Predictive and Reactive Grip Force Responses to Rapid Load Increases in People With Multiple Sclerosis.

OBJECTIVES: To determine the effects of multiple sclerosis (MS) on predictive and reactive grip force control in a catching task and on clinical tests of hand function. DESIGN: Case-control study with matched-pairs control group. SETTING: University prevention and rehabilitation center. PARTICIPANTS: Participants (N=30) consisted of people with multiple sclerosis (PwMS) (n=15) and healthy controls (n=15), matched for sex, age, and hand dominance. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Performance on the Expanded Disability Status Scale (EDSS), Nine-Hole Peg Test (9-HPT), Jebsen-Taylor Hand Function Test (JTHFT), and 2-point discrimination (2PD) was evaluated. To analyze grip force control, blindfolded subjects held a receptacle equipped with grip force and acceleration sensors in their hand. In a catching task, a weight was dropped from (1) the experimenter's hand unexpectedly into the receptacle (reactive force control); and (2) from the subject's opposite hand (predictive force control). Grip forces and time lags were analyzed. RESULTS: PwMS (mean EDSS ± SD, 4.2±1.86) had impairments in the 9-HPT and JTHFT (P<.001). The 2PD did not differ significantly between PwMS and controls. During reactive force control (catching task 1), PwMS showed significantly higher grip forces immediately after impact (P<.05), and a significant prolongation of the time from grip force increase until reaching the peak of grip force (P<.001). PwMS and controls did not differ during predictive force control (catching task 2; P>.1). CONCLUSIONS: Exaggerated grip force responses and alterations of timing after an unpredictable perturbation, combined with preserved grip force control during predictable conditions, is a characteristic pattern of fine motor control deficits in MS. Measures of reactive grip force responses may be used to complement neurologic assessments. Further studies exploring the usefulness of these measures should be performed in a broader community of PwMS.

A new assay format for NF-kappaB based on a DNA triple helix and a fluorescence resonance energy transfer.

Herein we report a feasibility study for a new concept to detect DNA binding protein NF-kappaB based on a DNA triple helix formation in combination with a fluorescence resonance energy transfer (FRET). The new principle avoids expensive antibodies and radioactivity and might have implications for assays of other DNA binding proteins.

Constitutive serum response factor activation by the viral chemokine receptor homologue pUS28 is differentially regulated by Galpha(q/11) and Galpha(16).

Expression of the human cytomegalovirus (HCMV)-encoded chemokine receptor homologue pUS28 in mammalian cells results in ligand-dependent and -independent changes in the activity of multiple cellular signal transduction pathways. The ligand-dependent signalling activity of pUS28 has been shown to be predominantly mediated by heterotrimeric G proteins of the G(i/o) and G(12/13) subfamilies. Ligand-independent constitutive activity of pUS28 causing stimulation of inositol phosphate formation has been correlated with the coupling of pUS28 to G proteins of the G(q) family. It is well known that activation of G(q) proteins by cell surface receptors is coupled to activation of the Rho GTPase RhoA. Activated RhoA regulates numerous cellular functions, including the activity of the transcription factor serum response factor (SRF). The marked activation of G(q) proteins by pUS28 in transfected and HCMV-infected cells prompted us to investigate its effect on SRF activity. The results presented herein demonstrate that expression of pUS28 in COS-7 cells caused a vigorous induction of SRF activity. This effect was observed in the absence of chemokines known to interact with pUS28, and was specifically mediated by endogenous G(q) and/or G(11) as well as RhoA and/or a closely related Rho GTPase. The stimulatory effect of pUS28 and Galpha(q/11) was independent of phospholipase C-beta (PLCbeta) activation and was markedly sensitive to inhibition by wild-type, but not by constitutively active Galpha(16), thus identifying Galpha(16) as a modulator of Galpha(q/11) function likely to act by competing with Galpha(q/11) for and thus uncoupling Galpha(q/11) from activation by pUS28.

Inhibition of GSK3 differentially modulates NF-kappaB, CREB, AP-1 and beta-catenin signaling in hepatocytes, but fails to promote TNF-alpha-induced apoptosis.

Glycogen synthase kinase-3 (GSK-3) is known to modulate cell survival and apoptosis through multiple intracellular signaling pathways. However, its hepatoprotective function and its role in activation of NF-kappaB and anti-apoptotic factors are poorly understood and remain controversial. Here we investigated whether inhibition of GSK-3 could induce apoptosis in the presence of TNF-alpha in primary mouse hepatocytes. We show that pharmacological inhibition of GSK-3 in primary mouse hepatocytes does not lead to TNF-alpha-induced apoptosis despite reduced NF-kappaB activity. Enhanced stability of IkappaB-alpha appears to be responsible for lower levels of nuclear NF-kappaB and hence reduced transactivation. Additionally, inhibition of GSK-3 was accompanied by marked upregulation of beta-catenin, AP-1, and CREB transcription factors. Stimulation of canonical Wnt signaling and CREB activity led to elevated levels of anti-apoptotic factors. Hence, survival of primary mouse hepatocytes may be caused by the activation and/or upregulation of other key regulators of liver homeostasis and regeneration. These signaling molecules may compensate for the compromised anti-apoptotic function of NF-kappaB and allow survival of hepatocytes in the presence of TNF-alpha and GSK-3 inhibition.

S100B modulates IL-6 release and cytotoxicity from hypothermic brain cells and inhibits hypothermia-induced axonal outgrowth.

Brain protection is essential during neonatal and pediatric cardiac surgery. Deep hypothermia is still the most important method for achieving neuroprotection during cardiopulmonary bypass. Previously, we could demonstrate that deep hypothermia induces substantial cytotoxicity in brain cells as well as increased release of the pro-inflammatory cytokine interleukin-6 (IL-6), which plays an important role in neuroprotection and neuroregeneration. Deep hypothermia is also associated with increased levels of the astrocytic protein S100B in the serum and cerebrospinal fluid of patients. Since S100B may modulate pro-inflammatory cytokines and may stimulate neurite outgrowth, we have tested the hypothesis that nanomolar concentrations of S100B may increase IL-6 release from brain cells and support axonal outgrowth from organotypic brain slices under hypothermic conditions. S100B administration substantially reduced neuronal and glial cytotoxicity under hypothermic conditions. In the presence of S100B hypothermia-induced IL-6 release in primary astrocytes was significantly increased but reduced in BV-2 microglial cells and primary neurons. Surprisingly, deep hypothermia increased axonal outgrowth from brain slices and--in contrast to our hypothesis--this hypothermia-induced neurite outgrowth was inhibited by S100B. These data suggest that S100B differentially influences cytokine release and cytotoxicity from distinct brain cells and may inhibit neuroregeneration by suppressing hypothermia-induced axonal outgrowth.

Methylprednisolone attenuates hypothermia- and rewarming-induced cytotoxicity and IL-6 release in isolated primary astrocytes, neurons and BV-2 microglia cells.

Brain protection is crucial during neonatal and pediatric cardiac surgery. The major methods for brain protection are the administration of steroids and deep hypothermia. Therefore, we have investigated the impact of methylprednisolone (MP) administration and deep hypothermia on neonatal mouse astrocytes, neurons and BV-2 microglia cells. Brain cells were pretreated with MP (100 mM) and incubated according to a deep hypothermia protocol mimicking temperature changes during cardiac surgery in children: deep hypothermia (2 h at 17 degrees C, phase 1), slow rewarming (2 h up to 37 degrees C, phase 2), and normothermia (20 h at 37 degrees C, phase 3). In all brain-related cell types cytotoxicity was investigated as well as the release of the pro-inflammatory cytokine interleukin-6 (IL-6), which plays a major role in neuroprotection and neuroregeneration. Deep hypothermia induces substantial cytotoxicity and the secretion of IL-6 by astrocytes, BV-2 microglia cells and neurons. MP administration has no influence on the cell survival and IL-6 release of normothermic astrocytes, BV-2 microglia cells and neurons, while hypothermia-induced cytotoxicity and IL-6 secretion are significantly suppressed by MP. These data suggest that MP increases cell survival after deep hypothermia but also suppresses important neuroprotective and regenerative processes induced by IL-6. Hence, more specific immune modulation than that provided by MP may be needed to protect the brain during neonatal and pediatric cardiac surgery.

Sesquiterpene lactones as inhibitors of IL-8 expression in HeLa cells.

Twenty-four structurally different SLs were studied for their inhibition on IL-8 production in HeLa229 cells and different IC50-values were obtained. QSAR analyses revealed that the alpha-methylene-gamma-lactone and the presence and reactivity of a second reaction center, expressed by LUMO2, are the most important descriptors for IL-8. Using two SLs as examples, we demonstrated that SLs prevent DNA binding of AP-1, which has binding sites in the IL-8 promoter together with NF-kappaB and C/EBP, and that this is probably due to directly targeting AP-1. p38 MAPK, which plays a role in AP-1 activation as well as in IL-8 regulation, was not influenced by SLs. These data show that NF-kappaB and AP-1, and consequently IL-8 may be interesting targets in antiinflammation research and that the small molecules of SLs may be powerful candidates with promising properties for therapeutic modulation of the inflammatory response.