Role of Kir2.1 in human monocyte‐derived foam cell maturation

The role of K⁺ channels in macrophage immunomodulation has been well‐established. However, it remains unclear whether K⁺ channels are involved in the lipid uptake of macrophages. The expression and function of the inward rectifier potassium channel (Kir2.1, KCNJ2) in Human acute monocytic leukemia cell line (THP‐1) cells and human monocytes derived macrophages (HMDMs) were investigated using RT‐PCR and western blotting, and patch clamp technique. The expression of scavenger receptors in THP‐1–derived macrophages was detected using western blotting. Expressions of Kir2.1 mRNA and protein in HMDMs were significantly decreased by 60% (P < 0.05) and 90% (P < 0.001) on macrophage maturation, but overexpressed by approximately 1.3 (P > 0.05) and 3.8 times (P = 0.001) after foam cell formation respectively. Concurrently, the Kir2.1 peak current density in HMDMs, mature macrophages and foam cells, measured at −150 mV, were −22.61 ± 2.1 pA/pF, −7.88 ± 0.60 pA/pF and −13.39 ± 0.80 pA/pF respectively (P < 0.05). In association with an up‐regulation of Kir2.1 in foam cells, the SR‐A protein level was significantly increased by over 1.5 times compared with macrophages (P < 0.05). THP‐1 cells contained much less lipids upon Kir2.1 knockdown and cholesterol ester/total cholesterol ratio was 29.46 ± 2.01% (P < 0.05), and the SR‐BI protein level was increased by over 6.2 times, compared to that of macrophages (P < 0.001). Kir2.1 may participate in macrophage maturation and differentiation, and play a key role in lipid uptake and foam cell formation through modulating the expression of scavenger receptors.     

Correlation between endogenous polyamines in human cardiac tissues and clinical parameters in patients with heart failure

Polyamines contribute to several physiological and pathological processes, including cardiac hypertrophy in experimental animals. This involves an increase in ornithine decarboxylase (ODC) activity and intracellular polyamines associated with cyclic adenosine monophosphate (cAMP) increases. The aim of the study was to establish the role of these in the human heart in living patients. For this, polyamines (by high performance liquid chromatography) and the activity of ODC and N¹‐acetylpolyamine oxidases (APAO) were determined in the right atrial appendage of 17 patients undergoing extracorporeal circulation to correlate with clinical parameters. There existed enzymatic activity associated with the homeostasis of polyamines. Left atria size was positively associated with ODC (r = 0.661, P = 0.027) and negatively with APAO‐N¹‐acetylspermine (r = −0.769, P = 0.026), suggesting that increased levels of polyamines are associated with left atrial hemodynamic overload. Left ventricular ejection fraction (LVEF) and heart rate were positively associated with spermidine (r = 0.690, P = 0.003; r = 0.590, P = 0.021) and negatively with N¹‐acetylspermidine (r = −0.554, P = 0.032; r = −0.644, P = 0.018). LVEF was negatively correlated with cAMP levels (r = −0.835, P = 0.001) and with cAMP/ODC (r = −0.794, P = 0.011), cAMP/spermidine (r = −0.813, P = 0.001) and cAMP/spermine (r = −0.747, P = 0.003) ratios. Abnormal LVEF patients showed decreased ODC activity and spermidine, and increased N¹‐acetylspermidine, and cAMP. Spermine decreased in congestive heart failure patients. The trace amine isoamylamine negatively correlated with septal wall thickness (r = −0.634, P = 0.008) and was increased in cardiac heart failure. The results indicated that modifications in polyamine homeostasis might be associated with cardiac function and remodelling. Increased cAMP might have a deleterious effect on function. Further studies should confirm these findings and the involvement of polyamines in different stages of heart failure.     

Progranulin deficiency leads to severe inflammation,lung injury and cell death in a mouse model of endotoxic shock

Progranulin (PGRN) is a crucial secreted growth factor involved in various kinds of physiologic and disease processes and often has a protective role in inflammatory diseases. This study was designed to investigate the protective effects of PGRN on endotoxic shock in a mouse model of PGRN deficiency. After lipopolysaccharide (LPS) injection to induce endotoxic shock in mice, PGRN levels were induced in wild‐type (WT) mice at 6 and 24 hrs. Survival rate analysis, haematoxylin and eosin staining, immunohistochemical staining, enzyme‐linked immunosorbent assay and in situ terminal deoxynucleotidyl transferase–mediated uridine triphosphate nick‐end labelling assay were used to reveal the susceptibility, lung injury, inflammatory cell infiltration, production of inflammatory mediators and lung cell death in mice after LPS injection. PGRN‐deficient (Grn ^−/−) mice were highly susceptible to LPS‐induced endotoxic shock, with decreased survival, severe lung injury, increased production of pro‐inflammatory mediators, and inflammatory cell infiltration and apoptotic death in the lung. Additionally, recombinant PGRN (rPGRN) administration before LPS stimulation ameliorated the survival of and abnormalities in both WT and Grn ^−/− mice. Altogether, these findings indicate that PGRN may be a novel biologic agent with therapeutic potential for endotoxic shock probably by inhibiting LPS‐induced systemic and local inflammation in mice for treating endotoxic shock.     

A RCT of a transdiagnostic internet-delivered treatment for three anxiety disorders: examination of support roles and disorder-specific outcomes

Background

Anxiety disorders share common vulnerabilities and symptoms. Disorder-specific treatment is efficacious, but few access evidence-based care. Administering transdiagnostic cognitive-behavioral therapy via the internet (iCBT) may increase access to evidence-based treatment, with a recent randomized controlled trial (RCT) providing preliminary support for this approach. This study extends those findings and aims to answer three questions: Is a transdiagnostic iCBT program for anxiety disorders efficacious and acceptable? Does it result in change for specific disorders? Can good clinical outcomes be obtained when guidance is provided via a Coach rather than a Clinician?

Method

RCT (N = 131) comparing three groups: Clinician-supported (CL) vs. Coach-supported (CO) vs. waitlist control (Control). Individuals met DSM-IV criteria for a principal diagnosis of generalized anxiety disorder (GAD), social phobia (SP) or panic disorder with or without agoraphobia (Pan/Ag). Treatment consisted of an 8-lesson/10 week iCBT program with weekly contact from a Clinician or Coach, and follow-up at 3-months post-treatment.

Results

Outcomes for the pooled treatment groups (CL+CO) were superior to the Control group on measures of anxiety, depression and disability, were associated with medium to large effect sizes (Cohen''s d = .76 – 1.44) (response rate = 89–100%), and were maintained at follow-up. Significant reductions were found on disorder-specific outcomes for each of the target diagnoses, and were associated with large effect sizes. CO participants achieved similar outcomes to CL participants at post-treatment, yet had significantly lower symptom severity scores on general anxiety, panic-disorder, depression and disability at follow-up (d = .45 – .46). Seventy-four percent of CO and 76% of CL participants completed the program. Less than 70 minutes of Clinician or Coach time was required per participant during the program.

Discussion

This transdiagnostic iCBT course for anxiety appears to be efficacious, associated with significant change for three target disorders, and is efficacious when guided by either a Clinician or Coach.

Trial Registration

Australian New Zealand Clinical Trials Registry ACTRN12610000242022     

Acupuncture for migraine prophylaxis: a randomized controlled trial

Background:

Acupuncture is commonly used to treat migraine. We assessed the efficacy of acupuncture at migraine-specific acupuncture points compared with other acupuncture points and sham acupuncture.

Methods:

We performed a multicentre, single-blind randomized controlled trial. In total, 480 patients with migraine were randomly assigned to one of four groups (Shaoyang-specific acupuncture, Shaoyang-nonspecific acupuncture, Yangming-specific acupuncture or sham acupuncture [control]). All groups received 20 treatments, which included electrical stimulation, over a period of four weeks. The primary outcome was the number of days with a migraine experienced during weeks 5–8 after randomization. Our secondary outcomes included the frequency of migraine attack, migraine intensity and migraine-specific quality of life.

Results:

Compared with patients in the control group, patients in the acupuncture groups reported fewer days with a migraine during weeks 5–8, however the differences between treatments were not significant (p > 0.05). There was a significant reduction in the number of days with a migraine during weeks 13–16 in all acupuncture groups compared with control (Shaoyang-specific acupuncture v. control: difference –1.06 [95% confidence interval (CI) –1.77 to –0.5], p = 0.003; Shaoyang-nonspecific acupuncture v. control: difference –1.22 [95% CI –1.92 to –0.52], p < 0.001; Yangming-specific acupuncture v. control: difference –0.91 [95% CI –1.61 to –0.21], p = 0.011). We found that there was a significant, but not clinically relevant, benefit for almost all secondary outcomes in the three acupuncture groups compared with the control group. We found no relevant differences between the three acupuncture groups.

Interpretation:

Acupuncture tested appeared to have a clinically minor effect on migraine prophylaxis compared with sham acupuncture.

Trial Registration:

Clinicaltrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00599586","term_id":"NCT00599586"}}NCT00599586About 6%–8% of men and 16%–18% of women in the United States and England experience migraines, with or without an aura.^1,2 A prevalence of 1% has been reported in mainland China,³ compared with 4.7% in Hong Kong and 9.1% in Taiwan.^4,5 A recent Cochrane meta-analysis suggests that acupuncture as migraine prophylaxis is safe and effective and should be considered as a treatment option for willing patients.⁶Although the specific effects acupuncture are controversial, acupuncture, as it is currently practised, clearly differentiates between real acupuncture points and nonacupuncture points. The Chinese Government launched the National Basic Research Program to obtain more data about the specificity of acupuncture points.⁷Trials from Italy and Brazil^8,9 showed that acupuncture was more effective than sham acupuncture in preventing migraines, but other trials have reported no differences.^10–13 There is no evidence that one acupuncture strategy is more effective than another for treating migraines. According to acupuncture theory, a headache on the lateral side is usually defined as a Shaoyang headache. In Jinkuiyi,¹⁴ migraines are said to affect the yang meridians (including the Taiyang, Yangming and Shaoyang meridians). In Lingshu,¹⁵ the Shaoyang meridians are said to go through the lateral side of the body, therefore the Shaoyang meridians are thought to be superior for treating migraines. Some points on the Shaoyang meridians are regarded as being more specific for migraines than other points.¹⁶Our aim was to investigate whether acupuncture at specific acupuncture points was more efficacious in preventing migraine than sham acupuncture at nonacupuncture points. We also investigated whether the efficacy varied when acupuncture points along different meridians or points along the same meridian were used.     

Human TLR8 senses UR/URR motifs in bacterial and mitochondrial RNA

Toll‐like receptor (TLR) 13 and TLR2 are the major sensors of Gram‐positive bacteria in mice. TLR13 recognizes Sa19, a specific 23S ribosomal (r) RNA‐derived fragment and bacterial modification of Sa19 ablates binding to TLR13, and to antibiotics such as erythromycin. Similarly, RNase A‐treated Staphylococcus aureus activate human peripheral blood mononuclear cells (PBMCs) only via TLR2, implying single‐stranded (ss) RNA as major stimulant. Here, we identify human TLR8 as functional TLR13 equivalent that promiscuously senses ssRNA. Accordingly, Sa19 and mitochondrial (mt) 16S rRNA sequence‐derived oligoribonucleotides (ORNs) stimulate PBMCs in a MyD88‐dependent manner. These ORNs, as well as S. aureus‐, Escherichia coli‐, and mt‐RNA, also activate differentiated human monocytoid THP‐1 cells, provided they express TLR8. Moreover, Unc93b1 ^−/−‐ and Tlr8 ^−/−‐THP‐1 cells are refractory, while endogenous and ectopically expressed TLR8 confers responsiveness in a UR/URR RNA ligand consensus motif‐dependent manner. If TLR8 function is inhibited by suppression of lysosomal function, antibiotic treatment efficiently blocks bacteria‐driven inflammatory responses in infected human whole blood cultures. Sepsis therapy might thus benefit from interfering with TLR8 function.     

Insights from the GC content analysis of 76genome survey sequences (GSS) from Elaeisoleifera

South American oil-palm (Elaeis oleifera) is not cultivated in tropical countries like Malaysia on large scale due to low yield of palm oil derived from its fruit mesocarp. However, its fruit mesocarp oil contains about 68.6 % oleic acid (C_18:1) which is more than double in comparison to commercially cultivated oilpalm, E. guineensis Jacq Tenera (hybrid of Dura (♀) x Pisifera (♂)). It is also known that E. oleifera is a good source of tocotrienols and carotenoids. Therefore, it is of interest to know the genome sequence of E. oleifera. The objective of this study is to generate genome survey sequences (GSS) to get GC content insight in the E. oleifera genome. The nuclear genomic DNA isolated from young leaf‐tissues was digested with EcoRI and NdeI/DraI restriction enzymes; and three genomic DNA libraries were constructed using Lambda ZAP‐II, pGEM®‐T Easy, and pDONR 222™ as cloning vectors. Generated 76 GSSs were analyzed by using Bioinformatics tools. The analysis result indicates that the adenine, cytosine, guanine and thymine content in generated GSSs are 30%, 20%, 20%, and 30% respectively. In conclusion, based on the precise GC content analysis of the randomly isolated 76 GSSs by using Bioinformatics tools we hypothesize that GC content in E. oleifera genome is 40%. The hypothesized 40% GC content in E. oleifera genome is expected to remain close to the GC content based on the whole genome analysis.^ψThe nucleotide sequence data reported in this paper have been submitted to dbGSS division of the international DNA database (GenBank/DDBJ/EMBL) under accession numbers: {"type":"entrez-nucleotide-range","attrs":{"text":"DX575945- DX575972","start_term":"DX575945","end_term":"DX575972","start_term_id":"94323534","end_term_id":"94323561"}}DX575945- DX575972 and {"type":"entrez-nucleotide-range","attrs":{"text":"EI798032-EI798079","start_term":"EI798032","end_term":"EI798079","start_term_id":"146199019","end_term_id":"146199066"}}EI798032-EI798079.

Abbreviations

gDNA - Nuclear genomic DNA, GSSs - Genome survey sequences K12, SAOP - South American oil‐palm Db1     

Effect of iron supplementation on fatigue in nonanemic menstruating women with low ferritin: a randomized controlled trial

Background:

The true benefit of iron supplementation for nonanemic menstruating women with fatigue is unknown. We studied the effect of oral iron therapy on fatigue and quality of life, as well as on hemoglobin, ferritin and soluble transferrin receptor levels, in nonanemic iron-deficient women with unexplained fatigue.

Methods:

We performed a multicentre, parallel, randomized controlled, closed-label, observer-blinded trial. We recruited from the practices of 44 primary care physicians in France from March to July 2006. We randomly assigned 198 women aged 18–53 years who complained of fatigue and who had a ferritin level of less than 50 ug/L and hemoglobin greater than 12.0 g/dL to receive either oral ferrous sulfate (80 mg of elemental iron daily; n = 102) or placebo (n = 96) for 12 weeks. The primary outcome was fatigue as measured on the Current and Past Psychological Scale. Biological markers were measured at 6 and 12 weeks.

Results:

The mean score on the Current and Past Psychological Scale for fatigue decreased by 47.7% in the iron group and by 28.8% in the placebo group (difference –18.9%, 95% CI −34.5 to −3.2; p = 0.02), but there were no significant effects on quality of life (p = 0.2), depression (p = 0.97) or anxiety (p = 0.5). Compared with placebo, iron supplementation increased hemoglobin (0.32 g/dL; p = 0.002) and ferritin (11.4 μg/L; p < 0.001) and decreased soluble transferrin receptor (−0.54 mg/L; p < 0.001) at 12 weeks.

Interpretation:

Iron supplementation should be considered for women with unexplained fatigue who have ferritin levels below 50 μg/L. We suggest assessing the efficiency using blood markers after six weeks of treatment. Trial registration no. EudraCT 2006–000478–56.The prevalence of fatigue ranges from 14% to 27% among patients in primary care.¹ In addition, 1%–2% of visits to general practices are because of fatigue, and women are three times more likely than men to mention fatigue.¹ Unexplained fatigue can be caused by iron deficiency.² Verdon and coauthors found an improvement in fatigue following iron supplementation in nonanemic women with unexplained fatigue.³ However, the hemoglobin levels of these patients were not available, which may have contributed to the ongoing debate about the appropriateness of reference limits defining anemia in women.^4,5 Thus, the effectiveness of iron supplementation in nonanemic menstruating women with major fatigue without an obvious clinical cause is unknown.⁶ Our main objective was to test the hypothesis that oral iron therapy for a short period may improve fatigue, hemoglobin, iron stores and quality of life in menstruating nonanemic women whose ferritin levels are below 50 μg/L. Our secondary objective was to evaluate whether this effect is dependent on the initial levels of hemoglobin, ferritin or transferrin saturation.     

Empirically derived maximal acceptable wait time for surgery to treat adolescent idiopathic scoliosis

Background

Prioritizing patients using empirically derived access targets can help to ensure high-quality care. Adolescent scoliosis can worsen while patients wait for treatment, increasing the risk of adverse events. Our objective was to determine an empirically derived access target for scoliosis surgery and to compare this with consensus-based targets

Methods

Two-hundred sixteen sequential patients receiving surgery for adolescent idiopathic scoliosis were included in the study. The main outcome was need for additional surgery. Logistic regression modeling was used to evaluate the relation between surgical wait times and adverse events and χ² analysis was used as the primary analysis for the main outcome.

Results

Of the 88 patients who waited longer than six months for surgery, 13 (14.8%) needed additional surgery due to progression of curvature versus 1.6% (2 of 128 patients) who waited less than six months for surgery (χ² analysis, p = 0.0001). Patients who waited longer than six months for surgery had greater progression of curvature, longer surgeries and longer stays in hospital. These patients also had less surgical correction than patients who waited less than six months for surgery (Wilcoxon–Mann–Whitney test, p = 0.011). All patients requiring additional surgeries waited longer than three months for their initial surgery. A receiver–operator characteristic curve also suggested a three-month wait as an access target. The adjusted odds ratio for an adverse event for each additional 90 days of waiting from time of consent was 1.81 (95% confidence interval 1.34–2.44). The adjusted odds ratio increased with skeletal immaturity and with the size of the spinal curvature at the time of consent.

Interpretation

A prolonged wait for surgery increased the risk of additional surgical procedures and other adverse events. An empirically derived access target of three months for surgery to treat adolescent idiopathic scoliosis could potentially eliminate the need for additional surgery by reducing progression of curvature. This is a shorter access target than the six months determined by expert consensus.Adolescent idiopathic scoliosis effects just over 2% of females aged 12–14 years.^1–3 Although only 10% of patients require surgery, spinal instrumentation and fusion for adolescent idiopathic scoliosis is the most common procedure done in pediatric orthopaedics.⁴ Patients who wait too long for scoliosis surgery may require additional surgery such as anterior release to achieve satisfactory correction of the spinal curvature. These patients may also need longer surgeries and may be at increased risk of complications such as increased blood loss, neurologic deficits or inadequate correction of the curvature.^5–14 Furthermore, as seen in other studies of wait times, patients and families can feel anxiety and prolonged suffering while waiting for treatment, which can negatively impact the quality of care.^15–19 Programs such as the Canadian Pediatric Surgical Wait Times Project have determined a maximal acceptable wait time for adolescent scoliosis through expert consensus (similar to how other surgical wait time targets have been determined).²⁰ Surprisingly, there has been little or no attention given to developing evidence-based access targets or maximal acceptable wait times for most treatments.²¹ The purpose of this study was to determine the maximal acceptable wait time for surgical correction of adolescent idiopathic scoliosis using an empirically based approach to minimize the possibility of adverse events related to progression of curvature.     

Internet-based cognitive behavior therapy vs. cognitive behavioral group therapy for social anxiety disorder: a randomized controlled non-inferiority trial

Background and Aims

Cognitive behavioral group therapy (CBGT) is an effective, well-established, but not widely available treatment for social anxiety disorder (SAD). Internet-based cognitive behavior therapy (ICBT) has the potential to increase availability and facilitate dissemination of therapeutic services for SAD. However, ICBT for SAD has not been directly compared with in-person treatments such as CBGT and few studies investigating ICBT have been conducted in clinical settings. Our aim was to investigate if ICBT is at least as effective as CBGT for SAD when treatments are delivered in a psychiatric setting.

Methods

We conducted a randomized controlled non-inferiority trial with allocation to ICBT (n = 64) or CBGT (n = 62) with blinded assessment immediately following treatment and six months post-treatment. Participants were 126 individuals with SAD who received CBGT or ICBT for a duration of 15 weeks. The Liebowitz Social Anxiety Scale (LSAS) was the main outcome measure. The following non-inferiority margin was set: following treatment, the lower bound of the 95 % confidence interval (CI) of the mean difference between groups should be less than 10 LSAS-points.

Results

Both groups made large improvements. At follow-up, 41 (64%) participants in the ICBT group were classified as responders (95% CI, 52%–76%). In the CBGT group, 28 participants (45%) responded to the treatment (95% CI, 33%–58%). At post-treatment and follow-up respectively, the 95 % CI of the LSAS mean difference was 0.68–17.66 (Cohen’s d between group = 0.41) and −2.51–15.69 (Cohen’s d between group = 0.36) favoring ICBT, which was well within the non-inferiority margin. Mixed effects models analyses showed no significant interaction effect for LSAS, indicating similar improvement across treatments (F = 1.58; df = 2, 219; p = .21).

Conclusions

ICBT delivered in a psychiatric setting can be as effective as CBGT in the treatment of SAD and could be used to increase availability to CBT.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00564967","term_id":"NCT00564967"}}NCT00564967     

Efficacy of oral metronidazole with vaginal clindamycin or vaginal probiotic for bacterial vaginosis: randomised placebo-controlled double-blind trial

Background

To determine if oral metronidazole (MTZ-400mg bid) with 2% vaginal clindamycin-cream (Clind) or a Lactobacillus acidophilus vaginal-probiotic containing oestriol (Prob) reduces 6-month bacterial vaginosis (BV) recurrence.

Methods

Double-blind placebo-controlled parallel-group single-site study with balanced randomization (1∶1∶1) conducted at Melbourne Sexual Health Centre, Australia. Participants with symptomatic BV [Nugent Score (NS) = 7–10 or ≥3 Amsel''s criteria and NS = 4–10], were randomly allocated to MTZ-Clind, MTZ-Prob or MTZ-Placebo and assessed at 1,2,3 and 6 months. MTZ and Clind were administered for 7 days and Prob and Placebo for 12 days. Primary outcome was BV recurrence (NS of 7–10) on self-collected vaginal-swabs over 6-months. Cumulative BV recurrence rates were compared between groups by Chi-squared statistics. Kaplan-Meier, log rank and Cox regression analyses were used to compare time until and risk of BV recurrence between groups.

Results

450 18–50 year old females were randomized and 408 (91%), equally distributed between groups, provided ≥1 NS post-randomization and were included in analyses; 42 (9%) participants with no post-randomization data were excluded. Six-month retention rates were 78% (n = 351). One-month BV recurrence (NS 7–10) rates were 3.6% (5/140), 6.8% (9/133) and 9.6% (13/135) in the MTZ-Clind, MTZ-Prob and MTZ-Placebo groups respectively, p = 0.13. Hazard ratios (HR) for BV recurrence at one-month, adjusted for adherence to vaginal therapy, were 0.43 (95%CI 0.15–1.22) and 0.75 (95% CI 0.32–1.76) in the MTZ-Clind and MTZ-Prob groups compared to MTZ-Plac respectively. Cumulative 6-month BV recurrence was 28.2%; (95%CI 24.0–32.7%) with no difference between groups, p = 0.82; HRs for 6-month BV recurrence for MTZ-Clind and MTZ-Prob compared to MTZ-Plac, adjusted for adherence to vaginal therapy were 1.09(95% CI = 0.70–1.70) and 1.03(95% CI = 0.65–1.63), respectively. No serious adverse events occurred.

Conclusion

Combining the recommended first line therapies of oral metronidazole and vaginal clindamycin, or oral metronidazole with an extended-course of a commercially available vaginal-L.acidophilus probiotic, does not reduce BV recurrence.

Trial Registration

ANZCTR.org.au ACTRN12607000350426     

Genetic Polymorphisms in Monoamine Systems and Outcome of Cognitive Behavior Therapy for Social Anxiety Disorder

Objective

The role of genetics for predicting the response to cognitive behavior therapy (CBT) for social anxiety disorder (SAD) has only been studied in one previous investigation. The serotonin transporter (5-HTTLPR), the catechol-o-methyltransferase (COMT) val158met, and the tryptophan hydroxylase-2 (TPH2) G-703Tpolymorphisms are implicated in the regulation of amygdala reactivity and fear extinction and therefore might be of relevance for CBT outcome. The aim of the present study was to investigate if these three gene variants predicted response to CBT in a large sample of SAD patients.

Method

Participants were recruited from two separate randomized controlled CBT trials (trial 1: n = 112, trial 2: n = 202). Genotyping were performed on DNA extracted from blood or saliva samples. Effects were analyzed at follow-up (6 or 12 months after treatment) for both groups and for each group separately at post-treatment. The main outcome measure was the Liebowitz Social Anxiety Scale Self-Report.

Results

At long-term follow-up, there was no effect of any genotype, or gene × gene interactions, on treatment response. In the subsamples, there was time by genotype interaction effects indicating an influence of the TPH2 G-703T-polymorphism on CBT short-term response, however the direction of the effect was not consistent across trials.

Conclusions

None of the three gene variants, 5-HTTLPR, COMTval158met and TPH2 G-703T, was associated with long-term response to CBT for SAD.

Trial Registration

ClinicalTrials.gov (ID-NCT0056496)     

Effectiveness of strengthened stimulation during acupuncture for the treatment of Bell palsy: a randomized controlled trial

Background:

The traditional Chinese theory of acupuncture emphasizes that the intensity of acupuncture must reach a threshold to generate de qi, which is necessary to achieve the best therapeutic effect. De qi is an internal compound sensation of soreness, tingling, fullness, aching, cool, warmth and heaviness, and a radiating sensation at and around the acupoints. However, the notion that de qi must be achieved for maximum benefit has not been confirmed by modern scientific evidence.

Methods:

We performed a prospective multicentre randomized controlled trial involving patients with Bell palsy. Patients were randomly assigned to the de qi (n = 167) or control (n = 171) group. Both groups received acupuncture: in the de qi group, the needles were manipulated manually until de qi was reached, whereas in the control group, the needles were inserted without any manipulation. All patients received prednisone as a basic treatment. The primary outcome was facial nerve function at month 6. We also assessed disability and quality of life 6 months after randomization.

Results:

After 6 months, patients in the de qi group had better facial function (adjusted odds ratio [OR] 4.16, 95% confidence interval [CI] 2.23–7.78), better disability assessment (differences of least squares means 9.80, 95% CI 6.29–13.30) and better quality of life (differences of least squares means 29.86, 95% CI 22.33–37.38). Logistic regression analysis showed a positive effect of the de qi score on facial-nerve function (adjusted OR 1.07, 95% CI 1.04–1.09).

Interpretation:

Among patients with Bell palsy, acupuncture with strong stimulation that elicited de qi had a greater therapeutic effect, and stronger intensity of de qi was associated with the better therapeutic effects. Trial registration: Clinicaltrials.gov no. NCT00685789.The use of acupuncture has gained increasing attention worldwide.^1,2 There is a long-held belief in the traditional theory and clinical practice of acupuncture that the intensity of the stimulus must reach a threshold to elicit de qi, which plays a pivotal role in achieving the best therapeutic effects.^3,4De qi — an internal compound sensation of soreness, tingling, fullness, aching, cool, warmth, heaviness and a radiating sensation at and around acupoints — is elicited by manipulation of the needles (rotated as well as being moved upward and downward).⁵ The concept of de qi, originating from the long-term accumulation of clinical experiences, has been explained to some degree by several scientific studies.^6–11 However, this long-held belief has not been confirmed by sufficient evidence from randomized controlled trials.⁵ Consequently, de qi and its related techniques have been neglected in acupuncture practice and research. As such, the therapeutic effects of acupuncture may be seriously compromised.^12–14In this study, we compared the efficacy of acupuncture with either strong (intended to elicit de qi) or weak stimulation among patients with Bell palsy.     

A head-to-head comparison of four artemisinin-based combinations for treating uncomplicated malaria in African children: a randomized trial

Background

Artemisinin-based combination therapies (ACTs) are the mainstay for the management of uncomplicated malaria cases. However, up-to-date data able to assist sub-Saharan African countries formulating appropriate antimalarial drug policies are scarce.

Methods and Findings

Between 9 July 2007 and 19 June 2009, a randomized, non-inferiority (10% difference threshold in efficacy at day 28) clinical trial was carried out at 12 sites in seven sub-Saharan African countries. Each site compared three of four ACTs, namely amodiaquine-artesunate (ASAQ), dihydroartemisinin-piperaquine (DHAPQ), artemether-lumefantrine (AL), or chlorproguanil-dapsone-artesunate (CD+A). Overall, 4,116 children 6–59 mo old with uncomplicated Plasmodium falciparum malaria were treated (1,226 with AL, 1,002 with ASAQ, 413 with CD+A, and 1,475 with DHAPQ), actively followed up until day 28, and then passively followed up for the next 6 mo. At day 28, for the PCR-adjusted efficacy, non-inferiority was established for three pair-wise comparisons: DHAPQ (97.3%) versus AL (95.5%) (odds ratio [OR]: 0.59, 95% CI: 0.37–0.94); DHAPQ (97.6%) versus ASAQ (96.8%) (OR: 0.74, 95% CI: 0.41–1.34), and ASAQ (97.1%) versus AL (94.4%) (OR: 0.50, 95% CI: 0.28–0.92). For the PCR-unadjusted efficacy, AL was significantly less efficacious than DHAPQ (72.7% versus 89.5%) (OR: 0.27, 95% CI: 0.21–0.34) and ASAQ (66.2% versus 80.4%) (OR: 0.40, 95% CI: 0.30–0.53), while DHAPQ (92.2%) had higher efficacy than ASAQ (80.8%) but non-inferiority could not be excluded (OR: 0.35, 95% CI: 0.26–0.48). CD+A was significantly less efficacious than the other three treatments. Day 63 results were similar to those observed at day 28.

Conclusions

This large head-to-head comparison of most currently available ACTs in sub-Saharan Africa showed that AL, ASAQ, and DHAPQ had excellent efficacy, up to day 63 post-treatment. The risk of recurrent infections was significantly lower for DHAPQ, followed by ASAQ and then AL, supporting the recent recommendation of considering DHAPQ as a valid option for the treatment of uncomplicated P. falciparum malaria.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00393679","term_id":"NCT00393679"}}NCT00393679; Pan African Clinical Trials Registry PACTR2009010000911750 Please see later in the article for the Editors'' Summary     

Heterologous Stacking of Prion Protein Peptides Reveals Structural Details of Fibrils and Facilitates Complete Inhibition of Fibril Growth

Fibrils play an important role in the pathogenesis of amyloidosis; however, the underlying mechanisms of the growth process and the structural details of fibrils are poorly understood. Crucial in the fibril formation of prion proteins is the stacking of PrP monomers. We previously proposed that the structure of the prion protein fibril may be similar as a parallel left-handed β-helix. The β-helix is composed of spiraling rungs of parallel β-strands, and in the PrP model residues 105–143 of each PrP monomer can contribute two β-helical rungs to the growing fibril. Here we report data to support this model. We show that two cyclized human PrP peptides corresponding to residues 105–124 and 125–143, based on two single rungs of the left-handed β-helical core of the human PrP^Sc fibril, show spontaneous cooperative fibril growth in vitro by heterologous stacking. Because the structural model must have predictive value, peptides were designed based on the structure rules of the left-handed β-helical fold that could stack with prion protein peptides to stimulate or to block fibril growth. The stimulator peptide was designed as an optimal left-handed β-helical fold that can serve as a template for fibril growth initiation. The inhibiting peptide was designed to bind to the exposed rung but frustrate the propagation of the fibril growth. The single inhibitory peptide hardly shows inhibition, but the combination of the inhibitory with the stimulatory peptide showed complete inhibition of the fibril growth of peptide huPrP-(106–126). Moreover, the unique strategy based on stimulatory and inhibitory peptides seems a powerful new approach to study amyloidogenic fibril structures in general and could prove useful for the development of therapeutics.Transmissible spongiform encephalopathies are neurodegenerative disorders in a wide range of mammalian species, including Creutzfeldt-Jacob disease in man, scrapie in sheep, and bovine spongiform encephalopathy in cattle. The deposition of aggregated prion protein fibrils on and in neurons is regarded to be the source of these neurodegenerative diseases and is frequently associated with occurrence of Congo red positivity (1–3). The fibrils are formed by the conformational change of the prion protein (PrP^c)² into the scrapie form (PrP^Sc). The misfolded conformer of the prion protein (PrP^Sc) is considered as the causative agent in these diseases according to the protein-only hypothesis (4). Studies have shown the toxicity of fibrils of the full-length recombinant mammalian prion protein as well as soluble β-rich oligomers to cultured cells and primary neurons (5).It is still unknown how much of the whole PrP^Sc molecule is involved in the fibril growth. It is shown that the N-terminal part of PrP, specifically residues 112–141, can go through conformational changes involving β-strand formation, which subsequently triggers fibril growth (6–8), and solid state NMR studies showed that residues 112–141 are part of the highly ordered core of huPrP-(23–144) (9). It was previously shown that peptides based on the 89–143 region of the human PrP protein can form fibrils rich in β-sheet structure which are biologically active in transgenic mice (10). Within this region it is the huPrP-(106–126) peptide that is the smallest known region of PrP that forms fibrils that are toxic and resemble the physiological properties of PrP^Sc (11–16). The formation of PrP^Sc is considered to be a two-step event; first, there is the binding between PrP^c and PrP^Sc and subsequently the conformational conversion from PrP^c into PrP^Sc occurs. Mutation studies in a prion-infected neuroblastoma cell line showed that in mouse PrP the regions 101–110 and 136–158 are crucial for the binding and conversion events, respectively (17). Because prevention of fibril growth is the prime therapeutic target, detailed structural knowledge of the fibril is essential for understanding the mechanism of fibril growth. However, structural analysis of amyloid fibrils is hampered by insolubility, isomorphism, and aggregation. X-ray diffraction of several amyloid fibrils revealed a so-called cross-β diffraction pattern which indicates that the fibrils contain β-strands perpendicular to the fibril axis and hydrogen bonds in parallel (18, 19). Thus, for fibril growth the β-strands have to stack on top of each other. Several structures have been suggested to explain the structure of the stacked β-strands; e.g. a parallel in register organization of stacked β hairpins (24) or the comparable dry steric zipper structure (25). Previously, we and other groups suggested that the β-sheet structures in the PrP^Sc fibril may be similar to the topologically most simple class of β-sheets; that is, the parallel left-handed β-helix (Fig. 1A) (6, 20, 21). The left-handed β helix is formed by triangular progressive coils (rungs) of 18–20 residues. Each rung is formed by three hexapeptide motifs, which results in an approximate 3-fold symmetry. Backbone-backbone hydrogen bonding and stacking of the side chains in adjacent rungs contribute to the folding of β-helical rungs. We suggested that each PrP^Sc monomer contributes two left-handed β-helical rungs to the fibril, comprising residues 105–124 and 125–143 (Fig. 1A). This two-rung structural model was recently confirmed for amyloid fibrils of the HET-s prion by NMR analysis (22). In contrast to fibrils which are composed of homologous stacks of identical peptides, e.g. the Aβ peptide (23), the PrP^Sc fibril is more complex because it is composed of heterologous stacks of at least two peptides. For homologous stacking of two identical peptides, the complementarity issue is relatively simple because the identical side chains are in register (e.g. Ile-Ile, Val-Val stacking, and Asn ladders). However, in the case of heterologous stacking, the side chains of the additional heterologous peptide needs to be complementary with the other peptide to allow fibril growth.Open in a separate windowFIGURE 1.A, theoretical model of the fibrillogenic core of PrP^Sc. In the PrP^Sc model based on the left-handed β-helix structure, each PrP^Sc monomer contributes two stacked rungs to the fibril (different color for each monomer). The protofibril is formed by consecutive stacking of the two windings. The stack of two rungs provides enough elevation to accommodate the remaining part (residues ∼ 146–253) of the PrP^Sc molecule (20). B, the left-handed β-helix structure of LpxA-based on x-ray crystallography. In the left-handed β-helix structure of LpxA (PDB code 1LXA) rungs 6 and 7 are indicated (red) that were used for the heterologous stacking studies. Linear and cyclized peptides based on rung 6 and rung 7 were modified to satisfy the ideal left-handed β-helix motif (see “LpxA Peptides” under “Results”) and tested for their intrinsic and cooperative fibrillogenicity. C, left-handed β-helical rung based on rung 6 of LpxA. The rung is formed by three hexapeptide motifs, which results in an approximate 3-fold symmetry. A left-handed β-helical rung can be cyclized by a disulfide bridge after the introduction of a cysteine at position 2 of the first hexapeptide and position 1 of the fourth hexapeptide (according to the numbering used for the hexapeptide repeats in the left-handed β-helix).To investigate whether the suggested rungs 105–123 and 125–143 from human PrP could be complementary (20), we studied the homologous stacking and the heterologous stacking of linear and cyclized prion protein peptides comprising the huPrP-(105–143) region (KTNMKHMAGAAAAGAVVGGLGGYMLGSAMSRPIIHFGS). Qualitative and semiquantitative analysis were done by electron microscopy and Congo red staining. The quantification of the fibril formation was assessed by thioflavin S staining, in which the addition of polyanions (e.g. heparin) enhance the β-sheet formation of peptides comprising the 82–143 region of PrP and improve the reproducibility of the fibril growth (24). This study provides first evidence of heterologous stacking by two isolated putative β-strand layers (or rungs) of the human prion protein with fibril formation as a result. The left-handed β-helix structure provided insight for the “stack-and-stop” approach. With this approach a mix of a stimulatory peptide and an inhibitory peptide could completely block fibril formation. The stimulatory peptide was based on the 125–143 region that was optimized to serve as a folding template for the consecutive stacking of the 106–126 peptide. This cooperative fibril growth was completely inhibited by the inhibitory peptide based on peptides 106–126 with strategic d-amino acid and/or proline substitutions. The findings in this study support models in which the sequential strands in a fibril must somehow spiral up- or downward along the fibril axis, e.g. like the hypothetical left-handed β-helical structure of PrP^Sc fibrils (20). Furthermore, it allows the development of well defined small protein modules which can be used for structure studies of the 82–143 domain of PrP^Sc and the development of therapeutics.     

Size matters for violent discharge height and settling speed of Sphagnum spores: important attributes for dispersal potential

Background and Aims

Initial release height and settling speed of diaspores are biologically controlled components which are key to modelling wind dispersal. Most Sphagnum (peat moss) species have explosive spore liberation. In this study, how capsule and spore sizes affect the height to which spores are propelled were measured, and how spore size and spore number of discharged particles relate to settling speed in the aspherical Sphagnum spores.

Methods

Spore discharge and spore cloud development were filmed in a closed chamber (nine species). Measurements were taken from snapshots at three stages of cloud development. Settling speed of spores (14 species) and clusters were timed in a glass tube.

Key Results

The maximum discharge speed measured was 3·6 m s⁻¹. Spores reached a maximum height of 20 cm (average: 15 cm) above the capsule. The cloud dimensions at all stages were related positively to capsule size (R² = 0·58–0·65). Thus species with large shoots (because they have large capsules) have a dispersal advantage. Half of the spores were released as singles and the rest as clusters (usually two to four spores). Single spores settled at 0·84–1·86 cm s⁻¹, about 52 % slower than expected for spherical spores with the same diameters. Settling speed displayed a positive curvilinear relationship with spore size, close to predictions by Stokes'' law for spherical spores with 68 % of the actual diameters. Light-coloured spores settled slower than dark spores. Settling speed of spore clusters agrees with earlier studies. Effective spore discharge and small, slowly settling spores appear particularly important for species in forested habitats.

Conclusions

The spore discharge heights in Sphagnum are among the greatest for small, wind-dispersed propagules. The discharge heights and the slow settling of spores affect dispersal distances positively and may help to explain the wide distribution of most boreal Sphagnum species.     

Efficacy of single-dose and triple-dose albendazole and mebendazole against soil-transmitted helminths and Taenia spp.: a randomized controlled trial

Background

The control of soil-transmitted helminth (STH) infections currently relies on the large-scale administration of single-dose oral albendazole or mebendazole. However, these treatment regimens have limited efficacy against hookworm and Trichuris trichiura in terms of cure rates (CR), whereas fecal egg reduction rates (ERR) are generally high for all common STH species. We compared the efficacy of single-dose versus triple-dose treatment against hookworm and other STHs in a community-based randomized controlled trial in the People''s Republic of China.

Methodology/Principal findings

The hookworm CR and fecal ERR were assessed in 314 individuals aged ≥5 years who submitted two stool samples before and 3–4 weeks after administration of single-dose oral albendazole (400 mg) or mebendazole (500 mg) or triple-dose albendazole (3×400 mg over 3 consecutive days) or mebendazole (3×500 mg over 3 consecutive days). Efficacy against T. trichiura, Ascaris lumbricoides, and Taenia spp. was also assessed.Albendazole cured significantly more hookworm infections than mebendazole in both treatment regimens (single dose: respective CRs 69% (95% confidence interval [CI]: 55–81%) and 29% (95% CI: 20–45%); triple dose: respective CRs 92% (95% CI: 81–98%) and 54% (95% CI: 46–71%)). ERRs followed the same pattern (single dose: 97% versus 84%; triple dose: 99.7% versus 96%). Triple-dose regimens outperformed single doses against T. trichiura; three doses of mebendazole – the most efficacious treatment tested – cured 71% (95% CI: 57–82%). Both single and triple doses of either drug were highly efficacious against A. lumbricoides (CR: 93–97%; ERR: all >99.9%). Triple dose regimens cured all Taenia spp. infections, whereas single dose applications cured only half of them.

Conclusions/Significance

Single-dose oral albendazole is more efficacious against hookworm than mebendazole. To achieve high CRs against both hookworm and T. trichiura, triple-dose regimens are warranted.

Trial Registration

www.controlled-trials.com ISRCTN47375023