Robust designation of meiotic crossover sites by CDK-2 through phosphorylation of the MutSγ complex.

Crossover formation is essential for proper segregation of homologous chromosomes during meiosis. Here, we show that Caenorhabditis elegans cyclin-dependent kinase 2 (CDK-2) partners with cyclin-like protein COSA-1 to promote crossover formation by promoting conversion of meiotic double-strand breaks into crossover­specific recombination intermediates. Further, we identify MutSγ component MSH-5 as a CDK-2 phosphorylation target. MSH-5 has a disordered C-terminal tail that contains 13 potential CDK phosphosites and is required to concentrate crossover­promoting proteins at recombination sites. Phosphorylation of the MSH-5 tail appears dispensable in a wild-type background, but when MutSγ activity is partially compromised, crossover formation and retention of COSA-1 at recombination sites are exquisitely sensitive to phosphosite loss. Our data support a model in which robustness of crossover designation reflects a positive feedback mechanism involving CDK-2­mediated phosphorylation and scaffold-like properties of the MSH5 C-terminal tail, features that combine to promote full recruitment and activity of crossover­promoting complexes.

GSK-3 promotes S-phase entry and progression in C. elegans germline stem cells to maintain tissue output.

Adult C. elegans germline stem cells (GSCs) and mouse embryonic stem cells (mESCs) exhibit a non-canonical cell cycle structure with an abbreviated G1 phase and phase-independent expression of Cdk2 and cyclin E. Mechanisms that promote the abbreviated cell cycle remain unknown, as do the consequences of not maintaining an abbreviated cell cycle in these tissues. In GSCs, we discovered that loss of gsk-3 results in reduced GSC proliferation without changes in differentiation or responsiveness to GLP-1/Notch signaling. We find that DPL-1 transcriptional activity inhibits CDK-2 mRNA accumulation in GSCs, which leads to slower S-phase entry and progression. Inhibition of dpl-1 or transgenic expression of CDK-2 via a heterologous germline promoter rescues the S-phase entry and progression defects of the gsk-3 mutants, demonstrating that transcriptional regulation rather than post-translational control of CDK-2 establishes the abbreviated cell cycle structure in GSCs. This highlights an inhibitory cascade wherein GSK-3 inhibits DPL-1 and DPL-1 inhibits cdk-2 transcription. Constitutive GSK-3 activity through this cascade maintains an abbreviated cell cycle structure to permit the efficient proliferation of GSCs necessary for continuous tissue output.

Pachytene piRNAs control discrete meiotic events during spermatogenesis and restrict gene expression in space and time.

Pachytene piRNAs, a Piwi-interacting RNA subclass in mammals, are hypothesized to regulate non-transposon sequences during spermatogenesis. Caenorhabditis elegans piRNAs, the 21URNAs, are implicated in regulating coding sequences; the messenger RNA targets and biological processes they control during spermatogenesis are largely unknown. We demonstrate that loss of 21URNAs compromises homolog pairing and makes it permissive for nonhomologous synapsis resulting in defects in crossover formation and chromosome segregation during spermatogenesis. We identify Polo-like kinase 3 (PLK-3), among others, as a 21URNA target. 21URNA activity restricts PLK-3 protein to proliferative cells, and expansion of PLK-3 in pachytene overlaps with the meiotic defects. Removal of plk-3 results in quantitative genetic suppression of the meiotic defects. One discrete 21URNA inhibits PLK-3 expression in late pachytene cells. Together, these results suggest that the 21URNAs function as pachytene piRNAs during C. elegans spermatogenesis. We identify their targets and meiotic events and highlight the remarkable intricacy of this multi-effector mechanism during spermatogenesis.

Antibacterial and anti-inflammatory activities of mycelia of a medicinal mushroom from Taiwan, Taiwanofungus salmoneus (higher Basidiomycetes).

Taiwanofungus salmoneus (T.T. Chang et W.N. Chou) Sheng H. Wu et al. (shiang-shan-chih), is a medicinal fungus indigenous to Taiwan. The mycelium was prepared from submerged culture and its ethanolic and hot-water extracts were used to study its antibacterial and anti-inflammatory activities. Gram-positive species (Bacillus cereus, Listeria monocytogenes, and Staphylococcus aureus) and gram-negative species (Escherichia coli and Salmonella typhimurium) of bacteria were used. In addition to the inhibitory zone, minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) also were determined. The ethanolic extracts showed higher inhibitory and bactericidal activities (MIC and MBC: 6.25-12.50 mg/ml) than the hot-water extracts (MIC and MBC: 25-50 mg/mL). In the anti-inflammatory test, the extracts were tested on lipopolysaccharide-induced nitric oxide and tumor necrosis factor- α production in RAW 264.7 cells. The values of the inhibition concentration at 50% of nitric oxide production were 18.2 and 14.2 µg/mL for the hot-water and ethanolic extracts, respectively. The 50% inhibitory concentration values of tumor necrosis factor- α production were 4.99 and 7.13 µg/mL for the hot-water and ethanolic extracts, respectively. On the basis of the results obtained, the mycelia could be used as an antibacterial and anti-inflammatory supplement.

Hydrophilic Metabolite Composition of Fruiting Bodies and Mycelia of Edible Mushroom Species (Agaricomycetes).

Mushrooms have two components, the fruiting body, which encompasses the stalk and the cap, and the mycelium, which supports the fruiting body underground. The part of the mushroom most commonly consumed is the fruiting body. Given that it is more time consuming to harvest the fruiting body versus simply the mycelia, we were interested in understanding the difference in metabolite content between the fruiting bodies and mycelia of four widely consumed mushrooms in Taiwan: Agrocybe cylindracea (AC), Coprinus comatus (CC), Hericium erinaceus (HE), and Hypsizygus marmoreus (HM). In total, we identified 54 polar metabolites using 1H NMR spectroscopy that included sugar alcohols, amino acids, organic acids, nucleosides and purine/pyrimidine derivatives, sugars, and others. Generally, the fruiting bodies of AC, CC, and HM contained higher amounts of essential amino acids than their corresponding mycelia. Among fruiting bodies, HE had the lowest essential amino acid content. Trehalose was the predominant carbohydrate in most samples except for the mycelia of AC, in which the major sugar was glucose. The amount of adenosine, uridine, and xanthine in the samples was similar, and was higher in fruiting bodies compared with mycelia, except for HM. The organic acid and sugar alcohol content between fruiting bodies and mycelia did not tend to be different. Although each mushroom had a unique metabolic profile, the metabolic profile of fruiting bodies and mycelia were most similar for CC and HE, suggesting that the mycelia of CC and HE may be good replacements for their corresponding fruiting bodies. Additionally, each mushroom species had a unique polar metabolite fingerprint, which could be utilized to identify adulteration.

Quality Evaluation of Shiitake Blanched and Centrifuged Broths as Functional Instant Drinks.

In the process of making mushrooms into vacuum-fried crisps, the resulting blanched broth (BB) and centrifuged broth (CB) are often discarded, thereby increasing the amount of wastewater and treatment costs. This study measured the proximate compositions, bioactive components, taste components, and minerals of freeze-dried BB and CB and then used functional indigestible dextrin (Fibersol-2) as a carrier to make these two broths into instant drinks. The solids of the BB and CB contained protein (16.88-19.21%), fat (0.01-0.23%), ash (12.89-13.50%), carbohydrate (67.28-70.00%), sugars and polyols (40.55-45.68%), free amino acids (6.58-6.69%), 5'-nucleotides (0.98-1.47%), and bioactive components, especially polysaccharides (4.53-7.45%), ergothioneine (both 0.19%), and total phenols (0.15-0.36%). The equivalent umami concentration of BB was 2.77-fold higher than that of the CB. Both BB and CB showed compositions and essential minerals that are rich in taste. Using a nine-point hedonic test, it was found that the solid contents of BB and CB in the instant drink affected the consumer's preference. The flavor and overall preference of instant drinks with 2.5% BB or CB were the best amongst consumers. Overall, the BB and CB were rich in nutrients and bioactive and taste components and could be developed as a functional food in the form of a drink.

Phosphorylated nuclear DICER1 promotes open chromatin state and lineage plasticity of AT2 tumor cells in lung adenocarcinomas.

KRAS/ERK pathway phosphorylates DICER1, causing its nuclear translocation, and phosphomimetic Dicer1 contributes to tumorigenesis in mice. Mechanisms through which phospho-DICER1 regulates tumor progression remain undefined. While DICER1 canonically regulates microRNAs (miRNA) and epithelial-to-mesenchymal transition (EMT), we found that phosphorylated nuclear DICER1 (phospho-nuclear DICER1) promotes late-stage tumor progression in mice with oncogenic Kras, independent of miRNAs and EMT. Instead, we observe that the murine AT2 tumor cells exhibit altered chromatin compaction, and cells from disorganized advanced tumors, but not localized tumors, express gastric genes. Collectively, this results in subpopulations of tumor cells transitioning from a restricted alveolar to a broader endodermal lineage state. In human LUADs, we observed expression of phospho-nuclear DICER1 in advanced tumors together with the expression of gastric genes. We define a multimeric chromatin-DICER1 complex composed of the Mediator complex subunit 12, CBX1, MACROH2A.1, and transcriptional regulators supporting the model that phospho-nuclear DICER1 leads to lineage reprogramming of AT2 tumor cells to mediate lung cancer progression.

Spatial single-cell sequencing of meiosis I arrested oocytes indicates acquisition of maternal transcripts from the soma.

Maternal RNAs are stored from minutes to decades in oocytes throughout meiosis I arrest in a transcriptionally quiescent state. Recent reports, however, propose a role for nascent transcription in arrested oocytes. Whether arrested oocytes launch nascent transcription in response to environmental or hormonal signals while maintaining the meiosis I arrest remains undetermined. We test this by integrating single-cell RNA sequencing, RNA velocity, and RNA fluorescence in situ hybridization on C. elegans meiosis I arrested oocytes. We identify transcripts that increase as the arrested meiosis I oocyte ages, but rule out extracellular signaling through ERK MAPK and nascent transcription as a mechanism for this increase. We report transcript acquisition from neighboring somatic cells as a mechanism of transcript increase during meiosis I arrest. These analyses provide a deeper view at single-cell resolution of the RNA landscape of a meiosis I arrested oocyte and as it prepares for oocyte maturation and fertilization.

The essential roles of FXR in diet and age influenced metabolic changes and liver disease development: a multi-omics study.

BACKGROUND: Aging and diet are risks for metabolic diseases. Bile acid receptor farnesoid X receptor (FXR) knockout (KO) mice develop metabolic liver diseases that progress into cancer as they age, which is accelerated by Western diet (WD) intake. The current study uncovers the molecular signatures for diet and age-linked metabolic liver disease development in an FXR-dependent manner. METHODS: Wild-type (WT) and FXR KO male mice, either on a healthy control diet (CD) or a WD, were euthanized at the ages of 5, 10, or 15 months. Hepatic transcriptomics, liver, serum, and urine metabolomics as well as microbiota were profiled. RESULTS: WD intake facilitated hepatic aging in WT mice. In an FXR-dependent manner, increased inflammation and reduced oxidative phosphorylation were the primary pathways affected by WD and aging. FXR has a role in modulating inflammation and B cell-mediated humoral immunity which was enhanced by aging. Moreover, FXR dictated neuron differentiation, muscle contraction, and cytoskeleton organization in addition to metabolism. There were 654 transcripts commonly altered by diets, ages, and FXR KO, and 76 of them were differentially expressed in human hepatocellular carcinoma (HCC) and healthy livers. Urine metabolites differentiated dietary effects in both genotypes, and serum metabolites clearly separated ages irrespective of diets. Aging and FXR KO commonly affected amino acid metabolism and TCA cycle. Moreover, FXR is essential for colonization of age-related gut microbes. Integrated analyses uncovered metabolites and bacteria linked with hepatic transcripts affected by WD intake, aging, and FXR KO as well as related to HCC patient survival. CONCLUSION: FXR is a target to prevent diet or age-associated metabolic disease. The uncovered metabolites and microbes can be diagnostic markers for metabolic disease.

Identification of protein domains on major pilin MrkA that affects the mechanical properties of Klebsiella pneumoniae type 3 fimbriae.

The Klebsiella pneumoniae type 3 fimbriae are mainly composed of MrkA pilins that assemble into a helixlike filament. This study determined the biomechanical properties of the fimbriae and analyzed 11 site-directed MrkA mutants to identify domains that are critical for the properties. Escherichia coli strains expressing type 3 fimbriae with an Ala substitution at either F34, V45, C87, G189, T196, or Y197 resulted in a significant reduction in biofilm formation. The E. coli strain expressing MrkAG189A remained capable of producing a normal number of fimbriae. Although F34A, V45A, T196A, and Y197A substitutions expressed on E. coli strains produced sparse quantities of fimbriae, no fimbriae were observed on the cells expressing MrkAC87A. Further investigations of the mechanical properties of the MrkAG189A fimbriae with optical tweezers revealed that, unlike the wild-type fimbriae, the uncoiling force for MrkAG189A fimbriae was not constant. The MrkAG189A fimbriae also exhibited a lower enthalpy in the differential scanning calorimetry analysis. Together, these findings indicate that the mutant fimbriae are less stable than the wild-type. This study has demonstrated that the C-terminal ß strands of MrkA are required for the assembly and structural stability of fimbriae.

Comparative study of contents of several bioactive components in fruiting bodies and mycelia of culinary-medicinal mushrooms.

Mushrooms have been consumed for thousands of years, and several bioactive components were found therein, including lovastatin, γ-aminobutyric acid (GABA) and ergothioneine. The study reported herein was to analyze these three bioactive components in 15 fruiting bodies and 9 mycelia of 19 species of mushrooms from genera Agaricus, Agrocybe, Auricularia, Boletus, Ganoderma, Hypsizygus, Inonotus, Lentinus, Morchella, Pleurotus, Tremella, Termitomyces, and Volvariella. The results show that Hypsizygus marmoreus contained the highest amount of lovastatin (628.05 mg/kg) in fruiting bodies and Morchella esculenta contained the highest amount (1438.42 mg/ kg) in mycelia. Agaricus brasiliensis contained the highest amount of GABA (1844.85 mg/kg) in fruiting bodies, and mycelia of Boletus edulis, Pleurotus citrinopileatus, and Termitomyces albuminosus contained extraordinarily higher amounts (1274.03, 1631.67, and 2560.00 mg/kg, respectively). Volvariella volvacea contained the highest amount of ergothioneine (537.27 mg/kg) in fruiting bodies and mycelia; Boletus edulis, Pleurotus ferulae, and P. salmoneostramineus contained relatively higher amount of ergothioneine too (258.03, 250.23, and 222.08 mg/kg, respectively). However, none of these components was detected in fruiting bodies of Inonotus obliquus. In conclusion, these three bioactive components were commonly found in most mushrooms, and these results might be related to their beneficial effects.

A Comparison of Serum and Plasma Blood Collection Tubes for the Integration of Epidemiological and Metabolomics Data.

Blood is a rich biological sample routinely collected in clinical and epidemiological studies. With advancements in high throughput -omics technology, such as metabolomics, epidemiology can now delve more deeply and comprehensively into biological mechanisms involved in the etiology of diseases. However, the impact of the blood collection tube matrix of samples collected needs to be carefully considered to obtain meaningful biological interpretations and understand how the metabolite signatures are affected by different tube types. In the present study, we investigated whether the metabolic profile of blood collected as serum differed from samples collected as ACD plasma, citrate plasma, EDTA plasma, fluoride plasma, or heparin plasma. We identified and quantified 50 metabolites present in all samples utilizing nuclear magnetic resonance (NMR) spectroscopy. The heparin plasma tubes performed the closest to serum, with only three metabolites showing significant differences, followed by EDTA which significantly differed for five metabolites, and fluoride tubes which differed in eleven of the fifty metabolites. Most of these metabolite differences were due to higher levels of amino acids in serum compared to heparin plasma, EDTA plasma, and fluoride plasma. In contrast, metabolite measurements from ACD and citrate plasma differed significantly for approximately half of the metabolites assessed. These metabolite differences in ACD and citrate plasma were largely due to significant interfering peaks from the anticoagulants themselves. Blood is one of the most banked samples and thus mining and comparing samples between studies requires understanding how the metabolite signature is affected by the different media and different tube types.

Elucidation of two new corticosteroids, betamethasone dibutyrate and betamethasone tributyrate.

Corticosteroids were used normally as anti-inflammatory drugs. However, in some area certain corticosteroids might be illegally used as growth promoting agent in feed, and as prohibited doping substances in game and sport for human or/and animal performance-enhancing. Synthesized structural similar corticosteroids were popular in black market because they can pass routine drug screening. In this study two new artificial synthesized corticosteroids were found in claimed hydrolyzed wheat product. Liquid chromatography coupled with high resolution mass spectrometry (HRMS, Orbitrap) was applied to separate and elucidate the corticosteroids in the sample. Two unknown peaks with optical spectra similar to corticosteroids were first screened out at the beginning, and then their accurate molecular weight (M + H+) m/z 533.29059 and m/z 603.33289 were detected by HRMS. Element formulas of unknowns were calculated by the accurate mass and isotopes abundance. Structures were proposed by their fragment ions at high energy collision dissociation (HCD, 10 eV) and compared with candidate standard compounds. The two unknowns shared similar molecular skeleton with steroid core structure and presented man made fluorine element in their molecule. As the results, the unknowns in the sample were artificial synthesized, and the sample product was not a real food. The detected corticosteroids were also synthesized as reference compounds for conformation. Two new corticosteroids named betamethasone dibutyrate and betamethasone tributyrate were found and first time reported in this work. The legality of structural similar/modified corticosteroids were blurry and their safety were unverified. The confirmed identifications of two new found corticosteroids, and their mass spectra were provided in this paper for the reference of drug detection.

A Pilot Study Comparing the Effects of Consuming 100% Orange Juice or Sucrose-Sweetened Beverage on Risk Factors for Cardiometabolic Disease in Women.

Overconsumption of sugar-sweetened beverages increases risk factors associated with cardiometabolic disease, in part due to hepatic fructose overload. However, it is not clear whether consumption of beverages containing fructose as naturally occurring sugar produces equivalent metabolic dysregulation as beverages containing added sugars. We compared the effects of consuming naturally-sweetened orange juice (OJ) or sucrose-sweetened beverages (sucrose-SB) for two weeks on risk factors for cardiometabolic disease. Healthy, overweight women (n = 20) were assigned to consume either 3 servings of 100% orange juice or sucrose-SB/day. We conducted 16-hour serial blood collections and 3-h oral glucose tolerance tests during a 30-h inpatient visit at baseline and after the 2-week diet intervention. The 16-h area under the curve (AUC) for uric acid increased in subjects consuming sucrose-SB compared with subjects consuming OJ. Unlike sucrose-SB, OJ did not significantly increase fasting or postprandial lipoproteins. Consumption of both beverages resulted in reductions in the Matsuda insulin sensitivity index (OJ: -0.40 ± 0.18, p = 0.04 within group; sucrose-SB: -1.0 ± 0.38, p = 0.006 within group; p = 0.53 between groups). Findings from this pilot study suggest that consumption of OJ at levels above the current dietary guidelines for sugar intake does not increase plasma uric acid concentrations compared with sucrose-SB, but appears to lead to comparable decreases of insulin sensitivity.

ERK phosphorylates chromosomal axis component HORMA domain protein HTP-1 to regulate oocyte numbers.

Oocyte numbers, a critical determinant of female reproductive fitness, are highly regulated, yet the mechanisms underlying this regulation remain largely undefined. In the Caenorhabditis elegans gonad, RAS/extracellular signal-regulated kinase (ERK) signaling regulates oocyte numbers; mechanisms are unknown. We show that the RAS/ERK pathway phosphorylates meiotic chromosome axis protein HTP-1 at serine-325 to control chromosome dynamics and regulate oocyte number. Phosphorylated HTP-1(S325) accumulates in vivo in an ERK-dependent manner in early-mid pachytene stage germ cells and is necessary for synaptonemal complex extension and/or maintenance. Lack of HTP-1 phosphorylation leads to asynapsis and persistence of meiotic double-strand breaks, causing delayed meiotic progression and reduced oocyte number. In contrast, early onset of ERK activation causes precocious meiotic progression, resulting in increased oocyte number, which is reversed by removal of HTP-1 phosphorylation. The RAS/ERK/HTP-1 signaling cascade thus functions to monitor formation and maintenance of synapsis for timely resolution of double-strand breaks, oocyte production, and reproductive fitness.

Long-term effects of western diet consumption in male and female mice.

Long-term consumption of a diet with excessive fat and sucrose (Western diet, WD) leads to an elevated risk of obesity and metabolic syndrome in both males and females. However, there are sexual dimorphisms in metabolism which are apparent when considering the prevalence of complications of metabolic syndrome, such as non-alcoholic fatty liver disease. This study aimed to elucidate the impact of a WD on the metabolome and the gut microbiota of male and female mice at 5, 10, and 15 months to capture the dynamic and comprehensive changes brought about by diet at different stages of life. Here we show that there are important considerations of age and sex that should be considered when assessing the impact of diet on the gut microbiome and health.

Pilot study of probiotic/colostrum supplementation on gut function in children with autism and gastrointestinal symptoms.

Over half of all children with autism spectrum disorders (ASD) have gastrointestinal (GI) co-morbidities including chronic constipation, diarrhea, and irritable bowel syndrome. The severity of these symptoms has been correlated with the degree of GI microbial dysbiosis. The study objective was to assess tolerability of a probiotic (Bifidobacterium infantis) in combination with a bovine colostrum product (BCP) as a source of prebiotic oligosaccharides and to evaluate GI, microbiome and immune factors in children with ASD and GI co-morbidities. This pilot study is a randomized, double blind, controlled trial of combination treatment (BCP + B. infantis) vs. BCP alone in a cross-over study in children ages 2-11 with ASD and GI co-morbidities (n = 8). This 12-week study included 5 weeks of probiotic-prebiotic supplementation, followed by a two-week washout period, and 5 weeks of prebiotic only supplementation. The primary outcome of tolerability was assessed using validated questionnaires of GI function and atypical behaviors, along with side effects. Results suggest that the combination treatment is well-tolerated in this cohort. The most common side effect was mild gassiness. Some participants on both treatments saw a reduction in the frequency of certain GI symptoms, as well as reduced occurrence of particular aberrant behaviors. Improvement may be explained by a reduction in IL-13 and TNF-α production in some participants. Although limited conclusions can be drawn from this small pilot study, the results support the need for further research into the efficacy of these treatments.

Effect of bovine milk fat globule membranes as a complementary food on the serum metabolome and immune markers of 6-11-month-old Peruvian infants.

This study builds on a previous study by this group in which 6-11-month-old Peruvian infants who were fed bovine milk fat globule membrane (MFGM) containing complementary food had significantly fewer episodes of infection-related bloody diarrhea relative to those consuming a control food (skim milk powder). Micronutrient deficiencies including zinc deficiency were prevalent in this study population. To understand the mechanism behind the health benefits of consuming MFGM, the serum metabolome and cytokine levels, as markers for systemic immune responses, were evaluated using 1H nuclear magnetic resonance-based metabolomics and a multiplex system, respectively. Combined with data on micronutrient status and anthropometry, a comparative analysis was performed. Supplementation with MFGM tended to improve micronutrient status, energy metabolism, and growth reflected as increased levels of circulating amino acids and weight gain, particularly in female infants compared to controls. Decreased levels of the microbial choline metabolite trimethylamine-N-oxide in the MFGM-supplemented group (both male and female infants) suggest a functional perturbation in the intestinal microbiota. A cytokine shift toward a less T helper type 1 response was observed in those receiving the MFGM supplement, which was mainly attributed to decreases in interleukin-2 levels. Our findings suggest that consumption of MFGM with complementary food may reverse the metabolic abnormalities found in marginally nourished infants, thereby improving metabolic regulation, which may lead to enhanced immunity.

Sucrose metabolism alters Lactobacillus plantarum survival and interactions with the microbiota in the digestive tract.

We investigated whether sucrose metabolism by probiotic Lactobacillus plantarum influences the intestinal survival and microbial responses to this organism when administered to mice fed a sucrose-rich, Western diet. A L. plantarum mutant unable to metabolize sucrose was constructed by deleting scrB, coding for beta-fructofuranosidase, in a rifampicin-resistant strain of L. plantarum NCIMB8826. The ScrB deficient mutant survived in 8-fold higher numbers compared to the wild-type strain when measured 24 h after administration on two consecutive days. According to 16S rRNA marker gene sequencing, proportions of Faecalibacterium and Streptococcus were elevated in mice fed the L. plantarum ΔscrB mutant. Metagenome predictions also indicated those mice contained a higher abundance of lactate dehydrogenases. This was further supported by a trend in elevated fecal lactate concentrations among mice fed the ΔscrB mutant. L. plantarum also caused other changes to the fecal metabolomes including higher concentrations of glycerol in mice fed the ΔscrB mutant and increased uracil, acetate and propionate levels among mice fed the wild-type strain. Taken together, these results suggest that sucrose metabolism alters the properties of L. plantarum in the digestive tract and that probiotics can differentially influence intestinal metabolomes via their carbohydrate consumption capabilities.

Bolus injections of novel thrombogenic site-targeted fusion proteins comprising annexin-V and Kunitz protease inhibitors attenuate intimal hyperplasia after balloon angioplasty.

BACKGROUND: Systemic administrations of conventional antithrombotics reduce neointima formation after angioplasty in experimental animals. However, clinical translation of these results has not been successful due to high risk for bleeding. OBJECTIVES: We sought to determine whether novel annexin-V (ANV)-Kunitz protease inhibitor fusion proteins, TAP-ANV and ANV-6L15, can specifically target to vascular injury site and limit neointima formation without inducing systemic hypo-coagulation in a rat carotid artery balloon angioplasty injury model. METHODS: Near infrared imaging was carried out after balloon-injury and injection of fluorescent ANV or ANV-6L15 to examine their bio-distributions. For peri-procedure treatment, TAP-ANV or ANV-6L15 was administered as i.v. boluses 3 times: 30-minutes before balloon-injury, immediate after procedure, and 120-minutes post-balloon-injury. For extended treatment, additional i.v. bolus injection was given on day-2, day-3 and every other day thereafter. Carotid arteries were collected on day-7 and day-14 for analysis. Blood was collected for measurement of clotting parameters. RESULTS: Near infrared imaging and immunochemistry showed that fluorescent ANV and ANV-6L15 specifically localized to injured carotid artery and significant amount of ANV-6L15 remained bound to the injured artery after 24-h. Peri-procedure injections of TAP-ANV or ANV-6L15 resulted in decrease of intima/media ratio by 56%. Extended injections of both yielded similar results. Both decreased the expression of PCNA on day-7 and increased the expression calponin on day-14 in the intima post-balloon-injury. CONCLUSIONS: TAP-ANV and ANV-6L15 can specifically localize to balloon injured carotid arteries after i.v. bolus injections, resulting in substantial attenuation of intimal hyperplasia without inducing a state of systemic hypo-coagulation.