 and followed up for 6- and 12-months in primary care settings in Hong Kong. Patients newly diagnosed with grade 1 hypertension (aged 40–70 years) who had no concomitant medical conditions requiring dietary modifications were consecutively recruited. Subjects were randomised to standard education (usual care) (n = 275), or usual care plus dietitian-delivered DASH-based dietary counselling in a single one-to-one session (intervention) (n = 281). Primary outcomes were the changes in estimated 10-year CV risk. Results Outcome data were available for 504 (90.6%) and 485 (87.2%) patients at 6 and 12 months, respectively. There was no difference in the reduction of 10-year CV risk between the two groups at 6 months (−0.13%, 95% confidence interval [95% CI] −0.50% to 0.23%, p = 0.477) and 12 months (−0.08%, 95% CI −0.33% to 0.18%, p = 0.568). Multivariate regression analyses showed that male subjects, younger patients, current smokers, subjects with lower educational level, and those who dined out for main meals for ≥4 times in a typical week were significantly associated with no improvements in CV risk. Conclusions The findings may not support automatic referral of newly diagnosed grade 1 hypertensive patients for further one-to-one dietitian counselling on top of primary care physician's usual care. Patients with those risk factors identified should receive more clinical attention to reduce their CV risk

Transplantation of canine olfactory ensheathing cells producing chondroitinase ABC promotes chondroitin sulphate proteoglycan digestion and axonal sprouting following spinal cord injury

Olfactory ensheathing cell (OEC) transplantation is a promising strategy for treating spinal cord injury (SCI), as has been demonstrated in experimental SCI models and naturally occurring SCI in dogs. However, the presence of chondroitin sulphate proteoglycans within the extracellular matrix of the glial scar can inhibit efficient axonal repair and limit the therapeutic potential of OECs. Here we have used lentiviral vectors to genetically modify canine OECs to continuously deliver mammalian chondroitinase ABC at the lesion site in order to degrade the inhibitory chondroitin sulphate proteoglycans in a rodent model of spinal cord injury. We demonstrate that these chondroitinase producing canine OECs survived at 4 weeks following transplantation into the spinal cord lesion and effectively digested chondroitin sulphate proteoglycans at the site of injury. There was evidence of sprouting within the corticospinal tract rostral to the lesion and an increase in the number of corticospinal axons caudal to the lesion, suggestive of axonal regeneration. Our results indicate that delivery of the chondroitinase enzyme can be achieved with the genetically modified OECs to increase axon growth following SCI. The combination of these two promising approaches is a potential strategy for promoting neural regeneration following SCI in veterinary practice and human patients