Aerobic exercise and its impact on musculoskeletal pain in older adults: a 14 year prospective,longitudinal study

We studied the long term impact of running and other aerobic exercise on musculoskeletal pain in a cohort of healthy aging male and female seniors who had been followed for 14 years. We conducted a prospective, longitudinal study in 866 Runners' Association members (n = 492) and community controls (n = 374). Subjects were also categorized as Ever-Runners (n = 565) and Never-Runners (n = 301) to include runners who had stopped running. Pain was the primary outcome measure and was assessed in annual surveys on a double-anchored visual analogue scale (0 to 100; 0 = no pain). Baseline differences between Runners' Association members and community controls and between Ever-Runners versus Never-Runners were compared using chi-square and t-tests. Statistical adjustments for age, body mass index (BMI), gender, health behaviors, history of arthritis and comorbid conditions were performed using generalized estimating equations. Runner's Association members were younger (62 versus 65 years, p < 0.05), had a lower BMI (22.9 versus 24.2, p < 0.05), and less arthritis (35% versus 41%, p > 0.05) than community controls. Runners' Association members averaged far more exercise minutes per week (314 versus 123, p < 0.05) and miles run per week (26 versus 2, p < 0.05) and tended to report more fractures (53% versus 47%, p > 0.05) than controls. Ever-Runners were younger (62 versus 66 years, p < 0.05), had lower BMI (23.0 versus 24.3, p < 0.05), and less arthritis (35% versus 43%, p < 0.05) than Never-Runners. Ever-Runners averaged more exercise minutes per week (291 versus 120, p < 0.05) and miles run per week (23 versus 1, p < 0.05) and reported a few more fractures (52% versus 48%, p > 0.05) than Never-Runners. Exercise was associated with significantly lower pain scores over time in the Runners' Association group after adjusting for gender, baseline BMI, and study attrition (p < 0.01). Similar differences were observed for Ever-Runners versus Never-Runners. Consistent exercise patterns over the long term in physically active seniors are associated with about 25% less musculoskeletal pain than reported by more sedentary controls, either by calendar year or by cumulative area-under-the-curve pain over average ages of 62 to 76 years.     

Dynamic imaging of protease activity with fluorescently quenched activity-based probes

Protease activity is tightly regulated in both normal and disease conditions. However, it is often difficult to monitor the dynamic nature of this regulation in the context of a live cell or whole organism. To address this limitation, we developed a series of quenched activity-based probes (qABPs) that become fluorescent upon activity-dependent covalent modification of a protease target. These reagents freely penetrate cells and allow direct imaging of protease activity in living cells. Targeted proteases are directly identified and monitored biochemically by virtue of the resulting covalent tag, thereby allowing unambiguous assignment of protease activities observed in imaging studies. We report here the design and synthesis of a selective, cell-permeable qABP for the study of papain-family cysteine proteases. This probe is used to monitor real-time protease activity in live human cells with fluorescence microscopy techniques as well as standard biochemical methods.     

Regulation of gap junctions by tyrosine protein kinases

Most of the gap junction proteins are regulated in part by post-translational phosphorylation. Phosphorylation has been shown to be important in gap junction assembly and turnover, and for channel function in the resting state. Connexin phosphorylation may be altered by the activation of intracellular signaling pathways in response to growth factors, tumor promoters, activated oncogenes, hormones and inflammatory mediators. In some instances altered phosphorylation has been associated with changes in connexin function and in other cases appears to be associated with changes in the levels of the connexin protein and/or mRNA. This review focuses on the role of tyrosine protein kinases in the regulation of gap junctions. The literature is most extensive for connexin43 and those studies are reviewed here. A great deal has been learned in recent years about how connexin43 is regulated by tyrosine kinase-dependent signaling pathways. These pathways are often complex and to some extent are cell type- and stimulus-dependent. Although considerable progress has been made in unraveling the cellular pathways that regulate connexin function, significant challenges remain to be addressed in identifying additional phosphorylation sites and determining the stoichiometries of the phosphorylation events that regulate connexin function and it's interaction with other cellular proteins.     

Mutations Altering a Structurally Conserved Loop-Helix-Loop Region of a Viral Packaging Motor Change DNA Translocation Velocity and Processivity

Many double-stranded DNA viruses employ ATP-driven motors to translocate their genomes into small, preformed viral capsids against large forces resisting confinement. Here, we show via direct single-molecule measurements that a mutation T194M downstream of the Walker B motif in the phage λ gpA packaging motor causes an 8-fold reduction in translocation velocity without substantially changing processivity or force dependence, whereas the mutation G212S in the putative C (coupling) motif causes a 3-fold reduction in velocity and a 6-fold reduction in processivity. Meanwhile a T194M pseudorevertant (T194V) showed a near restoration of the wild-type dynamics. Structural comparisons and modeling show that these mutations are in a loop-helix-loop region that positions the key residues of the catalytic motifs, Walker B and C, in the ATPase center and is structurally homologous with analogous regions in chromosome transporters and SF2 RNA helicases. Together with recently published studies of SpoIIIE chromosome transporter and Ded1 RNA helicase mutants, these findings suggest the presence of a structurally conserved region that may be a part of the mechanism that determines motor velocity and processivity in several different types of nucleic acid translocases.     

Predicting transmembrane beta-barrels and interstrand residue interactions from sequence

Transmembrane beta-barrel (TMB) proteins are embedded in the outer membrane of Gram-negative bacteria, mitochondria, and chloroplasts. The cellular location and functional diversity of beta-barrel outer membrane proteins (omps) makes them an important protein class. At the present time, very few nonhomologous TMB structures have been determined by X-ray diffraction because of the experimental difficulty encountered in crystallizing transmembrane proteins. A novel method using pairwise interstrand residue statistical potentials derived from globular (nonouter membrane) proteins is introduced to predict the supersecondary structure of transmembrane beta-barrel proteins. The algorithm transFold employs a generalized hidden Markov model (i.e., multitape S-attribute grammar) to describe potential beta-barrel supersecondary structures and then computes by dynamic programming the minimum free energy beta-barrel structure. Hence, the approach can be viewed as a "wrapping" component that may capture folding processes with an initiation stage followed by progressive interaction of the sequence with the already-formed motifs. This approach differs significantly from others, which use traditional machine learning to solve this problem, because it does not require a training phase on known TMB structures and is the first to explicitly capture and predict long-range interactions. TransFold outperforms previous programs for predicting TMBs on smaller (相似文献   

CD40 expression by microglial cells is required for their completion of a two-step activation process during central nervous system autoimmune inflammation

Microglial cells are monocytic lineage cells that reside in the CNS and have the capacity to become activated during various pathological conditions. Although it was demonstrated that activation of microglial cells could be achieved in vitro by the engagement of CD40-CD40L interactions in combination with proinflammatory cytokines, the exact factors that mediate activation of microglial cells in vivo during CNS autoimmunity are ill-defined. To investigate the role of CD40 in microglial cell activation during experimental autoimmune encephalomyelitis (EAE), we used bone marrow chimera mice that allowed us to distinguish microglial cells from peripheral macrophages and render microglial cells deficient in CD40. We found that the first step of microglial cell activation was CD40-independent and occurred during EAE onset. The first step of activation consisted of microglial cell proliferation and up-regulation of the activation markers MHC class II, CD40, and CD86. At the peak of disease, microglial cells underwent a second step of activation, which was characterized by a further enhancement in activation marker expression along with a reduction in proliferation. The second step of microglial cell activation was CD40-dependent and the failure of CD40-deficient microglial cells to achieve a full level of activation during EAE was correlated with reduced expansion of encephalitogenic T cells and leukocyte infiltration in the CNS, and amelioration of clinical symptoms. Thus, our findings demonstrate that CD40 expression on microglial cells is necessary to complete their activation process during EAE, which is important for disease progression.     

Identification of genes involved in the biosynthesis and attachment of Methanococcus voltae N-linked glycans: insight into N-linked glycosylation pathways in Archaea

N-linked glycosylation is recognized as an important post-translational modification across all three domains of life. However, the understanding of the genetic pathways for the assembly and attachment of N-linked glycans in eukaryotic and bacterial systems far outweighs the knowledge of comparable processes in Archaea. The recent characterization of a novel trisaccharide [beta-ManpNAcA6Thr-(1-4)-beta-GlcpNAc3NAcA-(1-3)-beta-GlcpNAc]N-linked to asparagine residues in Methanococcus voltae flagellin and S-layer proteins affords new opportunities to investigate N-linked glycosylation pathways in Archaea. In this contribution, the insertional inactivation of several candidate genes within the M. voltae genome and their resulting effects on flagellin and S-layer glycosylation are reported. Two of the candidate genes were shown to have effects on flagellin and S-layer protein molecular mass and N-linked glycan structure. Further examination revealed inactivation of either of these two genes also had effects on flagella assembly. These genes, designated agl (archaeal glycosylation) genes, include a glycosyl transferase (aglA) involved in the attachment of the terminal sugar to the glycan and an STT3 oligosaccharyl transferase homologue (aglB) involved in the transfer of the complete glycan to the flagellin and S-layer proteins. These findings document the first experimental evidence for genes involved in any glycosylation process within the domain Archaea.