Structural requirements of platelet chemokines for neutrophil activation

Using recombinantly expressed proteins and synthetic peptides, we examined the structural/functional features of the platelet chemokines, neutrophil-activating peptide-2 (NAP-2) and platelet factor 4 (PF4); that were important in their activation of neutrophils. Previous studies with the chemokine interleukin-8 (IL-8) had shown that the N- terminal region preceding the first cysteine residue was critical in defining neutrophil-activating properties. We examined whether NAP-2 and PF4 had similar structural requirements. In the Ale-glu-leu-arg (AELR) N-terminus of NAP-2, substitution of E or R abolished Ca2+ mobilization and elastase secretion. Unlike the parent molecule PF4, AELR/PF4, the hybrid formed by replacing the N-terminal sequence of PF4 before the first cysteine residue with the homologous sequence of NAP- 2, stimulated Ca2+ mobilization and elastase secretion. Furthermore, the effect of amino acid substitutions in the ELR motif differed from those seen with NAP-2 in that conserved substitutions of E or R in NAP- 2 abolished activity, but only reduced neutrophil activation in the hybrid. These studies show that just as with IL-8, the N-termini of NAP- 2 and PF4 are critical for high-level neutrophil-activating function. Desensitization studies provided information on receptor binding. NAP- 2, which binds almost exclusively to the type 2 IL-8 receptor (IL-8R), did not desensitize neutrophils to activation by IL-8 because IL-8 could bind to and activate via both type 1 and 2 IL-8R. AELR/PF4 appears to bind to both types of receptors because it desensitized neutrophils to NAP-2 activation; but was not desensitized by NAP-2, and because it desensitized to and was desensitized by IL-8. Thus, although NAP-2 and AELR/PF4 share approximately 60% amino acid homology, they have different receptor affinities. Studies were performed to define the role of the C-termini of these platelet chemokines in receptor binding. Heparin and a monoclonal antibody specific for the heparin- binding domain of PF4 both inhibited Ca2+ mobilization and elastase release, further suggesting that the C-terminus of these chemokines is important in receptor binding. Synthetic NAP-2(51-70) failed to mobilize Ca2+, whereas PF4(47-70) and PF4(58-70) induced Ca2+ mobilization and secretion of elastase at high concentrations. Pertussis toxin inhibited neutrophil activation by 40% to 50%, establishing a role for G-protein-coupled receptors such as the IL-8Rs in activation by the PF4 C-terminal peptides.(ABSTRACT TRUNCATED AT 400 WORDS)     

A humanised tissue‐engineered bone model allows species‐specific breast cancer‐related bone metastasis in vivo

Bone metastases frequently occur in the advanced stages of breast cancer. At this stage, the disease is deemed incurable. To date, the mechanisms of breast cancer‐related metastasis to bone are poorly understood. This may be attributed to the lack of appropriate animal models to investigate the complex cancer cell–bone interactions. In this study, two established tissue‐engineered bone constructs (TEBCs) were applied to a breast cancer‐related metastasis model. A cylindrical medical‐grade polycaprolactone‐tricalcium phosphate scaffold produced by fused deposition modelling (scaffold 1) was compared with a tubular calcium phosphate‐coated polycaprolactone scaffold fabricated by solution electrospinning (scaffold 2) for their potential to generate ectopic humanised bone in NOD/SCID mice. While scaffold 1 was found not suitable to generate a sufficient amount of ectopic bone tissue due to poor ectopic integration, scaffold 2 showed excellent integration into the host tissue, leading to bone formation. To mimic breast cancer cell colonisation to the bone, MDA‐MB‐231, SUM1315, and MDA‐MB‐231BO breast cancer cells were cultured in polyethylene glycol‐based hydrogels and implanted adjacent to the TEBCs. Histological analysis indicated that the breast cancer cells induced an osteoclastic reaction in the TEBCs, demonstrating analogies to breast cancer‐related bone metastasis seen in patients.     

The effectiveness of a treatment protocol for male lower urinary tract symptoms in general practice: a practical randomised controlled trial

BACKGROUND: Randomised controlled trials have shown the efficacy of several treatment modalities for lower urinary tract symptoms (LUTS) in selected populations. The effectiveness in daily practice has hardly been investigated, especially in primary care and is dependent on choices between all possible treatment options and best investigated in a comprehensive study, including all treatment modalities (watchful waiting, alpha-blockers, 5-alpha-reductase inhibitors, and surgery). AIM: Assessment of the effectiveness of a comprehensive treatment protocol for LUTS in primary care. DESIGN OF STUDY: Randomised controlled trial. SETTING: Fourteen general practices in the Netherlands. METHOD: Intervention: treatment protocol based on a formalised expert opinion. Control condition: usual care. Study population: 208 subjects with moderate to severe LUTS (IPSS > or =8, median = 13). OUTCOME MEASURES: symptom severity (IPSS [International Prostate Symptom Score]), bother score (Dan-PSS [Danish Prostate Symptom Score]), and maximum urinary flow (Q(max)); incidence of acute urinary retention and urinary tract infections. RESULTS: In the intervention group markedly more subjects used an alpha-blocker at end of follow-up than in the usual care group (24% versus 6%). No significant differences were found between intervention and control group in IPSS, Q(max) or Dan-PSS. CONCLUSION: alpha-blockers and watchful waiting are the most frequent treatment modalities for LUTS in primary care. Our study showed no evidence that a protocol using well-defined indications for all possible treatment modalities based on a formalised expert opinion procedure has added value. Based on our results, we cannot recommend a broadening of the indication for alpha-blockers, which, however, seems to be the current trend.     

Assessing the Need for Anesthesia and Sedation Services in Kuwaiti Dental Practice

The objective of this study was to examine the public health relevance of the prevalence of dental fear in Kuwait and the resultant barrier that it creates regarding access to dental care. The study analysis demonstrated a high prevalence of dental fear and anxiety in the Kuwaiti population and a perceived need for anesthesia services by dental care providers. The telephone survey of the general population showed nearly 35% of respondents reported being somewhat nervous, very nervous, or terrified about going to the dentist. In addition, about 36% of the population postponed their dental treatment because of fear. Respondents showed a preference to receive sedation and anesthesia services as a means of anxiety relief, and they were willing to go to the dentist more often when such services were available. People with high fear and anxiety preferred to receive some type of medication to relieve their anxiety. In conclusion, the significance and importance of the need for anesthesia services to enhance the public health of dental patients in Kuwait has been demonstrated, and improvements are needed in anesthesia and sedation training of Kuwaiti dental care providers.     

Minimal important differences in the Migraine-Specific Quality of Life Questionnaire (MSQ) version 2.1

To propose minimal important differences (MID) for the Migraine-Specific Quality of Life Questionnaire version 2.1 (MSQ v2.1). To our knowledge (to date), no published MID values exist for the MSQ v2.1 in any population. Analyses were performed on data from two pivotal clinical trials of topiramate for migraine prevention ( n  = 916), as well as from the QualityMetric National Headache Survey ( n  = 1016). Analyses included both distribution- and anchor-based MID techniques as well as group- and individual-level MID values. Group-level anchor-based MID values ranged from 3.2 [Role Restrictive domain (RR)] to 7.5 [Emotional Functioning domain (EF)], setting the minimum level of appropriate MID (which can also aid with power analysis). Individual-level distribution-based MID values resulted in highly similar estimates from two large databases: median MID of 8.5 for RR, 9.2 for Role Preventive (RP) and 12.0 for EF. Finally, individual-level anchor-based MID values ranged from 5.0 (RR and RP domains) to 10.6 (EF). For group-level purposes of calculating power for future studies, an MID of 3.2, 4.6 and 7.5 for RR, RP and EF is recommended. For within-group analyses for analysing clinical trial efficacy of each patient's change with responder analyses, 5 points is necessary for RR. For RP and EF, ranges are recommended: 5.0 to 7.9 for RP and 8.0 to 10.6 for EF. These latter two domains tend to have more error in the MID, and thus a sensitivity analysis with both ends of the range should be used to confirm significant differences in responder analyses.