Treatment satisfaction with pharmaceutical interventions in Japanese adults with osteoarthritis and chronic knee pain: an analysis of a web-based survey.

PURPOSE: Patient satisfaction is an important outcome in successful osteoarthritis (OA) treatment. The aim of this study was to evaluate treatment satisfaction for medication (TSM) in people with knee OA (KOA), identify the factors predictive of treatment satisfaction, and describe the burden of illness. PATIENTS AND METHODS: This cross-sectional, patient-reported study used an Internet-based survey and analyzed responses of respondents with KOA (N=400) on characteristics including pain sites and levels (including pain ratings using the Numerical Rating Scale and Short-Form McGill Pain Questionnaire), treatment satisfaction (Global, Effectiveness, and Convenience scores) based on the Treatment Satisfaction Questionnaire for Medication (TSQM-9), and quality of life (QoL; based on the Arthritis Impact Measurement Scale 2-Short Form). Respondents with only KOA (n=237) were compared with those having KOA and additional painful sites (KOA+; n=163). Factors predicting TSM were identified using multivariable linear regression analyses. RESULTS: Respondents with KOA were more likely to report intermittent pain for 3 months or more compared with those with KOA+ (58.6% vs 48.5%, respectively; P=0.044), while those with KOA+ were more likely to report consistent pain for 3 months or more (P=0.022). Respondents with KOA+ also had more difficulty due to their knee pain while sleeping (P=0.022) and resting (P=0.015). Reported TSM did not differ significantly across KOA vs KOA+ groups, with both groups reporting low satisfaction; all domains of QoL were worse for those with KOA+. Knee pain reduction by medication predicted higher satisfaction across domains, while lower pre-medication pain and post-medication pain matching expectations predicted higher TSQM-9 Global and Effectiveness scores. CONCLUSION: Medication treatment satisfaction rates were low among Japanese respondents with KOA. Given that lower pain, greater pain reduction post-medication, and meeting pain management expectations were predictive of higher satisfaction, treatment strategies that can better address pain may prove beneficial for overall patient satisfaction.

Gene expression profiling and functional assays of activated hepatic stellate cells suggest that myocardin has a role in activation.

BACKGROUND: Myofibroblast-like cells derived from transdifferentiated hepatic stellate cells (HSC) play a central role in scar formation that leads to liver fibrosis. The molecular mechanisms underlying this process are not fully understood. AIM: Our aim was to identify genes that are differentially regulated by HSC activation and to explore their function. METHODS: Using oligonucleotide microarrays, we performed transcriptional analysis of the human HSC cell line, LI90, cultured on Matrigel. Microarray data were validated by quantitative real-time polymerase chain reaction and Western blotting. The function of myocardin was assessed by myocardin RNAi and overexpression. RESULTS: Examination of Matrigel-induced deactivation of LI90 cells revealed marked downregulation of myocardin, an important transcriptional regulator in smooth and cardiac muscle development. Small interfering RNA-mediated suppression of myocardin expression in both activated LI90 and rat activated HSC resulted in loss of the phenotypic characteristics of myofibroblasts and significantly impaired the production of activated HSC markers, such as alpha-smooth muscle actin and extracellular matrix proteins like type I collagen. Overexpression of myocardin led to the upregulation of these marker genes. Myocardin was upregulated in rat primary HSC during in vitro activation and in the fibrotic liver of a dimethylnitrosamine-induced fibrosis rat model. CONCLUSIONS: This study demonstrates that myocardin is involved in the activation of HSC; myocardin may serve as a novel therapeutic target in the treatment of liver fibrosis.

Selective CB1 cannabinoid receptor antagonist, SR141716A, attenuates liver injury induced by Concanavalin A.

AIM: The aim of this study was to investigate the hepatoprotective activity of a selective cannabinoid receptor 1 (CB1) antagonist, SR141716A, in a Concanavalin A (Con A)-induced mouse liver injury model and to determine whether SR141716A has an effect on the production of inflammatory cytokines and chemokines induced by Con A. RESULTS: Injection of Con A (20 mg/kg) to mice developed hepatitis determined by plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) elevation and necrosis in the liver. Pretreatment with SR141716A (30 mg/kg) significantly reduced plasma AST and ALT level, protected against necrosis in the liver, and significantly reduced plasma cytokine and chemokine levels, including TNFalpha, IFN-gamma, CXCL9, MIP1-alpha, and IL-10 and no change decreased in IL-4. CONCLUSIONS: The selective CB1 antagonist, SR141716A, exerts a hepatoprotective effect on Con A-induced liver injury in mice by attenuating the increase in cytokine and chemokine levels and inhibiting hepatocyte injury. These findings raise the possibility of using CB1 antagonists as anti-inflammatory drugs for treating hepatitis as well as other inflammatory diseases.

Apoptosis signal-regulating kinase 1 is crucial for oxidative stress-induced but not for osmotic stress-induced hepatocyte cell death.

AIMS: In this study, we investigated the involvement of apoptosis signal-regulating kinase 1 (ASK1) in oxidative stress and osmotic stress-induced hepatocyte death. MAIN METHODS: Activation of ASK1-JNK/p38 cascade and resulting cell death induced by oxidative and osmotic stress was investigated by Western immunoblot analysis and cell toxicity assay using human hepatoma cell lines, Huh7 expressing high level of ASK1 and HepG2 cells expressing low level of ASK1. Gene knock-down of ASK1 using shRNA against ASK1 was conducted using mouse hepatocyte cell line, AML12. KEY FINDINGS: Activation of ASK1-JNK/p38 cascade and cell death in Huh7 expressing high level of ASK1 was markedly induced by the oxidative stress. HepG2 expressing low level of ASK1 was resistant to oxidative stress while cell death induced by osmotic stress was comparable between Huh7 and HepG2 cells. Although the phosphorylation of ASK1 was not observed by osmotic stress, the phosphorylation of p38 and JNK and resulting cell death was induced in both cell lines. The phosphorylation of ASK1 and p38/JNK in the mouse primary hepatocyte were also increased by oxidative stress. Knock-down of ASK1 mRNA in AML12 in vitro significantly reduced oxidative stress-induced cell death, however, knock-down of ASK1 in cells did not affect the osmotic stress-induced cell death. SIGNIFICANCE: This study revealed that ASK1 regulates oxidative stress- but not osmotic stress-induced hepatocyte death, suggesting ASK1 plays a critical role in oxidative-stress induced hepatocyte death. These results raise the possibility that an ASK1 may be a promising therapeutic target for liver diseases caused by oxidative stress.

Utility of plasma circulating mRNA as a marker to detect hepatic injury.

Utility of plasma circulating mRNA as a molecular marker to detect hepatic injury was evaluated. Total RNA was isolated from plasma of the rat liver fibrosis models at various time points, and plasma circulating mRNAs of major liver-derived genes, albumin and haptoglobin, were measured by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). Sensitivity and kinetics of plasma circulating mRNA were compared with those of plasma alanine aminotransferase (ALT) activity. We have found that the measurement of plasma circulating mRNA is more sensitive than plasma ALT activity, and enables early detection of hepatic injury. The plasma circulating mRNA will serve as a novel and highly sensitive molecular marker for hepatic injury.

5-Nitro-2-(3-phenylpropylamino)benzoic acid is a GPR35 agonist.

GPR35 is a Gi/o- and G16-coupled receptor abundantly expressed in gastrointestinal tissues and immune cells. Kynurenic acid (a tryptophan metabolite and ionotropic glutamate receptor antagonist) and zaprinast (a phosphodiesterase inhibitor) are GPR35 agonists. Here, we show that the chloride channel blocker 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB) is also a GPR35 agonist. NPPB activates the GPR35-Gi/o and GPR35-G16 pathways in human embryonic kidney 293 (HEK293) cells and induces intracellular calcium mobilization in a concentration-dependent manner in HEK293 cells coexpressing human, rat or mouse GPR35 and the chimeric G protein G(qi5). These results suggest a novel pharmacological activity of NPPB and will provide useful information to search for more potent and selective GPR35 agonists.

Visualizing sensory transmission between dorsal root ganglion and dorsal horn neurons in co-culture with calcium imaging.

Sensory information is conveyed to the central nervous system by primary afferent neurons within dorsal root ganglia (DRG), which synapse onto neurons of the dorsal horn of the spinal cord. This synaptic connection is central to the processing of both sensory and pain stimuli. Here, we describe a model system to monitor synaptic transmission between DRG neurons and dorsal horn neurons that is compatible with high-throughput screening. This co-culture preparation comprises DRG and dorsal horn neurons and utilizes Ca(2+) imaging with the indicator dye Fura-2 to visualize synaptic transmission. Addition of capsaicin to co-cultures stimulated DRG neurons and led to activation of dorsal horn neurons as well as increased intracellular Ca(2+) concentrations. This effect was dose-dependent and absent when DRG neurons were omitted from the culture. NMDA receptors are a critical component of synapses between DRG and dorsal horn neurons as MK-801, a use-dependent non-competitive antagonist, prevented activation of dorsal horn neurons following capsaicin treatment. This model system allows for rapid and efficient analysis of noxious stimulus-evoked Ca(2+) signal transmission and provides a new approach both for investigating synaptic transmission in the spinal cord and for screening potential analgesic compounds.

Zaprinast, a well-known cyclic guanosine monophosphate-specific phosphodiesterase inhibitor, is an agonist for GPR35.

We found that zaprinast, a well-known cyclic guanosine monophosphate-specific phosphodiesterase inhibitor, acted as an agonist for a G protein-coupled receptor, GPR35. In our intracellular calcium mobilization assay, zaprinast activated rat GPR35 strongly (geometric mean EC(50) value of 16nM), whereas it activated human GPR35 moderately (geometric mean EC(50) value of 840nM). We also demonstrated that GPR35 acted as a Galpha(i/o)- and Galpha(16)-coupled receptor for zaprinast when heterologously expressed in human embryonic kidney 293 (HEK 293) cells. These findings will facilitate the research on GPR35 and the drug discovery of the GPR35 modulators.

Identification of neurons that express 5-hydroxytryptamine4 receptors in intestine.

5-Hydroxytryptamine (5-HT) is an endogenous stimulant of intestinal propulsive reflexes. It exerts its effects partly through 5-HT4 receptors; 5-HT4 receptor agonists that are stimulants of intestinal transit are in clinical use. Both pharmacological and recent immunohistochemical studies indicate that 5-HT4 receptors are present on enteric neurons but the specific neurons that express the receptors have not been determined. In the present work, we describe the characterization of an anti-5-HT4 receptor antiserum that reveals immunoreactivity for enteric neurons and other cell types in the gastrointestinal tract. With this antiserum, 5-HT4 receptor immunoreactivity has been found in the muscularis mucosae of the rat oesophagus, a standard assay tissue for 5-HT4 receptors. It is also present in the muscularis mucosae of the guinea-pig and mouse oesophagus. In guinea-pig small intestine and rat and mouse colon, 5-HT4 receptor immunoreactivity occurs in subpopulations of enteric neurons, including prominent large neurons. Double-staining has shown that these large neurons in the guinea-pig small intestine are also immunoreactive for two markers of intrinsic primary afferent neurons, cytoplasmic NeuN and calbindin. Some muscle motor neurons in the myenteric ganglia are immunoreactive for this receptor, whereas it is rarely expressed by secretomotor neurons. Immunoreactivity also occurs in the interstitial cells of Cajal but is faint in the external muscle. Expression of the protein and mRNA has been confirmed in extracts containing enteric neurons. The observations suggest that one site of action of 5-HT4 receptor agonists is the intrinsic primary afferent neurons.

Uncoupling and endocytosis of 5-hydroxytryptamine 4 receptors. Distinct molecular events with different GRK2 requirements.

The 5-hydroxytryptamine type 4 receptors (5-HT4Rs) are involved in memory, cognition, feeding, respiratory control, and gastrointestinal motility through activation of a G(s)/cAMP pathway. We have shown that 5-HT4R undergoes rapid and profound homologous uncoupling in neurons. However, no significant uncoupling was observed in COS-7 or HEK293 cells, which expressed either no or a weak concentration of GRK2, respectively. High expression of GRK2 in neurons is likely to be the reason for this difference because overexpression of GRK2 in COS-7 and HEK293 cells reproduced rapid and profound uncoupling of 5-HT4R. We have also shown, for the first time, that GRK2 requirements for uncoupling and endocytosis were very different. Indeed, beta-arrestin/dynamin-dependent endocytosis was observed in HEK293 cells without any need of GRK2 overexpression. In addition to this difference, uncoupling and beta-arrestin/dynamin-dependent endocytosis were mediated through distinct mechanisms. Neither uncoupling nor beta-arrestin/dynamin-dependent endocytosis required the serine and threonine residues localized within the specific C-terminal domains of the 5-HT4R splice variants. In contrast, a cluster of serines and threonines, common to all variants, was an absolute requirement for beta-arrestin/dynamin-dependent receptor endocytosis, but not for receptor uncoupling. Furthermore, beta-arrestin/dynamin-dependent endocytosis and uncoupling were dependent on and independent of GRK2 kinase activity, respectively. These results clearly demonstrate that the uncoupling and endocytosis of 5-HT4R require different GRK2 concentrations and involve distinct molecular events.