Neuropeptide expression and morphometric differences in crushed alveolar inferior nerve of rats: Effects of photobiomodulation

Inferior alveolar nerve (IAN) injuries may occur during various dental routine procedures, especially in the removal of impacted lower third molars, and nerve recovery in these cases is a great challenge in dentistry. Here, the IAN crush injury model was used to assess the efficacy of photobiomodulation (PBM) in the recovery of the IAN in rats following crushing injury (a partial lesion). Rats were divided into four experimental groups: without any procedure, IAN crush injury, and IAN crush injury with PBM and sham group with PBM. Treatment was started 2 days after surgery, above the site of injury, and was performed every other day, totaling 10 sessions. Rats were irradiated with GaAs Laser (Gallium Arsenide, Laserpulse, Ibramed Brazil) emitting a wavelength of 904 nm, an output power of 70 mWpk, beam spot size at target ～0.1 cm², a frequency of 9500 Hz, a pulse time 60 ns, and an energy density of 6 J/cm². Nerve recovery was investigated by measuring the morphometric data of the IAN using TEM and by the expression of laminin, neurofilaments (NFs), and myelin protein zero (MPZ) using Western blot analysis. We found that IAN-injured rats which received PBM had a significant improvement of IAN morphometry when compared to IAN-injured rats without PBM. In parallel, all MPZ, laminin, and NFs exhibited a decrease after PBM. The results of this study indicate that the correlation between the peripheral nerve ultrastructure and the associated protein expression shows the beneficial effects of PBM.     

Diesel exhaust particulates affect cell signaling,mucin profiles,and apoptosis in trachea explants of Balb/C mice

Particulate matter from diesel exhaust (DEP) has toxic properties and can activate intracellular signaling pathways and induce metabolic changes. This study was conducted to evaluate the activation of extracellular signal‐regulated kinase (ERK) and c‐Jun N‐terminal kinase (JNK) and to analyze the mucin profile (acid (AB⁺), neutral (PAS⁺), or mixed (AB/PAS⁺) mucus) and vacuolization (V) of tracheal explants after treatment with 50 or 100 μg/mL DEP for 30 or 60 min. Western blot analyses showed small increases in ERK1/2 and JNK phosphorylation after 30 min of 100 μg/mL DEP treatment compared with the control. An increase in JNK phosphorylation was observed after 60 min of treatment with 50 μg/mL DEP compared with the control. We did not observe any change in the level of ERK1/2 phosphorylation after treatment with 50 μg/mL DEP. Other groups of tracheas were subjected to histological sectioning and stained with periodic acid‐Schiff (PAS) reagent and Alcian Blue (AB). The stained tissue sections were then subjected to morphometric analysis. The results obtained were compared using ANOVA. Treatment with 50 μg/mL DEP for 30 min or 60 min showed a significant increase (p < 0.001) in the amount of acid mucus, a reduction in neutral mucus, a significant reduction in mixed mucus, and greater vacuolization. Our results suggest that compounds found in DEPs are able to activate acid mucus production and enhance vacuolization and cell signaling pathways, which can lead to airway diseases. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 1297–1308, 2015.     

Mechanism of misfolding of the human prion protein revealed by a pathological mutation

The misfolding and aggregation of the human prion protein (PrP) is associated with transmissible spongiform encephalopathies (TSEs). Intermediate conformations forming during the conversion of the cellular form of PrP into its pathological scrapie conformation are key drivers of the misfolding process. Here, we analyzed the properties of the C-terminal domain of the human PrP (huPrP) and its T183A variant, which is associated with familial forms of TSEs. We show that the mutation significantly enhances the aggregation propensity of huPrP, such as to uniquely induce amyloid formation under physiological conditions by the sole C-terminal domain of the protein. Using NMR spectroscopy, biophysics, and metadynamics simulations, we identified the structural characteristics of the misfolded intermediate promoting the aggregation of T183A huPrP and the nature of the interactions that prevent this species to be populated in the wild-type protein. In support of these conclusions, POM antibodies targeting the regions that promote PrP misfolding were shown to potently suppress the aggregation of this amyloidogenic mutant.

The misfolding and aggregation of the human prion protein (PrP) is associated with a number of fatal neurodegenerative disorders designated as transmissible spongiform encephalopathies (TSEs), including Kuru, Creutzfeldt–Jakob disease, fatal familial insomnia, and mad cow disease (1). In its physiological form, the cellular PrP (PrP^C) is a 23-kDa monomeric glycosylated protein that is linked to the outer surface of the neuronal plasma membrane via a glycophosphatidylinositol (GPI) anchor (2). Under conditions associated with TSEs, PrP^C misfolds into a nonnative conformation (scrapie PrP^Sc) that is prone to aggregation into insoluble amyloid fibrils (3). The conversion into PrP^Sc has been proposed to be able to propagate from one cell to another (4). Genetic traits also exist linking point mutations of PrP and familial forms of TSEs. These pathological mutations are mostly located in the C-terminal globular domain of the PrP^C and have been shown to generally enhance the aggregation propensity of PrP (5, 6).While the pathological relevance of PrP is now established, its function remains highly debated. PrP^C has been shown to be involved in the maintenance of myelin on peripheral nerves (7), and evidence exists for a number of other putative physiological roles, including calcium modulation, copper sensing, long-term potentiation, and long-term memory (8). A well-characterized property of the physiological PrP^C is its native structure, which is composed of a disordered N-terminal flexible tail (residues 23–124) and a structured C-terminal region (125–230) composed of three α-helices and a short antiparallel β-sheet (9–11). Recently, structures of amyloid states of PrP generated in vitro have been resolved, providing insights about the possible in vivo form of PrP^Sc (12, 13). Despite the progress made in the structural characterization of PrP^Sc, there are major gaps in our understanding of the mechanisms that trigger the misfolding and aggregation of PrP^C under conditions associated with the insurgence and development of TSEs. A major challenge in this context is the study of metastable misfolding intermediates, which are elusive to conventional experimental techniques for structure determination because of their transient and heterogeneous nature (14). It is now generally acknowledged that the initial step for the misfolding of the C-terminal domain of PrP^C is the disruption of the native packing of the region formed by the two β-strands S1 and S2 and the α-helix H1, against the region composed of the helices H2–H3 (15). The detachment of these two subdomains, denoted as S1–H1–S2 and H2–H3, respectively, is prevented under native conditions by four main intramolecular “gatekeeper” interactions (D178–R164, T183–Y162, H187–R156, D202–R156). Each of these interactions is altered by specific pathological mutations associated with inherited forms of TSEs (D178N, T183A, H187R, D202N), suggesting a destabilization of the native interface between the two subdomains (15). Among these PrP variants, a mutation associated with very early-onset dementia and spongiform encephalopathy in patients, namely T183A (16), abolishes a crucial hydrogen bond between Y162 from the β-strand S2 and T183 from the α-helix H3. Under physiological conditions, this H-bond stabilizes the packing at the interface between the subdomains S1–H1–S2 and H2–H3 (SI Appendix, Fig. S1) (17), and its depletion by T183A induces the strongest destabilization of the PrP^C structure among the TSE-associated PrP variants (18).We addressed in the present study the fundamental early molecular mechanism of human PrP^C (huPrP^C) misfolding induced by T183A using NMR experiments in combination with biophysical investigations and enhanced molecular metadynamics simulations. The study compared the properties of wild-type (WT) and T183A huPrP^C and identified a misfolded intermediate species that acts as a precursor to the formation of amyloid aggregates by the C-terminal domain of the mutant (residues 125–230) despite the fact that this construct lacks the amyloidogenic region 106–126 (19). We provide conclusive evidence of this aggregation mechanism using POM antibodies (Abs) targeting the specific epitope that was here found to initiate the misfolding of T183A huPrP^C. Taken together, these results generate a new detailed understanding of the structural transitions in huPrP^C that trigger its amyloid formation and provide proof of principle for a structure-based identification of targeted molecular strategies to prevent huPrP^C misfolding and aggregation.     

Weekly docetaxel versus CMF as adjuvant chemotherapy for elderly breast cancer patients: safety data from the multicentre phase 3 randomised ELDA trial

Within an ongoing multicentre phase 3 randomised trial (ELDA, cancertrials.gov ID: NCT00331097), early breast cancer patients, 65-79 years old, with average to high risk of recurrence, are randomly assigned to receive CMF (cyclophosphamide 600 mg/m2, methotrexate 40 mg/m2, fluorouracil 600 mg/m2, days 1-8) or docetaxel (35 mg/m2 days 1-8-15), every 4 weeks. Here we report an unplanned safety analysis prompted by an amendment introducing creatinine clearance as a tool to adjust methotrexate dose. Before such change, 101 patients with a median age of 70 were randomly assigned CMF (53 patients) or docetaxel (48 patients). At least one grades 3-4 toxic event of any type was reported in 40 (75.5%) and 19 (39.6%) patients with CMF and docetaxel, respectively (p=0.0002). Grades 3-4 hematological events were observed in 37 (69.8%) vs. 4 (8.3%) cases (p<0.0001) and grades 3-4 non-hematological toxicity in 12 (22.6%) vs. 15 (31.2%) patients (p=0.11), with CMF and docetaxel, respectively. A higher incidence of anemia, neutropenia, thrombocytopenia and febrile neutropenia was reported with CMF. Constipation, mucositis, nausea and vomiting were more common with CMF; diarrhoea, abdominal pain, dysgeusia, neuropathy and liver toxicity were more frequent with docetaxel. No significant interaction was found between the occurrence of severe toxicity and baseline variables, including creatinine clearance and geriatric activity scales. In conclusion, weekly docetaxel appears to be less toxic than CMF in terms of hematological toxicity.     

Improving the role of echocardiography in studying the right ventricle of repaired tetralogy of Fallot patients: comparison with cardiac magnetic resonance

Right ventricular (RV) evaluation represents one of the major clinical tasks in the follow-up of repaired tetralogy of Fallot patients (rToF) with pulmonary valve regurgitation, as both severe RV dilatation and dysfunction are key factors in defining the need of pulmonary valve replacement. The aim of our study was to report the diagnostic accuracy of echocardiography in the identification of rToF patients with severely dilated and/or depressed RV as compared to cardiac magnetic resonance (CMR). Among our patients with rToF, a subgroup of 95 (17.6?±?6.8 years; 60% male), who underwent right ventricular qualitative and quantitative evaluation with CMR following echocardiographic suspicion of severe dilation/dysfunction, were included in the analysis. When comparing echocardiographic RV functional parameters to CMR findings, we found no association between CMR-ejection fraction (EF) and either tricuspid annulus plane systolic excursion (TAPSe) nor tissue Doppler systolic tricuspid excursion velocity (all p?=?ns). In contrast RVFAC was strongly associated with CMR-EF (r?=?0.44; p?<?0.01) as well as to longitudinal components of RV mechanics including tissue Doppler s′ (r?=?0.40; p?<?0.01) and TAPSE (r?=?0.36; p?<?0.01). When comparing echocardiographic and CMR structural parameters of the RV, we found that CMR RV volume was strongly related to echocardiographic measurements of RV end diastolic area (from the 4 chamber apical view) and with proximal parasternal short axis right ventricle outflow-dimension. Accordingly a regression model was derived from multiple regression analysis, which allows a more accurate estimate of CMR RV volume from echocardiography (r²?=?0.59, p?<?0.001). Our study demonstrates a significant, although imperfect, correlation between echocardiographic and CMR RV functional and geometrical parameters. Combining echocardiographic measures of RV inflow and RV outflow, we deliver a simple formula to estimate CMR-RV volume, improving the echocardiographic accuracy in RV volume quantification.     

Treatment of heterotopic caesarean scar pregnancy complicated with post termination increase in size of residual mass and morbidly adherent placenta

We report a case of a secundipara with heterotopic cesarean scar pregnancy (HCSP) treated with potassium chloride injection into the ectopic embryo followed by sac aspiration. The remaining “mass” increased in size threefold and was surrounded by a rich vascular network. An arteriovenous malformation was suspected; however, appropriate treatment was precluded because of the viability of the ectopic gestation. Sonographic examination revealed a morbidly adherent placenta, and attempt to resect the mass laparoscopically was complicated by bleeding that required hysterectomy. This case illustrates a complication of the intervention performed to preserve the intrauterine gestation in case of HCSP. © 2017 Wiley Periodicals, Inc. J Clin Ultrasound 46 :227–230, 2018     

Early PREdiction of sepsis using leukocyte surface biomarkers: the ExPRES-sepsis cohort study

Purpose

Reliable biomarkers for predicting subsequent sepsis among patients with suspected acute infection are lacking. In patients presenting to emergency departments (EDs) with suspected acute infection, we aimed to evaluate the reliability and discriminant ability of 47 leukocyte biomarkers as predictors of sepsis (Sequential Organ Failure Assessment score?≥?2 at 24 h and/or 72 h following ED presentation).

Methods

In a multi-centre cohort study in four EDs and intensive care units (ICUs), we standardised flow-cytometric leukocyte biomarker measurement and compared patients with suspected acute infection (cohort-1) with two comparator cohorts: ICU patients with established sepsis (cohort-2), and ED patients without infection or systemic inflammation but requiring hospitalization (cohort-3).

Results

Between January 2014 and February 2016, we recruited 272, 59 and 75 patients to cohorts 1, 2, and 3, respectively. Of 47 leukocyte biomarkers, 14 were non-reliable, and 17 did not discriminate between the three cohorts. Discriminant analyses for predicting sepsis within cohort-1 were undertaken for eight neutrophil (cluster of differentiation antigens (CD) CD15; CD24; CD35; CD64; CD312; CD11b; CD274; CD279), seven monocyte (CD35; CD64; CD312; CD11b; HLA-DR; CD274; CD279) and a CD8 T-lymphocyte biomarker (CD279). Individually, only higher neutrophil CD279 [OR 1.78 (95% CI 1.23–2.57); P?=?0.002], higher monocyte CD279 [1.32 (1.03–1.70); P?=?0.03], and lower monocyte HLA-DR [0.73 (0.55–0.97); P?=?0.03] expression were associated with subsequent sepsis. With logistic regression the optimum biomarker combination was increased neutrophil CD24 and neutrophil CD279, and reduced monocyte HLA-DR expression, but no combination had clinically relevant predictive validity.

Conclusions

From a large panel of leukocyte biomarkers, immunosuppression biomarkers were associated with subsequent sepsis in ED patients with suspected acute infection.

Clinical trial registration

NCT02188992.