The association between the inflammatory potential of diet and the risk of histopathological and molecular subtypes of breast cancer in northwestern Iran: Results from the Breast Cancer Risk and Lifestyle study.

BACKGROUND: This study explored the association between diet-associated inflammation and the risk of different molecular subtypes of breast cancer (BrCA) in a large, population-based case-control study conducted in northwestern Iran. METHODS: The study consisted of 1007 women with histopathologically confirmed BrCA and 1004 controls admitted to hospitals in Tabriz, northwestern Iran, for nonneoplastic conditions. Dietary Inflammatory Index scores and energy-adjusted Dietary Inflammatory Index (E-DII) scores, with and without supplements, were computed on the basis of dietary intake collected using a validated 136-item food frequency questionnaire. RESULTS: Women with the highest E-DII scores (quartile 4) versus those with the lowest E-DII scores (quartile 1) showed a significantly increased BrCA risk (odds ratio for quartile 4 vs quartile 1 [ORQ4vsQ1 ], 1.87; 95% confidence interval [CI], 1.42-2.47), particularly for lobular carcinoma (ORQ4vsQ1 , 3.07; 95% CI, 1.34-7.02). Findings were similar for premenopausal women diagnosed with luminal A BrCA (ORQ4vsQ1 , 2.71; 95% CI, 1.74-4.22) or luminal B BrCA (ORQ4vsQ1 , 2.86; 95% CI, 1.39-5.89). Women consuming the most proinflammatory diets were 3 times more likely to have triple-negative BrCA (ORQ4vsQ1 , 3.00; 95% CI, 1.002-8.96) while compared to luminal A BrCA. The BrCA risk for women consuming diets in the highest half of E-DII scores (E-DII > 0) was 59% greater than the risk for those in the lowest half (95% CI, 1.29-1.97). Also, higher E-DII scores that took into account supplements were associated with larger tumor sizes (T3 > 5 cm; P < .05). CONCLUSIONS: A proinflammatory diet, as indicated by higher E-DII scores, appears to increase the risk of BrCA in Iranian women. Large increases in risk were seen in invasive molecular subtypes of BrCA. Anti-inflammatory diets are suggested to prevent the risk of overall BrCA and more aggressive forms of BrCA in particular.

Topological analysis of the gp41 MPER on lipid bilayers relevant to the metastable HIV-1 envelope prefusion state.

The membrane proximal external region (MPER) of HIV-1 envelope glycoprotein (gp) 41 is an attractive vaccine target for elicitation of broadly neutralizing antibodies (bNAbs) by vaccination. However, current details regarding the quaternary structural organization of the MPER within the native prefusion trimer [(gp120/41)3] are elusive and even contradictory, hindering rational MPER immunogen design. To better understand the structural topology of the MPER on the lipid bilayer, the adjacent transmembrane domain (TMD) was appended (MPER-TMD) and studied. Membrane insertion of the MPER-TMD was sensitive both to the TMD sequence and cytoplasmic residues. Antigen binding of MPER-specific bNAbs, in particular 10E8 and DH511.2_K3, was significantly impacted by the presence of the TMD. Furthermore, MPER-TMD assembly into 10-nm diameter nanodiscs revealed a heterogeneous membrane array comprised largely of monomers and dimers, as enumerated by bNAb Fab binding using single-particle electron microscopy analysis, arguing against preferential trimeric association of native MPER and TMD protein segments. Moreover, introduction of isoleucine mutations in the C-terminal heptad repeat to induce an extended MPER α-helical bundle structure yielded an antigenicity profile of cell surface-arrayed Env variants inconsistent with that found in the native prefusion state. In line with these observations, electron paramagnetic resonance analysis suggested that 10E8 inhibits viral membrane fusion by lifting the MPER N-terminal region out of the viral membrane, mandating the exposure of residues that would be occluded by MPER trimerization. Collectively, our data suggest that the MPER is not a stable trimer, but rather a dynamic segment adapted for structural changes accompanying fusion.

Effects of glassing matrix deuteration on the relaxation properties of hyperpolarized 13C spins and free radical electrons at cryogenic temperatures.

Glassing matrix deuteration could be a beneficial sample preparation method for 13C dynamic nuclear polarization (DNP) when large electron paramagnetic resonance (EPR) width free radicals are used. However, it could yield the opposite DNP effect when samples are doped with small EPR width free radicals. Herein, we have investigated the influence of solvent deuteration on the 13C nuclear and electron relaxation that go along with the effects on 13C DNP intensities at 3.35 T and 1.2 K. For 13C DNP samples doped with trityl OX063, the 13C DNP signals decreased significantly when the protons are replaced by deuterons in glycerol:water or DMSO:water solvents. Meanwhile, the corresponding solid-state 13C T1 relaxation times of trityl OX063-doped samples generally increased upon solvent deuteration. On the other hand, 13C DNP signals improved by a factor of ∼1.5 to 2 upon solvent deuteration of samples doped with 4-oxo-TEMPO. Despite this 13C DNP increase, there were no significant differences recorded in 13C T1 values of TEMPO-doped samples with nondeuterated or fully deuterated glassing matrices. While solvent deuteration appears to have a negligible effect on the electron T1 relaxation of both free radicals, the electron T2 relaxation times of these two free radicals generally increased upon solvent deuteration. These overall results suggest that while the solid-phase 13C DNP signals are dependent upon the changes in total nuclear Zeeman heat capacity, the 13C relaxation effects are related to 2H/1H nuclear spin diffusion-assisted 13C polarization leakage in addition to the dominant paramagnetic relaxation contribution of free radical centers.

13C Dynamic Nuclear Polarization Using a Trimeric Gd3+ Complex as an Additive.

Dissolution dynamic nuclear polarization (DNP) is one of the most successful techniques that resolves the insensitivity problem in liquid-state nuclear magnetic resonance (NMR) spectroscopy and imaging (MRI) by amplifying the signal by several thousand-fold. One way to further improve the DNP signal is the inclusion of trace amounts of lanthanides in DNP samples doped with trityl OX063 free radical as the polarizing agent. In practice, stable monomeric gadolinium complexes such as Gd-DOTA or Gd-HP-DO3A are used as beneficial additives in DNP samples, further boosting the DNP-enhanced solid-state 13C polarization by a factor of 2 or 3. Herein, we report on the use of a trimeric gadolinium complex as a dopant in 13C DNP samples to improve the 13C DNP signals in the solid-state at 3.35 T and 1.2 K and consequently, in the liquid-state at 9.4 T and 298 K after dissolution. Our results have shown that doping the 13C DNP sample with a complex which holds three Gd3+ ions led to an improvement of DNP-enhanced 13C polarization by a factor of 3.4 in the solid-state, on par with those achieved using monomeric Gd3+ complexes but only requires about one-fifth of the concentration. Upon dissolution, liquid-state 13C NMR signal enhancements close to 20 000-fold, approximately 3-fold the enhancement of the control samples, were recorded in the nearby 9.4 T high resolution NMR magnet at room temperature. Comparable reduction of 13C spin-lattice T1 relaxation time was observed in the liquid-state after dissolution for both the monomeric and trimeric Gd3+ complexes. Moreover, W-band electron paramagnetic resonance (EPR) data have revealed that 3-Gd doping significantly reduces the electron T1 of the trityl OX063 free radical, but produces negligible changes in the EPR spectrum, reminiscent of the results with monomeric Gd3+-complex doping. Our data suggest that the trimeric Gd3+ complex is a highly beneficial additive in 13C DNP samples and that its effect on DNP efficiency can be described in the context of the thermal mixing mechanism.

Transition Metal Doping Reveals Link between Electron T1 Reduction and 13C Dynamic Nuclear Polarization Efficiency.

Optimal efficiency of dissolution dynamic nuclear polarization (DNP) is essential to provide the required high sensitivity enhancements for in vitro and in vivo hyperpolarized 13C nuclear magnetic resonance (NMR) spectroscopy and imaging (MRI). At the nexus of the DNP process are the free electrons, which provide the high spin alignment that is transferred to the nuclear spins. Without changing DNP instrumental conditions, one way to improve 13C DNP efficiency is by adding trace amounts of paramagnetic additives such as lanthanide (e.g., Gd3+, Ho3+, Dy3+, Tb3+) complexes to the DNP sample, which has been observed to increase solid-state 13C DNP signals by 100-250%. Herein, we have investigated the effects of paramagnetic transition metal complex R-NOTA (R = Mn2+, Cu2+, Co2+) doping on the efficiency of 13C DNP using trityl OX063 as the polarizing agent. Our DNP results at 3.35 T and 1.2 K show that doping the 13C sample with 3 mM Mn2+-NOTA led to a substantial improvement of the solid-state 13C DNP signal by a factor of nearly 3. However, the other transition metal complexes Cu2+-NOTA and Co2+-NOTA complexes, despite their paramagnetic nature, had essentially no impact on solid-state 13C DNP enhancement. W-band electron paramagnetic resonance (EPR) measurements reveal that the trityl OX063 electron T1 was significantly reduced in Mn2+-doped samples but not in Cu2+- and Co2+-doped DNP samples. This work demonstrates, for the first time, that not all paramagnetic additives are beneficial to DNP. In particular, our work provides a direct evidence that electron T1 reduction of the polarizing agent by a paramagnetic additive is an essential requirement for the improvement seen in solid-state 13C DNP signal.

Influence of Dy3+ and Tb3+ doping on 13C dynamic nuclear polarization.

Dynamic nuclear polarization (DNP) is a technique that uses a microwave-driven transfer of high spin alignment from electrons to nuclear spins. This is most effective at low temperature and high magnetic field, and with the invention of the dissolution method, the amplified nuclear magnetic resonance (NMR) signals in the frozen state in DNP can be harnessed in the liquid-state at physiologically acceptable temperature for in vitro and in vivo metabolic studies. A current optimization practice in dissolution DNP is to dope the sample with trace amounts of lanthanides such as Gd3+ or Ho3+, which further improves the polarization. While Gd3+ and Ho3+ have been optimized for use in dissolution DNP, other lanthanides have not been exhaustively studied for use in C13 DNP applications. In this work, two additional lanthanides with relatively high magnetic moments, Dy3+ and Tb3+, were extensively optimized and tested as doping additives for C13 DNP at 3.35 T and 1.2 K. We have found that both of these lanthanides are also beneficial additives, to a varying degree, for C13 DNP. The optimal concentrations of Dy3+ (1.5 mM) and Tb3+ (0.25 mM) for C13 DNP were found to be less than that of Gd3+ (2 mM). W-band electron paramagnetic resonance shows that these enhancements due to Dy3+ and Tb3+ doping are accompanied by shortening of electron T1 of trityl OX063 free radical. Furthermore, when dissolution was employed, Tb3+-doped samples were found to have similar liquid-state C13 NMR signal enhancements compared to samples doped with Gd3+, and both Tb3+ and Dy3+ had a negligible liquid-state nuclear T1 shortening effect which contrasts with the significant reduction in T1 when using Gd3+. Our results show that Dy3+ doping and Tb3+ doping have a beneficial impact on C13 DNP both in the solid and liquid states, and that Tb3+ in particular could be used as a potential alternative to Gd3+ in C13 dissolution DNP experiments.

Dietary inflammatory index and breast cancer risk: an updated meta-analysis of observational studies.

This updated meta-analysis sought to determine whether the pro-inflammatory potential of diet is a risk factor for breast cancer (BrCa) development, for the first time focusing on the effects of design heterogeneity. The search was performed using Scopus, PubMed, and Embase databases. Data were extracted from twenty-one eligible studies, including eleven cohorts (336,085 participants/20,033 incidence cases), and ten case-control studies (9,833 cases/12,752controls). The random-effect was used to calculate the relative risk (RR) using STATA 16 software. The highest dietary inflammatory index (DII) vs. the lowest category showed 16% increased risk of BrCa (95% CI: 1.06-1.26; I2 = 62.8%, P (I2) < 0.001). This was notable in post-menopausal status (RR = 1.13, 95% CI: 1.04-1.22), women with body mass index (BMI) ≥ 30 kg/m2 (RR = 1.35, 95% CI: 1.07-1.63), and study populations from developing countries (RR = 1.79, 95% CI: 1.12-2.47). Methodological covariates were subject to subgroup meta-analyses and showed stronger results among case-control studies (RR = 1.50, 95% CI: 1.20-1.80), studies considered age-matched controls (RR = 1.56, 95% CI: 1.19-1.93) and hospital-based controls (RR = 2.11, 95% CI: 1.58-2.64), and cohort studies identified by prolong follow-up durations (RR = 1.13, 95% CI: 1.03-1.22). This updated meta-analysis highlighted the pro-inflammatory diet as a risk factor for BrCa, especially among women in post-menopausal status, obese groups, and developing countries. Meta-analysis in methodological subgroups could improve results, less affected by heterogeneity, and suggested subclassification with important implications for future epidemiological designs and even clinical management.

Enhanced Efficiency of 13C Dynamic Nuclear Polarization by Superparamagnetic Iron Oxide Nanoparticle Doping.

Attainment of high NMR signal enhancements is crucial to the success of in vitro or in vivo hyperpolarized NMR or imaging (MRI) experiments. In this work, we report on the use of a superparamagnetic iron oxide nanoparticle (SPION) MRI contrast agent Feraheme (ferumoxytol) as a beneficial additive in 13C samples for dissolution dynamic nuclear polarization (DNP). Our DNP data at 3.35 T and 1.2 K reveal that addition of 11 mM elemental iron concentration of Feraheme in trityl OX063-doped 3 M [1-13C] acetate samples resulted in a substantial improvement of 13C DNP signal by a factor of almost 3-fold. Concomitant with the large DNP signal increase is the narrowing of the 13C microwave DNP spectra for samples doped with SPION. W-band electron paramagnetic resonance (EPR) spectroscopy data suggest that these two prominent effects of SPION doping on 13C DNP can be ascribed to the shortening of trityl OX063 electron T 1 as explained within the thermal mixing DNP model. Liquid-state 13C NMR signal enhancements as high as 20,000-fold for SPION-doped samples were recorded after dissolution at 9.4 T and 297 K, which is about 3 times the liquid-state NMR signal enhancement of the control sample. While the presence of SPION in hyperpolarized solution drastically reduces 13C T 1, this can be mitigated by polarizing smaller aliquots of DNP samples. Moreover, we have shown that Feraheme nanoparticles (~30 nm in size) can be easily and effectively removed from the hyperpolarized liquid by simple mechanical filtration, thus one can potentially incorporate an in-line filtration for these SPIONS along the dissolution pathway of the hyperpolarizer-a significant advantage over other DNP enhancers such as the lanthanide complexes. The overall results suggest that the commercially-available and FDA-approved Feraheme is a highly efficient DNP enhancer that could be readily translated for use in clinical applications of dissolution DNP.

Membrane Disruption Mechanism of a Prion Peptide (106-126) Investigated by Atomic Force Microscopy, Raman and Electron Paramagnetic Resonance Spectroscopy.

A fragment of the human prion protein spanning residues 106-126 (PrP106-126) recapitulates many essential properties of the disease-causing protein such as amyloidogenicity and cytotoxicity. PrP106-126 has an amphipathic characteristic that resembles many antimicrobial peptides (AMPs). Therefore, the toxic effect of PrP106-126 could arise from a direct association of monomeric peptides with the membrane matrix. Several experimental approaches are employed to scrutinize the impacts of monomeric PrP106-126 on model lipid membranes. Porous defects in planar bilayers are observed by using solution atomic force microscopy. Adding cholesterol does not impede defect formation. A force spectroscopy experiment shows that PrP106-126 reduces Young's modulus of planar lipid bilayers. We use Raman microspectroscopy to study the effect of PrP106-126 on lipid atomic vibrational dynamics. For phosphatidylcholine lipids, PrP106-126 disorders the intrachain conformation, while the interchain interaction is not altered; for phosphatidylethanolamine lipids, PrP106-126 increases the interchain interaction, while the intrachain conformational order remains similar. We explain the observed differences by considering different modes of peptide insertion. Finally, electron paramagnetic resonance spectroscopy shows that PrP106-126 progressively decreases the orientational order of lipid acyl chains in magnetically aligned bicelles. Together, our experimental data support the proposition that monomeric PrP106-126 can disrupt lipid membranes by using similar mechanisms found in AMPs.

Structure of an E. coli integral membrane sulfurtransferase and its structural transition upon SCN(-) binding defined by EPR-based hybrid method.

Electron paramagnetic resonance (EPR)-based hybrid experimental and computational approaches were applied to determine the structure of a full-length E. coli integral membrane sulfurtransferase, dimeric YgaP, and its structural and dynamic changes upon ligand binding. The solution NMR structures of the YgaP transmembrane domain (TMD) and cytosolic catalytic rhodanese domain were reported recently, but the tertiary fold of full-length YgaP was not yet available. Here, systematic site-specific EPR analysis defined a helix-loop-helix secondary structure of the YagP-TMD monomers using mobility, accessibility and membrane immersion measurements. The tertiary folds of dimeric YgaP-TMD and full-length YgaP in detergent micelles were determined through inter- and intra-monomer distance mapping and rigid-body computation. Further EPR analysis demonstrated the tight packing of the two YgaP second transmembrane helices upon binding of the catalytic product SCN(-), which provides insight into the thiocyanate exportation mechanism of YgaP in the E. coli membrane.

Identification of Surface-Exposed Protein Radicals and A Substrate Oxidation Site in A-Class Dye-Decolorizing Peroxidase from Thermomonospora curvata.

Dye-decolorizing peroxidases (DyPs) are a family of heme peroxidases, in which a catalytic distal aspartate is involved in H2O2 activation to catalyze oxidations in acidic conditions. They have received much attention due to their potential applications in lignin compound degradation and biofuel production from biomass. However, the mode of oxidation in bacterial DyPs remains unknown. We have recently reported that the bacterial TcDyP from Thermomonospora curvata is among the most active DyPs and shows activity toward phenolic lignin model compounds (J. Biol. Chem.2015, 290, 23447). Based on the X-ray crystal structure solved at 1.75 Å, sigmoidal steady-state kinetics with Reactive Blue 19 (RB19), and formation of compound II-like product in the absence of reducing substrates observed with stopped-flow spectroscopy and electron paramagnetic resonance (EPR), we hypothesized that the TcDyP catalyzes oxidation of large-size substrates via multiple surface-exposed protein radicals. Among 7 tryptophans and 3 tyrosines in TcDyP consisting of 376 residues for the matured protein, W263, W376, and Y332 were identified as surface-exposed protein radicals. Only the W263 was also characterized as one of surface-exposed oxidation sites. SDS-PAGE and size-exclusion chromatography demonstrated that W376 represents an off-pathway destination for electron transfer, resulting in the crosslinking of proteins in the absence of substrates. Mutation of W376 improved compound I stability and overall catalytic efficiency toward RB19. While Y332 is highly conserved across all four classes of DyPs, its catalytic function in A-class TcDyP is minimal possibly due to its extremely small solvent accessible areas. Identification of surface-exposed protein radicals and substrate oxidation sites is important for understanding DyP mechanism and modulating its catalytic functions for improved activity on phenolic lignin.

Prevalence of oral mucosal lesions in male smokers and nonsmokers.

Tobacco smoking is one of the most important risk factors for the development of oral mucosal lesions such as leukoplakia and hairy tongue. Controversy exists in the literature, however, about the prevalence of oral lesions in smokers. The aim of this study was to evaluate oral lesions in male smokers compared with nonsmokers in Hamadan. A total of 516 male participants were assessed, 258 of whom were smokers and 258 of whom were healthy nonsmokers. The prevalence of lesions was evaluated by clinical observation and biopsy. We found that the most prevalent lesions among smokers were gingival problems and coated tongue; smokers had significantly more lesions than did nonsmokers. Malignant and premalignant lesions were found in a higher age range. Among all participants in our study, we found a large number of oral mucosal lesions in smokers that had a strong correlation with smoking. Dental services need to implement care and health education for smokers to promote health.

Assessment of radiographic factors affecting surgical exposure and orthodontic alignment of impacted canines of the palate: a 15-year retrospective study.

OBJECTIVE: Impacted canines require a combination of both surgical and orthodontic management. In this study, patients treated for bone-impacted canines of the hard palatal were evaluated to assess which radiographic factors influenced the feasibility to move impacted maxillary permanent canines from the hard palate into the alveolar arch. MATERIALS AND METHODS: Eighty patients aged 12 to 24 (average 16 years) were treated surgically and orthodontically to align 146 bone-impacted canines of the hard palate (from 1994 to 2008). Factors such as age, sex, angulation of the canine to the midline (CAM), anomaly of the canine root (RA), overlap of the adjacent lateral incisor root (OALIR), and ratio of root formation (RRF) upon treatment were documented. Radiographic records and demographic data were assessed. The following radiographic measurements of canine position were made from the orthopantomogram (OPG): (1) angulation to the midline, (2) anteroposterior position of the root, (3) overlap of the adjacent incisor. RA or dilaceration was assessed from the OPG, maxillary occlusal (MO), and periapical (PA) radiographs. Whether the impacted canine had responded to surgical exposure and was orthodontically aligned, or surgically removed and discarded was also recorded. The data were analyzed to assess and correlate significance. RESULTS: Eighty patients aged 12 to 24 (19 males and 61 females) with 146 bone-impacted permanent canines of the hard palatal were treated. One hundred and three teeth (70.54%) had responded to surgical exposure and orthodontic alignment within 9 to 12 months. Forty-three impacted canine teeth (29.46%) had to be surgically removed because of ankylosis and no movement after 8 to 9 months using 50 to 60 g of traction force via elastic chains. Data analysis via chi-square and Pearson correlation tests showed that as the CAM increased (> 45 degrees), the canine was more likely to be unresponsive to treatment (P < .001). Increased overlap (> half the root) of the adjacent lateral incisor root (OALIR) via the canine crown influenced the treatment results negatively (P < .001). Additionally, presence of RA was also negatively influential (P < .001). However, the anteroposterior position of the canine did not influence the treatment results significantly, neither did age, sex, nor amount of root formation. CONCLUSION: Bone-impacted canines of the hard palatal are more likely to respond to surgical exposure and orthodontic management if AM is less than 45 degrees on the OPG; there is no RA found on OPG, periapical (PA), and maxillary occlusal (MO) radiographs; and OALIR by the canine crown is nonexistent or less than grade 2 (half the root) on the OPG.