Induced Mutagenesis in UGT74S1 Gene Leads to Stable New Flax Lines with Altered Secoisolariciresinol Diglucoside (SDG) Profiles

Flax secoisolariciresinol (SECO) diglucoside (SDG) lignan is an emerging natural product purported to prevent chronic diseases in humans. SECO, the aglycone form of SDG, has shown higher intestinal cell absorption but it is not accumulated naturally in planta. Recently, we have identified and characterized a UDP-glucosyltransferase gene, UGT74S1, that glucosylates SECO into its monoglucoside (SMG) and SDG forms when expressed in yeast. However, whether this gene is unique in controlling SECO glucosylation into SDG in planta is unclear. Here, we report on the use of UGT74S1 in reverse and forward genetics to characterize an ethyl methane sulfonate (EMS) mutagenized flax population from cultivar CDC Bethune and consisting of 1996 M2 families. EMS mutagenesis generated 73 SNP variants causing 79 mutational events in the UGT74S1 exonic regions of 93 M2 families. The mutation frequency in the exonic regions was determined to be one per 28 Kb. Of these mutations, 13 homozygous missense mutations and two homozygous nonsense mutations were observed and all were transmitted into the M3 and M4 generations. Forward genetics screening of the population showed homozygous nonsense mutants completely lacking SDG biosynthesis while the production of SMG was observed only in a subset of the M4 lines. Heterozygous or homozygous M4 missense mutants displayed a wide range of SDG levels, some being greater than those of CDC Bethune. No additional deleterious mutations were detected in these mutant lines using a panel of 10 other genes potentially involved in the lignan biosynthesis. This study provides further evidence that UGT74S1 is unique in controlling SDG formation from SECO and this is the first report of non-transgenic flax germplasm with simultaneous knockout of SDG and presence of SMG in planta

Persistence of anticancer activity in berry extracts after simulated gastrointestinal digestion and colonic fermentation

Fruit and vegetable consumption is associated at the population level with a protective effect against colorectal cancer. Phenolic compounds, especially abundant in berries, are of interest due to their putative anticancer activity. After consumption, however, phenolic compounds are subject to digestive conditions within the gastrointestinal tract that alter their structures and potentially their function. However, the majority of phenolic compounds are not efficiently absorbed in the small intestine and a substantial portion pass into the colon. We characterized berry extracts (raspberries, strawberries, blackcurrants) produced by in vitro-simulated upper intestinal tract digestion and subsequent fecal fermentation. These extracts and selected individual colonic metabolites were then evaluated for their putative anticancer activities using in vitro models of colorectal cancer, representing the key stages of initiation, promotion and invasion. Over a physiologically-relevant dose range (0–50 µg/ml gallic acid equivalents), the digested and fermented extracts demonstrated significant anti-genotoxic, anti-mutagenic and anti-invasive activity on colonocytes. This work indicates that phenolic compounds from berries undergo considerable structural modifications during their passage through the gastrointestinal tract but their breakdown products and metabolites retain biological activity and can modulate cellular processes associated with colon cancer

Amyotrophic lateral sclerosis-motor neuron disease, monoclonal gammopathy, hyperparathyroidism, and B12 deficiency: case report and review of the literature

<p>Abstract</p> <p>Introduction</p> <p>Amyotrophic lateral sclerosis (the most common form of motor neuron disease) is a progressive and devastating disease involving both lower and upper motor neurons, typically following a relentless path towards death. Given the gravity of this diagnosis, all efforts must be made by the clinician to exclude alternative and more treatable entities. Frequent serology testing involves searching for treatable disorders, including vitamin B12 deficiency, parathyroid anomalies, and monoclonal gammopathies.</p> <p>Case presentation</p> <p>We present the case of a 78-year-old Caucasian man with all three of the aforementioned commonly searched for disorders during an investigation for amyotrophic lateral sclerosis.</p> <p>Conclusions</p> <p>The clinical utility of these common tests and what they ultimately mean in patients with amyotrophic lateral sclerosis is discussed, along with a review of the literature.</p

Changes in body mass index by age, gender, and socio-economic status among a cohort of Norwegian men and women (1990–2001)

<p>Abstract</p> <p>Background</p> <p>Consistent with global trends, the prevalence of obesity is increasing among Norwegian adults. This study aimed to investigate individual trends in BMI (kg/m²) by age, gender, and socio-economic status over an 11-year period.</p> <p>Methods</p> <p>A cohort of 1169 adults (n = 581 men; n = 588 women) self-reported BMI during a general health interview twice administered in two regions in Norway.</p> <p>Results</p> <p>Average BMI increased significantly from 23.7 (SD = 3.4) to 25.4 (SD = 3.8), with equivalent increases for both genders. Proportion of obesity (BMI ≥ 30) increased from 4% to 11% for women and 5% to 13% for men. Of those already classified as overweight or obese in 1990, 68% had gained additional weight 10 years later, by an average increase of 2.6 BMI units. The greatest amount of weight gain occurred for the youngest adults (aged 20–29 years). Age-adjusted general linear models revealed that in 1990, women with a lower level of education had a significantly greater BMI than more educated women. In both 1990 and 2001, rural men with the highest level of household income had a greater BMI than rural men earning less income. Weight gain occurred across all education and income brackets, with no differential associations between SES strata and changes in BMI for either gender or region.</p> <p>Conclusion</p> <p>Results demonstrated significant yet gender-equivalent increases in BMI over an 11-year period within this cohort of Norwegian adults. Whereas socio-economic status exerted minimal influence on changes in BMI over time, young adulthood appeared to be a critical time period at which accelerated weight gain occurred.</p

Neonatal mortality risk for vulnerable newborn types in 15 countries using 125.5 million nationwide birth outcome records, 2000-2020.

OBJECTIVE: To compare neonatal mortality associated with six novel vulnerable newborn types in 125.5 million live births across 15 countries, 2000-2020. DESIGN: Population-based, multi-country study. SETTING: National data systems in 15 middle- and high-income countries. METHODS: We used individual-level data sets identified for the Vulnerable Newborn Measurement Collaboration. We examined the contribution to neonatal mortality of six newborn types combining gestational age (preterm [PT] versus term [T]) and size-for-gestational age (small [SGA], 90th centile) according to INTERGROWTH-21st newborn standards. Newborn babies with PT or SGA were defined as small and T + LGA was considered as large. We calculated risk ratios (RRs) and population attributable risks (PAR%) for the six newborn types. MAIN OUTCOME MEASURES: Mortality of six newborn types. RESULTS: Of 125.5 million live births analysed, risk ratios were highest among PT + SGA (median 67.2, interquartile range [IQR] 45.6-73.9), PT + AGA (median 34.3, IQR 23.9-37.5) and PT + LGA (median 28.3, IQR 18.4-32.3). At the population level, PT + AGA was the greatest contributor to newborn mortality (median PAR% 53.7, IQR 44.5-54.9). Mortality risk was highest among newborns born before 28 weeks (median RR 279.5, IQR 234.2-388.5) compared with babies born between 37 and 42 completed weeks or with a birthweight less than 1000 g (median RR 282.8, IQR 194.7-342.8) compared with those between 2500 g and 4000 g as a reference group. CONCLUSION: Preterm newborn types were the most vulnerable, and associated with the highest mortality, particularly with co-existence of preterm and SGA. As PT + AGA is more prevalent, it is responsible for the greatest burden of neonatal deaths at population level

Excess Circulating Alternatively Activated Myeloid (M2) Cells Accelerate ALS Progression While Inhibiting Experimental Autoimmune Encephalomyelitis

Circulating immune cells including autoreactive T cells and monocytes have been documented as key players in maintaining, protecting and repairing the central nervous system (CNS) in health and disease. Here, we hypothesized that neurodegenerative diseases might be associated, similarly to tumors, with increased levels of circulating peripheral myeloid derived suppressor cells (MDSCs), representing a subset of suppressor cells that often expand under pathological conditions and inhibit possible recruitment of helper T cells needed for fighting off the disease.We tested this working hypothesis in amyotrophic lateral sclerosis (ALS) and its mouse model, which are characterized by a rapid progression once clinical symptoms are evident. Adaptive transfer of alternatively activated myeloid (M2) cells, which homed to the spleen and exhibited immune suppressive activity in G93A mutant superoxide dismutase-1 (mSOD1) mice at a stage before emergence of disease symptoms, resulted in earlier appearance of disease symptoms and shorter life expectancy. The same protocol mitigated the inflammation-induced disease model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE), which requires circulating T cells for disease induction. Analysis of whole peripheral blood samples obtained from 28 patients suffering from sporadic ALS (sALS), revealed a two-fold increase in the percentage of circulating MDSCs (LIN(-/Low)HLA-DR(-)CD33(+)) compared to controls.Taken together, these results emphasize the distinct requirements for fighting the inflammatory neurodegenerative disease, multiple sclerosis, and the neurodegenerative disease, ALS, though both share a local inflammatory component. Moreover, the increased levels of circulating MDSCs in ALS patients indicates the operation of systemic mechanisms that might lead to an impairment of T cell reactivity needed to overcome the disease conditions within the CNS. This high level of suppressive immune cells might represent a risk factor and a novel target for therapeutic intervention in ALS at least at the early stage

Namilumab or infliximab compared with standard of care in hospitalised patients with COVID-19 (CATALYST): a randomised, multicentre, multi-arm, multistage, open-label, adaptive, phase 2, proof-of-concept trial

BACKGROUND: Dysregulated inflammation is associated with poor outcomes in COVID-19. We aimed to assess the efficacy of namilumab (a granulocyte-macrophage colony stimulating factor inhibitor) and infliximab (a tumour necrosis factor inhibitor) in hospitalised patients with COVID-19, to prioritise agents for phase 3 trials. METHODS: In this randomised, multicentre, multi-arm, multistage, parallel-group, open-label, adaptive, phase 2, proof-of-concept trial (CATALYST), we recruited patients (aged ≥16 years) admitted to hospital with COVID-19 pneumonia and C-reactive protein (CRP) concentrations of 40 mg/L or greater, at nine hospitals in the UK. Participants were randomly assigned with equal probability to usual care or usual care plus a single intravenous dose of namilumab (150 mg) or infliximab (5 mg/kg). Randomisation was stratified by care location within the hospital (ward vs intensive care unit [ICU]). Patients and investigators were not masked to treatment allocation. The primary endpoint was improvement in inflammation, measured by CRP concentration over time, analysed using Bayesian multilevel models. This trial is now complete and is registered with ISRCTN, 40580903. FINDINGS: Between June 15, 2020, and Feb 18, 2021, we screened 299 patients and 146 were enrolled and randomly assigned to usual care (n=54), namilumab (n=57), or infliximab (n=35). For the primary outcome, 45 patients in the usual care group were compared with 52 in the namilumab group, and 29 in the usual care group were compared with 28 in the infliximab group. The probabilities that the interventions were superior to usual care alone in reducing CRP concentration over time were 97% for namilumab and 15% for infliximab; the point estimates for treatment–time interactions were –0·09 (95% CI –0·19 to 0·00) for namilumab and 0·06 (–0·05 to 0·17) for infliximab. 134 adverse events occurred in 30 (55%) of 55 patients in the namilumab group compared with 145 in 29 (54%) of 54 in the usual care group. 102 adverse events occurred in 20 (69%) of 29 patients in the infliximab group compared with 112 in 17 (50%) of 34 in the usual care group. Death occurred in six (11%) patients in the namilumab group compared with ten (19%) in the usual care group, and in four (14%) in the infliximab group compared with five (15%) in the usual care group. INTERPRETATION: Namilumab, but not infliximab, showed proof-of-concept evidence for reduction in inflammation—as measured by CRP concentration—in hospitalised patients with COVID-19 pneumonia. Namilumab should be prioritised for further investigation in COVID-19. FUNDING: Medical Research Council

Rod and Cone Pathway Signalling Is Altered in the P2X7 Receptor Knock Out Mouse

The P2X7 receptor (P2X7-R) is expressed in the retina and brain and has been implicated in neurodegenerative diseases. However, whether it is expressed by neurons and plays a role as a neurotransmitter receptor has been the subject of controversy. In this study, we first show that the novel vesicular transporter for ATP, VNUT, is expressed in the retina, verifying the presence of the molecular machinery for ATP to act as neurotransmitter at P2X7-Rs. Secondly we show the presence of P2X7-R mRNA and protein in the retina and cortex and absence of the full length variant 1 of the receptor in the P2X7-R knock out (P2X7-KO) mouse. The role of the P2X7-R in neuronal function of the retina was assessed by comparing the electroretinogram response of P2X7-KO with WT mice. The rod photoreceptor response was found to be similar, while both rod and cone pathway post-photoreceptor responses were significantly larger in P2X7-KO mice. This suggests that activation of P2X7-Rs modulates output of second order retinal neurons. In line with this finding, P2X7-Rs were found in the outer plexiform layer and on inner retinal cell classes, including horizontal, amacrine and ganglion cells. The receptor co-localized with conventional synapses in the IPL and was expressed on amacrine cells post-synaptic to rod bipolar ribbon synapses. In view of the changes in visual function in the P2X7-KO mouse and the immunocytochemical location of the receptor in the normal retina, it is likely the P2X7-R provides excitatory input to photoreceptor terminals or to inhibitory cells that shape both the rod and cone pathway response

Electrophysiological Characterization of The Cerebellum in the Arterially Perfused Hindbrain and Upper Body of The Rat

In the present study, a non-pulsatile arterially perfused hindbrain and upper body rat preparation is described which is an extension of the brainstem preparation reported by Potts et al., (Brain Res Bull 53(1):59–67), 1. The modified in situ preparation allows study of cerebellar function whilst preserving the integrity of many of its interconnections with the brainstem, upper spinal cord and the peripheral nervous system of the head and forelimbs. Evoked mossy fibre, climbing fibre and parallel fibre field potentials and EMG activity elicited in forelimb biceps muscle by interpositus stimulation provided evidence that both cerebellar inputs and outputs remain operational in this preparation. Similarly, the spontaneous and evoked single unit activity of Purkinje cells, putative Golgi cells, molecular interneurones and cerebellar nuclear neurones was similar to activity patterns reported in vivo. The advantages of the preparation include the ability to record, without the complications of anaesthesia, stabile single unit activity for extended periods (3 h or more), from regions of the rat cerebellum that are difficult to access in vivo. The preparation should therefore be a useful adjunct to in vitro and in vivo studies of neural circuits underlying cerebellar contributions to movement control and motor learning