Lentiviral vectors pLKO.1 TRC (Addgene 10879) and pWPI.1 (Addgene 12254) were used for constructing

recombinant lentiviruses. Oligonucleotides encoding hairpin precursors for si259 (5′-TTTCCAGGAGCAACCA TAATT-3′, corresponding to nt259-279 of PROX1 ORF) and si1646 (5′-GGCTCTCCTTGTCGCTCA TAA-3′, corresponding to nt1646-1666 of PROX1 ORF) were used for generating short interference RNA (siRNA) constructs. A scrambled sequence (Scr) was used as a control. Synonymous mutations were introduced into the target sequence of si1646 (5′-GGCTCTCATTATCACTCATAA-3′, mutations underlined) in PROX1 ORF to generate si1646-resistant PROX1. pEGFP-Prox1 was generated by inserting PROX1 cDNA into pEGFP-C2 (ClonTech, Mountain View, CA). pGL2-HIF-1α C646 contained the promoter (−572/+284) of HIF-1α in pGL2-Basic (Promega, Madison, WI). Cell transfection and luciferase reporter assays were performed as described.[22] The procedures are detailed in the Supporting Methods. Co-IP, GST pulldown, and western blot were performed as described.[22] Antibodies used in western blot experiments are listed in Supporting Table S2. The procedures are detailed in the Supporting

Methods. Anti-PROX1 monoclonal antibody (mAb) (Cell Signaling Technology, Danvers, MA) was used to immunoprecipitate sonicated chromatins prepared from Huh7 or MHCC-97H cells. Preimmune IgG was used for specificity control. Immunoprecipitated DNA was GPCR Compound Library purchase quantitated for HIF-1α promoter (Ct[IP]) using qrtPCR (forward, 5′-GGTGAGGCGGGCTT GCGGGAG-3′; reverse, 5′-GAGGAGCTGAGGCAG CGTCAGGG-3′). DNA from 5% input was quantitated for HIF-1α promoter in parallel (Ct[Input]). The relative occupancy was calculated using the equation: 2^(Ct[Input]-Ct[IP]) *100%. The procedures are detailed in the Supporting Methods. Animal experimental protocols were approved by the Animal Ethics Committee of Shanghai Medical College, Fudan University. Eight-week-old nude mice (BALB/c) were randomly divided into groups

(six mice/group) before inoculation or injection. Cells were inoculated into the liver parenchyma of nude mice (BEL-7402 derived cells, 2.0 × 106 cells/mouse) or subcutaneously injected into nude mice (MHCC-97H derived cells, 1.0 × 107 cells/mouse). The mice were sacrificed after 8 weeks and the number of metastatic 上海皓元医药股份有限公司 tumors was assessed by double-blinded evaluation. Statistical analysis was performed with SPSS v. 13.0. Kaplan-Meier analysis was used for survival analysis and the log-rank test was chosen to compare the difference. Pearson χ2 test or Fisher’s exact test were employed to compare qualitative variables, while Student t test was used for quantitative variables. A Cox proportional hazards model was adopted for multivariate analysis. Receiver operating characteristic (ROC) curve analysis was applied to assess the predictive values of variables. P < 0.05 was considered statistically significant for all tests.

Lentiviral vectors pLKO.1 TRC (Addgene 10879) and pWPI.1 (Addgene 12254) were used for constructing

recombinant lentiviruses. Oligonucleotides encoding hairpin precursors for si259 (5′-TTTCCAGGAGCAACCA TAATT-3′, corresponding to nt259-279 of PROX1 ORF) and si1646 (5′-GGCTCTCCTTGTCGCTCA TAA-3′, corresponding to nt1646-1666 of PROX1 ORF) were used for generating short interference RNA (siRNA) constructs. A scrambled sequence (Scr) was used as a control. Synonymous mutations were introduced into the target sequence of si1646 (5′-GGCTCTCATTATCACTCATAA-3′, mutations underlined) in PROX1 ORF to generate si1646-resistant PROX1. pEGFP-Prox1 was generated by inserting PROX1 cDNA into pEGFP-C2 (ClonTech, Mountain View, CA). pGL2-HIF-1α Nutlin-3a cost contained the promoter (−572/+284) of HIF-1α in pGL2-Basic (Promega, Madison, WI). Cell transfection and luciferase reporter assays were performed as described.[22] The procedures are detailed in the Supporting Methods. Co-IP, GST pulldown, and western blot were performed as described.[22] Antibodies used in western blot experiments are listed in Supporting Table S2. The procedures are detailed in the Supporting

Methods. Anti-PROX1 monoclonal antibody (mAb) (Cell Signaling Technology, Danvers, MA) was used to immunoprecipitate sonicated chromatins prepared from Huh7 or MHCC-97H cells. Preimmune IgG was used for specificity control. Immunoprecipitated DNA was PS-341 in vivo quantitated for HIF-1α promoter (Ct[IP]) using qrtPCR (forward, 5′-GGTGAGGCGGGCTT GCGGGAG-3′; reverse, 5′-GAGGAGCTGAGGCAG CGTCAGGG-3′). DNA from 5% input was quantitated for HIF-1α promoter in parallel (Ct[Input]). The relative occupancy was calculated using the equation: 2^(Ct[Input]-Ct[IP]) *100%. The procedures are detailed in the Supporting Methods. Animal experimental protocols were approved by the Animal Ethics Committee of Shanghai Medical College, Fudan University. Eight-week-old nude mice (BALB/c) were randomly divided into groups

(six mice/group) before inoculation or injection. Cells were inoculated into the liver parenchyma of nude mice (BEL-7402 derived cells, 2.0 × 106 cells/mouse) or subcutaneously injected into nude mice (MHCC-97H derived cells, 1.0 × 107 cells/mouse). The mice were sacrificed after 8 weeks and the number of metastatic MCE tumors was assessed by double-blinded evaluation. Statistical analysis was performed with SPSS v. 13.0. Kaplan-Meier analysis was used for survival analysis and the log-rank test was chosen to compare the difference. Pearson χ2 test or Fisher’s exact test were employed to compare qualitative variables, while Student t test was used for quantitative variables. A Cox proportional hazards model was adopted for multivariate analysis. Receiver operating characteristic (ROC) curve analysis was applied to assess the predictive values of variables. P < 0.05 was considered statistically significant for all tests.

Instead, the vast majority is located in the cytoplasm,17, 32 although the underlying mechanism that regulates cytoplasmic and nuclear localization of Twist1 is not clear. It has been reported that the NLS in Twist1 can lead to the interaction between Twist1 and the nuclear membrane pore channel and the NLS can also induce Twist1 to enter the

nucleus and act as a transcription factor.33 In the present study we provide data to show that the up-regulation of Twist1 reaches its peak level 24 hours after hypoxia, whereas the expression of Twist1 decreased after 24 hours, as many of these selleck products cells die due to continued hypoxia. When hypoxia was relieved after 24 hours, the high expression level signaling pathway of Twist1 can be sustained for more than 24 hours. Interestingly, the antiapoptotic protein Bcl-2 also exhibited an expression peak and trend similar to Twist1 within the same period (24 hours). This result indicated that Bcl-2 and Twist1 possibly acted during the stress phase in the same cell and followed similar kinetics. Bcl-2 and its family members

have been found to mediate the apoptosis process. They have also been found to participate in protein modification and to form a complex with other proteins for participating in complicated processes of cell metabolism.2, 3, 34 In tumor tissues, Bcl-2 expression in the nucleus correlates with poor prognosis. Our data provide evidence that Bcl-2 may form a complex with Twist1 and synergistically to promote the transcription of downstream target genes which can lead a cascade changes in proliferation, adhesion, migration, infestation, clone formation, and tubal formation of tumor cells. The formation of Bcl-2 with Twist1 as a protein complex to stimulate the transcription was unexpected. Bcl-2

has long been considered as a mitochondrial membrane protein. However, reports on the effects of Bcl-2 and its other family members in more complex biological processes are limited. The present study revealed that specific amino acids within Bcl-2 and 上海皓元医药股份有限公司 Twist1 are involved in the binding of two proteins and form a novel functional complex and jointly enter the nucleus, which leads to changes of multiple downstream target genes. Such a heterodimer is more potent in stimulating the transcription of multiple downstream target genes than Twist1 alone. Although the detailed mechanisms for interaction between Twist1 and Bcl-2 are not clear at this time, we speculate that the following mechanisms may be involved. Bcl-2 may be initially associated with the nuclear membrane pore structure, and assists Twist1 in entering the nucleus. In the nucleus, Bcl-2 and Twist1 forms a protein complex and functions in synergy on the promoters of different target genes to regulate their transcription.

45, 46 Those markers were found to be down-regulated upon HO-1 induction, further indicating inhibition of proliferation. These results are in line with JAK2 inhibitors clinical trials our observation that HO-1 induction reduced early signs of dysplasia and indicate that HO-1 induction at early time points (e.g., during inflammation or fibrosis) might have consequences on subsequent progression to HCC. Preliminary results even indicate that early HO-1 induction might interfere with progression to HCC. HO-1 has been shown to be overexpressed in alcohol-induced HCC in patients.47 Moreover, HO-1 down-regulation via short interfering

RNA significantly decreased tumor growth, whereas it increased cellular damage and apoptosis.47 Therefore, in the liver, HO-1 overexpression seems to exert beneficial click here or detrimental effects, depending on pathological conditions (e.g., inflammation or solid tumor formation). However, tumor-promoting effects by early HO-1 induction are unlikely, because induced HO-1 protein is degraded within days after treatment, whereas anti-inflammatory effects of HO-1 induction seem to last for at least 8 weeks longer than treatment with CoPP. Therefore, follow-up experiments have to determine the consequences of early HO-1

induction on progression to HCC caused by chronic inflammation. The expert technical assistance by Elena Tassika, Christine Loscher, and Nicola Peters 上海皓元医药股份有限公司 is gratefully acknowledged. Additional Supporting Information may be found in the online version of this article. “
“Background and Aim:  The Frequency Scale for the Symptoms of Gastroesophageal Reflux Disease (FSSG) is the standard questionnaire used in

Japan for the diagnosis of gastroesophageal reflux disease (GERD) and assessment of the response to treatment. We modified the FSSG in order to assess dyspepsia symptoms, and evaluated the modified questionnaire. Methods:  We modified the FSSG by adding two questions on interdigestive and postprandial epigastric pain. We then assessed the modified FSSG with 100 new untreated symptomatic patients presenting to hospital and in 200 subjects undergoing health checks. Endoscopic assessment of the esophagogastric junction was performed according to the modified Los Angeles classification with addition of Grade N (normal appearance) and Grade M (minimal change). Endoscopic images were assessed by five experienced endoscopists blinded to the questionnaire results. Results:  The 100 new patients included 16 with erosive GERD (>Grade A), 12 with peptic ulcer, and two with gastric cancer. Among the 70 patients with no evidence of organic disease, the modified FSSG diagnosed functional dyspepsia (FD) in 41 and non-erosive gastric disease (NERD) in 29. A significant difference was seen in the dyspepsia score between patients with FD and NERD.

Methods: Data were drawn from the International Collaboration of Incident HIV and Hepatitis C in Injecting Cohorts (InC3 Study) including nine prospective cohorts of people who inject drugs. Factors associated with high HCV-RNA levels (>5.6 log IU/mL=400,000 IU/mL) during the first two months post-infection AP24534 were assessed. Among those with viral persistence, factors associated with high HCV-RNA levels (>5.6 log IU/mL) at one year (8-16 month window) post-infection were also assessed. Logistic regression was used in analyses. Results: Among participants with detectable HCV-RNA during the first two months post-infection (n=178), interferon lambda 3 (IFNL3) CC genotype (vs. TT/CT; adjusted

odds ratio [AOR]: 3.05; 95%CI: 1.48, 6.27; P=0.002) was the only factor associated with high HCV-RNA levels. Among those with persistent HCV infection (n=308), male sex (vs. female, AOR: 1.93; 95%CI: 1.01, 3.69; P=0.046), IFNL3 CC genotype (vs. TT/CT; AOR: 2.48; 95%CI: 1.42, 4.35; P=0.001), HIV co-infection (vs. no HIV; AOR: 3.27; 95%CI: 1.35, 7.93; P=0.009), and HCV genotype G2 (vs. G3; AOR: 5.40; 95%CI: 1.63, 17.84; P=0.006) were independently associated with higher HCV-RNA levels. HCV G1 (vs. G3; AOR: 1.87; 95%CI: 0.99,

3.55; P=0.054) trended towards being associated with higher HCV-RNA. Conclusion: HCV-RNA levels were independently associated with IFNL3 genotype during the first two months post-infection and IFNL3 genotype, sex, HIV co-infection, and HCV genotype find more at one year post-infection. During chronic infection, factors influencing HCV-RNA levels exert their effects as early as one year following infection. Further research is needed to understand the interplay between the role of gender, host genetics and viral genotype in the pathogenesis of HCV infection. Disclosures: MCE公司 Arthur Y. Kim – Consulting: Abbvie Pharmaceuticals, Gilead Pharmaceuticals; Grant/Research Support: Bristol-Myers Squibb, Gilead Pharmaceuticals Barbara H. McGovern – Employment: AbbVie Andrew R. Lloyd – Grant/Research Support: Merck Maria Prins – Speaking and Teaching:

msd, roche Gregory J. Dore – Board Membership: Bristol-Myers Squibb, Roche, Gilead, Merck, Janssen, Abbvie; Grant/Research Support: Janssen, Bristol-Myers Squibb, Vertex, Roche, Gilead, Merck, Abbvie; Speaking and Teaching: Roche, Merck, Janssen Jason Grebely – Advisory Committees or Review Panels: Merck, Gilead; Grant/ Research Support: Merck, Gilead, Abbvie, BMS The following people have nothing to disclose: Behzad Hajarizadeh, Bart P. Grady, Kimberly Page, Andrea Cox, Thomas M. Rice, Rachel Sacks-Davis, Julie Bruneau, Meghan D. Morris, Janaki Amin, Janke Schinkel, Tanya L. Applegate, Lisa Maher, Margaret Hellard, Ronald Geskus Background: Daclatasvir (DCV) is an HCV NS5A inhibitor with potent antiviral activity and broad genotype coverage. Exhaustive information about resistance-associated variants (RAVs) to DCV is available for GT1, but current migratory waves are increasing interest in GT4.

Early studies suggested that reproductive suppression in subordinate females was caused by chronic elevation of glucocorticoid adrenal hormones as a result of social ‘stress’ induced by regular aggression from dominants (Wasser & Barash, 1983). However, recent research has shown that the presence of dominant females, or cues signalling their presence, can, on their

own, prevent subordinate females from Carfilzomib in vitro mating or conceiving in the absence of direct interactions with dominant females (French, 1997; Young, 2009). For example, in naked mole rats, the presence of dominant females is sufficient to prevent subordinate females in coming into breeding condition (Faulkes et al., 1997) while proximity of dominant females is sufficient to inhibit mating in several primates (Townsend, Deschner, & Zuberbuhler, 2008; Overduin-de Vries et al., 2013). Moreover, in some species, cortisol levels do not vary consistently between subordinates

and dominants (Abbott et al., 2002; Starling et al., 2010), while, in other species, subordinates show lower glucocorticoid levels than dominants (Creel, 2001) and these results are commonly interpreted as evidence that glucocorticoid levels associated with aggression are not responsible for reproductive suppression. However, an alternative explanation is that the Talazoparib manufacturer relationship between social status and glucocorticoid levels depends on the structure of societies and the 上海皓元医药股份有限公司 relative costs of acquiring and maintaining dominance, as well as on the relative intensity and frequency of threats faced by subordinates from dominants (Goymann & Wingfield, 2004; Rubenstein

& Shen, 2009). Dominants may exhibit higher cortisol levels than subordinates in species where maintaining dominance requires frequent physical contests, but not where dominance is inherited and stable as in female spotted hyenas. In addition, the physiological costs of social status can even vary within species, in relation to fluctuations in the level of social conflict. For example, reproductive suppression may be induced by substantial increases of glucocorticoid levels in subordinates at times where they are attempting to breed and are the target of frequent aggression by dominants (Young, 2009). Although the role of aggression in reproductive suppression has attracted most attention, it is clear that several other factors can be involved.

Elderly patients and those with more functional limitations (with lower HAL scores) were significantly less active. Sports participation levels were in line with other PWH at 66%. Although 55% reported bleeds resulting from sport and 31% reported having had a significant injury attributable to sport, overall sport was viewed in a positive light. Irish PWH are physically active and play a wide range of sports. Further efforts are needed to achieve optimal safety and to ensure that maximum benefit is gained. “
“Summary.  Radiosynoviorthesis is a safe and easy method for synovectomy in haemophilic arthropathy. Palbociclib research buy Various agents have been used in radiosynoviorthesis,

especially newly developed Etoposide order agent Holmium-166-chitosan complex has good clinical outcome. This study analysed clinical results and radiologic evaluation of radioisotope synoviorthesis using Holmium-166-chitosan complex in haemophilic arthropathy. From March 2001 to December 2003, 58 radiosynoviorthesis were performed in 53 haemophiliacs. The average age at procedure was 13.8

years. The Arnold and Hilgartner stage of the patients was from I to IV. Holmium-166-chitosan complex was injected in 31 ankle joints, 19 elbow joints and 8 knee joints. Average follow-up was 33 months since primary procedure. The range of motion of each medchemexpress joint, frequency of intra-articular bleeding and factor dose used were analysed

for clinical assessment. There was no significant improvement of range of motion in affected joints. After procedure, the average frequency of bleeding of the elbow joint has decreased from 3.76 to 0.47 times per month, the knee joint from 5.87 to 1.12 times per month, and the ankle joint from 3.62 to 0.73 times per month respectively (P < 0.05). After treatment, the average coagulation factor dose injected was significantly decreased to 779.3 units per month from 2814.8 units per month before treatment (P < 0.001). Radioisotope synoviorthesis with Holmium-166-chitosan complex in haemophilic arthropathy is a very safe and simple procedure with the expectation of a satisfactory outcome without serious complication. It has excellent bleeding control effect on target joint and the need for substitution of coagulation factor concentrate can be reduced. "
“Factor-Eight-Inhibitor-Bypassing-Activity (FEIBA) is a bypassing-agent used to control spontaneous bleeding or cover surgical interventions in Haemophiliacs who develop neutralizing antibodies against FVIII/FIX. The market lot-release of FEIBA is dependent on specific clot-based assays, carried out by both the manufacturer and regulatory authorities, relative to manufacturer’s in-house standards, which are produced on a small-scale and are replaced frequently.

Until

recently, a salient exception appeared to be the stegosaurs, whose low diversity was anomalous to this general model. However, new discoveries of stegosaurs have increased our knowledge of their diversity: Carpenter this website (2001) estimates that at least five stegosaur species are now known from the Morrison Formation of the western United States (although Galton & Upchurch (2004b) recognize only three). This would appear to simplify the problem, but there is an additional caveat: the species are not all contemporaneous (K. Carpenter, pers. comm., 2004), and there may be geographic differentiation as well within the Morrison. The lack of contemporaneity could have several explanations, including insufficient stratigraphic sampling to establish that more of these species lived at the same time than it now appears. However, another approach is phylogenetic. If these five species turned selleck inhibitor out to be morphotypes of a single anagenetic lineage, there would indeed be no evidence for contemporaneity. Would the hypothesis of species recognition thereby be weakened (Fig. 7, left)? In fact, such a result would weaken a hypothesis of anti-hybridization, but it would not

weaken or test the hypothesis of positive assortative mating (Paterson, 1993). However, if phylogenetic analysis revealed that these species indeed represented different lineages, and their ‘ghost ranges’ indicated that they must have diverged from others

at an earlier time, then at one time the test of contemporaneous species would have been passed (Fig. 7, right). 上海皓元医药股份有限公司 It is not impossible that such a pattern could also indicate other processes than species recognition, such as sexual or social selection, but in concert with non-directional evolutionary change the indication would be rather more strongly in favor of species recognition. Phylogenetic analysis and further biostratigraphic sampling can test this hypothesis. Finally, we return to the test of the Mate Recognition Hypothesis that Sampson (1999) proposed. We found that in every criterion, mostly related to higher rates of speciation and habitat shifts, the concept of ‘species recognition’ could be substituted for the terms related to sexual selection without any apparent difference in results. The exception was his fourth criterion (speciation will often be correlated with vicariance events rather than the formation of peripheral isolates), which we suggest is untestable in the fossil record, and in any case would not discriminate between sexual selection and species recognition as a cause.

Polymorphisms in the genes coding for interleukin [IL]-10 [16], tumour necrosis factor-α (TNF-α) [17], cytotoxic T-lymphocyte antigen-4 (CTLA-4) have been identified as genetic factors in the context of the Malmö International Brother Study [18,19]. Specific major histocompatibility complex class human leucocyte antigen (MHC HLA) genes (class I and II alleles) may also be implicated in increasing the risk of inhibitor development but these results are controversial [20]. It has been suggested that such genetic

factors form either an ‘unsafe’ or ‘safe’ platform for the inhibitors to develop, depending on whether they constitute a more or less dangerous pattern that could be triggered by some environmental selleck products events (Fig. 1a and b) [19]. The risk of inhibitor development will be low in patients with a ‘safe’ platform, even in the case of challenges providing ‘danger signals’ for the immune system (Fig. 1a). Conversely, patients with an ‘unsafe’ platform might

experience challenges to the immune system that reach the threshold for inhibitors to develop (Fig. 1b). An additional factor related to inhibitor development risk is ethnicity, with a particularly high risk associated with patients of an African-American origin [13,21]. The influence of other ethnic groups is an unresolved issue that needs to be addressed in future clinical studies. Environmental influences that are implicated in increasing the risk of inhibitor formation can be viewed as modifiable risk factors. Identifying environmental Wnt inhibitor risk factors for increasing the probability of inhibitor development affords the potential to intervene, and thereby modify patient treatment and outcomes. This would allow for improved anticipation of disease progression and permit prophylaxis to be tailored to individual patients. Evidence

from studies varies with respect to the effect on inhibitor formation of high intensity MCE therapy and exposure to clotting factors at an early age [22–28]. Data from several studies have supported the idea that first replacement therapy at an early age may increase the risk of inhibitor formation [26–28]. Lorenzo et al. reported first that the estimated cumulative incidence of inhibitors at 3 years was significantly higher in those initiating therapy before 6 months of age compared with patients starting with treatment between 6 and 12 months or those treated at age >12 months (41% vs. 29% and 12% respectively, P = 0.03) [26]. These results have been supported by van der Bom et al. who reported that the earlier the exposure to FVIII in infancy (at the age of <6 months), the higher the risk of developing inhibitors later in life (P for trend = 0.03) [27]. Furthermore, Santagostino et al.

01). Conclusions.— Our data support the continuum concept of headache, one in which noxious cervical afferent information may well be significantly underestimated. The high incidence of reproduction of headache supports the evaluation of musculoskeletal

features in patients presenting with migrainous and TTH symptoms. This, in turn, may have important implications for understanding the pathophysiology of headache and developing alternative treatment options. “
“(Headache 2010;50:921-936) Objective.— To assess the efficacy, safety, and tolerability of onabotulinumtoxinA (BOTOX®) as headache prophylaxis in adults with chronic migraine. Background.— Chronic migraine is a prevalent, disabling, and undertreated neurological mTOR inhibitor disorder. Few preventive treatments have been investigated and none is specifically indicated for chronic migraine. Methods.— The 2 multicenter, pivotal trials in the PREEMPT: Phase 3 REsearch Evaluating Migraine Prophylaxis Therapy clinical program each included a 24-week randomized, double-blind phase followed by a 32-week open-label phase (ClinicalTrials.gov identifiers NCT00156910, NCT00168428). Talazoparib in vitro Qualified patients were randomized (1:1) to onabotulinumtoxinA (155-195 U) or

placebo injections every 12 weeks. Study visits occurred every 4 weeks. These studies were identical in design (eg, inclusion/exclusion criteria, randomization, visits, double-blind phase, open-label phase, safety assessments, treatment), with the only exception being the designation of the primary and secondary endpoints. Therefore, the predefined pooling of the results was justified and performed to provide a complete overview of between-group differences in efficacy, safety, and tolerability that may not have been evident in individual studies. The primary endpoint for the pooled analysis was mean change from baseline in frequency of headache days at 24 weeks. Secondary endpoints were mean change from baseline to week 24 in frequency of migraine/probable migraine MCE公司 days, frequency of moderate/severe headache days, total cumulative hours of headache on headache days, frequency of headache episodes, frequency of migraine/probable migraine episodes, frequency of acute headache

pain medication intakes, and the proportion of patients with severe (≥60) Headache Impact Test-6 score at week 24. Results of the pooled analyses of the 2 PREEMPT double-blind phases are presented. Results.— A total of 1384 adults were randomized to onabotulinumtoxinA (n = 688) or placebo (n = 696). Pooled analyses demonstrated a large mean decrease from baseline in frequency of headache days, with statistically significant between-group differences favoring onabotulinumtoxinA over placebo at week 24 (−8.4 vs −6.6; P < .001) and at all other time points. Significant differences favoring onabotulinumtoxinA were also observed for all secondary efficacy variables at all time points, with the exception of frequency of acute headache pain medication intakes.