Crohn’s Disease in Clinical Remission Is Marked by Systemic Oxidative Stress

Introduction: Crohn’s disease (CD) is characterized by chronic and relapsing inflammation of the gastro-intestinal tract. It is assumed that oxidative stress contributes to CD pathogenesis, but systemic biomarkers for oxidative stress in CD are not yet identified. A reduction in free thiol groups in plasma proteins (“plasma free thiols”) reflects systemic oxidative stress since they are prime substrates for reactive oxygen species. Here, we determined the concentrations of plasma free thiols in CD patients and healthy controls and studied the putative correlation with disease parameters.Methods: Free thiols were quantified in plasma of patients with CD in clinical remission [according to the Harvey Bradshaw Index (HBI)] and healthy controls and adjusted for plasma albumin. Albumin-adjusted free thiol concentrations were analyzed for associations with clinical and biochemical disease markers.Results: Mean plasma free thiol concentrations were significantly lower in patients with CD (n = 51) compared to healthy controls (n = 27) (14.7 ± 2.4 vs. 17.9 ± 1.8 μmol/g albumin; P < 0.001). Patients with CD with above-average free thiols had significantly lower CRP levels (median 1.4 [interquartile range] [0.4; 2.6] vs. 3.6 [0.6; 7.0] mg/L; P < 0.05) and BMI (23.6 ± 4.8 vs. 27.1 ± 5.2 kg/m2; P < 0.05). Patients with CD having solely colonic disease demonstrated markedly reduced plasma free thiol concentrations compared to patients with ileocolonic involvement (13.2 ± 1.8 vs. 15.2 ± 2.2 μmol/g; P < 0.05). Finally, plasma free thiol concentrations negatively correlated with biomarkers of inflammation, including hsCRP, SAA, IL-17A (all P < 0.05), and VEGF.Conclusion: Plasma free thiols are reduced in patients with CD in clinical remission compared to healthy controls. Thus, subclinical CD disease activity is reflected by systemic oxidative stress and plasma free thiols may be a relevant therapeutic target and biomarker to monitor disease activity in CD

Riboflavin Supplementation in Patients with Crohn's Disease [the RISE-UP study]

Background and Aims: Crohn's disease [CD] is characterised by chronic intestinal inflammation and dysbiosis in the gut. Riboflavin [vitamin B2] has anti-inflammatory, antioxidant and microbiome-modulatory properties. Here, we analysed the effect of riboflavin on oxidative stress, markers of inflammation, clinical symptoms, and faecal microbiome in patients with CD. Methods: In this prospective clinical intervention study, patients received 100 mg riboflavin [DSM, Nutritional Products Ltd] daily for 3 weeks. Clinical disease activity [Harvey-Bradshaw Index: HBI], serum biomarkers of inflammation and redox status [plasma free thiols], and faecal microbiome taxonomical composition and functionality [fluorescent in situ hybridisation: FISH; and metagenomic shotgun sequencing: MGS], were analysed before and after riboflavin intervention. Results: In total, 70 patients with CD with varying disease activity were included. Riboflavin supplementation significantly decreased serum levels of inflammatory markers. In patients with low faecal calprotectin [FC] levels, IL-2 decreased, and in patients with high FC levels, C-reactive protein [CRP] was reduced and free thiols significantly increased after supplementation. Moreover, HBI was significantly decreased by riboflavin supplementation. Riboflavin supplementation led to decreased Enterobacteriaceae in patients with low FC levels as determined by FISH; however, MGS analysis showed no effects on diversity, taxonomy, or metabolic pathways of the faecal microbiome. Conclusions: Three weeks of riboflavin supplementation resulted in a reduction in systemic oxidative stress, mixed anti-inflammatory effects, and a reduction in clinical symptoms [HBI]. FISH analysis showed decreased Enterobacteriaceae in patients with CD with low FC levels, though this was not observed in MGS analysis. Our data demonstrate that riboflavin supplementation has a number of anti-inflammatory and anti-oxidant effects in CD

SARS-CoV-2 infects the human kidney and drives fibrosis in kidney organoids

Kidney failure is frequently observed during and after COVID-19, but it remains elusive whether this is a direct effect of the virus. Here, we report that SARS-CoV-2 directly infects kidney cells and is associated with increased tubule-interstitial kidney fibrosis in patient autopsy samples. To study direct effects of the virus on the kidney independent of systemic effects of COVID-19, we infected human induced pluripotent stem cell-derived kidney organoids with SARS-CoV-2. Single cell RNA-sequencing indicated injury and dedifferentiation of infected cells with activation of pro-fibrotic signaling pathways. Importantly, SARS-CoV-2 infection also led to increased collagen 1 protein expression in organoids. A SARS-CoV-2 protease inhibitor was able to ameliorate the infection of kidney cells by SARS-CoV-2. Our results suggest that SARS-CoV-2 can directly infect kidney cells and induce cell injury with subsequent fibrosis. These data could explain both acute kidney injury in COVID-19 patients and the development of chronic kidney disease in Long-COVID

The K+ Channel Opener 1-EBIO Potentiates Residual Function of Mutant CFTR in Rectal Biopsies from Cystic Fibrosis Patients

BACKGROUND: The identification of strategies to improve mutant CFTR function remains a key priority in the development of new treatments for cystic fibrosis (CF). Previous studies demonstrated that the K⁺ channel opener 1-ethyl-2-benzimidazolone (1-EBIO) potentiates CFTR-mediated Cl⁻ secretion in cultured cells and mouse colon. However, the effects of 1-EBIO on wild-type and mutant CFTR function in native human colonic tissues remain unknown. METHODS: We studied the effects of 1-EBIO on CFTR-mediated Cl⁻ secretion in rectal biopsies from 47 CF patients carrying a wide spectrum of CFTR mutations and 57 age-matched controls. Rectal tissues were mounted in perfused micro-Ussing chambers and the effects of 1-EBIO were compared in control tissues, CF tissues expressing residual CFTR function and CF tissues with no detectable Cl⁻ secretion. RESULTS: Studies in control tissues demonstrate that 1-EBIO activated CFTR-mediated Cl⁻ secretion in the absence of cAMP-mediated stimulation and potentiated cAMP-induced Cl⁻ secretion by 39.2±6.7% (P<0.001) via activation of basolateral Ca²⁺-activated and clotrimazole-sensitive KCNN4 K⁺ channels. In CF specimens, 1-EBIO potentiated cAMP-induced Cl⁻ secretion in tissues with residual CFTR function by 44.4±11.5% (P<0.001), but had no effect on tissues lacking CFTR-mediated Cl⁻ conductance. CONCLUSIONS: We conclude that 1-EBIO potentiates Cl⁻secretion in native CF tissues expressing CFTR mutants with residual Cl⁻ channel function by activation of basolateral KCNN4 K⁺ channels that increase the driving force for luminal Cl⁻ exit. This mechanism may augment effects of CFTR correctors and potentiators that increase the number and/or activity of mutant CFTR channels at the cell surface and suggests KCNN4 as a therapeutic target for CF

SARS-CoV-2 infects the human kidney and drives fibrosis in kidney organoids

Kidney failure is frequently observed during and after COVID-19, but it remains elusive whether this is a direct effect of the virus. Here, we report that SARS-CoV-2 directly infects kidney cells and is associated with increased tubule-interstitial kidney fibrosis in patient autopsy samples. To study direct effects of the virus on the kidney independent of systemic effects of COVID-19, we infected human-induced pluripotent stem-cell-derived kidney organoids with SARS-CoV-2. Single-cell RNA sequencing indicated injury and dedifferentiation of infected cells with activation of profibrotic signaling pathways. Importantly, SARS-CoV-2 infection also led to increased collagen 1 protein expression in organoids. A SARS-CoV-2 protease inhibitor was able to ameliorate the infection of kidney cells by SARS-CoV-2. Our results suggest that SARS-CoV-2 can directly infect kidney cells and induce cell injury with subsequent fibrosis. These data could explain both acute kidney injury in COVID-19 patients and the development of chronic kidney disease in long COVID

Matrix-metalloproteinase activity in first trimester placental bed biopsies in further complicated and uncomplicated pregnancies

Trophoblast invasion is partly regulated by matrix-metalloproteinases (MMPs). Aberrations in MPAP-activity in early pregnancy are thought to play a role in the pathophysiology of pregnancy associated conditions like pre-eclampsia and intra-uterine growth restriction (IUGR). A direct relation however, has not been published. We tested the hypothesis that MMP activity in the decidua is compromised in the first trimester of pregnancies, which are complicated by hypertensive disorders or IUGR in later pregnancy. During chorionic villus biopsy, decidua is microscopically separated from the villi and stored. A selection of pregnancies complicated by pre-eclampsia or HELLP-syndrome or IUGR was made, with two matched controls each. Zymography was performed to identify the presence of MMPs, and subsequently immunohistochemistry for MMP-2 and -9 and cytokeratin 7 to examine differences between cases and controls. Next, a specific immuno-capture assay was used to determine the activity of MMP-1, -2, -3, -8, -9, and 13, total as well as active. Although presence of MMP-2 and MMP-9 AN as found, which corresponded with the immunohistochemistry, no significant differences could be demonstrated between activity of total MMP-2 and total MMP-9 in complicated and uncomplicated pregnancies. Activity of MMP-1, -3, -8 and -13 could not be detected. In conclusion: our study confirms the presence of MMP-2 and -9 in first trimester placental bed biopsies, but does not support the current concept of deranged NIMP-activity in early pregnancy in further complicated pregnancies. (C) 2003 Elsevier Ltd. All rights reserved

The effect of riboflavin supplementation on the systemic redox status in healthy volunteers:A post-hoc analysis of the RIBOGUT trial

Background : Riboflavin is a redox-active vitamin that plays a pivotal role in human energy metabolism. Riboflavin may have beneficial health effects by increasing extracellular antioxidant capacity, thereby alleviating oxidative stress. Reduced levels of free thiols in blood reflect systemic oxidative stress, since they are readily oxidized by reactive species. In this study, we aimed to study the potential of riboflavin supplementation to improve the systemic redox status in healthy volunteers. Methods : This study was a post-hoc analysis of the RIBOGUT study, a randomized, double-blind, placebo-controlled human intervention trial that investigated the effect of riboflavin supplements on the gut microbiota composition of healthy individuals. Serum free thiols were quantified before and after intervention and adjusted to serum albumin levels. Changes in albumin-adjusted free thiols were analyzed, as well as potential associations with routine laboratory parameters and fecal bacterial quantification by fluorescence in-situ hybridization (FISH). Results : Participants were randomized to either placebo (n=34), riboflavin 50 mg daily (n=32), or riboflavin 100 mg daily (n=33). At baseline, no significant differences in albumin-adjusted serum free thiols were observed. After intervention with either placebo or riboflavin, albumin-adjusted serum free thiols did not significantly change (P>0.05), however, observed changes were inversely associated with changes in C-reactive protein (CRP) levels (r= -0.22, P<0.05). At baseline, albumin-adjusted serum free thiols were positively associated with fecal relative abundances of Faecalibacterium prausnitzii (P<0.01). Conclusion : Riboflavin did not change the systemic redox status in healthy individuals as reflected by serum free thiols, but observed changes in albumin-adjusted free thiol levels were negatively associated with changes in CRP levels. Strikingly, albumin-adjusted free thiols were independently associated with relative abundances of fecal F. prausnitzii, which may suggest a potential host redox-microbiota interaction

Fluid hydration to prevent post-ERCP pancreatitis in average- to high-risk patients receiving prophylactic rectal NSAIDs (FLUYT trial): study protocol for a randomized controlled trial

Contains fulltext : 190882.pdf (publisher's version ) (Open Access)BACKGROUND: Post-endoscopic retrograde cholangiopancreatography (ERCP) pancreatitis (PEP) is the most common complication of ERCP and may run a severe course. Evidence suggests that vigorous periprocedural hydration can prevent PEP, but studies to date have significant methodological drawbacks. Importantly, evidence for its added value in patients already receiving prophylactic rectal non-steroidal anti-inflammatory drugs (NSAIDs) is lacking and the cost-effectiveness of the approach has not been investigated. We hypothesize that combination therapy of rectal NSAIDs and periprocedural hydration would significantly lower the incidence of post-ERCP pancreatitis compared to rectal NSAIDs alone in moderate- to high-risk patients undergoing ERCP. METHODS: The FLUYT trial is a multicenter, parallel group, open label, superiority randomized controlled trial. A total of 826 moderate- to high-risk patients undergoing ERCP that receive prophylactic rectal NSAIDs will be randomized to a control group (no fluids or normal saline with a maximum of 1.5 mL/kg/h and 3 L/24 h) or intervention group (lactated Ringer's solution with 20 mL/kg over 60 min at start of ERCP, followed by 3 mL/kg/h for 8 h thereafter). The primary endpoint is the incidence of post-ERCP pancreatitis. Secondary endpoints include PEP severity, hydration-related complications, and cost-effectiveness. DISCUSSION: The FLUYT trial design, including hydration schedule, fluid type, and sample size, maximize its power of identifying a potential difference in post-ERCP pancreatitis incidence in patients receiving prophylactic rectal NSAIDs. TRIAL REGISTRATION: EudraCT: 2015-000829-37 . Registered on 18 February 2015. ISRCTN: 13659155 . Registered on 18 May 2015

Prenatal Diagnosis and Carrier Detection of Duchenne Muscular Dystrophy with Closely Linked RFLPs

By the use of a series of closely linked DNA probes detecting restriction fragment length polymorphisms (RFLPs) distributed over the short arm of the X chromosome, a double crossover was detected in a Duchenne muscular dystrophy carrier and an affected male fetus was diagnosed at 12 weeks of gestation, with a probable accuracy of more than 99·0%. A new mutation was identified in another family with the same degree of reliability; three females in this family were thus deemed not to be DMD carriers. The eleven RFLP-markers presently available on the short arm of the X chromosome are useful in the diagnosis of DMD since they bridge the Duchenne locus at genetic distances varying between 3 and 20 cmo. Moreover, recombination within the set of markers provides an independent way of regionally mapping these probes relative to each other along the short arm of the X chromosome