Vertebrate centromeres in mitosis are functionally bipartite structures stabilized by cohesin.

Centromeres are scaffolds for the assembly of kinetochores that ensure chromosome segregation during cell division. How vertebrate centromeres obtain a three-dimensional structure to accomplish their primary function is unclear. Using super-resolution imaging, capture-C, and polymer modeling, we show that vertebrate centromeres are partitioned by condensins into two subdomains during mitosis. The bipartite structure is found in human, mouse, and chicken cells and is therefore a fundamental feature of vertebrate centromeres. Super-resolution imaging and electron tomography reveal that bipartite centromeres assemble bipartite kinetochores, with each subdomain binding a distinct microtubule bundle. Cohesin links the centromere subdomains, limiting their separation in response to spindle forces and avoiding merotelic kinetochore-spindle attachments. Lagging chromosomes during cancer cell divisions frequently have merotelic attachments in which the centromere subdomains are separated and bioriented. Our work reveals a fundamental aspect of vertebrate centromere biology with implications for understanding the mechanisms that guarantee faithful chromosome segregation.

cGAS-STING drives the IL-6-dependent survival of chromosomally instable cancers.

Chromosomal instability (CIN) drives cancer cell evolution, metastasis and therapy resistance, and is associated with poor prognosis1. CIN leads to micronuclei that release DNA into the cytoplasm after rupture, which triggers activation of inflammatory signalling mediated by cGAS and STING2,3. These two proteins are considered to be tumour suppressors as they promote apoptosis and immunosurveillance. However, cGAS and STING are rarely inactivated in cancer4, and, although they have been implicated in metastasis5, it is not known why loss-of-function mutations do not arise in primary tumours4. Here we show that inactivation of cGAS-STING signalling selectively impairs the survival of triple-negative breast cancer cells that display CIN. CIN triggers IL-6-STAT3-mediated signalling, which depends on the cGAS-STING pathway and the non-canonical NF-κB pathway. Blockade of IL-6 signalling by tocilizumab, a clinically used drug that targets the IL-6 receptor (IL-6R), selectively impairs the growth of cultured triple-negative breast cancer cells that exhibit CIN. Moreover, outgrowth of chromosomally instable tumours is significantly delayed compared with tumours that do not display CIN. Notably, this targetable vulnerability is conserved across cancer types that express high levels of IL-6 and/or IL-6R in vitro and in vivo. Together, our work demonstrates pro-tumorigenic traits of cGAS-STING signalling and explains why the cGAS-STING pathway is rarely inactivated in primary tumours. Repurposing tocilizumab could be a strategy to treat cancers with CIN that overexpress IL-6R.

Targeting TRIM37-driven centrosome dysfunction in 17q23-amplified breast cancer.

Genomic instability is a hallmark of cancer, and has a central role in the initiation and development of breast cancer1,2. The success of poly-ADP ribose polymerase inhibitors in the treatment of breast cancers that are deficient in homologous recombination exemplifies the utility of synthetically lethal genetic interactions in the treatment of breast cancers that are driven by genomic instability3. Given that defects in homologous recombination are present in only a subset of breast cancers, there is a need to identify additional driver mechanisms for genomic instability and targeted strategies to exploit these defects in the treatment of cancer. Here we show that centrosome depletion induces synthetic lethality in cancer cells that contain the 17q23 amplicon, a recurrent copy number aberration that defines about 9% of all primary breast cancer tumours and is associated with high levels of genomic instability4-6. Specifically, inhibition of polo-like kinase 4 (PLK4) using small molecules leads to centrosome depletion, which triggers mitotic catastrophe in cells that exhibit amplicon-directed overexpression of TRIM37. To explain this effect, we identify TRIM37 as a negative regulator of centrosomal pericentriolar material. In 17q23-amplified cells that lack centrosomes, increased levels of TRIM37 block the formation of foci that comprise pericentriolar material-these foci are structures with a microtubule-nucleating capacity that are required for successful cell division in the absence of centrosomes. Finally, we find that the overexpression of TRIM37 causes genomic instability by delaying centrosome maturation and separation at mitotic entry, and thereby increases the frequency of mitotic errors. Collectively, these findings highlight TRIM37-dependent genomic instability as a putative driver event in 17q23-amplified breast cancer and provide a rationale for the use of centrosome-targeting therapeutic agents in treating these cancers.

Vertebrate centromere architecture: from chromatin threads to functional structures.

Centromeres are chromatin structures specialized in sister chromatid cohesion, kinetochore assembly, and microtubule attachment during chromosome segregation. The regional centromere of vertebrates consists of long regions of highly repetitive sequences occupied by the Histone H3 variant CENP-A, and which are flanked by pericentromeres. The three-dimensional organization of centromeric chromatin is paramount for its functionality and its ability to withstand spindle forces. Alongside CENP-A, key contributors to the folding of this structure include components of the Constitutive Centromere-Associated Network (CCAN), the protein CENP-B, and condensin and cohesin complexes. Despite its importance, the intricate architecture of the regional centromere of vertebrates remains largely unknown. Recent advancements in long-read sequencing, super-resolution and cryo-electron microscopy, and chromosome conformation capture techniques have significantly improved our understanding of this structure at various levels, from the linear arrangement of centromeric sequences and their epigenetic landscape to their higher-order compaction. In this review, we discuss the latest insights on centromere organization and place them in the context of recent findings describing a bipartite higher-order organization of the centromere.

Convergence of flow cytometry and bacteriology. Current and future applications: a focus on food and clinical microbiology.

Since its development in the 1960s, flow cytometry (FCM) was quickly revealed a powerful tool to analyse cell populations in medical studies, yet, for many years, was almost exclusively used to analyse eukaryotic cells. Instrument and methodological limitations to distinguish genuine bacterial signals from the background, among other limitations, have hampered FCM applications in bacteriology. In recent years, thanks to the continuous development of FCM instruments and methods with a higher discriminatory capacity to detect low-size particles, FCM has emerged as an appealing technique to advance the study of microbes, with important applications in research, clinical and industrial settings. The capacity to rapidly enumerate and classify individual bacterial cells based on viability facilitates the monitoring of bacterial presence in foodstuffs or clinical samples, reducing the time needed to detect contamination or infectious processes. Besides, FCM has stood out as a valuable tool to advance the study of complex microbial communities, or microbiomes, that are very relevant in the context of human health, as well as to understand the interaction of bacterial and host cells. This review highlights current developments in, and future applications of, FCM in bacteriology, with a focus on those related to food and clinical microbiology.

Human milk immune factors, maternal nutritional status, and infant sex: The INSPIRE study.

OBJECTIVES: Breastfeeding is an energetically costly and intense form of human parental investment, providing sole-source nutrition in early infancy and bioactive components, including immune factors. Given the energetic cost of lactation, milk factors may be subject to tradeoffs, and variation in concentrations have been explored utilizing the Trivers-Willard hypothesis. As human milk immune factors are critical to developing immune system and protect infants against pathogens, we tested whether concentrations of milk immune factors (IgA, IgM, IgG, EGF, TGFß2, and IL-10) vary in response to infant sex and maternal condition (proxied by maternal diet diversity [DD] and body mass index [BMI]) as posited in the Trivers-Willard hypothesis and consider the application of the hypothesis to milk composition. METHODS: We analyzed concentrations of immune factors in 358 milk samples collected from women residing in 10 international sites using linear mixed-effects models to test for an interaction between maternal condition, including population as a random effect and infant age and maternal age as fixed effects. RESULTS: IgG concentrations were significantly lower in milk produced by women consuming diets with low diversity with male infants than those with female infants. No other significant associations were identified. CONCLUSIONS: IgG concentrations were related to infant sex and maternal diet diversity, providing minimal support for the hypothesis. Given the lack of associations across other select immune factors, results suggest that the Trivers-Willard hypothesis may not be broadly applied to human milk immune factors as a measure of maternal investment, which are likely buffered against perturbations in maternal condition.

Functional bacterial cultures for dairy applications: Towards improving safety, quality, nutritional and health benefit aspects.

Traditionally, fermentation was used to preserve the shelf life of food. Currently, in addition to favouring food preservation, well standardized and controlled industrial processes are also aimed at improving the functional characteristics of the final product. In this regard, starter cultures have become an essential cornerstone of food production. The selection of robust microorganisms, well adapted to the food environment, has been followed by the development of microbial consortia that provide some functional characteristics, beyond their acidifying capacity, achieving safer, high-quality foods with improved nutritional and health-promoting properties. In addition to starters, adjunct cultures and probiotics, which normally do not have a relevant role in fermentation, are added to the food in order to provide some beneficial characteristics. This review focuses on highlighting the functional characteristics of food starters, as well as adjunct and probiotic cultures (mainly lactic acid bacteria and bifidobacteria), with a specific focus on the synthesis of metabolites for preservation and safety aspects (e.g. bacteriocins), organoleptic properties (e.g. exopolysaccharides), nutritional (e.g. vitamins) and health improvement (e.g. neuroactive molecules). Literature reporting the application of these functional cultures in the manufacture of foods, mainly those related to dairy production, such as cheeses and fermented milks, has also been updated.

Arabinoxylan and Pectin Metabolism in Crohn's Disease Microbiota: An In Silico Study.

Inflammatory bowel disease is a chronic disorder including ulcerative colitis and Crohn's disease (CD). Gut dysbiosis is often associated with CD, and metagenomics allows a better understanding of the microbial communities involved. The objective of this study was to reconstruct in silico carbohydrate metabolic capabilities from metagenome-assembled genomes (MAGs) obtained from healthy and CD individuals. This computational method was developed as a mean to aid rationally designed prebiotic interventions to rebalance CD dysbiosis, with a focus on metabolism of emergent prebiotics derived from arabinoxylan and pectin. Up to 1196 and 1577 MAGs were recovered from CD and healthy people, respectively. MAGs of Akkermansia muciniphila, Barnesiella viscericola DSM 18177 and Paraprevotella xylaniphila YIT 11841 showed a wide range of unique and specific enzymes acting on arabinoxylan and pectin. These glycosidases were also found in MAGs recovered from CD patients. Interestingly, these arabinoxylan and pectin degraders are predicted to exhibit metabolic interactions with other gut microbes reduced in CD. Thus, administration of arabinoxylan and pectin may ameliorate dysbiosis in CD by promoting species with key metabolic functions, capable of cross-feeding other beneficial species. These computational methods may be of special interest for the rational design of prebiotic ingredients targeting at CD.

Genetic insights into the dark matter of the mammalian gut microbiota through targeted genome reconstruction.

Whole metagenomic shotgun (WMS) sequencing has dramatically enhanced our ability to study microbial genomics. The possibility to unveil the genetic makeup of bacteria that cannot be easily isolated has significantly expanded our microbiological horizon. Here, we report an approach aimed at uncovering novel bacterial species by the use of targeted WMS sequencing. Employing in silico data retrieved from metabolic modelling to formulate a chemically defined medium (CDM), we were able to isolate and subsequently sequence the genomes of six putative novel species of bacteria from the gut of non-human primates.

Novel methods of microbiome analysis in the food industry.

The study of the food microbiome has gained considerable interest in recent years, mainly due to the wide range of applications that can be derived from the analysis of metagenomes. Among these applications, it is worth mentioning the possibility of using metagenomic analyses to determine food authenticity, to assess the microbiological safety of foods thanks to the detection and tracking of pathogens, antibiotic resistance genes and other undesirable traits, as well to identify the microorganisms responsible for food processing defects. Metataxonomics and metagenomics are currently the gold standard methodologies to explore the full potential of metagenomes in the food industry. However, there are still a number of challenges that must be solved in order to implement these methods routinely in food chain monitoring, and for the regulatory agencies to take them into account in their opinions. These challenges include the difficulties of analysing foods and food-related environments with a low microbial load, the lack of validated bioinformatics pipelines adapted to food microbiomes and the difficulty of assessing the viability of the detected microorganisms. This review summarizes the methods of microbiome analysis that have been used, so far, in foods and food-related environments, with a specific focus on those involving Next-Generation Sequencing technologies.

Synthesis of fucosylated oligosaccharides with α-L-fucosidase from Thermotoga maritima immobilized on Eupergit® CM.

Fucosylated oligosaccharides present in human milk perform various biological functions that benefit infants' health. These compounds can be also obtained by enzymatic synthesis. In this work, the effect of the immobilization of α-L-fucosidase from Thermotoga maritima on the synthesis of fucosylated oligosaccharides was studied, using lactose and 4-nitrophenyl-α-L-fucopyranoside (pNP-Fuc) as acceptor and donor substrates, respectively, and Eupergit® CM as an immobilization support. The enzyme was immobilized with 90% efficiency at pH 8 and ionic strength of 1.5 M. Immobilization decreased enzyme affinity for the donor substrate as shown by a 1.5-times higher KM value and a 22-times decrease of the kcat/KM ratio in comparison to the unbound enzyme. In contrast, no effect was observed on the synthesis/hydrolysis ratio (rs/rh) when α-L-fucosidase was immobilized. Also, the effect of initial concentration of substrates was studied. An increase of the acceptor concentration improved the yields of fucosylated oligosaccharides regardless enzyme immobilization. The synthesis yields of 38.9 and 40.6% were obtained using Eupergit® CM-bound or unbound enzyme, respectively, and 3.5 mM pNP-Fuc and 146 mM lactose. In conclusion, α-L-fucosidase from Thermotoga maritima was efficiently immobilized on Eupergit® CM support without affecting the synthesis of fucosylated oligosaccharides.

Evolutionary development and co-phylogeny of primate-associated bifidobacteria.

In recent years, bifidobacterial populations in the gut of various monkey species have been assessed in several ecological surveys, unveiling a diverse, yet unexplored ecosystem harbouring novel species. In the current study, we investigated the species distribution of bifidobacteria present in 23 different species of primates, including human samples, by means of 16S rRNA microbial profiling and internal transcribed spacer bifidobacterial profiling. Based on the observed bifidobacterial-host co-phylogeny, we found a statistically significant correlation between the Hominidae family and particular bifidobacterial species isolated from humans, indicating phylosymbiosis between these lineages. Furthermore, phylogenetic and glycobiome analyses, based on 40 bifidobacterial species isolated from primates, revealed that members of the Bifidobacterium tissieri phylogenetic group, which are typical gut inhabitants of members of the Cebidae family, descend from an ancient ancestor with respect to other bifidobacterial taxa isolated from primates.

Oleanolic acid ameliorates intestinal alterations associated with EAE.

BACKGROUND: Multiple sclerosis (MS) is a chronic demyelinating autoimmune disease affecting the CNS. Recent studies have indicated that intestinal alterations play key pathogenic roles in the development of autoimmune diseases, including MS. The triterpene oleanolic acid (OA), due to its anti-inflammatory properties, has shown to beneficially influence the severity of the experimental autoimmune encephalomyelitis (EAE), a preclinical model of MS. We herein investigate EAE-associated gut intestinal dysfunction and the effect of OA treatment. METHODS: Mice with MOG35-55-induced EAE were treated with OA or vehicle from immunization day and were daily analyzed for clinical deficit. We performed molecular and histological analysis in serum and intestinal tissues to measure oxidative and inflammatory responses. We used Caco-2 and HT29-MTX-E12 cells to elucidate OA in vitro effects. RESULTS: We found that OA protected from EAE-induced changes in intestinal permeability and preserved the mucin-containing goblet cells along the intestinal tract. Serum levels of the markers for intestinal barrier damage iFABP and monocyte activation sCD14 were consistently and significantly reduced in OA-treated EAE mice. Beneficial OA effects also included a decrease of pro-inflammatory mediators both in serum and colonic tissue of treated-EAE mice. Moreover, the levels of some immunoregulatory cytokines, the neurotrophic factor GDNF, and the gastrointestinal hormone motilin were preserved in OA-treated EAE mice. Regarding oxidative stress, OA treatment prevented lipid peroxidation and superoxide anion accumulation in intestinal tissue, while inducing the expression of the ROS scavenger Sestrin-3. Furthermore, short-chain fatty acids (SCFA) quantification in the cecal content showed that OA reduced the high iso-valeric acid concentrations detected in EAE-mice. Lastly, using in vitro cell models which mimic the intestinal epithelium, we verified that OA protected against intestinal barrier dysfunction induced by injurious agents produced in both EAE and MS. CONCLUSION: These findings reveal that OA ameliorates the gut dysfunction found in EAE mice. OA normalizes the levels of gut mucosal dysfunction markers, as well as the pro- and anti-inflammatory immune bias during EAE, thus reinforcing the idea that OA is a beneficial compound for treating EAE and suggesting that OA may be an interesting candidate to be explored for the treatment of human MS.

Selection for Antimicrobial Resistance in Foodborne Pathogens through Exposure to UV Light and Nonthermal Atmospheric Plasma Decontamination Techniques.

This study was aimed at assessing whether the repeated exposure of 12 strains of Salmonella spp., Escherichia coli, and Listeria monocytogenes to alternative nonthermal decontamination techniques with UV light (UV-C) and nonthermal atmospheric plasma (NTAP) may cause the emergence of variants showing increased resistance to clinically relevant antibiotics (ampicillin, cefotaxime, ciprofloxacin, gentamicin, streptomycin, tetracycline, erythromycin, vancomycin, and colistin). UV-C and NTAP treatments were applied on the surface of inoculated brain heart infusion (BHI) agar plates. Survivors were recovered and after 24 h of growth in BHI broth were again subjected to the decontamination treatment; this was repeated for 10 consecutive cycles. A total of 174 strain/decontamination technique/antibiotic combinations were tested, and 12 variant strains with increased resistance to one of the antibiotics studied were identified, with the increases in the MICs in Mueller-Hinton broth ranging from 2- to 256-fold. The variant strains of Salmonella spp. isolated were further characterized through phenotypic screenings and whole-genome sequencing (WGS) analyses. Most changes in susceptibility were observed for antibiotics that act at the level of protein synthesis (aminoglycosides, tetracyclines, and glycylcyclines) or DNA replication (fluoroquinolones), as well as for polymyxins. No changes in resistance to ß-lactams were detected. WGS analyses showed the occurrence of sequence alterations in some antibiotic cellular targets (e.g., gyrA for ciprofloxacin-resistant variants, rpsL for a streptomycin-resistant variant), accompanied by variations in stress response regulators and membrane transporters likely involved in the nonselective efflux of antibiotics, which altogether resulted in a low- to medium-level increase in microbial resistance to several antibiotics.IMPORTANCE The emergence and spread of antibiotic resistance along the food chain can be influenced by the different antimicrobial strategies used from farm to fork. This study evidences that two novel, not yet widely used, nonthermal microbial decontamination techniques, UV light and nonthermal atmospheric plasma, can select variants with increased resistance to various clinically relevant antibiotics, such as ciprofloxacin, streptomycin, tetracycline, and erythromycin. Whole-genome analysis of the resistant variants obtained for Salmonella spp. allowed identification of the genetic changes responsible for the observed phenotypes and suggested that some antimicrobial classes are more susceptible to the cross-resistance phenomena observed. This information is relevant, since these novel decontamination techniques are being proposed as possible alternative green techniques for the decontamination of environments and equipment in food and clinical settings.

Meaning of empowerment in peritoneal dialysis: focus groups with patients and caregivers.

BACKGROUND: While peritoneal dialysis (PD) can offer patients more independence and flexibility compared with in-center hemodialysis, managing the ongoing and technically demanding regimen can impose a burden on patients and caregivers. Patient empowerment can strengthen capacity for self-management and improve treatment outcomes. We aimed to describe patients' and caregivers' perspectives on the meaning and role of patient empowerment in PD. METHODS: Adult patients receiving PD (n = 81) and their caregivers (n = 45), purposively sampled from nine dialysis units in Australia, Hong Kong and the USA, participated in 14 focus groups. Transcripts were thematically analyzed. RESULTS: We identified six themes: lacking clarity for self-management (limited understanding of rationale behind necessary restrictions, muddled by conflicting information); PD regimen restricting flexibility and freedom (burden in budgeting time, confined to be close to home); strength with supportive relationships (gaining reassurance with practical assistance, comforted by considerate health professionals, supported by family and friends); defying constraints (reclaiming the day, undeterred by treatment, refusing to be defined by illness); regaining lost vitality (enabling physical functioning, restoring energy for life participation); and personal growth through adjustment (building resilience and enabling positive outlook, accepting the dialysis regimen). CONCLUSIONS: Understanding the rationale behind lifestyle restrictions, practical assistance and family support in managing PD promoted patient empowerment, whereas being constrained in time and capacity for life participation outside the home undermined it. Education, counseling and strategies to minimize the disruption and burden of PD may enhance satisfaction and outcomes in patients requiring PD.

Employment of fucosidases for the synthesis of fucosylated oligosaccharides with biological potential.

Fucosylated oligosaccharides play important physiological roles in humans, including in the immune response, transduction of signals, early embryogenesis and development, growth regulation, apoptosis, pathogen adhesion, and so on. Efforts have been made to synthesize fucosylated oligosaccharides, as it is difficult to purify them from their natural sources, such as human milk, epithelial tissue, blood, and so on. Within the strategies for its in vitro synthesis, it is remarkable the employment of fucosidases, enzymes that normally cleave the fucosyl residue from the non-reducing end of fucosylated compounds, as these enzymes are also capable of synthesizing them by means of a transfucosylation reaction. This review summarizes the progress in the use of fucosidases for the synthesis of compounds that have potential for industrial and commercial applications.

Household composition and the infant fecal microbiome: The INSPIRE study.

OBJECTIVES: Establishment and development of the infant gastrointestinal microbiome (GIM) varies cross-culturally and is thought to be influenced by factors such as gestational age, birth mode, diet, and antibiotic exposure. However, there is little data as to how the composition of infants' households may play a role, particularly from a cross-cultural perspective. Here, we examined relationships between infant fecal microbiome (IFM) diversity/composition and infants' household size, number of siblings, and number of other household members. MATERIALS AND METHODS: We analyzed 377 fecal samples from healthy, breastfeeding infants across 11 sites in eight different countries (Ethiopia, The Gambia, Ghana, Kenya, Peru, Spain, Sweden, and the United States). Fecal microbial community structure was determined by amplifying, sequencing, and classifying (to the genus level) the V1-V3 region of the bacterial 16S rRNA gene. Surveys administered to infants' mothers identified household members and composition. RESULTS: Our results indicated that household composition (represented by the number of cohabitating siblings and other household members) did not have a measurable impact on the bacterial diversity, evenness, or richness of the IFM. However, we observed that variation in household composition categories did correspond to differential relative abundances of specific taxa, namely: Lactobacillus, Clostridium, Enterobacter, and Klebsiella. DISCUSSION: This study, to our knowledge, is the largest cross-cultural study to date examining the association between household composition and the IFM. Our results indicate that the social environment of infants (represented here by the proxy of household composition) may influence the bacterial composition of the infant GIM, although the mechanism is unknown. A higher number and diversity of cohabitants and potential caregivers may facilitate social transmission of beneficial bacteria to the infant gastrointestinal tract, by way of shared environment or through direct physical and social contact between the maternal-infant dyad and other household members. These findings contribute to the discussion concerning ways by which infants are influenced by their social environments and add further dimensionality to the ongoing exploration of social transmission of gut microbiota and the "old friends" hypothesis.

Synthesis of Fucosyl-Oligosaccharides Using α-l-Fucosidase from Lactobacillus rhamnosus GG.

Fucosyl-oligosaccharides are natural prebiotics that promote the growth of probiotics in human gut and stimulate the innate immune system. In this work, the release of α-lfucosidase by Lactobacillus rhamnosus GG, and the use of this enzyme for the synthesis of fucosyl-oligosaccharides were investigated. Since α-lfucosidase is a membrane-bound enzyme, its release from the cells was induced by addition of 4-nitrophenyl-α-l-fucopyranoside (pNP-Fuc). Enzyme activity associated with the cell was recovered at 78% of its total activity. Fucosyl-oligosaccharides where synthesized using α-l-fucosidase extract and pNP-Fuc as donor substrate, and D-lactose or D-lactulose as acceptor substrates, reaching a yield up to 25%. Fucosyllactose was obtained as a reaction product with D-lactose, and its composition was confirmed by mass spectrometry (MALDI-TOF MS). It is possible that the fucosyl-oligosaccharide synthesized in this study has biological functions similar to human milk oligosaccharides.

Improvement of the transfucosylation activity of α-L-fucosidase from Thermotoga maritima for the synthesis of fucosylated oligosaccharides in the presence of calcium and sodium.

The influence of CaCl2 and NaCl in the hydrolytic activity and the influence of CaCl2 in the synthesis of fucosylated oligosaccharides using α-L-fucosidase from Thermotoga maritima were evaluated. The hydrolytic activity of α-L-fucosidase from Thermotoga maritima displayed a maximum increase of 67% in the presence of 0.8 M NaCl with water activity (aw) of 0.9672 and of 138% in the presence of 1.1 M CaCl2 (aw 0.9581). In addition, the hydrolytic activity was higher when using CaCl2 compared to NaCl at aw of 0.8956, 0.9581 and 0.9672. On the other hand, the effect of CaCl2 in the synthesis of fucosylated oligosaccharides using 4-nitrophenyl-fucose as donor substrate and lactose as acceptor was studied. In these reactions, the presence of 1.1 M CaCl2 favored the rate of transfucosylation, and improved the yield of synthesis duplicating and triplicating it with lactose concentrations of 58 and 146 mM, respectively. CaCl2 did not significatively affect hydrolysis rate in these reactions. The combination of the activating effect of CaCl2, the decrement in aw and lactose concentration had a synergistic effect favoring the synthesis of fucosylated oligosaccharides.

Deletion of Fstl1 (Follistatin-Like 1) From the Endocardial/Endothelial Lineage Causes Mitral Valve Disease.

OBJECTIVE: Fstl1 (Follistatin-like 1) is a secreted protein that is expressed in the atrioventricular valves throughout embryonic development, postnatal maturation, and adulthood. In this study, we investigated the loss of Fstl1 in the endocardium/endothelium and their derived cells. APPROACH AND RESULTS: We conditionally ablated Fstl1 from the endocardial lineage using a transgenic Tie2-Cre mouse model. These mice showed a sustained Bmp and Tgfß signaling after birth. This resulted in ongoing proliferation and endocardial-to-mesenchymal transition and ultimately in deformed nonfunctional mitral valves and a hypertrophic dilated heart. Echocardiographic and electrocardiographic analyses revealed that loss of Fstl1 leads to mitral regurgitation and left ventricular diastolic dysfunction. Cardiac function gradually deteriorated resulting in heart failure with preserved ejection fraction and death of the mice between 2 and 4 weeks after birth. CONCLUSIONS: We report on a mouse model in which deletion of Fstl1 from the endocardial/endothelial lineage results in deformed mitral valves, which cause regurgitation, heart failure, and early cardiac death. The findings provide a potential molecular target for the clinical research into myxomatous mitral valve disease.