Characterization of human and rodent native and recombinant adenosine A2B receptors by radioligand binding studies

Adenosine A2B receptors of native human and rodent cell lines were investigated using [3H]PSB-298 [(8-{4-[2-(2-hydroxyethylamino)-2-oxoethoxy]phenyl}-1-propylxanthine] in radioligand binding studies. [3H]PSB-298 showed saturable and reversible binding. It exhibited a KD value of 60 ± 1 nM and limited capacity (Bmax = 3.511 fmol per milligram protein) at recombinant human adenosine A2B receptors expressed in human embryonic kidney cells (HEK-293). The addition of sodium chloride (100 mM) led to a threefold increase in the number of binding sites recognized by the radioligand. The curve of the agonist 5′-N-ethylcarboxamidoadenosine (NECA) was shifted to the right in the presence of NaCl, while the curve of the antagonist PSB-298 was shifted to the left, indicating that PSB-298 may be an inverse agonist at A2B receptors. Adenosine A2B receptors were shown to be the major adenosine A2 receptor subtype on the mouse neuroblastoma x rat glioma hybrid cell line NG108-15 cells. Binding studies at rat INS-1 cells (insulin secreting cell line) demonstrated that [3H]PSB-298 is a selective radioligand for adenosine A2B binding sites in this cell line

Regeneration of Soft Tissues Is Promoted by MMP1 Treatment after Digit Amputation in Mice

The ratio of matrix metalloproteinases (MMPs) to the tissue inhibitors of metalloproteinases (TIMPs) in wounded tissues strictly control the protease activity of MMPs, and therefore regulate the progress of wound closure, tissue regeneration and scar formation. Some amphibians (i.e. axolotl/newt) demonstrate complete regeneration of missing or wounded digits and even limbs; MMPs play a critical role during amphibian regeneration. Conversely, mammalian wound healing re-establishes tissue integrity, but at the expense of scar tissue formation. The differences between amphibian regeneration and mammalian wound healing can be attributed to the greater ratio of MMPs to TIMPs in amphibian tissue. Previous studies have demonstrated the ability of MMP1 to effectively promote skeletal muscle regeneration by favoring extracellular matrix (ECM) remodeling to enhance cell proliferation and migration. In this study, MMP1 was administered to the digits amputated at the mid-second phalanx of adult mice to observe its effect on digit regeneration. Results indicated that the regeneration of soft tissue and the rate of wound closure were significantly improved by MMP1 administration, but the elongation of the skeletal tissue was insignificantly affected. During digit regeneration, more mutipotent progenitor cells, capillary vasculature and neuromuscular-related tissues were observed in MMP1 treated tissues; moreover, there was less fibrotic tissue formed in treated digits. In summary, MMP1 was found to be effective in promoting wound healing in amputated digits of adult mice. © 2013 Mu et al

Identification of Allele-Specific RNAi Effectors Targeting Genetic Forms of Parkinson's Disease

Parkinson's disease (PD) is a progressive neurological disorder affecting an estimated 5–10 million people worldwide. Recent evidence has implicated several genes that directly cause or increase susceptibility to PD. As well as advancing understanding of the genetic aetiology of PD these findings suggest new ways to modify the disease course, in some cases through genetic manipulation. Here we generated a ‘walk-through’ series of RNA Pol III-expressed shRNAs targeting both the α-synuclein A30P and LRRK2 G2019S PD-associated mutations. Allele-specific discrimination of the α-synuclein A30P mutation was achieved with alignments at position 10, 13 and 14 in two model systems, including a heterozygous model mimicking the disease setting, whilst 5′RACE was used to confirm stated alignments. Discrimination of the most common PD-linked LRRK2 G2019S mutation was assessed in hemizygous dual-luciferase assays and showed that alignment of the mutation opposite position 4 of the antisense species produced robust discrimination of alleles at all time points studied. Discrimination at this position was subsequently confirmed using siRNAs, where up to 10-fold discrimination was seen. The results suggest that RNAi-mediated silencing of PD-associated autosomal dominant genes could be a novel therapeutic approach for the treatment of the relevant clinical cases of PD in future

Radical genome remodelling accompanied the emergence of a novel host-restricted bacterial pathogen

The emergence of new pathogens is a major threat to public and veterinary health. Changes in bacterial habitat such as a switch in host or disease tropism are typically accompanied by genetic diversification. Staphylococcus aureus is a multi-host bacterial species associated with human and livestock infections. A microaerophilic subspecies, Staphylococcus aureus subsp. anaerobius, is responsible for Morel’s disease, a lymphadenitis restricted to sheep and goats. However, the evolutionary history of S. aureus subsp. anaerobius and its relatedness to S. aureus are unknown. Population genomic analyses of clinical S. aureus subsp. anaerobius isolates revealed a highly conserved clone that descended from a S. aureus progenitor about 1000 years ago before differentiating into distinct lineages that contain African and European isolates. S. aureus subsp. anaerobius has undergone limited clonal expansion, with a restricted population size, and an evolutionary rate 10-fold slower than S. aureus. The transition to its current restricted ecological niche involved acquisition of a pathogenicity island encoding a ruminant host-specific effector of abscess formation, large chromosomal re-arrangements, and the accumulation of at least 205 pseudogenes, resulting in a highly fastidious metabolism. Importantly, expansion of ~87 insertion sequences (IS) located largely in intergenic regions provided distinct mechanisms for the control of expression of flanking genes, including a novel mechanism associated with IS-mediated anti-anti-sense decoupling of ancestral gene repression. Our findings reveal the remarkable evolutionary trajectory of a host-restricted bacterial pathogen that resulted from extensive remodelling of the S. aureus genome through an array of diverse mechanisms in parallel

Modular protein-RNA interactions regulating mRNA metabolism: a role for NMR

Here we review the role played by transient interactions between multi-functional proteins and their RNA targets in the regulation of mRNA metabolism, and we describe the important function of NMR spectroscopy in the study of these systems. We place emphasis on a general approach for the study of different features of modular multi-domain recognition that uses well-established NMR techniques and that has provided important advances in the general understanding of post-transcriptional regulation

Bacterial endosymbiont Cardinium cSfur genome sequence provides insights for understanding the symbiotic relationship in Sogatella furcifera host

Background: Sogatella furcifera is a migratory pest that damages rice plants and causes severe economic losses. Due to its ability to annually migrate long distances, S.furcifera has emerged as a major pest of rice in several Asian countries. Symbiotic relationships of inherited bacteria with terrestrial arthropods have significant implications. The genus Cardinium is present in many types of arthropods, where it influences some host characteristics. We present a report of a newly # identified strain of the bacterial endosymbiont Cardinium cSfur in S. furcifera. Result: From the whole genome of S. furcifera previously sequenced by our laboratory, we assembled the whole genome sequence of Cardinium cSfur. The sequence comprised 1,103,593 bp with a GC content of 39.2%. The phylogenetic tree of the Bacteroides phylum to which Cardinium cSfur belongs suggests that Cardinium cSfur is closely related to the other strains (Cardinium cBtQ1 and cEper1) that are members of the Amoebophilaceae family. Genome comparison between the host-dependent endosymbiont including Cardinium cSfur and freeliving bacteria revealed that the endosymbiont has a smaller genome size and lower GC content, and has lost some genes related to metabolism because of its special environment, which is similar to the genome pattern observed in other insect symbionts. Cardinium cSfur has limited metabolic capability, which makes it less contributive to metabolic and biosynthetic processes in its host. From our findings, we inferred that, to compensate for its limited metabolic capability, Cardinium cSfur harbors a relatively high proportion of transport proteins, which might act as the hub between it and its host. With its acquisition of the whole operon related to biotin synthesis and glycolysis related genes through HGT event, Cardinium cSfur seems to be undergoing changes while establishing a symbiotic relationship with its host. Conclusion: A novel bacterial endosymbiont strain (Cardinium cSfur) has been discovered. A genomic analysis of the endosymbiont in S. furcifera suggests that its genome has undergone certain changes to facilitate its settlement in the host. The envisaged potential reproduction manipulative ability of the new endosymbiont strain in its S. furcifera host has vital implications in designing eco-friendly approaches to combat the insect pest

Constitutive modelling of skin ageing

The objective of this chapter is to review the main biomechanical and structural aspects associated with both intrinsic and extrinsic skin ageing, and to present potential research avenues to account for these effects in mathematical and computational models of the skin. This will be illustrated through recent work of the authors which provides a basis to those interested in developing mechanistic constitutive models capturing the mechanobiology of skin across the life course

Pulsed radiofrequency treatment in interventional pain management: mechanisms and potential indications—a review

Item does not contain fulltextBACKGROUND: The objective of this review is to evaluate the efficacy of Pulsed Radiofrequency (PRF) treatment in chronic pain management in randomized clinical trials (RCTs) and well-designed observational studies. The physics, mechanisms of action, and biological effects are discussed to provide the scientific basis for this promising modality. METHODS: We systematically searched for clinical studies on PRF. We searched the MEDLINE (PubMed) and EMBASE database, using the free text terms: pulsed radiofrequency, radio frequency, radiation, isothermal radiofrequency, and combination of these. We classified the information in two tables, one focusing only on RCTs, and another, containing prospective studies. Date of last electronic search was 30 May 2010. The methodological quality of the presented reports was scored using the original criteria proposed by Jadad et al. FINDINGS: We found six RCTs that evaluated the efficacy of PRF, one against corticosteroid injection, one against sham intervention, and the rest against conventional RF thermocoagulation. Two trials were conducted in patients with lower back pain due to lumbar zygapophyseal joint pain, one in cervical radicular pain, one in lumbosacral radicular pain, one in trigeminal neuralgia, and another in chronic shoulder pain. CONCLUSION: From the available evidence, the use of PRF to the dorsal root ganglion in cervical radicular pain is compelling. With regards to its lumbosacral counterpart, the use of PRF cannot be similarly advocated in view of the methodological quality of the included study. PRF application to the supracapular nerve was found to be as efficacious as intra-articular corticosteroid in patients with chronic shoulder pain. The use of PRF in lumbar facet arthropathy and trigeminal neuralgia was found to be less effective than conventional RF thermocoagulation techniques