Vimentin is a target of PKCβ phosphorylation in MCP-1-activated primary human monocytes

Objective and design

We designed a study to detect downstream phosphorylation targets of PKCβ in MCP-1-induced human monocytes.

Methods

Two-dimensional gel electrophoresis was performed for monocytes treated with MCP-1 in the presence or absence of PKCβ antisense oligodeoxyribonucleotides (AS-ODN) or a PKCβ inhibitor peptide, followed by phospho- and total protein staining. Proteins that stained less intensely with the phospho-stain, when normalized to the total protein stain, in the presence of PKCβ AS-ODN or the PKCβ inhibitor peptide, were sequenced.

Results

Of the proteins identified, vimentin was consistently identified using both experimental approaches. Upon ³²P-labeling and vimentin immunoprecipitation, increased phosphorylation of vimentin was observed in MCP-1 treated monocytes as compared to the untreated monocytes. Both PKCβ AS-ODN and the PKCβ inhibitor reduced MCP-1-induced vimentin phosphorylation. The IP of monocytes with anti-vimentin antibody and immunoblotting with a PKCβ antibody revealed that increased PKCβ becomes associated with vimentin upon MCP-1 activation. Upon MCP-1 treatment, monocytes were shown to secrete vimentin and secretion depended on PKCβ expression and activity.

Conclusions

We conclude that vimentin, a major intermediate filament protein, is a phosphorylation target of PKCβ in MCP-1-treated monocytes and that PKCβ phosphorylation is essential for vimentin secretion. Our recently published studies have implicated vimentin as a potent stimulator of the innate immune receptor Dectin-1 as reported by Thiagarajan et al. (Cardiovasc Res 99:494–504, 2013). Taken together our findings suggest that inhibition of PKCβ regulates vimentin secretion and, thereby, its interaction with Dectin-1 and downstream stimulation of superoxide anion production. Thus, PKCβ phosphorylation of vimentin likely plays an important role in propagating inflammatory responses.     

Effects of a Web-Based Tailored Multiple-Lifestyle Intervention for Adults: A Two-Year Randomized Controlled Trial Comparing Sequential and Simultaneous Delivery Modes

Background

Web-based computer-tailored interventions for multiple health behaviors can have a significant public health impact. Yet, few randomized controlled trials have tested this assumption.

Objective

The objective of this paper was to test the effects of a sequential and simultaneous Web-based tailored intervention on multiple lifestyle behaviors.

Methods

A randomized controlled trial was conducted with 3 tailoring conditions (ie, sequential, simultaneous, and control conditions) in the Netherlands in 2009-2012. Follow-up measurements took place after 12 and 24 months. The intervention content was based on the I-Change model. In a health risk appraisal, all respondents (N=5055) received feedback on their lifestyle behaviors that indicated whether they complied with the Dutch guidelines for physical activity, vegetable consumption, fruit consumption, alcohol intake, and smoking. Participants in the sequential (n=1736) and simultaneous (n=1638) conditions received tailored motivational feedback to change unhealthy behaviors one at a time (sequential) or all at the same time (simultaneous). Mixed model analyses were performed as primary analyses; regression analyses were done as sensitivity analyses. An overall risk score was used as outcome measure, then effects on the 5 individual lifestyle behaviors were assessed and a process evaluation was performed regarding exposure to and appreciation of the intervention.

Results

Both tailoring strategies were associated with small self-reported behavioral changes. The sequential condition had the most significant effects compared to the control condition after 12 months (T1, effect size=0.28). After 24 months (T2), the simultaneous condition was most effective (effect size=0.18). All 5 individual lifestyle behaviors changed over time, but few effects differed significantly between the conditions. At both follow-ups, the sequential condition had significant changes in smoking abstinence compared to the simultaneous condition (T1 effect size=0.31; T2 effect size=0.41). The sequential condition was more effective in decreasing alcohol consumption than the control condition at 24 months (effect size=0.27). Change was predicted by the amount of exposure to the intervention (total visiting time: beta=–.06; P=.01; total number of visits: beta=–.11; P<.001). Both interventions were appreciated well by respondents without significant differences between conditions.

Conclusions

Although evidence was found for the effectiveness of both programs, no simple conclusive finding could be drawn about which intervention mode was more effective. The best kind of intervention may depend on the behavior that is targeted or on personal preferences and motivation. Further research is needed to identify moderators of intervention effectiveness. The results need to be interpreted in view of the high and selective dropout rates, multiple comparisons, and modest effect sizes. However, a large number of people were reached at low cost and behavioral change was achieved after 2 years.

Trial Registration

Nederlands Trial Register: NTR 2168; http://www.trialregister.nl/trialreg/admin/rctview.asp?TC=2168 (Archived by WebCite at http://www.webcitation.org/6MbUqttYB).     

Robotic radical prostatectomy as the initial step in multimodal therapy for men with high-risk localised prostate cancer: initial experience of 160 men

Study Type – Therapy (case series) Level of Evidence 4 What's known on the subject? and What does the study add? The choice of therapy with high‐risk localised prostate cancer is difficult and, in the stark absence of any randomised trials, comparative retrospective analyses of case series continue to be necessary. Radical surgery has been considered by many to be inferior to a combination of radiotherapy (RT) and androgen deprivation therapy (ADT), but this changing perhaps coincidentally with the widespread acceptance of robot‐assisted laparoscopic prostatectomy surgery (RALP). Further evidence has now described the long‐term toxicities related to ADT, and this has strengthened a desire amongst many to at least defer, if not avoid, ADT unless absolutely necessary. This article presents a single‐centre experience of RALP in the setting of high‐risk localised disease, and concludes that RALP incorporating the use of post‐operative RT represents a strong perhaps optimum management strategy.

OBJECTIVES

• ? To report the outcome of robotic‐assisted laparoscopic radical prostatectomy (RALP) for men with localised high‐risk prostate cancer at diagnosis.
• ? Although commonly managed by radiotherapy (RT) with prolonged androgen‐deprivation therapy (ADT), we hypothesize that initiation of multimodal therapy with RALP is oncologically efficacious and may allow many men to avoid ADT.

PATIENTS AND METHODS

• ? Between December 2003 and September 2010, 1480 men underwent RALP of whom 160 fulfilled the National Comprehensive Control Network criteria for high‐risk disease (prostate‐specific antigen (PSA) >20 ng/mL and/or clinical stage, cT ≥ 3 and/or biopsy Gleason score ≥8).
• ? Biochemical recurrence (postoperative PSA ≥ 0.2) was used to assess outcome after RALP monotherapy.
• ? Treatment failure was defined as either a rising PSA level after salvage RT or the initiation of ADT.

RESULTS

• ? The mean age ± standard deviation was 63.1 ± 6.3 years. Median PSA level was 9.95 ng/mL (interquartile range 6.0–21.4).
• ? Analysis of prostatectomy specimen showed Gleason 8–10 cancers in 65 (41%), and extracapsular disease, pT ≥ 3, in 96 (60%) of which seminal vesicle invasion was evident in 36 (23%). Downgrading by prostatectomy occurred in 64 (40% of total group) and five (3%) were downstaged to pT2 disease. By contrast, any upgrading occurred in 29 (18% of total group) and upstaging occurred in 68 (43%). The overall positive surgical margin rate was 38%, correlating with stage pT2 (15%) or pT3 (53%).
• ? With median follow‐up of 26.2 months (interquartile range 5.5–37.3), two non‐cancer‐related deaths have occurred (overall survival 98.8%; cancer‐specific survival 100%), and biochemical recurrence has occurred in 53 men (33%). RALP surgery has served as monotherapy (n= 117, 73%), or has been followed by salvage RT (n= 24, 15%) and/or ADT (n= 43, 27%). Overall 2‐year and 3‐year treatment failure was 31 and 41%, respectively.
• ? Serum PSA level was the only independent predictor of overall treatment failure (hazard ratio [HR] 1.02, P= 0.001) although a strong trend was observed for both clinical stage (HR 1.22, P= 0.058) and the number of positive biopsy cores on transrectal biopsy (HR 1.06, P= 0.057).

CONCLUSIONS

• ? RALP incorporating the use of postoperative RT is a good multimodal management strategy for men with this aggressive variant of prostate cancer.
• ? At median follow‐up in excess of 2 years, we found low rates of treatment failure enabling a high proportion of men to remain free of ADT.

     

Principles for designing future regimens for multidrug-resistant tuberculosis

Fewer than 20% of patients with multidrug-resistant (MDR) tuberculosis are receiving treatment and there is an urgent need to scale up treatment programmes. One of the biggest barriers to scale-up is the treatment regimen, which is lengthy, complex, ineffective, poorly tolerated and expensive. For the first time in over 50 years, new drugs have been developed specifically to treat tuberculosis, with bedaquiline and potentially delamanid expected to be available soon for treatment of MDR cases. However, if the new drugs are merely added to the current treatment regimen, the new regimen will be at least as lengthy, cumbersome and toxic as the existing one. There is an urgent need for strategy and evidence on how to maximize the potential of the new drugs to improve outcomes and shorten treatment. We devised eight key principles for designing future treatment regimens to ensure that, once they are proven safe in clinical trials, they will be clinically effective and programmatically practicable. Regimens should contain at least one new class of drug; be broadly applicable for use against MDR and extensively drug-resistant Mycobacterium tuberculosis complex strains; contain three to five effective drugs, each from a different drug class; be delivered orally; have a simple dosing schedule; have a good side-effect profile that allows limited monitoring; last a maximum of 6 months; and have minimal interaction with antiretrovirals. Following these principles will maximize the potential of new compounds and help to overcome the clinical and programmatic disadvantages and scale-up constraints that plague the current regimen.