PPARG is dispensable for bovine embryo development up to tubular stages.

Following blastocyst hatching, ungulate embryos undergo a prolonged preimplantation period termed conceptus elongation. Conceptus elongation constitutes a highly susceptible period for embryonic loss and the embryonic requirements during this process are largely unknown, but multiple lipid compounds have been identified in the fluid nourishing the elongating conceptuses. Peroxisome proliferator-activated receptors (PPARs) mediate the signaling actions of prostaglandins and other lipids and, between them, PPARG has been pointed out to play a relevant role on conceptus elongation by a functional study that depleted PPARG in both uterus and conceptus. The objective of this study has been to determine if embryonic PPARG is required for bovine embryo development. To that aim, we have generated bovine PPARG KO embryos in vitro by two independent gene ablation strategies and assess their developmental ability. In vitro development to Day (D) 8 blastocyst was unaffected by PPARG ablation, as total, inner cell mass and trophectoderm cell numbers were similar between WT and KO D8 embryos. In vitro post-hatching development to D12 was also comparable between different genotypes, as embryo diameter, epiblast cell number, and embryonic disc formation and hypoblast migration rates were unaffected by the ablation. The development to tubular stages equivalent to E14 was assessed in vivo, following a heterologous embryo transfer experiment, observing that the development of extra-embryonic membranes and of the embryonic disc was not altered by PPARG ablation. In conclusion, PPARG ablation did not impaired bovine embryo development up to tubular stages.

Integration of QTL and transcriptome approaches for the identification of genes involved in tomato response to nitrogen deficiency.

Optimising plant nitrogen (N) usage and inhibiting N leaching loss in the soil-crop system is crucial to maintaining crop yield and reducing environmental pollution. This study aimed at identifying quantitative trait loci (QTLs) and differentially expressed genes (DEGs) between two N treatments in order to list candidate genes related to nitrogen-related contrasting traits in tomato varieties. We characterised a genetic diversity core-collection (CC) and a multi-parental advanced generation intercross (MAGIC) tomato population grown in greenhouse under two nitrogen levels and assessed several N-related traits and mapped QTLs. Transcriptome response under the two N conditions was also investigated through RNA sequencing of fruit and leaves in four parents of the MAGIC population. Significant differences in response to N input reduction were observed at the phenotypic level for biomass and N-related traits. Twenty-seven (27) QTLs were detected for three target traits (Leaf N content, leaf Nitrogen Balance Index and petiole NO3- content), ten and six at low and high N condition, respectively; while 19 QTLs were identified for plasticity traits. At the transcriptome level, 4,752 and 2,405 DEGs were detected between the two N conditions in leaves and fruits, respectively, among which 3,628 (50.6%) in leaves and 1,717 (71.4%) in fruit were genotype specific. When considering all the genotypes, 1,677 DEGs were shared between organs or tissues. Finally, we integrated DEGs and QTLs analyses to identify the most promising candidate genes. The results highlighted a complex genetic architecture of N homeostasis in tomato and novel putative genes useful for breeding tomato varieties requiring less N input.

Vibrational signatures of dynamic excess proton storage between primary amine and carboxylic acid groups.

Ammonium and carboxylic moieties play a central role in proton-mediated processes of molecular recognition, charge transfer or chemical change in (bio)materials. Whereas both chemical groups constitute acid-base pairs in organic salt-bridge structures, they may as well host excess protons in acidic environments. The binding of excess protons often precedes proton transfer reactions and it is therefore of fundamental interest, though challenging from a quantum chemical perspective. As a benchmark for this process, we investigate proton storage in the amphoteric compound 5-aminovaleric acid (AV), within an intramolecular proton bond shared by its primary amine and carboxylic acid terminal groups. Infrared ion spectroscopy is combined with ab initio Molecular Dynamics (AIMD) calculations to expose and rationalize the spectral signatures of protonated AV and its deuterated isotopologues. The dynamic character of the proton bond confers a fluxional structure to the molecular framework, leading to wide-ranging bands in the vibrational spectrum. These features are reproduced with remarkable accuracy by AIMD computations, which serves to lay out microscopic insights into the excess proton binding scenario.

Follistatin-like 1 (FSTL1) levels as potential early biomarker of cardiovascular disease in a Mexican population.

Cardiovascular diseases (CVD) are the leading cause of death globally. In recent years, follistatin-like protein 1 (FSTL1) has been proposed as an emerging potential clinical biomarker of CVD, since its concentration is upregulated in heart failure. The aim of the present study was to evaluate the association of FSTL1 levels and classic biomarkers with the risk of CVD in Mexican population. A case-control study was carried out in patients with cardiovascular diseases (CVD), arterial hypertension, but not CVD (cardiovascular risk factor-CRF), and healthy controls (control group) from the Mexican Institute of Social Security. Lipid profile, homocysteine (Hcys), serum amyloid A (SAA), FSTL1 concentration, PON1 concentration and activities [Arylesterase (ARE), and Lactonase (LAC)] were evaluated. High levels of FSTL1 were found in the CRF group and a positive association of FSTL1 (OR = 4.55; 95% CI 1.29-16.04, p = 0.02) with the presence of arterial hypertension, as well as Hcys (OR, 3.09; 95% CI 1.23-7.76, p = 0.02) and SAA (OR, 1.03; 95% CI 1.01-1.05, p < 0.01) with the presence of CVD. LAC activity (OR, 0.26; 95% CI 0.07-0.94, p = 0.04) and PON1 concentration (OR, 0.17; 95% CI 0.05-0.62, p = 0.01) were associated with a decrease in OR belonging to the group with CVD. Our results suggest that FSTL1 may be a useful biomarker for monitoring cardiovascular risk in clinical settings. However, longitudinal studies are needed to evaluate how FSTL1 could influence the association of PON1 activity and Hcys with CVD.

High-resolution structural insights into the heliorhodopsin family.

Rhodopsins are the most abundant light-harvesting proteins. A new family of rhodopsins, heliorhodopsins (HeRs), has recently been discovered. Unlike in the known rhodopsins, in HeRs the N termini face the cytoplasm. The function of HeRs remains unknown. We present the structures of the bacterial HeR-48C12 in two states at the resolution of 1.5 Å, which highlight its remarkable difference from all known rhodopsins. The interior of HeR's extracellular part is completely hydrophobic, while the cytoplasmic part comprises a cavity (Schiff base cavity [SBC]) surrounded by charged amino acids and containing a cluster of water molecules, presumably being a primary proton acceptor from the Schiff base. At acidic pH, a planar triangular molecule (acetate) is present in the SBC. Structure-based bioinformatic analysis identified 10 subfamilies of HeRs, suggesting their diverse biological functions. The structures and available data suggest an enzymatic activity of HeR-48C12 subfamily and their possible involvement in fundamental redox biological processes.

The risk of incident depression when assessed with the Lifestyle and Well-Being Index.

OBJECTIVES: Novel findings indicate links between unhealthy lifestyles and depression based on active inflammatory processes. Thus, identifying participants with poor habits could reveal differences in trends of incident depression. This study aimed to examine the association between an objective lifestyle assessment, as measured by the Lifestyle and Well-Being Index (LWB-I), and incident depression in healthy participants of a Spanish cohort. STUDY DESIGN: This was a longitudinal analysis of a subsample of 10,063 participants from the Seguimiento Universidad de Navarra cohort study. METHODS: Group comparisons and Cox proportional hazard models were conducted using the LWB-I, which categorizes the sample into groups with healthy and unhealthy lifestyles and well-being. The main outcome was incident depression as well as secondary outcomes. RESULTS: Those classified to the transition category of LWB-I were associated with a hazard ratio of 0.67 (95% confidence interval: 0.52-0.87), and those in the excellent category showed a hazard ratio of 0.44 (95% confidence interval: 0.33-0.58), which in both groups reflects a significantly lower risk of incident depression compared with the group including those classified in the poor LWB-I level. Moreover, the available sensitivity analyses concerning time of depression diagnosis or antidepressant treatment further supported the role of nutrition and physical activity on incident depression. Interestingly, throughout the follow-up, incident depression was inversely related to healthier daily habits as measured by the LWB-I. CONCLUSIONS: A global assessment of lifestyles such as the LWB-I provides valuable insight into the complex relationship between lifestyle factors and their link to depression risk.

Cross-sectoral impacts of the 2018-2019 Central European drought and climate resilience in the German part of the Elbe River basin.

The 2018-2019 Central European drought was probably the most extreme in Germany since the early sixteenth century. We assess the multiple consequences of the drought for natural systems, the economy and human health in the German part of the Elbe River basin, an area of 97,175 km2 including the cities of Berlin and Hamburg and contributing about 18% to the German GDP. We employ meteorological, hydrological and socio-economic data to build a comprehensive picture of the drought severity, its multiple effects and cross-sectoral consequences in the basin. Time series of different drought indices illustrate the severity of the 2018-2019 drought and how it progressed from meteorological water deficits via soil water depletion towards low groundwater levels and river runoff, and losses in vegetation productivity. The event resulted in severe production losses in agriculture (minus 20-40% for staple crops) and forestry (especially through forced logging of damaged wood: 25.1 million tons in 2018-2020 compared to only 3.4 million tons in 2015-2017), while other economic sectors remained largely unaffected. However, there is no guarantee that this socio-economic stability will be sustained in future drought events; this is discussed in the light of 2022, another dry year holding the potential for a compound crisis. Given the increased probability for more intense and long-lasting droughts in most parts of Europe, this example of actual cross-sectoral drought impacts will be relevant for drought awareness and preparation planning in other regions. Supplementary Information: The online version contains supplementary material available at 10.1007/s10113-023-02032-3.

Transit along the vas deferens results in a high percentage of filiform spermatozoa with compacted chromatin in the rooster (Gallus domesticus).

The present work aimed to evaluate the chromatin compaction of rooster spermatozoa along the male reproductive tract, and to study the vas deferens lining cells, potentially involved in sperm maturation. Chromomycin A3 (CMA3) was used to determine the chromatin compaction of spermatozoa from testis (T), proximal (including epididymis, V1), intermediate (V2) and distal (V3) vas deferens, and ejaculate (E). Six Birchen Leonesa roosters were used. E was obtained in vivo by dorso-ventral massage. V1, V2 and V3 sperm were obtained post mortem (six pairs of vasa deferentia), by flushing. T was obtained by washing the testes, cut in halves. The fixed cells were stained with CMA3 and propidium iodide for flow cytometry assessment. Results showed higher (P P P.

Freezing chicken semen: Influence of base medium osmolality, cryoprotectants, cryoprotectant concentration, and cooling rate on post-thaw sperm survival.

A classical chicken semen diluent (Lake's 7.1 diluent) was modified to have lowered osmolalities (ranging from 290 to 410 mOsm/kg). The modified medium with physiological osmolality of 325 mOsm/kg allowed cold storage of fresh semen for several days with very little loss of membrane integrity and motility, while high osmolalities inhibited motility. This modified medium was then used as base for freezing medium to test effects of the type and concentration of cryoprotective agent (CPA), and the cooling rate (CR). A number of CPAs (methylformamide, methylacetamide, dimethylformamide (DMF), dimethylacetamide (DMA), diethylformamide, and propylene glycol) were first compared by freezing semen with 0.6 mol/l of the respective CPA at a cooling rate of 250 °C/min. Post-thaw motility and membrane integrity were highest with DMA and DMF. Finally, in more detailed factorial experiments, semen from individual cocks or pooled semen was frozen using CRs of 4, 50, 250, and 440 °C/min and DMA concentrations ([DMA]) of 0.4, 0.6, 1.0, and 1.5 mol/l. Straws from each semen sample x treatment combination were divided for semen assessment at three different research groups for sperm motility, membrane integrity, kinked tails, and DNA fragmentation, using microscopy, computer assisted motility analysis, and flow cytometry. There were clear effects of both CR and [DMA] and their interaction. CRs 50 and 250 °C/min gave best post-thaw sperm performance. Higher DMA concentrations gave better post-thaw membrane integrity, but concentrations above 1.0 mol/l can decrease sperm velocity or even inhibit sperm motility. Therefore [DMA] may best be 0.6-1.0 mol/l at a CR of 50-250 °C/min.

Self-adjustment of spectral components to object wave field extraction in off-axis digital holography.

In this paper, we introduce a positioning system to self-adjust the spectral components of holograms recorded by an off-axis digital holography setup. The incidence angle of the reference arm is carried out by two motorized actuators controlled by an algorithm that automatically adjusts it to avoid the overlapping of the spectral components. The right positioning of the spectral components allows selection of a spectral region to extract only the virtual component so that, after an inverse Fourier transformation, the object wave field can be obtained, thus eliminating the undesired components and increasing the image quality of the reconstructed image.

A meta-analytic review of attitudes towards the sexuality of adults with intellectual disabilities as measured by the ASQ-ID and related variables: Is context the key?

BACKGROUND: The attitudes of others towards the sexuality of people with intellectual disabilities are one of the main perceived barriers to them expressing their sexuality. Research on what influences these attitudes yields heterogeneous results. METHOD: A systematic review of the literature and a meta-analysis were carried out. RESULTS: Eleven studies using the Attitudes to Sexuality Questionnaire-Individuals with an Intellectual Disability (ASQ-ID) were included. Within the included studies, the country's socio-economic development and level of individualism were associated with attitudes towards the sexual rights, parenting and self-control of adults with intellectual disabilities. General population and staff samples held more favourable attitudes than family samples in terms of sexual rights and parenting. Age and gender did not yield significant results. CONCLUSIONS: Variables related to country context may underlie the differences observed between countries and therefore influence the population's general thinking and ideologies. Unexpectedly, no age differences were observed. Gender-related results may reflect rapprochement between genders in sexuality. These findings are relevant for researchers and practitioners, as they suggest the importance of considering contextual factors when developing effective interventions that aim to support adults with disabilities to live their sexuality.

Birchen and Blue Leonesa sperm cryopreservation: a new technique for evaluating the integrity of cockerel sperm membranes.

1. Birchen and Blue Leonesa are two endangered chicken breeds mainly raised in Curueño Valley in North Spain. The establishment of a germplasm bank to guarantee the preservation of these breeds is needed. However, cockerels from different breeder flocks can show variance in semen cryoresistance.2. The following work focused on the sperm characterisation and cryopreservation of Birchen and Blue Leonesa cockerels from four different breeders. A total of 30 semen pools were analysed. Besides conventional sperm analysis, including motility by computer-aided sperm analysis (CASA) and DNA fragmentation by TUNEL, the present study tested a double staining method (MitoTrackerTM Green FM/propidium iodide). This gave simultaneous assessment of plasma and acrosomal and mitochondrial membranes, which were previously validated by SYBR-14/PI, CASA, aniline blue and TUNEL.3. No significant differences were found among fresh semen variables between breeds and breeders. For post-thawed variables, significant differences (P < 0.05) were found between breeders in sperm viability (58.0 ± 1.90 breeder D vs. 35.2 ± 7.41 breeder A, 37.2 ± 4.09 breeder B and 22.3 ± 5.92 breeder C) and DNA fragmentation (62.4 ± 9.91 breeder C vs. 31.8 ± 7.08 breeder B and 24.5 ± 5.49 breeder D). The lowest DNA fragmentation values for semen from breeder D birds were coincident with higher integrity of the mitochondrial membrane.4. The results revealed higher sperm cryoresistance in the cockerels from one of the breeders, possibly due to differences in management system (e.g. diet, housing, control of stress elements and pathogens, reproduction practices or maintenance of genetic diversity). These differences may determine the sperm freezability, and thus the effectiveness of developing a germplasm bank.

Mood in Parkinson's disease: From early- to late-stage disease.

BACKGROUND: Although depression is known to be frequent in Parkinson's disease (PD), it is unclear how mood can change and/or impact on patient's quality of life (QoL) over time. Our aim was to analyze the frequency of depression, mood related factors and the contribution of mood to a patient's QoL perception in regard to disease duration. METHODS: PD patients recruited from the COPPADIS cohort from January 2016 to November 2017 were included in this cross-sectional study. Three groups were defined: <5 years (Group A); from 5 to <10 years (Group B); ≥10 years (Group C). Analysis with well-planned linear regression models was conducted to determine how different factors contribute to mood (Beck Depression Inventory-II [BDI-II] as dependent variable), to health-related QoL (39-item Parkinson's Disease Questionnaire [PDQ-39SI] as dependent variable) and to global QoL (European Health Interview Survey - Quality of Life Eight-Item Index [EUROHIS-QOL8] as dependent variable). RESULTS: Six hundred and sixty-three PD patients (62.6 ± 8.9 years old, 59.6% males) were included: Group A, 50.1% (n = 332); Group B, 33.3% (n = 221) and Group C, 16.6% (n = 110). There were no differences between the three groups in terms of the frequency of depressive symptoms nor the frequency of depression type (major vs. minor vs. subthreshold) (p = 0.729). However, the unique percent variance of PDQ-39SI and EUROHIS-QOL8 explained by BDI-II total score was 2 (23.7%) and threefold (26.9%), respectively, in Group C compared to the other two groups. EUROHIS-QOL8 total score provided the highest unique contribution to mood (16.8%). CONCLUSIONS: Although depression-type frequency does not appear to change over time in PD; the contribution of mood on QoL perception is greater in patients with longer disease duration.

Ultrasonographic and hemodynamic characteristics of patients with symptomatic carotid near-occlusion: results from a multicenter registry study.

PURPOSE: The ultrasonographic and hemodynamic features of patients with carotid near-occlusion (CNO) are still not well known. Our aim was to describe the ultrasonographic and hemodynamic characteristics of a cohort of patients with CNO. METHODS: A prospective, observational, nationwide, and multicenter study was conducted from January/2010 to May/2016. Patients with digital subtraction angiography (DSA)-confirmed CNO were included. We collected information on clinical and demographic characteristics, carotid and transcranial ultrasonography and DSA findings, presence of full-collapse, collateral circulation, and cerebrovascular reactivity (CVR). RESULTS: One hundred thirty-five patients were analyzed. Ultrasonographic and DSA diagnosis of CNO were concordant in only 44%. This disagreement was related to the presence/absence of full-collapse: 45% of patients with CNO with full-collapse were classified as a complete carotid occlusion, and 40% with a CNO without full-collapse were interpreted as severe stenosis (p < 0.001). Mean velocities (mV) and pulsatility indexes (PIs) were significantly lower in the ipsilateral middle cerebral artery compared with the contralateral (43 cm/s vs 58 cm/s, p < 0.001; 0.80 vs 1.00, p < 0.001). Collateral circulation was identified in 92% of patients, with the anterior communicating artery (73%) being the most frequent. CVR was decreased or exhausted in 66% of cases and was more frequent in patients with a poor or absent collateral network compared with patients with ≥ 2 collateral arteries (82% vs 56%, p = 0.051). CONCLUSION: The accuracy of carotid ultrasonography in the diagnosis of CNO seems to be limited, with significant discrepancies with DSA. Decreased ipsilateral mV, PI, and CVR suggest a hemodynamic compromise in patients with CNO.

Engraftment after autologous hematopoietic stem cell transplantation in patients mobilized with Plerixafor: A retrospective, multicenter study of a large series of patients.

Plerixafor (PLX) appears to effectively enhance hematopoietic stem-cell mobilization prior to autologous hematopoietic stem cell transplantation (auto-HCT). However, the quality of engraftment following auto-HCT has been little explored. Here, engraftment following auto-HCT was assessed in patients mobilized with PLX through a retrospective, multicenter study of 285 consecutive patients. Information on early and 100-day post-transplant engraftment was gathered from the 245 patients that underwent auto-HCT. The median number of PLX days to reach the stem cell collection goal (≥2 × 106 CD34+ cells/kg) was 1 (range 1-4) and the median PLX administration time before apheresis was 11 h (range 1-18). The median number of apheresis sessions to achieve the collection goal was 2 (range 1-5) and the mean number of CD34+ cells collected was 2.95 × 106/kg (range 0-30.5). PLX administration was safe, with only 2 mild and transient gastrointestinal adverse events reported. The median time to achieve an absolute neutrophil count (ANC) >500/µL was 11 days (range 3-31) and the median time to platelet recovery >20 × 103/µL was 13 days (range 5-69). At 100 days after auto-HCT, the platelet count was 137 × 109/L (range 7-340), the ANC was 2.3 × 109/L (range 0.1-13.0), and the hemoglobin concentration was 123 g/L (range 79-165). PLX use allowed auto-HCT to be performed in a high percentage of poorly mobilized patients, resulting in optimal medium-term engraftment in the majority of patients in whom mobilization failed, in this case mainly due to suboptimal peripheral blood CD34+ cell concentration on day +4 or low CD34+ cell yield on apheresis.

Imaging of bee honey sugar crystals by second-harmonic generation microscopy.

Bee honey is an exceptionally nutritious food with unique chemical and mineral contents. This report introduces the use of the second-harmonic generation (SHG) microscopy for imaging honey sugar crystals' morphology as an alternative for its authentication process. The crystals and their boundaries are clearly observed with SHG compared with bright-field microscopy, where the liquid honey avoids the visualization of a sharp image. Four different honey samples of Mexico's various floral origins and geographical regions are analyzed in our study. These samples are representative of the diversity and valuable quality of bee honey production. The SHG image information is complemented with Raman spectroscopy (RS) analysis, since this optical technique is widely used to validate the bee's honey composition stated by its floral origin. We relate the SHG imaging of honey crystals with the well-defined fructose and glucose peaks measured by RS. Size measurement is introduced using the crystal´s length ratio to differentiate its floral origin. From our observations, we can state that SHG is a promising and suitable technique to provide a sort of optical fingerprint based on the floral origin of bee honey.

Arterial Wall Stiffening in Caveolin-1 Deficiency-Induced Pulmonary Artery Hypertension in Mice.

BACKGROUND: Pulmonary artery hypertension (PAH) is a complex disorder that can lead to right heart failure. The generation of caveolin-1 deficient mice (CAV-1-/-) has provided an alternative genetic model to study the mechanisms of pulmonary hypertension. However, the vascular adaptations in these mice have not been characterized. OBJECTIVE: To determine the histological and functional changes in the pulmonary and carotid arteries in CAV-1-/- induced PAH. METHODS: Pulmonary and carotid arteries of young (4-6 months old) and mature (9-12 months old) CAV-1-/- mice were tested and compared to normal wild type mice. RESULTS: Artery stiffness increases in CAV-1-/- mice, especially the circumferential stiffness of the pulmonary arteries. Increases in stiffness were quantified by a decrease in circumferential stretch and transition strain, increases in elastic moduli, and an increase in total strain energy at physiologic strains. Changes in mechanical properties for the pulmonary artery correlated with increased collagen content while carotid artery mechanical properties correlated with decreased elastin content. CONCLUSIONS: We demonstrated that an increase in artery stiffness is associated with CAV-1 deficiency-induced pulmonary hypertension. These results improve our understanding of artery remodeling in PAH.

[Three-month functional prognosis of patients hospitalised due to acute ischaemic stroke in Aragon: Rregional analysis of the impact of COVID-19]. / Pronóstico funcional a 3 meses de pacientes hospitalizados por ictus isquémico agudo en Aragón: análisis regional de la repercusión de la COVID-19.

Introduction: The COVID-19 pandemic has had an impact on ischaemic stroke management, with a reported decrease in hospital admissions, and even disruptions in healthcare and increased in-hospital mortality. However, there is a lack of evidence on the impact of the pandemic on functional prognosis. The aim of this study is to analyse the effect of the COVID-19 pandemic on the 3-month functional outcomes of patients hospitalised due to acute ischaemic stroke in Aragon (Spain). Materil and methods: We reviewed the data of all patients admitted due to ischaemic stroke to any hospital in our regional healthcare system between 30 December 2019 and 3 May 2020. We compared modified Rankin Scale scores and mortality at 3 months in patients hospitalised before and after the declaration of a state of emergency due to the COVID-19 pandemic. Results: In total, 318 patients with acute ischaemic stroke met our inclusion criteria. No differences were observed between periods in global or specific characteristics, with the exception of a higher proportion of patients older than 80 years during the first period (42.2% vs. 29.0%, P=.028). In the comparative analysis, we found no significant differences in mortality (12.3 vs. 7.9, P=.465) or in the proportion of patients with modified Rankin Scale scores ≤ 2 (57.7% vs. 57.1%, P=.425) at 3 months. Conclusion: To our knowledge, this is the first study to analyse the impact of COVID-19 pandemic on the 3-month functional outcomes of patients with ischaemic stroke. In our region, there has been no increase in rates of mortality or disability at 3 months in patients admitted due to ischaemic stroke during the pandemic.

Ccm1p is a 15S rRNA primary transcript processing factor as elucidated by a novel in vivo system in Saccharomyces cerevisiae.

In Saccharomyces cerevisiae, the mitoribosomal RNA of the minor subunit, 15S rRNA, is transcribed as a bicistronic transcript along with tRNAW. 5' and 3' sequences flanking the mature transcript must be removed by cleavage at the respective junctions before incorporating it into the mitoribosome. An in vivo dose-response triphasic system was created to elucidate the role of Ccm1p in the processing of 15S rRNA: Ccm1p supply ("On"), deprivation ("Off"), and resupply ("Back on"). After 72 h under "Off" status, the cells started to exhibit a complete mutant phenotype as assessed by their lack of growth in glycerol medium, while keeping their mitochondrial DNA integrity (ρ+). Full functionality of mitochondria was reacquired upon "Back on." 15S rRNA levels and phenotype followed the Ccm1p intramitochondrial concentrations throughout the "On-Off-Back on" course. Under "Off" status, cells gradually accumulated unprocessed 5' and 3' junctions, which reached significant levels at 72-96 h, probably due to a saturation of the mitochondrial degradosome (mtEXO). The Ccm1p/mtEXO mutant (Δccm1/Δdss1) showed a copious accumulation of 15S rRNA primary transcript forms, which were cleaved upon Ccm1p resupply. The gene that codes for the RNA component of RNase P was conserved in wild-type and mutant strains. Our results indicate that Ccm1p is crucial in processing the 15S rRNA primary transcript and does not stabilize the already mature 15S rRNA. Consequently, failure of this function in Δccm1 cells results, as it happens to any other unprocessed primary transcripts, in total degradation of 15S rRNA by mtEXO, whose mechanism of action is discussed.

Nicotine modulates molecules of the innate immune response in epithelial cells and macrophages during infection with M. tuberculosis.

Smoking increases susceptibility to becoming infected with and developing tuberculosis. Among the components of cigarette smoke, nicotine has been identified as the main immunomodulatory molecule; however, its effect on the innate immune system is unknown. In the present study, the effect of nicotine on molecules of the innate immune system was evaluated. Lung epithelial cells and macrophages were infected with Mycobacterium tuberculosis (Mtb) and/or treated with nicotine. The results show that nicotine alone decreases the expression of the Toll-like receptors (TLR)-2, TLR-4 and NOD-2 in all three cell types, as well as the production of the SP-D surfactant protein in type II pneumocytes. Moreover, it was observed that nicotine decreases the production of interleukin (IL)-6 and C-C chemokine ligand (CCL)5 during Mtb infection in epithelial cells (EpCs), whereas in macrophages derived from human monocytes (MDMs) there is a decrease in IL-8, IL-6, tumor necrosis factor (TNF)-α, IL-10, CCL2, C-X-C chemokine ligand (CXCL)9 and CXCL10 only during infection with Mtb. Although modulation of the expression of cytokines and chemokines appears to be partially mediated by the nicotinic acetylcholine receptor α7, blocking this receptor found no effect on the expression of receptors and SP-D. In summary, it was found that nicotine modulates the expression of innate immunity molecules necessary for the defense against tuberculosis.