Regulation of peripheral inflammation by spinal p38 MAP kinase in rats.

BackgroundSomatic afferent input to the spinal cord from a peripheral inflammatory site can modulate the peripheral response. However, the intracellular signaling mechanisms in the spinal cord that regulate this linkage have not been defined. Previous studies suggest spinal cord p38 mitogen-activated protein (MAP) kinase and cytokines participate in nociceptive behavior. We therefore determined whether these pathways also regulate peripheral inflammation in rat adjuvant arthritis, which is a model of rheumatoid arthritis.Methods and findingsSelective blockade of spinal cord p38 MAP kinase by administering the p38 inhibitor SB203580 via intrathecal (IT) catheters in rats with adjuvant arthritis markedly suppressed paw swelling, inhibited synovial inflammation, and decreased radiographic evidence of joint destruction. The same dose of SB203580 delivered systemically had no effect, indicating that the effect was mediated by local concentrations in the neural compartment. Evaluation of articular gene expression by quantitative real-time PCR showed that spinal p38 inhibition markedly decreased synovial interleukin-1 and -6 and matrix metalloproteinase (MMP3) gene expression. Activation of p38 required tumor necrosis factor alpha (TNFalpha) in the nervous system because IT etanercept (a TNF inhibitor) given during adjuvant arthritis blocked spinal p38 phosphorylation and reduced clinical signs of adjuvant arthritis.ConclusionsThese data suggest that peripheral inflammation is sensed by the central nervous system (CNS), which subsequently activates stress-induced kinases in the spinal cord via a TNFalpha-dependent mechanism. Intracellular p38 MAP kinase signaling processes this information and profoundly modulates somatic inflammatory responses. Characterization of this mechanism could have clinical and basic research implications by supporting development of new treatments for arthritis and clarifying how the CNS regulates peripheral immune responses

AI is a viable alternative to high throughput screening: a 318-target study

: High throughput screening (HTS) is routinely used to identify bioactive small molecules. This requires physical compounds, which limits coverage of accessible chemical space. Computational approaches combined with vast on-demand chemical libraries can access far greater chemical space, provided that the predictive accuracy is sufficient to identify useful molecules. Through the largest and most diverse virtual HTS campaign reported to date, comprising 318 individual projects, we demonstrate that our AtomNet® convolutional neural network successfully finds novel hits across every major therapeutic area and protein class. We address historical limitations of computational screening by demonstrating success for target proteins without known binders, high-quality X-ray crystal structures, or manual cherry-picking of compounds. We show that the molecules selected by the AtomNet® model are novel drug-like scaffolds rather than minor modifications to known bioactive compounds. Our empirical results suggest that computational methods can substantially replace HTS as the first step of small-molecule drug discovery

Mortality from gastrointestinal congenital anomalies at 264 hospitals in 74 low-income, middle-income, and high-income countries: a multicentre, international, prospective cohort study

Summary Background Congenital anomalies are the fifth leading cause of mortality in children younger than 5 years globally. Many gastrointestinal congenital anomalies are fatal without timely access to neonatal surgical care, but few studies have been done on these conditions in low-income and middle-income countries (LMICs). We compared outcomes of the seven most common gastrointestinal congenital anomalies in low-income, middle-income, and high-income countries globally, and identified factors associated with mortality. Methods We did a multicentre, international prospective cohort study of patients younger than 16 years, presenting to hospital for the first time with oesophageal atresia, congenital diaphragmatic hernia, intestinal atresia, gastroschisis, exomphalos, anorectal malformation, and Hirschsprung’s disease. Recruitment was of consecutive patients for a minimum of 1 month between October, 2018, and April, 2019. We collected data on patient demographics, clinical status, interventions, and outcomes using the REDCap platform. Patients were followed up for 30 days after primary intervention, or 30 days after admission if they did not receive an intervention. The primary outcome was all-cause, in-hospital mortality for all conditions combined and each condition individually, stratified by country income status. We did a complete case analysis. Findings We included 3849 patients with 3975 study conditions (560 with oesophageal atresia, 448 with congenital diaphragmatic hernia, 681 with intestinal atresia, 453 with gastroschisis, 325 with exomphalos, 991 with anorectal malformation, and 517 with Hirschsprung’s disease) from 264 hospitals (89 in high-income countries, 166 in middleincome countries, and nine in low-income countries) in 74 countries. Of the 3849 patients, 2231 (58·0%) were male. Median gestational age at birth was 38 weeks (IQR 36–39) and median bodyweight at presentation was 2·8 kg (2·3–3·3). Mortality among all patients was 37 (39·8%) of 93 in low-income countries, 583 (20·4%) of 2860 in middle-income countries, and 50 (5·6%) of 896 in high-income countries (p<0·0001 between all country income groups). Gastroschisis had the greatest difference in mortality between country income strata (nine [90·0%] of ten in lowincome countries, 97 [31·9%] of 304 in middle-income countries, and two [1·4%] of 139 in high-income countries; p≤0·0001 between all country income groups). Factors significantly associated with higher mortality for all patients combined included country income status (low-income vs high-income countries, risk ratio 2·78 [95% CI 1·88–4·11], p<0·0001; middle-income vs high-income countries, 2·11 [1·59–2·79], p<0·0001), sepsis at presentation (1·20 [1·04–1·40], p=0·016), higher American Society of Anesthesiologists (ASA) score at primary intervention (ASA 4–5 vs ASA 1–2, 1·82 [1·40–2·35], p<0·0001; ASA 3 vs ASA 1–2, 1·58, [1·30–1·92], p<0·0001]), surgical safety checklist not used (1·39 [1·02–1·90], p=0·035), and ventilation or parenteral nutrition unavailable when needed (ventilation 1·96, [1·41–2·71], p=0·0001; parenteral nutrition 1·35, [1·05–1·74], p=0·018). Administration of parenteral nutrition (0·61, [0·47–0·79], p=0·0002) and use of a peripherally inserted central catheter (0·65 [0·50–0·86], p=0·0024) or percutaneous central line (0·69 [0·48–1·00], p=0·049) were associated with lower mortality. Interpretation Unacceptable differences in mortality exist for gastrointestinal congenital anomalies between lowincome, middle-income, and high-income countries. Improving access to quality neonatal surgical care in LMICs will be vital to achieve Sustainable Development Goal 3.2 of ending preventable deaths in neonates and children younger than 5 years by 2030

Regulation of peripheral inflammation by spinal p38 MAP kinase in rats.

BackgroundSomatic afferent input to the spinal cord from a peripheral inflammatory site can modulate the peripheral response. However, the intracellular signaling mechanisms in the spinal cord that regulate this linkage have not been defined. Previous studies suggest spinal cord p38 mitogen-activated protein (MAP) kinase and cytokines participate in nociceptive behavior. We therefore determined whether these pathways also regulate peripheral inflammation in rat adjuvant arthritis, which is a model of rheumatoid arthritis.Methods and findingsSelective blockade of spinal cord p38 MAP kinase by administering the p38 inhibitor SB203580 via intrathecal (IT) catheters in rats with adjuvant arthritis markedly suppressed paw swelling, inhibited synovial inflammation, and decreased radiographic evidence of joint destruction. The same dose of SB203580 delivered systemically had no effect, indicating that the effect was mediated by local concentrations in the neural compartment. Evaluation of articular gene expression by quantitative real-time PCR showed that spinal p38 inhibition markedly decreased synovial interleukin-1 and -6 and matrix metalloproteinase (MMP3) gene expression. Activation of p38 required tumor necrosis factor alpha (TNFalpha) in the nervous system because IT etanercept (a TNF inhibitor) given during adjuvant arthritis blocked spinal p38 phosphorylation and reduced clinical signs of adjuvant arthritis.ConclusionsThese data suggest that peripheral inflammation is sensed by the central nervous system (CNS), which subsequently activates stress-induced kinases in the spinal cord via a TNFalpha-dependent mechanism. Intracellular p38 MAP kinase signaling processes this information and profoundly modulates somatic inflammatory responses. Characterization of this mechanism could have clinical and basic research implications by supporting development of new treatments for arthritis and clarifying how the CNS regulates peripheral immune responses

Activity of immobilized Thermomyces lanuginosus and Candida antarctica B lipases in interesterification reactions: effect of the aqueous microenvironment

Lipases are enzymes used in numerous reactions of industrial interest. Depending on their aqueous microenvironment, lipases can catalyze hydrolysis or, conversely, organic synthesis like interesterification. This reaction can be used as a method to modify the physical and chemical properties of fats and oils, a basic process for production of "structured lipids". For such synthesis reactions, thermodynamic water activity (a(w)) of the catalyst is generally the most important parameter to control. Actually, it will directly determine the performance of the synthesis, namely its yield, selectivity and stability. Effect of the a(w) on the activity of immobilized Thermomyces lanuginosus and Candida antarctica B lipases in interesterification reactions was studied. Water sorption and desorption isotherms were determined, showing a phenomenon of hysteresis for the Thermomyces lanuginosus lipase. Evaluation of the influence of a(w) on reaction yields revealed that the IE activity tends to increase with the water activity of immobilized Thermomyces lanuginosus lipase. In contrast, a(w) had little influence in the case of the Candida antarctica B lipase

Effect of IT p38 Inhibitor on Paw Swelling in Adjuvant Arthritis

<p>Rats were immunized with CFA on day 0 and treated daily with IT p38 inhibitors (SB203580 at 8 μg/d), SC p38 inhibitor (SB203580 at 8 μg/d), or IT saline beginning on days 8–20. Paw swelling was measured by water displacement plethysmometry. The IT p38 inhibitor significantly decreased paw swelling (*<i>p</i> = 0.001 by ANOVA and Dunnett's post-ANOVA test; <i>n</i> = 5–6/group). This graph presents data from one of three separate experiments with similar results.</p

Phosphorylation of p38 MAP Kinase in the Spinal Cord

<div><p>(A) Western blot analysis of p38, P-p38, and COX2 in the spinal cord of rats with adjuvant arthritis. Rats were sacrificed from day (D) 0 through day 17. Note faint P-p38 staining on days 8 to day 17. GAPDH normalized expression = 0.22 ± 0.02 for prearthritis (days 0–5) and 0.32 ± 0.3 postarthritis (days 8–17; <i>p</i> = 0.0255).</p> <p>(B) Low-power view showing P-p38 expression in adjuvant immunized rats, especially in lamina II (CST, corticospinal tract).</p> <p>(C–E) High power view of P-p38 (C), OX-42 (D), and merged view (E) demonstrating expression of P-p38 in microglia of adjuvant immunized rats.</p> <p>(F–H) High-power view of P-p38 (F), NeuN (G), and merged view (H) demonstrating scattered neurons expressing P-p38 in adjuvant immunized rats.</p> <p>(I) and (J) show higher power view of (E) and (H), respectively.</p> <p>Arrow indicates positively staining cells in all photomicrographs.</p></div

Effect of IT p38 Inhibitor on Joint Damage and Gene Expression in Adjuvant Arthritis

<div><p>Rats were immunized with CFA on day 0 and treated daily with IT SB203580 (8 μg) or IT saline beginning on days 8–20.</p> <p>(A) Radiographs of the hind paws were evaluated using a scoring system that demonstrated decreased damage in the SB203580-treated group compared with control rats. *<i>p</i> = 0.036 for IT SB203580 (<i>n</i> = 6/group). Similar results were observed in a separate experiment with treatment beginning on day 8 (<i>n</i> = 5–6/group).</p> <p>(B) Representative radiographs illustrate decreased joint destruction in the treated group.</p> <p>(C) Representative hematoxylin and eosin-stained sections demonstrate decreased erosions, cartilage damage, and synovial inflammation in the IT SB203580-treated group.</p> <p>(D) Quantitative real-time PCR was performed on the hind paws as described in Methods. Normal rat joints are also shown as a control. IT SB203580 significantly decreased IL-1 (*<i>p</i> = 0.025), IL-6 (**<i>p</i> = 0.006), and MMP3 (+<i>p</i> = 0.001) gene expression (<i>n</i> = 5–6/group); TNFα gene expression was modestly reduced, but did not reach statistical significance (̂<i>p</i> = 0.089).</p></div

Effect of IT Etanercept on Adjuvant Arthritis

<div><p>Rats were immunized with CFA on day 0 and treated with IT etanercept (100 or 300 μg q.o.d.) beginning on day 1.</p> <p>(A) IT etanercept (Etan) significantly decreased paw swelling (*<i>p</i> = 0.001 for IT saline compared with either IT etanercept group) (<i>n</i> = 6/group).</p> <p>(B) IT etanercept significantly decreased joint damage (by radiographic score; *<i>p</i> = 0.026 for IT saline compared with either IT etanercept group) (<i>n</i> = 6/group).</p> <p>(C) Similar doses of etanercept given systemically (SC) beginning on day 1 had minimal effect on paw swelling (<i>p</i> = 0.398 for IT saline compared with either etanercept group) (<i>n</i> = 6/group).</p> <p>(D and E) IT etanercept decreased spinal P-p38 staining in adjuvant arthritis. Rats were immunized on day 0 and injected IT with 300 μg of etanercept or vehicle on days 7 and 10. They were sacrificed on day 11 and P-p38 was determined by Western blot (D) and immunofluorescence (E). Western blot data were normalized to GAPDH and demonstrated significantly lower levels of P-p38 after IT etanercept treatment (<i>n</i> = 6 for etanercept, 4 for saline, and 2 for naïve). *<i>p</i> = 0.023 compared with IT saline.</p></div