Plasticity in mammalian skeletal muscle.

While it has been recognized for many years that different limb muscles belonging to the same mammal may have markedly differing contractile characteristics, it is only comparatively recently that it has been demonstrated that these differences depend upon the motor innervation. By appropriately changing the peripheral nerve innervating a mammalian skeletal muscle, it is possible to change dramatically the contractile behaviour of the reinnervated muscle. The manner by which the motor innervation determines the nature of a muscle fibre's contractile machinery is not completely understood, but it appears that the number and pattern of motor nerve impulses reaching the muscle play an important role. The biochemical changes occurring within muscle fibres whose contractile properties have been modified by altered motor innervation include the synthesis of different contractile proteins.     

Structure and regulatory profile of the monkeypox inhibitor of complement: comparison to homologs in vaccinia and variola and evidence for dimer formation

The outbreak of monkeypox in the Unites States in the summer of 2003 was the first occurrence of this smallpox-like disease outside of Africa. This limited human epidemic resulted from cross-infection of prairie dogs by imported African rodents. Although there were no human fatalities, this outbreak illustrates that monkeypox is an emerging natural infection and a potential biological weapon. We characterized a virulence factor expressed by monkeypox (monkeypox inhibitor of complement enzymes or MOPICE). We also compared its structure and regulatory function to homologous complement regulatory proteins of variola (SPICE) and vaccinia (VCP). In multiple expression systems, 5-30% of MOPICE, SPICE, and VCP consisted of function-enhancing disulfide-linked homodimers. Mammalian cells infected with vaccinia virus also expressed VCP dimers. MOPICE bound human C3b/C4b intermediate to that of SPICE and VCP. Cofactor activity of MOPICE was similar to VCP, but both were approximately 100-fold less efficient than SPICE. SPICE and VCP, but not MOPICE, possessed decay-accelerating activity for the C3 and C5 convertases of the classical pathway. Additionally, all three regulators possessed heparin-binding capability. These studies demonstrate that MOPICE regulates human complement and suggest that dimerization is a prominent feature of these virulence factors. Thus, our data add novel information relative to the functional repertoire of these poxviral virulence factors. Furthermore, targeting and neutralizing these complement regulatory active sites via mAbs is a therapeutic approach that may enhance protection against smallpox.     

Application of immunoproteomics to rapid cytokine detection

Cytokines are pivotal to a balanced innate or cell-mediated immune response, and can be indicative of disease progression and/or resolution. Methods to measure key cytokines rapidly with accuracy, precision, and sensitivity are consequently important. The current assay technologies, which are based on RT-PCR, immunoassays, or bioassays, are limited in their use in the clinic, in particular because of the long time (1-3 h) required to carry out the assays. An alternative, semi-quantitative approach described here, uses an immunological capture step and a mass spectral readout. The goal of the assay is speed rather than sensitivity or precision.     

Deficiency in Th2 Cytokine Responses Exacerbate Orthopoxvirus Infection

Ectromelia virus (ECTV) causes mousepox in mice, a disease very similar to smallpox in humans. ECTV and variola virus (VARV), the agent of smallpox, are closely related orthopoxviruses. Mousepox is an excellent small animal model to study the genetic and immunologic basis for resistance and susceptibility of humans to smallpox. Resistance to mousepox is dependent on a strong polarized type 1 immune response, associated with robust natural killer (NK) cell, cytotoxic T lymphocyte (CTL) and gamma interferon (IFN-γ) responses. In contrast, ECTV-susceptible mice generate a type 2 response, associated with weak NK cell, CTL and IFN-γ responses but robust IL-4 responses. Nonetheless, susceptible strains infected with mutant ECTV lacking virus-encoded IFN-γ binding protein (vIFN-γbp) (ECTV-IFN-γbp^Δ) control virus replication through generation of type 1 response. Since the IL-4/IL-13/STAT-6 signaling pathways polarize type 2/T helper 2 (Th2) responses with a corresponding suppression of IFN-γ production, we investigated whether the combined absence of vIFN-γbp, and one or more host genes involved in Th2 response development, influence generation of protective immunity. Most mutant mouse strains infected with wild-type (WT) virus succumbed to disease more rapidly than WT animals. Conversely, the disease outcome was significantly improved in WT mice infected with ECTV-IFN-γbp^Δ but absence of IL-4/IL-13/STAT-6 signaling pathways did not provide any added advantage. Deficiency in IL-13 or STAT-6 resulted in defective CTL responses, higher mortality rates and accelerated deaths. Deficiencies in IL-4/IL-13/STAT-6 signaling pathways significantly reduced the numbers of IFN-γ producing CD4 and CD8 T cells, indicating an absence of a switch to a Th1-like response. Factors contributing to susceptibility or resistance to mousepox are far more complex than a balance between Th1 and Th2 responses.     

Disruption to the 5-HT<Subscript>7</Subscript> Receptor Following Hypoxia–Ischemia in the Immature Rodent Brain

It has become increasingly evident the serotonergic (5-hydroxytryptamine, 5-HT) system is an important central neuronal network disrupted following neonatal hypoxic–ischemic (HI) insults. Serotonin acts via a variety of receptor subtypes that are differentially associated with behavioural and cognitive mechanisms. The 5-HT₇ receptor is purported to play a key role in epilepsy, anxiety, learning and memory and neuropsychiatric disorders. Furthermore, the 5-HT₇ receptor is highly localized in brain regions damaged following neonatal HI insults. Utilising our well-established neonatal HI model in the postnatal day 3 (P3) rat pup we demonstrated a significant decrease in levels of the 5-HT₇ protein in the frontal cortex, thalamus and brainstem one week after insult. We also observed a relative decrease in both the cytosolic and membrane fractions of 5-HT₇. The 5-HT₇ receptor was detected on neurons throughout the cortex and thalamus, and 5-HT cell bodies in the brainstem. However we found no evidence of 5-HT₇ co-localisation on microglia or astrocytes. Moreover, minocycline treatment did not significantly prevent the HI-induced reductions in 5-HT₇. In conclusion, neonatal HI injury caused significant disruption to 5-HT₇ receptors in the forebrain and brainstem. Yet the use of minocycline to inhibit activated microglia, did not prevent the HI-induced changes in 5-HT₇ expression.