Alternative splicing of BCL-x is controlled by RBM25 binding to a G-quadruplex in BCL-x pre-mRNA.

BCL-x is a master regulator of apoptosis whose pre-mRNA is alternatively spliced into either a long (canonical) anti-apoptotic Bcl-xL isoform, or a short (alternative) pro-apoptotic Bcl-xS isoform. The balance between these two antagonistic isoforms is tightly regulated and overexpression of Bcl-xL has been linked to resistance to chemotherapy in several cancers, whereas overexpression of Bcl-xS is associated to some forms of diabetes and cardiac disorders. The splicing factor RBM25 controls alternative splicing of BCL-x: its overexpression favours the production of Bcl-xS, whereas its downregulation has the opposite effect. Here we show that RBM25 directly and specifically binds to GQ-2, an RNA G-quadruplex (rG4) of BCL-x pre-mRNA that forms at the vicinity of the alternative 5' splice site leading to the alternative Bcl-xS isoform. This RBM25/rG4 interaction is crucial for the production of Bcl-xS and depends on the RE (arginine-glutamate-rich) motif of RBM25, thus defining a new type of rG4-interacting domain. PhenDC3, a benchmark G4 ligand, enhances the binding of RBM25 to the GQ-2 rG4 of BCL-x pre-mRNA, thereby promoting the alternative pro-apoptotic Bcl-xS isoform and triggering apoptosis. Furthermore, the screening of a combinatorial library of 90 putative G4 ligands led to the identification of two original compounds, PhenDH8 and PhenDH9, superior to PhenDC3 in promoting the Bcl-xS isoform and apoptosis. Thus, favouring the interaction between RBM25 and the GQ-2 rG4 of BCL-x pre-mRNA represents a relevant intervention point to re-sensitize cancer cells to chemotherapy.

Growth, survival, digestive activity and respiratory metabolism of Octopus maya juveniles fed with prepared diets.

The effects of two prepared feeds were tested on growth, survival, enzymatic activity, nutritive reserves in the digestive gland and oxygen consumption of Octopus maya juveniles. For the first time, a semihumid paste (HD, control) and a dry diet, in pelleted form (PD, experimental) with the same formulation were used for this species. The experiment lasted for 42 days. Results indicate that growth rates were similar for both diets (p > 0.05); however, survival (70%) was higher with the PD compared to the HD (48%) (p < 0.05). The performance index was higher for octopuses fed the PD (p < 0.05). No differences in acid proteases activity were observed. However, a higher activity of alkaline proteases in the octopuses fed the PD was observed (p < 0.05). Ingestion rate was higher for octopuses fed the PD. Routine energy inversion was similar in both treatments (p > 0.05). A greater energy inversion was observed in octopuses fed the PD, whose active metabolism was double compared to the octopuses fed the HD. Results showed that the PD promoted similar growth compared to the HD diet but favored survival, and a greater investment in the active metabolism, reflected in the apparent heat increase.

Comparative analysis of periodontal pain and quality of life in patients with fixed multibracket appliances and aligners (Invisalign®): longitudinal clinical study.

BACKGROUNDS: The aim of this longitudinal clinical study is to analyse and compare according to location, degree and type, the pain presented by patients during their first year of treatment, as well as the quality of oral life after the placement of two types of orthodontic appliances: conventional brackets and removable Invisalign ® aligners. METHODS: The sample consisted of 140 patients grouped into 2 study groups of 70 patients each. The first group (brackets group- BG), with fixed multibracket appliances, using the MBT technique and a 0.022" slot. The second group (invisaling group- IG), in treatment with removable aligners (IG), using the Invisalign ® system. They were providen with a questionnare where they had to record the degree (mild, moderate or intense), the type and location of the pain monthly during the first year of treatment. The second form was the Spanish version of the OHIP-14, oral quality of life questionnaire, which was provided the twelfth month after the start of treatment. RESULTS: In both groups, we found that the most frequent location of pain occurred during the first phase: mandibular for the IG group and maxillary in the BG group. Throughout the whole analysis, the intensity was mild-moderate with lower values in the conventional brackets' group. The BG group reported acute pain while the IG group reported sensitive pain during the first month; later both reported sensitive pain. CONCLUSIONS: There are differences in terms of periodontal pain in its degree, location, and type according to the different orthodontic techniques used. TRIAL REGISTRATION: The study was approved by the bioethics committee of the University of Salamanca (USAL_20/516).

Acetophenyl-thienyl-aniline-Linked Nucleotide for Construction of Solvatochromic Fluorescence Light-Up DNA Probes Sensing Protein-DNA Interactions.

The synthesis of 2'-deoxycytidine and its 5'-O-triphosphate bearing solvatochromic acetophenyl-thienyl-aniline fluorophore was developed using the Sonogashira cross-coupling reaction as the key step. The triphosphate was used for polymerase synthesis of labelled DNA. The labelled nucleotide or DNA exerted weak red fluorescence when excited at 405 nm, but a significant colour change (to yellow or green) and light-up (up to 20 times) was observed when the DNA probes interacted with proteins or lipids.

Nucleotides bearing aminophenyl- or aminonaphthyl-3-methoxychromone solvatochromic fluorophores for the enzymatic construction of DNA probes for the detection of protein-DNA binding.

We designed and synthesized nucleosides bearing aminophenyl- or aminonaphthyl-3-methoxychromone fluorophores attached at position 5 of cytosine or thymine and converted them to nucleoside triphosphates. The fluorophores showed solvatochromic fluorescence with strong fluorescence at 433-457 nm in non-polar solvents and very weak fluorescence at 567 nm in alcohols. The nucleosides and nucleotides also showed only negligible fluorescence in alcohols or water. The triphosphates were substrates for DNA polymerase in the enzymatic synthesis of modified DNA probes that showed only very weak fluorescence in aqueous buffer but a significant light-up and blue shift were observed when they interacted with proteins (histone H3.1 or p53 for double-stranded DNA probes or single-strand binding protein for single-stranded oligonucleotide probes). Hence, nucleotides have good potential in the construction of DNA sensors for studying protein-DNA interactions. The modified dNTPs were also transported into cells using a cyclodextrin-based transporter but they were not incorporated into the genomic DNA.

Thiophene-linked tetramethylbodipy-labeled nucleotide for viscosity-sensitive oligonucleotide probes of hybridization and protein-DNA interactions.

Cytosine 2'-deoxyribonucleoside dCTBdp and its triphosphate (dCTBdpTP) bearing tetramethylated thiophene-bodipy fluorophore attached at position 5 were designed and synthesized. The green fluorescent nucleoside dCTBdp showed a perfect dependence of fluorescence lifetime on the viscosity. The modified triphosphate dCTBdpTP was substrate to several DNA polymerases and was used for in vitro enzymatic synthesis of labeled oligonucleotides (ONs) or DNA by primer extension. The labeled single-stranded ONs showed a significant decrease in mean fluorescence lifetime when hybridized to the complementary strand of DNA or RNA and were also sensitive to mismatches. The labeled dsDNA sensed protein binding (p53), which resulted in the increase of its fluorescence lifetime. The triphosphate dCTBdpTP was transported to live cells where its interactions could be detected by FLIM but it did not show incorporation to genomic DNA in cellulo.

Brightly Fluorescent 2'-Deoxyribonucleoside Triphosphates Bearing Methylated Bodipy Fluorophore for in Cellulo Incorporation to DNA, Imaging, and Flow Cytometry.

Synthesis of cytosine, uracil, and 7-deazaadenine 2'-deoxyribonucleosides and triphosphates (dNTPs) bearing hexamethylated phenyl-bodipy fluorophore attached at position 5 of pyrimidines or at position 7 of 7-deazapurine was developed. All the title labeled nucleosides and dNTPs displayed bright green fluorescence with very high quantum yields. The modified dNmBdpTPs were good substrates to diverse DNA polymerases and were used for in vitro enzymatic synthesis of labeled DNA by primer extension or PCR. In combination with cationic cyclodextrin-peptide-based dNTP transporter, the dNmBdpTPs were successfully used for staining of genomic DNA in living cells for applications in confocal microscopy and in flow cytometry. The best performing cytosine nucleotide dCmBdpTP was used to monitor mitosis in live cells.

Inhibition of non-templated nucleotide addition by DNA polymerases in primer extension using twisted intercalating nucleic acid modified templates.

A simple and elegant method for inhibition of non-templated nucleotide addition by DNA polymerases and for following DNA 3'-heterogeneity in enzymatic DNA synthesis by primer extension (PEX) is described. When template bearing ortho-twisted intercalating nucleic acid (ortho-TINA) at the 5'-end is used, non-templated nucleotide addition is reduced in both the A- and B-family DNA polymerases (KOD XL, KOD (exo-), Bst 2.0, Therminator, Deep Vent (exo-) and Taq). Formation of a single oligonucleotide product was observed with ortho-TINA modified template and KOD XL, KOD (exo-), Bst 2.0, Deep Vent (exo-) and Taq DNA polymerases. This approach can be applied to the synthesis of both unmodified and base-modified oligonucleotides.

Fluorescence quenching in oligonucleotides containing 7-substituted 7-deazaguanine bases prepared by the nicking enzyme amplification reaction.

Recently, we reported the use of the Nicking Enzyme Amplification Reaction (NEAR) for the enzymatic synthesis of short oligonucleotides (ONs) containing 5-substituted pyrimidine or 7-substituted 7-deazaadenine nucleotides. Since no oligonucleotide products were visible on agarose gels stained by an intercalating dye (GelRed), we assumed that the method did not work for 7-substituted 7-deazaguanine deoxyribonucleoside triphosphates. We revisited the work and found that the NEAR method works for 7-deazaguanine nucleotides as well but that the resulting modified ONs quench the fluorescence of DNA intercalators, rendering them invisible on gel electrophoresis stained by them. Here, we report on the modified methodology for the NEAR synthesis and analysis of G-modified ONs and on quantification of the fluorescence quenching.

Cephalopod culture: current status of main biological models and research priorities.

A recent revival in using cephalopods as experimental animals has rekindled interest in their biology and life cycles, information with direct applications also in the rapidly growing ornamental aquarium species trade and in commercial aquaculture production for human consumption. Cephalopods have high rates of growth and food conversion, which for aquaculture translates into short culture cycles, high ratios of production to biomass and high cost-effectiveness. However, at present, only small-scale culture is possible and only for a few species: the cuttlefish Sepia officinalis, the loliginid squid Sepioteuthis lessoniana and the octopuses Octopus maya and O. vulgaris. These four species are the focus of this chapter, the aims of which are as follows: (1) to provide an overview of the culture requirements of cephalopods, (2) to highlight the physical and nutritional requirements at each phase of the life cycle regarded as essential for successful full-scale culture and (3) to identify current limitations and the topics on which further research is required. Knowledge of cephalopod culture methods is advanced, but commercialization is still constrained by the highly selective feeding habits of cephalopods and their requirement for large quantities of high-quality (preferably live) feed, particularly in the early stages of development. Future research should focus on problems related to the consistent production of viable numbers of juveniles, the resolution of which requires a better understanding of nutrition at all phases of the life cycle and better broodstock management, particularly regarding developments in genetic selection, control of reproduction and quality of eggs and offspring.

Evaluation of Octopus maya enzyme activity of the digestive gland and gastric juice.

As the demand for Octopus maya grows, sustainable farming practices become essential to prevent overexploitation, so that farming can be developed as a sustainable alternative to traditional fishing. Understanding the digestive dynamics of the octopus is essential for devising optimal dietary formulations in aquaculture. Despite the progress in understanding cephalopod digestion, little is known about the specific functioning of the digestive enzymes responsible for breaking down protein substrates. This knowledge gap underscores the need for further research to support sustainable O. maya population management. In this paper, dietary formulations are identified for cephalopods by characterizing O. maya digestive enzymes present in the digestive gland and gastric juice. The investigation revealed that acidic proteases showed a peak activity at higher temperatures than alkaline proteases. Inhibitors confirmed the presence of H, L, and D cathepsins. The lower activation energy of alkaline enzymes compared to acidic ones observed highlights an intriguing aspect of O. maya's digestive physiology. This research provides valuable insights into O. maya digestive enzyme functions, representing a significant advancement in formulating diets crucial for successful octopus farming that may help to fully understand its physiology.

A New Digital Method to Quantify the Effects Produced by Carriere Motion Appliance.

The aim of this study was to analyze a novel digital technique to quantify the distal tooth displacement and derotation angle produced by the Carriere Motion Appliance (CMA). Twenty-one patients with a class II molar and canine relationship underwent orthodontic treatment with CMA. All patients were exposed before (STL1) and after the CMA placement (STL2), submitted to a digital impression, and afterwards, data were uploaded to specific cephalometric software to allow automatic mesh network alignment of the STL digital files. Subsequently, the distal tooth displacement of the upper canines and first upper molars, as well as the derotation angle of the first upper molars were analyzed using the Pearson correlation coefficient (ρ). Repeatability and reproducibility were analyzed using Gage R&R statistical analysis. An increase in canine displacement was correlated with an increase in contralateral canine displacement (ρ = 0.759; p < 0.000). An increase in canine displacement was correlated with an increase in molar displacement (ρ = 0.715; p < 0.001). An increase in upper first molar displacement was correlated with an increase in the contralateral upper first molar displacement (ρ = 0.609; p < 0.003) and the canine displacement (ρ = 0.728; p < 0.001). The distal tooth displacement showed a repeatability of 0.62% and reproducibility of 7.49%, and the derotation angle showed a repeatability of 0.30% and reproducibility of 0.12%. The novel digital measurement technique is a reproducible, repeatable, and accurate method for quantifying the distal tooth displacement of the upper canine and first upper molar, as well as the derotation angle of the first upper molars after using CMA.

Programming Positive Mechanofluorescence in Liquid Crystalline Elastomers.

Liquid single crystal elastomers (LSCEs) containing organic fluorophores within their polymeric network are attractive materials to detect forces with simple spectroscopic measurements. Hitherto, all mechanoluminescent LSCEs decrease their emission intensity upon mechanical stimulation; that is, they display negative mechanofluorescence. Such behavior is governed by the mechanically induced approximation of the quenching mesogenic units and the fluorophores. In this work, we propose the integration of fluorescent molecular rotors (FMRs), whose luminescence is not quenched by the mesogens, in LSCEs as a valuable strategy to conceive elastomeric materials programmed with exactly the opposite behavior, i.e., their fluorescence increases upon deformation (positive mechanofluorescence). Specifically, carbazole-indolenine dyes are interesting candidates for this purpose since their luminescence depends mainly on the degree of intramolecular rotation allowed by the local environment. On this basis, the uniaxial deformation of an LSCE, along its anisotropic direction, incorporating such FMRs will place the fluorophores in a more restricted medium, leading to the desired enhanced emission at the macroscale.

Nutritive value of diets containing fish silage for juvenile Litopenaeus vannamei (Bonne, 1931).

BACKGROUND: Fish wastes has been used for many years as an alternative in feeds for aquaculture. In the present study weight gain of juvenile white shrimp Litopenaeus vannamei fed diets including fish waste silage (WS), fish waste silage with soybean meal SBM (WS + S) or fish waste meal (WM) was compared. A conventional acidic silage process was applied to obtain from wastes (skin, heads, bones and viscera) of snapper (Lutjanus spp.), grunt (Haemulon plumieri), and grouper (Epinephelus spp.) an ingredient rich in protein. RESULTS: After 3 days ensilage more than 90% protein was hydrolysed. Waste material processed at pH 3.8 lost about 24% tryptophan. Butylated hydroxytoluene (BHT) prevented lipid oxidation, as shown after 45 days with malonaldehyde production. Shrimp fed WS + S diet gained 0.7 g per week higher than those fed WS and WM diets with 0.3 g per week (P < 0.05). CONCLUSION: WS processed with formic acid under conditions of low pH is beneficial for the white shrimp L. vannamei. It sustained reasonable weight gain combined with soybean meal in practical diets. On the other hand, BHT addition was beneficial in preventing oxidative action during silage preparation.

Changes in the Color and Brightness of White Spots Associated with Orthodontic Treatment 6 Months after the Application of Infiltrative Resins: Systematic Review and Meta-Analysis.

One of the risks that we find after orthodontic treatment is the secondary appearance of white spot lesions (WLS) after the removal of fixed multi-bracket appliances. Today, there are several treatment methods, resin infiltration being the most used in the most serious cases. The objective of this study is to carry out a systematic review and meta-analysis to determine the efficacy and stability in the variables of color and gloss, six months after resin infiltration. A comprehensive search was performed in the following databases: PubMed, Embase, Google Scholar, Scopus, Medline, and Web of Science. Articles published in the last 10 years were selected, including in vivo studies with a six-month follow-up. PRISMA guidelines were followed to carry out this systematic review. All studies where the application of resin was performed on carious lesions were discarded. Once the inclusion and exclusion criteria were applied, a final sample of four articles was obtained, on which the review and meta-analysis were carried out. Once examined, all authors considered that there was an immediate improvement in both variables. However, statistically significant differences were obtained in the color change outcome, but not in the brightness outcome in the subgroup analysis after six months of icon resin infiltration.

The different activities of RNA G-quadruplex structures are controlled by flanking sequences.

The role of G-quadruplex (G4) RNA structures is multifaceted and controversial. Here, we have used as a model the EBV-encoded EBNA1 and the Kaposi's sarcoma-associated herpesvirus (KSHV)-encoded LANA1 mRNAs. We have compared the G4s in these two messages in terms of nucleolin binding, nuclear mRNA retention, and mRNA translation inhibition and their effects on immune evasion. The G4s in the EBNA1 message are clustered in one repeat sequence and the G4 ligand PhenDH2 prevents all G4-associated activities. The RNA G4s in the LANA1 message take part in similar multiple mRNA functions but are spread throughout the message. The different G4 activities depend on flanking coding and non-coding sequences and, interestingly, can be separated individually. Together, the results illustrate the multifunctional, dynamic and context-dependent nature of G4 RNAs and highlight the possibility to develop ligands targeting specific RNA G4 functions. The data also suggest a common multifunctional repertoire of viral G4 RNA activities for immune evasion.

Novel Digital Measurement Technique to Analyze the Palatine Suture Expansion after Palatine Rapid Maxillary Expansion Technique.

The aim of the study was to validate a novel digital measurement method to quantify the volume of the midpalatal suture after rapid maxillary expansion (RME). MATERIAL AND METHODS: Eight patients with maxillary skeletal transverse deficiency were submitted to palatine suture expansion using the McNamara orthodontic appliance during a period of nine months. After 30 days of treatment, all patients were exposed postoperatively to a cone-beam computed tomography (CBCT) scan. Afterwards, the scans were uploaded into the three-dimensional orthodontic-planning software to allow the volumetric assessment of the palatine suture expansion through palatine rapid maxillary expansion using a McNamara appliance. The repeatability was analyzed by repeating twice the measures by a single operator and reproducibility was analyzed by repeating three times the measures by two operators, and Gage R&R statistical analysis was performed. RESULTS: The expansion of the midpalatal suture by means of the rapid maxillary expansion technique, in terms of digital volume measurement, showed a repeatability value of 0.09% and between the two operators a reproducibility value of 0.00% was shown. CONCLUSIONS: The novel measurement technique demonstrated a high repeatability and reproducibility rate for volume assessment of the palatine suture expansion through palatine rapid maxillary expansion technique.

Cu4MnGe2S7 and Cu2MnGeS4: two polar thiogermanates exhibiting second harmonic generation in the infrared and structures derived from hexagonal diamond.

The new, quaternary diamond-like semiconductor (DLS) Cu4MnGe2S7 was prepared at high-temperature from a stoichiometric reaction of the elements under vacuum. Single crystal X-ray diffraction data were used to solve and refine the structure in the polar space group Cc. Cu4MnGe2S7 features [Ge2S7]6- units and adopts the Cu5Si2S7 structure type that can be considered a derivative of the hexagonal diamond structure. The DLS Cu2MnGeS4 with the wurtz-stannite structure was similarly prepared at a lower temperature. The achievement of relatively phase-pure samples, confirmed by X-ray powder diffraction data, was nontrival as differential thermal analysis shows an incongruent melting behaviour for both compounds at relatively high temperature. The dark red Cu2MnGeS4 and Cu4MnGe2S7 compounds exhibit direct optical bandgaps of 2.21 and 1.98 eV, respectively. The infrared (IR) spectra indicate potentially wide windows of optical transparency up to 25 µm for both materials. Using the Kurtz-Perry powder method, the second-order nonlinear optical susceptibility, χ(2), values for Cu2MnGeS4 and Cu4MnGe2S7 were estimated to be 16.9 ± 2.0 pm V-1 and 2.33 ± 0.86 pm V-1, respectively, by comparing with an optical-quality standard reference material, AgGaSe2 (AGSe). Cu2MnGeS4 was found to be phase matchable at λ = 3100 nm, whereas Cu4MnGe2S7 was determined to be non-phase matchable at λ = 1600 nm. The weak SHG response of Cu4MnGe2S7 precluded phase-matching studies at longer wavelengths. The laser-induced damage threshold (LIDT) for Cu2MnGeS4 was estimated to be ∼0.1 GW cm-2 at λ = 1064 nm (pulse width: τ = 30 ps), while the LIDT for Cu4MnGe2S7 could not be ascertained due to its weak response. The significant variance in NLO properties can be reasoned using the results from electronic structure calculations.

Dietary Potassium Downregulates Angiotensin-I Converting Enzyme, Renin, and Angiotensin Converting Enzyme 2.

BACKGROUND: The importance of dietary potassium in health and disease has been underestimated compared with that placed on dietary sodium. Larger effort has been made on reduction of sodium intake and less on the adequate dietary potassium intake, although natural food contains much more potassium than sodium. The benefits of a potassium-rich diet are known, however, the mechanism by which it exerts its preventive action, remains to be elucidated. With the hypothesis that dietary potassium reduces renal vasoconstrictor components of the renin-angiotensin system in the long-term, we studied the effect of high potassium diet on angiotensin-I converting enzyme, renin, and angiotensin converting enzyme 2. METHODS: Sprague Dawley male rats on a normal sodium diet received normal potassium (0.9%, NK) or high potassium diet (3%, HK) for 4 weeks. Urine was collected in metabolic cages for electrolytes and urinary volume measurement. Renal tissue was used to analyze angiotensin-I converting enzyme, renin, and angiotensin converting enzyme 2 expression. Protein abundance analysis was done by Western blot; gene expression by mRNA levels by RT-qPCR. Renal distribution of angiotensin-I converting enzyme and renin was done by immunohistochemistry and morphometric analysis in coded samples. RESULTS: High potassium diet (4 weeks) reduced the levels of renin, angiotensin-I converting enzyme, and angiotensin converting enzyme 2. Angiotensin-I converting enzyme was located in the brush border of proximal tubules and with HK diet decreased the immunostaining intensity (P < 0.05), decreased the mRNA (P < 0.01) and the protein levels (P < 0.01). Renin localization was restricted to granular cells of the afferent arteriole and HK diet decreased the number of renin positive cells (P < 0.01) and renin mRNA levels (P < 0.01). High potassium intake decreased angiotensin converting enzyme 2 gene expression and protein levels (P < 0.01).No morphological abnormalities were observed in renal tissue during high potassium diet.The reduced expression of angiotensin-I converting enzyme, renin, and angiotensin converting enzyme 2 during potassium supplementation suggest that high dietary potassium intake could modulate these vasoactive enzymes and this effects can contribute to the preventive and antihypertensive effect of potassium.

Best compromise nutritional menus for childhood obesity.

Childhood obesity is an undeniable reality that has rapidly increased in many countries. Obesity at an early age not only increases the risks of chronic diseases but also produces a problem for the whole healthcare system. One way to alleviate this problem is to provide each patient with an appropriate menu that is defined by a mathematical model. Existing mathematical models only partially address the objective and constraints of childhood obesity; therefore, the solutions provided are insufficient for health specialists to prepare nutritional menus for individual patients. This manuscript proposes a multiobjective mathematical programming model to aid in healthy nutritional menu planning that may prevent childhood obesity. This model provides a plan for combinations and amounts of food across different schedules and daily meals. This approach minimizes the major risk factors of childhood obesity (i.e., glycemic load and cholesterol intake). In addition, this approach considers the minimization of nutritional mismatch and total cost. The model is solved using a deterministic method and two metaheuristic methods. Test instances associated with children aged 4-18 years were created with the support of health professionals to complete this numerical study. The quality of the solutions generated using the three methods was similar, but the metaheuristic methods provided solutions in a shorter computational time. These results are submitted to statistical hypothesis tests to be validated. The numerical results indicate proper guidelines for personalized plans for individual children.