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Thiago De Marchi,1,2 Fabiano Frâncio,1,3 João Vitor Ferlito,3 Renata Weigert,3 Cristiane de Oliveira,3 Ana Paula Merlo,3 Délcio Luis Pandini,3 Bolivar Antônio Pasqual-Júnior,3 Daniela Giovanella,3 Shaiane Silva Tomazoni,4,5 Ernesto Cesar Leal-Junior2,4,5 1University Center of Bento Gonçalves (UNICNEC), Bento Gonçalves, Rio Grande do Sul, Brazil; 2Laboratory of Phototherapy and Innovative Technologies in Health (LaPIT), Post-graduate Program in Rehabilitation Sciences, Nove de Julho University, São Paulo, Brazil; 3Hospital Tacchini, Bento Gonçalves, Rio Grande do Sul, Brazil; 4Physiotherapy Research Group, Department of Global Public Health and Primary Care, University of Bergen, Bergen, Norway; 5ELJ Consultancy, Scientific Consultants, São Paulo, BrazilCorrespondence: Shaiane Silva TomazoniPhysiotherapy Research Group, Department of Global Public Health and Primary Care, University of Bergen, Alrek helseklynge, blokk D, Årstadveien 17, Bergen, NO-5020, NorwayEmail shaiane.tomazoni@eljconsultancy.comPurpose: We aimed to investigate the effects of photobiomodulation therapy combined with static magnetic field (PBMT-sMF) on the length of intensive care unit (ICU) stay and mortality rate of severe COVID-19 patients requiring invasive mechanical ventilation and assess its role in preserving respiratory muscles and modulating inflammatory processes.Patients and Methods: We conducted a prospectively registered, triple-blinded, randomized, placebo-controlled trial of PBMT-sMF in severe COVID-19 ICU patients requiring invasive mechanical ventilation. Patients were randomly assigned to receive either PBMT-sMF or a placebo daily throughout their ICU stay. The primary outcome was length of ICU stay, defined by either discharge or death. The secondary outcomes were survival rate, diaphragm muscle function, and the changes in blood parameters, ventilatory parameters, and arterial blood gases.Results: Thirty patients were included and equally randomized into the two groups. There were no significant differences in the length of ICU stay (mean difference, MD = − 6.80; 95% CI = − 18.71 to 5.11) between the groups. Among the secondary outcomes, significant differences were observed in diaphragm thickness, fraction of inspired oxygen, partial pressure of oxygen/fraction of inspired oxygen ratio, C-reactive protein levels, lymphocyte count, and hemoglobin (p < 0.05).Conclusion: Among severe COVID-19 patients requiring invasive mechanical ventilation, the length of ICU stay was not significantly different between the PBMT-sMF and placebo groups. In contrast, PBMT-sMF was significantly associated with reduced diaphragm atrophy, improved ventilatory parameters and lymphocyte count, and decreased C-reactive protein levels and hemoglobin count.Trial Registration Number (Clinical Trials.gov): NCT04386694.Keywords: COVID-19, photobiomodulation, intensive care unit, respiratory muscles, ventilatory parameters, mechanical ventilation

In animal and clinical trials low-level laser therapy (LLLT) using red, infrared and mixed wavelengths has been shown to delay the development of skeletal muscle fatigue. However, the parameters employed in these studies do not allow a conclusion as to which wavelength range is better in delaying the development of skeletal muscle fatigue. With this perspective in mind, we compared the effects of red and infrared LLLT on skeletal muscle fatigue. A randomized double-blind placebo-controlled crossover trial was performed in ten healthy male volunteers. They were treated with active red LLLT, active infrared LLLT (660 or 830 nm, 50 mW, 17.85 W/cm(2), 100 s irradiation per point, 5 J, 1,785 J/cm(2) at each point irradiated, total 20 J irradiated per muscle) or an identical placebo LLLT at four points of the biceps brachii muscle for 3 min before exercise (voluntary isometric elbow flexion for 60 s). The mean peak force was significantly greater (p < 0.05) following red (12.14%) and infrared LLLT (14.49%) than following placebo LLLT, and the mean average force was also significantly greater (p < 0.05) following red (13.09%) and infrared LLLT (13.24%) than following placebo LLLT. There were no significant differences in mean average force or mean peak force between red and infrared LLLT. We conclude that both red than infrared LLLT are effective in delaying the development skeletal muscle fatigue and in enhancement of skeletal muscle performance. Further studies are needed to identify the specific mechanisms through which each wavelength acts. [ABSTRACT FROM AUTHOR]

STUDY DESIGN: Randomized crossover double-blinded placebo-controlled trial. OBJECTIVE: To investigate if low-level laser therapy (LLLT) can affect biceps muscle performance, fatigue development, and biochemical markers of postexercise recovery. BACKGROUND: Cell and animal studies have suggested that LLLT can reduce oxidative stress and inflammatory responses in muscle tissue. But it remains uncertain whether these findings can translate into humans in sport and exercise situations. METHODS: Nine healthy male volleyball players participated in the study. They received either active LLLT (cluster probe with 5 laser diodes; lambda = 810 nm; 200 mW power output; 30 seconds of irradiation, applied in 2 locations over the biceps of the nondominant arm; 60 J of total energy) or placebo LLLT using an identical cluster probe. The intervention or placebo were applied 3 minutes before the performance of exercise. All subjects performed voluntary elbow flexion repetitions with a workload of 75% of their maximal voluntary contraction force until exhaustion. RESULTS: Active LLLT increased the number of repetitions by 14.5% (mean +/- SD, 39.6 +/- 4.3 versus 34.6 +/- 5.6; P = .037) and the elapsed time before exhaustion by 8.0% (P = .034), when compared to the placebo treatment. The biochemical markers also indicated that recovery may be positively affected by LLLT, as indicated by postexercise blood lactate levels (P<.01), creatine kinase activity (P = .017), and C-reactive protein levels (P = .047), showing a faster recovery with LLLT application prior to the exercise. CONCLUSION: We conclude that pre-exercise irradiation of the biceps with an LLLT dose of 6 J per application location, applied in 2 locations, increased endurance for repeated elbow flexion against resistance and decreased postexercise levels of blood lactate, creatine kinase, and C-reactiveprotein. LEVEL OF EVIDENCE: Performance enhancement, level 1b.J Orthop Sports Phys Ther 2010;40(8):524-532; Epub 12 April 2010. doi:10.2519/jospt.2010.3294. [ABSTRACT FROM AUTHOR]

Three quarters of all breast cancers express the estrogen receptor (ER, ESR1 gene), which promotes tumor growth and constitutes a direct target for endocrine therapies. ESR1 mutations have been implicated in therapy resistance in metastatic breast cancer, in particular to aromatase inhibitors. ESR1 mutations promote constitutive ER activity and affect other signaling pathways, allowing cancer cells to proliferate by employing mechanisms within and without direct regulation by the ER. Although subjected to extensive genetic and transcriptomic analyses, understanding of protein alterations remains poorly investigated. Towards this, we employed an integrated mass spectrometry based proteomic approach to profile the protein and phosphoprotein differences in breast cancer cell lines expressing the frequent Y537N and Y537S ER mutations. Global proteome analysis revealed enrichment of mitotic and immune signaling pathways in ER mutant cells, while phosphoprotein analysis evidenced enriched activity of proliferation associated kinases, in particular CDKs and mTOR. Integration of protein expression and phosphorylation data revealed pathway-dependent discrepancies (motility vs proliferation) that were observed at varying degrees across mutant and wt ER cells. Additionally, protein expression and phosphorylation patterns, while under different regulation, still recapitulated the estrogen-independent phenotype of ER mutant cells. Our study is the first proteome-centric characterization of ESR1 mutant models, out of which we confirm estrogen independence of ER mutants and reveal the enrichment of immune signaling pathways at the proteomic level., (© 2024. The Author(s).)

Background: Immunohistochemical (IHC) PD-L1 expression is commonly employed as predictive biomarker for checkpoint inhibitors in triple-negative breast cancer (TNBC). However, IHC evaluation methods are non-uniform and further studies are needed to optimize clinical utility., Methods: We compared the concordance, prognostic value and gene expression between PD-L1 IHC expression by SP142 immune cell (IC) score and 22C3 combined positive score (CPS; companion IHC diagnostic assays for atezolizumab and pembrolizumab, respectively) in a population-based cohort of 232 early-stage TNBC patients., Results: The expression rates of PD-L1 for SP142 IC ≥ 1%, 22C3 CPS ≥ 10, 22C3 CPS ≥ 1 and 22C3 IC ≥ 1% were 50.9%, 27.2%, 53.9% and 41.8%, respectively. The analytical concordance (kappa values) between SP142 IC+ and these three different 22C3 scorings were 73.7% (0.48, weak agreement), 81.5% (0.63) and 86.6% (0.73), respectively. The SP142 assay was better at identifying 22C3 positive tumors than the 22C3 assay was at detecting SP142 positive tumors. PD-L1 (CD274) gene expression (mRNA) showed a strong positive association with all two-categorical IHC scorings of the PD-L1 expression, irrespective of antibody and cut-off (Spearman Rho ranged from 0.59 to 0.62; all p-values < 0.001). PD-L1 IHC positivity and abundance of tumor infiltrating lymphocytes were of positive prognostic value in univariable regression analyses in patients treated with (neo)adjuvant chemotherapy, where it was strongest for 22C3 CPS ≥ 10 and distant relapse-free interval (HR = 0.18, p = 0.019). However, PD-L1 status was not independently prognostic when adjusting for abundance of tumor infiltrating lymphocytes in multivariable analyses., Conclusion: Our findings support that the SP142 and 22C3 IHC assays, with their respective clinically applied scoring algorithms, are not analytically equivalent where they identify partially non-overlapping subpopulations of TNBC patients and cannot be substituted with one another regarding PD-L1 detection. Trial registration The Swedish Cancerome Analysis Network - Breast (SCAN-B) study, retrospectively registered 2nd Dec 2014 at ClinicalTrials.gov; ID NCT02306096., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Downregulation of the DNA repair protein WD40-encoding RNA antisense to p53 (WRAP53) has been associated with radiotherapy resistance and reduced cancer survival. The aim of this study was to evaluate WRAP53 protein and RNA levels as prognostic and predictive markers in the SweBCG91RT trial, in which breast cancer patients were randomized for postoperative radiotherapy. Using tissue microarray and microarray-based gene expression, 965 and 759 tumors were assessed for WRAP53 protein and RNA levels, respectively. Correlation with local recurrence and breast cancer-related death was assessed for prognosis, and the interaction between WRAP53 and radiotherapy in relation to local recurrence was assessed for radioresistance prediction. Tumors with low WRAP53 protein levels had a higher subhazard ratio (SHR) for local recurrence [1.76 (95% CI 1.10-2.79)] and breast cancer-related death [1.55 (1.02-2.38)]. Low WRAP53 RNA levels were associated with almost a three-fold decreased effect of radiotherapy in relation to ipsilateral breast tumor recurrence [IBTR; SHR 0.87 (95% CI 0.44-1.72)] compared with high RNA levels [0.33 (0.19-0.55)], with a significant interaction (P = 0.024). In conclusion, low WRAP53 protein is prognostic for local recurrence and breast cancer-related death. Low WRAP53 RNA is a potential marker for radioresistance., (© 2023 The Authors. Molecular Oncology published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.)

To review the effects of photobiomodulation therapy (PBMT) on pain intensity and disability in people with plantar fasciitis (PF) when compared with control conditions, other interventions, and adjunct therapies. Systematic searches were conducted in five database randomized controlled trials (RCT). We only included randomized controlled trials (RCTs) in adults with PF that compared PBMT to placebo, as well as RCTs that compared PBMT to other interventions; and as an adjunct to other therapies. The methodological quality and certainty were assessed through PEDro Scale and GRADE approach, respectively. The data of comparison were pooled and a meta-analysis was conducted when possible. Nineteen RCTs involving 1089 participants were included in this review. PBMT alone (MD =  - 22.02 [- 35.21 to - 8.83]) or with exercise (MD =  - 21.84 [- 26.14 to - 17.54]) improved pain intensity in short-term treatment. PBMT was superior to (extracorporeal shock wave therapy) EWST for relief of pain (MD =  - 20.94 [- 32.74 to - 9.13]). In the follow-up, PBMT plus exercise had a superior to exercise therapy alone (MD =  - 18.42 [- 26.48 to - 10.36]). PBMT may be superior to (ultrasound therapeutic) UST in medium- and long-term follow-ups for disability, but can be not clinically relevant. There is uncertainty that PBMT is capable of promoting improvement in disability. PBMT when used with adjuvant therapy does not enhance outcomes of interest. PBMT improves pain intensity with or without exercise. PBMT has been shown to be superior to ESWT for pain relief, but not superior to other interventions for pain intensity and disability. The evidence does not support PBMT as an adjunct to other electrotherapeutic modalities., (© 2023. The Author(s), under exclusive licence to Springer-Verlag London Ltd., part of Springer Nature.)

Background: The purpose of this review was to analyze the acute effects of low-load resistance exercise with blood flow restriction (LLE-BFR) on oxidative stress markers in healthy individuals in comparison with LLE or high-load resistance exercise (HLRE) without BFR., Materials and Methods: A systematic review was performed in accordance with the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) guidelines. These searches were performed in CENTRAL, SPORTDiscus, EMBASE, PubMed, CINAHL and Virtual Health Library- VHL, which includes Lilacs, Medline and SciELO. The risk of bias and quality of evidence were assessed through the PEDro scale and GRADE system, respectively., Results: Thirteen randomized clinical trials were included in this review (total n = 158 subjects). Results showed lower post-exercise damage to lipids (SMD = -0.95 CI 95%: -1.49 to -0. 40, I2 = 0%, p = 0.0007), proteins (SMD = -1.39 CI 95%: -2.11 to -0.68, I2 = 51%, p = 0.0001) and redox imbalance (SMD = -0.96 CI 95%: -1.65 to -0.28, I2 = 0%, p = 0.006) in favor of LLRE-BFR compared to HLRE. HLRE presents higher post-exercise superoxide dismutase activity but in the other biomarkers and time points, no significant differences between conditions were observed. For LLRE-BFR and LLRE, we found no difference between the comparisons performed at any time point., Conclusions: Based on the available evidence from randomized trials, providing very low or low certainty of evidence, this review demonstrates that LLRE-BFR promotes less oxidative stress when compared to HLRE but no difference in levels of oxidative damage biomarkers and endogenous antioxidants between LLRE., Trial Registration: Register number: PROSPERO number: CRD42020183204., Competing Interests: NR is the founder of THE BFR PROS, a BFR education company that provides BFR training workshops to fitness and rehabilitation professionals across the world using a variety of BFR devices. NR has no financial relationships with any cuff manufacturers/distributors. The remaining authors declare that they have no conflict of interests. This does not alter our adherence to all PLOS ONE policies on sharing data and materials., (Copyright: © 2023 Ferlito et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Ipsilateral breast tumor recurrence (IBTR) is a clinically important event, where an isolated in-breast recurrence is a potentially curable event but associated with an increased risk of distant metastasis and breast cancer death. It remains unclear if IBTRs are associated with molecular changes that can be explored as a resource for precision medicine strategies. Here, we employed proteogenomics to analyze a cohort of 27 primary breast cancers and their matched IBTRs to define proteogenomic determinants of molecular tumor evolution. Our analyses revealed a relationship between hormonal receptors status and proliferation levels resulting in the gain of somatic mutations and copy number. This in turn re-programmed the transcriptome and proteome towards a highly replicating and genomically unstable IBTRs, possibly enhanced by APOBEC3B. In order to investigate the origins of IBTRs, a second analysis that included primaries with no recurrence pinpointed proliferation and immune infiltration as predictive of IBTR. In conclusion, our study shows that breast tumors evolve into different IBTRs depending on hormonal status and proliferation and that immune cell infiltration and Ki-67 are significantly elevated in primary tumors that develop IBTR. These results can serve as a starting point to explore markers to predict IBTR formation and stratify patients for adjuvant therapy., (© 2023. The Author(s).)

This clinical trial aims to provide evidence about the effectiveness of the Pilates method on stress urinary incontinence (SUI), as well as to elucidate the effects of photobiomodulation therapy associated with static magnetic field (PBMT/sMF) alone or associated with the Pilates Method on Pelvic floor muscle (PFM) in women affected by SUI. For that, a three-arm, parallel randomized, double-blinded, placebo-controlled trial was conducted (NCT05096936). We recruited thirty-three women diagnosed with SUI, randomly allocated to three groups: placebo PBMT/sMF plus method Pilates, PBMT/sMF active plus method Pilates and only PBMT/sMF active. The evaluation consisted of anamnesis and physical examination, muscle strength, completion of the ICIQ-SF questionnaire, and urinary loss. The evaluation of muscle strength and filling the ICIQ-SF were performed on the first and last days, while the Pad test was applied in baseline, one month, two months, and three months of intervention. We observed an increase in strength ( p < 0.01), tone ( p < 0.01), and quality of life ( p < 0.01), in addition to a decrease in urinary lost ( p < 0.01) for all groups comparing the pre and post-intervention. The PBMT/sMF alone, the Pilates, and the combination of the two therapies proved to be effective in improving the signs and symptoms of women with SUI.

Objective: Aberrant activity of androgen receptor (AR) is the primary cause underlying development and progression of prostate cancer (PCa) and castration-resistant PCa (CRPC). Androgen signaling regulates gene transcription and lipid metabolism, facilitating tumor growth and therapy resistance in early and advanced PCa. Although direct AR signaling inhibitors exist, AR expression and function can also be epigenetically regulated. Specifically, lysine (K)-specific demethylases (KDMs), which are often overexpressed in PCa and CRPC phenotypes, regulate the AR transcriptional program., Methods: We investigated LSD1/UTX inhibition, two KDMs, in PCa and CRPC using a multi-omics approach. We first performed a mitochondrial stress test to evaluate respiratory capacity after treatment with MC3324, a dual KDM-inhibitor, and then carried out lipidomic, proteomic, and metabolic analyses. We also investigated mechanical cellular properties with acoustic force spectroscopy., Results: MC3324 induced a global increase in H3K4me2 and H3K27me3 accompanied by significant growth arrest and apoptosis in androgen-responsive and -unresponsive PCa systems. LSD1/UTX inhibition downregulated AR at both transcriptional and non-transcriptional level, showing cancer selectivity, indicating its potential use in resistance to androgen deprivation therapy. Since MC3324 impaired metabolic activity, by modifying the protein and lipid content in PCa and CRPC cell lines. Epigenetic inhibition of LSD1/UTX disrupted mitochondrial ATP production and mediated lipid plasticity, which affected the phosphocholine class, an important structural element for the cell membrane in PCa and CRPC associated with changes in physical and mechanical properties of cancer cells., Conclusions: Our data suggest a network in which epigenetics, hormone signaling, metabolite availability, lipid content, and mechano-metabolic process are closely related. This network may be able to identify additional hotspots for pharmacological intervention and underscores the key role of KDM-mediated epigenetic modulation in PCa and CRPC., (Copyright © 2022 The Author(s). Published by Elsevier GmbH.. All rights reserved.)

Oxidative stress induced by exercise has been a research field in constant growth, due to its relationship with the processes of fatigue, decreased production of muscle strength, and its ability to cause damage to the cell. In this context, photobiomodulation therapy (PBMT) has emerged as a resource capable of improving performance, while reducing muscle fatigue and muscle damage. To analyze the effects of PBMT about exercise-induced oxidative stress and compare with placebo therapy., Data Sources: Databases such as PubMed, EMBASE, CINAHL, CENTRAL, PeDro, and Virtual Health Library, which include Lilacs, Medline, and SciELO, were searched to find published studies., Study Selection: There was no year or language restriction; randomized clinical trials with healthy subjects that compared the application (before or after exercise) of PBMT to placebo therapy were included., Study Design: Systematic review with meta-analysis., Data Extraction: Data on the characteristics of the volunteers, study design, intervention parameters, exercise protocol and oxidative stress biomarkers were extracted. The risk of bias and the certainty of the evidence were assessed using the PEDro scale and the GRADE system, respectively., Results: Eight studies (n = 140 participants) were eligible for this review, with moderate to excellent methodological quality. In particular, PBMT was able to reduce damage to lipids post exercise (SMD = -0.72, CI 95% -1.42 to -0.02, I 2 = 77%, p = 0.04) and proteins (SMD = -0.41, CI 95% -0.65 to -0.16, I 2 = 0%, p = 0.001) until 72 h and 96 h, respectively. In addition, it increased the activity of SOD enzymes (SMD = 0.54, CI 95% 0.07 to 1.02, I 2 = 42%, p = 0.02) post exercise, 48 and 96 h after irradiation. However, PBMT did not increase CAT activity (MD = 0.18 CI 95% -0.56 to 0.91, I 2 = 79%, p = 0.64) post exercise. We did not find any difference in TAC or GPx biomarkers., Conclusion: Low to moderate certainty evidence shows that PBMT is a resource that can reduce oxidative damage and increase enzymatic antioxidant activity post exercise. We found evidence to support that one session of PBMT can modulate the redox metabolism.

The ergogenic effects of photobiomodulation therapy combined with a static magnetic field (PBMT-sMF) on exercises with characteristics similar to those of CrossFit® are unknown. This study was aimed at investigating the effects of PBMT-sMF applied at different times on recovery and physical performance in CrossFit® athletes by analyzing functional aspects, muscle damage, inflammatory processes, and oxidative stress. This was a prospectively registered, triple-blinded, placebo-controlled, crossover trial. CrossFit® athletes were recruited and assigned to receive one of the four possible interventions. Each intervention included protocols before and after the exercise (referred to as the workout of the day (WOD)). The four possibilities of intervention were as follows: placebo before and after WOD (placebo), PBMT-sMF before and placebo after WOD (PBMT-sMF before), placebo before and PBMT-sMF after WOD (PBMT-sMF after), and PBMT-sMF before and after WOD (PBMT-sMF before and after). The order of possibilities for the interventions was randomized. The primary outcome was the functional test performance. The secondary outcomes were the subjective perception of exertion, muscle damage, inflammation, and oxidative stress. The outcomes were measured before the WOD; immediately after the intervention; and 1, 24, and 48 hours after the WOD. Statistical analysis was performed using repeated measures ANOVA followed by the Bonferroni post hoc test to examine the differences between the interventions at each time point. Twelve participants were randomized and analyzed for each sequence. PBMT-sMF enhanced the performance on functional tests (calculated as a percentage of change) when applied before or after WOD in the assessment performed immediately post-WOD and at 24 and 48 hours later ( p < 0.05) compared to placebo and PBMT-sMF before and after WOD. In terms of the secondary outcomes, PBMT-sMF applied before or after WOD significantly decreased the creatine kinase, catalase, and superoxide dismutase activities and interleukin-6, thiobarbituric acid, and carbonylated protein levels (all p < 0.05) compared to the other possibilities of intervention. In addition, PBMT-sMF applied before and after WOD decreased creatine kinase activity at 24 hours and IL-6 levels at 24 and 48 hours compared to placebo ( p < 0.05). None of the participants reported any adverse events. PBMT-sMF enhanced the performance of functional tests, decreased the levels of biochemical markers of muscle damage and inflammation, decreased oxidative stress, and increased antioxidant activity in CrossFit® athletes when applied before or after WOD., Competing Interests: Professor Ernesto Cesar Pinto Leal-Junior received research support from Multi Radiance Medical (Solon, OH, USA), a laser device manufacturer. Shaiane Silva Tomazoni has a personal relationship with Ernesto Cesar Pinto Leal-Junior. The remaining authors declare that they have no support from any organization for the submitted work, no financial relationships with any organization with an interest in the submitted work within the last three years, and no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2022 Henrique Dantas Pinto et al.)

Acute myeloid leukemia (AML) arises when leukemia-initiating cells, defined by a primary genetic lesion, acquire subsequent molecular changes whose cumulative effects bypass tumor suppression. The changes that underlie AML pathogenesis not only provide insights into the biology of transformation but also reveal novel therapeutic opportunities. However, backtracking these events in transformed human AML samples is challenging, if at all possible. Here, we approached this question using a murine in vivo model with an MLL-ENL fusion protein as a primary molecular event. Upon clonal transformation, we identified and extensively verified a recurrent codon-changing mutation (Arg 295 Cys) in the ERM protein moesin that markedly accelerated leukemogenesis. Human cancer-associated moesin mutations at the conserved arginine-295 residue similarly enhanced MLL-ENL-driven leukemogenesis. Mechanistically, the mutation interrupted the stability of moesin and conferred a neomorphic activity to the protein, which converged on enhanced extracellular signal-regulated kinase activity. Thereby, our studies demonstrate a critical role of ERM proteins in AML, with implications also for human cancer.

The purpose of this study is to compare the effect of photobiomodulation therapy (PBMT) and cryotherapy (CRT) on muscle recovery outcomes. These searches were performed in PubMed, PEDro, CENTRAL, and VHL (which includes the Lilacs, Medline, and SciELO database) from inception to June 2021. We included randomized clinical trials involved healthy human volunteers (> 18 years) underwent an intervention of PBMT and CRT, when used in both isolated form post-exercise. Standardized mean differences (SMD) or mean difference (MD) with 95% confidence interval were calculated and pooled in a meta-analysis for synthesis. The risk of bias and quality of evidence were assessed through Cochrane risk-of-bias tool and GRADE system. Four articles (66 participants) with a high to low risk of bias were included. The certainty of evidence was classified as moderate to very low. PBMT was estimated to improve the muscle strength (SMD = 1.73, CI 95% 1.33 to 2.13, I 2  = 27%, p < 0.00001), reduce delayed onset muscle soreness (MD: - 25.69%, CI 95% - 34.42 to - 16.97, I 2  = 89%, p < 0.00001), and lower the concentration of biomarkers of muscle damage (SMD =  - 1.48, CI 95% - 1.93 to - 1.03, I 2  = 76%, p < 0,00,001) when compared with CRT. There was no difference in oxidative stress and inflammatory levels. Based on our findings, the use of PBMT in muscle recovery after high-intensity exercise appears to be beneficial, provides a clinically important effect, and seems to be the best option when compared to CRT., (© 2021. The Author(s), under exclusive licence to Springer-Verlag London Ltd., part of Springer Nature.)

Proteogenomic approaches have enabled the generat̲ion of novel information levels when compared to single omics studies although burdened by extensive experimental efforts. Here, we improved a data-independent acquisition mass spectrometry proteogenomic workflow to reveal distinct molecular features related to mammographic appearances in breast cancer. Our results reveal splicing processes detectable at the protein level and highlight quantitation and pathway complementarity between RNA and protein data. Furthermore, we confirm previously detected enrichments of molecular pathways associated with estrogen receptor-dependent activity and provide novel evidence of epithelial-to-mesenchymal activity in mammography-detected spiculated tumors. Several transcript-protein pairs displayed radically different abundances depending on the overall clinical properties of the tumor. These results demonstrate that there are differentially regulated protein networks in clinically relevant tumor subgroups, which in turn alter both cancer biology and the abundance of biomarker candidates and drug targets.

The efficacy and side effects of endocrine therapy in breast cancer (BC) depend largely on estrogen receptor alpha (ERα) expression, the specific drug administered, and treatment scheduling. Although the benefits of endocrine therapy outweigh any adverse effects in the initial stages of BC, later- or advanced-stage tumors acquire resistance to treatments. The mechanisms underlying tumor resistance to therapy are still not well understood, posing a major challenge for BC patient care. Epigenetic regulation and miRNA expression may be involved in the switch from a treatment-sensitive to a treatment-resistant state and could provide a valid therapeutic strategy for ERα negative BC. Here, a hybrid lysine-specific histone demethylase inhibitor, MC3324, displaying selective estrogen receptor down-regulator-like activities in BC, was used to highlight the interplay between epigenetic and ERα signaling. MC3324 anticancer action is mediated by microRNA (miRNA) expression regulation, indicating an innovative function for this molecule. Integrated analysis suggests a crosstalk between estrogen signaling, ERα interactors, miRNAs, and their putative targets. Specifically, miR-181a-5p expression is regulated by MC3324 and has an impact on cellular levels of ERα. A comparison of breast tumor versus healthy mammary tissues confirmed the important role of miR-181a-5p in ERα regulation and points to its putative predictive function in BC therapy.

Objective: To synthesize evidence on the effects of blood flow restriction (BFR) comparing with high (HLT) and low load (LLT), and on the influence of different forms of application in individuals with knee osteoarthritis., Data Sources: The CENTRAL, PEDro, PubMed and BVS, which include Lilacs, Medline and SciELO, until April 2020., Review Methods: A systematic review and meta-analysis of randomized trials used the PRISMA guidelines, whose main keywords were: Therapeutic Occlusion, Resistance Training, and Knee Osteoarthritie, blood flow restriction and Kaatsu training. Method quality was evaluated with the PEDro scale. When studies demonstrated homogeneity on outcome measures, the mean differences or standardized mean differences with 95% confidence interval were calculated and pooled in a meta-analysis for pooled synthesis., Results: Five articles were eligible in this review with moderate to low risk bias. Our results, showed no difference between BFR and HLT in knee strength (SMD = 0.00, 95% CI, -0.54 to 0.54, P = 1.00), function (SMD = -0.20, 95% CI, -0.45 to 0.06, P = 0.13), pain and volume. But, when compared BFR and LLT, the descriptive analysis demonstrated significant results in favor BFR to muscle strength (71.4% of measurement) and volume (MD = 1.66, 95% CI, 0.93 to 2.38, P < 0.00001), but not in pain or function., Conclusion: BFR can be used as a strategy in the rehabilitation of osteoarthritis due to gains in strength and volume with low mechanical stress. However, its application must be safe and individualized, since they can attenuate the stimuli offered by BFR.

In breast cancer, Lysine-specific demethylase-1 (LSD1) and other lysine demethylases (KDMs), such as Lysine-specific demethylase 6A also known as Ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX), are co-expressed and co-localize with estrogen receptors (ERs), suggesting the potential use of hybrid (epi)molecules to target histone methylation and therefore regulate/redirect hormone receptor signaling. Here, we report on the biological activity of a dual-KDM inhibitor (MC3324), obtained by coupling the chemical properties of tranylcypromine, a known LSD1 inhibitor, with the 2OG competitive moiety developed for JmjC inhibition. MC3324 displays unique features not exhibited by the single moieties and well-characterized mono-pharmacological inhibitors. Inhibiting LSD1 and UTX, MC3324 induces significant growth arrest and apoptosis in hormone-responsive breast cancer model accompanied by a robust increase in H3K4me2 and H3K27me3. MC3324 down-regulates ERα in breast cancer at both transcriptional and non-transcriptional levels, mimicking the action of a selective endocrine receptor disruptor. MC3324 alters the histone methylation of ERα-regulated promoters, thereby affecting the transcription of genes involved in cell surveillance, hormone response, and death. MC3324 reduces cell proliferation in ex vivo breast cancers, as well as in breast models with acquired resistance to endocrine therapies. Similarly, MC3324 displays tumor-selective potential in vivo, in both xenograft mice and chicken embryo models, with no toxicity and good oral efficacy. This epigenetic multi-target approach is effective and may overcome potential mechanism(s) of resistance in breast cancer.

The effects of preexercise photobiomodulation therapy (PBMT) to enhance performance, accelerate recovery, and attenuate exercise-induced oxidative stress were still not fully investigated, especially in high-level athletes. The aim of this study was to evaluate the effects of PBMT (using infrared low-level laser therapy) applied before a progressive running test on functional aspects, muscle damage, and inflammatory and oxidative stress markers in high-level soccer players. A randomized, triple-blind, placebo-controlled crossover trial was performed. Twenty-two high-level male soccer players from the same team were recruited and treated with active PBMT and placebo. The order of interventions was randomized. Immediately after the application of active PBMT or placebo, the volunteers performed a standardized high-intensity progressive running test (ergospirometry test) until exhaustion. We analyzed rates of oxygen uptake (VO 2 max ), time until exhaustion, and aerobic and anaerobic threshold during the intense progressive running test. Creatine kinase (CK) and lactate dehydrogenase (LDH) activities, levels of interleukin-1 β (IL-1- β ), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF- α ), levels of thiobarbituric acid (TBARS) and carbonylated proteins, and catalase (CAT) and superoxide dismutase (SOD) activities were measured before and five minutes after the end of the test. PBMT increased the VO 2 max (both relative and absolute values- p < 0.0467 and p < 0.0013, respectively), time until exhaustion ( p < 0.0043), time ( p < 0.0007) and volume ( p < 0.0355) in which anaerobic threshold happened, and volume in which aerobic threshold happened ( p < 0.0068). Moreover, PBMT decreased CK ( p < 0.0001) and LDH ( p < 0.0001) activities. Regarding the cytokines, PBMT decreased only IL-6 ( p < 0.0001). Finally, PBMT decreased TBARS ( p < 0.0001) and carbonylated protein levels ( p < 0.01) and increased SOD ( p < 0.0001)and CAT ( p < 0.0001) activities. The findings of this study demonstrate that preexercise PBMT acts on different functional aspects and biochemical markers. Moreover, preexercise PBMT seems to play an important antioxidant effect, decreasing exercise-induced oxidative stress and consequently enhancing athletic performance and improving postexercise recovery. This trial is registered with Clinicaltrials.gov NCT03803956., Competing Interests: Professor Ernesto Cesar Pinto Leal-Junior receives research support from Multi Radiance Medical (Solon, OH, USA), a laser device manufacturer. The remaining authors declare that they have no conflict of interests., (Copyright © 2019 Shaiane Silva Tomazoni et al.)

This study aimed to analyze PBMT effects on futsal player's performance and recovery in a non-controlled field environment. It is a randomized, triple-blinded, placebo-controlled, crossover clinical trial. The research included six professional athletes and in each match phototherapy treatments were performed before matches (40 minutes), blood samples were collected before treatments, and samples immediately after the end of the matches and 48 h after. Furthermore, videos were analyzed to quantify the time athletes spent on the pitch and the distance they covered. PBMT was performed at 17 sites of each lower limb (40 mins before matches), employing a cluster with 12 diodes (4 laser diodes of 905 nm, 4 LEDs of 875 nm, and 4 LEDs of 640 nm, 30 J per site). The performance of the athlete could be quantified considering the time on the pitch and the distance covered; the biochemical markers evaluated were creatine kinase, lactate dehydrogenase, blood lactate, and oxidative damage to lipids and proteins. PBMT significantly increased the time of staying in the pitch and a significant improvement in all the biochemical markers evaluated. No statistically significant difference was found for the distance covered. Pre-exercise PBMT can enhance performance and accelerate recovery of high-level futsal players.

Aim: Study the cardiovascular risk factors in a feminine population vulnerable to cardiovascular events particularly to evaluate the principal factors or possible confounding variables., Methods: This is a cross-sectional descriptive study. Were analysed all the female patients from the Cardiovascular Rehabilitation Institute of Sports Medicine of Caxias do Sul who had the complete information on cardiovascular disease history, comorbidities and habits and who knew the complete gynaecological history by a phone interview., Results: A group of 91 patients were analysed. About the comorbidities and habits, 45.2% of these patients presented some tobacco load, 82.4% are hypertensive, 61.5% are dyslipidemic, 25.3% are diabetic and the BMI average was 29.27 (overweight). Between the patients who undergone a hysterectomy and had an episode of the acute coronary syndrome (10 patients), 70% had the event after the procedure. Between the post-menopause women with at least one episode of the acute coronary syndrome, 80.5% (33 patients) had the first event after the menopause., Conclusion: We found multiple lifetime risk factors that predisposed the women of the sample to have cardiovascular disease. Between the women with specific to women risk factors and without, the prevalence of cardiovascular disease was very similar. This information supports the idea that these are just confounding factors of CVD and the principals involved are the genetic factors and habits. For this reason, the focus of CVD prevention and treatment should be directed towards these aspects.

Background: Photobiomodulation therapy (PBMT) has recently been used to alleviate postexercise muscle fatigue and enhance recovery, demonstrating positive results. A previous study by our research group demonstrated the optimal dose for an infrared wavelength (810 nm), but the outcomes could be optimized further with the determination of the optimal output power., Objective: The aim of the present study was to evaluate the effects of PBMT (through low-level laser therapy) on postexercise skeletal muscle recovery and identify the best output power., Materials and Methods: A randomized, placebo-controlled double-blind clinical trial was conducted with the participation of 28 high-level soccer players. PBMT was applied before the eccentric contraction protocol with a cluster with five diodes, 810 nm, dose of 10 J, and output power of 100, 200, 400 mW per diode or placebo at six sites of knee extensors. Maximum isometric voluntary contraction (MIVC), delayed onset muscle soreness (DOMS) and biochemical markers related to muscle damage (creatine kinase and lactate dehydrogenase), inflammation (IL-1β, IL-6, and TNF-α), and oxidative stress (catalase, superoxide dismutase, carbonylated proteins, and thiobarbituric acid) were evaluated before isokinetic exercise, as well as at 1 min and at 1, 24, 48, 72, and 96 h, after the eccentric contraction protocol., Results: PBMT increased MIVC and decreased DOMS and levels of biochemical markers (p < 0.05) with the power output of 100 and 200 mW, with better results for the power output of 100 mW., Conclusions: PBMT with 100 mW power output per diode (500 mW total) before exercise achieves best outcomes in enhancing muscular performance and postexercise recovery. Another time it has been demonstrated that more power output is not necessarily better.

In a previous study, we detected a significant association between phosphoserine aminotransferase 1 (PSAT1) hyper-methylation and mRNA levels to outcome to tamoxifen treatment in recurrent disease. We here aimed to study the association of PSAT1 protein levels to outcome upon tamoxifen treatment and to obtain more insight in its role in tamoxifen resistance. A cohort of ER positive, hormonal therapy naïve primary breast carcinomas was immunohistochemically (IHC) stained for PSAT1. Staining was analyzed for association with patient's time to progression (TTP) and overall response on first-line tamoxifen for recurrent disease. PSAT1 mRNA levels were also assessed by reverse transcriptase quantitative polymerase chain reaction (RT-qPCR; n = 161) and Affymetrix GeneChip (n = 155). Association of PSAT1 to biological pathways on tamoxifen outcome were assessed by global test. PSAT1 protein and mRNA levels were significantly associated to poor outcome to tamoxifen treatment. When comparing PSAT1 protein and mRNA levels, IHC and RT-qPCR data showed a significant association. Global test results showed that cytokine and JAK-STAT signaling were associated to PSAT1 expression. We hereby report that PSAT1 protein and mRNA levels measured in ER positive primary tumors are associated with poor clinical outcome to tamoxifen.

Context:   Recent studies suggest the prophylactic use of low-powered laser/light has ergogenic effects on athletic performance and postactivity recovery. Manufacturers of high-powered lasers/light devices claim that these can produce the same clinical benefits with increased power and decreased irradiation time; however, research with high-powered lasers is lacking., Objective:   To evaluate the magnitude of observed phototherapeutic effects with 3 commercially available devices., Design:   Randomized double-blind placebo-controlled study., Setting:   Laboratory., Patients or Other Participants:   Forty healthy untrained male participants., Intervention(s):   Participants were randomized into 4 groups: placebo, high-powered continuous laser/light, low-powered continuous laser/light, or low-powered pulsed laser/light (comprising both lasers and light-emitting diodes). A single dose of 180 J or placebo was applied to the quadriceps., Main Outcome Measure(s):   Maximum voluntary contraction, delayed-onset muscle soreness (DOMS), and creatine kinase (CK) activity from baseline to 96 hours after the eccentric exercise protocol., Results:   Maximum voluntary contraction was maintained in the low-powered pulsed laser/light group compared with placebo and high-powered continuous laser/light groups in all time points (P < .05). Low-powered pulsed laser/light demonstrated less DOMS than all groups at all time points (P < .05). High-powered continuous laser/light did not demonstrate any positive effects on maximum voluntary contraction, CK activity, or DOMS compared with any group at any time point. Creatine kinase activity was decreased in low-powered pulsed laser/light compared with placebo (P < .05) and high-powered continuous laser/light (P < .05) at all time points. High-powered continuous laser/light resulted in increased CK activity compared with placebo from 1 to 24 hours (P < .05)., Conclusions:   Low-powered pulsed laser/light demonstrated better results than either low-powered continuous laser/light or high-powered continuous laser/light in all outcome measures when compared with placebo. The increase in CK activity using the high-powered continuous laser/light compared with placebo warrants further research to investigate its effect on other factors related to muscle damage.

This study aimed to determine the effectiveness of photobiomodulation therapy (PBMT) and cryotherapy, in isolated and combined forms, as muscle recovery techniques after muscle fatigue-inducing protocol. Forty volunteers were randomly divided into five groups: a placebo group (PG); a PBMT group (PBMT); a cryotherapy group (CG); a cryotherapy-PBMT group (CPG); and a PBMT-cryotherapy group (PCG). All subjects performed four sessions at 24-h intervals, during which they submitted to isometric assessment (MVC) and blood collection in the pre-exercise period, and 5 and 60 min post-exercise, while the muscle fatigue induction protocol occurred after the pre-exercise collections. In the remaining sessions performed 24, 48, and 72 h later, only blood collections and MVCs were performed. A single treatment with PBMT and/or cryotherapy was applied after only 2 min of completing the post-5-min MVC test at the first session. In the intragroup comparison, it was found that exercise led to a significant decrease (p < 0.05) in the production of MVC in all groups. Comparing the results of MVCs between groups, we observed significant increases in the MVC capacity of the PBMT, CPG, and PCG volunteers in comparison with both PG and CG (p < 0.05). We observed a significant decrease in the concentrations of the biochemical markers of oxidative damage (TBARS and PC) in all groups and muscle damage (creatine kinase-CK) in the PBMT, PCG, and CPG compared with the PG (p < 0.01). The clinical impact of these findings is clear because they demonstrate that the use of phototherapy is more effective than the use of cryotherapy for muscle recovery, additionally cryotherapy decreases PBMT efficacy.

Aim: This study aimed to evaluate the medium-term effects of low-level laser therapy (LLLT or photobiomodulation) in postexercise skeletal muscle recovery and performance enhancement and to identify the optimal dose of 810 nm LLLT., Materials and Methods: A randomized, double-blind, placebo-controlled trial was performed, with voluntary participation of 28 high-level soccer athletes. We analyzed maximum voluntary contraction (MVC), delayed onset muscle soreness (DOMS), creatine kinase (CK) activity, and interleukin-6 (IL-6) expression. The assessments were performed before exercise protocols, after 1 min, and 1, 24, 48, 72, and 96 h after the end of eccentric exercise protocol used to induce fatigue. LLLT was applied before eccentric exercise protocol with a cluster with five diodes, and dose of 10, 30, or 50 J (200 mW and 810 nm) in six sites of quadriceps., Results: LLLT increased (p < 0.05) MVC from immediately after exercise to 24 h with 50 J dose, and from 24 to 96 h with 10 J dose. Both 10 J then 50 J dose decreased (p < 0.05) CK and IL-6 with better results in favor of 50 J dose. However, LLLT had no effect in decreasing DOMS. No differences (p > 0.05) were found for 30 J dose in any of the outcomes measured., Conclusions: Pre-exercise LLLT, mainly with 50 J dose, significantly increases performance and improves biochemical markers related to skeletal muscle damage and inflammation.

We have previously identified UMP-CMP kinase (CMPK1) as a prognostic marker for triple negative breast cancer (TNBC) by mass spectrometry (MS). In this study we evaluated CMPK1 association to prognosis in an independent set of samples by immunohistochemistry (IHC) and assessed biological pathways associated to its expression through gene set enrichment analysis (GSEA). A total of 461 TNBC paraffin-embedded tissues were collected from different academic hospitals in Europe, incorporated into tissue micro-arrays (TMA), and stained for CMPK1 expression. We also collected gene expression data of 60 samples, which were also present in the TMA, for GSEA correlation analysis. CMPK1 IHC staining showed both cytoplasmic and nuclear components. While cytoplasmic CMPK1 did not show any association to metastasis free survival (MFS), nuclear CMPK1 was associated to poor prognosis independently from other prognostic factors in stratified Cox regression analyses. GSEA correlation analysis of the nuclear CMPK1-stratified gene expression dataset showed a significant enrichment of extracellular matrix (ECM; positive correlation) and cell cycle (negative correlation) associated genes. We have shown here that nuclear CMPK1 is indicative of poor prognosis in TNBCs and that its expression may be related to dysregulation of ECM and cell cycle molecules.

Estrogen receptor (ER)-positive breast cancer represents the majority (∼70%) of all breast malignancies. In this subgroup of breast cancers, endocrine therapies are effective both in the adjuvant and recurrent settings, although resistance remains a major issue. Several high-throughput approaches have been used to elucidate mechanisms of resistance and to derive potential predictive markers or alternative therapies. In this review, we cover the state-of-the-art of endocrine-resistance biomarker discovery with regard to the latest technological developments, and discuss current opportunities and restrictions for their implementation into a clinical setting., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Laser-capture microdissection (LCM) offers a reliable cell population enrichment tool and has been successfully coupled to MS analysis. Despite this, most proteomic studies employ whole tissue lysate (WTL) analysis in the discovery of disease biomarkers and in profiling analyses. Furthermore, the influence of tissue heterogeneity in WTL analysis, nor its impact in biomarker discovery studies have been completely elucidated. In order to address this, we compared previously obtained high resolution MS data from a cohort of 38 breast cancer tissues, of which both LCM enriched tumor epithelial cells and WTL samples were analyzed. Label-free quantification (LFQ) analysis through MaxQuant software showed a significantly higher number of identified and quantified proteins in LCM enriched samples (3404) compared to WTLs (2837). Furthermore, WTL samples displayed a higher amount of missing data compared to LCM both at peptide and protein levels (p-value < 0.001). 2D analysis on co-expressed proteins revealed discrepant expression of immune system and lipid metabolisms related proteins between LCM and WTL samples. We hereby show that LCM better dissected the biology of breast tumor epithelial cells, possibly due to lower interference from surrounding tissues and highly abundant proteins. All data have been deposited in the ProteomeXchange with the dataset identifier PXD002381 (http://proteomecentral.proteomexchange.org/dataset/PXD002381)., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

We recently reported on the development of a 4-protein-based classifier (PDCD4, CGN, G3BP2, and OCIAD1) capable of predicting outcome to tamoxifen treatment in recurrent, estrogen-receptor-positive breast cancer based on high-resolution MS data. A precise and high-throughput assay to measure these proteins in a multiplexed, targeted fashion would be favorable to measure large numbers of patient samples to move these findings toward a clinical setting. By coupling immunoprecipitation to multiple reaction monitoring (MRM) MS and stable isotope dilution, we developed a high-precision assay to measure the 4-protein signature in 38 primary breast cancer whole tissue lysates (WTLs). Furthermore, we evaluated the presence and patient stratification capabilities of our signature in an independent set of 24 matched (pre- and post-therapy) sera. We compared the performance of immuno-MRM (iMRM) with direct MRM in the absence of fractionation and shotgun proteomics in combination with label-free quantification (LFQ) on both WTL and laser capture microdissected (LCM) tissues. Measurement of the 4-proteins by iMRM showed not only higher accuracy in measuring proteotypic peptides (Spearman r: 0.74 to 0.93) when compared with MRM (Spearman r: 0.0 to 0.76) but also significantly discriminated patient groups based on treatment outcome (hazard ratio [HR]: 10.96; 95% confidence interval [CI]: 4.33 to 27.76; Log-rank P < 0.001) when compared with LCM (HR: 2.85; 95% CI: 1.24 to 6.54; Log-rank P = 0.013) and WTL (HR: 1.16; 95% CI: 0.57 to 2.33; Log-rank P = 0.680) LFQ-based predictors. Serum sample analysis by iMRM confirmed the detection of the four proteins in these samples. We hereby report that iMRM outperformed regular MRM, confirmed our previous high-resolution MS results in tumor tissues, and has shown that the 4-protein signature is measurable in serum samples.

Tamoxifen therapy resistance constitutes a major cause of death in patients with recurrent estrogen receptor (ER) positive breast cancer. Through high resolution mass spectrometry (MS), we previously generated a 4-protein predictive signature for tamoxifen therapy outcome in recurrent breast cancer. ANXA1 and CALD1, which were not included in the classifier, were however the most differentially expressed proteins. We first evaluated the clinical relevance of these markers in our MS cohort, followed by immunohistochemical (IHC) staining on an independent set of tumors incorporated in a tissue microarray (TMA) and regression analysis in relation to time to progression (TTP), clinical benefit and objective response. In order to assess which mechanisms ANXA1 and CALD1 might been involved in, we performed Ingenuity pathway analysis (IPA) on ANXA1 and CALD1 correlated proteins in our MS cohort. ANXA1 (Hazard ratio [HR] = 1.83; 95% confidence interval [CI]: 1.22-2.75; P = 0.003) and CALD1 (HR = 1.57; 95% CI: 1.04-2.36; P = 0.039) based patient stratification showed significant association to TTP, while IHC staining on TMA showed that both ANXA1 (HR = 1.82; 95% CI: 1.12-3.00; P = 0.016) and CALD1 (HR = 2.29; 95% CI: 1.40-3.75; P = 0.001) expression was associated with shorter TTP independently of traditional predictive factors. Pearson correlation analysis showed that the majority of proteins correlated to ANXA1 also correlated with CALD1. IPA indicated that ANXA1 and CALD1 were associated with ER-downregulation and NFκB signaling. We hereby report that ANXA1 and CALD1 proteins are independent markers for tamoxifen therapy outcome and are associated to fast tumor progression.

Estrogen receptor (ER) positive tumors represent the majority of breast malignancies, and are effectively treated with hormonal therapies, such as tamoxifen. However, in the recurrent disease resistance to tamoxifen therapy is common and a major cause of death. In recent years, in-depth proteome analyses have enabled identification of clinically useful biomarkers, particularly, when heterogeneity in complex tumor tissue was reduced using laser capture microdissection (LCM). In the current study, we performed high resolution proteomic analysis on two cohorts of ER positive breast tumors derived from patients who either manifested good or poor outcome to tamoxifen treatment upon recurrence. A total of 112 fresh frozen tumors were collected from multiple medical centers and divided into two sets: an in-house training and a multi-center test set. Epithelial tumor cells were enriched with LCM and analyzed by nano-LC Orbitrap mass spectrometry (MS), which yielded >3000 and >4000 quantified proteins in the training and test sets, respectively. Raw data are available via ProteomeXchange with identifiers PXD000484 and PXD000485. Statistical analysis showed differential abundance of 99 proteins, of which a subset of 4 proteins was selected through a multivariate step-down to develop a predictor for tamoxifen treatment outcome. The 4-protein signature significantly predicted poor outcome patients in the test set, independent of predictive histopathological characteristics (hazard ratio [HR] = 2.17; 95% confidence interval [CI] = 1.15 to 4.17; multivariate Cox regression p value = 0.017). Immunohistochemical (IHC) staining of PDCD4, one of the signature proteins, on an independent set of formalin-fixed paraffin-embedded tumor tissues provided and independent technical validation (HR = 0.72; 95% CI = 0.57 to 0.92; multivariate Cox regression p value = 0.009). We hereby report the first validated protein predictor for tamoxifen treatment outcome in recurrent ER-positive breast cancer. IHC further showed that PDCD4 is an independent marker., (Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.)

We here describe two proteomic datasets deposited in ProteomeXchange via PRIDE partner repository [1] with dataset identifiers PXD000484 (defined as "training") and PXD000485 (defined as "test") that have been used for the development of a tamoxifen outcome predictive signature [2]. Both datasets comprised 56 fresh frozen estrogen receptor (ER) positive primary breast tumor specimens derived from patients who received tamoxifen as first line therapy for recurrent disease. Patient groups were defined based on time to progression (TTP) after start of tamoxifen therapy (6 months cutoff): 32 good and 24 poor treatment outcome patients were comprised in the training set, respectively. The test set included 41 good and 15 poor treatment outcome patients. All specimens were subjected to laser capture microdissection (LCM) to enrich for epithelial tumor cells prior to high resolution mass spectrometric (MS) analysis. Protein identification and label-free quantification (LFQ) were performed with MaxQuant software package [3]. A total of 3109 and 4061 proteins were identified and quantified in the training and test set, respectively. We here present the first public proteomic dataset analyzing ER positive recurrent breast cancer by LCM coupled to high resolution MS.

Targeted quantitative mass spectrometry of immunoaffinity-enriched peptides, termed immuno-multiple reaction monitoring (iMRM), is a powerful method for determining the relative abundance of proteins in complex mixtures, like plasma or whole tissue. This technique combines 1,000-fold enrichment potential of antibodies for target peptides with the selectivity of multiple reaction monitoring mass spectrometry (MRM-MS). Using heavy isotope-labeled peptide counterparts as internal standards ensures high levels of precision. Further, LC-MRM-MS selectivity allows for multiplexing; antibodies recognizing different peptides can be added directly to a single mixture without subjecting to interferences common to other multiple antibody protein assays. Integrated extracted ion chromatograms (XIC) of product ions from endogenous unlabeled "light" peptide and stable isotope-labeled internal standard "heavy" peptides are used to generate a light/heavy peak area ratio. This ratio is proportional to the amount of peptide in the digestion mixture and can be used to estimate the concentration of protein in the sample.

Recent studies with phototherapy have shown positive results in enhancement of performance and improvement of recovery when applied before exercise. However, several factors still remain unknown such as therapeutic windows, optimal treatment parameters, and effects of combination of different light sources (laser and LEDs). The aim of this study was to evaluate the effects of phototherapy with the combination of different light sources on skeletal muscle performance and post-exercise recovery, and to establish the optimal energy dose. A randomized, double-blinded, placebo-controlled trial with participation of 40 male healthy untrained volunteers was performed. A single phototherapy intervention was performed immediately after pre-exercise (baseline) maximum voluntary contraction (MVC) with a cluster of 12 diodes (4 of 905 nm lasers-0.3125 mW each, 4 of 875 nm LEDs-17.5 mW each, and 4 of 670 nm LEDs-15 mW each- manufactured by Multi Radiance Medical™) and dose of 10, 30, and 50 J or placebo in six sites of quadriceps. MVC, delayed onset muscle soreness (DOMS), and creatine kinase (CK) activity were analyzed. Assessments were performed before, 1 min, 1, 24, 48, 72, and 96 h after eccentric exercise protocol employed to induce fatigue. Phototherapy increased (p < 0.05) MVC was compared to placebo from immediately after to 96 h after exercise with 10 or 30 J doses (better results with 30 J dose). DOMS was significantly decreased compared to placebo (p < 0.05) with 30 J dose from 24 to 96 h after exercise, and with 50 J dose from immediately after to 96 h after exercise. CK activity was significantly decreased (p < 0.05) compared to placebo with all phototherapy doses from 1 to 96 h after exercise (except for 50 J dose at 96 h). Pre-exercise phototherapy with combination of low-level laser and LEDs, mainly with 30 J dose, significantly increases performance, decreases DOMS, and improves biochemical marker related to skeletal muscle damage.

Ferritin heavy chain (FTH1) is a 21-kDa subunit of the ferritin complex, known for its role in iron metabolism, and which has recently been identified as a favorable prognostic protein for triple negative breast cancer (TNBC) patients. Currently, it is not well understood how FTH1 contributes to an anti-tumor response. Here, we explored whether expression and cellular compartmentalization of FTH1 correlates to an effective immune response in TNBC patients. Analysis of the tumor tissue transcriptome, complemented with in silico pathway analysis, revealed that FTH1 was an integral part of an immunomodulatory network of cytokine signaling, adaptive immunity, and cell death. These findings were confirmed using mass spectrometry (MS)-derived proteomic data, and immunohistochemical staining of tissue microarrays. We observed that FTH1 is localized in both the cytoplasm and/or nucleus of cancer cells. However, high cytoplasmic (c) FTH1 was associated with favorable prognosis (Log-rank p = 0.001), whereas nuclear (n) FTH1 staining was associated with adverse prognosis (Log-rank p = 0.019). cFTH1 staining significantly correlated with total FTH1 expression in TNBC tissue samples, as measured by MS analysis (Rs = 0.473, p = 0.0007), but nFTH1 staining did not (Rs = 0.197, p = 0.1801). Notably, IFN γ-producing CD8+ effector T cells, but not CD4+ T cells, were preferentially enriched in tumors with high expression of cFTH1 (p = 0.02). Collectively, our data provide evidence toward new immune regulatory properties of FTH1 in TNBC, which may facilitate development of novel therapeutic targets., (© 2014 by The American Society for Biochemistry and Molecular Biology, Inc.)

Currently, treatment of muscle injuries represents a challenge in clinical practice. In acute phase, the most employed therapies are cryotherapy and nonsteroidal anti-inflammatory drugs. In the last years, low-level laser therapy (LLLT) has becoming a promising therapeutic agent; however, its effects are not fully known. The aim of this study was to analyze the effects of sodium diclofenac (topical application), cryotherapy, and LLLT on pro-inflammatory cytokine levels after a controlled model of muscle injury. For such, we performed a single trauma in tibialis anterior muscle of rats. After 1 h, animals were treated with sodium diclofenac (11.6 mg/g of solution), cryotherapy (20 min), or LLLT (904 nm; superpulsed; 700 Hz; 60 mW mean output power; 1.67 W/cm(2); 1, 3, 6 or 9 J; 17, 50, 100 or 150 s). Assessment of interleukin-1β and interleukin-6 (IL-1β and IL-6) and tumor necrosis factor-alpha (TNF-α) levels was performed at 6 h after trauma employing enzyme-linked immunosorbent assay method. LLLT with 1 J dose significantly decreased (p < 0.05) IL-1β, IL-6, and TNF-α levels compared to non-treated injured group as well as diclofenac and cryotherapy groups. On the other hand, treatment with diclofenac and cryotherapy does not decrease pro-inflammatory cytokine levels compared to the non-treated injured group. Therefore, we can conclude that 904 nm LLLT with 1 J dose has better effects than topical application of diclofenac or cryotherapy in acute inflammatory phase after muscle trauma.

Background: Recent studies involving phototherapy applied prior to exercise have demonstrated positive results regarding the attenuation of muscle fatigue and the expression of biochemical markers associated with recovery. However, a number of factors remain unknown, such as the ideal dose and application parameters, mechanisms of action and long-term effects on muscle recovery. The aims of the proposed project are to evaluate the long-term effects of low-level laser therapy on post-exercise musculoskeletal recovery and identify the best dose andapplication power/irradiation time., Design and Methods: A double-blind, randomized, placebo-controlled clinical trial with be conducted. After fulfilling the eligibility criteria, 28 high-performance athletes will be allocated to four groups of seven volunteers each. In phase 1, the laser power will be 200 mW and different doses will be tested: Group A (2 J), Group B (6 J), Group C (10 J) and Group D (0 J). In phase 2, the best dose obtained in phase 1 will be used with the same distribution of the volunteers, but with different powers: Group A (100 mW), Group B (200 mW), Group C (400 mW) and Group D (0 mW). The isokinetic test will be performed based on maximum voluntary contraction prior to the application of the laser and after the eccentric contraction protocol, which will also be performed using the isokinetic dynamometer. The following variables related to physical performance will be analyzed: peak torque/maximum voluntary contraction, delayed onset muscle soreness (algometer), biochemical markers of muscle damage, inflammation and oxidative stress., Discussion: Our intention, is to determine optimal laser therapy application parameters capable of slowing down the physiological muscle fatigue process, reducing injuries or micro-injuries in skeletal muscle stemming from physical exertion and accelerating post-exercise muscle recovery. We believe that, unlike drug therapy, LLLT has a biphasic dose-response pattern., Trial Registration: The protocol for this study is registered with the Protocol Registry System, ClinicalTrials.gov identifier NCT01844271.

Background: Clinical outcome of patients with triple-negative breast cancer (TNBC) is highly variable. This study aims to identify and validate a prognostic protein signature for TNBC patients to reduce unnecessary adjuvant systemic therapy., Methods: Frozen primary tumors were collected from 126 lymph node-negative and adjuvant therapy-naive TNBC patients. These samples were used for global proteome profiling in two series: an in-house training (n = 63) and a multicenter test (n = 63) set. Patients who remained free of distant metastasis for a minimum of 5 years after surgery were defined as having good prognosis. Cox regression analysis was performed to develop a prognostic signature, which was independently validated. All statistical tests were two-sided., Results: An 11-protein signature was developed in the training set (median follow-up for good-prognosis patients = 117 months) and subsequently validated in the test set (median follow-up for good-prognosis patients = 108 months) showing 89.5% sensitivity (95% confidence interval [CI] = 69.2% to 98.1%), 70.5% specificity (95% CI = 61.7% to 74.2%), 56.7% positive predictive value (95% CI = 43.8% to 62.1%), and 93.9% negative predictive value (95% CI = 82.3% to 98.9%) for poor-prognosis patients. The predicted poor-prognosis patients had higher risk to develop distant metastasis than the predicted good-prognosis patients in univariate (hazard ratio [HR] = 13.15; 95% CI = 3.03 to 57.07; P = .001) and multivariable (HR = 12.45; 95% CI = 2.67 to 58.11; P = .001) analysis. Furthermore, the predicted poor-prognosis group had statistically significantly more breast cancer-specific mortality. Using our signature as guidance, more than 60% of patients would have been exempted from unnecessary adjuvant chemotherapy compared with conventional prognostic guidelines., Conclusions: We report the first validated proteomic signature to assess the natural course of clinical TNBC.

Quantitative proteomics plays an important role in validation of breast-cancer-related biomarkers. In this study, we systematically compared the performance of label-free quantification (LFQ) and SILAC with shotgun and directed methods for quantifying breast-cancer-related markers in microdissected tissues. We show that LFQ leads to slightly higher coefficient of variation (CV) for protein quantification (median CV = 16.3%) than SILAC quantification (median CV = 13.7%) (P < 0.0001), but LFQ method enables ∼60% more protein quantification and is also more reproducible (∼20% more proteins were quantified in all replicate samples). Furthermore, we describe a method to accurately quantify multiple proteins within one pathway, that is, "focal adhesion pathway", in trace amounts of breast cancer tissues using a SILAC-based SRM assay. Using this SILAC-based SRM assay, we precisely quantified five "focal adhesion" proteins with good quantitative precision (CV range: 2.4-5.9%) in replicate whole tissue lysate samples and replicate microdissected samples (CV range: 5.8-16.1%). Our results show that in microdissected breast cancer tissues LFQ in combination with shotgun proteomics performed the best overall and is therefore suitable for both biomarker discovery and validation in these types of specimens. The SILAC-based SRM method can be used for the development of clinically relevant protein assays in tumor biopsies.

The aim of this work was to evaluate the effects of low-level laser therapy (LLLT) on exercise performance, oxidative stress, and muscle status in humans. A randomized double-blind placebo-controlled crossover trial was performed with 22 untrained male volunteers. LLLT (810 nm, 200 mW, 30 J in each site, 30 s of irradiation in each site) using a multi-diode cluster (with five spots - 6 J from each spot) at 12 sites of each lower limb (six in quadriceps, four in hamstrings, and two in gastrocnemius) was performed 5 min before a standardized progressive-intensity running protocol on a motor-drive treadmill until exhaustion. We analyzed exercise performance (VO(2 max), time to exhaustion, aerobic threshold and anaerobic threshold), levels of oxidative damage to lipids and proteins, the activities of the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), and the markers of muscle damage creatine kinase (CK) and lactate dehydrogenase (LDH). Compared to placebo, active LLLT significantly increased exercise performance (VO(2 max) p = 0.01; time to exhaustion, p = 0.04) without changing the aerobic and anaerobic thresholds. LLLT also decreased post-exercise lipid (p = 0.0001) and protein (p = 0.0230) damages, as well as the activities of SOD (p = 0.0034), CK (p = 0.0001) and LDH (p = 0.0001) enzymes. LLLT application was not able to modulate CAT activity. The use of LLLT before progressive-intensity running exercise increases exercise performance, decreases exercise-induced oxidative stress and muscle damage, suggesting that the modulation of the redox system by LLLT could be related to the delay in skeletal muscle fatigue observed after the use of LLLT.

In the last years, phototherapy has becoming a promising tool to improve skeletal muscle recovery after exercise, however, it was not compared with other modalities commonly used with this aim. In the present study we compared the short-term effects of cold water immersion therapy (CWIT) and light emitting diode therapy (LEDT) with placebo LEDT on biochemical markers related to skeletal muscle recovery after high-intensity exercise. A randomized double-blind placebo-controlled crossover trial was performed with six male young futsal athletes. They were treated with CWIT (5°C of temperature [SD ±1°]), active LEDT (69 LEDs with wavelengths 660/850 nm, 10/30 mW of output power, 30 s of irradiation time per point, and 41.7 J of total energy irradiated per point, total of ten points irradiated) or an identical placebo LEDT 5 min after each of three Wingate cycle tests. Pre-exercise, post-exercise, and post-treatment measurements were taken of blood lactate levels, creatine kinase (CK) activity, and C-reactive protein (CRP) levels. There were no significant differences in the work performed during the three Wingate tests (p > 0.05). All biochemical parameters increased from baseline values (p < 0.05) after the three exercise tests, but only active LEDT decreased blood lactate levels (p = 0.0065) and CK activity (p = 0.0044) significantly after treatment. There were no significant differences in CRP values after treatments. We concluded that treating the leg muscles with LEDT 5 min after the Wingate cycle test seemed to inhibit the expected post-exercise increase in blood lactate levels and CK activity. This suggests that LEDT has better potential than 5 min of CWIT for improving short-term post-exercise recovery.

The purpose of the present study was to determine the effect of low level laser therapy (LLLT) treatment before knee extensor eccentric exercise on indirect markers of muscle damage. Thirty-six healthy men were randomized in LLLT group (n = 18) and placebo group (n = 18). After LLLT or placebo treatment, subjects performed 75 maximal knee extensors eccentric contractions (five sets of 15 repetitions; velocity = 60° seg(-1); range of motion = 60°). Muscle soreness (visual analogue scale--VAS), lactate dehydrogenase (LDH) and creatine kinase (CK) levels were measured prior to exercise, and 24 and 48 h after exercise. Muscle function (maximal voluntary contraction--MVC) was measured before exercise, immediately after, and 24 and 48 h post-exercise. Groups had no difference on kineanthropometric characteristics and on eccentric exercise performance. They also presented similar baseline values of VAS (0.00 mm for LLLT and placebo groups), LDH (LLLT = 186 IU/l; placebo = 183 IU/l), CK (LLLT = 145 IU/l; placebo = 155 IU/l) and MVC (LLLT = 293 Nm; placebo = 284 Nm). VAS data did not show group by time interaction (P = 0.066). In the other outcomes, LLLT group presented (1) smaller increase on LDH values 48 h post-exercise (LLLT = 366 IU/l; placebo = 484 IU/l; P = 0.017); (2) smaller increase on CK values 24 h (LLLT = 272 IU/l; placebo = 498 IU/l; P = 0.020) and 48 h (LLLT = 436 IU/l; placebo = 1328 IU/l; P < 0.001) post-exercise; (3) smaller decrease on MVC immediately after exercise (LLLT = 189 Nm; placebo = 154 Nm; P = 0.011), and 24 h (LLLT = 249 Nm; placebo = 205 Nm; P = 0.004) and 48 h (LLLT = 267 Nm; placebo = 216 Nm; P = 0.001) post-exercise compared with the placebo group. In conclusion, LLLT treatment before eccentric exercise was effective in terms of attenuating the increase of muscle proteins in the blood serum and the decrease in muscle force.