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BACKGROUND: Status dystonicus (SD) is a life-threatening condition. OBJECTIVE AND METHODS: In a dystonia cohort who developed status dystonicus, we analyzed demographics, background dystonia phenomenology and complexity, trajectory previous to-, via status dystonicus episodes, and evolution following them. RESULTS: Over 20 years, 40 of 328 dystonia patients who were receiving DBS developed 58 status dystonicus episodes. Dystonia was of pediatric onset (95%), frequently complex, and had additional cognitive and pyramidal impairment (62%) and MRI alterations (82.5%); 40% of episodes occured in adults. Mean disease duration preceding status dystonicus was 10.3 \textpm 8 years. Evolution time to status dystonicus varied from days to weeks; however, 37.5% of patients exhibited progressive worsening over years. Overall, DBS was efficient in resolving 90% of episodes. CONCLUSION: Status dystonicus is potentially reversible and a result of heterogeneous conditions with nonuniform underlying physiology. Recognition of the complex phenomenology, morphological alterations, and distinct patterns of evolution, before and after status dystonicus, will help our understanding of these conditions. \textcopyright 2018 International Parkinson and Movement Disorder Society.

UDP-sugars are used as glycosyl donors in many enzymatic glycosylation processes. In bacteria UDP-N-acetylglucosamine (UDP-GlcNAc) is synthesized from fructose-6-phosphate by four successive reactions catalyzed by three enzymes: Glucosamine-6-phosphate synthase (GlmS), phosphoglucosamine mutase (GlmM), and the bi-functional enzyme glucosamine-1-phosphate acetyltransferase/N-acetylglucosamine-1-phosphate uridyltransferase (GlmU). In this work several metabolic engineering strategies, aimed to increment UDP-GlcNAc biosynthesis, were applied in the probiotic bacterium Lactobacillus casei strain BL23. This strain does not produce exopolysaccharides, therefore it could be a suitable host for the production of oligosaccharides. The genes glmS, glmM, and glmU coding for GlmS, GlmM, and GlmU activities in L. casei BL23, respectively, were identified, cloned and shown to be functional by homologous over-expression. The recombinant L. casei strain over-expressing simultaneously the genes glmM and glmS showed a 3.47 times increase in GlmS activity and 6.43 times increase in GlmM activity with respect to the control strain. Remarkably, these incremented activities resulted in about fourfold increase of the UDP-GlcNAc pool. In L. casei BL23 wild type strain transcriptional analyses showed that glmM and glmU are constitutively transcribed. By contrast, glmS transcription is down-regulated with a 21-fold decrease of glmS mRNA in cells cultured with N-acetylglucosamine as the sole carbon source compared to cells cultured with glucose. Our results revealed for the first time that GlmS, GlmM, and GlmU are responsible for UDP-GlcNAc biosynthesis in lactobacilli.

B6.Y(Tir) (mice with Y chromosome from a strain in Tirano, Italy, and autosomes and X-chromosomes from the B6 strain) mice provide an excellent model for analysing sex development that occurs during gonadal differentiation; however, the molecular mechanisms that contribute to sex reversal are unclear. Our aim has been to establish which molecular events participate in this sex reversal. The pattern of gene expression related to testicular [Sry (sex-determining region of the Y chromosome), Sox9 (Sry-related high-mobility group box gene 9) and Mis (Müllerian-inhibiting substance)] and ovarian [Wnt4 (Wingless-type MMTV (murine-mammary-tumour virus) integration site family, member 4), Rspo1 (cysteine-rich secretory protein containing a thrombospondin type 1 repeat) and Stra8 (stimulated by retinoic acid gene 8)] differentiation was analysed by applying immunofluorescence and real-time RT-PCR (reverse transcription-PCR), focusing on XY gonads from the B6.Y(Tir) mouse, but also analysing the normal strains CD-1 and C57BL/6J (B6). The expression of genes related to the process of sexual differentiation was altered in the case of the B6.Y(Tir) strain, both at the transcript and protein level, inducing differentiation of ovaries and ovotestes, but not the formation of the testes, which were normal. Our results indicate that the expression of testicular genes is inhibited at various levels, permitting the expression of ovarian genes such as Wnt4, Stra8 and Rspo1. However, their activity was not clear when the data were averaged. Correlation analysis indicated that an ovary differentiation pathway is activated when the testicular differentiation pathway is inhibited.

What’s known on the subject? and What does the study add? Bladder cancer susceptibility may be determined by genetic differences in the activity of glutathione S-transferases, enzymes that regulate the conversion of exogenous carcinogens to excretable hydrophilic metabolites by glutathione conjugation. The discrepancy of results regarding the association of common genetic polymorphisms and complex diseases such as cancer has raised scepticism in this area of research. Although the evidence generally supports the implication of GSTM1 and GSTT1 polymorphisms in bladder cancer, there is still some debate, with some studies in favour and some against. This study shows a greater risk of bladder cancer in individuals with GSTM1 null genotype, particularly women. This relationship is less evident with GSTT1 null genotypes. Null genotypes in both genes appear to be synergistic, particularly among smokers, and to increase the predisposition to more aggressive tumours. Nevertheless, the role of GSTM1 and GSTT1 polymorphisms in predisposition to bladder cancer should be viewed with caution, due to the multifactorial genetic origin of this condition and the need for long-term longitudinal studies to confirm these results. OBJECTIVE • To estimate the prevalence and importance of GSTT1 and GSTM1 genotypes (implicated in glutathione S-transferase activity) in bladder cancer, to determine whether smoking and occupational factors influence this relationship, and to identify the value of GSTT1 and GSTM1 genotypes as prognostic factors. PATIENTS AND METHODS • A cross-sectional study was conducted with a group of patients with bladder carcinoma and a control group with benign conditions and no history of tumours. The controls were selected and paired as subjects were recruited. • Sociodemographic variables, smoking, professional occupation, histological features and the presence of GSTT1 and GSTM1 polymorphisms by multiplex PCR techniques were assessed. RESULTS • GSTM1 genotypes were investigated in 201 patients and 193 controls and GSTT1 genotypes in 190 patients and 163 controls. • In the patients group, GSTT1 null genotype was observed in 22.1% (not significant) and GSTM1 null genotype in 54.2% (P= 0.008) (odds ratio, OR, 1.7); when considered together, 15.5% (P < 0.05; OR, 3.5) of patients had both null genotypes. In the multivariate analysis, the presence of GSTM1 null genotype remained in the model (OR, 2.1) in addition to smoking and age. • Subjects with bladder tumour and GSTM1 null genotype were younger than patients without gene deletion (P= 0.049). • Women with GSTM1 null genotype presented a higher OR than men (P= 0.024). When stratified by smoking habit, smokers with both null genotypes showed an OR of 4.7. • The percentage of patients with G3 tumours was higher in patients with GSTT1 null genotype (P= 0.013) and in patients with both null genotypes (P= 0.002). • A higher percentage of infiltrating tumours was also observed in patients with both null genotypes (P= 0.035). CONCLUSIONS • The data obtained in the present study suggest a higher risk of bladder cancer in individuals with the GSTM1 null genotype. • This risk is twofold higher when GSTM1 and GSTT1 null genotypes are both present and is also higher in smokers. • A greater predisposition for more aggressive tumours appears to exist, particularly when both null genotypes are combined. • Longer-term longitudinal studies are needed to confirm these results.

Electrostatic contributions to the conformational stability of apoflavodoxin were studied by measurement of the proton and salt-linked stability of this highly acidic protein with urea and temperature denaturation. Structure-based calculations of electrostatic Gibbs free energy were performed in parallel over a range of pH values and salt concentrations with an empirical continuum method. The stability of apoflavodoxin was higher near the isoelectric point (pH 4) than at neutral pH. This behavior was captured quantitatively by the structure-based calculations. In addition, the calculations showed that increasing salt concentration in the range of 0 to 500 mM stabilized the protein, which was confirmed experimentally. The effects of salts on stability were strongly dependent on cationic species: K(+), Na(+), Ca(2+), and Mg(2+) exerted similar effects, much different from the effect measured in the presence of the bulky choline cation. Thus cations bind weakly to the negatively charged surface of apoflavodoxin. The similar magnitude of the effects exerted by different cations indicates that their hydration shells are not disrupted significantly by interactions with the protein. Site-directed mutagenesis of selected residues and the analysis of truncation variants indicate that cation binding is not site-specific and that the cation-binding regions are located in the central region of the protein sequence. Three-state analysis of the thermal denaturation indicates that the equilibrium intermediate populated during thermal unfolding is competent to bind cations. The unusual increase in the stability of apoflavodoxin at neutral pH affected by salts is likely to be a common property among highly acidic proteins.

A retrospective study of a group of 51 patients who underwent surgery for squamous cell carcinoma of the pyriform sinus was performed. Primary tumors and lymph nodes were reviewed histologically. The primary tumors were also examined by flow cytometry for DNA ploidy and cell cycle analysis. Sixteen (33%) of the cases were aneuploid and 64% had a moderate or high S-phase fraction. The overall 3-year survival rate was 49% (25/51). In the univariate analysis, tumor size, lymphatic invasion, inflammatory infiltrate, presence of lymph node metastases, clinical and histologic N status, size and number of lymph nodes involved, and presence of extracapsular extension all correlated with survival. When multivariate analysis was used, the only independent prognostic factors were tumor size, lymphatic invasion, and histologic N status. Ploidy and S-phase fraction did not contribute further prognostic information.